diff --git "a/deduped/dedup_0532.jsonl" "b/deduped/dedup_0532.jsonl"
new file mode 100644--- /dev/null
+++ "b/deduped/dedup_0532.jsonl"
@@ -0,0 +1,595 @@
+{"text": "Pencil-and-paper examination formats, and specifically the standard, five-option multiple-choice question, have often been questioned as a means for assessing higher-order clinical reasoning or problem solving. This study firstly investigated whether two paper formats with differing number of alternatives (standard five-option and extended-matching questions) can test problem-solving abilities. Secondly, the impact of the alternatives number on psychometrics and problem-solving strategies was examined.Think-aloud protocols were collected to determine the problem-solving strategy used by experts and non-experts in answering Gastroenterology questions, across the two pencil-and-paper formats.The two formats demonstrated equal ability in testing problem-solving abilities, while the number of alternatives did not significantly impact psychometrics or problem-solving strategies utilized.These results support the notion that well-constructed multiple-choice questions can in fact test higher order clinical reasoning. Furthermore, it can be concluded that in testing clinical reasoning, the question stem, or content, remains more important than the number of alternatives. The assessment of problem-solving skills, and specifically diagnostic skills, was once reserved for examination formats such as free-response questions, patient management problems (PMPs) or oral examinations. These evaluation methods, however, are all resource-intensive, thus making it difficult to provide the representative sampling of problems necessary to circumvent the problem of case specificity, which predicts that success in solving one clinical presentation does not predict success in another . As a coPrevious literature has demonstrated that altering item stems tends to determine clinical challenge, while psychometric properties such as discrimination and difficulty tend to be affected by the number of answer options , hereby The first examination format studied is the five-option MCQ (see Appendix A for example). Although MCQs have always been considered an efficient and reliable testing tool, they have not always been perceived as ideal for the evaluation of higher-order thinking skills such as problem solving. Prevailing perceptions that MCQs assess lower levels of knowledge such as recall of isolated facts, and/or encourage trivialization, do exist in the medical education community . To the The second examination format the EMQ format, initially designed in response to some of the criticisms of the MCQs. EMQs (see Appendix A for example) were introduced in the 1990s in both the NBME and USMLE, amongst others. Case and Swanson  have beeThese studies have focused on psychometrics, whereas potential benefits, and possible reasons for such benefits, of the EMQ format over standard MCQs in eliciting higher order problem solving remain unclear. No study has formally used think-aloud protocols to assess whether a well-written MCQ differs from EMQs in challenging examinees to problem solve. There is little doubt that poorly written MCQs can encourage students to learn isolated facts by rote. In fact, all available evaluation methods potentially yield information on clinical reasoning if the content is appropriate, suggesting that content is more important than question type .The two examination formats will be tested for their ability to elicit the three different diagnostic reasoning strategies generally available to learners: hypothetico-deductive reasoning, pattern recognition, and scheme-inductive reasoning. Deductive reasoning (hypothetico-deductive)  is a \"toPattern recognition has been identified by other research as a very successful approach used by experts to solve clinical problems -19. BefoThe third strategy is scheme-inductive reasoning. \"Schemes\" are defined as a mental categorization of knowledge that includes a particular organized way of understanding and responding to a complex situation. They are drawn on paper like \"inductive trees\" or \"road maps\" to recreate the major divisions (or chunks) used by expert clinicians for both storage of knowledge in memory and its retrieval for solving problems ,20  were designed to be as similar as possible in length, difficulty, and the presence of distracters. The stems differed only in the presence of a few key different pieces of information that led to a different diagnosis. The stems were then randomly assigned to one of the examination formats described above, MCQ or EMQ. The alternatives list included the correct diagnosis, and two plausible 'competing alternatives' to the correct answer.The examination was administered to twenty experts in Gastroenterology in two centers, Calgary (15) and Ottawa (5), as well as twenty non-experts, final-year medical students at the University of Calgary. Candidates were considered experts if they were specialists who spent more than 80% of their clinical time in the practice of Gastroenterology.The subjects were first asked to answer the eight questions. The examinees were not given a time constraint to complete the examination, though most completed it in 45 minutes. After the completion of the eight questions, the subjects, with the examination paper in hand and any written notes made during the examination, were asked to explain how they arrived at each diagnosis. A panel of two judges (experts in the Gastroenterology presentations being tested and in the recognition of the diagnostic reasoning process) interviewed the examinees. With as little prompting as possible, the examinees were asked to think-aloud  and descA 'Process Score' of 3 was assigned if pattern recognition was used. Determination that \"pattern recognition\" was used occurred when the subject directly reached a single diagnosis with only perfunctory attention to the alternatives. A 'Process Score' of 2 was assigned if a well-structured and accurate scheme was predominantly used to guide the inductive inquiry. Determination that a scheme-directed diagnostic reasoning strategy was used occurred by analysis of the verbal discourse using modified propositional analysis . A propoA 'Process Score' of 1 was assigned if the examinee relied on hypothetico-deductive reasoning exclusively or predominantly. It was determined that hypothetico-deductive reasoning was the diagnostic strategy utilized when the subjects analyzed one by one each alternative diagnosis presented with the clinical vignettes prior to selecting the most likely diagnosis.The interviews were audio taped or videotaped for later review. Such reviews were required infrequently, but were found necessary when the two judges identified different reasoning strategies. The most frequent cause for differences in identification of diagnostic reasoning strategy was examinees' use of more than one strategy. For example, the candidate might initiate the diagnostic reasoning process using scheme-inductive inquiry, but resort to deductive reasoning immediately after. Disagreement between the two judges was resolved by discussion until concurrence about the diagnostic reasoning strategy was reached. The final assigned mark reflected the predominant diagnostic reasoning strategy utilized.A dichotomous score  was assigned in order to compute the format psychometric properties.Reliability of the process scores and formats was estimated using Cronbach's alpha coefficient. Item statistics were generated for each item including a discrimination index. Inter-rater reliability of diagnostic reasoning scores was estimated by a Pearson correlation coefficient.A logistic regression analysis was used to determine which of the three independent variables being studied  had an impact on diagnostic reasoning or 'process score' (the dependent variable). Specifically, the analysis will model the odds of using an 'expert' method of problem-solving, that is scheme-inductive or pattern recognition  in relation to the three independent variables of format, expertise and clinical presentation. An expertise effect, which would be expected, will lend evidence of construct validity to the 'process score'. Analysis was carried out using the Stata software system .The two judges found it easy to agree on the broad type of strategy used by the subjects . However, there was less agreement when the same subject used more than one diagnostic strategy. The initial diagnostic reasoning scores resulted in an agreement between the two judges of 0.84.Both formats demonstrated quite acceptable reliability and discrimination, as per Table The results of the logistic regression analysis are as follows, in Table Table The present study had two major goals. The first was to determine whether the two pencil-and-paper formats studies, the MCQ and EMQ, could in fact assess problem-solving skills. In Table The first research question of this paper was to investigate whether the examination formats used in this study, the standard five-option Multiple-choice and Extended-matching questions, were capable of testing problem-solving abilities. The observation from the data is that the two formats can potentially evoke more 'expert' methods of diagnostic reasoning processes such as scheme utilization or pattern recognition. Table In regards to the second main research question, the two question formats, with different number of alternatives, did not exert an independent effect on diagnostic reasoning strategy, as shown in the logistic regression analysis Table . ShortenWhile the findings presented do support ongoing use of the MCQ format, there is no denying that the EMQ format has demonstrated superior psychometric properties over the MCQ format in a number of studies mentioned earlier in this paper. Furthermore, in our own study, several non-expert and some expert examinees did comment that Extended-matching questions made it more difficult to go through the list of alternatives prior to answering the question. For examinees relying on hypothetico-deductive reasoning, the Extended-matching format, because of the inherent difficulty of reading through an extended alternatives list, may, at least subjectively, provide a better challenge than the Multiple-choice format.A significant limitation to the study is the manner in which the cognitive problem-solving process selected by the subjects was ascertained. Thinking aloud was used. After the completion of the examination, subjects were asked to verbally report their thinking method to two judges. The two judges independently noted the cognitive problem solving process the subjects had used in arriving at a diagnosis. Although agreement between the two judges on the process selected was identical in more than 85% of the think-aloud interviews, in the remaining 15% there was disagreement. The cognitive process was then decided by reviewing audiotapes and videotapes, so that 100% agreement could result. In other words, consensus and not initial judgements were used.This is the first study that has used this type of think-aloud analysis to directly assess the ability of pencil-and-paper examination formats to test higher order problem solving. The results failed to show a significant difference between the two formats used, but did show that both formats can potentially evoke higher order diagnostic thinking. The results have several potential implications for medical education. Firstly, the results are important to examination construction, by demonstrating direct evidence that problem solving can be tested by pencil-and-paper formats, and specifically change some of the presented misperceptions about the standard five-option MCQ format. Secondly, demonstrating that the two formats can evoke scheme utilization is important. There is evidence  that theLastly, this study demonstrates that testing higher order problem solving requires careful attention to question stem rather than question format or number of alternatives. A well-constructed stem will challenge examinees to choose the correct response, potentially using more expert reasoning strategies, prior to examining the alternatives. This has great potential impact on examination writers, who need not feel obliged to provide more than five alternatives, once they have carefully constructed a stem with a problem-solving task in mind.The author(s) declare that they have no competing interests.SC conceived of the study, participated in its design and coordination, and drafted the final manuscript. HM participated in the study conception, design, and revised the initial manuscript draft. PH participated in the study design and performed the statistical analysis. GF participated in the statistical analyses. All authors read and approved the final manuscript.A 35 year-old woman presents with a one year history of diarrhea. She describes her stools are 10 \u2013 12 profuse, watery, non-bloody bowel movements per day. She is eating well but has lost 7 kg over the last year. She has no abdominal pain. She is unsure if her stools are oily, but they are difficult to flush. She is otherwise perfectly well, with no previous surgeries. She smokes 1/2 pack a day but does not drink alcohol. She has never traveled, camped or drank well water. Her family history reveals an aunt with ulcerative colitis. Examination is unremarkable except for pallor. Stool C & S, O & P and C. difficile are all negative. Laboratory work shows a microcytic anemia , with low ferritin (4), but normal B12 and folate levels.1) What is the most likely diagnosis for this patient?A) Celiac diseaseB) Crohn's colitisC) Villous adenoma of rectumD) Pancreatic insufficiencyE) Bacterial overgrowth ANS:__________A 33 year-old woman presents with a one year history of diarrhea. She describes her stools as 10 \u2013 12 profuse, watery, non-bloody bowel movements per day. She is eating well, but has lost 9 kg over the last year. She has no abdominal pain. She sometimes sees oil droplets in her stool, and they are very difficult to flush. She had surgery for stomach ulcers at age 20, and had repeat surgery five years later for \"bile gastritis\". She is otherwise healthy. She smokes 1/2 pack per day but does not drink alcohol. She has not drank well water, and has not traveled or gone camping recently. Her family history is significant for two cousins with Crohn's disease. Examination is unremarkable. Stool C & S, O & P and C. difficile are all negative. Her CBC shows a macrocytic anemia  with a normal ferritin, but low B12 and elevated folate levels.Select the most likely diagnosis from the list below: __________________A) Bacterial overgrowthB) Celiac diseaseC) Collagenous colitisD) Crohn's colitisE) Crohn's ileitisF) Colonic carcinomaG) Factitious diarrheaH) GiardiasisI) Ischemic colitisJ) Irritable bowel syndromeK) Lactose intoleranceL) Pancreatic insufficiencyM) Shigella dysenteryN) Villous adenoma of rectumO) Viral gastroenteritisThe pre-publication history for this paper can be accessed here:"}
+{"text": "A 24-y-old woman was admitted to the emergency department having had a generalized seizure . She was on the sixth day of treatment with 300 mg daily of slow-release bupropion (Zyban SR) as an aid to smoking cessation. She had a past medical history of tonsillectomy and hay fever, for which she was taking budesonide nasal drops . She was on no other medication. There was no history of head trauma, liver disease, or alcohol withdrawal. Clinical examination, including neurological examination, was normal. The patient's weight was 48 kg. Her blood pressure was 130/80 mm Hg. Electrocardiogram showed a sinus tachycardia at 102 beats per minute. Radiography of the skull and a computed tomography scan of the brain without contrast were both normal. The patient's blood glucose, urea, electrolytes, and liver function tests were all normal. Her serum calcium was 2.01 mmol/l  and her hemoglobin was 116 g/l . The bupropion was discontinued, and the patient recovered without any further seizures or other neurological sequelae. Bupropion recently came onto the Mauritian market as an aid to smoking cessation. This case report is a useful reminder to clinicians of the risks of taking the drug Originally developed as an antidepressant, bupropion has more recently been licensed in many countries as an aid to smoking cessation. It came onto the Mauritian market as a smoking cessation aid in November 2003.www.bnf.org) recommends starting the drug one to two weeks before the target smoking stop date, initially at a dose of 150 mg daily for 6 d and then 150 mg twice daily. The maximum period of treatment is 7\u20139 wk; treatment should be discontinued if abstinence is not achieved by 7 wk.The British National Formulary .Bupropion is associated with a dose-related risk of seizure. The Medicines Control Agency states that the incidence of seizures is one in 1,000 based on doses up to the maximum recommended daily dose of 300 mg per day . The seiUp to 24 July 2002, in the UK there were 184 reports of seizures suspected as being associated with the use of bupropion [In the case we have presented, the patient had no current or previous history of seizure disorders, bulimia, or anorexia nervosa. She was on the sixth day of treatment with bupropion, taking 300 mg per day in two separate doses. She had no other predisposing risk factors for seizure. In particular she was taking no prescription medications, or over-the-counter medications (such as those containing ephedrine products) that are known to lower the seizure threshold. While systemic steroids can lower the seizure threshold, systemic absorption of the nasal steroid drops the patient was taking is an extremely unlikely cause for her seizure (she was not taking a high dose of high-strength drops). Her weight (48 kg) may have been an important factor, since bupropion reaches higher plasma levels in smaller individuals\u2014indeed clinical trials of the drug have generally excluded patients weighing under 100 lb (about 45 kg) .This case report is a useful reminder to clinicians that bupropion is associated with a dose-dependent risk of seizures.In about half of the reports of seizure associated with bupropion, there was a past history of seizures and/or risk factors for their occurrence.It is extremely important to adequately assess seizure risk before prescribing bupropion.Patients should be made aware of the possible adverse effects of the drug."}
+{"text": "Plasmodium falciparum malaria in Sudan has been in process of change since 2003. Preceding the change, this study aimed to determine which artemisinin-based combination therapies is more effective to treat uncomplicated malaria in Malakal, Upper Nile, Sudan.The treatment for P. falciparum infections in children aged 6\u201359 months, in a period of 42 days after treatment.Clinical trial to assess the efficacy of 2 antimalarial therapies to treat A total of 269 children were followed up to 42 days. Artesunate plus Sulfadoxine/Pyrimethamine (AS+SP) and Artesunate plus Amodiaquine (AS+AQ) were both found to be efficacious in curing malaria infections by rapid elimination of parasites and clearance of fever, in preventing recrudescence and suppressing gametocytaemia. The combination of AS+SP appeared slightly more efficacious than AS+AQ, with 4.4% (4/116) versus 15% (17/113) of patients returning with malaria during the 6-week period after treatment . PCR analysis identified only one recrudescence which, together with one other early treatment failure, gave efficacy rates of 99.0% for AS+AQ (96/97) and 99.1% for AS+SP (112/113). However, PCR results were incomplete and assuming part of the indeterminate samples were recrudescent infections leads to an estimated efficacy ranging 97\u201398% for AS+SP and 88\u201395% for AS+AQ.These results lead to the recommendation of ACT, and specifically AS+SP, for the treatment of uncomplicated falciparum malaria in this area of Sudan. When implemented, ACT efficacy should be monitored in sentinel sites representing different areas of the country. Plasmodium falciparum resistance against the two most commonly used antimalarials: chloroquine (CQ) and sulfadoxine/pyrimethamine (SP). CQ resistance in the northern and central part of the Sudan is near 50% [in vivo studies documenting of 0\u201311% resistance in Khartoum and the eastern part of the country [The health situation in Sudan continues to be affected by long-lasting conflict and related humanitarian emergencies such as food crises and epidemics. Malaria is one of the major causes of morbidity and mortality. In Sudan, an estimated 7.5 million patients suffer from malaria each year and 35,000 die from this disease, which accounts for up to 20% of hospital deaths . The pronear 50% , but wasnear 50% . Resista country  and vary country -6. AmodiP. falciparum malaria in Sudan at the time of this study was still CQ as first-line and SP as second-line. A change in protocol was however in progress, and the Sudanese Health authorities had indicated that artemisinin-based combination therapy (ACT) was the preferred new first-line treatment [The nationally recommended treatment protocol for reatment . The Worreatment ,11.in vivo therapeutic efficacy of the first two combinations were studied in Malakal town, Upper Nile State.In line with recommendations for several other African countries, two ACTs were proposed for Sudan: Artesunate (AS) +SP or AS+AQ. The third option, used in other African countries, Artemether-Lumefantrine (Coartem) was not seen as a viable option at the time, because of its cost. To provide information on the efficacy of ACT treatment in Sudan, the P. falciparum infection with a density of 2,000 to 200,000 parasites/\u03bcl but no signs of severe malaria , which is an area is of medium to high malaria endemicity (main transmission season: June-November). This open-label study was done in Malakal paediatric hospital supported by M\u00e9decins Sans Fronti\u00e8res, from September 2003 to January 2004. The methodology used was in accordance with standard WHO procedures for antimalarial drug efficacy assessment in high transmission settings ,13. ChilThe sample size of at least 116 inclusions per arm was calculated to detect a 12% difference between the two arms, assuming a 97% efficacy at day 28 for the AS+SP arm  and 85%msp1 and msp2 loci was used to distinguish re-infections from recrudescences [Slides were examined by two microscopists independently and 20% of slides were cross-checked in an external laboratory . The density of parasites was determined by simultaneous count of white blood cells and parasites, assuming a standard density of 8,000 WBCs per \u03bcl . Haemoglescences ,15. In cThe research protocol was reviewed and approved by the Sudan Research Directorate, the Malaria, Leishmaniasis and Schistosomiasis Directorate, as well as by the Ethical Review Board of MSF . During recruitment of the children in study, informed written consent was obtained from their parents/guardians.A total of 269 children were recruited during the 5-month study period. Of these, 134 were treated with AS+AQ and 135 with AS+SP. Baseline characteristics were similar in both treatment groups .Both artemisinin-combitherapies were highly effective to treat up Table . At day PCR analysis was unable to generate results for a large part of cases  parasitaemia in the AS+AQ and AS+SP groups, respectively. Gametocytes were found in 20% (52/262) of children on inclusion. During follow-up, 78% of these carriers at enrolment showed gametocytes again on one or more follow-up days (32/41), whereas 22% of the children without gametocytes at inclusion were later found to be gametocytemic (42/192). There was no difference between the age, temperature, parasitemia, haemoglobin on admission of the two groups, neither in rates of ACPR at study-endpoint. Gametocyte carriage was similar in both treatment groups and all gametocytes disappeared gradually from the blood during the first weeks after follow-up, and cleared by day 21 in 95% of cases  at day 1 and day 2 was 86% and 97% in the AS + AQ group and 83% and 93% in the AS+SP, and at day 3 all but one (on AS+SP) were free from fever. Average haemoglobin levels for the AS+AQ and AS+SP treatment groups increased from under 8.0 g/dl to near 10 g/dl 6 weeks after treatment. The proportion of children classified as moderately anaemic (haemoglobin from 5 to 8 g/dl) reduced from 51% to 11% during follow-up period, similar in both arms. No serious adverse events were observed or reported on routine clinical examination during follow-up and none of the failures developed complicated malaria.This is one of the first studies into the clinical outcomes of ACT combination therapy in Sudan and provides useful information for decision-makers working to ensure effective antimalarial protocols in this part of the country.P. falciparum malaria in this area of the Sudan. AS+SP appears to be the better treatment option on the basis of non-PCR corrected responses, showing a lower percentage of patients returning with parasitaemia. The PCR analysis indicates the true efficacy is comparable between both treatments (near 99%), but is, however, limited by a high proportion of indeterminate cases. More realistically, an efficacy between 97\u201398% can be expected for AS+SP and 88\u201395% for AS+AQ, acceptable levels after the long follow-up of 42 days after treatment. A rapid parasite clearance and fever reduction was found following treatment with both ACTs. The rise in haemoglobin values and the reduction of the proportion of (moderately) anaemic children after treatment confirms that the malaria parasites were effectively removed from the blood and red blood cell levels rose after treatment.Both of the artemisinin-based combination therapies tested here were found to be highly efficacious in the treatment of uncomplicated The ACTs tested also had an effect on gametocytes. In general, the gametocidal action of AS appears to work through preventing the development of new gametocytes rather than clearance of existing ones . In our The main limitations of the study were that the number of patient exclusions were higher than anticipated (15%) due to concomitant febrile illnesses and loss to follow-up, as well as the lack of results for PCR-analysis, caused by samples missing or an inability to provide sufficient DNA on amplification due to low densities in post-treatment samples. Increasing the number of inclusions compensated losses to follow-up. Repeated attempts at PCR-analysis for problematic samples were only partly successful and, therefore, lead to extrapolation about the findings for the missing samples.P. falciparum and SP has shown high efficacy in various areas of northern Sudan [At time of writing, a change of national protocols towards ACT in northern Sudan is in preparation \u2013 coordinated among health authorities, NGOs and other relevant actors. The first line treatment recommended country-wide is AS+SP, based on reported high efficacy of SP. Northern Sudan is the only country in Africa, which has chosen the option AS+SP . On the rn Sudan ,4,23. Itrn Sudan . The useAn advantage of AS+SP over AS+AQ is that it is more convenient to administer, as SP is given as a single dose and can be administered under observation in a health facility, whereas AQ requires 3 days to complete a course. SP tablets are also easier to take (AQ has a bitter taste). Blister packs of AS+SP, which combine the two drugs and clearly indicate daily tablets to be taken, are currently available for different age categories in Sudan. AS+SP is cheaper than AS+AQ.Implementation of the new national protocol with AS+SP will hopefully take place as quickly as possible to prevent rising morbidity and mortality. Vulnerable displaced populations in epidemic-prone areas and areas of high perennial malaria transmission should be prioritized. Introduction of ACT will have to go hand in hand with laboratory confirmed diagnosis (microscopy or rapid diagnostic tests) to prevent unnecessary use of valuable drugs (thus minimizing drug pressure) and ensure that non-malarial cases are appropriately treated. The change in guidelines should also filter through all health service providers, including the private sector and drug vendors to decrease the potential risk of SP monotherapy and incomplete dosages. The change will initially require more funds to be made available for malaria treatment. The international community \u2013 including many NGOs, the WHO, donors, and the Global Fund \u2013 has shown willingness to support countries to change antimalarial protocols ,25.At present, MSF has already started to pilot AS+SP treatment in its project areas in Northern Sudan, i.e. Darfur, Upper Nile and Gedaref, on behalf of the Ministry of Health. The implementation of treatment as well as the future efficacy of AS+SP should be monitored carefully in a number of dispersed sentinel sites, as there is a possibility that SP resistance may further rise before the combination has been made available countrywide.IB was responsible for overall supervision, data analysis and writing of the paper, RA for field-implementation, communication as well as data analysis and writing, MB for preparation and set-up of study, support and advocacy. PH was responsible for microscopy laboratory work and quality. EA participated as field supervisor of clinical work and data gathering. EBH and MAF were responsible for day-to-day study procedures and supervision of clinical teams. JM did the PCR analysis and FC was involved in design of study, data analysis and writing of manuscript. All authors read and approved the final manuscript."}
+{"text": "Artesunate-amodiaquine combination for the treatment of childhood malaria is one of the artemisinin combination therapies (ACTs) recommended by National authorities in many African countries today. Effectiveness data on this combination in young children is scarce.The effectiveness of three daily doses of artesunate plus amodiaquine combination given unsupervised (n = 32), compared with the efficacy when given under full supervision (n = 29) to children with falciparum malaria were assessed in an unrandomized study.61 patients analysed revealed a PCR-corrected day-28 cure rate of 86 %  in the supervised group and 63 %  in the unsupervised group. The difference in outcome between both groups was statistically significant (p = 0.04). No severe adverse events were reported.The effectiveness of this short course regimen in young children with falciparum malaria could be augmented by increased adherence and improved formulation. Plasmodium falciparum strains [P.falciparum have started instituting the ACTs as first line malaria treatment [Antimalarial chemotherapy has been the primary option in the fight against malaria. However, the burden of this disease is still very heavy partly due to the development of multi-drug resistant  strains -4. The m strains . Africanreatment ,7.Artesunate plus amodiaquine combination is one ACT recommended by the World Health Organization (WHO) for use in malaria control programmes and a first line treatment for African children with uncomplicated malaria . This reP. falciparum malaria in Gabonese children was 94% [Artesunate and amodiaquine are generally safe and well tolerated when used as treatment for malaria -18. In a was 94% , but theThere have been very few effectiveness studies on ACTs. Depoortere et al.  found thP. falciparum malaria was assessed.In the present study, we assessed the effectiveness of three-day artesunate plus amodiaquine combination administered unsupervised, compared with the efficacy when given under supervision to children with P. falciparum and the entomological inoculation rate is about 50 infective bites per person per year [This study was conducted a study in Lambar\u00e9n\u00e9 from July 2004 to June 2005 on a cohort of children participating in an ongoing trial at the Medical Research Unit of the Albert Schweitzer Hospital (HAS), Gabon. Lambar\u00e9n\u00e9 is a small town of approximately 30,000 inhabitants located near the equator in the Central African rainforest belt. Malaria transmission is moderate and perennial. 95% of all malaria infections are caused by per year ,23.The study subjects are a subgroup of participants in a prospective Intermittent Preventive Treatment intervention (IPTi) trial ,25 \u2013 admP. falciparum with a rectal temperature of at least 38.5 \u00b0C or a history of fever in the last 48 hours. Written or documented oral consent of the parents or the guardians of the children were obtained at enrolment into the main study, usually after birth in the maternity ward of the Albert Schweitzer Hospital or the Public Regional Hospital in Lambar\u00e9n\u00e9. Ethical clearance was obtained from the Ethics Committee of the International Foundation of the Albert Schweitzer Hospital in Lambar\u00e9n\u00e9.All participants of the IPTi trial were included who had uncomplicated malaria, defined as the presence of asexual parasitaemia of Two groups of children were compared; those that received artesunate plus amodiaquine combination under supervision and those who received the combination unsupervised. Assignment of the two groups was not randomized. According to the IPTi protocol, all study subjects with falciparum malaria were treated with three daily doses of artesunate plus amodiaquine combination from July 2004. The first dose was administered at the Medical Research Unit and the second and third doses at home. This treatment was classified as unsupervised. Then, from December 2004 all patients received all three doses of artesunate-amodiaquine combination under supervision at the Medical Research Unit. This treatment was classified as supervised.All cases that had (i) a mixed infection or (ii) received a sufficient dose of another antimalarial drug a week prior or during the 28-day post-malaria follow-up period or (iii) taken the IPTi study medication a week prior or during the 28-day post-malaria follow-up period were excluded from intention to treat (ITT) analysis. If they vomited the artesunate plus amodiaquine combination twice, they were excluded from the evaluability analysis.Treatment consisted of a daily oral dose of artesunate 4 mg/kg body weight plus amodiaquine 10 mg/kg body weight given for three days (ARSUCAM\u2122 provided by Sanofi Synth\u00e9labo). Artesunate and amodiaquine were supplied in tablets which were crushed then mixed with sugar into syrup and given orally. A full dose of artesunate-amodiaquine combination was re-administered after 30 minutes if the child either spat the medication out or vomited within one hour.The day of treatment was considered as day 0. Patients were invited for follow up visits on days 2 and 28. A clinical and laboratory assessment was done on these follow up visits. These included a thick blood smear and a full blood count. The Giemsa-stained thick smears which were read by at least two experienced microscopists. Parasitaemia was quantified (number/\u03bcL) by the Lambar\u00e9n\u00e9 method . A smearWe advised the parent or guardian to administer doses as seen or practiced with the first dose, and to report back to the hospital if the child vomited or refused to take the medication. We also encouraged the parents or guardians to return back to the hospital at any time in case the child's health appeared to deteriorate.Our primary outcome was defined as a parasitological cure on day 28. Failure was defined as persistent parasitaemia or reappearance of parasites during the follow-up period of 28 days. Our secondary outcome was the safety of the artesunate-amodiaquine combination drug measured as the frequency of adverse events. These were defined as any signs or symptoms or any abnormal laboratory value not present on day 0 or becoming worse during follow up and were judged by the clinicians in the study with respect to severity and relationship to study drug.We measured the haemoglobin (Hb), white blood cell count (WBC) and neutrophils count on days 0 and 28 with an automated analyser .Filter-paper blots  were taken on day 0 and on the day of recurrent parasitaemia for polymerase chain reaction (PCR). For optimal differentiation between strains, and as described before ,28, we gEfficacy was assessed by evaluability  and ITT (intention-to-treat) analysis. Cure rates were calculated from the number of patients with clinical and parasitological cure by day 28 divided by total number of patients in the ITT population or per protocol population respectively. The according-to-protocol population was defined as children who completed the 3-day regimen of daily artesunate plus amodiaquine and had a day 28-follow up visit (+/- 1 week). There were 61 cases eligible for ATP analysis. The intention-to-treat population was defined as children who took at least the first dose. There were 89 cases eligible for ITT analysis. Based on a previous randomized trial of artesunate plus amodiaquine in our study area , a PCR-cAny data inconsistencies were reconciled, and the data were analyzed with the statistical software JMP 5.0 and Stata 8.2 . A descriptive analysis of both observed and unobserved group was made, and continuous data  between groups by unpaired t-test and categorical variables with a chi-square test were compared.Figure The risk ratio in the ATP analysis indicates that supervised administration of artesunate plus amodiaquine combination to children aged below 30 months have a lower risk of failure to treatment compared to when unsupervised . Supervised administration of artesunate plus amodiaquine combination to children aged below 30 months is also positively associated with an increase in probability of achieving a cure from uncomplicated malaria . This translates into an increase of 27% benefit if administration of this drug is supervised. Equivalently, one more cure with artesunate plus amodiaquine can be achieved for every 4 patients treated under supervision.The median parasitaemia of our cohort was 7,500 parasites/\u03bcL. The mean haemoglobin concentration in the supervised and unsupervised groups on day 0 were 8.7 g/dL and 8.1 g/dL respectively (p = 0.16), and on day 28 were 9.5 g/dL and 8.9 g/dL respectively (p = 0.03). The mean changes were not different between the groups. However, on day 0, thirty-nine patients (64%) had anemia defined as haemoglobin between 5 and 9 g/dL. On day 28, eighteen patients (30%) fell under this definition of anemia. Overall the mean haemoglobin concentration increased from 8.4 g/dL on day 0 to 9.2 g/dL by day 28.There were 36 adverse events reported during the course of this study, the most reported were cough (18%), diarrhoea (12%), vomiting (5.6%), and skin infections (4.5%). There was no serious adverse event or death recorded. In the supervised group, two subjects were treated with the five-day artesunate monotherapy , becauseThis study shows that supervised administration of artesunate plus amodiaquine to Gabonese children aged less than 30 months with uncomplicated falciparum malaria substantially reduces the risk of treatment failure compared to unsupervised administration. The proportion of supervised children cured by day 28 was 86%, while that of unsupervised children was substantially lower (63%). In a previous randomized study carried out in Lambar\u00e9n\u00e9, supervised administration of artesunate plus amodiaquine achieved a PCR corrected cure rate of 94% by day 28 . The difArtesunate plus amodiaquine combination was well tolerated. There were no serious adverse events reported during the course of this study. The weakness of our study was that it was not randomized and the sample size was relatively small. Crushed tablets that are probably harder to administer than palatable syrup were used. There is a clear benefit of supervised compared to an unsupervised treatment shown in the ATP analysis which was less evident in the ITT analysis. Certainly the effectiveness of 63% under unsupervised conditions in this study is unacceptably low. This effectiveness might be higher with a fixed combination better tasting syrup given under supervised conditions.P.falciparum malaria [The therapeutic life of this first line antimalarial recommended for the treatment of uncomplicated  malaria ,31 can b malaria  and part malaria ,33.SO designed the study, collected data and prepared the manuscript.MP designed the study, collected data and prepared the manuscript.NGS contributed in the preparation of the manuscript.JD, ML and COS collected data and participated in the study design.JFK contributed to the molecular genetic studies and contributed to draft the manuscript.BL, PGK and MPG helped in designing the study and in data analysis and interpretation."}
+{"text": "Accurate identification and quantification of malaria parasites are critical for measuring clinical trial outcomes. Positive and negative diagnosis is usually sufficient for the assessment of therapeutic outcome, but vaccine or prophylactic drug trials require measuring density of infection as a primary endpoint. Microscopy is the most established and widely-used technique for quantifying parasite densities in the blood.Results obtained by 24\u201327 expert malaria microscopists, who had independently read 895 slides from 35 donors, were analysed to understand how reader technique contributes to discrepancy in measurements of parasite density over a wide range of densities.Among these 35 donations, standard deviations ranged from 30% to 250% of the mean parasite density and the percent discrepancy was inversely correlated with the mean parasite density. The number of white blood cells indexed and whether parasites were counted in the thick film or thin film were shown to significantly contribute to discrepancy amongst microscopists.Errors in microscopy measurements are not widely appreciated or addressed but have serious consequences for efficacy trials, including possibly abandoning promising vaccine candidates. Microscopy has been used to detect malaria parasites in the blood of infected patients since Laveran first identified the parasites in 1880 [Like all detection methods, microscopy is an imperfect technique. However, unlike other methods, such as PCR and immunochromatographic assays, it relies heavily upon the judgment and experience of the individual user. This was noted at least as early as 1930, when Knowles and White reported on the 'Training and experience of the observer, the personal factor in the diagnosis of malaria' .To understand how reader technique contributes to discrepancies in reporting parasite species and densities, results from 895 slides made from 35 blood donations were analysed. Some of the slides were parasite-positive and some were guaranteed parasite-negative donations. One slide from each donation was sent to each of 27 expert malaria microscopists for evaluation of parasite presence, species and density. Each participant was asked to report the number of sexual and asexual stages of each species present, but was permitted to choose the manner in which slides were read. There were considerable differences in the densities and species reported for each sample. Analysis of these results yields important insights into the sources of discrepancies between readers reported elsewhere [Details of the patient selection, sample collection and preparation, and reference reader selection and participation have been described in detail elsewhere  and are Donors were chosen from among symptomatic patients self-presenting to regional health clinics or involved in Internal Review Board-approved malaria research protocols in Cambodia and Indonesia and consent specific to this study was obtained from each participant prior to drawing blood. Malaria-negative donations were taken from individuals who are natives of non-endemic areas who had not been exposed to risk of malaria in the past two years. Approximately 3 ml of venipuncture blood was collected in an ethylene-diaminetetraacetic acid (EDTA)-filled tube and multiple slides (up to 150), with both a thick and a thin smear, were prepared from each donation within hours of sample collection. Thick films were made by spreading exactly 6 \u03bcl of blood in a circle of 12 mm in diameter. Two microliters of blood were spread using the edge of a clean slide to make a thin film. The thin film was fixed with methanol and the entire slide was stained with Giemsa solution using standard procedures. Negative donations were stained in separate containers which had never been used for positive donations. Only slides from the same donor were stained in the same batch. Slides were coverslipped to preserve them.Slides from 35 donations were sent to 27 reference readers from 13 countries who were considered skilled malaria microscopists by reputation and who accepted the invitation to participate. Each reader received a slide from each of the 35 donations, but each slide was unique (i.e. no reader observed exactly the same slide). Each reader was asked to record the density of asexual and sexual forms of each species observed on the slide. When it was not possible to distinguish between asexual forms in mixed infections, readers often reported the species observed and recorded a total asexual parasite density. No guidelines were given as to how to count the parasites, whether by co-counting white blood cells (WBC) or co-counting red blood cells (RBC), or how many reference RBCs or WBCs should be reported, although the spreadsheet used by each reader to record data included columns for number of WBCs and number of RBCs. The absence of a rigid counting protocol generated a heterogeneity of counting techniques which permitted us to analyse differences between them.6 RBCs per microliter [Results which were reported as percent parasitaemia, percent parasitized RBCs or had no information as to how many RBCs or WBCs were indexed were excluded from this analysis (approximately 2.5% of reads). The number of asexual parasites observed was divided by the number of RBCs or WBCs indexed and multiplied by the standard approximation of 8,000 WBCs per microliter or 4.5 \u00d7 10croliter . ParasitDifferences in results reported by individual readers evaluating different slides from the same donation include differences in individual judgment and technique, and differences which arise due to handling or random distribution of parasites in the blood. Figure To further dissect the source of reader discrepancy at very low density, one microscopist examined the entire thick film of two slides from the same donation. He identified 57 parasites on one slide and 49 parasites on the other, giving densities of 10 and 8 parasites/\u03bcl, respectively. The original results reported for these same two slides by two other independent readers were 80 parasites/\u03bcl and 0 parasites/\u03bcl, indicating that random chance in the selection of fields to examine may play a considerable role in reader discrepancy, particularly at low densities.Readers were permitted to use either the thick film or the thin film and to count either parasites per WBC or parasites per RBC. There were no guidelines given as to how many WBCs or RBCs should be indexed when measuring the parasite density. Some readers indexed as many as 8,600 WBCs and 150,000 RBCs or as few as 3 WBCs and 400 RBCs . 10.4% of all positive results were reported as parasites per RBC.In order to understand how the differences in the numbers of reference blood cells counted contributed to overall discrepancy among readers, the first analysis used only counts made from the thick film (WBCs) and excluded counts made from the thin film (RBCs). The difference between the individual reads and the group mean for a donation was scaled by the group mean . This scaled residual was plotted versus the mean density for every measurement of every donation Figure . For alli.e. that parasite densities reported as parasites per WBC were always counted in the thick film. There were nine donations for which reads (at least two) were reported from both the thick and thin film. These donations had a higher mean density than donations for which all microscopists counted the thick film , indicating that microscopists were more likely to choose to count parasites on the thin film as parasite density increased. There were no donations for which all the microscopists counted parasites in the thin film. The mean density among measurements taken from the thick film were compared to those taken from the thin film. Figure Next, the parasite densities reported based on readings from the thick film were compared to those from the thin film. RBCs are lysed during staining of the thick film; therefore, parasite densities reported against a reference number of RBCs could only have been counted in the thin film. The inverse was also assumed to be true, As demonstrated above, the precision or agreement of the density measurements is strongly dependent on the number of WBCs counted. To determine how uniform the distribution of WBCs is from field to field in the thick film, one microscopist counted the parasites per field and the WBCs per field over 30 high-powered fields of a single thick film. He found a total of 424 WBCs and 8,897 parasites in 30 fields. Figure There is a critical and long-standing need for tools to standardize the training and evaluation of malaria microscopists. Slide sets consisting of donations with three species individually or in mixed infections and with a wide range of densities were produced to serve as such a tool . BecauseDiscrepancy among readers decreased with increasing parasite density. This is in agreement with an analysis of field data from Peru and Thailand  and fromThe observation that the deviation from the mean decreased as the number of WBCs indexed by the reader increased is logical. In general, as the number of measurements in any experimental system increases, the sample mean approaches the true mean or the expectation of the mean. Since the parasite density was evaluated as a function of the number of parasites per WBC, the estimate of the true number of parasites per WBC improves as the number of measurements (i.e. WBCs) increases. The validity of this technique is limited by the implicit assumption that WBCs are evenly distributed in the film. To test this assumption, the number of parasites and the number of WBCs per field were counted in a single thick film. Over 30 fields, the WBCs were much less uniformly distributed than the parasites. This leads to the conclusion that, even if the readers identified every true parasite without error, the variation in the distribution of WBCs from field to field could significantly contribute to discrepancy among readers and errors in estimating true parasite density.Comparing parasite densities observed in thick films to those observed in thin films confirmed the finding of Dowling and Shute  and otheThis analysis of differences in reader technique suggests several important considerations when attempting to reliably estimate parasite density from blood films. To improve precision, or agreement among readers, a uniform counting protocol should be applied. Increasing the number of indexed cells (WBCs or RBCs) also improves agreement between readers. However, to improve both precision and accuracy, the use of WBCs could be eliminated in favour of a grid counting or HPF counting method which allows the parasites per volume to be counted directly and avoids the problem of unevenly distributed WBCs and an inaccurate conversion factor ,12. UsinAccurate and precise quantification of malaria parasites is a critical endpoint for many types of intervention trials. In a clinical setting, errors could endanger a patient. Recent studies and unpublished personal experience have shown that false positive and negative rates using microscopy in the field can be as high as 30% . This stThe author(s) declare that they have no competing interests.WPO contributed to the study design and data analysis and prepared the first draft of the manuscript. MB contributed to the design and execution of the study, data analysis, and writing of the manuscript. CW contributed to the design and execution of the study as well as the analysis and intellectual content of the manuscript. SM contributed to the design and execution of the study. JM contributed to the intellectual content of the manuscript. RJ contributed to the study design and analysis. WRP contributed to the study design, data analysis, and drafting of the manuscript. FEM contributed to the study design, data analysis, and drafting of the manuscript.The views and opinions are those of the authors and do not purport to represent those of the U.S. Navy or Department of Defense."}
+{"text": "The studies are conducted annually in order to provide timely and reliable information on the status of the recommended regimens for malaria case management. The findings of the therapeutic efficacy of an artemisinin-based combination therapy of pediatric artemether-lumefantrine  in children under five years of age, weighing less than 10 Kg. The study was conducted at two clinics, one in Chongwe (Lusaka Province) and Chipata (Eastern Province). The 28-day follow-up period was used coupled with PCR genotyping for MSP1 and MSP2 in order to differentiate recrudescence from re-infections for parasites that appeared after Day 14.\u00ae) was found to produce significant gametocyte reduction. The Adequate Clinical and Parasitological Response (ACPR) was found to be 100% .91/111 children enrolled in the study, were successfully followed up. Artemether-lumefantrine .Coartesiane The malaria situation analysis in Zambia shows that malaria is the leading cause of mortality and morbidity, and the disease burden is worse in the biologically vulnerable children, under five years of age, and in pregnant women . The devThe assessment and monitoring of antimalarial therapeutic efficacy is key to the provision of sound evidence for policy decisions making regarding which antimalarials can be adopted for malaria case management. The National Malaria Control Centre has over time developed a consistent system for sentinel site surveillance of the therapeutic efficacy of antimalarial drugs. These studies have played a major role in guiding the country's decision to move from monotherapies to Artemisinin-Based Combination Therapies (ACTs).P. falciparum strains may arise [The use of combination therapy is expected to have a positive impact on malaria transmission by lowering the rates of gametocytaemia after treatment. Artemisinin derivatives are advocated for use in antimalarial combination therapy because they quickly reduce the level of parasitaemia and hence the parasite pool from which resistant ay arise .\u00ae), because there was insufficient safety and efficacy data at the time of the policy change.But, as the new drug policy outlines, SP is the current recommended drug for intermittent presumptive treatment for malaria in pregnancy and for children weighing less than 10 Kg. These two groups were excluded from using the tablet formulation of artemether-lumefantrine . Other criteria used for inclusion were: absence of severe malnutrition, mono-infection with P. falciparum , with a parasitaemia in the range of 2,000 to 200,000 asexual parasites per micro litre of blood, absence of severe malaria or danger signs, axillary temperature \u2265 37.5\u00b0C at visit, absence of other febrile conditions, ability to come for follow-up visit and informed consent from parent or guardian.The patients included were children under five years of age, weighing less than 10 Kg  while the second film was for slow staining (30\u201345 minutes in 3% Giemsa stain). The quick stain was used for initial screening for the presence of parasites, while the second blood smear was used to calculate parasite density. Parasitaemia (per microlitre) = number of parasites \u00d7 8,000/number of leucocytes counted.Two laboratory technicians were engaged in the study and both read through all the slides independently to assure quality. All the slides for the patients on the study were collected and used in quality control by an independent laboratory.Assessment of Therapeutic Efficacy of Antimalarial Drugs for Uncomplicated Plasmodium falciparum malaria adapted to the Zambian situation to include studies with artemisinin-based combination therapy. The 28-day follow-up period was used.The study followed the standard World Health Organization (WHO) Protocol for the On enrolment day, Day 0, a brief history of each child was obtained, the patient was weighed and the axillary temperature was recorded. The patient was then sent for laboratory examination to confirm malaria diagnosis and also to identify if parasitaemia was adequate for enrolment. Upon fulfillment of all inclusion criteria, consent was obtained from the caretaker and the child was enrolled on the study.\u00ae is an oral suspension of artemether (180 mg/60 ml) and lumefantrine (1080 mg/60 ml) indicated for the treatment of malaria in children. It is presented in a yellow powder which can be diluted with water to form a 60 ml suspension. The dosing schedule is based on body weight and it is to be taken once a day for three days. The drug is manufactured by Dafra Pharma nv/sa. The drug is registered in Zambia for use in children for the treatment of uncomplicated malaria and is being used in the private sector.CoartesianeAll medication was given under the supervision of the study team. Patients were observed for a few minutes after administering the study drug to ensure that they did not vomit. Any child who vomited had the dose re-administered but if vomiting persisted, the child was dropped from the study and referred for further management. A photograph of the child was taken as an identification tool on follow-up days and also as an incentive upon completion of the study.Follow-ups were scheduled for Days 2, 3, 7, 14, 21 and 28 plus any other day that the child was taken to the clinic on unscheduled day. On each of these days clinical and parasitological assessments were performed. The community health worker ensured the directions to the home of the patient were well written for ease of tracking of patients.Filter paper samples of blood were taken on Day 0 and any other day when the parasites first reappeared in the blood of the patient. Polymerase Chain Reaction (PCR) genotyping for MSP1 and MSP2 was done to differentiate recrudescence from re-infections. DNA was extracted from the filter papers and analysed using standard methods as described elsewhere ,6. A recChildren were withdrawn from the study if the patient developed any signs of severe or complicated malaria or any of the general danger signs during the follow-up period; he/she was given the first dose of parenteral quinine and taken urgently to the appropriate health facility. For ethical reasons, all children with parasitaemia on Day 28, irrespective of symptoms were treated with the alternative antimalarial drug  at the end of the follow-up period.Incentives were given to the caretakers as a way to motivate them not to miss a scheduled visit to the study site. A 1 Kg packet of sugar was given to each patient on Day 3, a bar of laundry soap on Day 7 and a packet of salt on Day 14 and a photograph of the child on Day 28. Any child, who dropped from the study before Day 28, received the photo on the last day of being on the study. Sweets were given to the children as an incentive for the number of finger pricks they get on the study.The data was recorded depending on the day of the study. The screening Log sheet was kept in the laboratory and this had serial numbers and information of all the patients screened for the study irrespective of their parasitaemia and clinical condition. The Case enrolment form was used for the patient's case history and it also contained clinical and parasitological data for Day 0. The case record form was used to record the patients' study number, directions to home, study drug, and all the clinical and parasitological data for Day 0 to Day 28, and the final classification of therapeutic response.Three categories of therapeutic response, namely Early Treatment Failure (ETF), Late Treatment Failure (LTF) and Adequate Clinical and Parasitological Response (ACPR) were used. This is based on the WHO classification system . At all The statistical procedure adapted for the interpretation of the results allowed testing the hypothesis that the proportion of treatment failures was above a certain level in the study area, and, therefore, a decision to change is deemed necessary.Collaboration with the major institutions in the country ensured the quality of study results. The study team members came from the National Malaria Control Centre, Tropical Diseases Research Centre, Macha Malaria Research Institute, University Teaching Hospital and Chainama Hills College of Health Sciences and District Health Management teams (DHMTs).The study was conducted in accordance with the standards of good clinical practice.Ethical clearance was sought from The Tropical Disease Research Centre Institutional Research Committee (#00002911). The study was also cleared by the Central Board of Health Director General's Office and the participating districts. All participants were enrolled after a written consent was obtained from the caretaker. The participants also received incentives to help them during the long follow up days. All information obtained in the study was treated with the confidentiality it deserved.Out of the 111 patients enrolled in the two sites, 91 were successfully followed up. Baseline characteristics of participants are outlined in Table \u00ae was found to have excellent 28-day therapeutic efficacy among children weighing less than 10 Kg. This drug has the potential of providing a viable option for children who are not eligible to take Coartem\u00ae . There is need, therefore, to continue to evaluate the long-term safety profile of this drug in the younger children. The drug had 28-day ACPR rates superior to those of SP (100% vs 76.1%) being recorded in Zambia in children with similar age and weight characteristics.CoartesianeThe study is limited in nature because it was not a randomized clinical trial which would have been ideal. However, an open label was opted for because there is sufficient data on the efficacy of the tablet formulation of artemether-lumefantrine-11.Further studies are needed to estimate the long-term safety and pharmacological action of this pediatric formulation. This will provide more information for countries where the use of this drug may be contemplated. The gametocyte clearance characteristics were similar to those, which have been recorded with the tablet formulation of artemether-lumefantrine ,9. The u\u00ae was very effective in treating uncomplicated P. falciparum malaria in children weighing less than 10 Kg in Zambia.CoartesianeThe author(s) declare that they have no competing interests.PC was responsible for the proposal development, study design, training of research assistants, data analysis and developing of the manuscript. MH was involved in sampling, training of research assistants, field supervision and revision of the manuscript. MK participated in the writing of the manuscript and case management. NS was involved in supervision and manuscript development."}
+{"text": "Plasmodium falciparum to conventional antimalarials, such as sulfadoxine-pyrimethamine (SP); such resistance is widespread in southeast Asia, South America, and Africa. The inappropriate use of antimalarials during the past century has contributed to this increase in resistance. For example, there has been overreliance on quinolines (such as chloroquine) and antifolates (such as pyrimethamine) resulting in cross-resistance among these drug classes. However, in the past decade, a new group of antimalarials\u2014the artemisinin compounds, such as artesunate, artemether, and dihydroartemisinin\u2014have been deployed on an increasingly large scale.There are at least 300 million acute cases of malaria each year globally, resulting in more than a million deaths. Ninety percent of deaths due to malaria occur in Africa, south of the Sahara, mostly in young children. The number of deaths is increasing, and one key factor linked to this has been widespread drug resistance of These compounds produce a very rapid therapeutic response, are active against parasites resistant to multiple drugs, are well tolerated, and reduce gametocyte carriage. To date, no parasite resistance to these compounds has been detected.If used alone, the artemisinins will cure falciparum malaria in seven days, but studies in southeast Asia have shown that combinations of artemisinin compounds with certain synthetic drugs produce high cure rates after just three days of treatment. There is also some evidence that combinations of therapies could greatly retard development of resistance to the partner drug. Although combinations including artemisinins have been widely advocated, they are expensive and relatively untested in highly endemic areas.PLoS Medicine, Adoke Yeka and colleagues compared artemisinin-based compounds and other combination therapies in four districts with varying transmission intensity in Uganda in 2,160 patients aged six months or greater with uncomplicated falciparum malaria. The team tested the combination of chloroquine and SP, currently the first-line therapy in Uganda, the combination of amodiaquine and SP, a cheap regimen proven to be efficacious in previous trials, and the combination of amodiaquine and artesunate.In this month's During the 28-day study they collected data on the efficacy of the different regimens and examined the effect on recrudescence and new infections after therapy. Combined amodiaquine and artesunate was the most efficacious regimen for preventing recrudescence, but this benefit was outweighed by an increased risk of new infection. This result was probably due to artesunate being rapidly eliminated, leaving only amodiaquine to provide post-treatment prophylaxis. Considering all recurrent infections, the combination of amodiaquine and SP was at least as efficacious as the other combinations at all sites and superior at the highest transmission sites.In all, 72% of all recurrent infections were due to new infections, and with the two most efficacious regimens  this proportion was 80%. The identification of new infections stressed the need for other malaria control measures, such as bed nets, said the authors.They also suggested that antimalarials should be judged not just on their impact on recrudescence but also on their impact on the risk of new infections after therapy. Previous studies have suggested that patients who suffer recrudescence have a higher risk of complicated malaria and death. Artemisinins are highly attractive antimalarials, but when used as monotherapy, they have a high risk of recrudescence and hence must be combined with other antimalarials to achieve maximum efficacy. But whether the partner drug should be long or short acting remains unclear, said the authors.Altogether, artemisinin combinations offer great hope for Africa, the authors say, although the ideal combination regimen remains uncertain and cost is a problem. To compare the efficacy of the different therapies, bigger and longer controlled trials are needed in conditions of varied transmission intensity. Nevertheless, based on the results of this study and others, Uganda has chosen a combination of artemether and lumefantrine as its first-line therapy against malaria."}
+{"text": "Plasmodium falciparum resistance to chloroquine and sulphadoxine-pyrimethamine.The therapeutic efficacy of artesunate plus amodiaquine and artemether/lumefantrine were assessed in an area of Nigeria with high levels of P. falciparum infection and parasite density 1,000 to 200,000 parasites/\u03bcL enrolled following informed consent by parents.Children aged 6 to 59 months with uncomplicated Eligible children were randomly assigned to receive either a 3-day course of artesunate (4 mg/kg) plus amodiaquine (10 mg/kg) or 6-dose course of artemether/lumefantrine (20/120 mg tablets) over three days. Patients were followed up with clinical and laboratory assessments until day 14 using standard WHO in-vivo antimalarial drug test protocol.A total 119 eligible children were enrolled but 111 completed the study. Adequate clinical and parasitological response (ACPR) was 47 (87.0%) and 47 (82.5%) for artemether-lumefantrine (AL) and artesunate+amodiaquine (AAMQ) respectively . Early treatment failure (ETF) occurred in one participant (1.8%) treated with AAQ but in none of those with AL. Two (3.7%) patients in the AL group and none in the AAQ group had late clinical failure. Late parasitological failure was observed in 9 (15.8) and 5 (9.3%) of patients treated with AAQ and AL respectively. None of participants had a serious adverse event.Artemether-lumenfantrine and artesunate plus amodiaquine have high and comparable cure rates and tolerability among under-five children in Calabar, Nigeria. Plasmodium falciparum resistance to common affordable antimalarial drugs, chloroquine and sulphadoxine-pyrimethamine has reached very high levels, and noticably hampered malaria control efforts in the region [of P. falciparum resistance in all parts of the country [Malaria accounts for more than one million deaths of mostly African children yearly. Early detection and prompt treatment is a key component of the global strategy for malaria control . Across e region . In Nige country -5.P. falciparum is a problem [The World Health Organization recommends that combination treatment rather than monotherapy should be used in areas where multi-drug resistance to  problem . Combina problem ,8. Artem problem .P. falciparum infection is a problem [The artemisinin-based combination treatment regimens are more expensive than each drug used singly, but their advantages over monotherapy far outweigh the cost. Artemisinin-based combination treatments are recommended for use as first-line treatment for uncomplicated malaria, even in resource-poor areas where multi-drug resistant  problem . ACTs ha problem ,10.The options under consideration in Nigeria for ACTs are amodiaquine with artesunate, or artemether-lumefantrine. Artemether-lumefantrine has reportedly high cure rates in observational studies, but controlled trials comparing it against other artemisinin containing regimens are few . It has P. falciparum to combinations of artemether/lumefantrine and artesunate plus amodiaquine in treating uncomplicated childhood malaria. This paper describes one of these trials, conducted in the far-south geo-political zone within the Niger Delta region of the country. P. falciparum resistance to chloroquine was first documented in this region in 1987 and the current rate of resistance to chloroquine and sulphadoxine-pyrimethamine (SP) exceeds 80% [The Nigerian government supported randomized controlled trials in the six geopolitical zones of the country to assess the clinical efficacy and parasitological response of eeds 80% ,12.Efiks and Ejagham. The people's main occupations are subsistence farming, fishing and logging. Malaria is holoendemic in the study area with high and perennial transmission, especially in the rainy season (from April to November). The study was conducted in the months of September, October and November, 2004.This study was conducted in Ikot Ansa a rural community within the Calabar Municipal Local Council area of Cross River State, Nigeria. The area has a projected population of 222,100, with 44,420 being under five years of age. This area lies within the tropical rainforest belt of southeastern Nigeria. The annual rainfall is 2,000\u20133,000 mm, within the rainy season lasting from April to October. There are two predominant ethnic groups, Children were enrolled if they had uncomplicated malaria, and fulfilled the following inclusion criteria: residence in study area, age 6\u201359 months, axillary temperature \u226437.5\u00b0C and parasite density of 1,000\u2013200,000 asexual parasites/\u03bcl of blood, and informed consent by parents/guardian. Children with packed cell volume (PCV) <15%, severe malnutrition, other signs of severe malaria or illness including any \"danger sign\"  in blocks of ten (10) to receive either artemether-lumefantrine (AL) or artesunate+amodiaquine. Allocation sequence was concealed in opaque, sealed and serially numbered envelopes. Study treatment was started on the day of randomization (day zero) and completed on day two, while follow-up continued for 14 days. Clinical examination plus body weight and height were recorded on day zero. Follow-up visits were scheduled on days 1, 2, 3, 7 and 14. On each visit, clinical examination including axillary temperature and thick blood film specimen obtained to screen for presence of malaria parasites and density was assessed by light microscopy. Packed cell volume (PCV) was estimated on day zero and repeated on day 14.Artesunate and amodiaquine were given orally at 4 mg/kg and 10 mg/kg body weight respectively, once daily on days 0, 1 and 2 with amodiaquine reduced to 5 mg/kg on day 2. Artemether/lumefantrine was given as 20/120 mg tablets. Participants weighing 5\u201314 kg received one tablet (20/120 mg), two tablets (40/240 mg) for body weight 1524 kg and three tablets for those weighing 25 \u2013 35 kg. Each participant had a total of six doses of AL, with each dose at 12-hourly intervals for 3 days. Treatment was directly supervised by the research nurse and patients observed for 30 minutes. Drugs were re-administered to those who vomited within this period. Tepid sponging, exposure and administration of paracetamol at 15 mg/kg body weight were used to reduce high fever. Participants that did not return on schedule for follow-up were visited at home on the same day.P. falciparum asexual parasitaemia on the thick smear. Parasites were enumerated using thick film as described by Shute [Thick and thin blood smears were prepared and stained in 3% Giemsa solution for 30 minutes. The smears were read to 100 fields with quantification of by Shute . The par2), 't' test for continuous normally distributed variables and nonparametric Kruskal-Wallis was used to compare continuous not normally distributed variables.Data generated were recorded in a log book, and individual patients case record files. Data were entered and analyzed with EPI-Info version 6.4. Data were also exported to SPSS version 11.0 for further analysis. Differences between groups were assessed using chi-square  is to enhance cure rates and delay development of parasite resistance to component drugs . This stThe cure rates obtained for a 6-dose regimen of AL and a 3-day course of AAMQ were high > 80%) but fell short of therapeutic efficacy levels reported in other areas. Day-14 cure rates for AAMQ were 91%, 93%, and 98% in Kenya, Senegal and Gabon, with lower day-28 cure rates of 68%, 82% and 85%, respectively . Another0% but feThe 14-day cure rate obtained for a 6-dose regimen of AL in the present study is comparable with those reported for 4-dose regimen in other trials, but lower than cure rates obtained in other 6-dose trials ,11. A 42This study has demonstrated comparable therapeutic efficacy and tolerability for a 6 dose regimen of artemether-lumefantrine and a 3-day course of artesunate plus amodiaquine. This apparent lack of difference in effects may be because the study samples were small and lacked statistical power to detect significant difference at 95% confidence level.The short follow-up period of 14 days is another methodological limitation of this study. Recent observations have shown that treatment failures affecting artemisininbased combination treatment regimens tend to occur after 21 days . The minThe Nigerian government has reviewed the malaria treatment policy, replacing chloroquine and sulphadoxine-pyrimethamine with artemisinin-based combination treatment (ACT) as the first-line treatment for uncomplicated malaria. This change in policy has been informed by evidence from local studies like the one reported here . Larger Martin Meremikwu, Emmanuel Ezedinachi and Donald Ordu developed the protocol and supervised the trial. All the authors contributed to the preparation of the paper."}
+{"text": "Attention deficit hyperactivity disorder (ADHD) is the most common neurological condition in children. This pilot study evaluated the effects of high-dose eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) supplementation on the isolated plasma phospholipids and behavior in children with ADHD (primarily inattentive subtype and combined subtype).Nine children were initially supplemented with 16.2 g EPA/DHA concentrates per day. The dosage was adjusted dependent on the ratio of arachidonic acid (AA) to EPA in the isolated plasma phospholipids at four weeks to reach a level normally found in the Japanese population.At the end of the eight-week study, supplementation resulted in significant increases in EPA and DHA, as well as a significant reduction in the AA:EPA ratio . A psychiatrist (blind to supplement compliance or dosage modifications) reported significant improvements in behavior . There was also a significant correlation between the reduction in the AA:EPA ratio and global severity of illness scores.The findings of this small pilot study suggest supplementation with high-dose EPA/DHA concentrates may improve behavior in children with ADHD. Attention deficit hyperactivity disorder (ADHD) is a neurological condition characterized by the inability to concentrate in a sustained manner, to pay attention to tasks, and to control impulsive actions . It is eThe findings that children with ADHD have altered PUFA levels led to interventional studies that supplemented with these fatty acids. Hirayama et al.  found thDepression is often a co-morbidity of ADHD and an increased AA:EPA ratio has been shown to positively correlate with severity of depression . High-doThe interventional studies in children with ADHD demonstrate that some behavior may improve with PUFA supplementation ,8,10; hoThis was an eight-week, open-label, proof-of-efficacy pilot study. Nine children aged 8\u201316 were recruited from the patient population under treatment for ADHD-primarily inattentive subtype or ADHD-combined subtype at the Hallowell Center, Sudbury, MA. There were more boys (n = 6) than girls (n = 3). Two-thirds (n = 6) presented with ADHD-combined subtype, and one-third (n = 3) with ADHD-primarily inattentive subtype according to criteria of the Diagnostic Statistical Manual (DSM) IV . Two of At the start of the study, all participants were instructed to consume two tablespoons (30 mL) of a liquid EPA/DHA concentrate (supplied by the Inflammation Research Foundation) providing 16.2 g of LC omega-3 fatty acids (10.8 g EPA and 5.4 g DHA) per day. The EPA/DHA concentrate dosage was adjusted at week four based on the AA:EPA ratio in the isolated plasma phospholipids as follows: if the AA:EPA ratio was below 1.0, the dosage was decreased to 15 ml , if the AA:EPA ratio was between 1.0 and 1.5 the dosage was decreased to 20 ml . Isolated plasma phospholipids are not subject to daily fluctuations in dietary intake and are a reliable marker of fatty acid levels  and corrThe ADHD Symptom Checklist-4 (ADHD SC-4) was used to monitor behavioral changes by the psychiatrist at each meeting. Retrospectively the psychiatrist asked the parent/guardian and child each of the checklist questions and also observed symptoms during the process. This questionnaire categorizes behavior as inattention, hyperactivity, oppositional/defiant, and conduct disorder. There was also a section to monitor medication side effects. Inattention and hyperactivity scores can range from 0 to 27 each. Oppositional/defiant scores range from 0 to 24 and conduct disorder from 0 to 3 .The Clinical Global Impression Scale was used by the psychiatrist to rate participants' severity of illness . The scoSummary statistics are reported as mean \u00b1 SD and medians. Statistical analyses were performed using Stata for Mac . Fatty acids, ADHD SC-4, CPRS, and severity of illness were reported for baseline (week 0), midpoint (week four) and at the conclusion of the study (week eight). The non-parametric Friedman test was used to assess changes over time. If the Friedman test was statistically significant at the 0.05 level, then the Wilcoxon signed rank test was used as a post-hoc test to compare changes from baseline to four and eight weeks. Spearman correlations were used to identify a relationship between changes in the primary fatty acid outcome variable, the AA:EPA ratio, and the primary behavioral outcome variable, severity of illness.Blood was monitored throughout the study to ensure that the AA:EPA ratio was greater than 1.0 because of the potential concern with high-dose EPA/DHA supplementation on prolonged bleeding times; although, a recent study of the Japanese indicated no adverse effects related to EPA intake in patients whose AA:EPA ration was lowered to 0.8 . To mainOverall, at the conclusion of the eight weeks of supplementation, the average EPA and DHA levels in the isolated plasma phospholipids significantly increased by a factor of 9.5 and 2.4 respectively  related to stimulant medication prior to study initiation that continued throughout the study. Sleep disturbance is another know adverse effect of medication for the treatment for ADHD [All categories of the ADHD SC-4 significantly improved by week eight. Specifically, inattention, hyperactivity, oppositional/defiant behavior and conduct disorder all significantly improved over the course of the study Table . The CPRThere was a significant positive correlation of the percent change in AA:EPA ratio with the percent change in severity of illness . However, the AA:EPA ratio did not significantly correlate with the ADHD SC-4 or CPRS categories.Supplementation with high-dose EPA/DHA concentrate resulted in significant modifications of fatty acids, particularly a significant improvement in the AA:EPA ratio in the isolated plasma phospholipids and improvements in behavior assessed by a psychiatrist (blinded to protocol adherence and supplement dosage adjustments) in this small pilot sample of children with ADHD.At baseline fatty acid analysis of the isolated plasma phospholipids from the children in this study were similar to that of previous studies of children with ADHD and thirst/skin symptoms of EFA deficiency ,5,11; hoSupplementation of high-dose EPA/DHA concentrates resulted in marked changes in fatty acid levels of the isolated plasma phospholipids. EPA and DHA levels in the isolated plasma phospholipids were used to monitor compliance. We chose the AA:EPA ratio as an important marker because of it's relationship with depression , as deprAlthough the EPA and DHA supplementation dosages used in this study were high compared to previous studies with children, there was no serious adverse effect except one case of loose stools that was corrected with a lower dose. Young et al.  supplemeStevens et al.  supplemeThis study found a statistically significant improvement in the psychiatrist's report of inattention, hyperactivity, oppositional/defiant behavior and conduct disorder based on the ADHD SC-4 questionnaire. Scores for inattention, hyperactivity and oppositional/defiant behavior continued to improve from week four to eight, even with the EPA/DHA concentrate dosage adjustment. The dosage adjustment, however, did not bring the AA:EPA ratio above 3 suggesting the importance of monitoring fatty acids and the AA:EPA ratio in particular rather than EPA/DHA dosage alone. The severity of illness scale demonstrated a positive improvement from an average of moderately symptomatic to mildly symptomatic. This improvement was similar regardless of medication use or lack there of. The percent change in severity of illness also correlated with percent decrease in the AA:EPA ratio, suggesting a connection between the clinical improvement observed by the psychiatrist and the improvements in the AA:EPA ratio.Data from Stevens et al.  in childThere are a number of limitations to this pilot study and therefore interpretation of results requires caution. The study is limited in that there was no placebo group for reference comparisons as this was a pilot study to determine appropriate dosage for protocol adherence and to maintain AA:EPA levels between 1.5 and 3. Dietary intake was not recorded at baseline or monitored throughout the study; therefore, we are unable to decipher intake of fatty acids from the diet. Also related to diet, we advised the children to eat more fruits and vegetables and consume meals and snacks that are balanced with protein, carbohydrates (preferably fruit and vegetables) and \"healthy\" monounsaturated fats. Advice for following both a \"healthy diet\" and high-dose fish oil supplementation may have been confounding factors. However, the dose-response relationship between percent change in AA:EPA ratio and the reduction in the severity of ADHD suggest the behavioral changes were due to, at least in part, the intake of high-dose EPA/DHA concentrates. The lack of behavioral change or regression to pre-study status in those subjects who were least compliant to supplementation also suggest that behavioral changes were associated with intake of the LC omega-3 fatty acids.EPA/DHA concentrate dosage adjustments themselves can be viewed as a limitation since some, but not all participants' daily intake dosage was modified at week four. The supplement intervention adjustment was based on the AA:EPA ratio, therefore those whose AA:EPA ratio dropped below the goal range was adjusted upward and by using this ratio as our goal, we also avoided most adverse events. The lack of a proper means to monitor supplement intake, such as weight of returned bottles, was also a limitation of this study. However, this was compensated for by use of isolated plasma phospholipids levels as a means to monitor protocol adherence.Although this was a small one-arm study, the results are encouraging as they suggest that high-dose EPA and DHA (up to 16.2 g per day) can be given to children with good adherence. Also, our results concur with trends and significant findings of some, but not all studies of PUFA supplementation in children with ADHD or related symptoms ,10,11. TThe preliminary results found in this pilot study warrant future randomized, placebo-controlled, double blind studies that use participant's AA:EPA ratios to determine EPA/DHA supplementation dosage for adjunct treatment of ADHD in children.BS is a stockholder and president of Zone Labs Inc; HLH is a stockholder and employee of Zone Labs Inc.PJS was involved with the design of the study and carried out all psychological testing. EMH was involved with the design of the study. HLH performed the statistical analysis and drafted the manuscript. BS conceived the study and helped to draft the manuscript. All authors read and approved the final manuscript."}
+{"text": "The usual method of assessing the variability of a measure such as the ankle brachial index (ABI) as a function of different observer groups is to obtain repeated measurements. Because the number of possible observer-subject combinations is impractically large, only a few small studies on inter- and intraobserver variability of ABI measures have been carried out to date. The present study proposes a new and efficient study design. This paper describes the study methodology.Using a partially balanced incomplete block design, six angiologists, six primary-care physicians and six trained medical office assistants performed two ABI measurements each on six individuals from a group of 36 unselected subjects aged 65\u201370 years. Each test subject is measured by one observer from each of the three observer groups, and each observer measures exactly six of the 36 subjects in the group. Each possible combination of two observers occurs exactly once per patient and is not repeated on a second subject. The study involved four groups of 36 subjects (144), plus standbys.The 192 volunteers present at the study day were similar in terms of demographic characteristics and vascular risk factors: mean age 68.6 \u00b1 1.7; mean BMI 29.1 \u00b1 4.6; mean waist-hip ratio 0.92 \u00b1 0.09; active smokers 12%; hypertension 60.9%; hypercholesterolemia 53.4%; diabetic 17.2%. A complete set of ABI measurements (three observers performing two Doppler measurements each) was obtained from 108 subjects. From all other subjects at least one ABI measurement was obtained. The mean ABI was 1.08 (\u00b1 0.13), 15 (7.9%) volunteers had an ABI <0.9, and none had an ABI >1.4, i.e. a ratio that may be associated with increased stiffening of the arterial walls.This is the first large-scale study investigating the components of variability and thus reliability in ABI measurements. The advantage of the new study design introduced here is that only one sixth of the number of theoretically possible measurements is required to obtain information about measurement errors. Bland-Altman plots show that there are only small differences and no systematic bias between the observers from three occupational groups with different training backgrounds. Coronary heart disease and stroke as manifestations of atherosclerosis are among the leading causes of death. Consequently, there has been an intensive search for measures of atherosclerosis burden that would indicate the cardiovascular and overall risk of the individual patient . An earlAn early and reproducible diagnosis of PAD is important for identifying high-risk patients as early as possible. Only a minority of patients exhibit the symptoms of intermittent claudication. Data from the literature suggest that for every patient with intermittent claudication, there are at least three with asymptomatic PAD .The introduction of Doppler sonography has made it possible for general physicians to diagnose asymptomatic PAD by determining the ankle-brachial index (ABI), which represents the ratio of ankle to brachial systolic blood pressure (SBP) ,6. ABI vTo establish ABI determination as a screening procedure for asymptomatic PAD, the interobserver variability  and intraobserver variability  of ABI measures must be known. Of particular interest is how these two types of variability differ depending on the observer's level of specialization. The present study therefore set out to determine inter- and intraobserver variability of ABI measurements by three groups of observers  and with a sufficiently large number of randomly selected subjects, age 65 to 70, free of serious disease and without symptomatic PAD (the target group of ABI screening measurements). In order to avoid having to make very large numbers of measurements , a study design was developed that provides the necessary information with one sixth the number of Doppler measures that would otherwise have to be performed. The paper deals with the study methodology.Volunteers were eligible for inclusion in the study if they were members of a public health insurance plan (in this case BKK Hoechst), between the ages of 65 and 70, resident in Frankfurt-Hoechst, ambulatory without wheelchair or walking aids, able to understand the study and able to provide written informed consent. Exclusion criteria were: need for nursing care of any kind, serious disease such as cancer, any amputation of upper or lower extremities, and a history of stroke with hemiplegia. Figure A total of 382 people (36.0%) responded to the study center in Bochum, of whom 310 were interested in participating and 270 signed the statement of informed consent. To ensure that at least 144 subjects would be available for the ABI measurements on the day of the study (four groups of 36), the first 100 men and the first 100 women to respond to the invitation were asked to attend. As a result there were 14 more participants per group than the minimum of 36 required (stand-by). The four groups were asked to arrive at 1.5- to 2.5-hour intervals. Another 20 subjects whose data were not to be included in the analysis of results were asked to participate in a \"dress rehearsal\" one day before the actual study day. The study was conducted in accordance with the Helsinki Declaration and the Guideline for Good Epidemiological Practice (GEP) issued by the German Working Group on Epidemiology . The stuThe duplicate ABI measurements were performed by six angiologists, six general physicians (GPs) and six medical office assistants with special training in Doppler measurements (MAs). The angiologists  were selected from the \"centers of excellence\" (coordinating and training centers) of the German Epidemiological Trial on Ankle-Brachial Index (getABI) ,12. The On the study day December 12, 2005), all volunteers first underwent a physical examination by an angiologist or GP. This basic examination covered weight and height for calculating BMI , waist and hip circumference for calculating waist-hip ratio, and patient history regarding vascular-related comorbidities .For the ABI repeat measurements, each subject saw exactly one observer from each of the three observer groups , and each observer measured six of the 36 subjects in a group Table . Each poThe primary purpose of the study was the assessment of three sources of variability, namely the true differences in ABI between subjects, the measurement error arising from duplicate measurements of the same subject by the same observer (intraobserver variability), and the additional error arising from measurements by different observers (interobserver variability). Secondary outcome measures were body mass index, waist-hip ratio, and risk factors for atherosclerotic complications .The measurements in the study group began after a ten-minute rest period. Each observer was assigned an aide, who was responsible for guiding the observer to the numbered beds in the correct time sequence, as set out in a predetermined list, and for recording the readings on a standard form. The aide ensured the correct sequence of measurements and the blinding of the observer to his/her previous measurements on the same subject. The observers had no access to the measures. Additionally, controllers verified the accuracy of the measurement sequence and procedure at each bed.Over the course of about 90 minutes, each subject saw three observers, one from each of the occupational groups. Each observer was given a list of six subjects to measure, and always completed all six subjects on the list before returning (approximately 45 minutes later) to the first person on the list to repeat the measurements on the same subjects in the same sequence. The volunteers remained supine between observers, resulting in a resting period of 5\u201310 minutes between each set of readings (by different observers).The Doppler ultrasound device with which the ABI readings were taken was the same one used in the getABI study ,12  with two random factors and their interaction, the subjects (108 levels) and the observers (18 levels). Note that the inclusion of the patient as a factor makes it possible to account for and calculate intra-patient correlation. No repeated measures model was used.The mean variation of the ABI measures in the case of multiple measurements can be broken down into the following four variance components:between different subjects, i.e. variance of the ABI that is medically interpreted.a. Variance of the true ABI values based on mean ABI same subject by the same observer \u2013 the so-called intraobserver variance, assumed to be due to measurement error and physiological variation.b. Variance of ABI measurements when multiple measurements are performed on the same subject by different observers (first component of interobserver variance), due to a systematic measurement bias in the individual observer, e.g. an observer generally measuring higher or lower values than others (independent of the specific subject).c. Variance in ABI measurements when multiple measurements are performed on the same subject by different observers, caused by interaction between subject and observer (second component of interobserver variance), i.e. an observer measuring higher or lower values than others for specific subjects only. For example, some observers may have a harder time measuring obese subjects than do other observers.d. An additional variance of ABI measurements when multiple measurements are performed on the same subject, namely the intraobserver variance b and the two interobserver variance components c and d, one obtains the total variance for multiple measurements on the same subject by different observers:By adding the three variance components of multiple measurements on the Total variance of all ABI values for a single subject = b + c + dSimple addition is permissible when the individual variance components are independent of one another. The standard deviation of the ABI measurements for a single subject is the root of the total variance.In order to evaluate the overall quality of the ABI as a measurement technique for discrimination between different subjects, all four variance components must be considered. An intraclass correlation coefficient (ICC) is normally formed for this purpose. The ICC indicates the proportion (in percent) of the total variance in measurement results between subjects that can be explained by the \"biological\" or real variance between the subjects examined. A high ICC indicates that the measurements can be used to discriminate between individuals. ICC values \u2265 0.75 are said to be acceptable . The ICCThe intraobserver variability of each individual observer can also be determined. By taking the mean of these values for all observers within one occupational group  it is possible to assess the average quality of the measurements by each of the three groups.Bland-Altman plots were used to visualize measurement errors. For each subject, the differences between the first and second measurements were plotted against the mean of the two measures with 95% limits of agreement .All data analyses were performed using SAS\u2122 Statistical Software .Of the 200 subjects invited to participate, 192 (96%) appeared on the study day. The data for the pilot study  were not analyzed. Demographic data for all subjects who appeared on the study day are shown in Table 2, and mean waist-hip ratio of 0.92 (0.09). Active smokers made up 9.3% of the sample and former smokers 40.7%; 58.1% were hypertensive, 54.8% dyslipidemic, and 15.7% diabetic. Only one subject (0.9%) of Groups 1\u20133 reported having had peripheral vascular-widening measures in the past. In Groups 4 and 5 five subjects (5.9%) reported having had such measures.The 108 individuals  for whom complete duplicate measurements were obtained had a mean age of 68.7 (1.5) years, mean BMI of 29.0 (4.3) kg/mABI measurements were performed on 189 of the 192 subjects who enrolled in the study. One subject dropped out after the basic examination, before any ABI determinations were performed, and on two other subjects no ABI measures could be obtained for technical reasons (very obese subjects with upper arm too large for the blood pressure cuff).In Groups 1\u20133 (108 subjects), the ABI measurement program was completed as planned with full sets of duplicate measurements. Because of time restrictions, full sets of measurements were not obtained from all subjects in the fourth group. The analysis was based on the first three groups of subjects  for whom complete measurements were obtained.The results of the ABI measurements are shown in Table There are no significant differences in mean brachial SBP or mean ABI between Groups 1\u20133 and incomplete Group 4  existed on the inter- and intraobserver variability of ABI measures ,17-19. TFrom the fact that the mean differences between the ABI measurements in Figure Previous studies indicate both that the variability in ABI measures is highly dependent on the experience of the observers, and that the value of the ABI measures depends on the apparatus with which the measurements are performed ,17-19. HThe variability in ABI measures is of particular importance for patients moving from one observer to another, for example as the result of changing family doctors (interobserver variance). Moreover, the ABI should always be an average of several measurements. Under ideal measurement conditions, an ABI <0.9 is considered a readily obtained indicator for the possible presence of PAD ,22. The With the partially balanced incomplete block design, it was possible to test every possible observer combination exactly once while performing only 16.7% of all the repeat ABI measurements that would theoretically be required. Thus we established the conditions for assessing components of variance with large numbers of subjects and observers, which can be applied also to measures other than ABI.PAD: peripheral arterial diseaseABI: ankle-brachial index2BMI: body mass index in kg/mWHR: waist-to-hip ratioThe author(s) declare that they have no competing interests.All authors contributed to the manuscript and gave final approval of the article.HGE is principal investigator, responsible for the conception of the study and the study protocol. He participated in analysis and interpretation of data, and drafted the article.CH was responsible for carrying out the study on the study day, gave logistic support, and drafted the article.THL was responsible for the statistical analysis and interpretation of data, and critically revised the manuscript.HJT obtained the funding, provided statistical expertise and gave final approval of the article.The pre-publication history for this paper can be accessed here:"}
+{"text": "The WHO recommends that adults with uncomplicated P. falciparum successfully treated with a blood schizonticide receive a single dose of primaquine (PQ) 45 mg as a gametocytocidal agent. An earlier pilot study suggested that 75 mg of bulaquine (BQ), of which PQ is a major metabolite, may be a useful alternate to PQ.In a randomized, partial blind study, 90 hospitalized adults with Plasmodium falciparum malaria that was blood schizonticide-responsive and a gametocytemia of > 55/\u03bcl within 3 days of diagnosis were randomized to receive single doses of either PQ 45 mg or BQ 75 mg on day 4. We assessed gametocytemia on days 8, 15, 22 and 29 and gametocyte viability as determined by exflagellation (2\u00b0 end point) on day 8.On day 8, 20/31 (65%) primaquine recipients versus 19/59 (32%) bulaquine recipients showed persistence of gametocytes (P = 0.002). At day 15 and beyond, all patients were gametocyte free. On day 8, 16/31 PQ and 7/59 BQ volunteers showed gametocyte viability (p = 0.000065).Plasmodium falciparum gametocytocidal agent and may clear gametocytemia faster than PQ.BQ is a safe, useful alternate to PQ as a  Malaria remains the most important parasitic infection with 300\u2013500 million people affected yearly and 1.5\u20132.7 million deaths each year. World over, malaria control has focused on pharmacological intervention, vector control, curtailing irrational and indiscriminate use of antimalarials, and the development of vaccines. Of these strategies, pharmacological intervention remains the most effective way to combat malaria [Plasmodia that infect humans [P. falciparum. [et al found that the efficacy of PQ 45 mg as a falciparum gametocytocidal agent in patients sensitive to chloroquine in India to be approximately 77% at Day 29 follow up. [8 aminoquinolines like primaquine are unique antimalarials in that they exhibit activity against multiple life cycle stages of t humans . Primaquciparum.  In 1998,llow up.  During tllow up. . BulaquiThe protocol was approved by the institutional ethics committee and the Drugs Controller General of India. The study was conducted between January 2002 and April 2004.Plasmodium falciparum infection, provided written informed consent, and had a gametocyte count > 55/\u03bcl within 72 hours of diagnosis, regardless of asexual parasite counts, were eligible for enrollment. The minimal gametocyte count was chosen based on infectivity to mosquitoes [Plasmodium vivax, claimed an allergy to primaquine or bulaquine, or were G6PD deficient were excluded. On admission, patients were initially assessed by thick and thin blood films stained using the Jaswant Singh and Bhattacharji (JSB) field stain [Patients who were at least 16 years old with uncomplicated squitoes . Patientld stain . SubsequEnrolled patients were admitted to hospital on Day 1 and treated under observation with quinine orally 10 mg/kg/day thrice daily for a total of 7 days and doxycycline 100 mg once daily for 7 days. At day 4, consecutive patients were randomly allocated in a 1:2 fashion to receive an observed single dose of either PQ 45 mg or bulaquine 75 mg, based on a computer generated randomization code. Unequal allocation was used because of earlier studies suggesting the superiority of bulaquine. The test articles were administered on day 4 because the incidence of nausea and vomiting is higher in the first few days of schizonticidal therapy and this was given regardless of parasite clearance. on day 8, all patients were assessed for gametocytemia, discharged, and asked to follow up on days 15, 22, and 29 for further safety and parasitologic checks.Giemsa stained malaria blood smears during hospitalization were done twice a day for the first 72 hours and once a day thereafter until discharge and on the follow-up days. The slide readers were blinded to the treatment.Efficacy was assessed by gametocytemia (primary end point) and gametocyte viability (secondary end point) on admission and all follow up days. The latter was assessed by the modified Shute's technique . This teSafety was monitored by routine clinical hematological and biochemical laboratories and an electrocardiogram on days 1 and 8. Adverse event recording was focused only on nausea, vomiting, and epigastric distress and were recorded only if not reported on Day 1 or if a symptom worsened after Day 1.The estimated sample size was calculated using Casagrande's method based on a previous study by Gogtay et al comparing the two drugs . AssuminA total of 93 male patients were enrolled. Women with malaria are not inclined to get admitted, especially because of hardships related to hospitalization and supervised drug administration. The age of the patients ranged from 16\u201372 years: 31.47 \u00b1 11.62 years). There were three drop outs, two in the Bulaquine arm and one in primaquine arm. These patients did not return for any follow up visit after discharge from the hospital and were omitted from analysis.At admission, gametocytaemia between the 2 groups was similar. At day 8, 20/31 (65%) PQ recipients and 19/59 (32%) BQ recipients had gametocytes on blood smear (p = 0.002). At days 15, 22, and 29, all patients in both treatment groups were free of gametocytes.At day 8, 16/31 (52%) PQ recipients and 7/59 (12%) BQ recipients had viable gametocytes on exflagellation testing (Table)(p = 0.000065). All patients with viable gametocytes had smear positive gametocytemia.There were no important clinical hematology or biochemical laboratory values, and all electrocardiograms were within normal limits.Plasmodium falciparum malaria assessed the efficacy of bulaquine, the parent compound of primaquine. given as a single dose of 75 mg for its gametocytocidal effect. On day 8 of therapy, fewer patients with bulaquine had gametocytes as compared to those who received 45 mg primaquine. This suggests superior efficacy of bulaquine or its ability to clear gametocytes more rapidly, as all patients were free of gametocytes by day 15 and beyond.A single dose of 45 mg primaquine is given along with or after schizonticidal therapy in areas where malaria is endemic as a transmission blocking strategy and currently is the only option available for this indication. The present study carried out in 91 cases of uncomplicated This clearance of gametocytes completely by Day 15 in both groups in the present study contrasts with our previous study with 45 mg primaquine where persistent gametocytemia was seen in 23% patients responsive to chloroquine up to Day 29. [Based on the results of the present study, bulaquine in a single dose of 75 mg may represent yet another treatment option for gametocytocidal effect in addition to primaquine and the current licensing of the drug in the country could be changed to include gametocytocidal effect apart from anti-relapse effect. Whether increasing the dose of primaquine from 45 mg to 60 mg will improve efficacy needs to be addressed in future studies.In this study, the Shute technique, which measures the ability of the male gametocyte to exflagellate was used as a surrogate marker for assessing transmission blocking. Assessment of true gametocytocidal efficacy of any drug will depend upon demonstration of the ability to block transmission to mosquitoes. This in turn can be assessed by only checking for the presence of the oocyst and ookinete in the mosquito midgut , not donIn the first study where we first reported the declining efficacy of primaquine as a gametocytocidal agent, chloroquine versus coartemether was studied, since chloroquine then was (and still remains) the first line therapy for the country.  The resuThe pre-publication history for this paper can be accessed here:"}
+{"text": "Data concerning antimalarial combination treatment for uncomplicated malaria in Madagascar are largely lacking. Randomized clinical trial was designed to assess therapeutic efficacies of chloroquine (CQ), amodiaquine (AQ), sulphadoxine-pyrimethamine (SP), amodiaquine plus sulphadoxine-pyrimethamine combination (AQ+SP) and artesunate plus amodiaquine combination (AQ+AS).287 children between 6 months and 15 years of age, with uncomplicated falciparum malaria, were enrolled in the study. Primary endpoints were the day-14 and day-28 risks of parasitological failure, either unadjusted or adjusted by genotyping.All treatment regimens, except for CQ treatment, gave clinical cure rates above 97% by day-14 and 92% by day-28 (PCR-corrected). AQ+SP was as effective as AQ+AS. The risk of new infection within the month after therapy was generally higher for AQ+AS than AQ+SP.These findings show that the inexpensive and widely available combination AQ+SP may be valuable in for the treatment of uncomplicated malaria in Madagascar and could have an important role in this country, where much of the drugs administered go to patients who do not have malaria. Plasmodium falciparum to the inexpensive and widely used drugs, chloroquine (CQ) and sulphadoxine-pyrimethamine (SP) [Malaria remains one of the most serious health problems worldwide and a leading cause of childhood morbidity and mortality, especially in Africa . At leasine (SP) ,4. As a ine (SP) . Of the ine (SP) . ACTs giine (SP) . MoreoveMalaria treatment in Madagascar is in transition: for over 50 years, malaria was treated successfully using chloroquine (CQ) as first-line treatment. In 2005, the National Malaria Control Programme (NMCP) decided to revise its treatment policy and to replace CQ with artemisinin-based combination therapy  with the support of resources from the Global Fund. This choice was guided by the recommendations of WHO and data from the most recent clinical trial based on the WHO standard protocol : the triFew data is available concerning alternative or combinatory antimalarial treatment in uncomplicated malaria in Madagascar. Randomized clinical trial was designed (i) to update knowledge about the therapeutic efficacies of CQ , AQ and SP alone (used for intermittent preventive treatment in pregnant women), (ii) to compare the respective therapeutic efficacies of two combinatory treatments, AQ+SP and AQ+AS, and (iii) to generate data which could be used as a guide for designing a rational antimalarial treatment policy in Madagascar.Anopheles funestus and the number of infective bites associated with Plasmodium falciparum is estimated at < 10 per person per year. The study protocol was reviewed and approved by the Ethics Committee of the Ministry of Health of Madagascar (N\u00b0007/SANPF/2007).The study was conducted between February and June 2006 during and at the end of the rainy season, in primary health centres of Moramanga and Saharevo in the east foothill areas of the Highlands of Madagascar. In this area, malaria transmission is low and predominantly seasonal. The main vector is P. falciparum at a parasitaemia between 1,000 and 200,000/\u03bcl, (ii) axillary temperature \u2265 37.5\u00b0C, (iii) body weight > 5 kg, (iv) absence of severe malnutrition, (v) absence of febrile conditions caused by diseases other than malaria, (vi) absence of 'danger signs'  and of severe and complicated malaria, (vii) haemoglobin (Hb) \u2265 5 g/dl, and (viii) informed written consent of parents/guardians.Children between six months and 15 years of age presenting at primary health centres of Moramanga and Saharevo were enrolled in the study if they met the following inclusion criteria : (i) monExclusion criteria were: (i) known hypersensitivity to SP, AQ, or AS, (ii) detection, during follow-up of mixed malarial infections, and (iii) development of concomitant disease which would interfere with the classification of treatment outcome.Patients were randomly assigned to receive one of five oral therapies: CQ ; AQ ; SP (25 mg/kg sulphadoxine and 1.25 mg/kg pyrimethamine as a single dose on day 0); AQ + SP or AQ + AS . Randomization was performed in blocks of five, and treatment regimens were allocated by an independent individual not involved in the analysis of the study.All other study personnel were blinded to the treatment assignments, and patients were not informed of their treatment regimen. Patients were directly observed for 30 minutes after treatment, and the dose was readministered if vomiting occurred. Patients who repeatedly vomited their first dose of study medication were excluded from the study.Following enrollment, patients were asked to return for follow-up visits on days 1, 2, 3, 7, 14, 21, 28, and any other day if they felt ill. Blood was obtained by finger prick for thick blood smears and storage on filter paper on all follow-up days. Haemoglobin was determined on day 0 and on day 28. Treatment outcomes were classified according to 2003 WHO guidelines as Early Treatment Failure , Late Clinical Failure , Late Parasitological Failure , and Adequate Clinical and Parasitological Response  .Patients classified as having suffered treatment failure were treated with quinine (10 mg/kg three times daily for 7 days); however, their response to repeat therapy was not assessed. Patients were excluded after enrollment if any of the following occurred: (1) use of antimalarial drugs outside of the study protocol; (2) parasitaemia in the presence of a concomitant febrile illness; (3) withdrawal of consent; (4) loss to follow-up, (5) protocol violation, or (6) death due to a non-malaria illness.\u00a9, Anglholm, Sweden) was used for haemoglobin assays.Blood smears were stained with 4 % Giemsa for 20 min. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 200 WBCs , assuming a WBC count of 8,000/\u03bcl. A smear was considered negative if no parasites were seen after review of 100 fields. Thin blood smears were used to detect non-falciparum infections. Gametocytes were counted against 500 WBC for 287 children at enrolment and in 258, 257, 220, 217 specimens available for that purpose collected on day 7, day 14, day 21 and day 28, respectively. A portable spectrophotometer . If all of the MSP-2 alleles present on the day of failure were present at the time of treatment initiation or on day 1, genotyping was repeated using MSP-1. An outcome was defined as recrudescence if all MSP-1 and MSP-2 alleles present at the time of failure were present at the time of treatment initiation or on day 1, and defined as a new infection otherwise.Molecular genotyping techniques were used to distinguish recrudescence from new infection for all patients failing therapy after day 7. Briefly, filter paper blood samples collected on the day of enrollment, on day 1 and on the day of failure were analyzed for polymorphisms in the genes for merozoite surface protein-1 (MSP-1) and merozoite surface protein-2 (MSP-2) using nested-PCR as previously described . First, \u00a9 software , and analysed using MedCalc\u00a9 software version 9.1.0.1 .Data were entered and verified using EpiInfo 6.04Analysis of treatment outcome was per protocol, which only included patients with treatment outcomes. Frequencies were compared by chi-squared tests and Fisher exact tests, and continuous variables by Student's t-tests, Mann-Whitney U-tests, analysis of variance or Kruskal-Wallis tests as applicable. All reported p-values are two-sided, without adjustment for multiple testing, and were considered statistically significant if less than 0.05.695 patients were screened for inclusion in the study and 287 patients were randomized to receive CQ, AQ, SP, AQ + SP or AT + AQ Figure . The basClinical and parasitological monitoring was complete for 96.1% (276/287) until day 14 of follow-up and for 92.3% (265/287) until day 28. Seventeen patients (5.7%) were lost to follow-up, four patients withdrew consent (1.3%) and one patient (0.3%) had to be withdrawn because he was treated outside the study with drugs active against malaria  and AQ  than with AQ+SP   did not differ significantly between the five groups .Gametocyte prevalence at enrolment was 5.6% (16/287) with a geometric mean density of 124.7 gametocytes/\u03bcl . There were no significant differences at enrolment in gametocyte prevalence or density with respect to treatment regime  or freely distributed at primary public health facilities (Ody Tazomoka\u00ae) [The first step was to update data concerning therapeutic efficacies of CQ, AQ and SP alone, because CQ is has long been widely used in Madagascar and recommended by the NMCP for the home management of presumed malaria in children under five years of age (HMM). Pre-packaged presentations of chloroquine are currently available for children from six to 11 months of age and for children from 12 to 59 months of age and either sold at an affordable price of US $0.025  . SP has \u00a9, Sanofi-Synth\u00e9labo Groupe, Paris, France) from the beginning of 2008.No severe side-effects attributable to study medication were found during the follow-up period in any of the treatment groups. CQ was significantly less effective than SP, AQ, AQ+SP and AQ+AS in treating uncomplicated falciparum malaria, with overall treatment failure of 35.9% within 14 days of follow up and 44.4% within 28 days (PCR-corrected). These data show a higher prevalence of chloroquine resistance than reported in previous studies -21 and aThis study also confirms that the assumption that ACTs are always more effective than NACTs is not always true especially in settings, such as Madagascar, where SP or AQ monotherapies remain effective. It was observed that AQ+SP was as effective as AQ+AS, consistent with previous reports in Africa both in high transmission ,15,24 anAlthough AQ+AS produced faster parasite clearance than AQ+SP, the risk of new infection within the month after therapy was high with AQ+AS than AQ+SP treatment, even in a low transmission area such as Moramanga. This phenomenon has previously been observed in areas of high transmission ,24. AccoHowever, cost and availability of ACTs remain major concerns, and it appears that the sudden increase in demand for artemisinins may exacerbate these problems, at least in the short-term . AQ+AS (In conclusion, these results indicate that AQ+SP is as effective as AQ+AS, and thereby show that the inexpensive and widely available combination AQ+SP may still be appropriate for the treatment of uncomplicated malaria in Madagascar, an area where resistance to the drugs is relatively uncommon. This could have an important role in this country, where diagnostic services are not accurate and where much of the drugs administered go to patients who do not have malaria.Didier M\u00e9nard was involved in all stages of this study. Ars\u00e8ne Ratsimbasoa, Nohary Nina Harimanana Andrianina, Zakaherizo Ramiandrasoa, Arthur Randriamanantena and No\u00e9line Rasoarilalao performed the field work. Martial Jahevitra performed molecular genotyping. Luciano Tuseo and Andrianirina Raveloson helped to compose the manuscript and gave constructive advice."}
+{"text": "Plasmodium falciparum poses a major threat to malaria control. Combination antimalarial therapy including artemisinins has been advocated recently to improve efficacy and limit the spread of resistance, but artemisinins are expensive and relatively untested in highly endemic areas. We compared artemisinin-based and other combination therapies in four districts in Uganda with varying transmission intensity.Drug resistance in We enrolled 2,160 patients aged 6 mo or greater with uncomplicated falciparum malaria. Patients were randomized to receive chloroquine (CQ) + sulfadoxine-pyrimethamine (SP); amodiaquine (AQ) + SP; or AQ + artesunate (AS). Primary endpoints were the 28-d risks of parasitological failure either unadjusted or adjusted by genotyping to distinguish recrudescence from new infections.p < 0.01) after AQ + SP or AQ + AS . Compared to AQ + SP, AQ + AS was associated with a lower risk of recrudescence but a higher risk of new infection. The overall risk of repeat therapy due to any recurrent infection (recrudescence or new infection) was similar at two sites and significantly higher for AQ + AS at the two highest transmission sites .A total of 2,081 patients completed follow-up, of which 1,749 (84%) were under the age of 5 y. The risk of recrudescence after treatment with CQ + SP was high, ranging from 22% to 46% at the four sites. This risk was significantly lower .The registration number for this trial is ISRCTN67520427 ( Artemisinin-based combination therapy for malaria may be effective at treating primary infection, but may not be the best option in highly endemic areas because of risk of recurrence. Plasmodium falciparum to the inexpensive and widely used drugs, chloroquine (CQ) and sulfadoxine-pyrimethamine (SP) [Malaria remains one of the most serious global health problems and a leading cause of childhood morbidity and mortality, especially in Africa . Effortsine (SP) ,3. Use oine (SP) . Howeverine (SP) In 2000, Uganda replaced CQ with the combination of CQ + SP as the recommended first-line treatment for uncomplicated malaria, although efficacy data for this regimen were lacking. To support development of a rational antimalarial treatment policy, the Uganda Malaria Surveillance Project was formed as a collaborative effort between the Ugandan Malaria Control Program, the East African Network for Monitoring Antimalarial Therapy, and academic researchers to provide efficacy data on antimalarial therapies from multiple sites. In addition to providing efficacy data from diverse locales, this project offers the possibility of evaluating the impacts of varied malaria endemicities and drug resistance patterns on responses to antimalarial therapy.In a recent study from a relatively low-transmission site in Kampala, Uganda, combinations of amodiaquine (AQ) + SP and AQ + artesunate (AS) were found to be significantly more efficacious than CQ + SP. Directly comparing the two AQ-containing regimens over 28 d, AQ + SP was associated with a higher risk of recrudescence after therapy, and AQ + AS was associated with a higher risk of new infections, such that the overall risk of repeat therapy was similar . ConsideWe conducted single-blind, randomized clinical trials to compare the efficacy and safety of three combination antimalarial regimens at four district health centers, using a common study protocol. Study sites, selected for geographic diversity, were: Jinja , Arua , Apac , and Tororo  . The levP. falciparum monoinfection with parasite density 2,000/\u03bcl\u2013200,000/\u03bcl. Because laboratory results were generally not available until the following day, a patient could be excluded after randomization.Consecutive patients presenting to district health centers between November 2002 and May 2004 with symptoms suggestive of malaria and a positive-screening thick blood smear were referred to study physicians. Patients were enrolled if they fulfilled the following selection criteria: (1) age 6 mo or greater; (2) history of fever in the last 24 h or axillary temperature > 37.5 \u00b0C; (3) no history of serious side effects to study medications; (4) no evidence of a concomitant febrile illness; (5) provision of informed consent; (6) no history of treatment with an antifolate or AQ during the previous week; (7) absence of pregnancy-based on history of last menstrual period; 8) no danger signs or evidence of severe malaria [ no dangePatients were randomly assigned to receive one of three oral therapies: CQ  + SP (25 mg/kg sulfadoxine and 1.25 mg/kg pyrimethamine as a single dose on day 0); AQ  + SP; or AQ + AS . Patients in the CQ + SP and AQ + SP treatment arms also received lactose placebo tablets on days 1 and 2.Randomization codes were computer-generated by an off-site investigator for two age groups (6\u201359 mo and 5 y or older) and provided to a study nurse responsible for treatment allocation. All other study personnel were blinded to the treatment assignments, and patients were not informed of their treatment regimen. Patients were directly observed for 30 min after treatment, and the dose was readministered if vomiting occurred. Patients who repeatedly vomited their first dose of study medication were excluded from the study.Following enrollment, patients were asked to return for follow-up visits on days 1, 2, 3, 7, 14, 21, 28, and any other day when they felt ill. Blood was obtained by finger prick for thick blood smears and storage on filter paper on all follow-up days except day 1. Hemoglobin measurements were repeated on day 28 or the day of treatment failure.Treatment outcomes were classified according to 2003 World Health Organization guidelines as early treatment failure , late clinical failure , late parasitological failure , and adequate clinical and parasitological response  [Blood smears were stained with 2% Giemsa for 30 min. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 200 WBCs , assuming a WBC count of 8,000/\u03bcl. A smear was considered negative if no parasites were seen after review of 100 high-powered fields. Thin blood smears were used to detect nonfalciparum infections. Hemoglobin measurements were made using a portable spectrophotometer . Molecular genotyping techniques were used to distinguish recrudescence from new infection for all patients failing therapy after day 3. Briefly, filter paper blood samples collected on the day of enrollment and the day of failure were analyzed for polymorphisms in merozoite surface protein-2 (MSP-2) using nested-PCR as previously described . GenotypThe study was designed to test the hypothesis that treatment with AQ + SP or AQ + AS would change the risk of recrudescence compared to CQ + SP at each study site. We calculated that 176 patients were needed to be enrolled in each treatment arm at each site for a 0.05 two-sided type I error with 0.8 power, assuming a risk of recrudescence adjusted by genotyping of 15% in the CQ + SP group and 5% in the comparison groups..04 , and analyzed using Stata version 8.0 . Efficacy data were evaluated using a per-protocol analysis, which only included patients with treatment outcomes. Prior to completion of this study we decided that a per-protocol analysis would be more appropriate than an intention-to-treat (ITT) analysis. Our protocol was used to study CQ + SP versus AQ + SP at three additional sites prior to the completion of the studies presented here. During the analysis of these previous studies we decided that a per-protocol analysis provided better estimates of the true risk of treatment outcomes than an ITT analysis, as in an ITT analysis one must assign treatment outcomes to patients who did not complete the study. In addition, there were so few patients enrolled who did not complete the study at all our sites that the comparative results using a per-protocol analysis did not differ from that using an ITT analysis. Primary efficacy outcomes included the 28-d risks for all recurrent infection , recrudescence , and new infections (any LCF or LPF categorized as new infection based on genotyping results). Risks of recrudescence and new infection were estimated using the Kaplan-Meier product limit formula with censoring for the competing risk (new infections censored when estimating risks of recrudescence and vice versa). Secondary outcomes included the risk of recurrent infection unadjusted by genotyping at day 14, presence of fever on days 1\u20133, parasitemia on days 2 and 3, change in hemoglobin level between the day of enrollment and the last day of follow-up, presence of gametocytes during any follow-up day, and the incidence of adverse events. Pairwise comparisons of treatment efficacy were made using risk differences with exact 95% confidence intervals (CIs). Other categorical variables were compared using chi-squared or Fisher's exact test, and continuous variables were compared using the independent samples t-test. All reported p-values were two-sided, without adjustment for multiple testing, and were considered statistically significant if less than 0.05.Data were double-entered and verified using EpiInfo 6Of 2,684 patients who underwent screening, 2,270 were randomized to treatment, and 2,160 were enrolled in the studies . Primaryp < 0.001), consistent with more common presentation of older individuals with malaria at a lower transmission site. Patients from Jinja also had higher baseline temperatures (p < 0.001) and mean hemoglobin levels (p < 0.001) compared to the other sites. Parasite densities varied significantly across the sites  and decreased with increasing entomological inoculation rate estimates. The proportion of patients with gametocytes present in pretreatment samples varied considerably (p < 0.001), ranging from 10% (Jinja and Tororo) to 51% (Apac). The proportion of patients providing a history of recent CQ use ranged from 5% (Arua) to 28% (Jinja), with 90% of patients reporting a partial treatment course (less than three doses) prior to presentation at the district health centers.Baseline characteristics of the patients across the three treatment groups were similar at each site . Betweenp < 0.04 for all pairwise comparisons). The risk of recurrent infection (unadjusted by genotyping) at day 14 was significantly higher for CQ + SP at all four sites (16%\u201357%) compared to AQ + SP (1%\u201311%) or AQ + AS (5%\u201313%)  . The ris(5%\u201313%) . After a(5%\u201313%) . Comparep = 0.009) (p < 0.001) and Apac  (p = 0.005) and Apac   . In cont= 0.003) . Overall= 0.004) .Although some patients classified as failures had asymptomatic parasitemia on day 28 (LPF), the majority (67%) were symptomatic on the day of failure (ETF or LCF). Comparisons of treatment efficacy (both unadjusted and adjusted by genotyping) considering only those patients who were symptomatic on the day of failure were similar to those above (data not shown).p = 0.03), although the difference was marginal, and over 75% of both recrudescences and new infections occurred after 20 d of follow-up. No differences between recrudescences and new infections were found with respect to the proportion of patients who were symptomatic, the risk of complicated malaria, parasite density, or changes in hemoglobin. Gametocytes during follow-up were common with both outcomes, but more common with recrudescences .Comparisons of the characteristics of late clinical and parasitological failures due to recrudescence versus those due to new infections are presented in Treatment with AQ + AS was associated with a significantly lower risk of parasitemia on day 2 at all four sites (2%\u20139%) compared to CQ + SP (37%\u201374%) or AQ + SP (33%\u201358%); however, this was not consistently associated with clinical benefit as measured by presence of fever and hemoglobin level . The prop = 0.40). Serious adverse events included anemia , convulsion , dehydration (one AQ + AS), edema (one AQ + SP), malnutrition (one CQ + SP), mental status change (two AQ + SP), respiratory illness , vomiting (one CQ + SP), and weakness (one AQ + SP). One patient died of suspected severe malnutrition and one patient died of congestive heart failure due to a presumed congenital heart defect (both patients had received AQ + SP). All serious adverse events were deemed to be unlikely (ten events) or possibly (ten events) related to the study medications.Among 2,160 patients enrolled in the studies, 20 serious adverse events were reported in 16 patients  and to varied drug resistance patterns. Varied endemicity likely played an important role, as suggested by marked differences in baseline characteristics between the study populations. Considering varied drug resistance patterns, preliminary data analyzing parasites from the four sites suggest that the prevalences of mutations known to mediate resistance to CQ and SP were similar (unpublished data). Thus, the major factor mediating differences in outcomes between study sites appears to be differences in the antimalarial immunity of study populations.Antimalarial drug-efficacy studies that limit follow-up to 14 d or less may significantly underestimate the risk of recrudescence . Our stuFollow-up in this study was limited to 28 d. In a previous longitudinal study of SP alone or in combination with AQ or AS from Uganda, 72% of recrudescences occurred within 28 d and 81% within 42 d . Thus, tWe suggest that antimalarial therapies be judged not only by their impact on the risk of recrudescence, but also by their impact on the risk of new infections after therapy . HoweverThe classification of recrudescence and new infection depends on the genotyping methods used. In our study we used a highly specific definition of recrudescence that required all alleles present at the time of retreatment be present at the time of treatment initiation. As recurrent infections containing both new alleles and alleles present at the treatment initiation were classified as new infections, we may have underestimated the true risk of recrudescence. Defining recrudescence as having any allele at the time of retreatment present at the time of treatment initiation increased our point estimates of the risk of recrudescence. However, using this more sensitive (but less specific) definition of recrudescence did not have a significant impact on comparative efficacies. Indeed, we suggest that the most useful comparisons of the clinical consequences of different treatments should be based on treatment outcome results unadjusted by genotyping, including all retreatments for clinical illness.Artemisinin derivatives are highly attractive antimalarials because they act rapidly, are well tolerated, and are currently not limited by resistance . HoweverAlthough ACT is clearly better than standard monotherapies, the somewhat disappointing results of ACT in Africa probably relate to the high endemicity of malaria and the inclusion of inadequate partner drugs with the artemisinins. Considering endemicity, patients with symptomatic malaria from highly endemic areas such as Africa generally have higher pretreatment parasite densities compared to patients from lower endemic areas such as Southeast Asia, and higDespite the limitations noted, ACT and other antimalarial combination therapies offer great hope for Africa. However, the ideal combination regimens remain uncertain . As AfriProtocol S1(58 KB DOC).Click here for additional data file.Protocol S2(1.0 MB DOC).Click here for additional data file.As resistance to current malarial regimens grows, it is increasingly important to find new combinations of drugs that will not only treat an ongoing infection, but will also prevent recurrence of the malaria (either a new infection or reappearance of the treated infection). One type of drug that is being assessed for preventing recurrence is based on artemisinin, which was originally derived from a plant, sweet wormwood. However, this drug is more expensive than older drugsThey did four randomized clinical trials in Africa at the same time; two in areas where malaria occurs very frequently and two where it is less frequent. They compared a combination of artemisinin with two other combinations of older drugs and looked to see how well the treatments worked on the present infection, on preventing recurrences, and on whether there were any serious adverse events.They found that the combination including artemisinin worked the best at treating current infections. However, patients given the artemisinin-based treatment were overall more likely to get new infections. Where malaria was very common, people treated with the artemisinin-based combination were more likely to get recurrent infections overall.Although the artemisinin-based treatment worked very well on present infections, for recurrent infections it did not perform better than, and was considerably more expensive than, an older combination of drugs. Artemisinin-based treatment should not automatically be assumed the best treatment for uncomplicated malaria in Africa.The World Health Organization has a Web site on Africa with links to malaria control:http://www.afro.who.int/The United Kingdom Department for International Development has information on malaria control in Africa:http://www.lshtm.ac.uk/dfid/malaria/"}
+{"text": "Advances in computing and telecommunications have resulted in the availability of a range of online tools for use in pathology training and quality assurance. The majority focus on either enabling pathologists to examine and diagnose cases, or providing image archives that serve as reference material. Limited emphasis has been placed on analysing the diagnostic process used by pathologists to reach a diagnosis and using this as a resource for improving diagnostic performance.The ReplaySuite is an online pathology software tool that presents archived virtual slide examinations to pathologists in an accessible video-like format, similar to observing examinations with a double-headed microscope. Delivered through a customised web browser, it utilises PHP (Hypertext PreProcessor) to interact with a remote database and retrieve data describing virtual slide examinations, performed using the Virtual Pathology Slide (VPS).To demonstrate the technology and conduct a preliminary evaluation of pathologists opinions on its potential application in pathology training and quality assurance, 70 pathologists were invited to use the application to review their own and other pathologists examinations of 10 needle-core breast biopsies and complete an electronic survey. 9 pathologists participated, and all subsequently completed an exit survey.Of those who replayed an examination by another pathologist, 83.3% (5/6) agreed that replays provided an insight into the examining pathologists diagnosis and 33.3% (2/6) reconsidered their own diagnosis for at least one case. Of those who reconsidered their original diagnosis, all re-classified either concordant with group consensus or original glass slide diagnosis. 77.7% (7/9) of all participants, and all 3 participants who replayed more than 10 examinations stated the ReplaySuite to be of some or great benefit in pathology training and quality assurance.Participants conclude the ReplaySuite to be of some or of great potential benefit to pathology training and quality assurance and consider the ReplaySuite to be beneficial in evaluating the diagnostic trace of an examination. The ReplaySuite removes temporal and spatial issues that surround the use of double-headed microscopes by allowing examinations to be reviewed at different times and in different locations to the original examination. While the evaluation set was limited and potentially subject to bias, the response of participants was favourable. Further work is planned to determine whether use of the ReplaySuite can result in improved diagnostic ability. Diagnostic pathology involves the classification of disease, using tissues obtained during biopsies, operations or autopsies. Classification is based on a complex set of visual features identified with the aid of a microscope. To reach a confident diagnosis, pathologists are required to possess well-developed searching, perception and classification skills, acquired with the assistance of intensive one-on-one tutoring sessions with expert pathologists. The workloads of expert pathologists, however, often restrict the frequency of these interactions.While telepathology has yet to be incorporated into most pathology labs in a clinical role -4, advanThe majority of training tools comprise interactive tutorials that display a limited number of pre-selected images per case, often with accompanying notation -15. HoweIn contrast to static systems, virtual slides (static-dynamic hybrids) enable unrestricted examination of entire digitised tissue sections ,23-27. TIn order to elucidate the diagnostic process of examining pathologists, the VPS , a virtual slide viewer that records diagnostic behaviour, has been developed and validated . The VPSThe ReplaySuite is a web-based, user-friendly software tool that enables pathologists to replay virtual slide examinations, performed using the Virtual Pathology Slide (VPS) ,29,30. UThe VPS is an interactive microscope emulator that presents a complete digitised tissue section via the Internet or CD ROM, allowing both stepwise increases of magnification (16\u00d7 to 2000\u00d7 on standard 17\" monitor) and eight-way directional lateral-motion. During VPS examinations  web pages to display images and text, using PHP, a server-side scripting language. It communicates with an Oracle database, on which all user tracking data is stored, via OCI  to retrieve and write data. JavaScript is used to deliver dynamic client side functionality within the graphical user interface (GUI). The ReplaySuite is deployed in a Microsoft Foundation Class (MFC) application written in Visual C++, which acts as a customised web browser. A complete description of the ReplaySuite specification is available in the ReplaySuite technical document .Data describing an examination is retrieved from the remote server, via server side scripting, enabling communication between the application and the database. Structured Query Language (SQL) queries are embedded within PHP, interrogating the database and retrieving datasets. This data is used to determine which images to request from the server or CD-ROM, and how long to display these images. Server side scripting is also used to dynamically generate static HTML, which provides the GUI to display the images. During a replay, each image viewed during the examination is displayed in the order it was examined and for the duration it was examined. Diagnostic comments made during the examination are displayed adjacent to the appropriate field. Client side scripting (JavaScript) is used to dynamically alter the main field of view locator on the slide overview, indicate the number of seconds left until the next field of view is displayed and specify the upcoming action  performed to navigate to the next field.Users are required to login into the ReplaySuite, using a unique username and password. In order to maintain user anonymity, users are assigned unique VPS ID numbers. When examination or diagnostic data is displayed, these VPS ID numbers are used to identify the examining pathologist. Fig. Upon logging in, the ReplaySuite displays an Examination list of all VPS examinations performed by that user. Users may also view, on demand, an Examination list of examinations for a specific case or sub-classify to view all examinations returning a specific diagnosis for a specific case. Examination lists display summarised information about each VPS examination, and for each examination displays, from left to right, the examining pathologists identification number (ID), the slide examined, the examiners diagnosis, group consensus diagnosis for the slide and the degree of group consensus Fig . Highlig1. View an examination list of all examinations for a selected slide2. View Diagnostic concordance graphs for a selected slide3. Replay any users examination of a selected slideDiagnostic concordance graphs are graphical representations of the variation in classification concluded by all examiners of a slide. Only data relating to slides previously examined by the user may be accessed. Two graphs are displayed; the top graph showing the variation in classification, the bottom graph the variation in sub-classification for B5  diagnosis Fig. . The numClicking 'View' in the Replay column on the right of an Examination list replays an examination. During a replay, each field of view examined is displayed in chronological order, appearing on-screen for the duration it was originally examined. Unlike real-time observation of an examination, replays may be paused, fast-forwarded and rewound. A slide overview, located in the top left of the interface Fig , relatesA preliminary study was conducted to demonstrate the capabilities of the ReplaySuite to pathologists, and to evaluate their opinions on its use and potential application in training and quality assurance. Participants were provided with open access to the 10 needlecore biopsies examined during the VPS validation study , which tUsing the ReplaySuite, participants were permitted to view group concordance graphs, replay their own and others examinations, view summary report forms and view statistical breakdowns of individual examinations. Use of the ReplaySuite was monitored and recorded to the database, allowing the identification of light/heavy users and highlighting the most frequently used functions. Table The ten needle core biopsies examined during the VPS validation study  were obt70 pathologists were invited to participate in the study. 38 Irish participants were invited with the assistance of Professor Peter Dervan, Mater Miscordiae Hospital, Dublin, and the remaining invitees comprising the 32 members of the European Working Group of Breast Screening Pathology (EWGBSP). EWGBSP members are recognised as expert breast pathologists in their native countries and all member states of the European Union (EU) are represented within the group. 17 of the 70 invited participants previously participated in the VPS validation study , 4 of whSubsequent to the study, participants completed an electronic questionnaire on their use and impressions of the software and its potential applications. Participants were asked to submit their level of agreement/disagreement with 19 statements using a Likert scale , and answer Yes or No to a further 5 questions. A number of questions were inverted to mitigate against the well-known bias of positively phrased questions. The following are the pertinent questions asked:\u2022 Did replaying an examination by another pathologist with a diagnosis different to your own provide you with an insight into why that diagnosis was concluded?\u2022 Did replaying examinations performed by other pathologists make you reconsider your diagnosis for any slides?\u2022 The ReplaySuite is user-friendly\u2022 The diagnostic pathway followed by the examining pathologist was apparent\u2022 Being able to pause, fast-forward and rewind replays was useful\u2022 The information panel was not useful\u2022 Training\u2022 Quality AssuranceParticipants were also given the opportunity to provide open-ended feedback:\u2022 Add any further comments, or have any additional features you would like to see incorporated into the ReplaySuite.\u2022 If yes , please state the difficulty that occurredParticipants who used the ReplaySuite were resurveyed post-study in order to determine if they regularly participated in teaching. 3 questions were asked:\u2022 Are you, or have you been involved in providing undergraduate medical training on a regular basis?\u2022 Are you, or have you been involved in providing postgraduate pathology training on a regular basis?\u2022 If you are not currently involved but have previously been in either activity, please state how long ago you were involved.9 participants used the ReplaySuite to review examination data. All 9 completed the electronic questionnaire, and all but one had more than five years experience in pathology practice, the exception possessing three years experience. 4 of the 9 participants were members of the European Working Group of Breast Screening Pathology. Table 7 participants (77.7%) replayed at least one examination and of these, 3 participants replayed more than 10. All participants who replayed at least one examination agreed or strongly agreed that the ReplaySuite was user-friendly Fig.  and 85.7All participants who replayed at least one examination agreed or strongly agreed that the diagnostic pathway of the examining pathologist was apparent during a replay. Of the 7 participants who replayed their own or others examinations, 6 (85.71%) replayed another pathologists examination. Of these 6 participants, 5 (83.3%) agreed that it provided an insight into the examining pathologists diagnosis. Of those who replayed another's examination, 2 (33.3%) reconsidered their original diagnosis, 1 re-diagnosing concordant with group consensus and 1 re-diagnosing concordant with original glass slide diagnosis.After originally concluding an original diagnosis of B2 (Benign) for Slide 5, User 7 replayed his/her own examination, and then replayed an examination of Slide 5 by User 6. This examination (User 6) concluded a diagnosis of B5 , concordant with group consensus.User 10 concluded an original diagnosis for Slide 8 of B4 , which concurred with group consensus but not glass slide diagnosis, B3 . User 10 replayed their own examination, then an examination of the same slide that concluded the same diagnosis by User 5. Two examinations concluding a B3 diagnosis were then replayed (Users 87 and 55), concordant with glass slide diagnosis.When asked to rate the potential benefit of the ReplaySuite in pathology training, 6 out of 7 of participants who had replayed at least one examination (85.71%) and all of those who had replayed more than 10 examinations rated it as of some or of great benefit Fig. . Of the When asked to rate the potential benefit of the ReplaySuite in quality assurance, 5 out of 7 of participants who had replayed at least one examination (71.4%) and all participants who had replayed more than 10 examinations considered it of some or of great benefit  and client-side (JavaScript) open source scripting languages, it presents this data to users via an intuitive Graphical User Interface (GUI), which was considered easy to use by participants who replayed at least 1 examination Fig. . UtilisiUse of the VPS for diagnosis has already demonstrated \"substantial\" diagnostic agreement between users, 88.23% of examining pathologists obtaining a Kappa of between 0.97 and 0.65 [et al (2003) have reported similar difficulties with low participation rates [et al (2003) cited technical difficulties and pathologists workloads as principal factors for low participation [A number of caveats should be considered when reviewing study data, the first being small sample size. There is a significant difference between the number of pathologists invited to participate and those who completed the study (9/70). While not unique in studies involving pathologists use of telepathology systems, the small sample size may be considered a potential source of error. Bamford on rates , howeveron rates . Low parcipation . System cipation . As bothSecondly, it is not unreasonable to suggest that the positive evaluation of the ReplaySuite may in part be attributable to bias from participants 4/9) who also previously participated in the VPS validation study. Pathologists who participated in both VPS and ReplaySuite studies expressed greater satisfaction and confidence in the VPS during the VPS validation study than pathologists who participated only in the VPS study . However who alsoIt also goes without question that those who participate in studies of this nature are often early adapters and often possess a positive bias towards new technology. This is an issue when evaluating any new software, and in that context, bias is unavoidable. 20 of the 70 participants invited to participate had foreknowledge of the VPS, however none, had previously seen the ReplaySuite and, therefore, had no preconceived notions about the software.The concept of a virtual double-headed microscope is not new , howeverThe benefits of various online tools and their impact on diagnostic performance have already been highlighted, however the ReplaySuite possesses a number of practical advantages. Many online tools present the diagnostic opinion of one pathologist. In contrast, the ReplaySuite can replay examinations of the same slide by multiple experts, illustrating a number of different diagnostic pathways that corroborate the same diagnostic hypothesis. Alternatively, reviewing pathologists may observe examinations that disagreed with group consensus, in order to identify the possible sources of disagreement. This is of particular interest for disorders that suffer from a high degree of inter-observer variability. Additionally, interactive tutorials and annotated virtual slides require considerable time to create, however, individual authoring time with the VPS/ReplaySuite is around 6 minutes, per user, per slide.Figures The objective of the study was to develop the ReplaySuite technology, and to assess the opinions of pathologists on its use and potential application. Participants concluded the ReplaySuite to be of some or of great potential benefit to pathology training and quality assurance and considered the ReplaySuite to be beneficial in evaluating the diagnostic trace of an examination. Future evaluation of the ReplaySuite in a larger quality assurance study and training environment is anticipated.One-to-one human tutoring remains, and will remain for the foreseeable future, the pre-eminent means of nurturing pathologist's diagnostic skills. However, as factors such as expert availability and case diversity vary, from institution to institution, it is impossible to replicate the same training environment universally. Supplementary online training tools may well help to redress the balance, providing wide access to expert pathologists and a diverse range of cases from multiple institutions, enabling an educational diversity often unachievable in a single institution. The ReplaySuite is a unique tool that permits pathologists to learn from multiple experts in a manner not possible with current tools.DS and SC possess share holdings in Slidepath Ltd, a company delivering informatics solutions for use in pathology quality assurance. DJ and PD have share options in SlidePath Ltd.DS conceived the ReplaySuite and participated in the design of the study. DJ created the ReplaySuite, carried out the study and drafted the manuscript. SC created the VPS and participated in the design of the database. PD participated in the design of the ReplaySuite and provided a pathologist's insight.The pre-publication history for this paper can be accessed here:"}
+{"text": "Sulfadoxine-pyrimethamine (SP) is an antimalarial drug that acts on the folate metabolism of the malaria parasite. We investigated whether folate (FA) supplementation in a high or a low dose affects the efficacy of SP for the treatment of uncomplicated malaria in pregnant women.This was a randomized, placebo-controlled, double-blind trial.The trial was carried out at three hospitals in western Kenya.The participants were 488 pregnant women presenting at their first antenatal visit with uncomplicated malaria parasitaemia (density of \u2265 500 parasites/\u03bcl), a haemoglobin level higher than 7 g/dl, a gestational age between 17 and 34 weeks, and no history of antimalarial or FA use, or sulfa allergy. A total of 415 women completed the study.All participants received SP and iron supplementation. They were randomized to the following arms: FA 5 mg, FA 0.4 mg, or FA placebo. After 14 days, all participants continued with FA 5 mg daily as per national guidelines. Participants were followed at days 2, 3, 7, 14, 21, and 28 or until treatment failure.The outcomes were SP failure rate and change in haemoglobin at day 14.p = 0.8), and 27.1% (38/140) in the FA 5 mg arm . The haemoglobin levels at day 14 were not different relative to placebo .The proportion of treatment failure at day 14 was 13.9% (19/137) in the placebo group, 14.5% (20/138) in the FA 0.4 mg arm , normally 0.4 mg per day. There is good evidence from systematic reviews of controlled trials that folate supplementation around conception and early in pregnancy is effective in protecting against neural tube (spine and brain) defects; continued supplementation throughout pregnancy reduces the chance of anemia in the mother. In many African countries, including Kenya, the dose of folate used is 5 mg per day, because this dose is more easily available there. In Kenya, as well as elsewhere in Africa, sulfadoxine-pyrimethamine is also given twice or more after the first trimester to treat and/or prevent malaria infection . However, there is some evidence from laboratory experiments and clinical studies, none of which were done in pregnant women, suggesting that folate supplementation might reduce the effectiveness of sulfadoxine-pyrimethamine. Therefore, these researchers conducted a trial to test this hypothesis in 415 pregnant Kenyan women with malaria parasites in the blood but no severe symptoms. All were given standard sulfadoxine-pyrimethamine treatment. The women were randomized to receive either folate 5 mg daily, folate 0.4 mg daily, or placebo tablets for 14 days, after which all women reverted to the standard folate 5 mg tablets. The women were followed up for 28 days after the initial sulfadoxine-pyrimethamine dose and the principal outcome the researchers were interested in was the failure of sulfadoxine-pyrimethamine treatment, defined as fever and the presence of parasites in the blood  or the failure of parasites to clear from the blood or to reappear too soon .What this trial shows: In this trial, women receiving folate 5 mg daily were approximately twice as likely to fail treatment with sulfadoxine-pyrimethamine than women receiving folate 0.4 mg or placebo.  All the treatment groups had similar levels of blood hemoglobin at the end of the study. There did not seem to be any major differences in adverse events  among women taking part in the different study groups.Strengths and limitations: The randomization procedures were appropriate and procedures were used to blind participants and researchers to the different interventions, therefore reducing the risk of bias. Since the trial had a placebo arm, it was possible to conclude that the lower dose of folate (0.4 mg) did not significantly affect efficacy of sulfadoxine-pyrimethamine as compared with placebo. A limitation of the study is that the length of the intervention was short, since all women reverted to standard 5 mg folate after 14 days. It is therefore not clear whether a longer trial would have shown additional risks or benefits of the different doses of folate. Finally, PCR genotyping was not done on the parasites infecting women in the trial; this procedure could have distinguished between true treatment failures and new infections (but which would have been unlikely within 14 days).Contribution to the evidence: Other trials and observational studies have suggested that high doses of folate can reduce the efficacy of sulfadoxine-pyrimethamine in children and adults. However these studies have not examined the effect in pregnant women, for whom most national bodies recommend regular folate supplementation. The results from this trial supports the findings from previous studies and enables the evidence to be generalized to pregnant women. The study also found no evidence that 0.4 mg folate compromises the efficacy of sulfadoxine-pyrimethamine. The findings suggest that the lower level of folate dosing should be used in pregnancy, or that antimalarial treatments other than sulfadoxine-pyrimethamine be used. Plasmodium falciparum than nonpregnant women, affecting approximately 30 million pregnancies annually . Th. Th23]. Malaria was defined as the presence of asexual-stage parasite of any species in thick smears, independent of clinical signs. A parasite density in the highest tercile at enrolment of the total study population was defined as a high parasite density. A young age was defined as under 20 years. Because of the short duration of follow-up and limited sample size, no attempt was made to assess the effect of FA on megaloblastic anaemia.p-value of less than 0.01; we used day 14 and not day 7 because we considered day 14 a more appropriate time point for assessing SP resistance. Because this criterion was met, enrolment was stopped with 488 women enrolled. The investigators remained blind until data cleaning, analysis, and quality control of the blood smears were completed.We calculated that a sample size of 600 women\u2014200 in each arm\u2014would allow us to detect an increase from 5% to 15% in the parasitological failure rate at day 7 with 80% power and 95% confidence, allowing for a 25% loss to follow-up. However, we did not feel comfortable continuing the trial at an overall treatment failure rate of over 40% at day 28 and an intervention which may contribute to this, because it is recommended that first-line therapy be changed at a 25% failure rate ,24. An iOne of the investigators generated a randomization list with a block size of 12 using the statistical program SAS .All FA treatment and placebo tablets were prepared off site and were identical in appearance and taste (Laboratory and Allied). Medicine envelopes with 14 tablets of a treatment arm were prepacked and labelled with the arm by staff who were not involved in randomization. The medicine envelopes were put in sealed, opaque envelopes with consecutive numbers according to the randomization list by an investigator.A trained clinical officer or nurse randomized eligible women by assigning them the next envelope in order of enrolment. The envelope was opened by the participant, and the study arm was allocated by the study staff according to the arm indicated on the medicine envelope.All FA treatment and protocol tablets were prepared off site and were identical in appearance and taste (Laboratory and Allied). All study staff participants were blind to the treatment in each arm.p-value was 0.05 or more. The statistical program SAS (SAS system for Windows version 8) was used for all analyses. All tests were two-sided; p < 0.05 was considered significant, except for the efficacy between study arms when a p < 0.013 was considered significant to adjust for multiple comparisons and the interim analysis ; a confidence interval (CI) of 98.7% was used for the efficacy analysis.We analysed the data on an intention-to-treat basis using a pre-established analysis plan. Cumulative treatment failures by follow-up day were compared among treatment arms using the Chi-squared test. We used the Kaplan-Meier curve to examine differences in patterns between treatment arms, and Cox proportional hazards regression analysis to examine the effect of treatment arm on time to treatment failure after we confirmed that the Cox proportional hazard assumption was met. For day 14 post-treatment, we repeated the Cox proportional hazards regression while adjusting for potential confounders; factors examined included site of enrolment, the use of an ITN, ethnicity, education level, socioeconomic status, sickle cell status (carrier versus not a carrier), gravidity, young age, HIV status, and a high parasite density at enrolment. Analysis of covariance was used to assess the effect of treatment arm on haemoglobin level . FactorsPlasmodium falciparum (98.0%), nine were mixed P. falciparum/P. malariae, and one was pure P. malariae. During 1,671 (99.4%) of the 1,682 routine visits made at or before day 14, the participant reported that she took the FA daily; the tablets were brought at 1,454 of the routine visits (86.4%) and a correct count was established at 1,307 visits (89.9%).Between November 2003 and November 2005, a total of 4,524 women were screened; 488 met all enrolment criteria, and 415 (85%) women completed the study . The stup < 0.01 comparing the FA 0.4 mg arm or the FA placebo arm to the FA 5 mg arm). On day 14 the number of treatment failure was 38 out of 140 women (27.1%) in the FA 5 mg arm, 20 out of 138 women (14.5%) in the FA 0.4 mg arm, and 19 out of 137 women (13.9%) in the FA placebo arm (p = 0.005), whereas FA 0.4 mg did not affect treatment failure risk   .p = 0.3, and 0.14 g/dl; 98.7% CI, \u22120.21 to 0.49 g/dl; p = 0.4, respectively; adjusted for maternal HIV infection, location of residence, high parasite density infection, and haemoglobin at enrolment).We did not find an effect of treatment arm on haemoglobin levels at day 14 or day 28 among 288 women who completed 28 days of follow-up without treatment failure . Among 3During the course of the study, 20 participants (4.1%) developed rashes . No severe adverse skin reactions or maternal deaths occurred. Premature delivery was experienced by 14 participants (2.9%) , and eight participants (1.6%) had a stillbirth or early neonatal infant death during the study .This study shows that the combined use of SP and daily FA supplementation in a dose of 5 mg compromised the efficacy of SP for the treatment of malaria parasitaemia in pregnant women. A plausible biological mechanism is available. FA is required for DNA synthesis in both humans and protozoa. Malaria parasites can utilize exogenous FA  as well as synthesize FA de novo (biosynthesis) , though It is likely that FA supplementation affects other antifolate antimalarial combinations as well, such as chlorproguanil-dapsone, dapsone-pyrimethamine, and cotrimoxazole. Cotrimoxazole will increasingly be used as a prophylactic drug among HIV-positive pregnant women. Further study into the effect of concomitant FA supplementation in malarious areas is needed.We did not collect blood at enrolment and follow-up visits to be able to differentiate between recrudescent and new malaria infections. After day 14, all groups switched to FA 5 mg, so we were not able to assess the extent to which FA contributes to SP treatment failure after 14 days. Several studies indicate that FA supplements do not predispose to increased risk of malaria acquisition ,32, and Our results are supported by studies among symptomatic, nonpregnant persons in areas of different malaria endemicity. A randomized, placebo-controlled study in Gambia reported approximately twice as common SP treatment failures among children with symptomatic malaria supplemented with a high dose of FA  compared to the FA placebo group in an area with low seasonal malaria transmission . In a loSP is recommended for the treatment and prevention of malaria in pregnancy. Although our results are based on the treatment of uncomplicated malaria in pregnant women, they will have implications for the use of SP as IPTp as well. Many countries have introduced IPTp with SP . We cannResistance to SP was high in the study area. However, at present, no safe and efficacious alternative drug is available for the treatment and prevention of malaria in pregnancy. Kenya has moved now to artemisinin-based combination therapy for children and quinine as first-line therapy for clinical malaria in pregnancy, but IPTp with SP continues to be used for prevention. A recent review of the efficacy of IPTp with SP in the face of increasing SP resistance reported that in areas with parasitological failure as high as 30% at day 14 in children under 5 years of age, significant reductions in adverse effects of malaria in pregnancy were seen when IPTp was used \u20138. HowevCONSORT ChecklistClick here for additional data file.(50 KB DOC)Trial ProtocolClick here for additional data file.(298 KB DOC)"}
+{"text": "Falciparum malaria is an important cause of maternal, perinatal and neonatal morbidity in high transmission settings in Sub-Saharan Africa. Intermittent preventive treatment with sulphadoxine-pyrimethamine (SP-IPT) has proven efficacious in reducing the burden of pregnancy-associated malaria but increasing levels of parasite resistance mean that the benefits of national SP-IPT programmes may soon be seriously undermined in much of the region. Hence, there is an urgent need to develop alternative drug regimens for IPT in pregnancy. This paper reviews published safety and efficacy data on various antimalarials and proposes several candidate combination regimens for assessment in phase II/III clinical trials. Falciparum malaria is an important cause of maternal anaemia, intra-uterine growth retardation, intrauterine death, stillbirth, premature delivery, low birth weight (LBW), perinatal and neonatal morbidity and mortality -5 and povs mefloquine[vs SP plus artesunate[vs SP plus azithromycin[In areas of high or moderate transmission, most malaria infections in pregnant women are asymptomatic and infected women do not present for treatment. In such areas, the World Health Organization recommends a combination of interventions to prevent malaria in pregnancy including insecticide-treated bednets (ITNs), intermittent preventive treatment in pregnancy (IPTp) and effective case management and treatment,17. A smefloquine; SP alonrtesunate; and SP thromycin respectithromycin, but conthromycin,49 and wthromycin, are likThe future of SP for intermittent preventive treatment of malaria in pregnancy is tenuous, particularly in East and Southern Africa, and there are insufficient, reliable data on the safety and efficacy of alternative antimalarials for the prevention and treatment of malaria in pregnancy. Hence, \u00a9, Malorone\u00a9, Coartem\u00a9, Lapdap\u00a9, truncated forms and associated terms  were used where it was felt appropriate. Manufacturers' information sheets, guidelines and reports from the World Health Organization (WHO), Medicines for Malaria Venture (MMV), Gates Malaria Partnership (GMP) and other leading organizations in the field were also scrutinized.A two-stage, systematic search was carried out in PubMed and EMBASE using combinations of the keywords: pregnancy, malaria, antenatal, prophylaxis, intermittent, prevention, clinical trial. Alternative spellings and truncated forms were also used. ArticleMefloquine hydrochloride is a 4-quinolinemethanol derivative with a chemical structure related to that of quinine. In healthy, non-pregnant human volunteers, peak plasma concentrations of around 250 \u03bcg/L are observed 6\u201324 h following a single 250 mg oral administration. MefloquMefloquine has been used alone for prophylaxis in the pre-conception period and in all trimesters of pregnancy-39,41,60Mefloquine, at standard prophylactic doses of 250 mg/week appears safe and effective when taken before conception and during all trimesters of pregnancy,39,41. IHowever, an observational, non-comparative study among seventy-two, female US soldiers stationed in Somalia reported a risk of spontaneous abortion of 16.7% (12/72) among women who took a standard dose mefloquine prophylaxis in the first trimester. One molThere is good evidence that mefloquine is safe and effective in the treatment of falciparum malaria in pregnancy. A low dose mefloquine treatment regimen  given to 33 women in the second or third trimester in Zaria, Nigeria was well tolerated, safe and effective with no adverse maternal or foetal outcomes reported. In a laIn contrast, a retrospective study in Thailand reported a borderline statistical association between mefloquine treatment during the second and third trimester of pregnancy and excess odds of stillbirth compared to women treated either with quinine or other antimalarials. Among 2Several studies were identified which looked at the safety and efficacy of mefloquine in combination with other antimalarials. All were conducted among women in the second or third trimester. A randomized, controlled trial with 40 subjects in Nigeria compared artemether (3.2 mg/kg intramuscularly once at entry) plus low dose mefloquine  versus artemether alone (3.2 mg/kg at entry followed by 1.6 mg/kg/day for 4 days) for the treatment of multidrug-resistant falciparum malaria. No seriA randomized, open-label clinical trial among 108 second and third trimester pregnant women in northwest Thailand compared a three-day mefloquine-artesunate regimen  versus seven days treatment with quinine  for multidrug-resistant falciparum malaria. A numbeA phase III equivalence trial comparing intermittent SP and mefloquine alone for the prevention of malaria-related morbidity in pregnancy is currently underway in Benin.Azithromycin is a macrolide antibiotic derived from erythromycin which has an elimination half-life of 68 h in healthy human volunteers and is excreted largely unchanged in the bile following hepatic clearance. In a prAzithromycin has been used extensively to treat sexually-transmitted and other bacterial infections during pregnancy in total treatment doses of up to 1\u20132 g and is considered a safe alternative for the treatment of chlamydia, mycoplasma and group B streptococci in pregnant women who are allergic to beta-lactam antibiotics -73.P. falciparum in vitro[Azithromycin is not yet licensed for use as an antimalarial agent but has shown promising activity against  in vitro,75, in t in vitro and in r in vitro-79. A ph in vitro. No seri in vitro. The effTo date, no clinical trials have been conducted among pregnant women but a phase III clinical trial of intermittent SP plus azithromycin in pregnancy versus intermittent SP alone is currently underway in Malawi.The elimination half-life of proguanil is 12 \u2013 21 h in healthy adult volunteers and around 40\u201360% of the drug is eliminated by urinary excretion. Proguanil is metabolized by a cytochrome P450 isoenzyme to an active metabolite (cycloguanil), responsible for antimalarial activity. Late prProguanil has been used extensively for malaria chemoprophylaxis in travellers, including pregnant women and children and is considered one of the safest antimalarials currently available,42,58,82\u00a9 (GlaxoSmithKline). Atovaquone has an elimination half life of 2\u20133 days in adult volunteers and around 95% is excreted unchanged in the bile[Proguanil has also been used in fixed combination with atovaquone (100 mg/250 mg respectively) in the commercially available preparation Malarone the bile. A rando the bile. In an o the bile. In a sm the bile. Median A randomized open-label clinical trial among 81 second and third trimester pregnant women in Thailand compared quinine and artesunate-atovaquone-proguanil (AAP) for the treatment of uncomplicated falciparum malaria and found that women treated with AAP had shorter fever and parasite clearance times and lower treatment failure rates (5% vs. 37%) compared to those in the quinine group. The AAP\u00ae) has a similar mode of action and pharmacokinetic profile to proguanil but has a slightly longer elimination half-life (around 32 h)[\u00a9 (GlaxoSmithKline). Despite evidence from a number of clinical trials of the safety and efficacy of this combination in the treatment of uncomplicated malaria in children[Chlorproguanil (Lapudrineund 32 h). The dru children,87,88 an children, a recen children conclude children and in M children. A combi children and a mu children.Artemisia annua), a herb which has been used in traditional Chinese medicine for over 2000 years[Artemisinin compounds  are derived from sweet wormwood  have been reported,95. TherFollowing two expert consultations in 2002, WHO recommended that artemisinins should not be used in the first trimester except in life-threatening situations where other antimalarials were not suitable, should be used with caution in the second and third trimesters and that further research be conducted to establish the pharmacokinetics of artemisinins during pregnancy, mechanism of toxicity and the relevance of animal toxicity data to humans. Pharmac\u00a9/Riamet\u00a9), which is available at subsidized cost to malaria-endemic countries following a special pricing agreement established in 2001 between WHO and the manufacturer, Novartis[The only fixed-dose artemisinin combination therapy (ACT) currently available is artemether-lumefantrine  was developed through an international public-private partnership between Holleykin Pharmaceutical, China; Sigma-Tau Industries, Italy; the University of Oxford, UK; and the Medicines for Malaria Venture (MMV)[Dihydroartemisinin is a derivative of artemisinin and the principal active metabolite of artesunate and artemether with an elimination half-life of 40\u201360 minutes. Piperaqnd others. Dihydroure (MMV). DP has ure (MMV),105-107.ure (MMV) and the \u00a9 (Sanofi-Aventis). Clinical trials in Ghana[A variety of new ACTs are currently being evaluated in phase II and III clinical trials in children and non-pregnant adults. Amodiaquine plus artesunate is being developed in fixed dose co-formulation as Coarsucam in Ghana and Tanz in Ghana have rep in Ghana. Given u in Ghana,49, it i in Ghana-113.The efficacy of dihydroartemisinin-mefloquine (DHA-MQ) compared to DHA alone and chloroquine alone for the treatment of uncomplicated falciparum malaria was recently described in a small study among 75 children aged 2\u201313 years conducted in Nigeria but no pPyronaridine, a Mannich-base schizontocide structurally related to the aminoacridine drug quinacrine, which exerts its antimalarial action by forming complexes with haematin, is beinChlorproguanil-dapsone-artesunate (CDA), artesunate-atovaquone-proguanil (AAP) and artesunate-SP are other promising ACTs under development, as described above.P. falciparum-infected erythrocytes following their adherence to placental chondroitin sulphate A (CSA)[The pharmacokinetics, dosing regimen, efficacy and safety profile of antimalarials currently used to treat and prevent falciparum malaria in adults and children are well-described. This is not the case in pregnancy where the limited data available in each of these areas makes it difficult to predict which drugs are likely to be the best candidates for treating and preventing infection. Many antimalarials are contra-indicated in pregnancy , whilst the safety of several promising candidates remains unclear. In malae A (CSA) and hyale A (CSA), which ee A (CSA). Pregnane A (CSA)-123, whie A (CSA),8. Physie A (CSA). For exae A (CSA),101. Fore A (CSA),55. Simie A (CSA). Given te A (CSA), it seemHow then to decide which antimalarials are likely to be effective for IPT in pregnancy and which combinations should be prioritized for investigation in clinical trials Table ? If the \u00a9)[Based on these criteria, dihydroartemisinin-piperaquine appears an appropriate candidate for IPTp and should be considered for phase II/III clinical trials in second and third trimester pregnant women. The combination is relatively inexpensive ; combineArtemether-lumefantrine may also be a suitable candidate: a relatively inexpensive, fixed co-formulation is available and the partner drugs have different molecular targets. The 'prophylactic effect' conferred by lumefantrine may not be sufficient however, given its relatively short half-life compared to sulphadoxine, pyrimethamine, mefloquine or piperaquine.Mefloquine in combination with artemether or artesunate has the advantage of a similar elimination profile to dihydroartemisnin-piperaquine and the safety of such combinations in the treatment of pregnancy-associated malaria is reasonably well-established. Indeed, the extensive data available on mefloquine use in pregnancy compared to piperaquine suggest that dihydroartemisinin-mefloquine could enter phase III trials ahead of dihydroartemisinin-piperaquine. Cost and the lack of fixed dose co-formulations  are disadvantages. Combining mefloquine with the relatively long acting azithromycin is another option and likely to have additional health benefits by reducing maternal and perinatal infections, which may offset cost concerns. Sexually-transmitted infections (STIs) are common in women of childbearing age in Africa and may go undiagnosed and untreated during pregnancy. MonthlyPatterns of SP resistance vary across sub-Saharan Africa so that new artemisinin-containing combinations with SP may be appropriate in West Africa, given that these would fulfil many of the criteria above. Such combinations are, however, unlikely to succeed in East and Southern Africa, given prevailing levels of resistance.Plasmodium vivax infection also needs to be taken into consideration[P. vivax infection[Are different types of drug combination needed in different areas of malaria transmission? Evidence for the effectiveness of IPTp comes primarily from studies conducted in areas of intense perennial transmission , but there may be a case for evaluating IPTp in areas of low transmission . The World Health Organization recommends that IPTp be an integral part of national malaria control programmes in high/medium perennial or seasonal transmission areas but caution that there is insufficient evidence for introducing IPTp in low transmission settings where the principal focus should be on effective case management and insecticide-treated bednets. In areaideration. In addiinfection,131.AV participated in the conception and design of the review, coordinated the literature review and drafted the manuscript.LV participated in the conception and design of the review, conducted the literature search, obtained full text articles and helped draft the manuscript.JC participated in the design of the review and helped draft the manuscript.BG participated in the conception and design of the review, provided additional information e.g. conference proceedings and helped draft the manuscript.DC conceived the study, participated in its design and coordination and helped draft the manuscript.All authors have read and approved the final manuscript.The author(s) declare that they have no competing interests."}
+{"text": "A six-dose antimalarial regimen of artemether-lumefantrine (A/L) may soon become one of the most widely used drug combination in Africa, despite possible constraints with adherence and poor absorption due to inadequate nutrition, and a lack of pharmacokinetic and effectiveness data.Plasmodium falciparum transmission setting, plasma lumefantrine concentrations were measured in a subset of patients on day 3 (C [lum]day3) and day 7 (C [lum]day7) post-inclusion. Predictors of lumefantrine concentrations were analysed to show how both C [lum]day7 and the weight-adjusted lumefantrine dose affect 28-day recrudescence and re-infection risks. The implications of these novel findings are discussed in terms of the emergence of lumefantrine-resistant strains in Africa.Within a trial of supervised versus unsupervised A/L treatment in a stable Ugandan day3 and C [lum]day7 distributions among 241 supervised and 238 unsupervised patients were positively skewed. Unsupervised treatment and decreasing weight-adjusted lumefantrine dose were negatively associated with C [lum]day3. Unsupervised treatment and decreasing age showed strong negative associations with C [lum]day7. Both models were poorly predictive (R-squared < 0.25). There were no recrudescences in either arm, but decreasing lumefantrine dose per Kg resulted in up to 13-fold higher adjusted risks of re-infection. Re-infections occurred only among patients with C [lum]day7 below 400 ng/mL (p < 0.001).C [lum]Maintaining the present six-dose regimen and ensuring high adherence and intake are essential to maximize the public health benefits of this valuable drug combination. Plasmodium falciparum malaria. The three day, six dose regimen of A/L is currently prioritized by the World Health Organization as a replacement for failing antimalarial monotherapies, notably chloroquine and sulfadoxine-pyrimethamine (SP). Several countries  have now adopted A/L as the first line antimalarial. Clinical trials have shown A/L to be safe and highly efficacious  values > 5 ng/mL, with coefficients of variation of 1.8 to 4.2% .day3 and C [lum]day7 were correlated using Spearman's rho correlation coefficients. The association between baseline patient variables and C [lum]day3 or C [lum]day7 was then analysed in separate multivariate linear regression models, proceeding as follows. First, the dependent variable (lumefantrine concentration) was transformed into its square root (C [lum]day3) or natural logarithm (C [lum]day7) so as to normalize distributions. For each patient, the weight-adjusted lumefantrine dose prescribed in mg/Kg was then calculated (= number of tablets per dose \u00d7 6 doses \u00d7 120 mg/dose/bodyweight in Kg), and categorized as <50, 50\u201364, 65\u201379, and \u226580 mg/Kg, which corresponded almost exactly to 1 SD intervals away from the mean of 64 mg/Kg. Certain baseline parameters were selected as independent explanatory variables for inclusion in the models: (i) if they appeared likely to represent plausible biological or epidemiological predictors ; or (ii) if they were associated with the outcome at the p < 0.20 level in a univariate analysis . The regression was performed on all selected explanatory variables, and non-significant ones were then progressively removed. Dropped variables were then re-added one by one and retained if they affected the model or were associated significantly with the outcome. Weight-adjusted lumefantrine dose, age group and arm were forced into all final models . After searching for plausible interactions and assessing the effect of influential data points, model assumptions were tested, including normality of residuals, linearity between dependent and explanatory variables, lack of collinearity among explanatory variables, and constant variance (homoscedasticity).Data were double-entered on EpiData 3.0 , and analysed using Stata 8.2 . Lumefantrine concentrations were compared by arm within age groups using the unpaired t-test, and C [lum]The association between the weight-adjusted lumefantrine dose and the risk of re-infection during the 28 days of follow-up was then analysed by the Cox proportional hazards model, controlling for potential confounders. All enrolled patients who were analysable by intention to treat (ITT) were included in this analysis day3 results were available for 22 of these patients (4.6%), and no C [lum]day7 results for were available for 24 (5.0%). A further five patients with outlier values were excluded from day 7 analyses. Characteristics of patients with missing data did not significantly differ from those of the analysable sample (data not shown).Baseline characteristics of the 479 patients who had at least one pharmacokinetic sampling are shown in Table day3 and C [lum]day7 for all age groups and arms  was 13 200/857 (15.4-fold), 12 100/978 (12.4) and 10 700/692 (15.5) respectively for C [lum]day3 and 718/58 (12.4), 824/86 (9.6) and 870/101 (8.6) for C [lum]day7. Median supervised versus unsupervised C [lum]day3 values (ng/mL) were 7040 vs. 2700 (<5 y), 5965 vs. 3310 (5\u20134 y), and 5320 vs. 3490 (\u226515 y) . Corresponding median C [lum]day7 values were 330 vs. 156, 402 vs. 249, and 382 vs. 281 . C [lum]day3 and C [lum]day7 showed good correlation , with no differences across age groups or arms. 70/241 (29.0%) supervised and 140/238 (58.8%) unsupervised patients had a C [lum]day7 below 280 ng/mL.Lumefantrine concentration distributions were skewed to the right. There was marked inter-individual variability in C [lum] Figures  and 2. Uday3 .In multivariate linear models, unsupervised treatment had a strong negative association with C [lum]y3 Table . A histoday7 result, the probability of re-infection increased significantly as C [lum]day7 decreased, and no re-infections occurred in patients with levels \u2265400 ng/mL  and other explanatory variables. However, stratifying by age group, the only strong confounder detected by the Cox model, also showed a significant risk of re-infection with declining age  person-days, or 0.46 per person-year. All re-infections occurred on or after day 21, and 22/34 (64.7%) on day 28. After adjusting for age and arm, decreasing weight-adjusted lumefantrine dose was associated significantly with a higher hazard of re-infection, although confidence intervals were wide , principally by the liver and biliary system: children under five had significantly lower C [lum]day7, possibly because of enhanced metabolism and clearance. Interestingly, age was not a significant explanatory factor for the C [lum]day3 in this study, a contrasting finding with the A/L adherence study [day3 and dizziness at baseline with C [lum]day7 are harder to explain, and might be spurious results because of multiple significance tests. In Thai patients, age, weight, and gender were not associated with any of the main PK parameters but the trial did not include children of less than six years of age [C [lum]rst dose . As a clce study . The assDespite highlighting these interesting associations, on the whole our models did not satisfactorily explain the variance in C [lum]. Previous studies have shown that the large effect on lumefantrine PK of lipid intake, which our study was not designed to measure, far outweighs other factors .day3 results in our study are broadly consistent with those of Thai patients but lower than those in Chinese patients, even though patients in this study received the six dose regimen. Apart from food intake, pharmacogenetic factors  who in turn had higher values than European patients . Median tabolism ) and difIn the multi-drug resistant areas of western Thailand, a day 7 lumefantrine concentration below 280 ng/mL was a reasonable predictor of treatment failure. In this study, there were no treatment failures despite day 7 lumefantrine concentrations <280 ng/ml in 45% of all patients, a finding probably due to the high parasite sensitivity to lumefantrine in Uganda.By contrast, a day 7 lumefantrine concentration of <400 ng/ml and receiving a lower dose per Kg of lumefantrine were risk factors for re-infection during follow-up; this risk was thirteen-fold higher in patients receiving less than 65 mg/Kg compared to those receiving \u226580 mg/Kg. Most re-infections became visible in the blood 21 to 28 days after start of treatment, when lumefantrine levels would be expected to be very low, and sub-therapeutic. In our study almost 50% (134/286) of children under 5 years received less than 65 mg/Kg of lumefantrine; these children also experienced an additional age-related, eight-fold higher risk of re-infection compared to adults. Parasite re-infection has received far less attention than recrudescence. Nonetheless, in the era of highly efficacious ACTs like A/L, preventing re-infections may become an important public health issue, especially in children under five in stable high transmission areas and individuals during malaria epidemics; in both groups, re-infections cause high malaria morbidity and mortality. Our findings suggest that, in routine practice, low adherence and poor lipid intake could result in re-infection rates that would negate the benefits of high A/L efficacy.in vivo study extended to only 28 days. Had patients been monitored for 42 days, as recommended by some, we could have detected later recrudescences [This escences . The samde novo resistance to ACT combinations is clearly a crucial consideration that probably overrides the desirability of preventing novel infections through a post-treatment prophylactic effect [Despite the limitations of this study, it is clear that a high day 28 A/L cure rate can be achieved despite low plasma lumefantrine levels, even among unsupervised patients. This apparently favourable situation may, however, set up conditions for the selection of resistant parasites, especially in young children who not only harbour high biomass infections but also achieve the lowest day 7 lumefantrine levels. The prevention of c effect . In thisA/L is a precious tool in the available arsenal against malaria morbidity and mortality, especially in African children. Although it needs to be made widely available to populations in need, care should also be taken to avoid creating favourable conditions for the emergence of lumefantrine-resistant malaria strains . FurtherFC, PP, CF, FB, WRJT and JPG designed the trial. FC and PP analysed data. CF, FB, SB, FG, ER, JB and IK were in charge of trial recruitment and on-site supervision, and contributed to data analysis. JKi, JKy and LF designed and supervised laboratory procedures, with the exception of pharmacokinetic assays, and interpreted PCR data. FC, PP, WRJ and JPG interpreted findings and wrote this paper with the contribution of other authors. All authors read and approved the final manuscript."}
+{"text": "Paracheck Pf\u00ae, a malaria rapid diagnostic test (RDT) detecting histidine rich protein 2 was undertaken amongst children aged 6\u201359 months in eastern Democratic Republic of Congo.An assessment of the accuracy of This RDT assessment occurred in conjunction with an ACT efficacy trial. Febrile children were simultaneously screened with both RDT and high quality microscopy and those meeting inclusion criteria were followed for 35 days.358 febrile children were screened with 180 children recruited for five weeks follow-up. On screening, the RDT accurately diagnosed all 235 true malaria cases, indicating 100% RDT sensitivity. Of the 123 negative slides, the RDT gave 59 false-positive results, indicating 52.0% (64/123) RDT specificity. During follow-up after treatment with an artemisinin-based combination therapy, 98.2% (110/112), 94.6% (106/112), 92.0% (103/112) and 73.5% (50/68) of effectively treated children were still false-positive by RDT at days 14, 21, 28 and 35, respectively.Paracheck-Pf\u00ae is as sensitive as microscopy in detecting true malaria cases, a low specificity did present a high frequency of false-positive RDT results. What's more, a duration of RDT false-positivity was found that significantly surpassed the 'fortnight' after effective treatment reported by its manufacturer. Though further research is needed in assessing RDT accuracy, study results showing the presence of frequent false positivity should be taken into consideration to avoid clinicians inappropriately focusing on malaria, not identifying the true cause of illness, and providing unnecessary treatment.Results show that though the use of  In response to increased antimalarial drug resistance, the past decade has witnessed the introduction of artemisinin-based combination therapy (ACT) by many malaria-endemic countries. With the significantly higher cost of ACT over the previously used chloroquine and sulphadoxine-pyrimethamine, there has been a strong emphasis in avoiding any unnecessary use of ACT and minimizing opportunities for the development of parasite drug resistance. Though clinical diagnosis can be very sensitive, its limited specificity can lead to inappropriate treatment . As suchParacheck-Pf\u00ae , detects the histidine rich protein 2 (HRP2), a protein uniquely synthesized by Plasmodium falciparum and present in the bloodstream of an infected individual [Plasmodium species. The second group of RDT detects parasite lactate dehydrogenase (pLDH), an enzyme produced by all four human malaria species.Although there has been a marked increase in the number of RDTs commercially available, they can be roughly divided into two categories. One group of RDTs, including dividual . Some HRP. falciparum and can be less costly than the pLDH alternative, studies have shown that HRP2 remains in the bloodstream for an extended time following successful eradication of the parasite, contributing to false positive results and limited specificity. A study by Humar et al. (1997) on ParaSight F (early HRP2 version utilizing IgG antibodies) showed detectable levels of HRP2 in 27% of patients 28 days following successful treatment [Paracheck-Pf and two other RDTs, a 2002 study in Vietnam went further to report a strong correlation between extended duration of positivity and higher parasite density at presentation [Although HRP2 test kits have generally shown higher sensitivity for reatment . Though reatment ,6. Whilet al. 199 on ParaSParacheck-Pf\u00ae was assessed, comparing RDT diagnosis to high quality microscopy during initial screening and follow-up.To date, no comprehensive study of persistent HRP2 antigenaemia has been published from sub-Saharan Africa. Therefore, coinciding with an ACT efficacy study, the accuracy of Plasmodium falciparum accounts for 95% of the plasmodium species in this region. Study approval was obtained from the ethics committee of the DRC National Malaria Programme and the external Ethics Review Board used by M\u00e9decins Sans Fronti\u00e8res.This RDT assessment occurred parallel to a 28-day ACT efficacy trial that followed WHO guidelines . In brieP. falciparum malaria confirmed by microscopy, unmixed P. falciparum infection between 2,000 \u2013 200,000 parasites per \u03bcL of blood, no concomitant disease that could mask the response to antimalarial treatment) were recruited after their guardians provided written informed consent. Children were randomized to receive artesunate + amodiaquine or artesunate + sulphadoxine-pyrimethamine. Children were followed to assess both ACT efficacy and duration of false positive RDT results, as defined by positive RDT and continued negative microscopy after ACT treatment. For the purposes of the RDT assessment, follow-up was performed by study clinicians on days 7, 14, 21, 28 post-treatment or on any other day if the child was unwell. An additional visit at day-35 was added specifically to continue monitoring RDT results.The RDT assessment was performed from March to June 2004. After screening of febrile children six to 59 months of age, 180 children who met inclusion criteria for the ACT efficacy study , difference in malaria species, or a difference in parasite density greater than 25%. External controls were conducted by a Ministry of Health government reference laboratory in the provincial capital.P. falciparum and presence of only the control line indicated a negative result. In any case where the control line did not appear, the result was considered invalid and the test was repeated. The person recording the RDT result was unaware of corresponding microscopy results. Internal quality control included an immediate blind second reading of 100% of RDTs. In cases where there was discordance, the two technicians re-examined the RDT together and made a collective decision on the reading.Concurrent with blood smear collection, RDTs were completed with the same finger prick blood sample at time of screening and follow-up. Test kits were stored as directed by the manufacturer and quality of package desiccant was checked before use. The fresh blood sample was transferred directly to the sample pad by the provided sample applicator. All RDTs were labelled with patient ID numbers and results were recorded 15 minutes after adding 6 drops (300 \u03bcL) of clearing buffer. Presence of both the control and test lines indicated a positive result for Data was double-entered into Excel (Microsoft XP) and transferred to STATA  for further analyses. Data included in analysis for duration of false positive RDT results is only from children who remained slide-negative until at least 28 days after treatment.P. falciparum infection with both RDT and microscopy  of the RDT was 79.9% , i.e. 79.9% of the 294 positive RDT results were matched by positive microscopy results ; one child was lost to follow-up due to family movement; 20 children were not able to complete a day-28 clinic visit after forced evacuation of the study team due to political insecurity in the region. This evacuation prevented an additional 44 children from having a day-35 visit. In summary, for day-28 and day-35 clinic visits, 112 and 68 children attended, respectively.At days 7, 14, 21, 28, and 35 after effective treatment, 99.1% , 98.2% , 94.6% , 92.0% , and 73.5%  of the patients attending a clinical visit were still false-positive by RDT, respectively  showed that false positive results can be explained by the presence of blood samples with parasite density levels below the detection threshold for microscopy [Using PCR, Bell croscopy . Such suThese study results show that the duration of Paracheck false positivity can be more than 35 days after effective treatment. As mentioned in the introduction, past evidence strongly suggests that this is due, in large part, to the prolonged time it takes for HRP2 to be cleared from the blood following treatment of falciparum malaria. Though the mechanism of HRP2 clearance is not well understood, there are several feasible explanations for its long persistence after adequate therapy, as discussed below.Duration of RDT false positivity has previously been correlated to higher parasite density on admission. As secretion of the protein is proportional to parasite numbers , a higheP. falciparum requires a longer time (about 8\u201310 days) than for other malaria species [HRP2 is also released by early gametocytes  and deve species . Thus peEarlier reports suggested that persistence of HRP2 may result from a cross-reacting auto-antibody such as rheumatoid factor (RF) present in the blood . HoweverPersistent antigenaemia could result from slow or incomplete clearance of circulating or sequestered sensitive parasites. After treatment, some patients with relatively resistant parasites might have persistent sub-patent parasite density (<100 parasites/\u03bcl) and a recrudescence of their infection after day 28 or 35. However, late recrudescence is unlikely to have happened in such a large proportion of cases as the artemisinin-based drugs currently display near 100% efficacy. Singh and Shukla .A duration of Paracheck-Pf false-positivity was found that significantly surpassed the 'fortnight' suggested by its manufacturer. Though other studies have shown the same trend, none have shown such a high proportion of RDTs still positive 28\u201335 days after parasite clearance. While further studies are needed, these results reinforce the need for medical staff to incorporate a patient's clinical history when interpreting Paracheck results. This is especially true for patients with routine clinical visits that include malaria screening and in areas with high malaria endemicity.TDS Responsible for study design, field-implementation, supervision of the team in the field, data collection, analyses, writing of and finalising this paper. HC Responsible for supporting data analysis and writing of finalised paper. RKKS Contributed to laboratory protocol design and supported supervision of laboratory team. IvdB Contributed to study design, overall supervision of the study, and supported data analysis and writing of this paper. All authors read and approved the final manuscript."}
+{"text": "Our aim was to evaluate the safety, reactogenicity, and immunogenicity of an investigational malaria vaccine.This was an age-stratified phase Ib, double-blind, randomized, controlled, dose-escalation trial. Children were recruited into one of three cohorts (dosage groups) and randomized in 2:1 fashion to receive either the test product or a comparator.The study was conducted in a rural population in Kombewa Division, western Kenya.Subjects were 135 children, aged 12\u201347 mo.Imovax\u00ae rabies vaccine).Subjects received 10, 25, or 50 \u03bcg of falciparum malaria protein 1 (FMP1) formulated in 100, 250, and 500 \u03bcL, respectively, of AS02A, or they received a comparator (We performed safety and reactogenicity parameters and assessment of adverse events during solicited (7 d) and unsolicited (30 d) periods after each vaccination. Serious adverse events were monitored for 6 mo after the last vaccination.3,1047 = 10.78, or F3, 995 = 11.22, p < 0.001); however, the comparison of 25 \u03bcg and 50 \u03bcg recipients indicated no significant difference .Both vaccines were safe and well tolerated. FMP1/AS02A recipients experienced significantly more pain and injection-site swelling with a dose-effect relationship. Systemic reactogenicity was low at all dose levels. Hemoglobin levels remained stable and similar across arms. Baseline geometric mean titers were comparable in all groups. Anti-FMP1 antibody titers increased in a dose-dependent manner in subjects receiving FMP1/AS02A; no increase in anti-FMP1 titers occurred in subjects who received the comparator. By study end, subjects who received either 25 or 50 \u03bcg of FMP1 had similar antibody levels, which remained significantly higher than that of those who received the comparator or 10 \u03bcg of FMP1. A longitudinal mixed effects model showed a statistically significant effect of dosage level on immune response  on the surface of the blood-stage infectious form of the malaria parasite. In early-stage clinical trials performed in people not exposed to malaria  and in African adults who were exposed to malaria, this candidate vaccine has already been shown to be safe and to bring about an immune response. As part of the next stage in developing this vaccine, a group of researchers next wanted to see whether the vaccine was also safe and brought about an immune response in the population most in need of a vaccine: young children living in an African region with very intense malaria transmission. Therefore, as reported here, this group performed a small trial in western Kenya, recruiting 135 children under 5 y of age to receive either the FMP1/AS02A vaccine (at three different doses) or rabies vaccine for comparison . The outcomes that the researchers were interested in were primarily adverse events, which they categorized using a standard questionnaire at up to 7 d after vaccination; unsolicited events reported up to 30 d after vaccination; and, finally, any serious events occurring up to 8 mo later. The researchers also examined antibody responses to the FMP1/AS02A vaccine.What this trial shows: Participants who received the FMP1/AS02A vaccine (as compared to the rabies vaccine) experienced more immediate symptoms, such as pain and swelling at the injection site. Most participants reported unsolicited events during follow-up, but the proportion of participants with adverse events did not seem to be different between the FMP1/AS02A vaccine groups and the rabies vaccine group. Unsolicited outcomes that were reported included, for example, clinical malaria, upper respiratory tract infections, and a few events that were thought to be related to the vaccines, such as fever and eczema. A few serious adverse events occurred up to 8 mo after vaccination, but the numbers did not seem to be different between the FMP1/AS02A and rabies vaccine groups, and the events were not judged to be related to vaccination. Finally, participants who received the FMP1/AS02A vaccine raised an antibody response to the vaccine, which was highest in those who received the highest vaccine dose. The researchers concluded that this vaccine was safe and brought about an immune response in the group of malaria-exposed children studied.Strengths and limitations: The trial was conducted in a population that is likely to benefit from the vaccine, if it is shown to be effective in further studies. Therefore, the data obtained from this study will be informative in helping to design future trials on FMP1/AS02A. The randomization procedures used in this study were appropriate, and in particular participants of different ages were equally distributed to the different intervention groups, helping to minimize bias. Procedures were also set up to prevent participants and staff giving the vaccines and collecting data from knowing which interventions participants had received. However, the number of participants recruited into the trial was small, and it therefore was not powered to detect anything other than large differences in rates of adverse events between the study groups.Contribution to the evidence: This study extends evidence from prior trials on the safety and immunogenicity of the FMP1/AS02A vaccine to a population that is representative of those most in need of an effective vaccine\u2014young African children. The results suggest that the vaccine candidate should undergo further evaluation in trials examining vaccine efficacy in a similar population. Plasmodium falciparum infections occur worldwide, leading to between 1 and 2 million deaths in sub-Saharan Africans\u2014most of them children [P. falciparum merozoite surface protein 1 (MSP-1) is a 195-kDa protein that is proteolytically cleaved to yield four fragments, which are associated with each other through noncovalent interactions on the merozoite surface. Among them is the carboxy-terminal 42-kDa fragment known as MSP-142 [42 and anti-MSP-119 antibodies that inhibit parasite growth in vitro [Aotus monkeys with a recombinant MSP-142 and potent adjuvant can confer protection against blood-stage challenge with P. falciparum [42 as a potentially efficacious malaria vaccine is further supported by epidemiologic studies demonstrating that antibodies to the relatively conserved domains are associated with a diminution of P. falciparum disease severity [The world struggles to come to terms with the immensity of the public health, economic, and political consequences of malaria morbidity and mortality. Although no inhabited continent is unaffected, Africa is particularly hard hit. Annually, over half a billion children . The prochildren . Nearly children , and a pchildren ,7. An alchildren ,9. The P MSP-142 ,11. Seco MSP-142 . Individin vitro . Moreovelciparum . In humaseverity  and by tseverity .42 of the 3D7 clone of P. falciparum has been formulated into a final test product, termed the falciparum malaria protein 1 (FMP1) [The MSP-11 (FMP1) , and com1 (FMP1) . The saf1 (FMP1)  and in p1 (FMP1)  and MaliAnopheles gambiae complex and the infecting parasite species is usually (95%) P. falciparum [The trial was conducted in Kombewa Division in the Kisumu District of Nyanza Province of western Kenya. The study subjects were drawn almost exclusively from the Luo ethnic group living in low-land country near Lake Victoria. The Luo are settled agriculturalists of Nilo-Saharan origin, most of whom engage in low-income subsistence farming or fishing. In this region of holoendemic malaria transmission, most infections (90%) are transmitted by mosquitoes of the lciparum ,24. Tranlciparum  and montData from a previous comprehensive census of Kombewa Division, as well as from a more focused census done immediately prior to recruitment, were used to identify potential field stations. This fairly comprehensive demographic database established how many eligibles (on the basis of age) were present in the areas to be drawn from. All parents known to have children in the relevant age groups and living within a 1-mile (1.6-km) radius of each of 12 field stations were invited to participate in a briefing.>45 IU/L; serum creatinine of > 1.1 mg/dL; absolute lymphocyte count (ALC) for 1-y-olds of < 4.0 \u00d7 103/mm3, for 2-y-olds of < 3.0 \u00d7 103/mm3, and for 3-y-olds of < 2.0 \u00d7 103/mm3; platelet count of <100,000/mm3; hemoglobin of < 8 g/dL; homozygosity for sickle cell disease (SS) genotype (by protein gel electrophoresis); and malnutrition defined as weight for height of < \u22123 z-scores.Prior to enrollment in this study, a medical history, physical examination, and screening laboratory tests were performed on every subject to detect any underlying medical condition that might confound clinical evaluation or data analysis. Males and females were considered eligible for enrollment if they were 12\u201347 mo old at the time of screening and in good health and if a parent agreed to sign a written and witnessed informed consent form and to remain in the study area for 12 mo. Exclusion criteria included any evidence of acute or chronic medical conditions that might increase the risk to the participating subject. Specific exclusion criteria included prior receipt of an investigational malaria vaccine or of a rabies vaccine; recent or planned use of any investigational new drug (IND), vaccine, immunoglobulin, or any blood product; use of immunosuppressant drugs within the previous 6 mo; confirmed or suspected immunodeficiency; history of splenectomy; administration, or anticipated administration, of a vaccine not foreseen by the study protocol within 30 d of the first dose of vaccine (with the exception of tetanus toxoid); and concurrent participation in another clinical trial. Laboratory exclusion criteria were findings of serum alanine aminotransferase (ALT) of Participants were recruited under a human use protocol approved by and executed in accordance with the guidelines of the Office of the Surgeon General, United States Army; the Ethics Review Committee of the Kenya Medical Research Institute ; and the Human Subjects Protection Committee of the Program for Appropriate Technology in Health (PATH). Informed consent was obtained from all participants in accordance with all applicable guidelines.Escherichia coli\u2013produced, GMP-manufactured, FMP1 antigen have been described elsewhere [The expression, purification, biochemical, and immunological characterization of the lsewhere , as has lsewhere . ImmediaImovax\u00ae Rabies, produced by Aventis Pasteur, Lyon, France), has been described elsewhere [The comparator vaccine, an inactivated rabies vaccine  was appointed before the study began to review both the safety data reports as the trial progressed and the data analysis by Statistics Collaborative after study completion. DSMB membership included a statistician and four senior clinical research investigators with experience in conducting malaria vaccine trials, one of whom was a Kenyan. An experienced local clinician  served as the local medical monitor (LMM), reviewed all serious adverse events (SAEs) and safety data between dose escalations, and functioned as the patient advocate. In addition, the trial was monitored for regulatory compliance by representatives of the United States Army Medical Materiel Development Activity, GlaxoSmithKline Biologicals, and Pharmaceuticals Product Development , all of whom made several visits to the study site. Initial approval of the study protocol and of subsequent protocol amendments was granted to the investigators by the Ethical Review Committee of the Kenya Medical Research Institute, Nairobi; by the Human Subjects Protection Committee of PATH, Seattle, Washington, United States; and by the United States Army HSRRB. The study was done under a Food and Drug Administration IND and was compliant with all relevant International Conference on Harmonization guidelines.Vaccinations of the second and third cohorts were staggered from each other by 2 wk . A safetStopping rules were to be invoked when either of the following observations was made: (1) > 20% of subjects with \u201csevere\u201d (grade 3) general AEs  related to the vaccine; or (2) any SAE, including death, judged to be vaccine related.The primary objective was to assess the safety and reactogenicity of the FMP1/AS02A malaria vaccine in malaria-exposed 12- to 47-mo-old children living in western Kenya. The secondary objective was to assess the humoral immune response to the FMP1/AS02A malaria vaccine in malaria-exposed 12- to 47-mo-old children living in western Kenya.42 3D7 strain antibody) titers as determined by an enzyme-linked immunosorbent assay (ELISA) on study days 0, 14, 30, 44, 60, 74, 90, 180, 270, and 364.The primary endpoints were (1) occurrence of solicited symptoms (based on a standardized questionnaire) during a 7-d follow-up period after each vaccination ; (2) occurrence of unsolicited symptoms during a 30-d follow-up period after each vaccination; and (3) occurrence of SAEs during an 8-mo follow-up period following the first dose of study vaccine . The secondary endpoints were anti-FMP1  or until resolution. Both local (injection-site pain and swelling) and general/systemic  symptoms were assessed . After tA peripheral blood smear was obtained from any subject who presented to the Walter Reed Project's Kombewa Clinic with fever, a history of fever within 48 h, or an illness that the attending doctor suspected might be due to malaria infection. After Giemsa staining and examination by oil-immersion light microscopy, detection of asexual parasitemia of > 0 parasites/\u03bcL resulted in the diagnosis and treatment for malaria.Immunology samples were collected on study days 0, 14, 30, 44, 60, 74, 90, 180, 270, and 364; samples collected on study days 0, 30, and 60 were collected immediately prior to vaccination. Immune response to the FMP1/AS02A vaccine was determined by anti-FMP1 ELISA endpoint titers reported in optical density units (ODUs), the dilution yielding an ODU of 1.0 in our assay. This assay has been described in detail elsewhere .Imovax\u00ae Rabies vaccine comparison group enabled broad initial estimates of the incidence of local and general side effects in a population that suffers from significant comorbidity from exposure to endemic illnesses. The control cohort also served as a comparator for the longitudinal immune responses to the FMP1 antigen in a malaria-exposed population. Although comparative statistics for the safety variables were calculated, the study had low power to detect anything other than large differences in the incidence of local and general side effects between the vaccination groups.This trial represents the first time to our knowledge that this vaccine candidate has been evaluated in a pediatric population. Sample size was chosen after weighing the need to detect any possible vaccine associated AEs against the need to limit the number of subjects exposed to an investigational product. Incorporation of the With sample sizes of 15, 30, and 45 , we have 90% power to detect an AE that occurs in 14%, 7%, and 5% of the population, respectively. The study was not powered to detect an immune response because the primary focus of this trial was safety.A randomization list generated by Statistics Collaborative contained sequential codes linked to a study vaccine assignment. These codes were assigned to subjects in the order in which they presented to the clinic on the day of first vaccination. Blocked blinded randomization was used with stratification for age and dosage groups. Fifteen subjects from each age group were randomized into each of the three cohorts (10 receiving the test article and five receiving the comparator). Because of the very small sample sizes , these sThe only persons at the study site with access to the randomization assignments were the study drug manager, the clinic pharmacist, and his assistant; it was necessary that these individuals have access to the codes for preparation of test articles. Each randomization assignment was sealed in a unique, tamper-evident envelope, which was opened at the time a subject presented for the first vaccination. The LMM also kept one set of the randomization codes in a sealed envelope in the event that emergency unblinding became necessary.Subjects were randomized in the order in which they presented on the first day of vaccination in a 2:1 ratio to receive either FMP1/AS02A (90 subjects) or the comparator vaccine (45 subjects). The subjects were enrolled by the study drug manager, clinic pharmacist, and his assistant, and these individuals assigned the subjects to their groups.Because the color and volumes of the reconstituted FMP1/AS02A and comparator vaccines differed, the barrel of the syringe was covered with opaque tape to mask its contents and labeled with the subject identification number and randomization code. Subjects, parents, and the staff performing follow-up evaluations were all blinded. Immunizations were carried out simultaneously in four separate consultation rooms that were connected to a central pharmacy (the vaccine preparation room) by small, closable service hatches. On vaccination days, the prepared syringe was handed through a service hatch to a vaccinator for vaccine administration. For each subject, an identification number, a randomization code from a chart, and a randomization code on the syringe were recorded on a vaccination form. Following vaccine administration, subjects were assessed and follow-up visits conducted by a group of clinicians who had not been involved in the vaccinations. The study investigators became unblinded to treatment allocations in July 2005, after study completion (which was September 2004).10 scale and were modeled by longitudinal mixed models to assess the effect of dose and age on the mean level of the antibody responses over time. The models used a spatial covariance structure, which takes into account the number of study days between two measurements when determining the correlation between them.Statistical analyses were performed by Statistics Collaborative using SAS version 8.2 . The incidence of solicited, unsolicited, and serious AEs were compared using two-sided Fisher's exact tests without correction for multiplicity. Geometric mean titers (GMTs) were calculated to assess immunogenicity at each timepoint when titers were collected. The titer data were transformed to a logA total of 590 parents of children were briefed; of these, 436 parents consented to screening. Of the 320 children who were returned for screening, 135 (77 girls and 58 boys) were enrolled and randomized to one of the three dosage cohorts. Among these were 25 children who were initially disqualified owing to clinical malaria; however, they were subsequently enrolled after successful treatment and confirmed cure. Of the 135 subjects who received the first vaccination, five did not receive the second vaccination and another eight did not receive the third. Thus, 122 subjects received all three vaccinations; 83 of the 90 subjects were randomized to receive FMP1/AS02A and 39 of the 45 randomized to receive the rabies vaccine. The 13 incompletely vaccinated subjects were evenly distributed among the three dose cohorts . BecauseRecruiting and enrollment occurred from 25 July through 12 September 2003. The study duration was approximately 12 mo for a subject with the last study visit on 19 September 2004.The study groups were comparable in baseline demography, height, weight, and vital signs . Baseline clinical laboratory measurements other than ALC were consistent across study arms; most measurements fell within the local normal range. Both study arms had a number of subjects outside of the normal range for ALC at the sampled timepoints, including prior to receipt of the first vaccination, but none of these was deemed clinically significant. Age, sex, height, weight, clinical laboratory values, and antibody to FMP1 prior to the first vaccination are presented in This study randomized 135 children (aged 12\u201347 mo) into three cohorts of 45 subjects, each consisting of 30 children who received FMP1/AS02A  and 15 children who received the comparator vaccine. Each cohort contained 15 subjects, 10 receiving FMP1/AS02 and five the comparator, in each of three age groups  for a total of 45 subjects in each age group distributed among the three cohorts. The comparator groups received rabies vaccine . Each sup-value, < 0.001); the percentages of subjects experiencing local symptoms during second and third vaccinations were lower in both study arms . The most common local reaction at any time was pain at the site of injection. Up to 38% of subjects receiving FMP1/AS02A (percentages varied by dosage group) also experienced injection-site swelling; however, no subject receiving the comparator experienced swelling.Both the test article and comparator vaccines were well tolerated. No parent or child withdrew from the study for a vaccine-related side effect. No sequence- or dose-related trends were apparent for general solicited symptoms . The mosFew subjects experienced grade 3 symptoms . Grade 3Unsolicited symptoms were recorded for 30 d following each vaccine administration and were categorized by a modified World Health Organization Adverse Reactions Terminology (WHOART) AE coding system. There were no differences by group or by cohort in the proportions of subjects experiencing an unsolicited symptom. All but one of the enrolled children experienced at least one unsolicited symptom during a follow-up period, and most experienced an unsolicited symptom during each of the three follow-up periods (77%\u201397% for FMP1/AS02A and 80%\u2013100% for the comparator). The most common unsolicited symptoms were upper respiratory tract infections (URTIs) and malaria.Approximately 80% of subjects in each study arm experienced at least one URTI during the three 30-d postimmunization follow-up periods. There was no indication that vaccinated groups had any increased risk of developing clinical malaria.For unsolicited symptoms, vaccine relatedness was determined by temporal relationship to a vaccination with absence of any reasonably explanatory comorbidity. Very few vaccine-related unsolicited symptoms occurred during the postimmunization follow-up period . These symptoms were mostly limited to injection-site reactions that differed from those elicited during the immediate follow-up period. They included a case of hyperpigmentation in the comparator group and three cases of fever by history, one of induration, and one of eczema in the FMP1/AS02A group. The last mentioned was a nonpruritic skin rash developing in the right flank area about 50 min after a second vaccination of the 10-\u03bcg dose of FMP1/AS02A. Initially presenting with a plaque-like appearance, these lesions developed a follicular eczematous pattern that then resolved over a few days. This child had been noted to have similar  dermatitic or eczematous lesions prior to the first vaccination. A third vaccination was withheld from this subject. One grade 3 unsolicited symptom occurred during the postimmunization follow-up period, a 25-\u03bcg FMP1/AS02A subject experiencing grade 3 malaria beginning 2 d after receiving the first vaccination.  No subject in the study experienced a grade 3 vaccine-related, unsolicited symptom.Serious and unexpected AEs were collected on a real-time basis through study day 240. No SAE was judged causally related to a vaccination. Seven documented SAEs occurred during this period: four in the FMP1/AS02A group and three in the comparator group. In the former group, two children in the 10-\u03bcg dosage cohort experienced convulsions 17 and 189 d after second and third vaccinations, respectively; one child in the 50-\u03bcg dosage group experienced convulsions 12 d after the third vaccination. A 37-mo-old female acquired malaria 26 d after receiving a third dose of 10 \u03bcg of FMP1/AS02A, was treated, and recovered. She developed hepatitis and severe anemia 7 d later. She died 62 d after her third vaccination, following several hospitalizations and blood transfusions. In this subject, an extravascular hemolysis caused severe organomegaly with secondary thrombocytopenia and hypersplenism, along with a possible partial obstruction of biliary outflow. The death of this subject led to a thorough review of the case by the principal investigator, the DSMB, and a pediatric hematology consultant not previously associated with the study. After considering all potential etiologies  that could have precipitated such a clinical course and the clinical context , it was concluded that there was not a likely causal biological connection between the immunization and the preterminal course and death. In accordance with the study protocol, the principal investigator's judgment was that \u201cthere are other, more likely causes and administration of the study vaccine is not suspected to have contributed to the adverse event.\u201d Her death was judged not related to the study vaccine, but rather to autoimmune hemolytic anemia, most likely secondary to a viral infection, causing first hepatitis and then an unusual warm reactive immunoglobulin G. The clinical course and the temporal association are here reported so that in the event of similar instances, this information will be readily available.In the comparator group, one subject experienced febrile, malaria-associated convulsions; one, an episode of severe bronchospasm; and one, an intestinal obstruction due to ascariasis. One unexpected AE occurred during the study: a subject from the comparator group experienced a markedly elevated ALT level and was diagnosed with hepatitis A. The subject was excluded from the third vaccination to eliminate the possibility of confounding the assessment of safety following subsequent vaccinations. In addition, two subjects from the FMP1/AS02A group developed phimosis, leading to elective hospitalizations and circumcisions.Blood (venous samples from arms) for laboratory measurements  was collected at baseline (study day 0) and on study days 14, 30, 44, 60, 74, and 90. Group mean safety laboratory values were generally unchanged in the 30-d postimmunization follow-up periods. The majority of subjects had all values within normal range. However, both vaccine and comparator groups had a number of subjects outside of the normal range of ALCs, including timepoints prior to first vaccination; none was judged to be clinically significant.The protocol specified that each child was to receive 10 hemoglobin determinations occurring at the seven timepoints cited above and on study days 180, 270, and 364. None of the 90 children who received FMP1/AS02A had a hemoglobin level below the local lower limit of normal ; one of the 45 children enrolled in the comparator arm fell below this limit at one timepoint.As expected of a subject population that had been highly exposed to malaria prior to vaccination, no baseline anti-FMP1 titer fell below the limit of detection. Baseline GMTs were comparable across study arms and dosage cohorts, ranging from approximately 1,000 to 3,000 ODUs. The overall baseline GMT in thousands  was 2.3 . After three vaccinations, no increase in antibody response was observed in subjects who received the comparator; however, antibody response increased with increasing dosage level of FMP1/AS02A . GMT peaTiter ratio plots indicate the percentage of subjects experiencing specific fold rises in antibody response over baseline values . At stud10-transformed values of anti-FMP1. All subjects were included in the model (data not shown). Tests of main effects (dose and age group), the age group \u00d7 dosage level interaction, and dose response were performed. The results from the model for all randomized subjects showed the immune response differed among dosage levels . Neither the age group \u00d7 dosage level interaction nor the age group main effect is statistically significant , providing no evidence that a subject's age influences the vaccine's immunogenicity over time. A highly significant linear trend in dose response was observed . The 50-\u03bcg dosage group had a higher response than the 10-\u03bcg dosage group ; however, the 25- and 50-\u03bcg dosage groups did not show a significant difference .To assess the effect of dose and age group on the mean level of antibody responses over time, we estimated least-square means and standard errors of logThis phase Ib dose-escalation and safety trial provided clear evidence of the safety and tolerability of 10-, 25-, and 50-\u03bcg doses of FMP1/AS02A when given to young children subject to intense malaria transmission in western Kenya. Although subjects allocated to the test article arm experienced more solicited local symptoms, the proportion affected is comparable to the symptoms seen with another vaccine antigen formulated with the same adjuvant system . The freThe 25- and 50-\u03bcg doses generated a humoral immune response that was of greater magnitude than the 10-\u03bcg dose in this pediatric population. The previous FMP1/AS02A adult study at Kombewa  found a 10-transformed linear modeling for all randomized subjects showed statistically convincing evidence of a dose-response relationship to antibody response.The waning of antibody levels in the 25- and 50-\u03bcg dosage groups after study day 74 is congruent with previous experience with FMP1. Significantly, by study day 364, the 25- and 50-\u03bcg dosage groups maintained higher antibody levels than subjects in the 10-\u03bcg dosage and comparator groups. It is not clear what effect boosting from natural exposure had on the magnitude and persistence of the antibody response, but the anti-FMP1 response in the rabies comparator group did not significantly increase over the year-long observation period. The results from the logConsistent with our ultimate goal of allowing administration of a malaria vaccine as part of the World Health Organization's Expanded Programme on Immunisation (EPI) to infants at greatest risk of malaria, we have here followed up the previous year's trial of FMP1/AS02A in adults in Kombewa  with anoAotus monkeys subjected to prolonged P. falciparum parasitemia have raised concern that malaria vaccines eliciting immunity that controls, but does not eliminate, parasitemia might themselves increase the risk of anemia in endemic human populations [Reports of experimentally induced, malaria-associated anemia in ulations . In the P. falciparum infections at levels obtaining in the study area, but may not be generalizable to children under other transmission intensities.The study population selected for this trial was chosen as representative of the target population for a malaria vaccine: at-risk, malaria-experienced children living in an area of endemic, holoendemic, or epidemic malaria. As the primary objective of the trial centered upon evaluation of safety and reactogenicity, results should be broadly generalizable to children of the representative age groups. Results pertaining to the secondary objective  should be generalizable to children of these age groups exposed to Along with the previous year's adult trial, this trial accomplishes the first two of the three immediate goals of the clinical development plan  for FMP1CONSORT ChecklistClick here for additional data file.(255 KB PDF)Trial ProtocolClick here for additional data file.(1.0 MB DOC)"}
+{"text": "Plasmodium falciparum often leads to therapeutic failure in Indonesia. Combining CQ with other drugs, like SP, may provide an affordable, available and effective option where artemisinin-combined therapies (ACT) are not licensed or are unavailable.Chloroquine (CQ) or sulfadoxine-pyrimethamine (SP) monotherapy for This study compared CQ (n = 29 subjects) versus CQ + SP  for clinical and parasitological cure of uncomplicated falciparum malaria in the Menoreh Hills region of southern Central Java, Indonesia. Gametocyte clearance rates were measured with (n = 56 subjects) and without (n = 61) a single 45 mg dose of primaquine (PQ).Msp-2 genotyping permitted reinfection-adjusted cure rates for CQ and CQ combined with SP, 70% and 99%, respectively (p = 0.0006).After 28 days, 58% of subjects receiving CQ had cleared parasitaemia and remained aparasitaemic, compared to 94% receiving CQ combined with SP (p < 0.001). P. falciparum parasitaemia in this study.Primaquine exerted no apparent affect on cure of asexual stage parasitaemia, but clearly accelerated clearance of gametocytes. CQ combined with SP was safe and well-tolerated with superior efficacy over CQ for  Chloroquine (CQ) was the mainstay of antimalarial therapy from 1946 until the past decade when parasite resistance rendered it clinically ineffective in most areas of the world. Adult CQ therapy costs less than $0.20, is widely available almost anywhere in endemic areas, and has a generally good safety and tolerability profile. Resource-strapped health agencies in endemic areas struggle with the decision to abandon this drug. Alternative drugs, such as artemisinin combination therapies (ACT), carry significant limitations linked to cost, ease of compliance and safety in vulnerable populations like infants and pregnant women. Combining antimalarials of differing mechanisms of action may diminish risk of onset of parasite resistance[Plasmodium falciparum would require little more than a modification of malaria therapeutic policy and practice. In contrast, most other combined therapy options would require licensing, acquisition, socialization, and carry unfamiliar and perhaps more severe risks linked to safety or compliance. This is an especially salient problem for pregnant women and young children, for whom almost no safety data exists.CQ combined with sulfadoxine-pyrimethamine (SP) has been proven safe and effective in clinical trials Table  and has P. falciparum is prevalent [P. falciparum infections are resistant to CQ and 22% resistant to SP [Clinical data for CQ combined with SP is still lacking in many countries, including Indonesia, where CQ and SP-resistant revalent -6. This nt to SP . Since sThe study was conducted between July and October 2001 in Purworejo District in the Menoreh Hills near the southern coast of Central Java, a region experiencing resurgent malaria at that time. The social, economic and demographic characteristics of malaria in the region are described elsewhere ,7,8. Fal\u00ae, St. Louis, MO USA), and if female of child-bearing potential, a urine human chorionic gonadotropin (hCG) test . Inclusion criteria included age \u2265 15, asexual stage P. falciparum parasite density \u2265 400/\u03bcl on Giemsa thick smear examined by standard microscopy, and availability for 28-day follow-up. Exclusion criteria included pregnancy, breast feeding, body weight < 40 kg, G6PD deficiency, history of antimalarial or antibiotic consumption during the previous 7 days, severe or complicated malaria, history of allergy or adverse reaction to study medications, and P. vivax or mixed species infection. Any parasitemic potential study subject not meeting these criteria was referred back to the cooperating clinic for standard therapy. Those patients deemed to meet enrollment criteria and willing to participate were assigned a sequential study subject code by the screening physician. The study subject codes were pre-assigned to treatment arms by a random process and all treatment was pre-packaged accordingly.Subject enrollment is depicted in Figure \u00ae; chloroquine diphosphate tablets 150 mg base; PT Bayer, Indonesia) in three once daily doses of 10, 10 and 5 mg base/kg on days 0,1, and 2 respectively; SP  in a single dose on day 0; Primaquine phosphate  in a single dose of 3 tablets on day 0 or day 2.Subjects were randomized to one of four treatment arms: (A) CQ 25 mg base/kg (body weight) over three days, (B) CQ 25 mg/kg over three days + SP 25/1.25 mg/kg single dose, (C) CQ 25 mg/kg over three days + SP 25/1.25 mg/kg single dose + primaquine 45 mg single dose on day 0, or (D) CQ 25 mg/kg over three days + SP 25/1.25 mg/kg single dose + primaquine 45 mg single dose on day 2. Study investigators directly observed and documented administration of each dose of medication: CQ  to assess symptoms, clinical recovery, adverse events and to obtain finger-stick blood samples for preparation of Giemsa-stained blood smears. Subjects complaining of illness on any day of follow-up were immediately brought to clinic and evaluated. Blood blot specimens (100 \u2013 200 \u03bcl) were also prepared on days 0, 2, 7, 14, 21, 28 or day of recurrent parasitaemia from fingerstick blood samples collected by heparinized, 100 \u03bcl micro-capillary tubes and expelled onto Whatman No. 1 filter paper  for drying and storage for later analysis.P. falciparum and other plasmodial species, and recorded sexual and asexual parasite densities as number of parasites per 200 white blood cells. Parasite densities are reported as parasites/\u03bcL assuming an average white blood cell count of 8,000/\u03bcl.The microscopists in this study were certified as competent by a standardized, proctored examination. Microscopists examined all ocular fields of standard Giemsa  stained thick smears at high power (1000\u00d7 oil immersion) for asexual and sexual forms of Msp-2 genotyping was performed on blood blot samples from day 0 and day of recurrent P. falciparum parasitaemia following Felger et al. [r et al. . Recrudemsp-2 genotyping were reclassified as therapeutic successes with abbreviated follow-up (until the point of censure). These results were used to determine the \"adjusted\" efficacies of CQ and CQ + SP. EpiInfo 2000 1.1.2  and SPSS version 9.0  were used to calculate confidence intervals for odds ratios (OR) and the 95% confidence interval around those ratios, and for estimating the p-value using the Yates' corrected or Fisher's exact test when appropriate. Cumulative incidence of therapeutic failure was estimated by actuarial analysis as described elsewhere [Therapeutic efficacy  was defined as the proportion of subjects reaching day-28 without recrudescence. Therapeutic failures occurred when asexual parasitaemia increased between day 0 and day 2, failed to clear by day 4, or recurred between days 4 and 28. These data were used to determine the crude therapeutic efficacy of each regimen. Infections initially classified as therapeutic failures but later proven to be reinfections by lsewhere .Among 124 parasitaemic persons identified during mass blood screening and passive case detection from outpatient clinics, 117 were enrolled and randomized to one of the 4 treatment regimens. Seven subjects did not meet study eligibility criteria and were returned to the referring clinic for treatment for the following reasons: taking quinine (1), pregnancy (1), G6PD deficiency (1), low body weight (2), declined participation (1) and administrative error (1).Among the 117 subjects enrolled and randomly assigned, no statistically significant demographic differences between the four study arms were identified; age ranged from 16 to 65 years, and the male to female ratio was 1.2:1  subjects cleared parasitaemia by day 3 regardless of the therapy assignment, and all 38 subjects with fever prior to therapy had a normal axillary temperature by day 2 , so these groups were combined specifically for comparison with CQ monotherapy against asexual blood stages.P. vivax during the second and third week of post-therapy observation. Fourteen (48%) completed the study without recurrent parasitaemia. The remaining 11 (38%) subjects developed recurrent P. falciparum during the 28-day follow-up. The crude cumulative incidence of failure of CQ therapy for uncomplicated P. falciparum was 42%  withdrew from the study after day 18 and 3 (10%) developed intercurrent % withdreP. vivax (day 22). Seventy-six (86%) completed the study without recurrent parasitaemia. The remaining 5 (6%) subjects developed recurrent P. falciparum during follow-up, therefore providing a crude therapeutic efficacy of 94%. The crude 28-day cumulative incidence of failure of CQ + SP therapy was 6%  withdrew before day 28 (three declined further participation between days two and 21 and three moved away from the study site between days one and 18). One subject (1%) developed intercurrent 6% Table . The relP. falciparum DNA could not be amplified for msp-2 genotyping, leaving eight evaluable recurrent infections. One case of day 28 recurrent parasitaemia was reclassified as a cure with late reinfection based on differing day 0 and day of recurrence msp-2 genotypes. From the remaining seven recurrent infections, the parasite genotypes matched and their classification as recrudescence affirmed. The adjusted cumulative incidence of failure of CQ therapy for uncomplicated P. falciparum was therefore 30% (therapeutic efficacy 70%)  Table .msp-2 genotypes, three apparent therapeutic failures were reclassified as successful cures with reinfection. The 28-day cumulative incidence of failure of CQ+SP therapy for uncomplicated P. falciparum was therefore only 1% (therapeutic efficacy = 99%)  Table . SubjectFigure The combination of CQ+SP proved 99% effective among 87 subjects with uncomplicated falciparum malaria in Central Java, Indonesia. Similarly excellent efficacy has been reported from other regions ,11-14, iP. falciparum and other species of malaria in Indonesia.Despite the marked differences between day-28 cure rates, CQ vs. CQ+SP regimens had essentially identical parasite and fever clearance times. These two indices parallel clinical recovery and almost certainly exacerbate the persistent over-the-counter marketing and unsupervised use of CQ monotherapy. Indeed, surveys of knowledge, attitudes and practices in the region of study affirmedCQ+SP can not be suggested as a first-line treatment policy that would be supported by comprehensive national programs of patient awareness, health provider training, and widespread distribution and marketing. The efficacy of the combination may not be sufficiently long-lived to support these costly adjustments and measures. However, CQ combined with SP may be a feasible therapeutic option in regions with evidence of treatment efficacy where health care providers are unable to use ACT because of problems with availability, fear of side effects or counterfeit drug. Use should be limited to areas like Central Java, where data suggests that CQ+SP efficacy was better than 95%.P. falciparum. A single 45 mg dose reduces gametocyte clearance time to 2 to 3 days [P. falciparum malaria treatment policy in Indonesia includes administration of 45 mg of primaquine on day 0 of treatment in hypoendemic areas. This study supports its continued use in Indonesia, especially with combination regimens containing SP, which has been linked to sulfonamide-associated gametocyte proliferation [Primaquine has proven gametocytocidal activity against o 3 days , nearly o 3 days . The addo 3 days . Variouso 3 days ; howevero 3 days ,29. In oferation . Combiniferation ; similarferation .P. falciparum in Central Java, Indonesia. The regimen was safe and efficacious, and it is inexpensive and readily available throughout Indonesia. A single dose of 45 mg primaquine, either on day 0 or day 2, suppressed or quickly eliminated gametocytaemia. Given wide regional differences in efficacy of CQ+SP, this regimen should be evaluated where its use is advocated as a therapeutic option for falciparum malaria.In summary CQ+SP combination was highly effective for parasitological cure in subjects with uncomplicated EL and JM are infectious disease specialists and contributed to the data analysis and drafting of the manuscript, and supervision of the field site. JKB is a parasitologist and was the principal investigator of the protocol and senior editor of the manuscript. MJB is an entomologist and contributed to site supervision and manuscript editing. IE is a statistician who performed the data analysis and supervised the data collection in the field. KC, PS, IW and LE are Indonesian general practitioners with experience in caring for subjects with malaria; they performed the screening, recruitment and care of subjects during this study as well as drafting of the manuscript."}
+{"text": "As a result of rising levels of drug resistance to conventional monotherapy, the World Health Organization (WHO) and other international organisations have recommended that malaria endemic countries move to combination therapy, ideally with artemisinin-based combinations (ACTs). Cost is a major barrier to deployment. There is little evidence from field trials on the cost-effectiveness of these new combinations.An economic evaluation of drug combinations was designed around a randomised effectiveness trial of combinations recommended by the WHO, used to treat Tanzanian children with non-severe slide-proven malaria. Drug combinations were: amodiaquine (AQ), AQ with sulfadoxine-pyrimethamine (AQ+SP), AQ with artesunate (AQ+AS), and artemether-lumefantrine (AL) in a six-dose regimen. Effectiveness was measured in terms of resource savings and cases of malaria averted . All costs to providers and to patients and their families were estimated and uncertain variables were subjected to univariate sensitivity analysis. Incremental analysis comparing each combination to monotherapy (AQ) revealed that from a societal perspective AL was most cost-effective at day 14. At day 28 the difference between AL and AQ+AS was negligible; both resulted in a gross savings of approximately US$1.70 or a net saving of US$22.40 per case averted. Varying the accuracy of diagnosis and the subsistence wage rate used to value unpaid work had a significant effect on the number of cases averted and on programme costs, respectively, but this did not change the finding that AL and AQ+AS dominate monotherapy.In an area of high drug resistance, there is evidence that AL and AQ+AS are the most cost-effective drugs despite being the most expensive, because they are significantly more effective than other options and therefore reduce the need for further treatment. This is not necessarily the case in parts of Africa where recrudescence following SP and AQ treatment (and their combination) is lower so that the relative advantage of ACTs is smaller, or where diagnostic services are not accurate and as a result much of the drug goes to those who do not have malaria. A randomised effectiveness trial of antimalarial drug combinations used to treat Tanzanian children found artemether-lumefantrine to be the most cost-effective. For many years, malaria was treated with a course of a single drug. This type of treatment made it easy for malaria parasites to become resistant to antimalarial drugs. This is a major factor contributing to the continuing high death rate from the disease. However, although parasites can easily adapt to resist one drug, adapting to combinations of two or three drugs is much harder. Scientists have therefore developed combinations of antimalarial drugs. One component of these combinations is artemisinin\u2014derived from a Chinese shrub. However, these combination therapies are much more expensive than the older treatments.The regions worst affected by malaria\u2014Africa and Asia\u2014are also the poorest. And, in these areas, where both individual and government resources are scarce, antimalarial treatments must be cost-effective as well as clinically effective.Most of the estimated 1 million to 3 million people worldwide killed by malaria every year are young children in sub-Saharan Africa. Growing drug resistance, poor prevention programs, and a frequent inability of patients to pay for treatment mean that effective therapy is desperately needed in this part of the world. However, because of differences in drug resistance between regions, a drug combination will not work everywhere. In addition, because of low annual incomes (the average in Tanzania is US$120), heavy subsidies will probably be required to ensure that combination treatments are widely used. With several healthcare problems competing for resources, policymakers are likely to subsidize only the most cost-effective treatments. The researchers wanted to provide policymakers with information on how different combinations of malaria drugs compare in terms of costs, health effects, and cost-effectiveness, so that they can decide which treatment is best for their region.They compared three combinations that the World Health Organization recommends for countries when making the transition from single-drug therapy. The three combinations\u2014amodiaquine (AQ) and sulfadoxine-pyrimethamine (SP); AQ and artesunate (AS); and artemether-lumefantrine (AL)\u2014were used to treat Tanzanian children. The researchers wanted to find out how many cases of malaria each combination averted  and how much the treatment cost. They looked at costs and savings from the perspectives of both healthcare providers and patients.Compared with no treatment, AL proved to be the most cost-effective; although it cost more for the provider (US$3.01 at day 28 of treatment) than the others, its effectiveness in getting rid of the parasite meant it would save the cost of future treatment. By day 28, AL had averted 382 cases of malaria compared with 279 for AQ+AS, 181 for AQ+SP, and 57 for AQ alone. Also, higher proportions of inaccurate diagnoses of malaria led to lower cost-effectiveness of treatments.Despite being more expensive, newer drugs can be cost-effective where alternatives fail. Although AL was the most cost-effective in places (such as Tanzania) where the malaria parasites are highly resistant to SP and AQ, the picture is likely to change for other areas. In West Africa, for example, AQ resistance is lower, and AQ+SP and AQ+AS would probably be more cost-effective. And in areas where both these combinations are just as good as AL in preventing recurring disease, they would be more cost-effective than AL. However, since AQ and SP have been used singly for many years, the likelihood is that resistance to these drugs will continue to increase. Accurate diagnosis turns out to be very important for maintaining the cost-effectiveness of combination antimalarial therapies. This will be essential if they are to be incorporated as a sustainable part of local health policies. The researchers also point out that, depending on which perspective is taken (provider or patient), the cost-effectiveness of treatments differs, making it important to compare like with like.Although investing in costly AL treatments and improving diagnostic capabilities will be a challenge for African governments that currently spend less than US$5 per person per year on healthcare, it will be necessary if they are to seriously tackle the malaria epidemic.http://dx.doi.org/10.1371/journal.pmed.0030373Please access these Web sites via the online version of this summary at US enters for Disease Control and Prevention provides malaria information aimed at the general public, physicians, and health workersThe Wellcome Trust; also has malaria information for the general public and covers the science of malaria research, including a downloadable animation of the parasite's life cycleThe Medicines for Malaria Venture (MMV) is a charity created to develop new antimalarial drugs through public-private partnerships Rising drug resistance levels to conventional monotherapy has resulted in strong arguments for a move to combination treatment for all malaria in Africa, especially artemisinin-based combination therapies (ACTs) \u20132. The rStudies have not examined the cost-effectiveness of these drugs in areas of Africa with failing monotherapy in using original data, and in particular we are not aware of any that have compared the current treatment options available to public health and clinical decision-makers in Africa. These are essential data for rational policy-making if public funds  are likely to be involved. We therefore set out to compare the cost-effectiveness of the three currently available drug combinations to treat children with non-severe, slide-proven malaria compared to monotherapy (amodiaquine [AQ]). The combinations were AQ with sulfadoxine-pyrimethamine (AQ+SP), AQ with artesunate (AQ+AS) or artemether-lumefantrine . The study was undertaken in the context of a clinical trial where the effectiveness of the drugs taken unsupervised was measured directly . In thisThe study was conducted at Teule Hospital, a designated district hospital located in Muheza, a small town in the northeast district of Tanga, Tanzania with an EIR  of over 300 and where the average individual has around three clinical attacks of malaria a year . There aAn incremental approach was used to estimate costs and effects. This involved comparing each of the new combination medicines against the existing monotherapy of AQ. SP is currently first-line and AQ second-line treatment for uncomplicated malaria in Tanzania. Pilot testing for this trial revealed unacceptably high levels of resistance to SP, so AQ was selected as the monotherapy against which each combination medicine was compared. Economic costs were estimated using the bottom-up approach in which all resources required in the delivery of treatment are valued . Health Findings are presented from both a provider  and a societal perspective. For the provider perspective, only those costs and effects borne by the provider are considered; this is the cost-effectiveness from the point of view of a ministry of health. For the societal perspective, the costs and effects borne by patients and their families are combined with those of the provider, which is especially relevant when considering the case for subsidy.The main areas of resource use for the health service were medication, personnel, rent for hospital space, and use of a microscope. Published market drug prices were used \u201316, and Families of patients incurred both indirect and direct costs. Direct costs included out-of-pocket expenses such as hospital fees, transport, and medication, as well as miscellaneous expenditures such as informal payments made to medical staff, food, and blankets. Hospital fees were not charged for children under the age of 5, but fees were rendered if the registration card needed to be replaced. Medical costs included extra antibiotics or blood transfusions. Indirect costs included time lost as a result of caring for a sick child at home, travelling to hospital, and waiting at hospital for treatment. Time lost was valued at the prevailing minimum subsistence wage rate in Tanzania .Estimation of resources savings were based on the cost of current first-line treatment with SP. Current therapy differed from the intervention treatments in two key respects: type of medicine used and the time families spent seeking treatment. Current treatment with SP was valued using the hospital's procurement prices at the dispensary. Indirect costs were measured for patients taking the current treatment. These costs were the same as for AQ, as parasitological failure rates and drug cost are almost exactly the same as for SP in this setting . For theCosts incurred at the household level were collected through structured interviews at the hospital outpatient facility. Sixty-three parents of febrile children under the age of 5 diagnosed with malaria were selected between March and April 2005. Parents were purposively selected from different ethnic, socioeconomic, and rural/urban locations. All participants gave informed consent. All remaining costs were collected from the principle investigators of the trial and from hospital accounting records. Staff salaries were obtained from the standard payroll scales for Tanzania. All costs were converted from Tanzanian shillings to US dollars based on the exchange rate prevailing at the outset of the trial  and discounted at 3% per year to estimate their present value . ResearcThe programme costs for each treatment are shown in Cost-effectiveness data are shown in The greatest resource savings were associated with AL. From a societal perspective, total savings were US$10,261 at day 14 and US$7,919 at day 28. The equivalent figures from a provider perspective are US$2,520 at day 14 and US$1,944 at day 28. For all treatments, savings were higher at day 14 compared with day 28. The greatest reduction in savings to society between days 14 and 28 was reported for AQ (61%). Comparing the three combination treatments against monotherapy (AQ) showed that from a societal perspective AL was again most-cost-effective at day 14. By day 28 there was only a minor difference between AL and AQ+AS.relative to monotherapy by saving more resources and averting more cases of malaria. Changing the subsistence wage rate also led to notable changes in costs, but this, again, did not change the fact that combination treatments such as AL continued to be more cost-effective than monotherapy. Altering drug prices or the discount rate had little effect on the findings of this study.The results of the sensitivity analysis are shown in This is, to our knowledge, the first field-based study to compare the cost-effectiveness of the three available drug combinations recommended by a consultative group of WHO for deployment by countries when they move from monotherapy . For eacThis clear and striking finding has to be interpreted with three notes of caution, all of which are important in trying to generalise the messages of the study. Firstly, and obviously, AL is the most cost-effective drug despite being the most expensive because it is significantly more effective than other options and is therefore likely to reduce the need for further treatment. This will not, however, necessarily be the case in parts of Africa where recrudescence following SP and AQ treatment (and their combination) is lower, so the relative advantage of this ACT is smaller ,20. In pDiagnostic accuracy has important operational implications. Our results show that accuracy of diagnosis plays an important role in the estimation of effectiveness of combination therapy for malaria. If cost-effectiveness of ACTs is undermined in this way then political support for them is unlikely to be sustained. In this study, double-read research microscopy was used to confirm malaria and only those with a positive result received treatment. In practice, however, microscopes are not widely used in many parts of Africa and clinical diagnosis tends to be the norm. Moreover, recent studies in Tanzania and elsewhere provide evidence that, in practice, many patients with a negative result go on to be treated for malaria . DrawingWhile resource savings are a legitimate outcome of many interventions they tend inevitably to be less precise than estimates of programme costs. For example, adherence levels may be higher under trial conditions. The data from this study come from an effectiveness study conducted in a setting similar to outpatient practice in children, with drugs taken unsupervised at home, rather than efficacy data from trials where all doses are supervised, but families may have perceived trial-based treatment to be of a higher quality and this may promote adherence and, in turn, cases of malaria averted. The lack of payment for ACTs in the trial reflects the current policy in Tanzania. However, the influx of these drugs in the private sector may give families an incentive to save or to sell extra drugs instead of taking the correct dose . These dIn this study all cost and cost-effectiveness estimates are shown from two perspectives: societal and provider. Depending on the perspective taken the results differ substantially. For example, both the programme costs and the resource savings to patients and their families are greater than those to the health service. This translates into lower cost-effectiveness ratios under a societal perspective than under a provider one. When comparing these results with cost-effectiveness ratios for other malaria control or health interventions it is important to ensure that, as far as possible, like is being compared with like and the starting point for this should be the perspective of the evaluation. Again, we have shown estimates based on both perspectives to allow readers to adopt the one that most suits the context in which they are working.Finally, it has been suggested that a cost-effectiveness ratio less than double the annual income per capita might be an acceptable threshold value for most governments deciding which intervention to fund . The fin"}
+{"text": "Plasmodium falciparum non-complicated malaria patients.To establish the relationship between production of glutathione and the therapeutic response to amodiaquine (AQ) monotherapy in Therapeutic response to AQ was evaluated in 32 patients with falciparum malaria in two townships of Antioquia, Colombia, and followed-up for 28 days. For every patient, total glutathione and enzymatic activity  were determined in parasitized erythrocytes, non-infected erythrocytes and free parasites, on the starting day  and on the day of failure (in case of occurrence).p < 0.003). In addition, only on the day of failure, \u03b3-GCS activity of parasitized erythrocytes was higher, compared with that of healthy erythrocytes (p = 0.01). Parasitized and non-parasitized erythrocytes in therapeutic failure patients (TF) had higher total glutathione on the starting day compared with those of adequate clinical response (ACR) (p < 0.02). Parasitized erythrocytes of TF patients showed lower total glutathione on the failure day, compared with starting day (p = 0.017). No differences was seen in the GR and \u03b3-GCS activities by compartment, neither between the two therapeutic response groups nor between the two treatment days.There was found an AQ failure of 31.25%. Independent of the therapeutic response, on the starting day and on the day of failure, lower total glutathione concentration and higher GR activities in parasitized erythrocytes were found, compared with non-infected erythrocytes (P. falciparum therapeutic failure in humans through differences in glutathione metabolism in TF and ACR patients. These results suggest a role for glutathione in the therapeutic failure to antimalarials.This study is a first approach to explaining  Plasmodium falciparum [P. falciparum to antimalarial drugs since 1960. In Antioquia, Colombia, 97% and 30% of therapeutic failure to the 4-aminoquinolines, chloroquine (CQ) and amodiaquine (AQ) respectively, have been reported [Malaria is a parasitic disease, which has the highest morbidity/mortality rate in tropical countries . The caulciparum . In ColoPlasmodium during intraerythrocytic stages in a process where haem is released and converts itself into a toxic molecule, because this parasite lacks haem oxygenase and produces reactive oxygen species (ROS) [Plasmodium is endowed with a machinery to detoxify ROS. This antioxidant system comprises GSH and thioredoxin redox system, superoxide dismutase, NADPH and a vigorous pentose phosphate pathway, but it lacks catalase and glutathione peroxidase [Haemoglobin is degraded by es (ROS) . The pares (ROS) ,7Plasmodroxidase -10.Glutahione is synthesized by the step-limiting enzyme \u03b3-glutamyl cysteine synthetase (\u03b3-GCS) and by glutathione synthetase (GS). This thiol found in its reduced state inside all cells, contributes to maintaining intracellular redox state; besides, it has been suggested that GSH is involved in drug resistance both as a cofactor for enzymatic reactions and by helping to mediate resistance as a source of reductive detoxification of haem. After its reaction with a free radical, GSH changes its oxidation state (GSSG). Glutathione reductase (GR) NADPH is the enzyme responsible for keeping glutathione in its reduced state ,10.P. falciparum to the 4-aminoquinolines, such as CQ, has been shown to be associated with modification of drug transport by the membrane proteins P-glycoprotein homologue 1(Pgh1) and the P. falciparum chloroquine resistance transporter (pfcrt) [The resistance of  (pfcrt) . An addi (pfcrt) .et al. [P. falciparum CQ-sensitive strain (3D7) by L-buthionine--sulfoximine (BSO), a specific inhibitor of \u03b3-GCS and by methylene blue (MB), an inhibitor of the GR, was significantly more pronounced than inhibition of P. falciparum CQ-resistant strain (Dd2) growth by these drugs. These results correlate with the higher levels of total glutathione in P. falciparum Dd2. In addition, they suggest that maintenance of intracellular GSH in P. falciparum Dd2 is mainly dependent on GSH synthesis, whereas in P. falciparum 3D7 is regulated via GR [Meierjohann et al.  showed td via GR .in vivo on Plasmodium berghei-infected mice confirm that drugs which alter GSH intracellular concentrations, change Plasmodium susceptibility to the CQ [P. berghei is associated with a significant increase in parasite glucose 6-phosphate deshydrogenase (G6PD) activity and GSH. Combination of CQ with an inhibitor of G6PD or BSO significantly increased sensitivity of resistant parasites to CQ and increased the survival period of the infected mice [in vivo on P. berghei and Plasmodium vinckei-infected mice showed that drugs, such as acetaminophen, indomethacin and disulphiram, which produce an indirect decrease in GSH, potentiate the antimalarial action of CQ and AQ sub-curative doses [Studies o the CQ . The coro the CQ . It has ted mice . Other sve doses .P. falciparum and therapeutic failure to AQ, since CQ has not actually been used as a monotherapy. Possible variations of total glutathione levels and GR and \u03b3-GCS activities were determined in function of the therapeutic response in parasitized erythrocytes, non-infected erythrocytes and free parasites. It should help to understand the phenomena of therapeutic failure to the 4-aminoquinolines.This study examined whether there exists a relationship between the total glutathione level in patients with malaria by P. falciparum malaria, according to the WHO Protocol 2000 that classifies the response in adequate clinical response (ACR) and therapeutic failure (TF). Patients were from Turbo and El Bagre, two townships in Antioquia, Colombia. In this study, the therapeutic failure was higher than permitted (>26%) by National Health Ministry treatment guidelines in Colombia and, therefore, the study was carried out only with 32 patients. Initial malaria diagnosis was carried out with thick and thin smear; these were Field and Giemsa stained respectively. Parasitaemia was calculated by counting the number of asexual forms by 200 leucocytes [Within the framework of the evaluation of the resistance to antimalarics by RAVREDA , during 2003 and 2004, clinical and parasitological therapeutic response to monotherapy with AQ was assessed in patients with non-complicated ucocytes . SamplesA written informed consent from each patient was obtained and signed before being included in the research project. This study was approved by the Ethics Committee of the Facultad de Medicina of the Universidad de Antioquia.Blood samples were thawed according to the workshop protocol suggested in the course \"Molecular Approaches to Malaria\" \u2013 International Centre for Engineering and Biotechonology .et al 2003 with some modifications [In thawed samples, parasitized and non-parasitized red blood cells were separated by Percoll gradients according to the methodology described by Omodeo-Sale ications . The thaThe free parasites were obtained according to Hsiao protocol with some modifications : the pelThe enzymatic activity of glutathione reductase (GR) and \u03b3-glutamylcysteine synthetase (\u03b3-GCS), protein concentration and total glutathione content were determined in parasitized and non-parasitized erythrocytes and in free parasites, and were carried out in duplicate.This was carried out according to the method described by Calberg I and Mannervik B 1985 . GlutathThis was determined according to the method described by Estrada del Cueto et al. 1999 . This te\u00ae Calbiochem) by reversed-phase HPLC. The method suggested by Luersen et al. 2000 was followed with some modifications [4, in 0.066 M NaOH and 33% (v/v) dimethyl sulphoxide (DMSO), 6 \u03bcl of 2 mM EDTA and 1.65 mM dithiothreitol, 6 \u03bcl of octanol and 14 \u03bcl of 1.8 M HCl. After three min 70 \u03bcl of 1 M ethylmorpholine buffer (pH 8.5), 134 \u03bcl of deionized water and 14 \u03bcl of 5 mM thiolyte\u00ae were added. The derivatization was carried out at 70\u00b0C for 10 min in the dark and was terminated by adding 26 \u03bcl of 100% acetic acid. After 20 min on ice in the dark, the sample was extracted with 200 \u03bcl of dichloromethane and centrifuged at 10,000 rpm at RT for two minutes. The supernatant, which is the water-soluble phase, was taken and filtered through a 0.45-\u03bcm nylon membrane. The filtered samples were stored at -20\u00b0C, protected from light until their injection in the HPLC. 20 \u03bcl obtained from the water-soluble phase were injected on to a LiChroCART\u00ae 100 RP-18 (5.0 \u03bcm) reverse phase HPLC. The column was eluted at a flow rate of 0.55 ml/min by the following gradient of solvent A (0.25% acetic acid) and solvent B (100% acetonitrile): 0 min, 100% solvent A; 5 min, 90% solvent A; 20 min, 85% solvent A, 25 min, 0% solvent A. The effluent was monitored by a fluorescence spectrophotometer . Under these conditions, the glutathione-thiolyte\u00ae adduct had a retention time of 13.9 min and cysteine-thiolyte\u00ae of 10.07 min. Cysteine (Cys) and GSH were used as external standards with recovery rate of 123.8 \u00b1 6.6% and 74.8 \u00b1 8.2% respectively. Two calibration curves were elaborated due to concentration variation of glutathione in the samples and in doing the regression analysis . The obtained sensibility was 3.2 pmol, which is found in the quantity range detected when working derivatized compounds with monobromobimane [The concentration of total glutathione (oxidized and reduced) was analysed as mBBr derivatives  was applied to all estimates and statistical tests. The program SpSS 14 was used for the statistical analysis.The data were analysed by the one-way variance analysis (Mann-Whitney test) in order to compare mean of total glutathione concentration and GR and \u03b3-GCS activities in parasitized red blood cells, non-parasitized and free parasites in function of the therapeutic response (ACR and TF). A matching test was applied (Wilcoxon test) to compare the means in both days (starting and failure day) in the group of patients that had therapeutic failure. A correlation analysis and a simple linear regression were made to assess the association between GR and \u03b3-GCS activity in ACR and FT patients with parasitaemia, and glutathione concentration in all cellular compartments. A confidence level of 95%  old and 17 of them were male. The 32 patients evaluated were divided into two groups according to therapeutic response to AQ; 22 of them had ACR and 10 TF (31.25%). On the starting day (zero day), mean parasitaemia was 8,699  rings/\u03bcl for ACR patients and 9,944  rings/\u03bcl for TF patients; on the failure day, mean parasitaemia was 1,681  parasites/\u03bcl.The therapeutic response to monotherapy with AQ was assessed during 28 days in 32 patients with non-complicated malaria by p < 0.001 and TF = 21.636 \u00b1 4.149 nmol/mg protein vs 11.725 \u00b1 4.833 nmol/mg proteins, p = 0.003). Also for both therapeutic response groups was found that GR activity was statistically lower in the healthy erythrocytes compared to parasitized erythrocytes . Differences were not found in the \u03b3-GCS activity between both compartments for any of two groups of therapeutic response (p > 0.05) . There were no differences for GR and \u03b3-GCS activities for either therapeutic response group . Differences in GR and \u03b3-GCS activities were not found for either therapeutic response group.2 < 0.5, data non-shown).In correlation analysis and a simple linear regression, TF patients presented a 58% correlation between parasitaemia and GR activity of free parasites, and statistically significant correlations were not found in any of the cellular compartments for ACR patients  (Table p = 0.017) and there was no important changes in total glutathione of non-parasitized erythrocytes or in the GR and \u03b3-GCS activity  Independent of the therapeutic response and of the treatment day (starting or failure), the total glutathione was higher and GR activity was lower in the non-infected erythrocytes compared with parasitized erythrocytes.2) The higher GR activity in the infected erythrocytes on the failure day could be due to a compensation mechanism by lower total glutathione in this compartment.3) The patients with TF had higher total glutathione in parasitized and non-parasitized erythrocytes on the starting day compared with the ACR patients, indicating a possible contribution to the therapeutic failure.4) On failure day, the maintenance of glutathione in parasitized erythrocytes depends on synthesis and reduction, possibly by the increase in the oxidative stress that AQ generates.5) In the FT patients, the total glutathione found in infected erythrocytes was less on the failure day compared with starting day. This is possibly due to the larger amount of glutathione which is consumed in presence of the drug and also to lower parasitaemia.P. falciparum therapeutic failure through differences found in glutathione metabolism in FT and ACR patients. Our results may involve glutathione in the therapeutic response to antimalarials; nevertheless, further studies are needed to prove this relationship. It is also important to compare these results with samples of healthy individuals to rule out other possible differences. In addition, it is important to perform more accurate enzymatic assays to discard differences in the enzymatic activities between the treatment days and between the therapeutic response groups. It would be useful to do a follow-up of the glutathione during and after the malaric episode in patients with different therapeutic response to establish if the glutathion (GSH and GSSG) change during the infection and if they return to the same level when the parasite is not present.This study is the first attempt to explain LZ designed and managed the study, made the assays of separation cellular and evaluated enzymatic activity, analysed of data and wrote the first draft of the manuscript. AP designed study, analysed of data, contributed in the standardization of the techniques and cowrote the first draft of the manuscript. AO validated the method for quantification of total glutathione and made injection the samples to HPLC. CL coordinated the validated for quantification of total glutathione and analysed data. SB coordinated the study, analysed data and critically reviewed and suggested changes to the manuscript. All authors read and approved the final manuscript."}
+{"text": "Intermittent preventive treatment with sulphadoxine-pyrimethamine (IPT-SP) is currently the recommended regimen for prevention of malaria in pregnancy in endemic areas. This study sets out to evaluate the effectiveness of IPT-SP in the prevention of maternal and placental malaria in parturient mothers in Ibadan, Nigeria, where the risk of malaria is present all year round.During a larger study evaluating the epidemiology of congenital malaria, the effect of malaria prophylaxis was examined in 983 parturient mothers. Five hundred and ninety eight mothers (60.8%) received IPT-SP, 214 (21.8%) received pyrimethamine (PYR) and 171 (17.4%) did not take any chemoprophylactic agent (NC).The prevalence of maternal parasitaemia in the IPT-SP, PYR and NC groups was 10.4%, 15.9% and 17% respectively (p = 0.021). The prevalence of placental parasitaemia was 10.5% in the IPT-SP, 16.8% PYR and 17% NC groups, respectively (p = 0.015). The prevalence of maternal anaemia (haematocrit <30%) was 5.7% vs. 8.9% vs. 13.4% among the IPT-SP, PYR and NC groups respectively (p < 0.0001) while that of pre-term delivery (GA <37 weeks) was 10.5%, 19.2% and 25.3% among IPT-SP, PYR and NC groups respectively (p < 0.0001). Babies born to mothers in the IPT-SP, PYR and NC groups had mean birth weights of 3204 \u00b1 487.16, 3075 \u00b1 513.24 and 3074 \u00b1 505.92 respectively (\u03c1 < 0.0001). There was a trend towards a lower proportion of low birth weight babies in the IPT-SP group (p = 0.095).IPT-SP is effective in preventing maternal and placental malaria as well as improving pregnancy outcomes among parturient women in Ibadan, Nigeria. The implementation of the recently adopted IPT-SP strategy should be pursued with vigour as it holds great promise for reducing the burden of malaria in pregnancy in Nigeria. The pregnant woman runs a higher risk of contacting malaria than her non-pregnant counterpart -4. An esPlasmodium falciparum have, however, compromised the efficacy of these regimens [To prevent the adverse effects of malaria in pregnancy, antimalarial chemoprophylaxis is generally recommended. For a long time, prophylaxis with weekly pyrimethamine or chloroquine was widely adopted in many African countries . Poor coregimens ,12. Interegimens -17. Thisregimens ,15. Studregimens ,14,16,17Despite the well documented lack of efficacy of pyrimethamine for malaria chemoprophylaxis, many practitioners have continued to use this approach for pregnant women in Nigeria. This article reports the effectiveness of IPT-SP in the prevention of maternal and placental malaria in an urban hospital in Ibadan, Nigeria during a pilot study of the IPT-SP strategy.The study was conducted at the St Mary's Catholic Hospital Eleta Ibadan between May 2003 and October 2004. St Mary's Hospital is a secondary health care facility run by the Catholic mission. It is located in the heart of the ancient city of Ibadan. However, St Mary's Hospital attracts patients of various socio-demographic classes from the entire city as a result of the high quality of service it has provided over decades at reasonable cost. Ibadan is located in the rain forest belt in south-western Nigeria. Malaria transmission is intense year round with a peak during the rainy season months of May to October and a nadir during the dry season months of November to April. In a recently conducted national efficacy study, day 14 efficacy of SP in Ibadan in acute uncomplicated malaria was 85%  among chWomen who participated in a larger prospective observational study evaluating the epidemiology of congenital malaria in Ibadan, Nigeria were enrolled into the study . Only paSix months after commencement of the study, a pilot study on IPT-SP was introduced in some states in Nigeria. St Mary's Hospital was one of the hospitals selected for the IPT-SP pilot study in Oyo state. While other centers gave SP free to pregnant women, the study site gave IPT-SP at a cost of \u20a650 ($0.35) per dose for sustainability. Three tablets of SP containing 500 mg sulphadoxine and 25 mg pyrimethamine per tablet  were administered under supervision of midwives at the antenatal clinic and recorded in the patients' antenatal case notes. The use of IPT-SP was confirmed from the antenatal notes. Records were subsequently signed by the midwife responsible for its administration. Enquiries about untoward effects were made during the next clinic visit and were recorded. An investigator assisted questionnaire was administered to enrolled mothers at delivery to collect information on socio-demographic factors, malaria chemoprophylaxis, and occurrence of fever and malaria symptoms as well as anti-malaria drug use within two weeks of delivery. Details of other clinical and laboratory enrollment procedures are described in the report of the epidemiology of congenital malaria in Nigeria .Babies born before 37 weeks of gestation were considered pre-term while those born from 37 weeks of gestation and above were considered term. Babies were weighed to the nearest gram using an electronic weighing scale. For the purpose of this study LBW was defined as neonatal birth weight less than 2,500 g while a haematocrit reading of <30% was considered as anaemia.nd and 3rd trimester of pregnancy, where second trimester is defined as 14 to less than 28 weeks of gestation and 3rd trimester as 28 weeks and above. Adequacy of pyrimethamine was defined as the use of 25 mg pyrimethamine weekly two weeks after quickening until term while the adequacy of chloroquine was defined as the use of 300 mg (two tablets) chloroquine base weekly for the same time duration as pyrimethamine. To be considered adequately used, proguanil needed to be taken at a dose of 200 mg daily for the same duration as pyrimethamine and chloroquine.For the purposes of this study, all patients were categorized into three groups based on type of chemoprophylactic agent used. Patients in group 1 received IPT-SP, group 2 received pyrimethamine while those who took no chemoprophylactic agent were in group 3. Adequacy of IPT-SP use was defined as the use of at least two therapeutic doses of SP at least one month apart during the 2Data collected were recorded into pre-coded case record forms. Thereafter, the data was double entered by two data entry clerks using EPI-INFO 6.04d . Preliminary data analysis was done with EPI-INFO. Thereafter, the data were transferred to SPSS version 10  for analysis. Descriptive statistics such as means and standard deviations were used to summarize quantitative variables while categorical variables were summarized with proportions. Frequency tables and graphs were presented for relevant variables. The student t-test was used to compare two mean values while the one way analysis of variance (ANOVA) was used to compare mean values in more than two groups. The Chi-square test was used to investigate associations between two categorical variables and also to compare proportions. For significant associations, the odds ratio (OR) and 95% confidence intervals (CI) were computed. A p-value less than 0.05 was considered statistically significant.Over an 18 month period (May 2003 to October 2004), 983 mother-baby pairs were enrolled. The mean maternal age was 29.6 \u00b1 5.2 years. The youngest mother was 17 yrs of age while the oldest was 48 yrs. Only 11 (1.1%) of the mothers were less than 20 yrs of age. The socio-demographic characteristics of mothers in the IPT-SP, pyrimethamine (PYR) and no chemoprophylactic (NC) groups were similar  of study participants admitted to using at least one form of anti-vector measure. Mosquito screens on windows were the most commonly used (77.9%) followed by insecticide sprays (69.1%), coils (25%), herbs (6.6%), untreated bed nets (6.6%) and mosquito repellants (0.5%). Insecticide treated bed nets (ITNs) were used by only 1.1% of pregnant women participating in the study.Most  mothers admitted to using some form of antimalarial chemoprophylaxis. The remaining 143 (14.6%) did not use any malaria chemoprophylactic drug during the index pregnancy. More than half of the mothers  received IPT-SP while 21.8% (214/983) received pyrimethamine monotherapy. Other chemoprophylactic agents used by mothers enrolled into the study include chloroquine , herbs  and proguanil . Five hundred and five of 598 (84.4%) patients who received IPT-SP received at least two therapeutic doses as stipulated in the guideline while 94.4% (202/214) of those who took pyrimethamine claimed to have taken adequate doses. All the patients who used chloroquine for chemoprophylaxis received grossly inadequate doses of chloroquine. The only patient who used proguanil took 100 mg daily for less than two weeks. All the patients who took chloroquine, herbal preparations and proguanil were considered as having taken no chemoprophylactic agent and were categorized into the NC group bringing the number in that group to 171 (17.4%).A history of febrile illness in the two weeks preceding delivery was obtained in 9.5% (56/592), 23.5% (50/213) and 20.4% (34/167) of the mothers who received IPT-SP, PYR and NC respectively (p < 0.0001).At delivery, the prevalence of maternal parasitaemia was significantly lower in the IPT-SP group when compared to the PYR and NC groups Table . Both maThe mean haematocrit of parturient women who used IPT-SP was higher than for the PYR and NC groups (p = 0.006) Table . The meaThe mean birth weight of babies born to mothers who received IPT-SP was significantly greater than those babies born to mothers in the PYR and NC groups (p < 0. 0001) Table . When coSP was well tolerated. There was no incidence of pruritus, fixed drug eruptions or allergic reactions attributable to any of the chemoprophylactic agents and IPT-SP used by any of the mothers. Five mothers reported episodes of dizziness within 30 minutes of taking SP. Dizziness in all cases terminated within a few hours without any specific treatment. There were no congenital malformations or deaths  among the study participants.P. falciparum malaria is well recognized as an important cause of morbidity leading to maternal anaemia and LBW in the neonate [Pregnancy-associated  neonate -7,21. Th neonate . This re neonate ,14,16,17 neonate ,14,16,17 neonate . All of Challis et al  working The differences in the prevalence of maternal and placental malaria among IPT-SP users and non-users occurred despite the fact that the vast majority of parturient women used one anti-vector measure or another. The pattern of anti-vector use was similar in both groups to a large extent. Although ITNs have been shown to be effective in the control of malaria in pregnant women ,24, its Judging from the marked reduction in the incidence of clinically diagnosed malaria in the two weeks preceding delivery in our study, IPT-SP use significantly reduced episodes of febrile illnesses suspected to be malaria during the last two weeks of pregnancy compared to mothers who used pyrimethamine or no chemoprophylaxis. Although it would have been useful to evaluate the timing of the last dose of IPT-SP prior to delivery, these data were not captured in the database of the study reported here.LBW and prematurity are the greatest risk factors for neonatal mortality and a major contribution to infant mortality ,9. In thAnaemia is a well recognized consequence of malaria. Although maternal anaemia is multifactorial, malaria is known to contribute significantly to its occurrence in pregnancy. The prevalence of anaemia was least among parturient women who received IPT-SP during pregnancy when compared with the PYR and NC groups. Anaemia was earlier reported by workers in the same environment to have a negative impact on pregnancy outcome with still births occurring more often in anemic mothers . Aimaku Adverse events reported by study participants on IPT-SP and other chemoprophylactic agents were mild and did not necessitate any treatment. This is similar to the findings of previous workers ,17,22. SIPT-SP is highly effective in preventing maternal and placental malaria among parturient women in Ibadan, southwestern Nigeria as well as in improving pregnancy outcomes, including a lower prevalence of pre-term deliveries, bigger babies and lower prevalence of maternal anaemia. This study has once again confirmed the lack of efficacy of pyrimethamine for the prevention of maternal and placental malaria in pregnant Nigerian women . The Nat\u2022 Conception of the study and study design: CF, OM, DH\u2022 Conduct of the study: CF, BY, FF, OM, DH\u2022 Analysis of the data: BY\u2022 Interpretation of the data: CF, BY, FF, DH, LS\u2022 Drafting and critical review of the paper: CF, BY, FF, OM, DH, LS\u2022 Read and approved final version of the paper: CF, BY, FF, OM, DH, LS"}
+{"text": "Early diagnosis and effective treatment with an appropriate drug form the main components of the World Health Organization's strategy to reduce malaria related mortality. The few available drugs might be safeguarded if combined with artesunate. The addition of artesunate to a standard antimalarial treatment substantially reduces treatment failure, recrudescence and gametocyte carriage.Plasmodium falciparum malaria was investigated in four sentinel areas in Sudan, with different malaria transmission .During late 2004, the efficacy of artesunate  plus sulfadoxine-pyrimethamine  for the treatment of uncomplicated Two hundreds and sixty-nine patients completed the 28-day follow-up. On day one, 60 (22.3%) patients were febrile and 15 (5.5%) patients were parasitaemic. On day three, all the patients were afebrile and aparasitaemic. While two patients  showed late Clinical and Parasitological Failures, the rest (99.3%) of the patients demonstrated Adequate Clinical and Parasitological Response. A gametocytaemia were detected during the follow-up in one patient . Adverse drug effects were detected in 32 (11.9%) patientsP. falciparum malaria in Sudan.The study showed that AS plus SP is an effective, safe drug in the treatment of uncomplicated  Plasmodium falciparum malaria infections [There are almost 515 range 300\u2013660) million episodes of clinical fections . Early d00\u2013660 miP. falciparum. Coupled with early detection and confirmed diagnosis, this strategy represents the only way forward in the chemotherapy of malaria [The few available drugs might be safeguarded if combined with artesunate. The addition of artesunate to standard antimalarial treatments substantially reduces treatment failure, recrudescence and gametocyte carriage, preventing the emergence and spread of drug resistance and interrupting the transmission of  malaria -8.P. falciparum malaria in Sudan [P. falciparum malaria. The study aimed to investigate the efficacy of AS plus SP, as there is little published data in Sudan [Malaria causes between 7.5 to 10 million cases and 35,000 deaths every year in Sudan . Due to in Sudan , artesunin Sudan ,12.P. falciparum malaria [The study was conducted in October and November, 2004 at four health centres in different regions of Sudan  Figure . Three o malaria , who hadAfter obtaining informed consent from the patient or the child's parents, a fixed questionnaire including relevant socio-demographic characteristics, medical history, physical findings and investigations conducted was completed for each patient.Blood films were prepared, stained with Giemsa and 100x oil immersion fields were examined. The parasite density was counted against 200 leucocytes, assuming 6,000 leucocytes/\u03bcl. All the slides were double-checked blindly and only considered negative if no parasites were detected in 100 oil immersion fields. If gametocytes were seen, then the count was extended to 500 leucocytes.The patients were given the AS plus SP combination, with artesunate (4 mg/kg. day) given on days 0\u20132 and a single dose of SP (25 mg/kg)  given on day 0. The tablets were crushed and dissolved in water for children who were not able to swallow them. Subjects were observed for vomiting for one hour; the full dose was repeated for those who vomited within 30 min and half of the dose was repeated if vomiting occurred between 30 and 60 minutes.Patients were requested to come on days 1, 2, 3, 7, 14, 21 and 28 and at any time if they felt unwell. At each visit, body temperature was measured and blood films were prepared. During the follow-up the patients were asked if they suffered from side effects which can be expected from antimalarial treatment ; these symptoms were considered to be drug related if they had not been reported at the patient's first presentation in the clinic.Quinine was given for treatment failures. Early Treatment Failures (ETF) in case of significant parasitaemia at day 2 or 3 or parasites and fever at day 3. Late Clinical Failures (LCF) for cases with parasites and fever during follow-up after day 3 and Late Parasitological Failures (LPF) for parasite infections with/without fever during the follow-up. Cases which remained negative during follow-up were considered Adequate Clinical and Parasitological Responses (ACPR). These were modified WHO guidelines ,15.2 test. P < 0.05 was regarded significant.Data were entered into a computer database and SPSS software  was used for statistical analysis. The means  were calculated for all the patients and were compared between the patients in the different locations using one way analysis of the variance (ANOVA), when the data is normally distributed and by the Kruskal Wallis test if the data was not normally distributed. Percentages were calculated and compared for the patients in the four locations by an \u03c7The study received ethical clearance from the Sudanese National Malaria Administration.Two hundred and ninety (32.5%) out of 890 screened patients fulfilled the criteria and were enrolled in the study. Twenty-one (7.2%) of these were lost in the follow-up and 269 patients  completed the 28-day follow-up. Their different characteristics are shown in Table On day one, 60 22.3%) patients were febrile and 15 (5.5%) patients were parasitaemic. By day three all the patients were afebrile and aparasitaemic. There were two (0.7%) Late Clinical and Parasitological Failures (days 7 and 22) from Kassala, there was no Clinical and Parasitological Failures from other locations  were significantly different between the four locations, the study showed that two  out of 269 patients were found to have Late Clinical and Parasitological Failures. Since the parasite genotyping (PCR) was not conducted, the possibility of re-infection/recrudescence is still there. Hundred percent efficacy of AS plus SP was recently reported from eastern Sudan [P. falciparum strains were reported from eastern Sudan [The study investigated the efficacy of AS plus SP for the treatment of uncomplicated rn Sudan  and 99% rn Sudan . The higrn Sudan ,6. Howevrn Sudan ,16. The rn Sudan ,8.The adverse effects  might influence the adherence to AS plus SP, especially science this therapy is only available in the oral form, which is not the medication preferred by Sudanese patients . FurtherA post- treatment gametocytaemia was detected in one patient in Kassala area. High 20%) levels of gametocytaemia had been reported in the eastern Sudan following SP, quinine and mefloquine treatment -20. Howe% levels P. falciparum malaria in Sudan.The study showed that, As plus SP is an effective, safe drug in the treatment of uncomplicated SBE, EMM, TA, MTM, ESA carried out the study in the different sites and participated in the statistical analysis and procedures, AHK participated in the statistical analysis, IA coordinated and participated in the design of the study, statistical analysis and the drafting of the manuscript. All the authors read and approved the final version."}
+{"text": "Plasmodium falciparum. Four antimalarial drug combinations, artesunate plus amodiaquine (Arsucam\u00ae), artesunate plus mefloquine (Artequin\u00ae), artemether plus lumefantrine , and amodiaquine plus sulphadoxine-pyrimethamine, were studied in five health districts in Senegal.In view of the high level of chloroquine resistance in many countries, WHO has recommended the use of combination therapy with artemisinin derivatives in the treatment of uncomplicated malaria due to This is a descriptive, analytical, open, randomized study to evaluate the efficacy and tolerability of these four antimalarial combinations in the treatment of uncomplicated falciparum malaria using the 2002 WHO protocol.All drug combinations demonstrated good efficacy. On day 28, all combinations resulted in an excellent clinical and parasitological response rate of 100% after correction for PCR results, except for the four-dose artemether-lumefantrine regimen (96.4%). Follow-up of approximately 10% of each treatment group on day 42 demonstrated an efficacy of 100%.The combinations were well tolerated clinically and biologically. No unexpected side-effect was observed and all side-effects disappeared at the end of treatment. No serious side-effect requiring premature termination of treatment was observed.The four combinations are effective and well-tolerated. Approximately 35% of in-patient and out-patient clinic attendance in Senegal are due to malaria . StudiesThe development and rapid increase of chloroquine resistance in many countries led the World Health Organization to organize a technical consultation in April 2001, during \u00ae), artesunate plus mefloquine (Artequin\u00ae), and artemether plus lumefantrine . As a comparison, evaluation of combined therapy with amodiaquine plus sulphadoxine-pyrimethamine (SP) was also done; these latter antimalarial drugs are readily available in Senegal and were in use as a therapeutic strategy at the time of the study.In this study, the efficacy, safety and tolerability of three ACT for the treatment of malaria in Senegal were evaluated : artesunate plus amodiaquine , artesunate-amodiaquine, artemether-lumefantrine, amodiaquine-SP .This descriptive, analytical, open, randomized study was carried out to evaluate the following antimalarial combinations in the treatment of The study was carried out during the transmission periods of 2002 and 2003 in five observation sites distributed according to the epidemiology of malaria transmission in Senegal: Richard Toll and Podor in the north, Guediawaye and Kaolack in the central zone and Velingara in the south. The transmission of malaria is moderate in these different zones with a peak during the rainy season.Patients were recruited from those presenting at the healthcare centers with clinical signs suggestive of malaria. After a thick peripheral blood smear for screening, patients of all ages who fulfilled the inclusion criteria laid down by WHO/2002  were selAll patients gave their informed consent before participating in the study. The study protocol was approved by the ethics committee of the Ministry of HealthAfter inclusion, patients were weighed before being allocated to a treatment arm. All medication was administered under the direct supervision of a physician. If a patient vomited immediately after a dose, the same dose was re-administered within 30 minutes. Half of the dose was given if vomiting continued after the first 30 min. In the case of persistent vomiting, the patient was excluded from the study and sent to the medical team at the health center for management using the standard treatment in current use.Dosages of the study drugs were as follows:(1) artesunate-mefloquine: for adults, 200 mg/day artesunate plus 250 mg/day mefloquine, for 3 days; for children, 100 mg/day artesunate plus 125 mg/day mefloquine, for 3 days;(2) artesunate-amodiaquine: 4 mg/kg/day artesunate plus 10 mg/kg/day amodiaquine, for 3 days. The dosages were also adjusted according to the age of the patient;(3) artemether-lumefantrine: two dosages were administered: (i) in adults, four doses of four tablets repeated after 8, 24, 48 h; in children weighing 5\u201315 kg, one tablet repeated at 8, 24 and 48 h; (ii) six doses consisting of two daily doses for 3 days;(4) amodiaquine-SP: 10 mg/kg/day amodiaquine for 3 days and half a tablet per 10 kg of SP as a single dose only on the first day.All side-effects were monitored actively and passively during the study.Clinical evaluation and blood sampling from finger pricks for the preparation of thick peripheral blood smears were carried out on days 0, 1, 2, 7, 14, 21 and 28; 30% of the patients were also examined on day 42. Blood smears were stained with Giemsa and 200 leukocytes were read. Parasitaemia was expressed as the number of asexual parasites per microliter of blood according to the following formula: number of asexual parasites \u00d7 8,000/200.Twenty-five percent of the patients were selected randomly and blood cell counts and levels of transaminases  and creatinine were determined on days 0, 14 and 28 to determine hepatic and renal function.Biological tolerability was assessed by measuring haemoglobin , transaminases , and creatininaemia .Plasmodium. Genotyping was carried out in the Parasitology Laboratory, Facult\u00e9 de M\u00e9decine, Universit\u00e9 Cheikh Anta Diop, Dakar.Blood collection on filter paper was carried out for all patients on days 0 and 28, and on all other days in cases of clinical or parasitological failure for genotyping the strain of The primary end-point criteria used to determine the efficacy of treatment were absence of parasites and clinical signs on days 14 and 28, and good clinical and biological tolerability. The secondary end-point criteria were the time taken for the parasites to disappear, and for gametocyte clearance.The following definitions of clinical and parasitological failure were used:This was defined as a patient who presented with danger signs between days 1 and 3, accompanied by parasitaemia; a patient with an axilliary temperature of > 37\u00b0C accompanied by parasitaemia on day 3; parasitaemia on day 3 > 25% that on day 0; and a patient with parasitaemia on day 2 higher than that on day 0.This was defined as a patient presenting with danger signs between days 3 and 28, accompanied by parasitaemia; or a patient presenting with parasitaemia and a temperature > 37.5\u00b0C between days 3, 14 and 28.Parasitological failure was defined as a patient presenting with parasites on day 28.This was defined as the sum of therapeutic failures plus parasitological failures.ACPR was defined as the absence of parasites and clinical signs on day 28.Patients who were considered to have failed clinically were given second-line therapy with quinine at a dose of 25 mg/kg/day for 5 days. All patients were subjected to clinical and biological tests during the 7 days following the administration of second-line therapy.Data were collected and analysed using Epi info 6. Descriptive statistics were used to compare the demographic characteristics of the study population and their initial clinical and biological characteristics .Clinical and biological data were compared with a confidence interval of 95%. All the clinical and laboratory data collected were subjected to quality control. The study was carried out according to standard operating procedures following the guidelines of good clinical practice.A total of 955 patients who fulfilled the inclusion criteria were included in the study. All patients gave their informed consent before participating in the study. Treatment was allocated as follows: 360 were given artesunate-amodiaquine, 145 were treated with artesunate-mefloquine, 140 were given four doses of artemether-lumefantrine, 149 were given six doses of artemether-lumefantrine, and 161 received amodiaquine-SP (Am-SP).A comparison of the demographic and laboratory data at inclusion into the study showed that all treatments groups were practically identical Table .On day 14, the ACPR was 100% for artesunate-amodiaquine, artesunate-mefloquine and Am-SP. For artemether-lumefantrine given as four doses, one case of clinical failure was observed resulting in an ACPR of 99.3% Table . On day Figure All the strains responsible for clinical or parasitological failure between days 21 and 28 were compared with the strains collected on day 0 by PCR using the marker MSP 1. Differences in strains were assessed on agarose gels using the specific markers MAD 20, K1 and RO 33. This comparison of strains with genetic markers before and after 'clinical failure' resulted in a correction of the results. No case of recurrence was observed after treatment with artesunate-amodiaquine, artesunate-mefloquine and Am-SP and 100% of clinical 'failures' were shown to be the result of re-infection. In contrast, the level of re-infection with artemether-lumefantrine (four-doses) was 84% (20/24)  presented with a slight increase in creatinine levels without any serious clinical signs Table .The results of this study demonstrated that each of the five antimalarial drug regimens tested had good clinical and parasitological efficacy. When the efficacy results were corrected for PCR results, an ACPR of 100% was observed with artesunate-amodiaquine, artesunate-mefloquine, artemether-lumefantrine given as six doses, and Am-SP. Only artemether-lumefantrine given as four doses was associated with a failure rate of 3.6%. An initial study conducted in Senegal [In Asia, studies have demonstrated that in areas where transmission is high and where the population has a high level of immunity to malaria, a therapeutic approach based on four doses of artemether-lumefantrine gave good results. Nevertheless, in non-immune populations  this therapeutic approach was associated with a failure rate of approximately 20%. This failure rate was reduced to less than 5% by using a dosage schedule based on six doses . The faiP. falciparum encountered in this study were highly susceptible to mefloquine despite the small dose used (15 mg/kg mefloquine). However, to prevent a reduction in parasite susceptibility, an increase in the dosage of mefloquine to 25 mg/kg was necessary, as shown previously in other studies [The excellent efficacy observed with artesunate-mefloquine at the present dosage, seems to demonstrate good activity against the asexual form of the parasite. Nevertheless, under the endemic conditions in our region, transmission levels also confer a degree of protection which can act synergistically with a drug. No case of re-infection was observed after a follow-up period of 42 days. It appears, therefore, that the strains of  studies .The combination of Am-SP is still effective in Senegal and in central Africa , while iSince 2003, the Programme National de Lutte contre le Paludisme du S\u00e9n\u00e9gal  has used the Am-SP combination instead of chloroquine as a temporary strategy in the treatment of uncomplicated malaria. However, it is believed that the widespread use of this combination in addition to the use of SP alone in pregnant women together with population movements is likely to lead to the appearance of therapeutic resistance to these drugs. Indeed, a study carried out in Senegal on molecular markers of resistance to SP has demonstrated the presence of triple mutations for dhfr and dhps .The gametocidal effect of artemisinin derivatives against gametocytes has been demonstrated in other studies  and has Genotyping of resistant stains using MSP1 and the alleles K1, RO 33 and MAD 20 enabled us to differentiate cases of recurrence from new infections.All drug combinations were well tolerated in this study. Only minor adverse events were observed, as seen in other studies ,17 and tSimilarly, clinical and biological tolerability was good for all drug combinations. No unexpected adverse events were encountered and all disappeared at the end of treatment. No serious adverse event requiring premature termination of treatment was observed. However, it is important to carry out further pharmacovigilance studies before these combinations are used more widely.Artemisinin derivatives in combination with classical antimalarial drugs, which are still effective as monotherapy, represent, therefore, the best options for the treatment of malaria in situations where chloroquine resistance exists. Possible hurdles to their use as treatment strategies for uncomplicated malaria include their relatively high cost and their availability in the context of a high worldwide demand. Considerable commitment by the pharmaceutical industry and increased participation of international organizations are necessary to give the population of these poor countries access to antimalarial drug combinations based on artemisinin derivatives.On the basis of these results, Senegal has modified its approach to treatment of malaria since 2004 and has adopted artesunate in combination with amodiaquine as first-line therapy for the management of uncomplicated malaria. However, a regular monitoring of this strategy is essential because therapeutic failures were documented in the south of Senegal .FB : Co principal investigator. Supervised the study, the data's coll, made molecular biology study and wrote the paperNJL: Co investigator, Statistical analyserND: Molecular Biology advisor. Participate in the molecular biology studyDY: Co investigatorFO: Co investigatorGO: Co principal investigator. Initiated the study, revised the paperAll the authors have read and approved the last manuscript"}
+{"text": "Plasmodium falciparum in animal models and in small-scale human studies. We conducted a multicentric noninferiority trial to assess the safety and efficacy of Cotrifazid against drug-resistant malaria in Papua New Guinea.Previous studies of a fixed combination including cotrimoxazole, rifampicin, and isoniazid (Cotrifazid) showed efficacy against resistant strains of The trial design was open-label, block-randomised, comparative, and multicentric.The trial was conducted in four primary care health facilities, two in urban and two in rural areas of Madang and East Sepik Province, Papua New Guinea.Patients of all ages with recurrent uncomplicated malaria were included.Patients were randomly assigned to receive Cotrifazid, mefloquine, or the standard treatment of quinine with sulfadoxine\u2013pyrimethamine (SP).Incidence of clinical and laboratory adverse events and rate of clinical and/or parasitological failure at day 14 were recorded.P. falciparum asexual blood-stage) was higher for Cotrifazid than for mefloquine or quinine + SP .The safety analysis population included 123 patients assigned to Cotrifazid, 123 to mefloquine, and 123 to quinine + SP. The Cotrifazid group experienced lower overall incidence of adverse events than the other groups. Among the efficacy analysis population , clinical failure rate (symptoms and parasite load) on day 14 was equivalent for the three groups , but parasitological failure rate , malaria is an important cause of death and disease in both adults and children. But concerns exist about whether current antimalarial drugs will be viable for much longer, indicating that new treatments are urgently needed. In 2001, new recommendations on the first-line treatment for uncomplicated malaria were introduced in PNG. These recommendations specify treating older children and adults with the combination of chloroquine together with sulfadoxine\u2013pyrimethamine (SP). However, there is already evidence that malaria parasites in PNG are evolving resistance to this combination therapy. Therefore a group of researchers examined whether new combinations of existing drugs for other diseases could be applied to treatment of malaria in the region. They conducted a trial comparing three different therapies in adults and children over the age of six months who presented to primary care clinics with uncomplicated malaria. The therapies compared were mefloquine (Lariam), quinine taken together with SP, and Cotrifazid, a combination of three different drugs mainly used against tuberculosis. Participants in the trial were followed up for 14 days after treatment, and the main outcome the researchers looked at was treatment failure . The researchers also compared the rate of adverse events and presence of malaria parasites in the blood in the different treatment groups.What the trial shows: Clinical treatment failure at day 14 was very low (either 0% or close to 0%) and approximately equivalent in all three treatment groups. The researchers then compared presence of malaria parasites in the blood and found that a much higher proportion of patients treated with Cotrifazid than the other two treatments had parasites in the blood at day 14 . Overall the rate of adverse events was lower in the Cotrifazid group than in the other two treatment groups.Strengths and limitations: Studies like this one that examine novel antimalarial treatments are particularly timely, as there is an urgent need to find drugs that will treat malaria resistant to current therapies. In this trial the procedures for randomizing participants to the different treatments were appropriate. However, a key limitation is that patients were followed up for only 14 days, and longer follow-up (as many groups now recommend) might have allowed the researchers to more accurately detect differences in efficacy between the treatments being compared.Contribution to the evidence: Few properly randomized controlled trials have been conducted that look at the ability of Cotrifazid to treat malaria. The results of the trial presented here suggest that Cotrifazid is safe, and short-term clinical efficacy is approximately equivalent to mefloquine or quinine plus SP. However, since in this trial Cotrifazid-treated patients were more likely to have malaria parasites reappear in the blood, Cotrifazid does not seem to be a good alternative treatment in PNG. Plasmodium spp. strains. The increasing prevalence of strains of Plasmodium falciparum resistant to 4-aminoquinolines and antifolate drugs has created a crisis in the clinical treatment of malaria in many countries, including Papua New Guinea (PNG) [Despite widespread efforts to prevent and treat malaria, worldwide the burden of morbidity and mortality is still high, due in part to the spread of drug-resistant ea (PNG) . IntroduCompounds already on the market for specific indications can easily be screened for effects against neglected diseases. Based on promising preliminary data, a fixed combination including cotrimoxazole, rifampicin, and isoniazid (Cotrifazid) has been developed. It has the advantage of proven safety and tolerability at higher dosage and for longer duration (in tuberculosis).P. berghei malaria in rodents  [P. vivax when associated with primaquine [The rationale to use this combination is based on animal and human experiments. Rifampicin is active against ed data) . In humaimaquine . Numerouimaquine ,5. Thereimaquine . IsoniazThree studies on Cotrifazid have been conducted in humans in endemic areas, including infants under 6 months of age \u20139. TheseThe objective of this trial was to compare, in patients with chloroquine- or amodiaquine-resistant malaria, the efficacy and safety of Cotrifazid to that of mefloquine or quinine + SP. The MRAC  of Papua New Guinea asked that the mefloquine arm be included, to obtain in Papua New Guinean patients reliable safety and efficacy data on alternative drugs that had shown good potential elsewhere.All patients older than 6 months of age presenting at the centres (described below) who were diagnosed with malaria , and who had already been treated for malaria in the 28 days before, qualified for inclusion in the study if the individual or legal guardian (for children) gave informed consent and if the clinician in charge would have given the standard treatment for drug-resistant malaria independent of the study. A participant was excluded if the clinician preferred to use quinine + SP for any reason , or if the patient had one of the symptoms or signs of complicated or severe malaria , had contraindications for mefloquine (history of psychiatric disorder or epilepsy), or was pregnant.Plasmodium species), patients were screened for other inclusion and exclusion criteria and recruited when appropriate.All patients suspected on clinical grounds to have drug-resistant malaria were investigated by a nurse employed specifically for the trial who performed the OptiMAL test . If the latter was positive , was supplied as a coated tablet that could be stored at room temperature. The treatment dosage and schedule for patients was as follows. Patients weighing 40 kg or more: two tablets twice per day; patients under 40 kg and 20 kg or more: one tablets twice per day; patients under 20 kg: one-half tablet twice day. These doses were given every 12 hours for 7 days (days 0\u20136).The treatment regimen for mefloquine  was given at the usual dosage of 25 mg/kg in children, 1,250 mg for adults less than 60 kg, and 1,500 mg for adults over 60 kg, split in two doses at hours 0 and 12.Quinine  was given as usual: 10 mg/kg every 8 hours for 5 days (days 0\u20134). In this regimen, patients also received a single dose of 0.5\u20133 tablets of Fansidar  on day 0.The doses in the morning of days 0\u20133 were administered under the supervision of the appointed nurses at the study centres. They were responsible for confirming both the participant identification number and the label of the trial drugs. Intake of the afternoon doses on day 3 was checked by interview on day 4, and intake of doses on days 4\u20136 by interview on day 7.Follow-up assessments were done on days 1, 2, and 3 , 7 , and 14 , or more intensively in individual cases of persisting symptoms or pathological signs. Blood samples were taken by venipuncture (2 ml) on days 0 and 7 and by fingerpick on days 1, 2, 3, and 14. For details on assessment procedures, see \u201cLaboratory Procedures\u201d below.Treatment was changed to quinine + SP if the patient failed treatment with other drugs. There was no rescue treatment if the failure occurred after quinine + SP; a second course with the same drugs was to be given, as stated in the national guidelines at the time of the study.The specific objectives of the trial were (i) to compare the efficacy of Cotrifazid to that of the standard treatment for drug-resistant malaria in PNG (quinine + SP) and to another drug (mefloquine) that is being considered for future use, and (ii) to compare the tolerance of Cotrifazid with that of quinine + SP and mefloquine in the same population. The trial was designed to test the null hypothesis that the clinical cure rate with Cotrifazid is not inferior to that with the comparators (mefloquine or quinine + SP).The prime measurement of safety was incidence of clinical or laboratory adverse events (AEs). All patients were followed clinically every day for the first four days, and longer in cases of complication. All patients were seen on days 7 and 14 to identify late AEs or clinical failure. Laboratory measurements included serum glutamic-oxaloacetic transaminase (aspartate aminotransferase) (SGOT), serum glutamate-pyruvate transaminase  (SGPT), and creatinine on days 0 and 7, and Hb concentration on days 0 and 14.AEs were defined according to standard criteria, i.e., any adverse change from the participant's baseline (pretreatment) condition , irrespective of whether the event was considered related to the trial drug or not. The occurrence of serious AEs, as defined by standard criteria, was recorded and acted upon. There was no assessment of AE intensity or relatedness to the product investigated because of the confounding effect of malaria symptoms and signs.The primary parameter of efficacy was the clinical treatment failure rate on day 14, using a blood slide as gold standard for parasitology. Clinical treatment failure was defined as (i) the occurrence of severe malaria between day 1 and day 14, or (ii) the persistence or recurrence of symptoms or signs (including temperature >37.5 \u00b0C) associated with any parasitaemia between day 5 and day 14.Secondary parameters of efficacy were (i) parasitological failure rate on day 14, (ii) fever clearance time, (iii) parasite clearance time, (iv) symptom clearance time, (v) occurrence of complications  and Hb concentration (from samples collected on day 14).The overall sample size of 330  was chosen to test the hypothesis that Cotrifazid was not inferior to mefloquine or quinine + SP, assuming a rate of treatment success of 95% with mefloquine or quinine + SP and a clinically acceptable rate of treatment success of 86% or more in the Cotrifazid group, and a 10% loss to follow-up  .Assignment to treatment groups was done by a randomisation list , which had been computer-generated (SAS Software) by a statistician of the Swiss Tropical Institute not involved in the study. The original list was kept at the Swiss Tropical Institute and a copy at the PNG Institute of Medical Research headquarters.Sequential numerical codes  were written on the reverse side of sealed envelopes that had been prepared at the Swiss Tropical Institute and forwarded to the local investigator prior to the start of the study. Inside the envelope, the treatment group was concealed on a paper with either the letter \u201cL\u201d for Lariam (mefloquine), \u201cC\u201d for Cotrifazid, or \u201cS\u201d for standard (quinine + SP) written on it.Once a patient had met the inclusion criteria and his or her (or the guardian's) consent had been given, the patient was assigned the code number following the one of the previous patient. The envelope corresponding to that code number was opened by the research nurse, and the first dose of the allocated treatment administered under supervision. The entire process was to be completed before any procedure was started for the following patient.This was an open-labelled trial. Neither the research nurse nor the patients were blinded to the treatment given.The OptiMAL test was used at screening (day 0) to document malaria.On days 0, 1, 2, 3, 7, and 14 malarial parasites were assessed by microscopy. Thick and thin films were stained with Giemsa at pH 7.2. Before a slide was declared negative 100 thick film fields were examined by microscopy. The number of malaria parasites per 200 white blood cells was counted. The number of asexual forms per \u03bcl was then calculated using a mean white blood cell count of 10,000/\u03bcl in children below the age of 5 y and of 8,000/\u03bcl in persons aged 5 y or older. The standard quality control performed at the PNG Institute of Medical Research was applied .Hb concentration was determined using a photometer .SGOT, SGPT, and creatinine were measured with an automated dry chemical photometer . Regular quality controls were run for all the measurements.P. falciparum genotype profiles were assessed on samples of day 0 and day of failure using a combination of several molecular markers of the SP resistance genes dhfr and dhps.The trial was approved by the Medical Research Advisory Committee of PNG. Verbal informed consent from each participant in the trial or from his/her parents or guardian(s) was obtained in front of a witness after explanation of the aims, methods, benefits, and potential hazards of the trial.A trial monitor  reviewed all procedures and ensured complete adherence to the protocol.Data were double-entered using a specific program written in FoxPro software version 3.0 and analysed using Stata software version 8.2.All participants who received at least one daily dose of Cotrifazid or comparators and who presented to one follow-up visit were included in the safety analysis population. Comparisons of the incidence of AEs (following prompted questions) reported at follow-up contacts (days 1\u201314) were performed using a Poisson model. Axillary temperatures, respiratory rates, and Hb values are reported as mean \u00b1 standard deviation (SD).All participants who showed asexual parasites (any species) at baseline and who received the treatment with Cotrifazid, mefloquine, or quinine + SP for at least the first three days (days 0\u20132), and who presented to the follow-up visit on day 14, were included in the per protocol efficacy analysis population. Efficacy was estimated by comparing the proportions of complications and the rates of clinical and parasitological failures between the Cotrifazid and comparator groups, using the Mantel-Haenzel Chi-square test with \u03b1 = 0.05, one-tailed) or Fisher's exact test where appropriate. A Kaplan-Meier survival analysis of all asexual parasites, fever, and symptoms was performed, and treatment groups compared using the log rank test. The Student's t-test was used to compare means of Hb concentration between the groups.An intention-to-treat analysis was also performed to assess parasitological outcomes at day 14, counting all children who didn't appear for assessment on that day as failures.The safety analysis population included 137 patients in the Maprik hospital outpatient clinic, 49 in Kunjingini health subcentre, and 183 in Madang town outpatient clinic. No patient was admitted to the ward. The participant flow is detailed in In the efficacy analysis population for the outcome measures , 32 patients were lost to follow-up on day 14, which left 218 patients  see .The study was conducted from April 2000 to January 2003.The female:male ratio was 180:184 (five were unknown). In the Cotrifazid group the median age was 7.2 y (range 1.4\u201345 y), mefloquine 7.4 y (range 0.5\u201356 y), and quinine + SP 8 y (range 0.5\u201361 y).The prevalence of reported symptoms and observed signs at baseline for each treatment group is described in P. falciparum in the Cotrifazid group, 90% (75/83) in the mefloquine group, and 87% (71/82) in the quinine + SP group. The corresponding values for P. vivax were 8.2% (7/85), 9.6% (8/83), and 23% (19/83). A higher prevalence of mixed infections was found in the quinine + SP group (10% versus 4% in the Cotrifazid group and 4% in the mefloquine group). Geometric mean densities were equivalent.Parasitology at baseline showed that 94% (80/85) of the patients were infected with Full compliance with the treatment evaluated from direct observation in the first three days and history-taking thereafter was 93% (115/123) in the Cotrifazid group , 98% (121/123) in the mefloquine group (2 d), and 96% (118/123) in the quinine + SP group (5 d). Two patients interrupted their treatment due to AEs, both in the quinine + SP group; both were cured although none of them took an alternative drug.No deaths occurred in this study. Among the 369 patients included in the safety analysis, only one patient suffered from a serious AE. This 1.7-y-old child, who was enrolled in the Cotrifazid group, required hospital admission due to the persistence of high fever, cough, and vomiting on day 3. The child had 32,640 parasites/\u03bcl on day 0, 40 parasites/\u03bcl on day 2, and none from day 3 onwards. Thus, the fever was very likely of nonmalarial origin. The child was put on standard treatment with quinine + SP and antibiotics, and made a full recovery. One child in the Cotrifazid and two in the mefloquine group experienced an impairment of their level of consciousness, but none was severe enough to warrant rescue treatment with quinine + SP or hospital admission. One additional child developed chest indrawing during the follow-up in the Cotrifazid group. In the Cotrifazid group, four participants had yellow eye colour during follow-up , but none in the other groups. The few other non-prompted AEs\u2014weakness in four patients, red urine in three , sneezing in three, facial swelling in one, and swollen lymph nodes in one\u2014were considered isolated events, and none required the administration of ancillary treatment.p < 0.001); stomach ache (p < 0.05), vomiting (p < 0.01), dizziness (p < 0.05), and tinnitus (p < 0.01) were less common in the Cotrifazid than in the quinine + SP group. In contrast, participants in the Cotrifazid group were reported shortness of breath significantly more often than did those in the mefloquine group (p < 0.01) . However, due to the low sample size finally included in the efficacy analysis population, we had only 80% power to find a 4-fold increase in failure rate, assuming a failure rate of 5% in the quinine + SP group.Among the 218 patients retained in the efficacy analysis population, no patients developed signs of severe malaria based on the clinician's judgment, nor did any require secondary hospital admission due to malaria. No patients who were initially treated with Cotrifazid or mefloquine required rescue treatment with quinine + SP. No clinical treatment failures occurred with Cotrifazid or mefloquine, but one late treatment failure (patient was asymptomatic but had a temperature of 37.7 \u00b0C and parasite density of 80/\u03bcl on day 14) occurred in the quinine + SP group. The three treatment groups were therefore equivalent with regard to the clinical cure rate  are detailed in Plasmodium-positive patients was significantly higher in the Cotrifazid group at 15% (11/72) than in the mefloquine group, 0% (0/71), and the quinine + SP group at 4% (3/75) . The rates for P. falciparum on day 14 were 8% (6/72), 0% (0/71), and 3% (2/75) respectively .When all patients sampled on a given day were taken into account (irrespective of the other time points and species at baseline), there was no difference in parasite clearance on day 1, 2, 3 and 7 . However, on day 14, the prevalence of asexual blood-stage P. falciparum only at baseline and during follow-up were considered, 9% (6/67) had asexual blood-stage parasitaemia on day 14 in the Cotrifazid group versus 0% (0/63) in the mefloquine group and 3% (2/65) in the quinine + SP group . This difference was not significant when the Cotrifazid group was compared to the quinine + SP group  of the failures were due to new infections, which gives a PCR-corrected failure rate of 8% (versus 9% uncorrected). Patients in the Cotrifazid group were more likely to develop P. falciparum gametocytaemia on day 14 than those in the mefloquine or quinine + SP groups: 38% (18/48) versus 5% (2/44) and 8% (4/53), respectively  (When patients with SP group . Parasitct test) .P. vivax only at baseline and during follow-up were considered, the parasite prevalence on day 14 was 43% (3/7) in the Cotrifazid group, 0% (0/8) in the mefloquine group, and 0% (0/15) in the quinine + SP group .When patients with p < 0.001, Fisher's exact test). The rates for P. falciparum were 22%, 15%, and 11% respectively, and those for P. vivax 21%, 15%, and 10% . There was also a tendency for a slower clearance of P. falciparum in the Cotrifazid group as compared to the other groups, but the differences did not reach statistical significance.p = 0.77) or on day 14 (p = 0.73).The mean Hb concentrations on day 14 were increased compared to the pretreatment values on day 0: 10.4 \u00b1 1.9 (post-treatment) versus 9.54 \u00b1 2.1 g/dl (Cotrifazid group), 10.4 \u00b1 2.2 (post-treatment) versus 9.37 \u00b1 2.0 g/dl (mefloquine group), and 10.2 \u00b1 1.9 (post-treatment) versus 9.52 \u00b1 1.8 g/dl (quinine + SP group). There was no statistical difference between groups on day 0 , but is insufficient to clear all parasites, especially those of The overall incidence of prompted AEs in the Cotrifazid group was lower than that observed in the mefloquine and quinine + SP groups. The types of AEs recorded in these last two groups were consistent with those of the literature, which also validates the data recorded for Cotrifazid. Except for \u201cshortness of breath,\u201d none of the prompted AEs was more frequent in the Cotrifazid group . The preP. falciparum at baseline had recurrent parasites by microscopy on day 14. At the time of study design (1999), we decided on a 14-day follow-up since it was, and is still now, the standard duration chosen to determine policy change in areas of intense transmission [Although the final sample size for efficacy analysis was smaller than expected because the OptiMAL test used for screening was positive in the presence of gametocytes only, we were able to demonstrate equivalence of the three regimens in curing uncomplicated malaria (primary outcome). Indeed, all treatment groups had almost 0% clinical failure, a rate that was lower than the 5% expected in the sample size calculation. The short duration of our follow-up is likely to have impacted positively on treatment outcome, although it is not different from the one used in several recent trials. A follow-up of 28\u201342 d, as proposed by White  for longsmission . Despitesmission \u201324.Plasmodium species) on day 14 versus 4% for quinine + SP and 0% for mefloquine. These results reflect slower and less effective parasite clearance  were true recrudescence, as expected with a follow-up of 14 days, as well as from results of previous studies done in PNG [P. vivax, but a recurrence is more likely than a new infection or relapse in a time period of 14 days. Circulating asexual stages of P. vivax after blood schizonticidal therapy might originate from asexual parasites that survived therapy, from activated hypnozoites that led to a relapse, or from a new infection. Unlike with P. falciparum infections, in which true recrudescence can be distinguished from new infections by the use of genotyping methods, current molecular methods used for the genetic analysis of P. vivax do not allow the unambiguous classification of recurrent parasitaemia, and hence of true treatment failure.The situation is less promising when considering parasitological failures. Of the patients treated with Cotrifazid, 15% were parasitaemic  [Three studies were conducted with Cotrifazid in the past, all of rather small scale and one with serious methodological flaws  \u20139. The pWe believe that the design and power of our trial was optimized to accurately assess Cotrifazid safety and efficacy, and the results therefore should robustly support any conclusions on the usefulness of Cotrifazid as an antimalarial drug. Despite its very good safety profile, short-term clinical equivalence with effective drugs such as mefloquine, and low cost, Cotrifazid for malaria does not appear to be an ideal alternative therapeutic option in its current formulation and regimen: patients experience a slower parasite clearance, some recurrence of asexual forms, and higher gametocytaemia on day 14 than after mefloquine or quinine + SP treatment. However, in formulating new and affordable combinations of antimalarial drugs in the future, the clinical efficacy of the Cotrifazid components could be taken into account, especially if it is considered an advantage, as part of the Integrated Management of Childhood Illness strategy , to use CONSORT ChecklistClick here for additional data file.(45 KB DOC)Trial ProtocolClick here for additional data file.(133 KB DOC)"}
+{"text": "Plasmodium falciparum. The fixed combination of oral artemether-lumefantrine, an artemisinin combination therapy (ACT) is highly effective and well tolerated. It is the only registered fixed combination containing an artemisinin. The trial presented here was conducted to monitor the efficacy of the six-dose regimen of artemether-lumefantrine (ALN) in an area of multi-drug resistance, along the Thai-Myanmar border.The use of antimalarial drug combinations with artemisinin derivatives is recommended to overcome drug resistance in P. falciparum infections. The PCR adjusted cure rates were evaluated by survival analysis.The trial was an open-label, two-arm, randomized study comparing artemether-lumefantrine and mefloquine-artesunate for the treatment of uncomplicated falciparum malaria with 42 days of follow up. Parasite genotyping by polymerase chain reaction (PCR) was used to distinguish recrudescent from newly acquired P. falciparum malaria were randomly assigned to receive artemether-lumefantrine (n = 245) or artesunate and mefloquine (n = 245) and were followed for 42 days. All patients had rapid initial clinical and parasitological responses. In both groups, the PCR adjusted cure rates by day 42 were high: 98.8%  for artemether-lumefantrine and 96.3%  for artesunate-mefloquine. Both regimens were very well tolerated with no serious adverse events observed attributable to either combination.In 2001\u20132002 a total of 490 patients with slide confirmed uncomplicated Overall, this study confirms that these two artemisinin-based combinations remain highly effective and result in equivalent therapeutic responses in the treatment of highly drug-resistant falciparum malaria. Plasmodium falciparum is a major health problem in many countries and the number of drugs available, effective and affordable is very limited [P. falciparum has developed resistance to almost all available antimalarials [Artemisia annua), which together with its derivatives, artesunate and artemether are the most active antimalarial compounds to date [4 every 48 hours. In addition, they have a very short terminal elimination half-life of less than 2 hours [P. falciparum infections in Thailand [P. falciparum infections in adults and children on the western border of Thailand.Multi-drug resistance of  limited . Along talarials . As in talarials . The usealarials . An extealarials . Artemis to date . The art 2 hours . PreviouThailand . Oral arThailand . Both arThailand . HoweverPlasmodium vivax and multi-drug-resistant P. falciparum [This study was conducted in the Maela and Mawker Tai malaria clinics of the Shoklo Malaria Research Unit between July 2001 and June 2002. Patients were recruited from two populations: displaced people of the Karen ethnic minority and migrant workers living on the western border of Thailand. This is an area of low and unstable transmission of lciparum . The triP. falciparum malaria were included in the study, provided that they or their guardians gave fully informed written consent intheir own language, they were not pregnant, they had not received mefloquine in the previous 63 days and there were no other clinical or laboratory signs of severe illness and/or severe and complicated malaria [\u00ae 20/120, Novartis Pharma AG, Basel, Switzerland) or mefloquine  plus artesunate . At enrolment (Day 0), a medical history was obtained, a full physical examination was performed and blood was taken for quantitative parasite counts and routine haematology . All information was recorded on a standard case record form. All patients were examined and blood smears were taken daily until they became aparasitaemic, and then weekly for 6 weeks. At each visit a questionnaire on adverse events was completed. A blood smear was also taken from any patient complaining of fever or symptoms compatible with malaria during the follow-up period. Parasite counts were determined on Giemsa-stained thick and thin blood films. The person-gametocyte-weeks were calculated per 1,000 person-weeks after excluding the episodes on admission and during treatment.Patients >10 kg in weight who had slide-confirmed acute  malaria . If theyComputerized randomization was in blocks of ten. Patients allocated to artemether-lumefantrine group (ALN) received the tablets at 0 and 8 hours and twice daily for the following 2 days. Artemether-lumefantrine was dispensed as a fixed dose combination tablet. Each tablet contained 20 mg of artemether and 120 mg of lumefantrine. The number of tablets was given according to the body weight. The minimum dosage for patients weighing less than 15 kg was one tablet per dose; patients between 15 and 24 kg received two tablets, those between 25 and 34 kg received three tablets and patients 35 kg and above were treated with four tablets per dose. Patients allocated to artesunate-mefloquine group (MAS3) received artesunate, 4 mg/kg oncedaily for 3 days (day 0 was the first day of treatment), plus mefloquine, 15 mg/kg on day 1 and 10 mg/kg on day 2.Each patient was given antipyretics and cooled by tepid sponging if the tympanic temperature was equal or above 37.5\u00b0C before drug administration. Drug administration was observed in all patients and if vomiting occurred in less than 30 min, administration of the full dose was repeated, if vomiting occurred between 30 and 60 min, half the dose was repeated. Patients treated with artemether-lumefantrine were given a glass of chocolate milk (200 ml) with each dose to increase absorption .P. falciparum infections. P. falciparum infections were genotyped for allelic variation in three polymorphic antigen loci, merozoite surface proteins 1 and 2 (MSP-1 and MSP-2) and glutamate rich protein (GLURP), on admission and in case of parasite reappearance [The primary therapeutic outcome measure in this study was the incidence of microscopically and genotypically confirmedrecrudescent infections in both treatment groups by day 42. Parasite genotyping by the polymerase chain reaction (PCR) was used to distinguish recrudescent from newly acquired pearance ,16. SecoAdverse events were symptoms or signs that were not presenton admission and that developed after the start of treatment. All adverse events, including those probably related to malaria, were recording and compared among treatment groups. The rates of early vomiting (<1 h) after each dose and for each drug wererecorded and compared among the groups in the analysis.Patients with uncomplicated recrudescent infections were re-treated with artesunate, 2 mg/kg/day for 7 days; patients >8 years oldalso received doxycycline, 4 mg/kg/day for 7 days.t test. Data not normally distributed were log-transformed or compared using the Mann-Whitney U test. The relative risks were calculated using cross-tabulations. The rates of adverse events at three different periods  were compared among treatment groups. For each of the three periods, the events were counted only once . The PCR-adjusted cure rates were evaluated by survival analysis and compared using the log-rank test. Patients for whom PCR genotyping was either inconclusive or missing were censored in the survival analysis on the day of parasite reappearance. For all statistical tests the significance level (p) was set at 0.05.Data were analysed using SPSS for Windows, version 11. Categorical data were compared using the Chi-square test with Yates' correction or by Fisher's exact test, as appropriate. Continuous variables conforming to a normal distribution were compared using Student's P. falciparum infections were enrolled between July 2001 and May 2002. The artesunate-mefloquine and artemether-lumefantrine groups included 245 patients each, the age range for all patients was 2\u201372 years. Baseline characteristics were similar in both groups  of 227 patients in the artemether-lumefantrine recipients and three (1.3%) of 238 artesunate-mefloquine recipients still had a positive blood film (P > 0.05). Overall, 12.3 % of patients were anaemic (haematocrit <30%) on admission, 10.8% in ALN group and 13.8% in MAS3 group (P = 0.33). The mean (SD) decrease in haematocrit value at day seven from baseline was greater in the group receiving MAS3 than in the ALN group: 9.3%  compared with 6.7%  respectively (P = 0.023).The initial responses to the two treatment groups were similar. None of the patients developed severe malaria. On admission, 55.0% 133/242) of the patients on ALN and 57.9% (140/242) of the patients in the MAS3 group had a tympanic temperature \u2265 37.5\u00b0C. All except three patients had a normal temperature on day 3 (2 in ALN and 1 in MAS3). There was no difference in fever clearance times between the two treatment groups  (Table P = 0.08). PCR confirmed treatment failures were more likely in children aged below 15 years than in adults . The mean age was 13.6 years  in patients with treatment failure and 23.7 years  in successfully treated patients. In this trial, only age group was independently associated with treatment failure, but not other factors e.g. higher parasitaemia ; anaemia (haematocrit <30%); fever (tympanic temp \u2265 37.5\u00b0C) on admission; sites; treatment groups; early vomiting (within one hour following drug administration). The mediantime to recrudescence was comparable for MAS3 group  and ALN group .During the 42-day follow-up period, 27 new P. vivax parasitaemia detected during follow up. There were significantly fewer cases of vivax malaria in the MAS3 group (29 of 227) than in the ALN group (90 of 225) (P < 0.001). The median time to appearance of P. vivax parasitaemia was longer in the MAS3 group  than in the ALN group .Of 452 patients, 119 (26.3%) had Twenty patients (8.3%) in the artemether-lumefantrine group and 19 (7.9%) in the artesunate-mefloquine group had gametocytes detected during the first 3 days. All except one patient in ALN group cleared gametocytes within first week after start of treatment. After excluding these, gametocytes developed (between day 7 and 42) in 1.2% (3/241) of ALN group and 1.3 (3/240) of MAS3 group. The person-gametocyte weeks were low and similar: 2.7  per 1000 person-weeks for both groups.P = 0.45). The rates of early vomiting (within one hour) of the drugs were very low (around 2%) and did not differ among groups (one in each group on day 2).Both treatment regimens were well tolerated. No serious adverse events were reported. Overall, 5/242(2.1%) of the patients vomited one or more doses of medication in the ALN group and 2/242(0.8%) of the MAS3 treated patients  and central nervous system . Figure P. falciparum are found, artesunate-mefloquine combination therapy (MAS3) is the standard treatment regimen for uncomplicated falciparum malaria [The loss of affordable effective antimalarial drugs to resistance represents a major threat to the people of malaria endemic countries . Using i malaria . Early d malaria . Artemisfalciparum malaria. In Thailand, several trials have been conducted with this combination. The six-dose schedule provides sustained blood lumefantrine levels and thus improved cure rates [Artemether-lumefantrine has been introduced recently for oral treatment of uncomplicated re rates . Lumefanre rates .P. vivax infections and 12 days longer median time to appearance of P. vivax parasitaemia in the MAS3 group were most probably due to the longer terminal half-life of mefloquine compared to lumefantrine [The present trial reconfirmed the efficacy of the six-dose regimen of artemether-lumefantrine given over three days . Both trfantrine ,22. MoreRH carried out the study and analyzed the data. RH, EAA, RMG, PS, TJ, NJW, FN conceived the study, participated in its design and co-ordination and contributed to draft the manuscript. LP, KLT assisted in collection of data. AB performed the PCR experiments. All authors read and approved the final manuscript."}
+{"text": "P. falciparum gametocytes may persist after treatment with sulphadoxine-pyrimethamine (SP) plus artesunate (AS) and contribute considerably to malaria transmission. We determined the efficacy of SP+AS plus a single dose of primaquine  on clearing gametocytaemia measured by molecular methods.P. falciparum malaria with an asexual parasite density between 500\u2013100,000 parasites/\u00b5L were randomized to receive treatment with either SP+AS or SP+AS+PQ. P. falciparum gametocyte prevalence and density during the 42-day follow-up period were determined by real-time nucleic acid sequence-based amplification (QT-NASBA). Haemoglobin levels (Hb) were determined to address concerns about haemolysis in G6PD-deficient individuals.The study was conducted in Mnyuzi, an area of hyperendemic malaria in north-eastern Tanzania. Children aged 3\u201315 years with uncomplicated Pfs25 QT-NASBA gametocyte prevalence was 88\u201391% at enrolment and decreased afterwards for both treatment arms. Gametocyte prevalence and density were significantly lower in children treated with SP+AS+PQ. On day 14 after treatment 3.9% (2/51) of the SP+AS+PQ treated children harboured gametocytes compared to 62.7% (32/51) of those treated with SP+AS (p<0.001). Hb levels were reduced in the week following treatment with SP+AS+PQ and this reduction was related to G6PD deficiency. The Hb levels of all patients recovered to pre-treatment levels or greater within one month after treatment.108 individuals were randomized. PQ clears submicroscopic gametocytes after treatment with SP+AS and the persisting gametocytes circulated at densities that are unlikely to contribute to malaria transmission. For individuals without severe anaemia, addition of a single dose of PQ to an efficacious antimalarial drug combination is a safe approach to reduce malaria transmission following treatment.ISRCTN61534963Controlled-Trials.com  Plasmodium falciparum that are responsible for clinical disease and death. Sexual stage parasites, gametocytes, can also be present in infected individuals and are responsible for the transmission of the parasite to mosquitoes. Drugs specifically targeting these sexual stage parasites may affect the spread of malaria in the human population. Anti-gametocyticidal drugs are used by several countries to prevent onward transmission from clinical malaria cases The majority of anti-malarial drug treatments target the asexual blood stages of P. vivax malaria and actively clears mature P. falciparum gametocytes P. falciparum gametocytes detected by microscopy Artemisinin-based combination therapies (ACT) are advocated as first-line antimalarial treatment because of their high treatment efficacy P. falciparum gametocytaemia in an area of hyperendemic malaria in north eastern Tanzania. Possible haemolytic effects of PQ were determined in relation to G6PD status.Here, we determine the safety and efficacy of SP+AS plus a single dose of PQ on clearing submicroscopic levels of http://www.controlled-trials.com/ISRCTN61534963/. Registration was done after patient recruitment started due to communication problems.The protocol for this trial and supporting CONSORT checklist are available as supporting information; see P. falciparum mono-infection at a density between 500\u2013100,000 parasites/\u00b5L were eligible for recruitment. Exclusion criteria were: a haemoglobin (Hb) concentration measured by Hemocue\u00ae below 8g/dL, inability to take drugs orally, known hypersensitivity to any of the drugs given, reported treatment with antimalarial chemotherapy in the past 2 weeks, evidence of chronic disease or acute infection other than malaria, domicile outside the study area, signs of severe malaria and eligibility for other malaria studies conducted in the region.This study was conducted in the period July through September 2006 in Mnyuzi, a rural village in the Tanga Region, north eastern Tanzania. Malaria transmission intensity is high with an estimated entomological inoculation rate (EIR) of 91 infectious bites per person per year Participants enrolled were randomized to one of the two treatment regimes:plus artesunate (AS), 4 mg/kg once daily for three days  plus placebo once on the third day ;Sulphadoxine (25 mg/kg) and pyrimethamine (1.25 mg/kg) as a single dose  plus AS plus primaquine  as a single dose on the third day (0.75 mg/kg). Primaquine capsules were produced following regulations of the European Pharmacopeia.SP Treatment was administered by staff at the recruitment clinic. Each child was observed for 30 minutes after treatment, a replacement dose was given in case of vomiting. None of the children had repeated vomiting. Paracetamol (10 mg/kg) was given until symptoms had subsided. In case of parasitological treatment failure, rescue treatment with mefloquine was administered . All staff engaged in the trial were blinded as to the treatment group of each child, apart from the study physician who administered medication.P. falciparum gametocyte prevalence and density and to determine the safety of a single dose of PQ in glucose-6-phosphate-dehydrogenase (G6PD) deficient children.Our primary objectives were to determine the effect of SP+AS and SP+AS+PQ on submicroscopic The primary outcomes were gametocyte prevalence and density by real-time nucleic acid sequence-based amplification (QT-NASBA). The secondary outcome was haemoglobin concentration following treatment. Other outcomes that we evaluated were microscopic gametocyte prevalence, treatment efficacy and the occurrence of side effects.Participants were encouraged to attend the recruiting clinic at day 1, 2, 3, 7, 14, 28 and 42 after enrolment and at any time the child became unwell. On each day of follow-up, tympanic temperature was measured by electric thermometer and a finger prick blood sample was used for haemoglobin (Hb) measurement using a Hemocue photometer , a microscopic slide, a 50 \u00b5L-blood sample for real-time nucleic acid sequence-based amplification (QT-NASBA) and a filter paper sample. The presence of symptoms suggestive of anaemia  or allergic drug reactions (rash) was assessed verbally during every follow-up visit. Field assistants visited the homes of children who failed to show up to collect additional samples.Blood smears were stained for 10 minutes with 10% Giemsa and screened for asexual parasites and gametocytes at enrolment and on day 3, 7, 14, 28 and 42 after treatment. All slides were double-read by experienced microscopists and were declared negative if no parasites were observed in 100 microscopic fields. Readings were compared for validation and slides giving discordant results were read by a third reader. The majority result was taken as final in the case of positive versus negative results and geometric mean of the two closest values for density discordants . Asexual parasites and gametocytes were counted against 200 and 500 white blood cells, respectively and converted to parasites/\u00b5L by assuming a density of 8000 white blood cells/\u00b5L blood.P. falciparum parasite detection by QT-NASBA was performed as described previously in vitro cultured mature NF54 gametocytes was included in each run to ascertain gametocyte density et al. et al.2O and 50 \u00b5L of 20% chelex suspension in distilled water (pH 9.5) and centrifugated at 5000 rpm for 10 minutes. 1 \u00b5L of supernatant was used in the PCR reactions.DNA extraction from bloodspots on filter paper was carried out by the chelex-100 method as described by Wooden P. falciparum genes msp1  and msp2 (IC1 and FC27) was performed as described by Snounou et alet al. was followed in that indeterminate samples for which a majority of novel bands appeared for the post-treatment infection were scored as new infections To differentiate between recrudescent parasites i.e. those persisting from the initial infection and parasites from a new infection, a nested PCR amplification of the polymorphic regions of G6PD deficiency was determined by screening human DNA for single nucleotide polymorphisms in the G6PD gene  by a simple high throughput method using PCR, sequence specific oligonucleotide probes (SSOPs) and ELISA-based technology Pfs25 QT-NASBA gametocyte prevalence after treatment. Assuming a gametocyte prevalence in the SP+AS group of 50% on day 14 after treatment Pfs25 QT-NASBA gametocyte prevalence of 58% in SP+AS treated children The primary endpoint used for sample size calculation was Pfs25 QT-NASBA and haemoglobin concentrations were determined by technicians who were unaware of the treatment allocated to study participants.The randomization sequence was generated in Stata 8.0  using restricted randomization with a block size of 20. Treatment allocation was determined by opening pre-prepared randomization envelopes in sequence by the study physician. The same physician was involved in participant selection and clinical evaluation. Parasite carriage by microscopy and Pfs25 QT-NASBA gametocyte density versus time et al.0+g3)/2+(7\u22123)\u00d7(g3+g7)/2+(14\u22127)\u00d7(g7+g14)/2+(28\u221214)\u00d7(g14+g28)/2+(42\u221228)\u00d7(g28+g42)/2]/42; where gd represents Pfs25 QT-NASBA gametocyte density on day d. Gametocyte negative samples were included as zeroes. The measure was scaled by 42 so that it represents AUC per day and this was transformed by log10. Microscopic and QT-NASBA parasite densities were analysed after log10-transformation. Because we were interested in clearance of gametocytes of the original infection, slides from individuals in which PCR analysis defined a new infection were excluded from analyses on post-treatment gametocyte prevalence and density.Therapeutic outcome was classified as early parasitological treatment failure (ETF), late treatment failure (LTF), re-infection or adequate clinical and parasitological response (ACPR) Proportions were compared using the chi-squared statistic for a 2-by-2 contingency table. Normally-distributed continuous variables were compared using the Student t-test. Variables that were not normally distributed were compared using the Wilcoxon rank-sum test. Multiple linear regression models were used in case of continuous variables to adjust for potential confounding factors such as asexual parasite and gametocyte density at enrolment. Multiple logistic regression models with Generalized Estimating Equations (GEE) were used to test the influence of treatment on dichotomous variables with multiple observations per participant, such as gametocyte prevalence during follow-up. Estimates were adjusted for potential confounding factors and a random effect was included in the models to allow for correlations within individuals. Regression coefficients (\u03b2) were calculated for continuous dependent variables and odds ratios (OR) for dichotomous dependent variables, both with 95% confidence intervals (95% CI). Statistical analyses were performed using SPSS for Windows 12.0  and Stata 8.0 .2\u200a=\u200a1.70; p\u200a=\u200a0.64).Microscopic gametocyte prevalence at enrolment was 26.4% (14/53) for children treated with SP+AS and 18.9% (10/53) for children treated with SP+AS+PQ . MicroscPfs25 QT-NASBA was 88.2% (45/51) for SP+AS treated individuals compared to 90.6% (48/53) for SP+AS+PQ treated children  compared to those gametocyte-free by microscopy .Enrolment gametocyte prevalence defined by children . PredictPfs25 QT-NASBA gametocyte density was not initially defined as outcome measure, but densities were compared between the treatment arms post-hoc. At the time of enrolment Pfs25 QT-NASBA gametocyte density was 28.8 gametocytes/\u00b5L (IQR 6.9\u2013109.9 gametocytes/\u00b5L) for SP+AS treated children compared to 17.5 gametocytes/\u00b5L (IQR 1.1\u201376.9) for SP+AS+PQ treated children , p<0.001). The Pfs25 QT-NASBA gametocyte density in gametocyte positive samples was consistently lower for SP+AS+PQ treated children during follow-up  for SP+AS treated children and 10.8 g/dL (9.8\u201311.8) for SP+AS+PQ treated children . While HThe reduction in Hb shortly after SP+AS+PQ treatment was most pronounced in children with the G6PD A- variant  althoughThis study shows that a single dose of primaquine (PQ) is of significant additive value in clearing gametocytes in an area of high malaria endemicity in Tanzania. Only 3.9% of children treated with SP+AS+PQ had gametocytes on day 14 after successful treatment compared to 62.7% for SP+AS treated children. Gametocytes persisting on day 14 after SP+AS+PQ treatment circulated at densities well below the theoretical threshold for mosquito infection.Gametocyte carriage after treatment with SP+AS persisted for more than one month and more than two-thirds of the treated individuals harboured gametocytes on day 14. These data confirm previous findings Focusing on the first two weeks after initiation of treatment, PQ seems to decrease the number of gametocytes rapidly to a level where onward transmission may be arrested completely. Although analyses on gametocyte densities should be considered as post-hoc analyses and we did not directly determine post-treatment malaria transmission, it is clear that the transmission potential is reduced in children treated with SP+AS+PQ. Gametocyte prevalence and density were lower in children treated with SP+AS+PQ and gametocytes persisting on day 14 after treatment circulated at densities below 0.1 gametocyte/\u00b5L. When assuming an average mosquito blood meal size of 2\u20133 \u00b5L P. falciparum infections is to reduce post-treatment infectivity Click here for additional data file.Protocol S1Trial Protocol(0.21 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum glutathione reductase. In 2004, an uncontrolled dose-finding study on the combination MB-CQ was performed in 435 young children with uncomplicated falciparum malaria in Burkina Faso . Three serious adverse events (SAE) occurred of which one was probably attributable to the study medication. In the per protocol safety analysis, there were no dose specific effects. The overall clinical and parasitological failure rates by day 14 were 10% [95% CI ] and 24% [95% CI ], respectively. MB appears to have efficacy against malaria, but the combination of CQ-MB is clearly not effective in the treatment of malaria in Africa.The development of safe, effective and affordable drug combinations against malaria in Africa is a public health priority. Methylene blue (MB) has a similar mode of action as chloroquine (CQ) and has moreover been shown to selectively inhibit the  Plasmodium falciparum to existing safe and affordable drugs such as chloroquine (CQ) and pyrimethamine-sulfadoxine severely threatens the available options for malaria control in sub-Saharan Africa (SSA) [The increasing resistance of ca (SSA) . To maxica (SSA) . AlthougP. falciparum glutathione reductase, has the potential to reverse CQ resistance and it prevents the polymerization of haem into haemozoin similar to 4-amino-quinoline antimalarials [Methylene blue (MB) has already been used some 100 years ago against malaria but it disappeared when CQ and other drugs entered the market . MB, a salarials -7. HowevP. falciparum asexual parasites per \u03bcl blood), haemoglobin \u2265 8 g/dl, absence of severe malaria and other significant disease, and informed written consent. The study was conducted in accordance with the internationally established principles for GCP and controlled by a data safety monitoring board (DSMB). The protocol was approved by the Ethics Committee of the University of Heidelberg and the local Ethics Committee in Burkina Faso.A single centre uncontrolled trial with three dose levels was conducted during the rainy season 2004 at the district hospital of Nouna in north-western Burkina Faso, an area of intense malaria transmission . FebrileChildren were recruited for the three dose levels sequentially. In addition to receiving a total CQ dose of 25 mg/kg , study children received total doses of MB of 36, 54, and 72 mg/kg respectively. At each dose level children were block-randomized by envelope to two or four MB doses per day. MB  was given as a 2.3% solution with fruit flavouring and honey supplement to mask the bitter taste. CQ (tablets or syrup) was taken from the essential drug stock of the hospital. In case of vomiting within 30 minutes after intake, the drugs were re-administered once.The dose escalation process for a dosage regimen went into the next higher dosage level if the safety (i.e. one-sided 95% CI for the incidence of relevant adverse events below 0.1) and the efficacy criterion (i.e. one-sided 95% CI for the incidence of CF below 0.15) were fulfilled at most in one dosage level.Centre de Recherche en Sant\u00e9 de Nouna (CRSN) [Study participants were hospitalized for 72 hours. Treatment failures were managed according to national guidelines. Children were systematically examined on day 0, 1, 2, 3, 4 and 14. Blood samples were processed with standard methods in the laboratory of the a (CRSN) . Methaema (CRSN) .Treatment outcomes were classified according to the WHO guidelines from 2003 as adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) and clinical failure (CF = ETF+LCF) .\u00ae 8.2 was used to analyse the data. Continuous variables in two groups were compared with the nonparametric Wilcoxon-Mann-Whitney (WMW) or Kruskal-Wallis Test (KW), categorical variables with Chi-square-test (Chi). The clinical failure rates were analysed using the two fixed factors (group and level) in a logistic regression model, likelihood ratio tests (LR) were conducted.Assuming a 10% drop-out rate, 72 patients per group and dose level were needed to discover relevant safety and efficacy scenarios with a power of 80% and to avoid a false positive dose effect with probability of 95%. The sample size estimation assumed the independence of the efficacy outcome and the safety outcome. The null hypothesis had to be rejected in each level if the incidence of the relevant study adverse events fell below 10%. For treatment outcomes, the null hypothesis was rejected if the CF rate was below 15%. The safety analysis was based on the children who have received at least one dose of CQ-MB . Efficacy data were assessed in the population of children who received the full course of treatment . All data were double entered and SASChi = 0.0233), age , and in methaemoglobin . Overall 88/409 (21.5%) of FAS children were found to be genotypically G6PD deficient .Overall, 435 children were included into the study, 412 in the FAS analysis and 364 in the PP analysis . During dose level 2, one child progressed to severe malaria within 24 hours of inclusion into the study and one child died of diarrhoea on day 8. During dose level 3, in one G6PD deficient male child the Hb dropped from 8.7 g/dl at inclusion to 4.7 g/dl on day5 but improved afterwards on iron supplementation. Clinically there were no signs of haemolysis and total serum bilirubin was normal on day 3. In seven other children the Hb value dropped by more than 3 g/dl, three of these were found to be G6PD deficient. There were no major differences in the incidence of other adverse events between study groups and dose levels (data not shown).Efficacy outcomes are given in Table The results of this study provide some indirect evidence that MB, a cheap drug which is registered in most countries, could be effective as treatment of uncomplicated malaria in SSA. Compared to the low efficacy demonstrated in a previous study where a low dosage of MB (12 mg per kg over three days) in combination with CQ was used, the CQ-MB combination in this study appeared to be more effective at three to six times higher MB doses (36\u201372 mg per kg over three days) [There were no differences in efficacy between a two times and a four times daily regimen in this dose-finding study; this suggests that a more than twice daily MB regimen would have no benefits. MB has a rather short half-life which has been estimated at 5\u20136 hours . HoweverMethylene blue belongs to a group of drugs considered to potentially cause haemolysis when given to persons with G6PD deficiency . HoweverP. falciparum in vitro, which may further explain the impairment of efficacy observed [Given the potential of CQ resistance reversal, the combination of CQ with MB was studied in trials conducted in Burkina Faso ,6,7. Howobserved .In conclusion, this study has provided indirect evidence for efficacy of MB in the treatment of uncomplicated falciparum malaria in SSA but has clearly shown that the combination CQ-MB is not useful in the treatment of malaria in SSA.P Meissner and G Mandi contributed equally to the study. P Meissner, G Mandi, S Witte, U Mansmann, A Jahn, I Walter-Sack, H Schirmer and O M\u00fcller designed the study. P Meissner, G Mandi, B Coulibaly, J Rengelshausen, W Schiek, G Mikus, J Burhenne and KD Riedel conducted the laboratory and clinical work. S Witte, U Mansmann and T Tapsoba did the statistical analysis. All authors contributed to the writing of the paper. O M\u00fcller was the principal investigator.All authors have no commercial or other association that might pose a conflict of interest. W. Schiek is employed by DSM. The sponsors of the study had no role in study design, data collection and data analysis. The authors had full access to all the data and the corresponding author had final responsibility for the decision to submit for publication."}
+{"text": "Presented here are the results of a comparative trial on the efficacy of three artemisinin-based combinations conducted from May to October 2004, in Pool Province, Republic of Congo.The main outcome was the proportion of cases of true treatment success at day 28. Recrudescences were distinguished from re-infections by PCR analysis. A total of 298 children of 6\u201359 months were randomized to receive either artesunate + SP (AS+SP), artesunate + amodiaquine (AS+AQ) or artemether + lumefantrine (AL), of which 15 (5%) were lost to follow-up.After 28 days, there were 21/85 (25%) recurrent parasitaemias in the AS+SP group, 31/97 (32%) in the AS+AQ group and 13/100 (13%) in the AL group. The 28-day PCR-corrected cure rate was 90.1% [95% CI 80.7\u201395.9] for AS+SP, 98.5% [95% CI 92.0\u2013100] for AS+AQ and 100% [95.8\u2013100] for AL, thereby revealing a weaker response to AS+SP than to AL (p = 0.003) and to AS+AQ (p = 0.06). A potential bias was the fact that children treated with AL were slightly older and in better clinical condition, but logistic regression did not identify these as relevant factors. There was no significant difference between groups in fever and parasite clearance time, improvement of anaemia and gametocyte carriage at day 28. No serious adverse events were reported.Considering the higher efficacy of AL as compared to AS+SP and the relatively high proportion of cases with re-infections in the AS+AQ group, we conclude that AL is clinically more effective than AS+SP and AS+AQ in this area of the Republic of Congo. Implementation of the recently chosen new national first-line AS+AQ should be monitored closely. Plasmodium falciparum is responsible for 98% of malaria cases. The malaria incidence has increased over the past years, as a result of the deterioration of the health infrastructure and population movements during the country's recent episodes of conflict on the one hand and due to rising levels of resistance to common antimalarials on the other hand.In the Republic of Congo (RoC), malaria is the leading cause of morbidity and mortality in children under five years of age and the main reason for hospitalization. The total population of the country, totalling 3.2 million, is at risk of year-round highly endemic malaria. The percentage of medical consultations due to malaria is 30 to 50% ,2. AccorChloroquine (CQ) was still the national recommended first-line and sulfadoxine-pyrimethamine (SP) the second line for uncomplicated falciparum malaria in 2004, at the time of study. Most recent data on drug efficacy have indicated a resistance as high as 90% against CQ and 25% against SP, and this was in a 14-day sentinel survey without PCR to identify new infections . This seThe health authorities in the RoC were, at the time, preparing to change the national antimalarial treatment protocol, but still had to make a choice for the best suitable therapy. In most African countries, the first-line therapy has been revised recently and; at the time of writing, a total of 33 countries have chosen to implement artemisinin combination therapy or ACT, although only 14 have actually commenced implementation .Here, we investigated the efficacy of three ACTs, that were considered potential new first-line therapies for the country, namely artesunate (AS) + SP, AS + AQ and artemether-lumefantrine (AL), in Pool Province in the southeast of RoC.Kindamba is the principal town of Kindamba district in the South of the RoC, located in Pool province Figure . This isKindamba Centre de Sant\u00e9 Int\u00e9gr\u00e9, supported by MSF since 2003. This clinic saw about 500 to 600 cases each month of laboratory-confirmed malaria cases in 2004. The proportionate malaria morbidity among outpatients averaged to about 45% in the under 5's and 30% in older age-groups, while this was over 70% and 40% of in-patients for the two age groups, respectively.The health centre where the study was based is the in vivo drug efficacy studies in areas of high transmission was followed [P. falciparum mono-infection with density between 2,000 and 200,000 parasites per \u03bcl were included in the trial. Written informed consent was asked from the care-taker of the child. Excluded were cases presenting with one or more general danger signs or signs of severe or complicated malaria [The standard WHO protocol for followed . Briefly malaria , a serio\u00ae, La Roche, France; 25 mg S and 1.25 mg P per kg) and 3-day AS , (2) AS + AQ 3-day AQ  and AS (4 mg/kg per day) and (3) AL fixed combination . The dosage of AL (twice daily for three days) depended on body weight and followed the manufacturer instructions. All were treated orally under supervision in the clinic. However, in the AL group, for patients that were unable to stay or to come back to the clinic in the evening, the second daily dose was taken at home and observed by village fieldworkers who visited the patient at the time of treatment. At each dose, AL was given with some fatty food or a glass of milk. When a child was assigned to AL but weighed below 10 kg, the next possible treatment allocation was given and AL was given to the following study patient.Children were enrolled in the study and randomly assigned to one of three therapies: (1) AS + SP: single dose SP on the first day  before and after treatment. The rescue treatment in case of failure was quinine for 7 days.Patients were asked to come back for treatment the following 2 days and for follow-up visits on day 3, 7, 14, 21 and 28. They were clinically examined at all visits, both to check for signs of illness as for potential side-effects of treatment, as passively reported to the medical responsible, and had their blood-film taken on all these occasions except day 1. Blood films (thin and thick smears) were stained by Giemsa and examined by experienced microscopists. A quantitative parasite count of asexual forms was performed for positive cases (using the parasite/White Blood Cell Count) and the presence of gametocytes was noted. Based on clinical and laboratory findings, cases were classified as early treatment failures (ETF), late clinical failures (LCF), and late parasitological failures (LTF) or adequate clinical and parasitological responders . The final outcome parameter of the efficacy trial was the cure rate at the end of the follow-up period of 28 days in each of the three treatment arms. A true failure was defined as a recrudescence defined by PCR analysis eliminating re-infections. Haematological recovery was assessed by haemoglobin measurements .2 tests to compare categorical data . Continuous data were tested for normality . Normally distributed data were analysed with T-tests and ANOVA, and not-normally distributed data with Ranksum tests. Multivariate regression analysis was performed to determine whether baseline characteristics were associated with the outcome of treatment. Age, weight, temperature and parasite density were entered into the model as linear variable or classified in 3 to 5 categories and haemoglobin level in 3 classes: moderate (5\u20137.9 g/dl), mild (8\u201310.9 g/dl) or no anaemia.Laboratory quality control was carried out on 10% of the slides by qualified laboratory experts working in laboratories within the other MSF projects in the country . Slides were read blindly, and results were compared afterwards with those from the study laboratory. Data analysis was performed using Excel database and SPSS software. We used \u03c7The protocol received ethical clearance from the RoC Ministry of Health of the Republic of Congo and the external Ethical Review Board used by MSF. The necessary permission from the local authorities  in Kindamba, Pool District of study was obtained.From 27 May to 6 October 2004, a total of 298 children were enrolled in the study and 101 children received AS+AQ, 91 received AS+SP and 106 received AL.Due to the different weight-entry criteria for AL \u226510 kg) as compared to the other two ACT groups (\u22655 kg), the children in the three treatment groups were not equal for all basic demographic characteristics on admission. There was a significant difference in weight, age, MUAC and blood haemoglobin value between the AL group and the other two groups. 36.5% of children were below 2 years; the average age was 27 months , 11 were lost to follow-up (due to migration out of the area) and 5 were secondary exclusions (1 was retreated while not (yet) a failure, 4 were given drugs at home with antimalarial activity during follow-up; see Figure The proportion of recurrent parasitaemia's during the 4-week follow-up was higher in the AS+AQ group , and AS+SP group  than in the AL group . No ETF's were observed in any of the three treatment groups.A total of 57 patients were excluded after PCR analysis. From the 65 patients eligible for PCR analysis, only 50 PCR were performed (15 samples not taken during a security evacuation of part of the study team). Among these, only 38 had a PCR result available (12 indeterminate results), of which 30 were re-infections which were excluded from the final analysis , and AS+AQ (p = 0.06) whereas differences in efficacy between AL and AS+AQ were not significant (p = 0.4).Cases of recrudescence as well as cases of re-infection occurred at a relatively long time interval after treatment in all three groups. The median time to recurrent parasitaemia was 21 days for the AS+AQ group, 21 for AS+SP and 28 for AL.Multivariate regression analysis showed that for 'recurrent parasitaemia', none of the baseline characteristics age, sex, weight initial parasite count, temperature or haemoglobin level influenced the outcome significantly; independent of these, the treatment group was the primary determinant of recurrent parasitaemia at day 28. For the recrudescent cases (PCR corrected true failure rates), multiple factorial analysis was not possible, due to the low numbers of failures.On inclusion, 178/298 (60%) of patients had measured fever. The axillary temperature decreased rapidly after all three therapy regimens. At day 3, the number of febrile patients was similarly low in each arm . All children were parasite free on day 3.The proportion of cases with gametocytes increased during the first two days of treatment, but overall reduced during the four weeks of follow-up, from 23% to 3% in the AS+AQ group, from 26% to 5% in the AS+SP group and from 8% to 1% in the AL group  and did not differ among the three treatment groups. There were two cases of urticaria, one after AS+SP and one after AS+AQ treatment, but these developed after completion of the treatment, so it was not necessary to stop or interrupt the treatment.P. falciparum. This is one of the first studies on ACTs in this country. AL is the most efficacious one of the three (100% with 95% CI 96\u2013100); none of the children receiving this therapy had a recrudescent infection in the 4 weeks of follow-up. AS+SP showed a significantly lower efficacy than AL (90% with 95% CI 87\u201396), and AS+AQ (98.5 with 95% CI 92\u2013100). The secondary outcomes of rapid fever-, parasite- and gametocyte clearance and the beneficial effect on blood haemoglobin levels were similar for the three therapies.The study presented here shows that in the Pool region, Republic of Congo, AS+SP, AS+AQ and AL, are effective to treat uncomplicated However, a substantially large proportion of children showed a recurrent parasitaemia in 4 weeks after treatment: 32% in the AS+AQ group, 25% in the AS+SP group and 13% in the AL group. AL appeared to prevent re-infection better than AS+AQ. Infection rates can be assumed equal among the treatment groups. AS is cleared from the blood within hours, so any protective effect of combination therapy comes from the remaining drug levels of AQ, SP and lumefantrine. Higher re-infection rates may be related to reduced AQ susceptibility/developing resistance of the parasites.The interpretation of PCR analysis should be taken cautiously. Re-infections were confirmed in 46% of the recurrent infections, but there was a substantial part of PCR results unavailable (42% or 27/65), due to missing samples and indeterminate results. The available PCR results indicate that about 20% of recurrent infections were recrudescences; applying this 'rule' to the missing values, we calculated failure rates of 6% for AS+AQ (3 extra recrudescent cases assumed from 13 missing cases), 11% for AS+SP (1 added of 5 missing cases) and 2% for AL (2 added of 9 cases).The treatment allocation was randomized, however, the specific entry criteria of weight for the AL group (above 10 kg) may have influenced the outcome of the study. The children selected for treatment with AL were on average older, and healthier. This group therefore may have had a better general condition and probably acquired some immunity against malaria, preventing rapid recrudescence, leading to a higher measured efficacy of AL. We had to follow the AL prescription criteria for ethical reasons, but maintained the usual weight limit for the other two treatment arms, to keep this study comparable to others in the region. The limitation of weight for AL has now been lowered from 10 kg to 5 kg in December 2004 by WHO ,13.The true failure rates may have been higher. Treatment failures can occur more than 28 days after treatment. It has been estimated  that 28-The levels of efficacy compare to findings from other recent studies on ACTs in surrounding in southwest Africa. In Angola, AL, AS+AQ and AS+SP were found to be highly effective . In DRC,falciparum malaria than the other two ACTs in the study area in Congo Republic, and this is reflected by studies in the neighbouring countries as well. A careful implementation of AL is, therefore, recommended as the new protocol.Artemether-lumefantrine has shown more consistent efficacy to treat uncomplicated However, recent decisions at the National Malaria Programme in Brazzaville point towards AS + AQ as the new first-line antimalarial treatment, described for use at household as well as health centre level with basic symptomatic diagnosis (draft document PNLP). The second-line treatment defined is AL (or quinine or artemether) based on microscopy diagnosis.Malaria is a high burden on health expenditures in Congo, a country with an infection prevalence estimated at 50% or more , leadingImplementation of the new RoC anti-malarial policy should be accompanied by adequate population sensitisation and facilitation of clear prescriptions using available blister-packaging or co-formulation, to prevent misuse of valuable medication risking further development of AQ resistance. The efficacy of the artemisinin combination therapies should be monitored at 2-year intervals after implementation.I Van den Broek was overall study supervisor and co-author for the final report. C Kitz was the site investigator in the field and co-author. S Attas contributed to the study implementation and was also overall medical coordinator for the MSF field programme. F Libama reviewed the study protocol and provided technical laboratory assistance and support. M Balasegaram and JP Guthmann were both involved in the study design, providing technical assistance and contributed to the writing of the final report."}
+{"text": "To compare the efficacy and safety of artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) for treating uncomplicated falciparum malaria in Uganda.Randomized single-blinded clinical trial.Apac, Uganda, an area of very high malaria transmission intensity.Children aged 6 mo to 10 y with uncomplicated falciparum malaria.Treatment of malaria with AL or DP, each following standard 3-d dosing regimens.Risks of recurrent parasitemia at 28 and 42 d, unadjusted and adjusted by genotyping to distinguish recrudescences and new infections.Of 421 enrolled participants, 417 (99%) completed follow-up. The unadjusted risk of recurrent falciparum parasitemia was significantly lower for participants treated with DP than for those treated with AL after 28 d  and 42 d  of follow-up. Similarly, the risk of recurrent parasitemia due to possible recrudescence (adjusted by genotyping) was significantly lower for participants treated with DP than for those treated with AL after 28 d  and 42 d . Patients treated with DP had a lower risk of recurrent parasitemia due to non-falciparum species, development of gametocytemia, and higher mean increase in hemoglobin compared to patients treated with AL. Both drugs were well tolerated; serious adverse events were uncommon and unrelated to study drugs.DP was superior to AL for reducing the risk of recurrent parasitemia and gametocytemia, and provided improved hemoglobin recovery. DP thus appears to be a good alternative to AL as first-line treatment of uncomplicated malaria in Uganda. To maximize the benefit of artemisinin-based combination therapy in Africa, treatment should be integrated with aggressive strategies to reduce malaria transmission intensity. Background: The burden of death and disease caused by malaria is very high, particularly amongst young children in sub-Saharan Africa. Many African countries have adopted combinations of drugs for first-line malaria treatment that include a compound based on the plant-derived molecule artemisinin. In Uganda, the artemisinin-based compound therapy (ACT) adopted as first-line therapy is artemether-lumefantrine (AL). However, there are limitations to the use of this drug; dosing is not convenient and it needs to be given with fatty food. There are also concerns that even though AL and other ACTs successfully treat the initial infection, there is a risk of malaria recurring soon after therapy, particularly in areas where mosquito attack rates are high. Therefore, the researchers here wanted to compare AL treatment with another ACT, dihydroartemisinin-piperaquine (DP), for treatment of uncomplicated malaria in children who live in an area of Uganda where malaria is transmitted at a very high rate.What the trial shows: 421 children aged between six months and ten years arriving at a local health centre with uncomplicated malaria were recruited into the study. The children were randomly assigned to receive either AL tablets, given twice daily for 3 days, or DP tablets, given once daily for 3 days. The primary outcomes in the trial were the risks of clinical malaria recurring, and the risk of parasites reappearing in the blood but without any sign of clinical malaria, over 42 days of follow-up. The researchers also carried out tests to distinguish new malaria infections from a reappearance of the old infection.Overall, children who received DP treatment were less likely to have malaria parasites reappear in their blood than children who received AL treatment. Similarly, the risk of a recurrence of clinical malaria was lower in the DP arm than in the AL arm; 39% of patients treated with AL went on to develop malaria symptoms by 42 days whilst only 25% of patients treated with DP did. These findings were similar if the researchers only looked at children in whom the same infection reappeared, as opposed to those who experienced a completely new infection. There did not seem to be any differences between the study groups in overall rates of adverse events, most of which were mild or moderate in nature. The serious adverse events that occurred in this trial were not thought to be related to the drugs being studied.Strengths and limitations: This trial recruited enough participants to test whether there were any differences in efficacy between the two treatments studied, and randomization procedures and blinding of study physicians were appropriate. A further strength is the follow-up period of 42 days, in line with recent World Health Organization guidelines for such trials. This trial included children aged under 10 years, whereas children aged under 5 years are generally considered most at risk from malaria. However, the majority of children in this trial were in the under-5 age group, and the overall results were similar when only considering children in this age range. Finally, one possible limitation is that blinding of participants may not have been complete in this trial: children and parents were not informed of their treatment assignment, but the study drugs were different in appearance.Contribution to the evidence: Studies from South-East Asia have indicated that DP has efficacy in treating malaria resistant to other drugs. However, little data exists regarding the efficacy and safety of DP in Africa. This trial adds important safety data for DP in African children, and also provides evidence that DP could be considered as an alternative treatment to AL, the current first-line therapy in many African countries. In Africa, treatment of uncomplicated malaria is undergoing dramatic changes. In response to widespread resistance of the parasite to commonly used monotherapies, particularly chloroquine (CQ) and sulfadoxine-pyrimethamine (SP), many countries have recently adopted artemisinin-based combination therapy (ACT) as a first-line regimen for the treatment of uncomplicated malaria . In UganA concern with all antimalarial regimens, particularly in areas of high transmission, is frequent recurrence of malaria after therapy. This concern was highlighted in a recent study comparing efficacies of the ACTs AL and AQ + AS in Tororo, Uganda, an area with very high malaria transmission . In thatDihydroartemisinin-piperaquine (DP) is a fixed-dose ACT that has recently become available in Africa. In studies from Southeast Asia, DP appeared to be well tolerated and highly efficacious against multidrug-resistant falciparum malaria \u201311. HoweThe study was conducted at Aduku Health Centre, Apac District, Uganda. The district experiences perennial holoendemic malaria. The entomological inoculation rate in Apac, a measure of transmission intensity, was measured at 1,564 infectious bites per person per year . The stuPlasmodium falciparum monoinfection with parasite density 2,000\u2013200,000/\u03bcl of blood. Because laboratory results were generally not available until the following day, a patient could be excluded after randomization.Consecutive patients presenting to the health center with symptoms suggestive of malaria and a positive screening thick blood smear were referred to study physicians for further assessment. Patients were enrolled if they fulfilled the following selection criteria: (1) age 6 mo to 10 y; (2) weight \u2265 5 kg; (3) history of fever in the last 24 h or axillary temperature \u2265 37.5 \u00b0C; (4) no history of serious side effects to study medications; (5) no evidence of a concomitant febrile illness; (6) provision of informed consent by a parent or guardian; (7) no danger signs or evidence of severe malaria; and (8) At enrollment, we asked children's parents or guardians about prior antimalarial therapy, use of other medications, and presence of common symptoms. Axillary temperature and weight were measured, and a physical examination was performed. A brief neurological assessment, consisting of simple clinical tests for fine finger dexterity , was undertaken. We also obtained blood by fingerprick for thick and thin blood smears, for hemoglobin assessment, and to store on filter paper for molecular analysis.Patients were asked to return for follow-up on days 1, 2, 3, 7, 14, 21, 28, 35, and 42, and any other day that they felt ill. Follow-up evaluation consisted of a standardized history and physical examination, including neurological assessment on all days of follow-up. We obtained blood by fingerprick for thick blood smears and storage on filter paper on all follow-up days except day 1. Hemoglobin measurement was repeated on day 42 or the day of recurrent symptomatic malaria. If patients did not return for follow-up, they were visited at home.http://www.hemocue.com).Blood smears were stained with 2% Giemsa for 30 min. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 200 white blood cells (WBCs), or per 500 if the count was less than 10 parasites/200 WBCs, assuming a WBC count of 8,000/\u03bcl. A smear was considered negative if no parasites were seen after review of 100 high-power fields. We also assessed gametocytemia from thick blood smears. Thin blood smears were reviewed for non-falciparum infections. A second microscopist, who was unaware of the results of the first reading, re-read all slides. A third microscopist unaware of the first two readings resolved discrepant slides. Hemoglobin measurements were made using a portable spectrophotometer , administered according to weight as one (5\u201314 kg), two (15\u201324 kg), three (25\u201334 kg), or four (\u2265 35 kg) tablets given twice daily for 3 d; DP , targeting a total dose of 6.4 and 51.2 mg/kg of dihydroartemisinin and piperaquine, respectively, given in three equally divided daily doses to the nearest quarter tablet. We used a pill cutter to ensure that the tablet fractions were as close to the nearest quarter tablet as possible. Participants in the DP group also received placebo tablets administered in the evening over 3 d to simulate the AL dosing schedule. Study medications were administered with water, and patients were given a glass of milk after each dose of study medication.http://www.vestergaard-frandsen.com) on the day of enrollment, with instructions for one net to be used by the study patient.All treatment was directly observed. Participants were given the option either to wait at the clinic for the evening dose (lunch was provided) or to leave the clinic and return in the evening (transport was provided). After each dose, children were observed for 30 min, and the dose was readministered if vomiting occurred. All patients were provided with a 3 d supply of acetaminophen for treatment of febrile symptoms. Children with hemoglobin of less than 10 g/dl were treated according to Integrated Management of Childhood Illness guidelines with ferrous sulfate for 14 d and antihelminthic treatment if appropriate. Households of all patients were given two long-lasting insecticide-treated bed nets (ITNs)  guidelines as early treatment failure ; late clinical failure ; late parasitological failure ; or adequate clinical and parasitological response  . PatientThe primary efficacy outcomes were the 28- and 42-d risks of early treatment failure or recurrent parasitemia (LCF or LPF), unadjusted and adjusted by genotyping. Secondary efficacy outcomes included prevalence of fever and parasitemia during the first 3 d of follow-up, change in mean hemoglobin from day 0 to day 42 or day of repeat therapy, and prevalence of gametocytemia (presence of gametocytes on thick smears) during follow-up in participants lacking gametocytes at enrollment.merozoite surface protein (msp)-2, msp-1, and four microsatellites [Molecular genotyping techniques were used to distinguish recrudescent from new infections for all patients with LCF or LPF response. Briefly, parasite DNA was isolated from filter paper blood samples collected at enrollment and on the day of recurrent parasitemia using chelex extraction. Paired samples were genotyped in a stepwise fashion using tellites . If, forSecondary safety outcomes included risks of serious adverse events and common adverse events of any severity. An adverse event was defined as any untoward medical occurrence, irrespective of its suspected relationship to the study medications . At eachWe calculated sample size to test the hypothesis that the risk of recurrent parasitemia after 42 d would differ between the two treatment groups. Based on previous data, the risk of recurrent parasitemia (unadjusted by genotyping) after 42 d was estimated to be 50% after treatment with AL . Using tA randomization list was computer generated by an off-site investigator without the use of blocking or stratification. Sequentially numbered, sealed envelopes containing the treatment group assignments were prepared from the randomization list. The study doctors assigned treatment numbers sequentially and the study nurse allocated treatment by opening the envelope corresponding to the treatment number. The randomization codes were secured in a locked cabinet accessible only by the study nurse. Participants were enrolled by the study physicians, and treatments were assigned and administered by the study nurse.Only the study nurse was aware of treatment assignments. All other study personnel, including the study physicians and laboratory personnel involved in assessing outcomes, were blinded to the treatment assignments. Patients were not informed of their treatment regimen, but the color of the two study medications was not the same .http://www.stata.com). Efficacy and safety data were evaluated using a modified intention-to-treat analysis which included all patients who fulfilled enrollment criteria. Patients who were randomized to therapy but not enrolled in the study due to laboratory results available on day 1 were not included in the analysis. Risks of recurrent parasitemia at 28 and 42 d of follow-up (adjusted and unadjusted by genotyping) were estimated using the Kaplan-Meier product limit formula. Data were censored for patients who did not complete follow-up and for new infections when estimating outcomes adjusted by genotyping. Patients with LCF or LPF due to non-falciparum species were censored as non-failures at the time they were classified as LCF or LPF. The Z-test was used to compare the Kaplan-Meier estimates of treatment efficacy at fixed points in time between the treatment groups. Confidence intervals around the difference between Kaplan-Meier estimates were calculated using normal approximation and Greenwood's estimates of standard errors. Categorical variables were compared using Chi-squared or Fisher exact test and continuous variables were compared using the independent samples t-test. All reported p-values are two sided without adjustment for multiple testing and were considered statistically significant if below 0.05.Data were entered and verified using Epi Info version 6.04 and analyzed using STATA version 8.0 . Additionally, patients treated with AL had a higher risk of recurrent parasitemia due to non-falciparum species  compared to patients treated with DP . Results were similar when restricting the analyses to children under the age of 5 y, as 94% of patients enrolled were in this age range.There were no early treatment failures in the first 3 d following initiation of therapy. Episodes of recurrent parasitemia were first detected 14 d following therapy in the AL arm and 21 d following therapy in the DP arm ; Table 2p = 0.002).Patients with asymptomatic recurrent parasitemia (LPF) were not treated unless they developed symptomatic malaria or reached the end of the 42-d follow-up period. Among 121 patients with LPF occurring before day 42, only six (5%) spontaneously cleared their parasites without treatment, 86  went on to develop symptomatic malaria by day 42, 28 (23%) had persistent asymptomatic parasitemia at day 42, and one (1%) took other antimalarials prior to day 42. Overall, 81 (39%) of 210 patients treated with AL went on to develop recurrent symptomatic malaria, compared to 52 (25%) of 211 patients treated with DP (p = 0.05). However, among patients with recurrent parasitemia there was no difference in the prevalence of anemia (Hb <10 g/dl) on the day of failure in the AL group  compared to the DP group  (p = 0.87).The prevalence of fever (either subjective or documented) was similar over the first 3 d of follow-up in the two treatment groups. Both treatments produced rapid clearance of parasitemia with no parasites detected by day 3 . The appp = 0.47). There was also no difference in the proportion of patients who experienced common adverse events of any severity  and AL (42%) treatment groups Click here for additional data file.Trial Protocol(1.1 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria with sulfadoxine-pyrimethamine (SP) is followed by a sharp rise in the prevalence and density of gametocytes. We did a randomized trial to determine the effect of treatment of asymptomatic infections with SP or SP plus one dose of artesunate (SP+AS) on gametocyte carriage.Treatment of P. falciparum to receive antimalarial treatment or placebo, and recorded the prevalence and density of gametocytes over the next 2 mo.The study was a three-arm open-label randomized trial. We randomized asymptomatic carriers of The trial was conducted during the dry  season in four rural villages in Gambia.P. falciparum infection and confirmed free of clinical symptoms of malaria over a 2-d screening period.Participants were adults and children aged over 6 mo with asexual Participants were randomized to receive a single dose of SP or SP+AS or placebo.The outcome measures were the presence of gametocytes 7 and 56 d after treatment, and the duration and density of gametocytaemia over 2 mo.p = 0.87), and 7.1% in the SP+AS group . By day 56, gametocyte prevalence was 13% in the placebo group and 2% in both drug-treated groups. Gametocyte carriage (the area under the curve of gametocyte density versus time), was reduced by 71% in the SP group, and by 74% in the SP+AS group, compared to placebo. Gametocyte carriage varied with age and was greater among children under 15 than among adults.In total, 372 asymptomatic carriers were randomized. Gametocyte prevalence on day 7 was 10.5% in the placebo group, 11.2% in the SP group  in the blood of infected people. Gametocytes are the transmissible stage of the parasite that can infect the mosquito when it takes a blood meal from an infected person. In people with clinical signs of malaria, treatment with commonly used antimalarial drugs such as chloroquine or sulfadoxine-pyrimethamine is often followed by a rise in the numbers of gametocytes in the blood, but it is not known to what extent this is caused by the drugs themselves. The effects of different antimalarial treatments on gametocytes have generally not been well studied in trials, and particularly not in individuals who are infected with malaria but don't show clinical signs . However, it is important to know the effects of different malaria drugs on gametocytes, because this will help predict how parasites are likely to evolve resistance to drugs. In particular, the effects on asymptomatic carriers are interesting because of the potential benefit of giving preventive treatments to all children or adults (whether or not they have symptoms) for malaria control. The trial from Gambia reported here was conducted in adults and children over 6 mo of age who were infected with P. falciparum malaria but free from clinical symptoms. Participants were randomized to receive either sulfadoxine-pyrimethamine, sulfadoxine-pyrimethamine and artesunate, or placebo. The researchers wanted to look at the effect of these treatments on numbers of gametocytes in the participants' blood samples seven days after treatment (the primary outcome) and then up to 56 days after treatment.What this trial shows: Seven days after treatment, the researchers found that the proportion of participants carrying malaria gametocytes in the groups treated with sulfadoxine-pyrimethamine or sulfadoxine-pyrimethamine and artesunate was similar to the proportion in the placebo group. A secondary objective of the study was to look at overall gametocyte load over 56 days following treatment, by measuring the area under the curve plotting gametocyte density against time. For this secondary objective, the researchers found that gametocyte load was highest in the placebo group, and substantially lower in the groups treated with sulfadoxine-pyrimethamine or sulfadoxine-pyrimethamine and artesunate. Therefore, there was no evidence that treatment with sulfadoxine-pyrimethamine increased gametocytes in the blood of children infected with malaria but without symptoms.Strengths and limitations: In this trial, screening tests to detect malaria infection and symptoms were done twice prior to enrolling participants into the trial, with a two-day window between tests. This procedure increased the chance of detecting individuals with clinical malaria so they could be excluded from the trial and treated for malaria. The use of a placebo arm in the trial, which adds to the strength of the study, could therefore be defended ethically. The conduct of this trial in a region with seasonal transmission of malaria limits the extent to which the findings can be generalized to other regions.Contribution to the evidence: This trial provides data on the extent to which treatment with sulfadoxine-pyrimethamine or sulfadoxine-pyrimethamine and artesunate affects gametocyte levels in people infected with malaria, but without symptoms. It had previously been thought that sulfadoxine-pyrimethamine increased gametocyte carriage, but this had not been formally tested in a placebo-controlled study. The results show that both treatments reduce overall gametocyte load to very low levels over a period of 56 days after treatment, as compared to placebo. Plasmodium falciparum is responsible for 300 million clinical cases and up to 1 million deaths annually, mainly among children under 5 y of age. Treatment of P. falciparum malaria with commonly used antimalarial drugs such as chloroquine or sulfadoxine-pyrimethamine (SP) is followed by a marked increase in the density of gametocytes, the parasite stage that infects mosquitoes [P. falciparum infection without clinical symptoms of malaria to treatment with SP, a combination of SP plus one dose of artesunate (SP+AS), or placebo, to determine the effects of treatment on gametocyte prevalence, duration of gametocyte carriage, and gametocyte density.In sub-Saharan Africa, malaria caused by squitoes ; this issquitoes ,8. The esquitoes \u201314, and squitoes ,11,14,15squitoes ), but fesquitoes . To inveP. falciparum, and occasionally by P. malariae and P. ovale, and transmission is highly seasonal, with most morbidity due to malaria occurring from September to November [The climate in Gambia is characterised by a long dry season from mid-October to mid-June followed by a short rainy season. Malaria is predominantly caused by November . The stuNovember . ExclusiParticipants enrolled were randomized to receive SP alone, SP+AS, or placebo. Participants who had symptomatic malaria at screening were not randomized but were treated with SP+AS and followed up similarly, in order to check that the normal pattern of gametocyte carriage after treatment in clinical cases was observed. Open-label SP , artesunate , and calcium  were used.P. falciparum infection during the low-transmission season.We wanted to determine the effect of treatment with SP alone and with a single dose of artesunate on gametocyte carriage, in people with The primary outcomes were gametocyte prevalence and density on D7, and secondary outcomes were gametocyte prevalence on D56 after treatment, and gametocyte carriage, defined as the area under the curve (AUC) of gametocyte density against time. Participants were visited at home by field workers on D3, D7, D14, D28, and D56 post-treatment to check that the person was well, to record self-reported adverse events, and to collect finger prick blood for microscopy. The study was not blinded, but field workers and laboratory assistants responsible for microscopy did not have access to treatment allocation data and therefore were likely to be unaware of the treatment received by participants. Two thick blood films were prepared from each individual, dried overnight, and stained with Giemsa, and 200 HPFs were examined independently by two microscopists for asexual and sexual stage parasites. Where there was a discrepancy the slide was read a third time, by another microscopist. During screening, one blood film was stained with Field's stain, and 50 HPFs were examined to identify infected individuals for enrolment. PCV was determined by collecting finger prick blood samples into heparinised capillary tubes and spinning them using a micro-haematocrit centrifuge .The primary endpoint was gametocyte prevalence on D7. Assuming a gametocyte rate in the placebo group of 20%, a sample size of 120 individuals per group would allow over 80% power to detect an increase to 40% in the SP-alone group and a decrease to 5% or less in the SP+AS group, allowing for 10% drop-out and using a significance level of 0.025 .The randomization sequence was generated in Stata  using restricted randomization with a block size of 12. Treatment allocation was determined by opening pre-prepared numbered opaque randomization envelopes in sequence by a study nurse who took no part in participant selection or evaluation, but who acted as the drug dispenser throughout the trial. SP was administered as 25 mg/kg sulfadoxine/1.25 mg/kg pyrimethamine for children under 50 kg, while adults and children weighing 50 kg or more were given three tablets (each tablet contained 500 mg of sulfadoxine and 25 mg of pyrimethamine) . In the SP+AS group, artesunate was given at the same time as SP at a dose of 200 mg (four tablets at 50 mg) for adults and for children weighing 50 kg or more, while children under 50 kg received 4 mg/kg body weight. Calcium tablets were given as placebo. Children were observed for 1 h after treatment to ensure that the drug was not vomited, and treatment was repeated in those who vomited the first dose. Treatment failures in the SP and SP+AS groups were to receive rescue medication with quinine while those in the placebo group were to be rescued with chloroquine and SP (which is highly effective in Gambia ).p-Values are two-sided. We summarised the information on gametocyte carriage by calculating the AUC of the gametocyte density against time, a measure of transmission potential. This is a weighted sum of the gametocyte densities, with weights proportional to the difference in time between adjacent sampling points. Individuals with missing data were included in the analysis, with the AUC weighted according to the number of non-missing sampling points. Similarly, an AUC was calculated for the binary variable  against time. The mean of this AUC is the area under the curve of the proportion positive plotted against time, and is a measure of the average duration of gametocyte carriage. We compared the AUC between treatment groups using the Kruskal-Wallis test and between age groups using a non-parametric test for trend [Data were double entered and validated. Analysis was done using Stata. Fractional polynomial logistic regression  was usedor trend ; both teP. falciparum malaria). Of the remaining 1,048 participants, representing 46%  of the population of the study villages, 99 (9.4%) had gametocytaemia with a geometric mean density of 6.1  gametocytes per microlitre. A total of 616/1,048 (59%) did not have detectable asexual parasitaemia, and most of these (97.4%) were gametocyte negative. Gametocyte prevalence was high among children under 15 y and then decreased with increasing age, and gametocyte prevalence was similar in males and females and in each of the four villages , and 7.1% in the SP+AS group . Gametocyte prevalence showed a steady decline in the active drug treatment groups, and by D28, gametocyte prevalence was less than 3% in the SP and SP+AS groups , and addition of AS reduced this index to 16 . In the clinical cases, the AUC gametocyte index was 147, significantly greater than the asymptomatic group treated with the same drug regimen (p = 0.02) but not significantly different from the placebo group (p = 0.32).Asexual parasite clearance was similar in the SP and SP+AS groups A. GametoS groups B. In conS groups  was 62 iAmong the 290 randomized participants who were gametocyte negative at enrolment, gametocyte prevalences on D3, D7, and D14 were similar in the SP and placebo groups C, suggesMean AUC gametocyte index varied with age and was greatest among children under 15 y; this pattern was associated with greater asexual parasite densities in this age group at enrolment . The aveP. falciparum infection, and we found no evidence that SP treatment enhances emergence of gametocytes into the peripheral blood, but rather that, during the dry season, effective treatment with SP or SP+AS virtually eliminates gametocyte carriage over time. Gametocyte carriage was not significantly reduced further by the addition of a single dose of artesunate; three doses may be more effective [In areas of seasonal malaria transmission, intermittent treatment of children with antimalarial drugs or some form of chemoprophylaxis may have a role in malaria prevention ,16,18,23ffective  but coulffective . ChildreOur study, conducted in the dry season in asymptomatic carriers, does not rule out the possibility that in acute clinical cases, with much greater parasite densities, treatment with SP causes some release of sequestered gametocytes, but our data are compatible with the explanation suggested by Butcher , that thTreatment reduces gametocyte carriage to very low levels over a period of 1 mo; mass drug administration of SP+AS in the study area in June 1999 did not reduce overall malaria incidence despite achieving relatively high coverage , and whiCONSORT ChecklistClick here for additional data file.(48 KB DOC)Trial ProtocolClick here for additional data file.(229 KB DOC)"}
+{"text": "Plasmodium falciparum malaria. Although the safety and efficacy of AL have been extensively documented, no clinical trials had been carried out in Zambia.In Zambia, unacceptably high resistance to commonly used antimalarial drugs prompted the choice of artemether-lumefantrine (AL) as first line treatment for uncomplicated Nine hundred seventy one adult patients with uncomplicated malaria were randomized to either sulfadoxine-pyrimethamine (SP)(486) or AL (485) and followed up for 45 days. Outcome of treatment was defined according to the standard WHO classification. Recurrent parasitaemia were genotyped to distinguish between recrudescence and new infection.Fever at day 3 was significantly lower  and the mean haemoglobin at day 45 significantly higher  in the AL group. Almost all clinical symptoms cleared faster with AL. Early treatment failure was significantly higher in the SP (25/464) than in the AL (2/463) . The rate of new infections was similar in both groups (18 with SP and 19 with AL). Late clinical failure  and late parasitological failure  were significantly higher in the SP group. Total treatment failure was significantly higher in the SP group  as compared to the AL  group .P. falciparum malaria in adults. Data on safety and efficacy of AL in pregnant women are urgently needed.In Zambia, the new first line regimen AL is far more efficacious than SP in treating uncomplicated  Plasmodium falciparum malaria. Zambia was the first African country to adopt an artemisinin-based combination treatment as its national policy. The safety of AL has been extensively reviewed [\u00ae or Riamet\u00ae. The fixed tablet formulation helps to overcome problems of compliance associated with non-coformulated combinations.In Zambia, malaria treatment and control have been undermined by the emergence of resistance to commonly-used antimalarial drugs such as chloroquine (CQ) and sulfadoxine-pyrimethamine (SP). Unaccepreviewed and severreviewed -10. AL eThe results of a randomized clinical trial on the efficacy of SP compared to AL in adult patients with uncomplicated malaria in Ndola, Zambia, are reported below. Patients were followed up for 45 days.P. falciparum density of 25/200 WBC  or more were included. Exclusion criteria were the following: pregnancy, severe falciparum malaria [The study started in March 2003, during the antimalarial drug policy transition from SP to AL, and was completed in June 2005. Patients were recruited at 4 peri-urban health centers in Ndola, Zambia, an area of mesoendemic malaria where transmission is perennial with a seasonal peak from November to April. A reliable ambulance service is available and patients requiring hospitalization can be referred to Ndola Central Hospital, about 5 km away. All individuals aged 15\u201350 years attending any of the 4 peri-urban clinics and presenting with fever (body temperature \u2265 37.5\u00b0C), and/or history of fever in the previous 48 hours, and without any other obvious disease were screened for malaria infection (thick and thin blood film in duplicate for parasite density and species identification) and pregnancy (if applicable). Patients with a  malaria , documen\u00ae, Roche: 500 mg sulfadoxine/25 mg pyrimethamine tablets), single dose of 3 tablets (2.5 tablets if < 50 kg), or AL , 4 tablets immediately followed by 4 tablets at 8, 24, 36, 48 and 60 hours, taken with a creamy snack. Treatment allocation was concealed until final recruitment. SP treatment was directly observed and patients were monitored at the health centre for at least 30 minutes after treatment. For AL, the morning doses were directly observed over the 3 days of treatment, while the evening doses were given to the patients to be taken at home, and empty sachets returned as evidence of taking the drug. Paracetamol tablets (3 doses/day for 2 days) were provided, to be taken when needed.Patients were randomly allocated by blocks of 10 (according to a pre-established list) to receive either SP (Fansidar\u00ae). Hb was measured again at day 14 and 45. Patients were encouraged to attend the health facility outside scheduled visits if they felt ill. Patients treated with SP and classified as failures  were treated with AL, while those on AL received quinine. Adverse events were documented and treated accordingly.Clinical history, signs and symptoms, body temperature were recorded and a blood sample for parasitaemia (blood slide) and for molecular analysis (on Schleicher & Schuell filter paper) were collected at days 0, 3, 7, 14, 21, 28 and 45 or at any unscheduled visit. The blood sample on filter paper was dried at room temperature and stored at -20\u00b0C with silica gel. At day 0 (before treatment), a drop of venous blood was collected for Hb measurement  was determined. Parasite density per \u03bcl was computed taking into account the actual WBC counts. Internal quality control was organized as recommended by the WHO [\u00ae and noted with a precision of 0.1 g/dl. Blood samples collected on filter paper at enrolment and during follow up were used to genotype parasite strains; only those microscopically positive after day 14 were analysed. DNA was purified as described previously [P. falciparum genetic markers MSP1 and MSP2 repeat region was carried out. A recrudescent infection was defined as one that showed match in size of at least one allele for both the MSP1 and MSP2 genes between the first and second sample.All lab technicians were blinded to the patient's identity and all patient-related parameters. Thin blood films were fixed with methanol and thin and thick blood films were stained with 10% Giemsa. At the clinic, the number of asexual  the WHO . Hb was eviously  and a neOutcome of treatment was defined according to the standard WHO classification (WHO 2003): Early Treatment Failure (ETF) was defined as: i) danger signs of/or severe malaria on days 1, 2 or 3 with parasitaemia; ii) parasite density at day 2 greater than at day 0; iii) parasitaemia on day 3 with axillary temperature \u2265 37.5\u00b0C and iv) parasite density at day 3 equal or greater than 25% of that at day 0. Late Clinical Failure (LCF) was defined as danger signs of/or severe malaria and/or parasitaemia with axillary temperature \u2265 37.5\u00b0C between day 4 and day 45, without having been previously classified as ETF. Late parasitological failure (LPF) was defined as reappearance of parasitaemia between day 4 and day 45 without fever and without previously meeting any of the criteria for ETF or LCF. An adequate clinical and parasitological response (ACPR) was defined as absence of parasitaemia by day 45 without previously meeting any of the criteria for ETF, LCF and LPF. The overall rate of treatment failure  was computed as if the patient had an ETF, LCF or a LPF. Only parasitaemia confirmed by PCR as recrudescence was considered as treatment failure. Patients were also considered treatment failures if they received rescue treatment on or before day 45. As both drugs were registered and used in Zambia, no stopping rules were defined.All adverse events (AEs) were recorded on the Case Record Form (CRF). An AE was defined as \"any unfavorable and unintended sign, symptom, or disease temporally associated with the use of the drug administered\". A causality assessment of the AEs was done according to the guidelines of WHO-Uppsala Monitoring Centre (WHO-UMC).2 or Fisher's exact test (when required); Student's t-test was used for continuous variables. Paired t-test was used for within patient comparisons. All reported p-value are two-sided. Non-parametric tests  were used for non-normally distributed variables. For the intention-to-treat analysis the log-rank test and the Hazard Ratio (Cox regression) were estimated after testing for proportional hazard assumption (Schoenfeld test). Data of the patients excluded or lost to follow up were censored at the time of the last recorded visit. All possible interactions up to order two were tested.Data were double entered and cleaned in Epi-info . All analyses were performed using STATA statistical analysis software package version 8 . Proportions were compared using the \u03c7P < 0.001).Between March 2003 and June 2005, a total of 971 patients were randomized to either SP (486) or AL (485). At enrolment the 2 groups had similar demographic and clinical characteristics (Table vs SP: 72/485. P = 0.41). The main reason for withdrawal and loss to follow up (>80%) was population movement. Patients excluded or lost during follow up were younger  but other demographic and clinical characteristics were similar.By day three, 35 (3.6%) patients were lost to follow up and 120 (12.4%) by day 45; 12 patients withdrew and 9 were excluded . The percentage of lost to follow up were similar in both treatment arms (AL: 83/486 P < 0.0001) and fever  (P = 0.007) was significantly lower in the AL than in the SP group (Table P < 0.001). SP was also a risk factor for LCF  and LPF  (Table P < 0.001) . Average time to recrudescence was significantly different between treatment arms .SP and AL were generally well tolerated. Almost all clinical symptoms cleared faster with AL Table . At day up Table . PCR genP < 0.001). Gametocyte carriage during follow up was also significantly higher in the SP group (Table P < 0.001); AL: -2.3 g/l (P = 0.003). At day 45 mean Hb was significantly higher in the AL (134 g/l) than in the SP (130 g/l) group (P = 0.02). By day 45, compared to day 0, Hb had significantly increased in the AL (+ 2.7 g/l) (P = 0.02) but not in the SP group   and one treated with AL who developed a rash. Eleven patients (5 on AL and 6 on SP) had various symptoms possibly related to the study drugs but none serious enough to interrupt the treatment.This trial was conducted in Zambia where a six-dose regimen of AL has been recently implemented as the first line treatment for uncomplicated malaria in non-pregnant adults and children over 10 kg. However, SP was still the standard therapy in March 2003 and this treatment was used as control. AL was clearly better than SP for several outcomes such as parasitological clearance, resolution of symptoms and safety. Hb followed a similar pattern, the drop observed at day 14 was more pronounced and the hematological recovery by day 45 lower in the SP arm. As expected, SP-treated patients had a significantly higher prevalence of gametocytes even if some patients treated with AL had gametocytes at day 28. These results confirm previous findings ,13,14 anDespite the high cure rate, a similar rate of new infections was observed in both treatment groups. This is a concern as routine services do not make any distinction between recrudescence and new infections. In this mesoendemic study area, the ratio new infections/recrudescence was relatively low but in places where the malaria transmission is more intense it could increase substantially. The new infections and the residual number of parasites of the 'old' infection would be exposed to sub therapeutic doses of lumefantrine, the long acting partner drug in the AL combination, and this might increase the selection of resistant strains . FurtherIn Zambia, HIV-1 prevalence is estimated at 25.2 % among mothers attending the antenatal clinic . HIV-1 iIn conclusion, this study, despite the incompletely supervised treatment, confirmed the excellent efficacy and safety/tolerability of the six-dose AL in adults. Next to pharmacovigilance, further research is still needed to ensure its correct deployment, possibly to pregnant women.Modest Mulenga organized the collection of data, supervised the trial and contributed to the data interpretation and writing the paper. Jean-Pierre Van geertruyden contributed to the analysis plan, data interpretation, produced the final dataset, did the analysis and wrote the paper. Lawrence Mwananyanda organized the collection of data, supervised the trial and contributed to the data interpretation and writing the paper, Victor Chalwe supervised the trial and contributed to the data interpretation, Filip Moerman contributed to the study design and training. Roma Chilengi contributed to the study design and training. Chantal Van Overmeir and Jean-Claude Dujardin organized the PCR essays. Umberto D'Alessandro contributed to the study design, the analysis plan, data interpretation and the writing of the paper."}
+{"text": "Plasmodium falciparum in patients from Turbo (Urab\u00e1 region), El Bagre and Zaragoza (Bajo Cauca region), applying the 1998 protocol of the World Health Organization (WHO). Monotherapies using chloroquine (CQ), amodiaquine (AQ), mefloquine (MQ) and sulphadoxine-pyrimethamine (SP), and combinations using chloroquine-sulphadoxine-pyrimethamine (CQ-SP), amodiaquine-sulphadoxine-pyrimethamine (AQ-SP), mefloquine-sulphadoxine-pyrimethamine (MQ-SP) and artesunate-sulphadoxine-pyrimethamine (AS-SP), were examined.Evaluate the frequency of failure of eight treatments for non-complicated malaria caused by A balanced experimental design with eight groups. Samples were selected based on statistical and epidemiological criteria. Patients were followed for 21 to 28 days, including seven or eight parasitological and clinical evaluations, with an active search for defaulting patients. A non-blinded evaluation of the antimalarial treatment response  was performed.Initially, the loss of patients to follow-up was higher than 40%, but the immediate active search for the cases and the monetary help for transportation expenses of patients, reduced the loss to 6%. The treatment failure was: CQ 82%, AQ 30%, MQ 4%, SP 24%, CQ-SP 17%, AQ-SP 2%, MQ-S-P 0%, AS-SP 3%.The characteristics of an optimal epidemiological monitoring system of antimalarial treatment response in Colombia are discussed. It is proposed to focus this on early failure detection, by applying a screening test every two to three years, based on a seven to 14-day follow-up. Clinical and parasitological assessment would be carried out by a general physician and a field microscopist from the local hospital, with active measures to search for defaulter patients at follow-up. Plasmodium, mutations appear with a frequency of 9.5-71.2 \u00d7 10-10 mutations/site/year in coding regions [-4 mutations/site/year in microsatellites [After the use of any antimicrobial, treatment failure or resistance occurs in a percentage of cases, due to the variants or genetic mutants. In in vitro evaluation of resistance of Plasmodium to an antimalarial drug consists of assessing growth inhibition when exposed to different concentrations of the drug, and several methods have been described [in vivo evaluation, or therapeutic efficacy test (TET), measures the clinical and parasitological response of malaria patients to a specific antimalarial treatment. The clinical response is indicated by the clearance of symptoms and signs, and the parasitological response by the clearance of blood parasites, after oral administration of recommended doses of one or several drugs, under supervision of the sanitary staff, and with a follow-up during a determined period . The TET evaluation protocol, approved by the World Heath Organization (WHO) in 1998 [Plasmodium, which affect humans: Plasmodium vivax and, especially, Plasmodium falciparum; the latter has shown resistance to many antimalarial drugs [The escribed -6. The iIn Colombia, between 1961 and 2003  collects and summarizes the available information about treatment failure in Colombia; b) presents the results of the evaluation of TET for various treatment schedules, between 2000 and 2004, in the municipalities of Turbo (Urab\u00e1 region), El Bagre and Zaragoza (Bajo Cauca region), in the north-west and north-east of the state of Antioquia, Colombia; c) collects and summarizes information about use of the combination AQ-SP for the treatment of non-complicated falciparum malaria; and d) discusses the creation of an epidemiological surveillance system of antimalarial treatment failure as part of the malaria programmeme operating in the country.The TET with eight schemes was assessed in a sequential way, between 2000 and 2004. All throughout the study period at least two simultaneous groups were being studied. Patients were randomly assigned a specific treatment as they arrived; this was guaranteed by a blind lottery (tombola). The group of studies constitute an experimental design   with eigUrab\u00e1 and Bajo Cauca, two regions of the Antioquia state, generate 90% of malaria cases of this state. Urab\u00e1 is located on Urab\u00e1 Gulf in the Caribbean sea, near the borders with Panama; the territory has mostly a warm climate. Antioquian Bajo Cauca is located north of the state and has mostly a humid-very humid warm climate -20.2, while the Bajo Cauca region extends over 8,498 km2. In June 2002, Urab\u00e1 had 463,496 inhabitants and Bajo Cauca 219,951. The Urab\u00e1 population density was 36,4 inhabitants/km2 in 1998 and 39,7 in 2002; in Bajo Cauca it was 21.5 in 1993 and 25.8 in 2002 [The Urab\u00e1 region extends over 11,671 km in 2002 -20. In t in 2002  but thisIn Turbo, 60% of the population lives in rural areas , while in El Bagre only 38% lives in the rural area. The age structure of the two populations is similar . In 2002P. falciparum resistant to chloroquine and other drugs, as was the case on the borders of Thailand with Cambodia and Myanmar [Outwash mining is the principal economic activity of Bajo Cauca, followed by cattle farming and, to a lesser degree, rice and sorghum cultivation . In Urab Myanmar .The mean Annual Parasite Index (API) was 39 cases per 1,000 inhabitants in Turbo (1995\u20132000), 134 in El Bagre (1995\u20132000) and 33.5 in Zaragoza (1998\u20132000). For the period 2001\u20132003, the API was 19.5 in Turbo, 37.5 in El Bagre and 34.5 in Zaragoza (DSSA data). These data, with an API above 10/1,000, confirm the high-risk for malaria transmission in these regions.The calculated size sample was 42 for each of the eight treatment schedules, and for each region selected . The sample size was increased to 50 patients (a 20% increase) to compensate for losses during the follow-up period. This sample size was based on the 1998 WHO protocol . The conP. falciparum confirmed by thick smear. These patients attended the malaria diagnosis clinic at their local hospital. Males and non-pregnant females (confirmed by a fast pregnancy test), above one year of age, living in the rural or urban zone of their municipality, where they normally reside, were included in the study.The reference population was composed of patients with a history of fever for two to three days, with symptoms compatible with malaria, and a unique infection with Inclusion criteria, besides the features described in \"Reference Population\" were: a) to have non-complicated malaria ; b) to harbor between 250 and 50,000 asexual parasites/\u03bcL; c) exclusion of concomitant disease, after a clinical interview and examination carried out by the research team; d) voluntary participation in the study; e) commitment to attend the follow-up examinations.Criteria for exclusion of the study were: a) to evidence, at any moment, severe malaria or any other disease; b) appearance of serious adverse effects due to antimalarial drugs, to other medicines, or to other causes; c) failure to attend any of the first three follow-up examinations . The evidence of treatment failure to antimalarial therapy resulted in withdrawal from the study and recommendation of a rescue treatment, formulated according to the Colombian Health Ministry guidelines for malaria treatment .3, according to 1998 WHO protocol [Malaria diagnosis was carried out with thick smear according to WHO procedure  and routprotocol .Each patient, upon admission to the study and at each follow-up, was given: a) a medical evaluation to diagnose the presence of malaria; b) a parasitological evaluation . Special emphasis put on finding, identifying, classifying and defining the origin of adverse effects, which appeared during the 21 to 28 days of follow-up. This search for adverse effects was based on clinical assessment , rather than laboratory evaluation. The field team was composed, in each municipality, of a general physician and a clinical laboratory technician, interacting with the local malaria team, a microscopist and a health assistant. An active search for defaulting patients was carried out when needed in order to re-integrate them to the study.The drugs were taken with water, administered in the doses established by the Colombian Ministry of Health  Table 22. Each tThe follow-up period lasted 28 days in the three groups  and 21 days in the five other groups . TET was classified based on the 1998 WHO protocol  with min1. Early treatment failure, defined as the presence of any of the following:a) Development of danger signs or severe malaria on Day 1 Day 2 or Day 3b) Parasitaemia on day 2 higher than day 0;c) Parasitaemia on day 3 \u2265 25% of count on day 0.2. Late treatment failure, defined as the presence of any of the following:a) Danger signs or severe malaria in the presence of parasitaemia (same species of day 0) after day 3;b) Non-programmemed return of the patient between days 4 to 21  or 28 (for other), due to a clinical deterioration in the presence of parasitaemia (same species of day 0).c) Parasites (same species of day 0) on days 7, 14, 21, or 28.3. Adequate clinical-parasitological response: if the patient did not present either early or late failures.The EpiInfo 6.04 programmeme was used to make questionnaires and data bases and also to analyse information . SPSS 10.0 was also used for data analysis and graph construction.2); the means were compared with Wilcoxon or Kruskal-Wallis (K-W) tests. A significance level of 5% (\u03b1 = 0.05) was always established.Associations among groups or independent variables  were measured with square chi test  of Universidad de Antioquia.Of the 520 patients included in the study, 47% were recruited in Turbo (Urab\u00e1 zone) and 53% in The Bajo Cauca zone (El Bagre 37% and Zaragoza 16%). Data were analysed independently from the residence place of the patient since important differences were not found when this variable was considered.The patients exhibited the following characteristics:a) 61% were living in the rural zoneb) 69% were menc) 29% were under 15 years of age .TET could be measured in 93.7% of the patients (487 out of 520 patients). The reason for not evaluating TET in 33 patients was the impossibility of finding them (90%: 30/33) or voluntary withdrawal from the investigation (10%: 3/33). Failure to attend increased further into the follow-up time. Such defaulting was statistically different among treatments, and the four monotherapies had less defaulting overall (4%) than the four treatment combinations (8%). This percentage of defaulting per treatment varied from major to minor: a) in monotherapies: it was 9% with MQ, 3% with AQ, none with CQ and SP; b) in combinations: it was 12% with MQ-SP, 10% with AQ-SP and AS-SP, none with CQ-SP. Defaulting was higher (9.3%) in El Bagre-Zaragoza, which largely depends for communication on the Nech\u00ed River, than in Turbo (5.7%), in which basic transport is by land. The group of 33 non-attending patients did not differ from the non-defaulters (n = 487) in relation to the residence zone , sex, ethnicity, age, days of evolution of the current disease and initial parasitaemia.The mean of asexual parasitaemia in 487 patients before treatment administration was 7,700 parasites/\u03bcL, the median was 4,480, the 25 percentile was 2,000; the 75 percentile was 10,130 and the standard deviation was 8,472 parasites/\u03bcL. A 26% of patients had as many as 2,000 parasites/\u03bcL, 45% as many as 4,000, 50% up to 5,000, 75% up to 10,000 and only 8% over 20,000 parasites/\u03bcL.The mean asexual parasitaemia before treatment administration was not statistically equal among the treatments; it varied between 8,954 (MQ) and 5,203 parasites/\u03bcL (CQ-SP) (p < 0.05); between MQ and SP  there was a statistically significant difference. Gametocytes were found in 17% of patients before treatment, in 23% on day 1, 33% on day 2, 43% on day 3, 65% on day 7, 64% on day 14, 51% on day 21, and 15% on day 28. Before the treatment, the mean number of gametocytes ranged from 22 (AS-SP) to 140 gametocytes/\u03bcL (AQ-SP), with a general mean of 78 and a standard deviation of 421 gametocytes/\u03bcL .Monotherapy with CQ showed 82% failure, with AQ it showed 30%, with MQ 4%, and with SP 24%, while the combinations of SP with other drugs showed the following failure rates: CQ-SP 17%, AQ-SP 2%, MQ-SP 0% and AS-SP 3% (Table 4). There were 62 failures  in 487 patients in whom the final effect could be measured. Early failure contributed with 42% of the total failure (26/62) and the late failure accounted for 58% (36/62) of it. Considering defaulting patients as failures the percentage of failure reached 18% (62 observed failures + 30 supposed failures/520). The frequency of early and late failures changes from one treatment to another: a) with monotherapies, early and late failures were equal to the those with CQ, early failures were predominant with AQ (ratio 1:4), and the late ones were the majority with SP (2:1); b) with the CQ-SP combination, the late failures widely prevailed (9:1). With the rescue treatment, which varied according to the primary treatment, all treatment responses were successful; which meant no failure of two treatments was observed.Treatment failure was not associated with initial asexual parasitaemia (pre-treatment), although there were important differences as follows: those who successfully responded and those who had early failure had 7,760 and 7,951 parasites/\u03bcL, respectively, while those who had late failure showed 6,731 parasites/\u03bcL; despite that, the differences were not statistically significant. Neither was there any relation of TET with initial gametocytaemia. Thus, although initial asexual parasitaemia was not equal in all eight treatments, this fact did not have any effect in the TET.It should be pointed out that there was no significant association of the variable TET with any one of these explicative variables: municipality, zone, sex, age group, ethnic group, malaria antecedents, and hospitalization or transfusion antecedents.Antimalarial schemes administered  corresponded to those used for many years in Colombia and recommended by other authors ,26. DireThe protocol used for measuring TET was identical in most studies, with exception of the studies of 2000\u20132001, when CQ, SP and CQ-SP were evaluated over 21 days; in those undertaken between 2001 and 2004, the follow-up lasted 28 days.The number of patients in whom TET could not be measured (lost patients) was low (6% in the eight studies) and was not affected by the minimum sampling size. The strategy used to avoid defaulting consisted of monetary support  to cover transportation costs, as well as the Immediate Active Search (IAS) for any patient who missed a follow-up examination. In fact, defaulting rate has been higher than 40%, but was reduced to 6% through the application of that strategy. IAS was performed by the physician or clinical laboratory technician, accompanied by the health assistant, using motorcycles supplied by the local hospital. Even in the worst case scenario, i.e. if all patients in whom TET was could not be measured are considered as failures, the percentage of treatment failure for each treatment schedule was under 14%, except in the case of AQ, which had net failure (excluding defaulters) of 30%, and gross failure of 33% (when counting defaulters as failures).et al. [et al [From the evaluated schizonticide monotherapies , only CQ has been used in this way in Colombia, although associated to primaquine as gametocide (45 mg in a single dose). CQ had become useless in 1998 , dueThe accelerated growth of failure presented by the monotherapy AQ is serious because it increased from 7% in 1998 in El Bagre , ZaragozFailure of the monotherapy with SP also increased strongly, going from 12% in 1998 -30 to 24The AQ-SP combination is highly efficient (98%). The relatively scantly use of the AQ was a result of the adverse reactions which had been observed when AQ was used for prophylaxis , a situaA systematic review of 2001 compared AQ-SP and CQ-SP combinations for the treatment of non-complicated malaria; on the basis of seven studies in 1,277 patients, it concluded that there was no evidence of serious collateral effects with those combinations and that they, more than monotherapy with SP, \"could make people feel better more rapidly\", and caused a better steady elimination of blood parasites . In 2005According to the results obtained in this study and to the additional information presented here, treatment with AQ-SP should remain as the first line treatment of non-complicated falciparum malaria in Colombia, as it is at present , leavingP. falciparum in vitro resistance; c) P. falciparum y P. vivax genetic analysis and d) assessment of antimalarial blood concentrations in patients studied.Colombia should establish an \"Antimalarial Failure Epidemiological Surveillance System\" (AFESS). The AFESS should have, as primary objective, the early detection of failure by means of a screening test. For this, a selected screen test should be low cost, applicable in field conditions and have a high sensitivity. Our recommendation is to install an AFESS coupled to the antimalarial programmeme, to measure TET during a following up period of 7 \u2013 14 days and to install sentinel posts in strategic places of malaria regions. In addition the AFESS, should: a) carry out specific research projects aimed at confirmation of therapeutic failure after a following up period of 28 days or more, b) evaluation of"}
+{"text": "The study investigated the pharmacokinetics of fosmidomycin when given alone and in combination with clindamycin in patients with acute uncomplicated falciparum malaria.Plasmodium falciparum malaria who fulfilled the enrollment criteria were recruited from out-patient department of Mae Sot Hospital, Tak Province, Thailand. Patients were treated with monotherapy with fosmidomycin at the dose of 1,200 mg every 8 hours for 7 days (n = 15) or combination therapy with fosmidomycin (900 mg every 12 hours for 7 days) and clindamycin (600 mg every 12 hours for 7 days) (n = 18). Blood samples were taken for pharmacokinetic investigations of clindamycin and/or fosmidomycin and 24-hour urine samples were collected during dosing period. Efficacy assessments included clinical and parasitological evaluation. Safety and tolerability were assessed based on clinical and laboratory investigations.A total of 15 and 18 patients with acute uncomplicated z/F and CL/F, which were significantly smaller in the combination regimen. Plasma concentration-time profiles of both fosmidomycin and clindamycin were best fit with a one-compartment open model with first-order absorption and elimination and with absorption lag time. Steady-state plasma concentrations of fosmidomycin and clindamycin were attained at about the second or third dose. There was no evidence of dose accumulation during multiple dosing. Urinary recovery of fosmidomycin was 18.7 and 20% following mono- and combination therapy, respectively.Both mono- and combination therapy regimens of fosmidomycin were well tolerated with no serious adverse events. Combination therapy with fosmidomycin and clindamycin was proven highly effective with 100% cure rate, whereas cure rate of monotherapy was 22% (28-day follow up). Pharmacokientics of fosmidomycin following mono- and combination therapy were similar except VP. falciparum.Pharmacokinetic dose optimization of fosmidomycin-clindamycin combination therapy with the course of treatment of not longer than three days is required to obtain a regimen which is safe and produced 100% cure for multidrug-resistant  Plasmodium falciparum strains resistant to commonly used antimalarials. In addition to the high-grade chloroquine-resistant strains found in high frequencies in all endemic areas of the world, resistance to sulphadoxine/pyrimethamine is now common in large parts of Asia and South America and is spreading to Africa as well. Development of drug resistance and in some cases, concerns over safety, highlight the urgent requirement for new antimalarial drugs. Primarily directed towards the treatment of multidrug resistant P. falciparum, such drugs should possess novel modes of action while being of proven efficacy and safety.Malaria is one of the leading causes of morbidity and mortality in the tropics with an annual estimate of 500 million clinical cases and 2 million deaths . The treD-xylulose 5-phosphate (DOXP) reductoisomerase, an essential enzyme of the non-mevalonate pathway and therefore, selectively blocks the biosynthesis of isopentenyl diphosphate and the subsequent development of isoprenoids in P. falciparum  for FCTs and PCTs for mono- vs combination therapy were 56 (24\u2013104) and 56 (8\u201380) hr vs 40 (24\u201372) and 40 (16\u201380) hr, respectively. Concomitant medication included paracetamol, dimenhydinate, diphenhydramine and chlorpheniramime.Demographics, admission clinical and laboratory data of the groups of patients are presented in Table Observed and model-predicted (one-compartment model with first-order absorption and elimination with absorption lag time) median plasma concentrations of fosmidomycin and clindamycin following monotherapy and combination therapy are shown in Figures z/F and CL/F were significantly smaller following combination therapy; other pharmacokinetic parameters were comparable between the two treatment regimens.The pharmacokinetics of fosmidomycin given as monotherapy and combination therapy including pharmacokinetics of clindamycin given as combination therapy with fosmidomycin in individual patients analysed by non-compartmental pharmacokinetic analysis approach are summarized in Table Table vs 20.0 (6.9\u201343.3)%].Urinary concentrations of fosmidomycin were determined during the dosing period of 7\u201310 days and results showed that percentage of total amount of fosmidomycin excreted during 96\u2013120 hr were markedly high following both mono- and combination therapy of fosmidomycin. There was no significant difference in urinary recoveries of fosmidomycin (expressed as percent of dose administered) following monotherapy and combination therapy [median (95% C.I.): 18.7 (6.5\u201342.1) min-ss of fosmidomycin in two patients with successful treatment on day 28 following fosmidomycin monotherapy were 2.16 and 1.24 \u03bcg/ml, while the levels in 9 patients who developed recrudescence were 1.19, 1.42, 2.79, 0.67, 0.27, 4.57, 1.95, 0.06 and 1.47 \u03bcg/ml. In the group treated with fosmidomycin-clindamycin combination therapy, all were cured despite relatively low Cmin-ss of 0.20 \u03bcg/ml. The pharmacokinetics of fosmidomycin in patients who had successful or treatment failure following fosmidomycin monotherapy appeared to be similar.CFosmidomycin mono- and combination therapy with clindamycin were well tolerated with no serious adverse events, deaths or withdrawals due to adverse events. No neutropaenia or drop in haemoglobin were observed as previously reported in children . FollowiN-desmethyl clindamycin and clindamycin sulphoxide) [max-ss and Cmin-ss of fosmidomycin and clindamycin following multiple dosing regimens. The trough/peak plasma concentrations after both dosage regimens increased up to the second or third dose and remained at steady-state thereafter. Fosmidomycin was not metabolised and excreted unchanged in urine as a bioactive substance. The urinary recovery expressed as percent of the administered doses were relatively low (18 and 20% following mono- and combination therapy) comparing to those reported in healthy subjects (26%) [max-ss and Cav-ss values, the unchanged elimination half-lives after the first and last dose, and the constant urinary recovery during 0\u20137 days of dosing. In a previous study, saturated absorption of fosmidomycin after multiple dosing was suggested as there were a progressive decline in total urinary recovery from 35.9 to 22.4% during multiple dosing [A bioassay based on disk diffusion technique was applied for determination of fosmidomycin in plasma and urine as the drug is the only bioactive compound. The analysis of plasma concentrations of clindamycin was performed by HPLC since bioassay lacks ability to discriminate between clindamycin and its active metabolites (phoxide) -20. Fosmphoxide) . The biophoxide) ,19,22. Pphoxide) ,23,24. Its (26%) . This exe dosing .z/F and CL/F when clindamycin doses were added, while t1/2z remained unchanged. It is not clear whether these changes are contributed mainly by malaria infection or pharmacokinetic interaction with clindamycin or both. Reduction of renal clearance of fosmidomycin when clindamycin was given concurrently may account partly for this pharmacokinetic difference. Fosmidomycin is only eliminated by renal clearance while clindamycin is metabolised by CYP3A in liver to clindamycin sulphoxide and N-desmethyl clindamycin [1-acid glycoprotein [The non-compartment pharmacokinetic analysis of fosmidomycin monotherapy was investigated during the acute phase of malaria infection on the first day whereas its kinetics following combination therapy with clindamycin was investigated following the last dose on day 7. Data suggest that malaria infection may not have significant influence on the pharmacokinetics of both fosmidomycin and clindamycin. Although amount of fosmidomycin excreted in urine appears to be relatively low in patients with malaria compared with healthy subjects, overall pharmacokinetic profiles were similar. The pharmacokinetics of fosmidomycin when given alone or with clindamycin were generally comparable except for the significant paralleled reduction of Vndamycin . Plasma oprotein -23. Ther50 value for recombinant P. falciparum DOXP reductoisomerase is approximately 40 nM and varied from 300 to 1,200 nM in P. falciparum in vitro without obvious cross-resistance with other antimalarials [in vitro growth of P. falciparum is inhibited with an IC50 and IC90 of approximately 25 and 50 nM, respectively. As a consequence of its high activity but slow onset of activity, clindamycin is recommended for the treatment of asymptomatic malaria or in combination with other antimalarials. It has been suggested that time-dependent antimicrobial agents should remain above the MIC of pathogens for at least three malaria cycle of seven days. Unlike fosmidomycin, recent studies have suggested that clindamycin displays concentration-dependent bacteriocidal activity [in vitro post antibiotic effect (PAE) for certain period. In vivo, the PAE is generally longer and due to effects such as post antibiotic leukocyte enhancement and post-antibiotic sub-MIC effect. Frequent dosing interval of clindamycin regimen is therefore not necessary. The recommended oral dosing of clindamycin for bacterial infection was 600 mg every 6 hr or 900 mg every 8 hr. However, it was later shown that the pharmacokinetics and concentration-time profiles were comparable with the dosing regimen 1,200 mg every 12 hr which is more practical for clinical application. This may explain the sustained antimalarial efficacy when clindamycin was used with fosmidomycin despite low minimum plasma concentrations of fosmidomycin at steady-state. Whether the antimalarial activity of clindamycin bioactive metabolites should also be considered when performing a pharmacokinetic/pharmacodynamic evaluation has not yet been elucidated. In human, the formation of N-desmethyl clindamycin and its excretion in urine and faece have been confirmed [Fosmidomycin monotherapy was shown safe and effective blood schizonticide in the initial clearance of asexual parasitaemia. However, the high rate of recrudescence precludes its use in monotherapy ,10 Monotalarials ,26. Plasalarials ,28. As aactivity ,29 whileAUC Area under the plasma concentration-time curvemin-obs Observed trough concentrationCmax Maximum plasma concentrationCmax-ss Maximum plasma concentration at steady-stateCave-ss Average plasma concentration at steady-stateCCI Confidence intervalCL Total clearanceCV Coefficient of variationF BioavailabilityFCT Fever clearance timeHPCE High performance capillary electrophoresisHPLC High performance liquid chromatography50 Fifty percent inhibitory concentrationICMIC Minimum inhibitory concentrationPAE Post antibiotic effectPCT Parasite clearance timemax Time to maximum plasma concentrationtz Apparent volume of distributionVI Kesara Na-Bangchang was overall study supervisor and analyst of pharmacokinetic data including co-author for the manuscript. R Ruenweerayut was the site investigator and overall medical coordinator. J Karbwang and D Huchington reviewed the study protocol and manuscript and and provided technical support. A Chauemung was responsible for the analysis of pharmacokinetic samples (determination of fosmidomycin and clindamycin in plasma and urine).KN participated in the design of the study, performed pharmacokinetic and statistical analysis, as well as drafting the manuscript. RR was responsible for the conduct of clinical part of the study . AC carried out the analysis of fosmidomycin and clindamycin in plasma and urine samples. JK and DH conceived of the study, and participated in its design and coordination. All authors read and approved the final manuscript."}
+{"text": "To compare the efficacy and safety of artemisinin combination therapies for the treatment of uncomplicated falciparum malaria in Uganda.Randomized single-blind controlled trial.Tororo, Uganda, an area of high-level malaria transmission.P. falciparum malaria.Children aged one to ten years with confirmed uncomplicated Amodiaquine + artesunate or artemether\u2013lumefantrine.Risks of recurrent symptomatic malaria and recurrent parasitemia at 28 days, unadjusted and adjusted by genotyping to distinguish recrudescences and new infections.Of 408 participants enrolled, 403 with unadjusted efficacy outcomes were included in the per-protocol analysis. Both treatment regimens were highly efficacious; no recrudescences occurred in patients treated with amodiaquine + artesunate, and only two occurred in those treated with artemether\u2013lumefantrine. However, recurrent malaria due to new infections was common. The unadjusted risk of recurrent symptomatic malaria was significantly lower for participants treated with artemether\u2013lumefantrine than for those treated with amodiaquine + artesunate . Similar results were seen for the risk of recurrent parasitemia . Amodiaquine + artesunate and artemether\u2013lumefantrine were both well-tolerated. Serious adverse events were uncommon with both regimens.Amodiaquine + artesunate and artemether\u2013lumefantrine were both highly efficacious for treatment of uncomplicated malaria. However, in this holoendemic area, despite the excellent performance of both regimens in terms of efficacy, many patients experienced recurrent parasitemia due to new infections. Artemether\u2013lumefantrine was superior to amodiaquine + artesunate for prevention of new infections. To maximize the benefit of artemisinin combination therapy in Africa, treatment should be integrated with strategies to prevent malaria transmission. The impact of frequent repeated therapy on the efficacy, safety, and cost-effectiveness of new artemisinin regimens should be further investigated. Background: Malaria parasites have become resistant in much of Africa to many commonly used treatments, such as chloroquine and sulfadoxine\u2013pyrimethamine. Newer drugs, such as artemisinin-based combination therapies (ACTs), have been used extensively in Southeast Asia. Artemether\u2013lumefantrine (an ACT) has now been adopted as first-line malaria treatment in Uganda, with the combination of amodiaquine and artesunate as a backup treatment. There are two ways that successful treatment is measured; first by whether the treatment works in curing the infection; and second by whether it prevents the disease recurring\u2014either the same infection (known as recrudescence), or a new infection. The researchers wanted to look at how artemether\u2013lumefantrine compared with amodiaquine and artesunate for treating symptomatic malaria, and also at whether there were any differences in recurrence, either of clinical malaria, or of parasite infection without symptoms, for 28 days after treatment.What this trial shows: This randomized trial in young children with confirmed malaria in Uganda showed that both treatments resulted in a similar initial response to therapy, as measured by the risk of early treatment failure (within three days). The researchers found that artemether\u2013lumefantrine was more effective at reducing the risk and delayed the time to recurrence of malaria, compared with amodiaquine and artesunate. Additionally, treatment with artemether\u2013lumefantrine resulted in a reduced rate of parasite infection without malaria symptoms, as compared with amodiaquine and artesunate. Nearly all cases of recurrent malaria after either treatment combination arose from new infections, rather than from recrudescences.Strengths and limitations: The trial was correctly designed to test the questions of interest, and enough patients were recruited to properly examine the relative effects of the two treatment combinations. However, patients were only followed up for 28 days after treatment. A longer follow-up period might have revealed a higher rate of recurrence of malaria after treatment. In the trial, artemether\u2013lumefantrine was not administered with food, and it's known that lumefantrine is absorbed better when it is taken with a small amount of fat. It's possible that the effect of artemether\u2013lumefantrine seen in the trial could have been an underestimate of what might be achieved in ideal conditions.Contribution to the evidence: This trial adds information on the efficacy of artemether\u2013lumefantrine as compared with amodiaquine and artesunate in East Africa for treatment of uncomplicated  symptomatic malaria. The results are consistent with those of other trials on the efficacy of ACTs that have been performed in the region. The study also adds data on the risk of recurrent infections in an area where malaria occurs very frequently. Even following treatment with either ACT, the risk of recurrence was very high. Plasmodium falciparum to chloroquine and sulfadoxine\u2013pyrimethamine has required urgent introduction of alternative antimalarial therapies, including artemisinin-based combination therapies (ACTs)  tablets given twice daily for three days. Participants in the amodiaquine + artesunate group also received placebo tablets administered in the evening over three days, dosed similarly to weight-based guidelines for artemether\u2013lumefantrine. Study medications were administered with water. Although participants were encouraged to resume normal food intake, no food was provided with the medications.On day 0, patients were randomly assigned to receive amodiaquine + artesunate or artemether\u2013lumefantrine. A nurse administered study medications according to weight-based guidelines for administration of fractions of tablets (amodiaquine and artesunate) and the manufacturer's recommendations (artemether\u2013lumefantrine). We administered all drugs orally as follows: amodiaquine , 200 mg tablets, 25 mg/kg per treatment) 10 mg/kg on days 0 and 1, and 5 mg/kg on day 2; and artesunate , 50 mg tablets, 12 mg/kg per treatment, 4 mg/kg once daily for three days; artemether\u2013lumefantrine , 20 mg artemether/120 mg lumefantrine tablets, 3-d six-dose regimen administered according to weight, as one [10\u201314 kg], two [15\u201324 kg], three [25\u201334 kg], or four [All treatment was directly observed. Participants were given the option either to wait at the clinic for the evening dose (lunch was provided) or to leave the clinic and return in the evening (transport was provided). After each dose, children were observed for 30 min and the dose was readministered if vomiting occurred. Children who repeatedly vomited their first dose of study medication were excluded from the study and referred for further management. We provided all patients with a 3-day supply of paracetamol for treatment of febrile symptoms. Those with a concentration of hemoglobin of less than 10.0 g/dL were treated according to Integrated Management of Childhood Illness guidelines with ferrous sulfate for 14 days and given antihelminthic treatment if appropriate.The objectives of the study were to compare the efficacy and safety of amodiaquine + artesunate and artemether\u2013lumefantrine for the treatment of uncomplicated falciparum malaria in Uganda.The primary efficacy outcomes were the 28-d risks of recurrent symptomatic malaria  and recurrent parasitemia , unadjusted and adjusted by genotyping. Secondary efficacy outcomes included risk of fever and parasitemia during the first three days of follow-up, change in mean hemoglobin from day 0 to day 28, or day of repeat therapy, and risk of gametocytemia during follow-up in participants lacking gametocytes at enrollment.Treatment outcomes were classified according to 2003 WHO guidelines as early treatment failure , late clinical failure , late parasitological failure , and adequate clinical and parasitological response  . PatientMolecular genotyping techniques were used to distinguish recrudescent from new infections for all patients failing therapy after day 3. Briefly, filter paper blood samples collected on the day of enrollment and on the day of failure (LCF or LPF) were analyzed for polymorphisms in merozoite surface protein-1 (MSP-1) and \u22122 (MSP-2) using nested-PCR as previously described . First, Secondary safety outcomes included risk of serious adverse events, and risk of events of moderate or greater severity. At each follow-up visit, patients were assessed for any new or worsening event. An adverse event was defined as any untoward medical occurrence, irrespective of its suspected relationship to the study medications . All events were graded by severity  and relationship to study treatment  using guidelines from the World Health Organization  and the National Institutes of Health, Division of Microbiology and Infectious Diseases . A serious adverse event was defined as any adverse experience that resulted in death, life-threatening experience, inpatient hospitalization, persistent or significant disability or incapacity, or specific medical or surgical intervention to prevent serious outcome.http://statpages.org/proppowr.html.We calculated sample size to test the hypothesis that treatment with artemether\u2013lumefantrine would decrease the risk of recurrent symptomatic malaria by 15% at 28 days compared with amodiaquine + artesunate. The risk over 28 days of repeat therapy for symptomatic malaria (unadjusted by genotyping) with amodiaquine + artesunate was estimated as 50% based on previous data . Using tAn off-site investigator prepared computer-generated age-stratified randomization codes for two age groups (12\u201359 mo and 5\u201310 y) which were provided to a study nurse responsible for treatment allocation. The randomization list was secured in a locked cabinet accessible only by the study nurse. Participants were enrolled by the study physicians, and treatments were assigned and administered by the study nurse.Only the study nurse was aware of treatment assignments. All other study personnel, including the study physicians and laboratory personnel involved in assessing outcomes, were blinded to the treatment assignments. Patients were not informed of their treatment regimen.t-test. The non-parametric Wilcoxon rank sum test was used to analyze continuous data with a skewed distribution. All reported p-values are two sided without adjustment for multiple testing and were considered statistically significant if <0.05.Efficacy data were evaluated using a per-protocol analysis of patients with defined treatment outcomes. Analysis was performed using STATA version 8.0  statistical software program. Pair-wise comparisons of treatment efficacy were made using risk differences with exact 95% confidence intervals. Risks of recrudescence were estimated using the Kaplan\u2013Meier product limit formula with censoring for new infections. Secondary outcomes included the presence of fever on days 1\u20133, parasitemia on days 2 and 3, change in hemoglobin level between the day of enrollment and the last day of follow-up, presence of gametocytes during any follow-up day, and the incidence of adverse events. Other categorical variables were compared using Chi-squared or Fisher's exact test and continuous variables were compared using the independent samples Of 532 patients screened, 419 were randomized to treatment, and 408 were enrolled in the study . PrimaryThe study was conducted between December 2004 and July 2005.Among patients with efficacy outcomes, there was no difference in baseline characteristics between the two treatment groups .All 403 participants with unadjusted efficacy outcomes were included in the per-protocol analysis .The risk of recurrent symptomatic malaria  was significantly lower for participants treated with artemether\u2013lumefantrine than for those treated with amodiaquine + artesunate . Similar results were seen for the risk of recurrent parasitemia . In contrast, in the artemether\u2013lumefantrine group only one new infection occurred before day 21 .The time to recurrent malaria was significantly shorter in participants treated with amodiaquine + artesunate compared to those treated with artemether\u2013lumefantrine . In patiBoth treatments produced rapid clearance of parasitemia . AlthougThe risk of recurrent parasitemia and recurrent symptomatic malaria in this study varied over time . The risArtemether\u2013lumefantrine and artesunate + amodiaquine were both well-tolerated. Overall, 261 (65%) of participants experienced any adverse event of moderate or greater severity, and there was no difference between the two treatment groups . No abnoIn this randomized clinical trial, amodiaquine + artesunate and artemether\u2013lumefantrine were both highly efficacious for treatment of uncomplicated malaria in Tororo, an area with a very high level of transmission of malaria. No recrudescences occurred in patients treated with amodiaquine + artesunate and only two occurred in those treated with artemether\u2013lumefantrine, although additional recrudescences might have been detected if follow-up had been extended to at least 42 days. The performance of artemether\u2013lumefantrine was consistent with results from other recent studies from East Africa \u201316 and pIn this study, despite the excellent performance of amodiaquine + artesunate and artemether\u2013lumefantrine in terms of short-term efficacy, the proportion of patients experiencing recurrent parasitemia within one month was substantial in both treatment groups. Two-thirds of patients treated with amodiaquine + artesunate, and half of those treated with artemether\u2013lumefantrine, were parasitemic during 28 days of follow-up. Nearly all recurrent parasitemias were characterized as new infections. Nonetheless, even with the more efficacious regimen (artemether\u2013lumefantrine), over a quarter of patients had recurrent illness, demonstrating the importance of reinfection in areas of very high malaria transmission. This result highlights the need to re-evaluate the approach to treatment of recurrent episodes of malaria following initial ACT therapy. In our study, participants received quinine as second-line treatment. However it is unclear whether quinine, the same ACT, or a different ACT would be the optimal treatment. Given that nearly all recurrent parasitemias were due to new infections, it may be reasonable to retreat with the same ACT regimen, rather than with quinine. Our results also indicate that integration of approaches to control malaria is essential. In an area of South Africa with lower-intensity transmission, the combination of vector control measures and provision of artemether\u2013lumefantrine dramatically decreased the malaria burden . In areapfmdr1 86N allele, which may play a role in lumefantrine resistance [We found that artemether\u2013lumefantrine was superior to amodiaquine + artesunate for preventing new infections, consistent with recent results from Zanzibar . Becausesistance ,22. As AThe risk of recurrent parasitemia and symptomatic malaria had significant seasonal variation. Although malaria transmission occurs year-round in Tororo, the difference in transmission between dry and wet seasons had a substantial impact on risk of new infection, and therefore on the need for retreatment. The seasonal variation is important to recognize when comparing results of drug efficacy studies conducted at the same site. More broadly, these results highlight the importance of the level of transmission in determining risks of recurrent malaria after therapy. In a prior drug efficacy study conducted at the same site between September 2003 and March 2004, the unadjusted risk of recurrent infection at 28 days was 74% for amodiaquine + artesunate versus 59% for amodiaquine + sulfadoxine-pyrimethamine, with 12% versus 18% risk of recrudescence, respectively . For amop = 0.115), and that artemether\u2013lumefantrine provided greater protection against re-infection compared with amodiaquine + artesunate [A systematic review of the six-dose regimen of artemether\u2013lumefantrine published in 2005 identified nine randomized controlled trials, including four studies conducted in Africa  ,16,23\u201325Our study adds to the evidence base on the comparative efficacy of ACTs in Africa. The results from Tororo support the findings of the Zanzibar trial, although the overall risk of recurrent malaria at 28 days was substantially higher in our study (66% amodiaquine + artesunate versus 51% artemether\u2013lumefantrine) than in Zanzibar .Trial ProtocolClick here for additional data file.(821 KB DOC)."}
+{"text": "Artemisinin-based combination therapy is increasingly being adopted as first-line antimalarial therapy. The choice of appropriate therapy depends on efficacy, cost, side effects, and simplicity of administration.Plasmodium falciparum malaria in eastern Sudan.the efficacy of fixed co-formulated (f) artesunate-sulfamethoxypyrazine-pyrimethamine (AS+SMP f) administered at time intervals of 12 hours for a 24-hour therapy was compared with the efficacy of the same drug given as a loose combination (AS+SMP l) with a dose interval of 24 hours for 3 days for the treatment of uncomplicated seventy-three patients (39 and 34 in the fixed and the loose regimen of AS+SMP respectively) completed the 28-days of follow-up. On day 3; all patients in both groups were a parasitaemic but one patient in the fixed group of AS+SMP f was still febrile.P > 0.05) for the fixed and loose combination of AS+SMP respectively.Polymerase chain reaction genotyping adjusted cure rates on day 28 were 92.3% and 97.1% (Three (4.1%) patients (one in the fixed and two patients in the loose group of AS+SMP) in the study suffered drug-related adverse effects.Gametocytaemia was not detected during follow-up in any of the patients.P. falciparum malaria in eastern Sudan. Due to its simplicity, the fixed dose one-day treatment regimen may improve compliance and therefore may be the preferred choice.both regimens of AS+SMP were effective and safe for the treatment of uncomplicated  Plasmodium falciparum malaria infections [Plasmodium falciparum malaria in Sudan [P. falciparum malaria.There are almost 515 million episodes of clinical fections ,3. Malarfections . Due to P. falciparum malaria had been demonstrated previously [P. falciparum malaria. The manufacturer's  accomplished this combination and new treatment was considered for use as a single dose per day over 3 days. According to the pharmacokinetics property [P. falciparum malaria, since no published data exists yet concerning these drug combinations.The choice of an ACT in specific malaria-endemic areas needs to take into account their efficacy, side effects, price and mode of administration. Common ACT therapies last for 3 days and in some instances the patient has to swallow up to 24 tablets. Sulfamethoxypyrazine (SM) is chemically very close to sulfadoxine but has different pharmacokinetics ,7. The heviously -10. It iproperty ,7, it waP. falciparum malaria [The study was conducted in September-November 2005 at AL Sawagi ALganoubia health center in Kassala, eastern Sudan. The area is characterized by low malaria transmission . Febrile malaria  andno hiAfter obtaining informed consent from the patient or the child's parents, a fixed questionnaire including relevant socio-demographic characteristics, medical history, physical findings and investigations conducted was completed for each patient.Blood films were prepared, Giemsa-stained and examined by 100\u00d7 oil immersion fields. The parasite density was counted against 200 leucocytes, assuming 8000 leucocytes/\u03bcl. All slides were double-checked blindly and only considered negative if no parasites were detected in 100 oil immersion fields. If gametocytes were seen, then the count was extended to 500 leucocytes.P. falciparum merozoite surface protein 1&2 (MSP1 & MSP2) were detected [Three spots of blood were taken on filter-paper initially and later if parasites reappeared microscopically during the follow-up period to differentiate between re-infection and recrudescence using nested PCR where FC 27 and ICI alles of Primers from polymorphic detected ,14.The manufacturers' (Dafra Pharma NV) instructions were strictly followed. The patients were alternatively allocated either to the fixed or the loose combination of AS+SMP. The patients appointed to the fixed dose group were given 3 doses of AS+SMP f orally as a fixed combination . The fixed dose tablets were available in three different dosage strengths, each one containing AS/SM/P mg in different doses; 200/500/25 for adults (\u226540 kg), 100/250/12.5 for children and adolescents (16\u201339.9 kg) and 50/125/6.25 for babies and infants (\u226415.9 kg).The patients in the group receiving the loose combination were also given the treatment according to their weight. They received two different tablets which were colour-coded for ease of administration: pink SMP tablets and white AS tablets. Their treatment regimen lasted for three days with intervals of 24 hours.The tablets were crushed and dissolved in water for children who were not able to swallow them. Subjects were observed for vomiting for one hour; the full dose was repeated for those who vomited within 30 min and half of the dose was repeated if vomiting occurred between 30 and 60 minutes.Patients were requested to come back on days 1, 2, 3, 7, 14, 21 and 28 and at any time they felt unwell. At each visit, body temperature was measured and blood films were prepared. Haemoglobin was measured on day 0 and day 28. During the follow-up the patients were asked if they suffered from side effects which could be expected from the treatment ; these symptoms were considered to be drug related if they had not been reported at the patient's first presentation to the clinic.P. falciparum malaria in Sudan (Artemether-lumfantrine) was given for treatment failures and the patients were followed as above. Classification was done based on the following: Early Treatment Failures (ETF) in case of significant parasitaemia at day 2 or 3 or parasites and fever at day 3, Late Clinical Failures (LCF) for cases with parasites and fever during follow-up after day 3 and Late Parasitological Failures (LPF) for parasite infections with/without fever during the follow-up period. Cases which remained negative during follow-up were considered to be Adequate Clinical and Parasitological Responses (ACPR). These were modified WHO guidelines [The second-line treatment for uncomplicated idelines ,15.2 test. P < 0.05 was regarded significant.Data were entered into a computer database and SPSS software  was used for statistical analysis. The means  were calculated for all the patients and were compared between the patients in the two arms of the study using student t-test, when the data was normally distributed and by Mann-Whitney test if the data was not normally distributed. Percentages were calculated and compared for the patients by XThe study received ethical clearance from the Ethical board of the faculty of medicine, University of Khartoum.P. falciparum malaria. Seventy-three of them fulfilled the selection criteria and completed the 28-days follow-up; 39 and 34 in the fixed and the loose combinations of AS+SMP groups respectively. Eight patients, three in the fixed and five in the loose group were lost during the follow-up period because they changed their addresses, figure During the period of the study, eighty-one patients presented with uncomplicated Nine 12.3%) patients were children under five years of age; 5 (12.8%) vs. 4 (11.8%) of them in the fixed and the loose group of AS+MSP respectively. There was no significant difference in the presenting age, weight, temperature, parasite count and the haemoglobin level between the two groups, table 2.3% patiP > 0.05. However, these were mild and resolved spontaneously.Three (4.1%) patients (one in the fixed dose and two in the loose group of AS+SMP) in the study suffered drug-related adverse effects , P > 0.05 and one (2.6%) was parasitaemic in the fixed group of AS+SMP.On day 1; four (10.3%) vs. one (2.9%) patient was still febrile, On day 3; one patient in the fixed group of AS+SMP was still febrile but all the patients in both groups were aparasitaemic.P = 0.36) in the fixed and the loose group of AS+SMP respectively, showed late treatment and parasitological failures, the rest (89.7% vs. 97.1%) of the patients had an adequate treatment response.By day 28, four (10.3%) and one (2.9%) patient (The results of parasite genotyping indicated that four (one is the patient of loose and three patients in the fixed group of AS+MSP) of these five treatment failures were cases of recrudescence, where the parasites collected from each relapse parasitaemia appeared to be identical, in terms of their MSP1 & MSP2, to those collected pretreatment from the same patients. The fifth relapse in the fixed group (day 28) was caused by re-infection, where the parasite alleles were different.So PCR corrected cure rates were 92.3% and 97.1% (P > 0.05) for the two groups respectively.At the beginning of the study, gametocytaemia was detected in three patients (two in the fixed group), and it was not detected during the follow-up in any of the patientsP. falciparum malaria was reported in other African countries [This is probably the first publication on efficacy of AS+SMP in the treatment of uncomplicated falciparum malaria. The high cure rate demonstrated for AS+-SMP (>90%) in this study was expected because of the synergistic effect of artesunate and SMP and potentially because of the non-wide use of SMP as an antimalarial in this country. Previously high efficacy of SMP for the treatment of uncomplicated ountries -10.Although the loose combinations of AS+SMP was slightly (not significantly) more efficacious than the fixed one, the fixed regimen of AS+SMP is most promising since it permits reduction of the treatment duration to a maximum of 24 h. The mechanism of action of SM drug is the same as of sulfadoxine(S), but its protein binding and elimination half-life permit the new regimen to be developed in SM rather than in SP ,7,16. ThIn our study, the cure rates were 92.3% and 97.1 (P > 0.05) for both arms respectively. Treatment failures \u2013 even, AS+SP treatment failure \u2013 has recently been reported in the same area of the study . Fewer mThere was no gametocytaemia during the follow-up period. High levels of gametocytaemia had been reported in the eastern Sudan following non-artemisinin antimalarials , but it P. falciparum malaria in eastern Sudan. Due to its simplicity the fixed dose one-day treatment regimen may improve compliance and therefore may be the preferred choiceThis was a small-sized study, larger size studies are urgently needed to investigate the efficacy of this new formulated drugs combinations. Both regimens of AS+SMP were effective and safe for the treatment of uncomplicated IA, MM, MEO carried out the study and participated in the statistical analysis and procedures. IFK, MA participated in the genotyping of the parasite. IA coordinated and participated in the design of the study, statistical analysis and the drafting of the manuscript. All the authors read and approved the final version."}
+{"text": "Plasmodium falciparum malaria. Insufficient numbers of tablets and inadequate package inserts result in sub-optimal dosing and possible treatment failure. This study reports the case of three, non-immune, expatriate workers with P. falciparum acquired in Africa, who failed to respond to artemisinin-based therapy. Sub-therapeutic dosing in accordance with the manufacturers' recommendations was the probable cause.Artemisinin-containing therapies are highly effective against Manufacturers information and drug content included in twenty-five artemisinin-containing specialities were reviewed.A substantial number of manufacturers do not follow current WHO recommendations regarding treatment duration and doses.This study shows that drug packaging and their inserts should be improved. Plasmodium falciparum [In vivo resistance to this drug class has not yet been described [Artemisinin derivatives, such as artesunate, artemether and dihydroartemisinin, are the most parasiticidal of all the antimalarial drugs against lciparum . Their pescribed . Howeverescribed . If usedescribed -11. RecrP. falciparum malaria was recently diagnosed in three non-immune Europeans at the Geneva University Hospital. These three patients were initially treated in sub-Saharan Africa with artemisinin-based drugs. This motivated a thorough examination of package inserts of artemisinin-based drugs available in Africa. The potential association between treatment failure and inadequate information on drug leaflets are discussed in this study.Probable recrudescent P. falciparum-specific histidine-rich II protein. By inference, she was infected with P.falciparum either alone or mixed with another Plasmodium species. She self-prescribed oral artemether, obtaining the drug from the NGOs pharmacy. She did not recall the dose of artemether, but she reported full compliance with the five days course contained in the package. She improved on this treatment, but four days after stopping artemether, she developed fever again and had a positive malaria blood smear (species and parasitaemia unknown) consistent with a recrudescence of her primary infection. She was retreated with the same five days regimen with good effect. Some three and a half weeks later, she suffered a recurrence of fever associated with a positive malaria smear (species and parasitaemia unknown) suggesting either a recrudescence of her initial infection or a new infection. PCR genotyping was unavailable to differentiate her recurrent illness. She received artemether for seven days in combination with a single dose (three tablets) of sulfadoxine/pyrimethamine and had no further episodes of malaria while in Sudan. On review at the Geneva Hospital six weeks after her last febrile episode, she was well and afebrile.Patient 1 was a 27 year old Swiss female working in Sudan for an international NGO. She was not taking antimalarial chemoprophylaxis. She developed a febrile illness that was diagnosed locally as malaria by positive microscopic examination (species and parasitaemia unknown) and Paracheck\u2122, a rapid diagnostic test that detects P. falciparum was the likely diagnosis given the high prevalence of P. falciparum locally. However, a mixed infection cannot be formerly ruled out. She was treated at a local clinic with a single dose of intramuscular quinine in combination with the four doses regimen (four tablets \u00d7 4 over 48 hours) of artemether/lumefantrine (A/L); the latter is marketed as CoArtem\u00ae in malaria endemic countries. She made an uneventful recovery and then started daily chloroquine/proguanil (Savarine\u00ae) as prophylaxis. Twelve days after completing her A/L, she presented at Geneva Hospital with fever and had a positive malaria slide, with a P.falciparum asexual parasitaemia of 4,000/\u03bcL. She was treated successfully with three days of atovaquone/proguanil, remaining afebrile and aparasitaemic during 28 days of follow up.Patient 2 was a 35 years old Swiss woman working in rural C\u00f4te d'Ivoire for another international NGO. She was not on malaria prophylaxis. She developed fever and was found with a positive blood slide for malaria. Although the species was not specified, Plasmodium species but, considering the local malaria epidemiology, it was almost certainly P. falciparum. A local physician treated him with oral artesunate for three days and sulfadoxine/pyrimethamine, as a single dose of three tablets, with good effect. He did not recall the dose of artesunate but it was probably either 50 mg bid or 100 mg bid on the first day followed by 50 mg twice a day, consistent with the artesunate formulations likely to be available locally. However, 14 days after as the last dose of artesunate, he presented in Geneva with fever and his blood slide was positive for P.falciparum with an asexual parasitaemia of 190,000/\u03bcL, consistent with a probable recrudescence of his primary illness rather than a new infection (PCR unavailable). He was cured with the six doses regimen of artemether/lumefantrine.Patient 3 was a 30 years old Swiss man who was working in Guinea Conakry. He took weekly mefloquine as prophylaxis and reported good compliance. However, he developed an acute febrile illness that was diagnosed as malaria by blood smear. He was unaware of the falciparum malaria treatment failure after initial diagnosis and artemisinin-based treatment in Africa. It must be emphasized that the initial diagnosis could not be ascertained as malaria blood slides were not available for quality control. Considering the great variability in quality of diagnostic procedures in African laboratories, local diagnosis of malaria may not always be accurate.In summary, these three non-immune expatriate workers suffered of probable All patients reported having completed their treatment without episode of vomiting. The early timings of their recurrent malaria are consistent with recrudescence, but in the absence of PCR genotyping the possibility of early reinfections cannot be definitively excluded. Similarly, in the absence of defined plasmatic drug levels, none of these patients' recurrences meets the WHO definition of drug resistance . Sub-optSeveral artemisinin-containing therapies and their packaging were reviewed in order to assess the role of inadequate manufacturer information and insufficient drug content as possible causes of malaria treatment failure in sub-Saharan Africa.Packages of artemisinin-containing therapies currently sold in Africa were obtained from returning patients, WHO Roll Back Malaria, and the Internet. The list of twenty-five specialities presented here is not exhaustive Table , but canThe majority (10/17) of manufacturers of artemisinin monotherapies recommend five days of treatment instead of the WHO recommended seven days. Only three drug packages had paediatric dosing and none had specific recommendations for non-immune patients e.g. that 5 days of treatment is likely to be inadequate. Some packages contained less tablets than indicated in the package insert. The package inserts included in combination treatments were generally better, with doses both for children (7/8) and adults and with sufficient tablets to complete a therapeutic course. Of note was the use of the lower dose of mefloquine (15 mg/kg) rather than the higher dose (25 mg/kg) that is used in multidrug resistant areas of SE Asia.Sub-optimal antimalarial therapy results in increased morbidity and mortality of individuals living in malaria endemic areas, as well as travellers and expatriate workers ,14. WhilThis study shows that a substantial number of artemisinin-based drugs currently sold in Africa do not respect current WHO recommendations in term of treatment duration in case of monotherapy and many lack appropriate information regarding children dosage. Packages containing fewer pills than indicated in the manufacturers leaflet were also found. Concerning the use of low dose of mefloquine (15 mg/kg) in combination treatment, this may not be a problem in the short term in Africa where mefloquine as monotherapy or combined with artesunate has shown high efficacy in recent trials. Nevertheless, great care should be taken in monitoring cure rate, as this lower dose may potentially favour the development of mefloquine resistance -20.falciparum malaria'. Written information should avoid proposing different doses by trying to differentiate patients based on their presumed malaria immune status, as this may be confusing both for patients and prescribers. Similarly, packaging for combinations should be explicit, easy to use and contain the requisite number of tablets. Travellers should be aware that artemether/lumefantrine is known as Riamet\u00ae in temperate countries and CoArtem\u00ae in malaria-endemic countries. Adult Riamet\u00ae contains 24 tablets (six dose regimen) but adult CoArtem\u00ae contains only 16 tablets (4 dose regimen). Setting international standards on package inserts might be a useful step that could be championed by a relevant international body. The training of local drug sellers and community health workers in malaria endemic countries is another strategy that could be adopted.Travellers, NGO and other workers should be aware not only of the hazards of contracting malaria, but also how to obtain optimal treatment. This is especially relevant for those who may be located in remote rural areas. Drug manufacturers should also play their part in reducing the risks of treatment failure. It is suggested that all currently available artemisinin-based monotherapies should be packaged for adults and children with a complete therapeutic course of seven days of treatment, that package inserts should be improved e.g. warning that a full course of treatment must be taken even if patients feel well. In view of recent WHO recommendations, manufacturers should shift their production in favour of combined therapies rather than monotherapy to avoid the development of resistance for what is often called the 'last effective weapon against YJ: collected and analysed data and wrote the manuscriptFC: clinically managed the patients and co-supervised the study and the writing of the manuscriptLL: co-supervised the study and revised the manuscriptWT: collected data and co-supervised the writing of the manuscript"}
+{"text": "In the last five years, countries have been faced with changing their malaria treatment policy to an artemisinin-based combination therapy (ACT), many with no national data on which to base their decision. This is particularly true for a number of West African countries, including Guinea, where these studies were performed. Two studies were conducted in 2004/2005 in programmes supported by Medecins Sans Frontieres, when chloroquine was still national policy, but artesunate (AS)/sulphadoxine-pyrimethamine (SP) had been used in refugee camps for two years.In Dabola , 220 children aged 6\u201359 months with falciparum malaria were randomized to receive either AS/amodiaquine (AQ) or AS/SP. In vivo efficacy was assessed following the 2003 World Health Organization guidelines. In a refugee camp in Laine (south of Guinea), where an in vivo study was not feasible due to the unstable context, a molecular genotyping study in 160 patients assessed the prevalence of mutations in the dihydrofolate reductase (dhfr)  and dihydropteroate synthase (dhps)  genes of Plasmodium falciparum, which have been associated with resistance to pyrimethamine and sulphadoxine, respectively.In Dabola, after 28 days of follow-up, Polymerase Chain Reaction (PCR)-adjusted failure rates were 1.0% (95%CI 0\u20135.3) for AS/AQ and 1.0% (95%CI 0\u20135.5) for AS/SP. In the refugee camp in Laine, the molecular genotyping study found three dhfr mutations in 85.6% (95%CI 79.2\u201390.7) patients and quintuple dhfr/dhps mutations in 9.6% (95%CI 5.2\u201315.9).Both AS/AQ and AS/SP are highly efficacious in Dabola, whereas there is molecular evidence of established SP resistance in Laine. This supports the choice of the national programme of Guinea to adopt AS/AQ as first line antimalarial treatment. The results highlight the difficulties faced by control programmes, which have gone through the upheaval of implementing ACTs, but cannot predict how long their therapeutic life will be, especially in countries which have chosen drugs also available as monotherapies. In the last five years, countries have been faced with changing their malaria treatment policy to an artemisinin-based combination therapy (ACT), many with no national data on which to base their decision. This is particularly true for a number of West African countries; the Republic of Guinea is a good example. Indeed, when the National Malaria Control Programme (NMCP) of Guinea initiated its treatment policy change for treatment of non-severe malaria, replacing chloroquine (CQ) with an ACT, the only data available of antimalarial efficacy were results from two 14 day studies of CQ efficacy conducted in the South of Guinea (N'Zerekore) in 2000 and 2001, which reported failure rates of 24% and 28%, respectively . Nevertheless, high resistance to sulphadoxine-pyrimethamine (SP) and relatively high resistance to AQ were already documented in two neighbouring countries, as high as 52% and 46% for SP and 19% and 30% for AQ in Liberia and Sierra Leone, respectively [in vivo efficacy study which assessed the efficacy of two potential ACT candidates (AS/SP and AS/AQ) to replace the first line CQ treatment in Dabola prefecture  and the assessment of SP resistance from a genotypic analysis in the Lain\u00e9 Liberian refugee camp (South Guinea) where AS/SP was used since 2002 and where it was not possible due to security reasons to conduct an in vivo efficacy study  in two different regions of Guinea where the organization is supporting treatment of malaria are reported: an y Figure . The MinPlasmodium falciparum is the predominant species. Dabola reported 15,221 clinical malaria cases in 2003, accounting for 30% of the total outpatient consultations .Dabola prefecture  has an average altitude of 1,200 m and seasonal perennial malaria, with increased transmission between June and October. P. falciparum mono-infection and asexual parasitaemia between 2,000 and 200,000/\u03bcL were eligible for inclusion to the study. Exclusion criteria were (i) signs of severity or severe malaria according to WHO criteria, which included severe anaemia defined by haemoglobin < 5 g/dl [The efficacy study was based on the last WHO recommendations for the assessment and monitoring of antimalarial drug efficacy for area of high transmission . Childre< 5 g/dl .\u00ae, Roche, France); or amodiaquine 30 mg/Kg base divided into three daily doses of 10 mg/Kg ; each combined with artesunate at a daily dose of 4 mg/Kg on days 0, 1 and 2 . Drugs were crushed and mixed with water and sugar, given in a spoon or a syringe to small children. All doses were directly observed and repeated if vomiting occurred within 30 minutes. Patients were randomly allocated (blocks of 20), without concealment, to receive either AS/SP or AS/AQ.Study regimens consisted of either sulphadoxine-pyrimethamine 1.25-mg/Kg stat , children were re-assessed clinically and parasitologically on days 3, 7, 14, 21, and 28. Children, who were parasitaemic but asymptomatic during follow-up were asked to return to the clinic every three days. Gametocyte carriage was re-measured at day 28, and a second blood sample for PCR genotyping was collected in case of symptomatic recurrent parasitaemia occurring after day 9 , or at the end of follow-up in the presence of parasitaemia without symptoms (MSF/Epicentre guidelines). Rescue therapy (quinine hydrochloride 10 mg/Kg/8 hourly for seven days) was administered to treatment failures, orally or parenterally according to the patient's clinical condition.Children were withdrawn from the study in case of (i) vomiting any study dose twice, (ii) withdrawal of consent, (iii) onset of a serious febrile illness, (iv) intake of any drug with antimalarial properties, (v) missing any treatment dose, (vi) mixed species parasitaemia or (vii) any protocol violation. Patients who missed follow up visits and did not come on the successive day despite tracing were considered lost to follow up. Patients were classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) or adequate clinical and parasitological response (ACPR) as per WHO definitions .Capillary blood was obtained by fingerprick. Thick and thin films, prepared on the same slide, were stained with 10% Giemsa pH 7.2) for 15 minutes. Asexual parasitaemia was quantified against 200 to 500 leukocytes, assuming a white blood cell count of 8000/\u03bcL . Presenc.2 for 15\u00ae kits . Kits were air dried and stored in cool and dark boxes with desiccants. Genotypic analysis was performed at the Epicentre laboratory at Mbarara University (Uganda) according to a published method considering the three P. falciparum gene loci merozoite surface protein-1 (msp-1), merozoite surface protein-2 (msp-2), and glutamate rich protein (GLURP) [Blood samples for PCR analysis were collected on Isocode (GLURP) . Cases iAs no reliable estimates of antimalarial efficacy were available, all our calculations were based on an estimated 50% failure rate and a desired precision of 10%. Allowing for a type-1 error of 0.05 and considering a 15% drop out rate, 110 patients per study group were needed.\u00ae 11.0 for Windows  and EpiInfo 6.04b . Data entry errors were checked individually on each record. Appropriate statistical tests  over the total number of patients analysed. Losses to follow-up and withdrawn patients, as well as PCR-confirmed new infections and indeterminate PCR results were excluded from the analysis according to the last WHO guideline for assessment and monitoring of antimalarial efficacy [Records were entered in Epidata 3.0  and analysed on SPSSLain\u00e9 refugee camp  borders Ivory Cost and Liberia. Malaria transmission is perennial. A total of 10,541 malaria confirmed cases were recorded in 2004 . After DNA extraction, the PCR-Sequence Specific Oligonucleotide Probing (PCR-SSOP) method was used for molecular genotyping of point mutations in the dihydrofolate reductase (dhfr) and dyhydropteroate synthetase (dhps) genes [dhfr and dhps genes, which are fixed onto membranes and probed with SSOP designed to detect each of the single base-pair changes which code for substitutions at codons 51 (N to I), 59 (C to R), and 108  of dhfr and codons 436 , 437 (A to G), and 540 (K to E) of dhps. Since point mutations occur in a number of different configurations, each conferring differing degrees of resistance, data were to summarized in two ways. Firstly, by showing the allelic haplotypes (combination of mutations which are in the same gene), which were found among the parasites in Lain\u00e9. These can be determined easily in infections where there is a single genotype because the blood stage form of the parasite is haploid but are difficult or impossible to resolve in mixed infections [dhfr mutations plus the dhps 437 and 540 mutations. This particular genotype has been shown to be associated with SP treatment failure in a number of East African countries: Kenya, Malawi and Uganda but has only been reported rarely in West Africa [Patients from the outpatient clinic of the Lain\u00e9 refugee camp with measured fever (axillary temperature \u2265 37.5\u00b0C) were screened for falciparum malaria using the Paracheck-Pfs) genes ,10. Thist Africa -13.P. falciparum mono-infection and 220 (42.2%) were included  had a dhfr in 21 cases (11.6%) or of dhps in 45 cases (24.9%). One analysis was incomplete for the coding regions of dhps. Results were reported for 160 samples for dhfr mutations at all of three codons 108, 51 and 59, 135 samples for dhps mutations at codons 436, 437 and 540 and for 135 samples for combined dhfr-dhps mutations ; the proportion containing double mutant dhps  was 10.4% ; and the proportion containing all five was 9.6%  . In Dabola prefecture, based on the very good efficacy described here (above 95%) both AS/SP or AS/AQ are very good alternatives to CQ. Although important, these results provide information applicable only to the local area where the studies were conducted, and caution should be taken before extrapolating the results to the whole country. Site-specific patterns of health service utilization or malaria transmission intensity may result geographic differences in the efficacy of a drug [The f a drug .in vivo efficacy study was not feasible due to the unstable context related to security factors and where the combination AS/SP was already used since two years as first line treatment. Although, the genotypic findings cannot be correlated with any in vivo data from the area and that their predictive value for estimating how long the efficacy of SP used in combination with AS can be sustained is unknown, it was important to be able to document the presence and frequency of molecular markers of SP resistance in this setting. The frequency of the quintuple mutant dhfr-dhps genotype was quite high 7.3% . In Africa, the dhfr triple mutant genotype , which is a resistance marker of pyrimethamine has been shown to be strongly associated with resistance to SP, and this association is strengthened when the triple mutation is associated to the double dhps mutant , the quintuple mutation being the most predictive marker of treatment failure [in vivo studies in neighbouring Liberia and Sierra Leone [The results of the genotypic study in Lain\u00e9 are also important because they document the level of resistance to SP in a setting where day-28  failure . These rnd and 3rd trimesters. Therefore, considering the high levels of resistance to SP in neighbouring countries [In countries with no or very limited efficacy data of monotherapies (SP and AQ) or ACTs, such as in Guinea, choosing a standard ACT suitable for the whole country is difficult. The choice is usually based on a pragmatic decision, taking into consideration the potential risk of low efficacy based on data from neighbouring countries, such as Liberia and Sierra Leone in the Guinea case, availability and affordability of the ACT, which is still a limitation for artemether-lumefantrine and other advantages and disadvantages of the remaining ACT candidates such as dosing regimen and tolerance. The advantages of AS/SP and AS/AQ have been well described ,16. Theiountries  and the in vivo response. Genotypic studies are much simpler to perform than in vivo studies, particularly in countries in unstable situations. The predictive value of molecular results for estimating how long the useful therapeutic life of SP used in combination with artesunate should be further investigated.Concurrent in vivo and molecular studies of AS-SP are needed to see if there is a correlation between molecular findings and - MB designed the study, analysed and interpreted the data, wrote this paper- CR supervised the genotypic analysis, help to the interpretation of the molecular findings and revised the paper- MF implemented and coordinated the study laboratory procedures in Dabola and revised the paper- LC helped to design the study, revised the paper- GMG coordinated the study in the field, supervised all steps of the study including data entry and revised the paper- JPG supervised the analysis, the interpretation of data and revise the paper.All authors read and approved the final manuscript."}
+{"text": "P. falciparum infection in Sudan. However, there is limited information on the efficacy of ACTs in the country and only one report of PCR-corrected results for AS/SP only.A combination of artesunate (AS) plus sulphadoxine/pyrimethamine (SP) as first-line and artemether-lumefantrine (AL) as second-line treatment are currently recommended against uncomplicated msp-1 and msp-2 genes to differentiate recrudescence from reinfection.The WHO protocol for the assessment of antimalarial drug efficacy for the treatment of uncomplicated falciparum malaria was employed. Artesunate plus sulphadoxine/pyrimethamine (AS/SP) was compared to artemether-lumefantrine (AL) in a 28-day follow up. Samples that were classified as early treatment failure (ETF), late treatment failure (LCF) or late parasitological failure (LPF) were genotyped for A total of 178 patients were screened and 160 met the enrolment criteria and were recruited to the study of which 157 (98.1%) completed the follow up and had an analysed treatment outcome. On the AS/SP arm, three (0.038%) patients were lost during the follow-up, two on day 1 and one on day 7, and 77 (96.3) completed the study, while all 80 (100%) patients completed the follow up in the AL arm. In the per protocol analysis for AS/SP the treatment outcome for patients who completed the follow-up were as follows: adequate clinical and parasitological response (ACPR); 84.4% ETF; 1.3%, LCF; 3.9%, (LPF); 10.4%. For the AL arm the out come was as follows, ACPR; 90%, ETF; 0%, LCF; 6.3% and LPF; 3.8%. However, when PCR-corrected, 6.5% (5/77) of patients treated with AS/SP maintained parasites from their primary infection, while (7/80) in the AL group maintained their initial parasite genotype. Therefore, PCR-corrected efficacy was 93.5% in the AS/SP treated group and for AL it was 91.3%.Both AS/SP and AL are highly effective for the treatment of uncomplicated falciparum malaria in eastern Sudan. However, AS/SP appears to have a slightly higher efficacy than AL, this may be due to patient compliance with the repeated dose rather than drug efficacy. Plasmodium falciparum to chloroquine [Malaria remains one of the major public health problems in Africa. It is estimated that 7.5 to 10 million cases of malaria occur in Sudan every year [oroquine  and sulporoquine ,4, the NThe current treatment protocol for uncomplicated falciparum malaria in Sudan is artesunate plus sulphadoxine/pyrimethamine, as first-line and artemether-lumefantrine as second-line of treatment . There hA trial with 80 subjects in each arm would have at least 80% power (using a significance level of 0.10) to detect a difference between the two combinations if the failure rate detected after treatment is 20% in the AS/SP arm (based on the reported failure of SP of approximately 15 \u2013 20% in Sudan) and 5% in AL arm, allowing for 10% loss to follow-up.P. falciparum. Patients that satisfied the enrolment criteria according to the WHO protocol for the assessment and monitoring of antimalarial drug efficacy for the treatment of uncomplicated falciparum malaria [The study was carried out in three villages in eastern Sudan, southwest of Gedarif town . A community-based survey was conducted during the transmission seasons (October to December) of 2004 and 2005 to identify symptomatic patients that were microscopically diagnosed with mono infection of  malaria  were recFinger prick blood samples were collected and thick and thin smears were prepared on a patient's first attendance. When the patient was enrolled to the study, a finger prick sample was collected on days 0, 1, 2, 3, 7, 14, 21, and 28 on a glass slide and a parallel drop of blood was collected on filter paper. The filter paper was air-dried and stored in self-sealing plastic bags for molecular analysis.Thick and thin blood smears were stained with Giemsas' stain for ten minutes. The slides were read by two independent experienced microscopists. Parasite count was performed by counting asexual parasites against 200 white blood cells (WBC), and parasite density calculated assuming an average of 6,000 WBC in the population. Ten percent of the slides were read by a third microscopist in Khartoum for quality control. In case of disagreement the majority result was taken.Each patient was allocated randomly to either AS/SP or AL, treatment was administered under supervision for the AS/SP group, while only one dose of the AL was supervised and the second dose was given to the patient for self-administration. Patients were asked to return to the clinic on days 1, 2, 3, 7, 14, 21, and 28 or whenever they did not feel well. When a patient failed to come to the clinic on any scheduled day, a member of the team visited the home, gave them the dose and obtained the sample unless the patient was not found or refused to give the sample. Those who refused to give their sample (withdrawn) or missed more than one day of follow-up were excluded from the analysis.Patients, who were positive by microscopy and refused to participate in the study or did not meet the enrolment criteria, were provided with AS/SP for self-administration whereas pregnant women were referred to Gedarif Teaching Hospital as recommended by the national protocol.A per protocol analysis was performed employing the WHO excel sheet .Samples that were positive by microscopy on any of the follow-up days were classified as early treatment failure (ETF), late parasitological failure (LPF), or late clinical failure (LCF). Adequate clinical and parasitological response (ACPR) was defined as complete absence of microscopically detectable parasitaemia during the follow-up period, according to the protocol .msp-1 [msp-2 [Samples that were classified as ETF, LCF or LPF were genotyped by PCR as previously described for msp-1  and msp-1 [msp-2 ,14 on daThe study was reviewed and ethical clearance was obtained from the National Ethical Committee, Federal Ministry of Health, Sudan. Individual written informed consent was obtained from all participants or their legal guardians.178 patients were screened and 160 patients were enrolled and 157 completed the follow-up and had an analysable outcome. Of these, 77 patients were in the AS/SP arm and 80 were in the AL arm. Table A total of 77 patients completed the follow-up on the AS/SP group and 84.4% had an adequate clinical and parasitological response (ACPR). Of the 80 patients that received AL and completed the follow-up, 90% had successfully cleared their parasitaemia by day 28. Table P. falciparum on day 0 and who completed the follow up twenty patients had microscopically detectable parasitaemia on at least one day of the follow-up. Of these twelve received AS/SP and the remaining eight received AL. These samples were genotyped for msp-1 and msp-2 genes.Of the 157 patients diagnosed with In the AS/SP group, five patients (41.7 %) showed different allelic forms and were classified as reinfection, while seven (58.3) retained the same allelic pattern and were classified as recrudescence of parasites. Only one (1.25%) patient showed a different genotype in the AL group and the remaining 7 (87.5%) retained their initial parasite genotype. Hence, PCR-corrected efficacy was 93.5% and 91.3% for AS/SP and AL, respectively.Plasmodium development [Artemisinin derivatives are highly effective antimalarial compounds against different stages of elopment . In ordeelopment ,3,16.In Sudan, the current recommendation for uncomplicated malaria are AS/SP and AL as first and second-line treatment, respectively  with 100dhfr-108Asn, dhfr-59 and dhps-540Leu have rapidly increased in this area between 1993 to 1999 [In the present study, PCR-corrected results showed efficacy of AS/SP to be 93.5% in the Gedarif area. Although this is lower than the reports mentioned above, it is not surprising. Samples analysed in the present study were collected in 2004 and 2005, a time coinciding with increased use of SP mono-therapy due to the failure of CQ, which may have lead to increased levels of resistance to this drug. Although there is no recent report of SP clinical efficacy in our study area, alleles associated with SP failure have been reported to be increasing gradually. Triple mutants on  to 1999 . Data frThe partial failure of AL (8.9%) may be explained by inabsorption of lumefantrine due to low fat diet a situation that may be common in rural areas. In addition, recrudescence of AL-treated individuals has been reported in studies from other parts of the world -22.The present study has provided detailed PCR-corrected results of AS/SP and AL efficacy in eastern Sudan. Although the results may seem in disagreement with others, this is mainly due to the different transmission intensities in Sudan. In irrigated areas where there is perennial transmission such as Damazin, New Halfa, Malakal and Kosti Figure  there isArtemisinin-based combination therapies are considered to be highly effective antimalarials available in endemic countries. Artemisinin is highly effective in eliminating parasitaemia, however the partner drug is an important component in prolonging the usefulness of this combination. The results shown here call for continuous monitoring of the efficacy of ACT in eastern Sudan.EM carried out the sample collection, molecular analysis, data entry and contributed to drafting the manuscript, NG conceived of the study, contributed to its design, sample collection, data analyses and drafted the manuscript, SE contributed to the sample collection, microscopy and molecular analysis. IM and FM contributed to the sample collection and molecular analysis. AB participated in study design, data management and analyses. BE conceived of the study, participated in its design, analyses and helped to draft the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum to sulfadoxine-pyrimethamine (SP) across eastern and southern parts of Africa. This study assessed in vivo SP efficacy after two years of use as an interim first-line drug in Tanzania, and determined the rates of treatment failures obtained after 14 and 28 days of follow-up.Systematic surveillance for resistant malaria shows high level of resistance of P. falciparum  were treated with standard dose of SP. Treatment responses were classified according to the World Health Organization (WHO) definition as Adequate Clinical and Parasitological Response (ACPR), Early Treatment Failure (ETF), Late Clinical Failure (LCF) and Late Parasitological Failure (LPF) on day 14 and day 28.The study was conducted in the Ipinda, Mlimba and Mkuranga health facilities in Tanzania. Children aged 6\u201359 months presenting with raised temperature associated exclusively with Overall 196 (85.2%) of 230 patients had ACPR on day 14 but only 116 (50.9%) on day 28 (57.7% after excluding new infections by parasite genotyping). Altogether 21 (9.1%) and 13 (5.7%) of the 230 patients assessed up to day 14 and 39 (17.1%) and 55 (24.1%) of the 228 followed up to day 28 had clinical and parasitological failure, respectively.These findings indicate that SP has low therapeutic value in Tanzania. The recommendation of changing first line treatment to artemether + lumefantrine combination therapy from early next year is, therefore, highly justified. These findings further stress that, for long half-life drugs such as SP, establishment of cut-off points for policy change in high transmission areas should consider both clinical and parasitological responses beyond day 14. Others regard clearance of both symptoms and parasites over a much longer period as the most accurate measure of drug effectiveness [There is controversy over the therapeutic life of sulfadoxine-pyrimethamine (SP) when used alone for the treatment of uncomplicated malaria in Africa. Experts do not all agree on which drug efficacy measurements more accurately predict usefulness of a drug in a community. Some consider clearance of symptoms alone  or plus tiveness .th\u201315th August 2003, Harare, Zimbabwe).The assessment methodology has profound implications in terms of treatment policy strategies. Attempts have been made to define the cut-off points for changing first-line malaria treatment. Using the old treatment response classification criteria, the action period was due when a combined Early Treatment Failure (ETF) and Late Treatment Failure (LTF) were between 16 \u2013 24% . A couplPlasmodium falciparum resistance to SP across eastern and southern parts of Africa [in vivo SP efficacy after two years of widespread use in Tanzania.Systematic surveillance for efficacy of antimalarial drugs shows increasing levels of f Africa ,7. In 20P. falciparum parasitaemia between 1,000\u2013100,000 parasites per \u03bcl were recruited. Exclusion criteria and other procedures were as detailed in the protocol [\u00ae Roche), i.e 1.25 mg/kg of pyrimethamine and 25 mg/kg of sulfadoxine. The responses were classified according to the new WHO definition as ACPR, ETF, LCF and LPF at day 14 and day 28 [The study was conducted from July to November 2003 in the Ipinda (south-west), Mlimba (south-east) and Mkuranga (east) health facilities in Tanzania. Malaria transmission in these areas is perennial with peaks between May and July. A slightly modified WHO antimalarial drug efficacy testing protocol  was usedprotocol . Patientmsp2 locus to detect new infections. Extensive diversity in this locus has been observed with over 84 allelic variants in south-eastern Tanzania [msp2 alone may sufficiently discriminate recrudescence from reinfection in Tanzania. It has previously been shown that analysis of msp2 locus alone can effectively distinguish recrudescence from reinfection in Uganda [Treatment failures rates were corrected after genotyping the Tanzania ,10 and oTanzania . These on Uganda . The cliA total of 241 patients were recruited, of which 13 were lost to follow-up. Table The total clinical failure by day 14 and 28 was observed in 21 (9.1%) out of 230 and 39 (17.1%) out of 228 patients, respectively. 13 (5.7%) and 55 (24.1%) of the patients had LPF by day 14 and 28, respectively. Thus 34 (14.9%) out of 230 and 112 (49.1%) out of 228 patients had overall treatment failure by day 14 and 28, respectively. After genotyping recurrent infections day-28 treatment failures decreased to 85 (42.3%). In this study Mkuranga recorded the highest rates of both PCR-adjusted and unadjusted treatment failures followed by Mlimba and Ipinda.In 2001 Tanzania replaced chloroquine with SP as interim first-line antimalarial drug. Prior to this change, baseline clinical trials with SP had been conducted throughout the country using the 14 day follow-up protocol, and indicated an average efficacy of 86% on day 14. These findings paved the way for the malaria treatment policy change  and SP iRestricting our analysis to outcomes at day 14 would have led to misleadingly low clinical (9.1%) and parasitological (5.7%) treatment failure rates with the overall treatment failure (14.9%) being equal to that recorded at baseline prior to policy change. Using a shorter follow-up period, another study in parts of Tanzania also recorded an overall SP treatment failure of only 9.2% . AccordiOur observations show that SP efficacy in Tanzania is compromised and fully justify the recent decision to review the current malaria treatment policy from early next year in favour of artemether + lumefantrine combination therapy. This recommendation should be implemented at a large scale as soon as possible. Such a change would be welcome to protect the use of SP in its indication for the intermittent preventive treatment of pregnant women (IPTp). Indeed, it is at present the only drug that can be used for IPTp purpose because of its good safety profile. As far as methodology is concerned, the findings stress that cut-off points for malaria treatment policy change in high transmission areas should consider both clinical and parasitological responses beyond day 14, coupled with distinction of recrudescence from re-infection using molecular genotyping.B. Genton, H-P. Beck and K. Mugittu designed the study. S. Abdulla and K. Mugittu organised the clinical work. H. Mshinda was the supervisor in Tanzania. N. Falk, I. Felger, H-P. Beck performed molecular genotyping. H. Masanja carried out the statistical analysis. K. Mugittu and B. Genton wrote the article and all others contributed to it.The study was approved by the Tanzanian national and institutional ethics bodies"}
+{"text": "Plasmodium falciparum antimalarial drug resistance.Artemisinin-based combination therapy (ACT) is being widely promoted as a strategy to counteract the increase in A randomized, double-blind, placebo-controlled, clinical trial of the efficacy, effect on gametocytes and safety of the addition of artesunate/placebo (4 mg/kg/day \u00d7 3 d) to amodiaquine (10 mg/kg/day \u00d7 3 d) was conducted in Choco department, a low intensity transmission area in northwest Colombia.From 2,137 screened subjects, 85 entered the study: 43 in the amodiaquine plus placebo and 42 in the amodiaquine plus artesunate groups. Potentially eligible cases failed to qualify mostly because they were not available for follow-up visits (73%). Based on a per protocol analysis, the therapeutic response to both treatments was high: amodiaquine/placebo 35/36, 97.2% (95% CI 85.5\u201399.9), and amodiaquine/artesunate 32/32, 100% (89.1\u2013100) after PCR genotyping. The Kaplan-Meier survival estimates based on all eligible patients enrolled  were similar in the two study groups (P = 0.3). The addition of artesunate significantly decreased gametocyte carriage on Day 4 (OR = 0.1 95% CI 0.02\u20130.6), Day 7 (OR = 0.2 95%CI 0.04\u20130.9), Day 14 (OR = 0.09 95% CI 0\u20130.8), and Day 21 (OR95%CI 0\u20130.9). Most subjects in both groups (81% in amodiaquine/placebo and 75.6% in amodiaquine/artesunate) reported at least one drug related adverse event. Symptoms were generally mild and self-limiting and there was no serious adverse event. Two patients on amodiaquine/artesunate voluntarily withdrew from study because they could not tolerate the medication.Both drug regimens were effective in this area of Colombia. The addition of artesunate reduced gametocyte carriage and did not adversely affect tolerability. In this set of patients, the rate of adverse events was higher than in other studies. Patients' follow-up is problematic in areas with dispersed population and affects the conduct of clinical studies and monitoring of treatment effects. The results are discussed in the light of concurrent increase resistance to amodiaquine in other endemic areas in Colombia and the factors that may influence a change in the national antimalarial drug policy. Plasmodium falciparum malaria. Colombia accounted for 13.2%  of the 886,102 malaria cases reported in the Americas in 2004; of these, 44,437 (10.2%) were due to P. falciparum . Thi. Thi9]. ividuals ,19. One ividuals . However3) was the same as that used by Schellenberg et al, but neutropaenia was defined differently: neutrophils <2,000/mm3 versus <1,500/mm3 [Adverse events were frequent in both study groups but differences between placebo and AS were not detected Table . A meta-,500/mm3 . This st,500/mm3 . The hig,500/mm3 . ColombiP. falciparum and Plasmodium vivax), and logistics. Like Peru, Colombia may decide to have two different first-line regimens according to geographical areas, namely, one for the SP sensitive Pacific coast and another for the SP resistant Amazon region, or it could opt for an ACT that would have country-wide efficacy. On the one hand, the advantage of having two first line regimens would be to limit the use of second- and third-line drugs, like quinine combined with an antibiotic or with mefloquine  to areas of multidrug resistance. On the other hand, one potential disadvantage would be the need to identify the origin of infection before prescribing treatment, particularly in the major cities, where people from both the Amazon and Pacific Coast are seen and in areas of inter-regional migration.Besides AS plus AQ, the current WHO malaria treatment guidelines recommend three different ACTs: artemether-lumefantrine (six dose regimen), AS plus mefloquine and AS plus sulphadoxine/pyrimethamine (SP), that are being used in South America . This exBefore implementing ACTs in Colombia, it is important to highlight that, in spite of reports of therapeutic failures to both AQ and SP as monotherapies, the efficacy of their combined use  has been shown to be 97% or more in the Pacific Coast and northern Colombia . HoweverP. falciparum malaria in one endemic area in Colombia and reduced gametocyte carriage compared to AQ alone. However, the relative high frequency of adverse events attributable to AQ found and the reports of AQ resistance in other endemic areas of Colombia suggest that other ACT options should be considered for Colombia.AQ plus AS was efficacious in subjects with uncomplicated LO was the principal investigator and designed and managed the study, analysed the data, and wrote the first draft of the manuscript. IG coordinated the study and cowrote the first draft of the manuscript. PO and WRJT designed and coordinated the study and critically reviewed and suggested changes to the manuscript."}
+{"text": "Treatments for uncomplicated falciparum malaria should have high cure rates. The World Health Organization has recently set a target cure rate of 95% assessed at 28 days. The use of more effective drugs, with longer periods of patient follow-up, and parasite genotyping to distinguish recrudescence from reinfection raise issues related to the design and interpretation of antimalarial treatment trials in uncomplicated falciparum malaria which are discussed here.The importance of adequate follow-up is presented and the advantages and disadvantages of non-inferiority trials are discussed. The different methods of interpreting trial results are described, and the difficulties created by loss to follow-up and missing or indeterminate genotyping results are reviewed.To characterize cure rates adequately assessment of antimalarial drug efficacy in uncomplicated malaria requires a minimum of 28 days and as much as 63 days follow-up after starting treatment. The longer the duration of follow-up in community-based assessments, the greater is the risk that this will be incomplete, and in endemic areas, the greater is the probability of reinfection. Recrudescence can be distinguished from reinfection using PCR genotyping but there are commonly missing or indeterminate results. There is no consensus on how these data should be analysed, and so a variety of approaches have been employed. It is argued that the correct approach to analysing antimalarial drug efficacy assessments is survival analysis, and patients with missing or indeterminate PCR results should either be censored from the analysis, or if there are sufficient data, results should be adjusted based on the identified ratio of new infections to recrudescences at the time of recurrent parasitaemia. Where the estimated cure rates with currently recommended treatments exceed 95%, individual comparisons with new regimens should generally be designed as non-inferiority trials with sample sizes sufficient to determine adequate precision of cure rate estimates . For a patient ill with malaria, rapid resolution of illness without complications from the disease or its treatment is the first priority. Preventing return of the illness is a second priority. In a high transmission setting reinfection is inevitable, so the longer that subsequent illness can be delayed, the better. Those who deploy antimalarial drugs have similar objectives, but need to know more. In particular they need to know the efficacy of the individual treatment against the parasites which caused the infection. The key measure is the cure rate. The cure rate is defined as the proportion of treated patients whose symptoms resolve, parasitaemia becomes undetectable and in whom there are no recrudescences of infection with the genotypes which caused the original illness. This review discusses how the cure rate should be measured and reported.The treatment of malaria is changing for the better, but this has brought new challenges in the design and interpretation of efficacy assessments. In the past few years, it has become accepted that antimalarial treatments must have high cure rates, which ideally should exceed 90% . The cor1/2 > 1 week e.g. mefloquine, piperaquine) and a 28-day follow-up is the minimum for rapidly eliminated antimalarial drugs [1/2 is 1 day to 1 week) 42 days follow-up captures most of the recrudescences. WHO now recommends a minimum of 28 days follow-up in antimalarial drug in-vivo studies. [The efficacy of antimalarial drug treatment, in uncomplicated falciparum malaria is assessed by following patients after observed treatment for sufficient time to \"capture\" all or most of the treatment failures (failures to cure the infection leading to recrudescences) that could occur. As recrudescences result from persistent erythrocytic infection, these recurrent infections re-emerge within a defined time period following treatment ,4. This al drugs . For drustudies. . As incrPlasmodium vivax and Plasmodium ovale infections, a relapse). The use of PCR genotyping of Plasmodium falciparum has considerably improved the ability to conduct community-based clinical trials of antimalarial drugs in endemic areas [P. vivax malaria is more difficult as recurrence of the infection may result from recrudescence, reinfection or relapse. The relapses derive from persistent hypnozoites in the liver and are commonly with different genotypes to that identified in the acute infection [P. falciparum studies blood samples are taken, usually on filter paper, and the genotypes are compared in blood samples from the acute infection and any infection which recurs during the period of follow-up. P. falciparum genotyping is usually based on comparison of variable blocks within the polymorphic genes MSP1, MSP2, and also often GLURP, or by use of microsatellite typing. Sometimes blood samples from the recurrent infection or the acute infection are not available, or go missing, or there are technical reasons while the PCR comparison cannot take place. In higher transmission settings a genotype often cannot be ascribed confidently in multiple infections . How should these patients then be evaluated? There is no clear consensus \u2013 but calling all missing or indeterminate results treatment failures is clearly illogical. Ideally in a study, all participants in the trial should complete the study, follow the protocol, and provide data on all the outcomes of interest at all time-points. In reality, most trials have missing data. Data can be missing either because critical information, such as PCR genotyping, is missing or uninterpretable [Conducting large trials with extended patient follow-up is logistically demanding. In endemic areas it is not possible clinically to distinguish a recrudescence from a newly acquired infection  analysis. But in most trials there are patients who do not complete the follow-up period, yet these patients do contribute useful information before they leave the trial, and this can and should be used. If such a patient did not fail (i.e. remained aparasitaemic) when last observed, that patient's data are said to be 'censored' at the time they were last followed up. The appropriate analysis for such data is survival analysis. This analytical approach is well established in the assessment of cancer chemotherapy and increasingly in the assessment of anti-infective drugs, as survival analysis deals explicitly with censored values. Patients with different follow-up periods cannot be treated the same way \u2013 someone who is followed up for longer has a greater chance of being recorded as treatment failure than another patient followed up for a shorter time. Failure rates should be estimated using the Kaplan-Meier method [In antimalarial drug trials there are two or more groups of patients followed for a prespecified length of time after different antimalarial treatments. The cure rates, which means the proportions of patients who reach the end of this follow-up period without recrudescence of the infection are compared. In the past, antimalarial treatment efficacy was usually assessed on a particular day (often day 14 or day 28 after starting treatment) so only patients followed to that day were included in the analysis. This is often referred as a r method . This isr method .t who have not developed a recrudescence is estimated by the Kaplan-Meier method asThe proportion of subjects beyond any follow-up time i-di)/rip = \u220f , an additional four failures were recorded at day 63, and 19 patients were lost to follow-up between days 28 and 63.0.21.Estimation of the failure rate at day 28 is easy and equals 21/100 = What is the day 63 failure rate?If all patients had been followed up until day 63 then the failure rate would have been 0.25 (25/100), but it was not the case.In total 25 failures were observed but patients were followed up for different periods of time. An analysis done only on patients who completed the follow-up would have a denominator of only 60 as the early failures (N = 21) and the patients lost to follow-up (N = 19) are not included. It is obviously wrong to ignore these early failures, but if they are included in the analysis it assumes that patients excluded (i.e. the 19 lost to follow-up) had the same probability of failing during the entire follow-up period as those included, whereas those lost to follow-up did not fail in the period between 0 and 28 days (when 21 of the 25 observed failures did occur). The failure rate would be estimated as:25/(60 + 21) = 25/81 = 0.31 \u00a0\u00a0\u00a0 (A)The correct analysis is based on the Kaplan-Meier approach which incorporates these temporal changes in the probability of failing treatment.0.79.In the example, the failure rate at 28 days is 21/100, so the probability of not failing during 28 days is 1-(21/100) or 0.933.The 60 patients who did not fail until day 28 were followed up until day 63, and for them the probability of failing between day 28 and day 63 is 4/60 or 0.0667. Thus, the probability of not failing between days 28 and 63 is 1-0.0667 or Of course patients can only not fail in the second interval (28\u201363 days) if they did not fail in the first interval (0\u201328 days). Therefore, the probability of not failing during 63 days is equal to the product of the probability of not failing during 0 to 28 days and the probability of not failing between 29 and 63 days. This product is the treatment success rate and the failure rate is one minus the success rate.So the failure rate is estimated as :1-(1\u201321/100)*(1\u20134/60) = 0.27 \u00a0\u00a0\u00a0 (B)This estimate is significantly lower than the per-protocol estimate. Splitting follow-up into intervals and calculating failure rates for these intervals makes sense biologically. Recrudescences do not have the same probability of occurring across the entire period of follow-up.Figure et al have recently reported a comparison of per protocol and survival analysis [P. vivax are common, a very high proportion of patients may have a recurrent infection within the follow-up period and be lost to a PP analysis [For all three distributions of observed failures, the PP rate estimate is the same, but the KM estimate varies. The discrepancies between the two methods are greatest when most of the observed failures occurred before patients were lost to follow-up. Discrepancies also increase dramatically with the proportion of patients lost to follow-up. Guthmann analysis  in 13 paanalysis .i will be equal to the total number of patients when there are no censored observations, that is when all patients were followed until the recurrence or time ti. When there are censored observations then the effective sample size at time ti isStatistical software used for the calculation of Kaplan-Meier estimates will usually also provide confidence intervals; these will be Greenwood's confidence intervals or confidence intervals based on the asymptotic variance of -log-log transformation of the survival function . Howeveri-di)/p;N' = where \u00e1 is a significance level and N' is the effective sample size. An Excel spreadsheet for calculating the confidence intervals for proportions and their differences is freely available on the web .The simplest way of comparing and presenting the failure rates and failure times between the treatment groups is to plot the Kaplan-Meier survival estimates on the same axes. To distinguish between chance variation in failure rate estimates in the two groups and a real difference, a hypothesis test is required. For survival data, two tests are commonly used: the log-rank test and the Wilcoxon test. Both test the null hypothesis that there are no real differences between the two groups, so small values of the test statistics will correspond to the acceptance of the null hypothesis. Both are based on the sum of the differences between the observed and expected number of failures in each group over all time points, but in the Wilcoxon test this sum is weighted by the total number of individuals at risk at each time. The log-rank test is preferable when the Kaplan-Meier plots for the two groups do not cross, as this reflects a continuous proportional difference in failure rates between the two groups. If the survival curves do cross, the Wilcoxon test should be used.It is always useful to express the difference between the two treatment groups in a summary measure. As the risk of failure changes over time and the rates of change are usually different in the two treatment groups, only measures evaluated at a prespecified timepoint are recommended. These could be the absolute risk reduction or the risk ratio (or relative risk). Hazard ratios , which aThe absolute risk reduction is the difference between failure rates in the two treatment groups. The risk ratio (relative risk) is calculated as the ratio of those failure rates. In antimalarial trials, because of losses to follow-up, Kaplan Meier estimates of failure rates should always be used. The absolute risk reduction assesses the clinical importance of the treatment difference while the relative risk has the intuitive appeal as it measures the magnitude of the difference.However, there are pitfalls in using the relative risks \u2013 for uncommon events, large relative risks will result from differences of only a few failures between the treatments, for example 2% difference in failure rate between 2 treatments of efficacy rates of 97% and 99% gives relative risk of 3.In the past antimalarial drug trials have been powered to detect differences between drugs \u2013 usually with 95% confidence and 80% power. This is increasingly difficult at cure rates over 90% because of the exponential increase in the sample size required Figure . The higis a difference between the two groups  and it is greater than the \u03b4. Rejection of the null hypothesis indicates that there is no difference between the groups. The choice of an appropriate value for \u03b4 is a compromise based on current knowledge, clinical judgement, likely policy implications and the practicalities of conducting large trials. Values of 10% have been widely used in assessing antimicrobial agents, but are too wide for current requirements for antimalarial drug efficacy. Smaller equivalence margins require that studies have larger sample sizes that have been usual in the past [Non-inferiority trials aim to show that an experimental treatment is not worse than the active control (i.e. current treatment) by more than a specified amount \u2013 the equivalence margin (often denoted \u03b4). The null hypothesis being tested is that there the past . Each cothe past ,5. Therethe past -24. The the past . This emBlinding is often used to avoid bias in comparative trials although it is often difficult in antimalarial drug assessments because of differences in treatment regimens and the difficulties in masking the taste of the drugs. Compared with superiority trials, blinding does not protect against bias as well in non-inferiority trials because a biased investigator wishing to show non-inferiority can simply give all patients similar results! Analysis of non-inferiority trials requires a calculation of the difference between the failures rates in the treatment groups and a calculation of the confidence interval around this difference using appropriate methods  and 'effIntention-to-treat (ITT) analysis intends to include all patients randomized into the trial irrespective of what happened to them subsequently ,28. ThisIn the per protocol analysis (PP), on the other hand, drug trial results are often analysed simply in categorical tables comparing proportions of patients deemed to have been cured at the predefined trial time-point(s). As explained previously this ignores the contribution of information provided by patients whose follow-up was incomplete.The intention-to-treat analysis (ITT) analysis is widely used to generate the most conservative estimate of the efficacy of an antimalarial drug. In an ITT analysis all patients who do not complete the follow-up period successfully may be considered therapeutic failures. The results then provide a combined assessment of the trial conduct and the drug efficacies, with efficacy and follow-up failures both having equal weight in the analysis. In antimalarial drug trials with long follow-up drop-outs increase with time. Thus, even if a drug has 100% true efficacy, an ITT analysis will show progressively worse results as the duration of follow-up is extended. It is simply measuring loss to follow-up and indeterminate or missing genotyping results. Slowly eliminated drugs which require longer follow-up will always fare worse. The better the drug is (ie. the higher the true cure rate), the greater is the proportional difference between true cure rate and ITT estimate. This is illustrated in Figure Some patients miss a follow-up appointment and then attend subsequently. These patients should not be censored from efficacy assessments if they have remained well, as it is unlikely that a recrudescent malaria infection would have occurred during the missed appointment period which then rapidly and symptomlessly self-cured. If the patient presents after the missing appointment with a recrudescence then the history can be used to estimate the onset of recrudescence. Continuous data (e.g. gametocyte-time curves) present more of a problem, and these need to be assessed on an individual basis. There may be sufficient data to justify interpolation, or omission may be appropriate.The use of genotyping to distinguish reinfection from recrudescence is based on the relative probabilities of finding identical polymorphic malaria parasite alleles by chance from the parasite population. It is necessary to define these probabilities for each individual allele for the parasite population (and thus patient population) under study. For multiple alleles their individual distributions must be unlinked. If there are multiple genotypes present then qualitative assessments (without extensive sequencing) cannot ascribe an individual genotype, and although probabilities can still be ascribed to recurrent infections, statistical power is reduced. If a patient has a recurrent infection in an antimalarial drug trial conducted in an endemic area, but a paired sample is either unavailable or, for technical reasons , a comparison of genotypes cannot be made then a recrudescence (treatment failure) cannot be distinguished from a newly acquired infection. The interpretation of genotyping results in high transmission settings, and the possibility that minority (undetected) genotype populations may cause subsequent recrudescence is a subject of considerable interest and debate. This is important subject and advances both from a biological and statistical perspective can be expected in the near future. But for the purpose of this discussion it is simply accepted that a potentially large number of patients may have indeterminate results, and that there is uncertainty as to how these patients should be analysed.To present the different analytical approaches, first it is necessary to describe the following proportions:i, ti+1, etc) there are ni, ni+1, etc. patients at risk of having a recurrence of parasitaemia.At start of each interval  andci patients who were lost to follow-updtheni = ri + ai + ci + di........................................................................................................................................... (1)ni+1 = ni - (ri+ ai +di) as those patients with true recrudescences previously are no longer \"at risk\" (i.e. they cannot have another recrudescence), and both dropouts and those with new genotype infections are not followed up further (the latter group having been treated).Then ni) for whom genotyping results are available will be either recrudescences, or new infections, but for some of them the PCR results will be indeterminate and so their status will not be known. If indeterminate PCR genotyping results (ind) are unrelated to treatment failure rates, and are not more or less likely in recrudescences than in newly-acquired infections, then they will occur at a constant rate (i.e. a constant proportion (f) of the total recurrences with time):The observed recurrences (ori = f\u00b7(ri + ai)).....................................................................(2)indi = f\u00b7\u2211 (ri + ai)...........................................(3)and subsequently \u2211 indi confirmed recrudescences and (1-f) ai confirmed new infections and so as ori = ri + aiIn this notation, there are (1-f) ri = (1-f) (ri + ai) + indi .......................................................(4).then ori and indi are small so estimates of f may be inaccurate, and it may be better estimated from the equation (3) using the total number of indeterminate results and the total number of observed recrudescences.At each time interval, numbers ri + ai) indeterminate results. So how should indeterminate results be treated in the analysis of efficacy?The ratio of recrudescences to new infections will depend on the failure rate (which for new treatments should be less than 10%), the malaria transmission intensity, and time since the start of the treatment. In high-transmission settings, eventually all patients will be reinfected. This and the non-linear relationship between recrudescences and reinfections (reflecting the relationship between a discrete and a cumulative distribution) are illustrated in an example which follows and in Figures In this approach all indeterminate PCR results are treated as recrudescences (i.e. treatment failures).i to ti+1) is:i.e. the failure rate for interval (tA = ((1-f)ri + indi)/(ni)FUnless there are no reinfections this method overestimates the failure rate. This overestimation  is not so profound at high failure rates or at low levels of malaria transmission, but, unless rates of indeterminate results are low, it creates serious discrepancies at low failure rates in areas of medium to high transmission. At its most extreme, if all patients are followed until they acquire their next infection, then for a drug with 100% efficacy (no true failures) the ITT failure rate equals the rate of indeterminate PCR results. As this could well exceed 10% then even a completely (100%) efficacious antimalarial drug cannot achieve the level of >90% cure rate currently recommended by the World Health Organisation in the trial [i+1.In this approach no assumptions are made, and the patients are simply censored from the analysis. They can be censored at the time when the recurrent parasitaemia occurred, that is at the end of the interval, at time tB = (1-f) ri/niFPatients with the indeterminate results are treated in exactly the same way as patients who became smear negative after treatment and were then lost to follow-up.i at the beginning of the interval.But as the PCR result is not known, and therefore whether it is a recrudescence or reinfection cannot be determined, the standard approach in survival analysis to the patient's data would be to exclude them from the analysis in the interval when the recurrence took place, so they have also to be deducted from the number at risk nC = (1-f) ri/(ni-indi)Fi.This corresponds to censoring them at the end of the previous interval, at time tIt could be argued that this type of censoring is not non-informative but if the number of patients lost to follow-up is small, very little bias is likely to result from applying methods based on non-informative censoring .i) where gi is the proportion of recurrent infections at time ti which are recrudescences. Thus at each time point for a recurrent parasitaemia this probability of recrudescence (gi) and a probability of reinfection (1-gi) are determined for the study population from the valid PCR-genotyping results. Obviously this requires sufficient data for adequate characterization. This ratio of probabilities is then applied to any indeterminate results.This approach uses all the available data but relies on there being sufficient data to characterize the temporal changes in the probability of recurrent parasitaemia being a recrudescence (gD = ori gi/ni = (ori\u00b7(1-f) ri/(ori - indi))/ni = FB/(1-f)Fi = (1-f) ri/(ori - indi) is the proportion of recurrences with confirmed PCR results which are recrudecences at time ti.where gD is the most accurate provided we have a good estimate of g.Estimate FC is methodologically correct but its includes patients with indeterminate results only while they did not have recurrent infection. There is currently no consensus on which approach should be taken, or the precise modelling approach to the calculation of g.Estimate FThe assessment of efficacy of an antimalarial treatment is based on the observed cure rate and the confidence intervals around the estimate. In a comparison of two treatments we calculate the individual confidence intervals around the cure rates observed, then the difference between the cure rates, and then the confidence interval for the difference. The sample size determines the width of these confidence intervals. Confidence interval calculations based on the normal approximation (for estimation of the standard error) are not appropriate for very small proportions (i.e. very low failure rates). The methods of Wilson ,18 and NUpper limits for 95% confidence intervals using Wilson's method are presented in Figure The above calculations are based on the assumption that there are no losses to follow-up. But if there are losses, the 'effective' sample size of Peto , which iUnequal randomization, for example 2:1 randomization, may also be considered in trials assessing new treatments. Although unequal randomization sacrifices statistical power slightly  in the comparison, it increases the precision in the estimate of treatment efficacy of the new drug, and it also provides a better adverse-effect characterization for the new treatment. The \"control\" (i.e. current treatment) trial arm is still important in helping to distinguish \"trial\" from \"drug\" problems i.e. it helps to identify systematic trial-related confounders which lead to unusual efficacy or toxicity findings. For example, a trial in which non-inferiority was shown but both treatment arms performed poorly (i.e. less than 90% cure rates) would not warrant rejection of the new treatment if the prior information suggested much better efficacy of the established treatment. There might have been a problem in the conduct of the trial. Thus there is a statistical trade-off between both the characterization of the difference between the two regimens and the precision of the \"control\" arm estimate, and the characterization of the new treatment effects. But there are also biological and programmatic reasons why it may be important to have a larger sample size for the new antimalarial treatment; as cure rates asymptotically approach 100% the selective force that drives the emergence and spread of resistance weakens, and in low transmission areas, provided coverage is high, the incidence of malaria will fall. Precise characterization of very high cure-rates provides important information to the policy maker which will be taken into account with costs, simplicity of administration, tolerability, adverse effect profile, etc., to influence the difficult decision of whether or not to change treatment recommendations.It is evident from Figures If the existing recommended antimalarial treatment used in comparative assessments of new antimalarials is still highly efficacious then sample sizes in conventional superiority trials must be large. For example, if the true cure rate for a new antimalarial treatment exceeds 99%, then the total sample size required to show superiority over the existing treatment, if this has an efficacy of 95%, exceeds 650. This doubles to over 1,300 if the new treatment is 98% efficacious. A 2:1 randomization increases these numbers by <8%. Few single centre antimalarial drug trials enrol more than 650 patients in a study. This seems to be the limit for superiority testing. If a non-inferiority trial is conducted, and the differences are much larger than anticipated, then a significant difference may be demonstrated. If the new compound has a significantly lower cure rate this would argue against its introduction. For the established compound more investigation might be necessary to determine the reasons why it performed less well than expected.The outlook for antimalarial chemotherapy has improved in recent years with the introduction of several new highly effective drug combinations, and elevation in the cure rates now required of these new treatments. A cure rate of at least 90% in uncomplicated malaria assessed at 28 days is recommended by the World Health Organization . As a reThere has been a considerable improvement in the quality and quantity of antimalarial drug trials reported in recent years. The introduction of PCR genotyping  has alloWith the increasing efficacy of new treatments and requirement to aim for cure rates of > 90% and preferably \u2265 95%, comparative trials should generally be designed as non-inferiority trials in which the null hypothesis is that there is a difference between the two groups when existing treatment efficacy still exceeds 95%. These trials should be powered to give a predefined precision for estimates of cure rates. For consideration as a policy option the point estimate of the cure rate of the new drug treatment, and the currently recommended treatment should both exceed 90% as this is the threshold currently recommended by WHO. Large trials are required to provide adequate precision of these estimates.Antimalarial drug comparisons must be large enough to provide precise cure-rate estimates, and follow-up must be long enough to capture the majority of recrudescences. Non-inferiority trials may be necessary when standard treatment efficacy is high (cure rates over 90%), but these have weaknesses which may not be familiar to investigators used to superiority trials. The primary efficacy endpoint should be derived from survival analysis. Intention to treat and per protocol point-analyses should be reported also as secondary results. The interpretation of current PCR gel-electrophoresis derived genotyping results in high transmission settings is difficult. Indeterminate or missing results should not be classified as treatment failures, but should also analysed using a survival approach. Consensus recommendations on the interpretation and analysis of antimalarial drug trials would be of great benefit.\u03b1 \u2013 significance level\u03b4 \u2013 equivalence margin\u220f \u2013 multiplication symbol\u2211 \u2013 summation symbolA \u2013 cumulative probability of developing a patent new infectioni \u2013 patients with newly acquired patent infections at time tiai \u2013 patients without patent infections at time ticCI \u2013 confidence intervali \u2013 patients were lost to follow-up at time tidF \u2013 true failure ratef \u2013 proportion of the total recurrences which have an indeterminant PCR result ITT \u2013 failure rate estimated by intention-to-treat approachFP. falciparum glutamate rich proteinGLURP i \u2013 proportion of the total recurrences which are true recrudescences at time tigi \u2013 number of indeterminate PCR results at time tiindITT \u2013 Intention-to-treat analysisKM \u2013 Kaplan-Meier survival analysisP. falciparum merozoite surface protein 1MSP1 P. falciparum merozoite surface protein 2MSP2 N \u2013 total number of patientsN' \u2013 adjusted total number of patients (effective sample size)i number of patients observed at time tini \u2013 number of observed recurrences at time tiorPCR polymerase chain reactionp \u2013 proportion of subjects beyond any follow-up time t who have not developed a recrudescencePP \u2013 \"per protocol\" analysisR \u2013 cumulative probability of developing a patent recrudescencei \u2013 patients with recrudescent infections at time tir1/2 \u2013 antimalarial drug elimination half lifetThis was written jointly by the two authorsA priori a margin of clinical non-inferiority (\u03b4) had to be selected, which is defined as the largest reduction in efficacy which would be clinically acceptable.In a study, patients were randomly assigned treatment A or B and were followed up for 63 days. The aim of the study was to show that the new treatment (A) is not less effective than the standard treatment (B). As we are interested in non-inferiority of treatment A, the 95% CI for the difference in efficacy (A-B) should be less than the value \u2013 \u03b4.In this example a \u03b4 of 0.1 (i.e. 10%) was selected. There were 100 patients in each group. For those who were lost to follow-up or had a recurrence of infection we list the length of follow-up completed in the table below. Stars denote loss to follow-up. In treatment group A six failures were observed and six patients were lost to follow-up, while in group B four failures were observed and three patient were lost to follow-up.A; 14* 14* 22  22 * 28* 28* 28* 34 44 52 61 63B; 14* 14* 17  24   28*  29   43Using the Kaplan-Meier method we estimate the efficacy as 0.94 (0.86 to 0.97) for treatment A and 0.96 (0.89 to 0.98) for treatment B.A = 88/0.94 = 94 and nB = 93/0.96 = 97. Using Newcombe's formula we obtain a 95% CI for the difference of (-0.09 to 0.05).The difference in efficacy (absolute risk reduction) A-B is -0.02. To calculate the confidence interval around this difference we need to calculate the 'effective' sample sizes: nAs the confidence interval is more positive than the \u03b4 of \u2013 0.1 we conclude that treatment A is not inferior to treatment B. But it should be noted that even with 100 patients per group the trial is underpowered; the precision of the cure rate estimates is poor  \u2013 see Figure"}
+{"text": "An artemisinin-based combination therapy, artesunate (AS) plus sulphadoxine-pyrimethamine (SP), was compared to SP monotherapy to provide evidence of further treatment options in southern Mozambique.Plasmodium falciparum malaria were randomly allocated SP (25/1.25 mg per kg day 0) or AS/SP . Allocation was concealed, but treatment was open-label except to microscopists. The primary objective was the relative risk of treatment failure, which was assessed using World Health Organization response definitions modified to a 42-day follow-up.Between 2003 and 2005, 411 patients over one year and 10 kg with uncomplicated dhfr/dhps mutation increased the relative hazard of failure compared to fewer mutations  and baseline axillary temperature increased the relative hazard of failure by 50% for each \u00b0C increase .Of the 411 subjects enrolled, 359 (87.3%) completed the follow up period . A survival analysis including 408 subjects showed that the polymerase chain reaction-adjusted cure rates were 90.4%  and 98.0% (95% CI 94.8%\u201399.3%) for SP and AS/SP respectively. Multivariable analysis showed that treatment with AS/SP decreased the relative hazard of treatment failure by 80% compared to SP  and age over seven years decreased the relative hazard of failure by 70% , when compared to younger age. However, having a quintuple While both treatments were efficacious, AS plus SP significantly decreased the relative hazard of treatment failure compared to SP monotherapy Artesunate plus sulphadoxine-pyrimethamine, but not sulphadoxine-pyrimethamine monotherapy, met the current WHO criteria of >95% efficacy for policy implementation.NCT00203736 and NCT00203814 Malaria is responsible for a large health burden, including an estimated 19% of deaths among children under five years of age [Plasmodium falciparum malaria varies throughout the country according to the season, other environmental factors, and the use of vector control measures. A treatment policy with effective anti-malarials is a key component of malaria control programmes and data regarding resistance of parasites to anti-malarial drugs are now a critical factor in drug policy decision-making. While chloroquine was the national policy in Mozambique for the treatment of uncomplicated malaria as recently as 2003, evidence of its poor efficacy throughout Africa since the 1980s signalled a need for alternative anti-malarials [falciparum malaria provided the partner drug is efficacious. ACT deliverers a more rapid cure compared to non-ACT, reducing gametocyte carriage , and delaying the development of anti-malarial resistance [Mozambique  evaluation in Namaacha and Matatuine Districts, Maputo Province, during 2002 and data showed that a combined polymerase chain reaction (PCR)-adjusted cure rate exceeded the 80% level considered for it to be combined with other anti-malarials to sustain its useful therapeutic life [dhfr) and dihydropteroate synthetase (dhps), mutation across all study sites was 5\u20136% throughout the study period [[There was clear evidence that ACTs substantially reduced treatment failure, recrudescence and gametocyte carriage but this had not been reported for Mozambique . Howevertic life ,9. The p period [, Raman JThis multi-centre, open-label, parallel-group RCT was conducted in Maputo Province, Southern Mozambique in four public-sector health facilities in two phases: initially Catuane and Namaacha (2003), thereafter Boane and Magude (2004\u20132005) Figure . There hP. falciparum infection by rapid diagnostic test . Patients diagnosed with P. falciparum parasitaemia up to 500,000 asexual parasites/\u03bcml blood (on thick Giemsa-stained smear) and with axillary temperature greater than or equal to 37.5\u00b0C (or history of fever within the previous 24 hours) were suitable for inclusion. Patients receiving folate or anti-malarials  within seven days were excluded. Also excluded were patients with danger signs  or severely ill, those with a history of G6PD deficiency or allergy to the study (or related) drugs, or those with serious underlying disease [Male and non-pregnant, non-breastfeeding female patients with malaria symptoms, that were older than one year of age and weighed more than 10 kg, were invited to give informed consent prior to screening for  disease ,13.\u00ae. Roche, Gauteng, South Africa) or AS/SP  (Table Eligible subjects were randomized to treatment with SP monotherapy (Fansidara) Table . Subjectin vivo therapeutic efficacy of anti-malarials, with extension to a 42 day follow-up due to the long elimination half-life of SP [\u00ae), clinical signs and symptoms to capture adverse events, and concomitant medications [The visit schedule was based on the WHO protocol for assessment of fe of SP ,15. Afteications . A fingeP. falciparum DNA extracted from these dried samples based on variations in three highly variable proteins  was used to determine if treatment failure was due to a re-infection or recrudescence of the original infection [dhfr  and dihydropteroate synthetase, dhps  genes were detected using nested PCR and restriction endonuclease cleavage [Asexual parasite density was calculated using the number of asexual parasites and assuming 8,000 leukocytes/\u03bcl blood. Genotyping of nfection ,18. Infecleavage . DigestiThe primary study objective was to compare the risk of treatment failure between the treatment groups, adjusted for baseline characteristics. A secondary outcome compared the rate of parasite clearance between groups.Response to treatment was classified according to WHO definitions for low to moderate malaria transmission intensity areas, modified for a 42-day follow up ,15. SubjKey fields were verified by a study monitor and double-entered into an MS Access 2000  database. Laboratory data were recorded electronically. Data were imported into Stata/IC 10.0  when outcomes, time-to-event indicators and explanatory variables were programmed.Major protocol violations were defined as subjects: i) missing days 1, 2, 3 of the ACT arm or days 2,3 of the monotherapy arm; ii) taking folic acid or a concomitant medication with anti-malarial activity ; iii), whose day 42 visit was more than three days late; and iv) taking an incorrect dose or repeat dose for vomiting. Data were not censored for missed visits as it was assumed intensive AE monitoring would detect malaria symptoms or treatment. Missing data fields were dropped where appropriate and subjects lost to follow-up after day 0 were automatically dropped from the time-to-event analyses. An attempt was made to follow all subjects to day 42 despite protocol violations.Sample sizes were calculated assuming an adequate clinical and parasitological response (ACPR) of 75% for SP and 90% for AS/SP  indicating 100 per treatment group in each phase of the study .Time-to-event outcomes, which were considered the most appropriate for the data, were analysed using Kaplan Meier (KM) survival methods and Cox's Proportional Hazards Regression. This allowed data up to the point of loss to follow-up or withdrawal to be included and for the relative hazard to approximate the relative risk.KM distributions and survival curves for the two treatment groups were compared using a log rank test. Should model diagnostics suggest hazards were not proportional, parametric models were applied . Time toThe study was approved by the Ethics Committees of the Mozambican Ministry of Health and the University of Cape Town prior to commencement, and conducted in accordance with the South African Clinical Trials Guidelines 2000 . Staff mSubject disposition is shown in Figure dhfr/dhps mutation increased the relative hazard of failure 3.2 fold compared to fewer mutations. The proportional hazard assumption was satisfied overall; however there was an accelerated time to failure (approximately 6-fold) for subjects with a quintuple mutation compared to subjects with fewer mutations suggesting that early treatment failure is particularly associated with presence of a quintuple mutation. When the analysis was repeated after excluding or censoring for major protocol violations , results confirmed those found above.From the survival analysis, Figure The ACT group had faster parasite clearance rates compared to the SP monotherapy group (log-rank p = 0.001) Figure . Multiva3-105 parasites per asexual life cycle, compared to 10-103 for SP) [The aim of this study was to provide evidence to Mozambican health policy-makers regarding efficacy of SP with or without AS as treatment for uncomplicated malaria. In a survival analysis, both drug regimens were effective, however adding artesunate improved outcomes significantly with regard to clinical and parasitological cure, and peripheral parasite clearance time. These results are consistent with the literature, that suggests combining AS with SP improves efficacy, provided efficacy of SP is sufficiently high . In addiSubjects aged over seven years had a 70% lower risk of treatment failure than those seven and under, and were 30% more likely to clear parasites faster (the latter association was only found when protocol violators were removed). These findings are consistent with the knowledge that immunity to malaria infection increases with age . Howeverdhps) and dihydrofolate reductase (dhfr) enzymes respectively. Resistance to SP develops due to accumulation of point mutations in genes that encode these enzymes. The presence of 5 mutations, 3 in the dhfr gene (S108N/N511/C59R) and 2 in the dhps gene (A437G/K540E) are associated with treatment failure [st line treatment of uncomplicated malaria (AQ-SP replaced with AS-SP) and as intermittent preventive treatment for pregnant women (SP in 2007).Sulphadoxine and pyrimethamine are anti-folates that inhibit the dihydropteroate synthetase  protocol violators, results were very similar. Including data from non-adherent subjects or those taking prohibited concomitant medications is useful for assessing the effect of these common practices on treatment response. AS courses shorter than three days are not recommended due to their lower efficacy and the increased likelihood of anti-malarial drug resistance emerging if combined with the more slowly eliminated SP . SP may,dhfr 164 mutation, associated with high-level pyrimethamine resistance found in neighbouring Malawi [This study found that, while both SP monotherapy and in combination with AS were effective, the ACT was far superior, supporting its current use in Mozambique as the national policy for the treatment of uncomplicated malaria. However, it is recognized that combinations with SP may have a limited useful life due to the spread of parasite resistance, and inadequate drug levels young children achieve. It is also possible that SP efficacy has declined since this study due to the national implementation of SP as an intermittent preventive treatment policy for pregnant women during 2007. Of further concern is the presence of the g Malawi .The authors declare that they have no competing interests.EA participated in study design, coordinated the study, conducted the statistical analysis and drafted the manuscript, TC managed the Mozambique study teams, YC provided medical expertise to the study teams, FL participated in study design, calculated sample sizes and supervised the statistical analysis, JR conducted molecular analyses, AB supervised analysis and reporting. KIB, Principal Investigator of the South East African Combination Anti-malarial Treatment (SEACAT) Evaluation, conceived of, and designed the study, and contributed to the analysis. All authors read and approved the final manuscript."}
+{"text": "A fixed-dose pediatric formulation of artesunate and mefloquine (Artequin Pediatric) has been developed. In this open, non-comparative study in Cameroonian children with uncomplicated falciparum malaria, the safety and efficacy of this formulation was tested, with a particular emphasis on the risk of neuropsychiatric adverse events (AEs). In total, 220 subjects, weighing between 10 and 20 kg, were enrolled; 213 qualified for analysis. Artesunate-mefloquine was given once daily for 3 days. Overall, 13.1% of patients reported mild to moderate neuropsychiatric AEs (elicited through a structured questionnaire or reported spontaneously) out of which 3.8% (mainly insomnia) were considered drug-related. Other drug-related AEs were infrequent (< 3%). Polymerase chain reaction-corrected cure rate  determined by survival analysis at 28 and 63 days was 96.6%. New infections were observed in 11.2% of evaluable patients at 63 days. The new formulation was well tolerated and efficacious in the population investigated. Plasmodium falciparum malaria is a leading cause of mortality in tropical countries, with most of the deaths occurring in children from sub-Saharan Africa.P. falciparum malaria in all endemic areas because of its efficacy and potential to reduce the spread of drug resistance.9P. falciparum malaria.17Artesunate-mefloquine is an ACT, which has been widely used in Asia; but also in Africa this combination appears to be highly efficacious in patients with uncomplicated Because of insufficient safety and tolerability data for artesunate-mefloquine, there has been concern regarding the use of this combination in African children, which restricted its deployment in this setting.25Concerns have also been raised by preclinical observations of neurotoxicity with artemisinin derivatives. In animal studies, intramuscular oil-based artemether and arteether have been associated with neuronal damage, particularly in areas of the brainstem involved in hearing and gait controlP. falciparum malaria. The primary aim was to assess the neuropsychiatric safety of the new pediatric formulation. Major secondary outcomes included Day-28 and Day-63 polymerase chain reaction (PCR)-corrected cure rates.Against this background, the safety and efficacy of the new granule formulation of artesunate-mefloquine was investigated in young African children with uncomplicated P. falciparum malaria were recruited from the peri-urban area of Yaound\u00e9.The study was performed between December 2007 and March 2009 in Yaound\u00e9, Cameroon, where malaria transmission is intense and perennial.P. falciparum malaria with a density between 2,000/\u03bcL  and 250,000/\u03bcL blood, ability to take drugs by mouth and to comply with the study protocol, and provision of written informed consent by the parent or guardian. Exclusion criteria included presence of severe and complicated malaria  as defined by WHO,Male or female children presenting with signs and symptoms of malaria were screened for study eligibility. Inclusion criteria were body weight between 10 and 20 kg, fever  or history of fever in the preceding 24 hours, Before enrollment, written informed consent was obtained from the parents or legal guardians of the child. The trial protocol was approved by the National Ethics Committee, Yaound\u00e9, Cameroon and the Ethics Committee of the Technical University of Dresden, Medical Faculty, Germany. This study is registered with ClinicalTrials.gov as NCT00978172.N = 50) were included in a prospectively planned interim analysis. The data from these patients were reviewed by an independent malaria expert and a neuropediatrician to confirm or reassess the initially planned sample size of 600 patients, and to look for insufficient efficacy and potential safety issues.This was an open-label, single-center, single-arm study. Patients were enrolled consecutively and admitted to the health facility for the first day of a 3-day treatment phase, and then followed up until Day 63 after treatment. A follow-up period of 63 days complied with the WHO guideline from 2003 recommending this observation period for patients treated with artesunate-mefloquine to avoid an underestimation of recrudescence rates.The investigational therapy (Artequin Pediatric) consisted of artesunate 50 mg/day (2.5\u20135 mg/kg/day) and mefloquine 125 mg/day (6.25\u201312.5 mg/kg/day) given in equal oral doses simultaneously in a fixed-dose formulation (stickpack with a mixture of artesunate and mefloquine granules) once daily for 3 consecutive days. The entire content of one stickpack was directly administered on the child's tongue. In case of administration difficulties, the content was to be put on a spoon and administered with a small amount of liquid. The mouth was thereafter to be rinsed with some liquid, e.g., with milk or water, and remaining granules swallowed. Patients have been encouraged to resume normal food intake as soon as food could be tolerated. The first dose was administered under supervised conditions at the health facility. The second and third doses were given at home by the parent or guardian. The two stickpacks for administration at home had to be returned to the study site (used or not) to check for treatment compliance. Patients who vomited a dose within 30 minutes after intake received a full replacement dose. No more than one dose was to be replaced per day. The study medication was provided by Mepha Ltd., Aesch, Switzerland in Good Manufacturing Practice quality.Parents or guardians were asked to bring their children back to the health center on Days 4, 7, 28, and 63 or on any other day if the child did not feel well. Rescue therapy according to local practice was administered to patients with early or late treatment failures,Neuropsychiatric examinations took place at baseline and on Days 7, 28, and 63. These examinations included standardized questions  to the parent or guardian concerning their observations, questions to the children according to their age and development status, doctor's observations, and standardized neurological assessments.Giemsa-stained thick and thin blood films were examined before initiation of treatment and at every follow-up visit (scheduled or not). Blood films were considered negative if no parasites were seen in 100 oil-immersion fields in a thick blood film. The thin film was used for species determination. Two qualified microscopists independently read the slides. In case of discrepant results, slides were additionally read by a parasitologist.To distinguish between recrudescence and new infection, blood samples for PCR analysis were collected from every patient at baseline and on Days 4, 7, 28, and 63. Additional PCR samples were taken in case the patient spontaneously returned to the health unit for signs and symptoms of malaria. For PCR analysis, a common technical protocol was used.33An open-label, non-comparative design was adopted as done previously with ACTs in this vulnerable population.36The primary objective of the trial was to assess the neuropsychiatric safety of the pediatric granule formulation of artesunate-mefloquine based on the incidence of neuropsychiatric AEs. Efficacy outcomes represented secondary study endpoints. Therapeutic outcomes were classified according to the current WHO protocol (WHO 2003) and included the proportion of patients who showed an adequate clinical and parasitological response (ACPR) at 28 and 63 days (PCR corrected and uncorrected). The ACPR was defined as absence of parasitemia on Day 28 (or Day 63) irrespective of axillary temperature, without any previous occurrence of early treatment failure, late clinical failure, or late parasitological failure.N = 50), the study size was reduced from 600 to 200 patients because of the low incidence of neuropsychiatric AEs overall and the virtual absence of drug-related neuropsychiatric AEs.A conservative 40% rate of neuropsychiatric AEs was assumed for the study treatment based on AE frequencies (mainly dizziness) reported for mefloquine when the drug was combined with an artemisinin derivative in adults from Southeast Asia.This was a single-arm study; hence, no pre-defined statistical hypothesis was tested. Consequently, no adjustment for multiple testing was made and two-sided 95% CIs were calculated for the primary safety variable . Day-28 and Day-63 ACPR rates were analyzed by the Kaplan-Meier method.Data were analyzed on the basis of different populations. The analysis of the primary safety endpoint was based on a modified safety population, defined as all patients who took at least one dose of study medication and had a post-baseline neuropsychiatric assessment on Day 7 or who reported a drug-related neuropsychiatric AE at any time before a new malaria episode occurred. The safety population included all patients who received at least one dose of study drug and had at least one post-baseline assessment, regardless of whether scheduled or unscheduled. A modified intention-to-treat (ITT) population was used for analyzing ACPR rates, and comprised all patients who had positive parasitemia at baseline and took at least one dose of study medication. Patients who discontinued the study before Day 4 without previous occurrence of early treatment failure or who had a mixed strain infection (as assessed by PCR) were excluded.N = 213), and 38.9% of the patients (14 of 36) who prematurely discontinued the study participation did so at or after Day 28. Only two patients discontinued the study because of an AE , both occurred during the 3-day treatment period but were considered unrelated to the study drug. Most patients  adhered to the visit schedule throughout the study, and only a low proportion of patients (< 1%) returned unused stickpacks to the study site. The mean (range) actual antimalarial doses in mg/kg/day were 3.5 (2.5\u20135) for artesunate and 8.8 (6.25\u201312.5) for mefloquine.In total, 220 patients with uncomplicated falciparum malaria were enrolled; 83.6% of patients completed the 63-day study . Main reFor both, the modified safety population and the modified ITT population, 213 (96.8%) participants qualified for inclusion. The main reason for exclusion was missing follow-up data for any visit (2.3%). In total, 215 (97.7%) patients were included in the safety population. Baseline demographics and background characteristics are shown in Artesunate-mefloquine was well tolerated by the study participants. Only two children vomited during the 3-day treatment period. One patient vomited the study drug and the replacement dose on Day 1, another patient vomited the Day-2 study medication.N = 27). Almost all of the neuropsychiatric AEs were either mild or moderate in severity. One child suffered from severe convulsion, which was considered unrelated to study medication and caused by the underlying malaria disease. The event resolved within 2 days on treatment with intravenous quinine.In total, 28 patients reported 50 neuropsychiatric AEs over the study course , which rThe incidences of drug-related neuropsychiatric AEs (primary safety variable) are shown in A total of 169 AEs other than neuropsychiatric were reported in 112 patients (52.1%). The most frequently affected system organ classes were infections and infestations (27.0%) and blood and lymphatic system disorders (20.5%). The most frequent AEs were anemia (20.5%), bronchitis (7.0%), and pyrexia (6.5%); vomiting was reported by 4 (1.9%) patients. Most of these AEs were either mild or moderate in severity. The AEs were not unexpected in children suffering from acute uncomplicated malaria and were largely pre-existing at baseline. Eleven [reported in 10 (4.7%) patients] of the 169 non-CNS AEs were classified as related to the study drug. All of the AEs suspected to be drug-related were mild or moderate in severity. The most frequent drug-related AEs were anemia , diarrhea  and vomiting ; they all had an onset within the first week of the study and lasted for 1\u20133 days, except anemia, which lasted 3\u20134 weeks.Overall, six SAEs occurred in 5 (2.3%) patients. One patient suffered from two episodes of anemia requiring hospitalization. Both episodes occurred in the context of a malaria infection. Two patients were hospitalized for severe malaria in the context of re-infections, one child was hospitalized because of severe asthenia related to the baseline malaria infection, and one patient suffered from convulsions during the night after baseline assessment and was hospitalized. All six SAEs were classified as not related to the study drug and resolved by the time of the last follow-up visit. No patient died.Changes of vital signs and hematology parameters were in accordance with malaria recovery.Cure rates were high . The PCRP. falciparum infection up to Day 63 . Mean time to re-infection was 52 \u00b1 11 days (median: 56 days); there was only one re-infection by Day 28. The most frequently used rescue medication for treatment failure or new infection was artemether/lumefantrine (3.3% of patients).In the non-PCR corrected analysis, the ACPR rates were 96.1% for Day 28 and 85.4% for Day 63 . There were 22 patients with a PCR-confirmed new The assessment of the safety and efficacy of a new pediatric granule formulation of artesunate-mefloquine in the treatment of uncomplicated falciparum malaria in African children weighing 10\u201320 kg was the main purpose of this study. The new formulation was well tolerated and highly efficacious in the treatment of falciparum malaria in young Cameroonian children. Very few patients (3.8%) reported drug-related neuropsychiatric AEs out of which sleeplessness/insomnia of mild to moderate severity occurred most frequently. All non-drug-related neuropsychiatric events were likely to be related to the underlying malaria disease. Only 4.7% of patients reported non-CNS AEs suspected to be drug-related, all of mild or moderate severity; mainly anemia and gastrointestinal disorders, which also may have been attributable to the underlying malaria infection. Non-drug-related, non-CNS AEs were observed but all of them were expected in children suffering from acute uncomplicated malaria and were largely pre-existing at baseline. No new safety signal was detected.Data on neuropsychiatric effects of mefloquine in young children are limited. One study performed in Southeast Asia found no evidence for such effects in children less than 5 years of age (weighing > 5 kg), when mefloquine, either alone or combined with an artemisinin derivative, was administered to treat falciparum or vivax malaria.Vomiting of antimalarial drugs is an important consideration in treatment. Mefloquine may cause two types of vomiting: early vomiting (< 1 hour after intake) related to gastric intolerability and late vomiting (> 1 hour and up to Day 7) probably related to central effects and co-existence of disease-related symptoms.As expected, the efficacy of the new artesunate-mefloquine pediatric formulation was excellent, with PCR-corrected ACPR rates of 96.6% for both Day 28 and Day 63, thereby indicating the absence of recrudescence after Day 28. In adolescents and adults, high Day-28 cure rates have been previously reported for the artesunate-mefloquine co-blister preparation.43N = 270), amodiaquine+sulfadoxine-pyrimethamine (N = 507), amodiaquine+artesunate (N = 515), and artemether-lumefantrine (N = 519) was compared.It has been suggested that ACT partner drugs with longer prophylactic times could result in a larger impact in higher-transmission settings, although their long-term benefit must be evaluated in relation to the risk of parasite resistance.The administration of study medication was largely done under non-supervised conditions . This is close to normal outpatient practice. The limitations of the study included the absence of an active control arm, which makes it difficult to estimate relative safety and efficacy in comparison with other treatments. In addition, it was an open-label study, which is associated with the unavoidable risk of bias, particularly with regard to AE reporting. However, the use of a standardized questionnaire adapted according to age and developmental status, and the conduct of standardized neurological examinations lowered the risk of bias. Moreover, the fact that neuropsychiatric AEs were elicited (not only spontaneously reported) carried the risk for over-reporting of such AEs; whereas certain signs and symptoms, which are difficult to assess in very young children , may have been under-reported in this age group.P. falciparum malaria in young Cameroonian children weighing between 10 and 20 kg. A small proportion of patients reported drug-related neuropsychiatric AEs of which sleeplessness/insomnia occurred most frequently. Despite earlier restrictions, there seems to be no reason to withhold artesunate-mefloquine from this population.In conclusion, over a 2-month observation period, the new fixed-dose pediatric granule formulation of artesunate-mefloquine administered once daily for 3 consecutive days was well tolerated and highly efficacious for the treatment of acute uncomplicated"}
+{"text": "PfLSA-3-rec adjuvanted with aluminium hydroxide. Eleven weeks after the third and last immunization she was experimentally infected by bites of Plasmodium falciparum-infected mosquitoes. When the thick blood smear became positive, at day 11, she was treated with artemether/lumefantrine according to protocol. On day 16 post-infection i.e. two days after completion of treatment, she woke up with retrosternal chest pain. She was diagnosed as acute coronary syndrome and treated accordingly. She recovered quickly and her follow-up was uneventful. Whether the event was related to the study procedures such as the preceding vaccinations, malaria infection or antimalarial drugs remains elusive. However, the relation in time with the experimental malaria infection and apparent absence of an underlying condition makes the infection the most probable trigger. This is in striking contrast, however, with the millions of malaria cases each year and the fact that such complication has never been reported in the literature. The rare occurrence of cardiac events with any of the preceding study procedures may even support a coincidental finding.A 20 year-old healthy female volunteer participated in a clinical Phase I and IIa safety and efficacy trial with candidate malaria vaccine Apart from acute coronary syndrome, myocarditis can be considered as a final diagnosis, but the true nature and patho-physiological explanation of the event remain unclear. Experimental human malaria infections are a well accepted method for testing efficacy of candidate pre-erythrocytic malaria vaccines since 1971 . The cliHere, a cardiac event occurring shortly after treatment is described in a healthy volunteer participating in a malaria vaccine phase I/IIa trial.PfLSA-3-rec in 2007-2008. The volunteer's medical history was unremarkable, except for a tonsillectomy at the age of 4 and mild atopic symptoms for which she occasionally used a nasal corticosteroid. As medication, she also used a second generation oral contraceptive. Family history revealed a myocardial infarction in the paternal grandfather, when 43 years old, and a positive paternal family history for dyslipidemia. She never smoked or used illicit drugs. At inclusion, the blood pressure was 135/88 mmHg with a pulse of 64/minute and a body mass index of 20 kg/m2; the results of the physical examination were normal. Subsequent blood pressure measurements yielded values below 130/80 mmHg. ECG and laboratory tests were normal, including plasma glucose and lipid levels  participated in a double blind randomized phase I conditional IIa trial with the candidate malaria vaccine The volunteer received three immunizations with 30 \u03bcg of LSA-3 recombinant protein adjuvanted with aluminium hydroxide. Immunizations were administered subcutaneously in the upper arm at monthly intervals. Adverse events after immunization were injection site pain, and headache with increasing intensity after each immunization. The headache after the third vaccination was severe and accompanied by photophobia; it lasted for two days and made her keep bed rest.Plasmodium falciparum (strain NF54). The thick blood smear became positive eleven days after exposure  and treatment with 80/480 mg artemether/lumefantrine (Riamet\u00ae) was immediately initiated. The dose was repeated after 8, 24, 36, 48 and 60 hours. At the time of treatment onset the volunteer reported headache, fatigue and myalgia. These symptoms had started at day eight and nine after exposure. The day after treatment onset the volunteer reported intermittent fever, which had completely resolved two days later. The highest recorded sublingual temperature was 39.6\u00b0C. Artemether/lumefantrine was continued until day 14 and blood smears and PCR were negative on this day. On day 15 she experienced some shortness of breath when performing physical exercise, which she attributed to hay fever.Permission from the Independent Research Committee was obtained for proceeding with the phase IIa part of the trial. Eleven weeks after the last immunization the volunteer was exposed to the bites of five mosquitoes infected with On day 16, two days after completion of treatment and with a negative blood smear, the volunteer woke up at 5.30 a.m. with retrosternal chest pain characteristic of angina pectoris, not radiating, not related to respiration or change of position. Pain was severe during 30 minutes and at that time accompanied by nausea. The volunteer called the study doctor who referred her to the cardiologist. On admission, the volunteer appeared not in acute distress, blood pressure was 120/80 mmHg, pulse 75/min, oxygen saturation 100% while breathing ambient air and temperature was 37.9\u00b0C. The jugular venous pressure was normal, as was the examination of the heart, lungs and peripheral arteries. The electrocardiogram showed 1 mm ST elevation in leads II, III and AVF and a negative T-wave in lead V2 -laterally suggestive for myocarditis , whilst having oppressive, non-radiating pain on the chest  and after treatment .Click here for fileElectrocardiogram showing the electrocardiograms previous to admission , whilst having oppressive, non-radiating pain on the chest  and after treatment .Click here for fileElectrocardiogram showing the electrocardiograms previous to admission , whilst having oppressive, non-radiating pain on the chest  and after treatment .Click here for fileElectrocardiogram showing the electrocardiograms previous to admission , whilst having oppressive, non-radiating pain on the chest  and after treatment .Click here for fileCardiac Magnetic Resonance Images. MRI images (file MRI.gif), showing the MRI image made ten months after the event. Late contrast enhanced pictures show a small, patchy staining sub-epicardial laterally and postero-laterally and slight staining spots on the infero-septal - inferior border.Click here for file"}
+{"text": "Plasmodium falciparum. This phenomenon suggests that there is a synergy between drug treatment and acquired immunity. This hypothesis was examined in an area of moderately intense transmission of P. falciparum in Tanzania during a drug trail with sulphadoxine-pyrimethamine (SP) or amodiaquine (AQ).In malaria endemic areas children may recover from malaria after chemotherapy in spite of harbouring genotypically drug-resistant P. falciparum antigens were assessed and related to treatment outcome.One hundred children with uncomplicated malaria were treated with either SP or AQ and followed for 28 days. Mutations in parasite genes related to SP and AQ-resistance as well as human sickle cell trait and alpha-thalassaemia were determined using PCR and sequence-specific oligonucleotide probes and enzyme-linked immunosorbent assay (SSOP-ELISA), and IgG antibody responses to a panel of Parasitological or clinical treatment failure (TF) was observed in 68% and 38% of children receiving SP or AQ, respectively. In those with adequate clinical and parasitological response (ACPR) compared to children with TF, and for both treatment regimens, prevalence and levels of anti-Glutamate-rich Protein (GLURP)-specific IgG antibodies were significantly higher (P < 0.001), while prevalence of parasite haplotypes associated with SP and AQ resistance was lower . Interestingly, anti-GLURP-IgG antibodies were more strongly associated with treatment outcome than parasite resistant haplotypes, while the IgG responses to none of the other 11 malaria antigens were not significantly associated with ACPR.These findings suggest that GLURP-specific IgG antibodies in this setting contribute to clearance of drug-resistant infections and support the hypothesis that acquired immunity enhances the clinical efficacy of drug therapy. The results should be confirmed in larger scale with greater sample size and with variation in transmission intensity. Plasmodium falciparum resistance to commonly available antimalarial drugs such as chloroquine (CQ), amodiaquine (AQ) sulphadoxine-pyrimethamine (SP) is now widespread in most malaria-endemic areas, including Tanzania  \u00d7 100. The cut-of for a positive antibody response was defined as the mean level plus two standard deviations of the antibody reactivity among the negative controls.The plasma samples collected at day 0 were tested for the presence of IgG to the eleven antigens mentioned above by ELISA as previously described . Wells osample plasma - MFIbackground)/(MFIpositive plasma - MFIbackground)] \u00d7 100. The antibody positivity cut-of was determined as the mean antibody levels of the plasma from the mean of the six Danish donors plus two standard deviations.Plasma levels of IgG with specificity for variant surface antigens (VSA) on two parasite isolate lines with different VSA-expression profiles, 3D7 unselected (VSA1) and 3D7 selected on transformed human bone marrow endothelial cells (THrMEB) (VSA2)  were tes2 test and Fisher's exact test were used to compare differences in proportions of antibody responders, presence of parasite resistant genotypes and genes assessing haemoglobinopathies in patients with adequate clinical and parasitological response (ACPR) and treatment failure (TF). Non-parametric Mann Whitney rank sum test was used to analyse differences in median antibody levels between treatment groups. Associations between quantitative variables as age, parasite densities and haemoglobin levels were assessed by linear regression. For multivariable analysis, logistic regression models were used to assess predictors of treatment failure. In these models, parasite density (log parasite density pre-treatment), age and age-squared were used as continuous exposure variables, while presence or absence of parasite resistant genotypes, human antibodies and genes causing haemoglobinopathies were included as categorical exposure variables. Possible effect modification between the exposure variables was considered by testing for interaction between presence of parasite mutant haplotypes and IgG antibodies. P < 0.05 was defined as significant.Statistical analyses of data were performed with Stata/SE version 8.2 . \u03c7The baseline characteristics of the 100 patients enrolled in the study are shown in Table Of the 50 patients receiving SP, none presented with early clinical failure (ECF), three patients presented with late clinical failures (LCF), 31 patients presented with late parasitological failures (LPF) and the remaining 16 patients had an adequate clinical and parasitological response (ACPR). Of the 50 patients receiving AQ, one presented with ECF, two patients presented with LCF, 16 patients presented with LPF and the remaining 31 patients had an ACPR. All samples were adjusted for new infections by PCR genotyping. Before PCR corrections, 32 patients receiving SP and 22 patients receiving AQ presented with LPF, respectively. All patients with either ECR, LCF or LPF were grouped together as treatment failures (TF). Thus, the treatment failure rates were 68% in the SP group and 38% in the AQ group. There were no significant differences with respect to age, sex, temperature, and haemoglobin level between patients with ACPR or TF in any of the treatment groups. However, in patients receiving SP, the parasite density on day 0 was higher in the TF than ACPR group (P = 0.03). Levels of haemoglobin, parasite density and failure rates showed no relation to age (data not shown).P. falciparum dhfr gene, and codon 436/437, 540, 581 and 613 of the P. falciparum dhps gene, were determined on day 0 for parasite infections in patients receiving SP  compared to patients with ACPR (50%), . There was no significant association between dhps mutant haplotypes (single SGKAA and double SGEAA combined) and treatment outcome , nor between the quintuple dhfr-dhps mutant haplotype (CIRN-SGEAA) and TF . However, due to the low number of samples, we could not exclude a possible association between the quintuple mutant haplotype and treatment outcome.A higher prevalence of the triple mutant haplotype, CIRN in Pfcrt mutant haplotype CVIET (68%) and the remaining samples were the CVMNK wildtype haplotype. There was a trend towards higher prevalence of the CVIET haplotype in patients failing treatment (84%) compared to patients with ACPR (58%), although not statistically significant .The majority of parasites in patients receiving AQ expressed the dhfr haplotype in patients failing SP treatment and a tendency for a higher prevalence of the mutant CVIET haplotype in patients failing AQ treatment were observed, indicating some association between presence of parasite resistant genotypes and treatment outcome.Thus, a higher prevalence of parasites expressing triple mutant 2 = 20.9, P < 0.001) and anti-GLURP-R2 IgG  as well as higher levels of anti-GLURP-R0 and R2 IgG (P < 0.001) compared to patients with TF. Such statistically significant associations were not observed for any of the other antigen-specific IgG responses. Furthermore, the associations between ACPR and prevalence and levels of GLURP-R0 and R2 antibodies were significant when the SP and AQ groups were analysed separately . The prevalence of the combined IgG responses to GLURP-R0 and R2 was also correlated to ACPR in both the SP  and the AQ  group. By contrast, no association between antibody responses to any other antigen, analysed individually or in combination, and treatment outcome in neither the SP nor the AQ group was demonstrated. Approximately 25% of patients with IgG against GLURP R0 did not have IgG against R2 and vice versa, indicating that responding to R0 not necessarily means responding to R2. Thus, among the thirteen IgG specificities, only anti-GLURP-R0 and R2 IgG proved to be associated with ACPR.Prevalence and levels of IgG antibodies with specificity for the eleven malaria antigens and the surface of the two parasite isolates were measured on day 0 in patients treated with SP or AQ Figure . In univ+-thalassaemia was 38% and the frequency of sickle cell trait (HbAS) 12%. No sickle cell anaemic patients (HbSS) were found. The prevalence of these traits was similar in the two treatment outcome groups for both SP and AQ , but not for SP (P = 0.41).In the study population, the overall frequency of homozygote and heterozygote alphadhfr triple mutant haplotype in the SP group, and the CVIET haplotype in the AQ group), and presence or absence of IgG antibodies to each antigen. In Table Patients were divided into four groups based on infections with or without mutant haplotypes  were more likely to have ACPR compared to patients with no anti-GLURP IgG infected with a resistant parasite haplotype (group 4). Interestingly, there was a marked positive effect of having anti-GLURP IgG on treatment outcome in patients with resistant haplotypes , indicating that possessions of anti-GLURP IgG were associated with reduced risk of TF when infected with resistant parasites. Similar significant associations were seen for GLURP-R2 but not for any of the other antigens.In multivariate regression models, the effect of IgG antibodies, parasite haplotype and parasite density on treatment outcome was analysed Table . Regardldhfr (CIRN) irrespective of the dhps haplotype status as demonstrated in other studies   19 was n with CQ ,23 treat with CQ  and antireatment , and IgGreatment . In addireatment  treatmenreatment , althougreatment . FinallyInterestingly, at day 0, all patients had IgG antibodies to at least one of the antigens tested, but no patient had IgG antibodies to all the antigens. This may reflect substantial variation in response to infections. Previous immuno-epidemiological studies have demonstrated that GLURP-specific IgG antibodies are associated with protection against high parasitaemia , clinicaThis study suggests that presence or level of GLURP-specific IgG antibodies in some settings are better predictors of SP and AQ treatment outcome than the parasite density at enrolment and drug-resistant related parasite mutations. This confirms the hypothesis that acquired immunity enhances the efficacy of antimalarial treatment, and supports the use of drug trials to identify markers of immunological importance. In addition, the findings suggest that drugs with reduced efficacy, such as SP, may still be effective for treatment of uncomplicated malaria in individuals with some acquired immunity, such as older children and adults living in areas of intense malaria transmission.AE, LV, MT and MA conceived and designed the study. WN was responsible for the clinical trial and the supervision of patient enrolment and sample collection. AE performed the experiments and analysis with MA, MT, IB and LV. TS, AJ, IB and TT participated in manuscript preparation and design of the study. All authors read and approved the final manuscript."}
+{"text": "The artemisinin-based combination treatment (ACT) of dihydroartemisinin (DHA) and piperaquine (PQP) is a promising novel anti-malarial drug effective against multi-drug resistant falciparum malaria. The aim of this study was to show non-inferiority of DHA/PQP vs. artesunate-mefloquine (AS+MQ) in Asia.Plasmodium falciparum mono-infection or mixed infection were randomised with an allocation ratio of 2\u22361 to a fixed-dose DHA/PQP combination tablet  or loose combination of AS+MQ . The cumulative doses of study treatment over the 3 days were of about 6.75 mg/kg of DHA and 54 mg/kg of PQP and about 12 mg/kg of AS and 25 mg/kg of MQ. Doses were rounded up to the nearest half tablet. The primary endpoint was day-63 polymerase chain reaction (PCR) genotype-corrected cure rate. Results were 87.9% for DHA/PQP and 86.6% for AS+MQ in the intention-to-treat , and 98.7% and 97.0%, respectively, in the per protocol population (97.5% CI: >\u22120.39%). No country effect was observed. Kaplan-Meier estimates of proportions of patients with new infections on day 63 (secondary endpoint) were significantly lower for DHA/PQP than AS+MQ: 22.7% versus 30.3% . Overall gametocyte prevalence , measured as person-gametocyte-weeks, was significantly higher for DHA/PQP than AS+MQ . Fifteen serious adverse events were reported, 12 (1.6%) in DHA/PQP and three (0.8%) in AS+MQ, among which six (0.8%) were considered related to DHA/PQP and three (0.8%) to AS+MQ.This was an open-label, randomised, non-inferiority, 63-day follow-up study conducted in Thailand, Laos and India. Patients aged 3 months to 65 years with P. falciparum malaria in areas where multidrug parasites are prevalent. The DHA/PQP combination can play an important role in the first-line treatment of uncomplicated falciparum malaria.DHA/PQP was a highly efficacious drug for ISRCTN81306618Controlled-Trials.com  The bisquinoline piperaquine (PQP) was first synthesised in the 1960s independently by teams in China and France. In 1978, because of its greater potency and tolerability relative to chloroquine, PQP replaced chloroquine in the Chinese National Malaria Control Programme. Such was the success of the programme, that the following decade saw the use of PQP decrease because of the decrease in patients suffering from malaria.In 1990, Chinese scientists working to develop alternative anti-malarial treatments that might offer higher cure rates with better tolerability profiles included PQP phosphate in an artemisinin-based combination treatment (ACT), known as China-Vietnam 4 (CV4\u00ae), that also included dihydroartemisinin (DHA), trimethoprim and primaquine phosphate Since their introduction in the 1990s, ACTs have been found to be highly effective treatments for malaria In order to assess the safety and efficacy of the treatment combination of DHA and PQP in Asia, we conducted a randomised trial in Thailand, Laos and India comparing DHA/PQP with another ACT, artesunate (AS) plus mefloquine (MQ). All study sites were located in areas of notable chloroquine resistance. Dihydroartemisinin plus PQP was administered as a single tablet (DHA/PQP) and AS plus MQ were administered as separate loose tablets (AS+MQ). This study had the largest sample size to date of any study assessing DHA/PQP in South East Asia.The protocol and amendments for this trial and supporting CONSORT checklist are available as supporting information; see This study was conducted in six centres in Thailand , two centres in Laos  and three centres in India  over two malaria seasons, with patients screened from June 2005 to February 2007.Plasmodium falciparum and P. vivax in the region are symptomatic and P. falciparum is multi-drug resistant, with high levels of resistance to chloroquine The study regions provided a range of transmission conditions and standard treatments. Study regions in Thailand were located in malarious forest on the Thai-Myanmar border in regions of unstable, low and seasonal malaria transmission that used AS+MQ as a first-line treatment. Nearly all infections with P. falciparum  or mixed infection were eligible for the study. Local regulations were followed in India that required all recruited patients to be aged 18 years and over. Key exclusion criteria were: severe malaria, treatment with MQ in the 60 days prior to screening, treatment with DHA/PQP in the 3 months prior to screening and >4% parasitised red blood cells. Pregnant or lactating women were not eligible for the study.Males and females aged from 3 months to 65 years weighing at least 5 kg with fever or history of fever (\u226537.5\u00b0C) and microscopically confirmed mono-infection with This was a randomised, Phase III, open-label study with two treatment arms: DHA/PQP and the active comparator AS+MQ. At the time the study was designed, there was no single first line treatment in the countries involved in the study. However, AS+MQ was first line treatment in Thailand, was well characterized and was widely used in all the considered countries. This made AS+MQ suitable as the control treatment against which the efficacy and safety of DHA/PQP would be tested. As the study was not blinded, to limit bias, the following procedures were put in place: 1) Randomisation was conducted under blinded conditions: the blind to the investigator and patient in the randomisation process was maintained by the use of sealed envelopes. 2) Evaluation of the PCR test results was blinded . 3) The chairman and the statistician of the independent Data Monitoring Committee reviewed the most relevant safety data and participated in the final blinded data review meeting where all decisions about assessment of the primary outcome and patient allocation to the pre-defined populations were made under blinded conditions. The randomisation list was generated by an external contract research organisation (MDS Pharma Services) using the plan procedure of SAS . Patients were allocated to receive either DHA/PQP or AS+MQ following a 2\u22361 randomisation schedule ratio. The unbalanced ratio was chosen to increase the chance of detecting rare adverse reactions and to provide more precise estimates of cure rates in the DHA/PQP arm.Dihydroartemisinin/PQP  was given once daily, on days 0, 1 and 2 of the study, at the standard dosage of 2.25 mg/kg and 18 mg/kg per dose of DHA and PQP, respectively, rounded up to the nearest half tablet. To facilitate the correct dosing of DHA/PQP, two formulations were used (DHA 20 mg + PQP 160 mg and DHA 40 mg + PQP 320 mg). Over the 3 days, the cumulative doses were of about 6.75 mg/kg of DHA and 54 mg/kg of PQP. Artesunate and MQ  were administered as separate tablets containing AS 50 mg and MQ 250 mg; AS was administered at 24-h intervals on days 0, 1 and 2 with a daily dose of 4 mg/kg and MQ was administered at 15 mg/kg on day 1 and 10 mg/kg on day 2 at 24-h intervals, but was not administered on day 0.P. falciparum, patients were told about the study and offered the opportunity to participate. Eligible patients who agreed to participate were given detailed explanations of the trial from study staff, including the information that they would be given one of two different anti-malaria treatments on a random basis. Participating patients all provided written informed consent. Patients who declined to participate were provided with treatment for uncomplicated P. falciparum malaria, standard for the area in which they lived. Participating patients were managed as outpatients and doses were given under medical supervision, although at some centres patients could be hospitalised according to local practice, which ranged from 3 days at Mae Ramat Hospital, Thailand and at centres in Laos and India to 28 days at Mahidol University, Thailand. If not in hospital, patients were encouraged to return to the study centre for assessments on days 1, 2, 3, 7, 14, 21, 28, 35, 42, 49, 56 and 63, and they were also told that they could make unscheduled visits on any day on which they felt unwell. Patients were followed-up for 63 days.As part of their routine treatment, febrile patients who attended the study centres underwent a thick blood smear test for malaria before any anti-malarial treatment was administered. If the smear was found to be positive for In accordance with WHO (WHO 2003 and 2009) and standard practice in malaria clinical trials, the primary endpoint of the study was the PCR-corrected cure rate, based on the adequate clinical and parasitological response (ACPR), which was defined as the absence of parasitaemia irrespective of the patient's body temperature, with the patient not meeting any of the pre-defined criteria of early treatment failure or late clinical or parasitological failure (see below). In more detail, the primary endpoint was defined using two methods. The first method was based purely on the standard definitions of early/late clinical and parasitological failure as defined by the WHO (B), and Fridericia, QTc(F)).Secondary endpoints included PCR-corrected adequate clinical and parasitological response on days 28 and 42, PCR-uncorrected adequate clinical and parasitological response (PCR-uncorrected), proportion of patients with early and late treatment failure, proportion of aparasitaemic patients, proportion of afebrile patients, number of new infections, gametocyte carriage and the safety profile of the two treatments including adverse events and 12-lead electrocardiograph parameters /(number of leukocytes counted). For samples with higher levels of parasitaemia (>3 parasites/1000 red blood cells), parasite density was calculated from the thin film per 1000 red blood cells as: . Gametocyte prevalence was also evaluated. Both the thick and thin blood smear readings were done locally following the above described procedure, while the gametocyte assessments were carried out in accordance with standard practice at each individual site. All technicians who read the slides had undergone appropriate training in malaria-related microscopy and had at least 5 years experience in reading blood smears. A process of quality control was used to monitor the values being provided by the local laboratories. One in five of every samples was sent to a central, independent laboratory  for review.The presence of Three spots of blood were collected on 3MM filter paper  at the enrolment visit and at any visit after day 7 for PCR analysis. Filter papers were dried and individually stored in a plastic bag containing silica gel. All filter papers were subsequently transferred to the Institute of Tropical Medicine  where centralised genotyping was conducted; deoxyribonucleic acid was purified as described elsewhere (B)) and Fridericia's (QTc(F)) correction methods. Electrocardiogram results were tabulated as normal, borderline or prolonged according to gender and age normal ranges : normal: <430 msec; borderline: 430\u2013450 msec; prolonged: >450 msec. Adult females: normal: <450 msec; borderline: 450\u2013470 msec; prolonged: >470 msec).Twelve-lead electrocardiograms were recorded at days 0, 2, 7, 28 , 63 and on the day of any recurrent parasitaemia that occurred using the CarTouch device with CarTouch version 1.4.1 software . The electrocardiogram was viewed during recording and then transmitted via modem to MDS Pharma Services Central Telemedicine Department  for interpretation and reporting. The data were analysed using both Bazett's  and on the day of any recurrent parasitaemia.Study staff conducted all interviews with patients, and children's parents, in their native language and they explained their rights under International Conference on Harmonisation-Good Clinical Practice (ICH-GCP). When obtaining informed consent from patients the signature of a witness was obtained if patients were unable to write. Consideration was given to the ethical implications of the randomisation ratio assigning more patients to receive DHA/PQP than AS+MQ during the design of the study. As the safety and tolerability profile of DHA and PQP has been well characterised at the doses proposed for the present study, it was considered acceptable to adopt this approach in the present study that would result in more patients receiving DHA/PQP.http://www.controlled-trials.com/isrctn/trial/l/0/81306618.html.The study protocol was approved by the following ethics committees: Institutional Ethics Committee, National Institute of Malaria Research (ICMR), Delhi, India; Goa Medical College and Hospitals Local Ethics Committee, Goa, India; Institutional Ethics Committee, Kasturba Medical College, Mangalore, India; Laos National Ethics Committee for Health Research (NECHR), National Institute of Public Health, Vientiane, Laos; Oxford Tropical Research Ethics Committee (OXTREC), University of Oxford, Headington, United Kingdom; The Ethical Review Committee for Research in Human Subjects, Ministry of Public Health, Nontaburi, Thailand; Tropical Medicine Ethics Committee (TMEC), Mahidol University, Rachathewi District, Bangkok, Thailand. The Food and Drug Department, Government of Laos PDR, approved the use of DHA/PQP in that country. The trial was conducted under the provisions of the Declaration of Helsinki (1964 and its subsequent amendments up to 2002) and in accordance with ICH-GCP. A Study Steering Committee, a Data Monitoring Committee and a Clinical Development Committee were created prior to the beginning of the trial, and worked independently to harmonise and monitor the study. The trial was registered prior to the enrolment of the first patient in the International Standard Randomised Controlled Trials Register, number ISRCTN81306618, at The statistical analysis was conducted according to a pre-defined data analysis plan. Three populations were prospectively planned. The intention-to-treat (ITT) population was defined as all randomised patients who took at least one dose of study treatment. The per protocol population was defined as all randomised patients who were eligible according to the main (pre-defined) protocol inclusion and exclusion criteria, received at least 80% of the study medication, underwent the day 63 assessment, took no other anti-malarial drugs (excluding rescue therapies) and, in the presence of asexual parasite stages on thick or thin blood smears, had an evaluable PCR test. Patients who missed visits from day 0 to day 2 were excluded from the per protocol population. The third population, referred to as modified ITT, was midway between the two described populations. Although this was pre-defined as the co-primary population (together with the per protocol), results in this paper are presented only for the two most extreme populations, i.e., ITT and per protocol populations, because these are standard and the findings were very similar in all populations analysed.The analysis of the PCR-corrected and uncorrected ACPR . All sources of uncertainty  were censored. Survival analysis techniques were also applied to the analysis of time to parasite clearance and the estimation of the rate of new infections . For descriptive purposes, the proportions of early, late and true treatment failures were also computed as simple rates for each treatment arm together with the relevant CIs.By-country heterogeneity in cure rates was assessed by the Breslow-Day test or by logistic regression, when the former was not applicable. The study was conducted over two malaria seasons with different centres working in the two study periods. To evaluate how the 63-day PCR-corrected cure rates varied across the cohorts from the two seasons, a logistic regression model was fitted with cohort and treatment as the explanatory variables. The treatment by cohort interaction was evaluated as a candidate to enter this model through a residual score test. All tests for heterogeneity and interaction were evaluated at the 10% significance level.Rates of person-fever-days and person-gametocyte-weeks were calculated as the number of weeks in which fever was present or blood slides were positive for gametocytes, respectively, divided by the number of follow-up weeks and expressed per 100 person-weeks.The safety population, which coincides with the ITT population, was used for all safety assessments. Adverse events were coded using the MedDRA dictionary (MedDRA V8.1). Proportions of patients experiencing at least one adverse event were compared between treatments using the Pearson Chi square test.To determine the sample size of this study, the PCR-corrected cure rate at day 63 was estimated to be at least 92% for AS+MQ in the ITT population using a literature search One thousand two hundred and thirty-nine patients were screened, with 769 patients randomised to DHA/PQP and 381 patients randomised to AS+MQ. There were no notable differences in demographic parameters between treatment arms overall or by country . ChildreOne hundred and forty-four patients were excluded from the per protocol population . There wThe analysis of the PCR-corrected cure rate at day 63 confirmed that DHA/PQP was non-inferior to AS+MQ. For the ITT population, PCR-corrected cure rates were 87.9% for DHA/PQP and 86.6% for AS+MQ  with a CI >1.75%. This treatment difference was also statistically significant . Similar results were obtained for the other populations.Non-inferiority of DHA/PQP versus AS+MQ was also shown at days 28 and 42 in all study populations . On daysA logistic regression test to assess by-country heterogeneity in the PCR-corrected cure rates , while the Kaplan-Meier estimate of the rate of aparasitaemic patients at day 3 (i.e. 24 h after completion of study treatment) was 97.6% for DHA/PQP and 97.6% for AS+MQ. Similar results were obtained for the per protocol population. Proportions of afebrile patients showed very similar profiles to aparasitaemic patients, with profiles being virtually superimposable. Fever incidence measured in person-fever-days (per 100 person-days) was similar for DHA/PQP and AS+MQ: 1065/5315 (20.04%) versus 524/2636 (19.88%), p\u200a=\u200a0.929 , measured as person-gametocyte-week rates, was significantly higher in the DHA/PQP arm in all populations  serious events were judged unrelated to DHA/PQP by the investigator: two cases of pyelonephritis, one case of aspiration pneumonia, and three cases of P. falciparum malaria. In the AS+MQ group three (0.8%) events in three patients were judged to be serious and all were judged related to study treatment: one case of anaemia from day 8, one case of convulsion on day 1, and one case of encephalitis from day 16 to day 31.There were no deaths during the study. In the DHA/PQP group, there were 12 (1.56%) events judged by the investigator to be serious, which occurred in 12 patients. Six (0.78%) of these events were considered by the investigators to be related to DHA/PQP. These were two cases of anaemia, one from day 7 to day 90, the other from day 7 to day 35; one viral infection (possibly Dengue fever) from day 15, fully recovered; and one Wolf Parkinson White (WPW) syndrome from day 2 to day 90. Wolf Parkinson White syndrome is a congenital defect of accessory conduction pathways. Electrocardiograms for this patient were submitted to two cardiologists to provide expert opinions. Both considered that the failure to diagnose WPW at baseline could be attributed to the increased heart rate caused by the patients' fever which hid the electrocardiographic characteristics of WPW. Other events considered serious and related were one convulsion on day 0, and one encephalitis on day 45 which resulted in a left-sided hemiplegia. The patient was diagnosed with Adverse event profiles for DHA/PQP and AS+MQ were very similar in terms of type and frequency of events and were consistent with those expected in adult patients with acute malaria. Most patients in the study experienced adverse events: 69.4% (532/767) of patients in the DHA/PQP arm and 72.4% (276/381) of patients in the AS+MQ arm. There was no statistically significant difference between the treatments in the incidence of adverse events . The most frequently reported adverse events were malaria symptoms, with headache the most commonly reported . The fre12/L for women and 4\u22125.5\u00d71012/L for men), 50% for low haemoglobin , and 67% for low platelets . At the end of the study following treatment, these proportions had decreased to approximately, 20%, 40% and 17%, respectively. There was no apparent difference between treatments. Increases in mean haemoglobin levels were observed over the 63-day follow-up period. In the ITT population, mean (standard deviation) haemoglobin values on day 0 were 118.7 (24.4) g/L for DHA/PQP and 119.8 (23.4) g/L for AS+MQ. At day 63, mean (standard deviation) changes from day 0 were 12.8 (22.2) g/L for DHA/PQP and 14.21 (21.2) g/L for AS+MQ. Similar results were observed for the per protocol population. Other than elevated liver parameters, as might be expected in this population, there were no relevant changes in biochemistry parameters.As would be expected in patients with malaria, anaemia and thrombocytopenia were common in each arm, approximate proportions were 30% for low red blood cells  method, there was a statistically significant difference between treatments , with a higher proportion of patients in the DHA/PQP group having borderline QTc(B) values . No statistically significant difference between treatments was observed at baseline for QTc(F) (2.9% for DHA/PQP vs. 1.6% in AS+MQ). On day 2, there was a statistically significant difference between treatments (Mantel-Haenszel Chi-square test p<0.001), with a higher proportion of patients in the DHA/PQP group having borderline  or prolonged  QTc(B) intervals than in the AS+MQ group . This difference was also observed for the QTc(F) method: borderline, 13.0% for DHA/PQP vs. 5.3% for AS+MQ (p<0.001); prolonged, 4.7% for DHA/PQP vs. 5.3% for AS+MQ (p<0.001). By day 7, there was no difference between treatments.At baseline, using the QTc(B) increase >60 msec from baseline to day 2 was 0.9% for DHA/PQP vs. 0.8% for AS+MQ (not significant), and 4.6% for DHA/PQP vs. 2.9% for AS+MQ when the same increase was assessed with QTc(F) (p<0.001). However, QTc and QT prolongation were reported as adverse events by 43 (5.6%) patients in the DHA/PQP group and 16 (4.20%) patients in the AS+MQ group; these were judged by the investigator to be related to study treatment for 28 (3.65%) patients in the DHA/PQP group and 13 (3.41%) patients in the AS+MQ group.The proportion of patients with QTc(F)values on day 0 were 387.70 msec for DHA/PQP and 385.54 msec for AS+MQ. Mean increases from baseline to day 2 were 22.93 msec and 14.65 msec, respectively. This difference between treatments was statistically significant (p <0.001). On day 7, the mean increase from day 0 for DHA/PQP had fallen to 10.47 msec with the value for AS+MQ being 13.39 msec; this difference was not statistically significant (p\u200a=\u200a0.075).Mean QTcP. falciparum malaria. The day-63 PCR-corrected cure rates . The sites in Laos and Thailand were also in regions with high levels of chloroquine resistance The DHA/PQP combination exerted a significant post-treatment prophylactic effect in this study. This is supported by, first, significant reductions in the incidence of new infections for DHA/PQP compared with AS+MQ and second, by higher uncorrected cure rates in the DHA/PQP arm than AS+MQ, as the uncorrected cure rate endpoint includes new infections as well as recurrence of infection. This effect is thought to be modulated by levels of drug remaining in the blood because of the long half-life of PQP. The post-treatment prophylaxis was significantly better for DHA/PQP reflecting the differential terminal half lives of PQP and MQ (4\u20135 weeks for PQP and 14 days for MQ). The extended post-treatment prophylactic effect is of particular importance in countries with a high risk of new infection and can also reduce the risk of anaemia by allowing patients more time for haematological recovery between infections. The DHA/PQP combination, like all ACTs, rapidly reduces parasite biomass in the patient through the brief yet potent activity of DHA (the artemisinin component). Subsequent removal of uncleared parasites is achieved by the less active but more slowly eliminated partner drug; in this case PQP with a half-life of 4\u20135 weeks (F) interval were seen for both DHA/PQP and AS+MQ after the start of treatment, but a statistically significant increase from baseline was observed in QTc interval for DHA/PQP relative to AS+MQ on day 2. An important element in assessing the clinical importance of QTc prolongation is the extent of change from baseline: an increase in QTc interval from baseline of >60 msec could be clinically significant and this is the recognised threshold for drug-induced arrhythmias in non-cardiovascular indications (ICH E14 guideline). There was a statistically significant treatment difference for the proportion of patients with such an increase in QTc(F), while no treatment differences were observed for the same increase in QTc(B) and for the incidence of cardiac events. Finally, by day 7 there were no significant differences between treatments. Based on our results, it is difficult to attribute a particular clinical relevance to the QTc increase observed with DHA/PQP. In fact, it is expected that malaria shortens the QT interval during the acute illness, leading to increases after the start of treatment. Most studies of antimalarial drugs report some transient prolongation of the QT interval in the days after the start of treatment Although formal statistical comparison of the tolerability profile of the two combinations was not planned or conducted, some individual adverse events appeared to be reported more frequently in patients receiving AS+MQ than those receiving DHA/PQP. Mefloquine has been linked with adverse events of gastrointestinal and central nervous system origin. In this study, the frequency of reporting of nausea, vomiting and dizziness was 2\u20133 fold higher in the AS+MQ arm than the DHA/PQP arm. Vomiting soon after dosing is an important determinant of treatment efficacy as attaining and maintaining effective systemic levels of anti-malarial drugs is essential to disease outcome. This is of particular importance where there are powerful influences on adherence to dosing regimens. Increases in QTcFor patients to access DHA/PQP via public sector healthcare systems, this fixed dose combination requires regulatory approval. One requirement for registration is that the formulation must be manufactured according to GMP. This study was conducted as one of a series of pharmacokinetic and ICH-GCP-compliant randomised, controlled trials at sites across Africa and South East Asia using DHA/PQP manufactured according to GMP.P. falciparum malaria. The effects were observed across the three Asian countries in which the study was conducted, with no country effect. The combination of DHA/PQP provided greater protection against new infections than AS+MQ. Results from this study indicate that, although there may be evidence that suggests that the overall efficacy of ACTs may be falling, DHA/PQP can play an important role in, possibly, a new policy of multiple first-line treatments of uncomplicated falciparum malaria.In conclusion, the fixed dose DHA/PQP combination tablet in this study emerged as a highly efficacious treatment for Checklist S1CONSORT checklist.(0.19 MB DOC)Click here for additional data file.Protocol S1Trial protocol.(1.32 MB PDF)Click here for additional data file.Protocol S2Protocol amendment 1.(0.14 MB PDF)Click here for additional data file.Protocol S3Protocol amendment 2.(0.23 MB PDF)Click here for additional data file.Protocol S4Protocol amendment 3.(0.19 MB PDF)Click here for additional data file.Protocol S5Protocol amendment 3A.(0.08 MB PDF)Click here for additional data file.Protocol S6Protocol amendment 3B.(0.09 MB PDF)Click here for additional data file.Protocol S7Protocol amendment 4.(0.09 MB PDF)Click here for additional data file.Protocol S8Protocol amendment 5.(0.10 MB PDF)Click here for additional data file."}
+{"text": "Because of the emergence of chloroquine resistance in Mali, artemether-lumefantrine (AL) or artesunate-amodiaquine (AS+AQ) are recommended as first-line therapy for uncomplicated malaria, but have not been available in Mali until recently because of high costs.Plasmodium falciparum infections.From July 2005 to January 2006, a randomized open-label trial of three oral antimalarial combinations, namely AS+AQ, artesunate plus sulphadoxine-pyrimethamine (AS+SP), and amodiaquine plus sulphadoxine-pyrimethamine (AQ+SP), was conducted in Faladje, Mali. Parasite genotyping by polymerase chain reaction (PCR) was used to distinguish new from recrudescent Plasmodium falciparum malaria were enrolled, and followed for 28 days to assess treatment efficacy. Baseline characteristics were similar in all three treatment groups. The uncorrected rates of adequate clinical and parasitologic response (ACPR) were 55.7%, 90.8%, and 97.7% in AS+AQ, AS+SP, and AQ+SP respectively (p < 0.001); after PCR correction ACPR rates were similar among treatment groups: 95.4%, 96.9%, and 99.2% respectively (p = 0.17). Mean haemoglobin concentration increased across all treatment groups from Day 0 (9.82 \u00b1 1.68 g/dL) to Day 28 (10.78 \u00b1 1.49 g/dL) (p < 0.001), with the greatest improvement occurring in children treated with AQ+SP. On Day 2, the prevalence of parasitaemia was significantly greater among children treated with AQ+SP (50.8%) than in children treated with AS+AQ (10.5%) or AS+SP (10.8%) (p < 0.001). No significant difference in gametocyte carriage was found between groups during the follow-up period.397 children 6 to 59 months of age with uncomplicated P. falciparum infection in Mali and appears to have the added value of longer protective effect against new infection.The combination of AQ+SP provides a potentially low cost alternative for treatment of uncomplicated  Plasmodium falciparum resistance to CQ has challenged control efforts in Mali, where recent studies indicate that the adequate clinical and parasitological response (ACPR) to CQ is estimated to be less than 50% in children aged between six months and nine years [For more than five decades, chloroquine (CQ) has been the main drug for the treatment of uncomplicated malaria in Mali. The emergence of ne years ,2 in difne years . The twone years ,5.Unfortunately, the implementation of ACT among the most vulnerable populations living in highly malarious rural areas has been hampered by the limited availability and high cost of the new drugs. In Mali, the healthcare system operates on local cost-recovery, and current prices for ACT in public sector range from 700\u20131,500 CFA (US$1.50\u20134.0). Thus, the majority of patients did not have access to the newly recommended treatment and continued to use chloroquine despite its low efficacy. Therefore, there was a need to test other combination therapies already available in the country before ACT became more widely available. WHO recommendations allow for the use of non-artemisinin combination treatment regimens for the treatment of uncomplicated malaria in settings where the component drugs are efficacious and well tolerated. Data from several settings have demonstrated a similar efficacy between ACT and the combination of amodiaquine plus sulphadoxine-pyrimethamine (AQ+SP) ,7.in vivo efficacy of ~95%) according to two separate studies [in vivo efficacy study to evaluate this non-artemisinin containing combination and compare it with two artemisinin-containing combinations (AS+AQ and AS+SP).In Mali, SP is still efficacious  with a peak in October. Most of the population is of the Bambara ethnic group with agriculture as the main occupation.in vivo criteria [P. falciparum with a parasitaemia of 2,000 \u2013 200,000/\u03bcl, ii) axillary temperature of \u2265 37.5\u00b0C, iii) haemoglobin \u2265 5 g/dL, iv) absence of febrile illness caused by diseases other than malaria, vi) absence of danger signs . Allocation to treatment groups was done according to block randomization (block size of 20); patients were not informed of the drug received.This was a randomized single-blind trial, conducted between July 2005 and January 2006, that included children between six and 59 months of age living in Faladje. After written informed consent of patients' parents or guardians, study subjects were enrolled if they fulfilled the following WHO criteria : i) micrTreatment was administered according to body weight at the following doses: AQ: 10 mg/kg/day from day 0 to day 2; AS: 4 mg/kg/day from day 0 to day 2; SP: 25 mg/kg of sulphadoxine and 1.25 mg/kg of pyrimethamine in single dose on day 0. All drugs were administered directly by the study team and the child was observed for 30 minutes. If vomiting occurred before 30 minutes, the dose was repeated; for vomiting after 30 minutes, a half-dose was administered. In the case of persistent vomiting, the child was referred to a health center for rescue treatment with intramuscular or intravenous quinine and withdrawn from the study.After the day of enrollment (day 0), the child was assessed on days 1, 2, 7, 14, and 28. Parents/guardians were encouraged to come to the clinic at any time when the child was sick.Capillary blood was obtained by fingerpick, thick and thin blood films were prepared and stained with 4% Giemsa for 20 minutes. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 300 WBCs assuming a WBC count of 7,500/\u03bcl. The same estimation method was used to calculate gametocyte density. Slides were read by an experienced medical microscopist, who was blinded to the treatment allocation. A negative blood film was defined as a slide with no parasites after review of 100 fields. Thin blood films were read only for parasite species determination. Ten percent of slides were read by a second reader, blinded to initial results. Discordant slides were read by a third microscopist.Blood haemoglobin concentration was measured using a portable photometer  on days 0, 14, 28 and any day of failure. Anaemia was defined as haemoglobin < 11.0 g/dl.To distinguish recrudescence from new infection, molecular genotyping techniques were used for patients who failed after day 7. At enrollment and during the follow-up visits, blood spots were obtained on filter papers and evaluated at the Molecular Epidemiology and Drug Resistance Unit of the Malaria Research and Training Center in Bamako. Molecular analysis was done according to the following scheme: paired blots (from day 0 and the day of parasitaemia recurrence) were analysed by nested PCR of parasite merozoite surface proteins (MSP1 and MSP2) and microsatellite (CA1), to distinguish between new infection and recrudescence as previously described .Treatment outcomes were classified following the 2003 WHO antimalarial drug efficacy guidelines . Early TThe protocol was approved by the ethical committee of the University of Bamako Faculty of Medicine, Pharmacy, and Odonto-stomatology (FMPOS) and the Institutional Review Board of the United States Centers for Disease Control and Prevention before the study was started. Overall community and local authorities' permission were also obtained in addition to parent or guardian informed consent, according to procedures described earlier .U-test for non-normally distributed continuous variables. The primary analysis was per protocol. Proportion of patients failing treatment was compared across groups using the chi-square test. Paired t test was used to estimate haemoglobin increase from baseline to different time points of regular follow-up. To account for multiple comparisons the results of the tests were considered statistically significant when p < 0.017 (0.05 divided by 3).Data were double entered using Microsoft ACCESS and statistical analysis was performed using SPSS version 11.0 . Baseline characteristics of subjects among groups were compared using the chi-square test for categorical variables and Mann-Whitney From July 2005 to January 2006, 876 children 6 to 59 months of age were screened for study entry Figure ; 397 werAt day 14, the proportion of children with anemia was significantly higher in AS+SP group (77.5%) when compared with AQ+SP , but not with AS+AQ ; the difference between the AS+AQ and AQ+SP groups was not statistically significant  when compared with day 0 (9.84 \u00b1 1.68 g/dl) (p < 0.001); and the most significant increase was observed among children in AQ+SP (1.07 g/dl) group followed by AS+AQ (0.95 g/dl) and by AS+SP (0.79 g/dl) groups.The proportion of patients with fever decreased from Day 0 (100%) to Day 1 (5.8%), Day 2 (2%), and Day 3 (1.5%) across all treatment arms. No difference was found among treatment groups at scheduled follow-up points with regard to the proportion of patients with fever. There were sharp declines in the proportion of children with parasitaemia from day 0 to day 2 in the AS+AQ (10.5%) and AS+SP (10.7%) groups, but not in AQ+SP (50.7%), (p < 0.001). By day 3, 1.5%, 0.8%, and 6% of patients were still parasitaemic in the AS+AQ, AS+SP, and AQ+SP groups respectively (p = 0.018)  or day 28 (p = 0.16) . A greater risk of new infection following treatment with AS+AQ when compared with the AQ+SP has been reported earlier in low  and highP. falciparum infection in Mali, and appears to have the added value of longer protective effect against new infections.The combination of AQ+SP provides a potentially low-cost alternative for treatment of uncomplicated The authors declare that they have no competing interests.In Mali, KK was the principal investigator and HM was the co-principal investigator of the study. At CDC, RN, MM, and AH contributed to the study design, protocol development, and manuscript writing. OY, HT, YK, EG, and RS participated in data collection, clinical investigation, and laboratory work. BT, AD were involved as field supervisors and laboratory data analysis. OD provided overall supervision in Mali. All authors participated in the writing and review of the final manuscript."}
+{"text": "Artesunate plus amodiaquine is a coblistered ACT, given as a single daily intake. It has been suggested that, in view of the number of tablets to be taken (particularly in adults), it may be possible to improve compliance by allowing patients to divide the daily dose. The objectives of this randomized, comparative, open-label, multicentre study, conducted in Senegal and in Cameroon in 2005, was to demonstrate the non-inferiority and to compare the safety of artesunate plus amodiaquine, as a single daily intake versus two daily intakes.A three-day treatment period and 14-day follow-up period was performed in any subject weighting more than 10 kg, presenting with a malaria paroxysm confirmed by parasitaemia \u2265 1,000/\u03bcl, after informed consent. Patients were randomly allocated into one of the two regimens, with dosage according to bodyweight range. All products were administered by an authorized person, blinded to both the investigating physician and the biologist. The primary endpoint was an adequate response to treatment on D14 (WHO definition). The two-sided 90% confidence interval of the difference was calculated on intent to treat (ITT) population; the acceptance limit for non-inferiority was 3%. The safety was evaluated by incidence of adverse events.Three-hundred and sixteen patients were included in the study. The two patient groups were strictly comparable on D0. The adequate responses to treatment were similar for the two treatment regimens on D14, PCR-corrected . The statistical analyses demonstrated the non-inferiority of administering artesunate/amodiaquine as two intakes. The drug was well tolerated. The main adverse events were gastrointestinal disorders (2.5%) and pruritus (2.5%); safety profiles were similar in the two groups.This pilot study confirms the efficacy and good tolerability of artesunate plus amodiaquine, administrated either in one or in two daily intakes. Plasmodium falciparum attacks.Nearly all currently available antimalarial monotherapies are facing resistance, the severity and incidence of which may vary according to endemic areas. Only one class of antimalarials, artemisinin derivatives and artesunate in particular, has been able to escape this phenomenon so far, most probably owing to its recent introduction, its mechanism of action and its rapid elimination from the body. National and international experts, in agreement with the WHO , have prThe onset of resistance to this type of treatment combination is unlikely, partly because the combined agents have different mechanisms of action, and also because the artemisinin derivative very rapidly reduces the parasite biomass which enables lasting exposure of the few remaining parasites to a high concentration of the combined drug.A number of controlled phase III studies under the auspices of the TDR , togethe\u00ae trademark.Bitherapy with artesunate and amodiaquine has obtained a marketing authorization in several African countries, including Senegal and Cameroon, under the ArsucamArtesunate plus amodiaquine was evaluated as a single daily intake in all of the studies already performed. The number of tablets to be taken for each intake (particularly in adults: eight tablets per intake) suggests that compliance may be improved by allowing patients to divide tablet intake into two daily intakes. The objective of this multicentre, randomized, open-label phase IV study, was to demonstrate the non-inferiority, in terms of clinical and parasitological efficacy on D14, of the administration of artesunate/amodiaquine as a single daily intake versus two daily intakes. The safety and tolerability of treatment were also studied.The study took place in three sites, two in south Senegal, where the climate is soudanian and the rainy season concentrated between June and October and one site in central Cameroon with a sub-equatorial climate, the rainy season is from April to November.Any adults or children weighing \u2265 10 kg, able to receive oral treatment, suffering from uncomplicated malaria attack with parasitaemia ranging from 1,000 to 200,000/\u03bcl (moderate and high transmission area) and axillary temperature \u2265 37.5\u00b0C or history of fever within the previous 24 hours, was enrolled after signed informed consent. Patients could not be included in the study if at least one sign of severe malaria or clinical danger (WHO definition) was present: serious concomitant disease, allergy to one of the investigational medicinal products, pregnancy, or documented intake of an antimalarial at a suitable dosage within seven days prior to inclusion.Conseil National de Recherche en Sant\u00e9 in Senegal and the Comit\u00e9 National d'Ethique in Cameroon on 17 June 2005 and 23 May 2005, respectively.An ethical approval was given by the 1 = 99% as the envisaged response rate for artesunate plus amodiaquine single dose (reference), a non-inferiority limit of 3%, a type I (\u03b1) error probability of 5% and a type II (\u03b2) error probability of 20%, the number of patients per group corresponded to 136. In order to anticipate premature withdrawals, a decision was made to include 150 patients per group, i.e. a total of 300 patients. Each site was, therefore, required to recruit 100 patients. In order to avoid any imbalance between the different weight categories, the randomization list was drawn up by centre and by weight range.Based on previous studies, with the hypotheses PDuring the inclusion visit (D0), a physical examination was performed with recording of medical history, demographic data , weight and height, and measurement of vital signs , and axillary's temperature, together with evaluation of symptoms. Parasitological tests  together with PCR filter paper were also performed.Follow-up visits took place on D1, D2, D3, D7 and D14 during which a physical examination was performed, with evaluation of clinical safety and symptoms, measurement of vital signs, and parasitological tests (except on D1 and D2). Filter paper blood spot samples were prepared on D14, if the thick blood film result was positive, with a view to PCR analysis.Since the artesunate plus amodiaquine coblisters contain one tablet of 50 mg artesunate and one tablet of 153 mg amodiaquine, dosages had to be adapted to the patient weight range: patients weighing between 10 and 21 kg received one tablet of each product per day, patients weighing \u2265 21 and \u2264 40 kg received two tablets of each product per day and patients weighing >40 kg received 4 tablets of each product per day. According to randomization, tablets were given either once or twice a day. The duration of treatment was three days in the two groups.Each patient was randomly allocated to one of the study groups, according to a pre-defined randomization list. The tablets were administered with a small amount of water. For the younger children, the tablets were crushed and administered with water. All treatments were administered in the presence of nursing staff. The patients stayed at the centre for 30 minutes after administration, for observation. If the patient rejected the treatment during the observation period, the same dose was re-administered. In the event of repeated vomiting, the patient had to be withdrawn from the study and a replacement treatment had to be introduced. All possible steps were implemented in order to avoid losing patients to follow-up: active follow-up, even going as far as the patient's home, was carried out by a member of the principal investigator's team.P. falciparum form after examination of 300 leucocytes; 10% of the slides were re-read at the centre's reference Parasitology Department at Dakar University.Thick films together with thin blood films were prepared at each study visit with the exception of D1. A molecular biology analysis (PCR) was performed on inclusion and on D14 +/- 1d in the event of a positive thick film on D14 +/- 1d. Slide-reading and parasite counts were performed on site and considered as negative in absence of asexual msp-1, msp-2 and microsatellitesThe molecular biology analyses were performed at the parasitology laboratory at the Faculty of Medicine, Universit\u00e9 Cheikh Anta Diop Dakar \u2013 Senegal using filter paper blood spot samples prepared in the field. The distinction of reinfections or recrudescence cases was carried out using genetic markers for Independent duplicate data entry was performed using the SAS software program, version 8.2, in compliance with the standards stipulated by GCP guidelines. All of the qualitatives and quantitatives variables recorded were described according to treatment group and globally. The 95% confidence intervals were calculated.The non-inferiority of artesunate plus amodiaquine as two intakes versus artesunate plus amodiaquine as a single intake was studied on the basis of the adequate response to treatment on D14 (WHO definition). The two-sided 90% confidence interval of the difference in the proportions of responders on D14 between the two treatment groups was calculated. If the upper limit of the confidence interval was below the acceptance limit (d = 3%), the non-inferiority of artesunate plus amodiaquine in two intakes versus a single intake would be demonstrated. This analysis was conducted on the ITT and per protocol (PP) populations. The main analysis corresponded to the ITT population.The safety of artesunate plus amodiaquine was evaluated by the presence or absence of adverse events, type of adverse event, intensity and the causal relationship with the investigational product in the safety population.From June to October 2005, 316 patients were included in the study. 161 patients were assigned to the artesunate plus amodiaquine single-intake treatment group (reference) and 155 to the artesunate plus amodiaquine two-intake treatment group. Seven patients were withdrawn from the study: five patients in the single-intake group and two in the two-intake groups. Only one patient (in the single-intake group) was withdrawn due to an adverse event, five patients were withdrawn because of loss to follow up and one because of consent withdrawn . Irrespective of treatment regimen, more than 99% of patients showed an adequate clinical and parasitological response.The primary efficacy endpoint was the adequate response to treatment after PCR analysis (ACPRc) between the two treatment groups was:  was below the acceptance limit, d = 3%., assessing that the two intakes treatment regimen is non-inferior to the single intake treatment regimen on the ITT population. These results were confirmed on the PP population.The results of the evaluation of efficacy on D14 before PCR and after PCR on the ITT population are presented on Table The changes in mean temperature during the study were similar in the two treatment groups, with values for mean temperature below 37.5\u00b0C as from D1. The percentage of apyretic patients between D1 and D14 is shown in Figure Gametocyte-carrier patients were 12 on inclusion, D7 and D14, and 14 on D3. The mean values for gametocytaemia were halved between D1 and D7 in both groups. With the exception of anorexia wich mainly affected patients under seven years of age in the two treatment groups, and was on D1 statistically higher in the single-intake group . A similar and favourable outcome, with regard to the symptoms of malaria, was observed for the two patient groups during the study. The changes in the percentage of patients in each group presenting these symptoms are shown in the Table Certain symptoms attributable to malaria, but not present on inclusion, appeared during treatment: asthenia (5.7%), vomiting (4.7%), anorexia (4.7%), diarrhoea (2.8%), abdominal pain (1.9%) and dizziness (2.2%), without any differences between the treatment groups. The influence of the number of tablets per intake, or the total quantity of drug substance to be ingested, on the onset of these symptoms was the subject of a descriptive analysis only, owing to the limited sample size.With the exception of two patients prematurely withdrawn from the study in the single-intake group , who only received treatment for one day, all of the other patients received treatment for three days, as provided for in the protocol. The adverse events (AE) recorded during the study were coded in compliance with the MedDRA dictionary, and described for the safety population.Among the population included, the AEs reported during the study mainly corresponded to gastrointestinal disorders (2.5%) or pruritus (2.5%), irrespective of the treatment regimen. Three patients in the two-intake group reported AEs related to nervous system disorders (two cases of drowsiness and one case of dysgeusia \u2013 bitter taste) versus one patient in the single-intake group (one case of seizure). All of the gastrointestinal or nervous system disorders and cases of pruritus reported during the study were perceived as related to the investigational product, irrespective of treatment regimen. Intensity of adverse events reported as being related to the investigational product is shown in Table Two serious adverse events occurred during the study, one in each treatment regimen; they were both perceived as unrelated to the investigational product. No death occurred during this study.Rejection of treatment during the first half-hour after administration was observed in 10 patients, six in the artesunate plus amodiaquine single-intake group and four in the two-intake group, mainly occurring on D0 Table . Out of Fifteen patients presented episodes of vomiting during the treatment period although this symptom was not observed on inclusion: seven patients (4.3%) in the single-intake group and eight patients (5.2%) in the two-intake group. It should be noted that this description does not take early treatment rejection into account. All these patients presented at least one of vomiting in the previous 24 hours. The number of tablets taken by the 15 patients concerned is shown in the Table The episodes of vomiting during treatment appear to have been influenced by the number of tablets taken each day. Dose fractionation does not appear to have any impact on the number of patients having presented this symptom during treatment. However, no conclusions may be drawn owning to the limited sample size in these subgroups.The correct prescription of ACT by healthcare staff and the compliance of patients are essential for an effective management of malaria in Africa. Different previous studies had already shown the efficacy and tolerability of artesunate plus amodiaquine in sub-Saharan Africa -7.The objective of this pilot study was to verify the non-inferiority of two daily intakes versus a single intake (which can permit to decrease the number of tablet to be taken per dose) and to evaluate the impact of dose fractionation on clinical safetyThis study showed that adequate responses to treatment were similar for the two treatment regimens, and approaching 100% before and after PCR analysis on D14. The statistical analyses conducted on the ITT and PP populations demonstrated the non-inferiority of administering Artesunate plus amodiaquine as two intakes versus a single daily intake, in terms of clinical and parasitological efficacy on D14.It was also demonstrated that the two tested treatment regimen were well tolerated; the observed AEs were similar in terms of type and incidence in the two treatment groups. No changes were observed in the safety profile when dividing Artesunate plus amodiaquine treatment into two daily intakes. The events reported during the study were expected and have already been described with this type of antimalarial.Numerous papers described an impact of artemisinin derivatives on gametocyte carriage, by killing immature stages of gametocytes. During this study, the number of gametocyte-carrier patients remained stable during follow-up (from D0 to D14), but the mean values for gametocytaemia were halved between D1 and D7 in both groups. This can possibly be explained by the presence of different stage of gametocytes in the study populations.The duration of follow-up of this study was only 14 days, but the efficacy has been demonstrated in numerous other studies within a D28 follow-up , and theThe combination artesunate plus amodiaquine has already been adopted as first line treatment of uncomplicated malaria attack in 18 countries in sub-Saharan Africa, with a regimen of one daily intake for three days. Even if the tolerability profile remains good in the clinical trials, in real life, some patients (mainly adults) complain about the too important number of tablets to be ingested at once. The results of this study will permit to divide the daily intake into two administrations.A fixed dose combination of these two active components has already been developed and is currently being evaluated in sub-Saharan Africa. This new formulation, with a limited number of tablets per intake may allow a better adherence to an effective treatment for African communities.The author(s) declare that they have no competing interests.JLAN: designed the study, collected data, and prepared the manuscriptBF : collected data and contributed in the preparation of the manuscriptVL: designed the study and prepared the manuscriptPh B and ASE : participated in the design of the study and collected dataAMD, PAS, M C and T K ; collected data on fieldOG : designed the study and prepared the manuscript"}
+{"text": "Plasmodium falciparum malaria.Numerous trials have demonstrated high efficacy and safety of artemisinin-based combination therapy (ACT) under supervised treatment. In contrast, effectiveness studies comparing different types of ACT applied unsupervised are scarce. The aim of this study was to compare effectiveness, tolerability and acceptance of artesunate plus amodiaquine (ASAQ) against that of artemether-lumefantrine (AL) in Ghanaian children with uncomplicated \u00ae) or AL (Coartem\u00ae). Study participants received their first weight-adjusted dose under supervision. After the parent/guardian was advised of times and mode of administration the respective three-day treatment course was completed unobserved at home. Follow-up visits were performed on days 3, 7, 14 and 28 to evaluate clinical and parasitological outcomes, adverse events, and haematological recovery. Length polymorphisms of variable regions of msp1 and msp2 were determined to differentiate recrudescences from reinfections. Acceptance levels of both treatment regimens were assessed by means of standardized interviews.A randomized open-label trial was conducted at two district hospitals in the Ashanti region, Ghana, an area of intense malaria transmission. A total of 246 children under five years of age were randomly assigned to either ASAQ . Interestingly, more late clinical failures until day 28 occurred in AL-treated children than in those who received ASAQ .Haematological recovery and drug tolerability were not found to be significantly different in both study arms. The acceptance of treatment with ASAQ was higher than that with AL .Unobserved AL and ASAQ treatment showed high adequate clinical and parasitological responses, though AL was inferior in preventing late clinical failures. Plasmodium falciparum drug resistance  were given weight-adjusted  were administered twice daily as three-day, six-dose regimen according to body weight , the second dose 8 hours after the first dose, according to manufacturer guidelines.AL .Both study drugs were well tolerated, and no drug-related SAE occurred during the study. There were four SAEs , i.e. one was a mild acute asthmatic attack on day 28, one a febrile convulsion on day 1, one child received concomitant treatment for suspected enteritic bacterial coinfection on day 3 and one child was extensively monitored due to a measured haemoglobin level of 4.8 g/dl on day 3 but subsequently needed no blood transfusion. Adverse events of grade 1\u20133 were, regarding their nature, severity, and relation to the study drug, similarly distributed in the study groups . This resulted in a slightly higher overall rank-score for ASAQ, compared to AL , although packaging and administration of the study drugs were appraised similarly positive in both groups . The discontinuation was necessary through the stopping rules of the study protocol despite the fact that the occurrence of anemia was not different in both study arms and was not related to treatment. Accordingly, the analysis was performed with a decreased number of study participants providing a power of 65% to detect differences between the study arms. Therefore, the lack of a significantly different effectiveness in one of the study arms has to be interpreted with caution. Nevertheless, cure rates could be provided within the given confidence intervals and the findings of different LCFs in both arms are valid independent of the estimated power.Another limitation of the study is the restriction of the follow-up period to 28 days bearing the risk of underestimation of treatment failures. Although logistically difficult, only longitudinal clinical trials with follow-up time over several months are appropriate for comparing the benefits and risks of different antimalarial drugs, especially in highly endemic areas . RecentlP. falciparum parasitaemia with detection of at least one msp1/msp2 amplicon of identical length within the 28-day follow-up period compared to amplicons found on day 0. This definition was chosen very stringent to identify all possible treatment failures. Remarkably, although most infections in our study area are multiclonal and the genetic diversity is high [With respect to the interpretation of PCR-controlled cure rates, it has to be considered, that the definition of reinfections is critical for the assessment of drug efficacy and effectiveness. In the study presented here, a recrudescence was defined as a recurrent  is high , some am is high . The aut is high .In accordance with these assumptions and in addition to the study mentioned above a recent comparative efficacy study in Ghanaian children showed lower parasitological and clinical failure rates in the ASAQ compared to the AL arm . AnotherThese results are in contrast to most comparative efficacy trials from Central and East Africa. In the Republic of Congo, in Uganda and Tanzania including Zanzibar, AL showed higher efficacy than ASAQ, especially a higher potency in preventing reinfections -13,26-28To explain the observed differences, one has to consider that not only the prevalences of molecular markers for ASAQ and AL drug resistance vary in different geographical regions ,32, but The lower effectiveness of AL in our trial compared to previous studies might be explained by unreliable or incorrect drug intake when unobserved. Furthermore, intestinal absorption of lumefantrine, the long-acting partner drug in AL, can be critically influenced by concomitant intake of fat-containing food . NeverthThe trial was discontinued due to a high frequency of anemia cases in our study population. Nevertheless, anemia occurred equally often in both study arms and haematologic recovery was similar and adequate in both arms. The safety profiles of the study drugs were good and our findings are in agreement with previous drug safety monitorings in Senegal ,37. It iASAQ and AL were both well tolerated and highly effective for treatment of Ghanaian children with uncomplicated falciparum malaria. Together with recent efficacy data, the more effective prevention of LCFs after unobserved ASAQ compared to AL treatment, affirms its appropriate role as national first-line therapy in Ghana. Monitoring of efficacy, safety, and drug resistance markers are essential to guide national treatment policies, however, further comparative effectiveness studies of ACTs are needed in the West African region.msp: merozoite surface protein; n: number; p: probability; PCR: polymerase chain reaction; P. f.: Plasmodium falciparum; PP: per-protocol; SAE: serious adverse event; SD: standard deviation; WHO: World Health Organization.ACPR: adequate clinical and parasitological response; ACT: artemisinin-based combination therapy; AE: adverse event; AL: artemether-lumefantrine; AQ: amodiaquine; AS: artesunate; ASAQ: artesunate plus amodiaquine;\u00b0C: degree Centigrade; CI: confidence interval; CQ: chloroquine; dl: deciliter; DNA: desoxyribonucleic acid; DSMB: data and safety monitoring board; ETF: early treatment failure; g: gram; Hb: haemoglobin; HIV: human immunodeficiency virus; HR: hazard ratio; ITT: intention-to-treat; LCF: late clinical failure; LPF: late parasitological failure; mg: milligram; \u03bcl: microliter; The authors declare that they have no competing interests.All authors participated in design, implementation, analysis or interpretation of the study. RK designed the study. RK, PK and JM were involved in all phases of the study and have full access to all the data in the study. RK takes responsibility for the integrity of the data and the accuracy of the data analysis. DA was the Principal Investigator and supervised the study. PK, MB, SAD, WNT and SAM were responsible for conduction of field studies and coordination of study procedures. Further acquisition and analysis of data was performed by WL, JV, MN, BK and BH. Analysis of data was led by RK, WL and JM. The manuscript was drafted by RK and PK and substantial input came from all investigators. All authors read and approved the final manuscript.Questions asked after completion of the 3-day treatment course to evaluate treatment acceptance of ASAQ and AL. Answers of parents/guardians were scored with 1 point (difficult/bad), 2 (fairly difficult/fair), 3 (easy/good), or 4 (very easy/very good) points, and were compared using Wilcoxon rank sum tests.Click here for file"}
+{"text": "The use of artemisinin-based combination therapy (ACT) at the community level has been advocated as a means to increase access to effective antimalarial medicines by high risk groups living in underserved areas, mainly in sub-Saharan Africa. This strategy has been shown to be feasible and acceptable to the community. However, the parasitological effectiveness of ACT when dispensed by community medicine distributors (CMDs) within the context of home management of malaria (HMM) and used unsupervised by caregivers at home has not been evaluated.In a sub-set of villages participating in a large-scale study on feasibility and acceptability of ACT use in areas of high malaria transmission in Ghana, Nigeria and Uganda, thick blood smears and blood spotted filter paper were prepared from finger prick blood samples collected from febrile children between six and 59 months of age reporting to trained CMDs for microscopy and PCR analysis. Presumptive antimalarial treatment with ACT  was then initiated. Repeat finger prick blood samples were obtained 28 days later for children who were parasitaemic at baseline. For children who were parasitaemic at follow-up, PCR analyses were undertaken to distinguish recrudescence from re-infection. The extent to which ACTs had been correctly administered was assessed through separate household interviews with caregivers having had a child with fever in the previous two weeks.Over a period of 12 months, a total of 1,740 children presenting with fever were enrolled across the study sites. Patent parasitaemia at baseline was present in 1,189 children (68.3%) and varied from 60.1% in Uganda to 71.1% in Ghana. A total of 606 children (51% of infected children) reported for a repeat test 28 days after treatment. The crude parasitological failure rate varied from 3.7% in Uganda (C.I. 1.2%\u20136.2%) to 41.8% in Nigeria (C.I. 35%\u201349%). The PCR adjusted parasitological cure rate was greater than 90% in all sites, varying from 90.9% in Nigeria (C.I. 86%\u201395%) to 97.2% in Uganda (C.I. 95%\u201399%). Reported adherence to correct treatment in terms of dose and duration varied from 81% in Uganda (C.I. 67%\u201395%) to 97% in Ghana (C.I. 95%\u201399%) with an average of 94% (C.I. 91%\u201397%).While follow-up rates were low, this study provides encouraging data on parasitological outcomes of children treated with ACT in the context of HMM and adds to the evidence base for HMM as a public health strategy as well as for scaling-up implementation of HMM with ACTs. Artemisinin-based combination therapy (ACT) is currently the universally recommended first-line therapy for uncomplicated malaria . NumerouThe use of ACT at the community level has been advocated as a means to increase access to effective antimalarial medicines by high risk groups living in underserved areas, mainly in sub-Saharan Africa ,7,8. ThiIn addition, poor adherence to the treatment schedule by both caregivers and CMDs, or a failure to provide adequate storage conditions to ensure drug stability in the community could reduce the efficacy of ACT and facilitate the emergence of drug resistance. There is thus a need for research to provide the evidence base to accompany deployment of ACT beyond the health facility level ,17 incluTo provide additional evidence to support the use of ACT at the community level, the UNICEF-UNDP-WORLD BANK-WHO Special Programme for Research and Training in Tropical Diseases (TDR) funded studies in Ghana, Nigeria and Uganda, representing different health systems and epidemiological settings. Report on the feasibility and acceptability of incorporating ACT in HMM has been published . In thisA multi-centre study of the use of ACT within the context of HMM in three African countries was carried out. Details of the study sites and the implementation of HMM in those sites have been previously reported .Three sites participated in the evaluation of parasitological cure rates: Ejisu-Juaben district in Ghana, Badeku and Ojoku/Ajia districts in Nigeria, and Bugiri district in Uganda. All three sites are rural areas in which malaria is hyper-endemic, with perennial transmission accompanied by seasonal peaks. The entomological inoculation rate (EIR) is estimated to be of the order of 500 infective bites per person per year in Ejisu-Juaben district  and BadeThe study involved follow-up of children between six to 59 months of age, whose caregivers reported to CMDs with a complaint of fever. The CMDs in the study areas had been trained on the signs and symptoms of both severe and uncomplicated malaria, to educate caregivers on the use of ACTs, on the preparation of thin and thick blood films and blood spots on filter paper as well as on proper storage of the specimens (filter papers and stained blood smear slides).At the time of the first visit of the febrile child and after informed consent had been obtained from the caregiver , CMDs prepared blood slides for microscopy and collected blood on filter paper [3 mm Whatman] for PCR analysis. In line with the currently recommended HMM strategy ), antimaBlood slides were read daily in local research laboratories and results returned to the CMDs the next day. Thereafter, CMDs provided the result to the mothers/caregivers. Mothers/caregivers of children with microscopically confirmed malaria were instructed to return for follow-up on day 28. On day 28 the caregivers were asked about symptoms and repeat peripheral blood films and blood spots on filter paper were obtained from their children. Caregivers of children who were free of patent parasitaemia were advised to take their children to the health centre or any health facility of their choice.In accordance with the national drug policies of the participating countries, a fixed combination of 20 mg of artemether and 120 mg of lumefantrine (AL) was used in Uganda and Nigeria, and a loose combination of artesunate and amodiaquine (AS/AQ) was used in Ghana. The recommended dose for AL in children between six and 35 months of age was one tablet twice daily for three days, and two tablets twice daily for three days in children between 36 and 59 months of age. Caregivers were advised to administer the drug after meals, preferably fatty food. For the AS/AQ combination, blister packs containing co-packed artesunate and amodiaquine tablets of 50 and 153 mgs respectively were used. For children aged 12 to 59 months the recommended dose of AS/AQ was one tablet of each drug once per day. However, for children aged 6 to 11 months, tablets had to be broken in half and repackaged to comply with the recommended AS/AQ dose in that age group. In all countries, antimalarial medicines for the study were provided by the Ministry of Health free of charge.A two-day training workshop was organized for the field supervisors to acquaint them with the intervention activities and the data collection tools. They were trained on how to make slides, blood spots on filter paper for PCR analysis and on proper storage of the specimens (filter papers and stained blood smear slides) in order to adequately supervise the CMDs. The mechanisms of drug distribution and supervision were also discussed.A total of 47 CMDs were recruited , following selection by the respective communities. A three-day workshop  was held for the CMDs in each study site. The training covered the clinical features of malaria, diagnosis, preparation of blood smears on slides, spotting blood on filter paper and treatment including dosage regimen. The CMDs were also instructed on the administration of consent forms, filling of data collection forms and criteria for referral. Refresher training on slide and filter paper preparation was performed during supervision by the field staff in the first month after initial training.Adherence to prescribed treatment schedules was assessed through cross-sectional surveys involving household interviews with caregivers of children with fever in the previous two weeks. All households in the study communities were visited and caregivers who met the inclusion criteria [has a child aged 6\u201359 months with fever in the two weeks preceding survey and have been resident in the community for at least six months] were interviewed. In cases were more than one caregiver in a household met the inclusion criteria, balloting was carried out to select the one to interview. During the interview, caregivers were questioned about the timing of treatment, the number of doses administered and the number of days over which drug was given. Used packs of drugs were inspected where available. The correctness of prescription by the CMDs was assessed by inspection of CMDs' registers during supervisory visits.Thick blood films and Whatman filter papers with blood spots prepared by CMDs from finger prick blood samples were collected from the CMDs within 24 hours. Blood films were stained with 10% Giemsa stain for 30 minutes and screened microscopically under X100 oil immersion lens using a light microscope. Intensity of infection was estimated by counting the number of asexual parasites in relation to 200 white blood cells. Parasite density was calculated assuming a mean leukocyte count of 8,000/\u03bcl blood. Ten percent of the slides were selected at random and re-read by an independent microscopist at each centre. Disagreements in readings were found in less than 5% of cases in all sites.Plasmodium falciparum infections at enrollment and who were parasitaemic at day 28 follow-up. Three highly polymorphic loci (block 1 of merozoite surface protein 1 (msp1), block 3 of merozoite surface protein 2 (msp2) and region II of glutamate rich protein (glurp)) of P. falciparum were amplified by nested PCR using oligonucleotide primers, reaction conditions and procedures described previously [Molecular genotyping for all sites was performed in Ibadan, Nigeria. Parasite genomic DNA was extracted from filter paper blood spots using the chelex extraction method . Genotypeviously . Ten micP. falciparum infection was characterized on the basis of fragment sizes of PCR products for each locus. If an isolate had just one allele at each of the three loci, the clone number was taken to be one. Infections were defined as polyclonal if more than one allele from one or more genes was apparent, as previously described [msp-1, msp-2 and glurp) in pre- and post-treatment samples. A lack of allelic identity at any of the three loci in pre- and post-treatment samples was considered to indicate a newly acquired infection/re-infection. An \"unresolved\" result was recorded when the parasite DNA could not be amplified. These samples were excluded from the analysis.Each escribed  and currescribed . BrieflyData were entered onto microcomputer and analysed using SPSS version 12.0  and STATA 9.0 The study was approved by the Ethics Review Committees in all participating countries and at the World Health Organization (WHO).Over a period of 12 months, a total of 1740 children presenting with fever were enrolled across the study sites. Of these, 1,189 (68.3%) had patent parasitaemia prior to treatment, of whom 606 (51%) reported for follow-up 28 days after treatment. The prevalence of parasitaemia prior to treatment was lower in Uganda than in Nigeria and Ghana . The proportion of children followed up at Day 28 was lowest in Ghana (34%) compared with 60% in Nigeria and 78% in Uganda. The crude parasitological failure rate was 41.8% in Nigeria (C.I. 35%\u201349%), 27.8% in Ghana (C.I. 22%\u201334%) and 3.7% in Uganda (C.I. 1.2%\u20136.2%). The PCR adjusted cure rate was over 90% in all sites, ranging from 91% in Nigeria and Ghana (C.I. 86%\u201395% and 87%\u201395% respectively) to 97% in Uganda (C.I. 95%\u201399%)  of children were treated correctly in terms of drug dose and duration of administration. The level of adherence was lower in Uganda  compared to Nigeria  and Ghana   malaria prevalence exceeded 50%, ranging from 61% to 71%. This finding has important implications for policy at local level, as the usefulness and cost-effectiveness of rapid diagnostic tests (RDT) for malaria may vary with the intensity of transmission  and withP. falciparum, a finding that had already been reported in literature [Secondly, it provides the first data on the cure rate that can be obtained with the use of ACT prescribed by CMDs and administered unsupervised by caregivers at home, a strategy that is advocated to improve access to effective treatment by underserved communities living in sub-Saharan Africa. The PCR adjusted cure rate varied among the sites from 91% in Nigeria to 97% in Uganda, rates which are all above the threshold of 90% below which WHO recommends changing antimalarial drug policy . Howeverterature .Incomplete adherence by caregivers to the correct dosage and treatment regimen when unsupervised, has been suggested as a possible cause of decreased effectiveness of ACT ,31 and aThis seems an unlikely explanation in this study. Reported adherence by caregivers was consistently high (over 80%) across sites. However, adherence was assessed based on caregivers' recall only and the possibility that some caregivers did not report adherence accurately should be borne in mind. Day 7 plasma lumefantrine level has been reported to be a useful predictor of the bioavailability and compliance with the artemether lumefantrine co-formulation ,32. Thusmsp1, msp2 and glurp alleles used to correct the failure rate have not been thoroughly investigated in the study areas. As in many other studies [Higher crude parasitological failure rates were observed in Nigeria and Ghana compared to Uganda, despite the former areas reporting higher proportions of correctly treated children than the latter. Although the rate of recrudescence was drastically reduced in both Nigeria and Ghana after genotyping, the overall pattern remained unchanged Table . This pa studies -35, thes studies .In interpreting these results, it is important to consider some limitations of the study methods. The protocol used to determine the parasitological efficacy of ACT is simple, with collection of blood samples done only at baseline and after 28 days of treatment. This has limited the capacity to detect the timing at which treatment failures occurred and to analyse the relative contribution of recrudescences and re-infections over time. Also, the follow-up procedure of children with malaria positive microscopy at baseline was mainly based on spontaneous reporting, which may have introduced a selection bias in the group of children that were tested at Day 28, with important variations in the proportion of children reporting for follow up across study sites. Finally, adherence was measured by questionnaires administered to caregivers, providing potential for recall bias, and no determination of lumefantrine blood levels was done. Most of these limitations derive from the fact that the study was carried out as part of a larger study to determine feasibility and acceptability of ACT use within HMM, including caregiver adherence and treatment coverage. In this context there was a need to minimize interference with spontaneous behaviours and real life conditions of use of antimalarials. A strict follow up protocol would have influenced the behaviour of the caregivers and thus limited the generalizability of the findings of the main study.The community effectiveness of antimalarial treatment is known to be influenced by a variety of factors beyond parasitological efficacy. These include access to effective treatment, quality of prescription and caregiver adherence ,36-38. HThe authors declare that they have no competing interests.All the authors except SC and KM conceived the study; IA, ENB, and FB were principal investigators for their respective country's study site and together with CH, CF, GOG, BY, and SB participated in the research design and supervised data collection from the field. CH and DY were involved with the PCR analysis. BY, KM and SC contributed to data analysis and interpretation. FP, the WHO/TDR HMM research program manager monitored the three sites. All authors contributed to the draft. All authors read and approved the final manuscript."}
+{"text": "Artemisinin was first identified by Chinese researchers as the active anti-malarial constituent of A. annua and its derivatives were found to be the most potent of all anti-malarial drugs. Artemether acts rapidly, reducing the infecting parasite biomass by approximately 10,000-fold per asexual life cycle. Lumefantrine, the other active constituent of AL, acts over a longer period to eliminate the residual 100-100,000 parasites that remain after artemether is cleared from the body and thus minimizes the risk of recrudescence. The two agents have different modes of action and act at different points in the parasite life cycle and show a synergistic action against Plasmodium falciparum in vitro. The combination of artemether and lumefantrine reduces the risk of resistance developing to either agent, and to date there are no reports of resistance to AL combined therapy in the malaria parasite that infects humans. Following a unique partnership agreement between Chinese authorities and Novartis, the manufacturer of AL, over 20 sponsored clinical studies have been undertaken in various malaria endemic regions and in travellers. These trials have involved more than 3,500 patients , and led to identification of a six-dose, three-day regimen as the optimal dosing strategy for AL in uncomplicated falciparum malaria. AL has consistently shown 28-day polymerase chain (PCR)-corrected cure rates greater than 95% in the evaluable population, meeting WHO recommendations. More recently, Novartis and the Medicines for Malaria Venture have worked in partnership to develop Coartem\u00ae Dispersible, a new formulation designed specifically to meet the specific needs of children with malaria. The dispersible tablets have shown similar high response rates to those observed with crushed standard tablets of AL. A partnership agreement between Novartis and WHO has seen over 250 million AL (Coartem\u00ae) treatments (75% for children) being distributed to malaria patients in developing countries without profit, supported by training programmes and educational resources.Artemisinin, from which the artemether component of Coartem Since 2002 it has been included in the WHO Model List of Essential Medicines, an index of priority drugs that guides purchasing decisions by United Nations agencies and many developing countries, and in 2007 it was added to the WHO Model List of Essential Medicines for Children. In 2004, Coartem\u00ae became the first fixed-dose artemisinin combination therapy to be pre-qualified by the WHO, and received approval from the Food and Drug Administration in the US in April 2009.Since it first received international licensing approval in 1999, CoartemArtemisia annua). The first description of the use of sweet wormwood dates from the year 168 BC, when the plant was mentioned in a tomb of a member of the Han dynasty for the treatment of 52 diseases; in the year 1086 it was recommended in a Chinese compendium of medicines for the management of fevers and chills [A. annua [The rapid-acting component of AL, artemether, is a derivative of artemisinin, a potent anti-malarial agent that is effective against multi-drug resistant parasites . ArtemisArtemether Figure  and otheIn vitro, artemether and lumefantrine show a synergistic action against P. falciparum [Understanding the Pharmacokinetics of Coartem\u00ae', these characteristics work synergistically: the rapidly-absorbed artemether and DHA achieve a fast reduction in parasite biomass and prompt symptomatic improvement, while the lumefantrine concentrations that persist in the blood after the three-day treatment course eliminate any remaining parasites to prevent recrudescence [Despite the potency of artemether, 100-100,000 residual parasites remain when the drug is used alone for a three-day treatment course, and as a result up to 10% of patients experience recrudescence ,12. It wlciparum . Additiolciparum ) and arelciparum . Lumefandescence .in vitro, the authors commented that artemisinin resistance does not seem to be a widespread epidemiologic phenomenon and in a subsequent publication by White et al pointed out that malaria parasites from Western Cambodia are no more resistant in vitro than parasites from other regions [The combination of artemether and lumefantrine would also be expected to restrict the development of drug resistance to either agent. Combination therapy is now widely accepted as the way forward to slow the rapid emergence and spread of resistance to malaria, and to increase the useful therapeutic life of anti-malarial drugs. Indeed, in 2006 WHO called for manufacturers to stop selling monotherapy artemisinin treatments in an attempt to prevent malaria parasites developing resistance to the drug . Concomi regions . No case regions , because regions . The sho regions . While t regions , and pip regions .\u00ae), in the first joint development programme of its type in Chinese history. Preclinical studies indicated that a 1:6 fixed combination of artemether and lumefantrine was well tolerated even at doses 10 times higher than those used in subsequent clinical trials [During the 1980s and 1990s, researchers at the Beijing Academy of Military Medical Sciences investigated combined administration of artemether with lumefantrine. From 1994 onwards, this work was undertaken in collaboration with Novartis, the manufacturer of AL The clinical efficacy of artemether/lumefantrine  [Until recently, there was no WHO-endorsed paediatric formulation for the treatment of malaria. Working in partnership, Novartis and the Medicines for Malaria Venture (MMV) have developed a formulation of AL tailored to the needs of children with patients ,25,29, bdetails) .\u00ae) available without profit for distribution through WHO in malaria-endemic developing countries. More than 250 million treatments have been provided through this without-profit arrangement, 75% of which were for children. In 2008 alone, 74 million treatments were distributed. This unexpected demand for AL has necessitated a dramatic growth in commercial farming of high-quality A. annua plants in Kenya and China, as well as the establishment of extensive new production facilities in China, Africa and the US. In addition, the widespread adoption of AL has been supported by a series of training programmes and educational materials for health professionals and patients or carers, to help ensure optimal prescribing and adherence to the AL regimen.Following a ten-year agreement with WHO in 2001, the manufacturer Novartis made AL \u00ae treatments have now been delivered. A long series of clinical trials has assessed the efficacy, safety and pharmacokinetics of AL - and more recently the dispersible formulation of AL - in a wide range of geographic regions and patient types, achieving cure rates higher than 95% among evaluable patients, with good tolerability. Ongoing investigations continue to ensure optimal safety and to identify the best routes for prescribing and distribution of AL, while AL continues to be provided in developing countries through non-profit production from Novartis with distribution via WHO.AL has had a remarkable journey to the clinic, from an ancient Chinese herbal remedy to a modern medicine that meets the most stringent drug approval criteria, and over 250 million CoartemThe author would like to acknowledge that Novartis Pharma AG sponsored this supplement. However, the author does not work for, or represent in any way, Novartis Pharma AG.The author met International Committee of Medical Journal Editors criteria for authorship."}
+{"text": "Sulfadoxine-pyrimethamine was a common first line drug therapy to treat uncomplicated falciparum malaria, but increasing therapeutic failures associated with the development of significant levels of resistance worldwide has prompted change to alternative treatment regimes in many national malaria control programs.in vivo therapeutic efficacy trial of sulfadoxine-pyrimethamine at two locations in the Peruvian Amazon enrolling 99 patients of which, 86 patients completed the protocol specified 28 day follow up. Our objective was to correlate the presence of polymorphisms in P. falciparum dihydrofolate reductase and dihydropteroate synthase to in vitro parasite susceptibility to sulfadoxine and pyrimethamine and to in vivo treatment outcomes. Inhibitory concentration 50 values of isolates increased with numbers of mutations  and septuplet (BR/51I/108N/164L and 437G/540E/581G) with geometric means of 76 nM (35\u2013166 nM), 582 nM (49-6890- nM) and 4909 (3575\u20136741 nM) nM for sulfadoxine and 33 nM (22\u201351 nM), 81 nM (19\u2013345 nM), and 215 nM (176\u2013262 nM) for pyrimethamine. A single mutation present in the isolate obtained at the time of enrollment from either dihydrofolate reductase (164L) or dihydropteroate synthase (540E) predicted treatment failure as well as any other single gene alone or in combination. Patients with the dihydrofolate reductase 164L mutation were 3.6 times as likely to be treatment failures  while patients with the dihydropteroate synthase 540E were 2.6 times as likely to fail treatment (96.7% (540E) vs 37.5% (K540); relative risk\u200a=\u200a2.58; 95% CI: 1.88 \u2013 3.73). Patients with both dihydrofolate reductase 164L and dihydropteroate synthase 540E mutations were 4.1 times as likely to be treatment failures  compared to patients having both wild forms (I164 and K540).We conducted an dihydrofolate reductase 164L or dihydropteroate synthase 540E.In this part of the Amazon basin, it may be possible to predict treatment failure with sulfadoxine-pyrimethamine equally well by determination of either of the single mutations NCT00951106ClinicalTrials.gov NCT00951106  P. falciparum malaria declined in the late 1980s and early 1990s worldwide, sulfadoxine-pyrimethamine  was introduced as a replacement first line treatment. SP is an inexpensive fixed dose combination tablet which is well tolerated, highly efficacious, and can be administered in a single dose thus insuring compliance, making this drug ideal for first line therapy for uncomplicated malaria. However, drug resistance can occur rapidly As the therapeutic efficacy of chloroquine for treatment of uncomplicated de novo folic acid synthesis Pfdhfr followed by subsequent mutations at 50R, 51I, 59R and 164L Pfdhps, leading to resistance to sulfadoxine. Selection of antimalarial therapies that affect similar pathways within the host cell and malaria parasite is cause for concern but in the case of SDX and PYR, they appear to have little toxic effects on host cell survivability In vivo drug resistance in Peru was shown previously to be highly correlated with the presence of the DHFR haplotype 108N/51I/164L and DHPS haplotype 437G/540E/581G in vivo resistance The mode of action of SP is well understood in vitro drug susceptibility values obtained from ex vivo parasites de novo folate synthesis since most cell culture medias contain folic acid and would necessitate the use of folate free media, which is costly There are few published reports correlating patient outcomes following treatment with SP and in vivo and in vitro drug susceptibility of the same parasite isolates are needed to demonstrate key parasite specific factors that contribute to observed outcomes.Favorable treatment outcomes to antimalarials, including SP, are dependent on host immune responses and pharmacodynamics in vitro drug susceptibility levels of sulphadoxine and pyrimethamine and treatment outcome. In vivo outcomes for one of these trials have been previously reported The purpose of the present study was to evaluate the therapeutic efficacy of SP in two locations in the Amazon rainforest region of Peru, and to correlate the presence of molecular markers associated with drug resistance and the Bolivia repeat sequence with PfDHFR and PfDHPS and how they relate to clinical outcomes are summarized in We successfully genotyped all isolates collected during the enrollment process. The results of the genotyping produced three different two-locus haplotype for the DHFR/DHPS loci: single mutant (108N), sextuplet  or septuplet (BR/51I/108N/164L and 437G/540E/581G) with an overall frequency of 44%, 21% and 35%, respectively. No other two-locus haplotypes were identified. Genotypes of Of 159 patients assessed for eligibility, 99 were enrolled in the study  and 86 pPfmsp2, and DNA sequence identify for Pfmsp1 block 2, when compared to isolates collected on day of enrollment, providing evidence that treatment failure was due to recrudescence not reinfection. At least nine genotypes of Pfmsp2 as determined by PCR RFLP and seven genotypes of Pfmsp1 block 2 as determined by DNA sequencing have been shown previously to be circulating in this region of Peru Isolates collected on the day of failure from all patients that failed treatment had 100% identical RFLP patterns for In addition, age, gender, weight, number of episodes of malaria in last year, species of previous malaria case and subsequent treatment of previous malaria case were not associated with treatment outcomes (data not shown). Additionally, serum was collected on the day of failure from 12 patients classified as ETF (PCT\u200a=\u200a2\u20133 days) to determine drug levels of both SDX and PYR in vitro culture and available for in vitro drug susceptibility testing. Twenty of the isolates were from patients classified as having ACPR (44%), 14 were from the LPF group (30%), 11 (24%) were from the ETF group and 1 was from the LCF group (2%). 50 values of SDX and PYR according to clinical outcome 50 value for SDX was 62 nM (95% CI: 28\u2013139 nM) and 29 nM (95% CI: 18\u201347 nM) for PYR. For persons who were treatment failures the geometric mean IC50 values were 1892 nM (LPF), 5701 nM (LCF) and 5104 nM (ETF) for SDX and 145 nM (LPF), 156 nM (LCF) and 231 nM (ETF) for PYR  and one isolate from a patient classified as LPF had FIC values indicative of synergy (<0.6). The remaining parasites from the LPF, LCF, ETF groups and those classified as having ACPR with IC50 values greater than 85 nM had FIC values indicative of an additive effect between SDX and PYR (0.6\u20131.5). When FIC values were grouped based on the number of polymorphism , the majority of the parasites with a single mutation showed synergy between the drug combination while nearly all those containing the sextuplet and septuplet haplotypes showed addition . Studies from Africa have shown that the triple mutant in DHFR  is useful in predicting treatment failures but the widespread presence of 108N and the absence of 59R in Peru prevents the use of this haplotype combination for this means P. falciparum with the 50R mutation but its presence has since been documented in other South American countries In Peru, the presence of the septuplet haplotype (BR/51I/108N/164L and 437G/540E/581G) was a near absolute predictor of treatment failure. A limitation of the current study was the presence of only three two-locus haplotypes, making it impossible to discern the effects other haplotype combinations had on clinical outcomes in this region. In lieu of genotyping the entire DHFR and DHPS, we report that the presence of mutations at 164L of DHFR and 540E of P. falciparum and 2/6 reporting cases of P. vivax malaria. Two of the four reporting previous P. falciparum cases, reported receiving treatment with SP as first line therapy.We found that not all patients harboring isolates with multiple mutations failed therapy, which could be due to the presence of some degree of acquired immunity as shown in a patient populations living in highly endemic regions P. falciparum isolates collected in South America Pfdhfr and Pfdhps loci show that parasites from South America with mutations conferring mid- and high-level resistance to SP have a common origin It has been shown that in vitro drug susceptibility levels where the clinical outcomes are also known 50 values at or above 100 nM considered to be resistant 50 geometric mean for PYR of 33 nM, which is lower than that reported previously in vitro sensitive levels for PYR even though they have the 108N mutation, provides insight into the significance of this mutation and its usefulness at predicting in vivo efficacy in this region of the Amazon. The relationship between in vitro and in vivo resistance to SDX/PYR showed that resistance to SDX is directly related in most cases to treatment failure. This data is supported by the in vitro drug interaction studies where, as a group, the combination of SDX/PYR was synergistic in most of the parasites that were sensitive to therapy, whereas the drug combination was additive in all of the ETF/LPF/LCF outcomes  and by the Universidad Peruana Cayetano Heredia Ethics Committee report # CAR-017-DUIICT-99, under Protocol # 719. The protocol for this trial and supporting CONSORT checklist are available as supporting information; see A single in vivo drug efficacy study followed recommendations of the World Health Organization P. falciparum asexual monoinfections, a parasite density greater than 500 parasites/\u00b5L but less than 200 parasites per oil immersion field , oral temperature \u226538.5\u00b0C (101\u00b0F) and/or a history of fever in the last 72 hours, and available and willing to return for follow-up were consented and enrolled in the trial using an IRB approved written consent form. In a subset of enrolled patients, gametocyte densities were determined on days 0, 2, 3, 4, 7, 14, 21 and 28 by counting sexual parasites per 200 WBCs. Patients that exhibited signs of severe malaria such as not being able to drink or breast feed, repeated vomiting, convulsions, lethargic or unconscious state, unable to sit or stand up. Respiratory distress or jaundice were not enrolled or followed clinically in this study but were referred to the local clinical care provides for hospitalization and treatment. Children under the age of 6 months were referred to the health clinic for diagnosis and treatment.The procedure used for the \u00ae; Roche S.A., Basel, Switzerland) was administered in a single oral dose equivalent to 1.25 mg/kg of pyrimethamine (up to a max of 75 mg) on Day 0 and were then followed with clinical histories, physical examinations, oral temperature measurements, and thick blood smears on days 1, 2, 3, 4, 7, 14, 21 and 28 to verify response. Patients that failed treatment with SP were treated on day of failure with a combination of quinine and tetracycline, quinine and chloroquine or quinine, tetracycline and primaquine.Sulfadoxine-pyrimethamine , and sensitive CDC/Sierra Leone I (D6), Fresh clinical et al., 50's) were determined for each drug alone and for fixed concentrations ratios by non-linear regression software\u2013NFIT . IC50 values at or above 100 nM were considered indicative of resistance for both pyrimethamine and sulfadoxine 50's were used to calculate 50% fractional inhibitory concentrations (FIC50's) singly and in combination as described by Berenbaum et al.50's were expressed with the following equation FIC Drug A\u200a=\u200aIC50A(B)/IC50A, where IC50A (B) is the 50% inhibitory concentration of drug A in the presence of drug B. The sum of FICs (\u2211FICs) of drug A and B at 1\u22361 concentration ratio were used to determine the interaction of S/P in vitro. Isobolograms from a limited number of isolates defined the drug-drug interaction as follows: interactions were classified as synergism with sum FICs (\u03a3FICs) \u22640.6, as additive with \u03a3FICs 0.61\u20131.5 and antagonistic with \u03a3FICs >1.5 The drug susceptibility test was based on the method described by Desjardins, P. falciparum genomic DNA was used for a PCR reaction using Pfdhfr and Pfdhps specific primers Pfdhfr and S436A and A437G, K540E, A581G and A613T/S in PfdhpsParasite DNA was extracted from 200 \u00b5L of whole blood collected on the day of enrollment from a three ml Vacutainer (Becton Dickson) with EDTA using QIAamp\u00ae DNA blood mini kit  according to manufacturer's instructions. Five \u00b5l of 50 values, parasite counts). Due to frequent travel for work, and the uncertainty where the cases of malaria were contracted, treatment outcomes from each site were combined. Unpaired t-tests (or one way ANOVA) were used to compare mean values in two (or > two) independent groups. Drug IC50 values and parasite counts were log transformed (base10) before analysis. Fisher's exact test was used to compare two independent proportions. McNemar's was be used to compare two dependent (paired) proportions. Multivariate logistic regression was used to help assess statistically significant predictors of treatment success/failure For single categorical predictor variables the relative risk (RR) for treatment failure and corresponding 95% confidence intervals (CIs) were computed. In all cases the reference group for assessing increased risk of failure is the category with the assumed lowest risk of treatment failure. For example, in assessing risk in relation to the total number of possible haplotypes in Pfdhfr/dhps , the reference group is the category with a \u201csingle mutation\u201d. Because of the close agreement between the two WHO systems for assessing therapeutic outcomes only the results using the clinical outcomes as defined in the WHO 2003 system are reported. All reported p-values are two-sided and 95% CIs were computed for important comparisons (effect measures).Statistical analyses were carried out using Minitab (Vs 14) and StatXact (Vs 4.01). Mean or geometric mean values were used to summarize quantitative measurements , drug ICChecklist S1CONSORT Checklist(0.03 MB DOC)Click here for additional data file.Protocol S1Trial Protocol(7.16 MB PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria. Fixed-dose AS/AQ is new, but its safety and efficacy are hitherto untested.Artesunate (AS) plus amodiaquine (AQ) is one artemisinin-based combination (ACT) recommended by the WHO for treating P. falciparum-infected children from Burkina Faso aged 6 months to 5 years. Dosing was by age. Primary efficacy endpoint was Day (D) 28, PCR-corrected, parasitological cure rate. Recipients of rescue treatment were counted as failures and new infections as cured. Documented, common toxicity criteria (CTC) graded adverse events (AEs) defined safety.A randomized, open-label trial was conducted comparing the efficacy (non-inferiority design) and safety of fixed (F) dose AS (25 mg)/AQ (67.5 mg) to loose (L) AS (50 mg) + AQ (153 mg) in 750, Recruited and evaluable children numbered 750 (375/arm) and 682 (90.9%), respectively. There were 8 (AS/AQ) and 6 (AS+AQ) early treatment failures and one D7 failure (AS+AQ). Sixteen (AS/AQ) and 12 (AS+AQ) patients had recurrent parasitaemia . Fourteen patients per arm required rescue treatment for vomiting/spitting out study drugs. Efficacy rates were 92.1% in both arms: AS/AQ = 315/342 (95% CI: 88.7\u201394.7) vs. AS+AQ = 313/340 (95% CI: 88.6\u201394.7). Non-inferiority was demonstrated at two-sided \u03b1 = 0.05: \u0394 (AS+AQ \u2013 AS/AQ) = 0.0% (95% CI: -4.1% to 4.0%). D28, Kaplan Meier PCR-corrected cure rates  were similar: 93.7% (AS/AQ) vs. 93.2% (AS+AQ) \u0394 = -0.5 (95% CI -4.2 to 3.0%). By D2, both arms had rapid parasite  and fever  clearances.p = 0.59). One patient developed asymptomatic, CTC grade 4 hepatitis . Technical difficulties precluded the assessment and risk of neutropaenia for all patients.Both treatments were well tolerated. Drug-induced vomiting numbered 8/375 (2.1%) and 6/375 (1.6%) in the fixed and loose arms, respectively (P. falciparum malaria with continued safety monitoring.Fixed dose AS/AQ was efficacious and well tolerated. These data support the use of this new fixed dose combination for treating Current Controlled Trials ISRCTN07576538 Plasmodium falciparum has rendered many anti-malarial drugs ineffective with a consequential increase in childhood morbidity and mortality, especially in Africa ) assessments. Parasite density was determined by counting the number of asexual parasites per 200 leucocytes on a Giemsa-stained thick film and expressed as the number per \u03bcL, assuming a leucocyte count of 8,000 per \u03bcL. If up to 500 parasites had been counted before reaching 200 leucocytes, the counting process was stopped at the end of the last field. Gametocytes were counted and expressed as the number per 1,000 leucocytes (thick film). Routine haematology  based on a per protocol defined analysis population (see below). The criteria for treatment failure followed broadly those of the WHO : (i) sigmsp2), then merozoite surface protein 1 (msp1), and then glutamine-rich protein (glurp), according to a previously published method  or (ii) afebrile on D2 , and (ii) with gametocytes during follow up  aparasitaemic on D2 (F: 339/347 [97.7%] p = 0.61) to the percentage of patients reporting AEs in the AS+AQ group, 74.7% (280 of 375). None of these were probably or definitely related to the study treatment. The number of patients with at least one possibly related solicited AE was similar among the two groups (AS/AQ group: 22 of 375 [5.9%] vs. AS+AQ group: 21 of 375 [5.6%]). The most frequently solicited AEs were coughing, rhinitis, anorexia, diarrhoea and abdominal pain. In total, 14 of 750 (1.9%) patients were withdrawn from the study for drug-induced vomiting, 8 (2.1%) and 6 (1.6%) in the fixed and loose arms, respectively (p = 0.59). Of the 14 withdrawn, one received an AQ dose below the therapeutic range, seven received AQ doses within the therapeutic range, and six received AQ doses above the therapeutic range; doses in the six were (i) fixed: 16.9, 21.1 mg per kg per day, and (ii) loose: 15.3, 17, 19.1, and 23.5 mg per kg per day.Both drug regimens were well-tolerated. Through direct questioning, 76.3% (286 of 375) of patients in the AS/AQ group reported at least one AE over 28 days, which was similar  deaths. All SAEs occurred within the first three days of treatment except for one patient, who had gastroenteritis requiring rehydration in the hospital on D11. Four of these patients (1.1%) were in the AS/AQ group and five (1.3%) in the AS+AQ group (p = 0.74). In the AS/AQ group these were: (i) one death on D1 owing to severe malaria , (ii) two patients with convulsions  and (iii) one child with gastroenteritis. AS+AQ SAEs were: (i) one death on D1 owing to severe malaria (D0 parasitaemia 1651/\u03bcL), (ii) one patient with convulsions and anaemia secondary to severe malaria, (iii) one patient with severe prostration , and (iv) two patients with acute respiratory distress, consistent with malaria-induced metabolic acidosis or pneumonia. All SAEs were considered unrelated to the study drugs.p < 0.0001). The mean platelet counts rose from 179.2 to 337.6 \u00d7 109/L by D28 (p < 0.0001).Because the mean values and mean changes from baseline were very similar in both treatment groups, laboratory data from both arms have been combined. By D28, the mean haematocrit increased from 26.6 to 31.2%  broke down, resulting in the full blood count (FBC) being determined at a private laboratory, where the differential white cell counts (WCC) were determined manually in a total of 350 patients. The results are reported here for completeness, but are deemed to be inaccurate because of the clear difference in the rates of neutropaenia  and leucopaenia  between the two laboratories. The total number of children (2004 and 2005 combined) with D0 and D28 neutropaenia and D0 and D28 leukopaenia were: (i) 32 of 657 (4.9%) and 87 of 569 (15.3%), (ii) 22 of 674 (3.3%) and 34 of 569 (6.0%), respectively. Of these totals, 28 of 32 (87.5%), 85 of 87 (97.7%), 15 of 22 (68.2%) and 33 of 34 (97.1%) were reported in 2004 by the private laboratory. All children with reported neutropaenia (2004 or 2005) were well; two were mildly febrile and only required symptomatic treatment.During the first malaria season (2004), the Pentra 60p = 0.02) and D28 total bilirubin  values fell significantly. Mean serum ALT (32 vs. 29 IU/L) and creatinine (37 vs. 38 \u03bcmol per L) values changed little over time. Six of 529 (1.1%) patients with D28 ALT results had CTC grade 2 raised (>2.5 to 5 \u00d7 ULN) ALT concentrations (range 104 to 196 IU per L) and one 2-year-old child (0.19%) in the AS+AQ group had asymptomatic, CTC grade 4 hepatitis ). All seven children had normal ALT values at baseline, were clinically well and aparasitaemic with normal D28 total bilirubin values, but all had raised D28 AST (range 73 to 1052 IU per L).Compared to D0, the mean D28 AST  found no clinical resistance. Based on the results of this study, clinical resistance to AQ in children under five is probably low.The efficacy of AS and AQ depends on the level of background resistance to amodiaquine, which is also partially cross-resistant to chloroquine in vitro ,30. All in vitro . There aThe cure rate in this study compares well with AS+AQ in Angola, Sudan, the Democratic Republic of Congo, southern Senegal, Colombia, and Uganda, where cure rates exceeding 90% have been reported ,31-35. NConsistent with other trials, symptom and fever resolution were rapid. By D2, less than 3% of children were still parasitaemic and less than 4% were still febrile. Similarly, within one week, all but one child was asymptomatic. Gametocyte carriage at baseline was higher than the ~10% observed in an earlier trial in the same areas , reachedper se, strategies for reducing drug refusal should be explored. No child in the fixed combination group developed itching and, consistent with other studies (<1 to 2%), only two children (0.3%) reported itching in the loose combination group [AS+AQ was well-tolerated; only nine children suffered serious adverse events, which were all due to either the development of severe malaria or intercurrent illnesses. Two of the nine children died because of severe malaria, giving a case fatality rate of 0.27%. All children were assessed carefully on admission and the two children who died had low, baseline parasite counts. In such small children, a number may have been developing clinically silent features of severe malaria when first seen and their parasite counts may have been increasing despite the use of the rapidly acting artesunate. The parasitaemia threshold of 1,000/\u03bcL used is lower than the 2,000/\u03bcL recommended by the WHO for areas of intense malaria transmission and the parasite counts in children were consistent with many similar studies conducted in Africa. Nevertheless, as the use of ACT increases, trying to identify patients at risk of developing severe malaria may be a new avenue of research. Just over 2% of patients in this study suffered from AEs considered to be study drug related; most were due to drug-induced vomiting. A number of children persistently spat out their tablets, making drug administration challenging. Whilst this is not an AE on group ,16. Thison group -39.In this study, there was a downward trend in the mean AST and the total bilirubin, consistent with disease resolution, but mean ALT levels were stable over time, suggesting a small disease effect on this enzyme in these children. Using the D28 ALT values as a marker of possible AQ hepatocellular toxicity by study end, about 1% had moderate liver impairment and one child (0.19%) had severe biochemical hepatitis with liver enzymes of circa 1,000 IU per L. All of these children were well and none were jaundiced. These liver enzyme changes are also consistent with an intercurrent, viral-induced hepatitis, but serology was not measured. These findings have implications for the deployment of amodiaquine because repeat dose AQ as prophylaxis has caused severe hepatitis, sometimes in conjunction with neutropaenia . AQ will\u00ae machine. An audit of the external laboratory and a quality control of the differential white cell counts for a sample of the slides were carried out. It was concluded that the manual counts were not reliable and that the haematology data should be presented separately for 2004 and 2005. Consequently, the frequency of neutropaenia in children recruited in 2004 cannot be determined accurately. Taking the automated derived data from CNRFP, the rate of D28 neutropaenia  was low . Nevertheless, continued monitoring for neutropaenia and abnormal liver function in studies involving amodiaquine should be done to document their risks of occurrence and their clinical features and significance.The current study reported a total of 87 patients (15.3%) with neutrophil counts <1,000 per \u03bcL at D28 compared to 32 patients (4.9%) at D0. However, there was a significant difference between the differential white cell counts measured in 2004 and 2005. In 2004, the differential white cell count was done manually but after 2004, the count was done using a new Pentra 60P. falciparum malaria and should be assessed in other malaria-endemic countries.This new fixed-dose combination, dosed by age, was efficacious and well-tolerated. It is a good option for treating acute uncomplicated Initiative (DNDi). JRK was the DNDi project manager and is a co-author on this paper, but had no direct part in the design or implementation of the trial. PO is a current staff member of the WHO and is required to declare that the opinions in this paper are not to be interpreted as those of the WHO. WRJT was a WHO staff member at the time of study conduct, but is not compromised by this past association.The authors declare that they have no conflict of interest. This clinical study was funded by the Research Directorate General of the European Commission (Contract N\u00b0ICA4-CT-2002-10046) and the Drugs for Neglected Diseases BSS, WRJT, and PO designed the study and developed the protocol. BSS, ABT, AG, AO, and AKT executed the study. WRJT and JRK oversaw and coordinated the study. CCM analysed the data. BSS, WRJT, JRK, CCM, and PO interpreted the data. BSS, WRJT, and CCM wrote the first draft of the paper. All co-authors have seen and approved the manuscript."}
+{"text": "Plasmodium vivax has a dormant hepatic stage, called the hypnozoite, which can cause relapse months after the initial attack. For 50 years, primaquine has been used as a hypnozoitocide to radically cure P. vivax infection, but major concerns remain regarding the side-effects of the drug and adherence to the 14-day regimen. This study examined the effectiveness of using the directly-observed therapy (DOT) method for the radical treatment of P. vivax malaria infection, to prevent reappearance of the parasite within the 90-day follow-up period. Other potential risk factors for the reappearance of P. vivax were also explored.P. vivax by microscopy, were recruited. All patients were treated with the national standard regimen of chloroquine for three days followed by primaquine for 14 days. Patients were randomized to receive DOT or self-administered therapy (SAT). All patients were followed for three months to check for any reappearance of P. vivax.A randomized trial was conducted from May 2007 to January 2009 in a low malaria transmission area along the Thai-Myanmar border. Patients aged \u2265 3 years diagnosed with p = 0.021). Factors related to the reappearance of vivax malaria included inadequate total primaquine dosage received (< 2.75 mg/kg), duration of fever \u2264 2 days before initiation of treatment, parasite count on admission \u2265 10,000/\u00b5l, multiple P. vivax-genotype infection, and presence of P. falciparum infection during the follow-up period.Of the 216 patients enrolled, 109 were randomized to DOT and 107 to SAT. All patients recovered without serious adverse effects. The vivax reappearance rate was significantly lower in the DOT group than the SAT group (3.4/10,000 person-days vs. 13.5/10,000 person-days, P. vivax malaria infection. Implementation of DOT reduces the reappearance rate of the parasite, and may subsequently decrease P. vivax transmission in the area.Adherence to the 14-day primaquine regimen is important for the radical cure of  Plasmodium vivax is most prevalent [Plasmodium falciparum, P. vivax infection rarely develops into complicated malaria and death is unusual. However, P. vivax has a dormant stage (the hypnozoite) that persists in the human liver and may cause relapse weeks, months, or even years later. Therefore, P. vivax infection is considered to have greater impact on morbidity than mortality, resulting in significant social and economic burdens [P. vivax transmission, because gametocytes appear almost simultaneously with schizonts.Globally, over 3 billion people live in areas at risk of malaria infection; about one billion of these live in countries outside Africa, where malaria transmission is low and revalent ,2. Unlik burdens . MoreoveP. falciparum and P. vivax. The incidence ratio of P. falciparum malaria to P. vivax malaria is almost 1:1 [P. vivax parasites in the bloodstream and hypnozoites in the liver. Although chloroquine resistance has been reported in Papua New Guinea and Indonesia [In Thailand, most malaria cases are caused by most 1:1 . A threendonesia , it is sndonesia -8. Occasndonesia -15. Howendonesia , and abondonesia . Relapsendonesia .P. vivax, including inadequate primaquine dosage [P. vivax hypnozoites requires 14 days of primaquine therapy, adherence to the drug regimen may greatly affect the prevention of relapse. Unfortunately, the effect of patient adherence on 14-day primaquine treatment, and its relation to preventing parasite reappearance, is not well-documented.A number of factors are reportedly associated with relapse, or the reappearance of e dosage ,14, highe dosage , age, ane dosage ,14,19,20P. vivax in a low malaria transmission area along the Thai-Myanmar border. Other potential risk factors for the reappearance of P. vivax after radical treatment were also explored.Directly-observed therapy (DOT) is an effective strategy to ensure patient adherence to long-term chemotherapy. Currently, DOT is the main strategy for treating tuberculosis (TB) in many countries and its success has been widely reported . HoweverThe study was conducted between May 2007 and January 2009 at the Rajanagarindra Tropical Disease International Centre (RTIC), Tanaosri Subdistrict, Suan Phung District, Ratchaburi Province, Thailand. The area is situated 100 km west of Bangkok near the Myanmar border. The study area comprised seven hamlets with about 3,500 residents. The population in the area is composed of four ethnic groups, Karen (85%), Thai (14%), Mon and Burmese (1%). For a decade, the RTIC clinic has provided free malaria diagnosis and treatment for the local community. Patients from other hamlets also routinely seek malaria treatment here.P. vivax infection were included in the study provided they or their guardians gave informed consent. Patients with microscopically confirmed mixed infections of P. vivax with other Plasmodium species, severe malaria, pregnant or lactating were excluded. G6PD deficient patients were included in this study, since the most common G6PD mutations in Thailand are G6PD Mahidol and G6PD Viangchan [Patients aged \u2265 3 years with microscopically-confirmed iangchan , which hiangchan . This stTo compare the effectiveness of directly observed therapy (DOT) with self-administered therapy (SAT), study participants were randomly assigned to either a DOT group or a SAT group using the block randomization method with a block size of four. Patients assigned to the SAT group, upon diagnosis, were given anti-malarials for self-administration with standard instructions in taking the drugs. Patients assigned to the DOT group were visited daily at home by RTIC staff until the full course was taken.P. vivax parasitaemia reappeared within three months, the daily primaquine dose was increased to twice the initial dose.All patients were treated with the national standard regimen for vivax malaria, as recommended by the Malaria Control Program of Thailand . G6PD screening is not required before giving the primaquine treatment. Chloroquine and primaquine doses were adjusted accordingly for patients under 14 years of age: for 8-13 years old, chloroquine 900 mg base over three days, and primaquine 10 mg daily for 14 days; for 3-7 years old, chloroquine 750 mg base over three days, and primaquine 5 mg daily for 14 days. If Baseline demographic and clinical data were gathered by interview. Finger-prick blood samples were collected on filter paper to examine for G6PD mutations and parasite genotype. All patients were followed for 90 days with Day 0 defined as the first day of drug administration.P. vivax and other Plasmodium parasitaemia. Patients were instructed to visit RTIC if they experience any symptoms during the follow-up period. At Days 7 and 14, patients in the SAT group were asked whether they took their anti-malarials as instructed. Upon microscopic confirmation of P. vivax reappearance, a finger-prick blood sample was taken to determine the parasite genotype for comparison against baseline genotype(s).During the follow-up period, patients from both groups were visited by RTIC staff at Days 7, 14, 28, 60, and 90, to examine for DNA was extracted from finger-prick blood samples on filter paper, using a QIAamp DNA Mini Kit according to the manufacturer's instructions .et al [2; for PCR-RFLP, 10 \u00b5l of the PCR product were digested with an appropriate restriction enzyme  in a total volume of 20 \u00b5l.The two most common G6PD mutations in the Karen population, G6PD Mahidol and G6PD Viangchan, were examined using polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis of the PCR products (PCR-RFLP), as described by Nuchprayoon et al  with slipvcs and pvmsp3-\u03b1 genes were used. Nested PCR and PCR-RFLP were carried out, following the previously described procedure [pvcs gene, five size variants: 650, 680, 700, 720, and 750 base pairs were coded as a, b, c, d, and e, respectively. VK type was labelled as VK210 or VK247 according to the results of PCR-RFLP. For the pvmsp3-\u03b1 gene, three size variants: 1,900, 1,500 and 1,100 base pairs, were labelled A, B, and C, respectively. For PCR-RFLP analysis by Hha I, digestion types were labelled as h1-h14 . Digestion types by Alu I were labelled as a1-a24 .The rocedure ,26. For pvcs gene and the pvmsp3-\u03b1 gene. It was defined as the minimum number of combinations of MOI at each locus. In each gene, MOI was estimated by the number of bands on a PCR product gel picture. In the case of the pvcs gene, even if only one band was observed on the PCR product picture, it was estimated that the MOI was two when the results of PCR-RFLP showed both VK types: VK210 and VK247.Multiplicity of infection (MOI) was estimated by genotyping results in two loci: the U tests, depending on the type and distribution of the data. Age was categorized into three groups, 3-7 years, 8-13 years, and \u226514 years, according to the age-adjusted primaquine regimen. The cut-off level of the total primaquine dose per body weight (2.75 mg/kg) was used to indicate inadequate dose (< 2.75 mg/kg), as reported in a previous study [P. vivax reappearance observed among patients in the SAT and DOT groups were calculated and compared. Cox's proportional hazard regression modelling was used to examine factors related to the first reappearance of P. vivax malaria parasitaemia. The analysis was performed using Stata 8.0 computer package .The baseline characteristics of the patients in the SAT and DOT groups were compared using chi-square, Student's t, and Mann-Whitney U test: p = 0.004 for continuous age variable, chi-square test: p = 0.001 for age groups); other baseline characteristics were comparable between DOT and SAT groups  or the DOT group (109 patients). Over half of the patients  were enrolled into the study during the high malaria transmission season in the study area (April to June). G6PD mutations were found in 22% of the patients . All mutations were G6PD Mahidol. Despite the random allocation, patients in the DOT group were younger than those in the SAT group (Mann-Whitney P. vivax parasitaemia during follow-up was observed in 15 patients (8%). The overall incidence rates of P. vivax parasitaemia reappearance  at Days 28, 60, and 90, were 1.67, 4.78, and 8.43, respectively. The median (min-max) time to reappearance of P. vivax parasitaemia was 68 (13-90) days.Of the 216 patients, 187 (87%) completed their 90-day follow-up period; 90 (83%) patients and 97 (91%) patients were in the DOT and SAT groups, respectively. Reappearance of P. vivax reappearance in the DOT group was significantly lower than the SAT group . G6PD mutation status, infected P. vivax genotype (VK type), gender, presence of gametocyte at baseline, and P. vivax acquisition month did not show statistically significant associations.In the univariate analysis, treatment group, total primaquine dose per bodyweight, parasite count on admission, and multiple ia Table . There wP. vivax was significantly higher in the SAT group, compared with the DOT group . Children aged 8-13 years had the lowest risk of P. vivax reappearance. Interestingly, the reappearance rate appeared to be about six times lower among patients who received late treatment, i.e. pre-treatment duration of fever \u22653 days, compared with early treatment . Inadequate total primaquine dose was also associated with reappearance as those who received a total primaquine dose < 2.75 mg/kg were about 10 times more likely to have a subsequent P. vivax attack than those who received a total primaquine dose \u2265 2.75 mg/kg. In addition, parasite-related factors including baseline parasite count \u2265 10,000 /\u00b5l, multiple P. vivax-genotype infection, and P. falciparum detected during follow-up were positively associated with risk of P. vivax reappearance , were included in the multivariate model Table . The reaP. vivax parasitaemia than those who took a complete14-day course (13% vs. 9%), although this difference was not statistically significant.Adherence to primaquine treatment was determined by interviewing patients in the SAT group (response rate = 94%) at Days 7 and 14. Fifteen of 101 patients (15%) reported that they have not taken the drug at least once during the 14 days of treatment. Patients who missed a primaquine dose or doses tended to have a higher risk of reappearance of p = 0.030), whereas those who came to the clinic > 2 days after the initial attack did not show any difference in treatment adherence for Weeks 1 or 2 (7% vs. 9%). Non-adherence was more likely to be reported by males and in children aged 8-13 years, although the differences were not statistically significant.Non-adherence during Week 1 of treatment was reported by 6% of patients, whereas 12% of patients reported that they missed at least one dose of primaquine during Week 2. The median number of missed doses was more than four times higher in the Week 2 of treatment than Week 1. Interestingly, patients who received the treatment within two days of an acute attack tended to non-adhere to treatment in Week 2, compared with Week 1 , and the pvmsp3-\u03b1 gene size of the P. vivax parasite in the primary attack and in the reappearance of parasitaemia did not reveal any significant difference.Among the patients who experienced reappearance of P. vivax hypnozoites, especially after symptoms have subsided, is a serious concern. In this study we found the non-adherence rate in treatment Week 2 was double compared to Week 1. The 15% non-adherence rate is probably an underestimate since some patients may not have reported missing a dose. Moreover, the home visit on Day 7 among the SAT group may increase the adherence rate among patients in that group, which further underestimates the reported non-adherence rate, compared with the real situation. The DOT method ensures that patients complete the full course of treatment. The findings of this study suggest that, compared with self-administration, DOT increases the likelihood of radical cure in P. vivax malaria infection. Patients with supervised therapy were about six times less likely to have P. vivax reappearance within the 90-day follow-up period. The protective effect of the DOT is likely to be larger than the estimate, if the underestimation of non-adherence rate in the SAT group was taken into account.Adherence to a 14-day primaquine regimen for the radical treatment of P. vivax parasitaemia. This indicates that an inadequate dose of primaquine during the primary attack may increase the risk of a subsequent P. vivax attack. In practice, weight-independent doses are used because of the difficulties of using weight-adjusted doses in the field. Therefore, total primaquine dose per body weight varies considerably; e.g., patients who weigh 60 kg and 80 kg receive 3.5 mg/kg and 2.6 mg/kg, respectively, adhering to the adult regimen of \"15 mg of primaquine daily for 14 days\". Treatment failure due to an inadequate dose may be misinterpreted as drug resistance. This misinterpretation may occur not only between individuals, but also between areas because average body weight differs between countries and ethnic groups. The average bodyweight of patients in this study aged \u2265 14 years was 52.7 kg compared to 67.3 kg in a study in Brazil [P. vivax is present. In addition, weight-based dosing schedules for primaquine should be more appropriate than the existing nationwide age-adjusted guideline for primaquine treatment due to a wide range of average body weight among different populations.This study also showed that total primaquine dose per body weight < 2.75 mg/kg was associated with reappearance of P. vivax parasitaemia. This may be explained by the relatively high dose of primaquine that children aged 8-13 years received compared with the other age groups . Even if these children skipped a dose once or twice, the total primaquine dose would still be sufficient to constitute a \"therapeutic dose\".Poor adherence was more likely to be reported by males or children aged 8-13 years. While children aged 8-13 years showed the highest rate of non-adherence to primaquine treatment, they had the lowest rate of reappearance of P. vivax-genotype infections were associated with increased risk of the reappearance of P. vivax parasitaemia, which may be the result of acquired genotype-specific host immunity [P. vivax reappearance; however, the conventional PCR used to analyse the genotype in this study was unable to identify the dominant genotype within an individual host. Using real-time PCR to determine the proportion of each genotype in the primary attack may help understand whether the dominant or the low-level genotype actually reappears.Higher multiple immunity . PatientP. vivax in this study was related to the level of parasitaemia on admission, consistent with a previous study from Brazil [P. vivax parasite is the Chesson strain, which can produce about equal numbers of hepatic schizonts and hypnozoites [The risk of the reappearance of m Brazil . In Thainozoites . This ranozoites . TherefoP. vivax. Patients who received early treatment (\u2264 2 days) were more likely to develop a repeat P. vivax attack. This finding is possibly explained by the relationship between the timing of primaquine administration and the growth and stabilization of the hypnozoites. Research has not yet confirmed exactly how primaquine works on hypnozoites. However, primaquine is thought to affect the parasite mitochondrial electron transport chain [P. vivax, and the size of hypnozoites becomes stable 7-15 days after infection [P. vivax is 12-17 days, some hypnozoites may not have stabilized yet during the first few days after the acute attack. Therefore, if primaquine administration is started earlier after an initial attack, the drug's effects may be less than expected because the hypnozoites are not \"dormant\" enough. The association between relapse rate and timing of treatment may be explained by the difference in the non-adherence rate between patients treated early and those who had treatment delay. In this study, non-adherence was found in 13% of patients who received late treatment, as compared with 17% of those with early treatment. However, this difference was not statistically significant.The duration of fever before initial treatment was significantly associated with the reappearance of rt chain . Becausert chain . The hypnfection -34. HoweP. falciparum parasitaemia during the follow-up period was found to be a risk factor. However, the sample size was too small to obtain reliable results. Further research is needed to confirm these findings.The appearance of pvcs and pvmsp3-\u03b1 gene loci were used to compare the parasite genotypes in the primary attack and the reappearance of parasitaemia. The results showed a similar genotype pattern in the primary attack and the reappearance of parasitaemia. No particular genotypes were found to be associated with reappearance. Recently, P. vivax genome sequencing has revealed some genes that are likely related to dormancy or differentiation [The ntiation , and mayntiation . Since tP. vivax malaria infection; DOT should be considered for improving compliance to 14-day primaquine treatment, since it reduces the risk of relapse. In areas where both tuberculosis and vivax malaria are prevalent, integrating DOT for P. vivax with existing DOT system for treatment of tuberculosis, where health workers had been trained for house visit to deliver drugs, should have a major impact on reducing disease transmission and on the financial burden of these two diseases. Inadequate dose of primaquine is also one of the most important factors for radical cure of P. vivax; weight-adjusted doses should be considered and the most practical way to apply weight-adjusted doses should be sought.This study supports the supposition that poor adherence to 14-day primaquine treatment can affect the effectiveness of radical therapy for The authors declare that they have no competing interests.RT, MI, JKo, JKa, SPuk and PS designed the study protocol; SPua, NT, and WM collected the data; RT and MI conducted the PCR analysis; RT, JKa and PS analysed and interpreted the data; RT, SL, and NJD drafted the manuscript. All authors read and approved the final manuscript."}
+{"text": "A study carried out in 2003\u20132005 in Southern Benin showed a day-28 sulphadoxine-pyrimethamine (SP) monotherapy failure rate greater than 40%, while for SP combined with artesunate (SP-AS) the failure rate was 5.3%. Such a large difference could be explained by the relatively short 28-day follow-up period, with a substantial number of recurrent infections possibly occurring after day 28. This paper reports the treatment outcome observed in the same study cohort beyond the initial 28-day follow-up.dhfr gene (59) and the dhps gene (437 and 540) point mutations related to SP resistance.After the 28-day follow-up, children treated with either chloroquine alone (CQ), SP or SP-AS, were visited at home twice a week until day 90 after treatment. A blood sample was collected if the child had fever (axillary temperature \u226537.5\u00b0C). Total clinical failure for each treatment group was estimated by combining all the early treatment failures and late clinical failures that occurred over the whole follow-up period, i.e. from day 0 up to day 90. Pre-treatment randomly selected blood samples were genotyped for the dhfr Arg-59 with the dhps Gly-437 mutant and the dhps 540 wild type (85.5%). The dhps 540 mutation could be found in only three (8.3%) samples.The PCR-corrected clinical failure at day 90 was significantly lower in the SP-AS group  . The most prevalent haplotype was dhps 540 mutation, by the rapid parasite clearance with artesunate and by the level of acquired immunity.Combining artesunate to SP dramatically increased the treatment efficacy, even when extending the follow-up to day 90 post-treatment, and despite the high percentage of failures following treatment with SP alone. Such a good performance may be explained by the low prevalence of the  In Benin, the current national anti-malarial drug policy was established in 2004 and selected two artemisinin-based combination therapies (ACT), artemether-lumefantrine (AL) and amodiaquine-artesunate (AQAS) as the first- and second-line treatment, respectively ,2. This dhfr) gene (Asn-108 + Ile-51 + Arg-59) with 2 (Gly-437 + Glu-540) in the dihydropteroate synthetase(dhps) gene [dhfr mutations are selected in a stepwise manner, with the Asn-108 mutation occurring first, followed by the Ile-51 and then the Arg-59 mutations. Therefore, the Arg-59 mutation is considered as a surrogate marker for the dhfr triple mutation [In Africa, SP treatment failure is strongly related to the combination of 3 mutations in the dihydrofolate reductase  and for later genotyping was collected. Similar procedures were carried out when children of the cohort attended the health facilities within the study area. Children with fever (axillary temperature \u226537.5\u00b0C), a  malaria , danger Besides the first three days of treatment , children were seen at scheduled visits at days three, seven, 14, 21 and 28 . The 28-Thin blood films were fixed with methanol and were stained, together with thick films, with Giemsa 10% for 10 minutes. Parasite density was determined according to the number of parasites per 200 white blood cells (WBC), and assuming a total WBC count of 8,000/\u03bcl. If gametocytes were seen, the gametocyte count was extended to 1,000 WBC. Slide reading was blinded to patients' identity and treatment allocation. Packed cell volume (PCV), measured by micro-haematocrit centrifugation, was assessed at day 0, 14 and 28.Parasite genotyping was done at day 0 and the day of recurrent parasitaemia from blood collected and dried on filter paper (Whatman filter paper grade 3). DNA was purified and genotyping done by nested PCR for variable blocks within the merozoite surface protein 1 and 2 (msp1 and msp2) as described previously ,19. A reOnly children having received either SP or SP-AS were included in this analysis. Patients in the CQ group were excluded since the objective was to estimate the prevalence of molecular markers related to SP resistance and explain the high efficacy in the SP-AS arm. Patients classified as ETF were also excluded because of the previously reported lack of association between SP resistance molecular markers and ETFs ,5.dhfr gene (59), and two for the dhps gene (437 and 540) were analysed. Genotyping was carried out by mutation-specific nested polymerase chain reaction (PCR) and/or restriction digestions as described elsewhere [dhfr and dhps codon was characterized as wild-type (no mutation present), mixed (both wild and mutant genotypes in the same infection), or pure mutant (only mutant genotypes detected).Genotyping was done on blood samples collected before treatment at day 0 in all patients with recurrent parasitaemia and in about half of randomly selected patients having had ACPR between day 0 to 28. Considering the stepwise selection of the triple mutation, one point mutation for the lsewhere . Each dhThe follow-up time after day 28 was arbitrarily fixed at day 90 after the initial treatment. However, between day 29 and 90, blood slides were done only on children with fever so that only LCF but not LPF could be identified. Therefore, in order to make the first (day 0 to 28) and the second (day 29 to 90) follow-up periods comparable, only children experiencing either ETF or LCF between day 0 and day 90 were considered as clinical treatment failures; total clinical failure (TCF), for each treatment group was estimated by combining all the ETF and LCF that occurred over the whole follow-up period, i.e. from day 0 up to day 90. In addition, the risk of evolving towards a clinical episode was estimated for the children who during the first 28-day follow-up experienced a LPF identified as a recrudescence by PCR genotyping.dhfr/dhps mutations, infections were defined as wild type when no mutation could be detected, single mutant when only the dhfr 59 mutation was present, double mutant when the dhfr 59 mutation was present with either the dhps 437 or the 540 mutations, and triple mutant when all three mutations were detected.Concerning the Per Protocol and Kaplan Meier survival analysis at 42, 63 and 90 day. In the Kaplan Meier survival analysis, each patient contributed to the analysis for the time s/he was followed up. When estimating TCF, data were censored for subjects who ended follow-up prior to day 90 and for new P. falciparum (PCR corrected) infections. In the survival analysis, clinical failure risks were described by Kaplan Meier estimates and compared between groups with a log-rank test. A p-value p < 0.05 was considered as statistically significant. Pair-wise comparisons of treatment efficacy at day 90 were made with a Cox proportional hazards model.Data were analysed with STATA version 10 . Descriptive statistics were used to summarize baseline values and demography data. Data not normally distributed were compared by Wilcoxon Rank sum test or Kruskal-Wallis analysis of variance. Categorical data were compared using the chi-square or the Fisher's exact test when required. The risk of clinical failure was estimated both by The study was approved by the Minister of Health of Benin, the Ethical Committee of the Facult\u00e9 des Sciences de la Sant\u00e9, Cotonou, Benin and by the Ethical Committee of the Institute of Tropical Medicine, Antwerp. Written informed consent was obtained from all children's parents or guardians.Per Protocol analysis, TCF (PCR uncorrected) at day 90 was 13.7% (10/73) in the SP-AS, 44.6% (29/65) in the SP alone, and 47.9% (34/71) in the CQ group. After PCR correction, these rates were 2.7% (2/73), 41.5% (27/65) and 42.3% (30/71), respectively  was high, both in the SP alone  and in the SP-AS  groups (Table dhps 540 mutation was low in the SP group (8.3%) while no mutation was found in the SP-AS group. In both treatment groups, no significant association between any individual mutation and treatment outcome was found. Even though the haplotype 59M437M540W (Arg-59 + Gly-437 + Glu-540) (85.5%) was the most represented, it was not associated to treatment failure (p = 1.0) (Table DNA could be amplified for the majority of blood samples 63/65) and one additional sample was not interpretable after genotyping. The prevalence of the /65 and oBy the Day 28, 38 children experienced a LPF confirmed to be a recrudescence by PCR genotyping  . Among tAmong the 7 patients with LPF by day 28 and classified as new infection by genotyping (in the estimation of treatment outcome considered as ACPR), 2 evolved towards LCF  . A three-day treatment with AS is an incomplete treatment when resistance to the partner drug is already high. However, good efficacy of the combination SP-AS, over 90% at day 28, has already been reported from some African sites with high SP resistance -24, whildhfr) and dihydropteroate synthase (dhps) genes linked to SP treatment failure [dhfr mutations 108, 51 and 59 have been related to pyrimethamine resistance [dhps mutations 437 and 540 to sulphadoxine resistance [dhfr gene probably occurring first and the dhps mutations following later [dhps 540 mutations was a much stronger predictor of clinical treatment failure than the dhfr 59 mutation [dhps 540 wild-type associated with the dhfr 59 and dhps 437 mutations was the most prevalent haplotype but it was not associated with SP treatment failure, possibly because of the lack of mutations in other codons, e.g. 51 in the dhfr gene. Treatment failure with SP associated to the dhps 437 mutation is lower than that with the dhps 540 mutation [dhps 540 mutation with the dhps 437 wild type were found, similar to other reports in which the occurrence of the dhps 540 mutation without the dhps 437 mutation was uncommon [dhps 540 mutation. Additional elements to be considered are the rapid effect of AS on parasite load [dhps mutations occur in a stepwise fashion, first the dhps 437 and then the dhps 540 mutations, a process driven by drug pressure, and that indeed the double mutant is associated with a higher risk of failure, the efficacy of SP-AS may rapidly decrease over time if this treatment or SP alone are commonly used.The prevalence of the point mutations in the dihydrofolate reductase . Experiences from several African countries [Combining AS to SP dramatically increased the treatment efficacy, even when extending the follow-up to day 90 post-treatment, and despite the high percentage of failures following treatment with SP alone. Such a good performance may be explained by several factors including the low prevalence of the ountries -42 indicountries ,44.The authors declare that they have no competing interests.AN contributed to the study design, coordination and supervision of the field work, data entry, cleaning and analysis, and paper writing. AE contributed to the study design, protocol writing, data analysis and review of the paper. DA contributed to data management and analysis. DB contributed to data management and analysis. CVO contributed to blood sample analysis and manuscript review. JM contributed to statistical analysis. MA contributed to the study design, coordination & supervision of the field work, and paper reviewing. MC contributed to study design and paper reviewing. AM contributed to the study design, data analysis and interpretation as well as manuscript review. UDA contributed to study design, protocol writing, data analysis, interpretation and supervision, and manuscript review."}
+{"text": "High levels of drug resistance to SP have been reported from north-eastern Tanzania associated with mutations in parasite genes. This study compared the in vivo WHO antimalarial efficacy protocol was used in 6 to 59 months old symptomatic children and a modified protocol used in 2 to 10 months old asymptomatic infants. Enrolment was stopped early (87 in the symptomatic and 25 in the asymptomatic studies) due to the high failure rate. Molecular markers were examined for recrudescence, re-infection and markers of drug resistance and a review of literature of studies looking for the 581G dhps mutation was carried out. In symptomatic children PCR-corrected early treatment failure was 38.8% (95% CI 26.8\u201350.8) and total failures by day 28 were 82.2% (95% CI 72.5\u201392.0). There was no significant difference in treatment failures between asymptomatic and symptomatic children. 96% of samples carried parasites with mutations at codons 51, 59 and 108 in the dhfr gene and 63% carried a double mutation at codons 437 and 540. 55% carried a third mutation with the addition of a mutation at codon 581 in the dhps gene. This triple: triple haplotype maybe associated with earlier treatment failure.An open label single arm (SP) standard 28 day In northern Tanzania SP is a failed drug for treatment and its utility for prophylaxis is doubtful. The study found a new combination of parasite mutations that maybe associated with increased and earlier failure.NCT00361114ClinicalTrials.gov  Plasmodium falciparum resistance to SP has been recorded in Muheza, north-eastern Tanzania since 1994in vivo efficacy of SP in clearing parasites  in order to understand the relationship between the clinical efficacy and the protective efficacy of SP-IPTiSulphadoxine-pyrimethamine (SP) is one of the most widely used antimalarials worldwide. It is used as first line treatment for uncomplicated malaria alone or in combination with other antimalarials, although it has been replaced with other antimalarials in Southeast Asia and sub-Sahran Africa because of high levels of resistance. SP is also recommended for use as Intermittent Preventive Treatment of malaria in pregnancy (IPTp)in vivo protocolin vivo efficacy of SP in both symptomatic 6\u201359 month old children and asymptomatic 2\u201310 month old children, the target group for IPTi.Traditionally, antimalarial drug efficacy is measured using the standard WHO 28 day dhfr genedhps genedhfr mutations  together with two dhps mutations  prior to treatment is a significant predictor of SP P. falciparum treatment failuredhfr gene became established in south east Asia twenty years ago and was found to be associated with a high level SP resistance, as well as resistance to chlorproguanil-dapsonedhfr and dhps genes with resistance to SP we studied, molecular markers in symptomatic children and asymptomatic infants who had parasitological failures.Parasite susceptibility to SP is influenced by mutations in two genes. Resistance to pyrimethamine is determined by point mutations at codons 16, 50, 51, 59, 108 and 164 of the The protocol for this trial and supporting CONSORT checklist are available as supporting information; see The study was conducted in Hale Health Centre, Tanga Region, situated 32 km north of Muheza where SP resistance was first observed in Tanzania. The district experiences perennial, holoendemic malaria although, in recent years, transmission appears to have declined substantially. The entomological inoculation rate from the neighbouring district of Muheza was 148 infectious bites per person per year in 2000P. falciparum only with an initial parasite density of between 2,000 and 200,000 asexual parasites per microliter, (5) absence of general danger signs  or other signs of severe and complicated falciparum malaria according to WHO definitions, (6) measured axillary temperature \u226537.5\u00b0C, (7) ability to attend stipulated follow-up visits, (8) informed consent provided by parent/guardian; (9) absence of history of hypersensitivity reactions to SP and (10) no prior antimalarial use in the preceding 2 weeks.All children between the ages 6 and 59 months who attended Hale Health Centre during July\u2013August 2006 with a fever or history of fever during the study period were screened for malaria using a rapid diagnostic test (RDT) . Children with a positive RDT had a thick blood smear read and those with a positive blood smear were referred to the study clinician. Study inclusion criteria were: (1) weight of \u22654.5 kgs, (2) not -enrolled in the IPTi trial, (3) absence of severe malnutrition (weight-for-height <3 standard deviations from the norm), (4) slide-confirmed infection with P. falciparum infection. From those consented for screening, finger prick blood was obtained for the rapid diagnostic test, thick and thin blood smear preparation and filter paper samples for molecular studies. Children who had a positive blood slide were further assessed for their eligibility for inclusion into the study and enrolled in the drug sensitivity study after obtaining an informed consent. The eligibility criteria were the same as for the study of symptomatic children except that there was no history of fever in the last 48 hours, that measured axillary temperature should be less than 37.5\u00b0C and that the presence of P. falciparum parasitaemia at any density was acceptable.Consent was obtained from caretakers of 2\u201310 month old infants who attended the Maternal Child Health (MCH) clinic for immunization or weighing, for screening for In order to detect a 15% difference in adequate parasite clearance by day 28 between symptomatic 6\u201359 month old children and asymptomatic 2\u201310 month old children with 80% power at the 5% significance level using a ratio of 2 symptomatic cases to 1 asymptomatic case, we estimated that 292 symptomatic children and 146 asymptomatic infants would be required.Children were treated with SP  by weight . The content and solubility of the SP tablets were confirmed by solubility testing and high performance liquid chromatography at the London School of Hygiene and Tropical Medicine. The study drugs achieved the expected concentrations of SP in solution when compared to controls  purchased in the UK. SP was given under observation by study staff and children were observed for at least 1 hour after treatment. If a child vomited within 30 minutes of receiving the drug, the full dose was repeated. if a child vomited between 30 minutes and an hour, half the dose was repeated. If a study child vomited the study medication twice, the study child was given rescue treatment and excluded from the study. Rescue treatment for uncomplicated malaria was artemether- lumefantrine  and for severe malaria was parenteral quinine.Children in the symptomatic study were seen at the clinic on days 1, 2, 3, 7, 14, 21, and 28 after treatment and home visits were made for those who failed to report. Parents were encouraged to bring any child who became ill between specified visits to the clinic where they were evaluated and treated by a study clinician thoughout the study period. Malaria blood films and filter paper samples were obtained from children in the symptomatic group at all active and passive follow-up time points. Blood samples were not collected on days 1, 2, 3, and 21 from the asymptomatic infants if the infant remained well. However, blood samples were collected at any time if the infant became symptomatic.The primary end point, parasitological failure by day 28 was defined as (1) development of danger signs or severe malaria, (2) parasitaemia on Day 2 that was higher than that on Day 0, (3) parasitaemia on Day 3 \u226525% of the count on Day 0, (4) parasitaemia on or after Day 4. Failures were further divided into early treatment failures (within day 3 post treatment), late clinical failures (recorded fever plus parasitaemia from day 4 to day 28 post treatment), and late parasitological failure (parasitaemia at day 14 or day 28 post treatment in the absence of fever). In the symptomatic study children with parasitaemia on or after day 4 were treated with rescue treatment if they became symptomatic or until they reached day 28 when all parasitaemic children were treated. In the asymptomatic study any child on or after day 4 with parasitaemia was treated with rescue treatment.dhfr, dhps, sulphadoxine, sulfadoxine, pyrimethamine, Fansidar, Africa, prevalence, malaria and resistance. To be included in the review, articles had to include analysis of codon 581 of the dhps gene in isolates collected from study sites in Africa.The literature search for reports of the A581G mutation up to October 2007 was done using the National Library of Medicine search engines, Pubmed and Medline. The following terms were included in the search queries: Blood smears were stained with 20% Giemsa for 20 minutes and read by two independent microscopists for speciation, and quantification. Parasite density was estimated by counting parasites against 200 White Blood Cells (WBC). A blood smear was considered negative if no asexual forms were seen after observing 500 WBC. Discordant results (33% difference in quantification or positive/negative results) were read by a third microscopist; agreement between any two micoscopists and the average parasite density were deemed to be the correct finding. Parasite counts were adjusted assuming a standard WBC of 8000 per microlitre.dhfr and dhps were PCR amplified and point mutations at codons 51, 59, 108 and 164 of the dhfr gene and codons 436, 437, 540, 581, and 613 of the dhps gene were genotyped using a dotblot methodology previously described by Pearce et alDNA was extracted from bloodspots dried on filter papers by soaking overnight in 1 mL. of 0.5% saponin-1\u00d7 phosphate buffered saline (PBS). The segment was then washed twice in 1 ml of PBS and boiled for 8 min in 100 \u00b5L PCR quality water with 50 \u00b5L 20% chelex suspension (pH 9.5). We tested for the presence of mutations in addition to those at codon 436, 437, 540, 581, and 613 of dhps by direct sequencing. PCR products were purified using ExoSAP-IT\u00ae . Cycle sequencing was performed using Applied Biosystems BigDye V 3.1 and samples loaded on the ABI-3730 capillary system. Sequence reads were checked by eye and edited using the Seqman . The presence of SNPs was confirmed by reads through both forward and reverse strands.Recrudescent and new infections were differentiated first by typing size and sequence of the highly polymorphic repeat region of MSP2Multiplicity of infection (MoI) was assessed by examining the numbers of alleles detected at MSP2 and pfPK2. Where the number of alleles at these two loci differed, the higher of the two values was used since this is the minimum number co-infecting genotypes which can explain the observed diversity.Data were double entered into an Access  data base and analyzed in STATA 9.0 . Crude and PCR-corrected rates (excluding new infections and indeterminate PCR) were estimated. A survival analysisa priori were included in the logistic regression model: age, parasite density, study population (asymptomatic or symptomatic) and molecular markers. In addition, any factor that was significant at the 10% level in the crude analysis was included in the model.Logistic regression was used to determine factors associated with treatment failures (cases) compared to non-failures (controls). The following variables identified dhps gene.Further survival and regression analyses were carried out in order to examine the relationship with time to failure and the presence of the mutation 581G in the One hundred and fifty, 6\u201359 month-old febrile children were screened for malaria in July\u2013August 2006 and 87 children who fulfilled the inclusion criteria were enrolled see . RecruitBetween October 2006 and June 2007, 926 asymptomatic 2\u201310 month-old infants were screened for malaria and 25 infants with parasitaemia were enrolled into the study arm. This study was also stopped early due to a high failure rate with one child progressing to severe disease within 3 days post treatment. Apart from this serious adverse event no other adverse events were reported. The trial profiles are shown in The therapeutic efficacy of SP by day 14 and 28 post treatment is shown in New infections were detected at all time periods in the study. In the symptomatic group 4, 5 and 10 new infections were detected between days 1\u20133, 4\u201314 and 15\u201328 respectively and in the asymptomatic group 2 new infections were detected between days 4 and 14. Molecular typing revealed a high rate of multiple clone infection both pre and post treatment, with between 1\u20134 MSP2 alleles and 1\u20138 pfPK2 alleles found in individual infections. dhfr and dhps in both groups and prevalence of haplotypes in dhfr and dhps. There were no mutations found at codon 164 in the dhfr gene but 54% of parasites in the symptomatic group carried the 581 G mutation in the dhps gene on enrollment day. The ratio of 581 G mutation to 581 A wild type increased from enrollment day to day of failure (0.78 and 0.93 respectively) but this change was not statistically significant . Direct sequencing confirmed the presence of mutations at codons 436, 437, 540 and 581 and did not reveal any additional mutations. In the symptomatic group 96% of parasites carried the triple dhfr mutant haplotype   and were excluded from further analysis. From samples from the symptomatic study a significant trend for increased and earlier failure was seen with those carrying the 581G mutation prior to treatment   showing reatment . However density  and not P. falciparum isolates have been tested for the 581G mutation. The 41 references which describe them are listed in supplementary online material . In IfakaraIn Africa as a whole the A581G mutation has been observed in only 14 of a total of 106 surveys. The surveys that had observed A581G mutation span nine countries namely EthiopiaP falciparum malaria is dangerous in the area where our study was done. SP had no effect on the course of illness in four cases, three in the symptomatic group and one in the asymptomatic group, who developed severe disease within 3 days of post treatment. The efficacy of SP resulted in similarly poor parasitologic clearance in 2\u201310 month old asymptomatic infants when compared with 6\u201359 month old symptomatic children. The WHO recommends changing first line treatment when day 28 failure rates exceed 10%The results of this study demonstrate clearly that use of SP for the treatment of The primary purpose of this study was to determine if SP was more efficacious in asymptomatic parasitaemic infants than in symptomatic children even though the drug was failing as a treatment for the latter group. This has been shown to be the case for pregnant women in areas of moderate SP resistance where SP may still be effective for treatment or prevention even though it fails when used for the treatment of symptomatic childrenWithin 8 years the level of SP resistance in the study area has increased at an alarming rate. The day 28 ACPR has decreased from 55% in 1999 The molecular work was limited by the number of clones in the samples and the sensitivity of the methods to detect masked clones. In the symptomatic group 4 new infections were detected in the first 3 days post-treatment. This is a highly unlikely result and is probably due to masked clones resistant to SP becoming dominant post treatment, a common criticism of PCR-correction. This problem was also likely to interfere with the interpretation of new infections when comparing the symptomatic and asymptomatic groups as the symptomatic group was more likely to have heavier parasite densities and more clones. Thus, in the symptomatic group a higher proportion of recrudescences would be wrongly called new infections. The number of clones detected in samples was high and in many cases these represented multiple clones recrudescing reflecting the high transmission and high prevalence of resistance genotypes circulating in the study area.in vivo efficacy studies of SP are unjustifiable in areas where the level of SP resistance is known to be high. In such situations, studies of molecular markers of resistance play a key role in measuring patterns of resistance as there is strong evidence that mutations in dhfr and dhps genes are associated with P. falciparum resistance to SP. Previous reports from Africa have demonstrated an association between moderate levels of resistance to SP and triple mutation in the dhfrdhps genesdhfr genedhps gene and there was some evidence that the presence of this mutation in parasites at the start of treatment was associated earlier treatment failure. Biological plausibility of this is backed by the findings that the 581G single mutant offers resistance in vitro and that similar mutations in bacteria produce resistance to sulfa drugsdhps triple is recognized in South America, where it is associated with SP treatment failuredhps double and now the more resistant dhps triple mutation. This early failure may prove to be pertinent in the case of IPTp. ter Kuile and colleaguesdhfr and double dhps mutants, it is possible that the duration of prophylaxis given by SP will be further reduced and this may affect the protective efficacy of SP preventative strategies. Further work is needed to determine the geographical spread of this mutant and if the presence of this mutation in combination with the dhfr triple and dhps double confers increased and earlier failure of cases treated with SP.Further References S1References collected by systematic review to produce maps of the prevalence of the 581 dhps Mutation in (0.04 MB DOC)Click here for additional data file.Checklist S1CONSORT Checklist(0.06 MB DOC)Click here for additional data file.Protocol S1These are the 2 study protocols which were approved by the IRBs(0.01 MB ZIP)Click here for additional data file."}
+{"text": "Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common inherited human enzyme defect. This deficiency provides some protection from clinical malaria, but it can also cause haemolysis after administration of drugs with oxidant properties.P. falciparum malaria was evaluated according to G6PD deficiency in a secondary analysis of an open-label, randomized clinical trial The safety of chlorproguanil-dapsone+artesunate (CD+A) and amodiaquine+sulphadoxine-pyrimethamine (AQ+SP) for the treatment of uncomplicated In the first 4 days following CD+A treatment, mean haematocrit declined on average 1.94% (95% CI 1.54 to 2.33) and 1.05% per day (95% CI 0.95 to 1.15) respectively in patients with G6PD deficiency and normal patients; a mean reduction of 1.3% per day was observed among patients who received AQ+SP regardless of G6PD status (95% CI 1.25 to 1.45). Patients with G6PD deficiency recipients of CD+A had significantly lower haematocrit than the other groups until day 7 (p\u200a=\u200a0.04). In total, 10 patients had severe post-treatment haemolysis requiring blood transfusion. Patients with G6PD deficiency showed a higher risk of severe anaemia following treatment with CD+A  or AQ+SP .CD+A showed a poor safety profile in individuals with G6PD deficiency most likely as a result of dapsone induced haemolysis. Screening for G6PD deficiency before drug administration of potentially pro-oxidants drugs, like dapsone-containing combinations, although seldom available, is necessary.NCT00461578ClinicalTrials.gov  Glucose-6-phosphate dehydrogenase (G6PD) plays a critical role in protecting red cells from oxidant haemolysis. G6PD allows regeneration of NADPH, which is essential for glutathione recycling and protection against oxidative damage. G6PD deficiency is the most common inherited human enzyme defect, present in more than 400 million people worldwide. The gene encoding G6PD is located on the X chromosome; G6PD-deficient hemizygous males and homozygous females are vulnerable to oxidant haemolysis, whereas heterozygous females have variable (milder) deficiency determined by the degree of X-chromosomal inactivation.P. falciparum malaria There are numerous polymorphisms of the G6PD gene. Approximately 200 variant alleles have been described and 140 mutations, or combination of mutations, have been identified Pneumocystic carinii pneumonia Acute haemolytic anaemia is the most frequent clinical manifestation associated with G6PD deficiency. This can be precipitated by foods or drugs with oxidant properties. The degree of drug-induced G6PD deficiency related haemolysis depends on a number of factors including the G6PD variant, the drug and dosage, and poorly characterised disease factors. Sulphonamides and sulphones are still widely used in tropical countries, and may cause haemolysis in patients with G6PD deficiency. Dapsone is used as a treatment of leprosy, some skin conditions, and more recently P. falciparum malaria in children In 2005\u20132006 an open label, randomised clinical trial was carried out in Rwanda to evaluate the safety and efficacy of artesunate+chlorproguanil-dapsone and amodiaquine+sulphadoxine-pyrimethamine for the treatment of uncomplicated We evaluated the safety profiles of these two antimalarial treatments according to G6PD deficiency.P. falciparum malaria in Rwanda. The trial was open label; 800 patients, aged 6\u201359 months, with uncomplicated malaria were randomised to receive AQ+SP (400) or CD+A (400). Patients were hospitalized for 4 days, so that appropriate management, including blood transfusion, could be performed and followed\u2013up weekly until day 28 after treatment. Clinical and parasitological outcomes were recorded according to WHO guidelines The drug trial has been described in detail elsewhere The initial protocol of the clinical study included an arm with CD alone, but this arm was discontinued because of the remarkably high number of failures observed at an early stage and thereafter patients were allocated to CD+A or AQ+SP only.CD+A: CD was administered orally at doses of 2.0 mg/kg chlorproguanil and 2.5 mg/kg dapsone once daily for 3 days using commercial Lapdap\u00ae paediatric tablets containing 15 mg of chlorproguanil hydrochloride and 18.75 mg of dapsone  together with artesunate 4 mg/kg/day for 3 days using 50 mg tablets .AQ+SP: AQ was administered orally at a dose of 10 mg/kg daily for 3 days and SP was co-administered at a dose of 25 mg/kg sulphadoxine plus 1.25 mg/kg pyrimethamine the first day. AQ+SP and artesunate were both provided by Sanofi-Aventis .Early Treatment Failure (ETF) if she/he had any of the following: i) danger signs or severe malaria on days 1, 2 or 3 with parasitaemia; ii) parasite density at day 2 greater than at day 0; iii) parasitaemia on day 3 with axillary temperature\u226537.5\u00b0C and iv) parasite density at day 3 equal to or greater than 25% of that at day 0 Serious Adverse Event was defined as any untoward medical occurrence that at any dose results in death, requires inpatient hospitalisation or prolongation of existing hospitalisation, results in persistent or significant disability/incapacity or is life-threatening.A patient was defined as an The clinical protocol of the original study was approved by the Ministry of Health of Rwanda, the ethical Committees of the London School of Hygiene and Tropical Medicine, London, UK, and the Prince Leopold Institute of Tropical Medicine, Antwerp, Belgium. As soon as the analyses of the blood samples were available, the families of children with G6PD deficiency were traced and informed by a medical doctor with a health worker. An explanatory health-card was given to the families which could be presented to hospital.5\u2032- CTG GCC AAG AAG AAG ATC TAC CC-3\u2032 and REV 5\u2032- GAG AAA ACG CAG CAG AGC ACA G 3\u2032. DNA (50\u2013100 ng) was amplified in a total reaction volume of 30 \u00b5l consisting of reaction buffer, 2 mM MgCl2, 0.1 \u00b5mol of each primer, 0.1 mM dNTPs each and 1 U of BioTaq DNA polymerase (Bioline). A touchdown program was used to prevent non-specific amplification; an initial denaturation step at 95\u00b0C for 5 min was followed by 14 cycles at 95\u00b0C for 40 s, 68.5\u00b0C for 40 s, and at 72\u00b0C for 40 s, with annealing temperature decreasing 0.5\u00b0C per cycle; the programme was completed by 23 additional cycles with annealing temperature 61\u00b0C and a final extension step at 72\u00b0C for 10 min. Amplified fragments were digested with NlaIII (NEBiolabs) restriction enzyme at 37\u00b0C for 4 hours and analysed with 2.5% MetaPhor (Cambrex) gel electrophoresis.DNA extraction from Whatman filter papers was performed using standard Chelex method Data were double-entered using Microsoft Access 2003 and validated using Epi Info 3.3.2 . All analyses were performed using STATA statistical analysis software package version 10.0 .Patients were categorised into four groups for comparison: AQ+SP G6PD (B) (without G6PD deficiency), AQ+SP G6PD (A-) (with G6PD deficiency), CD+A G6PD (B) (without G6PD deficiency) and CD+A G6PD (A-) (with G6PD deficiency). Homozygous females and hemizygous males were considered as patients with G6PD deficiency. We also conducted a sensitivity analysis including the heterozygous G6PD females in the group of patients with G6PD deficiency.Continuous data with a normal distribution were compared using ANOVA and the non-parametric Kruskal-Wallis test was used to analyse continuous data with a skewed distribution. Proportions were compared using chi-squared or Fisher's exact test.The overall fractional reduction in haematocrit was defined as the difference between the patient's lowest level of haematocrit and that at baseline  divided by the haematocrit at baseline. The fractional reduction from day 0 to day 3 was calculated in a similar manner.The differences in the rate of decline in haematocrit levels from day 0 to day 3 were assessed in a separate model from the rate of recovery from day 7 to day 28. For each model, a regression line was fit for each patient and differences between groups as well as within group variation were adjusted for using random effects. The models were also adjusted for parasitaemia and temperature at presentation. The Odds Ratios (OR) and Risk Ratios (RR) were calculated with 95% Confidence Intervals (95% CI) using a 2-sided Fisher's exact test.In the original study 792 children were studied of whom 702 were successfully genotyped for the G6PD variant A- (at position 202). For the remaining patients  samples were not received or DNA extraction was not successful. The total G6PD allelic frequency in the study population was 9.6% in the males  and 7.5% in the females , p\u200a=\u200a0.3 . The PCVBetween day 1 and 3 the mean PCV decreased in all four groups. After day 7, mean PCV increased steadily in all groups . The larThe mean fractional reduction (SD) from day 0 to day 3 was also highest in the CD+A G6PD (A-) group: 19.8% (12.5) compared with 12.0% (10.6) in the AQ+SP G6PD (B) group, 11.6% (9.34) in the AQ+SP G6PD (A-) group and 10.0% (9.26) in the CD+A G6PD (B) group (p<0.001).Between day 0 and day 3 compared with patients in the AQ+SP G6PD (B) group, those in the CD+A G6PD (A-) group had a significantly higher rate of decline in PCV, after adjustment for (log) parasitaemia and temperature at day 0 (p\u200a=\u200a0.04); whereas PCV levels declined at the same rate in the AQ+SP G6PD (A-) group and CD+A G6PD (B) group when compared with patients in the AQ+SP G6PD (B) group . In the first 4 days, haematocrit declined, on average, 1.3% per day for all patients who received AQ+SP treatment, regardless of G6PD status (95% CI 1.25 to 1.45). For patients in the CD+A G6PD (B) group, haematocrit levels declined, on average, 1.05% per day (95% CI 0.95 to 1.15) whereas in the CD+A G6PD (A-) group, the mean haematocrit decline was greater, 1.94% per day (95% CI 1.54 to 2.33).From day seven, the haematocrit increased steadily in all groups. The rate of increase per day was similar (0.24%) for all patients treated with AQ+SP regardless of G6PD status (95% CI 0.22 to 0.25) and after adjustment for (log) parasitaemia at day 0. Haematocrit levels for patients in the CD+A (B) group increased 0.27% per day (95% CI 0.26 to 0.29) and 0.35% per day (95% CI 0.28 to 0.42) in patients in the CD+A G6PD (A-) group.When we included the heterozygous females in the deficient groups the findings remained the same. At day one after treatment the haematocrit was lower than at baseline, but still comparable in the four groups (p\u200a=\u200a0.12); after day 1 the reduction was again greater in patients in the CD+A G6PD (A-) group, which had a significantly lower mean PCV at days 2 and 3 (p\u200a=\u200a0.01 and p<0.001 respectively). After day 7, results were identical.In the multivariate analysis, those in the CD+A G6PD (A-) group had a significantly higher rate of decline in PCV compared with patients in the AQ+SP G6PD (B) group, (p\u200a=\u200a0.004) after adjustment for (log) parasitaemia and temperature day 0; whereas PCV levels in the AQ+SP G6PD (A-) group and CD+A G6PD (B) group declined at the same rate when compared with patients in the AQ+SP normal group (p\u200a=\u200a0.99 and p\u200a=\u200a0.34 respectively). From day 7, results were also comparable: the rate of increase per day was similar for all patients treated with AQ+SP regardless of G6PD status (p\u200a=\u200a0.28) and both CD+A G6PD (A-) and G6PD (B) patients had a higher increase compared to AQ+SP G6PD (B) patients (p<0.001) after adjustment for (log) parasitaemia at baseline.During the study 12 patients had a rapid fall of the haematocrit with other danger signs or severe malaria on days 1, 2 or 3 with parasitaemia . Nine of those received a blood transfusion within four days of recruitment along with parenteral quinine and other supportive therapies . Four paThose cases were classified as Early Treatment Failures (apart from the patient who developed severe anaemia at day 14) with possible Serious Adverse Events as clinically it was not possible to distinguish whether the haemolysis was a consequence of the malaria infection only or also drug related. It is possible that the poor efficacy of AQ+SP, which resulted in a higher number of Early Treatment Failures compared to CD+A treatment As mentioned in the Overall the risk of receiving a blood transfusion was about 7.6 times higher in G6PD (A-) patients, including heterozygous girls, (95% CI 2.2 to 25.6) compared to G6PD (B) patients regardless of the antimalarial treatment . If we cThe median WBC count was similar in the treatment groups at recruitment and at each day of the follow-up, 7, 14, 21 and 28 .P. falciparum malaria treated with CD+A experienced an estimated decline in haematocrit of nearly 2% per day  compared with 1% per day in patients who received AQ+SP, regardless of G6PD status.Malaria is a major cause of anaemia in tropical areas. In this study, conducted in a high transmission area, the haematocrit fell during acute malaria. This initial reduction was independent of the treatment received. Malaria causes anaemia through haemolysis of parasitized erythrocytes, accelerated clearance of uninfected erythrocytes, impaired compensation for this loss by bone marrow dyserythropoeisis and haemolysis due to oxidative stress The significantly steeper decline in haematocrit in G6PD-deficient children treated with CD+A is most likely to have resulted from the use of sulphone dapsone and its metabolite hydroxylamine. Artesunate may cause a temporary suppression of reticulocytosis which does not translate into anaemia, but would not explain the differential effect in G6PD-deficient subjects. Wootton et al., By the end of the 28 day follow-up the haematocrit of most patients, with or without G6PD deficiency, recovered to normal levels. Drug induced haemolysis is self-limiting in individuals with G6PD deficiency because only older red cells are destroyed during drug challenge as these are the cells which are most enzyme deficient, whereas newly produced erythrocytes have nearly normal levels of G6PD which enables them to resist drug-induced destruction In total, 10 patients had severe post-treatment haemolysis requiring blood transfusion and the risk of receiving a blood transfusion was higher in subjects with G6PD deficiency recipients of CD+A.In this trial we recruited children with a PCV\u226521%; we hospitalized and carefully monitored them for the whole administration of the drug, transfusing blood when necessary. Under normal conditions these children would have received the treatment at the health centre or the drug would have been self-administered at home with the consequent risk of life threatening haemolysis.In heterozygous females, the degree of deficiency is determined by the outcome of X-chromosomal inactivation and on average they have less severe clinical manifestations, although some develop severe acute haemolytic anaemia. In fact one heterozygous subject in this trial needed a blood transfusion.In 2002, a randomised clinical trial in Kenya In 2004, results of a randomised multicentre trial in Africa More recently, preliminary results of a multi-centre, double-blind Phase III trial of CD co-formulated with artesunate vs. artemether-lumefantrine The prevalence of G6PD deficiency in African populations varies from 28.1% in southwest Nigeria to 22.5% in Congo, 15.7% in Mali, 13.0% in Uganda and 9.0\u201315.5% in Gabon Pro-oxidant drugs, including antimalarial drugs associated with haemolysis in G6PD-deficient individuals are used commonly in malaria endemic areas. Screening for G6PD deficiency before drug administration of potentially pro-oxidants drugs although seldom available, is necessary.Chlorproguanil-dapsone+artesunate in Rwanda had a poor safety profile in G6PD deficient individuals most likely as a result of dapsone induced haemolysis. This drug combination has also been previously reported to have poor efficacy (70% Adequate Clinical and Parasitological Response at day 28 PCR corrected"}
+{"text": "Plasmodium falciparum resistance patterns over 10 years in southwest Vietnam in infected patients treated with artemisinin compounds.Artemisinin derivatives have been used for malaria treatment in Vietnam since 1989. Reported malaria cases have decreased from 1,672,000 with 4,650 deaths in 1991, to 91,635 with 43 deaths in 2006. Current national guidelines recommend artemisinin-based combination therapy (ACT), although artesunate is still available as monotherapy through the private sector. Recent reports suggest that effectiveness of ACT and artesunate monotherapy has declined in western Cambodia. This study examined P. falciparum malaria, and the length of time that artemisinin had been in use. In vivo and in vitro monitoring of P. falciparum susceptibility to anti-malarial drugs was conducted in 1998, 2001, 2004/5, and 2008/9. Patients with confirmed P. falciparum malaria received therapy with 5 or 7 days of artemisinin (1998 and 2001 respectively) or 7 days of artesunateThe study was conducted in two communes in Phuoc Long district, Binh Phuoc province, 100 km west of the Cambodian border. This was chosen as a likely site for emerging artemisinin resistance because of the high prevalence of P. falciparum to artemisinin in in vitro tests was stable during the period, except for a rise in EC90 and EC99 in 2001.In the four surveys, 270 patients were recruited and treated. The mean parasite clearance times differed between 1998, 2001 and 2004/5  but not between 1998 and 2008/2009. The mean parasite clearance times were correlated with parasite density at day 0 . Treatment failure rates after PCR adjustment were 13.8%, 2.9%, 1.2%, and 0% respectively. Susceptibility of P. falciparum sensitivity to artemisinin compounds in the two sites over a ten-year period. The introduction of ACT in this area in 2003 may have protected against the development of artemisinin resistance. Adherence to the latest WHO and Vietnamese guidelines, which recommend ACT as first-line therapy in all malarious areas, and continued monitoring along the Vietnam-Cambodia border will be essential to prevent the spread of artemisinin resistance in Vietnam.This study showed stable levels of  P. falciparum malaria be treated with artemisinin-based combination therapy (ACT) to prevent recrudescence and to delay the selection of resistant strains [The global burden of malaria has increased during the last decade and in 2006 there were an estimated 247 million cases and 881,000 deaths worldwide [ strains -5.In Vietnam, artemisinin derivatives have been used for malaria treatment since 1989. Reported malaria cases have diminished from 1,672,000 clinical cases with 4,650 deaths in 1991, to 91,635 with 43 deaths in 2006[in vivo and in vitro results are presented for drug resistance monitoring surveys in 1998, and 2001 in Phuoc Trung commune, and in 2004/5 and 2008/9 in Bu Gia Map commune.In 1998, the National Institute for Malariology, Parasitology and Entomology (NIMPE) and researchers at the University of Melbourne commenced monitoring in sentinel sites in Vietnam to examine drug resistance trends-12. In BThe study was conducted in Phuoc Long district, Binh Phuoc province, which is located 130 km north-east of Ho Chi Minh and 100 km west of the Cambodian border  , artemisinin for seven days in 2001  , artesunate for seven days in 2004/5  , and artesunate for seven days in 2008/9 . Naphaco is a Vietnamese manufacturer who was GMP certified after inspection by WHO in 2008. Health clinic staff directly observed all treatment doses.Patients were observed in the health clinic for the first seven days. If the patient vomited within 30 minutes after administration, the full dose was repeated; if vomiting occurred between 30 to 60 minutes after administration, half the dose was repeated. Follow-up appointments were scheduled for days 14, 21 and 28 or if they became unwell and consisted of a physical examination, a blood sample and completion of a standardised data collection form. If treatment failure occurred, patients were treated with CV8 after blood (50 \u03bcl \u00d7 2) was collected by finger prick and transferred onto 3 MM Whatman filter paper for DNA analysis.Treatment failure was defined according to WHO guidelines .In vitro micro-test plates (WHO plates) containing chloroquine and mefloquine were prepared by Sain Penang University, Malaysia. Piperaquine, dihydroartemisinin, and artemisinin plates were freshly prepared by NIMPE using WHO guidelines, and quinine plates were obtained from WHO [from WHO ,15,16. Tin vitro test plates and incubated for 24 to 36 hours[Blood was transferred to  36 hours. One of  36 hours,15,17.et al [msp-1 and msp-2 and 1.5% agarose for glurp. If the same genotype pattern was found in both samples A and B , the infection was most likely to be a recrudescence. In contrast, if the patterns of both samples differed, a re-infection was assumed although it is possible that a minor population of the original infection was responsible. It should also be noted that as specific parasite genotypes are highly common in this region, a subject who is infected with a genotype and then cured may be reinfected with the same genotype (and thus misclassified as a recrudescence).Nested PCR was performed as described by Foley et al  Nested PMean effective concentrations (EC) were calculated using a computer adapted probit analysis of log-dose responses  based ont test. Linear regression was used to assess for a trend in parasite clearance times. The correlation coefficients between the parameters of recent treatments were determined by Spearman rank correlation (r).Discrete data were compared using the chi-square test. Comparisons between two independent groups were made using the student's Consultation was undertaken between the NIMPE team and commune, district and provincial health staff. The team provided patients with information regarding the study and informed consent was documented at the time of enrolment for the surveys. The project was approved by the Human Research Ethics Committee of the National Institute of Malariology, Parasitology and Entomology , the Walter and Eliza Hall Institute of Medical Research  and Melbourne Health .P. falciparum malaria completed a five-day treatment course with artemisinin in the 1998 study, 69 patients completed seven days treatment with artemisinin in 2001, and 82 and 54 patients completed seven days treatment with artesunate in 2004/2005 and 2008/2009 respectively . Artemisinin given for seven days (2001) had a slightly longer parasite clearance time than artesunate for seven days in 2004/5  and artesunate given for seven days in 2004/5 had a longer parasite clearance time than in 2008/9  . Overall 36.9% 24/65) patients had recrudescent parasitaemia during the 28-day surveillance period in 1998 after treatment with 5 days of artemisinin. The earliest recrudescence occurred on Day 11. Six patients had a recrudescence between days 11 and 14, 13 patients between days 15 and 21, and five patients between days 22 and 26. Twelve of the 24 patients had a further parasite recrudescence after retreatment with a five-day course of artemisinin . In 2001 the treatment schedule was changed to artemisinin for seven days, and in 2004/2005 and 2008/2009, artesunate was given for seven days. In these surveys, recrudescent parasitaemia was confirmed microscopically in 7.2% (5/69), 7.3% (6/82) and 3.7% (2/54) patients respectively. The treatment failure rates after PCR confirmation were 13.8% (9/65), 2.9% (2/69), 1.2% (1/82), and 0% (0/54), in the four surveys respectively. The proportion of patients with persistent parasitaemia at day 3 was 6.2% in 1998, 8.7% in 2001, 3.7% in 2004/5 and 1.9% in 2008/9  but no difference between 1998 and 2008/9 (p = 0.127), and no difference between 2004/5 and 2008/9 (p = 0.511).The mean ECP. falciparum resistance patterns over 10 years in infected patients treated with artemisinin or artesunate in two sites in Phuoc Long district, Binh Phuoc province, Vietnam, which were thought to be likely sites for emerging resistance to artemisinin due to the length of time that artemisinin-based compounds had been in use. Seven day treatment with artesunate was found to be effective in this area in 2008/9 with a recrudescence rate from 0% to < 5% and no cases of early treatment failures. There were no significant differences in fever or parasite clearance times between the surveys in1998 and 2008/9. Treatment failure rates reduced during the period. These findings were supported by in vitro testing of P. falciparum isolates, although tests in 2001 raised the possibility of reduced sensitivity to artesunate.This study examined In the 1998 survey, patients were treated with five days of monotherapy with artemisinin compared to treatment with artemisinin or artesunate for seven days in later surveys. The recurrent parasitaemia rate of 36.9% and treatment failure rate of 13.8% confirms the lack of efficacy of this approach. Artesunin vitro tests in the 2001 survey raised the possibility of increasing P. falciparum tolerance to artemisinin, although the significance of reduced parasite sensitivity in EC90 and EC99 tests is not clear. Menard et al [The reduced sensitivity to artemisinin in rd et al  found thP. falciparum infection being detected in the later surveys.A more likely explanation for the increase in EC90 and EC99 in the 2001 survey was deterioration of artemisinin plates during the three months they were kept in the field during the survey. To overcome this possibility plates were replenished every two weeks in the 2004/5 and 2008/9 surveys. However, the reason for prolonged fever clearance time and the higher day three parasitaemia in the 2001 survey is not clear and the possibility remains that the parasites circulating at the time were more tolerant to artemisinin than in the earlier survey. The use of artesunate for treatment in the 2004/5 and 2008/9 surveys, and the introduction of ACT (CV8) as routine treatment in health centres in this district from 2003 may have decreased the chance of drug resistant In vitro resistance of P. falciparum field isolates to artemether and corresponding point mutations in the SERCA-type PfATPase6 have been reported in French Guiana and Senegal [P. falciparum resistance to artesunate in the Mekong region although treatment failures with ACT remain low. Senegal . Also, a Senegal  and in C Senegal . These sin vitro testing. There was also considerable loss to follow-up in the 2001 survey. This and the non-random selection of patients may have biased the results.The limitations of this study included the use of different formulations and treatment regimes with artemisinin and artesunate in the four surveys. However, the design of each survey was consistent with WHO protocols, and treatment of patients was consistent with national malaria treatment guidelines that were current at the time. The first two surveys were also conducted at a different commune than the last two surveys. The two communes were about 80 Km apart and although malaria endemicity is similar at the two sites, it is possible that different patterns of drug resistant malaria emerged at each site during the study period. Anecdotal evidence from a nearby district has suggested higher levels of persistent parasitaemia on day 3, and this is currently being investigated . Another weakness was that although an adequate sample of more than 50 patients was recruited in each survey not all parasite isolates were able to be established in culture for The main strengths of the study included the long follow-up period over 10 years and the careful manner in which the surveys were conducted. For each survey, the same experienced national team was involved in recruiting, treating and monitoring patients, who were given directly observed therapy and monitored closely for the first seven days. Artemisinin plates were replenished in the field every three months in the first two surveys and every two weeks in the last two surveys reducing the likelihood of plate deterioration.P. falciparum sensitivity to artemisinin compounds over a ten-year period in two communes in Phuoc Long district, Binh Phuoc province, Vietnam. The introduction of ACT in 2003 may have protected against the spread of artemisinin resistance in this area. Adherence to the latest WHO and Vietnamese malaria treatment guidelines, which recommend ACT as first line therapy in all malarious areas, and consistent malaria control measures such as early microscopic diagnosis, drug resistance monitoring, provision and use of bed nets, and health worker training will be essential if the development of resistance to artemisinin-based compounds is to be prevented in Vietnam.This study has shown relatively stable levels of The authors declare that they have no competing interests.NVT was responsible for planning, and implementation of the study and analysis of results, and preparation of the first draft of the manuscript in collaboration with other authors. AC provided training for NVT, supervision of the laboratory aspects of the project, and had input into the manuscript. NTT provided supervision for the NIMPE and provincial field team who implemented the project, and had input into data analysis. GC provided ongoing supervision in Hanoi on behalf of UOM, and had input into the manuscript. BP and TT had primary involvement in the implementation of field and laboratory aspects of the project. NH provided overall supervision of NIMPE staff and was involved in planning the project. BB was involved in planning the project, procurement of funding, supervision, and writing the manuscript.All authors read and approved the final manuscript."}
+{"text": "In the present study the therapeutic efficacy of artemether/lumefantrine for the treatment of uncomplicated P. falciparum infection at Kersa, Jima zone, South-west Ethiopia, has been assessed.Artemether/Lumefantrine , late clinical failure (LCF), late parasitological failure (LPF) and adequate clinical and parasitological response (ACPR).90 patients were enrolled and completed the 28 day follow-up period after treatment with artemether/lumefantrine. Cure rate was very high, 96.3%, with 95% CI of 0.897-0.992 (PCR uncorrected). Age-stratified data showed adequate clinical and parasitological response (ACPR) to be 100% for children under 5 and 97.4% and 87.3% for children aged 5-14, and adults, respectively. There was no early treatment failure (ETF) in all age groups. Fever was significantly cleared on day 3 (P < 0.05) and 98% of parasites where cleared on day 1 and almost all parasites were cleared on day 3. 72.5% of gametocytes were cleared on day 1, the remaining 27.5% of gametocytes were maintained up to day 3 and total clearance was observed on day 7. Hemoglobin concentration showed a slight increase with parasitic clearance (P > 0.05). No major side effect was observed in the study except the occurrence of mouth ulcers in 7% of the patients.The current study proved the excellent therapeutic efficacy of artemether/lumefantrine in the study area and the value of using it. However, the proper dispensing and absorption of the drug need to be emphasized in order to utilize the drug for a longer period of time. This study recommends further study on the toxicity of the drug with particular emphasis on the development of oral ulcers in children. P. falciparum has developed resistance to nearly all antimalarials in current use, although the geographical distribution of resistance to any single antimalarial drug varies greatly [Early diagnosis and treatment of cases are the most important strategies for the control and prevention of malaria. It is crucial for proper management of the disease and to prevent further complications.  greatly . World m greatly .P. falciparum to chloroquine (CQ) and sulfadoxine-pyrimethamine (SP) necessitated a change as first-line antimalarial drug for the treatment of P. falciparum. Consequently, Artemether/Lumefantrine (Coartem\u00ae) (AL) was adopted in 2004 [\u00ae is being used as the first-line drug for the treatment of uncomplicated malaria [In Ethiopia, the increased resistance of  in 2004 . Current malaria . A base-\u00ae) need be monitored with particular emphasis to monitoring its therapeutic efficacy [\u00ae) in the routine treatment of uncomplicated P. falciparum malaria at Serbo health center, Kersa district, south west Ethiopia.However, the trend of malaria changes over time, and thus the cure rate, tolerance, compliance and safety of AL  seasons.The study was conducted at Serbo Health Center, Kersa district which is located in Jima zone, Oromia region, 323 km south west of Addis Ababa Fig.  between The study subjects were recruited from febrile patients visiting Serbo Health Center based on WHO inclusion guideline for the assessment and monitoring of antimalarial drug efficacy for the treatment of uncomplicated falciparum malaria .P. falciparum, a parasitemia level of 1000 - 100,000/\u03bcl, absence of danger signs or signs of severe and complicated malaria according to the definition given by [The following inclusion criteria were used for the study: Mono-infection with given by  presence\u00ae (iii) concomitant presence of febrile condition with the potential to confound study outcome (Example: acute respiratory infection (ARI), measles, severe diarrhea, etc.) (iv) severe malnutrition and (v) pregnant and lactating women were not included in the study [Patients with: (i) severe signs of malaria according to WHO criteria, which included severe anaemia defined by haemoglobin < 5 g/dl, (ii) history of allergic reactions to the study drug Coartemhe study .Finger-prick blood samples were collected from consenting patients for malaria parasite identification and hemoglobin level measurement. Patients that satisfied the criteria, were enrolled to the study and followed up on days 1, 2, 3, 7, 14, 21, and 28 where finger-prick samples were taken for microscopic glass slides. A parallel drop of blood was collected on filter paper on day 0 during enrollment and on any unexpected visit. The filter paper was air dried and stored in a self sealing plastic bag with desiccators for further molecular analysis.Considering the very low or unknown proportion of clinical failure of artemether/lumefantrine resistance in Ethiopia, sample size was calculated with 10% precision, 95% confidence interval, 15% withdrawal and 28 day study period, 90 study participants were included in the study [Thick and fixed thin blood smears were stained with 10% Giemsa (pH 7.4) for ten min. Blood films were taken at least eight times for each patient during the study period  and on any unexpected visit by the patient. These smears were used in species identification and thick blood films were used to determine parasite density (100 high power field (HPF) were the number of asexual parasites per HPF were recorded, according to the method described in the WHO Protocols . A bloodSlides were read by two senior microscopists from the Ethiopian Health and Nutrition Research Institute (EHNRI) and one from Serbo health center. In case of disagreement the majority result was taken. All slides were properly documented for quality control.Finger-pick blood sample was used to measure hemoglobin using a portable spectrophotometer (Haemocue).\u00ae) was obtained from Novartis pharma AG, Basel, Switzerland, Bach No. T2005-59, through the WHO office, Addis Ababa. All eligible patients were treated with Coartem\u00ae at day 0. Dosing was a six dose regimen given twice daily for three days. Study medication was administered based on weight; the first dose was given under direct observation. The successive day's doses were given to the patient/guardian for self administration in front of health professionals in the area . Patients were followed for 30 min post treatment and if vomiting occurred, a second full dose was administered. If repeated vomiting occurred, patients were withdrawn from the study. Patients were asked to return to the clinic on days 1, 2, 3, 7, 14, 21, and 28 or were highly encouraged to return whenever they did not feel well. Patients, who failed to come to the clinic at the scheduled time, were visited in their home on the same day and the necessary blood sample, reports of adverse effects and temperature data were collected.Artemether-lumefantrine  vomiting the drug twice, (ii) withdrawal of consent, (iii) onset of a serious febrile illness, (iv) intake of any drug with antimalarial properties, (v) missing repeated treatment doses, (vi) mixed species parasitemia or (vii) any protocol violation. Patients who missed follow-up visits and did not come on the successive day despite tracing were considered as lost to follow-up. Patients withdrawn and with complications were referred to the health centers for proper treatment. Patients withdrawn for the re-appearance of Patients were classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) or adequate clinical and parasitological response (ACPR) as per WHO definitions .Development of danger signs for severe malaria on Day 1, Day 2 or Day 3, in the presence of parasitemia; parasitemia on Day 2 higher than Day 0 count irrespective of axillary temperature; parasitemia on Day 3 with axillary temperature \u2265 37.5\u00b0C; parasitemia on Day 3 \u2265 25% of count on Day 0.Development of danger signs for severe malaria after Day 3 in the presence of parasitemia, without previously meeting any of the criteria of ETF. Presence of parasitemia and axillary temperature >37.5\u00b0C (or history of fever) on any day from Day 4 to Day 28, without previously meeting any of the criteria of ETF.Presence of parasitemia on any day from Day 7 to Day 28 and axillary temperature > 37.5\u00b0C, without previously meeting any of the criteria of early treatment failure or late clinical failure.Absence of parasitemia on Day 28 irrespective of axillary temperature without previously meeting any of the criteria of ETF, LTF or LPF.Samples that were classified as ETF, LCF or LPF were genotyped to differentiate recrudescence from re-infection. Blood samples for PCR were collected on standard filter paper, air dried and stored in cool and dark boxes with desiccants. Genotypic analysis was performed at EHNRI as previously described for msp-1  and msp-The study protocol was reviewed and approved by Ethical Review Committee of Ethiopian Health and Nutrition Research Institute and Nation Ethical Review Committee of Ethiopia.Prior to the trial, a consent form was signed by the patient or by the parent/guardian after being translated and read in the vernacular language that the patients or the carers understood.All the data from recruited patients were imported into an Excel spreadsheet and the WHO designed Excel data analysis program was used for analysis and SPSS 11 were used for descriptive statistics and comparing data .P. falciparum mono-infections. Of the P. falciparum mono-infected patients, 236 could not be enrolled in the study for having a parasite load below 1000 parasites per microliter of blood. Of the remaining 125 patients who were eligible for the study, 35 were excluded for the following reasons: 27 were out of the reachable area, 2 were pregnant, another 4 took anti-malaria drugs before commencement of the study and the remaining 2 refused consent.During the study, 1628 febrile cases, clinically suspected to be infected with malaria, were screened for eligibility. Of these 428 (26.3%) had malaria-positive slides and 361 (22.2%) of them had P. falciparum positive and excluded as treatment failure and treated with quinine. On day 21, 84 slides were examined and two positive slides were found, one positive for P. falciparum and the other for P. vivax. The first was removed as treatment failure and the second withdrawn from the study and treated with chloroquine. On the final day (Day 28), slides from the remaining 82 patients were examined and one P. falciparum and three P. vivax cases were recorded. The first was considered a treatment failure and treated with quinine, and the later three were taken as negative slides for data analysis and were treated with chloroquine  patients (PCR uncorrected) were excluded during follow-up. And thus a total 82 patients were analyzed at the end-point (day 28) and the result was satisfactory compared with other studies. All patients completed the study properly and none of them missed the follow-up.Cure rate was very high in all age groups Table . The oveAs shown in Figure Gametocyte clearance declined with age in the early treatment days, 72.5% were cleared on day 1. However, the remaining, about 28% of gametocyte load was maintained up to day 3 irrespective of treatment. However, total clearance was observed on day 7 Figure .Hemoglobin concentration showed a slight increase with parasitic clearance from the blood, however, the mean change in hemoglobin concentration was not statistically significant between day 0, day 14 and day 28 (P > 0.05)  of which the most common ones were cough in 46% of cases, headache in 17%, anorexia in 9%, weakness/fatigue in 7%, abdominal pain in 5%, diarrhea and sleep disorder in 3%, and vomiting in 2% of the cases. There was also a considerable occurrence of inflammation of the oral mucosa (mouth ulcer) in 7% of patients.\u00ae, however, it was recorded in considerable number of patients (7%) in under 5 and 5-14 age groups.Most of adverse events disappeared with resolution of the disease, however cough prevailed after treatment. The occurrence of mouth ulcer  is not a commonly reported adverse effect of Coartem\u00ae) has a high cure rate. The results showed very high efficacy in the very young children less than five years of age followed by middle age children and adults. Results from the present study were consistent with previous studies conducted in Ethiopia and elsewhere [Our findings have indicated that artemether/lumefantrine  endemicity where inhabitants were considered to be non-immune to malaria. In accord with this, the child splenomegally rate of the district was found to be 7.3% (data not shown) confirming the hypo-endemicity of the study site. Hence our study area being one of an unstable seasonal malarious type, no such stratified immunity acquisition was expected. However, the high cure rate among children could be due to higher milk/fat content in their diet, which facilitates the absorption and hence efficacy of Coartemal route ). Nevert\u00ae was found to be very low at about 2.5%.Of the total of the three treatment failures (one LPF and two LCFs), PCR genotyping showed two of the three recurrent infections (one LPF and one LCF) were recrudescence and one of the LCF was PCR unresolved and excluded. The PCR analysis result increased the overall efficacy in to 97.5%, and the recrudescence rate with Coartem\u00ae clears fever and parasitemia in a very short period of time (less than three days), the concentration of hemoglobin fails to increase within the same period of time. Increments in hemoglobin level have been noted with clearance of fever and parasitemia, especially in those patients who had a lower level of hemoglobin concentration in the baseline of treatment than others. However, the difference was statistically insignificant (p < 0.05). Similar findings were also observed in other studies [In this study, although Coartem studies ,14. The  studies .\u00ae become afebrile within the first two or three days. An antipyretic (paracetamol) was given for febrile patients (body temperature above 38\u00b0C), however the number of patients who required paracetamol during the follow up period was significantly low compared to the baseline. A similar finding was noted in other studies [\u00ae, as a drug with fast resolution capacity of clinical symptoms.Fever is a manifestation of malaria that most frequently causes discomfort. Thus, it was encouraging that patients treated with Coartem studies -15,17,19th day and beyond [\u00ae supports the report that there was some effect of the artemisinin derivative on the sexual stage of the parasite [\u00ae decreases gametocytes by 6-8 fold compared with SP and CQ [\u00ae was reported to reduce gametocyte infectivity down to 40% [The study showed the decline of gametocytes with treatment. They were almost cleared from the patients after seven days. This rate of clearance was higher when compared with other studies, which reported the presence of gametocytes up to the 14d beyond ,14. It hd beyond ,20,21 anparasite ,14. Coar\u00ae most of which are similar to the symptoms of malaria itself. An adverse effect noted in 16.5% of all patients was dry, unproductive cough at baseline. This appears to be very likely a symptom associated with the disease. Interestingly, a further 45% of all patients developed this symptom after commencement of treatment. Similar results were reported in a study conducted in Uganda where, 37% and 40% of cough was observed before and after treatment, respectively [\u00ae and its etiology is not clear. Low levels of immunity due to concurrent diseases such as HIV could be one reason. But, the problem has been noted in 7.1% (6/85) of the patients. The clinicians in the area have also reported the same side effect of the drug in the past. It is therefore, very unlikely to consider the occurrence to be due to chance.Studies showed that the overall toxicity of artemether combinations is very low, the drug is well-tolerated and non-compliance is very low ,17,21. Aectively . HoweverP. falciparum in Kersa district south west, Ethiopia. Four years after its introduction as a first line drug, Coartem\u00ae has been proved to be effective, in rapidly clearing fever and parasites within 72 h in patients suffering from uncomplicated malaria. However, the proper absorption of the drug must be ensured and monitored through controlled nutritional interventions so that the parasites are not exposed to mal-absorbed low, tolerable level of the drug.This study has proved the excellent therapeutic efficacy profile of the combination drug, given as a three day multiple regimen of the optimized formulation for the treatment of This work also recommends further study on blood concentration of the drug and pharmaco-vigilance on the drug toxicity particularly the development of mouth ulcer in children, as drug adverse effects hamper compliance to the proper dosage which in turn enhances development of drug resistance.The authors declare that they have no competing interests.2 involved in preparing the manuscript. TM involved in data analysis, interpretation and critical revision of the manuscript for publication. All authors read and approved the final manuscript.AA was fully involved in all phases of the study, including data collection and monitoring both in the field and in laboratory during data analysis, interpretation, and writing the manuscript; MK designed the study project, supervised the study and revised the manuscript. GL and HM were involved in field data collection and molecular analysis. AA"}
+{"text": "Many countries have implemented artemisinin-based combination therapy (ACT) for the first-line treatment of malaria. Although many studies have been performed on efficacy and tolerability of the combination arthemeter-lumefantrine (AL) or dihydroartemisinin-piperaquine (DP), less is known of the effect of these drugs on gametocyte development, which is an important issue in malaria control.In this two-arm randomized controlled trial, 146 children were treated with either AL or DP. Both groups received directly observed therapy and were followed for 28 days after treatment. Blood samples were analysed with microscopy and NASBA. In comparison with microscopy NASBA detected much more gametocyte positive individuals. Moreover, NASBA showed a significant difference in gametocyte clearance in favour of AL compared to DP. The decline of parasitaemia was slower and persistence or development of gametocytes was significantly higher and longer at day 3, 7 and 14 in the DP group but after 28 days no difference could be observed between both treatment arms.Although practical considerations could favour the use of one drug over another, the effect on gametocytogenesis should also be taken into account and studied further using molecular tools like NASBA. This also applies when a new drug is introduced.Current controlled trials ISRCTN36463274 Plasmodium falciparum parasites to the commonly used drugs chloroquine (CQ) and sulphadoxine-pyrimethamine (SP), many African countries recently adopted artemisinin-based combination therapy (ACT) as first-line treatment for uncomplicated malaria. The combination artemether-lumefantrine (AL) proved to be highly effective and well-tolerated in several studies in Africa [In response to widespread resistance of n Africa -5. Disadn Africa -8 or at n Africa . In Ugann Africa .P. falciparum strains [Another ACT, dihydroartemisinin combined with piperaquine (DP), which was originally developed in China, is increasingly used in Southeast Asia ,10. Pipe strains . This re strains ,11-13 as strains ,14. Cons strains -17. In h strains .Adequate assessment of gametocytaemia is important. Quantitative nucleic sequence based amplification technique (QT-NASBA) has been shown to be much more sensitive for the detection of gametocytes than microscopy ,18-20. IP. falciparum malaria with initial parasitaemia between 1,000 and < 200,000 parasites/\u03bcl blood, axillary temperature \u2265 37.5\u00b0C  or a history of fever. Children with severe malaria, mixed infection or other underlying illness were excluded from the study. In total 146 children were recruited for the present study. Ethical approval for this study was obtained from appropriate local authorities and The Kenya Medical Research Institute  Ethical Steering Committee (SSC protocol No 948). The trial was registered as an International Standard Randomized Controlled Trial at current controlled trials (ISRCTN36463274).The study was conducted in Mbita, western Kenya, at the shores of Lake Victoria during the high malaria transmission season of April-July 2007. Mbita, is an area with highly variable transmission that depends on the local environmental circumstances that can support mosquito conditions. The EIR is calculated to be 6 infectious bites per person per month . Childrend dose of AL, at home.Following diagnosis (at day 0), the patients were randomly allocated to one of the two treatment groups following a computer generated randomization list. One group was assigned DP  once per day for three days. One tablet of the study drug contained 20 mg of dihydroartemisinin and 160 mg of piperaquine (paediatric formulation). Treatment was according to body weight as follows: children between 4\u20137 kg received half a tablet per dose, those between 7\u201313 kg 1 tablet, 13\u201324 kg 2 tablets per dose and children between 24\u201335 kg 4 tablets. The other group was assigned to AL . Each tablet contained 20 mg artemether and 120 mg lumefantrine. Patients received treatment according to bodyweight; i.e. children between 5\u201314 kg received one tablet per dose, those between 15\u201324 kg two tablets and those between 25\u201334 kg received three tablets per dose. Doses were given twice daily. All treatments were given with a glass of milk under direct supervision at the clinic or, for the 2Efficacy was assessed using the WHO in vivo test with a follow-up period of 28 days . At enroPatients showing complications or treatment failure were treated with appropriate supportive therapy. Children developing danger signs or severe malaria on day 1 or 2 of the study were withdrawn from the study, referred to the hospital, and given alternative treatment. Adverse events were recorded on the case record forms. An AE was defined as an unfavourable and unintended symptom, sign or disease. A serious adverse event (SAE) was defined as a symptom or sign that is temporally associated with the drugs administered to the patient that is life threatening or results in hospitalization, permanent and significant disability or death. SAE's were immediately reported to the ethical committee of KEMRI and the drug safety department of Sigma-Tau, Italy.Parasite clearance and gametocyte dynamics were assessed microscopically as well as with quantitative nucleic acid sequence based amplification assay, QT-NASBA.Giemsa-stained thick and thin smears were prepared according to WHO guidelines. Two independent experienced microscopists, who were blinded to the treatment and clinical status of the patient, examined the smears for the presence of parasites and identified the observed parasite species. Parasitaemia was determined by counting the number of parasites against 200 leukocytes for the asexual stages . The presence of gametocytes was examined against 500 leukocytes.et al [6 to 10 in vitro cultured parasites/ml was used as reference and processed and analysed with NASBA. Furthermore, to assess prevalence of gametocytes below the detection limit of microscopy, QT-NASBA targeting Pfs25 mRNA as described by Schneider et al [Finger prick blood (50 ul) for NASBA analysis was collected on Whatman 903 filter paper  and air-dried at room temperature. Nucleic acid extraction was performed as previously described by Boom et al . Real-tiet al . In ordeer et al  was usedet al [In order to discriminate between re-infection (RI) and recrudescence (RE), merozoite surface protein 1 and 2 (msp1 and msp2) and glutamate rich protein (GLURP) genotyping was performed as described by Snounou  on bloodet al . Molecul2-test and means were compared with the one-way ANOVA or Student t-test. A simplified trapezoid area under the curve (AUC) analysis using gametocyte data from days 0, 3, 7, 14 and 28, as a surrogate for the infectiousness of the participants in the different treatment groups, was performed.The aim of the study was to compare gametocytaemia after AL and DP and to compare assessment of gametocytaemia by microscopical examination versus QT-NASBA All data were entered in excel and analysed with SPSS for windows (version 12.0). Parasite densities were analysed after natural log-transformation. Where appropriate, proportions were compared with the \u03c7In total 1882 cases suspected of uncomplicated malaria were screened for eligibility into the study during an 8-week recruitment period in April and May 2007. 1,736 children were excluded because they did not meet the inclusion criteria  was 8.96 * 105 at 48 hours for the AL treatment and 2.06 * 104 at 48 hours for the DP treatment.There were no early treatment failures during the first three days of follow up. Only one patient in the AL arm had a recurrent parasitaemia  at day 28 of follow up. Genotyping analysis revealed that this patient had a reinfection with NASBA was also applied to monitor parasite dynamics below sub-microscopical level. Humidity in some of the filter papers degraded the RNA in the blood spots of some of the samples. This led to several extraction failures. In order to have a clear picture of parasite dynamics only those series with a full range of follow-up samples, i.e. 56 DP and 54 AL treated patients, were analysed. Both treatment arms showed a steep decline in parasitaemia from the day of enrollment (day 0) to day 1; 62% reduction after DP treatment and 89% reduction in the AL arm. At day 2, the level of parasitaemia was reduced to 1.2% in the AL group and 2.75% in the DP group.At baseline Hb levels in both treatment groups were comparable Table . At day P. falciparum reinfection on day 28 in the AL group and the child that developed broncho-pneumonia (case presented below). Furthermore the presence of fever at recruitment was no predictor for gametocyte carriage (p > 0.05)Fever clearance was defined as the time from receiving the assigned treatment to the time a normal body temperature was recorded (\u2264 37.5\u00b0C), in study cases who presented with fever. Fever clearance was rapid in both study groups. On day 1, 10 cases (13.7%) in the DP group presented with fever and six cases (8.2%) were observed in the AL group. In the DP group fever was observed on day 2 in four cases (5.5%) and three cases (4.1%) on day 3. In the AL group cases with fever also presented on day 2  and on day 3 . Fever was not observed during follow-up after day 7, with the exception of the child that presented with a Plasmodium falciparum infection with parasitaemia of 20,120 parasites/\u03bcl was diagnosed. There was a fever (38.6\u00b0C), but there were no other complaints and no signs of severe anaemia (Hb: 6.4 mmol/L). On day 3, there were no signs of illness. On day 7, the child presented with fever (38.6\u00b0C), cough and complaints of anorexia. There was no history of significant illness or allergies. After examination , broncho-pneumonia was diagnosed and the child was treated with oral phenoxymethylpenicillin for five days and Paracetamol syrup. On day 14, the child did not attend the follow-up visit, the parents reported that the child died a day before in a local health post. The event was assessed as unrelated to the study drug. Autopsy was not performed.Most adverse events were mild, self limiting and consistent with symptoms of malaria. There was no significant difference between the two study groups Table . One patThe presence of gametocytes in clinical samples was assessed by microscopy and NASBA and is presented in Table (day 0\u201328) of the two treatment groups was calculated to be 20.0 infectious persons/day for the DP treatment arm and 10.5 infectious persons/day for the AL treatment arm. Stratifying the data for age under and above 60 months showed no difference in either of the groups.NASBA analysis on 56 DP treated subjects and 54 AL treated subjects detected strikingly more gametocyte carriers at the start of the study compared to microscopy; i.e. 22 study subjects in the DP arm (39.3%) and 21 in the AL (38.9%) were harbouring gametocytes before. The pfs25 NASBA revealed that 34 cases (60.7%) were gametocyte positive in the DP group on day 3, of which 13 cases were newly identified compared to day 0. In contrast, a significantly lower number of patients  were gametocyte positive in the AL treatment group on day 3, of which six new cases. This trend was also observed on day 7: the DP arm had 33 (58.9%) gametocyte positive samples, whereas the AL treated group had a significantly lower number of gametocyte positive samples . However on day 14  and day 28  of follow-up no significant difference in gametocyte carriage was observed between both treatment groups. The AUC et al, who found a greater increase in the DP treated patients [Several studies have analysed the efficacy and tolerability of AL and DP and all show very good results ,12,14,25The effects on gametocytaemia and possibly malaria transmission deserve further study. Whereas asexual parasites were cleared in three days after the initiation of the two treatment schedules, gametocytaemia appeared different when assessed by microscopy as well as with NASBA. Gametocytes were present in low numbers throughout follow-up in both study groups. Artemisinin derivatives have in general a negative effect on gametocyte development and survival and thus influence malaria transmission, at least in low transmission areas -17. In tThe present study showed a limited effect of DP on gametocyte development in comparison with AL when a sensitive tool like NASBA is used for gametocyte detection, which could limit the usefulness of DP to areas with low transmission but this finding should be further investigated in larger studies in different study sites with different transmission intensities. It is not clear if plasma concentrations of dihydroartemisinin in the blood could play a role. Dihyrodartemisinin is the major and the active metabolite of artemether. So far, no studies have been performed that compare the plasma levels of dihydroartemisinin when given as such or after administration of artemether and subsequent metabolisation. This should be further investigated together with effect on gametocytogenesis, which should incorporate a sensitive detection tool for gametocytes such as NASBA.The effect that drugs can have on gametocyte clearance as measured with NASBA could have some implications for the introduction drugs and especially the introduction of new drugs. This study showed that with sensitive detection tools a difference in parasite clearance can be observed but these results should be confirmed in larger studies and in other study areas with different malaria transmission intensities. Transmission intensity varies significantly in the different African countries and within a country high and low transmission areas can often be identified. Malaria endemic countries generally have a national malaria drug policy for the whole country. Although this is logical from a practical and logistical point of view, it may not be the best approach for effective malaria control. It could, therefore, be more effective if a country develops specific drug policies to suit regional instead of national requirements.The authors declare not to have a conflict of interest. The organizations that provided financial support had no influence in the design, the actual field and laboratory work, analysis and writing.PM was involved in the conception of the study, carried out the molecular analysis of the samples, carried out the statistical analysis of the results and drafted the manuscript. PS coordinated the study in the field and was responsible for the clinical examination and treatment of the participants. SvA collected clinical data and blood samples in the field and did the clinical data analysis. SO coordinated the study in the field an arranged the logistics in the field. IV carried out the NASBA assays and the subsequent analysis.HS was involved in the design of the study and contributed to the drafting of the manuscript. PK as involved in the design of the study, critically read and improved the manuscript. All authors read and approved the final manuscript"}
+{"text": "Multi-drug resistant falciparum malaria is an important health problem in the Peruvian Amazon region. We carried out a randomised open label clinical trial comparing mefloquine-artesunate, the current first line treatment in this region, with dihydroartemisinin-piperaquine.P. falciparum uncomplicated malaria were recruited, randomized (260 with mefloquine-artesunate and 262 with dihydroartemisinin-piperaquine), treated and followed up for 63 days. PCR-adjusted adequate clinical and parasitological response, estimated by Kaplan Meier survival and Per Protocol analysis, was extremely high for both drugs (99.6% for mefloquine-artesunate and 98.4% and for dihydroartemisinin-piperaquine) . All recrudescences were late parasitological failures. Overall, gametocytes were cleared faster in the mefloquine-artesunate group (28 vs 35 days) and new gametocytes tended to appear more frequently in patients treated with dihydroartemisinin-piperaquine (day 7: 8 (3.6%) vs 2 (0.9%), RR: 3.84, 95%CI [0.82\u201317.87]). Adverse events such as anxiety and insomnia were significantly more frequent in the mefloquine-artesunate group, both in adults and children.Between July 2003 and July 2005, 522 patients with P. falciparum malaria but it is better tolerated and more affordable than mefloquine-artesunate . Therefore, it should be considered as a potential candidate for the first line treatment of P.falciparum malaria in Peru.Dihydroartemisinin-piperaquine is as effective as mefloquine-artesunate in treating uncomplicated NCT00373607ClinicalTrials.gov  P. falciparum malaria occurs in the Peruvian Amazon region where the current first line treatment, mefloquine associated with artesunate (MAS3), was introduced in 2001 following high resistance to chloroquine (CQ) and sulfadoxine/pyrimethamine (SP) P. falciparum, MAS3 is not well tolerated and its compliance is poor: 14% of treated patients experienced adverse events in a post-marketing surveillance study carried out in Peru In Peru, malaria is one of the most common parasitic diseases and an important public health problem. The highest incidence of P. falciparum malaria in Southeast Asia P. falciparum. However, its combination with dihydroartemisinin has shown to be active against multi-drug resistant P. falciparumversus 18.65 USD per adult treatment) Dihydroartemisinin-piperaquine (DHA-PPQ) is a new artemisinin-containing fixed-combination treatment (ACT) that has proved to be well tolerated and highly efficacious against uncomplicated vs the current MAS3 regimen in Peruvian patients with uncomplicated P. falciparum malaria and results are reported below.We have carried out the first clinical trial in Latin America evaluating the safety, tolerability and efficacy of DHA-PPQ The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Anopheles darlingiPlasmodium falciparum represents 20%\u201330% of all malaria infections, the rest being due to P. vivax. All age groups are at risk, with the majority of infections progressing towards symptomatic disease, though severe malaria cases are rare A randomised, open-label clinical trial was carried out between July 2003 and July 2005 in 9 health posts (HP) and 4 commune health centres (CHC) in the rural communities located south of Iquitos, the largest city in the Peruvian Amazon region  P. falciparum they were referred to the San Juan Health Centre where another blood slide was collected. They were enrolled in the study if they met, besides being in the required age range, the following inclusion criteria: fever (axillary temperature\u226537.5\u00b0C) or history of fever in the preceding 24 h and mono-infection with P. falciparum with asexual parasite density between 1,000 and 200,000/\u00b5l. Patients were excluded if they had at least one of the following exclusion criteria: severe malaria, pregnancy or lactation, any other concomitant illness or underlying disease, contra-indication to the trial drugs, history of treatment with mefloquine in the last 60 days or CQ, primaquine or quinine within the 14 days before the current episode.Patients aged 5\u201360 years old attending the health facilities and with suspected clinical malaria had a blood slide done (thick blood film). If positive for http://www.clinicaltrials.gov.Patients meeting the inclusion/exclusion criteria were enrolled in the study only after written informed consent was obtained from them or their carer. The study protocol was approved by the Ethical Review Committee of the Universidad Peruana Cayetano Heredia and that of the Institute of Tropical Medicine, Antwerp, Belgium. The research was performed in accordance with the ethical standards of the Peruvian Ministry of Health. The trial has been registered as an International Standard Randomised Controlled Trial, number NCT00373607 at TM, Hoffman-La Roche Co., Basel, Switzerland) and AS\u200a=\u200a4 mg/kg/day , DHA-PPQ\u200a=\u200a2.1 mg/kg/day DHA and 16.8 mg/kg/day PPQ . The number of pills was rounded off to the nearest quarter tablet. Patients were observed for 1 hour: if vomiting occurred within 30 min of dosing, a full dose was re-administered; if vomiting occurred between 30 min and 1 hour only half dose was re-administered. For young children, drugs were crushed, dissolved in water and squirted into their mouths.Patients were randomised in blocks of ten to receive either MAS3 or DHA-PPQ. Sealed opaque envelops containing the treatment allocations were opened only after the final decision to recruit the patient had been made. Drugs were administered once daily for 3 days according to the patient's body weight: MQ\u200a=\u200a8 mg/kg/day (LariamP.vivax infections were treated with chloroquine (25 mg/kg over 3 days).All enrolled patients were followed up daily until day 3 (24\u201348\u201372 h) and were asked to return on days 7, 14, 21, 28, 35, 42, 49, 56 and 63 or outside scheduled visits if any symptom occurred. If the patient did not report for scheduled visits, every effort was made to locate him/her at the home address. During each visit, a clinical assessment was done and a blood sample for parasitaemia taken by finger prick. At Day 0 and at any visit from Day 7 onwards, a blood spot on filter paper was also taken for later PCR analysis on recurrent parasitemia cases. PCV was measured at Day 0, 7 and 14; for those still in the follow up, a last measurement was done at day 63; anaemia was defined as a PCV\u226433%. A venous blood sample was taken at D0 and D7 for a complete blood count, liver and renal function tests. Rescue treatment was provided to patients with recurrent parasitaemia or with serious adverse events requiring treatment withdrawal. Patients treated with DHA-PPQ were given MAS3 and those on MAS3 received DHA-PPQ. Severe malaria cases were treated with parenteral quinine. Clinical examination systematically assessed body temperature (axillary temperature), heart and respiratory rate, blood pressure, liver and spleen size. All signs and symptoms were systematically recorded on a pre-coded standardized form.The thick and thin blood films were stained with Giemsa 10% for 10 minutes and reading for species determination and parasite density was blinded to treatment. Parasite density was calculated by counting the number of asexual forms for 200 white blood cells (WBC) and assuming 8,000 WBC/\u00b5l. A blood smear was declared negative if no asexual form was detected after reading 100 microscopic fields. If sexual forms were detected, a gametocyte count was performed against 1,000 WBC. Quality control was performed on 10% of all slides (random sample) at the National Hospital of the Cayetano Heredia University in Lima. The agreement between the first and second reading was 98%.PCV was measured using micro capillary tubes which were centrifuged and PCV determined following the Hawksley micro-haematocrit reading method 2) was taken on filter paper (Whatman n\u00b03) for later PCR analysis. Each PCR filter paper was dried and stored individually in a plastic bag containing silica gel. Parasite genotyping was done by a nested PCR for variable blocks within the merozoite surface protein 1 (MSP1), MSP2 and glutamate-rich protein (GLURP), as described previously A blood spot . Treatment outcome was established according to the World Health Organization and Pan American Health Organization guidelines for An adverse event (AE) was defined as \u201cany unfavourable and unintended sign, symptom, or disease temporally associated with the use of the drug administered\u201d.Data were double-entered and cleaned in ACCESS, Microsoft office 2003, and analysed in STATA version 8.0 . Descriptive statistics were used to summarise baseline clinical and biological values of all randomized patients. All PCR indeterminate results were considered as treatment failures, adopting a worst case scenario.ITT), Per-protocol (PP) and Kaplan Meier survival analysis to determine the cure rate (PCR-adjusted) at day 63 in each treatment group. In the ITT analysis, patients lost to follow up were considered as treatment failures (worst case scenario) and the denominators constituted by the patients initially randomized to each group. In the PP analysis, patients lost to follow-up and those excluded after enrolment were excluded from the analysis, and only patients with complete follow-up were included in the denominator. In the ITT and PP analysis, cure rates were compared using chi-square (\u03c72) test with Yates' correction or two-tailed Fisher's exact test as appropriate. In the Kaplan Meier survival analysis, each patient contributed to the analysis for the time s/he was followed up. Patients lost to follow up or with a new Plasmodium infection  were censored at the time they were last seen for the primary outcome. In the survival analysis, cure rates were described by Kaplan Meier estimates and compared between groups with a log-rank test. Gametocyte carriage rates were estimated as proportion of patients with gametocytemia at day 0, 7, 14, etc, until total clearance in both groups, and person-gametocyte-weeks were calculated to measure gametocyte carriage and transmission potential. The person-gametocyte-weeks rates were calculated as the number of weeks in which blood slides were positive for gametocytes during the 2 first weeks of follow-up after treatment divided by the number of all follow-up weeks and expressed per 1,000 person-weeks. The risk of adverse events during the first week after the start of the treatment was calculated for each group separately in children and adults. Relative risks and p-values were computed using chi-square (\u03c72) test with Yates' correction or two-tailed Fisher's exact test as appropriate.Data were analysed by Intention-to-treat  than in the DHA-PPQ (13%) group (p\u200a=\u200a0.31). The median PCV at day 0 was comparable in both treatment groups .versus 58.5%, p\u200a=\u200a0.03], though this difference tended to disappear at day 2 so that by day 3 all patients had cleared parasitaemia  in the DHA-PPQ and 12 (5%) in MAS3 group had recurrent parasitemia, among which ectively . In the PP analysis, 98.3% vs 99.6% . In the ITT analysis, the ACPR was obviously lower than the other estimates, 90.1% [95%CI: 85.8\u201393.4] for DHA-PPQ and 93.8% [95%CI: 90.2\u201396.4] for MAS3, but not significantly different . Results were very similar in the Before treatment, 35 patients (13%) in DHA-PPQ and 43 patients (17%) in the MAS3 group had gametocytaemia. After treatment, the gametocyte carriage rate decreased faster in MAS3 (gametocytes cleared by day 28) than in the DHA-PPQ group (gametocytes cleared by day 35) . Among pOverall during the first week of treatment, adverse events were more frequent among patients treated with MAS3 compared to those treated with DHA-PPQ , and theThe majority of adverse events were transient and resolved spontaneously with the only exception of a woman who complained of anxiety and insomnia during the whole follow-up period and needed the administration of sedatives and anxiolytic drugs. Serious drug-related adverse events requiring early treatment termination occurred in 3 patients treated with MAS3. A 30 years old man suddenly presented at day 4 with severe dizziness, loss of balance, unsteadiness, amnesia and severe anxiety. Immediately after the end of the treatment, he reported severe headache and insomnia. Clinical and biological examinations diagnosed a toxic metabolic encephalopathy, possibly due to MAS3. A 41 years old man had severe weakness, anxiety, and transient arrhythmia 30 minutes after the second dose of MAS3. The third patient, a 6 years old boy, presented with palpitations and chest pain, 12 hours after the second dose of MAS3.versus 18.65 USD per adult treatment) This first clinical trial carried out in South America on DHA-PPQ showed that this new fixed-dose ACT is as efficacious as MAS3 but better tolerated and with significantly less adverse events. These excellent results strongly support DHA-PPQ as a good alternative to MAS3, the current first line treatment in the Peruvian Amazon region. Moreover, compared to MAS3, DHA-PPQ is less expensive , while in the MAS3 group the only new infection detected occurred at day 21. However, a similar study (63-day follow up) carried out in Cambodia in a low transmission area and comparing DHA-PPQ with MAS3 did not report any difference in the incidence of new infections The occurrence of adverse events was generally lower and less severe in patients treated with DHA-PPQ than in those treated with MAS3, both in adults and children. Their high occurrence in the MAS3 group differ considerably from that observed in another trial carried out in Peru in 2000 P. falciparum malaria in the Amazon region of Peru. This co-formulation is safe and better tolerated than MAS3 and thus may have a better compliance. All these advantages combined to its low cost makes DHA-PPQ a suitable and even advisable first line treatment for P. falciparum malaria in Peru. Our encouraging results should prompt neighbouring endemic countries to conduct similar efficacy trials, and possibly change their anti-malaria treatment policy to this highly efficacious, safe, well tolerated and affordable ACT.In conclusion, our study confirmed that DHA-PPQ is a highly effective antimalarial treatment for Protocol S1Trial Protocol(0.13 MB DOC)Click here for additional data file.Checklist S1CONSORT Checklist(0.05 MB DOC)Click here for additional data file."}
+{"text": "The fixed dose antimalarial combination of dihydroartemisinin-piperaquine(DP) is a promising new artemisinin-based combination therapy (ACT). Wepresent an individual patient data analysis of efficacy and tolerability inacute uncomplicated falciparum malaria, from seven published randomizedclinical trials conducted in Africa and South East Asia using a predefinedin-vivo protocol. Comparator drugs were mefloquine-artesunate (MAS3) inThailand, Myanmar, Laos and Cambodia; artemether-lumefantrine in Uganda; andamodiaquine+sulfadoxine-pyrimethamine andartesunate+amodiaquine in Rwanda.In total 3,547 patients were enrolled: 1,814 patients (32%children under five years) received DP and 1,733 received a comparatorantimalarial at 12 different sites and were followed for 28\u201363days. There was no significant heterogeneity between trials. DP was welltolerated with 1.7% early vomiting. There were less adverseevents with DP in children and adults compared to MAS3 except for diarrhea;ORs (95%CI) 2.74 (2.13 to 3.51) and 3.11 (2.31 to 4.18),respectively. DP treatment resulted in a rapid clearance of fever andparasitaemia. The PCR genotype corrected efficacy at Day 28 of DP assessedby survival analysis was 98.7% (95%CI97.6\u201399.8). DP was superior to the comparator drugs in protectingagainst both P.falciparum recurrence and recrudescence. There was nodifference between DP and MAS3 in treating P. vivax co-infections and insuppressing the first relapse . Children under 5 y were at higher risk of recurrence for bothinfections. The proportion of patients developing gametocytaemia and thesubsequent gametocyte carriage rates were higher with DP  than MAS3 .DP proved a safe, well tolerated, and highly effective treatment ofP.falciparum malaria in Asia and Africa, but the effect on gametocytecarriage was inferior to that of MAS3. Plasmodium falciparum malaria Over 80 countries worldwide have now implemented WHO recommendations to useartemisinin-based combination therapy (ACT) as first-line treatment ofPlasmodium falciparum, andvivaxP.falciparum malaria. Therandomized trials included in this individual patient data analysis Piperaquine has a terminal half-life of several weeks The trials were conducted in North-western Thailand, Rakhine state, Myanmar, SouthernLaos, and Western Cambodia, where mefloquine combined with a three day course ofartesunate (MAS3) was the comparator drug, in Uganda where artemether-lumefantrine(AL) was the comparator, and in Rwanda, where artesunate+amodiaquine(AS+AQ) and a non-ACT group amodiaquine+sulfadoxine-pyrimethamine(AQ+SP) were the comparator antimalarial drugs. In Rwanda, following highlevels of chloroquine (CQ) resistance, the combination AQ+SP was adopted asthe first-line anti-malaria treatment in 2001. However, AQ+SP has alwaysbeen considered an interim strategy and different artemisinin-based combinationtreatments (ACT) have been tested in the past few years as possible alternatives.P.falciparum monoinfections were included in Laos, Rwanda, andUganda, while in Cambodia, Thailand and Myanmar patients with mixed infections wereincluded. Studies excluded pregnant or breastfeeding women, patients with HIV-AIDSor severe malaria. Malaria on admission and reappearance were confirmed bymicroscopy examination of blood smears. All sites screened actively for adverseevents until Day 28.Patients presenting with acute uncomplicated falciparum malaria were recruited intothe treatment studies provided they gave fully informed consent. Eligible patientswere 12 to 59 months old patients weighing more than 10 kg in Rwanda, 6 months to 10years old, weight >5 kg in Uganda, and all patients between 1 and 65 years inMyanmar, Laos, Cambodia, and Thailand. Only uncomplicated cases ofTrials were open label randomized comparative studies. Lengths of follow-upvaried from 28 days in Rwanda, to 42 days in Laos, Myanmar, Uganda, and 63 daysin Cambodia and Thailand. Patients received a total dose of approximately 7mg/kg bw dihydroartemisinin and 56 mg/kg bw piperaquine divided into 3 dailydoses, except in Cambodia and in the first trial in Thailand in which an earlierdose regimen was used where the same total dose was divided into 4 doses givenat 0, 8, 24 and 48 h. One tablet of DP  contained40 mg of dihydroartemisinin and 320 mg of piperaquine phosphate. Comparator drugdoses were MAS3 3 days: artesunate 4 mg/kg/day, and mefloquine 8 mg/kg/day, orAQ+SP: AQ 10 mg/kg/day for 3 days and SP 25 mg/kg of sulfadoxine and1.25 mg/kg of pyrimethamine the first day or AS+AQ 3 days: artesunate 4mg/kg/day, and amodiaquine 10 mg/kg/day, and AL, 20 mg artemether/120 mglumefantrine tablets according to weight as one (5\u201314 kg), two(15\u201324 kg), three (25\u201334 kg), or four (\u226535 kg)tablets given twice daily for 3 days.Drug administration was observed directly by study investigators, except for onearm in Myanmar where effectiveness was assessed Polymerase chain reaction (PCR) parasite genotyping was performed on pairedsamples for parasite genotyping to distinguish between new infections andrecrudescent cases. Allelic variation within MSP1, MSP2, and GLURP was used inAsia as described previously The clinical trials from each country were approved by appropriate authorities.The Thai studies were approved by the Faculty of Tropical Medicine, MahidolUniversity Ethical Committee . Approval for the Laos studywas granted by the Ethical Committee of the Faculty of Medical Sciences,National University of Laos; both of these studies were also approved by theOxford Tropical Research Ethics Committee (OXTREC), University of Oxford, UK.The protocol for the trial in Myanmar was approved by the Myanmar Department ofHealth and by the M\u00e9decins Sans Fronti\u00e8res (MSF) EthicalReview Board. In Rwanda, the study was reviewed and approved by the Ministry ofHealth of Rwanda and by the Ethical Committee of the Prince Leopold Institute ofTropical Medicine, Antwerp, Belgium. The Cambodian study received ethicalclearance from the Cambodian National Ethical Review Committee , and the MSF Ethical Review Board. In Uganda, approval camefrom the Makerere University Research and Ethics Committee, the Uganda NationalCouncil of Science and Technology, and the University of California, SanFrancisco Committee for Human Research.The databases of randomized controlled trials were sent by the investigators. Thefollowing aspects of the quality of trial methodology were evaluated: generationof the allocation sequence, adequacy of concealment of the allocation oftreatment, degree of blinding, and completeness of follow-up. Generation of theallocation sequence and allocation concealment was classified as adequate,inadequate, or unclear Blinding was classified as open, single or double. The proportion of patientslost to follow-up (regardless of failures) was computed and consideredacceptable if <10% within 28 days. Other markers of qualityassessed were whether a sample size was determined using power calculations andwhether an intention-to-treat (ITT) analysis could be computed.The analysis was by modified intention-to-treat where patients who did notcomplete the study were censored on their last day of follow-up, but they werenot regarded as a failure as in a \u201cpure\u201d ITT analysis.P.falciparum infection were censored for the primary outcome at the timethey were last seen. All studies followed patients for at least 28 days and theprimary endpoint was defined prospectively as the parasitological treatmentfailure .Treatment failure was considered as the sum of early and late treatmentfailures, as defined by the WHO The primary endpoint was the treatment efficacy by Day 28. Patients lost tofollow-up (or missing a weekly visit) or with a new The efficacy was measured using Kaplan-Meier survival analysis. We applied astatistical correction for cases where PCR genotyping gave an indeterminateresult or was unavailable, computing adjusted quotients by determining theprobability by site and at any time of a parasite reappearance being either arecrudescence or a new infection \u2003The risks of recrudescence (reappearance of the same genotype)in the DP groups compared to the comparator groups (stratified bystudy), using the different lengths of follow-up of each studies.\u2003The risks of recurrence (defined as recrudescence as above andnew infections) in the DP groups compared to the comparator groups(stratified by study), using the different length of follow-up of eachstudies.\u2003The risk of new infection (excluding recrudescences) in DPgroups compared to the comparator groups per study and overall result(stratified by study) at Day 28.The risks of treatment failure  were compared in multivariateanalysis (Cox regression models) individually and overall by stratifying bysite in an attempt to account for potential statistical heterogeneity.\u2003The predictors of recrudescence or new infection or recurrenceby Day 28 were assessed by Cox regression models. The covariatesexamined were: sex, age (continuous), food absorption with the drug,anaemia on admission (haematocrit <30%), parasitecount on admission (log transformed), elevated temperature on admission(temperature measured by any method at \u226537.5\u00b0). As theage groups were different between Africa (under 5 years old) and Asia(children and adults), we conducted this part of analysis separately forthe 2 continents.\u2003Gametocyte carriageThe predictors of gametocyte prevalence on admission weremeasured using a logistic regression and controlling forsite.The time to clearance of gametocytes already present on admissionwith data censored at the time of gametocyte clearance wascalculated based on the results of blood smears. Within eachstudy, the same sampling schedules were used for all thepatients, but between trials parasite counts were performed atdifferent times so any analysis was stratified by site toaccount for these differences. For patients who clearedgametocytaemia, time of the first negative count (followed byfurther negative counts) was taken as time of clearance.Differentials in gametocytes clearance were calculated usingKaplan-Meier method using logrank test, stratified by site, anda Cox regression model stratified by site measured the risks ofgametocytes carriage between treatment groups.The predictors of gametocyte appearance during the follow-up inpatients without gametocytaemia on admission were assessed by aCox regression model stratified by site. The presence ofgametocytaemia after starting treatment was analysed as a binaryvariable: the Mantel-Haenszel method and the homogeneity teststratified by site were used to estimate a combined odds ratiobetween treatments. One positive gametocyte count at any timeafter treatment during the follow-up period was enough to definegametocyte carriage, while a complete set of negative countsduring follow-up was required to confirm no carriage.Gametocyte carriage rates were measured inperson-gametocyte-weeks, using binary variable calculated within42 days of follow-up. Person-gametocyte-weeks (PGW expressed per1000) were defined as the number of weeks in which blood slideswere positive for gametocyte divided by the total number ofweeks followed up in patients with gametocyte results \u2003Haematological changes were measured using the paired t-test.Anaemia was defined as haematocrit <30%, and anaemiarecovery during the follow-up by the time for the haematocrit to reach30% or more.\u2003Adverse events: defined as any sign, symptom, or disease thatwas not present on admission and was associated with the use of amedicinal product, whether or not it was considered as related to themedicinal product. A serious adverse event was defined as a sign orsymptom that was fatal, life threatening or required admission tohospital. Adverse events were standardised and expressed as an incidencedensity, in person-days at risk within 28 days. The incidence rate ratiotest was used to compare the incidence of adverse events. We assumedthat young children (<5 years old) were unable to answerquestions about dizziness, nausea, headache, confusion, numbness,hearing disturbance, tinnitus or visual disturbance.P. vivax recurrences inthe Asian trials with longer follow-up (63 days) was calculated bycensoring the data at the time of P. vivax appearance(binary variable). The predictors of P.vivax appearancewere measured by using Cox regression, and the incidence density ofP. vivax appearance was calculated inperson-day.\u2003The effects of DP on 2 test.Chi-square, Mann-Whitney, Kruskall-Wallis tests were used as appropriate.Confidence intervals (CI) were measured at 95% by the binomialdistribution, or the Wilcoxon procedure, or the Taylor series estimate asappropriate Heterogeneity was assessed by the Cochran Q test, and IA total of 3,547 patients were enrolled in six countries from 12 different sitesbetween October 2003 and June 2006. Individually, the trials enrolled between 75and 303 patients treated with DP, 1,475 patients treated withother ACTs, and 258 in the non-ACT (AQ+SP) group . The pro2 test\u200a=\u200a26%,P\u200a=\u200a0.15, Cochran Q test for heterogeneity),and regarded as low In all the trials the methodological quality was high and the randomisationsequence was computer-generated. All trials were open label, and the basis forthe sample size studied was provided in all studies. In all studies, the primarytreatment outcome was the parasitological treatment failure. All trials reporteddata on haemoglobin levels during follow-up, and recorded gametocyte carriage atstudy enrolment and during follow-up. All studies assessed adverse events and 10different adverse events were documented in all trials. Although there weredifferences in geographical location, transmission intensity , age,treatment and supervision, heterogeneity between trials was not significant(IThe median age of the recruited patients was 13 years (range 1 to 65) . OverallOverall 54.6% of patients were febrile (\u226537.5\u00b0C) onadmission. This decreased to 8.8% at 24 h (i.e. median feverclearance <24 hours) and 2.1% at 48 h. No difference wasdetected between treatment groups. The median time for the spleen to be nolonger palpable was 14 days (range 1\u201342). There was no difference inthe time to resolution of splenomegaly between the treatment groups(P\u200a=\u200a0.70).Of the 1,814 DP treated patients, 126 patients (6.9%) were lost tofollow-up by Day 28. Of the 221 patients with recurrent infections, 9 cases hadindeterminate PCR genotyping results, and 3 results were not available (lostsamples). The results of parasitological efficacy per study site are shown inP. falciparum parasitaemia thecorresponding results were 96.1% (95%CI95.0\u201397.2) and 90.4% (95%CI 87.8\u201393.0)in children (P\u200a=\u200a0.001).In DP groups, the overall observed parasitological efficacy using results fromsurvival analysis at Day 28, corrected by PCR was 98.7%(95%CI 96.8\u201398.3). In children under 5 years old, thecorresponding efficacy was lower: 94.2% . For theoverall recurrence of Based on randomised comparisons by country and using the full length offollow-up of the different trials, DP recipients were at lower risk for aPCR confirmed failure compared to MAS3 in Thailand(P\u200a=\u200a0.001), AL in Uganda(P\u200a=\u200a0.004), and AQ+SP in Rwanda(P\u200a=\u200a0.001). OverallP. falciparum recurrencethan its comparator groups in Rwanda; compared to AQ+SP(P\u200a=\u200a0.006) and AS+AQ(P\u200a=\u200a0.006); to AL in Uganda(P\u200a=\u200a0.005); and MAS3 in Thailand(P\u200a=\u200a0.016).Using multivariate analysis and the full length of follow-up stratified bysite and controlling for age and anaemia, the overall risk of recurrence waslower in the DP groups than in the comparator groups. DP provThere was a longer interval from primary infection to recurrence (suggestinga greater duration of suppressive prophylaxis) compared to MAS3 in Thailand; in Uganda compared to AL; in Rwanda compared toAQ+SP , but was not different toAS+AQ (P\u200a=\u200a0.23).P. falciparum compared to the other treatments.At 28 days the risk of new infection was greater in African compared to Asiansettings (P\u200a=\u200a0.001) reflecting the highertransmission intensity. In these high transmission areas, patients treatedwith DP were at lower risk for a new infection within 28 days compared toAQ+SP ,AS+AQ  in Rwandaand AL  in Uganda.Overall, in the multivariate analysis stratified by site, DP had a greaterpost treatment prophylactic effect (against new infections) at Day 28against In the DP groups, using multivariate analysis stratified by site at Day 28,age (as continuous variable) was the only predictor of recurrence andrecrudescence in Africa or Asia when analyzed separately. In Africanchildren, younger patients (per 1 year increase in age) were at higher risksfor recurrence , and recrudescence, as well asin Asian patients .No significant predictors of new infections were detected in Africanchildren, but in Asian patients younger patients were at higher risks fornew infections .Admission pre-treatment gametocytaemia was present in a median (range) of6.1% (0.9\u201341.9) of the patients. Using multivariateanalysis in DP groups, and controlling by site, younger patients, admissionanaemia, and a lower admission parasite count were related to a higher riskof patent gametocytaemia .Clearance of gametocytaemia was slower in DP groups than in the comparators,overall and in individual sites. In Cambodia in the DP treatment arm,82(95%CI 55\u201394)% of patients presenting withgametocytaemia still had gametocytaemia on Day 3 compared to only27(7\u201354)% in the comparator arm. On Day 14 of thefollow-up in Thailand 24(10\u201339)% in DP arm and11(2\u201330)% in the other arm, in Uganda7.4(2.4\u201316)% in DP arm compared to1.8(0.3\u201313)% in the other arm, in Myanmar31(23\u201338)% in DP arm as compared to none in thecomparator arm. Overall, using multivariate analysis, the risk of gametocytecarriage by Day 14 was lower in comparators than in DP groups.Overall 178 (8.4%) out of 2125 patients presenting withoutgametocytes developed gametocytaemia after starting treatment. Recurrentparasitaemia as well as anaemia on admission were associated with gametocyteappearance .There wIn Cambodia, Myanmar, and Thailand, the overall gametocyte carriage ratewithin 42 days was greater in the DP  compared to MAS3 groups . In patients without gametocytaemia on admission and who developedgametocytaemia during the follow-up, the carriage rate was also greater withDP (11/1000 PGW) than MAS3 (6/1000PGW).When using available data there was no relationship between the dose of DHA actually received by thepatient and the proportion of patients remaining with gametocytaemia on Day14 (P\u200a=\u200a0.382).On admission, 537 out of 1,797 (29.9%) DP recipients withavailable data were anaemic (Hct<30%). Of these29.8% (74/537) had severe anaemia (Hct<20%).Using multivariate analysis, anaemia on admission was strongly associatedwith age and varied by country . ChildreIn Cambodia, Laos, Myanmar, Thailand, there was a relative mean pairedtransient decline of 6.3% (95%CI 5.7\u20137.3) inhaematocrit from admission to Day 7 in DP groups. No difference was detectedcompared to the MAS3 group. By contrast, in Rwanda, between Day 0 and Day14, the relative mean paired haematocrit difference was significantly higherin the AS+AQ group  compared to the DP group (P\u200a=\u200a0.021).In Uganda, patients treated with DP  had a higher relative paired mean increasein haemoglobin levels on Day 42 compared to AL group.Overall, vomiting on admission and before treatment administration was a riskfactor for vomiting the first dose of DP and for vomiting DP treatment over the three days. This was similar in every country. In patients who didnot present with vomiting on admission, the overall incidence of earlyvomiting (defined as vomiting the drug within 1 hour after intake) DP waslow; 1.7%  on Day 0. Over the 3 days of treatment, theoverall incidence rate of early vomiting ranged from 3.2 (95%CI2.1\u20134.8)% in Thailand to 9.9 (95%CI6.8\u201314.4)% in Rwanda . In DP gWe examined the incidence and prevalence of 24 other different adverse events(apart from early vomiting) in 1,267 individuals using available recordsfrom DP groups in five countries and 9 different sites. It was not possibleto calculate the adverse events duration in Laos, and Uganda. In theremaining DP groups, the five most commonly reported adverse events by Day28 were 23.3% for headache, 17.0% for dizziness,13.8% for sleep disturbance, 11.6% for anorexia,10.5% for nausea . HyperseIn the Asian trials, the only adverse event that was significantly morefrequent in DP treated patients compared to the MAS3 group was diarrhoea inall age groups ; among them 38% reported one, and26% two adverse events (without excluding patients under 5). Thetotal number of adverse events reported per patient was higher in olderpatients (P\u200a=\u200a0.001) and in anaemicpatients on admission (P\u200a=\u200a0.020 aftercorrecting for age).The incidence of adverse events was significantly lower in DP recipientscompared to MAS3 recipients . More patients treated with MAS3reported two or more AEs . In Rwanda, the risk of havingany adverse event was higher in the AQ+SP group and in the AS+AQ group compared to the DP group. No differences were detectedin Uganda in the AL group compared to the DP group.A child from Rwanda had a seizure and received a rescue treatment. Overall, 5deaths occurred in DP groups, all of which were considered to be unrelatedto the treatment, except in Thailand where a 43-year-old woman who died fromsevere malaria the day after entering the study, which might have beenrelated to a lack of efficacy of the drug. In Thailand, there were another 2deaths: a 13-year-old girl who died on Day 7 from probable bacterial sepsis,and a 21 year old male who died on Day 28 from gunshot wounds. In Laos, atwo year old child died on Day 37 from cerebral malaria after probablereinfection. In Myanmar, one 11-year-old child died after developing feveron Day 20 and had generalized seizures the next morning (Day 21). Themalaria smear was negative. In the comparator groups, one death occurred inMAS3 groups in Thailand involving a malaria-smear negative 13-year-old boy,who was clinically well by the third day of treatment. He was reported tohave deteriorated rapidly with worsening abdominal pain and distension,jaundice, and anuria, and he died within a few hours.P. vivax recurrence rates. In Cambodia, Laos, Myanmar, andThailand, 265 patients receiving DP (28.8/100 person-days within 63 days)had P. vivax parasitaemia detected during the follow-up(P. vivax parasitaemia was 49(14\u201363) days compared with 49 (28\u201363) days, in MAS3groups. Patients with P. vivax on admission (mixedinfections) were at higher risk of having a second P. vivaxepisode during the follow-up . Children were at increased riskof P. vivax recurrence when compared to adults;0\u20134 age group  and the 5\u201314 age group. Inpatients who had mixed infections on admission, the median time toP. vivax recurrence was 42 days (7\u201363),significantly shorter than in patients presenting with a P.falciparum monoinfection: 49 (16\u201370) days,(P\u200a=\u200a0.029). In Uganda patients treatedwith DP had a lower risk of recurrent parasitaemia due to P.malariae and P. ovale species compared topatients treated with AL No difference was detected between DP groups and the comparator(mefloquine-artesunate) in ollow-up. ThemedP. falciparum malaria along the Thai-Myanmar border, therehas been increasing use of ACTs throughout the malaria affected world.Dihydroartemisinin-piperaquine (DP) is a relatively new and very promising fixeddose ACT which has been extensively evaluated in the past few years. This analysisof 3,547 patients  in randomized comparative clinicaltrials includes 7 of the 22 published studies (until December 2008), but is broadlyrepresentative as it comprises a wide patient age range and derives from areas ofwidely differing intensity of malaria transmission. It is the first individualpatient data set compiled prospectively, based on randomized comparisons in studiesof generally similar overall design. It includes half of all the patients treated with DP in published clinical trials with parasitegenotyping corrected results and reports 43%  of allpatients included in published studies to date.Since initial deployment in 1994 of the mefloquine-artesunate (MAS3) combination totreat To present the results of this analysis of individual patient data we have used amethod similar to that used for a meta-analysis of trials (for instance inCochrane's review) with graphical representation of risks, recommended forcommunicating in medical research In the 12 different African and Asian sites with varying levels of backgroundantimalarial drug resistance, DP was well tolerated and highly efficacious. Like allACTs DP produces rapid therapeutic responses with swift resolution of symptoms andfever and clearance of parasitaemia. Efficacy (PCR corrected) exceeded96% in all sites except one in Rwanda, where no food was co-administeredwith the drug. Treatment with DP was associated with lower risks of overallrecurrence (P\u200a=\u200a0.001), recrudescence(P\u200a=\u200a0.001), and new infections(P\u200a=\u200a0.001) compared with the comparator drugs. Thesuperiority of DP was demonstrated in Rwanda, Uganda and Thailand against the localcurrent first-line treatments. This excellent efficacy and tolerability profilesuggests that DP could be considered as a potential alternative first-lineantimalarial treatment. This result based on studies of 1,814 adults and childrenwho received antimalarial treatment with DP, contrasts with the recently reportedfinding of a study of 100 young children in Papua New Guinea DP was less potent than mefloquine\u2013artesunate in suppressinggametocytaemia. Similar results were also observed in Kenya in comparison with ALP. falciparum speciesco-infections. Unlike P. falciparum infections, recurrence ofP. vivax (either relapses or failures) cannot be reliablydistinguished from a new infection P. vivax strains DP was effective in treating non-P. vivax reappearance in children would also support apharmacokinetic explanation. The Day 7 piperaquine level is a useful measure of drugexposure. Young children have lower piperaquine levels on Day 7 and higher treatmentfailure rates than older children and adults 1/2,\u03b1), and a more rapid fall in initial PQ plasma concentrationscompared to the population mean profile The higher risk of treatment failure in children treated with DP compared with adultsis similar to the pattern seen with other antimalarial drugs, and presumably resultsboth from lower immunity and lower blood piperaquine concentrations. The shortertime to The risk of early vomiting of DP was lower than following AQ+SP in Rwanda(P\u200a=\u200a0.001). No differences were observed with theother comparator treatments. Early vomiting was more frequent in young children,although it was not a risk factor for treatment failure, presumably because thesechildren were re-dosed successfully after vomiting the drug.The DP safety profile has been excellent in all published series. The overall safetyanalysis showed that the risk of the most common adverse events was significantlylower following DP treatment than in the comparator arms in both children(<15 years old) and adults. Adverse events were often related to the diseaseitself , although diarrhea wasapproximately twice as common following DP than with MAS3.The use of common protocols for data collection to assess antimalarial drug efficacyand tolerability allows combination of these data into larger internationaldatabases which can give us more information on the safety and efficacy of thesedrugs in different patient groups DP is not yet recognized internationally and its use has been limited by itsregulatory status. The formulation used in the trials was donated by Holley andmanufactured according to Chinese SFDA Good Manufacturing Practice (GMP) standards.Nevertheless, differences in efficacy might be related to the variability in thecomposition of the study drug.This antimalarial combination is currently under evaluation by the WHOpre-qualification process. DP is clearly an important new antimalarial drug. It iswell tolerated, highly effective and safe. The higher rates of gametocytaemiacompared with other ACTs and lower piperaquine levels early in the terminalelimination phase observed in children suggest that dosage may have to be increasedin this important patient group.Table S1for :Recurre(0.07 MB DOC)Click here for additional data file.Table S2for : Risks o(0.05 MB DOC)Click here for additional data file.Table S3for : Day 28(0.12 MB DOC)Click here for additional data file."}
+{"text": "Intermittent preventive treatment in infants (IPTi) is a new malaria control tool. However, it is uncertain whether IPTi works mainly through chemoprophylaxis or treatment of existing infections. Understanding the mechanism is essential for development of replacements for sulfadoxine-pyrimethamine (SP) where it is no longer effective. This study investigated how protection against malaria given by SP, chlorproguanil-dapsone (CD) and mefloquine (MQ), varied with time since administration of IPTi.A secondary analysis of data from a randomised, placebo-controlled trial in an area of high antifolate resistance in Tanzania was conducted. IPTi using SP, CD, MQ or placebo was given to 1280 infants at 2, 3 and 9 months of age. Poisson regression with random effects to adjust for potential clustering of malaria episodes within children was used to calculate incidence rate ratios for clinical malaria in defined time strata following IPTi. The short-acting antimalarial CD gave no protection against clinical malaria, whereas long-acting MQ gave two months of substantial protection (protective efficacy (PE) 73.1%  and 73.3%  in the first and second month respectively). SP gave some protection in the first month after treatment ) although it did not reduce the incidence of malaria up to 12 months of age. There was no evidence of either long-term protection or increased risk of malaria for any of the regimens.Post-treatment chemoprophylaxis appears to be the main mechanism by which IPTi protects children against malaria. Long-acting antimalarials are therefore likely to be the most effective drugs for IPTi, but as monotherapies could be vulnerable to development of drug resistance. Due to concerns about tolerability, the mefloquine formulation used in this study is not suitable for IPTi. Further investigation of combinations of long-acting antimalarials for IPTi is needed.NCT00158574Clinicaltrials.gov  Intermittent preventive treatment in infants (IPTi) is a new control strategy for reducing the malaria burden in endemic countries of sub-Saharan Africa where transmission of malaria is high and malaria is an important cause of mortality and morbidity in infants. There is now sufficient evidence that three courses of sulfadoxine-pyrimethamine (SP) IPTi has a protective efficacy of 20\u201330% against clinical malaria in the first year of life; therefore SP-IPTi may be adopted as a policy in some African countries Although the first IPTi study found a suggestion of sustained protection To investigate these issues, an individually randomised, placebo-controlled trial of SP and two alternative regimens  was undertaken in an area of high SP resistance in Tanzania It could be informative to investigate how protective efficacy in the above study varies with time since treatment. While it is possible that the finding of no benefit overall for SP or CD could mean that there is genuinely no benefit at any time, it is also possible that there is either a small benefit initially after treatment which is too small to remain detectable over the whole of infancy. Alternatively, it may be that an initial benefit of treatment is cancelled out by a subsequent increased risk of clinical malaria, either through loss of premunition The purpose of the present study was therefore to investigate how the protective efficacy of the three drug regimens used in the above trial changed with time since treatment, and thus clarify the mechanism of protection given by IPTi.www.clinicaltrials.gov identifier: NCT00158574), and was approved by the Ethics Review Board of the National Institute for Medical Research of Tanzania and the London School of Hygiene and Tropical Medicine. Briefly, a double-blind, individually randomised, placebo-controlled trial of IPTi using SP, CD and MQ was undertaken in two districts in Tanzania with a high prevalence of SP resistance. Full treatment doses of the study drugs were given to children enrolled in the trial at the time of the second diphtheria-pertussis-tetanus/polio vaccine (DPT/polio 2), DPT/polio 3 and measles vaccinations at approximately 2, 3 and 9 months of age. The separate IPT rounds are referred to in this paper as IPT1, IPT2 and IPT3. Children were followed-up until two years of age for incidence of the primary outcome of clinical malaria (temperature \u226537.5\u00b0C or history of fever within 48 hours plus parasitaemia of any density).This study uses data from the Kilimanjaro IPTi Drug Options Trial, described in detail elsewhere et al. in vivo efficacy study of SP in this region showed a day 28 adequate clinical and parasitological response of only 18% The analyses presented here use only data from the moderate transmission site in Korogwe, Tanga Region, Tanzania. B\u00f8dker Children who had received and successfully swallowed a given round of IPT  were included in the analysis. The endpoint was clinical malaria, as defined above, detected passively at one of the study clinics. Time at risk was calculated from time of IPT treatment until completion of five months (150 days) of follow-up, receipt of a subsequent dose of IPT or exit from the study , whichever occurred first. As in the main trial analysis, children who had a malaria episode or who were treated with an antimalarial had 21 days removed from the person time at risk. To enable month specific estimates of the protective efficacy by time since treatment, follow-up time was stratified by month (30 days) since treatment using lexis expansion Poisson regression was used to calculate incidence rate ratios for clinical malaria, allowing multiple episodes of malaria per child. To adjust for potential clustering of malaria episodes within children, Poisson regression models with gamma distributed random effects were used. Multivariate models were built using an additive step-wise approach, using the likelihood ratio test to compare models and aiming for the simplest model that adequately explained the data. Since the principal interest of this study was the interaction between intervention group and time since treatment, this interaction was modelled first. Covariates known to be associated with the incidence of malaria in this population from the main trial were then added to the models .To explore the changes in protective efficacy against clinical malaria over time, malaria incidence rate ratios for SP, CD and MQ versus placebo by month since IPT were calculated. Protective efficacy and its 95% confidence interval was then calculated as 1-rate ratio. To obtain more precise estimates of protective efficacy over time, a summary analysis was then performed combining time at risk and malaria episodes following all three IPT rounds into a single analysis that related malaria incidence to time since most recent IPT. To account for possible differences in efficacy between separate IPT rounds, the IPT round from which time at risk and incidence was taken was tested for inclusion in the multivariate regression models as an interaction term with treatment group and as a covariate.In the analysis of the clinical trial, IPTi with MQ was shown to provide a protective efficacy against clinical malaria of 38.1%  in infants aged 2\u201311 months of age. There was no statistically significant protective effect from SP \u22126.7%  or CD 10.8%  during infancy It was not possible to calculate protective efficacy by month after IPT1, since most children received IPT2 one month later (75% within 28 days and 95% within 42 days of IPT1) and incidence of malaria in the placebo group in this period was very low . InclusiThere was no statistically significant evidence that SP gave any protection in any specific month after IPT2 . HoweverThere was no evidence of protection from CD in most months after treatment . There wThe point estimates of protective efficacy suggested some protection in the two months following IPT2, but the confidence interval overlaps unity . Beyond In the summary analysis of all 3 IPT rounds, there was strong evidence of a high level of protection in the first month ; p\u200a=\u200a0.013). The point estimate of protective efficacy was similarly high in the second month, but this was only of borderline statistical significance ; p\u200a=\u200a0.050). There was a suggestion of protection in the third month but this was not statistically significant. There is thus no strong evidence of any benefit or detrimental effect in months three, four or five after IPT with MQ.Mefloquine provided a high level of protection against clinical malaria in the first two months after IPTi, consistent with its extended plasma half-life and its presumed therapeutic efficacy in Africa SP provided some protection against clinical malaria in the first month after IPT, despite the high prevalence of antifolate resistance in the study site Chlorproguanil-dapsone provided little protection against clinical malaria compared to placebo. While there was a suggestion of some protection in the third month after IPT, largely due to apparent protection after IPT2, we consider this likely to be a chance finding given the borderline statistical significance and the lack of a plausible biological mechanism for protection to be delayed to such a specific and finite window after treatment. Chlorproguanil-dapsone was shown to be effective in treating acute malaria episodes that had failed treatment with SP in Tanzania Due to the relatively low incidence rate of clinical malaria in this study population, estimates of the intervention effect are less precise than would be ideal for this type of analysis, and the time-stratified analyses are likely to be underpowered to detect effects in specific time strata. This is particularly the case when examining the protective efficacy of IPT1 and IPT2. The low incidence rate also limits the minimum size of time strata for which sensibly precise estimates of protective efficacy can be calculated to around one month. The analytical approach includes multiple comparisons and thus it is possible some spurious low p-values may have appeared due to chance. However, in the present analysis, the directions of trends can still be seen, which combined with understanding of the pharmacodynamics of the antimalarials allows a biologically plausible interpretation to be made. With a larger trial or a higher incidence rate of malaria, it would have been possible to stratify follow-up more finely to get a more detailed picture of how protection varies over time, as was possible in a similar analysis of the Navrongo IPTi trial To assess if our findings could have been affected by the decision to stratify follow-up time by month, the analyses were repeated with different sized time strata. This did not alter the estimate of duration of protection or the interpretation (data not shown). We did not find any evidence of interaction between IPTi drug group and IPTi round (i.e. there was no suggestion that efficacy of IPTi compared to placebo varied between the different rounds), but it would be preferable to have greater precision to look at separate rounds in isolation.Short-acting antimalarials are unlikely to be a suitable choice for intermittent preventive treatment in infants. Longer-acting antimalarials with a duration of action similar to that of mefloquine and SP are likely to be more efficacious, since they would provide a substantial period of post-treatment prophylaxis. However, the mefloquine formulation used in this trial is not likely to be suitable for IPTi because it is poorly tolerated The finding of protection limited to a shorter period immediately after IPTi with SP suggests that malaria episodes are only prevented when plasma concentrations of both sulfadoxine and pyrimethamine are greatest and the two components act in synergy; this period may last for 15 days for antifolate resistant parasites The overall protective efficacy of mefloquine up to 12 months of age in this trial was 38.1% , which compares favourably with most of the trials of SP If long-acting drugs offer the best protection when used for IPTi, this raises an immediate dilemma. Slowly eliminated antimalarial drugs may be most prone to development of drug resistance Mefloquine, the most efficacious regimen in this IPTi trial, protected children by giving a substantial period of post-treatment prophylaxis. SP provided protection against clinical malaria for approximately one month, but it appears that some malaria attacks were simply delayed until the following month once drug concentrations had waned. Given concerns about possible side-effects and tolerability of mefloquine, and exacerbation of drug resistance, novel combinations of long-acting antimalarials should be investigated for use in IPTi.Protocol S1Trial Protocol(1.08 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium vivax infection, the parasite responsible for the majority of its malaria burden. Chloroquine resistance in P. vivax is emerging in Asia. Therapeutic responses across Afghanistan have not been evaluated in detail.Afghanistan's national guidelines recommend chloroquine for the treatment of P. vivax mono-infections was conducted. Consistent with current national guidelines, primaquine was not administered. Subjects were followed up daily during the acute phase of illness (days 0-3) and weekly until day 56. The primary endpoint was the overall cumulative parasitological failure rate at day 56 after the start of treatment, with the hypothesis being that dihydroartemisinin-piperaquine was non-inferior compared to chloroquine (\u0394 = 5% difference in proportion of failures).Between July 2007 and February 2009, an open-label, randomized controlled trial of chloroquine and dihydroartemisinin-piperaquine in patients aged three months and over with slide-confirmed P. vivax, 536 were enrolled in the trial. The day 28 cure rate was 100% in both treatment groups. Parasite clearance was more rapid with dihydroartemisinin-piperaquine than chloroquine. At day 56, there were more recurrent infections in the chloroquine arm  than the dihydroartemisinin-piperaquine arm , a difference in cumulative recurrence rate of 6.1% (2-sided 90%CI +2.6 to +9.7%). The log-rank test comparing the survival curves confirmed the superiority of dihydroartemisinin-piperaquine over chloroquine (p = 0.003). Multivariate analysis showed that a lower initial haemoglobin concentration was also independently associated with recurrence. Both regimens were well tolerated and no serious adverse events were reported.Of 2,182 individuals with positive blood films for vivax malaria in Afghanistan. In a setting where radical therapy cannot be administered, dihydroartemisinin-piperaquine provides additional benefit in terms of post-treatment prophylaxis, reducing the incidence of recurrence from 4-8 weeks after treatment.Chloroquine remains an efficacious treatment for the treatment of The trial was registered at ClinicalTrials.gov under identifier NCT00682578. Plasmodium vivax is associated with less mortality than Plasmodium falciparum, but still exerts a considerable burden of disease globally [P. vivax infection, but chloroquine resistance is spreading [globally  and is aglobally . In endeglobally . Controlpreading . High lepreading  and Indopreading , but lowpreading .P. vivax malaria and is also used in combination with sulphadoxine-pyrimethamine for presumptive treatment of clinical malaria episodes when accurate laboratory diagnosis is unavailable. Radical treatment with primaquine is not currently recommended because of the risk of oxidant haemolysis in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency, as testing for G6PD deficiency is not available. Limited access to prompt diagnosis and treatment, self-medication with low quality medicines [P. falciparum [P. vivax [P. falciparum in Kunduz province [P. vivax resistance to sulphadoxine-pyrimethamine is present at a relatively early stage in Nangarhar and Herat provinces[Chloroquine was used for more than 50 years as the first-line drug for all forms of malaria in Afghanistan. Chloroquine continues to be first-line drug treatment as a mono-therapy for slide-confirmed edicines  and non-lciparum  but appeP. vivax ,10. Relaprovince  and moleprovinces.P. falciparum infections in Afghanistan. As the correct malaria parasite species may not be identified in patients presenting with fever, P. vivax infections in Afghanistan will inevitably be treated with ACT [P. falciparum [vivax malaria, with more long-lasting post-treatment prophylactic effects than other forms of ACT [Since 2004, artemisinin combination therapy (ACT) in the form of artesunate/sulphadoxine-pyrimethamine has been the recommended first-line treatment for with ACT . For opelciparum  and seves of ACT ,14,15. DFor the purpose of evidence-based decision making, and to obtain better information on the effectiveness of available anti-malarial drugs against vivax malaria in Afghanistan, the relative therapeutic efficacy of chloroquine and dihydroartemisinin-piperaquine in the treatment of vivax malaria was assessed at three sites in Afghanistan.P. vivax remains endemic and accounts for more than 90% of all malaria cases [The study was conducted from July 2007 to February 2009 at three provincial malaria control centres, one in the east and two in the north of the country. The epidemiology of malaria in Afghanistan is determined by the physical geography of the country, malaria being confined to lower lying areas with sufficient rainfall for mosquito survival consisting of the northern plains  , the JalJalalabad, capital of Nangarhar province, is a referral centre for the whole eastern region and lies adjacent to the Pakistani border where population movement in both directions takes place. The population is multi-ethnic with Pashtoon being the dominant group. The province as a whole  is generally semiarid but where possible rice is cultivated. Taloqan is the capital of the north-east province of Takhar with a population that is mainly Tajik with significant proportions of Uzbek and Pashtoon. Rice is cultivated on the plain and malaria is a well-known health problem . MaimanaThis was a prospective, open label, randomized controlled trial conducted in patients with microscopically confirmed symptomatic vivax malaria who were aged three months and older. All febrile patients over three months of age presenting to the sites were screened and enrolled according to preset criteria. A standard data sheet was used to record demographic information, details of symptoms and their duration, and previous anti-malarial medications. Clinical examination findings and vital signs were documented, including axillary temperature measured with a digital thermometer. Venous blood samples were obtained for haemoglobin measurement (Hemocue) and white cell count using the counting chamber method. Giemsa-staining of thick and thin blood films was performed to confirm parasite species and parasitaemia by a standard approximation method (40 \u00d7 number of parasites per 200 white blood cells on the thick film). 10% of slides were cross-checked in the MORU (Bangkok) parasitology laboratory. Approximately 20 \u03bcl capillary blood was collected on filter papers for subsequent confirmation of species by PCR and future molecular studies. Slide examination results were considered to be negative after examination of 30 high-power fields of the thick film.P. vivax, written informed consent by the patient or attending parent/guardian, and a negative urine pregnancy test in women of childbearing age. Exclusion criteria were any clinical or laboratory feature of severe malaria [Plasmodium infection, pregnancy (or unwillingness to undergo pregnancy testing), lactation, and anticipated inability or unwillingness to complete the 56 day follow-up.The inclusion criteria were slide-confirmed infection with asexual stages of  malaria , haemoglPatients were allocated to the two treatment arms based on a pre-generated randomization list made in blocks of 20 that was produced and held independently of the field teams by a statistician. The individual allocations were kept in sealed, opaque envelopes and opened only after enrolment by the field team. Patients and clinical field workers were, therefore, not blinded to the treatment arm after allocation. Microscopists were blinded to treatment allocation at follow-up examinations.\u00ae, each tablet containing 40 mg dihydroartemisinin and 320 mg piperaquine; Holleypharm, China) aiming for a target total dose of 6/48 mg/kg in divided doses over three days. According to the body weight, individual doses were either rounded to the nearest half or quarter tablet or, if necessary, tablets were crushed, mixed with 5 ml water and the appropriate volume was given. All doses were taken under supervision at the malaria treatment centre and patients were observed for 60 minutes after drug administration. Patients vomiting within this time received the same dose again. Patients who vomited the medication twice were withdrawn from the study.Patients received either quality-controlled chloroquine  aiming for a target total dose of 25 mg base/kg in divided doses over three days, or dihydroartemisinin-piperaquine  and followed up by standard protocols. Patients with mixed P. falciparum/P. vivax infection were also classified as treatment failures but treated with artesunate/sulphadoxine-pyrimethamine. Patients were censored from the analysis if there was stated withdrawal of consent at any stage, persistent vomiting during the acute phase  or occurrence during the follow-up of concomitant disease that would interfere with a clear classification of the treatment outcome . Patients failing to attend follow-up visits received a visit at home and were asked to reattend the centre for assessment. Transportation was provided for all follow-up visits for all subjects. All enrolled patients were given one long-lasting insecticide treated bed net.The subjects were seen on days 0 to 3 inclusive and weekly to 56 days, so that late recrudescence and relapses could be detected ,24. PatiP. vivax infection at or before day 56 between the two arms. Secondary outcomes were parasite and fever clearance times, recurrence at earlier time points, gametocytaemia at day 28 and 56, haemoglobin recovery, and safety and tolerability of the treatments.The primary outcome was the difference in the proportion of patients with parasitologically-confirmed recurrence of Given the relatively high predicted efficacy of chloroquine, a non-inferiority design was used  with theData were double entered into Microsoft Access. All analyses were conducted using STATA version 9.0. The Student's t-test, Mann-Whitney U and chi-squared (or Fisher's exact) tests were used for comparison of baseline variables, as appropriate. Parasitological failure rates were assessed by survival analysis using the Kaplan-Meier method and Wilson confidence intervals for the difference in efficacy proportions were calculated using the effective sample size . DihydroP. vivax and P. falciparum as described previously [DNA was extracted from filter-paper blood spots and nested PCR performed to detect eviously .http://www.clinicaltrials.gov/ct as NCT00682578. This description of the trial describes recruitment of patients with uncomplicated infection with either P. falciparum or P. vivax, with treatments depending on the species detected. Recruitment of P. falciparum cases proved much slower than for P. vivax and the ongoing P. falciparum study will be presented elsewhere.The study was approved by the Ethics Committee of the Faculty of Tropical Medicine, Mahidol University, Thailand, the Oxford Tropical Research Ethics Committee, Oxford University, UK and the Institutional Review Board of the Afghan Public Health Institute, Ministry of Public Health, Afghanistan. The trial was registered with the clinical trials website P. falciparum (n = 44) and severe malaria (n = 12). The majority of patients (314/536 (58.6%)) were recruited in Jalalabad; 119 (22.2%) patients were recruited in Taloqan and 103 (19.2%) in Maimana. 73 (13.6%) patients were less than five years of age, 258 (48.1%) were 5-14 years of age and 205 (38.2%) were 15 or older.From July 2007 to February 2009, 536 patients were enrolled in the study with 268 receiving chloroquine and 268 dihydroartemisinin-piperaquine Figure . The maiP. vivax monoinfection; the remaining slides were classed as Plasmodium infection of undetermined species because of slide quality. PCR confirmation was undertaken on an independent, randomly selected subset of paired blood samples from 10 patients with parasitological failure. All 10 baseline samples and 9 of 10 samples taken at recurrence contained P. vivax by this method. P. falciparum was not detected in any sample by either microscopic or molecular methods.At the reference laboratory 94% of baseline slides were confirmed as 9/l respectively, p = 0.02).Patient characteristics at baseline were broadly similar between the two treatment groups Table  except tPatients in the chloroquine group who completed treatment received a median total dose of 25.9 mg/kg base (IQR 25.0 - 27.3 mg/kg); for the dihydroartemisinin-piperaquine group the median total dihydroartemisinin dose was 6.0 mg/kg (IQR 5.45 - 6.67 mg/kg). Both treatments were generally well tolerated. Repeated vomiting of the study medication was not reported within 60 minutes of drug administration in any patient. No patient deteriorated or developed signs of severity. Five patients who received the first dose withdrew consent before completing treatment and were censored from analysis. Of the 531 individuals who completed treatment, 38 (7.2%) cases were lost to follow-up before day 56; 13 in the chloroquine arm and 25 in dihydroartemisinin-piperaquine arm (p = 0.04)  in the chloroquine group and 7 in the dihydroartemisinin-piperaquine group, giving a day 56 failure rate of 8.9% (95% CI 6.0 - 13.1%) in the chloroquine group and 2.8% (1.4 - 5.8%) in the dihydroartemisinin-piperaquine group  yielded a difference in cumulative parasitological failure rate between chloroquine and dihydroartemisinin-piperaquine of 1.5% (2-sided 90% CI -3.3 to +6.3%); as predicted there was a wider 90% confidence interval than that based on survival analysis but also a lower point estimate for the difference in failure rate between the chloroquine and dihydroartemisinin-piperaquine groups caused by the greater number of losses to follow-up in the dihydroartemisinin-piperaquine group. The lower bound of the 90% confidence interval was still higher than the prespecified non-inferiority margin (5%), but included zero, indicating that dihydroartemisinin-piperaquine was not inferior to chloroquine even in this considerably more conservative analysis   dihydroartemisinin-piperaquine was again superior Figure , with 29The day 28 cure rate was 100% in both arms with the first recurrences observed at day 35 in the chloroquine group and day 42 in the dihydroartemisinin-piperaquine group. Twenty-one of 30 (70%) recurrent parasitaemias were associated with gametocytaemia, with no significant difference between treatment arms in terms of proportion of recurrent patients with gametocytaemia .There were no serious adverse events recorded during the trial and no patient required hospitalization. Both study drugs appeared to be well tolerated. There was a significant increase in the proportion of patients with pruritus in the chloroquine arm compared to dihydroartemisinin-piperaquine on days 1, 2 and 3; on day 1, 15/268 subjects in the chloroquine arm had pruritus compared to 2/266 in the dihydroartemisinin-piperaquine arm , consistent with the known side-effects of chloroquine. None of the other signs or symptoms documented in the first three days of treatment were found to be significantly different between the groups. There were no significant differences between the two arms in haemoglobin concentration or occurrence of anaemia (haemoglobin < 10 g/dl) at any stage during follow-up .A univariate analysis undertaken to identify variables associated with failure showed that treatment allocation (chloroquine rather than dihydroartemisinin-piperaquine), study site , younger age and lower haemoglobin concentration at baseline were all significantly associated with recurrent infection. Admission parasitaemia, presence of gametocytes at admission and gender had no effect in this model Table .Multivariate analysis of the same seven variables entered into the univariate analysis indicated that treatment with chloroquine and lower haemoglobin concentration at baseline were the only two significant independent predictors of recurrent infection (Adjusted Hazard Ratio (AHR) 3.66, 95% CI 1.48-9.02, p < 0.001 for treatment allocation and AHR 0.50, 95% CI 0.37-0.66, p = 0.005 for haemoglobin concentration).P. falciparum [P. vivax [P. vivax remains sensitive to chloroquine in the study area, which included locations both north and south of the Hindu Kush divide of central Afghanistan. These data extend the geographic range of efficacy monitoring beyond Eastern Afghanistan where chloroquine has been shown previously to be efficacious in clearing P. vivax parasitaemia [P. vivax chloroquine-resistance manifests as frequent failure to clear parasites by day 4 [This trial in three separate malaria areas of Afghanistan confirmed that both chloroquine and dihydroartemisinin-piperaquine are well-tolerated and broadly efficacious treatments for vivax malaria. Initial parasite clearance was significantly faster after dihydroartemisinin-piperaquine, consistent with the pharmacodynamic properties of artemisinin-containing therapies observed in lciparum  and P. vP. vivax ,24. Nevesitaemia ,10. Thesby day 4 ,29-31 anby day 4 .P. vivax infections in the chloroquine arm, compared to 3% in the dihydroartemisinin-piperaquine arm, with all failures occurring on or after day 35. Recurrences of P. vivax may consist of a mixture of relapses from liver hypnozoites, recrudescences of the erythrocytic infection (due to inadequate drug levels or resistance), and reinfections acquired from additional innoculations. It is not possible with current methodologies to distinguish reliably between these possibilities [P. vivax hypnozoites. The rapid clearance of parasites in the chloroquine group, and the fact that failures were not seen before day 28, suggest that recrudescences associated with chloroquine-resistance did not contribute significantly to the number of recurrences. For these reasons, the majority of recurrences observed in this study are likely to have been relapses.Over the 56-day follow-up period dihydroartemisinin-piperaquine proved superior to chloroquine in terms of the prespecified survival analysis of recurrence rates. Approximately 9% of cases had recurrent bilities ,33. Howebilities -36. In tP. vivax is prevalent DHA-PPQ has been shown in several comparative trials to prevent recurrences of P. vivax over a six-week period more effectively than ACT containing partner drugs with shorter half-lives [The pattern of recurrences is consistent with the known properties of the drugs under study. Chloroquine is effective in preventing early relapses (<28 days) of vivax malaria in chloroquine-sensitive vivax malaria ,10,36-39lf-lives ,14,15. IP. vivax infection in Papua New Guinea, high levels of chloroquine-resistance were present, as evidenced by prolonged parasite clearance times in the CQ-SP arm. There was an 87% rate of recurrence in the first 42 days after CQ-SP compared to 30.6% in the dihydroartemisinin-piperaquine arm [P. vivax, and confirms the longer lasting post-treatment prophylactic effect of dihydroartemisinin-piperaquine compared to chloroquine in this setting. The observed proportion of chloroquine-treated patients with recurrence over 56 days is less than in a study undertaken in the Jalalabad Malaria Reference Centre in 2004 [In the only previous trial directly comparing dihydroartemisinin-piperaquine to chloroquine  for uine arm , a diffe in 2004  , younger age was found to be a risk factor for recurrence, a finding consistent with previous studies examining this effect over longer follow-up periods ,37. PossIt is possible that the action of dihydroartemisinin-piperaquine is simply to delay relapses of vivax malaria rather than prevent them; in other words the survival curves for recurrence with dihydroartemisinin-piperaquine and chloroquine will eventually merge. Studies with longer-term follow-up would be needed to determine if this is the case and if dihydroartemisinin-piperaquine offers tangible benefits in terms of long-term health of the individual (preventing relapse and associated anaemia) and possibly the community (by reducing transmission). Any health benefits have to be balanced against the relative cost of each treatment course, which is currently in favour of chloroquine use. These issues also have to be considered in the context of the challenges of treating patients with radical therapy in this region ,36.P. vivax malaria across three regions of Afghanistan. Consistent with the known pharmacological properties of the drugs, dihydroartemisinin-piperaquine provides a longer period of post-treatment prophylaxis, reducing the incidence of recurrence from 4-8 weeks after treatment.Both dihydroartemisinin-piperaquine and chloroquine are efficacious treatments for The authors declare that they have no competing interests.GRA, SP, MI, AMD, SJL, NPJD, PS, NJW and FK were involved in the conception and design of the study. GRA and FK were responsible for supervising patient recruitment and follow-up. MI undertook species confirmation by PCR. GRA, SP, CJW and SJL participated in the statistical analysis. GRA, SP, CJW, SJL and NJW drafted and critically revised the manuscript. All authors read and approved the final manuscript."}
+{"text": "Fixed-dose combination antimalarial drugs have played an increasingly important role in the treatment and chemoprophylaxis of falciparum malaria since the worldwide failure of monotherapy with chloroquine. Atovaquone-proguanil is one such combination drug used both for prophylaxis in travellers, and for treatment of acute malaria cases in European hospitals and clinics.pfcyt-b gene, previously found to be associated with failure of the atovaquone component.A series of eight atovaquone-proguanil treatment failures and two prophylaxis breakthroughs from four UK hospitals from 2004\u20132008 were analysed for evidence of mutations in the pfcyt-b mutations were found in five falciparum malaria patients with recrudescent parasitaemia occurring weeks after apparently successful treatment of a primary infection with atovaquone-proguanil. Four of these cases carried parasites with the Tyr268Cys mutation in pfcyt-b, previously reported in two French patients with malaria. In contrast, mutations in pfcyt-b were not found in three patients treated with atovaquone-proguanil who exhibited delayed clearance of the primary infection, nor in two returning travellers with malaria who had used the combination for prophylaxis. Using current and previously published data, mean time to recrudescence of parasites carrying pfcytb codon 268 mutations was estimated as 28.0 days after treatment (95% C.I. 23.0 \u2013 33.0 days), whereas treatment failures without codon 268 mutations received rescue treatment an average of 4.71 days after initial AP treatment (95% C.I. 1.76 \u2013 7.67 days).Parasites carrying pfcyt-b may have been overlooked in previous studies of atovaquone-proguanil treatment failure as it is not detected by current RFLP methods.Genetically-determined parasite resistance to atovaquone is associated with delayed recrudescence of resistant parasites three weeks or more after initial clearance of parasitaemia by atovaquone/proguanil therapy. The 268-Cys allele of  Plasmodium falciparum malaria [de novo mutations in the parasite's pfcyt-b gene that arise from increased oxidative damage to mitochondrial DNA generated by the action of atovaquone [\u00ae).The hydroxynaphthoquinone compound atovaquone was developed in the 1990s as an anti-protozoal drug, with demonstrated activity against cytochrome b in pathogen mitochondria. High rates of post-treatment recrudescence in patients treated with atovaquone alone for ovaquone . This lepfcyt-b mutations in African parasite isolates, mostly following atovaquone monotherapy or treatment with AP [pfcyt-b mutations Tyr268Ser or Tyr268Asn [in vitro sensitivity to atovaquone compared to that of parasites carrying codon 268 mutations in pfcyt-b [AP is widely used for malaria prophylaxis in travellers, and for treatment of clinical malaria cases in high income countries. It is not used as therapy in sub-Saharan Africa due to its high cost, but there are reports of  with AP -6. Malaryr268Asn ,7, altho pfcyt-b .P. falciparum cyt-b genes from malaria patients with recurrent parasitaemia following treatment with AP in four UK settings, or following use of AP prophylaxis in a malaria endemic area. The relevance of these results for malaria treatment policy in the UK is discussed.DNA sequencing analysis was performed on pfcyt-b gene was carried out as part of the surveillance remit of the Malaria Reference Laboratory (MRL), and performed either at the Department of Clinical Parasitology, Hospital for Tropical Diseases (HTD) or at the LSHTM Malaria Reference Laboratory (MRL), using identical PCR and sequencing protocols. The MRL is mandated by the UK Health Protection Agency to provide molecular surveillance in the form of genetic typing of P. falciparum for drug resistance-associated alleles, to assist in development of national policy on prevention and treatment of imported malaria. The work described was carried out under this mandate. All patient identifiers have been removed from this report.Parasites were isolated from three categories of patients with falciparum malaria: those whose clinical and parasitological response to AP was delayed in the first three days following AP treatment, those with microscopically confirmed recrudescent parasitaemia following initially successful treatment with AP, and three cases where AP had been taken as prophylaxis during travel in an area of malaria endemicity. Genotyping of the pfcyt-b gene was amplified using primers cyt-b1 (5'CTC TAT TAA TTT AGT TAA AGC ACA 3') and cyt-b2 (5' ACA GAA TAA TCT CTA GCA CC 3') by conventional PCR with standard regents. Reaction conditions were used as follows: samples were heated initially at 93\u00b0C for 10 minutes and then at 93\u00b0C for 50 s, 45\u00b0C for 50 s and 70\u00b0C for 1 min over 50 cycles. PCR products were purified by fractionation on a 1% agarose gel and elution with the MinElute Gel Extraction Kit .A 939 bp portion of the PCR products were sequenced using the ABI Prism Big Dye Terminator kit, using the amplification primers and a third primer, cyt-b7 (5'-CAA TTA CTA AAC CAG CTG G-3'), to prime the sequencing reactions. Electropherogram data were edited, collated and analysed using Chromas software . All DNA sequences were confirmed by at least two independent sequence reads.pfcyt-b gene was determined for each suspected atovaquone-resistant parasite isolate. These were obtained from the following patients:The sequence surrounding codon 268 of the Patient 1 was a 29 year old male, admitted in October 2004 with uncomplicated falciparum malaria (parasitaemia 1.1%). Although ethnically of African origin, he was born and resident in the UK. Approximately six weeks prior, this patient had been treated for falciparum malaria acquired in Sierra Leone with a full course of AP (four tablets daily for three days). His symptoms resolved at that time, although parasitological confirmation of cure was not obtained. The recrudescent infection was treated with quinine/Fansidar\u00ae (sulfadoxine/pyrimethamine). Parasite DNA was extracted from two sequential peripheral blood samples taken nine hours apart at the time of his recrudescent infection.Patient 2 was a 44 year-old male who presented in August 2004 with 1% P. falciparum parasitaemia after travel in Nigeria, where an unspecified antimalarial injection had been received some weeks earlier. His previous history of exposure and thus immune status with respect to malaria is unknown. Parasitaemia rose under treatment with AP to 4% with continuing symptoms on day 3 of admission. Treatment was then switched to intravenous quinine. DNA was extracted from a single pre-treatment sample.Patient 3 was a 60-year old female treated with a full course of AP for falciparum malaria in July 2004 following a trip to Nigeria. She was a UK resident who had been brought up in Nigeria. There were no clinical features of severe disease, although initial parasitaemia was 2.5%. Sixty-eight hours following the start of her course of AP she returned with persisting symptoms and a parasitaemia of 0.1%. She was commenced on a course of quinine therapy and rapidly improved. A blood film taken the following day was parasite negative.Patient 4, a five-year old boy, presented in October 2004 with a 0.1% P. falciparum parasitaemia following a trip to Nigeria. Although ethnically African, he had lived all his life in the UK. He made a prompt and unremarkable clinical recovery following a full course of AP. Symptoms recurred 23 days later and the child re-presented after a further two days with a 0.3% parasitaemia. DNA was extracted from a peripheral blood sample taken at the time of his recrudescent infection.Patient 5, a 21-year old male resident in London for seven years but having spent his childhood in Sierra Leone, presented with a P. falciparum parasitaemia of 0.8% in September 2006 following a visit to his country of origin. He reported full adherence to AP prophylaxis (one tablet daily).Patient 6 was a 56-year old female who presented to hospital with microscopy-confirmed falciparum malaria within a few days of returning to her home in London from Nigeria in August 2006. The patient informed hospital staff of a known allergy to quinine, suggesting suspected malaria infections had been treated in the past. She was treated with a full course of AP. Persisting parasitaemia at 1% after an additional 4th day of AP treatment precipitated a change to quinine with antihistamine cover, which cleared the parasites. A blood sample was sent for DNA sequencing analysis.Patient 7 was a 30-year old male born and brought up in Uganda but was a UK resident at the time of the malaria episode. He was diagnosed with falciparum malaria in Switzerland in September 2007 after travel to Uganda, and reported full adherence to a three day therapeutic course of AP. Parasite clearance was not confirmed microscopically. He presented again in London 21 days after treatment with 0.2% P. falciparum parasitaemia and was treated with a course of quinine. DNA was extracted for analysis from a blood sample collected at the time of his recrudescent infection.Patient 8 was a 36-year old male presenting with uncomplicated falciparum malaria, parasitaemia less than 0.1%, in June 2008 after visiting Uganda and Kenya without taking malaria prophylaxis. This individual was born and brought up in Uganda, but had been resident in the UK for 21 years. He was treated with a full 3-day course of AP and resolution of symptoms was reported over the following two days. His symptoms returned 21 days later and a repeat blood film confirmed recrudescent P. falciparum parasitaemia of less than 0.1%. Parasite material was retrospectively retrieved from a Giemsa-stained thin film from the primary episode and compared with results obtained from the blood sample collected at the time of recrudescence.Patient 9 was a 47-year old male presenting with uncomplicated falciparum malaria (parasitaemia < 0.1%) in July 2008 after visiting Sierra Leone. He reported good adherence to AP prophylaxis.Patient 10 was a 47-year old British female who worked in a rural school in The Gambia from March 2008 to June 2008, and taking mefloquine prophylaxis for that period. On 30th June, having discontinued chemoprophylaxis, she travelled to Senegal and within 3 days developed fever. She received a presumptive diagnosis of malaria, and took a 3 day course of AP from a back-up supply she carried with her, and reported that her symptoms resolved. This individual returned to the UK on the 8th of July. She took 1 further dose of mefloquine from her prophylaxis course on return to the UK. She attended her GP 4 weeks later with fatigue and headache, and a blood film was P. falciparum positive with a parasite count of 3%. The patient was treated successfully with quinine followed by doxycycline.pfcyt-b locus are presented for these ten patients in Table Sequencing data for the pfcyt-b gene were identified among the parasite isolates investigated: patient 4 carried parasites with the Tyr-268-Ser allele of pfcyt-b as previously described [P. falciparum infection at least three weeks after a primary malaria episode which initially resolved with AP treatment. No cases harbouring the Tyr-268-Asn allele of pfcyt-b were identified, and no mutations were identified at any other codons of the pfcyt-b gene. During his first (June) episode of malaria, Patient 8 harboured parasites with the wild-type Tyr at codon 268 of pfcyt-b, but 26 days later only parasites with Cys encoded at position 268 were identified.Two different mutations at position 268 of the escribed , whereasescribed . Each ofP. falciparum infections in the first 72 hours following initiation of AP treatment, precipitating a change in therapy (to quinine in each case). Each patient carried parasites with pfcyt-b genes that were wild-type at codon 268. Patients 5 and 9 reported good adherence to AP prophylaxis during visits to malaria endemic zones. Plasmodium falciparum parasites isolated from both of these patients were wild type at codon 268 of the pfcyt-b locus.Patients 2, 3 and 6 exhibited delayed clearance of pfcyt-b is associated with recrudescence of parasites at least 20 days after apparently successful AP therapy. To test this possibility, data from this series of eight patients for whom AP therapy failed were analysed together with ten such patients described by Musset et al. [pfcyt-b presented with recrudescent infections on average 28.4 days after treatment (95% C.I. 23.9 \u2013 32.9 days). Therefore, carriage of mutations in pfcyt-b is significantly associated with delayed recrudescent infections in these two studies of malaria patients presenting to European hospitals.These results suggest that genetically determined resistance to atovaquone at codon 268 of t et al. . ProphylP. falciparum infections. AP is also widely employed as first-line treatment for cases of uncomplicated falciparum malaria in a number of London hospitals. Reports of AP treatment failure in the UK and Europe describe substitution of Tyr with Asn, Ser or Cys at codon 268 of the pfcyt-b gene in recrudescent parasites, typically three to five weeks after the initial malaria infection has been cleared by AP treatment [P. falciparum recrudescence three to six weeks after initially successful AP treatment for uncomplicated malaria in travellers are described here. Parasites from one of these patients carried the Tyr-268-Ser mutant allele of pfcyt-b and the remaining four carried the Tyr-268-Cys allele. The 268-Cys allele has recently been reported in two French patients with P. falciparum malaria [cyt-b genes of drug-selected Plasmodium yoelii isolated from atovaquone-treated mice [Atovaquone-proguanil (AP) has been widely adopted in the UK as a chemoprophylactic agent for travellers likely to be exposed to reatment . Five fu malaria , but waspfcyt-b sequence of recrudescent parasites with that of parasites present 26 days earlier. As previously observed [pfcyt-b locus only among recrudescent parasites suggests that this mutation arose de novo in this individual due to AP selection. However, pfcyt-b mutations have been found among African parasite isolates without a history of atovaquone exposure [pfcyt-b loci, suggesting spread of a restricted number of mutant clones, would be instructive [In one case, patient 8, it was possible to compare the observed , the apptructive .pfcyt-b loci. The poor response to treatment in these patients infected by apparently atovaquone-sensitive parasites is consistent with the findings of Musset et al [in vitro sensitivity to atovaquone, and did not carry mutations at codon 268 of the pfcyt-b locus. Malabsorption of atovaquone is thus one possible cause of reduced drug plasma levels and poor AP treatment response in the first few days following initiation of therapy. Drug levels were not determined for any of the patients described here, but such measurements would have strengthened the study.In contrast, three patients exhibiting persistent parasitaemia after 60 \u2013 90 hours of AP treatment harboured parasites with wild-type et et al , who demP. falciparum infection despite AP prophylaxis-use harboured parasites with a wild-type pfcyt-b locus. It is unclear whether the failure of prophylaxis in these two cases was due to poor absorption of atovaquone, sub-optimal adherence to the prophylaxis regimen or other unknown parasitological or host factors. Measurement of plasma levels of atovaquone in these patients may have helped to resolve these possibilities. A third UK traveller who used mefloquine prophylaxis for only part of her time at risk of malaria infection, patient 10, reported self-treatment with AP for fever while in Senegal. Thus the parasitaemia diagnosed in the UK is almost certainly a recrudescence of the first infection treated with AP some weeks earlier and so this case is consistent with the general observation that pfcyt-b mutations are associated with late recrudescent P. falciparum parasitaemia. The partial use of mefloquine prophylaxis in this particular case history, including an additional dose after return to the UK, one month before the appearance of recrudescent parasites, raises the possibility these parasites were also mefloquine resistant.Two travellers presenting in the UK with pfcyt-b are associated with delayed recrudescence of parasites after AP therapy is only partly in agreement with the findings of a large multicentre study, in which several cases of AP treatment failure were not associated with pfcyt-b mutations [pfcyt-b. However, Wichmann and colleagues used the RFLP method of Schw\u00f6ebel et al [pfcyt-b mutations. Examination of the respective recognition site sequences verified that, firstly, the restriction enzymes employed in this method, NsiI and AlwNI, will not digest the Tyr-268-Cys allele found in three patients reported here and secondly, that this allele would be erroneously identified as a wild-type Tyr due to digestion only by the enzyme SspI. Therefore, it is possible that the remaining three individuals described by Wichman et al [pfcyt-b gene were ruled out by sequencing a 716 bp PCR fragment, no data are shown to indicate whether or not the Tyr-268-Cys allele was or was not present. The results of the present study, and those of Musset et al [The conclusion that mutations at codon 268 of utations . At leasel et al  to identan et al  as havinet et al , emphasiP. falciparum malaria from Jan 2004 to December 2007. During this period only three apparent treatment failures have been confirmed as carrying parasites with pfcyt-b mutations, suggesting a resistance rate of approximately 1%. It is also worth noting that two of these cases occurred in non-immune patients. Bearing in mind that approximately 90% of falciparum malaria cases at St. Thomas' hospital affect semi-immune individuals who were born and brought up in endemic areas, this suggests that the acquired immunity to malaria enjoyed by many malaria patients may reduce the risk of occurrence of recrudescent infection associated with pfcyt-b mutations, as previously observed in studies of atovaquone monotherapy [The findings reported here can inform both treatment policy and advice given to prospective travellers to malaria endemic areas. Current data suggest that AP affords highly effective prophylaxis for endemic region travel, and is an efficacious treatment for uncomplicated cases of imported malaria. However studies with adequate follow-up of both treated cases and prophylaxis users are needed to verify these observations. Denominators for our study are difficult to estimate and, in the absence of active follow-up, some treatment failures may have presented for re-treatment elsewhere or not at all. Estimates from one of the hospitals participating in this study (St Thomas') are that AP alone was used to treat approximately 280 patients with confirmed et al [in vivo.The results presented here also provide some support for the suggestion of Musset et al  that treet al . PrelimiP. falciparum isolates from malaria patients in the UK suspected of AP treatment (or prophylaxis) failure were tested for mutations in the pfcyt-b locus by direct DNA sequencing of PCR amplified products. Five patients with late, recrudescent treatment failure harboured mutations at position 268, and four of these were the recently described Tyr-268-Cys allele. This allele would not have been detected by restriction fragment analyses used in other studies. Three patients with apparent early treatment failure did not carry mutations, and may have failed to clear parasites due to low bioavailability of atovaquone. For purposes of drug resistance surveillance, DNA sequencing of the pfcyt-b locus is recommended in cases of late recrudescence of P. falciparum malaria, but may not be indicated where initial treatment response is delayed.A series of The authors declare that they have no competing interests.CJS conceived the study, performed experiments, analysed data and wrote the first draft of the paper. ML contributed to study design, performed experiments, analysed data and contributed to writing the paper. NP, QF and GP contributed patient samples or reagents and contributed to writing the paper. MB performed experiments and analysed data. JLK and PLC contributed to study design and analysis, provided samples and edited the second draft of the paper."}
+{"text": "Currently, there are few field-adapted analytical techniques for monitoring amodiaquine utilization in patients. This study evaluates the field applicability of a new method to determine amodiaquine and its metabolite concentrations in whole blood dried on filter paper.Artemisinin combination therapy (ACT) has been widely adopted as first-line treatment for uncomplicated falciparum malaria. In Uganda, amodiaquine plus artesunate (AQ+AS), is the alternative first-line regimen to CoartemTwelve patients aged between 1.5 to 8 years with uncomplicated malaria received three standard oral doses of AQ+AS. Filter paper blood samples were collected before drug intake and at six different time points over 28 days period. A new field-adapted sampling procedure and liquid chromatographic method was used for quantitative determination of amodiaquine and its metabolite in whole blood.The sampling procedure was successively applied in the field. Amodiaquine could be quantified for at least three days and the metabolite up to 28 days. All parasites in all the 12 patients cleared within the first three days of treatment and no adverse drug effects were observed.The methodology is suitable for field studies. The possibility to determine the concentration of the active metabolite of amodiaquine up to 28 days suggested that the method is sensitive enough to monitor amodiaquine utilization in patients. Amodiaquine plus artesunate seems effective for treatment of falciparum malaria. AL, however, has several limitations including twice-daily dosing regimen and recommendation to administer the drug together with fatty food [Artemisinin combination therapy (ACT) has been widely adopted as first-line treatment for uncomplicated falciparum malaria -3. Althotty food .Amodiaquine (AQ) is a 4-aminoquinoline drug that has demonstrated sustainable efficacy compared to other anti-malarial drugs -9. In viGenerally, most of the malaria endemic areas are rural. These areas usually suffer from shortage of resources, for example electricity, which is required during collection of samples such as plasma. Furthermore, adsorption and drying of whole blood samples on filter paper is known to reduce the risk of exposure to HIV, hepatitis B and C virus, and other infectious agents , thus maThus, in order perform clinical studies necessary for evaluation of AQ+AS impact in such areas, field-adapted sample collection and robust analytical techniques for monitoring AQ levels in blood and other body fluids are needed. Methods for determination of amodiaquine and its metabolite, desethylamodiaquine (DAQ) from plasma, whole blood and urine have been reported -18, but A new field-adapted filter paper method was recently developed for determination of amodiaquine from whole blood spotted on filter paper . The aim\u00ae from government for malaria treatment.The study was conducted in Aduku Health Centre, Apac District in Northern Uganda between January and February, 2008. Aduku is one of the sentinel sites that were chosen by Uganda Malaria Surveillance Project for evaluating the efficacy of available anti-malarial therapies and monitoring adverse drug events. It is surrounded by a large swamp and has very high malaria transmission intensity. The location is rural and, therefore, possibly has no drug pressure as there has been no supply of AQ or AS. The health centre receives free supplies of CoartemConsent forms were administered to all parents/guardians whose children presented with fever to the study site during study period. Patients that consented/assented to participate in the study underwent both clinical and laboratory examination for malaria. Those suffering from malaria were further screened to exclude cases of complicated malaria .Twelve patients aged between 1.5 to 8 years, having clinically-confirmed acute uncomplicated falciparum malaria, were recruited into the study and appropriate referrals were made for the rest. Study participants were followed-up for 28 days according to World Health Organization (WHO) protocol . Since tEthical approval for this study was given by the Makerere University Research and Ethics Committee and permission to conduct the study was given by Uganda National Council of Science and Technology.\u00ae)  plus AS (Arinate\u00ae)  under supervision. Both Amobin\u00ae and Arinate\u00ae are made according to Good Manufacturing Practice (GMP). The medication dosages were based on the age group as recommended by the Ministry of Health Uganda [At enrolment, a medical history was taken and a clinical examination was made. Two venous blood samples were obtained. One sample was used to obtain solid and consistent readings of the parasites and the other for day zero drug levels. The children were thereafter orally given AQ , about 200 to 300 \u03bcL of venous blood samples was collected before treatment and samples were collected daily thereafter, just before administration of AQ+AS. Aliquots of collected blood were mixed with equal amounts of phosphoric acid  from which 100 \u03bcL was spotted onto filter paper and allowed to dry at room temperature (23 \u2013 28\u00b0C). The filter paper samples were then stored in plastic envelopes till analysis. The analysis was carried out as described by Ntale et al .Thus, filter paper containing dried blood spots was cut into 4 to 6 small pieces and put into 12 mL polypropylene test tubes. Subsequently, 150 \u03bcL of I.S. solution (0.6 \u03bcM) and 1 mL of water were added and the contents were sonicated for 15 min. Sodium carbonate buffer , KOH  and 8 mL of di-isopropylether were added. Final pH of the water phase was between 9.17 \u2013 9.24. The samples were shaken for 20 min. and centrifuged for 10 min. at 3500 \u00d7 g. The organic (upper) phase was transferred to a new polypropylene test tube and back-extracted for 10 min. with 150 \u03bcL of sodium dihydrogen phosphate buffer . The phases were separated by centrifugation for 10 min. and the organic phase was removed by aspiration. The water phase (130 \u03bcL) was injected into the chromatograph.A total of twelve children participated in the study, with age ranging between 1.5 to 8 years. There was equal sex distribution but their weight varied between 7.7 Kg to 25.8 Kg. The AQ dosing was based on age, however, on a mg/Kg basis the dose ranged from 10.1 to 19.5 . Median AQ levels were highest on day 3, and DAQ levels followed a similar trend but were higher than those of AQ at all time points  in 12 whole blood samples collected from all the patients. The enrolled patients were followed-up for 28 days and during follow-up, none of the subjects dropped out. The failure to quantify AQ after seven days in all the subjects confirmed that the drug was rapidly metabolized to its active metabolite, DAQ as reported earlier .Although the AQ dose varied from 10.1 to 19.5, there were no adverse drug reactions observed. The observed variability in the reported concentrations of AQ and DAQ could be attributed to differences in body weight of the study participants and other factors such as nutritional status and inter-individual genetic variability. This finding is in agreement with existing literature, which also reveals considerable variability between patients and healthy subjects after intravenous infusion with AQ. Post-infusion plasma concentrations ranging between 231 to 2,352 nM for AQ have been reported in patients while values between 180 to 5,400 nM have been detected in healthy individuals ,26. In oDespite inter-individual variations in AQ and DAQ concentrations found in this study, the results demonstrate that both the sampling and analytical procedure of the recently developed filter paper method for determination of AQ and DAQ are applicable in the field. In addition, the analytical procedure is sufficiently sensitive for the analysis of low concentrations of both AQ and DAQ in whole blood dried on filter paper.in vivo [It has been suggested earlier that DAQ (and not AQ) must be monitored in vivo . TherefoAlthough the present study was not powered enough n = 12) to assess efficacy, it is encouraging that the 28-day follow up indicated 100% parasitological cure for the 12 patients treated with AQ+AS (table 2 to asseThis study has demonstrated that under field conditions it is possible to collect and analyse blood samples dried on filter paper thus making it possible to carry out large scale study of AQ pharmacokinetics in children with malaria infection, and to follow-up for any severe adverse reactions related to drug/metabolite concentrations in blood.The authors declare that they have no competing interests.MN was responsible for the conceptualization and design of the study, developing the research protocol, data collection and analysis and overall responsibility for the preparation of the manuscript. CO participated in the study design, protocol development, data collection, analysis and manuscript preparation. MM participated in experimental design, analysed all the samples and gave comments on the manuscript. JM, LLG, OB and JOO all contributed to the design of the study and to the preparation of the manuscript."}
+{"text": "Colin Sutherland discusses a Policy Forum article that explores the design and standardization of pre-licensure phase III antimalarial treatment trials. Subsequent investment in research and clinical trials, and new drug procurement arrangements through the Global Fund to Fight AIDS, Tuberculosis and Malaria, have achieved much to be proud of. Many African countries have now implemented new malaria treatment policies centred around artemisinin combination therapy (ACT) as the front-line treatment for malaria.It is almost ten years since a clarion call was sounded to malaria researchers, funding agencies, governments, and international organisations to help avert \u201ca malaria disaster\u201d . At thatsuperiority of the investigational product against a failing drug such as chloroquine or sulphadoxine-pyrimethamine, can no longer apply. There are now important questions about the design and standardisation of such pre-licensure phase III antimalarial treatment trials.There is now an urgent need to bring new combination therapies into the malaria drug development pipeline, to provide endemic country governments with alternative regimens suited to malaria transmission in their setting, and to minimise the global impact of resistance to ACTs, should it arise. However, in order to be granted licensure, such new combinations must be tested against current ACT-based regimens of high efficacy. Therefore the previously established phase III clinical trial designs, which test for PLoS Medicine:This Perspective discusses the following new article published in Plasmodium falciparum malaria. PLoS Med 5(11): e227. doi:10.1371/journal.pmed.0050227Borrmann S, Peto T, Snow RW, Gutteridge W, White NJ (2008) Revisiting the design of phase III clinical trials of antimalarial drugs for uncomplicated Steffen Borrmann and colleagues discuss appropriate endpoints and their measurement during phase III trials of new antimalarial drugs.and cure at least 90% of treated patients in a phase III study: violation of either criterion would mean the new therapy was not deemed of adequate efficacy.In a new Policy Forum on this topic, Steffen Borrmann and colleagues make important recommendations for such trials . In partHowever, we encounter dangerous waters when the question of efficacy endpoints arises\u2014what are the appropriate measures of antimalarial efficacy when testing for non-inferiority with an established regimen in phase III studies? A feature of falciparum malaria efficacy studies over the last decade has been the use of so-called \u201cPCR correction\u201d see  to distiAn alternative view is that uncorrected estimates of antimalarial efficacy, combining therapeutic and prophylactic effects in a single measure, should be given primacy. The main justification for this unfashionable opinion is 2-fold. Firstly, there is no evidence of a clinically meaningful difference between recrudescent and newly emergent infections . SecondlDelta margin: In the context of non-inferiority clinical trials, the delta margin sets the efficacy benchmark below which the test drug fails to be \u201cnon-inferior\u201d to the comparator drug. This benchmark is simply the measured efficacy of the comparator drug minus the delta margin; for a comparator with measured efficacy of, say, 96%, and a delta margin of 5%, the test drug must reach a measured efficacy of 91.2%.PCR correction: Use of genetic fingerprinting techniques, usually based on the polymorphic antigen loci pfmsp1 and pfmsp2, to determine whether P. falciparum parasites recurring in a patient's peripheral blood after antimalarial treatment are genetically identical to, or different from, the parasites present prior to treatment. If identical to pre-treatment parasites, the recurrent infection is considered to be a recrudescence; if different it is considered to be newly emergent from the liver and not a treatment failure, sensu stricto. There is disagreement in the literature as to the validity of this approach for correcting estimates of drug efficacy.Anopheles mosquitoes, provide early signals of developing P. falciparum resistance to antimalarial drugs, and could be more widely applied to comparative efficacy studies [There are other ways of monitoring the performance of antimalarial drugs not considered by Borrmann and colleagues. Transmission endpoints, such as gametocyte carriage or the infectivity of treated individuals to  studies ,6. Effic studies . This ap"}
+{"text": "Besides existing artemisinin-based combination therapies, alternative safe, effective and affordable drug combinations against falciparum malaria are needed. Methylene blue (MB) was the first synthetic antimalarial drug ever used, and recent studies have been promising with regard to its revival in malaria therapy. The objective of this study was to assess the safety and efficacy of two MB-based malaria combination therapies, MB\u2013artesunate (AS) and MB\u2013amodiaquine (AQ), compared to the local standard of care, AS-AQ, in Burkina Faso.Open-label randomised controlled phase II study in 180 children aged 6\u201310 years with uncomplicated falciparum malaria in Nouna, north-western Burkina Faso. Follow-up was for 28 days and analysis by intention-to-treat. The treatment groups were similar in baseline characteristics and there was only one loss to follow-up. No drug-related serious adverse events and no deaths occurred. MB-containing regimens were associated with mild vomiting and dysuria. No early treatment failures were observed. Parasite clearance time differed significantly among groups and was the shortest with MB-AS. By day 14, the rates of adequate clinical and parasitological response after PCR-based correction for recrudescence were 87% for MB-AS, 100% for MB-AQ (p\u200a=\u200a0.004), and 100% for AS-AQ (p\u200a=\u200a0.003). By day 28, the respective figure was lowest for MB-AS (62%), intermediate for the standard treatment AS-AQ , and highest for MB-AQ .MB-AQ is a promising alternative drug combination against malaria in Africa. Moreover, MB has the potential to further accelerate the rapid parasite clearance of artemisinin-based combination therapies. More than a century after the antimalarial properties of MB had been described, its role in malaria control deserves closer attention.NCT00354380ClinicalTrials.gov  Plasmodium falciparum to accessible and affordable first-line drugs such as chloroquine (CQ) and sulfadoxine/pyrimethamine (SP) in most countries of SSA Early diagnosis and prompt treatment with an effective antimalarial medicine remains the mainstay of malaria control in sub-Saharan Africa (SSA) in vivo by uncontrolled use of artemisinins or in combination with ineffective partner drugs Treatment with effective drugs as combinations has eventually become a paradigm in malaria control, with the particular aim to delay and possibly reverse the development of drug resistance P. falciparum glutathione reductase, it inhibits the heme polymerization within the parasite's food vacuole, and prevents methaemoglobinaemia in clinical malaria Methylene blue (MB), the first synthetic drug ever used against malaria in vitro study Amodiaquine (AQ) has remained remarkably effective in many SSA countries despite emerging CQ resistance Against this background, the safety and efficacy of MB combined with artesunate (AS) and of MB combined with AQ was studied in a controlled trial in Burkina Faso.The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Centre de Recherche en Sant\u00e9 de Nouna (CRSN) in Nouna Health District, north-western Burkina Faso. The area is highly endemic for malaria with most clinical cases occurring during or briefly after the rainy season which lasts from June until October The study was conducted in October/November 2006 in the urban research zone of the The study was designed as a randomized controlled phase II trial. The study was open label, with blinding only for the microscopists involved. The primary objective was to investigate the safety of the combinations MB-AS and MB-AQ in children with uncomplicated falciparum malaria. The secondary objective was to determine the efficacy of these MB-based combinations in the treatment of children with uncomplicated falciparum malaria. The primary end point of the study was the incidence of observed and self-reported adverse events over the 28 days observation period. Secondary end points were: treatment outcomes until day 14 and day 28 of follow-up, i.e. adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), fever clearance time, parasite clearance time, and hematocrit at D28 compared to baseline The community was informed on the project, and mothers with febrile children were invited to come to fever measurement points (specific private houses) in Nouna town . Children who were considered eligible for the study  were referred to the Nouna district hospital outpatient clinic for further examinations.P. falciparum asexual parasites per \u00b5L blood), and written informed consent given by the parents/caretakers. The age range criterion results from the absence - at the time of the study - of a liquid, taste-masked MB formulation, thus, patients needed to be able to take coated MB tablets . Exclusion criteria were signs of severe malaria, any apparent other disease, and malaria treatment \u2013 except CQ - with western drugs and/or antibiotics with antimalarial potency during the preceding week.Inclusion criteria were: age 6\u201310 years, ability to swallow tablets, uncomplicated falciparum malaria . A sealed envelope containing the identification numbers was opened after finalisation of the examination in the hospital and after informed consent was given.180 children were assigned to receive either MB-AS, MB-AQ, or AS-AQ, AS-AQ being the official first-line antimalarial in Burkina Faso. MB  was given at a dose of 10 mg per kilogram of body weight twice daily over three days. AS  was given at a dose of 4 mg per kilogram of body weight once daily over three days. AQ  was given at a dose of 10 mg per kilogram of body weight once daily over three days. The study drugs were administered under direct observation by study nurses  or field workers . In case of vomiting within the first 30 minutes the treatment was repeated. If vomiting occurred again, the patient was excluded and referred to the paediatrics department of the hospital. Children with fever \u226538.5\u00b0C received a standard dose of 10 mg per kilogram paracetamol tablets every 6 hours  until the symptoms subsided.P. falciparum parasites and were followed up on days 1, 2, 3, 7, 14, and 28. Mothers and caretakers were encouraged to come back at any time between scheduled visits in case of unforeseen symptoms.Follow-up of study children was for 28 days using a slightly modified version of the latest WHO protocol on antimalarial drug efficacy testing A finger-prick blood sample was taken on days 0, 2, 3, 7, 14, and 28, and during unscheduled visits. From this, malaria parasitaemia and haematocrit values were determined using standard CRSN procedures P. falciparum msp1 and msp2 genotype patterns in matched pairs of isolates obtained on admission and on the day of reappearance of parasitaemia From each blood sample, an aliquot was stored on filter paper. After shipment to the Institute of Tropical Medicine and International Health Berlin, DNA was extracted using commercial kits . Differentiation of recrudescences from new infections was achieved by comparing PCR-generated The study was designed to have a statistical power of 80% in order to detect a difference in the number of adverse events of at least 20% among study groups that was significant at the five percent level. Losses to follow-up and dropouts due to other reasons were considered treatment failures in an intention-to-treat manner.The Chi square test (Chi) was used to compare proportions, and the non-parametric Wilcoxon-Mann-Whitney test (WMW) to compare metric or ordinal data. When possible, estimates and the corresponding 95% confidence interval are given.The closed testing procedure Calculations were performed with SAS release 9.1 .The study protocol was approved by the Ethics Committee of the Medical Faculty at Heidelberg University and the National Ethics Committee of Burkina Faso. Caretakers were asked for their written consent after having received detailed information from a study nurse about all known risks and benefits of the study through translation of a detailed research consent form into the local language,a priori defined full analysis set (FAS) for the intention-to-treat analysis. There was one loss to follow-up due to out-migration from the study area . At enrolment, the demographic and clinical characteristics of the participants in the three study groups were similar, except a small but significant difference in weight  were included into the study . These cn weight .There was no case of death but one serious adverse event over the 28 days follow-up period. It occurred in the AS-AQ group as a brief hospitalisation late at night on day 0 due to severe vomiting. This child was not considered a treatment failure and thus not excluded from the further analysis.Adverse events are shown in Efficacy results are given in Similarily, by day 28, the rate of ACPR without correction was significantly lower in the MB-AS group (28%) compared to the groups MB-AQ (79%) and AS-AQ (66%). During the 28 days of follow-up, exclusively new genotypes, i.e. re-infections, were more frequent with MB-AS  than with MB-AQ  or AQ-AS . After PCR-based correction for recrudescence, the ACPR rates increased to 62% (MB-AS), 95% (MB-AQ), and 82% (AS-AQ), differences being significant among all groups .Fever was cleared rapidly in all three groups; only two patients remained febrile until day 2 in the MB-AS group, and two until day 1 in the MB-AQ group. However, there were significant differences in parasite clearance among study groups : While iThere were no significant changes in median haematocrit values over time and among groups (data not shown).Treatment with the MB-based combinations was associated with significantly more adverse events compared to AS-AQ, and this was caused by the frequent occurrence of vomiting and dysuria, symptoms already known to be associated with MB tablets Urolene Blue\u00ae is a taste-masked MB tablet used for other indications in patients who are able to swallow tablets. There is currently no paediatric taste-masked formulation on the market and the provision of non-masked liquid MB solutions or crushed tablets dissolved in water is not recommendable as the drug has a strong bitter-metallic taste Two main results with regard to the efficacy of the MB-based combinations are provided by the present trial. First, the efficacy of MB-AQ in the study area is high, and significantly higher than it is the case for the standard ACT regimen. With 95% ACPR after 28 days of follow-up, the efficacy of MB-AQ is in the range of various ACT regimens in SSA At 82% ACPR, the standard regime AS-AQ surprisingly did worse than expected and than it has been reported previously from other places in West Africa P. falciparum parasites than the other two treatment regimes. This provides evidence in humans for a clinically relevant synergy between an artemisinin derivative and MB, which has already been shown in vitroSecondly, although performing rather poor in terms of overall efficacy, MB-AS achieved a more rapid clearance of Likely related to the short elimination half-life of both MB and AS As MB is both available and affordable, the combination MB-AQ would be an interesting alternative antimalarial regimen when shown to be safe and effective in larger multi-centre phase III studies. Such studies are now planned for in the frame of a public-private-partnership by our group. Apart from the potential benefit of another effective antimalarial regimen contributing to a healthy competition of different regimens on the market, MB can be considered a re-emerging antimalarial with the potential to be a valuable partner drug in various ACT and non-artemisinin drug combinations.In conclusion, MB-AQ is a promising alternative drug combination against falciparum malaria in SSA. Moreover, MB has the potential to further accelerate parasite clearance when used in ACT. Further studies are needed to define the possible roles of MB in malaria treatment.Checklist S1CONSORT Checklist(0.06 MB DOC)Click here for additional data file.Protocol S1Trial Protocol(0.20 MB DOC)Click here for additional data file."}
+{"text": "N = 85) with acute Plasmodium vivax malaria and normal glucose-6-phosphate dehydrogenase screening were randomized to receive 30 mg or 60 mg primaquine daily for 7 days . The regimens were well tolerated and all patients recovered fully. Median fever clearance , mean \u00b1 SD parasite clearance times (87.7 \u00b1 25.3 hours), gametocyte clearance, and adverse effects were similar in the 2 groups. Two patients, 1 from each group, had a 30% reduction in hematocrit. The cumulative 28 day relapse rate  by Kaplan Meier survival analysis was 29% (16\u201349%) in the 30 mg group compared with 7% (2\u201324%) in the 60 mg group; P = 0.027. Comparison with previous data obtained at this same site suggests that the recurrences comprised approximately 17% recrudescences and 12% relapses in the 30 mg/day group compared with 3% recrudescences and 4% relapses in the 60 mg/day group. These data suggest that the dose-response relationships for primaquine's asexual stage and hypnozoitocidal activities in-vivo are different. A 1 week course of primaquine 60 mg daily is an effective treatment of vivax malaria in this region.Thai adult males ( Plasmodium vivax and P. ovale malaria. Primaquine at the dose most usually recommended in the past for radical cure (15 mg base adult dose daily for 14 days) is also weakly effective against asexual stages of Plasmodium vivax malaria.Plasmodium vivax has a 3 week interval between primary infection and the first relapse, and approximately half the patients experience at least 1 relapse.8- The schizontocidal activity of primaquineP. vivax, although the 2 week course currently recommended for radical cure remains a significant impediment to good adherence and therefore optimal efficacy. The dose-response relationships for primaquine's asexual stage and hypnozoitocidal activities have not been well characterized. To compare these 2 discrete activities and to assess efficacy and tolerability, we evaluated 7-day courses of 30 mg and 60 mg daily primaquine-only regimens for the treatment and radical cure of Plasmodium vivax malaria in Thailand.Primaquine is the only widely available hypnozoitocidal drug for Plasmodium vivax malaria admitted to the Bangkok Hospital for Tropical Diseases, Thailand between 1996 and 1998. The study was approved by the ethics committee of the Faculty of Tropical Medicine, Mahidol University, Bangkok. All patients gave full informed consent.The study was conducted in adult male patients with acute symptomatic All patients were screened and only those with normal G6PD testing were included. Other exclusion criteria were a history of drug hypersensitivity, having taken any antimalarial drugs within the previous 48 hours, or urine screening tests that were positive for sulfonamides (lignin test) or 4-aminoquinolines (Wilson Edeson test). Patients who had patent mixed infections or were unable to stay in the hospital until clearance of both fever and parasitemia were excluded from the study.1.Primaquine  0.50 mg base/kg once daily (adult dose 30 mg base/day) for 7 days2.Primaquine 1.0 mg base/kg once daily (adult dose 60 mg base/day) for 7 days.After clinical assessment and confirmation of the diagnosis from thick and thin blood smears, baseline blood samples were taken for routine hematology and biochemistry. Treatments were allocated according to double-blinded randomization codes. The treatment allocation was kept in serially numbered sealed envelopes that were opened only after the patient had been enrolled in the study. Patients were randomized to receive 1 of the following 2 regimens:Oral acetaminophen (0.5\u20131 g 4 hourly) was given for fever > 38\u00b0C.P. falciparum infection were treated with a 7-day course of quinine  combined with tetracycline (250 mg every 6 hours) and censored from that time in the subsequent assessment.Vital signs were recorded every 4 hours until resolution of fever and thereafter every 6\u201312 hours. Fever clearance time was defined as the time for body temperature to fall below 37.5\u00b0C and remain below this value for > 48 hours. Early treatment failure was defined as persistence of fever and parasitemia for > 7 days or persistence of parasitemia in the absence of fever for > 14 days. Reappearance of infection was assessed in patients who remained in Bangkok either in the hospital or at home  for at least 28 days. Patients who failed to respond to the treatment, or those who had recurrent vivax infections were treated subsequently with the standard regimen of chloroquine (25 mg/kg base over 3 days) and primaquine (0.25 mg base/kg/day for 14 days). Patients with delayed appearance of Assessment of gastrointestinal symptoms  and any other symptoms and signs was performed every day during and after primaquine treatment . Hematocrit was assessed daily during and after primaquine treatment. Laboratory investigations for full blood count and routine biochemical tests were performed before treatment (D0) and then on day 7, day 14, and day 28.P. vivax cannot be distinguished reliably.8Parasite counts of both sexual and asexual stages were measured every 2\u201312 hours in thin films and thick films until the parasitemia became detectable only in thick films, then every 12 hours until clearance, and thereafter daily for 28 days. Parasitemia was expressed as the number of parasites per microliter of blood . Gametocytes and asexual stages were counted separately. Gametocyte counts were calculated from the thick film only. Gametocyte carriage was expressed as the proportion of patients with patent gametocytemia either before treatment  or at any time after the start of treatment. The gametocyte clearance time was defined as the interval from the first detection of gametocytes in peripheral blood films either before or after treatment to the last detection of gametocytes. The variables describing parasite clearance were time taken until the asexual malaria parasite count fell by 50% (PC50), time taken to fall by 90% (PC90) of the pre-treatment admission value, and the time taken to fall below detectable levels in a peripheral blood smear (PCT); parasite reduction ratio (PRR) was the ratio of the parasite count before treatment to the counts at 24 hours and at 48 hours. Recurrence of vivax malaria could have resulted from recrudescence or relapse but not re-infection because these patients remained out of the transmission area throughout the follow-up period. In an individual patient whose parasitemia cleared, recrudescence or relapse of P. falciparum and were compared using the log rank test. The Spearman rank correlation coefficient was used to evaluate the association between gametocyte counts, parasite counts, fever clearance, and parasite clearance times. All statistical analyses were performed using the statistical computing package SPSS Version 16.0 for Windows .The data from each treatment group were compared by one-way analysis of variance. Non-parametric data were compared by the Kruskal-Wallis test. The cumulative cure rates were calculated by Kaplan-Meier survival analysis censoring for loss to follow-up or appearance of N = 74, 87.1%) and a high proportion  had had at least one malaria infection in the past year . All patients presented with a history of fever  before hospital admission. The geometric mean (range) peripheral blood P. vivax asexual parasite count was 6,205/\u00b5L . Overall 29% (N = 25) of patients had detectable gametocytemia at presentation. The median (range) gametocyte count was 41/\u00b5L (16\u2013110/\u00b5L). The majority of patients (88%) were thrombocytopenic  but none had significant abnormalities of other baseline hematology results (N = 43 and 42), serum glutamic-pyruvic transferase (sGPT) concentrations were significantly higher in the high dose group  but all baseline characteristics and other laboratory findings were similar in the 2 groups (P \u2265 0.24).There were 85 patients with acute vivax malaria 14\u201361 years of age (mean \u00b1 SD = 24.6 \u00b1 10.1 years). The majority of patients acquired malaria infection from the western border of Thailand ( results . BetweenP \u2265 0.18). Excluding the patient with acute tonsillitis, the median fever clearance time was 47 hours (range 4 to 130 hours). On admission, 55 patients (64.7%) had gastrointestinal symptoms i.e. nausea , vomiting , abdominal pain , and diarrhea . These symptoms disappeared after day 3 in the majority of patients  as the malaria infection subsided (P = 0.033). There was no other significant adverse effects or complications in the studied patients.All patients made a full clinical recovery after treatment. One patient developed acute tonsillitis on day 5 but responded well to antibiotics (amoxicillin). His fever cleared 201 hours after starting primaquine treatment. The overall mean (SD) parasite clearance time was 87.7 hours (25.3 hours). The overall median (range) parasite reduction ratios at 24 hours and 48 hours were 2 (0.1\u20134.1) and 14.4 (0.1\u20131376), respectively. Between the 2 treatment groups, there were no significant differences in PCT or parasite reduction ratios (subsided . Only onP < 0.001), then returned back to baseline values on day 14 , and became significantly higher than the baseline hematocrit on day 28 . The overall median hematocrit reduction was 8.8% (range 2.2\u201333.3%). A hematocrit drop of > 30% was observed in 2 cases; one from each group. The lowest recorded value was 30% on day 6 in one case and 28% on day 7 for the other. Neither was associated with symptoms and none of the studied patients needed a blood transfusion. Comparing the 2 treatment groups, there were no significant differences in the baseline hematocrit values , the proportion of patients with hematocrit reduction  or the magnitude of hematocrit reductions (P \u2265 0.11).The mean \u00b1 SD baseline hematocrit of all patients (36.3% \u00b1 5.6%) decreased significantly after day 2 to day 7 (N = 72/82). The mean \u00b1 SD baseline platelet count was 111,000 \u00b1 77,000/\u03bcL but then increased to above 200,000/\u03bcL in all subsequent follow-up assessments  in both groups  was found in the majority of patients  of patients, 29.4% (N = 25) on admission and a further 32.9% (N = 28) after starting primaquine treatment. The gametocyte carriage of all studied patients  was brief; median was 12 hours  and was not significantly different between the two treatment groups . The overall gametocyte carriage rates were not significantly different between the 2 groups either before (N = 15 and 10) or after treatment (N = 14 and 14) (P = 0.58).During hospital stay, patent gametocytemia was detected in 62.4% (P = 0.016).Gametocytemia before treatment and the ratio to asexual stage parasite densities were similar in the two treatment groups . In patiP = 0.62).Complete clinical and parasitological follow-up was obtained in 64 patients (75%). These patients were either followed-up outside the malaria transmission area for 28 days or remained in the hospital until the subsequent appearance of vivax or falciparum malaria. The remaining 25% of patients did not return after discharge. Their demographic and disease characteristics were similar to those successfully followed. There were no significant differences in proportion of loss to follow-up between the two treatment groups  of 29% (16\u201349%) in the 30 mg group compared with 7% (2\u201324%) in the 60 mg group; P = 0.027  had reappearance of vivax and another one (3%) had emergence of falciparum malaria after a full clinical recovery and parasite clearance of the primary vivax illness. The corresponding figures were 2/33 (6%) relapses and 3 (7%)  = 0.027 .www.ajtmh.org). The 60 mg/day recurrence rates in this study are similar to those in the previous studies, which suggests first that this dose gives nearly maximal hypnozoitocidal activity, and second that the significantly higher recurrence rate with the 30 mg/day primaquine monotherapy results predominantly from recrudescences  and with similar patient groups with acute vivax malaria, in which primaquine (30 mg base or 60 mg base per day) was given together with artesunate 5 or 7 days) for 3, 5, or 7 days. days forP \u2265 0.11). Regardless of treatment groups, patients with gametocytemia cleared asexual parasite much slower that those without gametocytemia (P \u2264 0.018) and also had slower fever clearance times although this was not statistically significant (P = 0.075). This was not attributable to higher parasitemia as there were no significant differences in parasitemia between those with and without gametocytemia. There were also no significant differences in the numbers of previous malaria attacks, duration of fever, or admission hematocrit values or reappearance rate of vivax malaria between patients with and without gametocytemia , it is seldom actually prescribed in endemic areas in the 14 day courses currently considered necessary for optimum effects.P. vivax is traditionally considered more difficult to eliminate than P. falciparum, current global initiatives against vivax malaria may be unsuccessful if primaquine remains only a theoretical recommendation. Chloroquine resistance in P. vivax is now established in several areas, notably Oceania and in Indonesia. As primaquine has significant asexual stage activity against P. vivax, the combination of chloroquine and primaquine would be expected to delay or prevent the emergence of resistance. India, which harbors the majority of the world's P. vivax infections, recommended an inadequate primaquine dose regimen (5 days only) for radical cure for nearly half a century, but perhaps the reason that chloroquine resistance did not emerge there was because of combination treatment against the asexual stage infection. If an effective radical cure could be given in a shorter duration than 2 weeks, then adherence would be expected to improve.Primaquine may be the most under-used antimalarial drug. Although widely recommended for the radical cure of relapsing malarias . Assessment of radical curative regimens with chloroquine therefore require long follow-up, and may underestimate true relapse rates. It is not known whether the suppressed relapse is prevented or delayed. The 8-aminoquinolines are the only class of drug with radical curative activity, although there is intriguing evidence that other quinoline antimalarials may synergize with them in this activity.Chloroquine is still the most widely used vivax malaria treatment in the world. Given alone for tropical frequent relapse From a practical therapeutic perspective the higher dose shorter course regimen, if generally well tolerated, may be a preferable component of combination treatment regimens for vivax malaria. Further studies to assess tolerability in a broader age range are now indicated."}
+{"text": "Steffen Borrmann and colleagues discuss appropriate endpoints and their measurement during phase III trials of new antimalarial drugs. Plasmodium falciparum caused at least 0.5 billion uncomplicated clinical attacks of malaria worldwide, particularly in non-immune young children living in Africa . Fr. Fr33]. In high-transmission areas, the composite effect size will largely be determined by the post-treatment prophylactic efficacy against secondary infections . The durRecrudescent primary infections, as well as re-infections, entail the risk of secondary malaria and/or hematological complications requiring re-treatment ,35. ClinThe risk that a patient with recurrent infection will be symptomatic at detection or succumb subsequently to secondary malaria during follow-up is related to the individual level of specific, if imperfect, acquired immunity (\u201csemi-immunity\u201d). For example, an analysis of antimalarial treatment trials in high-transmission areas found that children below one year were at least three times as likely to experience symptomatic recurrences than children aged more than three years . This dedelay instead of proportions of recurrent infections, should be undertaken.Based on the above considerations, whilst it is clear that, in areas of very high transmission, multiplicity of infection confers a significant and irreducible error in genotyping , parasitOptimal target profiles for new antimalarial drugs have been described elsewhere and used as guiding principles from early discovery through clinical development ,40. In adelayed ? Should the new drug be superior to or \u201cjust as good as\u201d the reference treatment? Or to use a hypothetical example, in times of continued shortage of viable therapeutic options, is it desirable to turn down a new once-daily regimen with a composite efficacy comparable to the recently introduced twice-daily regimen of artemetherlumefantrine, but which may be inferior to dihydroartemisinin-piperaquine because piperaquine is exceptionally slowly eliminated [To date, there is no guidance on the benchmark for the protective efficacy of a new antimalarial drug in preventing secondary infections, or the composite efficacy of curative and preventive effects. The establishment of a uniform benchmark is challenging: in addition to clearing more than 90% of primary infections, should a new drug prevent 30%, 40%, or more of secondary infections? Over which time frame\u2014four, six, or more weeks? Or, put more simply, for how long on average should re-infections or, more generally, recurrent infection be iminated ,42,43? AThe current use of day 28 estimates provides a single benchmark period to compare all new drugs, but we feel there is a need to revisit this single time-point for phase III trial endpoints, particularly for new investigational drugs or combinations containing at least one component with an intrinsically long plasma half-life.In phase III, the new candidate drug is compared in a randomised controlled trial  to a staSmall differences in parasitological cure rates demonstrated in superiority trials, especially between 95% and 100%, are important in reducing the emergence and spread of resistance , but frosuperiority of the post-treatment prophylactic efficacy of a new drug with comparatively higher anti-liver stage activity or prolonged plasma half-life can be best examined in areas with high re-infection rates because of the postulated differential impact on re-infection and/or secondary malaria rates [The ia rates . The comPLoS Medicine Perspective by Colin J. Sutherland:This Policy Forum is further discussed in a 10.1371/journal.pmed.0050228Sutherland CJ (2008) Comparing highly efficacious antimalarial drugs. PLoS Med 5(11): e228. doi:non-inferiority when in fact the new drug is inferior (type I error) [inferiority when in fact the new drug is not inferior (type II error). If the primary endpoint is measured as proportional point estimate , we recommend basing the analysis on the per-protocol population, which includes only patients with observed treatment responses and other informative outcomes, e.g., intake of outside-protocol antimalarial medication. The preferred method for comparing antimalarial drug efficacy, however, is survival analysis [The transition from superiority to non-inferiority trial designs has implications for the choice of the primary analysis population . Non-infI error) ,49. On tSurvival analysis techniques are increasingly used to assess failure rates in randomised phase III trials . The keyThe interpretation of a non-inferiority trial with a survival estimate of recrudescent infections can be based on two criteria: (1) the hypothesis test result, e.g., the 95% CI of the proportional hazard ratio test/control remains below the non-inferiority limit on the hazard ratio to support claims of efficacy ; and (2)P. falciparum malaria are designed and interpreted are only now being developed [Consensus-agreed regulatory guidelines on how phase III trials of antimalarial drugs for uncomplicated eveloped . This re"}
+{"text": "Benin has recently shifted its national antimalarial drug policy from monotherapies to combinations containing artemisinin derivatives. When this decision was taken, the available information on alternatives to chloroquine and sulphadoxine-pyrimethamine, the first- and second-line treatment, was sparse.In 2003 \u2013 2005, before the drug policy change, a randomized, open-label, clinical trial was carried out on the efficacy of chloroquine, and sulphadoxine-pyrimethamine alone or combined with artesunate, with the aim of providing policy makers with the information needed to formulate a new antimalarial drug policy. Children between six and 59 months of age, with uncomplicated malaria and living in the lagoon costal area in southern Benin, were randomly allocated to one of the three study arms and followed up for 28 days.Treatment failure (PCR corrected) was significantly lower in the artesunate + sulphadoxine-pyrimethamine group  than in chloroquine group or the sulphadoxine-pyrimethamine alone group  (p < 0.001). Despite high sulphadoxine-pyrimethamine failure, its combination with artesunate greatly improved treatment efficacy.In Benin, artesunate + sulphadoxine-pyrimethamine is efficacious and could be used when the recommended artemisinin-based combinations (artemether-lumefantrine and amodiaquine-artesunate) are not available. However, because sulphadoxine-pyrimethamine is also used in pregnant women as intermittent preventive treatment, its combination with artesunate should not be widely employed in malaria patients as this may compromise the efficacy of intermittent preventive treatment. In Benin, early malaria case management, both at health facility and community level, is the backbone of the national malaria control strategy . High chIn Benin, CQ resistance was first reported in 1986 -6 and, iSulphadoxine-pyrimethamine (SP) efficacy was assessed in 2002 in five different sites with varying intensity of malaria transmission . It had Anopheles gambiae and Anopheles melas are the main malaria vectors [This is a randomized trial conducted at three adjacent peri-urban sites: Ladji, Awansori and Toweta, situated in the coastal lagoon area of Cotonou.  vectors , and the vectors ,11. Prev vectors .The local authorities and the population were informed about the study and its objectives in 2001. A census was carried out in 2002\u20132003. Houses were numbered and information on the household's residents  collected. A list of six to 59 months old children without any obvious chronic illness, with parents willing to participate to the study, and unlikely to emigrate outside the study area was produced. Five hundred and fifty three children  were randomly chosen from this list to be part of the cohort under surveillance. Parents or legal guardians were asked before inclusion to sign an informed consent form. Refusal to participate did not affect access to basic health services. Parents/guardians were instructed to attend the nearby health facility, where a physician was always available, whenever their child was sick. They were also asked to administer to the child only drugs given or prescribed by the research physician.Plasmodium falciparum mono-infection with a density between 1,000\u2013200,000/\u03bcL. Children were allocated to one of the study treatment groups, i.e. CQ, SP or SP-AS if they met no exclusion criteria: PCV < 25%, severe malaria [Starting from July 2003 up to January 2005, each child was visited twice a week by the research team. Body temperature was measured routinely during home visits and, if the child was febrile (body temperature \u2265 37.5\u00b0C), a blood sample for parasitaemia (thick and thin blood film) and for genotyping at a later stage (filter paper Whatman grade 3) was collected. A similar approach was applied to children attending health facilities within the study area. An additional blood sample for PCV was collected in children with a  malaria , danger msp1and msp2) as described previously [Thick blood films were stained with Giemsa (10% for 10 minutes). Parasite density was determined on a thick blood film according to the number of parasites per 200 white blood cells (WBC), and assuming a total WBC count of 8,000/\u03bcl. If gametocytes were seen, the gametocyte count was extended at 1,000 WBC. Microscopy examination was totally blinded to the patient's identity and treatment group. PCV, measured by micro-haematocrit centrifugation, was assessed on days 0, 14 and 28. Genotyping, to distinguish between a new infection and a recrudescence, was done on blood samples collected before treatment and at the time of any recurrent parasitaemia and was performed at the Institute of Tropical Medicine, Antwerp, Belgium. DNA was extracted as described elsewhere. Parasite genotyping was done by nested PCR for variable blocks within the merozoite surface protein 1 and 2 . Descriptive statistics were used to summarize baseline values and demography data.P. falciparum infection when estimating outcomes corrected by PCR. In the survival analysis, cure rates were described by Kaplan Meier estimates and compared between groups with a log-rank test. Pair-wise comparisons of treatment efficacy at day 28 were made with a Cox proportional hazards model. Gametocyte carriage rates were estimated as proportion of patients with gametocytaemia and person-gametocyte-weeks (PGW) were calculated to measure gametocyte carriage and transmission potential. The PGW were calculated as the number of weeks in which blood slides were positive for gametocytes during the two first weeks of follow-up after treatment divided by the number of all follow-up weeks and expressed per 1,000 person-weeks. P < .05 was considered statistically significant.Analysis of variance (ANOVA) was used for normally distributed continuous data. Data not normally distributed were compared by Wilcoxon Rank sum or Kruskal-Wallis tests. Categorical data were compared by calculating the chi-square value or by Fisher's exact test. Data were analysed by Intention-to-Treat (ITT), Per-Protocol (PP) and Kaplan Meier survival analysis to determine the cure rate (PCR corrected) at day 28 in each treatment group. In the ITT analysis, patients lost to follow up were considered as treatment failures (worst case scenario) and the denominator was the number of patients initially randomized to each group. In the PP analysis, patients lost to follow-up and those excluded after enrolment were excluded from the analysis; only patients with a complete follow-up were included in the denominator. Cure rates, fever and parasite clearance were compared using logistic regression models. Odds ratio and 95% CI were estimated from these models. In the Kaplan Meier survival analysis, each patient contributed to the analysis for the time s/he was followed up. Data were censored for patients who did not complete follow up and for new The study was approved by the Minister of Health of Benin, the Ethical Committee of the Facult\u00e9 des Sciences de la Sant\u00e9, Cotonou, Benin and the Ethical Committee of the Institute of Tropical Medicine, Antwerp. Informed consent was obtained from parents or guardians.P. falciparum infection, of whom 237 fulfilled all inclusion criteria. These children were randomly assigned to one of the study groups: 79 to CQ; 77 to SP and 81 to the combination SP-AS , SP alone: 4/67 (6.0%), CQ: 6/62 (9.7%) (p = 0.02). Parasite clearance was also faster in the SP-AS than in the other two treatment groups. By day 3, the percentage of children without infection was 94.9% (75/79) in the SP-AS, 56.7% (38/67) in the SP and 56.5% (35/62) in the CQ group (p < 0.001).In the SP-AS group, no ETF was observed while there were 16 in the SP and 18 in the CQ group Table . By day By day 28, seven additional recurrent parasitaemia occurred in the SP-AS group (one LCF and six LPF), seven in the SP group (two LCF and five LPF), and 14 in the CQ group (two LCF and 12 LPF). Thus, the failure rate (PCR uncorrected) was 10.5% (8/76) for the SP-AS, 45.6% (31/68) for the SP alone and 74.6% (53/71) for the CQ group. After PCR correction, these rates were 5.3% (4/76), 44.1% (30/68) and 71.8% (51/71), respectively  were similar  to the PP analysis. Differences between study arms were highly significant  and the hazard of failure estimated by Cox regression was 20 times higher with CQ compared to SP-AS  (p < 0.001). For SP alone versus SP-AS, the assumption of proportional hazards was respected only after day 3: the hazard for treatment failure was almost six times higher with SP compared to SP-AS  (p = 0.002) . In total, 28 serious adverse events were identified  (p = 0.15). No death was reported. No severe malaria cases were observed in the SP-AS group while there were two cases of convulsions in the other two groups (one CQ and one SP) and five cases of severe anaemia (three CQ and two SP) Table . Seven pThe combination SP-AS was safe, well-tolerated and significantly more efficacious than the two other treatments. Children treated with SP-AS had also a faster fever and parasite clearance. At day 28, the PCR-corrected TTF for the SP-AS group was only 5.3%, an extremely low figure compared to the other two treatments. These findings are comparable to the 28-day PCR-corrected cure rates recently reported from Ghana, (94.5%)  and eastAs expected, gametocyte carriage in children treated with SP-AS was much lower than in the other treatment groups. A decrease in malaria transmission after the introduction of ACT has already been reported in Thailand and it has been suggested that this was linked to the lower gametocyte production in patients treated with ACT . HoweverBenin recently chose artemether-lumefantrine and amodiaquine-artesunate as recommended treatments for uncomplicated malaria. This decision was taken in 2004 but it is unfortunate to notice that these ACT are not available yet at peripheral health facilities. Other African countries such as Burkina Faso are in a similar situation, i.e. the national drug policy has been changed but not implemented yet . In Benin, considering that SP and AS are easily available and their combination has an acceptable efficacy, SP-AS may be used whenever the recommended standard treatments are hopefully temporarily unavailable. However, because SP is also used in pregnant women as intermittent preventive treatment (IPTp), its combination with AS should not be widely employed in malaria patients as this may compromise the efficacy of IPTp.The author(s) declare that they have no competing interests.AN: contributed to the study design, study coordination and supervision, field work, data entry, cleaning and analysis, and paper writing.AE: contributed to the study design, data analysis and review of the paper.DG: contributed to the study design and field work.CVO: contributed to the parasite genotyping.CA: contributed to the planning and implementation of the field work.HvL: contributed to the data management.JM: contributed to the statistical analysis.MA: contributed to the implementation of the study.MC: contributed to the study design and paper reviewing.AM: contributed to the study design, data analysis and interpretation and manuscript review.UDA: contributed to study design, protocol writing, data analysis, interpretation and supervision, and manuscript review."}
+{"text": "Results from trials of intermittent preventive treatment (IPT) in infants and children have shown that IPT provides significant protection against clinical malaria. Sulfadoxine-pyrimethamine (SP) given alone or in combination with other drugs has been used for most IPT programmes. However, SP resistance is increasing in many parts of Africa. Thus, we have investigated whether SP plus AQ, SP plus piperaquine (PQ) and dihydroartemisinin (DHA) plus PQ might be equally safe and effective when used for IPT in children in an area of seasonal transmission.During the 2007 malaria transmission season, 1008 Gambian children were individually randomized to receive SP plus amodiaquine (AQ), SP plus piperaquine (PQ) or dihydroartemisinin (DHA) plus PQ at monthly intervals on three occasions during the peak malaria transmission season. To determine the risk of side effects following drug administration, participants in each treatment group were visited at home three days after the start of each round of drug administration and a side effects questionnaire completed. To help establish whether adverse events were drug related, the same questionnaire was administered to 286 age matched control children recruited from adjacent villages. Morbidity was monitored throughout the malaria transmission season and study children were seen at the end of the malaria transmission season.All three treatment regimens showed good safety profiles. No severe adverse event related to IPT was reported. The most frequent adverse events reported were coughing, diarrhoea, vomiting, abdominal pain and loss of appetite. Cough was present in 15.2%, 15.4% and 18.7% of study subjects who received SP plus AQ, DHA plus PQ or SP plus PQ respectively, compared to 19.2% in a control group. The incidence of malaria in the DHA plus PQ, SP plus AQ and SP plus PQ groups were 0.10 cases per child year , 0.06  and 0.06  respectively. The incidence of malaria in the control group was 0.79 cases per child year .All the three regimens of IPT in children were safe and highly efficaciousNCT00561899ClinicalTrials.gov  Although significant advances have been made in controlling malaria during the last two decades, the disease remains a major cause of mortality and morbidity in sub-Saharan Africa. The most important recent steps in improving malaria control include the introduction of artemisinin-based combination therapy (ACT), intermittent preventive treatment (IPT) in pregnancy and increased coverage with ITNs. However, malaria is still estimated to cause directly nearly one million deaths a year and the vast majority of deaths occur among young children, especially among those living in remote rural areas of Africa The role of IPT in the prevention of malaria and anaemia in children has been evaluated in a limited number of trials. In Mali, an area of seasonal malaria transmission, two doses SP given to children aged 6 months to 9 years at an interval of two months gave a protective efficacy of 40% against clinical attacks of malaria We have, therefore, undertaken a clinical trial in which the safety, tolerability and efficacy of SP plus AQ, SP plus piperaquine (PQ) and dihydroartemisinin (DHA) plus (PQ) have been compared when used for intermittent preventive treatment in children in an area of seasonal transmission.The protocol for this trial and supporting CONSORT checklist are available as supporting information; see The study was based at the MRC Field Station, Basse in the Upper River Region of The Gambia and was conducted between May 2007 and December 2008. Malaria in The Gambia is highly seasonal, occurring almost exclusively during the rainy season (July to November) with greatest incidence in October to November The study was designed as an open-label, randomized trial. One thousand and eight children were individually randomized to receive SP plus AQ, SP plus PQ or DHA plus PQ (Duo-Cotecxin\u00ae) at monthly intervals on three occasions during the peak malaria transmission season . IPTc has been shown to reduce morbidity from malaria in children substantially. Thus, it was considered ethically unacceptable to include a randomized placebo group in the study. For this reason, an alternative design was chosen that relied upon recruitment of children aged 6\u201359 months from adjacent villages (less than 10 km from a study village). Using the data provided through an on-going Demographic Surveillance System (DSS) 286 children matched for age to children in the trial were randomly selected from 9 adjacent villages to form a control group.Meetings were held in the villages in the study area to provide information about the trial to parents of potential study participants and to seek community consent. Fourteen villages agreed to participate in the study; the average distance to the nearest health centre of these villages was approximately 5 km. A list of children living in the study villages and who would be aged between 6 and 59 months at the time of the first treatment in September 2007 was obtained from the Basse DSS. These children and their parents were invited for screening for potential participation in the trial between July and August 2007. Written informed consent was obtained from parents or guardians of all study subjects before screening. Children had their age and identities checked, and were examined to rule out any clinically significant disease. Finger-prick blood was collected for preparation of a thick blood smear and measurement of haemoglobin concentration. Children were considered eligible if they had no clinically significant chronic or acute disease. Exclusion criteria included known allergy to any antimalarial drug or presence of acute or chronic, clinically significant pulmonary, cardiovascular, hepatic or renal disease. Blood films were examined shortly after collection from any child with symptoms suggestive of malaria whilst remaining blood films were examined at a later date. Children with acute malaria were treated with Coartem\u2122 and excluded from the study.One thousand and eight eligible children whose parents consented were individually randomized into the SP plus AQ, SP plus PQ or DHA plus PQ treatment groups in a 1\u22361\u22361 fashion. A photo ID card was given to all participants to facilitate identification at every subsequent contact, at home or in the health centres. The randomization list was generated using permuted blocks of 12, using the random number generator of STATA version 10. Treatment assignments were placed in sealed opaque envelopes labelled with the randomization number, each child enrolled was assigned the next study number in sequence.Monthly treatment was given at the nearest health centre by a team of two nurses who played no part in the evaluation of safety or efficacy measurements and who did not communicate any information on group allocation to the team in charge of the evaluation of adverse events and morbidity. Children received half a tablet of SP [tablets containing 25 mg pyrimethamine and 500 mg sulphadoxine]  if they weighted 10 kg or less and a whole tablet if they weighed more than 10 kg. Amodiaquine [200 mg base tablets]  was given as follows; \u00bd, \u00bd and \u00bc tablets on days 1,2 and 3 for children who weighed 10 kg or less and 1,1 and \u00bd tablets on days 1,2 and 3 for children who weighed more than 10 kg. Piperaquine [tablets containing 250 mg]  was given at a dose of half a tablet for children who weighed 11 kg or less and as a whole tablet if a child's weight was more than 12 kg. Duo-Cotecxin\u00ae [tablets containing 40 mg of DHA and 250 mg PQ]  was given at a dose of half a tablet for children who weighed 13 kg or less and a whole tablet for children who weighed more than 13 kg.To ensure compliance with European regulations regarding drug quality, a sample of PQ tablets was tested for drug content and impurities by Sigma Tau Pharmaceutical company; drug concentration was within 5% of the nominal level and analysis of impurities indicated that although some impurities exceeded the ICH limit for known impurities , all these impurities were identified and none were of concern in terms of safety. The solubility and drug content of the SP and AQ tablets was confirmed by analysis with high performance liquid chromatography at the London School of Hygiene & Tropical Medicine, which was also used after the trial to confirm that drugs had been allocated to the correct group in accordance with the randomization code. Tablets were crushed, mixed with honey or suspended in water and given on a spoon. All participants were kept under observation for 30 minutes after drugs were taken. If vomiting occurred during this period, the full dose of treatment was re-administrated. If a child vomited the second administration treated was not repeated but he/she was allowed to stay in the trial.To determine the proportion of children who experienced an adverse event such as headache, fever, weakness, abdominal pain, anorexia, nausea, vomiting, diarrhoea, rash, itching or sleep disorder following drug administration, study participants were visited at home three days after the start of each round of drug administration and a side effects questionnaire completed to document and quantify adverse events that might have occurred since receiving the trial medication. Grading of the severity of adverse events was done by trained field workers. An adverse reaction was graded as mild (grade 1) if it was easily tolerated, moderate (grade 2) if it interfered with normal activity and severe (grade 3) if it prevented normal activity and required treatment.To help establish whether adverse events were drug related or coincidental, the same questionnaire was administered to children in the control group on completion of each treatment round.Passive surveillance for malaria was maintained throughout the transmission season for 16 weeks (September to December). Parents/guardians of children in the trial and controls were encouraged to take their child to the health centre identified as being closest to their home  at any time that their child became unwell. Project staff were based at each of these health facilities to identify children in the trial and to ensure that they were seen, properly investigated and treated promptly. At each clinic visit, axillary temperature was recorded using a digital thermometer and haemoglobin concentration measured. A dipstick for diagnosis of malaria  was used if fever (axillary temperature of \u226537.5\u00b0C) or a history of fever within the previous 48 hours was present. In such cases, a thick blood smear was also collected for subsequent confirmation of the diagnosis. Study subjects with documented fever or history of recent fever and malaria parasitaemia were treated with Coartem\u2122. The treatment of study subjects seen at the health centres for other conditions was carried out in accordance with national guidelines and a standardized form was used to document details of the illness and treatment received.Children enrolled in the study were seen at a health centre at the end of malaria transmission season for examination by a study physician and a finger-prick blood sample was obtained for preparation of a thick blood smear and determination of haemoglobin concentration. A standardized questionnaire was administered to the parents/guardians of the study subject to collect information regarding any illness that had occurred since the last visit, symptoms experienced, use of healthcare facilities and use of medicines.Thick smears were prepared in duplicate so that if the subject had symptoms of malaria, one smear could be stained with Field's stain and read promptly to guide treatment. The other smear was stained with Giemsa stain and 200 high power fields (HPF) were examined before a smear was declared negative. Only the Giemsa-stained slide readings were used for the trial analysis. Parasite density was expressed per \u00b5l with the assumption that 1 parasite per high-powered field (hpf) equals 500 parasites per \u00b5l Haemoglobin concentration was measured at recruitment, during morbidity surveillance and at the end of malaria transmission season surveys using a portable haemoglobinometer .The primary study endpoint was the proportion of children who reported an adverse event during at least one of the three follow up surveys. Secondary endpoints were the number of clinic attendances with malaria during the surveillance period and the mean Hb concentration and the prevalence of asexual parasitaemia recorded in the cross-sectional survey in December. Sample size was calculated for comparison of each treatment group to the group that received SP plus AQ, the best regimen identified in a previous trial conducted previously in Niakhar, Senegal The primary analysis included all individuals who were allocated to treatment provided and who were followed up at the three visits that they received at least one follow-up visit, regardless of the number of treatments received. Data were analysed according to the treatment group to which a child was allocated. The prevalence of each type of adverse event was computed for the different treatment and control groups and presented with an exact binomial confidence interval. Fisher's exact test was used to test for differences in prevalence between the treatment groups. The overall frequency of adverse events was computed separately for each month and a Chi-square test for trend was used to test for a time trend in the prevalence of adverse events.For the secondary analysis, malaria incidence was estimated for the treatment and control groups. To do this we calculated the total child years of follow-up in each group and the number of first cases of malaria. Rates were calculated with exact confidence intervals based on the Poisson distribution. For rate ratios, exact Poisson regression was used to obtain confidence intervals. All treated cases with parasitaemia at any density were included in the analysis of efficacy. Differences in mean Hb concentrations between treatment groups in December were estimated using analysis of covariance, adjusting for the Hb measurement at enrolment. All analyses were done with STATA software version 10 .The study was approved by the London School of Hygiene & Tropical Medicine ethics committee and by the joint MRC/Gambia Government ethics committee.Plasmodium falciparum parasitaemia was found infrequently at enrollment.One thousand two hundred children aged 6\u201359 months were screened and allocated to receive monthly IPT with SP plus AQ, SP plus PQ or DHA plus PQ. One thousand and eight children (84%) who were enrolled, treated and followed up for at least one visit were included in the primary analysis. The first dose of IPT treatment was given between 13\u201324 September 2007, the second dose between 16\u201326 October and the final dose between 13\u201324 Novembers. A cross-sectional survey was carried out between 6\u201313 December 2007. One of these 1008 children died in a road traffic accident and a further 23 were lost to follow up before the end of the surveillance period . Loss toField workers visited study subjects three days after the start of each treatment round to document the incidence of adverse reactions. A summary of solicited adverse events is provided in To test whether the risk of reported adverse events varied with the number of doses received, separate analyses were carried out for each of the treatment rounds and the results showed significant decrease in the risk of adverse events in the SP plus PQ group when measured after the 2nd and 3rd dose compared to the first round .The incidence of clinical attacks of malaria was generally low in all the treatment groups; overall incidence of malaria among children in the trial was 0.22 per child year . No child had more than one episode of malaria. The incidence of malaria in the DHA plus PQ, SP plus AQ and SP plus PQ groups was 0.10 , 0.06  and 0.06  respectively. The incidence of malaria in the non-randomized control group measured by passive case detection was 0.79   and 7. TNinety-seven percent (982/1008) of the study children  were seen during the end of the rainy season survey when blood samples for determination of parasitaemia and haemoglobin were collected. The prevalence of parasitaemia was very low . Mean Hb concentration was similar in the three groups  . ModeratIn this study, we have compared the safety, tolerability and efficacy of three potential drug combinations for intermittent preventive treatment of malaria in children. All three treatment regimens investigated, including the one used in the previous Senegalese trial (SP plus AQ), were safe and efficacious. The incidence of malaria during the trial and the prevalence of parasitaemia at the end of the malaria transmission season were very low in all the three treatment groups. These results are consistent with findings from previous trials of IPTc carried out in other West African countries which showed significant levels of protection against clinical attacks of malaria during the malaria transmission season All three combination investigated were safe. No serious adverse event attributable to study medication was observed. Minor adverse events observed in this trial occurred infrequently in all the treatment groups and were similar in the three treatment groups. Minor adverse events were more common in the control group than in the three treatment groups and these were very likely due to common childhood illnesses such as malaria as the incidence of malaria was higher in the control group compared to the three treatment groups. Previous studies of IPTc in Senegal suggested that significantly more minor adverse events occur in children who received amodiaquine-containing preparations than in those who received artesunate and SP AQ has been used widely for treatment of uncomplicated malaria in the past and is now used in combination with artesunate as first line treatment for malaria in several countries in Africa with no apparent problems over safety. A review of the use of AQ for treatment of malaria in Africa concluded that it is safe SP has been used widely for IPT in infants and in pregnant women because of its long half-life, safety, wide availability, low-cost, ease of delivery (a single dose treatment) and a good acceptability profile. The main concern about the use of SP for IPT is the propensity for this drug to cause severe skin reactions in a small proportion of subjects. No adverse cutaneous events were recorded in children in the current study. The main threat to the use of SP in IPT regimens comes from increasing resistance to SP, especially in East Africa. Thus, there is a need to find alternative drugs for chemoprevention which should, ideally, be used in combination to reduce the likelihood of the emergence of resistance.As there is now increasing evidence that IPT works in infants and children through its prophylactic effect Piperaquine is attractive as a candidate drug for IPT as it is safe, with a toxicity profile similar to that of chloroquine, and an estimated terminal elimination half-life of approximately 17 days. The combination of piperaquine with SP is a logical one for IPT as the two drugs have long half lives. However, currently there is no good manufacturing practice (GMP) product of piperaquine available for general use. Dihydroartemisinin-piperaquine is an artemisinin-containing, fixed-combination antimalarial drug, developed in China, which is safe and highly effective in the treatment of clinical malaria For IPTc to be an effective malaria control tool, a reasonable level of compliance with drug-taking must be maintained with minimum outside support. Whether this can be achieved will depend upon the attitude of both the health authorities who administer the drug and the mothers/guardians who take their children to the health centre or to the community worker to receive the tablets. In the current study, compliance was generally good for all the treatment regimens. However the number of children who completed all the three doses was significantly less in the SP plus AQ treatment group compared to the SP plus PQ and DHA plus PQ groups. It is not clear if the lower compliance was due to the reduced tolerability of AQ seen in other studies as the frequency of minor adverse events observed in all the treatments groups was similar.A potential weakness of the trial was that it was not a placebo-controlled trial, the optimum design for most clinical trials, but this approach was not considered desirable because a trial conducted along these lines would have resulted in some children not receiving IPTc. As IPTc has been shown previously to reduce morbidity from malaria, it was considered ethically unacceptable to include a control group. The possibility of unobserved selection bias and confounding cannot be ruled and the results provided on the differences in the incidence of clinical episodes of malaria in the treatment groups compared with the control group must be treated with caution. However, it appears that all treatment groups were highly efficacious.IPT in infants and pregnant women has a major advantage over IPT in older children in that a delivery system for drug administration, the reproductive and child health clinic (RCH) already exists. In contrast, the optimum delivery strategy for IPTc is not known. A number of studies are under way to identify ways in which this highly efficacious intervention could be most effectively delivered.There may be other groups of children who are at particular risk from malaria and anaemia who could benefit from IPT. These include children with sickle cell disease or HIV infection. In older children, seasonal IPT might improve school attendance and performance. Thus, there is a need to investigate whether SP plus AQ, SP plus piperaquine (PQ) and dihydroartemisinin (DHA) plus PQ might be equally safe and effective when used for IPT in these groups of children who are at particular risk of malaria.Protocol S1(0.17 MB DOC)Click here for additional data file.Checklist S1CONSORT checklist.(0.05 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria in several countries, as a mean of prolonging the effectiveness of first-line malaria treatment regimens. A three-day course of artesunate-mefloquine  has been adopted by Malaria Control Programme of Thailand as first-line treatment for uncomplicated falciparum malaria all over the country since 2008. The gametocytocydal anti-malarial drug primaquine is administered at the dose of 30 mg (0.6 mg/kg) on the last day. The aim of the present study was to assess patient compliance of this combination regimen when applied to field condition.Artemisinin-based combination therapy (ACT) is presently recommended by the World Health Organization as first-line treatment for uncomplicated Plasmodium vivax parasitaemia during follow-up were included into the study. The first dose of the treatment was given to the patients under direct supervision. All patients were given the medication for self-treatment at home and were requested to come back for follow-up on day 3 of the initial treatment. Baseline (day 0) and day 3 whole blood mefloquine and plasma primaquine concentrations were determined by high performance liquid chromatography.A total of 240 patients  who were attending the malaria clinics in Mae-Sot, Tak Province and presenting with symptomatic acute uncomplicated falciparum malaria, with no reappearance of Two patients had recrudescence on days 28 and 35. The Kaplan-Meier estimate of the 42-day efficacy rate of this combination regimen was 99.2% (238/240). Based on whole blood mefloquine and plasma primaquine concentrations on day 3 of the initial treatment, compliance with mefloquine and primaquine in this three-day artesunate-mefloquine combination regimen were 96.3% (207/215), and 98.5% (197/200), respectively. Baseline mefloquine and primaquine levels were observed in 24 and 16% of the patients.The current first-line treatment and a three-day combination regimen of artesunate-mefloquine provides excellent patient compliance with good efficacy and tolerability in the treatment of highly multidrug resistance falciparum malaria. Previous treatment with mefloquine and primaquine were common in this area. Plasmodium falciparum. Resistance has been observed to almost all currently used anti-malarials. The extensive deployment of anti-malarial drugs in the past fifty years has provided a remarkable selection pressure on malaria parasites to evolve resistance mechanisms. Due to the high resistance level of P. falciparum to the most affordable drugs, such as chloroquine and sulphadoxine-pyrimethamine, together with high recrudescence rates with monotherapy with artemisinin derivatives, artemisinin-based combination therapy (ACT) is presently recommended by the World Health Organization as first-line treatment for uncomplicated P. falciparum malaria in several countries as a mean of prolonging the effectiveness of first-line malaria treatment regimens . Since mefloquine is a long half-life drug (14-21 days in patients), the decrease in the concentrations within three days is insignificant and, therefore, day 3 concentrations in patients who had measurable baseline concentrations were estimated by subtraction of baseline concentrations from the measured day 3 concentrations. Figure st, 2nd and 3rd quatiles were 1,716, 2,359, and 3,059 ng/ml, respectively. Median (range) concentrations on day 3 were 89 (6-640) ng/ml, with a 95% confidence interval of 75-106 ng/ml. There was no patient with outlier primaquine concentrations, but undetecable concentration was observed in three cases. Whole blood mefloquine concentrations in these cases were as high as 2,669, 2,964 and 6,703 ng/ml. Based on primaquine concentrations on day 3 with exclusion of the pharmacokinetic factor, compliance with primaquine in this three-day artesunate-mefloquine combination regimen was 98.5% (197/200). There was no correlation between the pre-treatment concentrations of mefloquine and primaquine found in this study.Of the 214 patients whose primaquine concentrations were measured, 180 (84.1%) had undetectable baseline primaquine level, whereas 29 (13.6%), 4 (1.9%), and 1 (0.5%) had concentrations of less than 100, between 100 and 500, and more than 500 ng/ml, respectively [median (range): 0  ng/ml]. Figure Mefloquine was introduced first as a single-dose therapy for falciparum malaria in Thailand in 1984, but initial high cure rates were not sustained. In a proposition to halt the loss of anti-malarial monotherapies to resistance in rapid succession, the strategy of ACT with mefloquine was developed and was adopted in Thailand in 1994. The current regimen of a three-day combination of artesunate-mefloquine given in total adult dose of 600 mg (12 mg/kg body weight) artesunate and 1,250 mg (25 mg/kg body weight) mefloquine has been implemented by the Bureau of Vector Borne Disease, Department of Disease Control, Ministry of Public Health for the treatment of acute uncomplicated falciaprum malaria since 2008. The main objective was to obtain high cure rate of approaching 100% to prevent the spread of drug resistance. Prior to 2008, the same total dose of artesunate-mefloquine was given but as a shorter course of two days and two split doses of mefloquine  given on the same day at 6-8 hours apart instead of two separate days as the current regimen. With the current regimen, in addition to the achievement of high cure rate, the incidence of vomiting due to high dose mefloquine and thus treatment failure is expected to be reduced. Absorption of mefloquine in malaria patients is dose limited and is reduced in the acute phase of illness. Splitting the dose of 25 mg/kg mefloquine improves mefloquine absorption and bioavailability, and thus and the therapeutic response in the treatment of acute falciparum malaria -18.i.e. mefloquine with long half-life of 14-21 days [e.g. low dose phenobarbital, than the has been applied for monitoring compliance to short course treatment with anti-malarial regimens (5 day course of artesunate and 7 day course of the combination quinine-tetracycline) [max [In the present study, a good initial response was observed in all patients where parasitaemia was cleared within three days after the initial dose of artesunate and mefloquine. The present combination regimen of artesunate-mefloquine was shown to improve the cure rate from approximately 87% with the 2 day course to 96.3% in the same study area (Tak province) during the year 2001 to 2002 [-21 days  may not -21 days . Most me-21 days . In anotcycline) . The concycline) . It has ne) [max . The casne) [max . This exne) [max ,27. Co-fne) [max . NeverthIt is interesting to note that 24.2 and 15.9% of patients had mefloquine and primaquine levels at baseline pretreatment. Seven% had baseline mefloquine concentrations less than 100 ng/ml, 11.6% had concentrations between 100 and 500 ng/ml, and 13% had concentrations greater than 500 ng/ml. This implies that patients may have received previous treatment with mefloquine longer than 35-48, between 35-48, and within 14 days, respectively . In one The current first-line treatment three-day combination regimen of artesunate-mefloquine provides excellent patient compliance with good efficacy and tolerability in the treatment of multidrug resistance falciparum malaria in field setting. Previous treatment with mefloquine and primaquine was common in this area. The results obatined are useful for the malaria control programme of Thailand as baseline data for optimization of treatment regimen for uncomplicated falciparum malaria.The authors declare that they have no competing interests.KC participated in the design of the study and performed the statistical analysis. PB conceived of the study, and participated in study coordination. VM carried out the measurement of drug concentrations. KN partcipated in the design of the study, data analysis and drafted the manuscript. All authors read and approved the final manuscript."}
+{"text": "This resource will help guide rational drug policies that optimize antimalarial drug use, in the hope that the emergence and spread of resistance to new drugs can be, if not prevented, at least delayed.The proliferation of antimalarial drug trials in the last ten years provides the opportunity to launch a concerted global surveillance effort to monitor antimalarial drug efficacy. The diversity of clinical study designs and analytical methods undermines the current ability to achieve this. The proposed World Antimalarial Resistance Network (WARN) aims to establish a comprehensive clinical database from which standardised estimates of antimalarial efficacy can be derived and monitored over time from diverse geographical and endemic regions. The emphasis of this initiative is on five key variables which define the therapeutic response. Ensuring that these data are collected at the individual patient level in a consistent format will facilitate better data management and analytical practices, and ensure that clinical data can be readily collated and made amenable for pooled analyses. Such an approach, if widely adopted will permit accurate and timely recognition of trends in drug efficacy. This will guide not only appropriate interventions to deal with established multidrug resistant strains of malaria, but also facilitate prompt action when new strains of drug resistant plasmodia first emerge. A comprehensive global database incorporating the key determinants of the clinical response with  The emergence and spread of antimalarial drug resistance is one of the most important factors undermining malaria control programmes in most of the malaria endemic world. . FollowiThe number of clinical trials being conducted has risen dramatically over the last decade. This is primarily in response to an increasing appreciation of the major impact that antimalarial drug resistance has had on the health of tropical communities, the perceived need for the rational deployment of new regimens based on empirical evidence and a significant increase in the funding for these critical endeavours. Numerous groups have now generated a wealth of knowledge on different treatment regimens in different sites over a prolonged period of time. Unfortunately many studies remain unpublished. While each trial may have been designed with an immediate applicability for local policy makers or to test a specific hypothesis, an opportunity exists to extract information of global relevance.in vitro, molecular and pharmacokinetic information will directly inform decisions about which drugs should be used in specific settings to assure adequate cure rates for afflicted populations  Data from patients with different follow-up periods can be combined and efficacy estimates generated at different time points. These can then be compared between studies with different length of follow-up.2) All available data contributes in the analysis, thus increasing the precision of the derived estimates.3) The approach avoids systematic biases introduced by dropping patients from the analysis that do not complete follow-up  or classifying patients as failures who do not represent true biological failures .In survival data each patient is characterized by a period of observation and whether or not treatment failure occurred during this observation period (the \"status\"). This can be derived from four key parameters:1) The last day of follow-up2) The patient outcome on the last day of follow-up3) The parasite species on the day of failure4) The genotyping results The outcomes for all patients on the last day of follow-up need to be categorized into mutually exclusive groups that cover all possible endpoints Table . These c1) Patients who complete the study and do not meet criteria for treatment failure. For these patients, the last day of follow-up is the full duration of the study period.2) Patients who meet criteria for treatment failure; in this case the last day of follow-up is the day when treatment failure occurred.3) Patients who neither complete the full follow-up period nor meet the criteria for treatment failure . For these patients, the last day of follow is the day when the patient was last observed.Since the primary objective is to determine the prevalence of parasite resistance, failure is defined as early treatment failure during the first few days after the start of treatment or the first reappearance or persistence of parasitaemia during subsequent follow-up. These definitions are consistent with the current WHO guidelines.in vivo response into the four simple parameters listed above, allows one to generate in a transparent and flexible manner, the status of the patient required for survival analysis. Table Condensing the patients' The proposed system standardizes the process of data collection in a robust and yet flexible manner, allowing an analysis that accommodates the inevitable diversity of study methodologies and collection methods. Importantly it still retains the ability to present alternative analyses, such as proportions of failures on any particular day. Whatever analytical method is used the process should be transparent to analysts interpreting the findings. Such transparency is now evident from the coding table Table . This apData can be gathered retrospectively from completed studies but this may require recoding of the data to produce the key parameters outlined in Table Plasmodium falciparum recurrence (unadjusted and adjusted by genotyping) up to day 28 and day 42 and the cumulative risk of recurrence with Plasmodium vivax at day 28 and day 42. Subgroup analyses could be performed selecting patients according to certain age and parasitaemic limits to minimize the confounding of immunity and parasite biomass.The global database will initially focus on deriving six standardized efficacy estimates: the cumulative incidence of Although the risk of treatment failure is the main component of the therapeutic response, other parameters of interest can also be included, such as parasite and fever clearance times, haematological recovery, gametocyte carriage, and drug safety and tolerability Table . As dataTo demonstrate the utility of such a system data have been collected from 82 studies with 162 treatment arms from 25 countries. In total individual records from 25,214 patients have already been pooled and the day 28 recurrence rates estimated (PCR adjusted and unadjusted). The latter are available online  along wiThe goal of a global database is to characterize temporal and geographical trends in antimalarial efficacy, allowing information-based decision making among health policy developers and implementers. Retrospective analysis of the demise of old drugs could potentially provide useful information on how a similar fate might be avoided when deploying new treatments. Ultimately, the strength of an international resource lies in its ability to collate and report rapidly the current data on antimalarial efficacy. This will ensure that the international community has the best opportunity to detect and react to the earliest warnings of failing drug regimens. The additional benefit of documenting what is currently known is that it becomes apparent where important gaps in the data exist, thus helping to prioritize clinical and parasitological studies of antimalarial efficacy in these areas.Another important component of the World Antimalarial Resistance Network will be the development of additional resources to increase local capacity to conduct and interpret clinical trials. In this respect an open-access software program will be developed to provide a universal database system ready for sites to implement. This free software would provide a much needed service to clinical investigators conducting antimalarial efficacy trials in malaria endemic countries. At the same time, the data would be collected in a format that could easily be uploaded to the global database, at the discretion of the local study team. The system would also promote adherence to standardized protocols and facilitate the collection of the most critical variables for basic efficacy and safety measurements.in vitro susceptibility assays and pharmacokinetic parameters with the clinical data. This can be done both at the population (in time and place) or individual level. Such merging will allow the investigation and validation of surrogate markers of clinical efficacy. The power of a comprehensive data collection would allow a multivariate analysis to identify parameters, other than parasite resistance, associated with the therapeutic outcomes. This information could then be used to rationalize public health strategies (e.g. optimizing dosing recommendations and identification of groups of patients at particular risk).As the data from the other global databases proposed in this edition of the journal accrue, the opportunity will arise to cross-link molecular, There are numerous challenges to interpret the data that have already been gathered, and to develop acceptable and reliable methodologies that will improve study design and data collection and analysis in the future. The first step toward these goals is to ensure that research groups, policymakers, governing bodies and funding agencies perceive tangible benefits that can be accrued from sharing both data and methodologies. These benefits need to be balanced by the perceived risks to ownership and privacy when releasing individual patient data to a third party. Transparent protocols need to be developed that comply with international ethical standards, protect intellectual property rights, and ensure that patient data remain confidential.Five key variables are required to derive efficacy estimates for each treatment Table . SubgrouWhere drug efficacy surveillance systems are being developed the benefits of adopting established methodologies and analytical approaches are compelling. However it is often difficult to change a system that is already in place and appears adapted to local needs. The challenge is to ensure that the proposed changes in the coding of data are simple and easily incorporated into established practices. Importantly the uniform collection of the key variables advocated in this paper will circumvent the inevitable variations in protocol design and execution, which reflect local requirements and capacities. From the feedback of the participants in a pilot project, it appears that this is so, and that, importantly, the process of cleaning, validating and analyzing data is greatly improved. This is currently an evolving system and, therefore, there are likely to be problems that need to be resolved and areas that need further clarification. Feedback from those involved in conducting efficacy studies is welcomed and can be registered as a readers comment on this article.in vitro susceptibility testing central to detecting early warning signals of resistance to ACTs. Furthermore developing a global resistance database to track clinical efficacy will help rationalize treatment practices, ensure ongoing surveillance, and identify factors most conducive to maintaining parasite susceptibility to available treatment regimens.In the last ten years the emergence of multidrug resistant strains of plasmodia raised the spectre of untreatable malaria for poorly resourced communities. The development of ACTs has brought the hope of not only rescuing some compromised antimalarial drugs, but also delaying the emergence and spread of new resistant strains. In addition improved funding has paved the way for the discovery of novel antimalarial compounds, some of which should become available in the next two to ten years. Although the next ten years look much brighter for malaria treatment programmes, it would be a disaster if that led to complacency. Plasmodia have proved efficient at evolving resistance to novel agents and this is likely to continue. Hence it is vital that prospective monitoring of antimalarial drug efficacy remains an international priority, with estimates of clinical efficacy and The author(s) declare that they have no competing interests."}
+{"text": "Plasmodium falciparum malaria, are more effective against sexual stage parasites (gametocytes) than previous first-line antimalarials and therefore have the potential to reduce parasite transmission. The size of this effect is estimated in symptomatic P. falciparum infections.Artemisinin combination therapies (ACT), which are increasingly being introduced for treatment of Data on 3,174 patients were pooled from six antimalarial trials conducted in The Gambia and Kenya. Multivariable regression was used to investigate the role of ACT versus non-artemisinin antimalarial treatment, treatment failure, presence of pre-treatment gametocytes and submicroscopic gametocytaemia on transmission to mosquitoes and the area under the curve (AUC) of gametocyte density during the 28 days of follow up.ACT treatment was associated with a significant reduction in the probability of being gametocytaemic on the day of transmission experiments (OR 0.20 95% CI 0.16\u20130.26), transmission to mosquitoes by slide-positive gametocyte carriers (OR mosquito infection 0.49 95% CI 0.33\u20130.73) and AUC of gametocyte density (ratio of means 0.35 95% CI 0.31\u20130.41). Parasitological treatment failure did not account for the difference between ACT and non-artemisinin impact. The presence of slide-positive gametocytaemia prior to treatment significantly reduced ACT impact on gametocytaemia (p < 0.001). Taking account of submicroscopic gametocytaemia reduced estimates of ACT impact in a high transmission setting in Kenya, but not in a lower transmission setting in the Gambia.Treatment with ACT significantly reduces infectiousness of individual patients with uncomplicated falciparum malaria compared to previous first line treatments. Rapid treatment of cases before gametocytaemia is well developed may enhance the impact of ACT on transmission. Plasmodium falciparum malaria [P. falciparum infections.With resistance to chloroquine and sulphadoxine-pyrimethamine well-established in many malaria endemic countries, artemisinin combination therapies (ACT) are now being introduced for first line treatment of  malaria ,2. Clini malaria  and no s malaria -6. This  malaria ,8. Furth malaria . By redu malaria -12 and t malaria ,14. To aPrevious summary analyses of randomized ACT trials have shown a reduction in gametocyte prevalence at single timepoints during follow up but have focussed on clinical efficacy ,15. GameRaw data were pooled from six previously published randomized antimalarial trials in which participants were assessed for infectivity to mosquitoes after treatment Table  13,19-2-219-22. P. falciparum malaria with >500 asexual parasites/\u03bcl who sought treatment at a health centre were randomized to treatment groups. Altogether four non-artemisinin antimalarial regimens were tested: chloroquine (CQ), sulphadoxine-pyrimethamine (SP), CQ-SP, sulphadoxine-pyrimethamine-amodiaquine (SP-AQ) and four ACT: chloroquine-artesunate (CQ-AS), SP-AS1 (single dose of artesunate), SP-AS3 (3 days artesunate) and artemether-lumefantrine (AL). Dosage schedules were as follows: study 1: CQ: 10 mg/kg days 0\u20133, SP: 250 mg sulphadoxine +12.5 mg pyrimethamine if \u2264 10 kg, plus 125 mg sulphadoxine + 6.25 mg pyrimethamine per 5 kg increase in bodyweight day 0, SP-AS1: as SP, plus 4 mg/kg AS day 0, SP-AS3: as SP, plus 4 mg/kg AS days 0\u20132; study 2: SP & SP-AS3: as study 1; study 3: CQ: 10 mg/kg days 0\u20131, 5 mg/kg day 2, CQ-AS: as CQ, plus 4 mg/kg AS days 0\u20132; study 4: CQ: as study 3, SP: 25 mg/kg sulphadoxine, 1.25 mg/kg pyrimethamine day 0, CQ-SP: as CQ & SP; study 5: CQ-SP: CQ as study 1 days 0\u20132 plus SP day 0 as study 1, AL: 20 mg artemether plus 120 mg lumefantrine per 5 kg body weight up to 24 kg, or 120 mg artemether plus 720 mg lumefantrine for 25 kg+ 0 h, 8 h, 20 h, 32 h, 44 h, 56 h; study 6: SP: as study 4, SP-AQ: as SP plus 10 mg/kg AQ days 0\u20132, SP-AS3: as SP plus 4 mg/kg AS days 0\u20132, AL: as study 5.The study design and laboratory methods for each trial are described in detail elsewhere -22,24. IGiemsa-stained blood sample smears were made on a number of days during follow-up until day 14 or day 28 after treatment , or from a laboratory colony (studies 5 & 6), were fed on blood samples through an artificial membrane at 37\u00b0C. Mosquito midguts were dissected at 7\u20138 days and the number of oocysts in the mosquito midgut was determined by microscopy.Venous blood samples for human-to-mosquito transmission experiments were taken on a specified day of follow-up  the area under the gametocyte density-time curve (AUC), 2) the proportion of patients with slide-positive gametocytaemia on the day of mosquito feeding, 3) the proportion of mosquitoes infected with oocysts (the most readily quantifiable parasite stage in the mosquito), and 4) oocyst density in each dissected mosquito midgut. Asexual parasite and gametocyte densities are reported as arithmetic rather than geometric means to take into account zero values and the variability of the data. The AUC for each individual was calculated by taking the arithmetic mean of adjacent timepoints and multiplying by the intervening number of days, using data from days 0, 7, 14 and 28. Individuals missing gametocyte density readings on day 0, day 28, or at more than one other timepoint were not included in the AUC analysis.Individual treatments were grouped into ACT and non-artemisinin antimalarials. One treatment regimen which contained only 1 dose of an artemisinin derivative  was grouped separately. Antimalarials which were used in at least two studies were also examined individually; for the remainder, treatment effect could not be well distinguished from study effect. Gametocyte prevalence at day 0 and parasitological treatment failure were investigated as modifiers of ACT effect on all four outcomes.The impact of ACT and other covariates on the AUC (outcome 1) was analyzed using negative binomial regression, which models the log of the arithmetic mean and gives a good fit to the skewed distribution . RegressAll analyses were undertaken using Stata .Data were available on 3,174 patients in the six trials, of whom 529 contributed data to the human-to-mosquito transmission results  and OR of high oocyst density 0.49 (0.07\u20133.62)  compared to ACT-treated individuals  Figure . This maSlide-positive gametocytaemia on day 0, indicating the presence of mature gametocytes before treatment, was predictive of higher gametocytaemia during follow up. ACT impact on gametocytaemia was reduced among those with pre-treatment gametocytes (ratio of gametocyte AUC means 0.80 (95% CI 0.64\u20131.00) and OR of gametocyte prevalence on the day of feeding 0.41 (95% CI 0.25\u20130.67) compared to those negative for pre-treatment gametocytes (ratio of gametocyte AUC means 0.23 (95% CI 0.19\u20130.28) and OR of gametocyte prevalence on the day of feeding 0.13 (95% CI 0.10\u20130.17) (interaction p < 0.001 for both outcomes) Figure . The patIn study 6, the molecular QT-NASBA technique detected gametocytes in 311/332 (94%) of blood samples that were classified gametocyte-positive by microscopy, and furthermore in 615/959 (64%) of those classified negative, giving a higher estimated prevalence of gametocytes at all timepoints . GametocAmong patients who were tested by both QT-NASBA and microscopy (n = 249), ACT impact on gametocytaemia was smaller if submicroscopic gametocyte carriers were taken into account. The ratio of means of the area under the gametocyte prevalence-time curve comparing ACT to non-artemisinin antimalarials was 0.40 (95% CI 0.23\u20130.69 p < 0.001) using microscopy data, compared to 0.60 (95% CI 0.51\u20130.70 p < 0.001) with QT-NASBA data. This difference suggests bias is introduced by microscopy because a greater proportion of ACT-treated gametocyte carriers have a density in the submicroscopic range.In study 5 in The Gambia, 1/1165 (0.1%) mosquitoes were infected when feeds were performed on 33 individuals who were gametocyte slide-negative, compared to 19/882 (2.2%) mosquitoes who were infected by 26 gametocyte carriers in the same study. This suggests that infection of mosquitoes by submicroscopic gametocytaemia was rare in this setting. By contrast, gametocyte slide-negative individuals in study 6 in Kenya infected almost as high a proportion of mosquitoes as gametocyte slide-positive patients: 4.7% compared to 5.9%. In this study the impact of ACT was substantially smaller if \"being infectious\" was defined as infecting any mosquitoes (regardless of microscopic gametocytaemia), rather than as being gametocyte slide-positive on the day of feeding .These results demonstrate that gametocytaemia among ACT-treated patients is significantly lower than in individuals treated with non-artemisinin antimalarials during 28 days of follow up. ACT treatment is also associated with reduced human-to-mosquito transmission both among those with microscopically detectable gametocytes, and randomly selected patients. ACT appear to destroy a substantial proportion of immature, developing gametocytes while sequestered in the microvasculature, resulting in a significant reduction in the release of mature gametocytes into the peripheral blood. Their more rapid reduction of the asexual reservoir may also contribute.Transmission to mosquitoes was measured mainly at day 7 when ACT impact on gametocytaemia was at one of its highest points. Although gametocyte density is not the only factor determining transmission to mosquitoes , theHigher levels of parasitological treatment failure among non-artemisinin treated individuals did not account for any of the higher observed ACT impact on gametocytaemia and transmission to mosquitoes. However, parasitological failure did predict higher gametocytaemia and transmission to mosquitoes. Drug resistant parasite genotypes have been shown to enhance transmission by boosting gametocyte numbers, particularly in patients treated with monotherapies ,20. The Study 6 in Kenya  highlighThe grouping of different treatments into non-artemisinin and ACT categories in these results is simplistic since individual treatment regimens within these groups sometimes had significantly different associations with the outcomes Table . TherefoWhile it is useful to have a pooled estimate of ACT impact from these studies, the heterogeneity in results should not be overlooked. The laboratory methods used in each of the studies were mostly very similar, but there were some variations which may have influenced the estimated ACT impact, such as the number of days between treatment and measurement of human-to-mosquito transmission, and different microscopists. For example, the SP-AS3 treatment group in study 1 did not show reduced oocyst density among blood-fed mosquitoes compared to non-artemisinin treatments, but only 12 patients in this group participated in feeding experiments. Of the 6 individuals successfully infecting mosquitoes at day 4, two were already harbouring gametocytes at day 0, and a third had very high day 4 gametocytaemia. These individuals almost certainly carried gametocytes on day 4 representative of a substantial pre-treatment gametocyte population, and may have been less infectious if transmission experiments were performed 7 or 14 days after treatment, as in the other studies analysed. Differences in baseline characteristics between studies such as transmission setting could also have caused differences in ACT effects.These results are representative of ACT effects upon young symptomatic patients in a trial setting. In order to evaluate potential impact of ACT on population-level transmission, it will be crucial to know the proportion of infections which are asymptomatic and so untreated in different transmission settings, and their relative contribution to the infectious reservoir. Many other operational factors including the coverage of ACT relative to other antimalarials and levels of adherence to treatment regimens will also be important in determining such effects. Empirical observations on population gametocyte carriage and its evolutionary basis ,35 will ACT reduce gametocytaemia and onward transmission to mosquitoes significantly compared to previous first-line non-artemisinin antimalarials, although transmission from ACT-treated patients is not fully prevented. Malaria transmission intensity could potentially be reduced as ACT are scaled up in malaria-endemic countries, however it will be important to quantify the proportion of asymptomatic and therefore untreated infections in different populations. ACT impact is likely to be reduced by factors which increase the presence of mature gametocytes before treatment, such as lack of rapid access to treatment or high malaria transmission intensity. Future studies of different antimalarials should also measure the infectiousness of gametocyte carriers with submicroscopic densities, to further clarify the contribution of such individuals to post-treatment transmission.LCO designed the analysis with ACG, carried it out, and drafted the manuscript, ACG also revised the manuscript, CJD and CJS conceived the idea of a summary analysis, CJD, CJS and TB provided data from the original trials, revised the manuscript critically and aided the interpretation."}
+{"text": "Use of different methods for assessing the efficacy of artemisinin-based combination antimalarial treatments (ACTs) will result in different estimates being reported, with implications for changes in treatment policy.in vivo studies of ACT treatment of uncomplicated falciparum malaria were combined in a single database. Efficacy at day 28 corrected by PCR genotyping was estimated using four methods. In the first two methods, failure rates were calculated as proportions with either (1a) reinfections excluded from the analysis  or (1b) reinfections considered as treatment successes. In the second two methods, failure rates were estimated using the Kaplan-Meier product limit formula using either (2a) WHO (2001) definitions of failure, or (2b) failure defined using parasitological criteria only.Data from different Data analysed represented 2926 patients from 17 studies in nine African countries. Three ACTs were studied: artesunate-amodiaquine , artesunate-sulphadoxine-pyrimethamine  and artemether-lumefantrine .Using method (1a), the day 28 failure rates ranged from 0% to 39.3% for AS+AQ treatment, from 1.0% to 33.3% for AS+SP treatment and from 0% to 3.3% for AL treatment. The median [range] difference in point estimates between method 1a (reference) and the others were: (i) method 1b = 1.3% [0 to24.8], (ii) method 2a = 1.1% [0 to21.5], and (iii) method 2b = 0% [-38 to19.3].The standard per-protocol method (1a) tended to overestimate the risk of failure when compared to alternative methods using the same endpoint definitions (methods 1b and 2a). It either overestimated or underestimated the risk when endpoints based on parasitological rather than clinical criteria were applied. The standard method was also associated with a 34% reduction in the number of patients evaluated compared to the number of patients enrolled. Only 2% of the sample size was lost when failures were classified on the first day of parasite recurrence and survival analytical methods were used.in vivo study should be to provide a precise estimate of the risk of antimalarial treatment failure due to drug resistance. Use of survival analysis is the most appropriate way to estimate failure rates with parasitological recurrence classified as treatment failure on the day it occurs.The primary purpose of an  The protocol was revised in 1967 and 1972, with patients followed up for 28 days and kept in a mosquito-free environment to prevent re-infection. Subsequent modifications to the protocol in 1996 permitted a shorter length of follow-up of 14 days in areas of high transmission, where the main endpoint changed from parasitological clearance without subsequent recrudescence to adequate clinical response, i.e. a patient in whom parasites reappeared without symptoms was still regarded as 'cured'. In 2001, the length of follow-up was increased to 28 days in areas of intense transmission, if PCR genotyping was available, and the concept of late treatment failure incorporating clinical or parasitological failures was introduced. Asymptomatic patients with parasitological failure were followed from the day of reappearance to the last day of follow-up when they were treated if parasites were still present  be analysed using the traditional 'per-protocol' approach\", i.e. failure rates expressed as proportions with exclusion of patients lost from the study for any reason other than genotyping-confirmed recrudescence before the last day of follow-up. An Excel\u2122 programme was developed to enable users to get their results analysed with per-protocol and Kaplan-Meier methods. In practice, survival analysis has not been used widely, but consensus appears to be growing that it is the method of choice ,9.PCR genotyping allows the reappearance of parasites during follow-up to be classified as a new or recrudescent infection. However the technique is not without its limitations. Methods are not standardized with different laboratories using different numbers of genotypic markers and different methods of interpretation . Distingin vivo studies using the WHO protocol are used to guide changes in treatment policy. Current recommendations (WHO 2006) are that a new antimalarial drug combination should be 95% efficacious when deployed and change should be considered when efficacy falls to 90%  differences between these results and those obtained using method 1b were 1.3% [0\u201324.8], method 2a 1.1% [0\u201321.5] and method 2b 0% [-38\u201319.3].Classifying patients as failures on day 7 if initial parasitaemia had not cleared, or on the first day of parasite recurrence, regardless of symptoms (method 2b) increased the number of treatment failures (uncorrected by genotyping) by 75, from 1015 to 1090 but it should be noted that the majority of these patients (50) were from one site (Boende).Figure Recurrence of parasitaemia between days 14 and 28 occurred in 942 (32%) children. Of these, 77 (8.3%) patients could not be classified because: (i) 18 samples were missing, (ii) in 31 samples it was not possible to amplify any parasite DNA, (iii) DNA was amplified but no classification could be given (indeterminate) in 26 samples, and (iv) the reason was unknown for two samples.A total of 865 recurrences were categorized following PCR genotyping with: (i) 118 (12.5%) classified as recrudescent, (ii) 734 (77.9%) as re-infections, and (iii) 13 (1.4%) as recrudescence + re-infection, regarded as recrudescent for the purposes of this analysis. The use-effectiveness was 91.8% (865/942) and ranged from 60.0% for Congo-Kindamba to 100.0% for Angola-Kuito and Uganda-Kampala and Bobo-Dioulasso, Burkina Faso  and withdrawals (4%) was small over 28 days.If the results of the survival analysis using the same definitions of failure (WHO 2001) are compared to the results of the per protocol analysis the median difference in the point estimates for failure at day 28 was small at 1.1% but ranged from 0 to 21.5%. Again, the difference was greater at the sites with the highest numbers of parasite recurrences, whether related to transmission intensity and more reinfections or a higher number of recrudescent infections. The precision of those estimates generated by the per protocol analysis was lower. Comparisons of precision cannot be made against methods 1b or 2b because the endpoint classifications were different.The change in definition of failure from the WHO (2001) protocol to method 2b led to an increasing number of patients being classified as treatment failures (uncorrected by genotyping): 1090 compared to 1015. By classifying patients on day 7 or on the day of recurrence rather than by day 28 (method 2b vs. 2a), the results of the survival analyses changed little in most of the studies but markedly at certain sites where several patients who had been classified as ACPRs using the 2001 criteria were reclassified as late treatment failures using the 2005 definition, e.g. the Boende site in DRC where failure [95% CI] would have been estimated as 15.3% [8.7\u201325.9] when the study was done in 2003 and 56.3% [45.8\u201367.4] if it had been done 2 years later following the newer recommendations. This was because a relatively high proportion of patients who were still parasitaemic on days 7 (n = 29) and 14 (n = 21) but asymptomatic had cleared their parasites by day 28 without treatment.in vivo study depends on the study design and execution, the quality of the laboratory and the interpretation of the genotyping results. For the efficacy analysis, there is a growing consensus that the Kaplan-Meier survival analysis is the optimal method because it retains the maximal amount of data for analysis and avoids the inaccurate overestimation of failure rates that will occur when large numbers of patients are excluded from the analysis. This overestimation is likely to increase as a drug's efficacy declines with a consequent increase in the number of recrudescences and new infections. These results support expert opinion that survival analysis should become the standard way of analysing data from in vivo tests and should replace the traditional per-protocol method. The per-protocol method was associated with a 34% reduction in sample size, compared to a < 10% reduction for the other methods. However, it is true that while survival analysis maximizes the sample size, some of the patients will only have completed a trivial period of follow up. Regarding re-infections as treatment successes (method 1b) preserves the sample size but leads to an underestimation of the failure rate and is not recommended.The methods for assessing antimalarial drug efficacy continue to evolve. The quality of the data generated by an Efficacy of AS+AQ was below 90% at 7/15 sites and of AS+SP at 4/8 sites and the lower 95% CI was below this level at a further four and two sites, respectively. Efficacy of AL was high in the small number of patients studied. The reduced efficacy of AS+AQ and AS+SP could be explained by the use of the more slowly eliminated partner drug as monotherapy before and during the deployment of the combinations. An alternative explanation for these results is different methods of classification by PCR genotyping. The results reported here suggest that PCR genotyping performed well as a discriminatory test inasmuch as the majority of patients were classified, but the results are only as valid as the interpretation algorithms used to classify them. Differences in performance were observed between different laboratories, which did not appear to be related to the number of markers used. PCR outcomes defined as recrudescence + re-infection were classified as new infections by laboratories in certain studies but not in others as described in the methods. Current genotyping techniques, even using up to six markers, are not sufficiently discriminatory in areas of high transmission where multiple genotype infections are common . StandarThese results support the recommendation for a minimum follow-up period of 28 days in the assessment of efficacy of ACTs, with PCR-genotyping of all recurrences and the use of the Kaplan-Meier product-limit formula to estimate efficacy of treatment in all correctly randomized patients.Accurate reporting of methods by investigators is essential, in particular definitions of endpoints and criteria for censoring all categories of patients who do not complete follow-up. It is recommended that the classification of treatment failure is based on parasitological outcomes only and that all parasitological recurrences be treated straightaway in view of the high likelihood that patients will go on to develop symptoms . Furtherin vivo assessment to guide treatment policy. Such studies should be powered to estimate the effect size with adequate precision. However, if an efficacy assessment is being done because of a suspicion a drug is failing it makes sense to evaluate a potential replacement at the same time in an adequately powered, randomized, controlled trial. This will provide good evidence to guide national programmes in their drug selections.The WHO (2001) protocol was not designed to be a comparative study protocol, but rather a single-arm in vivo efficacy monitoring data should be used as the only determinant for policy change is debatable, if a choice of highly efficacious therapies becomes available. When comparing treatments of similar efficacy other factors determining programmatic effectiveness become more important to policy makers such as simplicity of the regimen, formulation, tolerability, adherence, cost and post-treatment prophylactic effect.Whether The authors declare that they have no competing interests.JPG, PO and WRT conceived of the study, and participated in its design and coordination and helped to draft the manuscript. EAA participated in its design, analysed some of the data and wrote the first draft of the manuscript. LP and ET did the statistical analysis and helped to draft the manuscript. GD, HB, IZ, IvdB, PPP, MvH, FC and SB collected data and revised the manuscript. All authors read and approved the final manuscript."}
+{"text": "Declining in clinical efficacy of artesunate-mefloquine combination has been documented in areas along the eastern border (Thai-Cambodian) of Thailand. In the present study, the clinical efficacy of the three-day combination regimen of artesunate-mefloquine as first-line treatment for acute uncomplicated falciparum malaria in Thailand was monitored in an area along the western border (Thai-Myanmar) of the country.Plasmodium falciparum malaria were included into the study. Patients were treated initially (day 0) with 4 mg/kg body weight artesunate and 15 mg/kg body weight mefloquine. The dose regimen on day 2 was 4 mg/kg body weight artesunate and 10 mg/kg body weight mefloquine. On day 3, artesunate at the dose of 4 mg/kg body weight was given with 0.6 mg/kg body weight primaquine. Whole blood mefloquine and plasma artesunate and dihydroartemisinin (active plasma metabolite of artesunate) concentrations following treatment were determined by high performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LCMS), respectively.A total of 150 Burmese patients  aged between 16 and 50 years who were attending the Mae Tao clinic, Mae-Sot, Tak Province, and presenting with symptomatic acute uncomplicated P. falciparum and reappearance of Plasmodium vivax in their peripheral blood during follow-up. The Kaplan-Meier estimate of the 42-and 28-day efficacy rates of this combination regimen were 72.58% (95% CI: 63.20-79.07%) and 83.06 (95% CI 76.14-94.40%), respectively. Parasite clearance time (PCT) and fever clearance time (FCT) were significantly prolonged in patients with treatment failure compared with those with sensitive response . Whole blood mefloquine concentrations on days 1, 7 and 14 in patients with sensitive and recrudescence response were comparable. Although plasma concentration of dihydroartemisinin at 1 hour of treatment was significantly lower in patients with recrudescence compared with sensitive response , the proportion of patients with recrudescence who had relatively low (compared with the lower limit of 95% CI defined in the sensitive group) was significantly smaller than that of the sensitive group.Thirty-four cases had recrudescence during days 7 and 42. Five and 5 cases, respectively had reinfection with P. falciparum to mefloquine contribute to some treatment failure following treatment with a three-day combination regimen of artesunate-mefloquine, results suggest that artesunate resistance may be emerging at the Thai-Myanmar border.Although pharmacokinetic (ethnic-related) factors including resistance of  Plasmodium falciparum, particularly the previous mainstay anti-malarial drug chloroquine, sulphadoxine-pyrimethamine and mefloquine is further aggravating the situation . In addition, the proportions of patients who had parasitaemia (PCT24 hr) and fever (FCT24 hr) cleared within 24 hours were significantly higher in the group with sensitive response (Table Patients with recrudescence (confirmed by PCR) and sensitive responses were well matched in regard to sex, age, weight, height, and parasite density on enrollment Table . The PCTvs 24.0 4.0-32.0 vs 9 (25%), 3 (33.3%) vs 2 (9%) and 5 (50%) vs 5 (23.8%) cases for concentrations on day 1, 7 and 14, respectively]  ng/ml]. For both artesunate and dihydroartemisinin, the proportions of patients in the recrudescence group who had the concentrations at 1 hour after the first dose lower than the lower limit of 95% CI for median concentrations defined in the sensitive group  was significantly smaller than the sensitive group  , 1, 7 and 14 of treatment are summarized in Table vs 9 5%, 3 (3324 hr) and fever (FCT24 hr) cleared within 24 hours. The efficacy of this combination was markedly dropped when compared with our recent investigation during 2008-2009 with the same regimen in the same area, where a 42-day cure rate of 99.2% was obtained  compared to the sensitive group [24.0 (14.0-32.0) hours]. Only 6 17.6%) and 7 (20.5%) patients with recrudescence response respectively, had parasitaemia . Dihydroartemisinin is the active plasma metabolite of artesunate, which has a longer half-life (1-2 hours) with potency 2-4 times greater than the parent drug [P. falciparum isolates in this area. There has been no defined threshold levels of artesunate/dihydroartemisinin for treatment of falciparum malaria but it is noted however that artesunate concentration (at 1 hour) of as low as 120 ng/ml was adequate in two patients with sensitive response, while dihydroartemisinin concentration (at 1 hour) of as high as 1,250 ng/ml was inadequate in one case with recrudescence. The later had also confirmed adequate mefloquine concentrations on days 1, 7 and 14. While a total of 12 patients had confirmed adequacy of whole blood/plasma concentrations of both mefloquine and dihydroartemisinin, in adequacy of the concentrations of mefloquine alone, dihydroartemisinin alone and both contributed to 13, 10 and one cases with treatment failure, respectively. None of the treatment failure vomited the doses of mefloquine or artesunate. It was reported that patients from Tak Province experiences a lower frequency of nausea and vomiting than other groups ['Genuine resistance' of treatment failure cases was confirmed by monitoring for the adequacy of whole blood mefloquine and plasma artesunate/dihydroartemisinin concentrations. Mefloquine concentrations on days 1, 7 and 14 of treatment in patients with recrudescence response tended to be lower than those with sensitive response and with larger proportion of cases with the concentrations  lower than the lower limit of 95% CI defined in the sensitive group. Blood concentrations of mefloquine on day 1 of treatment has been reported to be an important determinant of successful treatment . On the ent drug . This suin vitro sensitivity data of individual parasite isolate, it is difficult to clearly define whether reduced clinical efficacy of the combination is due to which of each partner drug. It is noted however that mefloquine concentration profiles in this patient population (Burmese) appear lower (by about 30%) than our previous reports in Thai patients following the same dose of mefloquine [in vitro susceptibility testing, pharmacokinetic or other host factors [in vitro susceptibility of the parasite isolates (baseline and the day of recrudescence) together with candidate molecular markers of resistance (amplification and SNPs of pfmdr1 and pfatp6) are under way, to clearly define, with support of drug concentration data, if the reduced artesunate-mefloquine efficacy observed is really due to a true decline in artesunate sensitivity or a further progression of mefloquine resistance or both [in vitro susceptibility of P. falciparum to mefloquine and artesunate/dihydroartemisinin in this area, and in particular vigilance to detect early emergence of higher levels of artemisinin resistance is requiredResults of clinical efficacy together with drug concentration profiles may suggest the possibility of contribution of pharmacokinetic factors as well as mefloquine and artesunate/dihydroartemisinin resistance to the observed high failure rate of the current three-day combination regimen. With the absence of floquine ,23. The  factors . Analysi or both ,26. Of pThe present study was the first observation of a marked decline in clinical efficacy of a three-day artesunate-mefloquine regimen in the area along the Thai-Myanmar border. Although pharmacokinetic (ethnic-related) factors including resistance of the parasite to mefloquine mainly contribute to some treatment failure, results suggest that artesunate resistance may be emerging at the Thai-Myanmar border.The authors declare that they have no competing interests.All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria in the Amazon region of Peru. Although MQ resistance has yet to be reported from the Peruvian Amazon, it has been reported from other countries in the Amazon Region. Therefore, continuous monitoring is warranted to ensure that the first-line therapy remains efficacious. This study examines the in vivo efficacy and pharmacokinetic parameters through Day 56 of three commercial formulations of MQ  given in combination with artesunate.Artemisinin-based combination therapy (ACT) is recommended as a means of prolonging the effectiveness of first-line malaria treatment regimens. Different brands of mefloquine (MQ) have been reported to be non-bioequivalent; this could result in sub-therapeutic levels of mefloquine with decreased efficacy. In 2002, mefloquine-artesunate (MQ-AS) combination therapy was adopted as the first-line treatment for uncomplicated P. falciparum mono-infection were randomly assigned to receive artesunate in combination with either (1) Lariam, (2) Mephaquin, or (3) Mefloquina AC. Patients were assessed on Day 0 , 1, 2, 3, 7, and then weekly until day 56. Clinical and parasitological outcomes were based on the standardized WHO protocol.Thirty-nine non-pregnant adults with Whole blood mefloquine concentrations were determined by high-performance liquid chromatography and pharmacokinetic parameters were determined using non-compartmental analysis of concentration versus time data.max) and interquartile range was 2,820 ng/ml  for Lariam, 2,500 ng/ml  for Mephaquin, and 2,750 ng/ml  for Mefloquina AC Farma. The pharmacokinetics of the three formulations were generally similar, with the exception of the Cmax of Mephaquin which was significantly different to that of Lariam (p = 0.04).By day 3, all patients had cleared parasitaemia except for one patient in the AC Farma arm; this patient cleared by day 4. No recurrences of parasitaemia were seen in any of the 34 patients. All three MQ formulations had a terminal half-life of 14\u201315 days and time to maximum plasma concentration of 45\u201352 hours. The maximal concentration (CAll three formulations had similar pharmacokinetics; in addition, the pharmacokinetics seen in this Peruvian population were similar to reports from other ethnic groups. All patients rapidly cleared their parasitaemia with no evidence of recrudescence by Day 56. Continued surveillance is needed to ensure that patients continue to receive optimal therapy. The World Health Organization recommends the use of artemisinin-based combination therapy (ACT) as a means of prolonging the effectiveness of first-line malarial treatment regimens -3. The eet al showed a significant difference in the bioequivalence of Mephaquin\u00ae (Mepha) compared with the reference product, Lariam\u00ae (Roche), with Lariam having nearly twice the maximum plasma concentration of Mephaquin (Cmax 1018 ng/ml vs. 656 ng/ml respectively) and requiring approximately 1/3 of the time to reach maximum concentration (tmax 13 vs 46 h) when pharmacokinetic parameters were assessed using plasma levels of MQ : Lariam = 8,159 , AC Farma = 4,280 , Mephaquin = 9,224 . There were no significant differences in the rate of parasite clearance between treatment arms (Table Thirty-nine adult subjects ranging in age from 18\u201361 years (mean 36 years) were initially enrolled in the study. Seventy-two percent of the patients were male. Each arm initially consisted of 13 individuals. A total of 5 subjects did not complete the full follow-up period. One subject was excluded from the Mephaquin arm on day 1 for hypertension requiring medication. Two were withdrawn due to adverse events: one from the Lariam arm on day 5, and one from the AC Farma arm on D28. Two were lost to follow-up: one from the AC Farma arm on day 35 and one from the Lariam arm on day 37. Thus, 11, 12, and 11 patients remained for full analysis of efficacy in the Lariam, Mephaquin, and AC Farma arms, respectively. The three groups were similar with regard to gender, age, and weight  following development of meningoencephalitis, presumed to be viral. Two other patients in the AC Farma arm had adverse events deemed unrelated to the study drug and remained in the study; one developed a subarachnoid haemorrhage and was hospitalized on study Day 48, the other required surgery for a traumatic spleen injury on study day 14.1/2) of 14.5\u201315 days and time to maximum plasma concentration (tmax) of 45\u201352 hours. The maximal concentration (Cmax), expressed as ng/ml was 2,820  for Lariam, 2,500  for Mephaquin, and 2,750  for Mefloquina AC Farma. The non-parametric Mann-Whitney test revealed a significant difference in the Cmax of Mephaquin relative to the reference (P < 0.05). There were no significant differences (P > 0.05) for all other parameters between the test and the reference groups. The coefficient of variation ranged from 16\u201337% for Cmax and AUC values.The pharmacokinetic analysis included all patients who had remained in the study until at least day 28, and thus excluded only two patients, one in the Mephaquin arm and one in the Lariam arm. Figure In this study, it was shown that although there is some manufacturer dependent variation in the drug levels of MQ, this did not appear to affect clinical outcomes. All patients rapidly cleared their parasitaemias with no evidence of recrudescence by Day 56. There was also no difference in occurrence of drug-related side effects between groups; all formulations were well-tolerated.1/2) of 14\u201315 days and time to maximum plasma concentration (tmax) of 45\u201352 hours. This is similar to previous reports in the literature [et al [et al [1/2 values of 16 and 11.5 days while tmax values were 46 and 12 hours, respectively. The tmax values may have corresponded more closely with Weidekamm et al [0-t, tmax, and t1/2between the three products. Although a statistically significant difference (P < 0.05) was observed in the Cmax of Mephaquin relative to the reference  for MQ pharmacokinetic parameters [et al discusses various approaches to deal with highly variable drugs [A large relative standard deviation (%RSD) was observed for the Crameters ,6,27. MaFollowing introduction of a new anti-malarial drug, it is important to continue to monitor drug efficacy. Although many studies use 28-days of follow-up, this length of follow-up may be inadequate for detecting early development of resistance. When combinations of drugs with discordant half-lives, such as mefloquine-artesunate, are used, the patient, in effect, receives monotherapy with the longer lasting drug for the tail end of therapy. This may lead to recrudescence, which may occur more than 4 weeks after administration of drug . It is cThe pharmacokinetics of three formulations of MQ administered in conjunction with artesunate to patients with uncomplicated malaria were similar to each other; in addition, it was shown that they are similar in the Peruvian population to that described previously in Southeast Asian populations. This is the first study to demonstrate the pharmacokinetics of mefloquine in a South American population. All three formulations appeared equally efficacious in non-pregnant adults, although the small sample size precludes a definitive evaluation of efficacy. Continued surveillance is needed to ensure that patients continue to receive optimal therapy.The views expressed in this article are those of the authors and do not necessarily reflect the official policy of the Department of the Navy, Department of Defense, nor the US Government. Some of the authors are military service members. This work was prepared as part of our official duties. Title 17 U.S.C. \u00a7 105 provides that \"Copyright protection under this title is not available for any work of the United States Government.\" Title 17 U.S.C. \u00a7 101 defines a U.S. Government work as a work prepared by a military service member or employee of the U.S. Government as part of that person's official duties. The study protocol was approved by the Naval Medical Research Center institutional review board in compliance with all applicable federal regulations governing the protection of human subjects.The authors declare that they have no competing interests.JG participated in analysis and interpretation of the data and drafted the manuscript. MG performed the pharmacokinetics studies, analysed and interpreted the results of the pharmacokinetics studies, and drafted the manuscript. SD was critical in enrolling patients and collecting clinical data and samples. OVR performed the quantification of the blood samples. BG performed the HPLC analysis of the samples to quantitate the concentration of the drug and assisted with the pharmacokinetic analysis and calculations. WMQ made substantial contributions to acquisition of data and samples. GCU made substantial contributions to the design and implementation of the study, secured funding for the protocol, assisted with sample collection, provided oversight of the study conduct, and participated in analysis and interpretation of data. LS participated in analysis and interpretation of the data and helped to draft the manuscript. TKR conceived the study, participated in the study design and revision of the manuscript. DJB oversaw the daily operation of the study and contributed to the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria and further define the CDA safety profile, particularly its hematological safety in glucose-6-phosphate dehydrogenase (G6PD) -deficient patients.Chlorproguanil\u2212dapsone\u2212artesunate (CDA) was developed as an affordable, simple, fixed-dose artemisinin-based combination therapy for use in Africa. This trial was a randomized parallel-group, double-blind, double-dummy study to compare CDA and artemether\u2212lumefantrine (AL) efficacy in uncomplicated P. falciparum malaria were randomized (2\u22361) to receive CDA 2/2.5/4 mg/kg once daily for three days (N\u200a=\u200a914) or six-doses of AL over three days (N\u200a=\u200a458). Non-inferiority of CDA versus AL for efficacy was evaluated in the Day 28 per-protocol (PP) population using parasitological cure (polymerase chain reaction [PCR]-corrected). Cure rates were 94.1% (703/747) for CDA and 97.4% (369/379) for AL . CDA was non-inferior to AL, but there was simultaneous superiority of AL (upper 95%CI limit <0). Adequate clinical and parasitological response at Day 28 (uncorrected for reinfection) was 79% (604/765) with CDA and 83% (315/381) with AL. In patients with a G6PD-deficient genotype , CDA had the propensity to cause severe and clinically concerning hemoglobin decreases: the mean hemoglobin nadir was 75 g/L  at Day 7 versus 97 g/L  for AL. There were three deaths, unrelated to study medication .The trial was conducted at medical centers at 11 sites in five African countries between June 2006 and August 2007. 1372 patients  with acute uncomplicated Although parasitologically effective at Day 28, the hemolytic potential of CDA in G6PD-deficient patients makes it unsuitable for use in a public health setting in Africa.ClinicalTrials.gov NCT00344006 P. falciparum resistance to these agents.Malaria is clearly an insupportable burden upon sub-Saharan Africa. Approximately 210\u2212300 million clinical malaria episodes and over 1 million deaths occur in this region annually Artemisinin-based combination therapy (ACT) is clinically effective, and may reduce malaria transmission and the potential for the development and spread of resistance Chlorproguanil\u2212dapsone\u2212artesunate (CDA) was developed as an affordable, fixed-dose ACT for use in Africa. Once-daily dosing over three days and good activity of the chlorproguanil\u2013dapsone (CPG\u2212DDS) component suggested a promising combination P. falciparum malaria and further define the CDA safety profile, in particular, its hematological safety in G6PD-deficient malaria patients. A randomized, parallel-group, double-blind, double-dummy, multicenter, Phase III study was conducted to demonstrate non-inferiority of CDA to AL based on parasitological cure rate (adjusted for polymerase chain reaction [PCR] genotyping) at study Day 28. Patients were young children and adolescents, representing the most clinically vulnerable population.Artemether\u2212lumefantrine (AL) is the current gold standard ACT. The six-dose regimen is highly effective and represents a challenging comparator for any new agent The protocol for this trial and supporting CONSORT checklist are available as supporting information; see This study was conducted in accordance with Good Clinical Practices, applicable regulatory requirements, and the Declaration of Helsinki. Approval was obtained from each participating center's ethics committee or institutional review board and the WHO Special Programme for Research and Training in Tropical Medicine. An Independent Data Monitoring Committee (IDMC) was convened and planned to conduct two interim safety analyses. The IDMC appointed an independent end-point reviewer (blinded to treatment assignment).P. falciparum malaria ; fever at screening or within the previous 24 h; weight \u22657.5 kg; hemoglobin \u226570 g/L or hematocrit \u226525%; willingness to comply with study procedures; written or oral witnessed informed consent from parent/guardian plus assent from patients \u226512 years old.Male and female patients (\u22651\u2013<15 years old) who met the following criteria were eligible for enrolment: acute uncomplicated P. falciparum malaria; known hypersensitivity/allergy to study treatments; known G6PD deficiency, methemoglobin reductase deficiency, hemoglobin M/E, or porphyria; neonatal hyperbilirubinemia; concomitant medication associated with hemolysis or hemolytic anemia; concomitant infection  or underlying disease that could compromise efficacy evaluation; malnutrition ; received treatment with another antimalarial that might have compromised study treatment evaluation, or an investigational drug within 30 days or 5 half-lives; or participated previously in the study. A negative pregnancy test was required from women of child-bearing age; breastfeeding mothers were excluded.Subjects were excluded if they had: severe/complicated At screening, a full medical history was obtained and a clinical examination undertaken. Eligible patients were randomized (2\u22361) to receive CDA  2/2.5/4 mg/kg/day once daily for three days, administered according to predefined dosing charts, or six-dose AL . One 20/120 mg AL tablet was given for patients weighing 5\u201315 kg, two for 15\u2013<25 kg, three for 25\u2013<35 kg and four for \u226535 kg, taken at study start (Day 0), 8 h later and then twice daily for two days. AL was dosed with milk. Matching placebos were administered to maintain blinding. Administration of therapy was directly observed and if vomiting occurred within 30 mins of dosing, the patient was re-dosed. If vomiting occurred again, the patient was treated with rescue medication  and followed until Day 42.Patients remained hospitalized until Day 3, returning for follow-up visits on Days 7, 14, 28 and 42. Field workers visited patients at home on Days 4, 5 and 6 to ensure they remained well. Clinical evaluation and temperature recording was performed before first dose, during therapy and at all clinic follow-up visits. Subjects failing therapy at any time during the study received rescue medication immediately and were followed until Day 42 or resolution.Venous blood samples (2 mL) were obtained for hematology assessments at screening and on Days 1, 2, 3, 7, 14, 28 and 42 and for standard clinical chemistry tests at pre-dose, on Days 3 and 42, and on Days 14 and 28 if previous assessment values were abnormal.P. falciparum msp-1, msp-2, and glurpAsexual parasite and gametocyte counts were performed at screening, at pre-dose, every 8 h during the in-patient stay until discharge on Day 3 and at follow-up visits on Days 7, 14, 28 and 42. At each time point, two thick and one thin film were prepared and parasite densities determined by examination of a thick blood slide (10 \u00b5L thumb prick), according to WHO methods A pre-dose blood sample (10 \u00b5L) was collected onto pre-printed filter paper for G6PD genotype analysis at two laboratories . Following DNA extraction, fragment amplification using PCR primers for loci 376 A\u2192G, 202 G\u2192A, 542 G\u2192T, 680 G\u2192T and 968 T\u2192C allowed recognition of the wildtype (G6PD B) and the common African mutations G6PD A and A\u2013 P. falciparum malaria and further define the CDA safety profile, in particular its hematological safety in G6PD-deficient patients.The objective of this trial was to compare CDA and AL efficacy in uncomplicated The primary efficacy endpoint was parasitological cure rate (PCR-corrected) at Day 28, defined as eradication of initial malaria infection by Day 7 plus aparasitemia through to Day 28. Parasitological cure was also evaluated at Days 14 and 42 as a secondary outcome.PCR-corrected parasitological cure was the standard method for assessing antimalarial efficacy at the time that the study was designed. However, publication of the WHO 2003 guidelines for evaluating antimalarials led to the protocol being amended to include adequate clinical and parasitological response (ACPR) as a secondary outcome Other secondary outcomes included: asexual parasite and gametocyte counts; mean parasite clearance time ; and mean fever clearance time .Adverse events were defined as any unfavorable and unintended sign , symptom, or disease (new or exacerbated) temporally associated with the use of a medicinal product. Adverse events were recorded at all clinic visits and coded using MedDRA (Version 10.1). Serious adverse events were defined as: death or life-threatening illness; resulting in hospitalization or prolongation of hospitalization; significantly disabling/incapacitating; congenital anomaly; hemoglobin decrease \u226540% versus baseline; hemoglobin <50 g/L; blood transfusion; methemoglobin \u226520% or \u226510 to <20% with associated clinical symptoms; or any other event considered significant by the investigator.A composite \u2018hemoglobin safety\u2019 endpoint was defined prospectively as: hemoglobin decrease of \u226540 g/L or \u226540% versus baseline or hemoglobin <50 g/L or blood transfusion.The study was designed with \u226590% power to test the hypothesis that CDA was non-inferior to AL for the primary outcome measure. Assuming efficacy of 93% for CDA and 95% for AL, 650 and 325 evaluable patients, respectively, were required using a one-sided hypothesis test at a 2.5% significance level and a 7% non-inferiority margin. Allowing for 30% loss, target recruitment was 930\u2236465 patients CDA:AL. No multiplicity was associated with this single primary comparison.GlaxoSmithKline generated the randomization schedule comprising blocks of six patients by center.Randomization was done at the study site by a telephone call to the automated Registration and Medical Ordering System (RAMOS). An individual patient's code could be accessed by an investigator via RAMOS in an emergency.Investigators, technicians performing microbiological assessments and patients were blinded to study treatments.The intent to treat (ITT) population included all randomized patients who received at least one dose of study medication. The PP population was a sub-set of the ITT population including patients who did not violate the protocol in a way that might impact the efficacy analysis.The primary analysis was performed on the Day 28 PP population. The difference in PCR-adjusted parasitological cure rates and the 95% confidence interval (CI) was calculated using the normal approximation to the binomial distribution. A similar analysis was performed for the ITT population and for ITT observed cases (i.e. excluding missing data). For the PP population, a logistic regression model was fitted to estimate treatment effect adjusted for country, age , and baseline parasitemia . Descriptive statistics were provided for secondary endpoints (PP and ITT populations) and safety outcomes (ITT population).For patients who were failures owing to new infections (determined using PCR), both PP and ITT analyses deemed these subjects to be successes at that time point. However, at subsequent time points, the PP analyses considered these patients as missing and the ITT analysis assumed that they were failures.Patients with missing data at the relevant time point were excluded from PP analyses of efficacy data . In the ITT analyses, treatment failure was imputed for missing data. For example, a patient with no Day 28 parasite count would be excluded from the primary endpoint and ACPR PP analyses but included as a treatment failure in the supporting ITT analyses. If a patient was missing a parasite assessment before Day 28, this would not affect assessment of Day 28 parasitological cure, whereas for ACPR, treatment failure could not be assessed and the patient would be excluded from the PP analysis with an imputation of failure in the corresponding ITT analysis.Comparisons of CDA and AL for the hemoglobin safety composite endpoint and changes in hemoglobin versus baseline were analyzed by the following G6PD genotype categories: G6PD normal, G6PD heterozygous females, and G6PD deficient . Logistic regression modeling was used to quantify the impact of G6PD genotype, weight, age, baseline hemoglobin, baseline parasitemia, and treatment on hemoglobin safety endpoints.Participant flow is shown in The trial was conducted between June 2006 and August 2007 at health centers in Bobo-Dioulasso, Burkina Faso; Kintampo, Ghana; Eldoret, Kilifi and Pingilikani, Kenya; Ibadan, Enugu, Jos and Calabar, Nigeria; and Bagamoyo and Kiwangwa, Tanzania.Baseline characteristics were similar between treatment groups; all patients were of Black African ethnicity . G6PD geDay 14, there were 787/914 (86%) evaluable patients in the CDA group and 393/458 (86%) in the AL group and at Day 42 661/914 (72%) and 332/458 (72%), respectively.Parasitological cure (PCR-corrected) at Day 28 in the PP population was 94.1% for CDA and 97.4% for AL , and no interaction with country, age or baseline parasitemia. The model provided overall adjusted cure rate of 95.2% for CDA and 97.9% for AL. The standard errors of these estimates were obtained using the delta method to provide the 95% confidence interval for treatment difference . This analysis supports the primary unadjusted analysis where CDA met the non-inferiority criterion of a lower 95%CI of \u2265\u22127%.Non-inferiority was maintained in the ITT analysis . The ITTP\u200a=\u200a0.834), age (P\u200a=\u200a0.479), or baseline parasitemia (P\u200a=\u200a0.179).For parasitological secondary endpoints , CDA wasThis study was not powered to compare treatments for individual countries. Per-country Day 28 cure rates (PP population) were as follows: Ghana/Burkina Faso, CDA 95.7% (110/115), AL 98.3% (58/59), treatment difference \u22122.7%, 95%CI \u22127.6, 2.3; Kenya, CDA 95.5% (150/157), AL 97.5% (77/79), treatment difference \u22121.9%, 95%CI \u22126.7, 2.8; Nigeria, CDA 94.1% (272/289), AL 98.0% (148/151), treatment difference \u22123.9%, 95%CI \u22127.4, \u22120.4); Tanzania, CDA 91.9% (171/186), AL 95.6% (86/90), treatment difference \u22123.6%, 95%CI \u22129.4, 2.2.ACPR at Day 28 in the PP population was 79% for CDA and 83% for AL; ACPRp was 93% and 94%, respectively . The proAsexual parasite counts decreased rapidly in both treatment groups; by 16 h post first treatment dose, parasite counts had declined by 99% in both groups. Mean (SD) parasite clearance time was 23.5 (11.0) h (N\u200a=\u200a909) for CDA and 26.2 (11.5) h (N\u200a=\u200a456) for AL (ITT population). For patients with baseline fever, mean fever clearance time (SD) was: 28.7 (23.3) h (N\u200a=\u200a538) and 26.6 (22.2) h (N\u200a=\u200a250) (ITT population).Gametocytes were present at baseline in 25/912 (3%) patients in the CDA group and in 10/458 (2%) in the AL group (ITT population). The proportion of gametoyctemic patients decreased throughout the study similarly in both treatment groups. For both the PP and ITT population analyses, the geometric means ranged between 1.0 and 1.1 parasites/\u00b5L throughout the study in both treatment groups. The median gametocyte counts were 0 prior to dosing and at each time point following dosing for both treatment groups.A subject was considered compliant with study treatment if they received the treatment to which they were randomized as well as the correct dose for their body weight on all three dosing days. Compliance with study medication was 94% for both treatment groups.The proportion of treatment-emergent adverse events due to any cause was similar between the treatment groups: 476/914 (52%) in the CDA group and 220/458 (48%) in the AL group. The majority of adverse events were mild to moderate in intensity . Adverse events reported by investigators as probably or possibly drug-related occurred in 184 (20%) patients in the CDA group and 86 (19%) in the AL group .Serious adverse events were reported for 63/914 (7%) patients in the CDA and 15/458 (3%) patients in the AL group. In the CDA group, serious adverse events most likely related to oxidative hemolysis occurred in 46/914 (5%) of patients versus 3/458 (<1%) in the AL group. In G6PD-deficient patients receiving CDA, 26/80 (33%) had such events  compared with 10/597 (2%) for G6PD normal  and 5/123 (4%) for female heterozygotes . In the AL group, malaria was the most common serious adverse event (7/458 [2%]). All serious adverse events in the AL group were in G6PD-normal patients, except one case of malaria in a heterozygous female.Overall in the ITT population, mean hemoglobin concentrations decreased versus baseline from Day 1 for both treatment groups , and werThe proportion of patients with a decrease in hemoglobin concentration versus baseline of \u226520 g/L was 305/914 (33%) with CDA and 112/458 (24%) with AL. By G6PD genotype, this endpoint occurred in 173/597 (29%) normal, 45/123 (37%) heterozygote and 45/80 (56%) deficient patients in the CDA group and in 77/322 (24%), 9/43 (21%) and 12/36 (33%), respectively in the AL group. Occurrences of the hemoglobin safety endpoint in G6PD-deficient patients were 35% (28/80) with CDA versus 0% with AL . A post-Fourteen blood transfusions were performed for a significant decrease in hemoglobin, all in the CDA group: 12 in G6PD-deficient patients, one in a heterozygous female, and one in a G6PD-normal patient. One subject in the CDA group received a transfusion for treatment of sickle cell crisis (G6PD status unknown). A further subject in the CDA group  was hospitalized on Day 6 requiring a blood transfusion (hemoglobin 38 g/L), though blood was not available. This patient subsequently recovered under close observation and supportive treatment with hematinics.Three deaths occurred during the study: two in the CDA and one in the AL group; none were attributed to study treatment by the investigator. In the CDA group, a 2-year-old female had severe malaria and severe herbal intoxication at Day 23 and 24 and died on Day 24; a 4-year-old male with sickle cell anemia with crisis and severe sepsis was inadvertently recruited into the study, withdrawn on Day 1 and transfused and died on Day 2. In the AL group, a 1-year-old male had severe pyrexia at Day 26 and died on Day 29. There were three other withdrawals because of adverse events, two with CDA (vomiting and febrile convulsion) and one with AL (hematuria), all of which resolved.For the primary endpoint of this trial, parasitological cure at Day 28 (PP population), CDA was highly efficacious (94%). CDA was non-inferior to AL, but AL was simultaneously superior to CDA. An adjusted logistic regression analysis (PP population) also showed AL superiority. Non-inferiority of CDA to AL for the primary efficacy endpoint was not maintained at Day 42. These results confirm the high efficacy of AL in Africa It was notable that there was no difference in reinfection rates between the two treatment groups. A recent comparison of CDA and AL also found no difference in reinfection rates Although this study was designed primarily to assess efficacy, the most clinically important findings relate to safety, specifically G6PD-related hemolysis in the CDA group. The 2\u22361 CDA:AL randomization schedule was chosen to provide adequate data for a meaningful analysis of CDA safety. Also, we defined a composite hemoglobin safety endpoint based on laboratory data for hemoglobin declines of clinical concern and blood transfusion events. Overall, the main risk factors associated with the composite endpoint were G6PD deficiency versus G6PD normal  and CDA treatment versus AL . In G6PD-deficient patients , 35% met the composite endpoint criteria with CDA versus none with AL. Of particular concern was that 12/15 patients requiring blood transfusion were G6PD-deficient, and all in the CDA group.This study was not powered to detect a difference in hematological safety in G6PD-deficient patients between treatment groups. However, subjects meeting the composite hemoglobin safety endpoint would clearly require hospital-based interventions. These are often difficult to access for the majority of malaria patients in sub-Saharan Africa. Based on the composite endpoint data, CDA has an unacceptable risk:benefit profile versus AL for the treatment of malaria in G6PD-deficient patients. Consequently, this population should not be exposed to CDA.The prevalence of G6PD deficiency in this study was 16% hemizygous males and 4% homozygous females; a substantial sub-population. The lowest prevalence of G6PD deficiency was 7.7% in Ibadan, though this is considerably lower than previous estimates, generally in excess of 20% G6PD phenotyping and genotyping are not generally available in Africa. Even if G6PD-deficient patients could be excluded, post-hoc regression analysis found that in G6PD-normal patients, the risk of a \u226520 g/L hemoglobin decrease versus baseline was slightly increased with CDA versus AL . The clinical relevance of this small difference is debatable, but further undermines CDA utility, given the availability of alternative agents. CDA was developed for use in Africa as a safe, affordable, effective and simple antimalarial. As these conditions cannot be met, the Joint Development Team decided to terminate CDA development and withdraw CPG\u2212DDS licenses.Although we knew that there would be some hemolytic effect of dapsone, the magnitude of the difference in this effect between CDA and AL in G6PD-deficient patients was unexpected In this study, comparison of the timecourse for hemoglobin decline with CDA and AL is instructive . For AL,Dapsone had been used in malaria and leprosy for many years with no major safety concerns et al., Day 3 and Day 14 data were sparse, but at Day 7 there was a significantly larger decrease in mean hemoglobin with CPG\u2212DDS than for the comparator sulfadoxine\u2212pyrimethamine (SP): treatment difference \u22124 g/L  et al. study becomes evident when the proportion of patients with hemoglobin decreases of \u226520 g/L at any time point is examined; this was 16% for both CPG\u2212DDS and SP In the CPG\u2212DDS safety study by Alloueche et al. study is considered. G6PD status was assessed retrospectively and only in a subset of patients, i.e. some patients with a \u226520 g/L hemoglobin drop and matched controls (by age and sex). There was no excess risk for a \u226520 g/L hemoglobin drop with CPG\u2212DDS versus SP in G6PD-normal or G6PD-deficient patients (classified as A\u2212 or A\u2212/A\u2212). However, for female heterozygotes, there was an excess risk with CD versus SP et al. results are in direct contrast to all of the G6PD-based analyses reported here, where G6PD-deficient patients were at greater risk of hemolytic events versus G6PD-normal and female heterozygote patients. Our study indicates that frequent hemoglobin assessment plus comprehensive G6PD genotype data are required for an adequate analysis of hemolytic potential in the G6PD-deficient sub-population.The position becomes even more complicated when the limited analysis of hemoglobin declines by G6PD genotype in the Alloueche A recently published study conducted in Rwanda comparing CDA versus amodiaquine (AQ) plus SP for the treatment of uncomplicated malaria is also informative One important question is whether the hemolysis observed here is new to CDA or would have been seen with CPG\u2212DDS versus AL. Preliminary results from a second Phase III clinical trial of similar design indicate no major differences in hematological safety between CDA and CPG\u2212DDS .There were some limitations in the hematological data collected in this study. A validated methemoglobin test, suitable for clinic use, was only available towards the end of the study. Consequently, only 32 patients from two centers  had screening and post-baseline methemoglobin data. Collection of reticulocyte data was not automated and there was no quality control process. On examination, reticulocyte percent and absolute data could not be interpreted and the results are not presented. G6PD phenotypic data were only available for 632/1372 (46%) patients because of logistical problems in transporting samples to a central laboratory within a narrow time window and data are not presented. The difficulties in obtaining sufficient G6PD phenotype data quickly hampered assessment of safety within the G6PD-deficient population by the IDMC during the recruitment phase. For the limited data available, concordance of a G6PD-deficient phenotype with genotype in hemizygous males was 271/313 (87%), and phenotype findings supported genotype findings with respect to both G6PD-deficiency prevalence and hemoglobin safety.To our knowledge, this is the first randomized controlled clinical Phase III multi-centre study to include a component investigating the relationship between hematological safety and G6PD deficiency. There is no accepted in vitro test or in vivo model to determine the potential for G6PD-related hemolysis. G6PD genotyping and separate analysis of hemizygous males/homozygous females versus heterozygote females and G6PD-normal patients was necessary to show the clinical importance of the interaction between CDA and G6PD deficiency for hematological safety. Our results highlight that new antimalarials, such as AL, may have significant safety as well as efficacy benefits over older therapies, illustrating the need to conduct comprehensive Phase III trials of antimalarial combination therapies against relevant comparators, even when the components are already approved individually. It also highlights the challenges of developing therapies for populations harboring G6PD deficiency and possibly other polymorphisms.Checklist S1CONSORT checklist(0.07 MB DOC)Click here for additional data file.Protocol S1Trial Protocol(0.44 MB PDF)Click here for additional data file."}
+{"text": "In Gabon, following the adoption of amodiaquine/artesunate combination (AQ/AS) as first-line treatment of malaria and of sulphadoxine/pyrimethamine (SP) for preventive intermittent treatment of pregnant women, a clinical trial of SP versus AQ was conducted in a sub-urban area. This is the first study carried out in Gabon following the WHO guidelines.dhps K540E mutation, as a molecular marker to predict SP-treatment failure.A random comparison of the efficacy of AQ (10 mg/kg/day \u00d7 3 d) and a single dose of SP (25 mg/kg of sulphadoxine/1.25 mg/kg of pyrimethamine) was performed in children under five years of age, with uncomplicated falciparum malaria, using the 28-day WHO therapeutic efficacy test. In addition, molecular genotyping was performed to distinguish recrudescence from reinfection and to determine the frequency of the P = 0.019) in the treatment of uncomplicated childhood malaria and for preventing recurrent infections. Both treatments were safe and well-tolerated, with no serious adverse reactions recorded. The dhps K540E mutation was not found among the 76 parasite isolates tested.The day-28 PCR-adjusted treatment failures for SP and AQ were 11.6%  and 28.2% , respectively This indicated that SP was significantly superior to AQ (The level of AQ-resistance observed in the present study may compromise efficacy and duration of use of the AQ/AS combination, the new first-line malaria treatment. Gabonese policy-makers need to plan country-wide and close surveillance of AQ/AS efficacy to determine whether, and for how long, these new recommendations for the treatment of uncomplicated malaria remain valid. Chloroquine (CQ) resistance is widespread in Gabon and this forced the health authorities to revise the national malaria treatment policy ,2. In JuPlasmodium falciparum CQ resistance was first reported in vitro in Gabon in 1983 [P. falciparum resistance to CQ [According to the data reported in the literature,  in 1983  and conf in 1983 . In 1987 in 1983 . Althoug in 1983 ,7, there in 1983 . It has ce to CQ . In Gaboce to CQ ,12, whilce to CQ ,14. A bece to CQ . A clinice to CQ .If the strategy of combining two or more antimalarial drugs is widely recommended as a means for improving treatment efficacy and delaying the development of resistance, a knowledge of the resistance level against individual components of the combination is essential. Policy-makers in Gabon urgently need to know whether the newly-adopted malaria treatment strategy is a viable option in terms of therapeutic efficacy. They also need to learn how best to use information from clinical trials for optimizing the policy or considering when to make a further change in treatment policy.dhps K540E mutation, previously defined as a predicting tool of SP-resistance [The objectives of this study were to determine the efficacy level of AQ and SP using the standard WHO 28-day therapeutic efficacy test, with PCR-genotyping to distinguish recrudescences from reinfections due to therapeutic failures. Additionally, the prevalence of the sistance , was alsGabon is a Central African country bordering Equatorial Guinea and Cameroon to the north, and the Republic of Congo to the east. It has a 800 km coastline, providing access to the Atlantic Ocean to the west. It is situated on the equator and, hence, has a hot and humid climate, with precipitation nine months out of twelve. A dense forest covers 85% of its territory. Officially, Gabon's population is approximately 1.5 million, with 80% of the population living in towns. There are eight main ethnic groups, which are spread out across the country, but more than forty different ethnic groups altogether. On an administrative level, Gabon is subdivided into nine provinces, namely: Estuaire; Haut-Ogoou\u00e9; Moyen Ogoou\u00e9; Ngouni\u00e9; Nyanga; Ogoou\u00e9-Ivindo; Ogoou\u00e9-Lolo; Ogoou\u00e9-Maritime; and Woleu-Ntem. The rainy season is from October to May, and the dry season is from June to September. Malaria transmission is perennial with seasonal fluctuations.Anopheles gambiae s.s is considered to be the most important vector of malaria, the Plasmodium index in children under 15 years of age is ranging from 65% to 73%, indicating an intense malaria transmission [This study was carried out between March-July 2005 in Oyem, a town of the Woleu-Ntem province of 35,000 inhabitants, near the northern borders with Cameroon and Equatorial Guinea, and 411 km away from Libreville  to the south-west. Malaria is hyperendemic and transmission is perennial. In this forest region, where smission .P. falciparum, parasitaemia \u2265 2,000 asexual parasites per \u03bcL of blood, free from severe malnutrition, absence of general danger signs or severe malaria, an axillary temperature of 37.5\u00b0C or above, absence of febrile conditions caused by diseases other than malaria, ability to come for the stipulated follow-up visits and easy access to the health facility, informed consent of parent/guardian, absence of history of hypersensitivity reactions to sulphonamides, and at least 5 g haemoglobin (Hb)/dL. Neither a history of previous antimalarial drug use, nor the presence of antimalarial drugs in the urine was an exclusion criterion in the WHO standard protocol [The study was conducted in fchildren with fever, below five years of age, attending the Out-patient Department of Oyem Hospital. Using the 28-day WHO therapeutic test for intense transmission areas of malaria , patientprotocol . Before Sample size estimations for the therapeutic test were performed by assuming that the treatment failures rate would be significantly less than 15% for SP and AQ, referring to previous clinical trials outcomes. Thus, according to WHO recommendations, a minimum of 50 patients was required for each therapeutic group .\u00ae tablets, Pfizer, Dakar, Senegal) 30 mg/kg body weight over three days (i.e. 10 mg/kg daily), or SP  as a single dose of 25 mg/kg of sulphadoxine/1.25 mg/kg of pyrimethamine.After obtaining informed consent from parents or guardians on day 0, the enrolled children were randomly allotted to one of two treatment groups, to receive either AQ base  was systematically given on day 0 and if needed on day 1 and 2. All tablets were administered orally by a nurse in the presence of the physician. For 30 min following drug administration, patients were observed for vomiting and other side-effects. The same dose was re-administered if vomiting occurred. On days 1 and 2, symptoms, other medications, temperature, and physical examination were recorded, but microscopy was not performed unless one or more of danger signs were present. The same clinical observation was repeated and parasitological examination was conducted on days 3, 7, 14, 21 and 28. On day 0, when the patient presented a fever without parasitaemia in the absence of another pathology the child was seen on the next day for intensive follow-up and microscopic diagnosis of malaria. All treatment failures were treated with quinine tablets (8 mg/kg base three times daily for seven days).The responses to drug treatment were classified according to the WHO protocol , as an a\u00ae filter papers  were dried and stored at room temperature in small separated and sealed plastic bags, prior to genotyping analysis at the National Malaria Reference Centre, Bichat-Claude Bernard Hospital, Paris, France. DNA was extracted with chelex-100 resin as previously reported [dhps K540E mutation.Finger-prick blood samples blotted on Isocodereported . Merozoireported , was ampreported . Genotypreported . The casreported , was perThe local health and institutional authorities approved the research protocol . Verbal and written informed consent for participation were obtained from parents or guardians, after thorough information on the study was provided in the local language.P-values of < 0,05 were calculated to demonstrate differences statistically.Data were analysed using version 2000 of the Epi-info software  and Graphpad Instat software . Proportions were compared by performing Fisher's Exact test. The therapeutic response at day-28 of follow-up with its corresponding 95% confidence intervals was calculated using an intention-to-treat analysis, which included all patients who fulfilled enrolment criteria. Standard deviation (SD) was generally indicated for means and Of the 632 febrile children with suspected malaria who were screened, three hundred-two  had confirmed malaria, of whom 154 met the inclusion criteria. Three hundred forty-four  were male; their mean age was 32.2 months.P = 0.0195). Of the 32 late treatment failures recorded in AQ group, detailed PCR-genotyping results were as following: 13 recrudescences, six reinfections, seven mixed results (recrudescence + reinfection) and six unconclusive results (PCR-amplification failure). Regarding SP group, detailed PCR-genotyping results among 14 late treatment failures were as following: eight true recrudescences and six reinfections. SP was superior to AQ for preventing recurrent infections  and parasitaemia. Overall, fourteen patients were excluded or lost to follow up (seven in AQ group and seven in SP group) during the follow-up period because of failure to follow the protocol  or failure to come for follow-up on the scheduled days . Details of the patient follow-up are reported in Figure dhps K540E mutation observed in the present study was already described in Gabon [dhps K540E mutation as a molecular marker predicting clinical SP-treatment failures is not true either in the existing literature or in the present study where 11.6% of SP resistance was found without a single instance of dhps K540E mutation. The AQ-resistance level reported in the study is too high, according to WHO guidelines (AQ-treatment failures >25%). The present results regarding AQ efficacy are consistent with other studies from Lambaran\u00e9 (Moyen-Ogoou\u00e9 province) and Bakumba (Haut-Ogoou\u00e9 province), reporting worse AQ efficacy with PCR-uncorrected resistance rates ranging from 34.7% to 47% on day 28 [The results of this study show, both clinically and parasitologically, that SP is significantly superior to AQ in terms of efficacy, with day-28 PCR-corrected failure rates of 11.6% and 28.2% respectively. SP also was more effective than AQ for preventing recurrent infections. Both regimens were safe and well-tolerated, with no serious adverse reaction recorded during the course of the study. Although protocols of previous studies were not in conformity with the standard WHO protocol, our data concerning SP efficacy are consistent with those reported in other parts of the country, such as in Bakoumba (Haut-Ogoou\u00e9 province) indicating a treatment failure rate of 14% at day 28 without PCR-adjustment in children under 10 years ,10,24. Tin Gabon  and in nin Gabon ,25,26. Tn day 28 ,14. In cn day 28 . Though n day 28 . Howevern day 28 ,30; thisIn conclusion, geographical differences of AQ efficacy discussed in this study are a major concern in Gabon now that the AQ/AS combination has been selected as the new malaria first-line treatment policy. Thus, further investigations carefully conducted in the respect of good clinical practices are necessary to ascertain AQ-resistance at the national level.P. falciparum infection. The results of this longitudinal prospective study will provide an essential baseline of the parasitological efficacy of those monotherapies and also to be able to understand, at a later stage, the natural history of the evolution of resistance to ACTs.In a country where ACTs are implemented, classical antimalarial drugs, including AQ and SP, must be monitored by recruiting persons with asymptomatic BN, VG, RD, JLB and MK designed the study and contributed to the discussion. MMM, JRMM, EN, MBA and RL participated to the clinical study. JRMM, BN, RD processed samples and analysed the data. BN wrote the first draft of the manuscript, then VG, RD, JLB and MK critically reviewed the manuscript. All authors read and approved the final manuscript.The author(s) declare that they have no competing interests."}
+{"text": "Introduction. To enhance effective treatment,african nations including Ghana changed its malaria treatment policy from monotherapy to combination treatment with artesunate-amodiaquine (AS+AQ). The major challenge to its use in loose form is adherence. Objective. The objectives of this study were to investigate adherence and treatment outcome among patients treated with AS+AQ combination therapy for acute uncomplicated malaria. Methodology. The study was conducted in two rural districts located in the middle belt of Ghana using quantitative methods. Patients diagnosed with acute uncomplicated malaria as per the Ghana Ministry of Health malaria case definitions were randomly allocated to one of two groups. All patients in both groups were educated about the dose regimen of AS+AQ therapy and the need for adherence. Treatment with AS+AQ was supervised in one group while the other group was not supervised. Adherence was assessed by direct observation of the blister package of AS+AQ left on day 2. Results. 401 participants were randomized into the supervised (211) and unsupervised (190) groups. Compliance in both supervised (95.7%) and unsupervised (92.6%) groups were similar (P = .18). The commonest side-effects reported on day 2 among both groups were headaches, and body weakness. Parasite clearance by day 28 was >95% in both groups. Discussion/Conclusions. Administration of AS-AQ in both groups resulted in high levels of adherence to treatment regimen among adolescent and adult population in central Ghana. It appears that high level of adherence to AS-AQ is achievable through a rigorous education programme during routine clinic visits. Plasmodium species to commonly-used anti-malarial drugs [P. falciparum malaria in Ghana [It is estimated that about 2.7 million malaria deaths occur annually with about 90% occurring in Africa . Betweenal drugs . Artemisal drugs  and redual drugs . Artesunin Ghana . In Ghanin Ghana . HoweverAdherence to ACTs could be enhanced by treatment supervision by trained community workers. Supervisory home visits by a trained community worker form a major component of the Community Health Planning and Services programme being implemented by the Ghana Health Service . This study assessed adherence to AS+AQ therapy and also the effect of adherence to AS+AQ therapy on clinical and parasitological outcomes.2 and are located in the forest-savannah transitional zones in the middle belt of Ghana. The total population based on the Health and Demographic Surveillance System (HDSS) of the Kintampo Health Research Centre (KHRC) was approximately 126000 in the 2007 census  and the main economic base is subsistence farming. The mean monthly temperature ranges from 18\u00b0C to 38\u00b0C, and the average rainfall is 1250\u2009mm per annum, creating optimal conditions for malaria transmission at a rate of about 269 infective bites per person per year [The study was carried out in the Kintampo North and South districts of Ghana. The two districts cover in total an area of 7162\u2009kmper year .Adolescent and Adult patients (age\u2009>\u200914 years) seen at community-based clinics and diagnosed as having uncomplicated malaria  were eligible for enrolment into this study. Patients who were unable to tolerate oral therapy, not resident in the study area or unwilling to take part in the study as well as pregnant women, were excluded from the study.Patients with signs and symptoms were assessed and recorded by a study clinician. Participants who satisfied the eligibility criteria  were recruited into the study and randomised into one of two groups, supervised or unsupervised groups.Artesunate and amodiaquine  were prescribed according to body weight  and administered twice daily for 72 hours. All participants in each group received the first dose of antimalarial under observation and were educated about AS+AQ dose regimen and the need for adherence as per Ghana's malaria treatment guidelines.Randomization was carried out in blocks of twenty by tossing a coin to determine the first group. The first block of twenty participants was allocated into the supervised group. The subsequent block of twenty participants was allocated into unsupervised group and these were alternated until the desired sample size was achieved. These large blocks were used to prevent overrepresentation of patients coming from the same community allocated to one group.Participants in the supervised group were visited at their homes by trained community workers to administer subsequent treatment regimen under direct observation on the second and third days of treatment, and post-treatment days . The participants in the unsupervised group were advised to self-administer the treatment and were visited at home on the third day for assessment of their adherence to the treatment regimen, then on the seventh, 14th, and 28th posttreatment days for parasite clearance and clinical resolution. At each visit a symptom assessment questionnaire was used to determine a patient's clinical outcome. Participants found not to be improving were referred to the health facility, reviewed further, and treated by the study clinician.Venous blood samples (1\u2009mL) were obtained prior to treatment to determine the haematological, biochemical, and malaria microscopy. Haemoglobin levels were determined using ABX Micros 60 Haematology Analyzer . Biochemical analyses were determined using Selectra E Clinical Chemistry analyser . Malaria parasite density was estimated from malaria parasite counts per 200 white blood cells and then extrapolated to malaria parasites per microlitre of blood with an assumption that a microlitre of blood contains 8000 leucocytes. Two hundred thick film fields were examined before assigning a negative malaria diagnosis. Prepared blood smears were read by expert microscopy readers in Kintampo Health Research Centre. Ten percent of positive and negative blood smears were randomly selected and confirmed by an independent blinded microscopist.P. falciparum parasitaemia by microscopy and clinical symptoms reported by day 7 determined as the proportion of study participants in each group with symptoms of malaria reported through a structured questionnaire by day 7.The main outcome measure was adherence to AS+AQ combination therapy. This was defined as the correct number of tablets of artesunate or amodiaquine left on the third day (day 2) of treatment when the treatment regimen was expected to be completed. This was assessed by direct observation of the blister package of artesunate-amodiaquine tablets. Secondary outcome measures included parasite clearance rates on days 14 and 28 determined as the proportion of study participants in each group without It was expected that about 65% of study patients would adhere to medication under supervision and about 50% would comply with the treatment without any supervision. A sample size of 182 per arm was estimated given 95% confidence interval and a power of 80. Assuming a 10% drop-out rate, about 202 participants were recruited per group.P-value of \u2264\u2009.05 was considered indicative of a statistically significant difference.Data were analysed using STATA 9 . Baseline characteristics and the endpoints were compared between the two groups. Point estimates were summarized as means and proportions and interval estimates as ranges and 95% confidence intervals. Biochemical and hematological indices were described according to the CTC AE v.3 grading classification. All statistical tests were two sided, and a The study was approved by ethics committees of the Ghana Health Service. The study participants were enrolled after obtaining informed written consent.A total of 520 participants were screened for inclusion into the study. 211 and 190 participants were randomized into the supervised and unsupervised groups, respectively . The average age of the participants in the supervised and unsupervised groups were 38.1 and 39.5 years, respectively, and there was no statistically significant difference in the baseline characteristics between the two groups. The mean weight among both groups was 49.6\u2009Kg (48.5\u201350.7) . The meaP = .18) There was no association between sex, age, or education and adherence to treatment regimen in the supervised and unsupervised groups  but decreased by Day 7 . Body weakness, bodily pains, joint pains, and drowsiness were the other commonly reported side effects after receiving treatment with AS-AQ. There were uncontrollable body movements among 10% and 5.9% of supervised and unsupervised participants, respectively. These were mainly mild uncontrollable extension of the neck and tongue. These cases resolved spontaneously without hospitalization.There were no marked differences in the reported side effects within the first one week of treatment between the two groups . HeadachP < .01). This pattern was similar in the unsupervised group . Majority of the participants haemoglobin levels decreased by a margin between 0.9\u2009g/dL and 2\u2009g/dL from day 0 to day 2 .The proportion of participants with anaemia (hb\u2009<\u200911.0\u2009g/dL) increased significantly between day 0 and day 2 in both groups. The proportion of anaemia among the supervised group increased significantly from 12.6% on day 0 to 38.8% on day 2 , Alanine Transaminase (ALT), Creatinine (Cr), and Urea (Ur) analyses were carried out on day 0 and day 2 to assess liver and renal functions. The proportion of high\u2009\u2009(\u2265\u2009grade 3 on CTC AE V3 classification) biochemical indices were similar among both groups on days 0 and 2 .P = .37) and day 2 . Though the proportion of high (\u2265\u2009grade 3 on CTC AE V3 classification) creatinine was similar among the two groups at day 0 and 2, there was a significant increase by Day 2 compared with day 0 among both supervised (P < .01) and unsupervised (P \u2264 .01) groups.The proportion of high creatinine levels were similar among both groups on day 0  among those supervised, compared with 99.5% (185/186) among those unsupervised. Day 28 parasite clearance was 95.3% (201/211) and 97.4% (185/190) among the supervised and unsupervised groups, respectively. PCR was not carried out to distinguish recrudescence from reinfections.In choosing a new drug or combination of drugs for treatment of malaria, one has to consider several factors such as therapeutic efficacy, side effects, patients' perception about the treatment regimen, cost, and availability and accessibility of the drug(s) in question. While side effects and treatment regimen are key determinants to patient's adherence, perception of illness, treatment-seeking behaviour, and acceptability and affordability of the drug will also influence adherence. Poor treatment practices may lead to suboptimal blood levels or one of the drugs could be given as monotherapy leading to population resistance as occurred with chloroquine. Thus, periodic evaluation of adherence to treatment regimen and optimum ways to ensure adherence among target populations is important.Adherence to the three-day treatment regimen of AS+AQ was very high in this study in both supervised and unsupervised participants. In a feasibility study of the use of artemether-lumefantrine which has a more complicated three day treatment regimen, >80% of care-givers in southern Ghana correctly administered the drug . A similAdherence to treatment is likely to be affected by factors such as age, number of tablets and duration of treatment. In the case of AS+AQ adult patients are expected to take between 12 and 24 tablets of AS+AQ tablets over three days. The introduction of the fixed dose of AS+AQ has halved the number of tablets and this is likely to enhance adherence . HoweverThe side-effects reported in this study are similar to those reported in controlled studies  with theSupervision of home treatment of malaria by health workers was perceived to be acceptable among participants in the supervised group. They were willing to allow health workers within their communities to supervise their drug administration. In areas where community-based health programme services (CHPSs) are available, public health nurses could supervise malaria treatment, observe the progress of the sick child, and refer where appropriate. This process will increase parasite clearance and prevent resistance as has been reported among children in Gabon . There was a significant increase in the proportion of anaemia among the study participants. A decrease in haemoglobin levels was observed on day 2 following treatment; this was similar to the pattern of haemoglobin dynamics among younger children treated with artemisinin combination drugs (including AS+AQ) in the same area. In these children, there was an initial drop in haemoglobin followed by an increase by day 14 . A similThe efficacy of AS+AQ in adult study group in this high malaria transmission area is comparable to AS+AQ efficacy results in other studies where malaria transmission is similarly high: 98.6% obtained in northern Ghana , 92.7% iIn conclusion, both supervised and unsupervised administration of AS+AQ resulted in high levels of adherence to treatment regimen among an adult population in a rural district in Ghana. Although this may have been influenced by the intensity of research related activities, it nevertheless gives a good indication of the potential to achieve high level patient adherence through rigorous routine patient education.Limitations. Adherence to therapy was assessed using the number of tablets left in the sachets which is dependent on what participants showed to the investigators. A better method could have been an estimation of drug levels in blood or urine. However, this test is expensive. In the absence of such estimates, the observation method was used. In communities were participants in both groups existed concurrently, adherence levels could have been influenced by contamination. However, this is unlikely due to the short duration between drug treatment and adherence assessment during which sick adolescents and adults are unlikely to move around to cause contamination. Additionally, randomisation was done in blocks of 20 to minimise contamination during the short treatment duration.K. P. Asante, R. Owusu, D. Dosoo, S. A. Etego, and S. Owusu-Agyei were involved in the design and implementation of the study. K. P. Asante, R. Owusu, D. Dosoo, E. Awini, G. Adjei, S. A. Etego, and S. Owusu-Agyei contributed to the analysis of the data collected. K. P. Asante, R. Owusu, E. Awini, G. Adjei, D. Chandramohan, and S. Owusu-Agyei contributed to the write up of the paper. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria as part of a multicentre study in Asia. Survival analysis and adjustment for re-infection showed that the 63-day cure rates  were 100% for AM and 99.5% (96.4\u201399.8%) for DP. The 63-day cure rates per protocol were 99% (97 of 98) for AM and 99.5% (196 of 197) for DP (P = 0.55). The difference (AM minus DP) in cure rates (95% CI) was \u22120.5% (\u22125.1 to 2.0%), which is within the 5% non-inferiority margin. The median fever and parasite clearance times were also similar for AM and DP. The proportion of patients with at least one recorded potential adverse event was significantly higher in the AM group  than in the DP group  . Dihydroartemisinin-piperaquine is not inferior to AM in the treatment of uncomplicated P. falciparum malaria in Laos and is associated with fewer adverse effects. The results of this study were similar to those of the larger multicentre study.We conducted an open, randomized clinical trial of oral dihydroartemisinin-piperaquine (DP) versus artesunate-mefloquine (AM) in 300 patients in Laos with uncomplicated  Plasmodium falciparum infection of 4.7\u201323.5/1,000 population.P. falciparum poses a public health threat. The Lao Government changed national policy for first-line antimalarial drug treatment of uncomplicated P. falciparum malaria to artemether-lumefantrine (AL) in 2005. This artemisinin-combination treatment (ACT) and artesunate-mefloquine both have high efficacies and good tolerability with 42-day failure rates of \u2264 6% in Laos.Malaria remains an important public health challenge in southern Laos with a median incidence of \u2122; Holleykin Pharmaceutical Co., Guangzhou, China) with artesunate  plus mefloquine  in southern Laos demonstrated that DP had a 100% cure rate (n = 110) assessed at 42 days-follow up.Dihydroartemisinin-piperaquine (DP) is a potential alternative; it can be taken once a day, does not have food-dependent bioavailability, and can be produced for a lower cost than AL. A trial comparing DP  in patients with acute, uncomplicated The study was conducted during June\u2013October in 2005 and 2006 at Phalanxay (10 beds) and Xepon (30 beds) District Clinics and during June\u2013October 2006 at Xepon District Clinic, Savannakhet Province. Phalanxay and Xepon districts (Savannakhet Province) are located 605 km and 665 km, respectively, southeast of Vientiane, the capital of Laos. Phalanxay  and Xepon  are inhabited predominantly by rice farmers of the Lao Theung ethnic group.12P. falciparum during follow-up.Patient inclusion and exclusion criteria have been reported.If the study inclusion criteria were met, the patients were randomly assigned to receive either 1) AM: artesunate , 4 mg/kg/day for 3 days (days 0\u20132) plus mefloquine , 15 mg base/kg on day 1 and 10 mg base/kg on day 2 or 2) DP: dihydroartemisinin-piperaquine (Sigma-Tau), 2.1/16.8 mg/kg in a single daily dose for 3 days. Each tablet contained 40 mg of dihydroartemisinin and 320 mg of piperaquine for adults and 20 mg of dihydroartemisinin and 160 mg of piperaquine for children < 15 years of age. The use of AM and DP in this study was reviewed and approved by the Food and Drug Department, Ministry of Health, Laos.The treatment choice was kept in a sealed opaque envelope, which was opened only after the decision to recruit had been made. An unequal randomization, 2:1 (DP:AM), was used to provide more precise estimates of DP cure rates and to provide more patients for the safety database of DP. Axillary temperature was measured every six hours. Study drug administration was observed directly by the study physicians. Study medications administered to older children (> 5 years of age) and adults were given as tablets or fractions of tablets orally with a glass of water and those given to young children (\u2264 5 years of age) were crushed, mixed with water, and administered as a slurry if the children were unable to swallow. Patients were observed for one hour to ensure that the medications were not vomited or regurgitated.12P. falciparum recurrence. Venous blood samples were obtained for hematologic and biochemical tests on days 28 and 63 if the results were abnormal on day 28 and were also performed on the day of P. falciparum recurrence.Patients were reviewed daily until parasite clearance was observed, then weekly for 63 days from the start of treatment, or at other times if he or she was ill. At each visit, finger prick blood was obtained for a malaria blood smear and hematocrit. Three blood spots were collected on filter paper from those with reappearance of asexual parasitemia for genotyping.P. falciparum parasitemia or treatment failure were withdrawn from the study, re-treated with oral artesunate (2 mg/kg/day) plus doxycycline (4 mg/kg/day)  for 7 days and followed-up. Those persons who received DP and had recurrent P. falciparum parasitemia or treatment failure were withdrawn from the study, re-treated with AM for three days, and followed-up. Patients in whom severe disease developedhttp://www.mdsps.com/).Patients who received AM and had recurrent Parasite counts in thick and thin blood films stained with Field's stain were obtained daily until parasite clearance and then weekly from day 7.15P. falciparum parasites.The primary end point was the PCR-corrected adequate clinical and parasitologic response PCR-ACPR at day 63 by using World Health Organization (2003) guidelines.P. falciparum parasites after checking \u2265 500 oil-immersion microscopic fields), 5) proportion of a febrile patients on days 1, 2, and 3, 6) fever clearance time (time in hours from the start of treatment at which the axillary temperature first decreased below 37.5\u00b0C and remained below 37.5\u00b0C for 48 hours), 7) gametocyte carriage (blood slide positive for gametocytes) after treatment, 8) fractional changes in hematocrit after antimalarial treatment, and 9) adverse events.Secondary end points were 1) crude or PCR-uncorrected adequate clinical and parasitologic response, 2) proportion of patients with treatment failure at or after day 7, 3) proportion of aparasitemic patients on days 1, 2, and 3, 4) parasite clearance time  and per-protocol (PP) populations.19Data were analyzed by using SPSS version 11.0 . Comparisons between two groups were made by using the Mann-Whitney U test, the Student's Gametocyte carriage was summarized as person-gametocytes-week rates calculated as the total number of weeks with gametocytes divided by the total number of weeks of follow-up and presented as per 1,000 person-weeks.The numbers of patients screened for malaria and enrolled in the trial is shown in P. falciparum malaria . She had one 5-minute convulsion 30 hours after receiving AM and remained unconscious (Glasgow Coma Score \u2265 9/15) for 70 minutes. She recovered after receiving supportive treatment. This patient was not excluded from the study and completed 63 days of follow-up and showed cure.Severe disease developed in a four-year-old girl who had uncomplicated P. falciparum reappearance in the AM and DP groups, respectively, PCR analysis indicated that only one patient, in the DP group, had a recrudescent infection. If we considered the four-year-old patient with the convulsion and coma after treatment as an early treatment failure, the 63-day cure rates per protocol, excluding patients who refused to swallow tablets or had persistent vomiting, or experienced re-infection, were 99% (97 of 98) for AM and 196 of 197 (99.5%) for DP. The difference (AM minus DP) in cure rates (95% CI) of \u22120.5% (\u22125.1 to 2.0%) is within the 5% non-inferiority margin. After censoring the patient who had convulsions, those experiencing re-infections and those who refused to swallow the study drugs or had repeated vomiting, the cure rates (95% CI) by survival analysis (Kaplan-Meier) were 100% for AM and 99.5% (96.4\u201399.8%) for DP (P = 0.43). Similarly, 28-day cure rates by intention-to-treat analysis were 99% (97 of 99) for AM and 97.5% (197 of 202) for DP. Intention-to-treat and per protocol analyses showed that the 63-day PCR-uncorrected cure rates for AM and DP were 89 of 98 (91%) versus 190 of 202 (94%) (P = 0.16), and 89 of 91 (91%) versus 185 of 197 (94%) (P = 0.33), respectively. The number of patients with P. vivax appearance in the AM and DP groups during follow-up was similar (Of 8 and 12 patients with subsequent ) for DP . If the  similar .P = 0.67), and all patients (except one in the DP group) had no parasites detected. The median fever and parasite clearance times were similar between the AM and DP groups  (P = 002). Mean leukocyte counts were significantly higher on day 28 than on day 0 in both treatment groups (P < 0.001).At presentation, 57% of all patients had fever, as defined by a body temperature \u2265 37.5\u00b0C. At day 2, 99% and 97.5% in AM and DP groups, respectively, were afebrile (P groups . Compari t-test) . The meaP = 0.04). A palpable spleen  and liver  were significantly more frequent in children than adults. The admission geometric mean parasitemia/\u03bcL was significantly higher in children than adults  versus 6,471 parasites/\u03bcL ) (P = 0.001), as were the proportions of patients with clinical anemia and vomiting at presentation . The frequency of the following admission symptoms and signs were significantly lower in children than adults: weakness , muscle pain , headache , nausea , pruritis , and tinnitus .The mean admission temperature was significantly higher in children (38.0\u00b0C [37.8\u201338.1\u00b0C]) than adults (37.7\u00b0C [37.4\u201337.9\u00b0C]) . Mean hematocrit values on admission and on all days between days 7 and 63 were significantly lower in children than in adults (P < 0.001) in both treatment groups.The median and range fever and parasite clearance times were not significantly different between children (23 [7\u201364] hours and 2 [1\u20133] days) and adults (21 [7\u201368] hours and 2 [1\u20132] days) (P = 0.48). The time to clearance of gametocytes was longer in the DP group than in the AM group. All patients in the AM group and only 8 (47%) patients in the DP group cleared their gametocytemias by day 7 . After treatment, gametocytemia developed on day 56 in one patient in the AM group who did not have gametocytemia on admission at the same time as a new P. falciparum infection. Gametocytemia also developed after admission (days 1\u20132), which cleared by day 14, in 6 other patients in the DP group who did not have gametocytemia on admission. The proportion of patients in whom gametocytemia developed in the AM and DP groups was not significantly different (P = 0.68). Person-gametocyte-weeks was estimated as 12 weeks/1,000 patients in the AM group and 22 weeks/1,000 patients in the DP group (P = 0.069). The proportion of patients with gametocytemia at any time point was higher in children  than in adults  , and the proportion of patients with gametocytemia after treatment was 4% (7 of 185) in children and 0% (0 of 92) in adults .Twenty-three (7.7%) patients had patent gametocytemias on admission, 6 (6%) in the AM group and 17 (8%) in the DP group (P > 0.05). The proportion of patients with at least one recorded potential side effect/adverse event after treatment was significantly higher in the AM group  than in the DP group  (P = 0.04). The incidence of post-treatment dizziness, nausea, insomnia, and anorexia were all significantly higher in AM group than in the DP group (P \u2264 0.01) (Adverse events were reported by 269 (89.7%) patients \u2265 3 years of age who were able to answer questions about these symptoms. A convulsion and coma developed in a four-year-old patient after treatment with AM (see above). Severe malaria or other life-threatening events after treatment did not develop in any other patients. The frequency of patients with symptoms and signs before treatment that may subsequently be confused with drug-related adverse events did not differ significantly between groups ( \u2264 0.01) . The freP < 0.001) than at admission (P \u2264 0.001): mean PR interval (95% CI) difference between baseline and days 2 and 7 were \u22129.5 (\u221213.1 to \u22126.0) and \u22125.7 (\u22129.1 to \u22122.4), respectively, for the AM group and \u22125.3 (\u22127.1 to \u22123.5) and \u22123.6 (\u22125.6 to \u22121.6), respectively, for the DP group. Four patients (two in the AM group and two in the DP group) had PR interval prolongation (> 200 ms) after treatment on day 2 or day 7 but the PR intervals on day 7 or day 28 were normal. The mean QRS interval was also significantly longer on day 2 than on admission (P = 0.001) but was similar to the baseline value by day 7 (P > 0.05) in both treatment groups  (P > 0.05) (P > 0.05) (Modeling the QT/RR relationship< 0.001) . The mea > 0.05) . The num > 0.05) ,24 were  > 0.05) .P. falciparum malaria in southern Laos and showed a 63-day follow-up cure rate of almost 100% for both drugs. Our previous study conducted in the same area, using GMP drugs from China, also demonstrated that the 42-day follow up cure rates were 99% and 100% in AM and DP groups, respectively.This study confirms that the fixed-dose, co-formulated ACT dihydroartemisinin-piperaquine is not inferior to the more expensive, not co-formulated AM in the treatment of uncomplicated P. falciparum malaria who were treated with DP, the 63 day PCR-corrected cure rate was 97.7%.53The 26 published clinical trials conducted in Asia, Africa, and South America that compared the efficacy of DP with other ACT regimens  show curAn important limitation of this trial for public health policy in Laos is that DP was compared with AM and not with AL, which is the current Lao national treatment policy. Artesunate-mefloquine was the comparator because this was required for the multicentre trial. In 2005, AM was the intended national treatment policy but sudden and unexpected unavailability of AM led the Government of Laos to change to AL. Artesunate-mefloquine is national policy in Cambodia, Myanmar (Burma), and Thailand, and DP is national policy in Vietnam and China. There have been six comparisons of DP versus AL, all in children from Africa, and the 28\u201363-day cure rates were not significantly different between AL (84\u201396%) and DP (93\u2013100%).30P. falciparum gametocyte prevalence (days 0\u201370) measured as person-gametocyte-weeks was significantly higher after DP than AM .In the multi-center study, overall Of 26 published clinical trials of either DP or DP plus trimethoprim  that inc12Although piperaquine has a chemical structure similar to chloroquine, which can cause lethal cardiologic effects,34P. falciparum malaria in pregnancy (PCR-adjusted cure rate at 63 days was 92%),In Laos, AL is provided by the Global Fund at a cost of < 1.4 U.S.$/adult treatment course and is free to patients through the public sector. When compared with AL, DP has potential benefits because it is less expensive to manufacture, may have better adherence, and can be used in a once a day regimen rather than a twice a day regimen. Although a study of the unsupervised effectiveness and supervised efficacy of DP and AM treatment in western Myanmar suggested good adherence to both regimens, there have been no studies directly comparing DP and AL adherence.P. falciparum malaria in Laos.In conclusion, DP demonstrated non-inferiority to the reference treatment of AM in Laos and in the overall three-country study. The high efficacy and relative safety of DP and its availability as a fixed combination and probable lower cost compared with other candidate ACTs suggests that it could play a significant role in the first-line treatment of uncomplicated"}
+{"text": "Plasmodium falciparum malaria, which has been reported in Nigeria. The objective of this multi-centre study was to evaluate the efficacy, safety and tolerability of the co-packaged formulation of artesunate and mefloquine in the treatment of uncomplicated malaria in two weight groups: those between 15 \u2013 29 kg and \u2265 30 kg respectively.The combination of artesunate and mefloquine has been reported to be effective against multi-drug resistant The trial was conducted in rural communities in the north-east, north-central, south-west and south-eastern parts of Nigeria. The WHO protocol for testing antimalarial drugs was followed. Outpatients having amongst other criteria, parasite density of \u22651,000 \u03bcl were enrolled. The co-packaged drugs were administered for 3 days at a dosage of artesunate, 4 mg/kg body wt/day and mefloquine, 25 mg/kg/body wt total) on days 0, 1 and 2. Patients were followed up for 28 days with the assessment of the parasitological parameters on days 1, 2, 3, 7, and 28.Four hundred and forty-six (446) patients were enrolled and 431 completed the study. Cure rates in both treatment groups was >90% at day 28. The mean parasite clearance times in treatment groups I and II were 40.1 and 42.4 hours respectively. The combination of artesunate and mefloquine showed good gametocidal activity, (gametocyte clearance time of 42.0 & 45.6 hours in treatment groups I and II respectively). There were no serious adverse events. Other adverse events observed were headache, dizziness, vomiting and abdominal discomfort. There was no significant derangement in the haematological and biochemical parameters.P. falciparum malaria in Nigeria.This co-packaged formulation of artesunate + mefloquine (Artequin\u2122) is highly efficacious, safe and well-tolerated. It is recommended for the treatment of uncomplicated  Malaria remains one of the greatest causes of morbidity and mortality in the world. Globally, there are between 300\u2013500 million cases of clinical malaria every year, with 85% of these from Africa . CurrentArtemisia annua) and the derivatives, namely, artemether, artesunate and dihydroartemisinin have now gained popularity as short acting drugs which could be used in combination with drugs which have longer half-life [Plasmodium falciparum [et al [The artemisinin drugs are developed from the Chinese wormwood  with longer acting mefloquine is expected to constitute a good ACT.The Drug Therapeutic Efficacy tests (DTET) conducted on two such combinations, namely, artesunate + amodiaquine and artemether + lumefantrine in 2004 showed adequate clinical and parasitological response (ACPR) of 94.6% and 96.8% respectively . The FedHowever, there is the challenge of availability and affordability of ACTs. To improve better access to ACTs at affordable prices, Roll Back Malaria partners in the pharmaceutical industries were encouraged to pre-package ACTs, which could be used if found effective, approved and duly registered by the regulatory authorities. One such combination drug is Artequin\u2122, a combination of artesunate and mefloquine, manufactured by MEPHA Ltd . Although this combination has been reported to be efficacious elsewhere, there is need to determine the efficacy, safety and tolerability of this ACT among Nigerians.This co-packaged formulation of artesunate and mefloquine has not been used before now in Nigeria. The outcome of therapeutic efficacy tests could be different in Nigeria, or even in different geographic zones of the country. It is, therefore, important to determine the efficacy, safety and tolerability of this co-packaged formulation of AM among Nigerian population. There is also the need to provide more options for malaria control in Nigeria.The objectives of the study were:in vivo test extended to 14 and 28 day follow-up period.\u2022 To evaluate therapeutic efficacy of a combination of artesunate plus mefloquine (AM) using the modified WHO seven-day P. falciparum malaria.\u2022 To determine the safety and tolerability of AM in the treatment of acute uncomplicated \u2022 To estimate gametocyte carriage and its reduction during treatment.This was a descriptive, open label, multi-centre, non-comparative trial of three-day regimen of a combination of AM for efficacy, safety and tolerability. Patients were stratified into two treatment groups according to their weights. Treatment group I consisted of those weighing between 15\u201329 kg, while treatment group II consisted of participants whose weight was \u2265 30 kg.This multi-centre study was conducted in four geographical zones of the country. In southwest Nigeria, 2 health facilities in Ijede, Lagos were used for the study. Ijede is a rural community in Ikorodu Local Government Area, Lagos State. The second site was in Borno, north eastern Nigeria, where a Specialist Hospital, a General Hospital and University Teaching Hospital were recruitment points. The third site was the Primary Health Centre in Ikot Ansa, Calabar, south eastern Nigeria. The fourth site was located in north central Nigeria where ECWA Evangel Hospital, Jos University Teaching Hospital and Plateau State Specialist Hospital were enrolment points. All sites were considered to be homogenous and high malaria transmission areas, hence their suitability for trials of this nature.P. falciparum parasitaemia in the range of 1,000 to 250,000 asexual parasites per \u03bcl of blood, presence of axillary temperature \u226537.5\u00b0C and/or history of fever in the preceding 24 hours, informed consent by parent/guardian (in the case of children), ability to come for the stipulated follow-up visits, and easy access to the health facility.Patients between 15\u201329 kg or \u2265 30 kg with mono-infection with a Patients with danger signs such as: unable to drink or breastfeed, unable to sit or stand up, vomiting everything, recent history of convulsion, lethargic or unconsciousness were excluded. Others excluded were those with signs of complicated falciparum malaria, such as severe anaemia (PCV \u2264 15%), shock, bleeding disorders, coke colored urine, jaundice, presence of severe malnutrition by clinical examination and weight for height measurement, history of allergy to study drugs and pregnant women.Patients who met the enrolment criteria were recruited. Written informed consent was obtained prior to enrolment. Day 0 was the day of screening, clinical assessment, initial malaria smears, haematological and biochemical assessments and enrolment. Temperature was measured in the axilla using digital electronic thermometer. Venous blood was collected from enrolled patients for baseline laboratory indices. For biochemistry, liver enzymes , total and conjugated bilirubin and serum creatinine were done. Haematological parameters such as haemoglobin, white blood cells (WBC), and erythrocyte sedimentation rate were also investigated. The patients were allocated to either treatment group I (15 to 29 kg) or treatment group II (\u2265 30 kg) and given the 3-day co-packaged drug at a dosage of artesunate  and mefloquine .The drugs were administered under supervision and patients were observed for 60 minutes. If vomiting occurred within 30 minutes of drug administration, the full dose was repeated. However, if it occurred 30\u201360 minutes, half the dosage was given again. Participants who vomited a second time were excluded from the study and referred for treatment with appropriate parenteral antimalarial regimen. Use of concomitant medications (including acetaminophen) were documented in the Case Report Form (CRF).The patients returned on days 1 and 2 to complete the drug administration and for clinical assessment. They were also given appointment for days 3, 7, 14 and 28 for clinical examination and blood smears. They were also asked to return to the clinic on any other day should they have new complaints, or any change in their condition. Patients that failed to report at the health centre for the scheduled visit were followed to their residence by trial field workers.Serious adverse events, loss of patient to follow-up, consent withdrawal or withdrawal as a result of treatment failure, were criteria for discontinuation of treatment.Primary treatment efficacy was determined based on parastological cure rates on days 2, 3, 7, and 28 and by the times to parasite and fever clearance and from the proportion of patients without gametocytes. The other outcomes assessed were early treatment failure (ETF), late clinical failure (LCF) and late parasitological failure (LPF). Recrudescence denoted clinical recurrence of malaria after the initial clearance of parasite from circulation. Parasite reappearance after day was interpreted as either true recrudescence or a new infection. Thus, treatment efficacy for cure rates in our context were described as uncorrected since no DNA polymerase chain reaction (PCR) analysis was performed in any of the four sites.All adverse events were monitored and recorded on the case report forms (CRFs). Haematological parameters, liver enzymes and creatinine were assessed for the purpose of detecting abnormal laboratory features that constitute adverse events. Efforts were made to assess patients that dropped out from the study for the 28 days of active follow up period for safety reasons.The sample size used for this drug trial was calculated from the table of anticipated proportion (WHO/HTM/RBM/2003) at 95% confidence level and 10% precision. Calculation was based on estimated cure rate for current artemisinin-based antimalarial drug treatment [2 or student t-test or analysis of variance (ANOVA) as appropriate. Pearson's correlation test was used to examine the relationship between selected variables.Data generated from this trial were entered into EPI-INFO version 6.04. Microsoft excel software was also used to plot simple graphs. Various aspects of the data were subsequently analysed using SPSS statistical package version 11. Descriptive statistics were produced for different parameters before figures representing various observations were compared using XAt screening, thick and thin blood films were collected. Thick film was examined with a binocular microscope with an oil immersion objective lens to quantify the parasitaemia. Parasitaemia was measured by counting the number of asexual parasites against a number of leukocytes in the thick blood film, based on a putative count of 8,000 leukocytes per microlitre of blood or an adequate mean WBC in the population under investigation. The number of asexual parasites was counted against 200 leukocytes using a hand tally counter. The number of parasites per microlitre of blood was calculated by using the formula:P. falciparum gametocytes were seen, a gametocyte count was performed against 1000 leukocytes (WHO/MAL/82.988).If Written approval was obtained by the Ethics Review Boards (IRBs) of the various institutions for the study. The Heads of communities and authorities of the various health facilities also consented to the conduct of the study. The study was carried out in accordance with the principles laid down by the World Health Assembly of 1975 on Ethics in Human experimentation and the Helsinki Declaration. The study adhered to Good Clinical Practices (GCP) and conformed to the TDR Standard Operating Procedures.Participants were informed of the aims, methods, anticipated benefits and potential hazards of the study. Written informed consent was obtained from each participant and/or parent/guardian of patients participating in the study. Subject were informed that they could withdraw consent to participate anytime without any consequence to them. This was written in English and the local dialects of the various communities.A total of 4,139 patients who had not taken any antimalarial medication in the previous seven days were screened in the four sites. Overall, 446 patients fulfilled the criteria for enrolment and were enrolled, but a total of 431 (96.6%) patients consisting of 203 in treatment group I (weight \u2265 30 kg) and 228 in treatment group II (weight 15 to 29 kg) completed the study. The demographic and clinical characteristics of the patients are shown in Table A total of 15 patients defaulted as a result of withdrawal/loss to follow-up and/or protocol violation. The trial profile is shown in Figure Results showed that on D1, 83(40.9%) of the 203 enrolled treatment group I (TG I) and 72 (31.6%) of the 228 enrolled treatment group II (TG II) no longer had any malaria parasites in their blood. The mean parasite densities in the remaining 120 in TG I and 156 in TG II were drastically reduced. The clearance rate dramatically increased to 92.1% and 91.7% for TG I and TG II respectively on D2 until total clearance was achieved in the remaining 16 in TG I and 19 in TG II on D3.-1 blood. It was not possible to ascertain whether this was recrudescence or of new infection. It was not possible to confirm recrudescence with polymerase chain reaction.In treatment group I, the geometric mean parasite density of the 203 participants enrolled on D0 was 6,890.8 parasites/\u03bcL of blood, which decreased to 30.74 parasites/\u03bcL on D1, giving a percentage rate of 99.6%. The rates on other days (i.e. using the geometric mean parasite densities) were as follows: D2 (99.98%), D3 (100%), D7 (100%) and D14 (100%). However, on day 28, one of those in TG I (in Lagos site) manifested low-grade parasite density of 360 parasite \u03bcLParasite clearance time (PCT) in 203 participants in TG I was determined from the spread sheet data (WHO/MAL/82.988). Parasitaemia completely cleared in 83 patients within 24 hrs, 104 cleared in 48 hrs while 16 patients were cleared in 72 hours. Time to parasite clearance was calculated as follows:In treatment group II, the geometric mean parasite density of the 228 enrolled patients on day 0 was 11,727.5 parasites/\u03bc of blood which reduced to 54.2 parasites on D1, giving a percentage rate of 99.2% on D1 , D3 (100%), D7 (100%) and D14 (100%). As in TG I, six patients in TG II in three sites manifested low-grade parasitaemia on day 28. Time to parasite clearance (PCT) in 228 patients in TG II was also determined from the spread sheet. Here, parasitaemia had cleared in 72 patients within 24 hrs (D1), 137 participants within 48 hrs (D 2) and in the remaining 19 within 72 hrs (D3).Time to parasite clearance was calculated as follows:Result showed that 109 of the 203 enrolled patients in TGI had temperatures below 37.5\u00b0C. The mean temperature of the 94 in TG I patients with temperatures \u2265 37.5\u00b0C was 38.44 \u00b1 0.75\u00b0C. The temperature dropped to a mean value of 36.4 \u00b1 0.67\u00b0C in 24 hrs. The mean temperature of 132 patients in TG II who were febrile (T \u2265 37.5\u00b0C) was 38.56 \u00b1 0.79\u00b0C. The temperature of these patients dropped to 36.57 \u00b1 0.81\u00b0C by D1 (24 hrs). The fact that many patients were afebrile pre-treatment made it inappropriate to calculate fever clearance time.Three of the four sites investigated the gametocyte carriage rate and the changes in geometric mean gametocyte densities (GMGD) in the patients. The gametocyte carriage rate for TG I and TG II were 5.9% and 4.4% respectively. In ten patients where gametocytes were found, the mean gametocyte count dropped from pre-treatment levels on day 1. The gametocytes cleared in four patients in 24 hours, three were cleared in 48 hours and three in 72 hours. Time to gametocyte clearance in TG II was calculated (as in parasite clearance time) to be 45.6 hours.In twelve TG I patients where gametocytes were found, the gametocytes cleared in five patients in 24 hours. It cleared in four patients in 48 hours and in the remaining three patients in 72 hours. The gametocyte clearance time in TG I was 42.0 hours.No serious adverse event (SAE) was reported during the study in all the four trial sites. Many adverse events (AE) of the antimalaria drugs were most likely related to the underlying malaria disease. Of the 39 reports, 10 patients  reported vomiting. The others were as follows: headache , dizziness  and abdominal discomfort . There was one report (0.2%) each of sleeplessness, fast breathing, weakness and back pain . Other haematological parameters, viz, lymphocytes, neutrophils, monocytes and eosinophils also had varying values which were not statistically significant (P > 0.05) from the D0 values Table .-l in TG I and 5.12 \u00b1 5.63 \u03bcmol-l in TG II. The level remained within this value from D0 to D28. There were also no significant decreases or increases (P > 0.05) in the values of serum alanine aminotransferase , serum aspartate amonitransferase  and alkaline phosphotase (ALK).The Biochemical values on D0 are also shown in Table The therapeutic efficacy study of co-packaged three day treatment preparation of artesunate and mefloquine in four different geographic areas of Nigeria showed that the combination is both effective and well tolerated in the treatment of acute uncomplicated falciparum malaria.nd day. This rapid clinical and parasitological response confirmed the previous findings of others [The geometric mean parasite density was drastically reduced in both treatment groups within 24 hours after treatment and completely cleared by day 3 in the remaining patients who completed the study. The parasitological response in both treatment groups were 100% on D3, D7, and D14 but 97.5% on day 28. The rapid clinical response was shown by a drop in temperature to normal values (viz. below 37.5\u00b0C) on the 2f others -19,21,22P. falciparum, AM therapy was also beneficial in inducing significant reduction in gametocyte rate and density. The data suggest that AM ultimately cleared gametocytes from peripheral blood. This shows that AM exhibits a gametocidal effect. The second observation was that patients with mixed infections of P. falciparum and Plasmodium malariae were cured parasitologically and clinically.Apart from the rapid clearance of asexual forms of et al [nd and 4th week post-therapy, attributed this phenomenon to new infections. There is a need to perform more molecular genotyping of the parasite strains in subsequent trials to confirm this observation.It was observed in the course of this trial, that parasitaemia appeared on day 28 in one patient of the TG I and six patients of TG II. Parasitaemia was not associated with fever or other symptoms of malaria. Since analysis of the parasite genotypes was not performed, it was not possible to determine whether these were new infections or recrudescence. It is possible that the observed day 28 parasitaemia could be re-infections rather than recrudescence. Falade et al , by genoBased on the experience of this study, AM is safe and well-tolerated. The laboratory values were not significantly different pre-and post-treatment. The marginal variations in liver function test results may be related to stabilization of the liver following successful treatment. The same result was observed with mean haemoglobin values which returned to normal after recovery. The slight reduction in mean platelet count (data not presented) was consistent with the reported findings of relative thrombocytopenia in 50 to 75% of patients with acute malaria .P. falciparum malaria in four sites in Nigeria. It also exhibited significant gametocidal activity. As observed by other workers, the rapid parasitological response corresponded to the fast clinical response.In conclusion, the results of this study have confirmed the efficacy of AM in the treatment of The authors declare that they have no competing interests.PUA, MMM, and IMW conceived the idea, wrote the protocol and supervised data collection and writing of this paper. PUA played a central role in coordinating the work of the 4 centres and is the contact author for this article. SO, FAO, IJO, VIE and OOA wrote and reviewed the paper before it was finally sent. VIE was a focal person for supply of the drugs. All authors made scientific contributions to this article."}
+{"text": "A recent study has shown that treatment of visceral leishmaniasis (VL) with the standard dose of 15 mg/kg/day of paromomycin sulphate (PM) for 21 days was not efficacious in patients in Sudan. We therefore decided to test the efficacy of paramomycin for a longer treatment duration (15 mg/kg/day for 28 days) and at the higher dose of 20 mg/kg/day for 21 days.This randomized, open-label, dose-finding, phase II study assessed the two above high-dose PM treatment regimens. Patients with clinical features and positive bone-marrow aspirates for VL were enrolled. All patients received their assigned courses of PM intramuscularly and adverse events were monitored. Parasite clearance in bone-marrow aspirates was tested by microscopy at end of treatment , 3 months  and 6 months after EOT (secondary efficacy endpoint). Pharmacokinetic data were obtained from a subset of patients weighing over 30 kg.42 patients (21 per group) aged between 4 and 60 years were enrolled. At EOT, 85% of patients  in the 20 mg/kg/day group and 90% of patients (95% CI: 69.6% to 98.8%) in the 15 mg/kg/day group had parasite clearance. Six months after treatment, efficacy was 80.0% (95% CI: 56.3% to 94.3%) and 81.0% (95% CI: 58.1% to 94.6%) in the 20 mg/kg/day and 15 mg/kg/day groups, respectively. There were no serious adverse events. Pharmacokinetic profiles suggested a difference between the two doses, although numbers of patients recruited were too few to make it significant .Data suggest that both high dose regimens were more efficacious than the standard 15 mg/kg/day PM for 21 days and could be further evaluated in phase III studies in East Africa.NCT00255567ClinicalTrials.gov  Visceral leishmaniasis (VL) is a parasitic disease transmitted through the bite of sandflies. The WHO estimates 500,000 new cases of VL each year, with more than 90% of cases occurring in Southeast Asia, East Africa, and South America. If left untreated, VL can be fatal. We had previously conducted a large multi-center study in Sudan, East Africa, to assess the efficacy of paromomycin (PM) alone or in combination with sodium stibogluconate. Clinical studies in India have shown that 15 mg/kg/day PM for 21 days was an effective cure. However, the same treatment regimen was not efficacious in two study sites in Sudan. Here, our aim was to assess two high-dose regimens of PM in Sudan: 15 mg/kg/day for 28 days and 20 mg/kg/day for 21 days. The results suggest that, at these total doses, PM is more efficacious than when given daily at 15 mg/kg for 21 days, and that high doses are required to treat VL in Sudan. Efficacy of 20 mg/kg/day PM for 21 days is currently being evaluated in a prospective, comparative phase III trial in East Africa. According to the WHO estimates, visceral leishmaniasis (VL) is a parasitic disease that affects more than 500,000 people globally each year Although drugs  currently exist to treat this parasitic infection, their use has been limited because of high cost, toxicity, or development of parasite resistance In an effort to identify an effective treatment for VL in East Africa, we had previously initiated a multi-center phase III study in Sudan, Ethiopia, and Kenya comparing the efficacy of PM alone at the dose shown to be efficacious in India (15 mg/kg/day for 21 days) against SSG alone (20 mg/kg/day for 30 days) and against a combination treatment of SSG and PM . PM monotherapy did not show adequate efficacy, particularly in Sudan where parasite clearance was below 50% in patients at 6 months after end of treatment (EOT), and the study had to be prematurely stopped In the current study, we sought to find an efficacious dose of PM for the treatment of VL in Sudan and to explore possible reasons for the failure of the drug at the previous dose studied of 15 mg/kg/day for 21 days. In our previous study using this dose, conducted in 5 sites in Ethiopia, Kenya and Sudan, we found an overall end of treatment cure of 67.4% and 6-month post-treatment cure of 63.8% Therefore a total dose increase of 33% was attempted through two possible regimens- an increased dose of 20 mg/kg for 21 days or a prolonged course of 15 mg/kg for 28 days. The former regimen has been evaluated in some clinical trials in India The main objective was to assess the efficacy of two dosing regimens of PM monotherapy for the treatment of VL: 20 mg/kg/day for 21 days and 15 mg/kg/day for 28 days. Secondary objectives were to assess the safety of PM and compare the pharmacokinetic (PK) profiles of the two groups in a subset of patients.Patients with clinical symptoms and signs suggestive of VL and confirmed by visualization of parasites in bone-marrow aspirates were eligible for enrollment according to the National VL guidelines for Sudan for treatment and control. To be included in the study, patients had to: be between 4 and 60 years of age; be able to comply with the protocol  and S3; 3/mm3; platelets less than 40,000/mm3; liver function test values more than three times the normal range; and serum creatinine values outside the normal range for age and gender.Patients were excluded from the study if they: had negative bone-marrow smears; were clinically contraindicated to having a bone-marrow aspirate; received any anti-leishmania drug in the past 6 months; had severe protein or caloric malnutrition (Kwashiorkor or marasmus); had previous hypersensitivity reaction to aminoglycosides; suffered from a concomitant severe infection, ie tuberculosis, HIV, or any other serious underlying disease ; suffered from other conditions associated with splenomegaly such as schistosomiasis; had previous history of cardiac arrhythmia or an abnormal electrocardiogram (ECG); were pregnant or lactating; or had pre-existing clinical hearing loss. If tuberculosis or schistosomiasis were suspected, these were screened through laboratory testing. Additionally, patients with the following laboratory values were excluded: hemoglobin less than 5 g/dL; white blood cell less than 10This was a two-arm, randomized, open-label, dose-finding study done at a single site in Sudan . This site participated in the previous study conducted on PM Parasitology slides were prepared from bone-marrow aspirates, read, and reported according to a standardized, approved WHO method For the PK analysis, the first six consenting patients weighing 30 kg or more were selected from each treatment group and had additional venous blood and urine samples on day 1 and day 14 in the 20 mg/kg/day group, and on day 1 and day 26 in the 15 mg/kg/day group. The timing for blood sampling was 0 (before treatment) and at 0.25, 0.5, 1, 2, 4, 6, 8, 12, and 24 hours after administration of the drug, and at 0\u20132, 2\u20134, 4\u20136, 6\u20138, 8\u201312, and 12\u201324 hours after administration of the drug for urine sampling.The trial was done in accordance with the Declaration of Helsinki (2002 version) for the conduct of research on human subjects and followed the International Committee for Harmonization guidelines for the conduct of clinical trials. All trial site personnel received relevant training in Good Clinical Practices.The Ethics Committee of the Institute of Endemic Diseases, University of Khartoum, and the Directorate of Health Research, Federal Ministry of Health, Sudan approved the study protocol , which was submitted as a protocol amendment  to our pAll participants or their parents or legal guardians gave their written informed consent before entry into the trial. Children were included in this study because they represent more than 50% of VL cases in this endemic area, and were included in the PK sampling if they met the weight criteria (>30 kg). This study was registered at ClinicalTrials.gov (registration number NCT00255567).The study medication was 1 g/2mL paromomycin sulphate . Doses in the study groups were 20 mg/kg/day paromomycin sulphate  and 15 mg/kg/day paromomycin sulphate . The rescue medication was AmBisome , which was reconstituted according to the manufacturer's instructions for a dosage of 3 mg/kg/day for 10 days.The primary efficacy endpoint was cure (parasite clearance) at EOT . The secondary efficacy endpoint was parasite clearance at 6 months follow-up.Efficacy was reported as the number and percentage of randomized patients in whom no parasites were detected by arm at each endpoint. Rescue medication before an endpoint indicates treatment failure at that endpoint. Clinical and biological parameters, such as temperature, weight, spleen size and hemoglobin, were used by clinicians to decide each patient's response to treatment and whether rescue medication was indicated. Relapse at 3-month follow-up was defined as patients with no parasites detected at EOT but in whom parasites were seen at 3-month follow-up. Slow response to treatment was defined as the presence of parasites at EOT but not at 6 months in the absence of rescue medication. Therefore, a treatment failure at initial test of cure (EOT) that did not receive rescue medication was defined as a slow responder so long as there was parasite clearance at 6-month follow-up.Safety analyses were based on the number of adverse events (AEs) that occurred during the study, including changes of pre-defined magnitudes in audiometric measurements .Analyses were carried out using Stata, version 10 . Analyses were done on the intention-to-treat (ITT) population. In case of missing data, efficacy analyses were by complete-case analysis, excluding patients with missing data, and by worst-case analysis, where missing efficacy data are assumed to be treatment failures. To account for different treatment durations and avoid overestimating harm in the longer treatment duration group, the treatment-emergent adverse event (TEAE) rate was calculated, per group, as the total number of adverse events divided by the total person-time at risk 104 patients with suspected VL were screened for entry into this study. Of these, 42 patients were enrolled in the study  . All trePM was well tolerated in this study. 48 AEs were reported in total; 20 in the 20 mg/kg for 21 days group and 28 in the 15 mg/kg for 28 days group , and nonAlthough six patients from each group should have taken part in the PK study, only data from three patients in the 20 mg/kg/day PM group and six in the 15 mg/kg/day PM group were obtained. Only one patient was a child (age of 12 years and weight of 39 kg in the 15 mg/kg group). The others were aged between 17 and 28 years.Mean plasma PM concentrations at the earlier time points were similar between the two treatment groups . NeverthTo date, the standard treatment for VL in East Africa still consists of antimonials. This study is part of the first large-scale multi-centre clinical trial to assess the efficacy of PM for the treatment of VL for the East African region. The initial study Although efficacy is normally assessed as parasite clearance at 6 months in trials for VL, in this study we chose to use parasite clearance at EOT as the primary endpoint because a chance of loss to follow-up of just a few patients would significantly affect the result. In addition to the small sample size, another potential limitation is the use of bone-marrow aspiration for diagnosis and test of cure. However, spleen aspiration remains contraindicated in rural hospitals in Sudan, making bone marrow the best viable alternative.At 6 months after treatment, efficacy was 80.0% (95% CI: 56.3% to 94.3%) and 81.0% (95% CI: 58.1% to 94.6%) in the 20 mg/kg/day and 15 mg/kg/day groups, respectively, compared with less than 50% (in Sudan) at 6 months observed in the previous study Serious safety issues that would limit the evaluation of PM at high doses were not identified in this study. Otoxicity, which has been seen as a transient side-effect of PM in other studies PK analyses showed that peak plasma PM concentration occurred 1\u20132 hours after administration and suggest that, at the high daily dose of 20 mg/kg, elevated plasma PM concentrations may be maintained for a longer period of time (up to 8 hours). Unpublished data  of PM administration (15 mg/kg) to healthy Sudanese volunteers showed peak PM plasma concentrations similar to those in American volunteers who received a similar dose Even though interpretation of our results is limited because of the small sample size, we identified what seems to be a more efficacious dose of PM than the one previously used in Sudan Checklist S1CONSORT checklist.(0.19 MB DOC)Click here for additional data file.Protocol S1Dose-ranging CSR.(0.56 MB PDF)Click here for additional data file.Protocol S2Clinical trial protocol.(0.23 MB PDF)Click here for additional data file.Protocol S3Protocol amendment.(0.06 MB PDF)Click here for additional data file."}
+{"text": "There is renewed acknowledgement that targeting gametocytes is essential for malaria control and elimination efforts. Simple mathematical models were fitted to data from clinical trials in order to determine the mean gametocyte circulation time and duration of gametocyte carriage in treated malaria patients.Data were used from clinical trials from East Africa. The first trial compared non-artemisinin combination therapy (non-ACT: sulphadoxine-pyrimethamine (SP) plus amodiaquine) and artemisinin-based combination therapy (ACT: SP plus artesunate (AS) or artemether-lumefantrine). The second trial compared ACT (SP+AS) with ACT in combination with a single dose of primaquine (ACT-PQ: SP+AS+PQ). Mature gametocytes were quantified in peripheral blood samples by nucleic acid sequence based amplification. A simple deterministic compartmental model was fitted to gametocyte densities to estimate the circulation time per gametocyte; a similar model was fitted to gametocyte prevalences to estimate the duration of gametocyte carriage after efficacious treatment.The mean circulation time of gametocytes was 4.6-6.5 days. After non-ACT treatment, patients were estimated to carry gametocytes for an average of 55 days (95% CI 28.7 - 107.7). ACT reduced the duration of gametocyte carriage fourfold to 13.4 days (95% CI 10.2-17.5). Addition of PQ to ACT resulted in a further fourfold reduction of the duration of gametocyte carriage.These findings confirm previous estimates of the circulation time of gametocytes, but indicate a much longer duration of (low density) gametocyte carriage after apparently successful clearance of asexual parasites. ACT shortened the period of gametocyte carriage considerably, and had the most pronounced effect on mature gametocytes when combined with PQ. The transmission of malaria depends on the presence of mature sexual stage parasites, gametocytes, in the human peripheral blood. Once ingested by a mosquito taking a blood meal, gametocytes develop through different mosquito-specific stages that ultimately render the mosquito infectious to humans. There is renewed acknowledgement that targeting gametocytes, either alone or as part of integrated control programmes, is essential for malaria control and elimination efforts -4.Plasmodium falciparum gametocytes are relatively insensitive to many anti-malarials [alarials  and circalarials ,7. The malarials , but alsalarials .These mean values do not reflect the broad range in the time that a single person carries gametocytes. Following efficacious clearance of asexual parasites, the source of gametocyte production, the duration of gametocyte carriage is determined by the maximum duration of gametocyte sequestration and the maximum circulation time following their release into the bloodstream. Maximum gametocyte sequestration time was previously estimated at 12 days; maximum circulation time at 22.2 days . As a coin vitro studies observed additional activity of artemisinins against mature gametocytes [In vivo studies commonly observe that mature gametocytes can persist after ACT treatment and result in post-treatment malaria transmission [Understanding the duration of gametocyte carriage is particularly important for determining the role of gametocytocidal anti-malarial drugs in the elimination of malaria . The impetocytes ,21 althoetocytes . In vivosmission ,22-24. Psmission ,11,19,25smission ,26.Here, by fitting simple mathematical models to data from two field trials which used sensitive molecular gametocyte detection methods, the mean gametocyte circulation time was determined and the full duration of gametocyte carriage was estimated in patients treated with non-ACT and gametocytocidal ACT and ACT-PQ drug combinations.P. falciparum infection at a density of 500 - 100,000 parasites/\u03bcL and no microscopical evidence of co-infection with other Plasmodium species. Two microscopists each read 100 high power microscopic fields for asexual parasites. The trial in Kenya included children aged 0.5-10 years and excluded children with an Hb < 5 g/dL; the trial in Tanzania included children aged 3-15 years and excluded children with an Hb < 8 g/dL. Parents or guardians provided written informed consent for enrolment and the trial protocols received ethical clearance in Kenya , Tanzania  and the UK .Data were used from two clinical trials. Characteristics of the studies are summarized in table Pfs25 mRNA that is expressed in mature gametocyte (stage V) gametocytes [Gametocyte carriage was determined on day 0, 3, 7, 14, 28 after treatment; treatment (d0-2) was completed 24 hours before the second time-point for gametocyte detection. For the trial in Tanzania, gametocyte carriage was also determined on day 42. In both studies, gametocyte prevalence and density were determined by examining 100 microscopic fields specifically for gametocytes and by quantitative nucleic acid sequence based amplification (QT-NASBA). The latter method is based on the detection of etocytes  and has etocytes . Enrolmeetocytes ,16.Pfs25 QT-NASBA at any time between day 0 and day 28 after enrolment, ii) were successfully cured of their asexual parasites  and had no detectable re-infection. SP-treated children experienced a very high treatment failure rate by microscopy (56%). Additional low level SP-treatment failures may have been missed by day 28 [This study aimed to determine the mean circulation time of gametocytes and the impact of ACT and PQ on existing gametocyte carriage. Therefore, individuals were selected who i) had gametocytes by y day 28  and the y day 28 . The SP y day 28 ,35 but ny day 28 ; iii) SPy day 28  and, they day 28  and PQ [y day 28 ,38 may hy day 28 . The iniet al [A simple deterministic compartmental model describing the gametocyte density per \u03bcL blood over time Figure  was fittet al . Both moG changes over time depending on the rate of release of gametocytes into the bloodstream \u03c1 from a sequestered gametocyte population S, and on the decay rate \u03bc of the circulating gametocytes. Exponentially distributed sequestration and decay times were assumed, so that 1/\u03c1 gives the mean sequestration time and 1/\u03bc is the mean circulation time per gametocyte. The differential equations describing this process are given by:In the density model, the gametocyte concentration in the bloodstream 0 Sis the density of the sequestered gametocyte population on day 0, and 0 Gis the density of circulating gametocytes on day 0:This is solved to obtain an expression for the expected gametocyte density in circulation at time t since treatment began, G(t), where The model was fitted to the log gametocyte densities using maximum likelihood methods, assuming a Normal distribution for the log transformed data and incorporating Normally-distributed random effects for each patient in overall density. Zero density values in the data were treated as values below the detection limit of the assays (0.02 gametocytes/\u03bcL for the QT-NASBA method) and the likelihood was specified to account for these censored observations. Each trial was fitted separately as there was a significant difference in initial log gametocyte density between trials and therefore other factors may also have differed between the two study populations. Gametocyte circulation time was allowed to vary by treatment type, since both ACT and PQ may have differing effects. The initial size of the sequestered gametocyte population was an uncertain parameter. As well as estimating this parameter from the model fitting, a sensitivity analysis was undertaken for a range of fixed values. Because ACT is likely to clear immature gametocytes before they can be released into sequestration, we fixed the sequestered gametocyte population to be 3 log lower than for the non-ACT treatment Table . The dur0I, the rate f at which initially gametocyte-negative proportion E move to the gametocyte-positive state I, and the rate r at which infectious individuals recover to become gametocyte negative. The equations describing this process are given by:The gametocyte prevalence model describes the appearance and disappearance of any circulating gametocytes according to the prevalence at day 0, t days after treatment, I(t), as:This can be solved to obtain an expression for the expected prevalence of gametocytes in the patients f, was estimated from the data, and also varied in sensitivity analysis [f and 1/r, respectively. To obtain the predicted marginal probabilities of gametocyte positivity at each time point for each treatment group from estimated parameters, 10,000 values were simulated from the random effects distribution using the estimated variance from the model, added to the logit transformed model prediction, then transformed back to the prevalence scale.The model was fitted to the individual patient prevalence data using maximum likelihood methods assuming a Binomial outcome distribution and random patient effects Normally distributed on the logit scale. The rate of loss of gametocytaemia, r, was allowed to vary by treatment type, while the other parameters were assumed to be constant between groups. The rate of becoming gametocyte positive, analysis . The timAll model fitting was carried out using the PROC NLMIXED procedure in SAS .The numbers and characteristics of individuals included in the analysis are summarised in table Pfs25 QT-NASBA gametocyte density data. The data indicated an approximately linear decrease in log-gametocyte density between day 3 and day 28  between day 0 and day 3. In the Tanzanian trial, the median reduction in gametocyte density between day 0 and day 3 after initiation of ACT treatment was 67.7% . There was no statistically significant change in gametocyte density in the first three days after initiation of efficacious non-ACT treatment (p = 0.10). The ACT-PQ combination had a stronger and longer impact on gametocyte carriage than ACT alone. The gametocyte density decreased very sharply between day 0 and day 3  and, rather than a gradual decline, continued to decrease sharply afterwards. Between day 3 and day 7, gametocytes were cleared for 76.9% (20/26) of the children who still had gametocytes by day 3. Only 4.9% (2/41) of all gametocytaemic children at enrolment still had detectable levels of gametocytes by day 14 after ACT-PQ treatment. Between day 14 and day 28, one of these two individuals remained gametocytaemic. Four others became gametocytaemic after being gametocyte free on day 3, 7 and 14 (n = 3) or on day 7 and 14 (n = 1), suggesting that their gametocytes were produced by a newly acquired infection or parasites that emerged from the liver or recrudesced after treatment was initiated.The variation between study drugs and trials was reflected in the estimated duration of gametocyte carriage after treatment Table . It was P. falciparum gametocytes were revisited using sensitive molecular gametocyte detection methods. The mean circulation time of gametocytes was 4.6-6.5 days, depending on the study site. Gametocyte carriage persisted for an estimated average of 55 days after the initiation of treatment that resulted in the apparent clearance of asexual parasites. ACT shortened the period of gametocyte carriage considerably, particularly when combined with PQ, a regimen that had the most pronounced effect on mature gametocytes.In the current study, previous estimates of the circulation time of Findings on circulation time of gametocytes confirm previous estimates that used microscopy for gametocyte detection. Although the molecular gametocyte detection method that was used was considerably more sensitive for detecting low-density gametocyte carriage ,6, the rPfs25 mRNA-based detection method only detects mature stage V gametocytes [P. falciparum parasites, it is possible to estimate the circulating and sequestered number of parasites by measuring plasma concentration of histidine-rich protein (PfHRP2) [P. falciparum gametocytes.While the non-ACT combination had no activity against mature gametocytes , both ACetocytes . Assuminetocytes , iii) thetocytes  and v) tetocytes ,22. Whiletocytes  or solel(PfHRP2) . A similAddition of PQ to ACT was found to strongly increase the clearance of remaining gametocytes . A recenPfs25 QT-NASBA. Undetected low density infections could have boosted gametocyte production [Pfs25 QT-NASBA gametocyte densities were higher in the Tanzanian trial, which may explain the observed difference, given that ACT is thought to impact less on mature gametocytes. Increased clonal complexity [The presented analysis has several limitations in addition to the fact it is based on only two studies. The confounding potential of re-infections was minimised by limiting the observation period to a time when re-infection was deemed unlikely. However, it is possible that some low-density re-infections or persisting parasitaemia occurred, as was suggested by the increase in gametocyte prevalence in the trial in Tanzania after gametocytes were cleared from all but one individual. There is currently no molecular detection tool that specifically detects asexual parasites with the same sensitivity as the gametocyte oduction  or resuloduction  and resuoduction  and the mplexity  and younmplexity  have botmplexity , but lowmplexity , where tThe current analysis indicates that gametocytes persist for an average of more than one month after clearance of asexual parasites. Artemisinins can shorten the duration of gametocyte carriage approximately fourfold. PQ is a more potent gametocytocidal drug than artemisinins and in led to a fourfold reduction in the duration of gametocyte carriage compared to ACT alone.The authors declare that they have no competing interests.TB and LO analysed the data and wrote the manuscript; TB, SS, SO, PS and RS were responsible for the original study design and data collection; JTG and ACG contributed in data analysis; CS, RS, ACG and CD contributed to data interpretation and manuscript preparation. All authors read and approved the final manuscript.0)Circulation time of gametocytes: sensitivity of model fits to assumed size of sequestered gametocyte population at day 0 (S. This table contains the outcomes of a sensitivity analysis where different fixed values of the sequestered gametocyte population were fitted. The impact on the estimated circulation time of gametocytes is presented for each trial and treatment arm.Click here for file"}
+{"text": "In order to improve the monitoring of the antimalarial drug resistance in Madagascar, a new national network based on eight sentinel sites was set up. In 2006/2007, a multi-site randomized clinical trial was designed to assess the therapeutic efficacy of chloroquine (CQ), sulphadoxine-pyrimethamine (SP), amodiaquine (AQ) and artesunate plus amodiaquine combination (ASAQ), the antimalarial therapies recommended by the National Malaria Control Programme (NMCP).Children between six months and 15 years of age, with uncomplicated falciparum malaria, were enrolled. Primary endpoints were the day-14 and day-28 risks of parasitological failure, either unadjusted or adjusted by genotyping. Risks of clinical and parasitological treatment failure after adjustment by genotyping were estimated using Kaplan-Meier survival analysis. Secondary outcomes included fever clearance, parasite clearance, change in haemoglobin levels between Day 0 and the last day of follow-up, and the incidence of adverse events.A total of 1,347 of 1,434 patients (93.9%) completed treatment and follow-up to day 28. All treatment regimens, except for the chloroquine (CQ) treatment group, resulted in clinical cure rates above 97.6% by day-14 and 96.7% by day-28 (adjusted by genotyping). Parasite and fever clearance was more rapid with artesunate plus amodiaquine, but the extent of haematological recovery on day-28 did not differ significantly between the four groups. No severe side-effects were observed during the follow-up period.These findings (i) constitute an up-dated baseline data on the efficacy of antimalarial drugs recommended by the NMCP, (ii) show that antimalarial drug resistance remains low in Madagascar, except for CQ, compared to the bordering countries in the Indian Ocean region such as the Comoros Archipelago and (iii) support the current policy of ASAQ as the first-line treatment in uncomplicated falciparum malaria. Plasmodium falciparum drug-resistance, especially with the spread of the parasite resistance to the inexpensive and widely used drugs, such as chloroquine (CQ) or sulphadoxine-pyrimethamine (SP) [Despite major efforts made by national and international health organizations, malaria remains the most widespread infectious parasitic diseases and one of the most serious global health problems in the world . In sub-ine (SP) ,3, and tine (SP) ,5.Therefore, substantial efforts have been made to encourage the monitoring and the evaluation of the antimalarial drugs resistance in the endemic countries for assessing regularly their antimalarial drug policies and ensuring a continued coverage of effective antimalarial treatment .\u00ae sold by the NGO \"Population Service International\" (PSI) or Ody Tazomoka\u00ae freely distributed at primary public health facilities by Malagasy Ministry of Health , as transitory measure until ACTs were available at community level [In Madagascar, since 2005, antimalarial treatment is in transition with the elaboration and the implementation of the new national policy for the fight against malaria . The maity level . At presin vitro assessment of P. falciparum sensitivity to antimalarial drugs and the evaluation of the frequency of genetic markers associated with P. falciparum drug resistance [In vivo studies were also carried out, but these studies were limited geographically, the sample sizes were small and used only a 14-day follow-up period, a series of constraints which may have significantly underestimated the true risk of treatment failure [Since 1999, the antimalarial drug resistance surveillance system in Madagascar is supported by a national network for the surveillance of malaria resistance (named RER \u2013 R\u00e9seau d'Etude de la R\u00e9sistance). The RER was formed as a collaborative effort between the MalMoH and the Malaria Research Unit of the Institut Pasteur de Madagascar (IPM). The strategy of monitoring was initially based on the use of the sistance . In vivo failure -14.In order to improve the monitoring of antimalarial drug resistance in Madagascar, a new RER was set up in 2006, with the support of Global Fund. According to the WHO recommendation , this neThe antimalarial drug efficacy trials were conducted in two steps in eight sentinel sites located in the four different malarious epidemiological strata throughout Madagascar: from February to June 2006, in Ejeda and Ihosy in the South Madagascar , in Maevatanana and Miandrivazo in the West Madagascar  and in Tsiroanomandidy and Moramanga in the foothills of the Central Highlands of Madagascar ; from March to August 2007 in Andapa and Farafangana in the East Madagascar  Figure .This was an open-label trial in which patients with microscopically-confirmed falciparum malaria were randomized into three or four treatment groups according to sentinel sites: CQ, amodiaquine (AQ), SP or ASAQ in Ejeda, Ihosy, Miandrivazo, Maevatanana and Moramanga; CQ, AQ and SP in Tsiroanomandidy and AQ, SP and ASAQ in Andapa and Farafangana.P. falciparum at a parasitaemia between 1,000 and 200,000/\u03bcl, (ii) axillary temperature \u2265 37.5\u00b0C, (iii) body weight > 5 kg, (iv) absence of severe malnutrition, (v) absence of febrile conditions caused by diseases other than malaria, (vi) absence of 'danger signs'  and of severe and complicated malaria, (vii) haemoglobin (Hb) \u2265 5 g/dl, and (viii) informed written consent of parents/guardians. Known hypersensitivity to SP, AQ, or AS was considered as an exclusion criterion. Once written informed consent was given, patients were enrolled in the study and assigned consecutive patient numbers. Randomization to treatment group was performed in blocks of three or four, and treatment regimens were allocated by an independent individual, not involved in the analysis of the study.All patients aged between six months and 15 years were eligible to be enrolled and were screened for malaria at the primary health centres in the sentinel sites on the basis of history of febrile illness. According to the slightly modified WHO 2003 protocol [Patients were administered either CQ , AQ , SP (25 mg/kg sulphadoxine and 1.25 mg/kg pyrimethamine as a single dose on day 0) or ASAQ . Patients were directly observed for 30 minutes after treatment, and the dose was readministered if vomiting occurred. Patients who repeatedly vomited their first dose of study medication were excluded from the study.Patients were assessed on days 1, 2, 3, 7, 14, 21 and 28, and any intervening day they were unwell for malaria infection. Blood was obtained by finger prick on all follow-up days and on any unscheduled day to use for analysis of thick and thin blood smears and for storage on filter paper. Thick and thin blood slides were examined by light microscopy for parasites on any day during the 28-day follow-up. Blood slides were read by a microscopist blind to treatment allocation. All slides were controlled by a second microscopist also blind to treatment group and previous diagnosis. Discordant slides were read, blind to treatment group and previous diagnosis, by a third microscopist. Haemoglobin was measured on Day 0 and Day 28 using a HemoCue haemoglobinometre .Treatment outcomes were assessed according to WHO 2003 guidelines as Early Treatment Failure , Late Clinical Failure , Late Parasitological Failure , and Adequate Clinical and Parasitological Response  . OverallPatients classified as having suffered treatment failure were treated with quinine (10 mg/kg three times daily for seven days); however, their response to repeat therapy was not assessed. Patients were excluded after enrolment if any of the following occurred: (i) use of antimalarial drugs outside of the study protocol; (ii) detection during follow-up of mixed malarial infections (iii) parasitaemia in the presence of a concomitant febrile illness which would interfere with the classification of treatment outcome; (iv) withdrawal of consent; (v) loss to follow-up, (vi) protocol violation, or (vii) death due to a non-malaria illness.Blood smears were stained with 10 % Giemsa for 10 min. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 200 WBCs , assuming a WBC count of 8,000/\u03bcl. A smear was considered negative if no parasites were seen after review of 100 fields. Thin blood smears were used to detect non-falciparum infections .msp-1) and merozoite surface protein-2 (msp-2) using nested-PCR as previously described [msp-2 genotyping patterns on the day of failure were compared with those at treatment initiation and on day 1, using Quantity One\u00a9 software . If all of the msp-2 alleles present on the day of failure were present at the time of treatment initiation or on day 1, genotyping was repeated using msp-1. An outcome was defined as recrudescence if all msp-1 and msp-2 alleles present at the time of failure were present at the time of treatment initiation or on day 1, and defined as a new infection otherwise.Molecular genotyping techniques were used to distinguish recrudescence from new infection for all patients failing therapy after day 7. Filter paper blood samples collected on the day of enrolment, on day 1 and on the day of failure were analysed for polymorphisms in the genes for merozoite surface protein-1 , and analysed using MedCalc\u00a9 software version 9.1.0.1 . As the proportion of treatment failures was unknown in the sentinel sites, a minimum of 50 patients was included for each treatment group and each site, according to available human and financial resources and the WHO recommendations [Data were entered and verified using EpiInfo 6.04ndations .2 or Fisher's exact test, and continuous variables were compared using an independent samples t test.Efficacy data were assessed with a per-protocol analysis that included all patients who completed the study. An age-stratified analysis for patients less than five years of age and patients five years or older was planned. This study was not designed for the primary analysis to be stratified by site. Parasite densities were normalized using logarithmic transformation. Categorical variables were compared using \u03c7P values. A P value (two-tailed) of less than 0.05 was considered statistically significant.The primary efficacy outcomes were 14-day and 28-day clinical and parasitological failure risks both unadjusted and adjusted by genotyping. Risks of clinical and parasitological treatment failure after adjustment by genotyping were estimated using Kaplan-Meier survival analysis techniques in accordance with the new WHO protocol . With suThe study protocol was reviewed and approved by the Ethics Committee of the Ministry of Health of Madagascar (N\u00b0007/SANPF/2007). An informed written consent was provided by the parents/guardians of all patients before they were included in the study.P. falciparum (22.4%). Of these patients, a total of 1,434 patients were enrolled in February-June 2006 and March-July 2007 (76.6%). A total of 320 patients were randomized to CQ, 385 to AQ, 383 to SP, and 346 to ASAQ. The main reasons for patients with P. falciparum not being included were non-consent, inability to attend follow-up or concurrent disease. The flow of patients through the trial is shown in Figure In the eight sentinel sites, 8,363 patients were screened, with 1,873 patients positive for lciparum 2.4%. Of A total of 1,347 of 1,434 patients (93.9%) completed treatment and follow-up to day 28 . Eighty eight patients were either lost to follow-up (n = 68) or excluded/withdrawn from the trial (n = 20). Of these patients, 18 withdrew consent, two were withdrawn due to adverse events. The baseline characteristics of patients across the treatment groups were similar at each site (Table The results of the treatment efficacy are presented by treatment group and day of follow-up in Table P < 0.0001).All treatment regimens, except for the CQ treatment group, resulted in clinical cure rates above 97.6% by day-14 and 96.7% by day-28 (adjusted by genotyping). Considering all sites, the OTF of the CQ group was significantly higher than in the other treatment group as well as day 14 or day 28 unadjusted or adjusted by genotyping (P < 0.0001). The risk of treatment failure adjusted by genotyping in children less than five years old was almost twice as frequent than in older patients . The risk of treatment failure by day-14 was significantly lower than by day-28 adjusted by genotyping .In the CQ group, ETF was uncommon with only 10.6% of the OTF . Most of the treatment failures adjusted by genotyping were occurred in the second/third week of follow-up mainly as LPF (63.6%). Among the eight sites, the OTF adjusted by genotyping of CQ was significantly different ranging from 19% in Ihosy to 64% in Ejeda (P = 0.08), especially because of the small number of the patients included per site and the small number of treatment failure.In the SP group, treatment failures were rare with an OTF adjusted by genotyping of 3.3%, but ETF was accounting for more than one third of the treatment failures: Two patients from Ejeda developed danger signs despite decreasing parasite density, one patient from Tsiroanomandidy presented at day 3, a parasite density > 25% of count on day 0 and the last patient from Maevatanana, a parasite density with an axillary temperature \u2265 37.5\u00b0C. The distribution of the treatment failure adjusted by genotyping was not significantly different among the sites (P = 0.24 for AQ and P = 0.12 for ASAQ).In the AQ and ASAQ groups, treatment efficacies were globally very high throughout Madagascar. Treatment failures adjusted by genotyping were not significantly different among the sentinel sites . Fever clearance was delayed with CQ and SP, the proportion of febrile patients being significantly lower with ASAQ and AQ until day-2. The details are shown on Figure No severe side-effects attributable to the study medication were observed during the follow-up period. Minor side effects as vomiting were reported between day 1 and day 3 in seventeen patients (1.26%): 8 in the AQ group, 4 in the CQ and the SP groups and 1 in the ASAQ group.in vivo drug efficacy. Seven additional sites were secondly incorporated in the RER for monitoring in vitro drug resistance (in vitro drug sensitivity assay and molecular markers) to provide complementary data and warning signals of the possible emergence of resistance or trends of declining drug efficacy.As effective case management remains the cornerstone of malaria elimination , the monP. falciparum parasitaemia on inclusion was between 1,000 and 200,000/\u03bcl. The protocol included randomized, 28-day follow-up with genotyping to discriminate recrudescence from new infections, and assessment of safety and tolerability. Despite the complexity of conducting a multi-site study in Madagascar, an excellent completion rates was achieved (> 93%) and high-quality data were collected.The objective of the new RER was firstly to establish up-dated baseline data on the efficacy of the antimalarial drugs recommended by the MalMoH in its new Madagascar national policy. The methodology used was based on the WHO 2003 protocol , with soThe four main antimalarial drugs used and recommended by the NMCP were evaluated in five sites. In Tsiroanomandidy, ASAQ was not evaluated because this drug was already assessed in another clinical trial  and in ATwo methodological points could be underline by the results of the study: (1) the necessity to extend the follow-up to a minimum of 28 days (even more for drugs with a prolonged duration of action) to avoid the underestimation of the risk of treatment failure , i.e. foin vivo drug efficacy study performed in Madagascar since 1983 [et al [This multi-site study represents the first extent nce 1983 -24. The 3 [et al , even wiPf-dhfr and Pf-dhps [The results concerning the SP efficacy were more informative, contrasting with the good effectiveness reported in previous studies in 2003\u20132004 ,14. The  Pf-dhps  will be This study also confirmed that ASAQ, the first-line treatment of the uncomplicated malaria recommended in Madagascar was very effective, particularly because of the excellent effectiveness of AQ, the partner drug associated with artesunate. Although it was not feasible to monitor for the potentially serious adverse effects associated with amodiaquine such as neutropenia or hepatotoxicity , ASAQ was well tolerated in the study population. This combination produced also faster parasite and fever clearances than the other antimalarial treatments. These findings constitute important baseline data on the efficacy of this combination, a key tool within the framework of the elimination of the malaria .Pfcrt) or SP-resistance (Pf-dhfr or Pf-dhps) or the in vitro response of parasites for drug for which no validate molecular marker are available such as amodiaquine, artemisinin derivatives or amino alcohols . These complementary indicators should allow the early detection of the spread of P. falciparum drug resistance, especially those introduced from the Comoros Islands [The results from this multi-site study show that antimalarial drug resistance remains low in Madagascar, except for CQ, compared to the bordering countries in the Indian Ocean region such as the Comoros Archipelago ,27. The  Islands . Indeed,DM contributed to the design and coordination of the study, supervised the enrolment and follow up of the patients, assisted with data entry and interpretation and prepared the manuscript. ARat supervised the enrolment and follow up of the patients. LR, ARan and RR performed field work in Tsiroanomandidy and Moramanga. M-AR and ER were involved in laboratory work. MJ and VA carried out molecular genotyping. MR, L-PR, LR, OD, LT and ARav helped to compose the manuscript and gave constructive advice."}
+{"text": "This study compared the safety and tolerability of three combination antimalarial regimens in a cohort of Ugandan children.Combination antimalarial therapy is recommended for the treatment of uncomplicated A longitudinal, single-blind, randomized clinical trial of children was conducted between November 2004 and May 2007 in Kampala, Uganda. Upon diagnosis of the first episode of uncomplicated malaria, participants were randomized to treatment with amodiaquine + sulphadoxine-pyrimethamine (AQ+SP), artesunate + amodiaquine (AS+AQ), or artemether-lumefantrine (AL). Once randomized, participants received the same regimen for all subsequent episodes of uncomplicated malaria. Participants were actively monitored for adverse events for the first 14 days after each treatment, and then passively followed until their next study medication treatment, or withdrawal from study. Outcome measures included the risk of adverse events at 14 and 42 days after treatment.Of 601 enrolled children, 382 were diagnosed with at least one episode of uncomplicated malaria and were treated with study medications. The median age at treatment was 6.3 years (range 1.1 \u2013 12.3 years). At 14 days of follow-up, AQ+SP treatment was associated with a higher risk of anorexia, weakness, and subjective fever than treatment with AL, and a higher risk of weakness, and subjective fever than treatment with AS+AQ. Treatment with AL was associated with a higher risk of elevated temperature. Repeated episodes of neutropaenia associated with AS+AQ were detected in one participant. Considering only children less than five years, those who received AQ+SP were at higher risk of developing moderate or severe anorexia and weakness than those treated with AL , or AS+AQ . Extending the analysis to 42 days of follow-up had little impact on the findings.This study confirms the safety and tolerability of AS+AQ and AL in Ugandan children, and suggests that AQ+SP is safe, but less well-tolerated, particularly in younger children. As newer antimalarial regimens are deployed, collecting data on their safety and tolerability will be essential.Current Controlled Trials Identifier ISRCTN37517549 Combination antimalarial therapy is now advocated for treatment of uncomplicated falciparum malaria in Africa ,2. Of thAlthough combination therapy is highly effective, some concerns about the safety and tolerability of these regimens remain . SeriousRecent studies of the safety of newer combination regimens have been reassuring, identifying no serious concerns, but to date, assessments of drug safety and tolerability have been limited -22. MostA longitudinal, single-blind, randomized controlled trial assessing the efficacy and safety of AQ+SP, AS+AQ and AL for all episodes of uncomplicated malaria was begun in November 2004 in a cohort of children in Kampala, Uganda . An analThe study was conducted between November 2004 and May 2007 in the Mulago III parish of Kampala. Mulago III parish is primarily residential with a high population density, typical of an urban slum. The parish is located near the study clinic at Mulago Hospital, the main tertiary referral hospital for Uganda.A census project was conducted prior to the onset of the study to generate a sampling frame of households with appropriately aged children for recruitment and to gather basic demographic information about the target population . ExperieThe study received ethical approval from the Uganda National Council of Science and Technology, the Makerere University Research and Ethics Committee, and the University of California, San Francisco Committee on Human Research.Parents and guardians were asked to bring participants to the study clinic for all medical care. At each participant encounter a standardized system was used to assess participant symptoms, physical exam findings and laboratory abnormalities based on grading of severity . In general, events were classified as mild if no therapy was required, as moderate if minimal intervention and/or monitoring were required, as severe if medical care and possible hospitalization were required, and as life-threatening if active medical intervention or hospitalization were required. Weakness, anorexia, vomiting, diarrhoea, cough, pruritus, and coryza were actively assessed in all participants. Headache, abdominal pain, and nausea were also assessed in children over three years of age. A standardized physical exam was also conducted, which included assessment of hearing and fine finger dexterity . Any additional symptoms reported by the participant or exam findings were also assessed and graded accordingly.If a participant presented with fever, a blood smear was taken, and malaria diagnosed if any of the following criteria were met: 1) complicated malaria  and any parasitaemia; or 2) fever (documented tympanic temperature \u226538.0\u00b0C and/or history of fever in the previous 24 hours) and any parasitaemia . FollowiStudy medications were administered according to weight-based guidelines. The study medications were dosed as follows: amodiaquine , 10 mg/kg on first two days then 5 mg/kg on third day, sulphadoxine-pyrimethamine , sulphadoxine 25 mg/kg and pyrimethamine 1.25 mg/kg as a single dose on first day; artesunate , 4 mg/kg on all three days; artemether-lumefantrine , 20/120 mg tablets given twice a day for three days according to weight: 5\u201314 kg, one tablet per dose; 15\u201324 kg, two tablets per dose; and 25\u201334 kg, three tablets per dose. Participants in the AQ+SP and AS+AQ groups also received placebo tablets administered in the evening over three days, dosed similarly to weight-based guidelines for AL. Administration of each first daily dose of medication was directly observed by the study nurses, and each second daily dose of medication or placebo was given to the participant's parent or guardian to administer at home in the evening. All study personnel involved in outcome assessment were blinded to treatment allocation and participants were not informed of their treatment assignments.Participants randomized to treatment with study medications were assessed for adverse events beginning with the first malaria episode. An adverse event was defined as any untoward medical occurrence, irrespective of its suspected relationship to the study medications as per International Conference of Harmonization (ICH) guidelines . A serioOn the day that treatment was initiated (day 0), a baseline assessment was conducted consisting of the standardized history and physical examination, and laboratory testing . Participants were asked to return for completion of treatment and standardized follow-up assessment on days 1, 2, 3, 7, 14, or any other day they felt ill. Complete blood count and ALT measurement were repeated on day 14.At each follow-up encounter, adverse events were identified by evaluating for any new or worsening symptoms, physical exam findings, or laboratory abnormalities, as compared to the day 0 baseline assessment. Participants with abnormalities present on day 0 were not classified as experiencing an adverse event unless the symptom worsened from baseline, or resolved and then recurred. For adverse events of moderate or greater severity, additional information was captured, including the suspected relationship of the event to the study treatment , event outcome, and date of resolution. Participants who experienced a serious adverse event were placed on study treatment hold until the relationship of the event to the study medications could be established. If the event was deemed unrelated or only possibly related to the study medications, participation in the study was continued. Patients with clinical treatment failure (according to 2006 WHO criteria) within 14 days of initiation of therapy were treated with quinine, and active surveillance for adverse events was continued for an additional 14 days .Following the initial 14-day period, participants were asked to return to the clinic only when they desired medical attention. Passive surveillance for adverse events of moderate or greater severity continued until the participant was retreated with study medications, at which time a new cycle of adverse event assessment and reporting began, until the end of the study period, or until premature withdrawal from the study. Any case of symptomatic malaria diagnosed more than 14 days after a previous episode was considered a new malaria episode (for treatment purposes) and managed accordingly. Once randomized, participants received the same treatment regimen for all subsequent episodes of uncomplicated malaria.Complete blood counts and alanine aminotransferase measurements were performed by a College of American Pathologists accredited laboratory using a Coulter AcT 5 diff CP (Beckman Coulter) for haematology tests and Cobas Integra 400 plus (Roche) for chemistries.Details on sample size calculation are presented elsewhere . Data weData were analysed in a stepwise approach. All adverse events of any severity or relationship at 14 days of follow-up were first considered, then the analysis was restricted for adverse events of moderate or greater severity, and finally restricted for adverse events of possible, probable, or definite relationship to study medications. Risks of adverse events at 14 days were estimated using simple proportions as nearly all patients completed 14 day follow-up. Data at 42 days of follow-up was limited to adverse events of moderate or greater severity. Risks of adverse events at 42 days of follow-up were estimated using the Kaplan-Meier product limit formula, censoring for participants who did not complete 42 day follow-up either due to repeat treatment with study drugs or premature withdrawal from the study.Pairwise comparisons of adverse events for individual episodes of malaria were made at 14 and 42 days of follow-up with generalized estimating equations, adjusting for repeated measures in the same study participant using exchangeable correlation and robust standard errors. A p-value of < 0.05 was considered statistically significant.Details of recruitment and the study population are presented elsewhere ,26. BrieA total of 1120 study treatments were administered during 605 person-years of follow-up  were of moderate or greater severity. Of these events, 143 (19%) were classified as not related, 541 (72%) as possibly related, and 65 (9%) as probably related to study medications. No adverse events were considered to be definitely related to the study medications.Statistically significant differences between the treatment groups were found for 4 adverse events: anorexia, subjective fever, weakness, and elevated temperature Table . Within 3), one was moderate (660 cells/mm3), and one was severe (330 cells/mm3). For nine of the 10 treatments, the initial day 0 neutrophil count was within normal limits. All episodes of neutropaenia resolved spontaneously. The participant was subsequently excluded from the study due to concern about a causal association between AS+AQ and the neutropaenic episodes. Moderate grade neutropaenia was only seen in one other participant, after treatment with AQ+SP. This participant developed neutropaenic episodes after two of 15 study treatments, with one mild, and one moderate decrease in neutrophil count.At 14 days, there were no statistically significant differences in the risk of laboratory-related adverse events between treatment groups  occurred within the first 14 days of follow-up , 10 episodes of moderate or severe anaemia, and one episode of severe thrombocytopaenia. One participant treated with AL developed severe ALT elevation at 14 days of follow-up, which was deemed possibly related to study medications, although an infectious hepatitis was suspected.We assessed for factors which might modify the effect between treatment and risk of adverse events. Neither the number of prior treatments, nor duration since prior treatment, significantly impacted the risk of adverse events. However, when the analysis of the association between treatment and risk of adverse events was stratified by age, significant differences were found Table . The assPassive surveillance for adverse events of moderate or greater severity continued until participants were diagnosed with another episode of uncomplicated malaria, or study follow-up ended. During 15\u201342 days of follow-up after study medication treatment, an additional 440 adverse events were recorded, representing 15% of total adverse events captured. No life-threatening adverse events were observed. No events suspected to be probably or definitely related to study medications were detected. During 15\u201342 days of follow-up, an additional three SAEs were reported. Of these, one event was a febrile convulsion; another was elevation of liver enzymes secondary to acute hepatitis A infection, and the last a skin abscess.At 42 days, patients treated with AQ+SP remained at significantly higher risk of anorexia and weakness than AL-treated participants; however, the difference in risk of subjective fever was not sustained (Table In this randomized, blinded, longitudinal clinical trial, the safety and tolerability of three commonly used antimalarial combinations in Ugandan children was evaluated. Although all three regimens were found to be safe, participants treated with AQ+SP, particularly young children, were significantly more likely to develop anorexia and weakness than participants treated with AL or AS+AQ. Focusing the analysis on events of moderate or greater severity strengthened the observed associations, but redefining outcomes based on suspected relationship to study medications was of limited utility. Extending follow-up for adverse events from 14 to 42 days had little impact on the findings. Overall, in our cohort of children in Kampala, AL and AS+AQ appear to be superior to AQ+SP due to improved tolerability and greater efficacy .ACTs are generally considered to be safe and well-tolerated, which is supported by this study ,28,29. TAlthough no significant differences in safety were found between AS+AQ and AL in the cohort, the one case of repeated neutropaenic episodes associated with AS+AQ was concerning given the known propensity of amodiaquine to cause blood dyscrasias and agranulocytosis . A recenIn this study, a higher risk of anorexia and weakness associated with AQ+SP when compared with the other regimens was found, consistent with prior reports of severe weakness in Rwandan children treated with AQ+SP ,28. HoweNo standardized system for adverse event monitoring in antimalarial clinical trials currently exists, and the approach to monitoring may differ between sites. Guidelines for use of laboratory testing in antimalarial drug safety monitoring are also lacking.  In this study, events were actively assessed at all follow-up visits, with interviews, a standard physical exam, and laboratory tests at standard intervals. Although time intensive, the system allowed events related to drug tolerability, such as anorexia and weakness, to be captured. Relying on spontaneous reporting of symptoms by participants, or assessing only for objective measures, such as serious adverse events and laboratory abnormalities, may have missed differences in tolerability. The duration of follow-up for monitoring of adverse events after antimalarial therapy is variable, often coinciding with the recommended length of follow-up for efficacy outcomes of 28 or 42 days ,19,20,28There were several potential limitations to this study. Although the assessment of adverse events was designed to be as objective as possible by using grading scales for severity and relationship, the results are limited by the subjective interpretations of symptom reports. In addition, active surveillance continued only for the 14 days after study treatment was initiated. Extending the duration of active surveillance may have captured additional events of interest. Laboratory values were only assessed at day 0 and day 14 of study treatment, routinely every 90 days, and at the discretion of the study clinicians. Some abnormalities may not have been captured by this schedule. Although 382 patients treated with study medications over multiple treatments were followed, the relatively small sample size limited the ability to detect uncommon events. Finally, multiple comparisons in the risk of adverse events associated with treatment groups were made, increasing the likelihood that statistically significant differences in risks could have been detected just by chance.This study confirms the safety and tolerability of AS+AQ and AL in Ugandan children, and suggests that AQ+SP is safe, but less well-tolerated than the other regimens, particularly in young children. The higher risk of elevated temperature observed in association with AL could impact on the perception of its effectiveness and may need to be addressed. Although no significant differences in safety were found between AS+AQ and AL, the one case of repeated neutropaenic episodes associated with AS+AQ was concerning. As newer ACTs are deployed, evaluating for safety and comparative tolerability will be essential. Standardized guidelines for monitoring adverse events in antimalarial clinical trials are needed.PJR, GD, MRK, and SGS conceived and designed the study, CMS, TDC, DNM, BN, and SGS participated in data collection, GD, TDC, PJ, and VY participated in the data analysis, CMS, PJ, TDC, AOT, PJR, GD, and SGS participated in interpretation of the data. All authors participated in the writing of the manuscript, and read and approved the final manuscript. None of the authors declare any potential conflicts of interest, including financial interests and relationships and affiliations relevant to the subject of the manuscript."}
+{"text": "New antimalarial regimens, including artemisinin-based combination therapies (ACTs), have been adopted widely as first-line treatment for uncomplicated malaria. Although these drugs appear to be safe and well-tolerated, experience with their use in Africa is limited and continued assessment of safety is a priority. However, no standardized guidelines for evaluating drug safety and tolerability in malaria studies exist. A system for monitoring adverse events in antimalarial trials conducted in Uganda was developed. Here the reporting system is described, and difficulties faced in analysing and interpreting the safety results are illustrated, using data from the trials.Between 2002 and 2007, eleven randomized, controlled clinical trials were conducted to compare the efficacy, safety, and tolerability of different antimalarial regimens for treatment of uncomplicated malaria in Uganda. The approach to adverse event monitoring was similar in all studies. A total of 5,614 treatments were evaluated in 4,876 patients. Differences in baseline characteristics and patterns of adverse event reporting were noted between the sites, which limited the ability to pool and analyse data. Clinical failure following antimalarial treatment confounded associations between treatment and adverse events that were also common symptoms of malaria, particularly in areas of lower transmission intensity.Despite prospectively evaluating for adverse events, limitations in the monitoring system were identified. New standardized guidelines for monitoring safety and tolerability in antimalarial trials are needed, which should address how to detect events of greatest importance, including serious events, those with a causal relationship to the treatment, those which impact on adherence, and events not previously reported.Although the World Health Organization has supported the development of pharmacovigilance systems in African countries deploying ACTs, additional guidance on adverse events monitoring in antimalarial clinical trials is needed, similar to the standardized recommendations available for assessment of drug efficacy. Since 2004, antimalarial treatment in sub-Saharan Africa has changed dramatically. As a result of widespread resistance to chloroquine and sulphadoxine-pyrimethamine, most sub-Saharan African countries have recently revised their antimalarial drug policies, selecting new artemisinin-based combination therapies (ACTs) as first-line treatment for uncomplicated malaria . AlthougGuidelines for assessing the efficacy of antimalarial treatments have been provided by the World Health Organization (WHO) -9. HowevTo evaluate the safety and tolerability of newer combination antimalarial therapies, a system for monitoring adverse events in clinical trials conducted in Uganda since 2002 was developed. This system has been used in studies evaluating treatment of single episodes of uncomplicated malaria, and in a longitudinal cohort study evaluating repeated antimalarial treatments. Here the reporting system is described, and difficulties faced in analysing and interpreting the safety data are discussed. The challenges of adverse event monitoring and key areas that need to be addressed in developing guidelines for assessing the safety and tolerability of antimalarial drugs are highlighted.From 2002 to 2007, eleven randomized, controlled clinical trials were conducted to compare the efficacy, safety, and tolerability of different combination antimalarial regimens for treatment of uncomplicated malaria in Uganda Table . Ten of Ethical approval for the studies was obtained from the Uganda National Council for Science and Technology ; the University of California, San Francisco Committee on Human Research ; the Makerere University Research and Ethics Committee ; and the University of California, Berkeley .Kampala is an urban center where malaria is meso-endemic . The UMSP study sites, selected for geographic diversity, included: Kanungu , Kyenjojo , Mubende , Arua , Apac , Tororo , and Jinja  , and creatinine . In the UMSP studies, blood was obtained by finger-prick for thick and thin blood smears, and haemoglobin measurement.Participants were randomly assigned study treatment in all trials, although procedures varied slightly between studies. Details on randomization, treatment allocation and administration, and blinding are included in the initial study reports ,15-18,20The standardized history and physical examination was repeated at all scheduled and unscheduled follow-up visits. Follow-up laboratory assessments in the Kampala studies included thick blood smears, complete blood count and ALT , and creatinine . In the UMSP studies, repeat assessments included thick blood smears, and haemoglobin on the final day of follow-up or the day of treatment failure.The approach to adverse event monitoring was similar in all studies. An adverse event was defined as any untoward medical occurrence, irrespective of its suspected relationship to the study medications as per International Conference of Harmonization (ICH) guidelines for Good Clinical Practice (GCP) . A serioAt each participant encounter a standardized system was used to assess symptoms, physical exam findings, and laboratory test results based on grading of severity  would not be classified as experiencing an adverse event unless the symptom worsened from baseline, or resolved and then recurred. For all adverse events of moderate or greater severity, additional information was captured, including the suspected relationship of the event to the study treatment. In the ten studies evaluating treatment of single episodes of malaria, relationship was classified as not related, or unlikely, possibly, probably, or definitely related to study medications. In the Kampala longitudinal study, the classification was similar, but the 'unlikely' category was excluded. Data were double-entered individually for each study into EpiInfo version 6.04 or Access  and analysed using Stata version 8.0 . Categorical variables were compared using the chi-square test or Fisher's exact test. Continuous variables were compared using two-sample t-tests.A total of 5,614 study treatments were evaluated in 4,876 patients enrolled in the eleven clinical trials  were classified as unlikely or possibly related to the study medications; classification of an event as 'probably' related to the study treatment occurred rarely. This likely reflects the difficulty in ruling out alternative etiologies for an event, or an understandable reluctance on the part of physicians to commit to more definite relationship classes. Assessment for causality requires careful consideration of various factors including the timing of the event, the timing of the treatment, response to antimalarial therapy, presence of concomitant illnesses, and administration of concomitant medications and herbal therapies. Algorithms based on a decision tree considering these factors could be developed to help standardize assessment of the relationship between adverse events and antimalarial treatment.Antimalarial clinical trials are typically designed and powered to test hypotheses about efficacy. As a result, the sample size of trials generally limits the ability to detect uncommon events. The safety data collected at the different sites was to be pooled in order to increase the power to detect differences between the study treatments. However, significant differences in baseline characteristics, and patterns of drug use, were observed between the study sites. In addition, despite using a standardized protocol and similar training, differences in adverse event assessment and reporting at the different sites were observed, which complicated the analysis. The potential for such variation in baseline characteristics and adverse event monitoring may significantly impact on the ability to effectively combine data collected in different studies and sites, even if standardized protocols are used. Analysis of safety data is also challenged by the impact of multiple comparisons on hypothesis testing. The likelihood that significant differences between treatments may be found just by chance increases with the number of comparisons made, which may need to be taken into account when designing studies and interpreting results.In the studies of single episodes of malaria, clinical treatment failure appeared to confound the association between study treatment and certain adverse events, particularly in areas of lower transmission intensity. A study participant who fails initial therapy and develops early treatment failure or recurrent clinical malaria will present with the signs and symptoms of acute illness, which will be captured as adverse events. In areas of high transmission, where participants are at risk of developing new infections within the study follow-up period, this will be an issue even if the drugs are highly efficacious. Thus, the efficacy of an antimalarial regimen for treating the initial infection, and for preventing new infections, will be associated with the risk of adverse events. One approach, which was considered, is to restrict the analysis by excluding treatment failures. However, excluding treatment failures limits the available data, particularly in areas of high transmission. Controlling for treatment failure and other relevant factors in a multivariate analysis is another approach. Alternatively, this problem could be addressed in the study design. In the longitudinal study of repeated malaria treatment episodes, a new cycle of adverse event reporting began with each new episode of malaria, and symptoms noted on that day were not considered adverse events but rather baseline for the new episode. However, the cohort study design may not be optimal for all settings.Further studies are needed to establish the safety profiles of newer antimalarial regimens in Africa, particularly with widespread use. Despite prospectively evaluating for adverse events using standardized methods modelled on currently available international guidelines, limitations in the system were identified. New guidelines for monitoring for safety and tolerability in antimalarial trials are needed. Although the WHO has acknowledged the need for pharmacovigilance, and has taken the lead in supporting the development pharmacovigilance systems in African countries deploying ACTs, additional guidance on adverse events monitoring in antimalarial clinical trials would be welcomed ,26. SuchSGS, PJR, GD, MRK, FWM, and AT conceived and designed the studies, SGS, AM, AY, HB, KB, CMS, TDC, DNM, BN and GD participated in data collection, SGS, PJ, HB, KB, and GD participated in the data analysis. All authors participated in the writing of the manuscript, read and approved the final manuscript. None of the authors declare any potential conflicts of interest, including financial interests and relationships and affiliations relevant to the subject of the manuscript.Guidelines for physical examination.Click here for fileSeverity grading guidelines.Click here for file"}
+{"text": "Artemisinin combination therapies (ACTs) are currently the preferred option for treating uncomplicated malaria. Dihydroartemisinin-piperaquine (DHA-PQP) is a promising fixed-dose ACT with limited information on its safety and efficacy in African children.P. falciparum malaria was tested in five African countries . Patients were randomised (2\u22361) to receive either DHA-PQP or AL. Non-inferiority was assessed using a margin of \u22125% for the lower limit of the one-sided 97.5% confidence interval on the treatment difference (DHA-PQP vs. AL) of the day 28 polymerase chain reaction (PCR) corrected cure rate. Efficacy analysis was performed in several populations, and two of them are presented here: intention-to-treat (ITT) and enlarged per-protocol (ePP). 1553 children were randomised, 1039 receiving DHA-PQP and 514 AL. The PCR-corrected day 28 cure rate was 90.4% (ITT) and 94.7% (ePP) in the DHA-PQP group, and 90.0% (ITT) and 95.3% (ePP) in the AL group. The lower limits of the one-sided 97.5% CI of the difference between the two treatments were \u22122.80% and \u22122.96%, in the ITT and ePP populations, respectively. In the ITT population, the Kaplan-Meier estimate of the proportion of new infections up to Day 42 was 13.55% (95% CI: 11.35%\u201315.76%) for DHA-PQP vs 24.00% (95% CI: 20.11%\u201327.88%) for AL (p<0.0001).The non-inferiority of DHA-PQP versus artemether-lumefantrine (AL) in children 6\u201359 months old with uncomplicated DHA-PQP is as efficacious as AL in treating uncomplicated malaria in African children from different endemicity settings, and shows a comparable safety profile. The occurrence of new infections within the 42-day follow up was significantly lower in the DHA-PQP group, indicating a longer post-treatment prophylactic effect.ISRCTN16263443Controlled-trials.com  Artemisinin-based combination therapies (ACTs) are highly efficacious and fast acting antimalarial medicines. The World Health Organization (WHO) recommends their use for treating uncomplicated malaria Several trials The protocol for this trial and supporting non-inferiority adapted http://www.controlled-trials.com/isrctn.The study protocol was approved by the Institutional Review Board of the Institute of Tropical Medicine, Antwerp, the Ethical Committee of the Antwerp University Hospital, the University of Heidelberg Ethics Committee and by the National Ethics Review Committee or Institutional Review Board at each trial site. The trial was conducted under the provisions of the Declaration of Helsinki and in accordance with Good Clinical Practices guidelines set up by the International Conference on Harmonization. A Study Steering Committee, a Data Monitoring Committee and a Clinical Development Committee were created prior to the beginning of the trial, and worked independently to harmonise and monitor the study. The trial was registered prior to the enrolment of the first patient in the International Standard Randomized Controlled Trials Register, number ISRCTN 16263443, at Between August 2005 and July 2006, a randomised, open-label, multicentre clinical trial was carried out in five African sites . Characteristics of the five sites are summarized in Plasmodium falciparum mono-infection with asexual parasite densities between 2,000 and 200,000/\u00b5l; fever (axillary temperature \u226537.5\u00b0C) or history of fever in the preceding 24 h. Patients were not recruited if they met at least one of the following exclusion criteria: severe malaria Children 6\u201359 months old attending the health facilities with uncomplicated malaria were included in the study if they fulfilled the following inclusion criteria: body weight >5 kg; microscopically confirmed Patients were individually randomised according to a 2\u22361 (DHA-PQP\u2236AL) scheme, so as to have more patients in the DHA-PQP arm to provide better estimates for its cure rates and more cases for the integrated safety data base. A randomisation list stratified by country was generated by an independent off site contract research organisation (CRO), with each treatment allocation concealed in opaque sealed envelopes that were opened only after the patient's recruitment.Both drugs were administered under direct supervision during 3 consecutive days, according to the patient's body weight. AL  was administered twice a day  according to the following dosage: weight 5\u201314 kg: one tablet per dose; weight 15\u201324 kg: two tablets per dose; weight 25\u201334 kg: three tablets per dose. DHA-PQP  was given once daily, at the standard dosage of 2.25 mg/kg and 18 mg/kg per dose of DHA and PQP, respectively, rounded up to the nearest half tablet. To facilitate the correct dosing of DHA-PQP, two formulations were used (DHA 20 mg + PQP 160 mg and DHA 40 mg + PQP 320 mg). In case of vomiting, a full dose was repeated if this occurred within the first half an hour, or half a dose if it occurred between 30 minutes and 1 h. AL was administered concomitantly with milk (as recommended by the manufacturer) while for DHA-PQP no specific instructions regarding co-administration with food were given. For infants, drugs were crushed, mixed with water and administered as slurry.Both patient allocation to the different analysis populations and assessment of the primary end-point were made by staff blinded to the treatment assignment and before availability of the PCR results.All children were kept at the health facility for the 3-day dosing period. The mother/guardian was asked to return with the child for scheduled visits on days 7, 14, 21, 28, 35 and 42 post-treatment, or if any symptoms occurred. Field workers traced patients missing any visit. For each visit, a physical examination was performed by the study clinicians, vital signs were recorded, and axillary temperature measured with an electronic thermometer. Adverse events and serious adverse events were recorded and monitored throughout the study. A 12-lead electrocardiogram (ECG) was performed at enrolment and repeated on days 2 and 7 to assess any QT/QTc interval prolongation. Any ECG abnormality detected at enrolment requiring urgent management was considered an exclusion criterion. All ECG records were transmitted daily online to a central cardiologist  who interpreted them in a blinded manner, and feedback was sent to the sites as soon as available. The QTc interval (ms) was evaluated after correcting for the heart rate with Bazett's or Fridericia's formulae and classified according to the following categories: Normal <430 ms; Borderline: 431\u2013450 ms; Prolonged >450 ms.The study was supervised by monthly monitoring visits. Rescue treatment for recurrent parasitaemia was according to local national guidelines. All participants, with the exception of those in Kilifi, received a free insecticide-treated bed net at recruitment.Capillary or venous blood was taken at every visit. Thick and thin blood films were prepared, dried and Giemsa-stained, and parasite density estimated by counting the number of asexual parasites in 200 white blood cells (WBC), assuming a standard WBC count of 8,000/\u00b5l. Quality control was performed in blind conditions on 20% of all the slides at a central laboratory. Samples for haematology (full blood count) and biochemistry  were taken at enrolment, at days 3, 28 and 42, and at any other visit if judged necessary by the clinician. For PCR analysis, three blood spots were collected on filter paper (Whatmann 3 MM) at enrolment and at any visit after day 7. Each filter paper was dried and individually stored in a plastic bag containing silica gel. All filter papers were subsequently transferred to the Institute of Tropical Medicine  where centralised genotyping was conducted. Purification of DNA was conducted as previously described The primary endpoint was the PCR-corrected adequate clinical and parasitological response (ACPR) at day 28; secondary efficacy outcomes included PCR-corrected cure rates at days 14 and 42, PCR-uncorrected cure rates at days 14, 28 and 42; parasite and fever clearance times, presence and clearance of gametocytes, and haemoglobin (Hb) changes from baseline to day 28. All standard safety outcomes such as incidence of adverse events, changes from baseline on haematology and clinical chemistry parameters, ECG findings and vital sign variation during the study were also evaluated.Treatment outcome was analysed in two ways. The first, based on a pre-defined (in the protocol) procedure further developed with the Data Monitoring and the Clinical Development Committees, complemented the WHO definitions (see below) with a set of rules allowing the evaluation of each individually randomised patient, e.g. patients having taken not-allowed anti-malarial drugs or with halfway missing data such as blood parasitaemia . Such anSeveral populations were defined for the analysis and two of them , despite not being the primary ones as defined by the protocol, are presented here on the basis of their comparability with the populations discussed in previous published studies, their clinical relevance and the fact that conclusions are similar on all considered populations. The ITT included all randomised patients having taken at least one dose of the study treatments. The ePP population included all randomised patients fulfilling the protocol eligibility criteria, having taken at least 80% of the study medication when not previously classified as early treatment failures, completing the day 28 assessment and having an evaluable PCR in case of recurrent parasitaemia. Primary efficacy analysis was based on a 97.5% (one-sided) confidence interval (CI) computed on the difference between the day 28 PCR-corrected cure rates Rates of person-gametocyte-weeks for measuring gametocyte carriage and transmission potential were calculated as the number of weeks in which blood slides were positive for gametocytes divided by the total number of follow-up weeks and expressed per 1,000 person-weeks.All safety variables were analysed in the ITT population.This study was designed as a non-inferiority trial. Assuming 80% statistical power, a one-sided \u03b1 level of 2.5%, and adopting an unequal 2\u22361 randomisation ratio, 1,500 patients  were needed to show that the difference of the day 28 PCR-corrected cure rates between DHA-PQP and AL was within \u22125%, assuming a response rate for AL of at least 92%.Overall, 2,001 patients were screened, and 1,553 recruited and randomised to receive the study drugs  . Five paRandomisation generated comparable groups between countries and overall .DHA-PQP was as efficacious as AL. The day 28 PCR-corrected cure rate was 90.4% (ITT) and 94.7% (ePP) in the DHA-PQP group, and 90.0% (ITT) and 95.3% (ePP) in the AL group . The lower limits of the one-sided 97.5% CIs on the differences between the two treatments were \u22122.80% and \u22122.96%, in the ITT and ePP populations, respectively . The anaThe day 28 PCR-corrected cure rates in infants (6\u201311 months-old) were similar to those in older children and above 90% in both treatment groups .The uncorrected cure rates were significantly higher in the DHA-PQP group, both at day 28  and at day 42 . This was mainly due to fewer late failures later classified as new infections in the DHA-PQP as compared to the AL arm. In the ITT population, new infections until day 42 occurred significantly less in the DHA-PQP group  than in the AL group  . SimilarWhen the day 28 PCR-corrected cure rates were analysed by country, the heterogeneity test was borderline significant at the 10% level only in the ePP population , suggesting some minor differences among sites . HoweverWhen considering the WHO standard definition of TTF, the two treatment groups were similar at day 28 , while at day 42 TTF tended to be lower in the AL group .Parasite clearance was rapid in both treatment groups . About 60% of patients had fever at baseline while at day 2 more than 97% of patients were afebrile in both treatment groups. Gametocyte prevalence at recruitment was similar in both study arms . However, gametocyte carriage measured as rate of person-gametocyte-weeks was significantly higher in the DHA-PQP group than in the AL group, both for the ITT  and the ePP  populations.Haemoglobin changes from baseline to day 28 were comparable between treatment groups (data not shown) while the change from baseline to the last available data was significantly higher in the DHA-PQP group than in the AL group .Both DHA-PQP and AL were well tolerated with the majority of adverse events of mild or moderate severity, and consistent with symptoms attributable to malaria . There wECG was performed in more than 98% of patients at day 0 and day 2, always before the administration of the treatment . In the DHA-PQP group, the proportion of patients with borderline (29.1%) and prolonged (9.1%) QTc interval at day 2 corrected by the Bazett's method was higher than in the AL group (19.8% and 6.9%) (p<0.001). However, this was not confirmed when applying the Fridericia's correction as the corresponding proportions were 1.0% and 0.2% in the DHA-PQP group and 1.2% and 0.2% in the AL group (p\u200a=\u200a0.76). In addition, a \u226560 ms increase of the QTc interval between day 0 and day 2 (Bazett's correction) was observed in just 2.7% (DHA-PQP) and 2.0% (AL) patients; only two patients per group showed a QTc at day 2 higher than 500 ms. When considering the occurrence of the AE \u201cElectrocardiogram QT prolonged\u201d, similar percentages  were observed in the two treatment groups .No other difference between groups was observed during the follow up (data not shown).Two deaths (one per group) occurred during the study. In Uganda, a 3 year-old girl died 24 h after commencing treatment with DHA-PQP. Sepsis or severe malaria was considered by the investigating clinician as the most likely cause. In Mozambique, an 18 month-old girl died 7 h after the first dose of AL. Severe malaria was considered the most likely cause of death, although other aetiologies such as sepsis, hypoglycaemia, heart conditions or bronco-aspiration could not be excluded. No autopsy could be performed in these two children and a causal relationship with the treatment could not be ruled out.in vitro against chloroquine-resistant strains The fulfilment of the non-inferiority criterion on all analysis populations and the confirmation that in this study the comparator treatment had the expected efficacy One hundred twenty nine infants aged 6\u201311 months treated with DHA-PQP responded as well to treatment as older children, though the study was not powered to confirm non-inferiority between the two treatment groups. Infants represent a special group as they are more at risk of malaria and of receiving inadequate doses of antimalarial treatments. In Papua, Indonesia, the PQP plasma concentration at day 7 was the major determinant of the therapeutic response to DHA-PQP Patients treated with DHA-PQP had a significantly higher rate of person-gametocyte-weeks compared with those having received AL. This contrasts with a previous study in Papua, Indonesia, which showed no difference in gametocyte carriage between DHA-PQP and AL Dihydroartemisinin-piperaquine was well tolerated, with few adverse events of clinical relevance. A higher frequency of abdominal pain and diarrhoea has previously been reported for DHA-PQP compared with mefloquine-artesunate It has also been previously reported that the only potentially serious adverse effects of artemisinin derivatives are rare type 1 hypersensitivity reactions In conclusion, DHA-PQP is a safe, efficacious, tolerable and affordable new antimalarial treatment option in Africa. Its longer PTP period may be particularly useful in areas where transmission is intense, though it may exert an important drug pressure on the parasite populations, possibly selecting resistant strains. The deployment of several ACTs as multiple first line treatments may overcome this problem. Indeed, assuming that different treatments are used in equal amounts in the host population, the use of multiple first line therapies would have two main benefits, i.e. the inability of the parasite to adapt to a variable environment and the reduced drug pressure as the rate at which a given treatment is used would be lower than if it was the only one available Protocol S1Trial protocol(0.46 MB DOC)Click here for additional data file.Checklist S1CONSORT checklist(0.36 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum gametocyte carriage in three regions of differing malaria endemicity.The aim of this study was to identify and compare factors associated with Retrospective data from Thailand, The Gambia and Tanzania were used. The data came from large prospective field-based clinical trials, which investigated gametocyte carriage after different anti-malarial drug treatments.Gametocytaemia was detected during the observation period in 12% of patients (931 out of 7548) in Thailand, 34% (683 out of 2020) in The Gambia, and 31% (430 out of 1400) in Tanzania (p < 0.001). Approximately one third  of the patients with gametocytaemia during the observation period, already had patent gametocytaemia at enrolment (day 0 or day 1): 35% (318/931) in Thailand, 37% (250/683) in The Gambia, 26% (112/430) in Tanzania. Maximum gametocytaemia was usually observed on or before the seventh day after starting treatment . Lowest gametocyte carriage rates were observed following treatment with artemisinin derivatives, while sulphadoxine-pyrimethamine (SP) was associated with significantly greater development of gametocytaemia than other drug treatments (p < 0.001). The duration of gametocyte carriage was shorter in Thailand by 86% and Tanzania by 65% than in The Gambia. Gametocyte carriage was 27% longer among people presenting with anaemia, and was shorter in duration among patients who received artemisinin derivatives, by 27% in Thailand and by 71% in Tanzania and The Gambia.This study confirms the independent association of gametocytaemia with anaemia, and the significantly lower prevalence and duration of gametocyte carriage following treatment with an artemisinin derivative. The large differences in gametocyte carriage rates between regions with different levels of malaria transmission suggest that drug interventions to prevent transmission will have different effects in different places. Plasmodium falciparum infections, it remains unclear whether gametocyte production is programmed early on after hepatic schizogony or is a response to stimuli acting upon the parasite population. The proportion of parasites committed to sexual stage development may change during the course of an infection. The developing sexual stages (stages I to IV) remain sequestered in the microvasculature for approximately 10 days before appearing as morphologically distinct male and female stage V gametocytes in the peripheral blood. One male (containing eight microgametes) and one female (macro-gamete) are required per mosquito blood meal (approx 2 \u03bcL) for infection to occur. Thus gametocyte densities of 1 per \u03bcL are theoretically sufficient to infect mosquitoes, a density beneath the limit of detection for most routine microscopy. This explains malaria transmission from subjects without apparent gametocytaemia.Malaria control rests traditionally on two strategies; vector control (reducing the numbers of anopheline vectors and reducing the probability of being bitten), and drug treatment. Effective anti-malarials reduce morbidity, prevent mortality, and reduce asexual parasite biomass. This, in turn, reduces the numbers of sexual (transmissible) parasites (gametocytes). In In areas of low or unstable transmission most malaria infections are transmitted by people who are ill, or recovering from symptomatic malaria. In such areas asymptomatic infections are unusual, so treatment-seeking behaviour and the pharmacokinetic and pharmacodynamic properties of the anti-malarial drugs used are important determinants of transmission. But low transmission settings often contain small foci of higher transmission and the few asymptomatic individuals in these areas are important in sustaining malaria through the dry season. In higher transmission settings the situation is more complex. Anti-disease controlling immunity is acquired which results in an increasing proportion of infections stabilizing at relatively high parasite densities. These may be either asymptomatic or tolerated in older patients who are less likely to seek treatment. These infections may still be transmissible . In addiRecrudescence (treatment failure), and the subsequent extended duration of infection is an important source of transmission particularly of drug-resistant parasite genotypes . The facThe aim of this study was to identify factors associated with gametocyte carriage at different levels of malaria transmission and thus clinical epidemiology. Retrospective data from study sites in three different endemic areas were used. The data come from large prospective field-based clinical trials which investigated gametocyte carriage after different anti-malarial drug treatments.P. falciparum, although Plasmodium malariae and Plasmodium ovale infections also occurred occasionally. Entomological inoculation rates (EIRs) were between 200 and 300 infective bites per person per year, with no seasonal variation. In this region, there were a large number of government dispensaries and chloroquine was widely available in local shops. This study on gametocyte dynamics was part of a larger study investigating natural transmission-blocking immunity in this area of intense perennial transmission  = 0.01 [0.0002\u20130.80]) as compared to those with gametocytes at enrolment (0.05 [0.001\u20132.67]); p < 0.001. In patients who had any parasitaemia assessments between day 0 and day 7, 79% had maximum parasitaemia recorded on day 0. In the others, the ratio between maximum parasitaemia and enrolment parasitaemia was median (90% range) = 2.6 (1\u201350).Overall, 12% (250/2018) of The Gambian patients had gametocytaemia recorded on enrolment compared to 8% (112/1400) in Tanzania and 4% 318/7502) in Thailand  overall developed gametocytes carriage during follow up after treatment with artemisinin derivatives. This proportion varies across sites. It was highest in The Gambia; 15% (96/645) compared with 0% (0/100) and 1% (26/3338) in Tanzania and Thailand. However, it should be noted that 10 patients in Tanzania and 109 in Thailand had gametocytaemia recorded between days 1 and 6 but as their readings were negative on day 7 and 14, they were included in these comparisons as not gametocytaemic. Sulphadoxine-pyrimethamine (SP) was associated with the greatest carriage rates (64% 221/348) and this effect was similar in Tanzania and The Gambia . The corresponding overall rates for developing patent gametocytaemia on day 7 or day 14 after the other treatments were 29% (114/390) following chloroquine, 3% (10/349) following halofantrine, 6% (53/878) after mefloquine, and 22% (34/153) after quinine.P. falciparum infection were associated with increased risk of gametocytaemia. In the multivariate analyses  and Gambian patients with high parasitaemia on enrolment (OR [95%CI] = 1.503 [1.177\u20131.919]) were at increased risk of developing gametocytaemia In all countries, treatment with artemisinin derivatives alone or combined with other drugs resulted in the lowest risk of gametocytaemia. In Tanzania, apart from the treatment, only young age was associated with gametocyte carriage after enrolment. Different sampling schedules prohibited us from combining data from different sites.In the univariate analyses Table , statistes Table , in ThaiOf the 2027 patients with gametocytes before day 28, 12 (0.6%) had three and 164 (8%) had two separated episodes of gametocytes carriage, with a negative count between positive counts. Among a total of 2213 episodes, only 58 (3%) had one missing measurement and 7 (0.3%) episodes had 2 or more missing measurements between the first and last counts. Quality of the follow-up information varied between sites, with 68% censored observations in The Gambia, 61% in Tanzania and 19% in Thailand, although one third of these in The Gambia and one quarter in Tanzania were censored after day 14. The proportion of patients who had missing measurements before the first positive gametocyte count was 7%, 6% and 2% in Tanzania, The Gambia and Thailand.Gametocyte carriage rates (95% CI) after 7 and 14 days were smallest in Thailand (Table In Thailand other risk factors for prolonged carriage included high enrolment parasitaemia (p = 0.005), a prolonged history of illness (p = 0.038) and anaemia (p = 0.034). These associations were not apparent in the other sites.Site, presence of gametocytaemia on enrolment, haematocrit and treatment were all independent predictors of the duration of gametocyte carriage in the multivariate analysis of the combined data from Thailand and The Gambia Table . In ThaiWhen a common model was fitted for all three data sets using common covariates (anaemia could not be included since it was missing for majority of patients in Tanzania), gametocyte carriage in Tanzania was estimated to be shorter by 65% than gametocyte carriage in The Gambia. Effects of gametocytaemia on admission and treatment with artemisinin derivatives in Tanzania were estimated to be the same as in The Gambia.In Thailand, the maximum gametocyte density was increased in patients with gametocytaemia on enrolment (Incidence Rate Ratio (IRR) [95%CI] = 1.261 [1.081\u20131.430], p = 0.001, n = 925), severe anaemia  and a prolonged history of illness (> 3 days) . The two children age groups (< 5 years and 5\u201315 years) had similar levels of maximum gametocytaemia which were significantly higher than in adults . The effects of these covariates were the same for patients presenting with gametocytaemia on enrolment and patients who developed gametocytaemia later, as tested by the interaction term in the model. No covariates were significant in the multivariate analysis.In The Gambia, the only determinant of maximum gametocyte density was drug treatment, but this effect was different for patients presenting with gametocytaemia on enrolment and patients who developed gametocytaemia later . In both cases, patients treated with SP produced higher number of gametocytes than patients treated with artemisinin derivatives .In Tanzania, higher gametocyte densities were found in patients with high parasitaemia on enrolment , females , anaemic patients , and symptomatic patients . Children aged 5\u201315 years  and adults  had significantly lower gametocyte densities compared to children less than 5 years of age (n = 427). In the multivariate model only age  and parasitaemia  remained independent determinants of gametocyte density. Anaemia was not investigated in the multivariate analysis due to the large number of missing values. Since multivariate analyses in each site resulted in models with no common covariates no analysis was performed on the combined data set.The putative patient infectivity was estimated from the serial gametocyte counts. Among patients with gametocytes measurements available on days 0, 7, 14 and 28, all four blood film examinations were negative for gametocytes in 92% 3,643/3,970) of Thai patients, compared to 63% (177/281) of Tanzanian patients and 54% (373/685) of patients from the Gambia; p < 0.001. These patients were excluded from the analysis of infectivity. In nearly all cases (1012/1017) the area under the infectivity curve (AUIC) calculated using Method A  was medi43/3,970 P. falciparum gametocyte carriage. Some of this heterogeneity is explained by known factors such as the differences in age (a proxy for cumulative exposure and thus immunity), levels of anaemia, and type of anti-malarial drug used [This comparison between three malariaous regions with very different epidemiological characteristics detected considerable heterogeneity in rug used . On enroAdmission gametocytaemia was associated independently with lower haematocrit, lower parasitaemia, and lower temperature. Slowing the expansion of asexual parasitaemia may have less effect on the persistence of gametocytaemia, as gametocytes have greater longevity than the asexual stages. In Thailand, where duration of illness was recorded, gametocyte prevalence was associated with a longer period of illness before presentation. In low transmission settings the majority of gametocytaemic individuals will be symptomatic and seek treatment during the acute phase of the illness, and the majority of gametocyte carriage will occur after starting treatment. In the high transmission settings contribution of asymptomatic carriers is significant and it has been demonstrated that carriers with submicroscopic densities of gametocytes are capable of infecting mosquitoes ,9. The dHowever, transmission intensity often varies greatly over small geographic distances, and low transmission settings often contain microfoci of much higher transmission intensity. For example even in Thailand, there is evidence of an untreated reservoir of infectious individuals in some areas , althougIn nearly all patients with gametocytaemia, densities peaked in the first week after starting treatment (day 0\u20137). The day 7 gametocyte density proved a good surrogate for the area under the gametocytaemia-time curve. The factors associated with post-treatment gametocytaemia were similar to those associated with enrolment gametocytaemia. There were large differences between treatment regimens; those containing an artemisinin derivative were associated with lower and shorter carriage, and treatment with SP being associated with higher carriage rates and longer duration. Although there is a relationship between the administration of anti-malarial drugs and gametocyte density, these data cannot be translated directly into transmission potential because of the different effects of the drugs on gametocyte viability ,16,38-40As drug resistant parasites become more prevalent, the duration of malaria infections lengthens and the proportion of recrudescent infections increases. Recrudescent infections are cumulatively of longer duration than primary infections, and are associated with higher rates of anaemia and gametocyte carriage . Drug reThis study has several limitations. Data sets from several studies were combined for the purpose of this analysis. This was a retrospective analysis so inclusion criteria for the trials were not standardized which resulted in the differences in the populations participating in the three sites. For example, in The Gambia only children were enrolled while in the other two sites a broader age spectrum was enrolled. Different covariates were assessed and laboratory methods were not standardized between the sites. Different detection limits for gametocytes affected the estimated densities and rates of carriage. It is also possible that the apparent differences in gametocyte carriage rates, which are dependent on the frequency of observation, are influenced by these differences in trial conduct. Gametocytes counts were collected at slightly different sampling schedules and there was a considerable number of missing values (20\u201340%) at visits scheduled for all three sites. The estimates of infectivity are illustrative, being based on two data-sets which characterized the sigmoid relationship between gametocyte densities and infectivity to an anopheline vector. This hides considerable inter-individual variability and the confounding effects of immunity and other factors. Much more data are needed to characterize and quantify the sources of variance in this assessment and produce more valid assessments necessary to model the impact of interventions on malaria transmission.This study confirms the independent association of increased rates of gametocyte carriage with anaemia ,44, and P. falciparum: Plasmodium falciparum; P. malariae: Plasmodium malariae; P. ovale: Plasmodium ovale; P. vivax: Plasmodium vivax; SP: Sulphadoxine-pyrimethamine.ACT: Artemisinin combination therapy; AUIC: Area under the infectivity curve; AUC: Area under the gametocyte density time curve; EIR: Entomological inoculation rate; GCR: Gametocyte carriage rate; IQR: Inter-quartile range; IRR: Incidence Rate Ratio; K-M: Kaplan-Meier; OR: Odds ratio; CI: Confidence interval; The authors declare that they have no competing interests.KS analysed the data and drafted the manuscript; RNP, CJS, CJD, LS, FN provided the data and contributed to the manuscript writing, NJW contributed to the manuscript writing and conceptually to data analysis."}
+{"text": "Malaria in pregnancy is serious, and drug resistance in Africa is spreading. Drugs have greater risks in pregnancy and determining the safety and efficacy of drugs in pregnancy is therefore a priority. This study set out to determine the efficacy and safety of several antimalarial drugs and combinations in pregnant women with uncomplicated malaria.Pregnant women with non-severe, slide proven, falciparum malaria were randomised to one of 4 regimes: sulfadoxine-pyrimethamine [SP]; chlorproguanil-dapsone [CD]; SP+amodiaquine [SP+AQ] or amodiaquine+artesunate [AQ+AS]. Randomisation was on a 1\u22362\u22362\u22362 ratio. Women were admitted for treatment, and followed at days 7, 14, 21, 28 after the start of treatment, at delivery and 6 weeks after delivery to determine adverse events, clinical and parasitological outcomes. Primary outcome was parasitological failure by day 28.1433 pregnant women were screened, of whom 272 met entry criteria and were randomised; 28 to SP, 81 to CD, 80 to SP+AQ and 83 to AQ+AS. Follow-up to day 28 post treatment was 251/272 (92%), and to 6 weeks following delivery 91%. By day 28 parasitological failure rates were 4/26  in the SP, 18/77  in the CD, 1/73 (1% 95%CI 7\u20130.001) in the SP+AQ and 7/75 (9% 95%CI 4\u201318) in the AQ+AS arms respectively. After correction by molecular markers for reinfection the parasitological failure rates at day 28 were 18% for CD, 1% for SP+AQ and 4.5% for AQ+AS. There were two maternal deaths during the trial. There was no apparent excess of stillbirths or adverse birth outcomes in any arm. Parasitological responses were strikingly better in pregnant women than in children treated with the same drugs at this site.Failure rates with monotherapy were unacceptably high. The two combinations tested were efficacious and appeared safe. It should not be assumed that efficacy in pregnancy is the same as in children.NCT00146731ClinicalTrials.gov  Using an effective antimalarial drug for prevention and treatment is essential. Understandable concerns are raised however by using any new drug in pregnancy; older drugs have a better known safety profile in pregnancy, but are likely over time to become less effective due to the emergence and spread of drug resistance. Although there are no human data to suggest that artemisinins are teratogenic, animal data for use of artemisinins early in pregnancy have raised concerns of teratogenicity Sulfadoxine-pyrimathamine (SP) has been assessed for safety in pregnancy, is now recommended for intermittent preventive treatment in pregnancy (IPTp), and remained Tanzanian national policy for treatment until November 2006 and for Intermittent Preventive Therapy in pregnancy in most countries (IPTp) to date Despite concerns from animal studies both artesunate and artemether have been given to many pregnant women (often inadvertently), and current published data demonstrate no evidence of human teratogenicity The present study compared the efficacy, tolerability and safety of standard SP-monotherapy to CD that had been registered in UK at the design of the study and to two drug combinations for which good efficacy data was available from East Africa. AQ+SP has proved effective both in Ugandan and Tanzanian children 21. Throughout the study period SP (defined as monotherapy for the purposes of this paper) was national first-line treatment for malaria, and this was taken as the comparator arm for this study.This open-label study was conducted among pregnant women who attended Muheza Designated District Hospital (Muheza DDH). The lowlands of Muheza district experience hyperendemic to holoendemic malaria. The day 28 parasitological failure rates to AQ monotherapy in a recent effectiveness trial in children under 5 years was 76%, with the comparative rates of 61% and 40% for AQ+SP and AQ+ASPregnant women with mild-moderate, slide proven, falciparum malaria were recruited from the Antenatal wing (ANC) of the Reproductive and Child Health (RCH) clinic at Muheza Designated District Hospital. Pregnant women from Muheza Township and surrounding villages attend this clinic for their medical care. Nurses at the RCH identified all febrile pregnant women with a fever or recent history of fever (within 48 hours), symptoms compatible with anaemia or malaria and referred them to the study team. All referrals were re-interviewed and examined by a medical officer from the study team to exclude concomitant infection(s). Duplicate thick and thin blood smears were Giemsa stained and examined microscopically for malaria parasites.either a positive blood smear for P.falciparum with at least 800 asexual parasites/\u00b5L in an asymptomatic woman or any of the following symptoms within 2 days prior to consultation; history of fever; headache, vomiting, chills/rigors, and/or any of the following signs: temperature \u226537.5\u00b0C and <39.5\u00b0C, Hb\u22657 and <9 g/dl) together with P.falciparum parasitaemia at any density. Additionally, all cases had to be 14\u201334 gestation weeks pregnant on the day of attending the clinic, have a viable foetus defined by the presence of foetal heartbeat by sonicaid or pinnard, able to take drugs orally, able to attend follow up clinic, and gave written informed consent to participate or a finger-print witnessed consent for women unable to read.Inclusion criteria were pregnancy with Plasmodium or major protocol violation.The main exclusion criteria were; severe and complicated forms of malaria Women who met the inclusion criteria were randomised to one of 4 regimes: three tablets of sulfadoxine-pyrimethamine (500 mg sulfadoxine/25 mg pyrimethamine per tablet) orally at once [SP] in line with the national policy; chlorproguanil-dapsone (1.2 mg/kg and 2.4 mg/kg respectively for 3 days) [CD]; SP 3 tablets once+amodiaquine (10 mg/kg for 3 days) [SP+AQ]; amodiaquine (10 mg/kg for 3 days)+artesunate (4 mg/kg for 3 days) [AQ+AS]. Randomisation was on a 1\u22362\u22362\u22362 ratio for SP, CD, SP+AQ and AQ+AS to maximise information about the drugs whose use in pregnancy is less known; it was assumed the difference in outcome would be greatest for SP compared to other arms so the size of this arm could be smaller. Randomisation was in blocks of random sizes, and conducted in London using Stata 7. Treatment allocations were placed in a sealed opaque envelope, with pregnant women picking their own envelope. Patients were allocated a study number sequentially, and after consenting to participate, participants picked an envelope in front of the attending clinician. Opening the envelope constituted entry to the trial and analysis was conducted on that basis .All women were admitted to a ward dedicated to research for the first three days to facilitate supervised drug administration and to monitor clinical response and adverse events. Drugs were administered by study nurse-midwives employed by the project. After each administration, the patient was observed for 45\u201360 minutes. The dose was repeated if vomiting occurred within the observation period. Vomiting the second dose was registered as an adverse event and led to withdrawal from the study. Such cases were treated with parenteral quinine. Patients were treated for symptoms with standard medications e.g. paracetamol for fever. Women continued to receive routine antenatal medicaments of iron supplements, folic acid (5 mg), and tetanus toxoid (TT) given by the RCH. In addition to daily clinical observations and laboratory tests, foetal viability  was monitored daily during admission using a Doppler machine, and at each follow-up visit.Adverse events were classified by severity and potential causal relationship to study drugs. All serious adverse events (SAEs) were independently investigated by a local safety monitor. Subjects with an AE were followed up until the condition had disappeared or stabilized.Parasite counts on Giemsa-stained blood films were performed daily during admission, repeated on days 7, 14, 21 and 28, and on any other day(s) of complaints. Counts were made against 200 white blood cells (WBC) on a thick blood smear. All slides had a second reading done in an independent research laboratory. Discordant results were read by a third reader, with the majority taken as the definitive outcome. All microscopists were blind to treatment allocation. A separate read for gametocytes was undertaken counting against 500 white blood cells. To quantify the effect of treatment on gametocyte carriage, we determined the area under the curve (AUC) of gametocyte density over time which incorporates both the magnitude and the duration of gametocyte carriage Blood samples for haematology and clinical chemistry, and in anaemic women stool microscopy for intestinal helminths were obtained on admission. Blood and urine samples were repeated at day 3 and where indicated at day 7. Haemoglobin, total and differential white blood cell count, platelet count, creatinine, total bilirubin, alanine aminotransferase (ALT) and albumin were measured (using CBC machine for haemogram and Reflectron for biochemistry) on days 0, 3 and 7, and whenever else indicated. Pre-test counselling for HIV-testing was undertaken, and where consent was given an HIV test was performed. HIV-positive mothers were referred to the HIV care unit of Muheza DDH for counselling and for consideration of antiretroviral drugs.On discharge from the ward, patients were followed up on days 7, 14, 21 and 28 post initiation of treatment and at any time they felt unwell before day 28. At the end of each clinic, members of the study team followed all non-attendees to their homes to establish and record reasons for non-attendance and to collect a blood smear. Patients with either early or late treatment failure following treatment with any of the 4 study regimens were treated with quinine 10 mg/kg 8 hourly for 7 days as rescue therapy.Birth outcome and Dubowitz assessment were recorded for all deliveries taking place at the hospital. Mothers and their newborns attended follow up clinics 6 weeks after delivery. All non-attendees (whether delivered at Muheza DDH or not) were followed up at home. Assessment at this time included a further check of the newborn by the paediatrician for any abnormality that may have been missed at birth or for any serious problem that may occur after birth such as kernicterus. Whenever the mother had moved from the study area, all possible efforts were made to ascertain birth outcome verbally from close relatives. A DSMB reviewed all SAEs, which were notified as they occurred.Blood for PCR was collected on glass-fibre membranes from all patients at enrolment and at each follow-up. The polymorphic repetitive regions were amplified by nested-PCR for block 3 of msp2 The primary end-point of the trial was parasitological failure by day 28. This was defined as any of: a need for rescue treatment due to clinical deterioration defined by altered sensorium, seizures, persistent vomiting, renal impairment, respiratory distress, a fall in Hb below 7 g/dl, or in cases where the initial haemoglobin dropped 20% or more from baseline Hb, at any time during admission; persistence of fever with parasitaemia on day 3; increased parasite density on day 2 or 3 compared with baseline density; failure to clear parasites on day 7; rescue medication for recurrent malaria before day 28; slide parasite positivity at day 14, 21 or 28.Major secondary endpoints were: clinical failure by day 28 , parasitological or clinical failure by day 14, incidence of foetal death during treatment, defined as absence of foetal heartbeat assessed by Doppler; change in haemoglobin from baseline on day 14; incidence of perinatal and neonatal mortality, assessed 4\u20136 weeks after due date of delivery; clinically apparent neonatal abnormality 4\u20136 weeks after due date of delivery; preterm delivery and other adverse events during treatment.Initially the study was powered to detect a 4 fold difference in treatment failure between SP+amodiaquine and amodiaquine+artesuante groups (8% vs 2%) with 95% precision and 80% power, which would require a samples size of 80 women in the SP+placebo group and 240 women in each of the other three groups. Vigorous measures to protect pregnant women in the district from malaria on a general background of reduced transmission of malaria in this area fortunately led to substantial reductions in the number attending the antenatal clinic with clinical malaria. The data from this and other sites was reviewed on October 2004 and it was decided that given the absence of other data, the question was important enough and of sufficient public health priority that a trial able to detect a larger difference would still be of public health importance. A revised sample size was calculated to detect a difference from 1% (the best likely failure rate in any arm) and 15% (above which no drug could be deployed). This gave a sample size of 72 in each arm when \u03b1 was .05 and \u03b2 0.8. The unbalanced sample size (1\u22362\u22362\u22362) was by this time established and could not be revised retrospectively although the statistical rationale for it was not present with the revised design.Data were double entered into Microsoft Access, and analysed using Stata 8. The analytical plan was finalised before the analysis was undertaken. For primary and major secondary outcomes, proportions with confidence intervals were calculated. Odds ratios were calculated for the difference between all arm and the best and worst arms for parasitological failure unadjusted, and adjusted for the predefined risk factors age, parity, HIV serostatus, initial parasitaemia and initial haemoglobin.Ethical permission was granted by the ethics committees of the National Institute for Medical Research, Tanzania, and the London School of Hygiene & Tropical Medicine and conducted in accordance with the Declaration of Helsinki. All participants gave written informed consent (or witnessed where whey could not read). The trial was monitored by an independent external clinical monitor and was prospectively registered on ClinicalTrials.gov No. NCT00146731. The protocol for this trial and supporting CONSORT checklist are available as supporting information: see The trial ran from Jan 2004\u2013Sept 2006. 1433 pregnant women were screened, of whom 272 were enrolled, 28 to the SP, 81 to the CD, 80 to the SP+AQ and 83 to the AQ+AS arms respectively. The slight variation from the planned 1\u22362\u22362\u22362 randomisation was due to random variation in the smallest arm because the trial did not reach the originally planned sample size on which the randomisation was based. Reasons for exclusion and flow through the trial are outlined in By day 14, the parasitological failure rates (including both symptomatic and asymptomatic cases) were 1/24 (4%) in the SP, 1/78 (1.3%) in the CD, 0/71 (0%) in the SP+AQ and 0/77 (0%) in the AQ+AS arms respectively. By day 28 the equivalent parasitological failure rates were 4/26  in the SP, 18/77  in the CD, 1/73 (1% 95%CI 0.001\u20137) in the SP+AQ and 7/75 (9% 95%CI 4\u201318) in the AQ+AS arms respectively. After correction by molecular markers for reinfection, the parasitological failure rates at day 28 were 18% for CD, 1% for SP+AQ and 4.5% for AQ+AS; numbers in the SP arm were considered too small to be reliable. Full data are shown in At day 14, restricting data to patients with no gametocytes at baseline, 5/16 women (31%) in the SP arm, 12/50 (24%) in the CD arm, 5/54 (9%) in the SP+AQ arm and 3/56 (5%) in the AQ+AS arm were gametocytaemic . The oddThere were two maternal deaths during the trial. One woman in the CD arm had mild malaria both clinically and parasitologically when she entered the trial, but developed hyperparasitaemia (>20% parasitaemia) and severe malaria over 48 hours. Review of initial blood films showed that the initial parasite count was correct, but all parasites were synchronous pre-schizonts. She came from a mountain area with little malaria transmission. The second woman in the SP+AQ arm made an initial response but then deteriorated despite clearing her parasites. Consent for determining her HIV serostatus was not given, but other clinical factors suggest it is likely she died from an immunosupression related illness. In neither case was the direct effect of study drugs thought likely to have been the cause, although CD may have failed to stop progression of severe disease in the first case. No other maternal SAEs were recorded; non-severe maternal adverse events are recorded in No foetal deaths occurred within 28 days of administration of drugs except in the two women who died. There was one macerated stillbirth in the AQ+AS arm. Other adverse birth outcomes, largely relating to complications related to asphyxia are shown in Balancing the risk-benefit of antimalarial drugs in pregnancy is not easy 22. It is possible that the dose of chlorproguanil that was used in CD was not high enough to attain adequate therapeutic levels in pregnancy. It was one of two generally used dosing regimens, the other being one based on 2 mg/kg chlorproguanil per day. The lower dose was chosen because of fear of possible risk of drugs in pregnancy, and a higher dose might have proved more effective CD had a similar failure rate to SP, a difference from the results of a similar study conducted in children under five years of age in the same area 5 years ago, where CD treated malaria that had failed to respond to SPIt is encouraging, as seen in As in children, the artemisinin-based combinations had a far greater impact on gametocyte carriage than non-artemisinin combinations, even when (as was the case with SP+AQ) the drug combination was itself highly efficacious. The relative immunity which is the likely cause of the different impact on efficacy between pregnant adults and children does not seem so marked for gametocytes. This is potentially important when considering the likely impact of ACTs on transmission, since a substantial proportion of the transmission of malaria is from adults.Tanzania has a moderate prevalence of HIV in pregnancy, but unfortunately the numbers in this trial are not big enough to answer the question as to which drugs are likely to be most appropriate in HIV-infected pregnant women. There is a complex interaction between HIV and malaria in pregnancy; HIV both increases the risks of side-effects of some drugs and reduces the efficacy of antimalarials This study adds to the existing data, mostly from studies in Southeast Asia, demonstrating no evidence of teratogenicity when artemisinins are used in the last two trimesters. This is reassuring now that ACTs are being rolled out. It cannot settle the question about the safety of artemisinins early in the first trimester, which is the period that has raised most concern on safety in animal studies. There is no evidence from human studies that artemisinins are teratogenic, but data is still too sparse to rule this out. It may be that non-artemisinin combinations, especially for IPTp, remain a sensible option in some settings for the interim. The investigators decided at the outset of the trial not to study artemether-lumefantrine (Coartem), despite being an excellent antimalarial, because there was no data on safety in African pregnant women. One study conducted in Thailand indicated that the pharmacokinetics of Coartem is deranged in pregnancy The major limitations of the study are the fact that the size was smaller than anticipated, and that women without typical symptoms of malaria are likely to be under-represented (as they do not present), and women with placental malaria but no peripheral parasites by definition cannot be included. Bias is unlikely to be a major issue as this is a randomised trial, although in smaller trials important random variations between arms can occur.Despite the fact that the sample size is small for definitive conclusions on safety, it is reassuring both that newer drugs and combinations, including the artemisinin combinations, are tolerable and efficacious in pregnant women in East Africa. In this area, monotherapy with SP or CD for malaria in the last two trimesters of pregnancy, whilst it has antimalarial parasitological failure rates substantially lower than in children, is unacceptably high and should be abandoned. The risks of malaria in pregnancy are too great to continue to use drugs with appreciable parasitological failure rates.CONSORT S1CONSORT Checklist(0.06 MB DOC)Click here for additional data file.Protocol S1Trial Protocol(0.16 MB PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum is common in many endemic and other settings but there is no clear recommendation on when to change therapy when there is delay in parasite clearance after initiation of therapy in African children.Drug resistance in The factors contributing to delay in parasite clearance, defined as a clearance time > 2 d, in falciparum malaria were characterized in 2,752 prospectively studied children treated with anti-malarial drugs between 1996 and 2008.pfmdr 1/pfcrt mutants. Delay in clearance significantly increased gametocyte carriage (P < 0.0001).1,237 of 2,752 children (45%) had delay in parasite clearance. Overall 211 children (17%) with delay in clearance subsequently failed therapy and they constituted 72% of those who had drug failure, i.e., 211 of 291 children. The following were independent risk factors for delay in parasite clearance at enrolment: age less than or equal to 2 years , presence of fever , parasitaemia >50,000/ul , and enrolment before year 2000 . Following treatment, a body temperature \u2265 38\u00b0C and parasitaemia > 20000/\u03bcl a day after treatment began, were independent risk factors for delay in clearance. Non-artemisinin monotherapies were associated with delay in clearance and treatment failures, and in those treated with chloroquine or amodiaquine, with Delay in parasite clearance is multifactorial, is related to drug resistance and treatment failure in uncomplicated malaria and has implications for malaria control efforts in sub-Saharan Africa. Plasmodium falciparum is a major obstacle to successful chemotherapeutic control of the disease. Resistance to chloroquine (CQ) and sulphadoxine-pyrimethamine (SP) is now widespread in sub-Saharan Africa, South Asia and South America [P. falciparum multidrug resistance gene 1 (Pfmdr 1) in asexual and sexual parasites following treatment of infections with artemether-lumefantrine (AL) [The emergence and spread of multidrug resistance in  America ,2 and th America -6, may s America -9 and inine (AL) , there iP. falciparum to CQ and SP deteriorated steadily over the past ten years.Despite increasing drug treatment failure, there is no clear guidelines, at least in Nigeria, about the time to change anti-malarial drug treatment if parasites do not clear quickly from peripheral blood following treatment of uncomplicated acute infections in African children. It is postulated in the present study that, parasite clearance exceeding two days is associated with risk of treatment failure and resistance and could be used as a criterion to change therapy in very young children. The present study reports the relationship between delay in parasite clearance and anti-malarial treatment failure in children with falciparum malaria in an area of intense transmission in south-western Nigeria, where resistance in The studies took place between July 1996 and July 2008 in patients presenting at the University College Hospital in Ibadan, a hyperendemic area for malaria in south-western Nigeria . EthicalP. falciparum, parasitaemia \u2265 2,000 asexual forms/\u03bcl blood, negative urine tests for anti-malarial drugs 4-aminoquinolines and sulphonamides , absence of concomitant illness, no evidence of severe malaria [Briefly, children with symptoms compatible with acute falciparum malaria who fulfilled the following criteria were enlisted in the study: age 144 months or below, mono-infection with  malaria , and wriThick and thin blood films prepared from a finger prick were Giemsa-stained and were examined by light microscopy under an oil-immersion objective, at \u00d7 1,000 magnification, by two independent assessors. Parasitaemia in thick films was estimated by counting asexual parasites relative to 1,000 leukocytes, or 500 asexual forms, whichever occurred first. From this figure, the parasite density was calculated assuming a leukocyte count of 6,000/\u03bcl of blood. Gametocytes were also counted in thick blood films against 1,000 leukocytes assuming an average leukocyte count of 6,000/\u03bcl of blood -20. HaemResponse to drug treatment was assessed using World Health Organization (WHO) criteria  as folloPfmdr 1 and pfcrt genotypes were assessed by PCR using methods described previously [pfmdr 1 and pfcrt were detected by nested PCR-RFLP methods.At enrolment, 100 \u03bcl of capillary blood was collected on to 3 MM Whatman\u2122 filter paper for resistance markers determination in 2000 and 2006 in those treated with CQ or AQ. eviously ,10. Sing2 with Yates' correction. Normally distributed, continuous data were compared by Student's t-tests and analysis of variance (ANOVA). Data not conforming to a normal distribution were compared by the Mann-Whitney U-test and the Kruskal-Wallis test (or by Wilcoxon rank sum test). A multiple logistic regression model was used to test the association between parasite clearance > 2 d (yes or no at presentation or during follow up) and factors that were significant at univariate analysis: age, presence of fever, asexual parasitaemia at presentation or during follow-up, a history of vomiting, and drug treatment. Because the study was conducted over a period of 12 years, time in years since the commencement of trials was included as a covariate in the model for pretreatment delay in parasite clearance. P-values of < 0.05 were taken to indicate significant differences.Data were analysed using version 6 of the Epi-Info software , and theP. falciparum. There were 1,716 under five-year olds. The geometric mean parasitaemia at enrolment was 34,044/\u03bcl   were enrolled into the drug studies. All were recruited into prospective randomized studies and all had primary infections with 2 for trend = 155.8, P < 0.0001).Overall 57 of the 2,752 children (2.1%) had treatment failure by day 7, and this rose to 113/2,695 (4.2%) by day 14, and 62/796 (7.8%) by day 21 . There was no significant difference in the proportions of children with delay in clearance in those treated with AS (3 of 120), AQAS (14 of 142), AL (6 of 90), and AMQ (10 of 174) . The addition of AS to MQ (AMQ) significantly reduced the proportion of patients with delay in clearance . Similarly, the addition of AS to AQ (AQAS) significantly reduced the proportion of patients with delay in clearance .Overall, delay in parasite clearance occurred in 1237 of the 2752 children (45%)  in those treated with CQ  and the proportions of children with delay in clearance between 1996 and 2004, or between 2000 and 2006, in patients treated with CQ (n = 316), SP (n = 218), or AQ (n = 573) were evaluated. There was a significant increase in PCT from 3.08 \u00b1 1.06 d (n = 53) in 1996 to 4.23 \u00b1 2.08 d (n = 76) in 2004 (P < 0.0001), but there was no difference in proportion with delay in parasite clearance during the same period  in those treated with SP  in 1996 to 4.21 \u00b1 1.84 d (n = 71) in 2004 (P = 0.0001) and a significant increase in proportion with delay in parasite clearance during the same period  in those treated with AQ  in 2000 to 2.87 \u00b1 1.39 d (n = 118) in 2006 (P = 0.0001), but there was no difference in proportion with delay in parasite clearance during the same period  to 2003 (20% 15/72), \u03c72 for trend = 0.075, P = 0.78. In those treated with AQ gametocyte carriage at enrolment also did not increase from 2000  to 2006 (13% 16/120), \u03c72 for trend = 0.55, P = 0.46.Overall 10%  of the children had patent gametocytaemia on presentation. In those treated with CQ gametocyte carriage at enrolment did not increase from 1996  to 2003 (22% 14/62), \u03c72 = 44.69, P < 0.0001) . In those who did not carry gametocyte at enrolment, gametocyte carriage within two weeks of commencing therapy was related to parasite clearance time; it rose from 13%  among children who cleared their parasitaemia on day 1 or on day 2 to 24.5%  among those who cleared their parasitaemia on day 3 or on day 4 , 36% (51/139) and 13% (15/139), respectively in 2000. In 2006, the corresponding prevalence was 50% (59/118), 36% (42/118) and 14% (17/118). In the 256 children, 13% (18/129), 4% (4/93), and 5% (2/34) with mutant, mixed, and wild alleles, respectively were treatment failures. The difference between these proportions was significant . The proportions of 3 to < 5 year-olds with parasite clearance > 2 d in these groups were: 75% (48/64), 51% (28/54) and 63% (12/19), respectively for mutants, mixed and wild type. The difference in these proportions was significant .A total of 260 isolates from 260 children were selected for genetic analysis of P. falciparum, in particular to CQ and SP, in south-western Nigeria has been relatively slow but steady [Pfmdr-1 gene following anti-malarial treatment [The emergence of drug resistance in t steady -14,25,26reatment ,10. The reatment  should hreatment . The latResistant infections at case management and community levels are difficult to manage and early identification and prompt treatment of patients at risk of subsequent treatment failure would improve patient care and community management of resistant infections. In the present study, a systematic evaluation of the pre-treatment and during-treatment factors contributing to delay in parasite clearance, the relationship between delay in parasite clearance and treatment failure, and effects of non-artemisinin and artemisinin mono- or combination therapy on parasite clearance in children were done. The results showed that delay in parasite clearance is multifactorial in origin with the host, parasite and drug factors contributing almost equally to delay in clearance and subsequent failure of treatment in those with delay in parasite clearance.The age-and pretreatment parasite population size- dependent delay in parasite clearance are likely linked to treatment outcome by affecting both the immune-dependent ability to clear and the probability of survival of a subpopulation of asexual blood parasite stages, respectively. In cases of polyclonal infections, a common finding in this endemic area ,29,30, tInterestingly, pre-treatment and intra-treatment elevated body temperatures were independent risk factors for delay in clearance. Fever is considered a crude measure of unspecific immune responsiveness . If the In general, progressive and significant elongations in parasite clearance times over time were striking features of treatment outcomes in children treated with CQ, and SP, the two most widely and longest used drugs in south-western Nigeria, and for AQ, a drug that is less frequently used, but is a partner drug for artesunate. These elongations were not accompanied by significant increases in proportions of patients with delay in parasite clearance during the same periods in those treated with CQ and AQ, suggesting similar mechanism(s) for development of resistance in the parasite to both drugs. In addition, the absence of increase in proportion with delay in clearance over time could have been due to the fact that these proportions were already above 50% when the studies began in 1996 and 2000, respectively for CQ and AQ. This would suggest that the 50% threshold may be the time to consider change in treatment policy at the community or national level in this endemic area.Non-artemisinin monotherapy and non-artemisinin combination therapy were associated with significantly increased proportions of children with delay in clearance. These findings call for a new strategy to determine the maximum point in time at which to effect change in therapy in policy formulation with the widespread use of ACT in Africa. Perhaps this time point may be when 50% of a population of under-three to under-five year olds show delay in parasite clearance. This is best exemplified by the results from the children treated with amodiaquine, where over 50% showed a delay in clearance and 65% of those who were treatment failures were those with demonstrable delay in clearance. These findings contrast sharply with those found in children treated with artesunate and artemisinin-based combination therapy and strongly support the WHO recommendation of ACT  as firstDelay in parasite clearance was associated with increased gametocyte carriage and, therefore, presumably with a potential for increased transmissibility of drug resistant phenotype. This is of public health import since delay in parasite clearance, by virtually all anti-malarial drugs, including ACT, is associated with an increased risk of gametocytaemia -38.pfmdr1/pfcrt mutants and mixed mutants/wild was above 50%, and with little or no increase between 2000 and 2006. Intriguingly, this was associated with increase in parasite clearance time in CQ and AQ-treated patients. Carriage of mutant parasites was associated with an increased risk of treatment failure. Additionally, the proportions of under five-year-olds with delay in parasite clearance increased with genotype classification suggesting that delay in parasite clearance was associated with drug resistance and treatment failure in young children. However, there are other factors that may affect susceptibility that were not evaluated in these children.Over the study period the prevalence of There is need to justify the definition of delay in clearance used in this study: one asexual cycle of the blood stage infection takes approximately 48 h. Thus after two days, the parasites appearing in blood and visible by microscopy when the patient presents, should be the one to appear in blood 48 h after treatment began if the drug does not kill all the parasites . Thus thA better understanding of the multifactorial causes and mechanisms of delay in parasite clearance following anti-malarial treatment regimens can provide vital information for setting policy recommendations at case management, community, and national levels and prolonging the clinical life of the currently useful anti-malarials.The authors declare that they have no competing interests.AS led the design, conduct, data analysis and manuscript preparation. EOA was involved in manuscript preparation. GOG and CTH were involved in design, conduct, and preparation of the manuscript. AS was involved with data analysis. OAF, TMO and OSM were involved with conduct of study. All authors read and approved the manuscript."}
+{"text": "The use of anti-malarial drug combinations with artemisinin or with one of its derivatives is now widely recommended to overcome drug resistance in falciparum as well as vivax malaria. The fixed oral dose artemisinin-naphthoquine combination  is a newer artemisinin-based combination (ACT) therapy undergoing clinical assessment. A study was undertaken to assess the safety, efficacy and tolerability of ANQ combination in areas of multi-drug resistance to generate preliminary baseline data in adult population of Papua New Guinea.The clinical assessment was an open-labeled, two-arm, randomized study comparing ANQ combination as a single dose regimen and three days regimen (10 mg/kg/day) of chloroquine plus single dose sulphadoxine-pyrimethamine (CQ+SP) for the treatment of uncomplicated falciparum malaria with 28 days follow-up in an adult population. The primary outcome measures for efficacy were day 1, 2, 3 7, 14 and 28-day cure rates. Secondary outcomes included parasite clearance time, fever clearance time, and gametocyte carriage. The main outcome measures for safety were incidences of post-treatment clinical and laboratory adverse events.P. falciparum were randomly assigned to receive ANQ and CQ+SP, only 100 patients (51 in ANQ group and 49 in CQ+SP group) were evaluated for clinical and parasitological outcomes. All the patients treated with ANQ and CQ+SP showed adequate clinical and parasitological response with 28 days follow-up. The cure rate for ANQ on day 1, 2, 3, 7, 14, and 28 was 47%, 86%, 92%, 94%, 94% and 94%, respectively. Recrudescence account for 6%; all were cleared on day 21. For CQ+SP treated group the cure rates were 24%, 67%, 82%, 82%, 84% and 88%, respectively. Recrudescence accounted for 10%; all were cleared on day 28 except for one patient. Both regimens were well tolerated with no serious adverse events. The proportion of gametocyte carriers was higher in CQ+SP treated group than ANQ treatment .Between June 2005 and July 2006, 130 patients with confirmed uncomplicated P. falciparum malaria in the adult population of Papua New Guinea and deserves further clinical evaluation.While these data are not themselves sufficient, it strongly suggests that the ANQ combination as a single dose administration is safe and effective for the treatment of uncomplicated  Plasmodium falciparum  and Plasmodium vivax (P. vivax)  for adults/amodiaquine for children) have been used as the first-line anti-malarial drugs for malaria treatment for many years. However, the therapeutic efficacy of the 4-aminoquinolines, particularly of chloroquine, has declined over the years ,6 since nt genes -12, whicnt genes -15. Althnt genes ,17, PNG nt genes . The ANQnt genes ,20. The P. falciparum and P. vivax infections with high prevalence of multi-drug resistant P. falciparum are experienced in both regions [P. falciparum infection (ii) age 14 years and over, (iii) agreed to 28-day follow-up, (iv) parasitaemia of 500 parasites/\u03bcL and over (later it was changed to \u2265 250 parasites/\u03bcL due to slow in recruiting patients), and (v) signed informed consent form. The exclusion criteria were: (i) danger signs of severe malaria, (ii) pregnancy, (iii) history of allergy to study drugs, (iv) taken anti-malarial drugs in the last seven days, and (v) medical history or concurrent systemic medical conditions. The exiting criteria from the study were: (i) completion of the 28-day period, (ii) identification of mixed species infection in the course of study, (iii) development of serious drug related toxicity, (iv) progression of signs and symptoms indicative of impending severity of malaria, and (v) developed non-malaria infections during the course of the study. The study was designed as an open-label, two-arm, randomized study comparing a single dose ANQ , China) with three days CQ and single dose SP (CQ+SP), the current first line treatment for malaria in PNG. The Research Committee of the School of Medicine and Health Sciences, University of Papua New Guinea, and the Medical Research Advisory Committee (MRAC) of the PNG National Department of Health (MRAC #:1529) gave the ethical approval for the study.Three hospitals were selected for the study; two in Port Moresby, southern region, and Madang in the northern region Figure . Moderat regions -6,21. ThThe trial patients were admitted to the hospital for the first three days (72-hrs). On admission (day 0), a medical history and a full physical examination were performed on all patients. Venous blood (~10 ml) was collected for biochemistry, haematology, and renal function tests. Blood smears for parasite microscopy were done on 0, 8, 16, 24, 32, 40, 48, 60 and 72 hrs before patients were discharged from hospital. Thereafter patients were asked to return to the hospital on days 7, 14, 21, and 28. On each visit a full physical examination, blood smears and collection of venous blood (5 ml) were done for each patient; blood smears for parasite microscopy and venous blood for biochemistry, haematology, and renal function tests. In addition, a questionnaire on adverse events was completed for each patient. Any patients complaining of fever or symptoms consistent with malaria, a blood smear was taken and microscopically examined during the follow-up period.The malaria parasite counts were determined on 10% Giemsa stained thick blood smears under high power magnification (10 \u00d7 100). Parasitaemia was measured by counting a number of asexual parasites against 200 white blood cells (WBC). However, where parasites decreased following treatment such that parasites may fall below 10 parasites per 200 white blood cells, count was done against 500 WBC. The parasitaemia (parasites/\u03bcl of blood) was calculated based on the number of asexual forms seen in 200 WBC multiplied by 8000 . The bloGeigy Scientific Tables [The trial patients were randomized to either of the two treatments using a table of random numbers adopted from c Tables . PatientA & FCTB), FCTA defined as time taken for the temperature to fall for the first time below 37.5\u00b0C from first dose of the medication; FCTB defined as time taken to become and remain afebrile during the period of the study; and proportion of patients who had gametocytes post-treatment.The primary outcome measured for efficacy was cure rates (the proportion of cases of true treatment success judged by two negative blood smears on two consecutive occasions without recrudescence parasitaemia) on day 1, 2, 3, 7, 14, and 28. Recrudescence was defined as reappearance of peripheral parasitaemia after two or more consecutive negative blood smears. The secondary measures were, parasite clearance time (PCT) defined as time taken from the first dose of the medication to the first of two successive negative blood smears; fever clearance times  were calculated for each patient's parasitaemia. Mean parasitaemia values were converted to percentage, baseline parasitaemia being 100%. Using Excel database and SPSS software the percentages of mean parasitaemia were plotted against time to assess the response pattern. Student's P. vivax, five patients excluded for violation of the randomization order by one physician, one patient had a history of psychiatric illness, one patient had renal impairment, one patient was found be pregnant, two patients absconded after admission before treatment commenced. One hundred (n = 100) patients fulfilled the criteria for final analysis; fifty-one (n = 51) patients in ANQ group and forty-nine (n = 49) in the CQ+SP group. Adherence to the protocol was generally good: around 94% (n = 94/100) completed follow-up; 6% (n = 6) patients, two (n = 2) patients failed follow-up investigation on day 21 in ANQ group and four (n = 4) failed follow-up investigations on 28 in the CQ+SP group.One hundred and thirty (n = 130) patients were recruited into the study with positive blood smears for malaria parasites between June 2005 and July 2006. Thirty (n = 30) patients were excluded for the following reasons: 10 patients had The baseline characteristics were similar in both treatment groups Table . The patA was 13.6 \u00b1 9.6 hr in ANQ group versus 17.2 \u00b1 9.9 hr in CQ+SP (p > 0.05). The mean FCTB was 19.8 \u00b1 12.7 hr in ANQ group versus 31.7 \u00b1 21.5 hr in CQ+SP group (p > 0.05).Only 76 patients of 100 patients were evaluated for FCT; remaining 24 patients had a temperature <37.5\u00b0C on admission. However, within 24 hrs, 50% of the afebrile patients in each group developed fever (\u226537.5\u00b0C). The general profile of temperature resolution in both treatment groups is shown in Figure The cure rates for ANQ on day 1, 2, 3, 7, 14, and 28 were 47%, 86%, 92%, 94%, 94% and 94%, respectively. Recrudescence account for 6% (n = 3/51) and the recrudescence parasitaemia cleared in all on day 21. For CQ+SP treated group, the cure rates were 24%, 67%, 82%, 82%, 84% and 88%, respectively. Recrudescence accounted for 10% (n = 5/49) and the recrudescence parasitaemia cleared in all on day 28. One patient  in CQ+SP group remained parasitaemic throughout the study period. Figure The parasite clearance profile for the first 72 hrs is shown in Figure Pre-treatment gametocytaemia was observed in 6 patients . After the start of the two treatments, 12% (n = 6/51) of the patients in ANQ treated group developed gametocytes versus 41% (n = 20/49) in CQ+SP group (p < 0.05). The post-treatment gametocyte density was not evaluated as this was not part of the study design.Both treatment regimens were well tolerated with no serious adverse events being reported or detected. Post-treatment emergent symptoms that worth noting were transient deafness , itchiness , skin rash , and dark urine . The rate of vomiting after initiating treatment was low  and did not differ greatly between the two treatment groups. Several patients reported treatment emergent symptoms such as anorexia, nausea, dizziness, abdominal pain and headache within 24-hrs of admission. However, it was difficult to discern which of the symptoms was malaria related and which were due to drug treatment. One of the patients developed psychosis ~5 hours after oral administration of ANQ. The patient had low plasma glucose , which was corrected with dextrose saline. The patient underwent psychiatric assessment (by a psychiatrist) and organic causes were excluded. The patient's condition during post-treatment follow-up was uneventful. Hypoglycaemia rather than ANQ medication could have been responsible for the psychosis.Both treatments in this study cleared parasitaemia adequately and reliably. However, ANQ had the more rapid anti-malarial action, resulting in almost 50% of the trial patients being cleared of parasites in the first 24 hours. Moreover, ANQ treatment significantly suppressed gametocytogenesis than the CQ+SP treatment. What makes this ACT unique is the claim that it is effective when administered as a single dose, whereas other ACTs require multiple dosing ,27. The The ANQ combination possesses benefits of both short-acting artemisinin, and long-acting naphthoquine combined into one tablet. The administration of 8 tablets in a single dose seems an optimal dosage for the trialed population in treating uncomplicated falciparum malaria infections. The dosage is safe and effective in clearing the blood parasitaemia. Rapid reduction in blood parasitaemia by ANQ, without post-treatment surge in parasitaemia, is critical for preventing potential complications or progression to severe and complicated malaria. However, whether rapid clearance of parasites by ANQ makes ANQ a more effective drug combination than CQ+SP is to be ascertained with further studies. Interestingly, CQ+SP combination still remain relatively effective at dosage given (30 mg/kg over three days), despite reports of seemingly moderate to high levels of multi-drug resistance in the study areas -6,21. HoOne of the key elements in any drug development and evaluation is the issue of safety of the population for which the drug is intended. Data from this study indicate that the use of ANQ combination is relatively safe, both from the clinical and laboratory perspectives. However, the quinine-like transient deafness reported by several patients who received ANQ combination requires further investigation. Although the sample size in this study is insufficient, larger studies are needed to define the safety and efficacy in different populations including children and pregnant women and in different ethnic settings. This ACT is uniquely novel in that it can be administered less frequently (as a single one day dose or twice a day dose) and yet produces almost equivalent cure rates to more frequently administered drug regimens such as CQ+SP combination in this study or artemether-lumefantrine combination in other studies . Hence, In conclusion, although both combinations assessed in this study provided relatively equivalent therapeutic responses (efficacy), tolerability, and safety profiles at day 28, the anti-malarial action of ANQ was more rapid than CQ+SP. The ANQ combination is a potential new generation ACT, which deserves further clinical evaluation.The authors declare that they have no competing interests.FWH designed the study protocol, supervised the study and prepared the manuscript. The design of the study protocol was assisted by DL, AS, AM, who were also involved in the conduct of the study. RJ and CK were responsible for the conducted of the study in Port Moresby while ST and JG were responsible for the conduct of the study in Madang. IK and MS were independent clinical assessors while GH was appointed as an independent clinical monitor and provided logistical support to the study. All the authors agreed to the content of the text."}
+{"text": "The efficacy of artemisinin-based combination therapy has already been demonstrated in a number of studies all over the world, and some of them can be regarded as comparably effective. Ease of administration of anti-malarial treatments with shorter courses and fewer tablets may be key determinant of compliance.Patients with uncomplicated falciparum malaria and over six months of age were recruited in Cameroon, Mali, Rwanda and Sudan. 1,384 patients were randomly assigned to receive artesunate-sulphamethoxypyrazine-pyrimethamine (AS-SMP) three-day (once daily for 3 days) regimen (N = 476) or AS-SMP 24-hour  regimen (N = 458) or artemether-lumefantrine (AL), the regular 6 doses regimen (N = 450). The primary objective was to demonstrate non-inferiority (using a margin of -6%) of AS-SMP 24 hours or AS-SMP three days versus AL on the PCR-corrected 28-day cure rate.The PCR corrected 28-day cure rate on the intention to treat (ITT) analysis population were: 96.0%(457/476) in the AS-SMP three-day group, 93.7%(429/458) in the AS-SMP 24-hour group and 92.0%(414/450) in the AL group. Likewise, the cure rates on the PP analysis population were high: 99.3%(432/437) in the AS-SMP three-day group, 99.5%(416/419) in the AS-SMP 24-hour group and 99.7(391/394)% in the AL group. Most common drug-related adverse events were gastrointestinal symptoms (such as vomiting and diarrhea) which were slightly higher in the AS-SMP 24-hour group.AS-SMP three days or AS-SMP 24 hours are safe, are as efficacious as AL, and are well tolerated..NCT00484900  Over the last few years artemisinin-based combination therapy (ACT) is widely accepted as an appropriate treatment for malaria. This disease remains an important killer, particularly in childhood, in sub-Saharan Africa -3. ACT oThe efficacy of ACT has already been demonstrated in a number of studies all over the world -7, and s\u00ae), taken once daily (two tablets simultaneously) over three days, was studied in Mali [\u00ae), was subsequently developed. Since preliminary experiments with these tablets indicated that the dosing interval could be reduced to 12 hours enabling a 24-hour therapy [\u00ae (AL FDC), available as a six-dose regimen, as standard therapy for assessing AS-SMP FDC. The primary objective was to demonstrate the non-inferiority of AS-SMP 24 hours or AS-SMP three days versus AL on the PCR-corrected 28-day cure rate.In a recent communication, WHO experts announced that the ideal anti-malarial drug should have an efficacy of at least 95% as measured over 28 days of follow-up. They recommend that re-infection in that period should be minimal and that, ideally, the treatment should be restricted to a few pills administered as a single dose and should have a short treatment duration [ in Mali , Ivory C in Mali , and is  therapy , it was This multi-centre study, which took place in four African countries: Cameroon, Mali, Rwanda and Sudan. In Cameroon, the study took place in the capital city, Yaound\u00e9. The study site was the Cameroon Baptist Convention clinic in the peri-urban district of Biyem-Assi. Transmission in Yaound\u00e9 occurs in two peak periods at the start of the rainy season in March/April, and in October/November as the rains cease. Chloroquine resistance is above 45%, amodiaquine resistance is about 10% and resistance to sulphadoxine/pyrimethamine (SP) is higher than 10% in Yaound\u00e9 or other parts of Cameroon ,15.Plasmodium falciparum is the predominant plasmodium species, accounting for more than 95% of malaria cases. Chloroquine resistance is above 25% [In Mali, the study took place at Bancoumana health center and surrounding villages (Kolle and Samako). The study area is located at about 60 km south-west of Bamako. The main population activity is farming. Malaria is hyperendemic and the transmission is mainly seasonal from June to December. bove 25% ,17 and rPlasmodium falciparum is the predominant plasmodium species, accounting for more than 95% of malaria cases. Resistance to SP exceeded 25% [In Rwanda, the study took place in Rwamagana and Muhimai. The study area is located at about 50 km east of Kigali. The main population activity is farming. Malaria is hyperendemic. eded 25% .of P. falciparum resistance to chloroquine (>70%) and the reported SP resistance in Sudan is ranging from 5% to 30% [In Sudan the study took place at Alhara Alola health center in New Hlafa, eastern Sudan. This area is located at 500 km from Khartoum in the middle of the second largest irrigated agricultural scheme in Sudan. Cotton and wheat are the main crops cultivated during the winter season. The area is characterized by a high level % to 30% ,20.P. falciparum. Inclusion criteria were: above six months of age, weight above5 kg, infection with P. falciparum  at screening, fever (axillary temperature \u226537.5\u00b0C or a history of fever in the preceding 24 hours), resident of study site, able to take oral treatment. Exclusion criteria were: symptoms or signs of severe malaria [This study recruited patients presenting at the health center with typical symptoms of malaria and who had a blood smear positive for  malaria , seriousThis was a prospective in vivo efficacy study. It was a randomized open label trial comparing fixed dose AS-SMP 24-hour or three-day regimen to AL as the standard reference treatment. The laboratory personnel assessing the parasitaemia was kept blinded. The study was carried out according to current WHO 2003 Protocol .\u00ae, Dafra Pharma, Belgium), as a 24-hour or three-day regimen, or AL . The randomization code was computer-generated by a third party not involved in patients' outcome assessment. It was a block randomization procedure stratified by country. Study codes were sealed in individual envelopes and securely stored. Randomized patients were assigned a study number in numerical sequence by the investigators. All drugs were manufactured according to Good Manufacturing Practice (GMP). AL tablets were a fixed combination, each containing 20 mg of artemether and 120 mg of lumefantrine. AL was administrated according to body weight  as six consecutive doses: The first dose at diagnosis and the second dose eight hours later on Day 0, and then two doses at 12 hourly intervals for the subsequent two days.Enrolled patients were randomly assigned a fixed dose of AS-SMP . Patients allocated to the 24-hour treatment with AS-SMP FDC were given tablets for three consecutive intakes at an interval of 12 hours (0 h \u2013 12 h \u2013 24 h).All study drug doses were administrated at the heath center by the study team. A full drug dose was re-administrated if the patients either spat out or vomited the study drugs within 30 minutes. Half the drug dose was re-administrated if the patient vomited the study drugs between 30 minutes to one hour. If the patient rejected again, he/she received another anti-malarial drug in conformity with the National Malaria Control Programme of his/her respective country, and was then excluded from the study.Follow-up examinations were made on days 0, 1, 2, 3, 7, 14, 21 and 28 or at any time if patient felt unwell. Blood from a finger prick was obtained to make a thick smear and a filter paper dot during each day of follow-up. The patients or guardians were asked about drug consumption since the last clinic visit. Individuals for whom treatment failed were treated according to National Malaria Control Programme. Giemsa-stained thick smears were read by an experienced microscopist blinded to study arm. Parasitaemia was quantified by a standard approximation method (40 \u00d7 number of parasites per 200 leucocytes on the thick film). Slide quality control was done by masked re-reading of 10% of slides, selected randomly. Haematological tests (Complete Blood Count) and biochemistry analyses  were done at baseline and at day 7 and then on day 28. Patients at Cameroon site and few patients at Sudan site did the blood test on day 14 instead of day 7. The tests were done any time for any subject if clinically recommended or in case of significant abnormality in tests during scheduled lab visits. Venous whole blood collected in anticoagulant tube was used for haematological tests, while venous blood collected in serum separator tube was used for biochemistry analyses. Less than 10 ml of blood was needed for these tests.msp1 and msp2) and microsatellite (CA1), to distinguish between re-infection and recrudescence as described previously [msp1 and msp2 and the microsatellite CA1 gene loci.For participants with recurrent parasitaemia after day 7, paired polymerase chain reaction (PCR) blots (from day 0 and the day of parasitaemia recurrence) were analysed for parasite merozoite surface protein 1 and 2 genes  re-infection, if the alleles of the pre- and post-treatment samples are distinct for any one of these loci; (iii) mixed recrudescence and re-infection, if similar alleles are found in the pre- and post-treatment samples for all the markers as mentioned above, but with additional distinct alleles identified; (iv) indeterminate, if either or both the pre- and post-treatment samples could not be amplified. Mixed recrudescent and re-infection cases were computed as recrudescent.Possible outcomes were: (i) recrudescence, if the alleles of the pre- and post-treatment samples were the same for The classification of the therapeutic outcome was done according to the current 28-day WHO protocol . The priThe non-inferiority of the AS-SMP 24 hours or AS-SMP 3 days to AL on the 28-day PCR-corrected cure rates was assessed by constructing a one-sided, lower limit, asymptotic 97.5% confidence interval (CI) on the difference in cure rates between AS-SMP 24 hours or AS-SMP 3 days and AL. Non-inferiority was declared if the lower limit of this CI was greater than -6% (for AS-SMP 24 hours or AS-SMP three days minus AL). The non-inferiority margin of 6% was chosen from a cure rate of 94% at day 28 with AL reported in the literature [nQuery Advisor 5.0 software was used for sample size calculation. On the basis of the above assumption, 900 patients (300 per treatment group) would be needed to demonstrate non-inferiority stated above with approximately 80% power. Assuming a 15% non-evaluability rate (e.g. lost to follow-up) it was planned to enroll 1035, rounded up to 1044 (348 per treatment arm). That total sample size was increased later to a total of 1,384 subjects included into the study increasing the precision of the parameters estimate (the efficacy proportion as well as of the secondary objective outcomes such as adverse events). Data from all sites were pooled and analyzed based on different populations.The intent-to-treat (ITT) population analysis included all randomized subjects. Lost to follow-up and withdrawn subjects were considered as treatment failure cases.The per protocol (PP) population analysis included all subjects who took study medication and made study visits up to the time of treatment failure or to the end of the study (28 days). Lost to follow-up and withdrawn subjects including protocol violation cases were excluded in this population. The primary analysis was based on the 28-day PCR corrected efficacy of both ITT and PP populations.The baseline and safety analysis were done using ITT population. Data were double-entered, validated using Microsoft Access and analysed with STATA version 10.0 . Chi-square test with Fisher correction exact test was used as appropriate to compare categorical variables. Parametric or non-parametric tests were computed as appropriate to compare continues data between the three treatments arms. P value less than \u03b1 = 0.05 was considered as statistically significant.The protocol was reviewed and approved by: the National Ethical Committee, Yaound\u00e9 (for Cameroon),, the Ethical Committee of the Faculty of Medicine, Pharmacy and Dentistry at the University of Bamako , the National Ethical Committee, Kigali (for Rwanda), and the Ethical Review Board of the Academy of Medical Science and Technology, Faculty of Medicine (for Sudan). Each patient (or their guardian or parent) gave fully informed written consent prior the enrollment.The first patient was enrolled in August 2006 and the study was completed in May 2007.Of the 1,384 enrolled participants  in the AS-SMP three-day group, 91.1%(417/458) in the AS-SMP 24-hour group and 89.6%(403/450) in the AL group in the ITT population. Likewise, the PCR uncorrected 28-day cure rates were similar among treatment groups: 95.9% (422/440) in the AS-SMP three-day group, 96.0%(404/421) in the AS-SMP 24-hour group and 96.0%(380/396) in the AL group in the PP population. The reinfection rates were similar among treatment groups: 2.8% in the AS-SMP three-day group, 3.1% in the AS-SMP 24-hour group and 3.6% in the AL group in the PP population  and 2/387 (0.3%) in the AS-SMP 24-hour group and in the AL group on day 2 had fever and parasite while the fever clearance was total in all treatment groups on day 3. Gametocyte carriage decreased from baseline to day 28. At baseline on Day 0, 6/432 patients (1.4%) in the AS-SMP three-day group, 8/394 (2.0%) and 16/418 (3.8%) in the AS-SMP 24-hour group were carrying gametocytes. Only 2/427 patients (0.5%) in the AS-SMP 3 group, 0/387 (0%) and 10/410 (2.4%) in the AS-SMP 24-hour group were carrying gametocytes on day 7; p = 0.001.There was one patient with ETF in the AL group. Few patients developed severe malaria: four patients in the AL group and two in the AS-SMP 24-hour group, all treated successfully with other anti-malarias treatment (by injection).The adverse events probably related to study drug were gastrointestinal signs/symptoms , dizziness, rash and weakness. Gastrointestinal signs/symptoms, such as vomiting and diarrhoea, were slightly higher in AS-SMP 24-hour group: for vomiting, it was 7.0% (n = 458), 4.6% (n = 476) and 2.2% (n = 450) for AS-SMP 24-hour group, AS-SMP three-day group and AL group respectively; p = 0.003. For diarrhoea it was 3.3% (n = 458), 0.6% (n = 476) and 1.3% (n = 450) for AS-SMP 24-hour group, AS-SMP three-day group and AL group respectively; p = 0.006 (Table \u00ae) problems such as two doses per day over three days and high re-infection rate in areas with high transmission intensity are a disadvantage [Malaria remains a major health problem characterized by high mortality and serious morbidity in particular with children less than five years of age -3. Malardvantage . The optdvantage .The current study brings interesting information regarding a new ACT. Previous studies had shown AS-SMP to be adequate -13 and tThe study shows that the efficacy of the AS-SMP 24-hour or AS-SMP three-day fixed dose treatment in patients with acute uncomplicated falciparum malaria was not inferior to AL, and had a comparable safety profile except for vomiting slightly higher in the AS-SMP 24-hour group.PCR-corrected 28-day cure rates were high in both ITT and PP analysis in the different treatment groups. The efficacy rate was comparable across the countries, although the study was not powered enough for comparing the three treatment groups within country. Moreover, there was no difference among the three treatment groups in term of clearance of fever or parasite  or moreover AS-SMP 24-hour treatment (0\u201312 h-24 h) has been developed. This study shows that AS-SMP three-day or AS-SMP 24 hours is as efficacious as the AL three-day treatment (two doses a day).\u00ae co-blister) gave a 96.6% ACPR. The high cure rate of AS/SMP compared to AS/SP in the Rwanda study is also confirmed in this multicentre study; confirming the chemical activity differences between SMP and SP [In this study, it was clearly demonstrated that the shortening of the dose interval between tablets from three days to one day (24 hours) did not compromise the outcome of the malaria course. However, the second dose of the 12-hour interval administration drug of AS-SMP and second dose administration drug of AL may fall outside the regular working hours of the health centers and also may fall to patients' bed time and therefore may impact on treatment compliance. In term of efficacy, there was no difference between treatment groups and the result in each treatment group approaches 100% in the PP analysis (ACPR = 99%). This is rather remarkable since the study was carried out in areas where resistance to SP is high. This holds for Rwanda where the SP resistance is estimated to exceed 25% . Also inP and SP ,32. ThisP and SP . The stuAS-SMP three days or AS-SMP 24 hours are safe, are as efficacious as AL, and are well tolerated, although vomiting and diarrhoea were slightly higher in the AS-SMP 24-hour group than other groups.The authors declare that they have no competing interests except for FHJ who is an employee of Dafra Pharma. No other author received any honoraria or salary from Dafra Pharma.All authors contributed to the design of the study and assisted with data interpretation. IS, WM, RS and IA coordinated the study and supervised the enrollment and follow-up of patients. FH J, AD, OKD, and AD participated in study design. IS and AD contributed in data management and analysis. AD did the molecular analysis in determining the status of recrudescence or reinfection. KS, HM, OBT, ND and YTD collected the data. All authors participated in the preparation of the manuscript and approved the final version.v.s Sulfadoxine-PyrimethamineSulfamethoxypyrazine-Pyrimethamine . History and comparison of Sulfamethoxypyrazine-Pyrimethamine_vs_Sulfadoxine-PyrimethamineClick here for file"}
+{"text": "Plasmodium falciparum, because this antigen can persist after effective treatment, giving false positive test results in the absence of infection. An assessment of the accuracy of Malaria Pf\u2122 immuno-chromatographic test (ICT) and description of persistent antigenicity of HRP2 RDTs was undertaken in a hyperendemic area of Uganda.Parasite-based diagnosis of malaria by microscopy requires laboratory skills that are generally unavailable at peripheral health facilities. Rapid diagnostic tests (RDTs) require less expertise, but accuracy under operational conditions has not been fully evaluated in Uganda. There are also concerns about RDTs that use the antigen histidine-rich protein 2 (HRP2) to detect Using a cross-sectional design, a total of 357 febrile patients of all ages were tested using ICT, and compared to microscopy as the gold standard reference. Two independent RDT readings were used to assess accuracy and inter-observer reliability. With a longitudinal design to describe persistent antigenicity of ICT and Paracheck, 224 children aged 6\u201359 months were followed up at 7-day intervals until the HRP2 antigens where undetectable by the RDTs.P. falciparum. ICT had an overall sensitivity of 98%, a specificity of 72%, a negative predictive value (NPV) of 98% and a positive predictive value (PPV) of 69%. ICT showed a high inter-observer reliability under operational conditions, with 95% of readings having assigned the same results .Of the 357 patients tested during the cross-sectional component, 40% (139) had positive blood smears for asexual forms of In children followed up after successful antimalaria treatment, the mean duration of persistent antigenicity was 32 days, and this duration varied significantly depending on pre-treatment parasitaemia. In patients with parasite density >50,000/\u03bcl, the mean duration of persistent antigenicity was 37 days compared to 26 days for parasitaemia less than 1,000/\u03bcl .ICT is an accurate and appropriate test for operational use as a diagnostic tool where microscopy is unavailable. However, persistent antigenicity reduces the accuracy of this and other HRP2-based RDTs. The low specificity continues to be of concern, especially in children below five years of age. These pose limitations that need consideration, such as their use for diagnosis of patients returning with symptoms within two to four weeks of treatment. Good clinical skills are essential to interpret test results. Prompt and accurate diagnosis of malaria is the key to effective case management and a major component of Uganda's malaria control strategy. Clinical diagnosis is the least expensive and most widely practised method and the one generally used for self treatment. However, the overlapping of malaria symptoms with other tropical diseases impairs its specificity and can promote the indiscriminate use of anti-malarials and compromise quality of care for patients with non-malarial fevers in endemic areas . TherefoConventional light microscopy is the \"gold standard\" for routine parasite-based diagnosis because it is sensitive, inexpensive to perform, can differentiate malaria species and quantify parasite load. Microscopy requires well-trained technicians, a microscope in good working order and a well-functioning quality assurance system. The challenges for governments to ensure that such prerequisites are in place mean that microscopy is unavailable at many peripheral health facilities in resource limited settings, where most cases are managed in the public health sector. Rapid diagnostic tests (RDTs) require less expertise to be conducted correctly, and non-specialized staff can be quickly trained to use them. Malaria diagnostic accuracy may be strengthened by the use of RDTs where microscopy is not available or feasible to maintain.Uganda changed its malaria drug policy in 2004 from chloroquine plus sulphadoxine-pyremethamine (CQ+SP) to artemisinin-based combination therapy (ACT) using Coartem\u2122 (artemether-lumefantrine) as the first-line treatment. ACT is a more expensive treatment than the previous drug combination of CQ+SP, and the Ministry of Health (MoH) is thus keen to shift malaria diagnosis to become parasite-based rather than clinical. This is to ensure that only confirmed cases of malaria receive anti-malarials and differential diagnosis is utilized to identify the cause of fever in cases giving a negative result for malaria. In this way, the quality of care of fever cases will be greatly improved and the wastage of anti-malarials will be minimized.Plasmodium falciparum, Plasmodium lactate dehydrogenase (pLDH), and Plasmodium aldolase. The current policy guideline in Uganda recommends use of HRP2, due to the very high proportion of P. falciparum infections [A review of studies evaluating diagnostic tools for malaria found very high accuracy and reliability of RDTs . The tesfections .P. falciparum infections, are heat stable[HRP2-based RDTs are very sensitive in detecting at stable and geneat stable. Howeverat stable-9. Low sP. falciparum only, unlike previously evaluated RDTs that detect all four human malaria species. There have been a number of other studies on malaria RDTs done in Uganda [Currently, there is limited information on the accuracy of Malaria Pf\u2122 ICT (ICT) marketed by ICT Diagnostics, South Africa. The data available ,7,10-16 n Uganda -21.This study assesses the operational accuracy and ease of use (inter-observer and intra-test reliability) of ICT in a district hospital setting. It also describes persistent HRP2 antigenicity of both ICT and Paracheck Pf (another HRP2-based RDT) in children aged 6\u201359 months treated for malaria in a hyperendemic setting in Uganda.P. falciparum infection using microscopy as the gold standard. In addition, a longitudinal design was used to describe persistent antigenicity using two HRP2 tests (ICT and Paracheck). Both components were undertaken in Soroti Regional Referral Hospital, Uganda. Soroti town is 347 kilometres north of the capital Kampala and is in an area of hyperendemic malaria with a parasite prevalence of 85% in children below nine years of age [The study was conducted as two independent serial components. A cross-sectional assessment was performed to determine the accuracy of ICT for For both study components, consecutive patients presenting to the OPD were screened for symptoms/history suggestive of malaria and evaluated for eligibility into the study. During the cross-sectional component, patients who fulfilled the following criteria were enrolled: 1) aged six months and above; 2) history of fever in the last 24 hours or axillary temperature \u2265 37.5\u00b0C; 3) no evidence of a concomitant febrile illness; 4) provision of informed consent; 5) no danger signs or evidence of severe malaria.Plasmodium falciparum monoinfection on blood smear; 2) positive RDT test results; 3) no evidence of a concomitant febrile illness; 4) no danger signs or evidence of severe malaria; 5) provision of informed consent including extended follow-up; and 6) negative blood smear for asexual-stage peripheral parasitaemia on day-3 after anti-malarial therapy.For the longitudinal component, only children aged 6\u201359 months and residing within seven kilometres from the hospital were recruited. The additional inclusion criteria were as follows: 1) At screening, patients or guardians were asked about prior anti-malarial use and presence of common symptoms. Weight and axillary temperature were measured and a physical examination performed and patients that provided consent were referred to the laboratory for investigation.In the laboratory, blood was obtained by finger prick for thick and thin blood smears, for RDT and a drop was taken on a filter paper for possible future molecular analysis.Plasmodium species. Patients were managed based on microscopy results and national treatment guidelines. Medication was dispensed at a general OPD pharmacy unsupervised by the study team.Blood smears were stained with 10% Giemsa for 30 minutes. Thick smears were examined for parasites including gametocytes and parasite densities were determined by counting the number of asexual parasites per 200 white blood cells (WBC), or per 500 WBC if the parasite count was less than 10 parasites per 200 WBC, assuming a WBC count of 8,000/\u03bcl. A slide was considered negative if no parasites were seen after reviewing 100 high power fields. Thin smears were read to determine the A RDT result was interpreted as positive when both the test line and control line showed pink, negative when only the control line showed pink or invalid when the control line did not appear regardless of the test line. Two independent readings were graded based on visual assessment as \"strong\" or \"faint positive\" for reactive tests and \"negative\" for non-reactive ones. A test result was graded \"strong\" if the test line was as intense as the control line and a \"faint\" result was a line that could only be seen in good light. The reason for this distinction on the positive line intensity is that a strong result is very clear, but faint positive lines can easily be missed in situations where the lighting is poor or the operator has poor eyesight. Grading was also used to determine the source of variability in results reported by two independent readers. Any invalid RDT tests were not repeated.During the longitudinal component of the study, patients were evaluated simultaneously using Paracheck and ICT RDTs at screening and on every follow-up day. Only patients with a negative day-3 smear were enrolled for the longitudinal component. Enrolled patients were asked to return for follow-up on day-7 and every subsequent seven days or any other day that they felt ill. Follow-up evaluation consisted of taking finger prick blood for RDT tests and blood smears. Enrolled patients with complaints were also clinically evaluated and managed. The follow-up period ended when an outcome was determined as either smear-positive or RDT-negative. Patients with positive smears were referred to the clinician for evaluation and treatment. with an alarm set at 37\u00b0C.Local hospital staff were involved in the day-to-day conduct of the study. Both laboratory and clinical teams were trained to perform the RDT. The training was followed with a pilot study of about 20 patients to ensure that all study procedures were clearly followed and that the study did not cause any unnecessary interference to the normal functioning of the clinics and the laboratory. Laboratory technologists were assigned work according to hospital duty register. All study slides were read by first and second microscopists who were blinded to each others' results. The same second microscopist was maintained throughout the longitudinal component to ensure consistency. Reading were considered discordant for the following scenarios: 1) positive/negative discordance for asexual stages; 2) species discordance; 3) asexual discordance of more than 50% difference between the parasitaemia; 4) positive/negative discordance for gametocytes. A third, external microscopist, unaware of the first two results, resolved any discordant results and read a further random 10% for external quality control purposes. Similar to microscopy, RDT cassettes were labelled and read by two laboratory technologists independently 15 minutes after adding buffer. Temperature and humidity for the storage conditions for RDTs during study period were monitored using Tinytag\u2122 Data Loggers The outcome measures were RDT and microscopy results for the cross-sectional component. During the longitudinal component, the possible outcomes at each follow-up evaluation were; 1) positive RDT and negative smear interpreted as persistent antigenicity; 2) both RDTs and smear negative interpreted as cleared antigen; and 3) positive RDT and positive smear interpreted as a recurrence of asexual parasites (possible recrudescence or re-infection). Patients with persistent antigenicity at any time point were given a 7-day follow-up appointment. Patients were excluded from further follow-up if they withdrew consent or both RDT readings were invalid or not interpretable.et al [To determine accuracy of ICT, sample size was calculated considering a sensitivity of 90% and a prevalence of malaria in the study population of 50% (among children under 10 years). Using the nomogram by Carley et al , at a prFor the longitudinal component, to describe and determine predictors of persistent antigenicity, a sample of 224 participants that completed follow-up showed >80% power of detecting a difference between low parasite density , and high  day-zero parasitaemia. and analysed using SPSS version 12.0 . Overall agreement, as a measure of reliability of RDT readings was calculated using a kappa statistics. Using microscopy as the reference, sensitivity of the RDT was measured as the proportion of RDT positives over the total positives determined by microscopy and specificity as proportion of RDT negatives over the total negatives determined by microscopy. The positive predictive value (PPV) is the number of RDT true positives divided by the number all RDT positive test results . The negative predictive value (NPV) is the number of RDT true negatives divided by the number all RDT negative test results .Data were entered and verified using EpiData 3.01Kappa statistics. For analysis purposes, day-zero parasite density was categorized into three groups; <1,000/\u03bcl, 1,000\u201350,000, and > 50,000/\u03bcl. The mean duration was determined for persistent antigenicity  and compared equality of survival distribution for categorized day-zero parasite density using log rank test statistics.Duration of persistent antigenicity was measured as time in days, when RDT remains positive after treatment and the blood slide remains negative. Data from patients with recurrent parasitaemia were included in the analysis of persistent antigenicity until the day the microscopy smear became positive and an outcome measure of the previous time-points assigned. Inter observer reliability of ICT and intra-test reliability of ICT and Paracheck were calculated using P. falciparum, predominately in the children under five years, where 102 (73%) were positive  in two components. During the cross sectional component, 364 patients were recruited into the study, but seven patients were excluded from this analysis because of unreadable blood smears. Of the 357 patients evaluated, the median (IQR) age was 11 (1\u201328) years, 46% were under the age of five, and 60% female. A total of 139 (39%) had positive blood smears for asexual forms of P. falciparum monoinfection were screened with both ICT and Paracheck RDTs on day-zero. On day-3, participants were evaluated to determine anti-malarial treatment response. Of the 310 patients screened on day-zero, 51(16.4%) were not enrolled in the follow-up component due to the following reasons: sixteen still had asexual parasitaemia (early treatment failures); three children with negative smears on day-zero had been included in violation of the protocol, while 32 children who missed day-3 evaluation were excluded because anti-malarial treatment response could not be determined in time. Out of the 259 children successfully enrolled for follow-up after day-3 evaluation, three patients withdrew consent citing unwillingness to continue with a hospital based follow-up and thirty-two others failed to honour appointments during the course of follow-up  and specificity of 72% (95% CI: 65\u201377). The specificity of ICT was significantly lower in the <5 years at 54% (95% CI: 41\u201367) compared to 78% (95% CI: 71\u201385) in the older age group. Overall, ICT test had a NPV of 98% (95% CI: 95\u2013100) and PPV of 69% (95% CI: 62\u201375). The 78% (95% CI 58\u201370) PPV in the under fives is significantly higher than 51% (95% CI 39\u201364) in the above fives , showing a very high operational test reliability.For the cross-sectional component of the study, the results of the first and second ICT readings were highly comparable with 336/354 (95%) concordant reading  and further analysis below is presented simply as HRP2 results and does not distinguish between the two RDTs.During the longitudinal component of the study, all tests were performed using ICT and Paracheck RDTs. Results where graded as \"strong\" positive, \"faint\" positive and negative. Comparing ICT and Paracheck paired tests done for the same patient during the follow-up component showed 1488/1559(95%) concordance between the two RDT results. The major difference between the two HRP2 tests was in terms of line intensity as \"strong\" versus 'faint\" positives. There were 17 discordant pairs, for which ICT showed 14 as negative while Paracheck showed \"faint positive\". Two \"faint positive\" ICT tests were negative on Paracheck. Only one negative ICT showed \"strong positive\" with Paracheck. Overall, there were eight invalid ICT tests Table . When alFor all the 310 children screened on day-zero, there was no significant difference at baseline of gender, age, temperature, parasite density, presence of gametocytes or recent anti-malarial use between those who completed follow-up and the ones that did not  were followed up until the antigenicity was no longer detectable while 62 (28%) had recurrent peripheral parasitaemia detected by microscopy during the course of the follow-up time at which point they were no longer followed-up. Persistent antigenicity was documented in 98% of the cases on day-7 and this decreased to 14% by day-35  discordant records were assigned different interpretations as \"faint positives\" versus negative results. There was a high level of result concordance between ICT and Paracheck tests during longitudinal component. The two HRP2 tests were of comparable accuracy and performance in an operational setting.This implies that given minimum training, it is feasible for field workers to interpret and report consistent results.P. falciparum parasite density in Ugandan children. Overall, persistence of HRP2 antigenicity was detectable in the majority of children at three weeks following successful treatment. The mean duration of 32 days is particularly worrying because HRP2 persistent antigenicity reduces the accuracy of the RDT test. This is more of concern in hyperendemic areas, where frequent malaria infections mean that children are likely to have the antigen in their blood even if their fever is not caused by malaria. There is a danger of over-diagnosis of malaria based on the interpretation of HRP2 test results. Misdiagnosis results in poor quality of care as the appropriate illness may not be addressed in time. It may also result in misuse of anti-malarials, poor resource allocation and, clinicians doubting the efficacy of the medicine used. Clinical skills in taking history and eliciting signs are essential when interpreting RDT results in an effort limit misuse of anti-malarials. Misuse of anti-malarial contributes to drug pressure and spreading of drug resistant P. falciparum [The study also described the duration of ICT false positivity due to persistent antigenicity and its relationship to pre-treatment lciparum , resultilciparum .Previous studies from Asian countries have also reported persistent antigenicity after malaria clinical cure and the levels reported have varied widely, with ranges of 29\u201365% on day-7 ,29-31. IThe causes of persistent antigenicity after malaria therapy are still debatable. Previous studies have listed causes of persistent antigenicity as parasite density levels below detection threshold for microscopy, delayedThe study participants were drawn from a pool of all persons suspected of having malaria, regardless of history, clinical status, demographic characteristics or other factors that may affect sensitivity and specificity in a routine health clinic. Inclusion of a large number of individuals improved the precision of study estimates. Therefore, the results obtained here reflect the performance of ICT in the operational setting.There were some limitations in the conduct of the study. The evaluation was performed using microscopy as a gold standard, fully aware of its limitations in detecting low parasite densities . To imprThe study team neither observed therapy nor assessed for adherence to medication for the children during the longitudinal component. Poor adherence to treatment may have led to recurrent parasitaemia as a result of treatment failure.RDTs devices are new technologies that resemble HIV test kits formats previously used in this setting. During the follow-up, a significant number of patients that had consented to participate failed to honour all the scheduled appointments. Only three patients formally withdrew consent mentioning that their families were suspicious that these were actually HIV studies. It is likely that others may have kept away from the study with similar misinformation. All patients' data collected during the evaluation of persistent antigenicity was utilized to mitigate any effect (bias) caused by loss to follow-up.P. falciparum infection in a highly endemic region of Uganda. However, persistent antigenicity reduces the accuracy of this and other HRP2-based RDTs. The low specificity continues to be of concern, especially in children below five years of age, if they return with symptoms within four weeks of previous antimalarial treatment. The study highlights important practical issues that need consideration when using RDTs for diagnosis. Good clinical skills are essential to interpret test results given the persistent antigenicity and specificity variation for age in endemic areas. There is a need to explain to the end user the best way to use the tools for diagnosis.In conclusion, this study shows that ICT is an accurate test for diagnosis of malaria in an operational setting where microscopy is not available. It is a reliable test in detecting The authors declare that they have no competing interests.HC, JKT, JBR and DJK participated in designing the study. DJK and HC directed the field study. GWO participated in the data acquisition and the on-site supervisor. DJK and HC verified data, DJK and JKT analysed the data. DJK and HC participated in the manuscript preparation. All authors read and approved the final manuscript."}
+{"text": "Artemisinin-combination therapy (ACT) is recommended as first-line treatment of falciparum malaria throughout the world, and fixed-dose combinations are preferred by WHO; whether a single gametocytocidal dose of primaquine should be added is unknown. We aimed to compare effectiveness of four fixed-dose ACTs and a loose tablet combination of artesunate and mefloquine, and assess the addition of a single gametocytocidal dose of primaquine.Plasmodium falciparum malaria or mixed infection, who were older than 6 months, and who weighed more than 5 kg. Treatments were randomised in equal numbers within blocks of 50 and allocation was in sealed envelopes. All patients were also randomly assigned to receive either a single dose of primaquine 0\u00b775 mg base/kg or not. Patients were followed up for 63 days. Treatment groups were compared by analysis of variance and multiple logistic regression. The primary outcome was the 63 day recrudescence rate. This study is registered with clinicaltrials.gov, number NCT00902811.In an open-label randomised trial in clinics in Rakhine state, Kachin state, and Shan state in Myanmar (Burma) between Dec 30, 2008, and March 20, 2009, we compared the effectiveness of all four WHO-recommended fixed-dose ACTs  and loose artesunate\u2013mefloquine in Burmese adults and children. Eligible patients were those who presented to the clinics with acute uncomplicated P falciparum infections, a rate significantly higher than for artemether\u2013lumefantrine , fixed-dose artesunate\u2013mefloquine , loose artesunate\u2013mefloquine , and dihydroartemisinin\u2013piperaquine . Hazard ratios for re-infection (95% CI) after artesunate\u2013amodiaquine were 3\u00b72 (1\u00b73\u20138\u00b70) compared with the two artesunate\u2013mefloquine groups (p=0\u00b701), 2\u00b76 (1\u00b70\u20136\u20130) compared with artemether\u2013lumefantrine (p=0\u00b704), and 2\u00b73 (0\u00b79\u20136\u00b70) compared with dihydroartemisinin\u2013piperaquine (p=0\u00b708). Mixed falciparum and vivax infections were common: 129 (16%) had a mixed infection at presentation and 330 (41%) patients had one or more episodes of Plasmodium vivax infection during follow-up. The addition of a single dose of primaquine (0\u00b775 mg/kg) reduced P falciparum gametocyte carriage substantially: rate ratio 11\u00b79 (95% CI 7\u00b74\u201320\u00b75). All regimens were well tolerated. Adverse events were reported by 599 patients, most commonly vomiting and dizziness. Other side-effects were less common and were not related to a specific treatment.155 patients received artesunate\u2013amodiaquine, 162 artemether\u2013lumefantrine, 169 artesunate\u2013mefloquine, 161 loose artesunate\u2013mefloquine, and 161 dihydroartemisinin\u2013piperaquine. By day 63 of follow-up, 14 patients  on artesunate\u2013amodiaquine had recrudescent P falciparum infections in this region.Artesunate\u2013amodiaquine should not be used in Myanmar, because the other ACTs are substantially more effective. Artesunate\u2013mefloquine provided the greatest post-treatment suppression of malaria. Adding a single dose of primaquine would substantially reduce transmission potential. Vivax malaria, not recurrent falciparum malaria, is the main complication after treatment of M\u00e9decins sans Fronti\u00e8res (Holland) and the Wellcome Trust Mahidol University Oxford Tropical Medicine Research Programme. Artemisinin-based combination therapy (ACT) is recommended by WHO for the treatment of uncomplicated falciparum malaria.Artemisinin and its derivatives reduce gametocyte carriage,Our aim was to compare the efficacy of the four available fixed-dose ACTs and the currently used loose tablet combination of artesunate with mefloquine and to assess the effectiveness of adding a single gametocytocidal dose of primaquine.Plasmodium falciparum malaria (parasite density 500\u2013200\u2008000 parasites per \u03bcL) or mixed infection were enrolled into the study after fully informed consent was obtained from them or their carer. Patients were excluded if they were pregnant, had severe malaria, had severe acute malnutrition , had taken antimalarial drugs within the past 48 h, had taken mefloquine during the past 9 weeks, or had known history of hypersensitivity to any of the study drugs.Between Dec 30, 2008, and March 20, 2009, we recruited patients into our open-label randomised study at three clinics in Rakhine state in western Myanmar (Burma),After patients were screened and enrolled into the study, they were stratified prospectively into three age groups . Patients were randomly assigned in equal numbers to receive one of the five different treatments. They were then randomly assigned either a single dose of primaquine 0\u00b775 mg base/kg  or not. Treatment allocations were put in sealed envelopes in blocks of 50 for each age-group, and random assignment was achieved by patients drawing an envelope from a box after enrolment. When the box was empty, another 50 envelopes were added.For the four fixed-dose combinations the standard dosage instructions of the manufacturer for weight and age ranges were followed. For the loose tablets regimen of artesunate plus mefloquine, the target dose was artesunate 4 mg/kg per day for 3 days  plus mefloquine 25 mg base/kg on day 0; the number of pills given was rounded off to the nearest quarter of a tablet. For fixed dose artesunate\u2013mefloquine hydrochloride we used 25 mg plus 55 mg or 100 mg plus 220 mg tablets, the target dose was 4\u00b70 mg/kg per day plus 8\u00b78 mg/kg per day for 3 days. For fixed-dose artemether\u2013lumefantrine we used 20 mg plus 120 mg tablets twice daily for 3 days, the target dose was 3\u00b73 mg/kg per day plus 19\u00b78 mg/kg per day; patients were advised to consume some fatty food (or mothers were encouraged to breastfeed treated infants) before each dose. For fixed-dose dihydroartemisinin\u2013piperaquine we used 40 mg plus 320 mg or 20 mg plus 160 mg tablets, the target dose was 2\u00b75 mg/kg per day plus 20 mg/kg per day. For fixed dose artesunate\u2013amodiaquine we used 25 mg plus 67\u00b75 mg, 50 mg plus 135 mg, or 100 mg plus 270 mg tablets, the target dose was 4 mg/kg per day plus 10\u00b78 mg base/kg per day.The first dose was taken under supervision. For children, tablets were crushed and syrup was added. All subsequent doses were self administered. Patients received sealed plastic bags containing the remaining doses and were instructed clearly about their subsequent treatment, emphasising the importance of taking primaquine after food, and taking their medicines even when their symptoms had subsided. Patients were asked to return weekly for 9 weeks for assessment, and at any other time if they became ill. Microscopists examining blood films were unaware of treatment allocation. Haemoglobin was measured on day 63.Patients with recurrent falciparum malaria were treated with artesunate\u2013mefloquine (loose tablets). Patients who had already received this treatment were given dihydroartemisinin\u2013piperaquine. PCR genotyping was used to distinguish recrudescence from re-infection, by assessment of variable blocks within merozoite surface proteins 1 and 2, and glutamate-rich protein.Plasmodium vivax or Plasmodium malariae infections received chloroquine (25 mg base/kg) and continued follow-up. If the patients were again positive for P vivax 14 days or later, they were retreated with chloroquine and defined as having a new P vivax episode .Patients who developed intercurrent A sample size of 160 patients (80 with primaquine) per study group  allowed a cure rate (100\u2013recrudescence rate) of 95% per group to be estimated with 5% precision, and allowed estimation of effectiveness equivalence with a maximum allowable difference of 10% (90% power and 95% confidence) between groups with a follow-up drop-out rate of up to 20%.Treatment groups were compared by two-way factor analysis: ANOVA for continuous variables and multiple logistic regression for categorical data. A test for trend was used for comparisons across age groups. Time-to-event outcomes, including time to first recurrence , time to recrudescence (new infections censored), and time to new infection (recrudescent infections censored) were assessed by the Kaplan-Meier method. Patients lost to follow-up were censored at the last time seen. When failure rates were zero, CIs were estimated with the exact binomial method with the effective sample size. All other treatments were compared against the current standard of care, which is artesunate plus mefloquine (loose tablets), and then against fixed-dose artesunate\u2013mefloquine hydrochloride, with the log-rank test, or the Wilcoxon-Breslow test of equality if survival lines crossed. Cox regression was used to quantify risks.clinicaltrials.gov, number NCT00902811.Person-gametocyte weeks (PGW) were defined as the number of weeks in which gametocytaemia was patent (excluding at admission) divided by the duration of follow-up, expressed per 1000 person-weeks. The protocol was approved by the Myanmar Department of Health and by the M\u00e9decins sans Fronti\u00e8res ethics review board. This study is registered with The sponsor of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication.P falciparum on day 7 and so were treated again. Of the 808 eligible patients, 397 (49\u00b71%) also received primaquine  more common in children younger than 5 years (13 [15%] of 87 children) than in those aged 5\u201314 years (28 [9%] of 298), or in adults (28 [7%] of 423).All patients cleared parasitaemia by day 7. 69 patients had recurrence of  63 days . Recurre 63 days . Thus re38 new infections and 20 recrudescent parasitaemias occurred by day 63. Results for nine patients were indeterminate , and data for two patients (one artesunate\u2013amodiaquine and one loose artesunate\u2013mefloquine) were missing. If indeterminate or missing results are treated as censored findings, then the treatment failure rate after fixed-dose artesunate\u2013amodiaquine was significantly higher than for the other treatments  and more than twice that with artemether\u2013lumefantrine  and dihydroartemisinin\u2013piperaquine .P vivax infection during follow-up  had subsequent P vivax malaria compared with 95 (74%) of those with mixed infection initially (p<0\u00b70001). Fewer cases of P vivax were recorded in patients who received fixed-dose artesunate\u2013mefloquine than in patients who received fixed-dose artesunate\u2013amodiaquine, fixed-dose artemether\u2013lumefantrine, or the loose tablet regimen of artesunate plus mefloquine . Thus 12 of the 14 cases of recrudescence of P falciparum infection after artesunate\u2013amodiaquine happened before the median time to P vivax relapse for this group  than for adults . Children were nearly three times more likely to develop intercurrent vivax malaria than were adults, after adjusting for treatment and mixed infections at admission . Single-dose primaquine did not affect P vivax rates (data not shown).129 patients (16%) had mixed infections at presentation, and these were more common in children (102 [27%] of 385) than in adults . All patients with mixed infections responded to ACT treatment. 330 patients had ollow-up : 259 hadfloquine . The medis group , which aP falciparum, because patients were censored at the time of any recurrence of malaria. The day 63 malaria-free rate was highest in the fixed-dose artesunate\u2013mefloquine  and the dihydroartemisinin\u2013piperaquine  groups. The other three treatment groups had malaria-free rates of less than 50%, and all were significantly worse than fixed-dose artesunate\u2013mefloquine .Pooling all recurrences of falciparum or vivax malaria together provided a combined measure of efficacy and post-treatment prophylaxis, and it also addresses any possible confounding by weak activity of chloroquine against recrudescent vs ten of 268; 0\u00b710, 0\u00b701\u20130\u00b776; p=0.006).264 patients (33%) presented with patent gametocytaemia, which was more common in children younger than 5 years (46 [53%] of 87) than in children aged 5\u201314 years (121 [41%] of 298) and in adults . Independent of age, gametocytaemia was more common among moderately anaemic patients  than among patients with higher values . Without primaquine the treatment regimens had widely different gametocyte carriage rates . With prP vivax, but was slightly reduced by primaquine .246 patients (30%) had a haemoglobin concentration of less than 10\u00b70 g/dL (mild anaemia), and 70 (9%) a concentration of less than 8\u00b70 g/dL (moderate anaemia) at presentation. Young children were more likely to be anaemic at presentation (28 [32%] of 87) than older children (26 [9%] of 298) and adults . On day 63, eight (1%) of 693 patients were moderately anaemic and 72 (10%) were mildly anaemic. The mean increase of haemoglobin was similar among the five treatment groups, was unaffected by intercurrent 599 patients (74%) reported adverse events during the study . 13 patiControl and ultimately elimination of falciparum malaria depends on providing effective and well tolerated medicines and minimising transmission. In Myanmar, our large study shows that artesunate\u2013mefloquine, the first-line treatment in the main study region since 1996, remains very effective. The new fixed-dose artesunate\u2013amodiaquine was well tolerated, but was not effective enough to be recommended as a first-line treatment. The other available ACTs were all well tolerated and highly effective, suggesting good adherence to the prescribed treatments, although only the first dose was observed . Addition of a single dose of primaquine to ACT regimens has a very large additional effect on gametocytaemia, and therefore on malaria transmission potential, making any residual differences in gametocyte carriage after the primary infection between the five ACT regimens insignificant.From an operational perspective, fixed-dose combinations are simple to give and avoid monotherapy, thereby protecting against resistance. Findings from our previous studies have shown that artesunate\u2013mefloquine, the first-line treatment in Myanmar since 1996, is very effective.P falciparum gametocytaemia, and substantial discrepancy between cure rates in adults and children.Malaria transmission in Myanmar is seasonal and generally low, but access to effective drugs is limited, and the clinical epidemiology is similar to that of higher-transmission settings with a high prevalence of anaemia, high rates of The 8-aminoquinolines have a rapid and powerful sterilising effect on mature gametocytes.P falciparum and P vivax infections were common. 330 patients (41%) had at least one episode of vivax malaria in the 63 day follow-up. The risk was particularly high in young children\u201456 children (64%) younger than 5 years had vivax malaria after acute falciparum malaria. This compares with P falciparum recrudescence rates of less than 10%. Treatment of vivax relapses with chloroquine might have provided weak additional activity against P falciparum, suppressing some late recrudescence, particularly for the less effective artesunate\u2013amodiaquine. Although because most recrudescence happened within 4 weeks, and most vivax infections were not noted until after 5 weeks, this effect is unlikely to have been large. Thus for the four highly effective ACT regimens the main clinical problem after effective treatment of acute falciparum malaria in Myanmar is recurrent P vivax malaria not P falciparum.Mixed P vivax malaria. Adverse effects were generally mild, despite the high doses received by some patients, and did not substantially affect adherence. Although giving the entire 25 mg/kg dose of mefloquine as loose tablets was directly observed, suppression of recurrent vivax malaria was significantly worse than with the fixed combination, suggesting better absorption of mefloquine with the latter.This is, to our knowledge, the first comparison of all currently available fixed-dose artemisinin combination treatments for falciparum malaria. Our findings suggest that artesunate\u2013mefloquine, artemether\u2013lumefantrine, or dihydroartemisinin\u2013piperaquine are all good treatments of falciparum malaria in Myanmar. The new fixed dose artesunate\u2013mefloquine had the highest cure rates, the lowest rates of gametocyte carriage, and the most effective suppression of"}
+{"text": "Plasmodium falciparum infections as well as CQ monotherapy for P. vivax infections. Patients with P. falciparum failing therapy were re-treated with unsupervised quinine\u00a0\u00b1\u00a0doxycycline therapy and those with P. vivax with either unsupervised quinine\u00a0\u00b1\u00a0doxycycline or amodiaquine. In total, 143 patients were enrolled in the study (103 treated with CQ+SP and 40 with CQ). Early treatment failures occurred in four patients (4%) with P. falciparum and six patients (15%) with P. vivax. The failure rate by Day 28 for P. vivax was 65% (95% CI 49\u201381). After PCR correction for re-infections, the Day 42 recrudescence rate for P. falciparum infections was 48% (95% CI 31\u201365). Re-treatment with unsupervised quinine\u00a0\u00b1\u00a0doxycycline resulted in further recurrence of malaria in 48% (95% CI 31\u201365) of P. falciparum infections and 70% (95% CI 37\u2013100) of P. vivax infections. Eleven patients with recurrent P. vivax were re-treated with amodiaquine; there were no early or late treatment failures. In southern Papua, a high prevalence of drug resistance of P. falciparum and P. vivax exists both to first- and second-line therapies. Preliminary data indicate that amodiaquine retains superior efficacy compared with CQ for CQ-resistant P. vivax.To determine the level of antimalarial drug resistance in southern Papua, Indonesia, we assessed the therapeutic efficacy of chloroquine plus sulfadoxine\u2013pyrimethamine (CQ+SP) for  DespitePlasmodium falciparum in Indonesia was first documented over 30 years ago (P. vivax was only described in 1989 in northern Papua Province (formerly Irian Jaya) on the eastern border of the archipelago -resistant ears ago , the emehipelago . Within P. vivax . Other srn Papua  and elsern Papua . High ralciparum  and for P. vivax . The comlciparum  and P. vP. vivax .P. falciparum had high-grade resistance (RII or RIII) to CQ monotherapy . The corresponding failure rates of CQ plus 3 days of primaquine resulted in a Day 28 cure rate of 13% for P. vivax infections.Only one study has been published from southern Papua, which demonstrated that in 1992 52% of patients with otherapy . In thisAs part of a series of studies to rationalise antimalarial protocols in this region, we undertook a series of chemotherapeutic trails to determine the efficacy of protocols for uncomplicated falciparum and vivax malaria that were prevailing in 2004.22.1Anopheles koliensis, A. farauti and A. punctulatus . Owing to economic migration, the ethnic origin of the local population is diverse, with highland Papuans, lowland Papuans and non-Papuans all resident in the region. In view of the high number of infections in non-immune patients, local protocols recommend that all patients with patent parasitaemia are given antimalarial therapy.The study was carried out at an established rural outpatient clinic west of the city of Timika in southern Papua, Indonesia. This forested lowland area has unstable malaria transmission associated with three mosquito vectors: ctulatus . The ann2.2P. falciparum (CQ+SP) and P. vivax (CQ) in children and adults with uncomplicated symptomatic malaria. The study was based on the 2003 WHO in vivo antimalarial drug sensitivity protocol  with any parasitaemia and a fever or a history of fever during the preceding 48\u00a0h presenting to the outpatient clinic were enrolled in the study. Exclusion criteria included pregnant or lactating women, children under 10\u00a0kg, patients with signs of severity  count. Parasite counts were determined on Giemsa-stained thick films as the number of parasites per 200 WBCs. All slides were read by a certified microscopist with 10 years experience. A thick film was considered negative on initial review if no parasites were seen in 100 high power fields. A thin film was also examined to confirm parasite species and used for quantification if parasitaemia was >200 per 200 WBCs. All slides were cross-checked by another experienced microscopist.P. vivax infections) or 6 weeks . Patients were also encouraged to return to the clinic on any other day that they felt unwell. At each clinic visit a full physical examination was performed, the symptom questionnaire was completed and blood was taken to check for parasite count and haemoglobin level using a battery-operated portable photometer . Blood spots on filter paper (Whatman 3\u00a0mm chromatography paper) were also collected on Day 0 and on the day of failure.Patients were examined daily thereafter until they became afebrile and aparasitaemic. At each visit, a blood film was taken and a symptom questionnaire was completed. Patients were then seen weekly for 4 weeks (2.5n\u00a0=\u00a06) and 2.5% and 5.3% at 100\u00a0ng/ml (n\u00a0=\u00a06). The limit of quantitation was 5\u00a0ng/ml for CQ and 2.5\u00a0ng/ml for DCQ. The minimum effective concentration (MEC) of CQ+DCQ was defined as 30\u00a0ng/ml plasma for P. falciparum and 15\u00a0ng/ml for P. vivax  were assayed by HPLC as described previously . The intP. vivax . These cP. vivax .2.6P. falciparum  or CQ for pure P. vivax infections were administered. The CQ+SP regimen consisted of CQ  and SP was given as a single dose on Day 0 (25\u00a0mg/kg sulfadoxine and 1.25\u00a0mg/kg pyrimethamine). All drug administrations were supervised and participants were observed for 30\u201360\u00a0min to exclude adverse reactions and to ensure the medication was not vomited. If vomiting occurred within 60\u00a0min, the whole dose was repeated once. If vomiting occurred again within 60\u00a0min, the patient was withdrawn from the study. Paracetamol was given if the axillary temperature was \u226538\u00a0\u00b0C.Standard treatment courses of CQ+SP for P. vivax within 28 days were given a 3-day course of supervised amodiaquine if they consented . Primaquine (15\u00a0mg base/kg of body weight for 14 days) was administered to those individuals with P. vivax infection or mixed infection on Day 28 of their participation in the study.Those patients failing therapy were re-treated with quinine  plus doxycycline (100\u00a0mg twice a day for 7 days) if \u22658 years age and not pregnant. From May 2004, those patients with reappearance of 2.7P. vivax infections and by Day 42 for P. falciparum infections. The WHO criteria  at 24\u00a0h (parasitaemia on day 1/parasitaemia on admission) were assessed using the receiver operator curve and Youden's Index calculated as sensitivity plus specificity minus 1.Data were double entered and validated using EpiData 3.02 software  and analysis was performed using SPSS for Windows . The Mann\u2013Whitney 2.9http://www.clinicaltrials.gov/ct) as NCT 00157859.The study was approved by the Ethics Committee of the National Institute of Health Research and Development, the Indonesian Ministry of Health , the Ethics Committee of Menzies School of Health Research  and the Oxford Tropical Research Ethics Committee. Informed consent was obtained from all adult participants and from parents of children. An independent Data Safety Monitoring Committee (DSMC) was established prior to commencing the study and was asked to review the progress of the study after 100 patients had been enrolled. The trial was registered with the clinical trials website  had pure P. vivax infections and were treated with CQ monotherapy. Baseline characteristics are given in Between April and September 2004, 143 patients with uncomplicated malaria were enrolled in the study. In total, 103 (72%) had 3.1P\u00a0=\u00a00.003). In total, 6.3% of patients (9/143) were unable to tolerate their medication owing to recurrent vomiting (seven in the CQ+SP group and two in the CQ group), only one of whom was an adult. Five of these patients tolerated changing medication to oral quinine, but two children required i.v. quinine. A further 7.0% of patients (10/143) withdrew consent prior to completion of therapy (four children and six adults). One patient with P. vivax infection on admission was treated with CQ but was found to have mixed P. falciparum and P. vivax infection on Day 1 and treatment was changed to quinine. The study profile is shown in Early vomiting within the first hour of drug administration occurred in 8% of patients (8/103) treated with CQ+SP and 10% of patients (4/40) treated with CQ. Vomiting after CQ administration did not differ between the species of infection, but was 7.2-fold (95% CI 1.7\u201331) higher in children compared with adults: 19% (13/68) and 3% (2/75), respectively  in the CQ+SP group and 26.3\u00a0mg/kg (SD 2.6\u00a0mg) in the CQ group. Follow-up to Day 28 or day of failure in these patients was achieved in 90% (77/86) of those treated with CQ+SP and 84% (31/37) following CQ administration.3.2P. falciparum infection compared with 6.7% of patients (1/15) with mixed infection (P\u00a0=\u00a00.5). Although ETF was more likely in patients with pure P. vivax infections treated with CQ , the difference did not reach statistical significance (P\u00a0=\u00a00.06). Four patients developed warning signs or markers of severity and required rescue with i.v. quinine. One adult with P. falciparum developed convulsions and coma, and one child with falciparum malaria developed respiratory distress on Day 1. An adult and a child, both with vivax malaria, developed severe vomiting and diarrhoea on Day 1. The remaining six patients were categorised as having ETF due to delayed parasite clearance. Early treatment failure was 8.8-fold (95% CI 2.7\u201328) higher in infants compared with older children and adults (P\u00a0<\u00a00.001).Early treatment failure (ETF) with CQ+SP occurred in 4.2% of patients (3/71) with P. falciparum and 7.5 (range 1\u2013180) for P. vivax (P\u00a0<\u00a00.001). By Day 3, 29% of patients (8/28) infected with pure P. vivax had failed to clear their parasitaemia compared with 38% (5/13) of patients with mixed infection and 7% of patients (4/56) with pure P. falciparum (P\u00a0=\u00a00.004). However, fever clearance times were generally rapid, with 97% of patients (30/31) having defervesced within 48\u00a0h.The median PRR at 48\u00a0h was 57 (range <1\u20131512) for 3.3P. falciparum, 18 had P. vivax and 8 had mixed infections  compared with 74% (95% CI 50\u201398) for patients with mixed infections and 65% (95% CI 49\u201381) for pure vivax infections  of children compared with 59% (95% CI 38\u201380) of adults . The same trend was apparent 28 days after treatment of vivax malaria: 82% (95% CI 64\u2013100) of children and 49% (95% CI 24\u201374) of adults (P\u00a0=\u00a00.004). Survival plots of the cumulative incidence rate of therapeutic failure are presented in The cumulative Day 28 failure rate for fections . By Day P. falciparum or mixed infections on enrolment and at treatment failure. In 17 cases the recurrent isolate was identified as a true recrudescence, in 10 cases as a new infection and in 6 cases the result was indeterminate. The failure rate of P. falciparum by Day 42, corrected by PCR and including ETFs, was 48% (95% CI 31\u201365).Parasite genotyping was performed in 77% of patients (33/43) with P. vivax but not with P. falciparum. The PRR of P. vivax 48\u00a0h after starting CQ therapy was 20 (range 4\u2013180) for those vivax successfully treated compared with 7 (range 1\u2013148) in those with recurrence of P. vivax by Day 28 (P\u00a0=\u00a00.05). The PRR of P. vivax at 24\u00a0h was significantly correlated with the day of recurrence . Using Youden's Index, the best cut-off at 24\u00a0h was a PRR of 7.4; parasite reduction below this level predicted subsequent treatment failure with 70% sensitivity and 63% specificity.The initial speed of parasite reduction was correlated with subsequent treatment failure in patients infected with 3.4P. falciparum or mixed infections and in 27% of patients (7/26) presenting with vivax malaria, plasma CQ concentrations were above the MEC. The plasma CQ concentrations on Day 7 were significantly lower in children  compared with adults  (P\u00a0=\u00a00.0001) but did not differ by species of infection. The difference was still apparent after excluding those with early vomiting. Three patients (one adult and two children) had undetectable CQ concentrations on Day 7; all of these patients failed therapy within 21 days.On admission, 39% of patients (35/89) had measurable concentrations of plasma CQ see . In 29% P. falciparum or mixed infections. In these patients, the median concentration of CQ+DCQ was 43\u00a0ng/ml (range 0\u2013242\u00a0ng/ml), exceeding the MEC in 61% of cases (14/23). In patients with P. vivax, CQ concentrations were measured in 7 (39%) of 18 patients during follow-up. The median CQ+DCQ concentration on the day of recrudescence was 44\u00a0ng/ml (range 25\u2013108\u00a0ng/ml), with all of these patients having concentrations in excess of the MEC.Plasma samples were available for CQ analysis in 49% of patients (23/47) who recrudesced with 3.5P. falciparum compared with 18% (7/40) with P. vivax and 38% (5/13) with mixed infections (P\u00a0=\u00a00.004). By Day 7, none of the patients with P. vivax infections had gametocytes present on blood film examination compared with 47% (32/68) following treatment of P. falciparum and mixed infections (P\u00a0<\u00a00.001). Following P. falciparum infection, gametocyte carriage was still present in 29% of patients (7/24) on Day 28. At the time of recrudescence, gametocytes were present in 34% of patients (22/64) failing therapy, with no difference between species of infection.On admission, gametocytes were present in 9% of patients (8/87) infected with 3.6P. falciparum or mixed infections had treatment failures (2 delayed parasite clearance and 47 with late recurrence). Four patients refused re-treatment. A child aged 5 years who had a reappearance of P. falciparum that was associated with vomiting and tachypnoea on Day 12 was treated with i.v. quinine as an inpatient. In total, 44 patients agreed to be re-treated with 7 days of unsupervised quinine . There were no ETFs but 67% of patients (22/33) successfully followed-up had a reappearance of parasitaemia by Day 28 . By Day 28 the uncorrected failure rate of unsupervised quinine\u00a0\u00b1\u00a0doxycycline for P. falciparum was 48% (95% CI 31\u201365) in this cohort. There was no significant difference in those receiving monotherapy or additional doxycycline.Overall, 49 patients with P. vivax treatment failures (4 delayed parasite clearance and 18 with late recurrences). A 16-year-old male re-presented on Day 22 with a recurrence of P. vivax associated with tachypnoea and a 3-day history of vomiting everything; he was treated as an inpatient. One adult refused re-enrolment. In total, 20 patients (15 children and 5 adults) agreed to re-treatment and follow-up. Nine patients were re-treated with 7 days of unsupervised quinine, of whom six were successfully followed-up: five had a further reappearance of P. vivax and one had an adequate parasitological response. Eleven patients were re-treated with amodiaquine (30\u00a0mg base/kg) over 3 days. There were no ETFs. Three were successfully followed to Day 28: two had an adequate parasitological response and one child had a further reappearance of P. vivax on Day 31.Twenty-two patients had 4P. falciparum or mixed infections treated with CQ+SP and 16% of those infected with P. vivax treated with CQ alone required early rescue therapy. The failure rate for P. vivax had risen to 65% by Day 28 and that for P. falciparum or mixed infections had risen to 71% by Day 42. When this degree of resistance became apparent, the study was referred to an independent DSMC who felt that it was inappropriate to continue using these treatment regimens and the study was stopped. These failure rates were based on a per-protocol analysis to define resistance levels of the parasite. In practice a further 6% of patients were intolerant of CQ and another 8% failed to complete therapy. If one assumes that all these patients would have had a recurrence of malaria during the follow-up period, then the overall Day 42 cure rate during the study period would have been nearer 32% for P. falciparum and mixed infections and 24% for P. vivax. These levels of drug resistance are undermining the local malaria control strategy and contributing a considerable burden of mortality and morbidity upon the local communities.This study highlights that the antimalarial policy being used in 2004 in southern Papua, Indonesia, was no longer effective. Almost 5% of patients with P. falciparum 2.4-fold (95% CI 1.3\u20134.3) higher compared with adults. This may be attributed in part to children being less tolerant of CQ therapy as demonstrated by 7.2-fold (95% CI 1.7\u201331) higher rates of early vomiting and 60% lower plasma CQ levels on Day 7. However, as in other endemic areas, the lower failure rates in adults are also likely to represent a degree of immunity helping to clear resistant blood-stage parasites from the bloodstream  more likely to suffer an ETF. Older children also had an increase risk of late treatment failure of odstream .P. falciparum and P. vivax, but until recently the degree of resistance of P. vivax has tended to lag behind that of P. falciparum. Our data from southern Papua show CQ resistance in P. vivax was significantly worse than that of P. falciparum. The 48-h PRR of 7.5 for P. vivax was significantly lower than the 57 observed for P. falciparum. These figures compare with 438 for CQ-sensitive P. vivax isolates from Thailand . This adds further evidence that the failure rates observed in our study represent true CQ-resistant P. vivax rather than poor drug absorption.Conducting an in vivo study in an endemic area is confounded by re-infection. However, in this study PCR genotyping confirmed that 63% of the e stages . A revie 28 days . In this isolate . In our P. falciparum, it is effective against all stages of P. vivax and in combination with CQ may improve efficacy against blood stages of P. vivax , a regimen to which adherence is poor in practice. Furthermore, in Papua it is unlikely that co-administration of an unsupervised 14-day course of primaquine would have prevented relapse of resistant isolates at the tail end of CQ treatment because of poor compliance with the lengthy primaquine regimen . In our study, the numbers completing a 28-day follow-up following re-treatment were small but highlighted the 48% failure rate for pervised . QuinineP. vivax . In totaP. falciparum and P. vivax in southern Papua. The current level of resistance is likely to be the most important factor in the inability of the established malaria control programme to decrease the large burden of disease. Alternative strategies involving the use of artemisinin-based combination therapies are in progress.In summary, our findings highlight the high levels of treatment failure both of first- and second-line therapies for The authors have no conflicts of interest concerning the work reported in this paper."}
+{"text": "We reviewed the efficacy of a 3-day regimen of mefloquine and artesunate regimen (MAS3. The proportion of patients with parasitaemia persisting on day-2 increased significantly from 4.5% before 2001 to 21.9% since 2002 (p<0.001). Delayed parasite clearance was associated with increased risk of developing gametocytaemia . Gametocytaemia on admission and carriage also increased over the years . MAS3 efficacy has declined slightly but significantly , although efficacy in 2007 remained well within acceptable limits: 96.5% . The in vitro susceptibility of P. falciparum to artesunate increased significantly until 2002, but thereafter declined to levels close to those of 13 years ago . The proportion of infections caused by parasites with increased pfmdr1 copy number rose from 30% (12/40) in 1996 to 53% (24/45) in 2006 .3,264 patients were enrolled in prospective treatment trials between 1995 and 2007 and treated with MASArtesunate-mefloquine remains a highly efficacious antimalarial treatment in this area despite 13 years of widespread intense deployment, but there is evidence of a modest increase in resistance. Of particular concern is the slowing of parasitological response to artesunate and the associated increase in gametocyte carriage. Plasmodium falciparum parasites in this region have developed significant resistance to commonly used antimalarials, except the artemisinin derivatives. The treatment of uncomplicated P. falciparum malaria in Thailand has been modified several times during the past 30 years to counter the rapid emergence and spread of drug resistance 2, (artesunate 6 mg/kg repeated once 24 hours later), was introduced in Thailand in 1995, under strict supervision, exclusively in three provinces (including the northwestern border of Thailand) where failure rates with mefloquine (15 mg/kg) -sulfadoxine-pyrimethamine (MSP) had risen above 50% 2 was extended to all provinces along the borders with Myanmar and Cambodia, following evidence from these areas of inadequate clinical cure rates with mefloquine alone, or MSP The Thai-Myanmar border harbors some of the world's most drug resistant malaria parasites. 3, to ensure that artesunate covered two parasite asexual life-cycles, thereby reducing the parasite biomass exposed to mefloquine alone 3, one of the ACT regimens currently recommended by the WHO, is also used in clinics serving the migrant population, located along the Thai-Myanmar border, and since January 2008 has replaced MAS2 in the Thailand National protocol.In camps for displaced persons located along the Thai-Myanmar border, mefloquine and artesunate combination therapy has been evaluated since 1991 and deployed systematically since 1994 3 efficacy in the thirteen years between January 1995 and December 2007 in camps for displaced people and clinics for migrant workers located along the northwestern border of Thailand.This report presents a continuous description of the in vivo, in vitro and molecular correlates of MASP. falciparum and P. vivax are the two predominant species, P. malariae is occasionally found, and P. ovale is rare. All age groups are affected and nearly all the P. falciparum infections are symptomatic. Since 1995, the Shoklo Malaria Research Unit (SMRU) has conducted 7 prospective randomized open-label controlled chemotherapeutic trials of new antimalarial drugs in which MAS3 was one of the treatment arms 3 drug efficacy The epidemiology of malaria in the area has been described in detail elsewhere 3 efficacy: in vivo, in vitro and molecular assessment of pfmdr1 copy number. The in vivo efficacy of MAS3 has been monitored annually Three methodologies were use to evaluate MASP. falciparum parasites from 1995 onwards pfmdr1 gene copy number is the major molecular determinant of mefloquine susceptibility and provides an important tool for monitoring drug resistance In vitro antimalarial drug susceptibility studies were carried out on fresh isolates of P. falciparum malaria or a mixed infection, were enrolled after giving written  or witnessed verbal consent . Severely ill patients and hyperparasitaemic patients  were treated with different regimens and were not included in this analysis. Patients who had received treatment with mefloquine in the previous 63 days or who had failed their last antimalarial drug treatment, pregnant women, children <5 kg in body weight, and patients with signs of severity or concomitant disease were excluded from the studies. All patients were examined daily until fever subsided  and until parasite clearance , and then followed weekly for 6 to 9 weeks. In the event of parasite reappearance during follow up patients were re-treated and followed-up again. The clinical trials conducted at SMRU were all reviewed by the Ethics Committee of the Faculty of Tropical Medicine Mahidol University and the Oxford Tropical Research Ethics Committee.Specific study procedures have been described previously for each randomized controlled trial Patients received oral artesunate  or Arsumax\u00ae  4 mg/kg/day for 3 days once daily, and mefloquine 25 mg base/kg, Lariam\u00ae  or Mequin\u00ae . Children who were unable to swallow complete tablets received the same weight-adjusted dosage of crushed tablets mixed with water. Each drug administration was observed in all patients.Patients received mefloquine either as a single dose of 25 mg/kg , as a split dose of 15 mg/kg and 10 mg/kg given 24 hrs apart on the second and third day of treatment (the current protocol), or as a split dose of 8 mg/kg given once a day for 3 days in two trials In the 2006 dose-assessment study P. falciparum parasite on a blood smear between day-5 and 45 of follow-up (42 days+3 days to take into account a delay in the last follow-up). Recrudescent infections were differentiated from new infections using 3 loci genotyping as described previously Recurrent infection was defined as the presence of asexual forms of In view of the importance of the parasite reduction ratio P. falciparum malaria infection attending the research clinics. Only primary infections were assessed. Isolates were collected if the parasite density was at least 5 parasites/1,000 red blood cells (RBC), from patients attending the clinic, whether enrolled in one of the studies or not. Venous blood (5 ml) was collected into a sterile Vacutainer\u00ae tube containing 0.05 ml Potassium-EDTA. Samples were kept at room temperature and transported within the next 4\u20136 hours to the main laboratory, where they were set up in continuous culture immediately. The minimum parasitaemic threshold required for parasite growth in culture, as well as the necessity to take 5 ml of blood limited the evaluation of P. falciparum drug susceptibility to patients with higher parasitaemia who agreed to have a venopuncture .Parasites isolates were obtained from patients with acute 50 measurements was assessed regularly using cloned K1 isolates of P. falciparum.Drug susceptibility testing by hypoxanthine uptake has been described previously by Brockman pfmdr1 copy number was available from whole blood , or from 50 \u00b5l of capillary blood transferred to Whatman\u2122 filter paper. In order to investigate changes in resistance markers over time parasite genotypes were assessed each year since 1996, with the exception of 1998, 2000 and 2007, using venous blood taken before treatment from a random selection of patients with primary P. falciparum infections. Randomisation was stratified by age and sex to represent the population seeking care for malaria in the outpatient's clinics.DNA for molecular analysis of Pfmdr1 copy number was assessed by quantitative PCR  using the methods described previously pfmdr1 were detected by nested PCR-RFLP methods as described P. falciparum parasites. PCR results were deemed indeterminant if only one of the three gene locus was amplified, or if analysis of genetic frequency of alleles in the population revealed a recrudescent pair as statistically insignificant p<0.10), as well as other potential risks factors such as age, sex, admission parasitaemia, mixed parasite infection, and type of drug regimen.Data were analyzed using SPSS for Windows\u2122  and STATA . Differences in settings, sequence randomization, mefloquine treatment timing, and study procedures were included in the assessment of heterogeneity between studies using the Cochran Q test and the I2 test Gametocytaemia was considered as a binary variable (present/absent) and analyzed on admission, at day-1, 2, 3, 7, and weekly thereafter until day-42. Gametocyte carriage rates were presented as person-gametocyte-weeks , and expressed per 1,000 weeks of follow-up. Prevalence rate on admission, overall carriage rate and among patients without gametocyte on admission were calculated yearly.50 values, and coefficients of variation calculated. Temporal trends for mefloquine and artesunate IC50 were analyzed on the ln-transformed data.In vitro dose-response curves were analyzed by fitting the data to an inhibitory E-max pharmacokinetic model using WINNONLIN  and the IC50 and the pfmdr1 copy number using the isolates for which both the IC50 and the copy number were available.Regression analysis on the ln-transformed values was performed to investigate association between the ICpfmdr1 copy number, gametocyte carriage rates, and the proportions of patients remaining febrile and/or parasitaemic on day 2\u20134 over the years were analyzed by \u03c72 test for trend.Changes in 3 combination was evaluated in 3,264 patients with uncomplicated P. falciparum malaria infections, enrolled in 9 trials, between 1995 and 2007 .The efficacy of the MASand 2007 Table 1.n\u200a=\u200a625) of patients did not complete the 42-day follow-up period. The main reasons for loss of follow-up were change of address and employment obligations (592/625). The drug combination was generally well tolerated and few patients reported having serious adverse events 3 and required parenteral treatment; three deaths were reported . None were related to malaria or to an adverse drug event. There were no significant differences in age, sex and presence of mixed infections on admission between patients lost to follow up and those who completed it.The admission characteristics of patients are presented in Changes in fever clearance times were evaluated in the 2,029 (62%) of patients who presented with fever Table 3.p<0.001, test for trend). Before 2001, 95.5%  were aparasitaemic within 48 hours compared to 78.1%  between 2001 and 2007 . By day-3, 99.8%  patients had cleared their parasitaemia before 2001, but this fell to 95.9%  thereafter, (p<0.001), (p\u200a=\u200a0.007).Parasite clearance times were available for 3,218 patients (98.6%), Table 3.P. falciparum+P. vivax) infection, and haematocrit on admission, the increase in parasite clearance time over the 13 year study period remained significant  with a 0.05% annual increase of patients with delayed parasitaemia. In the same multivariate analysis, parasitaemia on admission , gametocytaemia on admission , and site of recruitment  were significantly associated with failure to clear parasitaemia within 48 hours.After accounting for age, sex, presence of a mixed , resulting in a significant increase in gametocyte carriage rate over the study period , .Overall 6.3%  of patients had patent gametocytaemia on admission and this proportion increased from 1.2% (6/482) in 1995 to 7.2% (10/139) in 2007 ( trend), Table 4.p\u200a=\u200a0.009), or on day-3 or later , ; 95% CI, 7\u201313) than those whose parasitaemia was cleared on day-2 , Figure 2p\u200a=\u200a0.082, test for trend), but was related to the parasite clearance time; it increased from 2.9%  among patients who cleared their parasitaemia on day-1 or on day-2 to 5.9% (26/437) among those who cleared their parasitaemia on day-3 or later, p\u200a=\u200a0.001.The majority of patients without gametocytaemia on admission, who later developed gametocytaemia, did so on day-1 (43%). The proportion of patients with gametocyte appearance during follow-up did not increase significantly over the years (p\u200a=\u200a0.018), parasitaemia on admission , year of recruitment , and site of recruitment , patients who had a delayed parasite clearance (3 days or longer) remained at a higher risk of developing gametocytaemia .In the multivariate analysis, after controlling for gametocytaemia on admission  were new infections, 108 (32%) were recrudescent infections, and in 48 (15%) cases the PCR result was inconclusive (n\u200a=\u200a15) or samples were not available (n\u200a=\u200a33). The median time to recrudescence occurred at 21 days of follow-up (range 7\u201342 days); although time to recrudescence has increased by one week since 2006, it has not changed significantly over the 13 year period, and neither has the time to re-infection .Parasitological efficacy (non PCR-adjusted) increased significantly over time as the risk of p<0.001). However the efficacy of MAS3 in 2007 remained above 95%: 96.5% , .A small but statistically significant decline in PCR-adjusted parasitological efficacy (assessed at day-42) was observed over the study period , Figure 3P. falciparum isolates were assayed for in vitro drug susceptibility during the study period; artesunate IC50 could be determined satisfactorily in 1,179 (75%) isolates and mefloquine IC50 in 1,195 (76%). Between 1995 and 2001 there was a significant decrease in the IC50s of both artesunate  and mefloquine . From 2002 this trend reversed for artesunate with IC50s increasing 41% per year (p<0.001) until 2006, although decreasing in 2007  to 37.1 nM/l , and the IC50 of artesunate from 6.47 nM/l  to 4.19 nM/l .In total, 1,581 pfmdr1 copy number successfully determined in 84% (444/531) of these isolates. The proportion of infections with two or more pfmdr1 copy number rose from 30% (12/40) in 1996 to 53% (24/45) in 2006 ,  were infected with parasites with increased pfmdr1 copy number. Infections with increased pfmdr1 copy number were at greater risk of recrudescence by day-42 compared to single copy number infections . There was no difference in the proportion of patients parasitaemic on day-2 in those with single  or increased pfmdr1 copy number , even after controlling for baseline parasitaemia.Among 408 patients treated with MASpfmdr1 was associated with a 2.9 fold increase in mefloquine IC50  and a 1.7 fold increase in artesunate IC50 . The rise in IC50 associated with increased pfmdr1 copy number was greater during the period 1996\u20132001 compared to thereafter for both artesunate  and mefloquine . In contrast there was no change in the risk of treatment failure associated with increased copy number over the study period.In the 394 isolates with in vitro and molecular data, amplification of 3) as first line treatment for uncomplicated P. falciparum malaria in 1994 in the camps for displaced persons. Despite the prevailing high levels of mefloquine resistance, cure rates increased to more than 90% and MAS3 had a spectacular impact on the incidence of P. falciparum malaria cases in the camps P. falciparum isolates to mefloquine improved significantly until 1999, attributed to the eradication of the more mefloquine resistant \u201cstrains\u201d and the return of more mefloquine-sensitive \u201cwild-type\u201d parasites 3, which are less fit than their mefloquine-sensitive counterparts The rapid development of mefloquine resistance in Tak province on the Thai-Myanmar border in the early 1990s led to the introduction of the three day combination of artesunate and mefloquine  and within the accepted drug sensitivity levels. Mefloquine susceptibility has fluctuated widely, and its association with the pfmdr1 copy number seems to have changed over the years. These changes in antimalarial activity in vitro have not been associated with a sharp reduction in cure rates, as happened when mefloquine was used as monotherapy pfmdr1 copy number pfmdr1 copy number, and yet there are indications that artemisinin efficacy has declined as evidenced by an increase in the parasite clearance time. The decrease in in vitro susceptibility to artesunate after 2002 could not be explained by mechanisms associated with mefloquine resistance (predominantly pfmdr1 amplification). The correlation between artesunate and mefloquine IC50s evident before 2002 was no longer evident afterwards. This suggests the emergence of a new factor affecting artesunate susceptibility, which could include either novel mutations within the pfmdr1 gene, increased gene expression or alternate genetic events.Over the subsequent 13 years of continuous MAS3 are more concerning. Rapid parasite clearance is the pharmacodynamic hallmark of artemisinin and its derivatives. This is associated with reduced gametocyte carriage compared with other antimalarial drugs. There has been a small but significant slowing of parasite clearance, reflected by the increasing proportion of patients with parasitaemia persisting on the second and subsequent days following the start of treatment. Up until 2001 less than 7% of patients were parasitaemic on the second day of follow-up; however this figure has more than doubled in recent years. The delay in parasite clearance has been associated with a small increase in treatment failure rates, presumably reflecting a slight reduction in initial parasite reduction ratios, which results in a greater number of parasites for mefloquine to remove in the third and subsequent post treatment replication cycles P. falciparum infections rather than infections mixed with P. vivax on admission, a parasitaemia over 40,000 parasites per \u00b5l on admission and site have been found to be independent risk factors for treatment failure following MAS3The changes in the early in vivo parasitological responses to MASP. falciparum population resulting in small but significant increases in parasite tolerance to the artesunate component of MAS3. These are considerably less in magnitude than those reported recently from Western Cambodia pfmdr1 amplification is a contributor to artemisinin susceptibility, the main cause of this reduction in artesunate susceptibility is not known. Of particular concern is the increased transmissibility of these tolerant parasites. The changes are small, but they justify a very close monitoring of the susceptibility of P. falciparum in this area, and in particular vigilance to detect early emergence of higher levels of artemisinin resistance. The development of resistance to artemisinin and its derivatives would be a global disaster for malaria control as current treatment regimen are dependant on this class of antimalarial drugs.Despite some worrying signs, which could only have been detected by these very large prospective studies, the 3-day mefloquine-artesunate combination is still more than 95% effective in Tak province, 13 years after its introduction in the camps for displaced persons. Considering that resistance to mefloquine monotherapy had reached very high levels before this combination was deployed, and that surrounding areas deployed different dose regimens, its longevity is remarkable. Our data suggest that changes have occurred in the prevalent"}
+{"text": "Objectives. The objectives of the study were (i) to evaluate the efficacy of combination drugs, such as artesunate + sulphadoxine-pyrimethamine (AS\u2009+\u2009SP) and amodiaquine + sulphadoxine-pyripethamine (AQ\u2009+\u2009SP) in treatment of uncomplicated falciparum malaria (ii) to differentiate recrudescence from reinfection by analysing msp-1 and msp-2 genes of Plasmodium falciparum in treatment failure cases. Methods. We carried out an in vivo study in the year 2005 in 206 children between 6 to 59 months age groups. Of the 206, 120 received AQ\u2009+\u2009SP, and 86 received AS\u2009+\u2009SP. A clinical and parasitological followup during 14 days was undertaken. Finger-prick blood sample from each patient was taken on Whatman filter paper (no. 3) on days 0, 7, 14 and also the day when the parasite and symptoms reappeared for PCR analysis. Results. Late treatment failure was observed in 3.5% (4/114) with AQ\u2009+\u2009SP, and 2.5% (2/79) with AS\u2009+\u2009SP. The success rate was 96.5% with AQ\u2009+\u2009SP and 97.5% with AS\u2009+\u2009SP. No deaths and severe reactions were recorded. Out of the 6 treatment failure cases, one was reinfection as observed by PCR analysis of msp-1 and msp-2 genes on day 14. Discussion. Both the combinations found to be efficacious and safe and could be used as a first-line treatment for uncomplicated falciparum malaria in Equatorial Guinea.  All of thefailure cases received rescue treatment with oral quinine and cured see . After tBoth combinations were well tolerated, no deaths werenotified and no severe reactions were recorded, except 3 patients (2.5%) hadcomplaints of itching in AQ + SP treatment group, they were given antihistamine. Fourpatients (3.3%) in AQ + SP and 1 patient (1.2%) in AS + SP treatment groups vomited,which could not be confirmed whether this was the consequence of malaria or thetreatment effect.parasiteclearance (P < .001). In AS + SP treatment group, 77.2% (61/79) and in AQ + SP treatmentgroup, 94.7% (108/114) presented parasitemia on day 1. On day 2, 3.8% (3/79) ofAS + SP and 55.3% (63/114) of AQ + SP treatment groups were parasitemic. By day 3,all of them were negative.The learance  was fastgametocyte carriage was less than 5% in both arms. After receivingtreatment, gametocyte density was found to be higher in AS + SP patients thanAQ + SP. With the first combination, by day 2, 12% of children had gametocytes onthe thick film, compared with the 7.5% with AQ + SP. By day 3, the 7.8% withAS + SP and the 6% with AQ + SP had gametocytes. By day 14, gametocytemia was under2% with both the combinations.Baseline Haematocrit at enrolment showed that 37%of the children had moderate anaemia with haematocrit 24\u201329, 49% had mildanaemia with haematocrit 30\u201334, and 14% were nonanaemic. After treatment themoderate anaemia decreased from 37% to 23%, and nonanaemic cases increased from14% to 22%. Molecular correction was done to distinguish recrudescent from reinfection in 6treatment failure cases: 4 cases from Bata, 3 in the AQ + SP combination,and 1 in the AS + SP, and 2 in Malabowith twocombinations (AQ + SP and AS + SP).msp-1 allelic family, and msp-2 detected in the six treatmentfailure cases. We could confirm the resistantcases from Bata by molecular analysis; because the parasite genotypes detectedby PCR on day 14 were similar with those on day 0. However, in the AQ + SP treatmentfailure cases from Malabo,one of them was reinfection because a different set of genotypes appeared onday 14 as shown in falciparum malaria. Keeping this in view, present study was undertaken on thedrug sensitivity of two different therapeutics combinations. This study wouldbe helpful in National Malaria Control Program of Equatorial Guinea tointroduce a new combination therapy for malaria treatment.There is a need to give a thought about any monotherapy as afirst-line treatment in case of patients report with uncomplicated Two combinations were assessed, the combinations AS + SP, wherethe AS is an efficacious drug and reduces the parasitaemia faster than others,and the second combination, AQ + SP, is also an effective combination but with therisk to develop resistant in due course of time, but more accessible to thepopulation when compared with artemisinin derivatives.falciparum malaria in Equatorial Guinea. These results are similar to other studies madein different African's areas. A study carried out in Tanzaniain an area of high malariatransmission compared the efficacy and safety of AQ and SP in monotherapy andAQ + SP as a combination. The study demonstrated that AQ + SP combination was safe, and the combination of the two drugs had higher efficacy (96.2%) thanmonotherapy [The study demonstrated that both combinations are safe andefficacious for the treatment of nonsevere otherapy . In a sotherapy . A studotherapy . These fficacy 9.2% thanIn general terms, drugs were well tolerated, with a lowpercentage of adverse effects. In the group that received AQ + SP, a small numberof children required retreatment after vomiting after treatment. Severereactions with amodiaquine as neutropenia have been described also in otherpublications. In one study, it was shown that the risk was associated with amodiaquine used as a weeklyprophylaxis, not like a treatment . In a sClearance of parasites was faster in case of combinationwith artemisinin derivatives, due to the mode of action of artemisine, whichreduces parasitaemia by a factor of 10 in each cycle. In Uganda, wasdescribed the difference in parasitemia clearance using a combination with andwithout artemisinin derivatives. By day two, in the first case just 3% werepositive compared to the second group, where 48% were positive . These Contrary to other studies, we have not seen anydifference in gametocytes carriage. From this study, we could not say that usingof artemisinin-based combination therapies may reflect in decrease of malariatransmission in the area. One study in Tanzania, where different drugs inmonotherapy and combinations were compared, demonstrated that AQ + SPgametocytes on day 14 were 25.7% in children, which was double than usingthe combination with AS, where just 11.9% of children had gametocytes .Both the combinations had SP. It is necessary to assess other therapieswithout this drug due to the following reasons. First, SP is the only safe drugactually to be used as Intermittent Preventive Treatment for pregnant women. Following WHO recommendations, we need to protect the future development ofresistance to this drug. And the second reason is that SP has a resistancearound 18% in monotherapy . Drugs that have been used for a long time as monotherapy and havealready demonstrated resistance have the risk to develop resistance incombination in due course of time.msp-1 and msp-2 genes of P. falciparum, or other polymorphic markersas microsatellites [From the results of this study, we may suggest that inall in vivo studies a molecular assay for discriminating recrudescencefrom reinfection should be done. The molecular correction is the only methodthat allows us to give a real data about the level of resistance detected in anendemic area. This way, it is possible to analyse different markers like tellites .We need to continue the epidemiological surveillanceand monitoring resistance in order to give enough information to the National MalariaProgram of Equatorial Guinea for revising malaria treatment policy in thecountry. This should be done as per the recommendation of WHO before the drug resistancereaches at levels not allowed to be used.falciparum malaria. To choose one ofthem will depend on economic resources of Equatorial Guinea in the moment tochange Malaria Policy of the country; the molecular correction givescomplementary information to strengthen the findings of in vivo studies carriedout in areas with a high level of transmission.The conclusions of this study are that both the combinations are safe and efficaciousfor their use as a first-line treatment in case of nonsevere"}
+{"text": "Sulphadoxine-pyrimethamine (SP) is the only single dose therapy for uncomplicated malaria, but there is widespread resistance. At the time of this study, artemether-lumefantrine (AL) and chlorproguanil-dapsone (CPD), both multi-dose regimes, were considered possible alternatives to SP in Malawi. The aim of this study was to investigate the impact of poor adherence on the effectiveness of AL and CPD.Children \u226512 months and adults with uncomplicated malaria were randomized to receive AL, CPD or SP. Adherence was measured using a questionnaire and electronic monitoring devices, MEMS\u2122, pill bottles that recorded the date and time of opening. Day-7 plasma dapsone or lumefantrine concentrations were measured to examine their relationship with adherence and clinical response.841 patients were recruited. The day-28 adequate clinical and parasitological response (ACPR) rates, using intention to treat analysis (missing data treated as failure), were AL 85.2%, CPD 63.7% and SP 50%. ACPR rates for AL were higher than CPD or SP on days 28 and 42 . CPD was more effective than SP on day-28 (p = 0.01), but not day-42.Very high adherence was reported using the questionnaire, 100% for AL treated patients and 99.2% for the CPD group. Only three CPD participants admitted missing any doses. 164/181 (90.6%) of CPD treated patients took all their doses out of the MEMS\u2122 container and they were more likely to have a day-28 ACPR than those who did not take all their medication out of the container, p = 0.024. Only 7/87 (8%) AL treated patients did not take all of their doses out of their MEMS\u2122 container and none had treatment failure.Median day-7 dapsone concentrations were higher in CPD treated patients with ACPR than in treatment failures, p = 0.012. There were no differences in day-7 dapsone or lumefantrine concentrations between those who took all their doses from the MEMS\u2122 container and those who did not. A day-7 lumefantrine concentration reported to be predictive of AL treatment failure in Thailand was not useful in this population; only one of 16 participants with a concentration below this threshold (175 ng/ml) had treatment failure.This study provides reassurance of the effectiveness of AL, even with unsupervised dosing, as it is rolled out across sub-Saharan Africa. Self-reported adherence appears to be an unreliable measure of adherence in this population. In 2007, Malawi switched its first line therapy for uncomplicated malaria from sulphadoxine-pyrimethamine (SP) to artemether-lumefantrine, a fixed-ratio combination of artemether (AM) with lumefantrine (LU). The need to take AL twice daily for three successive days means there is a larger potential for poor adherence with AL compared to the single dose regimen of SP.As part of efforts to optimize the operational use of AL, it is important to assess the effect of poor adherence to (compliance with) this regime on clinical outcome. This paper describes a large single centre open label randomized clinical trial to investigate the effect of adherence upon the effectiveness of AL and chlorproguanil-dapsone  in comparison to SP in a 'real-life' setting in adults and children with uncomplicated malaria. The study was initially designed to focus primarily on CPD, an anti-folate combination, previously shown to be effective in Africa in areas with high SP resistance . SubsequP. falciparum monoinfection on the blood slide, iv) Hb \u22657 g/dl, and v) no features suggesting severe malaria or a concomitant illness. Pregnant or breastfeeding women, women with a positive pregnancy test and patients with known G6PD deficiency or allergies to sulphonamide or dapsone were excluded. Written informed consent was required from the participant, or from the parent or guardian in the case of children. Approval was obtained from the research ethics committees of the College of Medicine, University of Malawi and Liverpool School of Tropical Medicine. A data and safety monitoring board was appointed. The study was registered with the Clinical Trials register, ISRCTN 12285821.The study took place at Zingwangwa health centre, in the city of Blantyre, Malawi, where malaria transmission is perennial, peaking during December to April. Between May 2004 and April 2006, adults and children presenting with an illness suggesting falciparum malaria were screened for malaria parasitaemia by a finger-prick thick blood film. Patients willing to enter the trial were additionally screened for eligibility by a questionnaire and capillary blood sample to check their haemoglobin concentration [Hb]. Women \u226512 years had a urine pregnancy test. Those with a positive blood film were eligible for enrolment in the study if they fulfilled the following criteria: i) age \u22656 months, ii) weight \u226510 kg, iii) Patients meeting inclusion criteria were recruited and randomized to AL, CPD or SP on a 1:2:1 basis. Randomization was performed in blocks of 20 using an off-site computer-generated code. Sealed sequentially numbered envelopes designating treatment group were opened for each participant following entry to the trial.CPD was dosed by height; paediatric and adult tablet formulations were available. A total of three doses of CPD were prescribed, one daily for three days. AL was dosed by weight and six doses were prescribed; on recruitment and after 8, 24, 36, 48 and 60 hours. The single SP dose was based upon age. The dosing schedules are shown in Table The first dose of medication was given at the clinic and further doses of CPD and AL were taken at home. Verbal instructions on how to take the remaining doses were given with advice to take AL with food or milk. A three-day supply of paracetamol (10 mg/kg) was given to take as required. Participants were cautioned against taking other anti-malarial drugs. Participants were observed for one hour to ensure that the medication was not vomited: if vomiting occurred within the first hour, the dose was repeated. Vomiting of the repeated first-dose resulted in removal from the trial, treatment with parenteral quinine and transfer to hospital for assessment.Formal follow up took place on days 7, 14 and 28 but participants could present to the clinic on any other day if unwell. In the second half of the study, follow up was extended to day-42 to detect treatment failures occurring after 28 days. Patients who did not attend for follow up were visited at home by study personnel (subject to consent) to ensure their safety. At each follow up visit, symptom histories were taken, Hb was checked and a malaria blood slide examined. In addition, on day-7, a venous blood sample was collected for pharmacokinetic analysis (see below). Clinical outcome was assessed using modified World Health Organization (WHO) criteria . ModificAdherence with the SP regimen was assured by direct observation of the single dose therapy. Adherence with AL and CPD was measured in two ways:1. Use of simple drug dosing questionnaires on day-7; participants or guardians were asked how many tablets had been taken and when.2. Use of MEMS\u2122  capped pill bottles which recorded electronically the date and time when the medication bottle was opened. Adherence was estimated by comparing the MEMS\u2122 number of openings with the prescribed dosing regime. MEMS\u2122 were used in a sub-group of patients according to availability at the time of recruitment. The MEMS\u2122 data were interpreted as follows; for CPD, participants who opened their pill bottle one time or more on each of the two consecutive days after recruitment were assumed to have taken all their prescribed doses of medication. Those who failed to open their pill bottles on the two consecutive days after recruitment were assumed to have missed some doses. For AL, if the MEMS\u2122 recorded at least one opening of the bottle on the day of recruitment and at least two more on each of the next two days, it was assumed that all the doses were taken.\u00ae Hb estimates and malaria slides were performed from capillary blood sample at each follow up visit. Blood slides were stained with Field's stain and parasite densities estimated from thick films by counting the number of parasites per 200 white blood cells assuming a total count of 8,000/\u03bcl. The concentrations of dapsone (DDS) and lumefantrine (LU) were measured from venous blood collected on day-7 to determine whether they were associated with treatment compliance or outcome. The blood was spun at 2,000 g for 10 minutes in a refrigerated centrifuged and the plasma removed and stored at -80\u00b0C for the pharmacokinetic analyses.HemocueDapsone (DDS) was quantified from 0.5 ml of plasma using HPLC-UV (high performance liquid chromatography - ultra-violet) detection using a method developed and validated in Liverpool . Chromatographic separations were carried out using Surveyor HPLC system (Thermo Fisher Scientific). Internal standard  was added to the plasma samples along with 0.5 ml sodium hydroxide (0.1 M). The samples were then vortexed for 10 seconds followed by the addition of 5 ml dichloromethane and again vortexed for a further 20 seconds. The samples were then centrifuged at 1,600 g for 10 minutes after which the organic (bottom) layer was transferred to a clean tube. A further 5 ml of dichloromethane was added to the aqueous layer, vortexed for 20 seconds and centrifuged again at 1,600 g for 10 minutes with the organic layer collected once more. The pooled organic layers were evaporated to dryness under a steady stream of nitrogen at 30\u00b0C. Samples were reconstituted in 100 \u03bcl of mobile phase of which 50 \u03bcl was injected onto column.\u00ae 100Rp-18, 5 \u03bc guard column. The mobile phase comprised of acetonitrile: triethylamine (0.1% adjusted to pH 3 with ortho-phosphoric acid): methanol (7:70:23 v/v), flowing at 1 ml/min. Column and tray temperature control was set at room temperature. The UV detector was set at 254 nm. Data was captured and processed using Xcalibur 1.4 software (Thermo Fisher Scientific). The lower limit of quantification (LLOQ) of the dapsone assay was 100 ng/ml and the calibration curve was linear in the range 0-2500 ng/ml.Chromatographic separation was achieved using a Phenomenex Synergi 4 \u03bc MAX RP C12  column, fitted with a LiChrosphereLumefantrine was quantified from 0.25 ml of plasma using HPLC-UV detection following a previously described method . The LLOThe primary aim of the study was to investigate the effect of incomplete adherence on the effectiveness of AL and CPD. Effectiveness was assessed by comparing the day-28 \"adequate clinical and parasitological response\" (ACPR) rates. The ACPR rates were not corrected using the polymerase chain reaction (PCR) to distinguish recurrent parasitaemias due to reinfections and recrudescence (true treatment failures). Secondary aims were to compare the effectiveness of AL, CPD and SP on days 28 and 42, to compare the different methods for measuring adherence, to investigate the associations between day-7 drug concentrations and adherence and treatment outcome and to compare serious adverse events (SAEs) and Hb changes after the three treatments.An approximate sample size of 1,000 participants was calculated, with 500 participants in the CPD group and 250 in the SP and AL groups. Assuming 50% of the participants were fully adherent, this sample size would detect a difference between a cure rate of 97% in the CPD arm and 90% in the SP arm with 95% confidence and a power of 80%. Because the initial focus of the study was on CPD, we planned to recruit twice as many children into that treatment group than the SP or AL groups. Data were double entered and validated prior to the analyses.Data analysis was performed using Stata 8: the primary analysis was on an intention to treat (ITT) basis. Binomial regression was used to obtain risk differences between treatments and 95% confidence intervals. Fisher's exact p-values were reported. Tests of significance were performed using the 0.05 level to infer significance for the planned analyses. For pair wise comparisons between combination therapies, we adjusted the significance level to 0.017 (i.e. 0.05/3) using Bonferroni's approach.841 of the 7,536 patients screened met the inclusion criteria and were enrolled. Common reasons for non-recruitment included negative blood slides (n = 6150), Hb <7 g/dl (n = 189) and the child weighing <10 kg (n = 63). Baseline characteristics of the recruited patients are shown in Table The results of the day-28 and day-42 ITT analyses with missing outcomes treated both as failures and successes and the per protocol (PP) analyses are shown in Tables Table There were 11 serious adverse events (SAEs) in the study, including one death, with eight SAEs in the CPD group and three in the SP treatment group. SAE rates are similar as 422 patients were recruited to the CPD group and 210 to the SP group. SAEs were defined as any occurrence resulting in death or admission to hospital for participants under follow up in the study. The three-year old girl who died had a recruitment parasite count of 129,000 per \u03bcl, Hb 11.7 g/dl and packed cell volume (PCV) 34%. She received her first dose of CPD in the clinic. The following day, she had a convulsion and further fevers and on admission to hospital was found to a positive malaria blood slide, PCV 30% and normal blood glucose concentration. She was treated with intravenous quinine, chloramphenicol and penicillin. Convulsions continued despite treatment with lorazepam and she died two days after her admission to hospital with presumed cerebral malaria.Salmonella typhimurium bacteraemia . One patient developed features of severe malaria (convulsions and fever with a positive blood slide) 20 days after treatment with CPD, having had negative slides on days 7 and 14. Two patients developed severe anaemia within a week of treatment with CPD and required admission to hospital for blood transfusion. The first was a 30-year male, recruited with a Hb of 8.9, who complained of dizziness but no fever on day-7 and had a Hb of 4.8 g/dl and a negative malaria slide. The second, an 11 year boy, was recruited with a Hb of 11.6 g/dl (PCV 34%), but admitted to hospital on day-3 with vomiting, a PCV of 21% and a negative malaria slide. By day-4 the PCV had fallen to 13%  necessitating a blood transfusion. Neither patient became jaundiced, clinically or biochemically, or had a palpable spleen. Three patients taking SP aged 2, 4 and 15 years had SAEs, being admitted to hospital on days 1, 2 and 10 respectively with fever and found to have positive malaria blood slides. All three were diagnosed with severe malaria and received parenteral quinine before discharge. The 2 year old child required blood transfusion due to a fall in their PCV from 27% to 13% by day-2.SAEs in the seven remaining CPD treated patients included two patients (aged 8 and 20 years) who developed vomiting and abdominal pain and two adults admitted with fever one week after recruitment and found to have This was completed by 371/442 (88%) participants who took CPD and 185/209 (88.5%) who took AL. Missing data was due to patient withdrawals from the study or loss to follow up. All 185 participants in the AL group who completed questionnaires said that they had taken all six of their prescribed doses. Only three (0.8%) CPD participants admitted missing any doses.Data were available for 181 patients in the CPD arm; 164 (90.6%) took all their doses out of the MEMS\u2122 container while 17 (9.4%) did not. Participants who took all their doses were more likely to have an ACPR on day-28, p = 0.024 (Fishers exact). MEMS\u2122 data were available for 87 AL treated patients; 80 (92%) took all their doses out of the MEMS\u2122 container and seven (8%) did not. None had treatment failure by day-28 and so the association between the MEMS\u2122 data and day-28 outcome could not be tested.Day-7 PK samples were analysed from 348/422 (82.5%) of the CPD patients. Missing data were due to patient withdrawals before day-7 and samples not collected or lost. The DDS concentration in their day-7 sample was < LLOQ of the assay in 174 (50%) of the participants. Participants whose day-7 DDS concentration was < LLOQ were younger, shorter in height and received a lower dapsone dose per kg body weight  and were less likely to have an ACPR outcome on day-28 than those \u2265 LLOQ (104/178 (59.8%) vs. 143/174 (82.2%), p < 0.0001). For the 174 (50%) participants with quantifiable DDS concentrations, median day-7 DDS concentrations were higher in the ACPR group than the treatment failure group, 248.3 ng/ml vs. 152.3 ng/ml respectively, p = 0.012 (Mann-Whitney). There was no difference in the median day-7 DDS concentration between participants who took all their CPD doses and those who did not according to the MEMS\u2122.Day-7 samples was analysed for LU in 167/209 (79.9%) AL patients. Only 4/167 (2.4%) participants had day-7 LU concentrations < LLOQ of the assay. The median (IQR) LU concentrations for the 163 participants with quantifiable LU concentrations were 214 ng/ml (118-321). The small numbers of treatment failures or patients with samples < LLOQ does not permit meaningful comparisons of lumefantrine levels in these groups. The median (IQR) day-7 LU concentration in the 61 patients who took all their doses out of the MEMS\u2122 container was 193.0 ng/ml (105.0 - 273.0), compared to 127.0 ng/ml (42.0 - 390.0) in the five who did not, .A previous study from Thailand reported that patients with LU levels <175 ng/ml on day-7 are more likely to experience recrudescence by day-42 [Adherence to therapy is one of the cornerstones of successful treatment and the more complex a treatment regime is, the more likely it is that patients will fail to adhere properly. Single dose therapy - SP is the only example - is ideal, but widespread resistance means that SP is no longer a useful option for control programmes. All of the current treatment regimes promoted by the WHO are multi-dose and there is no single dose therapy on the horizon. The impact of poor adherence on treatment effectiveness is difficult to measure but is an important consideration for health policy makers. Methods for measuring adherence to therapy including drug questionnaires, pill counting, assessment of pharmacy repeated prescriptions, MEMS\u2122 and drug level monitoring. None of these is perfect, and measurement of adherence may itself lead to a change in adherence behaviour of the individual being monitored .AL is currently the most widely deployed artemisinin-combination therapy (ACT) in the world. By the end of 2008, it had been adopted as first line treatment policy for uncomplicated malaria in 23 countries in Africa . TreatmeMEMS\u2122 provide a more objective assessment of adherence, although the interpretation of their results can be difficult. They have been used extensively to monitor patients on long-term treatment such as anti-retroviral therapy. MEMS\u2122 record the date and time at each opening of the pill bottle. The numbers of tablets removed each time or whether the tablets are actually taken cannot be recorded. Patients taking AL may open the bottle once in the morning and remove all the tablets required for both the morning and evening doses at that time and this would be interpreted as poor adherence due to the missed evening dose. Patients are also known to \"play\" with their pill bottles and, as a result, multiple openings are recorded each day when no drug has actually been taken.In this study, over 90% of the patients receiving CPD took all their doses out of the MEMS\u2122 container and these patients were more likely to have an ACPR on day-28 than those participants who did not, p = 0.024. For AL, the MEMS\u2122 data suggested that 92% of the patients took all of their six doses. There were only three treatment failures in the AL treatment group by day-28 making it impossible to examine the association between the MEMS\u2122 results and treatment response.et al after a mean total dose of 61.2 mg/kg of LU [The median day-7 LU concentration reported here was 214 ng/ml, after a mean total LU dose of 63.9 mg/kg. This is lower than the median of 528 ng/ml reported by Price Other studies have addressed adherence with AL. In Bangladesh, an adherence rate of 93% was reported using pill counting and questionnaires and there was no difference noted in day-7 LU concentrations in poorly adherent patients . AverageThe measurement of the day-7 drug concentration has been proposed as a way to predict treatment outcome in malaria treatment studies . A studyMalawi was the first country in Africa to introduce SP as its first line treatment for uncomplicated malaria in 1993. In 2007, it was withdrawn by the national Malaria Control Programme, and replaced by AL. The results reported here confirm that SP was insufficiently effective for a first-line malaria treatment in Malawi at the time of this study; the day-28 ACPR rate for SP of 62.5% by PP analysis. In contrast, the PP day-28 and day-42 ACPR rates for AL were 98.3% and 95.4% respectively, in keeping with rates reported from other countries in the region including Mali, Ghana and Uganda, whether AL is taken supervised in an efficacy trial or unsupervised -16. ThesThe PP day-28 ACPR rate with CPD was 75.6% and was lower than had been expected. A longitudinal study from Malawi in 1999 reported a day-14 efficacy of around 96% [There were no SAEs in the AL treatment group, consistent with data from other studies reporting that this combination therapy is well-tolerated and highly efficacious. Three patients in the SP group required hospital admission with features of severe malaria within 10 days of recruitment to the study, probably reflecting the poor efficacy of SP. One child died having developed features of severe malaria a day after recruitment and treatment with CPD; it is impossible to know whether this resulted from poor treatment efficacy or was an inevitable consequence of that child's infection. The production of CPD was halted and the drug withdrawn because of concern about haemolysis. Two patients developed severe anaemia within a week of treatment with CPD. In both, the Hb fell by around 50% despite parasite clearance, making it unlikely that malaria was responsible. Dapsone-induced haemolysis is more common in G6PD deficient individuals but the G6PD status of participants in this study is unknown.SP has failed in Malawi and should no longer be used as a treatment for uncomplicated malaria. Its continued role in intermittent presumptive therapy in pregnancy programmes needs to be evaluated. In this study, AL was highly effective, even with unsupervised dosing and was well tolerated. Adherence to the six-dose regime was good and treatment was effective, even in those who did not properly comply with the dosing schedule. Self-reported adherence appears to be an unreliable measure of adherence in this population and may lead to over-estimation of the true level of adherence. This study provides strong reassurance of the effectiveness of AL as it is rolled out across sub-Saharan Africa.PAW and SAW were unpaid members of the MMV-GSK product development team for chlorproguanil-dapsone-artesunate. None of the other authors declare any conflicts of interest.The study was conceived and designed by DGL, PAW, SAW and MEM. The study was set up and run by DJB, DW, NK and PC. Laboratory analyses were performed by JM, DAH, DJB and DW. DJB, MM and DGL analysed the data. DJB, DGL, PAW and MEM wrote the paper. All authors read and approved the final manuscript.This work was funded by a grant from the Department for International Development, UK (Project number: RES 8074). The funders had no role in the study design or execution, or in the analysis and presentation of the data. GlaxoSmithKline provided the CPD medication without charge."}
+{"text": "Artesunate-amodiaquine (AS+AQ) and artemether-lumefantrine (AM-L) are efficacious artemisinin combination therapy (ACT) regimens that have been widely adopted in sub-Saharan Africa. However, there is little information on the efficacy of these regimens on subsequent episodes beyond 28 days, or on the safety of repeated treatments.Children aged six months to 14 years with uncomplicated malaria were randomly assigned to treatment with AS+AQ (n = 116), or AM-L (n = 111). Recruited subjects were followed-up, initially for 28 days, and then monthly for up to one year. All subsequent attacks of uncomplicated malaria after 28 days were treated with the same regimen as at randomization. Investigations aimed at determining efficacy and side effects were conducted.Adequate clinical and parasitological response in subjects with evaluable end-points were, 97.1% (100/103) and 98.2% (107/109) on day 14, and 94.2% (97/103) and 95.3% (102/107) on day 28 in the AM-L and AS+AQ groups, respectively. Similar results were obtained after PCR correction. The incidence of malaria attacks in the year following recruitment was similar between the two treatment groups (p = 0.93). There was a high incidence of potentially AQ-resistant parasites in the study area. The incidence of adverse events, such as pruritus, fatigue and neutropaenia were similar in the two treatment groups. No patient showed signs of hearing impairment, and no abnormal neurological signs were observed during one year of follow-up. Other adverse events were mild in intensity and overlapped with known malaria symptomatology. No adverse event exacerbation was observed in any of the subjects who received multiple treatment courses with these ACT regimens during one year follow-up.AS+AQ and AM-L were efficacious for treatment of children with uncomplicated malaria in Ghana and drug-related adverse events were rare in treated subjects during one year of follow-up. The high prevalence of potentially AQ resistant parasites raises questions about the utility of AQ as a partner drug for ACT in Ghana. The efficacy of AS+AQ in Ghana requires, therefore, continuous monitoring and evaluation.NCT 00406146  Artemisinin-based combination therapy (ACT) may slow the development of resistance and reduce malaria transmission . ArtesunThe efficacy of an ACT may depend on the level of pre-existing resistance to the non-artemisinin component , and it in vitro and in vivo evidence suggests that AQ resistance is linked to the T76 single nucleotide polymorphism in the Plasmodium falciparum chloroquine resistance transporter (Pfcrt) [Pfcrt T76 prevalence has been reported from Ghana [The molecular basis underlying the mechanism of AQ resistance has not been fully elucidated, but  (Pfcrt) ,11. Receom Ghana , raisingom Ghana .In spite of accumulating clinical experience and increasing use of ACT, aspects of the safety of these regimens remain a concern. With respect to artemisinins, high dose parenteral administration of the oil-soluble derivatives is associated with dose-dependent toxicity to areas of the brain stem involved in coordination and balance in animals ,15. No cWith respect to AQ, prophylactic use of this drug has been associated with fatal cases of agranulocytosis and hepatitis, mainly in non-immune adults ,22. AlthPfcrt alleles in the study population.This study, conducted to generate efficacy and safety data on AS+AQ and AM-L to support the new ACT policy in Ghana, was a clinical trial with extended follow-up . The desThe study was conducted in two primary health facilities, Korle Bu Polyclinic (KBP) and Mamprobi Polyclinic (MP). KBP is attached to a tertiary referral hospital, Korle Bu Teaching Hospital (KBTH). The two facilities are situated in Ablekuma sub-district in Accra, Ghana. Both facilities serve a mixed, predominantly low socio-economic urban population. The study was conducted between October 2004 and December 2006.Plasmodium falciparum, and if the study criteria were fulfilled.Children presenting with a history of fever to the facilities were referred to the study team. A project physician examined the child to exclude concomitant illnesses. Subjects were recruited if a blood film was positive for P. falciparum mono-infection with parasite density between 2,000 \u2013 200,000/\u03bcL; and willingness to comply with the follow-up schedule. Exclusion criteria were, symptoms or signs of severe malaria; obvious clinical evidence of chronic malnutrition or other severe disease; known intolerance or allergy to study medications; and reported treatment with any of the drugs under study one month preceding enrolment. Children aged five to 14 years were included to permit objective neurologic examination and audiologic assessment. Schoolchildren aged five to 14 years from two local primary schools were enrolled as controls for the audiometric study described below. Written informed consent was obtained from parents or guardians of all subjects. Ethical approval for the study was granted by the Ethical and Protocol Review Committee of the University of Ghana Medical School.Inclusion criteria for the study were, age six months to 14 years; signs and symptoms of acute uncomplicated malaria, including axillary temperature \u226537.5\u00b0C; confirmed A computer-generated simple randomization scheme was prepared in advance. Allocated treatments were kept in sealed opaque envelopes. After completion of formal enrolment procedures, allocated treatments were administered by project nurses. All study personnel (except project nurses), were unaware of the assigned treatments.\u00ae, Pfizer, Dakar, Senegal), 10 mg/kg body weight, single daily dose for three days, was administered with AS , 4 mg/kg body weight, single daily dose, for three days. AM-L  was administered at 0 and 8 hours on the first day, and then twice daily for two subsequent days according to body weight: 5\u201314 kg, one tablet/dose; 15\u201324 kg, two tablets/dose; 25\u201334 kg, three tablets/dose; 35 kg and over, four tablets/dose.AQ  in the clinic. Parents administered the evening dose of Coartem at home ,28. ChilSubjects were followed-up on days 1, 2, 3, 7, 14 and 28 for clinical and laboratory assessments described below. Children who missed their day 14 or day 28 follow-up appointments were seen on days 15 or 29. After 28 days, children were visited at home every month for up to one year. During the monthly follow-up visits, parents or caregivers were asked about the child's well-being, behavioural and developmental concerns, as well as open questions about possible side event occurrence since the last visit. A study physician examined children during the follow-up visits. Follow-up was terminated if parents relocated. Each time a subject presented with an acute febrile illness during follow-up, thick films for malaria parasitaemia and a thorough examination was done, before and after the episode. All episodes of uncomplicated malaria diagnosed more than 28 days after treatment were treated with the same pre-assigned regimen as at randomization.Neurological examinations were done before treatment, and then on days 1, 2, 3, 7, 14 and 28. The neurological examination was focused on detecting abnormalities in areas that have been implicated in artemisinin toxicity in animals, i.e. on coordination and balance. In assessment for nystagmus, the target was always kept within binocular vision; the direction of the fast phase of nystagmoid eye movements was recorded; and it was noted if nystagmus was reduced by fixation or not. Any associated characteristics, such as vertigo, nausea, vomiting, and tinnitus were noted and recorded. Gait analysis, Romberg's test, speech assessment and the finger-nose test were done to evaluate cerebellar function. The finger-nose test was done for children aged six years or older. The responses for the finger-nose test were scored numerically for, 1) quality, i.e. smoothness and signs of intention tremor during the movement, and ii) adequacy, i.e. the successful implementation of the given task. The scores assigned for smoothness were, 0 = no tremor; 1 = slight tremor towards the end of the movement; and 2 = marked tremor increasing towards the end of the movement. The scores ascribed for adequacy were, 0 = the subject puts fingertip/pen top correctly at tip of nose each time; 1 = subject misses the tip of his/her nose/pen top once or twice; 2 = subject misses the tip of his/her nose/pen top each time. A score of zero for smoothness and adequacy was considered optimal for children aged six years and above, and all tests were scored with reference to the child's developmental age .Pure tone, air conduction audiometry was done with a portable screening audiometer  for children aged five years and above who could cooperate with the test, on days 0, 3, 7, and 28. Hearing assessment was repeated after a follow-up period ranging from nine months to one year. Hearing thresholds were determined for each ear at the following frequencies: 125 Hz, 250 Hz, 500 Hz, 750 Hz, 1000 Hz, 1500 Hz, 2 KHz, 3 KHz, 4 KHz, 6 KHz and 8 KHz.Venous blood was collected into EDTA and heparinized tubes on days 0, 3, 7, 14, 28 and on any day of recurrent symptomatic parasitaemia. Parasite counts were determined in Giemsa-stained blood films relative to 200 white blood cells (WBC) and the measured WBC count. Total WBC and differential counts were measured by automated haematology analyzer . Serum aminotransferase and total bilirubin were measured by a clinical chemistry analyzer .msp-1 and msp-2, to distinguish between recrudescent and new infections, as described previously [Pfcrt gene, as described previously [A nested PCR method was used to analyse polymorphisms in the merozoite surface protein (MSP) genes, eviously . Nested eviously .The primary end-point was the unadjusted and PCR-adjusted cure rates at days 14 and 28. Efficacy outcomes were classified as, adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF) and late parasitological failure (LPF) .Secondary endpoints were proportion of subjects with fever and parasitaemia 24 and 48 hours after treatment, the total number of subsequent malaria attacks in each arm, and incidence of adverse events before and after 28 days.All adverse events were graded by severity , and relationship to study medication  .The sample size was estimated on the assumption of a negligible (3%) risk of treatment failure with AM-L (because AM-L was new in the study area), and a 15% risk of AS+AQ treatment failure (because of prior extensive use of AQ monotherapy in the area). A sample size of 103 subjects was required to detect a 12% difference in cure rates between AS+AQ and AM-L with 95% confidence and 80% power. Categorical variables were compared with the chi square test with Yates correction or the Fisher's exact test. Continuous variables were compared with the student's t test or one-way analysis of variance. A logistic modelling (GENMOD\u2122) procedure was used to compare the total number of treatments in the two arms for the year of follow-up. Risk differences and 95% confidence intervals were calculated with CIA\u2122. Other analyses were done with Sigma stat  and SAS .The primary efficacy outcome was analysed by a per-protocol (PP), and an intention-to-treat (ITT) method. In the ITT analysis, subjects lost to follow-up were included; these, as well as those withdrawn because of non-tolerance of oral medication, were considered treatment failures. In the PP analysis, only subjects with an assigned outcome were included. The follow-up time of subjects who did not complete one year of follow-up were censored at their last follow-up date. P values < 0.05 were considered significant.Salmonella typhi resulting in worsening clinical condition after parasite clearance (n = 1); and loss to follow-up (n = 5). The reasons for non-completion of follow-up in the AS+AQ arm were, withdrawals due to persistent vomiting (n = 2); withdrawal due to worsening clinical condition despite rapid parasite clearance in an infant who was subsequently admitted and managed clinically for suspected septicaemia (n = 1); and loss to follow-up (n = 6). The flow of subjects through the study (up to day 28) is outlined ; culture positive co-infection with ance n = ; and losance n = ; and losThe proportion of subjects  with ACPR were, 97.1% (100/103) and 98.2% (107/109) on day 14 (p = 0.67); and 94.2% (97/103) and 95.3% (102/107) on day 28 (p = 0.94) in the AM-L and AS+AQ groups, respectively  in the AS+AQ group after 24 hours (5.3%) than in the AM-L (14.6%) group (p = 0.06). The trend had disappeared after 48 hours (p = 0.32).After one year of follow-up, there were 34 further episodes of symptomatic uncomplicated malaria in 93 subjects  in the AS+AQ group, and 28 episodes in 82 subjects  in the AM-L group . Similar results were obtained, when logistic modelling (p = 0.94), or survival analysis (p = 0.76) was used to compare incidence in the two arms. All subjects who received repeat treatment courses responded appropriately to treatment. The time to a repeat treatment episode  was longer in the AS+AQ  arm than the AM-L  arm. There were three sibling pairs among those who were treated for repeat infections. These siblings (each randomized to receive a different ACT) reported simultaneously with a repeat attack of uncomplicated malaria.The \"CVIET\" (mutant) haplotype was present in 72.1% (75/104) of pre-treatment samples in subjects treated with AS+AQ. The effect of the \"CVIET\" haplotype prevalence on treatment outcome could not be evaluated because of the low number of subjects with treatment failure.Audiometry could be done in 72 subjects . Hearing thresholds were significantly elevated in treated subjects than in age- and sex-matched control children on days 0, 3, 7, and 28, but there were no differences between hearing thresholds of subjects and controls after nine to 12 months. The full details of the audiometric study will be presented in another report.Neurological examination could be done for 168 children . Nystagmus was observed in two subjects  between days one and seven. A detailed history indicated that one of these two subjects  was admitted as a neonate into neonatal intensive care (NICU) because of probable birth asphyxia. This subject was still attendant at pre-school at seven years of age , and demonstrated excessive emotional lability. An electroencephalography (EEG) done for this subject because of a history suggesting the possibility of a seizure disorder, reported the presence of \"epileptiform loci and generalized cerebral dysfunction\". The other subject with nystagmus , also had a history of NICU admission for macrosomia (birth weight 4.5 kg) and possible neonatal hypoglycaemia. This subject had poor academic performance (had repeated first and second grades in school) and possible cognitive impairment. A routine EEG done for this subject was normal. A positive Romberg's test (which may suggest sensory ataxia) was noticed on day 3 in an 11-year old female (AM-L). This child was not attending school (despite her age) and had a history suggestive of possible cognitive impairment. No other abnormal neurological findings were observed in the remaining children during the 28-days follow-up, monthly follow-up visits, or for subjects who received multiple treatments.There were no differences in the mean total WBC or absolute neutrophil counts between the two groups on days 0, 3, 7, 14 or 28  elevated, but there were no differences between the two treatment groups. The liver enzymes did not increase in response to treatment, but showed a trend towards normalization (reduction) as clinical symptoms improved (data not shown).Within the first three days of treatment, complaints of dizziness were reported by 14 (12%) and eight (7.2%) subjects in the AS+AQ and AM-L groups, respectively (p = 0.31). Complaints of fatigue and excessive sleepiness were reported by five (4.3%) and four (3.6%) subjects in the AS+AQ and AM-L groups, respectively (p = 1.0). Pruritus was reported by three  subjects. A history of vomiting and nausea was part of the presenting complaint by 44.8% (52/116) and 40.5% (45/111) subjects respectively in the AS+AQ and AM-L groups on admission (p = 0.74). Between the first three days of treatment, 27.5% (32/116) and 12.6% (14/111) subjects respectively complained of vomiting and or nausea in the AS+AQ and AM-L groups (p = 0.01). Severe anaemia (haemoglobin 4.8 g/dl) occurred on day 14 in a 1.5 year old subject (AS+AQ). Severe anaemia and dizziness were considered unlikely to be drug related, but pruritus and excessive sleepiness were considered possibly drug-related. Other reported adverse events were mild in intensity, and overlapped with known malarial symptomatology. These were mostly classified as unrelated to study medications.This study provides efficacy data on AS+AQ and AM-L, the two most widely adopted ACT regimens in sub-Saharan Africa, for Ghanaian children. The study also provides longitudinally collected safety data on neutrophil counts and neurological findings for Ghanaian children with uncomplicated malaria treated with repeated courses of these two ACT regimens.The results showed a high cure rate for both regimens after a standard 14- or 28-day follow-up. These results are consistent with efficacy results reported from several sub-Saharan African countries , indicatPfcrt mutant allele prevalence has been detected [Pfcrt allele prevalence on treatment outcome in the group treated with AS+AQ could not be assessed because of the low incidence of treatment failures in this group, AS+AQ has been associated with a high cure rate in an area of even higher (90%) Pfcrt K76T mutant prevalence [in vivo efficacy of AS+AQ from one area to another, highlighting the need to generate local efficacy data for these newly introduced treatment regimens in Ghana. Furthermore, the high mutant Pfcrt allele prevalence also provides additional evidence justifying the treatment policy change in Ghana to ACT.The finding of a high prevalence of potentially AQ-resistant parasites in this southern Ghanaian population raises questions about the long-term utility of AQ as a partner drug for ACT in Ghana, a question that has also been raised by other investigators from northern Ghana , where adetected . Althougevalence . This suevalence . Thus, iPrevious studies that have compared AS+AQ with AM-L over a follow-up duration of 28-days or longer have demonstrated a lower re-infection rate for AM-L . Other sThere was a drop in neutrophil counts in both arms after treatment. This pattern of neutrophil count reduction, as well as the absence of differences in neutrophil count profiles between children who received single course and those who received multiple courses, suggests the possibility of a disease-rather than drug-related effect.Malaria-associated neutropaenia has been reported for subjects treated with AQ, either as monotherapy, or in combination with AS , as wellThis is one of the few studies to conduct systematic longitudinal neurological examinations for children treated with artemisinin-based regimens. A total follow-up lasting 12 months did not reveal abnormal neurological signs that could be ascribed to artemisinin-based treatment in this study. The two subjects with observed nystagmus had underlying conditions that could easily explain the noted abnormalities. Furthermore, the observed nystagmus was not associated with other cerebellar signs, suggesting these were unlikely to be of clinical significance, or could be false positive due, for instance, to excessive lateral eye abduction. However, nystagmus has been reported in artemisinin-treated subjects without reported underlying neurological abnormality , as wellThe incidence of gastrointestinal adverse events was higher in the AS+AQ group compared to the AM-L group, similar to a recent report comparing AS+AQ with dihydroartemisinin-piperaquine . This maThe limitations of this study include its limited statistical power to detect rare side effects, as well as the difficulties associated with evaluating neurological function in the younger children below five years of age. Further studies of the nystagmoid eye movements, apart from the clinical observations done, would also be useful.AS+AQ and AM-L were efficacious for the treatment of Ghanaian children with uncomplicated malaria. After 12 months of detailed systematic follow-up, no overt clinical evidence of neurotoxicity or neutropaenia was observed. However, the sample size was limited for conclusive evidence on safety; therefore, post-marketing surveillance, with particular emphasis on neurological and haematological changes, is still indicated. In view of the high prevalence of potentially AQ-resistant mutant parasites in the study area, the efficacy of the AS+AQ combination, the first-line treatment for uncomplicated malaria in Ghana, requires continuous monitoring and evaluation.GOA, BQG, OPR and JALK designed the study; GOA, BQG, OPR, EVB, EDK and RL did the clinical work. GOA, LCGH and MA did the laboratory analyses. GOA and JALK did the data analysis. GOA drafted the manuscript, and all authors contributed significantly to the final draft."}
+{"text": "Plasmodium falciparum malaria. Fixed-dose combinations are more adapted to patients than regimens involving multiple tablets and improve treatment compliance. A fixed-dose combination of artesunate + amodiaquine (ASAQ) was recently developed. To assess the efficacy and safety of this new combination and to define its optimum dosage regimen (once or twice daily) in the treatment of uncomplicated P. falciparum malaria, a multicentre clinical study was conducted.The use of artemisinin derivative-based combination therapy (ACT) such as artesunate plus amodiaquine is currently recommended for the treatment of uncomplicated P. falciparum malaria were randomized to receive ASAQ orally once daily (ASAQ1), ASAQ twice daily (ASAQ2) or AL twice daily (AL) for three days. The primary outcome was PCR-corrected parasitological cure rate and clinical response.A multicentre, randomized, controlled, investigator-blinded, parallel-group study was conducted in five African centers in Cameroon, Madagascar, Mali and Senegal from March to December 2006. Efficacy and safety of ASAQ were assessed compared to those of artemether + lumefantrine (AL). The WHO protocol with a 28-day follow-up for assessing the drug therapeutic efficacy was used. Patients suffering from uncomplicated Of 941 patients initially randomized and stratified into two age groups , 936 (99.5%) were retained for the intent to treat (ITT) analysis, and 859 (91.3%) patients for the per protocol (PP) analysis. Among ITT population, up to D28, PCR-corrected adequate parasitological and clinical response rates were 95.2% in the ASAQ1 group, 94.9% in the ASAQ2 group and 95.5% in the AL group. Moreover, the cure rate evaluated among PP population was \u226598.5% in both ASAQ therapeutic arms. Therapeutic response rates did not display any significant differences between age groups or between one geographical site and another. Altogether, this demonstrates the non-inferiority of ASAQ1 regimen compared to both ASAQ2 and AL regimens. During follow-up mild and moderate adverse events including gastrointestinal and/or nervous disorders were reported in 29.3% of patients, with no difference between groups in the nature, frequency or intensity of adverse events.P. falciparum malaria both in adults and children. Implications of such findings are of primary importance in terms of public health especially in African countries. As most national policies plan to strengthen malaria control to reach the elimination of this disease, anti-malarial drugs such as the artesunate + amodiaquine fixed-dose ACT will play a pivotal role in this process.The non-inferiority of ASAQ compared with AL was demonstrated. The fixed-dose combination artesunate + amodiaquine (ASAQ) is safe and efficacious even in young children under 5 years of age. Whilst administration on a twice-a-day basis does not improve the efficacy of ASAQ significantly, a once-a-day intake of this new combination clearly appears as an effective and safe therapy in the treatment of uncomplicated The protocol was registered with the www.clinicaltrials.gov open clinical trial registry under the identifier number NCT00316329. Plasmodium falciparum malaria or the past decade [\u00ae) was designed and marketed in 2002. The results of different clinical trials confirmed the efficacy of Arsucam\u00ae in Senegal [Combination therapy has been on the path to become the treatment of choice for uncomplicated t decade . The uset decade . Since t Senegal ,4, the C Senegal , Mali [6 Senegal .P. falciparum infections [P. falciparum malaria.The 2006 WHO guidelines for malaria treatment recommend the combination of AS and AQ in a ratio of 2.5  or 3.1 (corresponding to the co-blister's ratio). However, the use of AS and AQ in multiple tablet combinations is unsatisfactory in terms of treatment compliance. It involves the risk that only one of the two drugs be received or that the drugs be administered at an inadequate dose. Since it is recognized in the medical fields that the availability of single-tablet formulations (containing drugs in a fixed-dose combination) reduces these risks and improves treatment compliance -10, suchfections . In ordeP. falciparum malaria. The study was conducted in five African sites where malaria transmission is perennial: Bancoumana , Yaound\u00e9 (Cameroon), Tsiroanomandidy (Madagascar), Keur-Soc\u00e9 and Mlomp . Patients were included between March and December 2006. The study was conducted during the highest malaria transmission period for each site. This protocol was registered with the www.clinicaltrials.gov open clinical trial registry under the identifier number NCT00316329.This was a multicentre, randomized, controlled, investigator-blinded, parallel-group study comparing three ACT regimens in patients with uncomplicated P. falciparum residing in the study area throughout the planned period and according to the following criteria: weighing \u2265 10 kg; displaying a parasitaemia from 1,000 to 200,000 asexual forms per microliter of blood; presenting an axillary temperature \u2265 37.5\u00b0C or suffering from fever within the last 24 hours and capable of receiving oral treatment.The study included patients infected with Patients with any clinical sign of severe malaria  or any sPatients were randomized to one of three treatment groups, namely ASAQ bi-layer fixed-dose combination tablets once daily intake (ASAQ1), ASAQ twice daily (ASAQ2), and AL twice daily (AL). Randomization was stratified according to patient age using separate randomization lists. The two age strata were children under five years of age (as the primary WHO target population) and patients aged five years and over. Treatment dosages were determined according to patient's body weight. Treatment duration was three days. An eight hour time-lapse was requested in case of twice-daily regimens . Tablets were orally administered with a small amount of drinking water; and patients were advised to resume a normal diet as soon as possible. All patients were monitored for 30 minutes after administration in order to ensure that the drug was not lost after an eventual vomiting episode. When this occurred, the same dose was re-administered. If vomiting re-occured, the subject was withdrawn from the study and a replacement treatment with another effective anti-malarial, generally quinine, was provided. In order to ensure blinding, all patients in the same body weight range received the same number of tablets per intake, which was defined by the recommended dose of AL. Patients in the ASAQ arms were given placebo tablets to match this number. Only the person administering the treatment was aware of the nature of the drug taken by each participant, whereas investigators evaluating safety and efficacy were kept blind.The WHO protocol with a 28-day follow-up  to assesin vivo WHO protocol. Clinical safety was monitored through regular patient interviews with regards to the occurrence of adverse events following the previous visit.Clinical efficacy was assessed by grading the pre-existing clinical signs and combining them with the temperature values. Parasitological efficacy was based on asexual parasitaemia, although a gametocyte count was also systematically performed. The following clinical symptoms were assessed systematically at each visit and graded as absent, mild, moderate, severe or very severe: perspiration, headache, chills, pain (specifying topography), jaundice, asthenia, dizziness, anorexia, skin fold, skin rash, hepatomegaly, pruritus. Splenomegaly was estimated according to the Hackett scale . VomitinP. falciparum parasites until 300 leukocytes were observed and then converting this figure into parasites (trophozoits and/or gametocytes) per microliter of blood, assuming an average leukocyte count of 7,500/\u03bcL. For quality control purposes, a random sample of at least 10% of all slides analyzed at each site, as well as all slides detected as positive, were re-evaluated using a blinded procedure with respect to the original result and to the source study center at the Malaria Research and Training Center (MRTC), Bamako, Mali.Fingerpick blood samples were collected for parasitological checking at enrolment and at successive follow-up visits. Thin and thick smears were obtained and stained with May Gr\u00fcnwald-Giemsa. Filter paper dried blood spot samples were retained on filter paper for later PCR analysis in case of recurrent parasitaemia. Blood samples for parasitological count were analyzed at local centers under their own procedures. Giemsa-stained thick blood smears were read by experienced microscopy practitioners who were blinded to treatment allocation. Parasite densities were calculated by counting the number of asexual and sexual msp-1, msp-2, and microsatellite Ca1 gene loci were compared. The PCR was performed using the following primers pairs MSP1: O1 = 5'-CACATGAAAGTTATCAAGAACTTGTC-3' and O2 = 5'-GTACGTCTAATTCATTTGCACG-3' for PCR1, N1 = 5'-GCAGTATTGACAGGTTATGG-3' and N2 = 5'-GATTGAAAGGTATTTGAC-3' for PCR2; MSP2: S3 = 5'-GAAGGTAATTAAAACATTGTC-3' and S2 = 5'-GAGGGATGTTGCTGCTCCACAG-3' for PCR1, S1 = 5'-GAGTATAAGGAGAAGTATG-3' and S4 = 5'-CTAGAACCATGCATATGTCC-3' for PCR2; and MICROSATELLITE Ca1: Ca1-1L = 5'-GCTGTAAAACGTGAACAAAAA-3 and Ca1-1R = 5'-CAATTCTGCTTCAGTTGGATT-3' for PCR1 and Ca1-L = 5'-ATTATGAACAATTCAGAC-3' and Ca1-R = 5'-GTTGTTATAGCTAATGAG-3' for PCR2. One to five microliters of DNA was amplified for 30 cycles during the primary PCR.For participants with recurrent parasitemia after day 7, paired polymerase chain reaction (PCR) blots (from day 0 and the day of parasitemia recurrence) were analyzed for parasite merozoite surface proteins (MSP-1 and MSP-2) and microsatellite CA1 to distinguish between reinfection and recrudescence . DNA wasOne to two microliters of the product of that amplification was used for another 30 cycles of PCR. Each PCR was performed with 1 \u03bcM of each primer, 200 \u03bcM of each dNTP, and 1.5\u20133 mM MgCl2.msp-1, msp-2, and Ca1; ii) reinfection, if the alleles of the pre-treatment and post-treatment samples were distinct for any of these three loci; iii) mixed recrudescence and reinfection, if similar alleles were found in the pre-treatment and posttreatment samples for all the markers as mentioned above, but with additional distinct alleles identified; and iv) indeterminate, if either or both the pre-treatment and post-treatment samples could not be amplified. Mixed recrudescent and re-infection cases were computed as recrudescent.Possible outcomes were i) recrudescence, if the alleles of the pre-treatment and post-treatment samples were the same for A priori power calculations were used to determine the sample size required to demonstrate non-inferiority between the ASAQ group and the AL group (inter-group difference <5%) in both the overall population and in a target subpopulation of children under five years of age with a type I error of 5% and a power of 80% to be able to detect such a difference. On the basis of previous studies with AL [ with AL ,18, a re2 test or Fisher's exact test, normally distributed quantitative variables with Student's t-test and other quantitative variables with the Wilcoxon test. Missing data were not replaced. All statistical analyses were performed centrally using SAS version 8.2 software .The primary efficacy endpoint corresponded to parasitological and clinical cure on Day 28 determined in the ITT population. The difference in the proportions of responders between the ASAQ group and AL group was estimated using a non-inferiority analysis with respect to a 5% significance level, according to the Binomial distribution, using the STATXACT software program with the \"noninf\" option (non-inferiority delta of 5%). Categorical variables were compared with the \u03c7This study was conducted according to the Declaration of Helsinki and existing national legal and regulatory requirements. The protocol was submitted and approved by the appropriate ethics committees and institutional review boards in each participating country. Written informed consent was obtained from each participant or their parent or guardian in case of minors. If patients were unable to sign, a fingerprint was applied on the consent form. In case of patients unable to read, the information was read and explained to them in the appropriate language. In such cases, the presence of a witness who also signed the consent form to confirm that the patient had freely given consent was required.Overall, 941 patients were randomized to be included in one of the three therapeutic arms. Since one patient (0.1%) did not receive any study medication, the safety population consisted of 940 patients. Table PCR-corrected clinical and parasitological cure rates on Day 28 in the ITT population are shown in Table Subgroup analyses were performed to assess treatment efficacy in three different age groups Table . The PP For all five sites Table , more thAt enrolment, patients in the three treatment groups had similar mean parasite densities. Parasitic clearance rates were at similar levels at any time and independently of the treatment groups considered: 1.74 \u00b1 0.60; 1.69 \u00b1 0.60 and 1.73 \u00b1 0.59 days in the ASAQ1, ASAQ2 and AL groups respectively , asthenia on Day 2 (higher in ASAQ2 group) and headache on Day 3 (higher in the ASAQ1 group).2 test, p = 0.01) was observed in case of mild to moderate somnolence although this was reported only in ASAQ groups in Madagascar (15 children under 14 years of age and two adult patients). Treatment-related adverse events (88.5%) \u2013 mainly of mild or moderate intensity \u2013 essentially concerned the nervous  and gastrointestinal systems (16.7%). Overall, 122 (13.0%) treated patients experienced vomiting or rejection episodes in the first half-hour after treatment administration from Day 0 to Day 2. No significant difference was recorded between the treatment groups (p = 0.26) although the subgroup including patients under five years of age was significantly more affected by vomiting and rejection. Two cases of rashes reported by investigators as treatment-related were submitted to dermatologists for expertise, with no clear diagnosis established. With respect to clinical symptoms occurring under treatment, a few cases of vomiting were reported in 7.4% of patients, pain in 2.5%, anorexia in 1.8%, pruritus in 1.7%, diarrhoea in 1.3%, asthenia in 0.9%, headache in 0.6% and chills in 0.4%.Overall, 275 patients (29.3%) in the safety population presented at least one emergent adverse event, of which nature, incidence and intensity were similar between the three treatment groups. Among them, the most frequently reported ones included intestinal parasitic infections , bronchitis (4.7%) and diarrhoea (3.0%). Investigators considered 20.4% of the emergent adverse events as treatment-related . A significant difference between treatment groups . For AST and ALT, the proportion of patients with abnormal values decreased during the study from 31.5% on Day 0 to 20.9% on Day 28 for AST, and from 9.2% to 6.0% for ALT. Abnormal values reported for these enzymes were of mild intensity. Except for anemia recorded as an adverse event in seven patients, no abnormal laboratory values were reported as adverse events by the investigators.Two patients died during the study: one patient in the ASAQ1 group passed away after a lung infection and anemia on Day 3 while one patient in the AL group fell into a fatal coma of unknown cause on Day 1. Investigators did not attribute those deaths to study treatment. One other serious adverse event was reported in the ASAQ1 group, corresponding to a case of severe anemia requiring hospitalization (related to treatment according to the investigator). This event occurred on the fifth day after the last intake of treatment and the patient recovered after hospitalization.Three patients in the ASAQ1 group discontinued the study due to adverse events . These events were considered by the investigator to be severe and probably related to treatment. All patients recovered though.P. falciparum malaria after treatment with this new formulation. These results are consistent with those from a previous study where AS+AQ were routinely used as multiple tablets (loose combination) in Mali [By demonstrating the non-inferiority of the therapeutic efficacy of ASAQ with respect to AL, this study shows that remarkably accurate clinical and parasitological response can be obtained in patients suffering from uncomplicated  in Mali , Senegal in Mali  and in M in Mali . In the The two ASAQ1 and ASAQ2 regimens showed similar efficacy and tolerability. This indicates that both regimens are suitable from a clinical point of view, even though ASAQ1 dosing can be expected to improve general compliance thanks to an easier dosage compared to ASAQ2. Considering the present ASAQ packaging, an appropriate dosing may rely on the intake of one tablet per day for children and teenagers, and two tablets once a day for adults. Such a simple administration should help the communities better understand and comply with the treatment.et al, [Plasmodium sp.In agreement with previous studies on AS + AQ mentioned above, all three ACT regimens tested in this work induced a rapid parasitic clearance within three days and a 50% decrease of gametocytaemia in two weeks to reach the complete gametocytes elimination in most of the patients at the end of the study (Day 28). These results confirm those from Sowunmi et al,   mother or caretakers facing a \"sleepy\" episode. Seven cases of severe transient and asymptomatic neutropenia occurred during this study independently of the treatment group. In a cohort of 382 Ugandan patients treated with AS + AQ, AQ + SP or AL, Maiteki-Sebuguzi et al  reportedIn Africa, the current national policy approach for malaria control is to strengthen the fight through a global plan of elimination, involving reduction of malaria incidence to a very low predetermined level . BesidesP. falciparum malaria in adults and children. Effectiveness studies are required now to estimate whether combining a limited number of tablets with a single daily dosage will have a real positive impact on patient compliance. The successful deployment of ASAQ fixed-dose requires operational pharmacovigilance programs which should be conducted to assess its long-term safety profile.The ASAQ fixed-dose combination administered once daily provided clinical and parasitological efficacy comparable to AL. Non-inferiority of ASAQ with respect to AL was also demonstrated in children under five years of age. Dividing the ASAQ intake into two daily doses does not confer any advantage in terms of efficacy or safety. The ASAQ fixed-dose combination thus offers an effective and safe option to treat uncomplicated The authors declare that they have no competing interests. All authors received a stipend from the study sponsor (sanofi-aventis) to cover participation in the study. Valerie Lameyre is employed by sanofi-aventis.All authors contributed to study design and implementation, patients' inclusion and following-up and data interpretation. All authors read and approved the final manuscript. Especially manuscript revision was due to the commitment of MR, PB and VL. MR and PB are guarantors of the paper.\u00ae (artesunate + amodiaquine fixed-dose combination tablets). The sponsor was involved in the study design, the writing of the report, the decision to submit the paper for publication and providing editorial support for the first draft of the manuscript.This study was initiated and funded by sanofi-aventis, manufacturer of Coarsucam\u2122/Artesunate Amodiaquine WinthropAn independent Study Supervisory Committee was established to oversee design and implementation of the study. This committee was consulted before finalizing the protocol and was regularly informed on study progression. It was consulted for notification of adverse events and for classification of protocol deviations. This committee could review any case report forms containing conflicting or questionable efficacy or safety data under blinded conditions. It was consulted for discussion of the results of the study and provided input for finalization of the study report."}
+{"text": "By 2003 higher levels of SP resistance were recorded, prompting an urgent need for replacing the first line drug with ACT, as currently recommended by the World Health Organization. Despite this recommendation country-specific evidence-based data to support efficacy and safety profile of ACT is still limited. A study on the efficacy and safety of artesunate plus amodiaquine (AS+AQ) and artemether plus lumefantrine (AL). Patients were randomized to received either SP or amodiaquine monotherapy or treated with standard doses of AS+AQ in Mlimba and Coartem in Kyela and followed-up for 28 days to assess treatment responses. This study reports results of the combination therapies.An A total of 157 children (76 in Mlimba and 99 in Kyela) who were enrolled in to the study and treated with either AL or AS+AQ were successfully followed-up. Both combinations were tolerated and effected rapid fever and parasite clearance. The crude ACPRs were 80 (87%) and 41 (63%) for AL and AS+AQ respectively. However, after PCR adjustments the corresponding figures raised to 100% (n = 86) and 93.8% (n = 45) in AL and AS+AQ groups, respectively. The mean haemoglobin improved moderately from day 0 to day 28 by 1 g/dl in AL and 0.4 g/dl in AS+AQ treatment group and was statistically significant (p < 0.001 both).These findings provide substantial evidence that AL is highly efficacious in areas of high resistance of SP and supported the country's decision to switch from SP monotherapy to AL. Plasmodium falciparum resistance to commonly used antimalarials such as chloroquine (CQ) and sulphadoxine/pyrimethamine (SP) has posed major challenges to malaria control in sub-Saharan Africa. In the face of escalated resistance to these widely used and long utilized antimalarial the World Health Organization (WHO) currently recommends the use of artemisinin combination therapies (ACTs) as the first line treatment of malaria in sub-Saharan Africa [The emergence and spread of n Africa -3. Severn Africa .invivo studies on efficacy of SP and other newly registered antimalarials geared to increase choices and preparedness should the need for policy revision arise [invivo study was carried out on the efficacy of some ACT drugs with a view of supporting the National Malaria Control Programme (NMCP) in reviewing the antimalarial drug treatment policy in Tanzania.In 2001, the Tanzanian Ministry of Health and Social Welfare switched its first line drug from CQ to SP Prior to this change, CQ has remained in use as the first line drug for over 45 years and had recorded day 14 parasitological cure rates of 10.3% ,7. In adon arise -10. In tinvivo efficacy testing framework of the Tanzania NMCP/East African Network for Monitoring Antimalarial (EANMAT). Two health facilities took part in the study; Ipinda in Kyela District at the boarder with Malawi and Mlimba in Kilombero District in South-eastern Tanzania. This study was conducted in 2004 between January \u2013 June and March \u2013 October at Ipinda and Mlimba health facilities, respectively. At both sites malaria transmission is perennial and peaks between May and July, after the long seasonal rainfall.The study was conducted within the P. falciparum count of 50\u20135,000/200 white blood cells (WBC) assumed to be 2,000\u2013200,000 parasites/\u03bcl. Patients were excluded from the study if they present with repeated convulsion, inability to take anything orally, severe anaemia i.e. Hb \u2264 5 gm/dl, difficult in breathing or patient with signs consistent with renal failure and patient's parent/guardian unwillingness to participate.The WHO standardized protocol for the assessment of therapeutic efficacy of antimalarial drugs (WHO 2000) was used and the study included sick children who were 6\u201359 months age if they presented with history of fever in the past 24 hours or axillary temperature of \u226537.5\u00b0C, and mono-infection of \u00ae) was used as the ACT while artesunate and amodiaquine (AS+AQ) in an ad-hoc as ACT and the same monotherapies were given at Mlimba. This study reports only the outcome of both combination therapies. AL was given to children according to bodyweight as follows 20/120 mg tablet to those weighing 5\u201314 kg and two tablets 40/240 mg to children with 15\u201324 kg. The full course of treatment for all study patients in this group consisted of 6-doses of AL that was given at 0, 8 and 16 hours, the remaining doses were given at 12-hourly intervals for a total of three days. At Mlimba sentinel site AS+AQ was given at a dose of 4 mg/kg and 10 mg/kg body weight respectively. AS+AQ was given on day 0, 1 and 2 with only amodiaquine's dose reduced to 5 mg/kg body weight on day 2. All treatments were supervised by study nurse and patients were observed for 30 minutes in the aftermath of drug intake. All patients who vomited within 30 minutes intervals were re-administered another full dose of the same medicine. All treated patients were followed for 28 days to assess clinical and parasitological responses. Patients that did not turn up for scheduled dose were visited at home by the study nurse on the same day. Treatment outcomes were classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) and adequate clinical and parasitological responders (ACPR) using the (WHO 2003) guidelines. Clinical therapeutic outcomes were adjusted by genotyping the P. falciparum merozoite surface protein 2 (msp2) and glutamate rich protein (glurp) on admission (Day 0) and any day of infection recurrences . Recrudescence was differentiated from new-infections as described by Mugittu et al 2006 [The patients were randomized to receive either monotherapy or the ACT according to the NMCP schedule of sentinel testing. At Ipinda sentinel site SP and amodiaquine were used as the monotherapies and artemether/lumefantrine (AL) and the Hb of \u22645 g/dl was considered as severe anaemia. Parasitological examinations involved preparation of thick and thin blood smears from each patient and examined by specialized microscopist from Ifakara Health Research and Development Centre (IHRDC). Parasitaemia was expressed as count per 200 WBC of blood assuming a normal leucocytes level of 8,000/\u03bcl.Patient follow up were scheduled on days 1, 2, 3,7,14, 21 and 28. On each visit, including day 0 under which patient were enrolled. Clinical, parasitological and haematological examinations were performed. Haemoglobin level (Hb) was assessed using HemoCue\u00ae database software version 7  at Ifakara Health Research and Development Centre (IHRDC). Data analysis was performed using STATA\u00ae statistical analysis software package version 8 . Descriptive analysis was done and differences in proportions of treatment outcome were compared using chi square test for proportions. Student's t-test was applied for continuous variables. Data on patients that were excluded for different reasons and those that were loss to follow up were not considered in the final analysis.Data generated in patient's case record forms were entered on FoxProA total of 99 and 76 patients were enrolled at Ipinda and Mlimba health facilities, respectively. Table Both drugs were tolerated; there was no report of significant Adverse Drug Reaction (ADR). Table The mean haemoglobin improved moderately from day 0 to day 28 by 1 g/dl in AL and 0.4 g/dl in AS+AQ treatment groups. The mean haemoglobin recovery after day 28 was statically significant in both groups (p < 0.001 both).invivo ACT efficacy study was to support the establishment of evidence-based results that can be used to change malaria treatment policy in Tanzania. This study has demonstrated high therapeutic efficacy and tolerability for a six dose regimen of AL and a 3-day course of AS+AQ in southern Tanzania. Both crude and PCR-corrected ACPRs for AL (87% and 100%) were higher than those recorded in the AS+AQ (63.2% and 93.8%) arm. The AL crude and PCR-corrected cure rates are more or less similar to those recorded in Muheza Tanzania in 2005 (79% and 97.2%), [The main goal of this  97.2%),  and in m 97.2%), .The testing of each drug independently for each site was due to the arrangements in place that required testing each individual drug as part of antimalarial nationwide drug testing allocation. This allows accumulation of evidence of the performance of the ACT in different settings. It can be argued that there are differences from place to place related to the response to treatment that is observed but treatment policies are formulated at regional or sub-regional level hence the need to get regional summary estimates. The sentinel system is a good approach toward addressing that need. Secondly it will be very difficult to sample all possible places where variation is being expected and under the current malaria transmission intensity in Tanzania, it is hard to follow up enough patients for all treatment groups in a single site during the same transmission year .An interpretation of the comparative efficacy of the two drugs was not considered in our analysis. The reasons for this approach are; first this study was not designed to measure the differences between two test drugs but only to get point estimates of the efficacy which also allows monitoring of their performance overtime. Second, both ACT drugs have high efficacy profiles in the region that in principal would require huge sample size to be able to show a comparative difference ,15. For According to the current malaria treatment policy revision guidelines (changing policy at >10% failure rate), both AL and AS+AQ would be considered suitable drugs for first line purpose since both recorded high PCR-corrected efficacies i.e. 100% and 93.8%, respectively. However, it was more rational to adopt AL as its efficacy is far above the cut-off point, whereas AS+AQ efficacy is just 3 units higher from the policy revision cut-off point and its useful therapeutic life might be compromised sooner following widespread use. In addition, the policy revision process took into account the suggestion that apart from reasonably high efficacies of partner drugs, an effective combination therapy should comprise of drugs that have not been deployed previously in the area for use as monotherapies . Prior tAM was responsible for the protocol development, study design, field trial set up data analysis and developing of the manuscript. SA was responsible for study design, training research assistants and developing the manuscript. KM participated in manuscript development and for comments on the earlier version. DS participated in molecular genotyping of recurrent infections. RM & AM were responsible for study design."}
+{"text": "Plasmodium vivax accounts for about 40% of all malaria infection in Ethiopia. Chloroquine (CQ) is the first line treatment for confirmed P. vivax malaria in the country. The first report of CQ treatment failure in P. vivax was from Debre Zeit, which suggested the presence of chloroquine resistance.in vivo drug efficacy study was conducted in Debre Zeit from June to August 2006. Eighty-seven patients with microscopically confirmed P. vivax malaria, aged between 8 months and 52 years, were recruited and treated under supervision with CQ (25 mg/kg over three days). Clinical and parasitological parameters were assessed during the 28 day follow-up period. CQ and desethylchloroquine (DCQ) blood and serum concentrations were determined with high performance liquid chromatography (HPLC) in patients who showed recurrent parasitaemia.An P. falciparum infection during follow-up. A total of 83 (95%) of the study participants completed the follow-up. On enrolment, 39.8% had documented fever and 60.2% had a history of fever. The geometric mean parasite density of the patients was 7045 parasites/\u03bcl. Among these, four patients had recurrent parasitaemia on Day 28. The blood CQ plus DCQ concentrations of these four patients were all above the minimal effective concentration (> 100 ng/ml).Of the 87 patients recruited in the study, one was lost to follow-up and three were excluded due to P. vivax parasites are emerging in Debre Zeit, Ethiopia. A multi-centre national survey is needed to better understand the extent of P. vivax resistance to CQ in Ethiopia.Chloroquine-resistant  Plasmodium vivax malaria is the most geographically widespread and the second prevalent cause of malaria globally. Among 2.6 billion people at risk of malaria infection, annual estimates of P. vivax cases range from 130 to 435 million [P. vivax malaria on the continent, accounting for approximately 40% of all infection in the country [P. vivax malaria in most parts of the world. However, CQ resistance in P. vivax has compromised its use since the first reported cases from Papua New Guinea in 1989 [P. vivax has been reported from the north-eastern coast of Indonesian Papua in 2003 [Chloroquine (CQ) is the first line treatment for  in 1989 . Since t in 1989 , Myanmar in 1989 ,7, India in 1989 , and par in 1989 -13, Colu in 1989 , Vietnam in 1989 , and Per in 1989 . Resista in 1989 . To date in 2003 .P. falciparum in studies conducted at 18 sites between 1997/98 [falciparum malaria in 1999 [CQ has also been the first line drug in Ethiopia for uncomplicated malaria since 1950. Following the findings of CQ treatment failure as high as 65% for  in 1999 . SP was  in 1999 . HoweverP. falciparun and P. vivax CQ treatment failure in Debre Zeit, was in 1995, when only 4 of 29 (14%) P. falciparum patients cleared their asexual parasites by day 7 and 2% of 225 P. vivax patients who were followed for seven days failed to respond to CQ treatment [P. vivax being more prevalent (70%) than P. falciparum (30%). Malaria transmission is generally more intense following the main rainy season, which occurs from September to December. The aim of the present study was to determine the rate of CQ treatment failure in P. vivax and confirm whether treatment failure is due to drug resistance or sub-therapeutic antimalarial drug concentrations.The first report of 2. It is located in Oromia region 2 P (1-P) was used for calculation. Assuming a loss-to-follow-up rate of 20% over a 28-day study, a total of 87 study participants were enrolled.The sample size was calculated based on the expected proportion of P. vivax mono-infection with \u2265250 parasites/\u03bcl of blood and elevated axillary temperature (\u226537.5\u00b0C) or a history of fever within the previous 48 hours and who gave informed consent, or parent's or guardian's consent and assent (for children aged 12\u201317 years) were enrolled in the study. Patients were excluded if they had signs of severe malaria according to WHO criteria [Patients were recruited according to the WHO protocol for monitoring anti-malarial drug resistance . Patientcriteria , had anoFollowing the treatment guidelines of the Federal Ministry of Health of Ethiopia (FMOH), 25 mg/kg of chloroquine phosphate  was administered over a 3-day period (10 mg/kg on days 0 and 1 and 5 mg/kg on day 2) . All druTreatment success was defined as clearance of parasitaemia with no reappearance within the 28 days of follow-up period. Treatment failure due to CQ resistance was defined as parasitaemia recurred during the follow-up period in the presence of therapeutic concentrations of CQ and DCQ (i.e \u2265 100 ng/ml as determined using HPLC) ,28.The study was approved by the Addis Ababa University Ethical Review Committee, the AHRI/ALERT Ethical Review Committee and the National Ethical Review Committee.Thick and thin blood film slides were prepared using 10% Giemsa solution for 30 min. The stained slides were examined under a light microscope using 100\u00d7 oil immersion by an experienced laboratory technician. Parasitaemia was calculated per 200 white blood cells (WBC) assuming 8000 WBC/\u03bcl of blood . A totalDrug disintegration, dissolution, identification and assay tests were done to assess the quality of the drug used in the study . The tested batch of CQ tablets was determined as per the British Pharmacopoeia (BP) .et al [Blood concentrations of CQ and DCQ were measured using the HPLC method described by Bell et al , with so\u00ae, Macclesfield, Cheshire, UK), preceded by a guard column . A variable wavelength UV-visible detector  set at 340 nm was used. Chromatographic data were recorded on a ChromoJet CH1 data integrator .The chromatographic system consisted of a gradient delivery pump  connected to a syringe loading injector  equipped with a 50-\u03bcl loop. Chromatographic separation was performed using a stainless steel analytical column . STATA version 7.0  was used for analysis. The survival analysis method of A total of 2,422 consecutive patients self-reported to have symptoms of malaria, 87 patients who satisfied the inclusion/exclusion criteria were recruited. The median age of study participants was 16 years (range: 8 months to 52 years). Twenty point seven percent (n = 18) of the study participants were under five years of age. The mean \u00b1 standard deviation (SD) for duration of illness was 3.15 \u00b1 1.85 days. Among the study participants, 62.1% had a history of fever and 37.9% had documented fever at the time of diagnosis. The geometric mean parasite density at day 0 was 6,614.6 parasites/\u03bcl , headache (86.2%) and chills/rigor (78%). Seven percent (n = 7) of the study participants vomited the first dose of CQ and they were retreated with a similar dose. None of them vomited a second time.P. falciparum infection during follow-up. One was found positive for asexual stages of P. falciparum on day 21 and the other two were positive for P. falciparum gametocytes on day 1 and 3. Only one participant was lost to follow-up while the remaining 83 study participants completed the 28-day follow-up.Among the 87 study participants, three patients were excluded from the study due to Based on therapeutic failure risk calculated by the Kaplan-Meier survival analysis, the number of patients at risk on day 0 was 87 and these fell to 83 at day 28. Among the 83 patients, 42% (n = 35) had cleared their parasitaemia by day 1 and 98% (n = 81) by day 3 while 100% parasite clearance was observed by day 7. However, 4.6% (n = 4) of CQ treatment failure was observed during the follow-up period. This was observed in four children below seven years of age who developed symptomatic recurrent parasitaemia at day 28. This puts the risk of CQ failure on day 28 at 4.6%  observed from the parasitic density observed at day 0 in the patients who cleared their parasitaemia.To monitor CQ absorption, CQ plus DCQ (CQ-DCQ) serum concentration of 83 samples (four patients with recurrent parasitaemia and 79 patients with treatment success) was determined on day 2. The median CQ-DCQ concentration was 882.3 ng/ml (range: 166.8\u20136714.6 ng/ml). There was no significant difference observed in the CQ-DCQ concentration at day 2 between the patients who showed recurrent parasitaemia and patients who had cleared their parasitaemia within the 28 days follow-up period .In order to confirm the cause of clinical treatment failure observed in the present study, blood drug concentration of CQ-DCQ was determined. The concentrations of CQ-DCQ in the blood at day 28 for patients who showed recurrent parasitaemia were all above the minimal effective concentration  Table . Furtheret al [PCR was run on five paired samples (four from patients who had shown recurrent parasitaemia and one patient who had cleared parasitaemia during the follow-up period). However, it was only possible to amplify five isolates: four day 0 samples and one day 28 sample using primers published by Bruce et al . CompariP. vivax in Debre Zeit, Ethiopia. This was confirmed with a quantitative analysis of the blood concentrations of CQ-DCQ, which were above MEC (100 ng/ml of blood). In addition, the presence of CQ-DCQ concentrations on day 2  showed that CQ was well absorbed from the gastrointestinal tract indicating that there were no other factors that might have influenced the kinetics of the drug including rapid excretion or poor absorption. Although only three samples from patients with successful treatment response were tested, blood concentrations of CQ-DCQ of these patients were above the MEC, which is in accord with a previous report by Baird et al [According to the WHO protocol, resistance is defined as the presentation of signs of severe malaria within the first two days after supervised treatment or the presence of parasitaemia and axillary temperature > 37.5\u00b0C between day 3 and 28 or presence of parasitaemia on any day between day 7 and day 28, irrespective of clinical conditions . In addiP. vivax from Africa. A previous report by Tulu et al in Debre Zeit has showed the presence of 2% (255) treatment failures in using seven days in vivo efficacy test [This is the first report of CQ resistance in acy test . HoweverP. vivax. In present study, msp 3-alpha was used to genotype P. vivax isolates from patients who showed parasitaemia. The primers failed to amplify parasite isolates which could be due to the mismatch at the 3' end of the primers, thus not attached to the parasite DNA which, lead to an inconclusive result for the genotype. One limitation of this study is that it did not explore other marker genes, including Pvcsp and Pvmsp1, which had been applied elsewhere for genotyping in P. vivax parasite isolates to obtain additional information on the diversity of the resistant isolates [In addition to the determination of drug concentrations in patient blood samples, molecular genotyping is recommended in studies of anti-malarial drug efficacy . This meisolates .P. vivax as this method assumes that the primary parasite population is the same as in recurrent parasitaemia due to relapse [P. vivax, clinical follow-up in combination with determination of CQ-DCQ concentrations in the blood at a time of recurrent parasitaemia will give a definitive diagnosis rather than follow-up in combination with genotyping. Since mixed parasite strains were observed in day 28 samples, when compared to day 0 sample, it was difficult to categorize as recrudescence/relapse from new infection.Though genotyping is recommended in differentiating relapse/recrudescence and re-infection, it has shortcomings when used for  relapse . However relapse  in which relapse , thus le relapse . MoreoveP. falciparum. However, there have been no systematic studies for P. vivax. The present data confirmed the emergence CQ resistant P. vivax in the Debre Zeit. Even though the level of resistance is not so high as to require a treatment policy change, the finding will be useful in alerting the responsible authorities to monitor the level of drug resistance and to determine the extent of this problem in all parts of the country, bearing in mind that P. vivax malaria is responsible for 40% of malaria cases in Ethiopia.In Ethiopia, there is a surveillance system for assessing the efficacy of first line drug for the treatment of P. vivax parasites are emerging in Debre Zeit, Ethiopia. There is a need for regular monitoring of the pattern of resistance to antimalarial drugs in the country.Chloroquine-resistant The authors declare that they have no competing interests.HT was involved in all aspects of the project, data collection, analysis, interpretation, supervising the project at the study site, and determined CQ-DCQ concentrations using HPLC and in writing of the manuscript. BP, LY, HE, AA have made a contribution in the design, data interpretation, work supervision and in critically revising the manuscript. GT has made a contribution in identifying the problem and on the concept of the study. SM has assisted in conducting the HPLC method validation and determination of CQ-DCQ concentrations in the patients' blood samples, and helped in the analysis of the results. GK has facilitated the HPLC laboratory work at the Kenyan Medical Research Institute. IE has contributed ideas for the project and revised the manuscript. All authors read and approved the final version of the manuscript."}
+{"text": "Vivax malaria remains a major cause of morbidity in the subtropics. To undermine the stability of the disease, drugs are required that prevent relapse and provide reservoir reduction. A 14-day course of primaquine (PQ) is effective but cannot safely be used in routine practice because of its interaction with glucose-6-phosphate dehydrogenase (G6PD) deficiency for which testing is seldom available. Safe and effective use of PQ without the need for G6PD testing would be ideal. The efficacy and safety of an 8-week, once weekly PQ regimen was compared with current standard treatment  and a 14-day PQ regimen.Plasmodium vivax patients were randomly assigned to either once weekly 8-week PQ (0.75mg/kg/week), once weekly 8-week placebo, or 14-day PQ (0.5mg/kg/day) in North West Frontier Province, Pakistan. All patients were treated with a standard chloroquine dose and tested for G6PD deficiency. Deficient patients were assigned to the 8-week PQ group. Failure was defined as any subsequent episode of vivax malaria over 11 months of observation. There were 22/71 (31.0%) failures in the placebo group and 1/55 (1.8%) and 4/75 (5.1%) failures in the 14-day and 8-week PQ groups, respectively. Adjusted odds ratios were: for 8-week PQ vs. placebo-0.05  and for 14-day PQ vs. placebo-0.01 . Restricted analysis allowing for a post-treatment prophylactic effect confirmed that the 8-week regimen was superior to current treatment. Only one G6PD deficient patient presented. There were no serious adverse events.200 microscopically confirmed A practical radical treatment for vivax malaria is essential for control and elimination of the disease. The 8-week PQ course is more effective at preventing relapse than current treatment with chloroquine alone. Widespread use of the 8-week regimen could make an important contribution to reservoir reduction or regional elimination where G6PD testing is not available.NCT00158587ClinicalTrials.gov  Plasmodium vivax is a common cause of malaria in the subtropics. Estimates put the global burden at 70\u201380 million cases per year P. vivax causes relatively few deaths there is increasing evidence suggesting that severe and life-threatening complications are more common than previously thought P. vivax has major deleterious effects on development and economic performance both at individual and national levels P.vivax is the predominant species Conventional transmission control methods targeting the mosquito vector are imperfect owing to the infectious reservoir; dormant hypnozoites in the liver produce episodes of relapse for several years after initial infection A truncated 5-day course of PQ for vivax malaria is used commonly in South Asia to reduce the risk of haemolysis and enhance adherence to treatment Studies from the 1960s provide evidence suggesting that successful PQ therapy is not a function of the length the treatment course, nor of the concentration of drug, but of the total dosage administered. Alving et al The aim of the study was to test whether an 8-week PQ regimen is effective at radical cure without the associated risk of haemolysis in G6PD deficient individuals. This regimen may be appropriate for deployment in resource-poor settings where G6PD testing is unavailable. It was compared to current standard treatment (chloroquine) given with 8-week placebo and to a regimen known to be effective (chloroquine plus 14-day PQ). Since treatment with chloroquine alone is current standard treatment, superiority of the 8-week course over placebo was the primary comparison.P. vivax (85\u201395% of cases) The study was conducted in Adizai, Baghicha and Khagan villages, close to Peshawar, Northwest Frontier Province, Pakistan where Afghan refugees have been resident for more than 20 years. Malaria transmission is seasonal and predominantly due to Ethical approval for the study was granted by the Pakistan Medical Research Council Committee on Bioethics and the London School of Hygiene and Tropical Medicine ethics committee. Permission was also obtained from local government agencies and the United Nations High Commissioner for Refugees. The study was prospectively registered at clinicaltrials.gov (number: NCT00158587). Sponsors and funding agencies  had no role in study design; collection, analysis, and interpretation of data; writing of the paper; or decision to submit for publication.P. vivax infection were asked to participate in the study following informed consent and if they met the inclusion and exclusion criteria. Consent was obtained in writing by patients or their guardians, and in the case of those unable to read was witnessed by a literate person. Inclusion criteria were: Patients diagnosed with P. vivax parasitaemia at study BHUs; Patients over 3 years of age; Patient permanently resident in the village. Exclusion criteria were: pregnancy or lactation; severe clinical anaemia (<7g/dl); P. falciparum and/or P. vivax (mixed infections); intake of any antimalarial drug in the 2 weeks prior to consultation; patients unavailable for the duration of follow up (11 months); patients with concomitant infections or disease likely to mask treatment response.Patients attending the basic health units with symptoms compatible with malaria had Giemsa stained thick and thin blood films obtained by finger prick and examined by trained microscopists. Those diagnosed with The study was designed as an open label, randomised, placebo controlled study. Patients were randomly allocated to one of the three treatment groups by study staff in the clinic once informed consent was received and inclusion/exclusion criteria assessed. For practical reasons, two randomisation methods were used. In Baghicha and Khagan villages, patients were randomised by household, whereas in Adizai randomisation was at the individual level. Randomisation lists for each village were generated using a random number list  by staff not involved in patient recruitment. Patients were randomised on enrolment by study staff in the BHUs based on house number or sequential patient numbers, depending on the study site.All patients were treated with initial 3-day chloroquine  for acute disease. This was accompanied by either supervised weekly placebo once per week for 8 weeks; supervised PQ treatment, daily for 14 days (0.5mg/kg per day); or supervised PQ once per week for 8 weeks (0.75mg/kg per week). All patients were tested for G6PD deficiency at enrolment using a colorimetric test . Those with G6PD deficiency were not randomised, but assigned to the 8-week PQ group in order to follow closely to assess safety.Patients received directly observed treatment according to the dosing schedule for each treatment group. Each patient was given an appointment card and if the patient was not present on the morning of their scheduled dose they were visited in the afternoon at their home. If the patient was not present they were treated on the next day. If they were absent on this second day, they were counted as losses to follow-up. Each dose was recorded with a signature or finger print of the patient. Patients were monitored during the eight weeks of treatment and for nine additional months by active surveillance (being visited in their homes every two weeks) and passively on presentation at the basic health centre. Patients presenting at the basic health unit with suspected treatment failure (febrile illness) were assessed by thick and thin blood smear. If positive they were classified as a treatment failure and re-treated with the original treatment given at enrolment. Blood slides were double read by two independent microscopists, blinded to the others result.The primary outcome was the occurrence of any episode of microscopically confirmed vivax malaria over the 11 month observation period which classified the patient as a treatment failure. Sample size estimation was based on the assumption of 32% and 49% failure in treated (14-day PQ) and untreated individuals, as recorded previously in other studies over a 9 month post treatment follow-up a priori basis and were included in multivariate analysis. Each treatment group was compared in turn to assess superiority (or otherwise) using logistic regression analysis, adjusting for potential confounders. Kaplan-Meier survival analysis using time to first relapse as the endpoint was used to calculate cumulative probability of treatment failure . Losses to follow-up were treated as censured data in the analysis. Data was recorded by trained health workers on patient record forms, double entered using Excel XP  and analysed using STATA v10.0 .Analysis was conducted on an intention-to-treat basis. In addition to the primary outcome, secondary outcome variables included the number of subsequent episodes and anaemia rates during and up to 2 weeks post-treatment as well as any notable adverse events. Univariate logistic regression analysis using the primary outcome provided crude odds ratios (OR). Potential confounders  were identified on an The protocol for this trial and supporting CONSORT checklist are available as supporting information; see th September 2004 until 17th July 2006. Follow-up was completed on 16th June 2007. The number (%) recruited into the study at each site was: Adizai 100 (50.0%), Baghicha 79 (39.5%), and Khagan 21 (10.5%). Ten (5.0%) were either lost to follow-up (9) or withdrawn owing to protocol violation (1). Seventy one (35.5%) patients were enrolled to 8-week placebo, 55 (27.5%) to 14-day PQ and 74 to 8-week placebo (37.0%). 200 patients were recruited from 13There were 22/71 (31.0%) failures in the placebo group and 1/55 (1.8%) and 4/75 (5.1%) failures in the 14-day and 8-week PQ groups respectively, giving rates of failure (per 1000/pers months) of 37.3 (95%CI: 24.6\u201356.6), 1.7 (0.2\u201312.0) and 5.5 (2.1\u201314.7), respectively. Treatment group and village were independently associated with treatment failure, with highest failure rates recorded in the placebo group and in Adizai village . Sex wasSince a prophylactic effect of PQ administered over the 8 week treatment period may confound true failures, two restricted analyses were conducted which included only those who had failed treatment in the post treatment period. The first was restricted to failures occuring in months 2\u201311. The second took account of the time the drug was potentially providing a prophylactic effect, and included the period of administration of active drugs plus an additional 30 day period, post-treatment. For this analysis data were censured at 33 days for the placebo group, 44 days for the 14 day PQ group and 91 days for the 8 week PQ group . The dat2\u200a=\u200a22.1, p<0.001) (p<0.001) . Figure There were no reported serious or notable non-serious adverse events and all treatments were well tolerated. Only 1 G6PD deficient patient, a 13 year-old male, was detected during this trial. On days 7 and 14 (prior to the second and third PQ doses), Hb in this individual fell below the confidence limits of the mean Hb in G6PD normal females aged 12\u201314 (n\u200a=\u200a13). On day 7 Hb was 10.0g/dl vs 12.6g/dl (95%CI: 11.8\u201313.4) and on day 14 Hb was 10.6 vs 12.6 (95%CI: 12.0\u201313.3). By day 21 (prior to the fourth dose) Hb in the patient was within the limits of age and sex matched G6PD normal individuals (hb\u200a=\u200a12.7 vs 12.6 [95%CI: 11.8\u201313.5]). The haemoglobin profile of the three treatment arms in the whole sample did not differ. No patient became seriously anaemic (Hb<7.0 g/dl), and no observed anaemia was clinically significant.The 8-week PQ regimen in combination with chloroquine is effective at curing acute vivax malaria and preventing relapses. Over a period of 11 months, new episodes of vivax malaria were more frequent in the group administered with placebo than in either PQ treated group and the superiority of 8-week PQ over chloroquine alone is demonstrated. The analysis provides information on the effect of an initial PQ treatment as well as repeated doses. The use of incidence of first relapse as the primary outcome provides evidence that the 8-week course is superior to placebo in prevention of episodes of relapse when given as a single course. Patients who presented with a second (or further) episodes were treated with the same regimen as at enrolment. Incidence of recurrent episodes was lower in both the PQ groups; only one recurrent episode was seen in any patient, whereas up to five recurrent episodes were seen in one patient in the placebo group.The restricted analysis, which used two approaches to exclude failures during the treatment period (when any prophylactic effect would be seen), confirms that both PQ treated groups are superior to placebo in preventing recurrent episodes. The pattern of failure in the 8-week PQ group, where failures occurred between two and three months post treatment, provides possible evidence of a prophylactic effect; the timing and pattern of the failures matches that of the placebo arm once the drug (and metabolites) have cleared. It could be, therefore, that failures (relapses) are simply delayed by a prophylactic effect of 8-week PQ therapy. If this were the case, however, failed patients treated with PQ for an additional eight weeks would be expected to have differing failure rates when censured at different time-points to account for the prophylactic effect. This effect is not seen; none of the 8-week PQ group had second relapse episodes . The perA recent meta-analysis of 14-day PQ trials describes regional variation in PQ efficacy in India, Brazil and Thailand Efficacy and safety data on 8-week PQ is scarce, based on small sample sizes with no comparison group An important issue in broadening access to 8-week PQ therapy is adherence to the regimen. Ensuring adherence to a 2 month, once weekly regimen will require proven interventions at the delivery level. Unsupervised 14-day PQ, accompanied by strong health education messages was equally effective to supervised therapy in terms of treatment outcome amongst Afghan refugees in Pakistan Since the idea of malaria elimination (or eradication) has again reached prominence Checklist S1CONSORT Checklist.(0.06 MB DOC)Click here for additional data file.Protocol S1Trial Protocol.(0.20 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria. There are few reports evaluating co-artemether in very young Nigerian infants and children. Results of the evaluation of the six-dose regimen in very young infants and children in Nigeria are presented in this report.The six-dose regimen of artemether-lumefantrine (AL) is now considered the gold standard for the treatment of uncomplicated As part of a larger African study, this open label, non-comparative trial, assessed the efficacy and safety of six-dose regimen of AL tablets in 103 Nigerian infants and children weighing between five and 25 kg suffering from acute uncomplicated malaria. Treatment was administered under supervision over three days with children as in-patients. 12-lead ECG tracings were taken pre-treatment and at day 3.Ninety-three infants and children completed the study as stipulated by the protocol. Mean fever and parasite clearance times for the intent to treat population (ITT) were 24.9 h \u00b1 (1.28) and 26 h \u00b1 (4.14) and the corresponding figures for the per-protocol population (PP) were 19.24 h \u00b1 13.9 and 25.62 h \u00b1 11.25 respectively. Day 14 cure rates for the ITT and PP were 95.1% and 100% respectively while day 28 cure rates were 91.3% and 95.7% respectively. The overall PCR corrected day 28 cure rate was 95.1% for the ITT. The six-dose regimen of AL was well tolerated with no drug-related serious adverse events. Although six patients recorded a QTc prolongation of > 60 ms on D3 over D0 recording, no patient recorded a QTc interval > 500 ms.The six-dose regimen of AL tablets is safe and effective for the treatment of acute uncomplicated malaria in Nigerian infants and children weighing between five and 25 kg.NCT00709969 Plasmodium falciparum [P. falciparum was rare at the time, such as China and India [The deleterious consequences of malaria continue to increase in endemic areas as a result of the emergence and widespread dissemination of drug resistant lciparum ,2. Childnd India ,4, but nnd India . Subseqund India ,7. The snd India .The safety and efficacy of the six-dose regimen of AL in infants and children weighing between five and 25 kg for the treatment of acute uncomplicated falciparum malaria was evaluated in this study. Special attention was paid to the response to treatment by non-immune infants between five and 10 kg. Infants weighing between five and 10 kg, who had never had an episode of malaria prior to the current episode were considered to be non-immune to malaria. Evaluations for neurotoxicity were included in this study based on reports of neurotoxicity associated with artemisinins ,10. In a\u00ae Treatment Group\" of the multicentre study evaluating the efficacy and safety of AL in the management of malaria in African infants and children [This report is based on data from the \"Nigerian Coartemchildren .The study was conducted at the General Out Patient Clinic and paediatric wards of the University College Hospital, Ibadan, south-western Nigeria between August 2002 and February 2003. Malaria transmission is intense throughout the year in the study area with peak transmission occurring during the rainy season months of May to October. Ethical approval for the study was obtained from the Joint University of Ibadan/University College Hospital Institutional Review Committee and the ethical review board of the World Health Organization, Geneva, before commencement. The study was conducted according to the declaration of Helsinki and its amendments. Good clinical practice was also adhered to and a written informed consent or witnessed verbal informed consent was obtained from the parent or guardian of each enrolled child before any study related procedure was performed.\u00ae). An open label study design was chosen because at the time there were no affordable products approved by regulatory authorities for the treatment of malaria in children weighing < 10 kg; thus no suitable comparator was available. The study population was stratified into three body weight groups as follows: 5-<10 kg, 10-<15 kg and 15-\u226425 kg as body weight groups (BWG) 1, 2 and 3 respectively. Children weighing 5-<10 kg (BWG1) constituted half of the study population in the design so as to have enough data for evaluation in this group of children.In an open label, non-comparative study design, children weighing between five and 25 kg suffering from confirmed acute uncomplicated falciparum malaria were treated with AL  was carried out. All worsening, and newly occurring signs and symptoms of malaria after baseline, whether considered drug-related or not, but before recurrence of parasitaemia were defined as TESS. Thick and thin blood films were prepared from a finger prick and stained with 10% freshly prepared Giemsa stain and examined using x1,000 magnification of a light microscope for parasite detection, quantification and morphology. Parasite density was calculated using individual patient's white blood cell count. Blood (3.5 ml) was collected for haematology and clinical chemistry evaluation on days 0, 3 and 28. Haematology evaluation included haematocrit, haemoglobin, red blood cell count, white blood cell count with differential, platelet count and glucose-6-phosphate dehydrogenase activity, while blood chemistry evaluation included blood glucose, serum bilirubin, creatinine, alanine transaminase (ALT), aspartate transaminase (AST) and \u03b3-glutamyl transferase. Urine measurements were performed when considered necessary and parameters evaluated included specific gravity, haemoglobinuria, proteinuria and sediment. Blood spots were blotted on isocode stix for polymerase chain reaction (PCR) before commencement of drug therapy and at recurrence of parasitaemia for genotyping of isolates to distinguish between recrudescence and re-infection . In addi\u00ae  which consists of artemether (20 mg) and lumefantrine (120 mg) per tablet supervised by a doctor or nurse. Children weighing 5 to < 15 kg received 6 doses of 1 tablet each of AL at 0, 8, 24, 36, 48 and 60 hours while children weighing 15 to < 25 kg received 2 tablets of study drug at similar time points as above. Study drug was administered with sweetened condensed milk for older children and feeding encouraged soon after dosing while younger children took the study drug with water and were breast-fed immediately following drug administration. Patients were subsequently seen as outpatients daily for four days (up to day 7) and then on days 14, 21 and 28 during the post treatment phase.The study consisted of two phases: a treatment phase and a post treatment phase. During the first four days, termed the treatment phase, patients were admitted to the ward for drug therapy. Enrolled patients received CoartemEfficacy evaluation was based on parasitological cure rates at days 7, 14 and 28. Cure rates at days 7, 14 and 28 were defined as the proportion of patients cleared of asexual parasitaemia within the specific time intervals of initiation of treatment with AL without recrudescence within the specified days. Re-infection was distinguished from true recrudescence by genotyping using PCR . In addi3vRRms). Values < 430 and 450 ms were classified as normal for males and females respectively, 431\u2013450 and 451\u2013470 ms, borderline while > 450 and > 470 ms were classified as prolonged.Safety evaluation was based on the intent to treat population (ITT). Safety assessment consisted of monitoring and recording of all adverse events whether volunteered, discovered by questioning or detected by examination in addition to clinically significant changes in haematology, blood chemistry and urine values. Adverse events information was collected by evaluating TESS. An adverse event was regarded as any undesirable sign, symptom or medical condition occurring after starting the study drug, whether the event was considered to be related to study drug or not. Any signs and symptoms that appeared newly or worsened from baseline were recorded as adverse events. Standard 12-lead electrocardiographic tracings were done pre-treatment on day 0 and at day 3 as part of safety evaluation. Primary data as well as changes from baseline were derived from an independent blinded review of electrocardiographic results. Corrected QTc interval values were calculated according to Bazett's formula (QTc = QT/vRR) and Fridericia's formula . The PP population was made up of ITT patients who received all six doses of medication and who adhered to the study protocol. The variables were summarized using descriptive statistics  by body weight group and overall. PCT and FCT were derived by means of the Kaplan-Meier method. 95% confidence intervals for the cure rates were determined using the exact Pearson-Clopper limits. Time to fever clearance and time to parasite clearance were evaluated using Cox's proportional hazards regression analysis.One hundred and three consecutive children screened between August 2002 and February 2003 at the study centre and who fulfilled the inclusion criteria were enrolled in the study. The distribution of the ITT population consisted of 50 in BWG 1, 37 in BWG 2 and 16 in BWG 3  during the study. Biochemical parameters were not adversely affected by the study drug.The study drug was well tolerated and no patient was withdrawn due to recurrent vomiting or poor tolerability. Anaemia (n = 59) was the most frequently reported adverse event by laboratory evaluation, while cough (n = 33) was the most often volunteered adverse event reported. Respiratory infection (n = 31), diarrhoea (n = 20), vomiting (n = 18), rash (n = 17), clonus (n = 13) and insomnia (n = 11) were the next six most common treatment emergent adverse events recorded after baseline but before recurrence of parasitaemia among study participants. Other adverse events recorded among patients included hypothermia (n = 6), hyperreflexia (n = 6) and distended abdomen (n = 1). Five of the six episodes of hypothermia occurred among children in BWG1. Mean haemoglobin level decreased from 93.3 \u00b1 16.04 g/L on day 0 to 83.2 \u00b1 13.43 g/L on day 3 and increased to 103.2 \u00b1 14.76 g/L on day 28 while the number of children with low neutrophil count on day 0 decreased from 87 to 40 at day 28. In addition there was no record of eosinophilia, intravascular haemolysis or leucopaenia  developed similar illnesses and were brought to the study team for treatment. The siblings were diagnosed as having cholera. They responded well to treatment and went home well recovered. A verbal autopsy was conducted to identify the cause of death of the study participant.No patient reported any symptomatic cardiac adverse event. Mean heart rate and RR intervals at enrollment were 140.8 \u00b1 21.41/min and 437.0 \u00b1 70.15 ms while day 3 figures were 119.8 \u00b1 21.36/min and 518.4 \u00b1 100.8 ms respectively. There was a slight increase in mean PR and QTc intervals between day 0 and day 3. The mean PR interval increased from 117.7 \u00b1 14.84 ms at enrollment to 126.9 \u00b1 19.52 ms on day 3. The mean QTc values on days 0 and 3 were 400.6 \u00b1 23.64 ms and 410 \u00b1 21.31 ms respectively. There was no record of a QTc interval greater than 500 ms using Bazett's formula. However 6 patients  recorded QTc interval prolongation of over 60 ms on day 3 when compared with day 0. All were asymptomatic.P. falciparum malaria in Nigerian children and infants weighing as little as 5 kg. The youngest child in this study was five weeks of age. Three of the enrollees were less than six months of age while 48.5% were less than two years of age  by more than five fold. This underscores the importance of educating health care workers, patients and caregivers of the importance of compliance and the need to administer AL with food that contains fat, be it conventional milk, soya milk or some appropriate local food.The superior efficacy of the six-dose regimen used in this study when compared with that of the four-dose regimen is consistent with findings in similar studies that evaluated the four-dose and six-dose regimens in the western border of Thailand, in Tanzania, Kenya and the Gambia ,12,21. Met al  working l [et al  did not l [et al , in a reArtemisinin containing anti-malarial combinations is known to induce rapid parasite clearance and high parasitological cure rates. This expectation was met by the response profile obtained in this study. Approximately 75% of the PP population was free of patent parasitaemia 24 hours after initiation of therapy while none of the study population had detectable peripheral parasitaemia by day 3.In conclusion, the six-dose regimen of AL is safe and effective in the management of acute uncomplicated falciparum malaria in Nigerian infants and children weighing as little as five kg, including non-immune children in an area of worsening drug resistant falciparum malaria. The results of this study suggest that if properly deployed, AL could lead to a reduction and may also contribute to a significant extent in halting the worsening morbidity and mortality from malaria on the African continent ,2. The FAL: Artemether-lumefantrine; ALT: Alanine transaminase; AST: Aspartate transaminase; BWG: Body weight group; CI: Confidence interval; FCT: Fever clearance time; IEC: Information, education and communication; ITT: Intent to treat population; PCT: Parasite clearance time; PP: Per protocol population; TESS: Treatment emergent signs and symptoms; /\u03bcL: per micro litre;COF, OOO, HOD-A, AGF, AMJO and LAS declare no competing interests. PH and MV are employees of Novartis Pharma AG and PidP was an employee of Novartis at the time this work was undertaken.COF developed the study design, participated in the study conduct and interpretation of the data, drafted and critically reviewed the paper. OOO participated in the study conduct and interpretation of the data, and edited the draft manuscript. HOD-A and AGF participated in the study conduct and edited the draft manuscript. PIdP, PH, MV, AMJO and LAS participated in interpretation of data and critically reviewed the draft manuscript. All authors read and approved the final version of the manuscript."}
+{"text": "Plasmodium falciparum malaria. However, oral paediatric formulations are urgently needed, as the tablets are difficult to administer to young children, who cannot swallow whole tablets or tolerate the bitter taste of the crushed tablets.Artemether/lumefantrine (AL) has been adopted as the treatment of choice for uncomplicated malaria in Kenya and other countries in the region. Six-dose artemether/lumefantrine tablets are highly effective and safe for the treatment of infants and children weighing between five and 25 kg with uncomplicated \u00ae) or three-dose of artemether/lumefantrine suspension (Co-artesiane\u00ae) for uncomplicated falciparum malaria in western Kenya. The children were followed-up with clinical, parasitological and haematological evaluations over 28 days.A randomized, controlled, open-label trial was conducted comparing day 28 PCR corrected cure-rates in 245 children aged 6\u201359 months, treated over three days with either six-dose of artemether/lumefantrine tablets (CoartemNinety three percent (124/133) and 90% (121/134) children in the AL tablets and AL suspension arms respectively completed followed up. A per protocol analysis revealed a PCR-corrected parasitological cure rate of 96.0% at Day 28 in the AL tablets group and 93.4% in the AL suspension group, p = 0.40. Both drugs effectively cleared gametocytes and were well tolerated, with no difference in the overall incidence of adverse events.\u00ae) was not superior to six-dose artemether-lumefantrine tablets (Coartem\u00ae) for the treatment of uncomplicated malaria in children below five years of age in western Kenya.The once daily three-dose of artemether-lumefantrine suspension (Co-artesianeClinicalTrials.gov NCT00529867 As in mpy (ACT) .Previous studies in Africa have shown that the six-dose regimen of artemether-lumefantrine (AL) tablets is safe, efficacious and effective in the treatment of uncomplicated malaria in children less than five years of age, residing in areas with high levels of CQ and SP resistance -8. SubseAlthough children less than five years of age are the major target of anti-malarial drug therapy in malaria endemic regions, the available oral paediatric formulations of ACT are not optimal for this high-risk population; young children cannot swallow whole tablets and sometimes spit out the drug, because of the bitter taste of the crushed tablets. In addition, AL tablets are not recommended for patients weighing less than 5 kg. These drug administration problems influence prescription of ACT by health workers and patient adherence, resulting in either under-dosing or over-dosing. Within the context of home-based management of malaria, 10 \u2013 20% of participants were either non-adherent or administered incorrect doses of AL tablets -14. TherP. falciparum, with the remaining being mixed P. falciparum with Plasmodium malariae and rarely Plasmodium ovale infections. In vivo drug resistance to SP in children <5 years is 74.5% [This study was conducted at Chulaimbo Health Centre in Kisumu District in western Kenya. The facility serves a predominantly rural population in an area with high perennial malaria transmission in the lowlands around Lake Victoria. Transmission peaks during the long rains (March\u2013May) and the short rains (October\u2013December). The annual entomological inoculation rate in the surrounding area was 31.1 infectious bites/person/year for 12 months ending June 2004 . More this 74.5% ,20 and 5is 74.5% .P. falciparum, with parasitaemia in the range of 2,000\u2013200,000 asexual parasites per microlitre of blood; no other cause for fever than suspected malaria; and, no general danger signs or signs of severe and complicated falciparum malaria as per WHO guidelines [A 2003 WHO protocol for the assessment and monitoring of anti-malarial drug efficacy for the treatment of uncomplicated falciparum malaria was used . Childreidelines .A randomized, controlled, open-label trial design was used. PCR-corrected cure rate by day 28 after first dose was the primary endpoint used for computing sample size. Assuming a 94% cure rate with artemether-lumefantrine tablets and 99.9% with the artemether-lumefantrine suspension, a sample of 134 children was required  in each treatment arm to detect this 6% difference in the parasitological cure rates, with 80% power using a two-sided alpha of 0.05. The randomization code was computer-generated without stratification from which treatment groups were assigned.At enrolment, a medical history was obtained from parents/guardians including presenting symptoms, current medications, previous anti-malarial use and bed net use. A physical examination was performed, weight and axillary temperatures recorded and finger prick blood obtained for malaria smears, haemoglobin and blotted on filter paper for parasite genotyping.\u00ae, Novartis AG), crushed, mixed with water and administered at hours 0, 8, 24, 36, 48 and 60 over 3 days while the second group received artemether/lumefantrine powder for suspension (Co-artesiane\u00ae Dafra Pharma NV) containing 15 mg artemether and 90 mg lumefantrine per 5 ml after reconstitution. A bottle of suspension was prepared for each child and administered once daily at hour 0, 24 and 48 over 3 days. Treatment doses were calculated based on patient weight and the administration directly observed under in-patient care. All children were given a glass of milk or breast fed (for those still breastfeeding) after drug administration since treatment times did not always coincide with meals. They were then observed for 30 minutes after drug administration for vomiting. If vomiting occurred, the whole treatment dose was re-administered. If the re-treatment dose was vomited, rescue treatment with parenteral quinine was administered and the child was withdrawn from the study.Consecutively eligible children were randomly assigned into one of two treatment groups according to the randomization code. A study nurse prepared and administered the study medications, according to the treatment assignment. One group received 6 doses of artemether/lumefantrine 20/120 mg tablets  were given to the children on completion of the study.All treatment failures were treated with oral or parenteral quinine for 7 days depending on clinical presentation. Paracetamol was administered to children with temperature \u2265 38.0\u00b0C or at the clinician's discretion. Antihelminthics were given to all children >1 year who had not received any in the past 3 months while children with haemoglobin <10 g/dl were treated with ferrous sulphate for 14 days. Long lasting insecticide treated bed nets  and at the study clinic during scheduled visits on days 7, 14, and 28, after initiating study treatment, or on any other day if the child was unwell. During each scheduled visit, a brief clinical history was obtained and a physical examination was performed; blood smears for malaria parasites and filter paper spot samples were obtained. Patients who did not return to the clinic for scheduled visits by mid day were visited at home by the social worker and asked to come to the health facility. Patients were excluded from the study if they; 1) withdrew consent, 2) left the study area, or 3) reported taking anti-malarial medication during follow-up.\u00ae  on Days 0, 7, 14 and 28. Filter paper blood spots were collected on Days 0, 14 and 28 or any other day of recurrent parasitaemia. Paired filter paper samples of children who had parasitaemia during follow-up were used to extract parasite DNA for PCR to distinguish recrudescent from new infections as described by Snounou et al. [Blood was drawn from a finger prick to prepare thick and thin blood smears on days 0, 1, 2, 3, 7, 14, 28 and on any other unscheduled visit. The smears were air dried, stained with 3% Giemsa for 30 \u2013 45 minutes and read independently by two technologists. Parasite density was calculated by counting asexual parasites against a 500 leucocytes assuming a leukocyte count of 8,000/\u03bcL of blood to obtain number of parasites and gametocytes per microlitre (\u03bcL). Thin smears were examined for plasmodium parasite speciation. A slide was considered negative after scanning 100 high power fields. A third microscopist independently read discrepant slides. Haemoglobin was measured using HemoCueu et al. . Block 2This study was approved by the Kenya National/Kenya Medical Research Institute Ethics Committee. Written informed consent was obtained from all parents or guardians of eligible children prior to enrolment.t-test and analysis of variance (ANOVA). For skewed data, medians were computed and comparisons made using the Kruskal-Wallis one way ANOVA. Two tailed p-values < 0.05 were considered statistically significant.Data from case report forms were checked, double entered and verified for errors using Epi Info 2002 . Data was analysed using SPSS  and Epi-Info 2002. The primary efficacy endpoint was day 28 PCR-corrected parasitological cure rate, defined as the proportion of patients without asexual parasitaemia within 7 days after beginning treatment, without recrudescence within 28 days after beginning treatment and who demonstrated no need for rescue treatment for signs of clinical malaria within 28 days after initiation of study treatment. Secondary end-points included PCR-corrected parasitological cure rate on day 14, gametocyte carriage, fever and parasite clearance rate. The intent to treat (ITT) population defined as all randomized patients who took at least one dose of study medication was used for safety analysis. Proportions were compared between treatment groups using the chi-square test. Normally distributed variables were compared using the Student's Two hundred and sixty seven of the 1327 children screened for malaria were enrolled into the trial; 134 in the AL suspension arm and 133 in the AL tablets arm. Seventeen (6.4%) children withdrew or were lost to follow up. Three (1.1%) children, all from the AL suspension arm, were excluded from analysis due to age >59 months, low weight for age and treatment with amodiaquine within three days of enrolment. Two children in the AL suspension arm were withdrawn from treatment due to repeated vomiting on Day 0. Data from 245 children was analysed; 124 in the AL tablets arm and 121 in the AL suspension arm  early treatment failures. One child in the AL tablets arm developed severe anaemia (Hb < 5.0 g/dl) on Day 1 and was withdrawn from treatment while another child in the AL suspension arm developed convulsions on Day 0 and received treatment for severe malaria. Both children recovered after treatment. The PCR corrected cure rate for day 14 was 100% in the AL tablets arm and 98.4% in the AL suspension arm while the day 28 cure rates were 96% (95% 95% CI 90.8 \u2013 98.7) in the AL tablets arm and 93% (95% CI 87.4 \u2013 97.1) in the AL suspension arm p = 0.40 (Table The proportions of children with fever (temperature \u2265 37.5\u00b0C), and parasitaemia over the first four days, and anaemia (haemoglobin < 10 g/dl) over the 28 day follow-up period were similar in both treatment groups Figures . FourteeThere was no difference in the incidence of vomiting between the treatment groups as a proportion of treatment doses administered over the first 3 days of treatment: AL tablets arm, 21/807 (2.6%) and AL suspension arm, 12/406 (3.0%), p = 0.72. Two patients (one in each arm) had diarrhoea, which was suspected to be due to the drugs.Seven Serious Adverse Events (SAE) were recorded during the trial, four in the AL tablets arm and three in the AL suspension arm, none of which was considered related to the study drugs. Three (2.4%) children in the AL tablets arm developed severe anaemia (haemoglobin<5.0 g/dl) during the study. One child developed severe anaemia on Day 1 and was withdrawn from the intervention, while the other two had severe anaemia detected on Day 7 during a routine follow-up visit. All three children with severe anaemia were enrolled with haemoglobin between 5\u20137 g/dl and were treated with iron supplementation only. The fourth child (0.8%) developed severe malaria on day 26 during follow-up. In the AL suspension arm, one child developed severe malaria and meningitis on Day 0 and was withdrawn from treatment, he subsequently developed neurological sequelae. The other children each developed severe malaria (day 22) and severe pneumonia (day 28) and recovered completely. Other adverse events are summarized in Table The 28 day PCR corrected efficacy of AL suspension was 93.4% compared with 96.0% for AL tablets (p = 0.40). Previous trials reported efficacy rates between 98 \u2013 100% for AL suspension in various parts of Africa -17. The The process of changing national treatment policies for malaria is complex and the costs involved are substantial ,25,26. CThe major motivation to evaluate this suspension was the limitations of administering hard tablets to children especially the very young, by mothers and health workers. These difficulties notwithstanding, AL tablets are efficacious and effective in this age-group. Although not widely available, an AL dispersible tablet for paediatric patients is currently under review by regulatory authorities. AL suspension and probably the AL paediatric dispersible tablet will be useful alternatives and a milestone in the treatment of uncomplicated malaria in young children who are unable to take AL tablets. Further studies are needed to evaluate the effect of widening the weight-based dosing ranges of AL suspension for ease of dispensing and administration. This study did not focus on the safety by body weight and future studies should investigate this aspect , malaria (11.2%) and skin infections (8.6%), none attributed to the study medications.This trial had several limitations. The open label design could bias the results because the parents of the children were aware of their children's treatment assignment. Children were not followed up to 42 days as recommended by WHO guidelines . However\u00ae) was not superior to 6-dose artemether-lumefantrine tablets (Coartem\u00ae) for the treatment of uncomplicated malaria in children below 5 years of age in western Kenya.The once daily 3-dose of artemether-lumefantrine suspension  of Dafra Pharma NV, Belgium in 42 + 6 healthy adult human subjects.Short report by the sponsor Dafra Pharma nv/sa. A relative bioavailability study for Fixed Dose Combination (FDC) comparing Coartem Tablets (containing Artemether 20 mg and Lumefantrine 120 mg) of Novartis Pharmaceuticals Limited, EU with Co-ArtesianeClick here for fileDiscussion Dosing Al. Following the WHO guidelines for the treatment of malaria  the recommended dose for artemether/lumefantrine tablets (Coartem\u00ae) when used for children between 5 and 14 kg is 1 tablet at time 0 h and 1 tablet at time 8 h followed by two tablets a day for two days . Calculated as mg artemether per kg body weight (bw), this means that for a child of 5 kg a dose of 8 mg artemether/kg per day spread over two doses is given. Evidence that this high dose of more than 4 mg/kg bw of artemether is needed for small babies (\u00b1 5 kg) is scanty or non existent. Moreover, this \"overdosing\" situation is in contrast with the adult dose of 8 pills a day divided over two intakes. This means that an adult of 50 kg takes a dose of 3.2 mg/kg of artemether and an adult of 75 kg takes one of 2.1 mg/kg per day. There is no dose finding study published that we could find to justify this variation in dosing.Click here for file"}
+{"text": "Its two components have different modes of action that provide synergistic anti-malarial activity. It is indicated for the treatment of infants, children and adults with acute, uncomplicated infection due to Plasmodium falciparum or mixed infections including P. falciparum. A formulation with improved palatability has been developed especially for children (Coartem\u00ae Dispersible), which rapidly disperses in a small amount of water for ease of administration.Current World Health Organization (WHO) guidelines for the treatment of uncomplicated falciparum malaria recommend the use of artemisinin-based combination therapy (ACT). Artemether/lumefantrine is an ACT prequalified by the WHO for efficacy, safety and quality, approved by Swissmedic in December 2008 and recently approved by the USA FDA. CoartemThe efficacy of the six-dose regimen of artemether/lumefantrine has been confirmed in many different patient populations around the world, consistently achieving 28-day PCR (polymerase chain reaction)-corrected cure rates of >95% in the evaluable population, rapidly clearing parasitaemia and fever, and demonstrating a significant gametocidal effect, even in areas of widespread parasite resistance to other antimalarials. Plasmodium falciparum resistance to monotherapies prompted the World Health Organization (WHO) 2006 guidelines for the treatment of malaria to recommend that combinations of antimalarials be used to treat malaria caused by P. falciparum . Al. Al10]. Since the efficacy was similar between the 96 and 60-hour six-dose regimens, the 60-hour regimen was chosen for further development to facilitate better compliance with a shorter treatment duration.These two randomized, open-label studies were both conducted in Thailand and included mefloquine plus artesunate (MAS) as an active control arm ,12.Study 026 enrolled patients \u2265 2 years of age , and StuIn Study 026, 150 patients received AL and 50 patients received mefloquine and artesunate. Patient characteristics were comparable between treatment groups, and treatment comparisons were not affected by a difference in baseline parasite density. The 28-day PCR-corrected cure rates were >95% in the evaluable population for both treatment groups  in sub-Saharan Africa  . In the The overall 28-day cure rate, corrected for reinfection, was 93.9% for the intent-to-treat population . The medTo facilitate administration of AL to children, a sweet-tasting dispersible formulation has been developed. The primary objective of Study B2303, which enrolled 899 African children (5-<35 kg body weight), was to demonstrate non-inferiority of the dispersible tablet to the crushed tablet based on the 28-day PCR-corrected cure rate in the evaluable population. The median age of children enrolled was 36 months, ranging from two months to 12 years, and the median body weight was 13 kg .\u00ae Dispersible) . Th. Th15]. A total of 599 adult patients (>16 years) and 877 children (<16 years) were treated with the six-dose AL regimen in the above studies. The 28-day PCR-corrected cure rate in evaluable patients consistently met WHO criteria for efficacy (>95%) in both adults and children. Rapid clearance of parasitaemia and fever was also evident, although noticeably quicker in children.A large number of independent studies have compared the efficacy of AL with other anti-malarial treatments. These studies conclusively show that AL is highly efficacious and at least as effective as other ACTs and combinations of antimalarials over a wide range of geographical areas and in a variety of populations. A summary of these trials is presented in Tables In studies reported from Africa, summarized in Table Several of the publications report the potent effects of ACT treatment on gametocyte carriage. One study investigated gametocyte carriage in detail and it was found that in AL-treated patients, not only was the rate of carriage of gametocytes significantly reduced as compared with that in patients receiving CQSP, but the carriage time of gametocytes and the infectivity of the gametocytes for mosquitoes were also significantly decreased . These fet al [Piola et al  conducte28-day cure rates were very similar . The only difference between supervised and unsupervised treatment was that blood lumefantrine levels in the unsupervised patients were statistically significantly lower (p < 0.001) at Days 3 and 7.P. falciparum malaria, with high parasitological cure rates and rapid clearance of parasitaemia and resolution of fever. In one study in which P. vivax infections were included, the comparator ACT (DP) was significantly more effective in terms of reducing the risk of recurrent P. vivax infection, probably due to the long half-life of piperaquine offering protection against re-infection for longer than lumefantrine: in the same study, PCR-corrected P. falciparum parasitological cure rates were almost identical for AL and DP [The studies reported from South-East Asia, summarized in Table L and DP .Studies from South Asia show high parasitological cure rates and rapid clearance of fever and parasitaemia with the six-dose AL regimen, comparable with those for MAS in the one study where this combination was used as a comparator. AL was more effective than non-ACT comparators in these studies.A meta-analysis of 32 published randomized studies performed predominantly in Africa but also in South America and Asia has evaluated the efficacy of a number of different forms of ACT using a Bayesian random effects approach. The analysis showed that AL was one of the most effective ACT with a 28-day PCR-corrected parasitological cure rate of 97.4%  in both adults and children in randomized, controlled trials in malaria-endemic regions -15. EffiP. falciparum malaria [Effectiveness studies comparing supervised and unsupervised treatment  also dem malaria .A large number of independent studies show the six-dose AL regimen is at least as effective as other artemisinin-based combination therapies and combinations of anti-malarials. A meta-analysis of 32 comparative trials showed AL to be one of the most effective ACTs currently available .in vivo has been reported in Africa [The combination of artemether and lumefantrine also has good gametocidal properties, which may help to limit the spread of resistance ,12. To dThe authors would like to acknowledge that Novartis Pharma AG sponsored this supplement. However, none of the authors works for, or represents in any way, Novartis Pharma AG.All authors met International Committee of Medical Journal Editors criteria for authorship."}
+{"text": "Plasmodium falciparum malaria in pregnancy. Evidence on the safety and efficacy of ACTs in pregnancy is needed as these drugs are being used increasingly throughout the malaria-affected world. The objective of this study was to compare the efficacy, tolerability, and safety of artemether-lumefantrine, the most widely used fixed ACT, with 7 d artesunate monotherapy in the second and third trimesters of pregnancy.To date no comparative trials have been done, to our knowledge, of fixed-dose artemisinin combination therapies (ACTs) for the treatment of P. falciparum malaria in the second and third trimesters of pregnancy. The primary endpoint was efficacy defined as the P. falciparum PCR-adjusted cure rates assessed at delivery or by day 42 if this occurred later than delivery, as estimated by Kaplan-Meier survival analysis. Infants were assessed at birth and followed until 1 y of life. Blood sampling was performed to characterise the pharmacokinetics of lumefantrine in pregnancy. Both regimens were very well tolerated. The cure rates  for the intention to treat (ITT) population were: AS7 89.2% (82.3%\u201396.1%) and AL 82.0% (74.8%\u201389.3%), p = 0.054 (ITT); and AS7 89.7% (82.6%\u201396.8%) and AL 81.2% (73.6%\u201388.8%), p = 0.031 (per-protocol population). One-third of the PCR-confirmed recrudescent cases occurred after 42 d of follow-up. Birth outcomes and infant (up to age 1 y) outcomes did not differ significantly between the two groups. The pharmacokinetic study indicated that low concentrations of artemether and lumefantrine were the main contributors to the poor efficacy of AL.An open-label randomised controlled trial comparing directly observed treatment with artemether-lumefantrine 3 d (AL) or artesunate monotherapy 7 d (AS7) was conducted in Karen women in the border area of northwestern Thailand who had uncomplicated The current standard six-dose artemether-lumefantrine regimen was well tolerated and safe in pregnant Karen women with uncomplicated falciparum malaria, but efficacy was inferior to 7 d artesunate monotherapy and was unsatisfactory for general deployment in this geographic area. Reduced efficacy probably results from low drug concentrations in later pregnancy. A longer or more frequent AL dose regimen may be needed to treat pregnant women effectively and should now be evaluated. Parasitological endpoints in clinical trials of any antimalarial drug treatment in pregnancy should be extended to delivery or day 42 if it comes later.Trial Registration: Current Controlled Trials ISRCTN86353884 Rose McGready and colleagues show that an artemether-lumefantrine regimen is well tolerated and safe in pregnant Karen women with uncomplicated falciparum malaria, but efficacy is inferior to artesunate, probably because of low drug concentrations in later pregnancy. Plasmodium falciparum, a mosquito-borne parasite that causes malaria, kills nearly one million people every year. Although most deaths occur among young children, malaria during pregnancy is also an important public-health problem. In areas where malaria transmission is high (stable transmission), women acquire a degree of immunity. Although less symptomatic than women who lack natural protection, their babies are often small and sickly because malaria-related anemia (lack of red blood cells) and parasites in the placenta limit the nutrients supplied to the baby before birth. By contrast, in areas where malaria transmission is low (unstable transmission or sporadic outbreaks), women have little immunity to P. falciparum. If these women become infected during pregnancy, \u201cuncomplicated\u201d malaria  can rapidly progress to \u201csevere\u201d malaria , which can be fatal to the mother and/or her unborn child unless prompt and effective treatment is given.P. falciparum becoming resistant to either drug. The most widely used fixed-dose ACT is artemether\u2013lumefantrine (AL) but, although several trials have examined the safety and efficacy of this treatment in non-pregnant women, little is known about how well it works in pregnant women. In this study, the researchers compare the efficacy, tolerability, and safety of AL with a 7-d course of artesunate monotherapy  in the treatment of uncomplicated malaria in pregnancy in northwest Thailand, an area with unstable but highly drug resistant malaria transmission.Malaria parasites are now resistant to many of the older antimalarial drugs . So, since 2006, the World Health Organization (WHO) has recommended that uncomplicated malaria during the second and third trimester of pregnancy is treated with short course (3 d) fixed-dose artemisinin combination therapy . Artemisinin derivatives are fast-acting antimalarial agents that are used in combination with another antimalarial drug to reduce the chances of The researchers enrolled 253 women with uncomplicated malaria during the second and third trimesters of pregnancy into their open-label trial . Half the women received each type of treatment. The trial's main outcome was the \u201cPCR-adjusted cure rate\u201d at delivery or 42 d after treatment if this occurred after delivery. This cure rate was assessed by examining blood smears for parasites and then using a technique called PCR to determine which cases of malaria were new infections  and which were recurrences of an old infection (classified as treatment failures). The PCR-adjusted cure rates were 89.7% and 81.2% for AS7 and AL, respectively. Both treatments were well tolerated, few side effects were seen with either treatment, and infant health and development at birth and up to 1 y old were similar with both regimens. Finally, an analysis of blood samples taken 7 d after treatment with AL showed that blood levels of lumefantrine were below those previously associated with treatment failure in about a third of the women tested.Although these findings indicate that the AL regimen is a well tolerated and safe treatment for uncomplicated malaria in pregnant women living in northwest Thailand, the efficacy of this treatment was lower than that of artesunate monotherapy. In fact, neither treatment reached the 90% cure rate recommended by WHO for ACTs and it is likely that cure rates in a more realistic situation  would be even lower. The findings also suggest that the reduced efficacy of the AL regimen in pregnant women compared to the efficacy previously seen in non-pregnant women may be caused by lower drug blood levels during pregnancy. Thus, a higher-dose AL regimen  may be needed to successfully treat uncomplicated malaria during pregnancy.http://dx.doi.org/10.1371/journal.pmed.0050253.Please access these Web sites via the online version of this summary at MedlinePlus encyclopedia contains a page on malaria (in English and Spanish)The malaria , and their 2006 Guidelines for the Treatment of Malaria includes specific recommendations for the treatment of pregnant womenInformation is available from the World Health Organization on malaria and on malaria during pregnancy (in English and Spanish)The US Centers for Disease Control and Prevention provide information on Roll Back Malaria Partnership on malaria during pregnancy, on artemisinin-based combination therapies, and on malaria in ThailandInformation is available from the  Plasmodium falciparum infections in pregnancy account for a high proportion of maternal mortality in malaria-endemic countries ) than in the AL (11.1% [13/117]) group delivered before day 42 of the follow-up, p = 0.20 and 2 (1\u20135) d, for both groups, p = 0.95, respectively.Patients in the AS7 and AL groups responded rapidly to treatment and had similar median (range) fever and parasite clearance times: 1 (1\u20132) d versus 2 (1\u20133) d, P. falciparum, of which 30 (41%)  were recrudescent infections, 44 (59%)  were novel infections (Before delivery (or day 42 if later), there were 74 episodes of first reappearance of fections  and nonefections ; Table 4p = 1.000 for both comparisons. The censored patients included two with late parasite reappearance: one (AS7 group) PCR-confirmed recrudescent infection on day 51 and one (AL group) PCR-confirmed new infection on day 140. PP PCR-adjusted cure rates by delivery (or day 42 if later) were again higher for AS7 than AL ,  than AL  and like than AL .p = 0.015. There was no significant difference in time to recrudescence between the AS7 and AL groups 32 (19\u201398) d and 23 (14\u201363) d respectively, p = 0.14 (The median (range) interval to recrudescent infection, 27 (14\u201398) d, was shorter than the time to reinfection, 37 (15\u2013147) d, p = 0.14 . Nearly p = 0.14 .n = 142) had recurrent infections at enrolment: of whom most had one previous infection (n = 127), 14 had two, and one had three previous episodes of P. falciparum within the same pregnancy but before entry to the study . Patients with unknown results were omitted from this subgroup analysis. The PCR-adjusted cure rate for the patients who entered the study with a recrudescent infection was significantly higher for AS7 compared to AL  of the patients who entered the trial had a primary infection. The remaining patients  and 29.6% (37/125) of patients treated with AS7 and AL, respectively (p = 0.23), at a median (range) of 29 (14\u2013125) d and 34 (14\u2013105) d following treatment (p = 0.47). After patients with P. vivax infection during follow up were censored, the PCR-adjusted cure rates for P. falciparum infection at delivery (or day 42 if later) using survival analysis were improved slightly: AS7 92.2%  versus AL 83.8% (95% CI 76.1%\u201391.5%), p = 0.045  appeared during follow-up to delivery (or day 42 if later) was considered a \u201cfailure\u201d resulted in low infection-free rates of AS7 31.9% (95% CI 20.1%\u201343.7%) and AL 38.1% (95% CI 28.1%\u201348.1%), p = 0.795, which illustrates the limited post-treatment prophylactic effects of the two regimens. By day 50, 47.8% (121/253) of patients had a reappearance of P. falciparum or P. vivax  between the AS7 (3.1% [4/128]) and AL (2.4% [3/125]) groups in the proportion of patients treated with other antimalarials before censoring. The trends in cure rates using survival analysis were unchanged when these patients were censored on the day of receiving these drugs (unpublished data).There were 11 patients treated with drugs with antimalarial activity ; in four of these cases the treatment occurred after the day of censoring for the primary study endpoint. There was no significant difference (p = 0.037), but there was no significant difference by day 14 (p = 0.058) through to day 42 (p = 0.621). Nine patients (3.6%) were transfused (six AS7 group and three AL group) for malaria-related anaemia.Nearly half the patients were anaemic at enrolment, with similar proportions in the two groups . The absp = 0.13.There was no significant difference in the proportion of patients with gametocytaemia on admission . There wP. falciparum infections (n = 14) was 387 (164\u2013551) ng/ml and 379  ng/ml in those with recurrent novel infections (n = 17), and 423  ng/ml in patients who had no parasite reappearance (n = 54), p = 0.26  lumefantrine concentrations in patients who had subsequent recrudescent p = 0.26 .The lumefantrine concentration (LC) is known to be higher in capillary plasma (CP) than venous plasma (VP). This relationship has been described previously by the following equation derived from data obtained at this study site :In total 35% 30/85) of patients had day 7 capillary plasma concentrations < 355 ng/ml , a threshold previously associated at this site with increased treatment failure rates in non-pregnant patients  to day 1: 5.4% IRBC). This patient was monitored with blood smears every 6 h for 24 h and, as the parasitaemia then reduced, she did not require rescue treatment, although she went on to have a PCR-confirmed recrudescence on day 22. For the remaining 140 SAEs (64 AS7 and 76 AL), the relationship to the treatment drug was rated as \u201cpossible\u201d to \u201cnot related\u201d. The most common SAE in this study was hospitalization (as required by protocol) to supervise the treatment of a reappearance of One patient experienced a mild allergy  after the second dose of artesunate which responded to oral chlorpheniramine and stopping artesunate. Early vomiting occurred in 1.2% (3/253) of patients, two AL and one AS7, all with the first dose and for each the repeat dose was tolerated.p < 0.05). There was no significant difference between the groups' haematological and biochemical parameters at baseline. In the individual paired analysis there was no significant difference in the proportion of patients in each group who developed abnormal values on day 14 when they were not present on enrolment.The patients who had blood counts and biochemistry evaluated had a history of less fever and less symptoms on admission, than the patients who were not studied (There was no difference in the baseline characteristics of patients who volunteered for ECG examination and those who did not have these performed. There were 51 patients treated with AL who had paired ECG examination (prior to treatment and 1 h after the last dose) with no clinically relevant abnormalities.p = 0.006 (Fisher's exact). Tinnitus in these seven patients was short-lived (< 1 wk) and resolved in all cases.The only symptom that differed significantly between the groups in follow-up, when it was not present on enrolment, was tinnitus, which was more frequent in the AS7 group 8.5% (7/82) compared to 0% (0/86) in AL group, Data on birth outcome were documented for 96.8% (244/252) of women and not available for 3.2% (8/252) who were lost to follow-up before the end of pregnancy. Amongst the women with a known outcome there were 99.6% (243/244) births and 0.4% (1/244) abortions. There were 239 singleton births including two stillbirths and four twin births .Escherichia coli urinary tract infection, and the other (AL group) possibly related to consanguinity (marriage to a second cousin).There were no significant differences in the major birth indicators between the groups including birth weight, gestation, congenital abnormalities, and the newborn neurological optimality score . The aboP. falciparum in peripheral and placental specimens was very high , one on day 2 of treatment. All infants were peripheral blood negative (microscopy and PCR) at birth. Two of the cord blood\u2013positive infants remained negative by weekly peripheral smear by day 28. The remaining infant, who became ill on day 20 of life, was P. falciparum positive by microscopy and PCR. Parasite genotyping of the infant blood matched the mother, cord, and placental DNA of the infection at the time of delivery.Malaria smear and blood spots for PCR genotyping on maternal peripheral blood, cord blood, placental blood, and the baby's peripheral blood were reported for 70% (169/243) of birth outcomes (including twins and excluding the abortion case). Concordance between microscopy and PCR detection of ery high . All theP. vivax parasitaemia at delivery, and one of these had P. vivax positive placental tissue by microscopy. No cord or baby malaria smears were positive for P. vivax by microscopy.Three women had peripheral blood p = 0.32. There was no significant difference in the median months of follow-up (unpublished data). A total of nine infants died among the 237 infants followed to 1 y, an overall infant mortality rate of 38 per 1,000 live-born singletons. Significantly more infant deaths occurred in the AS7, 6.7% (8/120) than AL 0.9% (1/117) group, p = 0.036. Five infant deaths occurred in the first month of life (day 0\u201332), three of which were related to prematurity , while the other two were from pneumonia in the first 2 wk of life in term infants. Of the remaining infants one was premature (33.1 wk) and died of sepsis at 3 mo. The mother was diagnosed with TB and HIV postpartum. One child died from diarrhoea at 4 mo, one from suspected intraventricular haemorrhage on a background of Allagile's syndrome at 7.5 mo (AL group), and one from suspected beri-beri at 10.5 mo. There was no difference between the infants in the groups in growth parameters or developmental parameters assessed by the Shoklo Developmental Scores at 3, 6, 9 (unpublished data), and 12 mo (All four sets of twins (eight babies) survived to 1 y and were developing normally. Of the singleton infants, 82.2% (195/237) were followed up for \u2265 9 mo. The main reason for loss to follow-up was leaving the study area. Overall 77.5% (93/120) of AS7 and 72.6% (85/117) of AL infants were evaluated at 12 mo, nd 12 mo .p = 0.61. The median (range) age of first P. falciparum was 29.9 (2.9\u201351) wk and 30.1 (3.4\u201344.3) wk for the first P. vivax infection. There was also no significant difference in the species, the median days to the first episode, or the total number of episodes in the first year of life (unpublished data), according to the mother's treatment group. The relative risk for recurrent malaria in the first year of life was significantly higher with P. vivax than P. falciparum: 2.9 (95% CI 1.2\u20137.0); 67.7% (21/31) versus 23.5% (4/17), p = 0.008.Of 668 morbid episodes treated in 196 infants in the first year of life, acute respiratory infection 51.9% (347), skin infection 18.3% (122), and malaria 13.0% (87 including one twin) were the most common diagnoses. Malaria affected approximately one-fifth of infants in the first year of life: AS7 16.1% (19/118) and AL 19.0% (22/116), P. falciparum infection only, as there had been no previous experience to establish the safety of AL in pregnancy. This trial was not originally designed to compare cure rates in recrudescent with those in new or primary infections , and parasitaemia are the two main determinants of the therapeutic response in studies conducted in non-pregnant patients . Thus paThe need in malaria-endemic areas for a safe and effective drug with an adequate post-treatment prophylactic effect in pregnancy is clearly demonstrated by the fact that half of the patients in this study had recurrent malaria infection within 50 d. All women who had positive placental blood at delivery had parasite reappearance after randomization.P. falciparum parasites sequester in the placenta and so the parasite burden is underestimated, antiparasitic immunity is compromised, and antimalarial drug levels are generally lower. This study shows that guidelines for defining antimalarial efficacy [Several factors contribute to the poorer treatment responses during pregnancy; efficacy ,41 in noefficacy ,28 and mefficacy . The durP. falciparum PCR genotyping of maternal peripheral and placental blood at delivery did not detect any submicroscopic infections. All cases of placentas that were positive by PCR were found in mothers with a positive peripheral blood smear or very recently treated positive peripheral blood smear. Early detection and treatment of malaria by active weekly screening of maternal peripheral blood smears appears to be a very reliable method to detect P. falciparum malaria in pregnancy in this area of low transmission.In contrast to high-transmission settings, microscopy of the placental smear did not add to the diagnostic yield. There were few adverse events in this study and these were mainly unrelated to treatment. Anaemia is a significant adverse effect of malaria in pregnancy . ArtemisP. falciparum infections before and during this study. This provides further reassuring safety data on the use of artemisinins in the second and third trimesters of pregnancy; however, further large-scale assessments are needed to confirm this safety profile.No pregnant women or newborns in this study died from malaria. The newborn anthropometric results and neurological scores  were verOne of the limitations of this trial includes the lack of blinding to antimalarial treatment administration. A placebo for this trial would have been difficult as the two regimens were very different. The embedded pharmacokinetic sampling would have been impossible without unblinding the trial or sampling women from both groups to maintain blinding, which would be unethical. The ad-hoc analysis shows differences within the strata, rather than simply between groups, which suggests the lack of blinding did not influence the efficacy results.P. falciparum is relatively resistant and transmission intensity (and therefore immunity) is low, the cure rates with AL in pregnancy are unsatisfactory. A dosage increase is needed. A major drawback is that lumefantrine absorption has been shown to be dose-limited [P. falciparum malaria in pregnant women has yet to be found.Immunity contributes significantly to treatment outcomes. It is probable that treatment responses to AL would be better in higher-transmission settings. But in this setting where -limited , so simp-limited . A 5 d rOur results do not indicate that the conventional regimen of AL cannot be used to treat pregnant women with uncomplicated falciparum malaria in other endemic regions. However, they show that in an area of multidrug resistance, the lower lumefantrine plasma levels found in pregnant women will result in a lower parasitological efficacy. In areas of higher endemicity where multigravid pregnant women have some protection from acquired premunition, the conventional regimen of AL, i.e., two doses per day for 3 d, may be more effective, and studies (including pharmacokinetics) are in progress at the time of writing.There are no data on a direct comparison of the pharmacokinetics of lumefantrine in pregnant and non-pregnant women. The only comparable pharmacokinetic data from the same population and the same pharmacology laboratory are from adults , and these were used in the present discussion. The results of the population kinetic study of lumefantrine in the pregnant women of this study will be presented elsewhere. Preliminary analysis confirms the findings of the initial pharmacokinetic study  and showThe current standard six-dose artemether-lumefantrine regimen was well tolerated and safe in pregnant Karen women with uncomplicated falciparum malaria, but efficacy was inferior to 7-d artesunate monotherapy and was unsatisfactory for general deployment in this area. Reduced efficacy probably results from lowered drug concentrations in later pregnancy in the presence of relatively resistant parasites. A higher-dose regimen may be needed to treat pregnant women effectively and should now be evaluated. Longer follow-up is needed when assessing antimalarial drugs in pregnancy as recrudescences may occur months after administration of rapidly eliminated drugs.Text S1(115 KB PDF)Click here for additional data file.Text S2Artemether-lumefantrine treatment of multidrug-resistant falciparum malaria in pregnancy: a pilot study.(198 KB DOC)Click here for additional data file."}
+{"text": "Analytical approaches for the interpretation of anti-malarial clinical trials vary considerably. The aim of this study was to quantify the magnitude of the differences between efficacy estimates derived from these approaches and identify the factors underlying these differences.Data from studies conducted in Africa and Thailand were compiled and the risk estimates of treatment failure, adjusted and unadjusted by genotyping, were derived by three methods (intention to treat (ITT), modified intention to treat (mITT) and per protocol (PP)) and then compared.29 clinical trials (15 from Africa and 14 from Thailand) with a total of 65 treatment arms (38 from Africa and 27 from Thailand) were included in the analysis. Of the 15,409 patients enrolled, 2,637 (17.1%) had incomplete follow up for the unadjusted analysis and 4,489 (33.4%) for the adjusted analysis. Estimates of treatment failure were consistently higher when derived from the ITT or PP analyses compared to the mITT approach. In the unadjusted analyses the median difference between the ITT and mITT estimates was greater in Thai studies (11.4% [range 2.1\u201331.8]) compared to African Studies (1.8% [range 0\u201311.7]). In the adjusted analyses the median difference between PP and mITT estimates was 1.7%, but ranged from 0 to 30.9%. The discrepancy between estimates was correlated significantly with the proportion of patients with incomplete follow-up; p < 0.0001. The proportion of studies with a major difference (> 5%) between adjusted PP and mITT was 28% (16/57), with the risk difference greater in African (37% 14/38) compared to Thai studies (11% 2/19). In the African studies, a major difference in the adjusted estimates was significantly more likely in studies in high transmission sites (62% 8/13) compared to studies in moderate transmission sites (24% 6/25); p = 0.035.Estimates of anti-malarial clinical efficacy vary significantly depending on the analytical methodology from which they are derived. In order to monitor temporal and spatial trends in anti-malarial efficacy, standardized analytical tools need to be applied in a transparent and systematic manner. Plasmodium falciparum and to the introduction of new treatment regimens such as artemisinin combination therapy (ACT). In addition, study designs have evolved to include a longer duration of follow-up and the inclusion of genotyping to distinguish recrudescence from new infection ) and moderate transmission sites (median 4.5% [range 2.2\u201312.8]).In Thailand, studies with a longer duration (42 days or more) had a greater proportion of patients with incomplete follow-up compared to studies with only 28-days of follow-up. This was apparent for both the unadjusted  and adjusted analyses . In Africa, where there was variation in transmission intensity, incomplete follow-up in the adjusted analyses was significantly higher in areas of high transmission (median 47% [range: 32.2\u201368.6]) compared to studies in moderate transmission areas . Patients with new s = 0.721, p < 0.0001), although this does not reach significance in the Thai studies ; Figure The unadjusted risk of treatment failure derived by ITT analysis was consistently higher than that derived by mITT analysis (median difference = 4.7% -0.3\u201331.8%]) (Table .8%)  and was correlated significantly with the proportion of patients with incomplete follow-up  of the treatment arms had a difference in the unadjusted risk estimates (PP-mITT) of greater than 5%; these studies were all from Thailand (19% 5/27), with none (0/38) conducted in Africa; p = 0.01. In the adjusted analyses the proportion with a major difference (> 5%), rose to 28% (16/57), with the risk greater in African (37% 14/38) compared to Thai studies (11% 2/19). In Africa the likelihood of a major difference in adjusted estimates was significantly greater in studies conducted in high transmission sites (62% 8/13) compared to moderate transmission sites (24% 6/25); p = 0.035.Anti-malarial drug clinical trials are conducted both to monitor anti-malarial drug resistance and to compare treatment regimens. As in all clinical trials, protocol violations and incomplete patient follow-up challenge the analysis and interpretation of the results. Malaria studies are, by their nature, logistically difficult, often being conducted in poorly resourced communities and prone to varying patient adherence to protocols. In addition to problems related to protocol adherence, anti-malarial clinical trials are also confounded by interrupted follow-up resulting from recurrent infections, either by the same or different malaria species. The statistical approach to deal with these challenges can vary according to the rationale of the study ,12. For P. falciparum infections or relapse of P. vivax infection, generally require retreatment and termination of the primary study. Even in study populations with the highest adherence rates, these proportions can often exceed a third of all patients enrolled  was generally low, this rose to as high as 36%. Interrupted follow-up was greater in the Thai studies compared to those conducted in Africa, in part because of the longer duration of study follow-up in Thailand. The occurrence of new The proportion of patients with incomplete follow-up has significant implications for the derived estimates of treatment efficacy. Both the ITT and PP methods consistently over-estimated the risk of failure when compared to the preferred mITT method, the discrepancy in risk estimates varying from trivial to highly significant. For example, in the comparison of the unadjusted ITT and mITT failure estimates, 46% (30/65) of the difference in estimates exceeded 5%, with one study having a difference of 31.8%. The bias was most pronounced in Thailand due to the high percentage of patients with incomplete follow-up. These findings highlight that although the ITT method of analysis has utility for conservatively comparing treatment arms within a comparative drug trial, it is significantly biased when deriving point estimates of efficacy, for comparison over time or geographical location.P. falciparum constituted an additional confounding factor for the adjusted analyses (PP-mITT). Whereas individuals with such infections are removed from the PP analysis, they are censored in the mITT analysis after contributing a period of observation to the cumulative risk during which treatment failure was not observed. As a consequence, the PP analysis consistently overestimates treatment failure compared to that derived by the mITT survival analysis (median = 1.7% [IQR 0.5\u20135.6]). In 28% (16/57) of cases this difference exceeded an absolute value of 5%. The discrepancy was particularly apparent in the high transmission sites in Africa where reinfections were highest. The differences in risk estimates were lower for the unadjusted analyses, although in Thailand, high relapse rates with P. vivax and greater loss to follow-up resulted in 18.5% (5/27) of PP estimates deviating by more than 5% from the mITT estimate.New infection with P. vivax relapse, or a high incidence of new P. falciparum infections. Furthermore, since the proportion of reinfections and relapses observed in clinical trials is dependent upon the efficacy of the drug and its pharmacokinetic properties, the potential bias introduced by methodologies has implications for the comparative analysis of antimalarials.Survival analysis is being used increasingly to derive estimates of anti-malarial treatment efficacy. Although the ease of calculating the simple proportions in the PP analysis retains its appeal, and these estimates continue to be reported frequently in the literature, caution is needed when generating temporal and geographical trends using different analytical methods. This is particularly true for studies with poorer patient adherence to follow-up, higher incidence of in vivo efficacy. First, survival analysis allows for all available data to contribute to the analysis, thus increasing the precision of the derived estimates. Second, it avoids systematic biases introduced by dropping from the analysis patients who do not complete follow-up (PP) or classifying failures as patients who do not complete follow-up (ITT). Finally, survival analysis allows for data from patients with different follow-up periods to be combined to generate efficacy estimates at different time points, thus enabling direct comparison between studies with different lengths of follow-up [Given the variations in study methods, survival analysis remains the preferred approach for monitoring ollow-up .in vivo efficacy data need to be collated at an individual patient level and standardized analytical tools applied in a transparent and systematic manner [), aims to do precisely that; gather global anti-malarial efficacy data and provide open access to their uniform interpretation.Over the last decade it has become evident that the wider availability of highly effective anti-malarial regimens must be an integral part of any realistic approach to achieving the global elimination of malaria . Currentc manner . The recThe authors declare that they have no competing interests.GD, FN and RNP supervised the clinical studies. WV, GD and RNP analysed the data. All authors contributed to the drafting of the manuscript.Characteristics and treatment outcomes of the clinical trials. Abbreviations: AL = artemether-lumefantrine; AM = artemether; AP: atovaquone-proguanil; AQ = amodiaquine; AS = artesunate; CQ = chlorquine; DP = dihydroartemisinin-piperaquine; MQ = mefloquine; SP = sulfadoxine-pyramethamine; ACPR = adequate clinical and parasitological response; ETF = early treatment failure; LCF = late clinical failure; LPF = late parasitological failureClick here for fileEstimates of the risk of failure according to treatment arm, derived by intention to treat (ITT), modified Intention to Treat (mITT) and per protocol (PP) analysis methods. Abbreviations: AL = artemether-lumefantrine; AM = artemether; AP: atovaquone-proguanil; AQ = amodiaquine; AS = artesunate; CQ = chlorquine; DP = dihydroartemisinin-piperaquine; MQ = mefloquine; SP = sulfadoxine-pyramethamineClick here for file"}
+{"text": "Both artemether and artesunate have been shown to be superior to quinine for the treatment of severe falciparum malaria in Southeast Asian adults, although the magnitude of the superiority has been greater for artesunate than artemether. These two artemisinin derivatives had not been compared in a randomized trial.A randomized double blind trial in 370 adults with severe falciparum malaria; 186 received intramuscular artesunate (2.4 mg/kg immediately followed by 1.2 mg/kg at 12 hours then 24 hours then daily) and 184 received intramuscular artemether (3.6 mg per kilogram immediately followed by 1.8 mg per kilogram daily) was conducted in Viet Nam. Both drugs were given for a minimum of 72 hours.There were 13 deaths in the artesunate group (7 percent) and 24 in the artemether group (13 percent); P = 0.052; relative risk of death in the patients given artesunate, 0.54; . Parasitaemia declined more rapidly in the artesunate group. Both drugs were very well tolerated.Intramuscular artesunate may be superior to intramuscular artemether for the treatment of severe malaria in adults. Over the past 25 years, the artemisinin derivatives have been used for the treatment of severe malaria. They are the most rapidly acting and potent of all the anti-malarial drugs. They can be given once daily and are safer and easier to administer compared to quinine. Artemisinin derivatives are now part of recommended first-line combination treatment for uncomplicated falciparum malaria everywhere in the tropical world . RecentlThis was a single centre double blind comparison of intramuscular artesunate and intramuscular artemether in patients admitted to the Hospital for Tropical Diseases, Ho Chi Minh City with severe falciparum malaria. It was conducted between May 1996 and June 2003. This study was approved by the Ethical and Scientific Committee of the Hospital for Tropical Diseases, Ho Chi Minh City. All study procedures were essentially as reported in previous studies performed in the Hospital for Tropical Diseases in patients with severe malaria .Plasmodium falciparum and had at least one of the following severe complications: cerebral malaria (Glasgow Coma Score was less than 11), renal acute failure (oliguria and serum creatinine > 250 \u03bcmol/L), jaundice  with a parasite count of more than 100,000/\u03bcL or with serum creatinine > 250 \u03bcmol/L, hypoglycaemia (blood glucose < 2.2 mmol/L), anaemia (haematocrit < 20%) with a parasite count of more than 100,000/\u03bcL, hyperparasitaemia , hyperlactataemia (plasma lactate > 4 mmol/L), metabolic acidosis  and shock (systolic blood pressure < 80 mmHg with cool extremities).Patients were included in the study if their peripheral-blood smears had asexual forms of Informed consent was provided by all studied patients (or accompanying relative if patient was unconscious).Patients were not included if they were younger than 14 years, were pregnant in the first trimester, were known intravenous drug abusers, had received more than 3 g of quinine or 2 doses of any artemisinin derivatives in the previous 48 hours before admission, had a past history of allergy to any artemisinin derivatives, or if known to be HIV positive.A full history was recorded from either the patient or their relatives. Patients were examined fully on admission. Baseline blood samples were taken for full blood count, biochemistry tests , blood culture and malaria parasite counts. Arterial pH and blood gases were also measured. Chest X-ray was also performed on admission. Hydration and neurological status were fully assessed. A urinary catheter was inserted if necessary. Lumbar puncture was done if Glasgow Coma Score was below 14. A Gram stain and bacterial culture was performed on all cerebrospinal fluid samples and was analyzed for protein, glucose, lactate levels and cell counts.Patients were treated as per standard recommendations and as previously reported . BrieflyPatients were randomized to treatment with either intramuscular artesunate or intramuscular artemether. Artesunate  was given in a dose of 2.4 mg/kg body weight on admission, then 1.2 mg/kg was given daily. Artemether  was given in a dose of 3.2 mg/kg body weight on admission, followed by 1.6 mg/kg daily. Both drugs were given intramuscularly to the anterior thigh until oral medication could be taken reliably. Oral artesunate was given in a dose of 2 mg/kg/day to complete a total course of seven days , providing a total cumulative dose between 17 and 18 mg/kg over seven days.Treatment failure was considered if parasitaemia did not fall by >75% of the admission value within 48 hours and intravenous quinine (20 mg/kg followed by 10 mg/kg 8 hourly) was given.The randomization was generated from random number tables. Labels with the name of drug for each patient were put in coded sealed opaque envelopes, and the envelopes were randomized in blocks of 20. Once a patient was enrolled in the study the envelope was opened. An independent team of nurses, not otherwise involved in the study or responsible for the care of these patients, open the envelope, randomized the patient and prepared the injection. Neither the treating physicians, study doctors and nurses, or patients knew which anti-malarial drugs was administered.The study was reviewed continuously by an external monitor. Results were entered into a database . Categorical data were analyzed with a statistical software package . Chi- square or Fisher's exact test was used to compare proportions as appropriate. Risk verification was expressed with odds ratio (OR) and 95% confidence interval (CI). The original sample size was calculated on an assumption of a 20% mortality in the artemether arm and a 50% reduction with artesunate. Assuming 80% power and 0.05 significance the study would have needed to recruit 400 patients into each arm . From the year 2000 onwards, the number of patients with severe malaria declined dramatically and a decision was made prior to unblinding to stop the study on the grounds of future futility. At this stage 370 patients had been recruited, 186 in the artesunate arm and 184 in the artemether arm. Assuming a 20% of mortality in the artemether arm the study at the time of stopping therefore had a power of 70% to demonstrate an approximate 50% reduction in absolute mortality.Between May 1996 and June 2003, 370 patients were recruited in the study. There was a gradual rise in number of severe malaria patients admitted to the HTD with peak in 2000, but after this date there was a sharp drop of severe malaria patients and a dramatic decline in the number of malaria patients admitted in the subsequent two years. Because of this sustained fall in the number of patients, a decision was made by the investigators on the grounds of future futility to terminate the study. This was done prior to unblinding of the treatment allocation. The baseline characteristics of patients on admission are shown in Table The overall mortality rate was ten percent (37 of 370 patients). There were 13/186 (7%) deaths in the artesunate (ARTS) treated patients compared with 24/184 (13%) deaths in the artemether (ARTM) treated patients unadjusted relative risk of death for artesunate = 0.54, 95% CI: 0.28 - 1.02, p = 0.052 ; 28 cases had persistent intractable shock; 20 cases had spontaneous bleeding mostly from gastro-intestinal tract; 12 cases had suspected nosocomial co-infection of lung and urinary tract; and eight cases had pulmonary oedema.Assessment of standard markers of recovery from severe malaria did not reveal any significant differences between the treatment groups , range (0-70%) and in the ARTM group: 21.4%, IQR (12.9-34.7), range (0-70%) p = 0.25), or in the number of times required blood transfusions (p = 0.19).After treatment hypoglycaemia was documented in three patients in the artesunate group and 5 in the artemether group. No local abscesses, urticaria or rashes were found in both groups. There were no severe unexpected adverse events.The treatment of severe malaria has changed in recent years with the introduction of the artemisinin derivatives. Until recently, artemether was favoured by World Health Organization and, therefore, most studies had been conducted with this derivative. In an individual patient data meta-analysis of randomized trials in severe falciparum malaria artemether was superior to quinine in Southeast Asian adults, but not in African children . UnfortuThe ARTS group was associated with lower risk  of developing coma after initiation of therapy than the ARTM group  RR = 0.45 CI 95%: 0.19 - 1.05, p = 0.056. The parasite clearance times were faster in the ARTS treated patients (median 90% reduction PCT (IQR) (hrs): 16 (12-30) in ARTS and 24 (12-36) in ARTM). These data may be explained by the pharmacokinetic properties of artesunate compared with artemether and the faster bioavailaibilty of the active metabolites with artesunate especially in severely ill patients with marked metabolic acidosis. It also suggests that the first few hours of effective treatment are very important and the faster the active drug can be got to the site of action (the blood) the better. As outlined above after intramuscular injection concentrations of artesunate peaked within 30 minutes of injection, and artesunate was hydrolysed immediately and completely to the biologically active dihydroartemisinin (DHA). Maximum artemether concentrations occurred at a median of 10 hours and was associated with a much more erratic and slower conversion to DHA. Importantly, in a particular subgroup of patients with severe metabolic acidosis the absorption and conversion to DHA was erratic, limited and delayed. This hypothesis would be consistent with previous pharmacokinetic work undertaken at the Hospital for Tropical Diseases [The trial was overtaken by events. Improved malaria control in Vietnam led to a dramatic decline in the incidence of severe malaria. The Hospital for Tropical Diseases is a referral hospital, which in the early 1990s was receiving approximately three to five patients each day with severe malaria. A decade later this had fallen to approximately one patient per month. This was insufficient to sustain this trial and so the trial was terminated before reaching the pre-defined end points. The trial was stopped with the investigators blind to the allocations and prior to any analysis of the results.The result after 370 patients had been randomized showed a borderline difference in favour of artesunate . The superiority of artesunate over artemether is supported by the larger differences in mortality rates when artesunate was compared with quinine , versus artemether (a 26% reduction), although these trials were conducted in different sites, with different protocols and different drug dosages, so are not strictly comparable [Both drugs were very well tolerated with little or no discomfort at the injection site, no serious local or systemic reactions, a low rate of hypoglycaemia (presumably disease related), and no neurological sequelae. Although artemether is easier to administer than artesunate, which is an advantage in busy epidemic settings, it is probably an inferior drug. Artesunate, administered as an emergency, should be the treatment of choice for severe falciparum malaria.The patients recruited to this trial all had severe falciparum malaria as defined by the World Health Organization. In terms of standard prognostic clinical markers of severity these patients were more severe than previously reported patients recruited into similar studies in Vietnam or compared with those patients entered into regional trials (data not shown) ,4. In thIntramuscular artesunate may be superior to intramuscular artemether for the treatment of severe malaria in adults. Artesunate, administered as an emergency, should be the treatment of choice for severe falciparum malaria.ARTS: artesunate; ARTM: artemether; DHA: dihydroartemisinin.The authors declare that they have no competing interests.NHP, ND, NW, JF and TTH participated in the design of the study. NHP, NTHM, TTHC, LVC, DXS, TTH and JF were involved in recruiting the patients and collecting clinical data. PQT performed the statistical analysis. NHP, ND, NW, JF and TTH were primarily responsible for drafting the manuscript. All authors approved the final version."}
+{"text": "Very few data on anti-malarial efficacy are available from the Democratic Republic of Congo (DRC). DRC changed its anti-malarial treatment policy to amodiaquine (AQ) and artesunate (AS) in 2005.The results of two in vivo efficacy studies, which tested AQ and sulphadoxine-pyrimethamine (SP) monotherapies and AS+SP and AS+AQ combinations in Boende , and AS+SP, AS+AQ and SP in Kabalo (Katanga province), between 2003 and 2004 are presented. The methodology followed the WHO 2003 protocol for assessing the efficacy of anti-malarials in areas of high transmission.Out of 394 included patients in Boende, the failure rates on day 28 after PCR-genotyping adjustment of AS+SP and AS+AQ were estimated as 24.6% [95% CI: 16.6\u201335.5] and 15.1% [95% CI: 8.6\u201325.7], respectively. For the monotherapies, failure rates were 35.9% [95% CI: 27.0\u201346.7] for SP and 18.3% [95% CI: 11.6\u201328.1] for AQ. Out of 207 patients enrolled in Kabalo, the failure rate on day 28 after PCR-genotyping adjustment was 0 [1-sided 95% CI: 5.8] for AS+SP and AS+AQ [1-sided 95% CI: 6.2]. It was 19.6% [95% CI: 11.4\u201332.7] for SP monotherapy.The finding of varying efficacy of the same combinations at two sites in one country highlights one difficulty of implementing a uniform national treatment policy in a large country. The poor efficacy of AS+AQ in Boende should alert the national programme to foci of resistance and emphasizes the need for systems for the prospective monitoring of treatment efficacy at sentinel sites in the country. The Democratic Republic of Congo (DRC) is a vast country covering more than two million square kilometres and with an estimated population of 60 million people. In 2006, the malaria case fatality rate in children below five years of age was 0.61 .in vivo efficacy studies conducted independently in DRC in 2003\u20132004, when the National Malaria Control Programme (NMCP) was in the process of changing treatment policy to an artemisinin combination therapy (ACT) are presented. The NMCP introduced AS+AQ as their first-line regimen in 2005 to replace SP monotherapy for the treatment of uncomplicated malaria. For one study, results on the molecular markers of SP resistance are also presented. A good correlation has been shown between mutations in the dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) genes of Plasmodium falciparum and resistance to pyrimethamine and sulphadoxine, respectively [The results of two The first study was located in Boende, Equatorial province, in the northwest of the country and the second study in Kabalo, Katanga province, in the southeast of DRC Figure . MalariaThe two studies were open-label. The study methodology was based on the WHO protocol for assessing efficacy of anti-malarials in areas of high transmission . In BoenP. falciparum parasitaemia between 2,000 and 200,000/\u03bcl. In Kabalo, due to the unstable security situation and recruitment difficulties, it was decided to extend the age group for inclusion to between six months and 10 years and to accept P. falciparum parasitaemia between 1,000 and 200,000/\u03bcl. Only children living less than half an hour's drive from the study clinic in Kabalo city were enrolled. In both studies, signs of severe malaria or any other serious health condition, intake of antimalarial treatment in the last seven days and mixed malaria infection were exclusion criteria.In Boende, eligibility criteria were age between six and 59 months, axillary temperature \u2265 37.5\u00b0C, living in or within 5 km of Boende city and Sample size was calculated using Epi Info 6 software . For the study in Boende, a sample size of 106 patients per arm was calculated using an estimated failure rate of 15%, a risk \u03b1 of 5%, a precision of 7% and assuming 10% lost to follow-up. In Kabalo, it was 60 patients per arm, using 15% failure rate, 5% risk \u03b1, 10% precision and 15% lost to follow-up. In Boende, monotherapies and combination therapies were evaluated sequentially because of the concern that the study would be terminated early due to security concerns. Therefore, children were initially randomly allocated to SP or AQ, and then to AS+SP or AS+AQ, using a ratio of 1:1 each time. In Kabalo, patients were allocated randomly to AS+AQ, AS+SP or SP using a ratio of 1:1:1. In both studies, randomization was in blocks of 12 and treatment allocations were concealed in sealed opaque envelopes, opened after consent was obtained.\u00ae, Roche, Basel, Switzerland) was given as a single dose of approximately 1.25 mg/kg of pyrimethamine and 25 mg/kg of sulphadoxine; (ii) AQ  was given at a dose of 10 mg/kg/day once daily for three days; and (iii) AS  was given at a dose of 4 mg/kg once daily for three days. All doses were directly observed at the health centre. Patients who vomited within half an hour had the full dose replaced; if vomiting occurred between 30 minutes and one hour after intake, only half the dose was replaced. Study drugs manufactured according to Good Manufacturing Practice standards were purchased for the purpose of the studies. A capillary blood sample was also collected onto Whatman grade 3 filter paper and stored on site for genotypic analysis if needed.After randomization, haemoglobin, asexual parasitaemia and presence of gametocytes were recorded at day 0, and patients received the first dose of treatment: (i) SP  vomiting any dose of study drug twice (at which point intravenous quinine was given), (ii) serious allergic reaction to the study drug, (iii) other serious illness, (iv) intake of any drug with anti-malarial properties during the period of follow-up, (v) withdrawal of consent or (vi) failure to attend scheduled visits. Treatment failures were treated with quinine hydrochloride. The WHO classification of treatment outcomes was used with patients categorised as early treatment failures (ETF), late clinical failures (LCF), late parasitological failures (LPF) on day 28 or adequate clinical and parasitological response (ACPR) . In the In Boende, thick blood films were stained with Giemsa 10% for 10 to 20 minutes and with Giemsa 5% for 25 minutes in Kabalo. Parasite density was determined as the number of parasites per 200 white blood cells (WBC), assuming a total WBC count of 8,000/\u03bcl. If less than 10 parasites were read per 200 WBC, the count was extended to 500 WBC. Microscopists reading the slides were blind to the treatment allocation. In the two studies, there was an independent double reading of all slides and resolution of discordant results (negative/positive/species or parasitaemia discordance above 50%) by a third senior reader. The third reading was taken as the final result. In addition, an external quality control of 10% of slides selected randomly from the screening, inclusion and follow-up slides was performed. From the study in Boende, slides were sent to the Institute of Tropical Medicine (IMT), Antwerp (Belgium). Slides from Kabalo were sent to the Amikuvu Regional Public Health Laboratory of Goma and the National Institute of Biomedical Research of Kinshasa (DRC). Haemoglobin was measured using the Lovibond technique .msp1) and merozoite surface protein-2 (msp2) P. falciparum gene loci [P. falciparum gene loci: msp1, msp2, and glutamate rich protein (glurp) [glurp alleles were first compared between baseline and post-treatment samples. If they were different, the recurrence was attributed to a reinfection. If they were identical, then the msp1 and msp2 alleles were compared between the 2 samples. If there was a common band on one or two msp markers, then the recurrence was considered as recrudescence. If not, it was due to a reinfection.PCR genotyping to distinguish recrudescence from reinfection was performed in two different laboratories. For Boende, the analysis was performed at the ITM of Antwerp, Belgium using merozoite surface protein-1 (ene loci . Infecti (glurp) . The gludhfr) and in codons 437 and 540 of the dihydropteroate synthase (dhps) genes of P. falciparum was performed in the ITM of Antwerp, Belgium using a real mutation-specific nested PCR and/or restriction digestions [dhfr and dhps codon was characterized as 'wild type' (no mutation present), 'mixed' (both wild and mutant genotypes clearly present in the same infection), or 'pure mutant' (only mutant genotypes detected). The dhfr genotypes for each infection were categorized as follows: 'wild type': no mutation detected; 'single': infection involving parasites with a single mutation; 'double': infection involving parasites with a double mutations and 'triple': infection involving parasites with all three mutations detected. The dhps genotypes were defined as wild type; single or double mutations. Infections by parasites with a triple dhfr mutation and double dhps mutation were categorized as 'quintuple' mutation infections. These results have been reported in detail elsewhere [In Boende, additional blood samples were collected from the children treated with SP as part of a multi-centre genotyping study to look for molecular markers of antifolate resistance. Analysis of mutations in codons 51, 59 and 108 of the dihydrofolate reductase . Failure rates were estimated using Kaplan-Meier survival analysis, in which withdrawals, losses to follow-up and reinfections were censored on the last day seen. Patients with missing or indeterminate PCR results were censored on the closest prior visit when the malaria smear result was negative.Data were entered in Microsoft ExcelA total of 1,005 children were screened between September 2003 and February 2004, of whom 394 (39.2%) were enrolled  had positive-negative discordant results during follow-up, 27 (65.7%) of them with very low parasitaemia (<100/\u03bcL). In three cases (1.7%), the discordance affected classification of the patient's outcome: patients with adequate therapeutic response were reclassified as ETF (two cases) and LPF (one case) based on the results of the external quality control.dhps and dhfr mutation, one with pure (1%) and two with mixed mutations, respectively.The results of the molecular markers of antifolate resistance are presented in Table A total of 880 children were screened between June and December 2004, of whom 207 23.5%) were enrolled Figure . Variati.5% were No child developed severe malaria after enrolment and there were no deaths. There was one serious adverse event \u2013 a child diagnosed with acute severe malnutrition during the first week. Of 182 slides sent for external quality control, 26 (14.3%) had positive-negative discordant results during follow-up; none of them affected patients' outcomes.dhfr mutation, which is known to be associated with an increased risk of treatment failure [These two independent studies document anti-malarial efficacy in 2004 from locations in the east and west of DRC, a country from which few such data have been reported previously. Efficacy of SP was poor for both sites. Efficacy of the AS-SP and AS-AQ combinations was poor in Boende but high in Kabalo. The comparison between the results of the two studies is limited by the difference in study methodology. The age and parasitaemia enrolment criteria were different and different genotypic methods were used to discriminate recrudescence from reinfection. Indeed, it is possible that the chance of being classified as a recrudescence was higher in Boende where a two-loci genotypic method was used compared to Kabalo, where a three-loci method was used. The high failure rate of SP in Boende is consistent with the molecular results for antifolate resistance, showing 61% had the triple  failure ,4.The threshold of resistance to SP at which it is no longer useful to add artesunate is not well defined. In Kabalo, the day 28-failure rate of SP was close to 20%, dropping to zero when given with three days of AS. Comparable results have been found in studies from Bie province in Angola where the PCR-adjusted 28-day failure rate was reduced from 25.3% for SP alone to 1.2% after addition of AS and in Southern Benin where the PCR-corrected failure rate of AS+SP at 28 days was reduced from 44.1% to 5.3% when combining AS to SP ,15. In cIt is not that surprising that in a large country, such as DRC, considerable differences in anti-malarial efficacy between provinces were observed. Site-specific patterns of treatment-seeking behaviour or malaria transmission intensity may explain geographical differences in the efficacy of anti-malarial treatments -18. The A limitation of these studies is the disappointing external quality control results, which suggest a failure of the internal quality control system in place. In the case of the Boende study, it is important to note that for the majority of discordant results the parasitaemia detected by one of the laboratories was less than 100/\u03bcL. Also, these results affected the classification of outcome in only three cases in Boende and none in Kabalo. The delay of some months before the EQC will have resulted in a deterioration in slide quality, which may partially explain these poor results. Having an adequate standard of microscopy even in these circumstances is critical. The experience at these two sites suggests that internal quality control measures in place were not adequate to guarantee this standard. External quality control at regular intervals may be the answer but is expensive, labour-intensive and the second reading needs to be timely to avoid excessive deterioration of slides, or slides need to be mounted. Sending a random sample of slides seems an obvious approach but leads to problems when agreement is not high. It is then recommended that all the slides be sent, by which time several months may have elapsed since the study has finished.in vivo efficacy studies in remote regions, in which there is political instability, is a major challenge. Proxy markers for resistance, such as molecular markers are invaluable tools to assess anti-malarial resistance in such a context, being less resource demanding than an in vivo study. Unfortunately, there are no markers, which reliably predict clinical resistance to drugs other than SP.Conducting Data from neighbouring Congo-Brazzaville to the west indicate high level SP and AQ resistance ,20. Effi. It is important that concomitant monitoring of anti-malarial efficacy is supported in order to inform policy makers [Since these studies were performed WHO pre-qualified anti-malarials have become available, e.g. artemether-lumefantrine and AS+AQ fixed-dose combination (FDC). Dihydroartemisinin-piperaquine and the FDC of artesunate-mefloquine are in the process of being registered. Both artemether-lumefantrine and dihydroartemisinin-piperaquine have been evaluated in a number of African countries and appear highly efficacious ,25. The y makers .The authors declare that they have no competing interests.MB was the principal investigator of the study conducted in Boende. She wrote the study protocol, coordinate supervised the study and wrote the paper. IVDB was the principal investigator of the Kabalo site. She wrote the study protocol, coordinate supervised the study and revised the manuscript. MVH participated to the development of the study protocol for Boende, gave a technical support to the investigators for the Boende site and revised the manuscript. PPPU participated to the development of the study protocol for Kabalo, gave a technical support to the investigators for the Kabalo site and revised the manuscript. CVO performed the genotypic analysis and coordinated the external quality control of the reading of malaria slides for the Boende site. She revised the manuscript. JK performed the genotypic analysis for the Kabalo site and revised the manuscript. CNN was the director of the National Malaria Control Programme when the two studies were discussed. He participated to the development of the study protocols and revised the manuscript. EA revised the statistical analysis of the 2 studies and revised the manuscript. JPG gave technical support to the principal investigators during the protocol writing, analysis and writing of the manuscript.All authors have read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria in Cambodia in 2000. However, recent clinical trials performed at the Thai-Cambodian border have pointed to the declining efficacy of both artesunate-mefloquine and artemether-lumefantrine. Since pfmdr1 modulates susceptibility to mefloquine and artemisinin derivatives, the aim of this study was to assess the link between pfmdr1 copy number, in vitro susceptibility to individual drugs and treatment failure to combination therapy.The combination of artesunate and mefloquine was introduced as the national first-line treatment for P. falciparum-infected patients enrolled in two in vivo efficacy studies in north-western Cambodia: 135 patients were treated with artemether-lumefantrine (AL group) in Sampovloun in 2002 and 2003, and 140 patients with artesunate-mefloquine (AM group) in Sampovloun and Veal Veng in 2003 and 2004. At enrollment, the in vitro IC50 was tested and the strains were genotyped for pfmdr1 copy number by real-time PCR.Blood samples were collected from pfmdr1 copy number was analysed for 115 isolates in the AM group, and for 109 isolates in the AL group. Parasites with increased pfmdr1 copy number had significantly reduced in vitro susceptibility to mefloquine, lumefantrine and artesunate. There was no association between pfmdr1 polymorphisms and in vitro susceptibilities. In the patients treated with AM, the mean pfmdr1copy number was lower in subjects with adequate clinical and parasitological response compared to those who experienced late treatment failure . This was not observed in the patients treated with AL . The presence of three or more copies of pfmdr1 were associated with recrudescence in artesunate-mefloquine treated patients (hazard ratio (HR) = 7.80 [95%CI: 2.09\u201329.10], N = 115), p = 0.002) but not with recrudescence in artemether-lumefantrine treated patients .The pfmdr1 copy number is a molecular marker of AM treatment failure in falciparum malaria on the Thai-Cambodian border. However, while it is associated with increased IC50 for lumefantrine, pfmdr1 copy number is not associated with AL treatment failure in the area, suggesting involvement of other molecular mechanisms in AL treatment failures in Cambodia.This study shows that  Plasmodium falciparum. In Cambodia, chloroquine resistance prompted the change to sulphadoxine-pyrimethamine (SP) as first-line treatment in the 1970s. Resistance to SP occurred and the first-line treatment was then changed to mefloquine monotherapy in 1993. However, by the mid-1990s, resistance to mefloquine exceeded 25% (RI-RIII) in western Cambodia -hypoxanthine  was used to assess parasite growth. Each isolate was tested once in duplicate in the microplates with serial dilutions of drugs. Drug response was quantified by monitoring [3H]hypoxanthine uptake in a Wallac MicroBeta Trilux counter . At the end of the incubation period, the plates were frozen at -20\u00b0C and thawed to lyse the cells. After collection on glassfiber filter paper using a cell harvester, the amount of [3H]-hypoxanthine incorporated into the parasites nucleoprotein was determined. The results of in vitro assay are expressed as the 50% inhibitory concentration (IC50), defined as the concentration at which 50% of the incorporation of [3H]-hypoxanthine was inhibited, as compared with in the drug-free control wells. Parasite growth was measured by using a log probit approximation to determine the IC50values.The in vitro drug sensitivity of the Cambodian isolates was assessed by use of a classical isotopic 48 h test . Briefly\u00ae) following the manufacturers protocol. Blood stored on filter paper was extracted using QIAamp DNA Mini Kit (Qiagen) , p = 0.002. After controlling for baseline parasitaemia and gender in a Cox's regression model, the hazard ratio was 12.28 [95%CI: 3.19\u201347.30], p < 0.001. There was no association between increased copy number and treatment failure in the AL group [HR = 1.03 [95%CI: 0.24\u20134.46], p = 0.962] (Figure pfmdr1 in anti-malarial resistance has been suspected since the 1990's but practical use of this information had to wait for methodological improvements and the availability of high quality quantitative PCR in endemic areas. This is now possible in a basic molecular laboratory. In this study, increased pfmdr1 copy number was associated with AM recrudescence. The association between pfmdr1 copy number and treatment failure to mefloquine-based regimens has been consistently found in both Cambodia and Thailand [pfmdr1 copy number within a host [pfmdr1 copy number was also associated with decreased in vitro susceptibility to mefloquine, lumefantrine and, to a lesser degree artesunate, which is consistent with previous studies [pfmdr1 copy number and AM failure is mediated by resistance to mefloquine alone or in conjunction with decreased sensitivity to artesunate. Further studies of this question are required.The involvement of Thailand -7. In adn a host ,19. Incr studies -22. It i50s may not be representative of the full set of samples studied for clinical outcomes and molecular features.In vitro drug sensitivity was successfully assayed in only 44% of the blood specimens. This high failure rate is likely to result from low parasite counts. However, some other factors such as poor transport condition from field sites to laboratory, or prolonged storage of infected blood at 4\u00b0C or presence of drug trace in the sample may have similarly interfere with the performance of the test [pfmdr1 copy number had apparently no impact on clinical AL failure even though it was associated with decreased in vitro susceptibility to both artemether and lumefantrine [pfmdr1-associated increase of IC50's for these drugs (Table pfmdr1 copy number and decreasing in vitro susceptibility to lumefantrine was at the same time observed, suggesting that AL failure could indeed result from decreased susceptibility to lumefantrine. Because of the exponential growth of Coartem\u00ae use in the past 5 years, high selection pressure could also be acting in Africa. The monitoring of drug resistance to lumefantrine is thus important for national anti-malarial treatment policy particularly in African countries.Interestingly, fantrine ,22. Thispfmdr1 copy number is a molecular marker of AM treatment failure in falciparum malaria on the Cambodia-Thailand border. However, while it is associated with increased IC50 for lumefantrine, pfmdr1 copy number is not associated with AL treatment failure in the area, suggesting involvement of other factors or molecular mechanisms in AL treatment failures in Cambodia.This study shows that The authors declare that they have no competing interests.pfmdr1 assay. CB and NK sequenced the samples. The clinical studies were coordinated and supervised by SD, MBD and PY. The study of molecular resistance markers was initiated by OMP, TF, SI and CW. FA, PR, JLB conceived the study, participated in study design. FA performed quality control of data and helped to draft the paper. PR is a staff member of the World Health Organization. This author alone is responsible for the views expressed in this publication and they do not necessarily represent the decisions, policy or views of the World Health Organization. All authors contributed to the critical review of the manuscript and agree to submission.PL did molecular investigation, carried out in vitro susceptibility testing of the field samples and drafted the paper. APA, NKS, and SRM developed the"}
+{"text": "Artemisinin-based combination therapy (ACT) is the treatment of choice for uncomplicated falciparum malaria. Artemether-lumefantrine (AL), a fixed dose co-formulation, has recently been approved for marketing in India, although it is not included in the National Drug Policy for treatment of malaria. Efficacy of short course regimen (4 \u00d7 4 tablets of 20 mg artemether plus 120 mg lumefantrine over 48 h) was demonstrated in India in the year 2000. However, low cure rates in Thailand and better plasma lumefantrine concentration profile with a six-dose regimen over three days, led to the recommendation of higher dose globally. This is the first report on the therapeutic efficacy of the six-dose regimen of AL in Indian uncomplicated falciparum malaria patients. The data generated will help in keeping the alternative ACT ready for use in the National Programme as and when required.msp-1 and msp-2 were used to differentiate the recrudescence and reinfection among the study subjects. In addition, polymorphism in pfmdr1 was also carried out in the samples obtained from patients before and after the treatment.One hundred and twenty four subjects between two and fifty-five years of age living in two highly endemic areas of the country (Assam and Orissa) were enrolled for single arm, open label prospective study. The standard six-dose regimen of AL was administered over three days and was followed-up with clinical and parasitological evaluations over 28 days. Molecular markers pfmdr1 gene.The PCR corrected cure rates were high at both the sites viz. 100% (n = 53) in Assam and 98.6% (n = 71) in Orissa. The only treatment failure case on D7 was a malnourished child. The drug was well tolerated with no adverse events. Patients had pre-treatment carriage of wild type codons at positions 86  and 184  of pfmdr1 gene is not co-related with clinical outcome. However, treatment failure can also occur due to incomplete absorption of the drug as is suspected in one case of failure at D7 in the study. AL can be a viable alternative of artesunate plus sulphadoxine/pyrimethamine (AS + SP), however, the drug should be used rationally and efficacy needs to be monitored periodically.AL is safe and effective drug for the treatment of acute uncomplicated falciparum malaria in India. The polymorphism in  Plasmodium falciparum to commonly used anti-malarial drugs, especially chloroquine, is being increasingly recognized in India [P. falciparum to anti-malarials and to achieve rapid resolution of parasitaemia and morbidity [Resistance in in India . More thorbidity . Accordiet al [Artemether-lumefantrine (AL) is a co-formulation of artemether and lumefantrine . The reported 28 day cure rate for six- dose regimen ranges between 89.6 and 98.5% and 42 day cure rate ranges between 87 and 94.8% -5. In InThe present study was conducted to evaluate the efficacy of AL with six-dose regimen in two endemic regions of India following the WHO (2003) therapeutic efficacy protocols .The study was conducted in Kamrup district (Assam) in north-eastern India and Keonjhar district (Orissa) in eastern part of India Figure , where tIt was an open label, single arm prospective trial based on the therapeutic efficacy protocols of WHO . The priP. falciparum malaria , with fever  or history of fever in preceeding 24 h, who gave voluntary consent were enrolled in local clinics. Pregnant or lactating women and children under 5 kg bodyweight were excluded. Patients with other febrile conditions, danger signs [Patients with microscopically confirmed er signs  or severAfter obtaining the informed written consent from patients or guardian, a medical history including presenting symptoms, current medications and previous anti-malarial use was obtained. A complete physical examination was performed and case record form was completed for each patient. Clinical history, examination and other investigations were all recorded. Blood was collected for parasitology and molecular biology studies.\u00ae, Novartis Pharmaceuticals Corporation, Suffern, New York, USA; Batch No.: F0443) according to body weight with biscuits and glass of water. Patients weighing 10\u201315 kg received one tablet (20 mg artemether plus 120 mg lumefantrine) per dose, those weighing 15\u201325 kg received two, those of 25\u201335 kg received three, and those >35 kg received four tablets. In total, six doses were administered at hours 0, 8, 24, 36, 48, and 60. Treatment with both the doses, every day, was directly observed by the study team. No patient had vomiting and, therefore, re-administration of the drug was not required.Patients were given AL  according to the manufacturer's instructions and used for a polymerase chain reaction (PCR). To distinguish between recrudescent and new infections PCR method was used to analyse polymorphisms in the merozoite surface protein (MSP) genes,  earlier .pfmdr1, one spanning codons 71\u2013242, and other covering 945\u20131308 codons were amplified to analyze SNP's at codons 86, 184, 1034, 1042 and 1246. In pfcrt SNP's at codons 72\u201376 were studied by amplifying region-spanning codons 44\u2013177. Primers and cycling conditions used for amplification of fragments of pfmdr1 are shown in Table pfcrt analysis was same as described earlier [. About 50\u2013250 ng of the purified DNA was used for sequencing PCR using ABI Big Dye Terminator Reaction Ready Kit Version 3.1. The sequencing PCR was performed in a volume of 20 \u03bcl with 1 \u03bcl of Terminator Ready Reaction Mix (TRR), 3.2 pmol of gene-specific primer. Cycling conditions for the sequencing PCR included 25 cycles of denaturation at 96\u00b0C for 10 sec, annealing at 50\u00b0C for 5 sec, and extension at 60\u00b0C for 4 min. Templates were purified and sequenced on an ABI Prism 310 Genetic Analyzer (PE Applied Biosystems).Two fragments of  earlier . The PCRData was entered in WHO TM software. The cumulative risk of failure was assessed by survival analysis with the Kaplan Meier method.msp-1 and msp-2 confirmed one of them as reinfection and one as recrudescence. This led to PCR corrected cure rates of 98.6% in Orissa. The cumulative risk of failure (PCR corrected) by Kaplan Meier analysis was 0.014 (95% CI 0.002\u20130.097) in Orissa while it was zero in Assam. The response was similar in all age categories and the only patient who had recrudescence on D7 was a seven year-old child, weighing 11 kg. The parasite count on D7 was 48/\u03bcl  in this patient. PCR analysis confirmed that it was a single clone infection and D0 and D7 samples displayed the matching allele bands indicating a case of recrudescence. Molecular analysis of pfmdr1 revealed 86Y in both D0 and D7 samples of this patient. Pfcrt haplotype as determined by DNA sequencing was C72M74N75K76 in both D0 and D7 samples reiterating the case as recrudescence.Patients were screened between July and October 2007 at Kamrup District, Assam and August to October 2007 at Keonjhar District, Orissa. A total of 124 patients were enrolled. Baseline characteristics and age category is shown in table pfmdr1 and pfcrt genes. In pfmdr1, 58 isolates (26 from Assam and 32 from Orissa) could be analyzed for both the fragments. Fragment 1 covering codons 86 and 184 was amplified from 91 isolates (27 in Assam and 64 in Orissa) while only 63 isolates (36 from Assam and 27 from Orissa) were successfully amplified and sequenced for pfcrt region covering codons 72\u201376. Out of the five pfmdr1 codons analysed , mutation was observed only at 86 and 184 codons while others remained wild type among all isolates. There were a total of 3 different pfmdr1 alleles among the isolates. Genotype Y86Y184S1034N1042D1246 was more prevalent  than wild type (N86Y184S1034N1042D1246) and the single mutant (N86F184S1034N1042D1246) alleles which were present in 31.03 and 6.89% of isolates, respectively. At codon 86, pre-treatment carriage of N86 was 41.7% while 86Y was 58.3%. However, at codon 184 most of the isolates showed 184Y . Mutation in pfcrt was observed at 72 (14.89% n = 9), 74, 75 (68.6% n = 43) and 76 (70% n = 53) codons among 63 isolates with C72I74E75T76 triple mutant allele being most predominant (68.6%) while isolates with C72M74N75T76 were rare (1.4%). Only 14.9% isolates were found to contain the wild type C72M74N75K76or double mutant S72M74N75T76alleles.A total of 124 clinical isolates (53 from Assam and 71 from Orissa) were analysed for genetic polymorphism in pfmdr1 were present in both Assam (n = 26) and Orissa (n = 32). Single mutant Y86Y184S1034N1042D1246 genotype was more prevalent in both regions as compared to N86Y184S1034N1042D1246 and N86F184S1034N1042D1246 genotypes (Table pfcrt triple mutant (C72I74E75T76) genotype was present in highest proportion in both Assam (72.2% n = 36), and Orissa . Proportionately wild type C72M74N75K76genotype was more prevalent in Orissa (25%) than in Assam (8.3%). Also single mutant C72M74N75T76 was found only in Orissa (3.5%). S72M74N75T76 genotype was present in both Assam (19.4%) and Orissa (7.1%).All the three genotypes of issa n = 2. Singlemsp-1 and msp-2 genotyping confirmed one of them as reinfection and other as a recrudescence. Another case in Orissa, reported on D38 with positive parasitaemia which was reinfection and as such not included in analysis of 28 day failure rate. Out of these cases of reinfection, one had Y86Y184S1034N1042D1246 and another showed N86Y184S1034N1042D1246 haplotype at D0. However, on the day of failure, both cases had different pfmdr1 genotypes, N86Y184S1034N1042D1246 (D28) and N86F184S1034N1042D1246 (D38) respectively  of 117 high risk districts and 256 chloroquine resistant PHCs of 48 districts for the treatment of falciparum malaria. The ACT recommended by the National Programme is AS+SP, which is highly effective but available as blister pack and not a fixed dose formulation. AL is the only fixed dose combination that has been approved for marketing recently and there is no data on safety and efficacy of recommended six-dose regimen in India. The present study was carried out to assess the efficacy of AL in India and its correlation with molecular markers. The drug was well tolerated and produced rapid parasite clearance. Majority of patients, pfcrt haplotype C72M74N75K76 and pfmdr1 Y86Y184S1032N1042D1246 haplotype in both D0 and D7 Samples. In vivo exposure to AL props up the selection of pfmdr1 86N in new inoculation and is suggested to be associated with tolerance to lumefantrine [A case of treatment failure on D7 has been observed for the first time with the six-dose regimen of AL in the study. There is only one earlier report of failure on D9 in a three-year old child in Comoros . TreatmeThe likelihood of resistance to AL in India is low, since the drug is not included in the National Drug Policy for the treatment of malaria till date. However, recently, marketing permission has been given to few companies. In this scenario, the reappearance of parasites on D7 led us to investigate the possible misuse of the drug. Six brands of AL were available in the town since 2006 and the drug was dispensed without prescription by the local chemists. Discussion with doctors also revealed that drug was being prescribed in various private clinics. This case highlights the need to study the kinetics of drug in malnutrition. In addition, over the counter availability and possible misuse of the drug raises concern of accelerating resistance in near future to one of the valuable ACT.pfmdr1 and pfcrt alleles in Indian P. falciparum isolates was also studied. High prevalence of 184Y (93.3%) in pre-treatment samples was observed. Genotypes indicate that majority of isolates carried Y86Y184S1034N1042D1246 (62.06%) followed by N86Y184S1034N1042D1246 (31.03%) and only 6.8% isolates had N86F184S1034N1042D1246genotype. In pfmdr1 mutation at only codons 86 and 184 have been found which is similar to earlier report [In addition to clinical outcomes, pre-treatment prevalence of r report .Y86Y184Y1246 genotype of pfmdr1, and selection of N86F184D1246 genotype after AL treatment. Results of the present study indicate baseline prevalence of Y86Y184D1246 haplotype in India. Also 6.8% pre-treatment isolates that carry N86F184D1246genotype were drug sensitive. Two clinical failure cases were classified as reinfection which selected N86Y184S1034N1042D1246and Y86Y184S1034N1042D1246 haplotype, whereas one case of recrudescence has not shown any change in pfmdr1 haplotype, i.e. it remained as Y86Y184S1034N1042D1246. Since there were very few clinical failure cases, it is difficult to conclude any specific selection of a particular genotype after AL treatment. Presence of pfcrt C72I74E75T76 genotype in highest proportion indicates the preexistence of high level of CQ resistance among the parasite population [Previous studies with AL in Africa  showed hpulation .The main limitation of the study is that co-relation of molecular markers with treatment failure was difficult due to very low number of treatment failure cases and drug concentration in this case could not be done.Considering the prevalence of molecular markers of resistance to sulphadoxine/pyrimethamine (SP) especially in north-eastern states in India , the nonin vivo studies and further molecular studies may help in developing markers to predict impending resistance.Lack of correlation of molecular markers for resistance with clinical outcome makes it difficult to monitor and predict resistance to this combination. Efficacy of AL needs to be monitored using Artemether-lumefantrine is safe and effective drug for the treatment of uncomplicated falciparum malaria in India. The efficacy of this ACT needs to be carefully monitored periodically since the treatment failures can occur due to resistance as well as subtherapeutic levels due to inadequate absorption especially if not administered with fat or in malnutrition. As such malnutrition and poor uptake of fat is common in poor tribal population. However, since AL is the only fixed combination available in India till date, it can be a viable alternative to AS+SP for the treatment of uncomplicated falciparum malaria.AL: artemether-lumefantrine; SP: sulphadoxine/pyrimethamine; ACT: artemisinin-based combination therapy.The authors declare that they have no competing interests.NV: PI, Development of protocol, Execution, quality control of study and preparation of manuscript. PS, PM: Molecular studies and Preparation of manuscript. Ruchi Singh: Molecular studies. SSM, PKT and KP: Patient enrollment and Parasitology. SKS and VD: Supervision and Co-ordination of field work. APD: Critical review of manuscript. YDS: Protocol and review for molecular studies and manuscript."}
+{"text": "In areas of seasonal malaria transmission, treatment of asymptomatic carriers of malaria parasites, whose parasitaemia persists at low densities throughout the dry season, could be a useful strategy for malaria control. We carried out a randomized trial to compare two drug regimens for clearance of parasitaemia in order to identify the optimum regimen for use in mass drug administration in the dry season.A two-arm open-label randomized controlled trial was conducted during the dry season in an area of distinct seasonal malaria in two villages in Gedarif State in eastern Sudan. Participants were asymptomatic adults and children aged over 6 months, with low-density P. falciparum infection detected by PCR. Participants were randomized to receive artesunate/sulfadoxine-pyrimethamine (AS+SP) combination for three days with or without a dose of primaquine (PQ) on the fourth day. Parasitaemia detected by PCR on days 3, 7 and 14 after the start of treatment and gametocytes detected by RT-PCR on days 7 and 14 were then recorded. 104 individuals who had low density parasitaemia at screening were randomized and treated during the dry season. On day 7, 8.3% were positive by PCR in the AS+SP+PQ group and 6.5% in the AS+SP group . At enrolment, 12% (12/100) were carrying gametocytes. This was reduced to 6.4% and 4.4% by day 14 (Risk difference 1.9% (95%CI \u22129.3% to +13.2%) in AS+SP+PQ and AS+SP groups, respectively.Addition of primaquine to artemisinin combination treatment did not improve elimination of parasitaemia and prevention of gametocyte carriage in carriers with low-density parasitaemia in the dry season.NCT00330902ClinicalTrials.gov  Plasmodium falciparum infections are asymptomatic and remain untreated. In eastern Sudan people who become infected during the wet season may retain chronic sub-microscopic asymptomatic infections throughout the dry season In malaria endemic countries of sub-Saharan Africa, the majority of Plasmodium vivax, as a prophylactic against all malaria species, and as a gametocytocidal drug against P. falciparumP. falciparum malaria to reduce transmission Artemisinin combination treatment is highly effective in eliminating asexual parasitaemia, the source of merozoites committed for gametocyte production P. falciparum infections P. falciparum parasitaemia before the transmission season in an area of marked seasonal transmission in eastern Sudan in order to identify the optimum regimen for use in mass drug administration during the dry season.Individuals with low gametocyte densities undetectable by microscopy may still be infectious to mosquitoes The protocol for this trial and supporting CONSORT checklist are available as supporting information; see The study was carried out in two villages in eastern Sudan. Abunaja Juama located at about 18 km south west of Gedarif and Kanara at 7 km south of Gedarif.P. falciparum and the main mosquito vector is Anopheles arabiensisP. falciparum. Signed informed consent forms were obtained from all participants and from the parents or guardians of children under 15 years. Exclusion criteria included pregnancy, history of allergy to sulpha drugs, fever or other signs or symptoms of illness but none of the persons screened had any of these conditions. After screening individuals were excluded if, on microscopic examination of a blood film, Plasmodium species other than falciparum were detected or the sample was negative by PCR.The whole area is characterized by seasonal malaria with transmission confined to three months of the year, October \u2013 December. The main malaria parasite is Participants were randomized to receive a standard dose of AS and SP (AS+SP) over three days, or AS+SP over the first three days plus primaquine (PQ) administered on the fourth day.P. falciparum parasitaemia, in order to identify the optimum regimen for use in mass drug administration in the dry season.We wanted to determine the efficacy of AS+SP compared to AS+SP+PQ in clearing sub-patent parasitaemia and gametocyte carriage in persons with asymptomatic sub-microscopic P.falciparum parasitaemia detected by PCR on days 3, 7 and 14 and presence of gametocytes detected by RT-PCR on days 7 and 14. For these endpoints, a sample by finger prick was obtained on days 0, 3, 7 and 14 on glass slides and on filter paper, and on days 7 and 14 venous blood was collected in K3EDTA vacutainers. Adverse events were recorded on days 1, 2, 3, 7 and 14 and packed Cell Volume was measured on days 0, 7 and 14. Parasite DNA was extracted by the Chelex-Resin method P. falciparum gametocyte specific pfS25 gene was amplified in a nested PCR to increase sensitivity of the detection The outcomes were In a previous trial 3EDTA vacutainer tubes (Becton Dickinson) for gametocyte detection by RT-PCR, and a drop of blood was collected on a glass slide for microscopic diagnosis and another drop on filter paper for PCR.A cross sectional survey was carried out in the middle of the dry season (June) 2004 to identify sub-patent parasite carriers. A finger prick sample was collected on a glass slide for microscopic diagnosis and on filter paper for molecular analysis. Persons who were positive by PCR in June and eligible were enrolled in the study and randomly allocated into one of the two treatment arms. The list of carriers was sorted according to village and age to ensure that the treatment groups were balanced with respect to these two variables. The random allocation of this ordered list into the treatment arms was then created using restricted randomization with a block size of 12 in Stata version 7 . In August, a clinical examination was performed; no participants were excluded due to illness or other exclusion criteria. Before administering the first treatment dose (day 0), a venous blood sample was obtained from each individual in KTreatment was administered under medical supervision. SP was administered on day 0 as 25 mg/kg sulfadoxine/1.25 mg/kg pyrimethamine for children under 50 kg, while adults and children weighing 50 kg or more were given three tablets (each tablet contained 500 mg of sulfadoxine and 25 mg of pyrimethamine). Artesunate was given on days 0, 1 and 2 at a dose of 200 mg (two tablets at 100 mg) for adults and for children weighing 50 kg or more, while children under 50 kg received 4 mg/kg body weight. Participants randomized to receive primaquine were, in addition, given a single dose of 0.75-mg/kg body weight on day 3. After each treatment dose, patients were observed for one hour, if the drug was vomited during the first half hour a full dose was repeated, if vomiting occurred after one hour half a dose was repeated.The trial was not blinded; however laboratory staff performing the PCR assays were unaware of the treatment allocation.95% confidence intervals were calculated for the difference between the proportions with parasitaemia and gametocytaemia in the two groups on days 7 and 14, using the modified score method of Newcombe . 114/615 (19%) had PCR detectable P. falciparum parasitaemia (including the 9 which were positive by microscopy) and were therefore eligible for enrolment into the trial. The three who were positive by microscopy for other Plasmodium species were PCR negative for falciparum.Out of a total of 615 asymptomatic individuals screened by microscopy in June 2004 (the dry season) 12 were positive for malaria parasites  when 104 eligible participants were enrolled and 10 were absent. Of these 4 (4.2%) were positive for asymptomatic th to 20th August, and followed up until 3rd September 2004.Participants were enrolled from 17The demographic data for the study group is shown in Four (4%) individuals from those who were randomized in the AS+SP group were found to be negative by PCR on day 0. Of these 3 were lost to follow up on day 14 and one withdrew consent by day 3. They have been excluded from the analysis of safety and efficacy. The remaining withdrawn and lost to follow-up cases were also excluded. Data analysis was performed according to the protocol.P. falciparum infection. However, only 20 (20.4%) out of 98 participants seen on day 3 post-treatment were found to harbour P. falciparum infection detectable by PCR. On day 7 only seven (7.4%) out of 94 participants retained PCR detectable P. falciparum infection, 4/48 (8.3%) in the AS+SP+PQ group and 3/46 (6.5%) in the AS+SP group . Similarly, five out of 92 seen on day 14 had PCR detectable parasitaemia (On recruitment (day 0) 100 (96%) out of 104 individuals carried asymptomatic sub-patent sitaemia .With regard to gametocytes, on enrolment, twelve (12%) out of 100 individuals carried gametocytes detectable by RT-PCR. None of them were gametocyte positive on day 7 or day 14. The same persons who were detected positive by PCR on days 7 and 14 were also positive for gametocytes by RT-PCR. On day 7, four (8.3%) out of 48 on AS+SP+PQ and three (6.5%) out of 46 individuals in AS+SP were found to have RT-PCR detected gametocytes . Similarly on day 14, 6.4% in AS+SP+PQ and 4.4% in AS+SP had sub-microscopy gametocytaemia (risk difference 1.9% (95%CI \u22129.3% to +13.2%).Two out of the seven gametocyte carriers detected on day 7 retained their gametocyte producing infection until day 14. However, gametocytes reappeared on day 14 among three individuals who were gametocyte negative on day 7.Packed cell volume was similar in both groups on day 7, mean 34.6% (15\u201344%) in the AS+SP group and 34.2% (26\u201342%) in the AS+SP+PQ group, difference between groups adjusted for baseline 0.78  P\u200a=\u200a0.315. Similar results were seen on day 14 .No serious or severe adverse events were reported. Four of the participants (3.8%) vomited after the first treatment dose of AS+SP; 1 on AS+SP and 3 on AS+SP+PQ arm. One of these vomited immediately, refused to take the drug again and withdrew consent. The other three cases (2.9%) vomited after more than eight hours after taking the drug and the dose was therefore, not repeated for them. Two people complained of insomnia and another two complained of itching.P. falciparum parasitaemia that persist in the dry season, as a potential control strategy in areas of seasonal transmission. After treatment with AS+SP alone 80% (78/98) of subjects with asymptomatic sub-patent P. falciparum infections became PCR negative by day 3 (before administration of PQ). However, only three (6.4%) in AS+SP and four (8.3%) in AS+SP+PQ group had sub-patent gametocytes on day 7, and two (4.4%) and three (6.4%) on day 14, respectively.We compared the efficacy of AS+SP and AS+SP+PQ in clearing asymptomatic sub-patent P. falciparum was found to be less than 1\u223610 P. falciparum gametocyte carriers in nature usually harbour less than 100 gametocyte \u00b5l\u22121 blood and there is evidence that 1\u201310 gametocytes \u00b5l\u22121 blood are infectious to mosquitoes P. falciparum to Anopheles mosquitoes P. falciparum infection to infectivity of Anopheles mosquitoes The present results are consistent with previous findings that asymptomatic sub-patent parasitaemia PfS25 gene, which is exclusively expressed by mature gametocytes Our results show the efficacy of artesunate against asymptomatic sub-patent gametocyte carriage. Gametocytes present before treatment were most probably mature since we used the P.falciparum parasitaemia in the dry season without the need for the addition of primaquine. These results, for infections near the limit of detection, may not apply to higher density infections where detectable gametocytes may persist for several weeks in spite of effective treatment of asexual parasitemia The clearance of sub-patent parasitaemia and gametocyte carriage by day 7 after treatment with AS+SP indicates that this may be an effective treatment for use to clear low density The combination AS+SP without primaquine is effective in eliminating the dry season sub-microscopic parasitaemia and gametocyte carriage. Therefore, this regimen could be recommended for use in mass drug administration in the dry season to control malaria in areas of seasonal transmission.However, it may be necessary to complete the drug administration before the rainy season and resurgence of the mosquitoes so as to avoid transmission of mature gametocytes.Protocol S1Trial Protocol(0.16 MB DOC)Click here for additional data file.Checklist S1CONSORT Checklist(0.04 MB DOC)Click here for additional data file."}
+{"text": "We report the findings from baseline plasma sampling of malaria patients prior to inclusion into an in vivo study in Tanzania and discuss the implications of residual concentrations of antimalarials in this setting.Repeated antimalarial treatment for febrile episodes and self-treatment are common in malaria-endemic areas. The intake of antimalarials prior to participating in an in vivo study conducted in a rural area of Tanzania in 2008, baseline plasma samples from patients reporting no antimalarial intake within the last 28 days were screened for the presence of 14 antimalarials (parent drugs or metabolites) using liquid chromatography-tandem mass spectrometry. Among the 148 patients enrolled, 110 (74.3%) had at least one antimalarial in their plasma: 80 (54.1%) had lumefantrine above the lower limit of calibration (LLC\u200a=\u200a4 ng/mL), 7 (4.7%) desbutyl-lumefantrine (4 ng/mL), 77 (52.0%) sulfadoxine (0.5 ng/mL), 15 (10.1%) pyrimethamine (0.5 ng/mL), 16 (10.8%) quinine (2.5 ng/mL) and none chloroquine (2.5 ng/mL).In an in vivo studies to allow for accurate assessment of treatment outcome. Furthermore, persisting sub-therapeutic drug levels of antimalarials in a population could promote the spread of drug resistance. The knowledge on drug pressure in a given population is important to monitor standard treatment policy implementation.The proportion of patients with detectable antimalarial drug levels prior to enrollment into the study is worrying. Indeed artemether\u2013lumefantrine was supposed to be available only at government health facilities. Although sulfadoxine\u2013pyrimethamine is only recommended for intermittent preventive treatment in pregnancy (IPTp), it was still widely used in public and private health facilities and sold in drug shops. Self-reporting of previous drug intake is unreliable and thus screening for the presence of antimalarial drug levels should be considered in future  The standard World Health Organization (WHO) protocol for monitoring antimalarial drug efficacy does not exclude patients with a history of previous antimalarial drug use or the presence of antimalarial drugs in the urine or blood The intake of antimalarial drugs prior to inclusion in an Policy makers in malaria endemic countries are faced with the difficult problem of ensuring easy and early access to effective and high quality antimalarials, while preventing their uncontrolled and unnecessary use, which would increase drug pressure on the parasites and encourage parasite resistance. Thus, knowledge of drug use in a specific area could help decisions makers to assess how treatment policies are implemented.Plasmodium falciparum malaria recruited in an in vivo study in Tanzania. Samples were analysed for the presence of 14 currently in-use antimalarials in a single run using a liquid chromatography-tandem mass spectrometry assay Here we report the findings of the analysis of baseline samples from patients with All the applied protocols and related documents were approved by the Ethikkommission beider Basel (EKBB), the Institutional Review Board of the Ifakara Health Institute and the National Institute for Medical Research Review Board. Blood samples were obtained after written informed consent in Swahili from the participants or their responsible guardians.et al., in preparation). In the private sector these drugs could be purchased over the counter without a doctor's prescription.The study was performed in a rural area with moderate to high malaria transmission intensity  during the main rainy season from March to May 2008. At the time of the study, artemether\u2013lumefantrine (AL) had recently been introduced as first-line treatment and was only available at government health facilities to ensure controlled prescription. Before 2006, the official first-line treatment in Tanzania was SP, which had in turn replaced CQ in 2001. In 2008, amodiaquine, SP and quinine were widely available in the private sector in the study area. Artesunate, dihydroartemisinin, and halofantrine could also be found sporadically in a few drug shops  Febrile patients were recruited at the Kibaoni Health Center, 6 km from Ifakara down town, that serves a population of 26,261. The population lives in villages with good coverage of government health facilities and licensed drug stores  for the presence of Plasmodium falciparum. Parasite count and specification of Plasmodium were done by microscopy. The patients with a positive result were then seen by a clinical officer, who invited them to participate in the study if they did not present with danger signs of complicated malaria or severe concomitant illness, and if they reported not having taken antimalarials in the previous 28 days. The latter information was checked against the patient's care log book when available. Consenting patients had a baseline sample  taken to check for potential residual antimalarials and correct pharmacokinetic analyses later on. Treatment with the standard first-line treatment AL was then initiated, according to body weight and age category.The trial was designed to assess the effects of the individual pharmacogenetic profiles on the disposition of standard antimalarials (to be reported elsewhere). It was based on the standard WHO protocol for N-desethyl-amodiaquine, lumefantrine, desbutyl-lumefantrine, piperaquine, pyronaridine, mefloquine, chloroquine, quinine, pyrimethamine and sulfadoxine, were determined by liquid chromatography coupled to tandem triple stage mass spectrometry (LC-MS/MS) Blood samples were kept on ice for no longer than 6 h after bleeding (venipuncture) and then aliquoted into whole blood, plasma and pellet and immediately stored at \u221280\u00b0C. Plasma concentrations of 14 antimalarial drugs and their metabolites, i.e. artemether, artesunate, dihydroartemisinin, amodiaquine, Summary statistics, Chi-square tests, multivariate analysis and graphs of residual plasma concentrations of antimalarials found prior to treatment were produced using Stata\u00ae . Logistic regression analysis was used to investigate the influence of body weight, sex and distance from health facilities or pharmacies on the presence of antimalarials at study entry. The distance between patient home and health facility or pharmacy was defined as \u201cclose\u201d or \u201cfar\u201d depending on the distance in kilometers, also taking into account ease of access, i.e. main road and river (according to 0C) and on Day 7 (7C) after a complete treatment with AL for the same patients. We included only patients for whom we had both samples and who complied with the three-day, six-dose AL treatment schedule. Assuming a terminal elimination half-life of \u00bdt\u200a=\u200a3.3 days for lumefantrine, an inter-individual variability of 40% To estimate the probable timing of drug intake, we compared the plasma concentrations of lumefantrine at baseline  according to body weight: 1500 mg for >45 kg, 1000 mg for 31\u201345 kg, 750 mg for 21\u201330 kg, 500 mg for 11\u201320 kg and 250 mg for 5\u201310 kg. Approximate averages of pharmacokinetic parameters, inter-individual variability and intra-individual  variability were derived from a literature review  and 2. Tin vivo study. Two patients (one from the Kibaoni HC area and one from Kining'ina) were excluded from the analyses , leaving 148 patients with a valid baseline sample, of whom 64 (43.2%) were male and 84 (56.8%) female (3 (2.0%) pregnant in 3rd trimester). Patients' ages ranged from 1 to 78 years (median 9 years). 51 (34.5%) patients were children under the age of 5, and 94 (63.5%) were <12 years old.A total of 1672 patients of all age were screened, of whom 389 (23%) had a positive malaria test and 150 were eligible and willing to participate in the \u03c72\u200a=\u200a5.82, P\u200a=\u200a0.016) than among older children and adults .The presence of antimalarial drug was detected in the plasma of 111 (74.3%) patients: 80 (54.1%) had lumefantrine above the lower limit of calibration (LLC\u200a=\u200a4 ng/mL), 7 (4.7%) desbutyl-lumefantrine (LLC\u200a=\u200a4 ng/mL), 77 (52.0%) sulfadoxine (LLC\u200a=\u200a0.5 ng/mL), 15 (10.1%) pyrimethamine (LLC\u200a=\u200a0.5 ng/mL), 16 (10.8%) quinine (LLC\u200a=\u200a2.5 ng/mL) and none chloroquine (LLC\u200a=\u200a2.5 ng/mL) or any other antimalarials tested. Summary statistics are shown in \u00bdt translates into 90% confidence intervals extending from 74% to 144% of estimates (median).For lumefantrine, among the 59 eligible patients the median plasma concentration (range) was 18.3 ng/mL (4.4\u2013181.8 ng/mL) on Day 0 and 413 ng/mL (37.3\u20131402 ng/mL) on Day 7. This means that, to account for the levels observed on Day 0, a similar dosage level should have been administered at a median of 21 days  before study entry. In 2 patients (3%) this estimate was >28 days. The variability in Back-estimation of the most likely times for sulfadoxine intake indicated a median of 108 days before blood sampling at study entry . Two patients had concentrations >78 mg/mL, compatible with same day intake. In 70 patients (91%), the estimate exceeded 28 days. The evaluation of uncertainty around individual dose intake times showed 90% confidence intervals extending from 49% to 202% of estimates (median).\u03c72\u200a=\u200a9.06, P\u200a=\u200a0.03 in the first analysis; likelihood ratio \u03c72\u200a=\u200a9.60, P\u200a=\u200a0.02 in the second analysis), patients with lower body weight being more likely to show residual AL levels. However, this was not the case for SP or SP and AL taken together. Furthermore, neither sex nor distance from health facilities or pharmacies showed a significant effect on residual levels of SP, AL or both at study entry.The investigation of the influence of body weight, sex and distance to health facilities or pharmacies on the probability of residual antimalarials at study entry (details not presented) showed only a significant relationship between body weight and residual AL levels  must have taken the drug within 28 days prior to treatment. Furthermore, it is also possible that patients might have taken a sub-therapeutic dose of AL more recently. However, as indicated by the wide variability in elimination half-lives, these values represent only rough estimates. Nevertheless, these findings suggest that at least half of the patients included into our study had taken AL, mostly within the previous month . The Day 7 values observed in this study are comparable with those of a study in Thai patients [median plasma concentration (range) of lumefantrine was 528 ng/mL (49\u20135175 ng/mL) after 6 doses of AL over 60 h according to body weight et al., in preparation), especially so when clinicians were doubtful about the diagnosis of malaria.SP was still found in approximately half of the patients, although it had been officially abandoned as first-line treatment since 2006. Assuming that the patients with residual SP in their plasma had taken a single dose of sulfadoxine according to body weight, most patients must have taken the drug \u223c3.5 months (median 108 days) prior blood withdrawal. However, 2 patients had sulfadoxine plasma concentrations indicating very recent exposure, and another 8 exposure during the last 4 weeks. Furthermore, it is also possible that patients might have taken a sub-therapeutic dose of SP more recently. These estimates are approximate, as indicated by the wide confidence interval explained by the fair degree of inter-individual variability in clearance, volume of distribution and residual error. The LC-MS/MS method used for the determination of sulfadoxine (LLC\u200a=\u200a0.5 ng/mL) would theoretically make it possible to detect traces of sulfadoxine up to 4 months (127 days) after a single dose of 25 mg/kg of sulfadoxine. These findings are sufficient to conclude that a significant number of patients had taken SP for a previous febrile episode, which is against the standard treatment recommendations. Recent surveys on availability of antimalarials have shown that sick people were getting SP from drugs shops, public and private health facilities . An epidemiological study which used the Explanatory Model Interview Catalogue (EMIC) reported that 87.5% (78.2\u201393.8%) of all fevers in children in our study area were treated with one of the recommended antimalarials  et al., in preparation).Why was the number of patients with residual antimalarials so high? Through the demographic surveillance system (DSS) data and a treatment seeking survey in the Ifakara area it was found that approximately 8% of children had fever in the previous 2 weeks when seen between January and April 2008  or information recorded in the care log book (self-treatment not documented) are thus unreliable at least in this population and for lumefantrine.Whatever the reason is for the large number of malaria patients with antimalarials in their blood at study baseline, the fact remains that these patients are the usual subjects investigated in Previous drug intake may affect the current treatment in several ways. Higher drug exposure resulting from cumulative levels may lead to better efficacy or more toxicity. The parasites causing the disease at the time of enrolment may be of a less sensitive population selected by the previous treatment. Thus, previous antimalarial intake may impact on the outcome of the treatment under investigation, and this study shows that only baseline drug concentration measurement in the blood can reliably be used to account for this effect. Our LC-MS/MS assay covers 14 antimalarials in a single run. We can confidently exclude a lack of specificity and false positives as we included blank plasma samples as negative controls and systematically repeated the measurement on a new chromatographic column. Furthermore, we have demonstrated that all drugs are stable for up to 48 h in plasma stored at 4\u00b0C 10 , values which are in line with those reported for symptomatic cases in malaria-endemic areas There is abundant literature on the effects of inadequate antimalarial treatment on the emergence and spread of resistance. Here, we do not know if the residual drug levels found were from a full or incomplete treatment, if the person had parasites at the time of the previous drug intake, and whether the parasites causing the current episode are the same or a new infection. Be it as it may, the residual levels were not enough to control parasite replication and clinical symptoms. This means that these parasites have been exposed to inadequate drug levels for some time. The chances of drug resistant parasites to be selected depends on several factors, and is higher for patients with no immunity (e.g. young children), drugs with long residence times and resistance being conferred through single point mutations, and for infections with a large parasite biomass The findings of this study must be confirmed in other settings as they have potential implications for both clinical research and surveillance (treatment efficacy and safety outcome) and control ."}
+{"text": "There are no data on the long term use of an artemisinin combination treatment in moderate or high transmission areas of Africa.Plasmodium falciparum-infected patients of all ages in the Oussouye district, Casamance, Senegal, over a period of six years (2000 to 2005). Efficacy, by Kaplan Meier survival analysis (n = 966), and safety  were determined over 28 days. A weight-based dosing regimen was used for the loose tablets during 2000\u20132003 (n = 731) and a commercially available co-blister was used during 2004\u20132005 (n = 235).Artesunate plus amodiaquine (AS+AQ) was used to treat slide-proven Annual crude (non PCR corrected) rates remained stable over the study period . Nine co-blister treated patients (0.9%) withdrew because of drug-related adverse events; seven had gastrointestinal complaints of whom two were hospitalized for vomiting. By Day 28, the mean total bilirubin (n = 72), AST (n = 94) and ALT (n = 95) values decreased. Three patients had Day 28 AST/ALT values > 40 < 200 IU/L. Changes in white cell counts were unremarkable (n = 87).AS+AQ in combination was highly efficacious and well-tolerated in this area and justifies the decision to use it as first line treatment. Long-term monitoring of safety and efficacy should continue. Artemisinin-containing combination therapies (ACTs) are now being deployed in some 42 malaria endemic countries and a further 26 have agreed to adopt ACTs following the World Health Organization (WHO) recommendation that ACTs should be the first line drugs for treating uncomplicated falciparum malaria .Artesunate plus amodiaquine (AS+AQ) is one of the currently available ACTs and is in use in Indonesia and 18 African countries , Zanzibar).Plasmodium falciparum while the trend of increasing in vitro mefloquine resistance has been reversed  in 2000, 94.0%  in 2001, 95.7%  in 2002, 94.9%  in 2003, 88.5%  in 2004 and 95.9%  in 2005. There were no differences (p = 0.12) in cure rates between years by the log rank test for homogeneity over time  between the loose (n = 731) and the co-blistered (n = 235) products: 95.1%  vs. 93.1% , respectively. This held true whether dosing was weight (n = 761) or age (n = 205) based: 94.8%  vs. 94.2% , respectively (p = 0.51) and whether treatment was supervised (n = 810) or unsupervised (n = 156): 94.5%  vs. 95.9% , respectively (p = 0.75).Year of study and product used were non significant contributors to failure in a Cox proportional hazard model. Using a saturated model containing year, product, site, age, sex, weight, dose AS and dose AQ, the contribution to hazard of failure was border-line for year (p = 0.06) but significant for the total daily dose of AS (p = 0.004) for a hazard of failure of 0.993 . Only year 2004 was statistically different from the reference year 2000 (p = 0.003): the hazard of failure was four times greater in 2004 than in 2000 .Complete safety records are available for 752 patients enrolled during 2001\u20132005. At presentation (Day 0), all patients reported fever or had a measured fever in the clinic. Other malaria associated symptoms/signs on presentation were weakness [n = 296 (29%)], headache [n = 291 (29%)], vomiting [n = 133 (13%)] and nausea [n = 113 (11%)]. There were no significant differences in the frequency of symptoms/signs between the different age groups or sexes.After treatment, 69 patients (7.1%) experienced at least one treatment emergent sign/symptom (TESS) which was either not present pre-treatment or worsened post-treatment: 54 patients suffered one TESS, 14 had two and 1 had three for a total of 85 TESSs: 36 were vomiting, 19 vertigo, 11 asthenia, 8 pruritus without a rash, 5 abdominal pain, 3 diarrhoea, 2 headaches and 1 nausea , were withdrawn from the study because of a TESS for an overall, crude withdrawal rate of 0.9%: 3.8% . All evFor the laboratory investigations, pretreatment results were available in 33%, 22% and 17% of patients for haematocrit (Hct), total white blood cells (WBC) and biochemistry, respectively Table . No CTC There were no shifts between Day 0 and Day 28 from grades 0 \u2013 2 to grades 3 or 4 Table . Out of There were no significant changes in mean values between Day 0 and Day 28 for the haematocrit  and total WBC  but there were significant decrease from Day 0 to Day 7  and increase from Day 7 to Day 28  for haematocrit. The mean AST, ALT and bilirubin decreased significantly while there was a small (0.16 mg/dL) but statistically significant increase in the mean creatinine values between Day 0 and Day 28 ; however, the latter induced more TESS and all the treatment withdrawals due to intolerance. The use of the co-blister did result in patients receiving doses of both drugs, in mg/kg, that were close to the recommended, weight based, mg/kg dosing regimen, though the co-blistered AQ mg/kg dose looked to be a little higher than the loose AQ and had a broader spread of the 95% CIs. Gastrointestinal complaints accounted for seven of the nine withdrawals and were probably AQ rather than AS related, given that AS has excellent tolerability. Five patients received AQ doses \u226520% than the weight based dose of 10 mg/kg and four were within the 15 mg/kg upper limit of the newly defined therapeutic window. Never the less, the AQ dose may have contributed to their gastro-intestinal complaints.The results obtained when patients are dosed more loosely by age are important because more practical regimens may expose patients to drug doses that are outside of the recommended, strictly defined, weight based doses. A more refined practical dosing has been developed for a new, aged dosed, fixed dose combination of AS+AQ . It is gNo cases of hepatitis or severe leukopenia (as a surrogate marker of neutropaenia) were detected, the two amodiaquine-associated toxicities that have caused fatalities in the past, when used as prophylaxis in travelers . HoweverThis study confirms that, in this area of moderate/intense transmission, the risk of clinical malaria is present throughout life and that children between 6\u201315 years of age are most affected. This is important as clinical studies in these areas normally enroll patients up to 10 years old, and thus would miss an important segment of the patient population. In this study, although treatment was statistically less efficacious in children under 11 years of age than in older patients, efficacy rates were still high in both, 95% and 97%, respectively.To conclude, this field study has shown a high and stable cure rate for AS+AQ even when dosed by age. Tolerability was good but continued intensive safety monitoring is still required in large numbers of patients for this ACT. Further research is planned to continue on the AS/AQ fixed dose combination.The author(s) declare that they have no competing interests.All authors read and approved the final manuscript.\u2022 P Brasseur was the Principal Investigator of the study. He contributed to the concept, protocol, analysis and reporting of the study, and contributed to the preparation of the manuscript. He personally contributed to the treatment, follow-up of patients and quality control of the study.\u2022 P Agnamey contributed personally to the treatment and follow-up of patients.\u2022 O Gaye participated in designing the concept and protocol of the study and supervised study conduct.\u2022 M Vaillant designed and conducted the analyses, contributed to the preparation of the manuscript.WRJ Taylor and P Olliaro contributed to the concept of the project; design of the protocol and analyses, reporting of the study, and to the preparation of the manuscript."}
+{"text": "Presumptive treatment for malaria is common in resource-limited settings, yet controversial given the imprecision of clinical diagnosis. We compared costs of diagnosis and drugs for two strategies: (1) empirical treatment of malaria via clinical diagnosis; and (2) empirical diagnosis followed by treatment only with Giemsa smear confirmation.Patients with a diagnosis of clinical malaria were recruited from a medical center in southwestern Nigeria. The patients received free Giemsa thick (diagnosis) and thin (differentiation) smears, but paid for any antimalarial treatment. Clinical diagnosis was made on clinicians' judgments based on symptoms, including fever, diarrhea, headache, and body-aches. The pediatric regimen was artesunate (6-9 tablets of 3mg/ kg on day one and 1.5mg/kg for the next four days) plus amodiaquine (10mg/kg day 1-2 and 5mg/kg on day three in suspension). Adults were given two treatment options: option one  and option two (4.5 50mg artesunate tablets on day one and nine for the next four days plus nine 200mg tablets of amodiaquine at a dose of 10mg/kg day 1-2 and 5mg/kg on day three). We calculated the costs of smears/drugs from standard medical center charges.Doctors diagnosed 304 patients (170 adults ages \u226516 years and 134 pediatric) with clinical malaria, prescribing antimalarial drugs to all. Giemsa thick smears were positive in 115/304 (38%). The typical patient cost for a Giemsa smear was 550 Naira (US$3.74 in 2009). For children, the cost of testing all, but treating only Giemsa positives was N888($6.04)/child; cost of empiric treatment of all who were clinically diagnosed was lower, N660($4.49)/child. For adults, the cost of testing all, but treating only Giemsa positives was N711($4.84)/adult for treatment option one (artesunate and sulfadoxine/ pyrimethamine) and N730($4.97)/adult for option two (artesunate and amodiaquine). This contrasts to lower costs of empiric treatment for both options one (N610=$4.14/adult) and two (N680=$4.63/adult).The most cost-effective approach to malarial management was empiric treatment, though many uninfected Nigerians will receive antimalarial drugs. If diagnostic costs declined, the cost-effectiveness calculus would change accordingly."}
+{"text": "Plasmodium falciparum and Plasmodium vivax malaria was assessed in Chumkiri, Kampot Province, Cambodia.Resistance to anti-malarial drugs hampers control efforts and increases the risk of morbidity and mortality from malaria. The efficacy of standard therapies for uncomplicated P. falciparum infected subjects, PCR genotyping of msp1, msp2, and glurp was used to distinguish treatment failures from new infections. Treatment failure rates at days 28 and 42 were analyzed using both per protocol and Kaplan-Meier survival analysis. Real Time PCR was used to measure the copy number of the pfmdr1 gene and standard 48-hour isotopic hypoxanthine incorporation assays were used to measure IC50 for anti-malarial drugs.One hundred fifty-one subjects with uncomplicated falciparum malaria received directly observed therapy with 12 mg/kg artesunate (over three days) and 25 mg/kg mefloquine, up to a maximum dose of 600 mg artesunate/1,000 mg mefloquine. One hundred nine subjects with uncomplicated vivax malaria received a total of 25 mg/kg chloroquine, up to a maximum dose of 1,500 mg, over three days. Subjects were followed for 42 days or until recurrent parasitaemia was observed. For P. falciparum infected subjects, 47.0% were still parasitemic on day 2 and 11.3% on day 3. The PCR corrected treatment failure rates determined by survival analysis at 28 and 42 days were 13.1% and 18.8%, respectively. Treatment failure was associated with increased pfmdr1 copy number, higher initial parasitaemia, higher mefloquine IC50, and longer time to parasite clearance. One P. falciparum isolate, from a treatment failure, had markedly elevated IC50 for both mefloquine (130 nM) and artesunate (6.7 nM). Among P. vivax infected subjects, 42.1% suffered recurrent P. vivax parasitaemia. None acquired new P. falciparum infection.Among 50 suggest that artesunate resistance may be emerging on a background of mefloquine resistance.The results suggest that artesunate-mefloquine combination therapy is beginning to fail in southern Cambodia and that resistance is not confined to the provinces at the Thai-Cambodian border. It is unclear whether the treatment failures are due solely to mefloquine resistance or to artesunate resistance as well. The findings of delayed clearance times and elevated artesunate IC Plasmodium falciparum has complicated efforts to control malaria, and can lead to unnecessary mortality if ineffective drugs remain the standard of care after drug-resistant strains become established  hypoxanthine , and the mixture (200 \u03bcl per well) was distributed into the 96-well test plates pre-coated with anti-malarial drugs. Each plate included two drug-free control wells and one control well without parasites. The plates were incubated for 48 h at 37\u00b0C in a 5% CO2 atmosphere and the cells then lysed by freeze-thawing. After collection on glass-fiber filter paper using a cell harvester, the amount of [3H] hypoxanthine incorporated into the parasites' nucleoprotein was determined using a Wallac MicroBeta Trilux counter . A log probit approximation was used to determine the 50% inhibitory concentration (IC50), defined as the concentration at which 50% of the incorporation of [3H] hypoxanthine was inhibited, as compared with the drug-free control wells.The in vitro drug sensitivity of the our test . In brieP. falciparum or P. vivax parasitaemia. Subjects who met World Health Organization criteria for early treatment failure  [P. falciparum malaria, the results of both per protocol and Kaplan-Meier analysis are reported. In the per protocol analysis the proportion of treatment failures was calculated by dividing the total number of subjects with recurrent parasitaemias (with or without PCR correction for re-infection) by the number of subjects who either suffered recurrent parasitaemia or completed the full 42 day follow-up period. In the Kaplan-Meier analysis, subjects were censored from the point at which they 1) were lost to follow-up or 2) acquired a P. vivax infection. The proportion of treatment failures for both day 28 and day 42 is reported. In the case of subjects with uncomplicated P. vivax malaria the analysis was identical, except that no attempt was made to distinguish between recrudescence, re-infection, and relapse from the liver. Subjects were considered to have cleared parasitaemia if there were at least two sequential negative smears. The day on which the first such negative smear was observed was defined as the day of clearance. Because smears were not taken on days 4\u20136, subjects with a reported clearance day 7 may actually have cleared their parasitaemia on any day between day 4 and 7. All statistical analysis was performed with StataSE 8.0.The primary outcome was recurrence of n day 3)  but who P. falciparum malaria were enrolled. One subject withdrew before completing treatment. Of the 150 who completed treatment, 127 (84.7%) still had P. falciparum parasitaemia on day 1, 71 (52.7%) on day 2, and 17 (11.3%) on day 3. All subjects were free of parasitaemia by day 7. Of these 150 subjects, seven (4.7%) were lost to follow-up before completing 42 days follow-up. Among the remaining 143 subjects, 31 (20.7%) developed recurrent P. falciparum parasitaemia. None of the recurrent parasitaemias occurred before day 14. Based on PCR genotyping for MSP1, MSP2, and GLURP, 4 of these parasitaemias were judged to be new infections and 27 to be recrudescences. Of the 27 recrudescences, 24 were accompanied by fever (axillary temperature \u2265 37.5\u00b0C) and were classified as late clinical failures; the remaining 3 were classified as late parasitological failures. Four subjects met criteria for early treatment failure[P. vivax parasitaemia, one on day 32 and seven on day 42. Figure The baseline characteristics of the study subjects are shown in Table P. vivax/P. falciparum infection. Of the remaining 109 subjects, 79 (72.5%) still had P. vivax parasitaemia on day 1, 5 (4.6%) on day 2, and 1 (0.9%) on day 3. All subjects were free of parasitaemia by day 7. Two subjects were lost to follow-up, at 28 and 25 days. Among the 107 subjects who completed follow-up, 45 (42.1%) suffered recurrent P. vivax parasitaemia; all recurrent parasitaemias occurred between day 28 and day 42. No attempt was made to distinguish between recrudescence, relapse from the liver, and re-infection. None of the P. vivax subjects acquired P. falciparum infection during follow-up.One hundred ten subjects with uncomplicated vivax malaria were enrolled. One subject was withdrawn from the study on day 1 when he was found to have a mixed 50 for mefloquine and artesunate in samples from only 51 P. falciparum subjects, 38 subjects who did not suffer a recrudescence and 13 who did. Figure 50 for mefloquine and artesunate for each of these samples. The mean mefloquine IC50 in subjects who suffered a recrudescence was 34 nM (95% CI 10\u201357) higher than in those who did not, 90 nM (95% CI 65\u2013115 nM) versus 56 nM (95% CI 45\u201367 nM). The mean artesunate IC50 was also higher in subjects who suffered recrudescence, 1.7 nM (95% CI 0.7\u20132.7 nM), than in those who did not, 1.2 nM (95%CI 1.0\u20131.5 nM), but the difference was not statistically significant.For logistical reasons, it was possible to measure the ICpfmdr1 gene has been associated with mefloquine resistance [pfmdr1 copy number and the proportion of samples with estimated copy number > 1.5 in the samples from day 0 and from the day of recrudescence. Samples from the day 0 parasitaemia in subjects who ultimately suffered a recrudescence had a higher mean pfmdr1 copy number than those from subjects who did not . The relative risk for recrudescence in subjects with pfmdr1 copy number > 1.5 was 3.5 (95% CI 1.4\u20138.8). Recrudescent samples had a higher mean copy number than the corresponding day 0 samples .Amplification of the sistance . Table 3pfmdr1 gene, and the initial parasite density. Longer time to parasite clearance was associated with greater risk of subsequent recrudescence, a greater proportion of pfmdr1 amplification, and higher initial parasitaemia. Table pfmdr1 copy number were independently associated with higher risk of recrudescence.Table An unexpectedly high failure rate for artesunate-mefloquine treatment of uncomplicated falciparum malaria was found at a site in Cambodia far from the Thai-Cambodian border, frequently considered a hot spot for the emergence of anti-malarial drug resistance. The failure rates at 28 and 42 days, calculated using Kaplan-Meier survival analysis and PCR correction for re-infection, were 13.1% (95% CI 8.5\u201319.7%) and 18.8% (95% CI 13.3\u201326.2%), respectively. Although it was not possible to measure circulating drug levels during therapy, all doses were directly observed, and the drugs were obtained through the Cambodian Ministry of Health from suppliers following current Good Manufacturing Practices. Thus, while it is formally possible that the unexpectedly high failure rates are due to adulterated, defective, or counterfeit anti-malarials, it seems unlikely. The study used the then current maximum dose of mefloquine, 1,000 mg. It is possible that efficacy would have been higher had we used a maximum of 1,250 or 1,500 mg. If treatment failures were related to a low dose/weight of mefloquine, it would have been expected that heavier subjects would be at greater risk of failure. The mean weight, however, of subjects who had an ACPR , was not significantly different from those who did not ; similarly the mean administered mefloquine dose per kg did not differ significantly between ACPR and failure . The risk of failure was the same in subjects < 40 kg, 40\u201350 kg, or > 50 kg, and in regression analysis there was no relationship between either weight or actual mefloquine dose per kg and time to clearance or risk of recrudescence. It therefore seems unlikely that increasing the maximum mefloquine dose would have significantly changed the observed failure rates.P. vivax malaria and treated with chloroquine acquired new P. falciparum infection during the 42 day follow-up period, suggesting that the P. falciparum attack rate is relatively low. PCR genotyping may miss minor parasite populations present on day 0 which may be selected during treatment, thus incorrectly characterizing treatment failures as new infections. That such selection occurred is suggested by the observation  and 21.4% (95% CI 15.5\u201329.0%), respectively.It is possible that PCR correction in fact led to an underestimation of the failure rate. Malaria transmission is not intense at this study site; most infections (> 80%) contained only a single genotype (data not shown). None of the 109 subjects enrolled with uncomplicated on Table  that theUse of non-study anti-malarial drugs during follow-up might also lower the measured treatment failure rate. Although subjects were instructed not to take anti-malarial drugs during the follow-up period, except those provided as treatment for documented recurrences by the study team, and although none reported having done so, we cannot exclude the possibility that some subjects may have taken non-study anti-malarial drugs. Such unrecorded treatment might have reduced the apparent treatment failure rate. In that case the failure rates reported herein would be an underestimate.P. falciparum has a high risk of late recrudescence [pfmdr1 gene and elevated mefloquine IC50 were predictive of treatment failure. It may be that all the observed treatment failures are due solely to mefloquine resistance. There was, however, some evidence suggestive of resistance to artesunate. First, pure mefloquine resistance should not greatly affect the time to parasite clearance in subjects treated with artesunate-mefloquine combination therapy. Trials of artesunate monotherapy [P. falciparum infections. Similarly, in a study carried out in northern Lao in 2003 [50 values were not strongly predictive of treatment failure, one sample with markedly elevated IC50 for both artesunate (6.7 nM) and mefloquine (130 nM) was identified. It thus appears possible that, at this study site, artesunate resistance is beginning to emerge on a background of pre-existing mefloquine resistance. If recent reports of declining efficacy of artesunate-mefloquine in Trat, Thailand [It is not clear whether the resistance observed here is due entirely to resistance to mefloquine, or whether there is also a contribution of resistance to artesunate. It is well known that a 3 day regimen of artesunate monotherapy for descence ,18. It iotherapy ,20 and aotherapy  have genotherapy , treatme in 2003 , the sam in 2003 , even ar in 2003 ,20. In cThailand , and PaiP. vivax parasitaemia by day 42  was found. All of the recurrent parasitaemias occurred after day 28 and all but 2 after day 35. Given the late timing of the recurrences, the absence of radical treatment to eliminate hypnozoites from the liver, and the lack of blood chloroquine levels on the day of recurrence, it is impossible to draw any conclusions about chloroquine resistance. It is clear, however, that subjects with vivax malaria treated with chloroquine alone are at substantial risk of recurrent morbidity; further studies including primaquine treatment and measurement of drug levels will be required to determine whether chloroquine resistance, reported elsewhere in south-east Asia, is a problem in Cambodia.Current Cambodian Ministry of Health treatment guidelines do not include primaquine treatment for vivax malaria because of the difficulty in screening for G6PD deficiency before treatment. It is not surprising that frequent recurrent The authors declare that they have no competing interests.pfmdr1 copy number assays and performed quality control on the data; TT examined and performed follow-up of study subjects after treatment; PC participated in the in vitro drug resistance assays; PL participated in the in vitro drug resistance assays; SM participated in study design; DS participated in study design; FA oversaw the in vitro drug resistance and pfmdr1 copy number assays and participated in data analysis; CW conceived the study, participated in study design, and performed quality control of data. All authors contributed to the critical review of the manuscript and agree to submission.WOR participated in study design and data collection, analyzed the data and drafted the manuscript; RS performed the"}
+{"text": "With the availability of new preventive and curative interventions, global malaria control has been strengthened significantly in recent years. Drugs effective in reducing malaria gametocytaemia might contribute to local elimination and possible long-term eradication. We here report on the effects of methylene blue (MB)-based malaria combination therapy on gametocytaemia during a randomised-controlled trial in Burkina Faso.An open-label randomised controlled phase II study in 180 children aged 6\u201310 years with uncomplicated falciparum malaria was conducted in Nouna, north-western Burkina Faso. Children were randomised to MB\u2013artesunate (AS), MB\u2013amodiaquine (AQ), and AS-AQ . Overall follow-up was for 28 days, follow-up for gametocytaemia was for 14 days.P. falciparum gametocytaemia at baseline.The treatment groups were similar in baseline characteristics and there was only one loss to follow-up. Compared to AS-AQ, both MB-containing regimens were associated with significantly reduced gametocyte carrier rates during follow-up days 3, 7, and 14. This effect was seen both in patients with and without P. falciparum gametocytes. MB-based combination therapy thus has the potential to reduce transmission of P. falciparum malaria in endemic regions, which has important implications for future elimination and eradication strategies.MB reveals pronounced gametocytocidal activity which appears to act against both existing and developing NCT00354380ClinicalTrials.gov  Malaria remains the most important parasitic disease globally, and in Sub-Saharan Africa (SSA) in particular P. falciparum disulfide reductases and in addition inhibits the parasite's heme detoxification Gametocytocidal antimalarials, and the artemisinines in particular, may accelerate such progress in that they reduce post-treatment transmission The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Centre de Recherche en Sant\u00e9 de Nouna (CRSN) in Nouna Health District, north-western Burkina Faso. The area is highly endemic for malaria with most clinical cases occurring during or briefly after the rainy season which lasts from June until October The study was conducted in October/November 2006 in the urban research zone of the The study was an open label randomized controlled phase II trial on safety and efficacy of MB-artesunate (AS) and MB-amodiaquine (AQ) in children with uncomplicated falciparum malaria, with blinding only for the laboratory technicians involved. The primary objective was to investigate the safety of the combinations MB-AS and MB-AQ in children with uncomplicated falciparum malaria. The secondary objective was to determine the efficacy of these MB-based combinations in the treatment of children with uncomplicated falciparum malaria. The study was designed to have a statistical power of 80% in order to detect a difference in the number of adverse events of at least 20% among study groups that was significant at the five percent level Post-treatment gametocyte carriage was initially not planned to examine and specified later as an additional secondary endpoint. The methodology of the trial has been published elsewhere P. falciparum asexual parasites per \u00b5L blood), and written informed consent given by the parents/caretakers. Exclusion criteria were signs of severe malaria, any apparent other disease, and malaria treatment \u2013 except CQ \u2013 with western drugs and/or antibiotics with antimalarial potency during the preceding week.Only children from Nouna town were participating in this study. Inclusion criteria were: age 6\u201310 years, ability to swallow tablets, uncomplicated falciparum malaria  was given at a dose of 10 mg per kilogram of body weight twice daily over three days, AS  at a dose of 4 mg per kilogram of body weight once daily over three days, and AQ  at a dose of 10 mg per kilogram of body weight once daily over three days. Children with fever \u226538.5\u00b0C received a standard dose of 10 mg per kilogram paracetamol tablets every 6 hours  until symptoms subsided.Follow-up of study children was for 28 days using a slightly modified version of the latest WHO protocol on antimalarial drug efficacy testing A finger-prick blood sample was taken on days 0, 2, 3, 7, 14, and 28, and during unscheduled visits. From this, malaria parasitaemia and haematocrit values were determined using standard CRSN procedures P. falciparum gametocytaemia prevalence and density, both in patients with ACPR and in patients without ACPR. In view of the short half-life of both AS and MB, these drugs would have no effect on new infections emerging after day 14. Thus, gametocytaemia occurring after day 14 was not considered.In early 2008, all slides from the trial (only day 0 until day 14) were systematically re-examined by two experienced laboratory technicians supervised by one of the investigators (BC) for The effects of different treatment arms on gametocytaemia were determined using per protocol analysis. The Chi square test (Chi) was used to compare proportions, and the non-parametric Wilcoxon-Mann-Whitney test (WMW) to compare metric or ordinal data. In order to use the closed testing procedure for multiple testing adjustment when comparing three groups, a global test is needed which compares three groups for differences. This is done by an exact chisquare test which is also valid for small sample sizes. When possible, estimates and the corresponding 95% confidence interval are given. Calculations were performed with the program SAS release 9.1 . Multiple testing between single arms in a three-armed trial is handeled by the closed testing procedure: If a difference between all three arms is assessed by a global test on significance level 5%, it is possible to use the same level for the three two group comparisons. This procedure keeps the family wise error rate.The study protocol was approved by the Ethics Committee of the Medical Faculty at Heidelberg University and the local Ethics Committee of Burkina Faso.One hundred and eighty children  were included in the study . There wThe prevalence of gametocytes declined following treatment, regardless of the drug administered. However, both MB-AS and MB-AQ were significantly more effective in this regard than AS-AQ during follow-up . No gameSimilar findings were observed among patients who exhibited gametocytaemia at enrolment .P. falciparum gametocytes on day 0 and/or day 2, both MB-based treatment regimes yielded lower P. falciparum gametocyte prevalence on day 3 and day 7 compared to AS-AQ  gametocytes and newly developing (younger) ones. However, due to the limited sample size these observations are based on small and not always statistically significant numbers and need to be interpreted with some caution. Moreover, P. falciparum gametocytaemia was not included as a specific endpoint a priori. This does not change the reliability of our findings as all slides from the trial were safely stored in the CRSN laboratory and re-examined by blinded laboratory technicians. Measuring the effects of the different treatments on submicroscopic gametocytaemia could have been another option to increase the power of this study. For instance, in a recent trial in Tanzanian children, gametocytes were detected in 23% and 89% by microscopy and real-time PCR assays, respectively Treatment of malaria with MB-based regimens not only is more effective than AQ-AS in this part of Burkina Faso P. falciparum gametocytes except on the more mature stages of their development ACT has been shown to be highly effective against P. falciparum gametocytes has long been known. In Sudanese children in the early 1930s, plasmoquine given prophylactically at a dosage of 0.02 mg/kg twice weekly, i.e. at a fraction of the therapeutical dose, almost completely prevented the occurrence of gametocytes plus AS reduced gametocyte carriage by more than 80% as compared to SP-AS alone P. falciparum are not affected by therapeutical doses of primaquine, and a reduced gametocytocidal activity has been reported from India P. falciparum treatment, intended to block post-treatment transmission. Yet, MB shows promising features in this regard including strong activity against asexual parasites, developing and mature gametocytes, as well as synergism with artemisinine derivatives The mechanism by which MB acts against gametocytes is obscure so far. Rapid elimination of asexual parasites (including fractions committed to gametocytogenesis) by MB-based combination treatment may be involved in preventing subsequent gametocytaemia. However, MB-AQ exhibited superior gametocytocidal effects but slower clearance of asexual parasites as compared to AQ-AS P. falciparum gametocytes. A combination of MB with an artemisinin derivate may thus maximise the gametocytocidal effects of the latter. These findings add to already existing in vitro and in vivo evidence of a synergy between MB and artemisinins P. falciparum parasites.In summary, this study provides evidence that suggests a high efficacy of MB-based combination therapy against Checklist S1CONSORT Checklist(0.06 MB DOC)Click here for additional data file.Protocol S1Trial Protocol(0.20 MB DOC)Click here for additional data file."}
+{"text": "The objective of this study was to determine the appropriate dose of artesunate for use in a fixed dose combination therapy with chlorproguanil\u2212dapsone (CPG\u2212DDS) for the treatment of uncomplicated falciparum malaria.Plasmodium falciparum malaria. Subjects were randomized into 4 groups to receive CPG\u2013DDS alone or plus 4, 2 or 1 mg/kg of artesunate once daily for 3 days. Assessments took place on Days 0\u22123 in hospital and follow-up on Days 7 and 14 as out-patients. Efficacy was evaluated in the Day 3 per-protocol (PP) population using mean time to reduce baseline parasitemia by 90% (PC90). A number of secondary outcomes were also included. Appropriate artesunate dose was determined using a pre-defined decision matrix based on primary and secondary outcomes. Treatment emergent adverse events were recorded from clinical assessments and blood parameters. Safety was evaluated in the intent to treat (ITT) population.Open-label clinical trial comparing CPG\u2212DDS alone or with artesunate 4, 2, or 1 mg/kg at medical centers in Blantyre, Malawi and Farafenni, The Gambia. The trial was conducted between June 2002 and February 2005, including 116 adults (median age 27 years) and 107 children (median age 38 months) with acute uncomplicated In the Day 3 PP population for the adult group (N\u200a=\u200a85), mean time to PC90 was 19.1 h in the CPG\u2212DDS group, significantly longer than for the +artesunate 1 mg/kg , 2 mg/kg  and 4 mg/kg  groups. For children in the Day 3 PP population (N\u200a=\u200a92), mean time to PC90 was 21.1 h in the CPG\u2212DDS group, similar to the +artesunate 1 mg/kg group , though the +artesunate 2 mg/kg and 4 mg/kg groups had significantly shorter mean times to PC90 versus CPG\u2212DDS; 14.4 h  and 12.8 h , respectively. An analysis of mean time to PC90 for the Day 14 PP and ITT populations was consistent with the primary analysis. Treatment emergent, drug-related adverse events were experienced in 35.3% (41/116) of adults and 70.1% (75/107) of children; mostly hematological and gastroenterological. The nature and incidence of adverse events was similar between the groups. No dose-related changes in laboratory parameters were observed. Nine serious adverse events due to any cause occurred in five subjects including two cases of hemolysis believed to be associated with drug treatment . One adult died of anaphylactic shock, not associated with investigational therapy.CPG\u2013DDS plus artesunate demonstrated advantages over CPG\u2013DDS alone for the primary efficacy endpoint (mean time to PC90) except in children for the 1 mg/kg artesunate dose. Based on a pre-defined decision matrix, the primary endpoint in the child group supported an artesunate dose of 4 mg/kg. Secondary endpoints also supported a 4 mg/kg artesunate dose to take forward into the remainder of the development program.NCT00519467ClinicalTrials.gov  Plasmodium spp. has undermined the effectiveness of many of the antimalarials used commonly, particularly chloroquine Over one million people die of malaria every year, mostly children P. falciparumIn consideration of these points, in 2004 the World Health Organization (WHO) Roll Back Malaria group recommended that first-line treatment of uncomplicated malaria should be with artemisinin-based combination therapy Plasmodium falciparumP. falciparum compared with the more slowly eliminated combination sulfadoxine-pyrimethamine with no increase in the incidence of retreatment Chlorproguanil\u2212dapsone  was developed through a public\u2212private partnership as a low-cost, fixed dose combination for the treatment of uncomplicated malaria due to A clinical program to develop CPG\u2212DDS\u2212artesunate (CDA) is currently in progress. CDA is an interesting combination for investigation as the three compounds have well-matched pharmacokinetics, i.e. all are relatively rapidly eliminated. Thus, there is a reduced risk of parasite exposure to any single compound after elimination of the partner drug, potentially decreasing the risk of resistance emergence.This study was a phase II trial to determine the most appropriate dose of artesunate for use in combination with CPG\u2013DDS. Four dosing groups were included: CPG\u2013DDS alone or with 4, 2, or 1 mg/kg artesunate. The WHO-defined \u2018adequate clinical and parasitological response\u2019 with CPG\u2013DDS alone has been reported as 96% in African children at 14-days' follow up As artesunate rapidly clears parasites from the blood, the protocol development committee concluded that parasitemia-derived indices of therapeutic response, in particular time to reduce parasitemia by 90%, would be the most relevant alternative outcomes to detect differences between the artesunate dose groups and CPG\u2212DDS alone. A suite of other outcomes was also included. Children are the primary population of interest because of the higher mortality risk compared with adults This novel study design was a pragmatic approach to evaluating the possible differential effect of adding artesunate at three doses to CPG\u2212DDS in children and adults. The aim of the trial was to determine the dose of artesunate that would be included in the CDA fixed dose combination for further testing in Phase III studies.This study was conducted in accordance with the International Conference on Harmonization Good Clinical Practice Guidelines and all applicable regulatory requirements and the guiding principles of the Declaration of Helsinki. Participants were recruited under a human use protocol approved by and executed in accordance with the ethics committees of the Malawi College of Medicine, the Joint Gambian Government/Medical Research Council and the Liverpool School of Tropical Medicine. The CONSORT checklist and trial protocol for this paper are available as supporting information; see P. falciparum asexual parasitemia, based on microscopic screening of stained blood films at a density of 10,000\u2212100,000 \u00b5L\u22121 in adults and 25,000\u2212100,000 \u00b5L\u22121 in children. All patients or their parent/guardian provided written or oral witnessed consent and agreed to comply with the requirements of the protocol. A negative pregnancy test was required for all women \u226512 years of age on enrolment.Eligible participants included adults aged 18\u221260 years or children aged 12\u2212120 months presenting to a healthcare facility with probable uncomplicated clinical malaria. Inclusion criteria were: weight between 5 and 85 kg; and presence of P. ovale and P. malariae parasitemia) or any other underlying disease that would compromise the diagnosis and the evaluation of the response to the study medication . Women who had a positive pregnancy test, did not take a pregnancy test or were breastfeeding during the study were also excluded.Subjects were excluded from the study if they had features of severe or complicated falciparum malaria, a known allergy to sulphonamides, evidence of any concomitant infection at the time of presentation , atovaquone\u2212proguanil, artemisinins, co-artemether, tetracycline or clindamycin within 7 days prior to screening; or treatment for 5 half-lives prior to screening with drugs that have potential antimalarial activity. In addition, subjects could not have used an investigational drug within 30 days or 5 half-lives (whichever was longer) prior to screening, or participated previously in the current study.A full history and examination was performed at the initial patient assessment. Study participants who fulfilled the inclusion/exclusion criteria were randomized to one of four treatment groups: CPG\u2013DDS alone; CPG\u2212DDS plus 4 mg/kg artesunate; CPG\u2212DDS plus 2 mg/kg artesunate or; CPG\u2212DDS plus 1 mg/kg artesunate. CPG\u2212DDS was provided as Lapdap\u2122 15/18.75 mg or 80/100 mg tablets  and patients were given half, one, one and a half or two tablets to achieve a target dose as close as possible to 2 mg/kg CPG and 2.5 mg/kg DDS. Artesunate was provided as 1 mg, 10 mg, 25 mg or 50 mg tablets . All drug treatments were administered according to predefined dosing charts.All patients received study drug once daily for 3 days , during which time they were hospitalized for study-related procedures. Patients were discharged from hospital on Day 3 and asked to return for outpatient follow-up visits on Days 7 and 14.Blood samples for parasitemia blood slide (10 \u00b5L) were obtained from adults via cannula for the first 24 h and by venepuncture thereafter and from children using finger-prick. In adults, sampling was performed on Day 0 at the time of first drug dose then at 1, 2, 3, 4, 6, 8, 12, 18, 24, 30, 36, 42, 48 and 72 h during therapy. In children, blood samples were collected at the time of first dose and at 6, 12, 18, 24, 36, 42, 48 and 72 h during therapy. Samples were also collected from all patients at the follow up visits on Days 7 and 14.For other outcomes, a 100 \u00b5L blood sample was taken at Day 0 at the time of first study dose in adults and children then at 1, 2, 4, 8, 12 and 24 h during therapy in adults and at 12 h only in children. The 12-h sample was the key timepoint used for the parasite viability assessment.Samples for biochemical and hematological analysis (2 mL of blood), including reticulocyte analysis, were taken on Day 0, Day 3 and Day 7 from all patients and on Day 14 if any value was abnormal on Day 7. Samples were analyzed locally at each investigational center.P. falciparum malaria.The objective of this trial was to determine the most appropriate dose of artesunate to use in combination with CPG\u2013DDS for the treatment of uncomplicated \u22121, according to WHO protocol WHO/HTM/RBM/2003.50 The primary endpoint was the mean time to achieve a 90% reduction in parasitemia versus baseline (PC90), determined from Giemsa-stained blood smears. The quality-control procedure for conducting the slide counts involved checking every seventh or tenth slide and eliciting an additional expert reading in the event of significant discrepancies. Screening parasitemia density was calculated using the nominal white blood cell count (WBC) value of 8,000 \u00b5Lin vivo. It was determined using the method of Murphy, et al. Parasite viability was considered a key secondary endpoint; defined as the percentage of ring-form parasites obtained from the patient 12 h after the first dose of study drug that developed into trophozoites when cultured The mean time to achieve a 99% reduction (PC99) and 50% reduction (PC50) in parasitemia versus baseline were also included as secondary outcomes and calculated using similar methods as mean time to PC90. Slides prepared for parasitology were also read for the presence or absence of gametocytes per 200 white blood cells at baseline and Days 3, 7 and 14.msp1 and msp2 polymerase chain reaction genotyping The number of early or late treatment failures based on the WHO 2002 definitions was also determined The most appropriate dose of artesunate was determined according to a pre-defined decision matrix, summarized as:Choose 2 mg/kg artesunate if all artesunate groups have efficacy greater than CPG\u2013DDS alone and there is no difference in the primary endpoint or any other difference in efficacy or safety between the artesunate treatment groups versus CPG\u2212DDS alone.Choose 4 mg/kg artesunate if 1 mg/kg or 2 mg/kg do not have greater efficacy than CPG\u2212DDS alone for the primary endpoint or if there is a difference in the primary endpoint or any other efficacy or safety endpoint between the artesunate 4 mg/kg and the 1 or 2 mg/kg groups versus CPG\u2212DDS alone (in favor of the 4 mg/kg artesunate dose).In addition, if data from the clinically more vulnerable child group suggested a higher artesunate dose than data from adults, then the higher dose would be chosen. The rationale for the design of the outcome decision matrix can be found in the Discussion section of this paper.per se except for the development of clinically severe malaria, including a parasite count of >250,000 \u00b5L\u22121 and severe anemia (hemoglobin <5.0 g/dL).Adverse events and severity  were determined by the investigators and defined as any untoward medical occurrence temporally associated with the use of study drug, whether or not considered to be related to the study drug. Adverse events were recorded and coded using the latest version of MedDRA. Lack of efficacy was not included as an adverse event A serious adverse event was defined as one which resulted in death, was life-threatening, resulted in hospitalization or prolongation of hospitalization, was disabling, a congenital anomaly or any other event considered significant by the investigator. The investigator was to follow all serious adverse events until resolution, the condition stabilized, the event was otherwise explained or the subject was lost to follow-up.A sample size of eighty-eight evaluable subjects (22 per treatment group) would provide 90% power at a 5% two-sided significance level to detect a difference of 9 h in the mean time to PC90 between the groups treated with CPG\u2212DDS plus 4, 2, or 1 mg/kg of artesunate versus CPG\u2013DDS alone. Allowing for a 25% drop-out rate, a target sample size of 120 adults and 120 children was calculated.The adult group was stratified by gender to prevent a bias. The child group was stratified by age to ensure that both younger and older children were represented.The randomization code was generated by computer at GlaxoSmithKline, Greenford, UK. Different codes were allocated for adult sex strata and child age strata and distributed using scratch cards bearing unique study numbers.This was an open-label study.Scratch cards were provided to clinicians and were selected consecutively on patient enrolment to reveal treatment allocation.In order to achieve accurate artesunate dosing, this study was conducted open label to the treating physician and nursing staff. However, the technicians performing the slide reading and parasite viability analysis were blinded to the treatment group. Subjects assigned to receive CPG\u2212DDS alone did not receive placebo to replace artesunate.\u22121 for adults and \u226512,500 \u00b5L\u22121 for children.The intent to treat (ITT) population included all randomized subjects who received at least one dose of study medication. The per-protocol (PP) population was a sub-set of the ITT population including those patients who were compliant with all doses of study medication, had no major protocol violations, took no prohibited concomitant medications during the treatment period and had a baseline actual parasitemia level of \u22655,000 \u00b5LThe primary analysis was conducted on the Day 3 PP population who had completed all visits through to the assessment at Day 3. Additional sensitivity analyses were prepared using the ITT population and the Day 14 PP population; a sub-set of the Day 3 PP population including patients that completed all visits during the study.The null hypothesis was that there would be no difference in the primary endpoint (mean time to PC90) between CPG\u2212DDS alone and CPG\u2212DDS plus artesunate at each of the three doses.\u2212\u03b2(x\u2212\u03bc)) where \u03b1\u200a=\u200athe lower asymptote, \u03b1+\u03bb\u200a=\u200athe upper asymptote, \u03b2\u200a=\u200athe parasitemia reduction rate, \u00b5\u200a=\u200athe time of maximum rate of reduction (i.e. point of inflexion), x\u200a=\u200atime from first dose (in hours); y\u200a=\u200alog[1+P(time\u200a=\u200ax)] and (P\u200a=\u200aparasite count), hence P\u200a=\u200aey\u20131. The model of best fit was selected for each subject by two independent statisticians blinded to treatment group based on a set of pre-defined evaluation criteria: possible data errors; outliers affecting model fit and whether to exclude them; estimated modeled baseline and fit of model in baseline region; estimate of PC90 and fit in these regions; and closeness of model fit with observed data. Any subjects where the logistic curve fitted to derive time to PC90 was not reliable were omitted in order to preserve the integrity of the analysis, termed \u2018poor model fit\u2019. Poor model fit was categorized as: the model being inadequate to fit to the data; there being insufficient data to fit to the model; or the subject not achieving PC90 by 72 h based on the model. The mean time to PC90 was determined from the derived times to PC90s of the available subjects.The treatment difference was calculated from the mean values of time to PC90. The principal analysis of mean time to PC90 was performed for the subjects with confirmed 90% reduction of parasitemia by 72 h based on a logistic model. This approach was taken as it was not feasible to draw blood samples more frequently and yet it was desirable to try to determine the time to PC90 that likely occurs between sampling time points. For each subject, observed parasite count data from 0\u221272 h were log-transformed. A logistic model was used to estimate the change in parasite count between observations. The logistic curve had the following form: y\u200a=\u200a\u03b1+\u03bb/ (children were enrolled at one center only).No adjustment for the multiple comparisons to the CPG\u2013DDS alone arm was made. However, in order to preserve the overall significance level at 5%, a step-down closed testing procedure was used whereby the 4 mg/kg artesunate group was compared to CPG\u2212DDS alone and only if there was a statistically significant difference at the 5% level was the 2 mg/kg compared with CPG\u2212DDS alone; only if this showed a significant difference was the 1 mg/kg dose compared with CPG\u2212DDS alone.The study was powered for the primary analysis. Additional analyses were presented for descriptive purposes, and no further adjustments for multiplicity have been implemented. Summary statistics  for parasite counts were tabulated by treatment group for adults and children. The proportion of subjects with gametocytes at each timepoint was also summarized along with 95% confidence intervals. Analyses of the secondary endpoints of mean time to PC50 and mean time to PC99 were conducted in a similar manner to those performed for PC90.The ITT population was used for safety analyses. Safety analyses included summaries of the proportion of patients with adverse events and serious adverse events by treatment group. For laboratory parameters, summary statistics  for each timepoint were presented by treatment group. In addition, the proportion of subjects with decreases in hemoglobin (\u22652 g/dL and \u22654 g/dL) was tabulated by treatment group.Participant flow for the adult and child groups is shown in There were 116 adults enrolled in the study. The Malawi centers contributed 62 patients; 14 were randomized to CPG\u2212DDS alone, and 17, 17 and 14 to +artesunate 1, 2 or 4 mg/kg, respectively. The Gambia center contributed 54 adult patients, 15 randomized to CPG\u2212DDS alone and 13 patients each randomized to +artesunate 1, 2 or 4 mg/kg. All of the 107 children enrolled into the study were recruited from the Malawi center.All randomized subjects were included in the ITT population. Reasons for withdrawal from the study are shown in The trial was conducted between June 2002 and October 2004 at the Queen Elizabeth Central Hospital, Blantyre and six local health centers in Malawi and between September 2004 and February 2005 at the AFPRC General Hospital, Farafenni, The Gambia.The baseline demographic and clinical characteristics of the Day 3 PP population are shown in For adults, the ratio of females to males was similar in all treatment groups except the CPG\u2013DDS +artesunate 4 mg/kg group, where the ratio was 6:12. These results were consistent with the 2:3 adult female:male randomization .For children, the median age in the CPG\u2013DDS +artesunate 1 mg/kg group was 46 months, compared with 29\u221241 months in the other groups. The ratio of females to males was similar in all treatment groups except the CPG\u2013DDS +artesunate 2 mg/kg group, where the ratio was 6\u223617 .The number of patients analyzed in the Day 3 PP population and the Day 14 PP population are shown in In addition, six patients were excluded from the principal analysis due to \u2018poor model fit\u2019. These exclusions were decided by statisticians blind to treatment group. Two adult patients (one in the CPG\u2212DDS and one in the +artesunate 2 mg/kg group) and four children  were excluded as the model was inadequate to fit their data. No patients were excluded due to not achieving PC90 by 72 h. Patients excluded from the principal analysis were included in the sensitivity analysis.P\u200a=\u200a0.002), 2 mg/kg  and 1 mg/kg  groups  groups , Table 2P\u200a=\u200a0.010) and the 2 mg/kg  groups (P\u200a=\u200a0.222)  groups , Table 2P\u200a=\u200a0.057)  . An anal=\u200a0.057) .P\u200a=\u200a0.024) and 2 mg/kg  groups versus CPG\u2013DSS alone (P\u200a=\u200a0.104).In adults at 12 h after the first dose of study drug, mean parasite viability was 12.4% in the CPG\u2212DDS group. There was a significant decrease in viability observed for the +artesunate 4 mg/kg , \u221268.1% (P\u200a=\u200a0.0002) and \u221257.3% (P\u200a=\u200a0.0021) for the +artesunate 4, 2, and 1 mg/kg groups, respectively  and \u22125.9 h (P\u200a=\u200a0.012), respectively (P\u200a=\u200a0.054).In adults, for the Day 3 per-protocol population, the mean time to PC50 for CPG\u2212DDS alone was 13.3 h (SD\u00b110.8 h). Statistical analysis showed that in adults, addition of artesunate 4 mg/kg or 2 mg/kg resulted in a significant reduction in mean time to PC50 versus CPG\u2212DDS alone; treatment difference \u22125.4 h (ectively . There wP\u200a=\u200a0.001) and \u22127.4 h (P\u200a=\u200a0.005), respectively (P\u200a=\u200a0.089).In children, the mean time to PC50 was 17.0 h (SD\u00b19.0) for CPG\u2212DDS alone. As for adults, in children the +artesunate 4 mg/kg and +artesunate 2 mg/kg groups had a significantly shorter mean time to PC50 than CPG\u2212DDS alone; treatment difference \u22129.0 h (ectively . There wP\u200a=\u200a<0.001), 2 mg/kg  and 1 mg/kg  versus CPG\u2212DDS alone.The mean time to PC99 in adults with CPG\u2212DDS alone was 25.0 h (SD\u00b111.5) . There wP\u200a=\u200a0.018).The mean time to PC99 in children with CPG\u2212DDS alone was 24.5 h (SD\u00b19.8) . In contIn adults, the percentage of gametocytemic patients in the CPG\u2013DSS alone group increased during the study to a maximum of 29.4% of patients assessed at Day 14 . In the In children, the percentage of gametocytemic patients in the CPG\u2013DDS group increased during the study to a maximum of 47.1% at Day 14 . In the The trial was underpowered for statistical analysis of clinical endpoints. Using the modified WHO 2002 definitions Owing to the wide confidence intervals, there were no clear differences in treatment failures observed between the artesunate dose groups and CPG\u2013DDS alone in either adults or children or between adults and children .The number of patients in the ITT population in whom the initial dose or re-dose of study drug(s) was successful on all 3 days (administration compliance) was 94.2% (210/223). For adults, administration compliance was 89.7%, 96.7%, 90.0%, 96.3% for the CPG\u2212DDS only, and +artesunate 1, 2 and 4 mg/kg groups, respectively; corresponding numbers for children were 95.8%, 96.6%, 100% and 89.3%, respectively.and they received the correct mg/kg/day target doses of study drug as per the randomization schedule. For adults, 89.7%, 96.7%, 86.7% and 96.3% of patients were totally compliant in the CPG\u2212DDS only, and +artesunate 1, 2 and 4 mg/kg groups, respectively; corresponding numbers for children were 95.8%, 93.1%, 96.2% and 82.1%, respectively.Overall, in the ITT population, 91.9% (205/223) of subjects were totally compliant, i.e. the initial dose or re-dose of study drug(s) was successful on all 3 days This study was not powered to conduct formal comparisons of safety between treatment arms. The observed nature and incidence of adverse events were similar between the groups  and wereThere were some differences in the reporting of adverse events by center (for adults). In Malawi, adverse events were recorded for 11/14 (78.6%) of adults receiving CPG\u2212DDS alone and 11/17 (64.7%), 12/17 (70.6%) and 11/14 (78.6%) for those receiving 1, 2 or 4 mg/kg of artesunate respectively. In The Gambia, the number of adverse events reported was much lower than in Malawi: 7/15 (46.6%) patients for CPG\u2212DDS alone and 5/13 (38.5%), 3/13 (23.1%) and 4/13 (30.7%) for those receiving 1, 2 or 4 mg/kg of artesunate respectively. However, in both centers there were no apparent differences in the nature or incidence of adverse events between the CPG\u2212DDS alone group and the artesunate groups.Two non-fatal treatment-emergent serious adverse events were observed in two adults (one each in the +artesunate 4 and 2 mg/kg groups) and six were observed in three children (two in the CPG\u2212DDS group and one in the +artesunate 4 mg/kg group). In addition, one child experienced vomiting before and after dosing with study treatment and required an additional night in hospital. Six of the serious adverse events were not thought to be related to study medication and included one case of sepsis in an adult patient and in the child group two cases of sepsis and one each of malnutrition, malaria and cerebral malaria.Two of the serious adverse events were thought to be related to study drug. One adult case of severe hemolytic anemia in the +artesunate 2 mg/kg group presented at Day 9 with a hemoglobin concentration of 6.6 g/dL (baseline 15.4 g/dL), was treated with two units of blood and the patient recovered. However, this patient subsequently went on to experience anaphylactic shock (day 14 after recruitment) which resulted in death. The investigator considered this event to be related to herbal preparations that the subject had taken and not to study medication.The second drug-related event was a report of anemia in a 10 year old child in the +artesunate 4 mg/kg group who presented with a hemoglobin concentration of 3.8 g/dL at Day 7 (baseline 12.0 g/dL), was treated with blood transfusion and antibiotics and recovered.No artesunate dose-related changes in laboratory blood parameters were noted . There wBaseline hemoglobin levels were lower in children than in adults, and decreased further at Day 3 . HoweverAll subjects experienced a rise in methemoglobin levels by Day 3, though in all subjects this had decreased to approximately baseline levels by Day 7 . There wThe addition of artesunate to CPG\u2013DDS demonstrated advantages over CPG\u2013DDS alone for the primary efficacy endpoint (mean time to PC90) and for the majority of secondary endpoints in the adult group.This study achieved very accurate artesunate dosing to the target groups. However, the final fixed dose triple combination tablet needs to allow for a wider range of eventual mg/kg doses based on the weight of the individual patient. For example, for a target dose of 2 mg/kg artesunate, some patients would be expected to actually receive only 1 mg/kg. Thus, the outcome decision matrix was constructed conservatively; i.e. to confidently select the +artesunate 2 mg/kg target dose for the fixed dose combination, the +artesunate 1 mg/kg dose would have needed to be statistically different from CPG\u2013DDS alone for all endpoints.In children, +artesunate 1 mg/kg was not significantly different from CPG\u2212DDS alone for mean time to PC90 (the primary endpoint) and for mean time to PC50 and PC99. In adults, although +artesunate 1 mg/kg was significantly different from GPG\u2212DDS alone for the mean time to PC90 (the primary endpoint) in the principal (logistic model) analysis, this was not supported by the sensitivity analysis using observed data. In addition, +artesunate 1 mg/kg was not significantly different from CPG\u2212DDS alone for mean time to PC50 in adults. Although patient numbers were small there was no statistical difference in the effect on parasite viability of +artesunate 1 mg/kg versus CPG\u2212DDS alone in adults. There were no apparent differences in the safety profile of CPG\u2212DDS alone or in combination with artesunate in this study and no artesunate dose-related changes in laboratory parameters were observed. Thus, 4 mg/kg artesunate was selected as the dose to be combined with CPG\u2212DDS in the fixed dose combination tablet for further investigation in Phase III studies.We examined the differential effect of adding artesunate to CPG\u2212DDS alone. This study was powered to determine the effect of the artesunate doses versus CPG\u2212DDS alone and not to compare the different artesunate doses against each other. In this study a mean time to PC90 of 9 h was assumed as the difference between the artesunate groups and CPG\u2212DDS alone. When comparing between artesunate doses, we would expect a smaller difference in mean time to PC90, and as the artesunate dose increases so the planned difference decreases. As a consequence, the required sample size increases for each comparison to the point where a study of this type would be impractical at Phase II.The 14 day follow up used in this study was sufficient to demonstrate differences between the artesunate groups and CPG\u2212DDS alone for the parasitological endpoints. The phase III studies with CDA will include a 28-day follow up, based on recent WHO guidelines \u22121 was used; lower parasite counts may have resulted in such rapid parasite clearance so as to preclude any comparison of artesunate effect. The maximum parasite load of 100,000 \u00b5L\u22121 has been used throughout the CPG\u2212DDS and CDA development plans. This is a compromise between having a robust test of the drug while minimizing the number of patients that might have a parasite count exceeding 250,000 \u00b5L\u22121 at any point, i.e. the safety cut off whereby a patient must be withdrawn from the study.The patients in this study were not selected to be necessarily representative of the typical patient presenting with malaria in this setting. The inclusion and exclusion criteria were stringent and selected particularly to identify patients in whom the necessary data for parasitiological endpoints could be collected in order to achieve the study aim, i.e. to identify the effective dose of artesunate. In particular, the cut-offs for parasite density were narrower than commonly used in studies that use ACPRp as a primary outcome. In children, an initial parasite density of at least 25,000 \u00b5LThe narrow selection criteria in this study and the requirement to stay in hospital contributed to the screening of a very large number of patients. Unfortunately, recording all the patients that were screened and the reasons for not enrolling them in the study was not practical at all the centers. We believe that screening data would have confirmed that the study patient population was not generally representative of patients presenting with malaria.Patients in this study were required to stay in hospital for four days and three nights. This had a negative impact on recruitment of this study in Malawi, in particular for adults, necessitating the opening of a second center in The Gambia. As a result, no discrete analysis by center was planned, though center was included in the model for analysis of efficacy. Adverse events were reported less frequently for all treatment groups (for adults) at The Gambia study center compared with the Malawi center. These adverse events were recorded based on the discretion of the investigator, and the observed differences could reflect different clinical perspectives or cultural differences between the two sites. However, as there were no differences in the safety outcomes for CPG\u2212DDS versus the artesunate groups for each center, differences between centers did not impact the overall safety outcome of the study.3 had a higher incidence of malaria than those with counts >500 cells/mm3 though the prevalence of malaria was not influenced by CD4 count Patients in this study were excluded if they had overt signs of immunosuppression, but no HIV testing was performed. In 2006, the estimated proportion of the adult population living with HIV was 14.1% in Malawi and 2.4% in The Gambia and in 2003, it was 14.2% and 2.2%, respectively For children, the median age in the CPG\u2013DDS +artesunate 1 mg/kg group was 46 months, compared with 29\u221241 months in the other groups. This difference may have affected the results as younger children tend to get more severe malaria. However, the direction of any effect would have been to make 1 mg/kg artesunate appear better than it would have given a more equal balance of ages across the groups, and so this would not have affected the outcome of the decision matrix for dose selection.When this trial was conceived, it was unknown whether the sparse blood sampling achievable in children would be sufficient to show any treatment differences. An adult group was, therefore, included to provide a more closely sampled data set. However, it seems that despite less frequent sampling in children, the parasitological endpoints were sensitive enough to show differences between the artesunate groups when each was compared to CPG\u2212DDS alone.It was hoped that parasite viability would provide a robust key secondary endpoint to inform interpretation of the mean time to PC90 data. However, a systematic error in the technique that was later corrected meant that the number of patients included in this analysis was less than we had planned; 40/85 (47.1%) of adults and 13/92 (14.1%) of children were excluded. Thus, the confidence intervals around the mean treatment difference were too large to make relevant comparisons of the differential effect of the artesunate doses versus CPG\u2212DDS alone. However, the data are still interesting. As would be expected, parasite viability did show the potent schizonticidal activity of artesunate, demonstrating the value of adding this drug to the already clinically effective CPG\u2212DDS combination. Unexpectedly, even for the CPG\u2212DDS group, where no effect on parasite viability would be anticipated from this anti-folate inhibitor, parasite viability in adults was only 12.4% versus 71.2% for children. It may be that in adults there is some residual, long-lived immune-related process contributing to the impairment of ex-vivo parasite viability. This observation requires further investigation.It was not practical/ethically justifiable to include an artesunate monotherapy arm in the current study due to the long (7-day) course of treatment required and the risk of developing resistance to artemisinins. The ACPRp at Day 28 for 4 mg/kg artesunate monotherapy has been reported as 72% (36/50) for 3-day dosing A number of clinical studies have evaluated artesunate combination therapy in the treatment of uncomplicated falciparum malaria in African children. However, data comparing different doses of artesunate within combination therapy have been lacking.on average, acceptable efficacy. This strategy aims to reduce the potential for resistance emergence and spread by achieving maximal parasite reduction in as many treatments as possible.A study conducted in Thailand by Angus et al. in patients of at least 15 years of age found that 2 mg/kg artesunate was the lowest dose that, on average, achieved maximum effects on parasite clearance outcomes P. falciparum and are thus a more sensitive population in which to test the effect of different artesunate doses. Children are also a more vulnerable population clinically and it is important to minimize the possibility of breakthrough parasitemia and recrudescence in this group. Although the children in the study were all over 13 months of age, we believe the differences in outcome observed between adults and children were due primarily to differences in immunological response to P. falciparum. This is supported by the observation of far higher levels of parasite viability in children than in adults at 12 h after the first dose of study treatment for CPG\u2212DDS alone and the need for higher doses of artesunate to effectively suppress parasite viability in children versus adults. A subsequent study is planned to determine the safety and efficacy of CDA in children of \u226412 months.There appeared to be a dose\u2212response effect in this study for parasite clearance with artesunate up to 4 mg/kg in children, but not in adults, though this was not tested formally. Children have immature immune systems and more limited exposure to Hematological effects are a known adverse event associated with the DDS component Erythrocyte survival is further shortened in subjects given DDS who are glucose-6-phosphate (G6PD) deficient Although this study was not powered to make a formal assessment of safety outcomes between the treatment groups, these did not appear to be influenced by artesunate dose. Safety outcomes will be investigated more fully in a phase III trial of CDA versus CPG\u2212DDS alone.Artemisinin-derived compounds, such as artesunate, are included in combination therapy to rapidly reduce parasite load, for their gametocytocidal effect\u2212potentially reducing transmission\u2212 and to preserve antimalarial effectiveness by reducing the potential for the development of parasite drug resistance. Clinically, they reduce fever rapidly. However, where the other components of combination therapy are effective, artemisinins have little impact on the WHO definition of adequate clinical and parasitological response at Day 14 or Day 28.This study demonstrates the use of parasitemia-related outcomes, specifically mean time to PC90, in a clinical trial to make decisions regarding the appropriate artesunate dose to use with an already clinically effective antimalarial combination therapy (CPG\u2212DDS). Based on the findings of this study, Phase III clinical trials with CDA are progressing with a fixed dose combination of 2 mg/kg CPG, 2.5 mg/kg DDS and 4 mg/kg artesunate.Protocol S1Trial Protocol(0.74 MB PDF)Click here for additional data file.Checklist S1CONSORT Checklist(0.05 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum histidine-rich protein 2 (PfHRP2) or Pf lactate dehydrogenase (PfLDH) was undertaken in children aged between six and 59 months included in an anti-malarial efficacy study in Benin.An assessment of the accuracy of two malaria rapid diagnostic tests (RDT) for the detection of In Allada (Benin), 205 children aged 6-59 months with falciparum malaria received either artesunate-amodiaquine (ASAQ), artemether-lumefantrine (AL), or sulphadoxine-pyrimethamine (SP). Children included in the study were simultaneously followed by both RDT and high-quality microscopy for up to 42 days.PfHRP2-based tests were positive in 203 children (99%) and PfLDH-based tests were positive in 204 (99.5%). During follow-up, independent of the treatment received, only 17.3% (28/162) of children effectively cured were negative with the PfHRP2 RDT at day 3, with a gradual increase in specificity until day 42. The specificity of antigen detection with the PfLDH test was 87% (141/162) on day 3, and between 92% and 100% on days 7 to 42. A statistical difference was observed between the persistence of PfHRP2 and PfLDH antigenaemia during follow-up in children treated with artemisinin-based combination therapy (ACT) but not with SP.At the time of inclusion, PfHRP2 RDT had very low specificity during follow-up until day 28. On the other hand, the PfLDH test could be used to detect failures and, therefore, to assess anti-malarial efficacy.Although both RDTs are as sensitive as microscopy in detecting true malaria cases, the  In response to increased anti-malarial drug resistance, artemisinin-based combination therapy (ACT) is recommended in Africa . Due to PfHRP2), uniquely synthesized by Plasmodium falciparum present in the bloodstream of infected individuals. The second group of RDTs detects parasite lactate dehydrogenase (PspLDH), an enzyme produced by all four Plasmodium species responsible for human malaria, and may also detect species-specific LDH, such as PfLDH, specific of P. falciparum, or PvLDH, specific of Plasmodium vivax.The numerous commercially available RDTs fall into two categories . One groPfHRP2 test kits have generally shown higher sensitivity for P. falciparum and can be less costly than the PspLDH alternative, studies have shown that PfHRP2 remains in the bloodstream for an extended time following successful eradication of the parasite, thus contributing to false-positive results and limiting specificity  were estimated using microscopy as the denominator for comparison. Sensitivity was defined as the percentage of positive tests among the total number of positive blood slides. Specificity was defined as the percentage of negative tests among the total number of negative blood slides. The positive predictive value (PPV) was defined as the percentage of positive blood slides among the total number of positive tests. The negative predictive value (NPV) was defined as the percentage of negative blood slides among the total number of negative tests. The percentage of false-positive (FP) tests was defined as the percentage of tests that remained positive during follow-up once the blood sample was negative by microscopy for asexual-stage parasites. The presence of gametocytes alone was not considered to be a positive microscopy result. Categorical variables were compared using the Fisher or XP. falciparum parasitaemia above 1,000/\u03bcL were included in the study. Parasite densities ranged between 1,000 and 525,000 parasites/\u03bcl.Between March and September 2007, 205 children presenting with fever and a The children were divided into three groups, according to their treatment allocation: 81 were treated with the AL combination, 80 with the ASAQ combination, and 44 with the SP combination. During the six-week follow-up period, 80 children were followed to day 42 and 40 children were followed to day 28. Among the children included in the three groups, 72 were given additional treatment within 35 days, due to malaria recrudescence or re-infection (and were excluded from the RDT study after this second treatment), and 13 children were lost to follow-up. Detailed results of the efficacy study have been published . Seven gPfHPR2-based tests were positive for 203 children  and the PfLDH-based tests were positive for 204 . PspLDH and PfLDH detection yielded similar results during the study. Since only children with P. falciparum malaria were included in the study, only PfLDH detection results are presented and compared with PfHRP2 detection.On day 0, similar sensitivities were observed with both RDTs. The PfHPR2-based test (from 100% on day 3 to 67% on day 42) and the PfLDH-based test (from 80% on day 3 to 67% on day 42) , except for PfLDH-based tests carried out on day-28 samples  was stratified into low , middle , and high . The proportion of children with low parasite density was lower than the proportion with high parasite density from day 7 to day 35 for PfHRP2-positive antigen detection. Twenty-eight days after effective treatment, 8%  of the children with low parasite density had PfHRP2 false-positive results, compared to 42.9%  of those with high parasite density (p = 0.0004). In contrast, on any day during follow-up, the proportion of false-positive PfLDH antigen detection results was low and similar in all three groups . Moreover, after SP treatment, the proportion of false-positive results obtained with the PfLDH-based test was high until day 14 . No correlation was found between the presence of gametocytes and the duration of RDT positivity, except on day 7, when three among the 13 results were false-positive (13/164) with the PfLDH-based test in gametocyte carriers.Independent of the treatment received, PfHPR2 and PfLDH detection. PCR-amplified exon 2 PfHRP2 fragments were obtained on day 0 from the two negative isolates with the PfHPR2-based test; therefore, the parasites were not antigen-deficient. False-negative results obtained during follow-up with both the PfHPR2- and PfLDH-based tests could be explained by low parasite densities, comprising between 16 and 800 parasites/\u03bcl.Possible reasons for negative results included low parasite density and/or deficient in antigen parasites for both PfLDH-based test is below 90% only for a short time, up to seven days after effective treatment. During days 3 to 28 post-treatment, PfLDH detection was more specific than PfHRP2, as has been demonstrated in other studies [PspLDH and PfLDH are both related to parasite viability [PfLDH in blood after treatment may be due to parasite death following adequate treatment [PfLDH-based test positivity after treatment. Following ACT, the persistence of PfLDH was of shorter duration than that of PfHRP2. In addition, PfLDH false-positivity lasted longer following SP treatment than with ACT (until day 14). The rapid action of artemisinin derivatives on all parasite blood stages could be the reason for the fast clearance of PfLDH in blood [The results of this study reveal that the specificity of the  studies ,13. Sinciability , the rapreatment . In addiin blood .PfHPR2-based test can be as long as 28 days after effective treatment. As mentioned above, past evidence strongly suggests that, to a large extent, this is due to the increased time it takes to clear PfHPR2 from the blood following P. falciparum clearance. The duration of false positivity observed in this study with the PfHPR2-based test has been correlated to higher parasite density on admission. Since secretion of the protein is proportional to parasite numbers [PfHPR2 to be cleared from blood. The results of the present study further support this point, indicating a strong correlation between the duration of PfHPR2-based test positivity and parasite density at admission. Although the mechanism of PfHRP2 clearance is not well understood, there are several explanations for its long persistence following adequate therapy. As previously suggested by Singh and Shukla [PfHRP2 in blood after efficacious treatment was influenced by the treatment. Indeed, artemisinin derivatives are active during the Plasmodium ring stage, leading to the rapid disappearance of parasites in blood [PfHRP2, which is generated at all Plasmodium stages, could be stopped earlier with ACT than with SP, and a parallel quick clearance of PfHRP2 could occur [PfHRP2 after treatment with either artesunate or artemether (fast-acting artemisinin compounds) or SP. In addition, PfHRP2 false-positives determined by comparison with microscopy results are observed in identical proportions in all three treatment groups during follow-up.The duration of false positivity of the  numbers , a highed Shukla , the prein blood . The prold occur . HoweverThe reference method in this study was the microscopy because according WHO recommendations, efficacy is defined according to clinical and microscopy criteria . ProbablPfHRP2 is also released by early gametocytes [PfHPR2 positivity [PfHRP2 or PfLDH detection positivity, due to the small number gametocytes carriers. The ACT effect on gametocytes did not account for the shorter persistence of PfHRP2 compared to the effect of SP therapy.etocytes . Therefositivity . In the PfHPR2- and PfLDH-based test sensitivities compared with microscopy at the time of screening. In contrast with other studies [PfLDH-based test, equivalent to that of the PfHPR2-based test. Previous studies may have been carried out using reagents whose quality had been altered by heat and/or humidity. Reagent quality has now been improved and conservation is better, even under drastic conditions [PfHPR2-based test, but negative with PfLDH-based test, and the third was negative with the PfHPR2-based test but positive with PfLDH-based test. No alteration was found by PCR in the DNA of PfHRP2 and PfLDH genes that could explain the false-negative RDT results obtained for these samples [The results of this study reveal high  studies ,22, a hinditions . The hignditions . Discrep samples ,26.PfHPR2-based test and 96.2% for the PfLDH-based test against P. falciparum samples at parasites densities more than 2,000/\u03bcl [PfLDH-based test during follow-up were sufficient to consider a positive result to be a strong presumption of relapse, especially after ACT treatment.In this study, for each day of follow-up, the sensitivities of the tests to detect failures were between 67% and 100%, but the numbers of relapsing patients were low (between 3 and 16), and the parasitaemia in positive samples not detected by the tests was low. Whereas, for the diagnosis, the performance of both RDT tests used in the study is good as demonstrated by WHO with a sensitivity of 100% for the 2,000/\u03bcl . However\u00ae IT allows detection of both PspLDH and PfLDH and can be used during follow-up of an in vivo drug sensitivity test to confirm the effectiveness of anti-malarial treatment, especially after ACT or other quick-acting drugs, or to confirm suspected relapse. Immunoquick\u00ae Malaria allows detection of PfHRP2 and is a good test for ruling out a diagnosis of malaria, but should not be used during follow-up prior to day 35.In conclusion, OptimalThe authors declare that they have no competing interests.SH, JLB, PD, and JFF conceived and designed the study. SH, MDB, and JFF participated in data collection. SH and JLB participated in the data analysis. All authors participated in the writing of the manuscript, read, and approved the final manuscript."}
+{"text": "Plasmodium falciparum, defined as the proportion of gametocytes that are male, may influence transmission but little is known of the effects of mefloquine or artesunate-mefloquine on gametocyte sex ratio and on the sex ratio of first appearing gametocytes.The gametocyte sex ratio of P. falciparum malaria were enrolled in prospective treatment trial of mefloquine or artesunate-mefloquine between 2007 and 2008. Gametocytaemia was quantified, and gametocytes were sexed by morphological appearance, before and following treatment. The area under curve of gametocyte density versus time (AUCgm) was calculated by linear trapezoidal method.350 children with uncomplicated v 12.8%, P = 0.01). Gametocyte clearance was significantly faster with artesunate-mefloquine . AUCgm was significantly lower in the artesunate mefloquine group (P = 0.008). The pre-treatment sex ratio was male-biased, but post-treatment sex ratio or the sex ratio of first appearing gametocytes, was significantly lower and female-biased two or three days after beginning of treatment in children given artesunate-mefloquine.91% and 96% of all gametocytes appeared by day 7 and day 14, respectively following treatment. The overall rate of gametocytaemia with both treatments was 31%, and was significantly higher in mefloquine than in artesunate-mefloquine treated children if no gametocyte was present a day after treatment began (25.3% Addition of artesunate to mefloquine significantly modified the emergence, clearance, and densities of gametocytes and has short-lived, but significant, sex ratio modifying effects in children from this endemic area. Gametocytes are crucial for malaria transmission from the human host to the mosquito vector. Although the exact mechanisms of gametocytogenesis are unclear and many individual mosquitoes that ingest gametocytes do not support their development to sporozoite stage , anti-maPlasmodiun falciparum gametocytes has been estimated to vary from 3-21 days [The persistence, in peripheral blood, of -21 days -6. HowevPlasmodium spp gametocyte sex ratio is female biased, but this ratio can vary significantly during the course of individual infections [In natural populations, the fections -9. Althofections ,11, the fections ,9,12,13.Artesunate-mefloquine combination is increasingly used on the African continent for the treatment of acute falciparum malaria. However, the individual components of artesunate-mefloquine are readily available and used in Africa; it is desirable to evaluate the effects of both mefloquine and artesunate mefloquine on gametocyte carriage and sex ratio. In addition, it is important to assess how these drugs influence the first appearance of gametocytes in African children following treatment of uncomplicated infections. The information from these evaluations may assist in planning control measures.The aims of the present study are to evaluate the emergence and clearance of gametocytes; the rate of gametocytaemia; the temporal changes in sex ratio; and the sex ratio of first appearing gametocytes in children treated for acute falciparum malaria with mefloquine or artesunate plus mefloquine in an endemic area of southwestern Nigeria.P. falciparum malaria in Ibadan, southwestern Nigeria, an endemic area of malaria [P. falciparum parasitaemia \u2265 2000 asexual forms/\u03bcL, a body (axillary) temperature > 37.4\u00b0C or history of fever in the 24-48 h preceding presentation, absence of other concomitant illness, no history of anti-malarial use in the two weeks preceding presentation, and negative urine tests for anti-malarial drugs (Dill-Glazko and lignin). Patients with severe malaria [The study was conducted between July 2007 and August 2008 in children aged ten years or below with acute uncomplicated  malaria . Fully-i malaria , severe After clinical assessment, blood was obtained for haematocrit determination and for quantification of asexual and sexual parasitaemia. Patients were randomized to (i) a single dose regimen of mefloquine 25 mg/kg on presentation (day 0), or (ii) artesunate at 4 mg/kg daily for three days (day 0-2) plus mefloquine given as in (i) above. All drugs were given orally and all patients waited for at least three hours after to ensure the drug was not vomited. If it was, the patient was excluded form the study.Oral paracetamol (acetaminophen) at 10-15 mg/kg 6-8 hourly was given for 12-24 h if body temperature was > 38\u00b0C. Patients were seen daily, at approximately the same time of the day for the first eight days (days 0-7) and then on days 14, 21, 28, 35 and 42 after treatment had begun. At each visit, patients were assessed clinically and thick and thin blood smears were obtained for quantification of parasitaemia. The fever clearance time was defined as the time taken for body temperature to fall to below 37.5\u00b0C and remain below this value for > 48 h.Thick and thin blood films prepared from a finger prick were stained with 10% Giemsa for 30 minutes and were examined by light microscopy under an oil-immersion objective, at 1000\u00d7 magnification by two independent assessors who did not know the drug treatment of the patient. A senior member of the study team reviewed the slides, if there was any disagreement between the microscopists. In addition, the slides of every third child enrolled in the study were reviewed by this senior member.Asexual parasite and gametocyte counts were measured daily for the first eight days (days 0-7) and thereafter on days 14, 21 and 28. Quantification of asexual and sexual parasites in Giemsa-stained thick blood films was done against 500 leukocytes in the case of asexual parasitaemia, and against 1,000 leukocytes in the case of gametocytes. From this figure, the parasite density was calculated assuming a leukocyte count of 6,000/\u03bcl of blood. Asexual parasite clearance time was the time interval from the start of anti-malarial treatment until the asexual parasite count fell below detectable levels in a peripheral blood smear. Gametocyte clearance time was the interval between the first and last positive smears for gametocytes. Capillary blood, collected before and during follow-up, was used to measure packed cell volume (PCV). PCVs were measured using a microhaematocrit tube and microcentrifuge . Routine haematocrit was done on days 0, 3, 7, 14, 21 and 28.Gametocyte sex determination was based on the following criteria ,17: maleversus time (AUCgm) were determined by a non-compartmental method using the computer programme Turbo Ken  as previously described [gm was obtained, using the linear trapezoidal rule from time zero  to the time of gametocyte clearance, or if there was no clearance, until 1008 h (day 42). The final gametocyte density at the time of clearance was assumed to be 0.001 sexual forms/\u03bcl blood (a level assumed to be below microscopic detection).For each patient, gametocyte densities were plotted against time. The areas under the curve of gametocytaemia escribed . BrieflyP. falciparum gametocytes and trophozoites. Proportions were compared by calculating \u03c72 with Yates' correction or by Fisher exact or by Mantel Haenszel tests. Normally distributed, continuous data  were compared by Student's t-tests and analysis of variance (ANOVA). Data not conforming to a normal distribution  were compared by the Mann-Whitney U-tests and the Kruskal-Wallis tests (or by Wilcoxon ranked sum test). Kaplan-Meier plots are also presented to compare gametocyte carriage rates following treatment. Differences in survival time were assessed by inspection of Kaplan-Meier curves and log-rank tests. All tests of significance were two-tailed. P-values of < 0.05 were taken to indicate significant differences. Data were (double)-entered serially using the patients codes and were only analysed at the end of the study.Data were analysed using Epi-Info version 6 , and theThree hundred and fifty patients were recruited: 174 in atresunate-mefloquine group and 176 in mefloquine group. All children completed at least 14-21 days of follow up. The baseline characteristics of children with gametocytaemia are summarized in Table Table Gametocytes were detected in peripheral blood in 108 children (31%) from the two treatment groups. Thus the overall proportions of children with gametocytaemia was 31%. The proportion was 12.8% and 25.3% for artesunate-mefloquine and mefloquine alone, respectively if no gametocytes were present by day 1 after enrolment  versus 22 of 171 (12.6%), in the mefloquine and artesunate-mefloquine groups, respectively, P = 0.71. After treatment, the emergence of gametocytes was significantly less frequent in the artesunate-mefloquine group than in the mefloquine alone group: 22 of 171 (12.6%) versus 44 of 171 (25%), respectively, P = 0.004. Post-treatment, gametocyte carriage was significantly higher than pre-treatment in the mefloquine alone group  but not in the artesunate-mefloquine group .The probability of mosquito infectivity is related to gametocyte density and the duration of carriage by the host. Figure gm was determined in patients who carried gametocytes at least on three occasions within 14 days. The mean AUCgm were (501.7 \u00b1 115.6 (SEM), 95% confidence interval (95% CI) 261.4-742.0 sexual forms/\u03bcl.d in mefloquine-treated children and (186.7 \u00b1 71.2 (SEM), 95% CI 22.4-351.0 sexual forms/\u03bcl.d in artesunate-mefloquine treated children  .Only seven children, all treated with mefloquine alone, had gametocytes on or after day 14. Overall, gametocyte clearance was significantly faster in the artesunate-mefloquine group than in the mefloquine alone group . However, sex ratio values 2 d after treatment began were significantly lower and more female-biased in the artesunate-mefloquine group compared with the mefloquine alone group (0.35 \u00b1 0.15 (SEM) v 0.76 \u00b1 0.07, P = 0.02). In addition, this value for artesunate-mefloquine was significantly lower than pre-treatment mean sex ratio (P = 0.04). In mefloquine treated children, the corresponding values were similar (P = 0.93).The variations in sex ratio following treatment are shown in Figure Overall, the sex ratio of first appearing gametocytes was male-biased in mefloquine treated children. In artesunate-mefloquine treated children, sex ratio of first appearing gametocytes three days after treatment began became female-biased and was significantly lower (P = 0.04) in the artesunate-mefloquine than in the mefloquine alone group Figure .The overall gametocyte carriage rate of 12% at enrolment is consistent with previous findings in this and other endemic areas ,11,21-23Only 40% of gametocytaemic patients were identified on presentation. This proportion is significantly lower than 72% recorded in an area of lesser intensity of transmission in Thailand . The reaThe risk of gametocyte carriage following treatment with anti-malarials varies considerably; the risk is often higher with monotherapy than with artemisinin combination therapy ,11,22,28The significantly higher post-treatment gametocyte carriage in children treated with mefloquine alone suggest that the slow clearance of asexual parasitaemia (independent of recrudescence) may be primarily responsible for the progression of committed asexual parasites to develop to gametocytes. Recrudescence in both mefloquine and artesunate-mefloquine is associated with gametocytaemia ,29. The The study evaluated the effects of treatment with mefloquine or artesunate-mefloquine on gametocyte clearance and sex ratio in children who were gametocytaemic before, during or after treatment commenced. To our knowledge, this is the first study of the effects of these drugs on gametocyte clearance and sex ratio in African children. In these children, artesunate-mefloquine cleared gametocytamia significantly more rapidly than mefloquine alone -30 and mMonocytes and leukocytes phagocytose early and late stages of gametocytes, respectively ,34. It iP. falciparum, an evaluation of the sex ratio of first-appearing gametocytes was made. The assumption was that the sex ratio of first-appearing gametocytes as opposed to sex ratio of all gametocyte present in circulation would be a more sensitive indicator of the effects of chemotherapy. Based on this assumption, the finding was that, overall, the sex ratio of first appearing gametocytes was male-biased except on the third day after treatment began in the artesunate-mefloquine group, when it became significantly female-biased. Since the gametocytes emerging immediately following treatment were already allocated to sex several days before treatment began, the female-biased sex ratio observed with artesunate-mefloquine would suggest that the drug has no effect on sex allocation strategy of the parasite. Therefore, it is likely that the observed female-biased sex ratio is due to preferential release, into peripheral blood, of female gametocytes or a selective killing effect on male gametocytes. The effect (s) on emergence, however, was short-lived, and may have been blunted by the relatively long half-life of mefloquine of 16-33 days [Gametocyte sex ratio may be influenced by a number of factors including anaemia, the number of and the competition between different parasite genotypes and other biological variables such as parasite density and transmission rates -9,12,13.-33 days  exerting-33 days . Another-33 days ,39. StudP. falciparum increased its investment significantly in gametocyte production but not in sex allocation suggesting that induction of gametocytogenesis and sex determination may have different mechanisms.Plasmodium species may increase their chances of transmission by investing in gametocyte production or in sex allocation . It woulgm was approximately three-fold higher in mefloquine than in artesunate-mefloquine treated children. Assuming sex ratio is related to mosquito infectivity, the higher value, coupled with a higher AUCgm treatment for mefloquine should presumably put patients treated with mefloquine at a higher risk for mosquito infectivity and transmissibility if the gametocytes were viable.In addition to the density of peripheral gametocytes  and the There are potential implications of the findings of the present study: primaquine is gametocytocidal and can shorten gametocyte clearance time ,41 but iThe authors declare that they have no competing interests.AS led the design, conduct, data analysis and manuscript preparation. OON and TMO were involved with conduct and data analysis. GOG and CTH were involved in design, conduct, and preparation of the manuscript. EOA was involved with data analysis and manuscript preparation. All authors read and approved the manuscript."}
+{"text": "Combination therapy has become a new paradigm in malaria treatment. Amodiaquine is a common partner drug in different malaria combination therapies used or investigated in sub-Saharan Africa, but data on its efficacy as a single drug are scarce.The objective of the study was to determine the efficacy of amodiaquine against falciparum malaria in neighbouring rural and urban areas of north-western Burkina Faso. The study was designed as an uncontrolled trial in children aged 6\u201359 months with uncomplicated falciparum malaria in the Nouna Health District.Plasmodium falciparum crt K76T mutation not predict AQ failure, but was selected in parasites re-appearing following treatment. No serious adverse events occurred and only 16 other adverse events were recorded.During the rainy season 2005, 117 children were enrolled, 62 from the rural and 55 from the urban study area. The crude adequate clinical and parasitological response (ACPR) rate was 103/117 (88%) by day 14 but decreased to 28/117 (24%) by day 28. After PCR correction for reinfections, ACPR rates were 108/117 (92%) and 71/117 (61%) by day 14 and day 28, respectively. There were no significant differences in efficacy between urban and rural areas. The Compared to chloroquine, amodiaquine is more effective against uncomplicated falciparum malaria in Burkina Faso. However, a considerable degree of amodiaquine resistance already exists and it is currently unclear how this resistance will develop when amodiaquine in combination with other drugs is used on a large scale.Current Controlled Trials ISRCTN73824458. Plasmodium falciparum to existing and affordable first-line drugs such as chloroquine (CQ) or sulphadoxine/pyrimethamine (SP) in most SSA countries  and 92% [88\u201395%], respectively, during four weeks of follow-up , which ce latter . This fireatment ,45,47. Rsistance . In ongoEfficacy and useful therapeutic life span of any antimalarial drug combination critically depend on the choice of the partner drugs and preexisting resistance patterns ,51. As AIn this trial from north-western Burkina Faso, AQ treatment of children with uncomplicated malaria achieved an unsatisfactory cure rate of 61%. Impaired AQ efficacy, may shorten the useful life span of the first-line drug AQ-AS and of further AQ-based combination therapies. Continued monitoring of the efficacy of AQ is warranted.The authors declare that they have no competing interests.OM, GM and PM designed the study. GM, BC and FPM conducted the laboratory and clinical work. OM, GM and PM analysed the data. All authors contributed to the interpretation of the findings and to writing of the paper."}
+{"text": "Uganda recently adopted artemether-lumefantrine (AL) as the recommended first-line treatment for uncomplicated malaria. However, AL has several limitations, including a twice-daily dosing regimen, recommendation for administration with fatty food, and a high risk of reinfection soon after therapy in high transmission areas. Dihydroartemisinin-piperaquine (DP) is a new alternative artemisinin-based combination therapy that is dosed once daily and has a long post-treatment prophylactic effect. We compared the efficacy and safety of AL with DP in Kanungu, an area of moderate malaria transmission.Patients aged 6 months to 10 years with uncomplicated falciparum malaria were randomized to therapy and followed for 42 days. Genotyping was used to distinguish recrudescence from new infection. Of 414 patients enrolled, 408 completed follow-up. Compared to patients treated with artemether-lumefantrine, patients treated with dihydroartemisinin-piperaquine had a significantly lower risk of recurrent parasitaemia  but no statistically significant difference in the risk of treatment failure due to recrudescence . Patients treated with dihydroartemisinin-piperaquine also had a lower risk of developing gametocytaemia after therapy . Both drugs were safe and well tolerated.DP is highly efficacious, and operationally preferable to AL because of a less intensive dosing schedule and requirements. Dihydroartemisinin-piperaquine should be considered for a role in the antimalarial treatment policy of Uganda.ISRCTN75606663Controlled-Trials.com  With the emergence of widespread resistance to chloroquine (CQ) and sulfadoxine-pyrimethamine (SP), most African countries have adopted artemisinin-based combination therapy (ACT) as first-line treatment for uncomplicated malaria. Although several ACTs exist, currently only two have been widely adopted into policy in Africa: artesunate-amodiaquine (AS/AQ) and artemether-lumefantrine (AL), each of which is the recommended therapy for uncomplicated malaria in over a dozen countries Dihydroartemisinin-piperaquine (DP) is a fixed-dose ACT which has recently become available in Africa. This drug is relatively inexpensive and has the advantages of once a day dosing and a long post-treatment prophylactic effectThere have been three published studies of DP in Africa. In all of these studies DP was associated with excellent safety and efficacy, as well as a lower risk of recurrent malaria compared to AS/AQ In contrast to our previous study done in an area of high transmission intensity, this trial was conducted in an area of moderate transmission intensity in Uganda. We compare the efficacy and safety of DP with the current first-line therapy, AL, testing the hypothesis that the risk of recurrent parasitaemia would differ between the two treatment arms. We also discuss the policy implications of these findings in the context of a growing body of evidence for the ACTs in Africa.The protocol for this trial and supporting CONSORT checklist are available as supporting information; see The study was conducted at Kihihi Health Centre, Kanungu District, Western Uganda. This district experiences perennial mesoendemic malaria; the entomological inoculation rate was measured at 7 infectious bites per person per yearP. falciparum mono-infection with parasite density 2,000\u2013200,000/\u00b5l of blood. Because laboratory results were generally not available until the following day, a patient could be excluded after randomization.Consecutive patients presenting to the health center with symptoms suggestive of malaria and a positive screening thick blood smear were referred to study physicians for further assessment. Patients were enrolled if they fulfilled the following selection criteria: 1) age 6 months to 10 years; 2) weight \u22655 kg, 3) history of fever in the last 24 hours or axillary temperature \u226537.5\u00b0C; 4) no history of serious side effects to study medications; 5) no evidence of a concomitant febrile illness; 6) provision of informed consent by a parent or guardian; 7) no danger signs or evidence of severe malaria; and 8) The study protocol was approved by the Makerere University Research and Ethics Committee, the Uganda National Council of Science and Technology, and the University of California, San Francisco Committee for Human Research. Parents or guardians of all participating children provided written informed consent before children could be enrolled into the study.A nurse administered study medications according to weight-based guidelines for fractions of tablets. We administered all drugs orally as follows: AL , administered as one (5\u201314 kg), two (15\u201324 kg), three (25\u201334 kg), or four (\u226535 kg) tablets given twice daily for 3 days; DP , targeting a total dose of 6.4 and 51.2 mg/kg of dihydroartemisinin and piperaquine, respectively, given in 3 equally divided daily doses to the nearest \u00bc tablet (cut with a pill cutter). Participants in the DP group also received placebo tablets administered in the evening over 3 days to simulate the AL dosing schedule. Study medications were administered with water, and patients were given a glass of milk after each dose of study medication. All treatment was directly observed at the study clinic. Participants were given the option either to wait at the clinic for the evening dose (lunch was provided) or to leave and return to the study clinic in the evening (transport was provided). After each dose, children were observed for 30 minutes, and the dose was re-administered if vomiting occurred. All patients were provided with a 3-day supply of acetaminophen for treatment of febrile symptoms. Children with haemoglobin of less than 10 gm/dL were treated according to Integrated Management of Childhood Illness guidelines with ferrous sulfate for 14 days and antihelmintic treatment if appropriate. Households of all patients were given two long-lasting insecticide treated bednets  on the day of enrollment, with instructions for one net to be used by the study patient.Patients were asked to return for follow-up on days 1, 2, 3, 7, 14, 21, 28, 35 and 42, and any other day that they felt ill. Follow-up evaluation consisted of a standardized history and physical examination, including neurological assessment on all days of follow up. We obtained blood by fingerprick for thick blood smears and storage on filter paper on all follow-up days except day 1. Haemoglobin measurement was repeated on day 42 or the day of recurrent symptomatic malaria. If patients did not return for follow-up, they were visited at home.Treatment failures received quinine . Patients with evidence of severe malaria or danger signs  were referred for treatment with parenteral quinine. Patients were excluded during follow-up for use of antimalarial drugs outside of the study, serious adverse events requiring a change in treatment, withdrawal of informed consent, or loss of follow-up (not located within 24 h Days 1\u20133 or 48 h Days 4\u201342).Initial screening blood smears were stained with 10% Giemsa for 10 minutes. Thick and thin blood smears were stained with 2% Giemsa for 30 minutes. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 200 white blood cells (WBCs), or per 500 if the count was less than 10 parasites per 200 WBCs, assuming a WBC count of 8,000/\u00b5l. A smear was considered negative if no parasites were seen after review of 100 high-power fields. We also assessed for the presence of gametocytes from thick blood smears. Expert microscopists reviewed thin blood smears for non-falciparum infections using known defining characteristics to differentiate between species msp-2, msp-1, and four microsatellites Molecular genotyping techniques were used to distinguish recrudescent from new infections for all patients with a late clinical failure (LCF) or late parasitological failure (LPF) response. Parasite DNA was isolated from filter paper blood samples collected at enrollment and on the day of recurrent parasitaemia using chelex extraction. Paired samples were genotyped in a stepwise fashion using The objective of the study was to compare the efficacy and safety of DP with the current first-line therapy, AL, for treating uncomplicated falciparum malaria in an area of moderate transmission intensity in Uganda.Treatment outcomes were classified according to 2006 WHO guidelines as early treatment failure , LCF , LPF (presence of asymptomatic parasitaemia on days 7\u201342) or adequate clinical and parasitological response  after 42 days was estimated to be 50% after treatment with AL based on previous dataA randomization list was computer generated by an off-site investigator. Sequentially numbered, sealed envelopes containing the treatment group assignments were prepared from the randomization list.The study number and assigned treatment were printed on a card and securely sealed in opaque envelopes. Sealed opaque envelopes containing the study number and assigned treatment were secured in a locked cabinet accessible only by the study nurse.The nurse administered treatment by opening an envelope with a matching treatment number sequentially assigned by the study physician.Only the study nurse was aware of treatment assignments. All other study personnel, including the study physicians and laboratory personnel involved in assessing outcomes, were blinded to the treatment assignments. Patients were not informed of their treatment regimen, but the color and taste of the two study medications were not the same .Data were entered and verified using Epi-Info version 6.04 and analyzed using STATA version 8.0 . Efficacy and safety data were evaluated using a modified intention-to-treat analysis which included all patients who fulfilled enrollment criteria. Patients who were randomized to therapy but were not enrolled in the study due to laboratory results not being available on day 0 were not included in the analysis. Risks of treatment failure at 28 and 42 days of follow-up (adjusted and unadjusted by genotyping) were estimated using the Kaplan-Meier product limit formula. Data were censored for patients who did not complete follow-up and for new infections when estimating outcomes adjusted by genotyping. Patients with LCF or LPF due to non-falciparum species were censored as non-failures at the time they were classified as LCF or LPF. Comparisons of treatment efficacy were made using risk differences (RD) with exact 95% confidence intervals. Categorical variables were compared using chi-squared or Fisher's exact test and continuous variables were compared using the independent samples t-test. All reported p-values are two sided without adjustment for multiple testing and were considered statistically significant if <0.05.Of 463 patients screened, 2 were excluded during screening and 47 were excluded after treatment assignment, but before enrollment . ReasonsStudy participants were enrolled from August 2006 to April 2007.Among patients with treatment outcomes, there were no significant differences in baseline characteristics between the two treatment groups .Efficacy and safety data were evaluated using a modified intention-to-treat analysis, which included all 414 patients who fulfilled enrollment criteria .P. falciparum and non-falciparum species. The risk of treatment failure unadjusted by genotyping was significantly lower for participants treated with DP than for those treated with AL after 28  and 42 days of follow up   . Most ep0\u201328.8%) . After c0\u201328.8%) .The proportion of patients with fever on Day 1 was significantly lower in participants treated with DP , but was similar over the second and third days of follow up in the two treatment groups. Both treatments produced rapid clearance of parasitaemia, with no parasites detected by day 3 . The appNo patients were withdrawn from the trial for vomiting that would have required alternative treatment. Adverse events, broadly defined as any untoward medical occurrences, occurred commonly. Most adverse events were of mild or moderate severity and consistent with symptoms due to malaria. The most commonly reported adverse events in both treatment groups were cough, coryza, abdominal pain, anorexia, weakness, diarrhea and pruritus . AdverseIn this study, we compared locally relevant antimalarial drug combinations in a randomized trial of children with uncomplicated malaria, and followed up patients for 42 days. AL is currently the first line treatment for malaria in UgandaP. falciparum infections in children. However patients treated with DP had a significantly reduced risk of treatment failure due to recurrent parasitaemia and a lower risk of recurrent parasitaemia due to recrudescence.DP is a newer fixed combination regimen that is registered in Uganda. Both regimens were highly efficacious in clearing initial The risk of treatment failure unadjusted by genotyping, which was significantly different between the two treatment arms, largely reflects a difference in rates of reinfection, rather than recrudescence. DP clearly offered a better post treatment prophylactic effect following therapy compared to AL. The significant lower risk of recurrent parasitaemia after treatment with DP is likely explained by differences in pharmacokinetics of the non-artemisinin partner drugs. Piperaquine, a bisquinoline, is estimated to have an elimination half-life of 2\u20133 weeks Five years after the call for deployment of artemisinin combination therapy for treating malaria first gained momentum This study was conducted in a moderate transmission area, but the results are consistent with those from studies from high transmission sites in Africa Our results add to recent data comparing DP to other artemisinin combination treatments of interest. In this study, DP is clearly superior to AL at preventing new malaria infections; is at least as safe as AL; and with simpler dosing requirements and lower cost, it appears to be a preferable alternative.This study adds data from a moderate transmission site to two prior studies at high transmission sites in Africa Protocol S1Trial Protocol(1.07 MB DOC)Click here for additional data file.Checklist S1CONSORT Checklist(0.06 MB DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum. Its use is strongly recommended in most sub-Saharan African countries, namely Cameroon, where resistance to chloroquine is widespread and antifolate resistance is emerging.The use of drug combinations, including non-artemisinin-based and artemisinin-based combination therapy (ACT), is a novel strategy that enhances therapeutic efficacy and delays the emergence of multidrug-resistant P. falciparum malaria according to the standard World Health Organization protocol at four sentinel sites between 2003 and 2007. A total of 1,401 children were enrolled, of whom 1,337 were assigned to randomized studies and 64 were included in a single non-randomized study. The proportions of adequate clinical and parasitological response (PCR-uncorrected on day 14 and PCR-corrected on day 28) were the primary endpoints to evaluate treatment efficacy on day 14 and day 28. The relative effectiveness of drug combinations was compared by a multi-treatment Bayesian random-effect meta-analysis.Studies were conducted in Cameroonian children with acute uncomplicated The results based on the meta-analysis suggested that artesunate-amodiaquine (AS-AQ) is as effective as other drugs . AM-LM appeared to be the most effective with no treatment failure due to recrudescence, closely followed by DH-PP.Although AM-LM requires six doses, rather than three doses for other artemisinin-based combinations, it has potential advantages over other forms of ACT. Further studies are needed to evaluate the clinical efficacy and tolerance of these combinations in different epidemiological context. Plasmodium falciparum is now widespread in Africa, and antifolate-resistant P. falciparum is emerging in some regions in Africa [P. falciparum infections, respectively, between 2002 and 2004.Chloroquine-resistant n Africa . In CameTo overcome drug-resistant malaria, malaria experts advocate the use of combination therapy ,3. The mCameroonian health authorities recommend AS-AQ for the treatment of uncomplicated malaria since 2004. AM-LM is an alternative therapy in Cameroon since 2006. In the previous studies, the results of the nationwide evaluation of the current therapeutic efficacy of monotherapies  were presented . The aimP. falciparum parasites/\u03bcL of blood, without other Plasmodium species [Clinical studies were conducted at four different urban centres situated in different geographic area in Cameroon. Malaria transmission is intense and continuous throughout the year in the country, except for the northern (Garoua) and far-northern provinces (Maroua), where transmission is low and seasonal. Children were enrolled after free and informed consent of the parents and/or legal guardians if the following inclusion criteria were met: age \u2264 five years of age, fever at the time of consultation, parasite density \u2265 2,000 asexual  species . As reco species . Each suPatients were randomized to two or three treatment groups, with the exception of the study conducted in Maroua where only AS-AQ combination was evaluated. Separate concealed-random list based on random number tables was prepared for each trial by the principal investigator. Patients were consecutively allocated by the local investigator according to the corresponding list. Amodiaquine (AQ) was administered at a standard dose of 10 mg base/kg body weight on days 0, 1, and 2. Sulphadoxine-pyrimethamine  was administered in a single dose. The dosage of AQ-SP was the same as that of monotherapies. The first doses of AQ and SP were administered simultaneously on day 0, followed by AQ alone on days 1 and 2.\u00ae) 6.4 mg/kg body weight of DH and 51.2 mg/kg body weight of piperaquine in 3 divided daily doses. Six doses of AM-LM (Coartem\u00ae) were administered as recommended by the manufacturer. For the AS-CD combination, the dose of chlorproguanil-dapsone (Lapdap\u00ae) was given once daily for three days, as recommended by the manufacturer. Paracetamol (30 mg/kg body weight/day) was administered to all patients.AS was administered at a total dose of 12 mg/kg body weight  for all ACTs containing AS. The following dosages of ACT were administered: AS-AQ  on days 0, 1, and 2; AS-SP, (SP on day 0); AS-MQ ; and DH-PP , merozoite surface antigen-1 (msa-2), and glutamine-rich protein (glurp) genes of the pre-treatment and recrudescent samples were amplified, as recommended by a group of malaria experts [Fingerprick capillary blood was collected for blood smear and DNA analysis at the time of treatment or parasitological failure occurring on day 7 or after. The polymorphic merozoite surface antigen-1 , significant difference between AQ, SP, AQ-SP arms was tested using ANOVA for the binary variable ACPR 1/treatment failure 0. The test of the efficacy trend of AQ-SP between 2003 and 2006 was performed by comparing the rate of ACPR in 2003 to the rate in 2006 using the odds ratio (OR) on day 14.For each 28-day follow-up study (2005-2007), the ORs and 95% confidence intervals were calculated. The Yusuf and Peto method was used for the 2006-study (AS-AQ versus AM-LM) as the AM-LM arm showed 100% ACPR patients after PCR adjustment [Unlike the classical random-effect meta-analysis, where there is the same reference treatment or placebo across the trials, a pooled effect and summary OR versus a reference treatment could not be directly estimated since treatments were not the same from one study to the other . As the The treatment response per subject was viewed as a binary variable, i.e. 1 for ACPR and 0 for failure. Data were agglomerated on day 14 for all studies, and on day 28 based on both PCR- uncorrected and corrected results when the follow-up reached 28 days or more. The model estimated i) posterior OR for each treatment compared to the AS-AQ treatment, ii) the variability among subjects within sub-trials and iii) the variability among treatment groups, starting with reasonable prior distribution for each parameter.Data were analysed using the statistical software R . For theP < 0.05). Even among patients with relatively high initial parasitaemia , the mean pre-treatment haematocrit (28.5 \u00b1 5.4%) increased to 34.6 \u00b1 4.0% on day 14, i.e. increase by 6.2% , attesting the general benefit of an effective combination therapy.A total of 1,401 patients were enrolled in our series of studies . All analysed data are presented in a CONSORT format in Fig. P = 0.01), with an overall cure rate of 87%  on day 14. In 2003, the efficacy of AQ-SP was not statistically different from AQ monotherapy (P > 0.05). There was no significant difference in the efficacy of AQ-SP between 2003 (145/156 or 93%) and 2006 (64/67 or 96%) on day 14 .With the AQ-SP treatment, the overall cure rate, i.e. ACPR, was 93.0% on day 14 and 78% on day 28 before PCR correction and 91% after PCR correction . AS-AQ combination was less effective  than DH-PP  . After PCR adjustment, the cure rates on day 28 were 92.7% and 88.0% for DH-PP and AS-AQ, respectively . Parasite clearance time was longer with DH-PP than AS-AQ (P < 0.05). Based on the ITT analysis, the AS-CD combination was less effective  than AS-SP  . After PCR adjustment, the cure rates on day 28 were 91.7% and 76% for AS-SP and AS-CD, respectively .From 2005 to 2007, the efficacy of artemisinin derivatives combined with a partner drug was assessed on day 28. The treatment outcomes of combination therapies, before and after PCR adjustment of the number of ACPR, are summarized in Tables P < 0.05). However, on day 3 (see Table P = 0.13). Moreover, more than 90% of patients cleared their parasitaemia on day 3, except for the AQ-SP combination and AQ monotherapy.Based on the PCR-corrected proportions of ACPR on day 28, there was no statistical difference in the efficacy of AQ-SP and AS-MQ combinations . The decrease in parasitaemia was more rapid with AS-MQ than AQ-SP on day 2 . Three patients were excluded due to repeated vomiting associated with AS-MQ administration.Treatment failure occurred in one out of 61 patients  treated with AS-MQ on day 28. Failure was observed in five additional patients  between day 29 and day 42. Vomiting in children treated with AS-MQ occurred more frequently than in the AQ-SP group, on day 1 (10.3% vs 1.5%), day 2 (10.8% vs 0) and day 3 (3.3% vs 1.6). Significant difference was observed on day 1 and 2 (P < 0.05) than that of AQ-SP, at both endpoints on day 14 and day 28. AM-LM and DH-PP were significantly more effective than AQ-SP on day 28.The logistic regression on pooled individual patient data (PCR-uncorrected) comparing the efficacy of ACT to that of AQ-SP showed that the efficacy of AQ-SP is not statistically different from that of AS-AQ, AS-MQ, AS-SP, and DH-PP on day 14 Table . HoweverThe results of the multi-treatment Bayesian random-effects meta-analysis based on individual data of children are shown in Figure The present work concerned a global analysis of a series of randomized studies of anti-malarial treatment efficacy conducted in Cameroon between 2003 and 2007. Following comparison between arms within each study, a multi-treatment Bayesian random-effects meta-analysis of the binary outcome, ACPR/failure as a marker of efficacy, was carried out both on day 14 and day 28. The latter used PCR-uncorrected and PCR-corrected data. This global approach increased the power for detecting differences between treatments, while controlling the type-1 error.Anti-malarials were AQ and SP monotherapies, their combination AQ-SP, and new drugs included in ACT. AQ monotherapy is still effective in Cameroon but should be protected with artesunate (or SP) to delay the emergence of resistance. The current trend in Africa is to reserve SP for the intermittent preventive treatment in pregnant women . During In Cameroon, AS-AQ and AM-LM have being used nationwide since 2007 although AM-LM is relatively less prescribed due to its low supply in the public sector. The present study indicates that AS-AQ is well-tolerated and highly effective, confirming the results of an earlier multicentric study conducted in Africa . Currenti.e. AS-SP, AS-CD, AS-MQ, DH-PP, that require once daily dose for three days, did not show a significant difference with AS-AQ. Previous studies have shown that AS-SP is a highly effective ACT [The forms of ACT, tive ACT . Howevertive ACT -22.P. falciparum infections for more than a decade [AS-MQ combination has been widely used in some Southeast Asian countries to treat multidrug-resistant a decade . Its effa decade -26. In ma decade ,28. The a decade ,29.Piperaquine, an 'old' bisquinoline synthesized in the 1960s and used extensively in China, has been found to be a suitable partner of dihydroartemisinin . Recent P. falciparum in Central Africa is not well defined at present. In countries where SP is largely employed for intermittent preventive treatment in pregnant women, it may not be advisable to use AS-SP for malaria treatment of the general population. Further studies are required to evaluate the optimal dosing of AS-MQ for African children. At present, it is probably too early to recommend AS-MQ in Africa as an alternative to other existing forms of ACT, which are better tolerated than AS-MQ. There are other novel forms of ACT, including artesunate-pyronaridine and artesunate-atovaquone-proguanil, that have not been evaluated in the present study. Clinical efficacy and tolerance of these combinations need to be evaluated and compared with those of AS-AQ and AM-LM in Central Africa.The results of these studies on the efficacy of AS-AQ and AM-LM are in agreement with those conducted elsewhere in Africa . The varThe authors declare that they have no competing interests.SWY developed the analysis plan and carried out both the statistical analyses and software implementations under the close supervision of HG and JCT. All participated in the interpretation of data. RT was responsible for checking the data and performing molecular techniques. VFN supervised the enrolment and follow-up of patients and participated in data entry and collection. GS designed the studies and assisted with data interpretation. LKB was responsible for overall scientific management and drafted the manuscript. All authors participated in the preparation of the report and approved the final version.Table S1: Pre-treatment clinical and laboratory characteristics of enrolled children who completed the 14-day or 28-day follow-up. 1 Patients were followed-up for 14 days in studies conducted in 2003 and for 28 days in studies performed in 2005-2007. Patients assigned to artesunate-mefloquine group were followed for 42 days. AQ, amodiaquine; SP, sulphadoxine-pyrimethamine; AS, artesunate; MQ, mefloquine; AM, artemether; LM, lumefantrine; CD, chlorproguanil-dapsone; DH, dihydroartemisinin; PP, piperaquine. 2 Number of patients enrolled . 3 The numbers of children aged > 60 months old (and/or adults for Maroua) are 18/57 in Garoua 2003 AQ, 16/58 in Garoua 2003 SP, 27/58 in Garoua 2003 AQ-SP, and 18/64 (28.1%) in Maroua (none at other study sites). Garoua and Maroua are situated in northern Cameroon where malaria transmission is seasonal. 4 The following number of patients had > 200,000 asexual parasites/\u03bcL of blood: 2 (1 in AQ group and 1 in SP group) in Yaound\u00e9 2003; 5 (2 in AQ group and 3 in SP group) in Bertoua 2003; 3 (2 in AQ group and 1 in AQ-SP group) in Garoua 2003; 10  in Yaound\u00e9 2005; 1 in Maroua; 4  in Yaound\u00e9 2006a; 5  in Yaound\u00e9 2006b; 7 (5 in AS-SP group and 2 in AS-CD group) in Yaound\u00e9 2007a; and 11 (5 in DH-PP group and 6 in AS-AQ group) in Yaound\u00e9 2007b.Click here for file"}
+{"text": "Plasmodium falciparum malaria. The objectives of the current analysis were to compare the efficacy and safety of AL across different body weight ranges in African children, and to examine the age and body weight relationship in this population.Artemisinin-based combination therapy, including artemether-lumefantrine (AL), is currently recommended for the treatment of uncomplicated P. falciparum malaria in infants and children in Africa were analysed according to body weight group.Efficacy, safety and pharmacokinetic data from a randomized, investigator-blinded, multicentre trial of AL for treatment of acute uncomplicated The trial included 899 patients (intent-to-treat population 886). The modified intent-to-treat (ITT) population (n = 812) comprised 143 children 5 to < 10 kg, 334 children 10 to < 15 kg, 277 children 15 to < 25 kg, and 58 children 25 to < 35 kg. The 28-day PCR cure rate, the primary endpoint, was comparable across all four body weight groups . There were no clinically relevant differences in safety or tolerability between body weight groups. In the three AL body weight dosing groups , 80% of patients were aged 10-50 months, 46-100 months and 90-147 months, respectively.Efficacy of AL in uncomplicated falciparum malaria is similar across body weight dosing groups as currently recommended in the label with no clinically relevant differences in safety or tolerability. AL dosing based on body weight remains advisable. Plasmodium falciparum malaria in virtually all African countries. Artemether-lumefantrine (AL), the first fixed-dose ACT to be prequalified by the WHO, has consistently shown PCR-corrected cure rates > 95% against this species, with prompt resolution of parasitaemia and fever, rapid gametocyte clearance and good tolerance in populations of adults and children [P. falciparum infections to ACT (including AL) from non-Asian areas have been reported [Growing resistance to conventional anti-malarial drugs and the associated resurgence in infection rates and malaria-related morbidity and mortality, particularly in sub-Saharan Africa , has ledchildren -9, even children ,9-11. Chreported . While creported ,14. Due reported , AL is areported  throughoreported . A threereported  and thusreported . High lereported .Since AL was first licensed in 1999, it has been recommended that the drug be dosed according to body weight ranges, i.e. 5 to < 15 kg, 15 to < 25 kg, 25 to < 35 kg and > 35 kg . This raP. falciparum malaria in infants and children [This was a randomized, investigator-blinded, multi-centre trial undertaken to compare the efficacy, safety and tolerability of AL using a six-dose regimen of the dispersible tablet formulation versus standard crushed tablets in the treatment of acute uncomplicated children . The stuchildren . This stP. falciparum malaria. Key inclusion criteria were age \u226412 years, body weight \u22655 kg and < 35 kg, the presence of fever  or a history of fever in the preceding 24 h, P. falciparum malaria (mono or mixed infection) with an asexual blood density \u22652,000/\u03bcL and < 200,000/\u03bcL, and the absence of severe signs of complicated malaria as defined by WHO [The study population comprised infants and children with microscopically confirmed acute uncomplicated d by WHO . Key excEligible patients were randomly allocated to either dispersible or crushed tablets on a 1:1 ratio within each of the three body weight dosing groups: 5 to < 15 kg, 15 to < 25 kg, or 25 to < 35 kg. AL was administered twice daily for 3 days  with dosage determined according to body weight: one tablet per dose for children 5 to < 15 kg, two tablets per dose for those 15 to < 25 kg, and three tablets per dose for those 25 to < 35 kg. One tablet contains 20 mg artemether and 120 mg lumefantrine. Dosages according to weight show important variability between the different weight groups, with the minimum and maximum levels of drug intake both occurring in the 5-15 kg group, in which artemether intake ranges from 1.33 to 4 mg/kg and lumefantrine intake ranges from 8 to 24 mg/kg for each dose administered. Variability in the other two weight groups is much lower.Patients were hospitalized during the three-day treatment phase and followed on a weekly basis up until day 42 after initiation of treatment. Treatment was administered under supervision and consumption of some food or drink  was recommended after dosing, to increase drug absorption. Patients who vomited a dose within 1 h of treatment were given a full replacement dose, but no more than two doses were to be replaced during the three-day treatment period.Antipyretics, such as paracetamol, were used to control high fever. In the event of severe malaria or danger signs, the patient was hospitalized and received intravenous quinine. Rescue therapy according to local treatment guidelines was also administered in cases of early or late treatment failure  or if a Follow-up visits took place on days 3, 7, 14, 28, and 42 after enrolment or at any time point whenever the child was sick. Patients who prematurely discontinued either study drug or the study were scheduled for a final visit before or on day 42. If they did not attend the day 42 visit, the final outcome of the malaria episode was determined where possible. Vital signs and body temperature were assessed at regular intervals during the study. A 12-lead electrocardiogram (ECG) was recorded at baseline and on day 3 (6-10 h after the last dose) and QTc calculated using Bazett's and Fridericia's QT correction formulae. Clinical signs and symptoms were assessed at baseline and before dosing, and any symptom that worsened or started after baseline was recorded as a new adverse event. An adverse event was defined as the appearance or worsening of any undesirable sign, symptom or medical condition occurring after starting the study drug even if the event was not considered to be related to study drug. An adverse event was considered serious if it was fatal or life-threatening, resulted in persistent or significant disability/incapacity or required inpatient hospitalization or prolongation of existing hospitalization. During the hospital stay and at every follow-up visit, adverse events were assessed for severity and association with study medication.Thick and thin Giemsa-stained blood slides were prepared before each dose was administered and at every follow-up visit. Slides were examined by two independent microscopists and considered negative if no parasites were seen after examination of 200 oil-immersion fields in a thick blood film. Species determination was made based on assessment of thin films. Blood samples for PCR analysis were collected from every patient at baseline and at days 14, 28 and 42. PCR was performed centrally for all cases of recurrent parasitaemia after day 7 to distinguish recrudescence from reinfection.max)). Cmax was determined from these two concentrations, whichever was larger for a given patient. For determination of lumefantrine concentration, a sampling scheme was employed whereby samples were taken 6 h after dose 3 in 10% of patients, 6 h after dose 5 in 10% of patients, 6 h after dose 6 in 50% of patients (i.e. the expected time to Cmax), 24 h after dose 6 in 10% of patients, on day 7 in 10% of patients and on day 14 in 10% of patients. A population mean concentration-time curve was constructed by averaging all concentrations available for specified sampling intervals and taking the mean of the actual sampling times in each interval. Cmax and AUC0-last were derived from this population mean concentration-time curve.The methodology has been described previously . In brieThe primary efficacy outcome was the 28-day PCR-corrected parasitological cure rate, defined as the proportion of patients with clearance of asexual parasitaemia within 7 days of initiation of study treatment, without recrudescence within 28 days and without use of rescue medication. Secondary outcomes included the 14-day and 42-day PCR-corrected parasitological cure rate, time to parasite, fever and gametocyte clearance, and safety and tolerability profiles.P. falciparum malaria who received \u22651 full dose of study drug and underwent \u22651 relevant post-baseline efficacy assessment while on treatment. The modified intent-to-treat (mITT) population consisted of all ITT patients who completed 28 days with a valid PCR assessment (if parasitaemia was present at day 28) or who were classified as treatment failures (i.e. discontinued study drug and received rescue treatment), but excluded patients who (i) received anti-malarials or anti-malarial antibiotics before day 28 for reasons other than rescue medication; (ii) had two replacement doses and vomited a subsequent dose within 1 h or vomited the replacement dose within 2 h; (iii) switched to rescue medication therapy during the three-day treatment period; or (iv) experienced a PCR-confirmed reinfection by day 28 or had missing/unclear PCR results at day 28. The safety population comprised all patients who received \u22651 dose of study medication and underwent at least one post-baseline safety assessment.The intent-to-treat (ITT) population comprised all randomized patients with confirmed The primary end-point of 28-day PCR-corrected cure rate was assessed in the mITT population and presented with 95% confidence interval (CI) values. Time to parasite and fever clearance was evaluated by Kaplan-Meier estimates. Pharmacokinetic parameters are presented descriptively. For this evaluation, data from patients receiving the dispersible formulation of AL or standard crushed tablets were combined since the two formulations have been shown to be similar in terms of all measured efficacy, safety and pharmacokinetic parameters .Data are presented according to the following weight groups: 5 to < 10 kg, 10 to < 15 kg , 15 to < 25 kg and 25 to < 35 kg.A total of 899 patients were randomized, of whom 168 weighed 5 to < 10 kg, 379 weighed 10 to < 15 kg, 289 patients (32.1%) weighed 15 to < 25 kg, and 63 patients (7.0%) weighed 25 to < 35 kg. The ITT population included 886 patients (98.6%) and the mITT population included 812 patients (90.3%)  completed the six-dose treatment regimen. One hundred and eighty-four patients provided two blood samples each for pharmacokinetic assessment of artemether and its major active metabolite DHA, and 625 patients provided one sample each for measurement of lumefantrine concentration. As reported elsewhere , there w% completFigure Each body weight group showed a 28-day PCR-corrected cure rate > 97% . The 28-day PCR-corrected cure rate was comparable across all three body weight groups, with overlapping 95% CI values, i.e. there was no indication that outcome was affected by body weight group Table . PCR curThere was a tendency towards longer time to parasite and fever clearance in the lower body weight groups Table . A meaniP. falciparum infection (20.8%) and vomiting (16.8%), the majority of which could be attributed to malaria. Including adverse events related to malaria, the cumulative rate of adverse events to day 42 was 74.4%, 69.9%, 67.8% and 61.9% in patients weighing 5 to < 10 kg, 10 to < 15 kg, 15 to < 25 kg and 25 to < 35 kg, respectively. There was a decreasing incidence of vomiting and pyrexia and an increasing incidence of headache and P. falciparum infection with higher body weight. The cumulative incidence of these adverse events to day 42 in the 5 to < 10 kg, 10 to < 15 kg, 15 to < 25 kg and 25 to < 35 kg cohorts was, respectively: vomiting, 32.1%, 16.4%, 11.4% and 3.2%; pyrexia, 42.3%, 38.3%, 34.9% and 23.8%; headache: 0.6%, 3.7%, 13.5% and 19.0%; P. falciparum: 16.7%, 20.1%, 23.2% and 25.4%. Other than vomiting , no adverse event was reported to have a suspected relation to study drug in more than five patients. Serious adverse events were rare, occurring in 1.4% patients (n = 13) overall: 4.8% in the 5 to < 10 kg group, 0.8% in the 10 to < 15 kg group, 0.7% in the 15 to < 25 kg group and none in the 15 to < 25 kg group. In the 5 to < 10 kg group, serious adverse events (14 events in eight patients) included two episodes of initially severe malaria and two late malaria episodes on days 26 and 29 . No serious adverse events were suspected to be related to study drug.The majority of adverse events were symptoms related to malaria . Across the total study population, the most frequent adverse events were pyrexia (36.9%), cough (24.2%), Study medication was discontinued due to adverse events in 20 patients (2.2%). Vomiting was the most frequent of these events (n = 17/22 adverse events). The most frequent reason for discontinuation of the study was adverse events  and loss to follow-up . The type of adverse event leading to study discontinuation was not recorded, but the protocol stipulated that any patient who required a second replacement dose of study drug was to be discontinued.P. falciparum infection after clearing parasites within 24 h of the initial infection (5 to < 10 kg group), one on day 3 due to an unspecified infection accompanied by severe dehydration after discontinuing AL due to vomiting on day 2, at which point the patient was free of infection (5 to < 10 kg group) and one on day 7 due to haemorrhage after scarification by a witch doctor (10 to < 15 kg group). No death was suspected to be related to the study drug.Three deaths occurred: one on day 31 due to a new and severe In this large population of African infants and children with uncomplicated falciparum malaria, the high efficacy of AL was confirmed across all body weight groups. The 28-day PCR-corrected cure rate, the primary endpoint, exceeded 97% in each body weight group with no indication that outcome differed between groups. These findings confirm that a fixed-dose strategy according to certain body weight groups (i.e. use of discrete dosing for a continuous weight variable) does not result in under-dosing with reduced efficacy or, alternatively, overdosing and risk of toxicity, at the extremes of each body weight window. Thus, the current body-weight dosing recommendations for AL appear appropriate. Specifically, dosing advice for the lowest body weight group (5 to < 15 kg) and, indeed, for the smallest patients within this category (5 to < 10 kg) is adequate, as confirmed by the efficacy and safety findings. Additionally, analysis of age by body weight group in this population of children from sub-Saharan African countries showed there to be only a minor overlap of age ranges between body weight groups based on the 10-90th percentiles.In endemic areas, effective anti-malarial treatment is particularly critical in young children, who are in the process of acquiring partial immunity and present with high parasite densities and acute clinical episodes that can progress rapidly into severe, life-threatening malaria . AL expoP. falciparum infection. The higher rate of vomiting and pyrexia in smaller children is expected, and in accordance with previous reports [P. falciparum infection reported as an adverse event most likely represents new infections, due to the long follow-up period, i.e. 42 days. The assessment of adverse events in cases of malaria, however, remains challenging because of the high background of malaria signs and symptoms [Tolerability of AL was good, with no new safety concerns observed. The most frequently reported events were symptoms of malaria i.e. pyrexia, vomiting, cough and  reports  it is po reports  and the symptoms . Here, cThe study methodology was robust. The population was large and the age distribution, in which approximately 60% of patients fell into the 5 to < 15 kg body weight group, reflects the fact that young children are the most vulnerable to malaria morbidity and mortality . As specThe efficacy of AL in treating uncomplicated falciparum malaria was similar across body weight dosing groups in infants and children, with no clinically relevant differences in safety or tolerability even when the lowest body weight group was further divided into patients weighing only 5 to < 10 kg. Analysis of age by body weight group in this population of children from sub-Saharan African countries indicates that the AL dosage scheme is appropriate even in the youngest children.Novartis Pharma AG funded the study and provided editorial support for the manuscript. QB has received speaker fees and payments from Novartis to attend meetings relating to this trial. VW, ON, GL and CK are employees of Novartis. DU is employed by Medicines for Malaria Venture (MMV). The other authors have no conflicts of interest to declare.All authors contributed to the design of the study and assisted with data interpretation. QB, CM, RG, SM, AN, JL, HM, AM, MB, PO, DU and BO coordinated the study and supervised the enrolment and follow-up of patients. VW provided biostatistical support and GL conducted the pharmacokinetic evaluation. ON and CK acted as medical advisors. All authors participated in the preparation of the manuscript and approved the final version."}
+{"text": "P. vivax and P. falciparum malaria. This study compared the efficacy and safety of pyronaridine-artesunate with that of chloroquine for the treatment of uncomplicated P. vivax malaria.New antimalarials are needed for P. vivax mono-infection were randomized (1\u22361) to receive pyronaridine-artesunate (target dose 7.2\u22362.4 mg/kg to 13.8\u22364.6 mg/kg) or chloroquine (standard dose) once daily for three days. Each treatment group included 228 randomized patients. Outcomes for the primary endpoint, Day-14 cure rate in the per-protocol population, were 99.5%,  with pyronaridine-artesunate and 100%  with chloroquine. Pyronaridine was non-inferior to chloroquine: treatment difference \u22120.5% , i.e., the lower limit of the 2-sided 95%CI for the treatment difference was greater than \u221210%. Pyronaridine-artesunate cure rates were non-inferior to chloroquine for Days 21, 28, 35 and 42. Parasite clearance time was shorter with pyronaridine-artesunate (median 23.0 h) versus chloroquine , as was fever clearance time . Kaplan-Meier estimates of post-baseline P. falciparum infection incidence until Day 42 were 2.5% with pyronaridine-artesunate, 6.1% with chloroquine . Post-baseline P. vivax or P. falciparum infection incidence until Day 42 was 6.8% and 12.4%, respectively . There were no deaths. Adverse events occurred in 92/228 (40.4%) patients with pyronaridine-artesunate and 72/228 (31.6%) with chloroquine. Mild and transient increases in hepatic enzymes were observed for pyronaridine-artesunate.This phase III randomized, double-blind, non-inferiority trial included five centers across Cambodia, Thailand, India, and Indonesia. In a double-dummy design, patients (aged >3\u2013\u226460 years) with microscopically confirmed P. vivax malaria was at least that of chloroquine. As pyronaridine-artesunate is also efficacious against P. falciparum malaria, this combination has potential utility as a global antimalarial drug.Pyronaridine-artesunate efficacy in acute uncomplicated NCT00440999Clinicaltrials.gov  Plasmodium vivax malaria, with up to 400 million clinical cases annually P. vivax malaria had been underappreciated; it can even be fatal P. vivax malaria live in the tropical areas across South and South East Asia, the remainder mostly live in the Western Pacific and Eastern Mediterranean P. vivax in areas where it had previously been eradicated, such as Korea and China, is of major concern An estimated 2.85 billion people are at risk of P. vivax treatment in most areas of the world for the past 50 years. However, P. vivax susceptibility to chloroquine has declined in some regions,  and new approaches are required urgently The blood schizontocide chloroquine and the tissue schizontocide primaquine have been the mainstay of Plasmodium vivax is highly sensitive to artemisinins P. vivax, artemisinin-based combination therapy (ACT) is recommended by The World Health Organization P. falciparum malaria P. vivax and P. falciparum are sympatric, ACT should have good efficacy (>95%) against both parasites, as differential diagnosis may not be feasible or reliable The blood stage of P. vivax and P. falciparum malaria. Pyronaridine is especially interesting as a component of ACT because in vitro studies using recent isolates indicate potent activity against chloroquine-resistant P. vivax and P. falciparumP. vivax similar to that of chloroquine P. falciparum malaria at Day 28 were >99%, and the combination was generally well tolerated Pyronaridine-artesunate is being developed as an ACT for use against P. vivax malaria. The primary objective of this clinical study was to compare the efficacy and safety of the fixed combination of pyronaridine-artesunate with that of standard chloroquine therapy in adults and children with acute, uncomplicated P. vivax malaria.This is the first pyronaridine-artesunate Phase III study reported for the treatment of The protocol for this trial and supporting CONSORT checklist are available as supporting information; see This study was conducted according to the principals of Good Clinical Practice and the Declaration of Helsinki and complied with all relevant regulatory requirements. The trial protocol was approved by institutional review boards in Cambodia , Thailand , India , and Indonesia . All patients or their guardians provided written informed consent and assent was required from children old enough to understand the study.This was a multi-center, randomized, double-blind, double-dummy, parallel-group comparative, non-inferiority trial conducted between March 2007 and March 2008 in local hospitals across five centers: Pailin, Cambodia; Mae Sot and Mae Ramat, Thailand; Mangalore, India; and Maumere (Island of Flores), Indonesia. There were no changes to the protocol after study commencement.P. vivax mono-infection confirmed by positive microscopy of P. vivax with a parasite density \u2265250 \u00b5L\u22121 of blood  and fever or documented history of fever in the previous 24 h, and with a body weight between 20 and 90 kg  were eligible for enrollment. Patients were excluded from the study if they had: any other condition requiring hospitalization; anemia (hemoglobin <8 g/dL); hepatic or renal impairment; presence or history of clinically significant disorders; known hypersensitivity to study drugs or excipients; known active hepatitis A IgM, hepatitis B surface antigen, hepatitis C antibody or seropositive for HIV antibody; used an antimalarial within the previous two weeks (urine test required); used an antibacterial with anti-malarial activity within the previous two weeks; received an investigational drug within the past four weeks; previously participated in the study. Women who were pregnant or lactating were also excluded. Agreement to use an acceptable method of contraception during the study was required from all women of child-bearing age.Male or female subjects between three and 60 years of age with acute uncomplicated Study drugs were pyronaridine-artesunate tablets 180\u223660 mg, chloroquine tablets 155 mg, and matching placebos . Study drugs were given orally with water once daily for three days  under direct observation of study staff. Vomiting of the first dose of study drug within 30 minutes of administration resulted in re-dosing. Vomiting of subsequent doses resulted in withdrawal from the study and treatment with rescue medication . For pyronaridine-artesunate, drug dose was based on body weight: 20\u201325 kg, 1 tablet; 26\u201344 kg, 2 tablets; 45\u201364 kg, 3 tablets; and 65\u201390 kg, 4 tablets, i.e. a pyronaridine-artesunate target dose of between 7.2\u22362.4 mg/kg and 13.8\u22364.6 mg/kg. The chloroquine dose for adults was 620 mg on Days 0 and 1 and 310 mg on Day 2. The chloroquine target dose for children was 10 mg/kg on Days 0 and 1 and 5 mg/kg on Day 2.All subjects remained hospitalized for Days 0\u20133, returning for follow-up assessments on Days 7, 14, 21, 28, 35 and 42. A medical history was taken at screening and physical examination performed at screening, Days 3, 28 and 42. Vital signs were monitored at screening, on Days 1, 2, 3 and 7 and at other visits if indicated. Clinical signs and symptoms of malaria were assessed at screening, Days 1, 2 and 3 and at other visits if indicated. Body temperature was taken every 8 h over at least 72 h following the first study drug administration or temperature normalization for at least two readings 7\u201325 h apart, then at each visit. Venous blood was drawn for hematology at screening, Days 3, 7, 28, and 42, and for clinical laboratory tests at screening, Days 3, 7, and 28, and Day 42 if clinically indicated. Urinalysis was performed on the same schedule. Adverse events were monitored throughout the study. 12-lead electrocardiograms were performed at screening, Day 2 and at Days 7, 14 and 42 if clinically indicated.Blood samples for testing glucose-6-phosphate dehydrogenase (G6PD) deficiency were collected on pre-printed filter paper. A semi-quantitative fluorescent spot test was used to distinguish between G6PD-deficient samples and those with normal or intermediate activity . Patients completing the study to Day 28 with normal G6PD activity were given a 14-day course of primaquine (15 mg/day for adults and 0.3 mg/kg/day for children) starting on Day 28 after all required assessments had been performed. G6PD-deficient patients completing the study to Day 28 were treated as per country policy.Plasmodium species, duplicate thin blood smears were taken at screening, pre-dose on Day 1 and at visits from Day 7 whenever parasitological blood samples were taken. Patients experiencing treatment failure were administered rescue antimalarial therapy as per local practice. All procedures scheduled for Day 28, including preparation of thin and thick films were performed before rescue medication was administered.Parasite density was determined by duplicate Giemsa-stained thick blood films examined independently by two microscopists; the mean of the two readings was recorded. In the case of discrepancy (i.e. >30%), a third microscopist reviewed the slides and the principal investigator made the final classification. Thick films were taken every 8 h until at least 72 h or until a negative smear was recorded at least 7\u201325 h apart, then at each visit, or if the patient returned unscheduled. Asexual parasite and gametocyte counts were recorded separately at screening; total parasite counts were recorded at all other assessments. Parasites were enumerated against 200 white blood cells (WBC) using 1,000\u00d7 magnification. When the number of asexual parasites dropped below 10 per 200 WBC, counting was done against at least 500 WBC. A blood slide was considered negative if no asexual parasites were observed per 1,000 WBC. External quality control, was conducted by a central independent laboratory, blinded to treatment . All slides from patients with treatment failure were re-examined, plus the first 5 slides from each site and 5% of all subjects randomly. Results confirmed that procedures were being conducted as per the defined protocol. To determine P. vivax malaria.The primary objective of this clinical study was to compare the efficacy and safety of the fixed combination of pyronaridine-artesunate (180\u223660 mg) with that of standard chloroquine therapy in adults and children with acute, uncomplicated P. vivax illness requiring hospitalization in the presence of parasitemia; or presence of parasitemia and axillary temperature \u226537.5\u00b0C any time between Days 3 and 14; or presence of parasitemia on any day between Days 7 and 14, irrespective of clinical condition. Subjects presenting with non-P. vivax malaria after clearance of P. vivax parasites were withdrawn from the study and given treatment for the new infection. If non-P. vivax malaria occurred on or after Day 14, the subject was considered a success for Day 14.Efficacy endpoints followed WHO 2002 guidelines for monitoring antimalarial drug efficacy, as were relevant at the time that the study was designed Secondary efficacy measures included: Day 28 cure rate; parasite clearance time, defined as the time from first treatment dose to aparasitemia for two consecutive negative readings taken between 7\u201325 h apart; fever clearance time, defined as time from first treatment dose to apyrexia for two consecutive normal temperature readings taken between 7\u201325 h apart; the proportion of patients aparasitemic on Days 1, 2, and 3; and the proportion of patients apyretic on Days 1, 2, and 3. The cure rates on Days 21, 35 and 42 were included as exploratory endpoints.Safety outcomes included: the frequency, nature and severity of adverse events; the frequency of serious adverse events; the frequency of adverse events considered by the investigator to be related to study medication; the reasons for study withdrawals; abnormal electrocardiograms; and abnormal hematology and clinical chemistry results. An adverse event was defined as any unfavorable and unintended sign, symptom, syndrome, or illness that developed or worsened during the period of observation in the clinical study. A serious adverse event was defined as one that resulted in: death or was life threatening; hospitalization or a prolongation of hospitalization; a congenital abnormality or birth defect; a persistent or significant disability or incapacity; or was judged by the investigator to be serious. Drug-related adverse events were determined by the investigator, based on the temporal relationship to drug therapy, any corroborating laboratory data, and improvement when drug was discontinued.Assuming a 90% cure rate on Day 14 in both treatment groups and a non-inferiority limit of \u221210%, a sample size of 410 evaluable subjects randomized in a 1\u22361 ratio (205 subjects in each treatment group) would provide 90% power to demonstrate non-inferiority of pyronaridine-artesunate compared to chloroquine, using a 2-sided 95% confidence interval (CI). Assuming a dropout rate of 10%, 456 subjects would need to be randomized to the study (228 subjects in each treatment group). Assuming a Day-14 cure rate of 95%, this sample size would provide >99% power to demonstrate non-inferiority of pyronaridine-artesunate compared to chloroquine.A computer-generated randomization scheme was provided by the sponsor. Subjects were randomized 1\u22361 within each study site in blocks of six to receive either pyronaridine-artesunate plus matching chloroquine placebo or oral chloroquine plus matching pyronaridine-artesunate placebo. Randomization numbers were assigned in ascending order to each subject according to the order recruited. The subject was allocated an individually numbered treatment pack, which contained sufficient tablets for 3 days' therapy plus an overage bottle containing tablets in case the subject vomited the first dose. Study drugs were administered on a double-blind, double-dummy basis. The investigator calculated the appropriate dose and study drug was administered by a different member of staff, designated by the investigator. All study investigators, laboratory technicians and patients were blind to treatment assignment. Active drugs and placebos were packaged similarly. Sealed opaque envelopes containing the study medication assignment for each subject were provided to the study site investigator for use in an emergency; no code breaks were required.The intent-to-treat (ITT) population was defined as all randomized subjects who received any amount of study medication. The per-protocol (PP) population included all randomized patients who completed a full course of study medication, had a known primary efficacy endpoint at Day 14 and did not violate the protocol in a way that would compromise efficacy evaluation, i.e. did not use prohibited concomitant medication; have a concomitant medical condition or infection that may have affected treatment outcome; or have any major protocol deviation.For this non-inferiority study, the primary efficacy endpoint of Day-14 cure rate was assessed in the PP population. Non-inferiority was demonstrated if the lower limit of the 2-sided 95% confidence interval (CI) for the difference in Day-14 cure rates was greater than \u201310%. The CI was calculated according to the Newcombe\u2013Wilson method without continuity correction. Furthermore, exact (Pearson\u2013Clopper) 95% CIs were calculated for the Day-14 cure rate in each of the two treatment groups. A similar analysis was repeated for the proportion of subjects with cure on Days 21, 28, 35 and 42.Parasite clearance time and fever clearance time were summarized using Kaplan-Meier estimates (log-rank test). Subjects without confirmed parasite clearance time or fever clearance time within 72 h after the first dose of study drug were censored at that time point.P. falciparum infection and for time to reappearance of any parasite  were also conducted using Kaplan-Meier estimates. All statistical evaluations were performed using SAS\u00ae, Version 9.1.3 in a UNIX environment.Post-hoc analyses of time to P. falciparum infection compared with 5/228 (2.2%) in the pyronaridine-artesunate group. More patients were excluded from the PP population in the chloroquine group, 19/228 (8.3%), than the pyronaridine-artesunate group, 10/228 (4.4%), mainly because of missing efficacy data and insufficient drug therapy.Patient disposition is shown in Patient baseline demographic and clinical characteristics are shown in Outcomes for the primary endpoint, cure rate at Day 14 in the PP population, were 99.5%,  in the pyronaridine-artesunate group and 100%  in the chloroquine group. Pyronaridine was non-inferior to chloroquine for the primary endpoint: treatment difference \u22120.5% . All 27 children \u226412 years were cured at Day 14. Day-14 cure rates were 100% in all study centers, except for the one failure in the pyronaridine-artesunate group, which occurred in a patient from Maumere, Indonesia . Results for Day-14 cure rate in the ITT population were supportive of the primary analysis; 95.2%;  with pyronaridine-artesunate and 93.0%  with chloroquine, treatment difference 2.2% . At all efficacy assessments from Day 14 until Day 42, pyronaridine-artesunate was non-inferior to chloroquine, with cure rates >95% .Using Kaplan\u2013Meier analysis of the ITT population, time to parasite clearance was shorter in the pyronaridine-artesunate group compared with the chloroquine group  patients in the pyronaridine-artesunate group and 179/228 (78.5%) patients in the chloroquine group. Therefore, evaluation of the anti-pyretic activity of pyronaridine-artesunate and chloroquine cannot be fully assessed. However Kaplan-Meier analysis of the ITT population showed a shorter time to fever clearance with pyronaridine-artesunate than with chloroquine  patients experienced a treatment-emergent adverse event of any cause compared with 72/228 (31.6%) in the chloroquine group . IncreasThere were no deaths during the study. Two patients, both in the pyronaridine-artesunate group, had serious adverse events ; neither was considered drug related by the investigator. Two patients, both in the chloroquine group, had adverse events leading to drug discontinuation and study withdrawal ; all considered possibly related to drug treatment by the investigator.Baseline hemoglobin levels were similar in the two treatment groups . A transBiochemistry laboratory observations were generally similar in both treatment groups, with the exception of alanine transaminase (ALT) and aspartate transaminase (AST) . From DaClinically significant electrocardiograms were recorded for 1/226 (0.4%) patients in the pyronaridine-artesunate group and 6/222 (2.7%) in the chloroquine group. All cases were recorded as QTc prolongation. All except one event (in the chloroquine group) was considered unrelated to study treatment by the investigator.P. vivax malaria with a Day-14 cure rate of 99.5% that was non-inferior to that of chloroquine (100%). Non-inferiority of pyronaridine-artesunate to chloroquine was maintained throughout the study. However, primaquine treatment was initiated in most cases after Day 28, making comparisons after this time point difficult. There are no comparative data of pyronaridine-artesunate in P. vivax malaria. Results for chloroquine were comparable to other studies in Thailand and India (100% efficacy) P. vivaxPyronaridine-artesunate was highly efficacious in the treatment of uncomplicated P. vivax malaria have also demonstrated rapid clearance of parasites and fever The clinical efficacy of the artemisinin derivatives is characterized by an almost immediate onset of activity and rapid reduction of parasitemia P. vivax or P. falciparum post-baseline infection was lower with pyronaridine-artesunate versus chloroquine, mainly because of the difference in P. falciparum post-baseline infections. Post-baseline infection with P. falciparum was less frequent and occurred later in the study with pyronaridine-artesunate versus chloroquine. Given the potential severity of P. falciparum malaria, this difference is clinically important. P. falciparum chloroquine resistance is widespread in vitro activity against chloroquine-resistant P. falciparumP. falciparumP. falciparum infections in the chloroquine group. However, we cannot confirm this as susceptibility or molecular testing of isolates was not performed.The incidence of P. falciparum malaria Adverse events of any cause with pyronaridine-artesunate in this study were consistent with reports for pyronaridine and artemisinins as monotherapy in falciparum malaria P. vivax and P. falciparum infection and P. falciparum reinfection are common in areas endemic for P. vivax. Practical difficulties in reliably distinguishing between the two pathogens means that, in practice, many malaria cases will be diagnosed clinically and treated empirically P. vivax/P. falciparum infection were excluded. However, our design does reflect the clinical situation in the study region where known P. falciparum malaria would not be treated with chloroquine because of the high risk of failure from widespread resistance to this agent P. vivax and mixed P. vivax/P. falciparum infection and would compare pyronaridine-artesunate to another ACT.Mixed P. vivax chloroquine resistance. In vitro data indicate high activity for pyronaridine against chloroquine-resistant P. vivaxIn this Phase III trial, the study population was restricted in order to allow comparison of the two treatments and probably does not reflect the baseline clinical and demographic characteristics of the general population of malaria patients in the region. Further studies are required to assess pyronaridine-artesunate in the wider clinical setting. Further data are also required to assess pyronaridine-artesunate efficacy in areas of high P. vivax malaria, particularly because this group is the most vulnerable to severe outcomes Our study included only 27 children, all over 7 years of age (probably because of the 20 kg minimum weight requirement). Pyronaridine-artesunate efficacy and safety data are required in very young children with P. vivax. Post-baseline P. falciparum infection was reduced and delayed with pyronaridine-artesunate versus chloroquine. Based on the results of this study in P. vivax and those conducted against P. falciparumIn conclusion, pyronaridine-artesunate had non-inferior efficacy and faster parasite and fever clearance compared with chloroquine against Checklist S1CONSORT checklist.(0.23 MB DOC)Click here for additional data file.Protocol S1Trial Protocol.(0.62 MB PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria. Data from seven studies supported by Novartis (1996\u20132007), including 647 adults  and 1,332 children  with microscopically confirmed uncomplicated P. falciparum malaria and treated with the recommended regimen of AL, were pooled. The 28-day polymerase chain reaction\u2013corrected parasitologic cure rate (primary efficacy endpoint) was 97.1% (495 of 510) in adults and 97.3% (792 of 814) in children . Gametocytemia prevalence after day was 4.2% (23 of 554) in adults and 0.9% (8 of 846) in children. No noteworthy safety signals were observed. Serious adverse events occurred in 1.4% of the adults and 1.3% of the children. This study is the largest data set to date assessing AL therapy for treatment of acute uncomplicated P. falciparum malaria. Artemether-lumefantrine showed high cure rates and rapid resolution of parasitemia, fever, and gametocytemia in adults and children, and showed an excellent safety and tolerability profile.Randomized trials have confirmed the efficacy and safety of artemether-lumefantrine (AL) for treatment of uncomplicated  Plasmodium falciparum malaria.6The World Health Organization (WHO) recommends that artemisinin-based combination therapies (ACTs) be used as first-line treatment of uncomplicated \u00ae, Novartis Pharma AG, Basel, Switzerland) became the first ACT to be pre-qualified by WHO, and it is now used as first-line treatment for uncomplicated Plasmodium falciparum malaria in many regions worldwide.P. falciparum malaria should be at least 90% and preferably > 95%.1In 2004, artemether-lumefantrine \u2013corrected parasitologic cure rate for AL at day 28 was > 95% in all the trials reviewed.6A recent Cochrane analysis evaluated the efficacy of ACTs for the treatment of uncomplicated P. falciparum malaria during 1996\u20132007. The analysis population was derived from six studies undertaken in Asia and Africa and one study in patients from Europe and Colombia. This, it encompasses regions with varying endemicity and transmission patterns.We report efficacy and safety findings for AL in a post-hoc pooled analysis of data from the Novartis study database, which includes approximately 2,000 children and adults treated for uncomplicated P. falciparum malaria was based on microscopically confirmed evaluation of Giemsa-stained blood slides. Patients with severe or complicated malaria were not included in this analysis. Study B2303 used an investigator-blinded design and A025 used a double-blind design; the remaining studies were open label. Six of the studies were performed in malaria-endemic countries Data from clinical trials within the Novartis database were included in this unplanned pooled analysis if they enrolled patients treated with the recommended regimen of AL, administered twice a day for three days, and diagnosis of In all studies, AL dosing was based on body weight: 5\u2013< 15 kg, 1 tablet per dose; 15\u2013< 25 kg, 2 tablets per dose; 25\u2013< 35 kg, 3 tablets per dose; \u2265 35 kg, 4 tablets per dose. Patients who vomited the first dose within one hour of treatment received a full replacement in all studies. The protocols of A2403 and B2303 specified that no more than two doses were to be replaced during the entire treatment period.P. falciparum genes encoding merozoite surface protein 2, merozoite surface protein 1, and glutamate-rich proteinSix of the studies  recorded the 28-day parasitologic cure rate, corrected for re-infection by PCR in studies A025, A026, A028, A2403, and B2303 , consistent with draft guidance from the FDA. The PCR analysis was performed by analysis of size polymorphisms in the The main efficacy variable was the 28-day PCR-corrected parasitologic cure rate, which was defined as the proportion of patients with clearance of asexual parasitemia within 7 days of the first dose of AL and without subsequent recrudescence within 28 days. Recrudescence was defined as reappearance of the original parasite strain as confirmed by PCR-based genotyping. Other efficacy endpoints were the 28-day non\u2013PCR-corrected parasitologic cure rate, time to parasite clearance, time to fever clearance, and proportion of patients with gametocytes at specific time points. The times at which patients were assessed for parasite clearance and fever clearance varied between studies. The presence of gametocytes was recorded at baseline, > 0\u201372 hours, > 72 hours\u2013day 7, and > day 7.Safety data included the incidence, type, and study drug relationship of adverse events. Adverse events were defined as the appearance or worsening of any undesirable sign, symptom, or medical condition after starting the study drug, even if the event was not considered to be related to the study drug, according to the International Conference on Harmonization Guidelines. A serious adverse event was defined as any untoward medical occurrence that resulted in death, was life-threatening, required inpatient hospitalization or prolongation of existing hospitalization, resulted in persistent or significant disability/incapacity, or was a congenital anomaly/birth defect. Other safety data included the incidence of death and serious adverse events, clinical laboratory assessments, and vital signs. The availability of laboratory data varied between studies in terms of parameters measured and time of sample collection. Creatinine clearance was used as a measure of renal function, and was calculated according to the Cockcroft-Gault formula in adults40Patients > 16 years of age were included in the adult analysis populations; patients \u2264 16 years of age were included in the pediatric analysis population. Efficacy and safety data were taken from the original study databases.P. falciparum malaria who received at least one dose of study drug. This definition was selected to comply with FDA draft guidance.post hoc analysis of the primary efficacy endpoint, 28-day PCR-corrected parasitologic cure rate, was performed on the basis of data only from study A2401 by using the last observation carried forward technique.Efficacy analyses were based on the modified intent-to-treat (mITT) population, which was generally defined as all patients with parasitologically confirmed The 28-day PCR-corrected parasitologic cure rate was also reported for subpopulations of the evaluable population, according to the following variables: 1) baseline parasite count ; 2) sex; 3) baseline renal function based on creatinine clearance  ; 4) baseline hepatic function ; and 5) age (pediatric population only)  and body weight .All data are presented descriptively. One-sample 95% confidence intervals (CIs) for the 28-day PCR-corrected cure rate were calculated by using the Pearson-Clopper method. Kaplan-Meier estimates of parasite clearance time and fever clearance time were calculated. Laboratory data are presented as standard summary statistics. Because of the variation between studies in the time points used for laboratory evaluations and deviations from schedules, time windows were defined. If a patient had more than one value within a given interval, the mean was used for summarization. Summary statistics for laboratory data should be interpreted carefully in view of the variation between time windows and the numbers of patients with available data. Safety analyses were based on all randomized/enrolled patients who received at least one dose of study drug. Statistical analyses were performed using the SAS software system .All studies were conducted according to the Declaration of Helsinki, and informed consent was obtained from all patients (or parents/guardians where appropriate) according to appropriate institutional review board approval.For adults, the mITT population comprised 599 patients, of whom 437 took part in studies from malaria-endemic regions , and 162 were travelers from non-endemic regions (study A2401) . The evaPatient demographics and baseline disease characteristics are shown in The full recommended regimen of AL was taken by 96.9% of adults (n = 627) and 97.1% of children  (safety populations). Seven adults (1.1%) and 140 children (10.5%) vomited at least one dose of AL.post hoc analysis using the last observation carried forward technique showed a 90.7% cure rate (147 of 162).Parasitologic cure rates on day 28 among the pooled efficacy population are summarized in One potential explanation for treatment failures may be underdosing of larger patients. In the evaluable population of study A2401, the cure rate was 93.0%  in patients weighing \u2265 70 kg versus 100%  in patients < 70 kg. Of the five patients weighing \u2265 70 kg who were classified as treatment failures, one patient discontinued treatment after the second dose because of signs and symptoms of severe malaria that were already present at baseline, one missed an assessment at day 6 but was clear of parasites at day 7, and three had recrudescence during days 21 and 28. In addition, of four patients with body weights >100 kg, three were clear of parasites at day 28, and one did not have a day 28 blood assessment, but was parasite free at 58 and 154 hours.In the pediatric population, the 28-day PCR-corrected parasitologic cure rate was 97.3%  in the evaluable population and 93.4%  in the mITT population. No differences were observed between age groups or different body weight categories, including patients \u2264 2 years of age or those weighing < 10 kg .Baseline parasite density did not appear to affect the 28-day PCR-corrected cure rate in either adults or children . There wThe median time to parasite clearance was 42.3 hours (95% CI = 41.5\u201343.2 hours) in the adult mITT population, and parasite clearance was achieved within 48 hours in 78.6% of patients . For chiP. falciparum count  was associated with slightly shorter fever clearance time  compared with those with parasite counts > 100,000/\u03bcL . For study A2401, the median time to fever clearance among patients with fever at baseline was 36.5 hours (95% CI = 27.8\u201339.5 hours in the mITT population).In adults, the median time to fever clearance was 28.5 hours (95% CI = 22.3\u201334.0 hours). Median time to fever clearance was shorter in patients with a baseline parasite count \u2264 100,000/\u03bcL  compared with those with a baseline count > 100,000/\u03bcL . The median fever clearance time in children  was markedly shorter than that in adults . As in adults, a low baseline P. falciparum gametocytes at baseline was 9.7% (58 of 596), which decreased to 4.2% (23 of 554) after day 7  of 647 adult patients. The most frequently reported adverse events were non-specific  and consistent with symptoms of acute malaria . AdverseP. falciparum infection, which was reported by investigators as a serious adverse event in two patients. Of the 22 serious adverse events, 18 occurred in study A2401, in which prolongation of hospitalization required a classification of serious adverse event. There were no cases of urticaria or anaphylactoid reaction reported as serious adverse events. Seven serious adverse events in three patients had a suspected relation to AL . An additional 13 serious adverse events were not suspected to be related to AL, and the relationship with AL was unknown for the final two events.No adult patient died. Nine patients (1.4%) experienced 22 serious adverse events. No type of serious adverse event occurred in more than one patient except for The overall incidence of adverse events in patients with normal renal function or mild impairment was 77.8% (175 of 225) and 78.3% (112 of 143), respectively. The small number of patients with moderate renal impairment (n = 10) each reported one or more adverse events. Subgroup analysis by hepatic function at baseline showed that the incidence of adverse events in adult patients with normal hepatic function or mild, moderate, or severe hepatic impairment at baseline was 71.9% (120 of 167), 79.3% (96 of 121), 83.3% (50 of 60), and 100% (17 of 17), respectively. Many of the adverse events that occurred more frequently in the presence of hepatic impairment were well-recognized symptoms of malaria.One or more adverse events were reported in 970 (72.8%) of 1,332 children; pyrexia (28.6%) and cough (22.7%) occurred most frequently . AdverseP. falciparum infection on day 28 (new infection or recrudescence was undetermined) in a patient who was parasite free \u2264 24 hours and still parasite free on day 13; gastroenteritis on day 8 with diarrhea treated with oral rehydration therapy; an unspecified infection with pyrexia and dehydration treated with quinine, paracetamol, metoclopramide, and amoxicillin; and hemorrhage on day 5 after scarification by a traditional healer; all deaths were considered unrelated to AL by investigators. Thirty serious adverse events were reported by investigators in 17 patients (1.3%), including P. falciparum infection (7), convulsion (3), pyrexia (2), and anemia (2). None of the three cases of convulsion was considered to be related to AL; each was reported together with malaria or pyrexia, and may have represented febrile convulsions. All other serious adverse events were reported in no more than one patient. One serious adverse event was reported as having a suspected relationship with AL. This event was a case of urticarial rash in a four-year-old patient that resolved \u2264 5 days after antihistamine treatment.Four (0.3%) children died. Deaths were caused by severe P. falciparum infection plus anemia in one patient, P. falciparum infection plus iron deficiency anemia in another patient, a lower respiratory tract infection in one patient, and urticaria in one patient as described above.Seventy-one children (5.3%) left the study prematurely because of adverse events. Seventy of these 71 children were enrolled in study B2303, in which the protocol specified that any patient who required a second replacement dose of study drug was to be removed from the study. Four patients stopped treatment because of an adverse event other than vomiting. These events were Pediatric patients with mild, moderate, or severe renal impairment at baseline had more adverse events than those without renal impairment . Children with mild or moderate hepatic impairment had a higher incidence of adverse events than those with normal function  but numbers of patients with moderate (n = 24) or severe (n = 2) hepatic impairment were low.9/L, 3.9 \u00b1 1.9 \u00d7 109/L, and 130 \u00b1 75 \u00d7 109/L, respectively, in adults, and 9.3 \u00b1 1.7 g/dL, 9.3 \u00b1 3.8 \u00d7 109/L, 4.8 \u00b1 2.8 \u00d7 109/L, and 181 \u00b1 99 \u00d7109/L in children. At days 26\u201339, the corresponding values were 12.8 \u00b1 1.7 g/dL, 7.4 \u00b1 2.2 \u00d7109/L, 3.9 \u00b1 1.7 \u00d7 109/L, and 231 \u00b1 81 \u00d7 109/L in adults and 10.4 \u00b1 1.4 g/dL, 9.1 \u00b1 3.3 \u00d7 109/L, 3.5 \u00b1 1.9 \u00d7 109/L, and 310 \u00b1 130 \u00d7 109/L in children. Renal function (assessed by creatinine clearance) increased from baseline and liver function values  decreased from baseline, which is consistent with malaria and its resolution. Hemolysis-related adverse events were not reported in adults. Four potential hemolysis-related adverse events occurred in the pediatric population (0.3%), all of which were increased reticulocyte counts.Hematogic parameters improved during follow-up, which is consistent with resolution of acute malaria. Mean values \u00b1 SD for hemoglobin level, leukocyte count, neutrophil count, and platelet count at baseline were 12.3 \u00b1 2.5 g/dL, 6.1 \u00b1 2.4 \u00d7 10P. falciparum malaria, AL achieved high cure rates and rapid symptom relief in adults and children and a good safety and tolerability profile. The efficacy of AL was similar for patients of all ages in malaria-endemic regions. In the only study of travelers from non-endemic regions, analysis was complicated by poor follow-up rates, but efficacy of AL also appeared to be excellent in the per protocol population.In this pooled analysis, the largest of its type so far to assess AL therapy for the treatment of acute uncomplicated The observed day 28 PCR-corrected cure rate in evaluable patients was approximately 97% in adults and children, which exceeded the recommendation of WHO that antimalarial drugs should show a cure rate of 95%.Recent reports that showed decreased rates of parasite clearance after treatment with artesunate and ACTs in western Cambodia are of concern.Parasite and fever clearance were achieved rapidly after treatment with AL in adult and pediatric populations. The slower fever clearance observed in study A2401 may have been caused by less frequent evaluation in this study and the relatively small proportion of patients (approximately 20%) who used antipyretics compared with other trials .P. falciparum malaria. In our analysis, the proportion of patients with gametocytemia after day 7 decreased by more than 50% from baseline in adults and by approximately 80% in children. Less than 1% of treated children were positive for gametocytes after day 7.The reduction in gametocytemia after treatment with AL is noteworthy. Clearance of gametocytes, the parasite form that infects mosquitoes, breaks the cycle of transmission, and thus contributes to malaria control. Artemisinins have gametocytocidal properties and reduce gametocyte carriage,P. falciparum malaria.Our analysis showed no unexpected safety concerns in almost 2,000 patients treated with AL. These results are consistent with those of other studies that demonstrated excellent safety and tolerability of AL.Unforeseen differences were not observed in the adverse event profile or laboratory results between subgroups. The higher rate of adverse events reported in patients with hepatic or renal impairment is likely to reflect more severe malaria in these groups. Cough was more frequent in children than in adults, which is not unexpected because respiratory infections are common in children in Africa with malaria.This pooled analysis had some limitations. First, as an unplanned pooling of data, the seven studies used various schedules for assessment and procedures for reporting adverse events. Accordingly, the reported rates of events cannot reliably be compared between trials. However, it is unlikely that this limitation impaired detection of any novel safety signals. Second, three of the seven studies were undertaken before 2000, i.e., more than 10 years ago, although there is no evidence to suggest that the efficacy and safety of AL have changed in the past decadeP. falciparum malaria exceeded 95% in adults and children, and showed a good safety and tolerability profile and rapid reduction in gametocytemia. Although data for non-immune persons remain limited, our findings are consistent with the recommendation that AL be used as first-line treatment of uncomplicated P. falciparum malaria in patients of all ages, whether they are semi-immune residents or non-immune visitors in malaria endemic or non-endemic regions.In conclusion, in this pooled analysis of almost 2,000 patients, the PCR-corrected 28-day cure rate for AL treatment of uncomplicated"}
+{"text": "Each year, thousands of cases of uncomplicated malaria are imported into Europe by travellers. Atovaquone-proguanil (AP) has been one of the first-line regimens used in France for uncomplicated malaria for almost ten years. While AP\u2019s efficacy and tolerance were evaluated in several trials, its use in \u201creal life\u201d conditions has never been described. This study aimed to describe outcome and tolerance after AP treatment in a large cohort of travellers returning from endemic areas.Between September 2002 and January 2007, uncomplicated malaria treated in nine French travel clinics with AP were followed for 30\u00a0days after AP initiation. Clinical and biological data were collected at admission and during the follow-up.A total of 553 patients were included. Eighty-eight percent of them were born in Africa, and 61.8% were infected in West Africa, whereas 0.5% were infected in Asia. Migrants visiting friends and relatives (VFR) constituted 77.9% of the patients, the remainder (32.1%) were backpackers. Three-hundred and sixty-four patients (66%) fulfilled follow-up at day 7 and 265 (48%) completed the study at day 30. Three patients had treatment failure. One-hundred and seventy-seven adverse drug reactions (ADR) were reported during the follow-up; 115 (77%) of them were digestive ADR. Backpackers were more likely to experiment digestive ADR compared to VFR . Twenty patients had to be switched to another regimen due to ADR.This study seems to be the largest in terms of number of imported uncomplicated malaria cases treated by AP. The high rate of reported digestive ADR is striking and should be taken into account in the follow-up of patients since it could affect their adherence to the treatment. Beside AP, artemisinin combination therapy (ACT) is now recommended as first-line regimen. A comparison of AP and ACT, in terms of efficacy and tolerance, would be useful. In 2010, the World Health Organization (WHO) estimated that 216 million cases of malaria occurred and were responsible of 650,000 deaths . In EuroAP has been licensed in France since 1997 and marketed since late 2001 and it remains the principal treatment of acute malaria. Since the National Experts Committee recommended in 2007 that AP, together with artemether-lumefantrine, be used as first-line regimen, \u2018old drugs\u2019 such as quinine or mefloquine were downgraded to second-line treatment . AtovaquPlasmodium falciparum malaria cases. It provided interesting data on the use of AP with a relatively large series (n\u2009=\u2009253), but heterogeneity of practices, due to \u2018centre effects\u2019 between participating European centres may limit the interpretation of the study ) (Table\u00a0Assessment of tolerance by patients for the 437 who answered to the questionnaire was classified as good for 304 (69.6%), satisfactory for 116 (23.8%), bad for 41 (8.4%) and very bad for 27 (5.5%). In 20 cases, a switch to another drug was reported mainly because of vomiting , confusion , headache , cutaneous eruption , and suspected resistance because of a positive smear at day 3 .P. falciparum malaria seems to be the largest series assessing the use of AP in the field of imported malaria. Patient profile  are similar to those observed in the majority of studies on imported malaria [This \u2018real life condition\u2019 prospective, observational study of 553 patients treated by AP for uncomplicated  malaria -24,26-28Not surprisingly, the rate of chemoprophylaxis and exposure prophylaxis was low. Clinical and biological presentation had no specificity but it is of interest to note that nearly one third of patients had no fever at admission, which might be misleading for non-experienced practitioners. Compared to some other studies describing non-comparative cohorts of malaria patients, follow-up, even too low, might be considered satisfactory given such a \u2018real life\u2019 design for this cohort, since outcome data are available for the majority at day 3 and since nearly half of patients were seen one month after treatment -21,26-29With regard to efficacy, if a majority of patients (95%) were fever-free at day 3, nearly a third of them were still parasitaemic confirming that AP is slow-acting ,20,30,31Even though rare under AP and comparable to other malaria treatment (1% in this study\u2019s series), the risk of relapse has to be considered by physicians given the potential severe outcome at a time when the diagnosis of malaria may be omitted (long delay after travel in endemic area) ,21,23. GPerceived efficacy of AP was satisfying. This evaluation is not as valid as parasitological efficacy but it has not been studied in Africa before, both for curative treatment and for prophylaxis in travellers. It is considered as a significant predictor for compliance to treatment and preventative behaviours ,37.With regard to tolerance of AP, data of this study highlighted a high rate of digestive ADR, mainly at day 3, especially nausea and vomiting. Digestive ADR represented 77% of ADR reported at day 3 and 72% of the totality of ADR reported during follow-up. It seems that digestive ADR were more frequent in backpackers compared to migrants, with no particular explanation, and no comparable data were found in the literature. In 18% of cases (n\u2009=\u200915), vomiting was severe enough to justify a change to second-line treatment. This high occurrence of digestive ADR could be partially explained by the population enrolled in the study since black people are known to have a lower clearance rate of atovaquone compared to white people . Yet theSurprisingly, these digestive side effects are one of the most important from studies. A review of ten trials comparing Atovaquone-Proguanil (AP) with other anti-malarial drugs for uncomplicated malaria report a median rate of nausea and/or vomiting (inter quartile range) of 15.6% (5.2 \u2013 25.0) for Atovaquone-Proguanil whereas other studies did not report this ADR ,20,30,31Nevertheless, data from this series on digestive ADR are comparable to the results of a recent cohort study showing a higher switch rate to second-line treatment in patients treated with AP, compared to others .The design of this study does not enable to guide physicians on their choice of anti-malarial drug. However, the significant risk of vomiting associated with AP needs to be taken into account, especially in patients already complaining of nausea and vomiting at the time of diagnosis. The need for fat intake with AP, as is recommended to improve absorption, may be an additional risk factor for vomiting . PrescriAs observed in the literature, this study did not reveal any liver toxicity. Moderate variations in transaminase level observed at day 3 and 7 were not significant and were possibly due to malaria parasite itself ,14,30,41A drop in haemoglobin level, as observed here, was commonly reported after initiation of treatment of acute malaria due to malaria haemolysis . The absIn 2007, French experts\u2019 consensus recommended both AP and artemether-lumefantrine as first-line treatment for acute uncomplicated malaria . As a coWith the introduction of ACT in Europe  for uncomplicated acute malaria onset, the use of AP will probably decrease in favour of ACT, due to its prompt efficacy and good tolerance.This observational series of 553 cases, the largest to date in number of patients, describes a large experience in using AP for imported uncomplicated malaria in real life conditions. Despite a non-comparative design, it appears that its efficacy is good and comparable to other similar drugs. The study confirms that AP is a valuable treatment option, while ADR, such as vomiting and its limited absorption in some cases, may be a limitation for its use.The authors have declared that they have no competing interests.HC, PR and OB participated in the design of the study and performed the statistical analysis. PR and OB conceived of the study, and participated in its coordination. HC, JC, SM and OB contributed to the draft manuscript. OF, PB, PC, HG, PHC, NG and MB participated in the recruitment of included patients. All authors read and approved the final manuscript."}
+{"text": "The predominance of chloroquine-susceptible falciparum malaria in Malawi more than a decade after chloroquine's withdrawal permits contemplation of re-introducing chloroquine for targeted uses. We aimed to compare the ability of different partner drugs to preserve chloroquine efficacy and prevent the re-emergence of resistance.pfcrt T76 and of anemia. Of the 640 children enrolled, 628 were included in the intention-to-treat analysis. Malaria incidence  was 0.59 (.46\u2013.74), .61 (.49\u2013.76), .63 (.50\u2013.79) and .68 (.54\u2013.86) episodes/person-year for group randomized to receive chloroquine alone or in combination with artesunate, azithromycin or atovaquone-proguanil respectively and the differences were not statistically significant. Treatment efficacy for first episodes was 100% for chloroquine monotherapy and 97.9% for subsequent episodes of malaria. Similar results were seen in each of the chloroquine combination groups. The incidence of pfcrt T76 in pure form was 0%; mixed infections with both K76 and T76 were found in two out of 911 infections. Young children treated with chloroquine-azithromycin had higher hemoglobin concentrations at the study's end than did those in the chloroquine monotherapy group.Children with uncomplicated malaria were enrolled at a government health center in Blantyre, Malawi. Participants were randomized to receive chloroquine alone or combined with artesunate, azithromycin or atovaquone-proguanil for all episodes of uncomplicated malaria for one year. The primary outcome was incidence of clinical malaria. Secondary endpoints included treatment efficacy, and incidence of the chloroquine resistance marker Sustained chloroquine efficacy with repeated treatment supports the eventual re-introduction of chloroquine combinations for targeted uses such as intermittent preventive treatment.NCT00379821ClinicalTrials.gov  In the long struggle to control and ultimately eliminate malaria, the development of resistance to antimalarial drugs has caused repeated setbacks. Chloroquine and sulfadoxine-pyrimethamine (SP) have been retired from use as first-line treatment for malaria in most areas due to widespread resistance. There is evidence that reduced susceptibility to the leading class of antimalarial drugs that replaced chloroquine and SP, the artemisinins, has emerged in Southeast Asia If chloroquine is reintroduced, whether for routine treatment or for targeted prevention, it will be prudent to administer it with a partner drug to protect against the re-emergence of resistance. Combination therapy is recommended to effectively treat individual infections and to prevent the emergence and spread of resistance To gain insight into the pharmacokinetic and pharmacodynamic properties that might protect against the re-emergence of chloroquine resistance, we selected three partner drugs to use in combination with chloroquine. Artesunate is a highly effective, very short-acting medication that rapidly decreases parasite biomass, but has a duration of action of only a few hours P. falciparum parasites are predominantly susceptible to chloroquine, a drug which, unlike the artemisinins, has established molecular markers of drug resistance Malawi provided a unique opportunity to evaluate the effect of drug combinations on the evolution of drug resistance. In this small southern African country, This study was designed as a longitudinal trial, rather than as the evaluation of efficacy of a treatment regimen against a single episode of malaria. For pivotal studies that examine the application of new drugs and treatment strategies, we have advocated for the use of longitudinal studies Ndirande is a peri-urban hillside township of Blantyre, the second largest city in Malawi. Malaria transmission is year-round, with a seasonal peak from December to March corresponding to the rainy season. The Ndirande Health Center is the sole government health facility serving the township's estimated population of 200,000. Children attending the pediatric clinic who were suspected of having malaria were offered screening and enrollment in this study.P. falciparum mono-infection with parasite density 2,000\u2013200,000/mm3; planning to remain in the study area for one year; willingness to return for four-weekly routine visits and unscheduled sick visits; and parental/guardian consent. Exclusion criteria included danger signs of severe malaria ; known allergy to a study drug; on chronic medication with a drug that has antimalarial activity; abnormal liver enzymes or renal function; and evidence of severe malnutrition or chronic disease. Children with known HIV infection were not enrolled because trimethoprim-sulfamethoxazole, an antimicrobial with antimalarial activity, is recommended for all HIV-infected children in Malawi.The protocol for this trial and supporting CONSORT checklist are available as supporting information: see All parents or guardians provided written informed consent prior to the initiation of any study-related procedures. The study protocol was reviewed and approved by the University of Malawi College of Medicine Research and Ethics Committee and the Institutional Review Board of the University of Maryland, Baltimore. The study was registered on ClinicalTrials.gov (NCT00379821).This study was a randomized longitudinal open-label drug efficacy trial among children who presented to the Ndirande Health Center with uncomplicated falciparum malaria. Participants were randomized with equal allocation to the four drug treatment arms: chloroquine monotherapy , chloroquine plus artesunate , chloroquine plus azithromycin , and chloroquine plus atovaquone-proguanil . Doses were rounded to the nearest one quarter tablet. Treatment was not blinded because of the use of tablets and liquids in different arms as well as the frequent need to crush tablets (to administer to young children) precluded over-encapsulation. At each malaria episode during the 12-month study period, participants received the same treatment as the one assigned at enrollment. Study medications were administered under direct observation. For all episodes of malaria, drug efficacy was assessed for 28 days. After the participant was enrolled in the study, an episode of malaria that occurred more than 14 days after a diagnosis of malaria was considered a subsequent episode and treated with the assigned treatment regimen. Participants returned to the clinic every four weeks for active follow-up and were encouraged to come to the clinic whenever they were ill. At every visit, the parents were questioned about medical care received outside the study and the health records were reviewed. If a participant was treated for malaria outside the study, the episode was recorded as a malaria illness and included in the intention-to-treat analysis. Participants who missed two consecutive 4-weekly routine visits were discontinued from the study.Full blood count with automated thre-part differential, ALT and creatinine were obtained at day 0 and day 14 of every clinical malaria episode and also at study weeks 16, 32 and 52. Normal values for hematological and biochemical evaluations were based on data collected from healthy children in rural Mozambique At enrollment and during subsequent episodes of uncomplicated malaria , diagnosis and treatment initiation occurred on day 0 and all doses of medication were administered under direct observation. Participants were followed actively on days 1, 2, 3, 7, 14, 21 and 28 at the study clinic, and by daily availability of a study clinician to evaluate and treat as needed. Malaria smears, dried blood spots on filter paper (Whatman 3MM) and hemoglobin measurement  were obtained at every follow up visit except day 1. The 2004 World Health Organization definitions of treatment outcomes were used P. falciparum chloroquine resistance transporter gene (pfcrt); pfcrt T76 is the primary marker for chloroquine resistance msp2), as described previously http://medschool.umaryland.edu/malaria/protocols.asp).All specimens from day 0 of each episode of malaria illness underwent pyrosequencing to determine the allele at position 76 of the The primary objective of the study was to compare the annual incidence of malaria clinical episodes between the CQ monotherapy arm and each of the three combined treatment arms. We hypothesized that the annual incidence rates of malaria episodes will differ between the group treated with chloroquine monotherapy and the group treated with chloroquine combined with a partner study drug.The secondary objectives were to assess the anti-malarial drug efficacy at first and subsequent administrations by treatment arm, to measure the effect of each treatment arm on anemia and the prevalence of chloroquine resistance and to assess the safety of repeated use of the drugs in each study arm.We estimated a mean rate of 1.5 malaria episodes per year with chloroquine alone. We derived the sample size required to detect a decrease in the mean to 1.0 episodes per year assuming a Poisson distribution and using a 2 sided test with alpha\u200a=\u200a0.05/3\u200a=\u200a.0166. The test size was adjusted to accommodate the 3 treatment versus control comparisons. With the above requirements and for a 2 treatment contrast, 120 subjects with complete observation were required per treatment arm to have 90% power. To account for migration and other causes of attrition, we enrolled an additional 33% (40 subjects) in each regimen for a total of 160 per arm, and an overall sample size of 640 subjects.The randomization sequence was generated by the trial statistician in SAS using blocked randomization; assignments were concealed using a pull-tab treatment list until participants were enrolled. This was an open-label trial. The laboratory technicians who read the malaria smears were blinded to study drug allocation.P. falciparum infections and episodes diagnosed outside of the study clinic, were included. Per-protocol analysis excluded subjects who did not receive the full course of treatment and included only those malaria episodes subsequent to initial treatment caused by P. falciparum mono-infection as diagnosed at the study clinic. Follow-up time was from enrollment to date of last contact, or until receipt of off-protocol anti-malarial medication. Time at risk excluded time under treatment for malaria.The primary endpoint for the study was the incidence of clinical malaria episodes per year of follow-up. A Poisson regression model offset by follow-up time was used to estimate the annual incidence of malaria in each treatment arm and to compare each combination arm to the chloroquine monotherapy arm. Results were reported using both the intention-to-treat and per-protocol populations. Intention-to-treat analysis used all evaluable subjects. All malaria episodes subsequent to initial treatment, including non-Kaplan-Meier estimates were used to calculate the probability of malaria-free survival by treatment arm. Cox proportional hazards were used to estimate the risk of malaria after initial treatment in each treatment arm and the incidence of severe malaria.Rates of adverse events and serious adverse events were compared by the Kruskal-Wallis test. For hemoglobin concentration and other laboratory tests conducted for safety, a generalized estimating equations (GEE) model with Gaussian link and exchangeable correlation structure was fit to estimate the effect of repeated use of study drugs on the mean clinical laboratory value, adjusted for treatment arm and age. Data were analyzed in SAS version 9.2  and R version 2.10.0.Nine hundred thirty-two potential participants underwent screening and 640 were enrolled between February 2007 and August 2008; follow-up was completed in 2009  cohort for initial malaria illness episode. The PP population for subsequent episodes is shown in Six hundred twenty eight subjects who returned for at least one visit after day 0 and were included in the intention to treat (ITT) analysis. Six hundred eleven evaluable subjects with a malaria episode caused by There were no statistically significant differences in the incidence of malaria between the chloroquine monotherapy arm and any of the combination therapy arms in the ITT or PP populations . TreatmeThe time from initial to first subsequent malaria episode was the same for all arms . FollowiAmong all age groups, there was no difference in the mean concentration in the mean hemoglobin at the end of the study by treatment arm (p\u200a=\u200a0.60 for GEE). Hemoglobin concentration at the end of the study period was higher among children \u22643 years of age in the chloroquine-azithromycin arm  compared to monotherapy arm  . There wpfcrt. Among the evaluated samples, the chloroquine-susceptible form (pfcrt K76) was present as the only detectable genotype in 909 (99.8%); the remaining two were mixed infections . No infections contained the chloroquine-resistant genotype exclusively. One mixed infection occurred 28 days after an initial treatment with chloroquine-artesunate and was classified as a new infection based on genotyping. The participant was subsequently treated with chloroquine-artesunate and had an adequate clinical and parasitological response at 28 days and no further episodes of malaria during the study. The second mixed infection occurred in a participant in the chloroquine-azithromycin arm. The treatment was the third the participant had received: the first occurred at enrollment, the second occurred three months after enrollment, and the episode with the mixed pfcrt genotype occurred 3.5 months later. The participant had an adequate clinical and parasitological response and had no further malaria episodes during the study.Nine hundred twenty-four episodes of malaria, including seven episodes of late treatment failure, occurred. DNA extracted from 911 filter paper specimens was amplified successfully to evaluate the allele at codon 76 of There were 49 serious adverse events (SAEs) reported in 46 subjects. This included eight evaluable cases of severe malaria in the study, five in the chloroquine monotherapy arm, one in the chloroquine-azithromycin group and two in the chloroquine-atovaquone-proguanil group. The difference between the treatment groups was not significant given the low number of events; however, there were no cases of severe malaria in the chloroquine-artesunate arm. Five SAEs were considered to be possibly associated with study product vomiting . Three dMinor differences between treatment groups were seen in drug-related adverse events. The rate of treatment-related pruritis was lowest in the chloroquine-artesunate group, with 2.5% of participants experiencing the event compared to a range of 12.5\u201316.3% (p\u200a=\u200a0.0005). Neutropenia, possibly related to the study drug based on temporal proximity, occurred in seven participants in the azithromycin arm; all resolved. Although neutropenia occurred infrequently in other treatment groups, none of those episodes was suspected of being associated with the study treatment. No difference in the results of the neurological assessments was detected between the groups. In the GEE models of the safety laboratory results, the effect of episode and the interaction of episode with treatment were not significant in any of the models, suggesting that repeated use of any study treatment did not affect the results.In the controlled setting of this clinical trial, the annual incidence of malaria was the same in all treatment arms, including chloroquine alone. Single episode treatment efficacy of chloroquine, both at the initial infection and among the participants who had subsequent episodes of malaria was high and sustained when it was administered repeatedly either alone or as part of combination therapy over twelve months in Blantyre, Malawi. These results further confirm the molecular and clinical evidence demonstrating the complete predominance of chloroquine-susceptible malaria in Malawi for the last ten years, and suggest that chloroquine could have a future role for malaria treatment and prevention in Africa. In this longitudinal study, where parasites were susceptible and in which effective drugs were administered appropriately, chloroquine-resistant parasites did not re-emerge despite repeated treatment in approximately half of the study participants over the course of a year. Because clinical treatment failure may be a relatively insensitive tool for the detection of drug resistance, especially in a population with some pre-existing immunity, we also tested for the molecular marker of chloroquine resistance. Our molecular assay was sensitive enough to detect a minority genotype representing at least 20% of a mixed infection. However, we found only two mixed infections containing chloroquine-resistant parasites among more than 900 episodes of malaria, and no examples of predominantly or exclusively resistant infections despite repeated use of a drug with a half-life of one to two months. This is evidence that resistant parasites are almost entirely absent in this human population.a priori.Because the annual incidence of malaria was lower than we had expected, it is possible that the study did not have sufficient power to detect differences in either the annual incidence of infection or treatment efficacy during subsequent episodes. However, using the observed rate of malaria in both the ITT and PP populations, we should have been able to detect meaningful difference between the chloroquine monotherapy group and any of the combination groups, if they existed. Based on an annual incidence of malaria of 0.6 in the chloroquine monotherapy group, we had 88% power to detect a difference of 0.2 episodes per person year between the two groups, with a type 1 error rate of 0.0166. For the PP analysis, we had 90% power to detect a change from 0.5 to 0.3 episodes per year. The number of treatments required to permit the re-emergence has not been estimated and as a result the study may have not had power to produce sufficient selective pressure. However, we did not have data to make these assumptions Because clinical efficacy and susceptibility were the same in all treatment arms, we had the opportunity to examine additional secondary benefits and the safety of the various combinations, independent of their antimalarial effect. In the chloroquine-artesunate group, there were no episodes of severe malaria. Severe malaria was uncommon in this clinical trial, probably because illnesses were diagnosed and treated promptly, and given their rarity, the study did not have adequate power to detect differences between groups. Nevertheless, these findings suggest that the addition of artesunate, the most rapid-acting of all the drugs included in this study, may further protect against severe disease, even when case detection and treatment are optimal.Anemia is another important morbidity associated with repeated malaria infections. We found a significantly higher hemoglobin concentration in children aged \u22643years in the chloroquine-azithromycin group. This observation is not attributable to more effective malaria treatment or longer duration of prophylaxis, as treatment outcomes and incidence of malaria infections were the same in all groups. The medication was given at a high dose and has a long half-life, so recipients were potentially protected from bacterial infections for weeks at a time. The higher hemoglobin concentration may thus be due to the prevention or treatment of bacterial infection; alternatively, decreasing the bacterial burden in the gastro-intestinal tract that may lead to improved absorption of nutrients or decreased levels of chronic inflammation. Further studies will investigate the detailed etiology of this phenomenon. Azithromycin was also associated with transient neutropenia, but with no adverse clinical outcomes or long term sequelae.There were few safety concerns. Participants in all groups experienced pruritis, as commonly occurs in dark-skinned individuals treated with chloroquine. The significantly lower rate of pruritis in the chloroquine-artesunate group was unexpected. It is possible that artesunate accelerates chloroquine metabolism, thus decreasing the adverse effects Several factors warrant caution before serious consideration is given to returning to chloroquine use as a wide-scale treatment policy. Although this study was large for an antimalarial drug therapeutic efficacy study, the proportion of malaria parasites under chloroquine selective pressure, relative to the entire parasite population in this community, was very small. How much drug pressure, either in total dose or duration of exposure, is required to lead to a detectable change in the distribution of drug-resistant malaria is not known. As is frequently the case in urban settings in Africa, the population was highly mobile; participants were censored from the study if they missed more than eight weeks of follow up. Losses-to-follow-up were similar across study groups, and there was no evidence to suggest that censoring was related to the outcomes of interest.All treatments were administered under directly observed therapy, conditions that do not reflect real world use. As a result, conclusions about the efficacy of different treatment regimens and the risk of rapid selection of resistance may be different when compliance with the full treatment regimen requires the administration of two medications at home for two additional days. Moreover, while this single-site study convincingly demonstrates the absence of chloroquine-resistant falciparum malaria from this location and the immediate environs where study participants acquired malaria during the study, much larger, regional multi-site studies would be required to inform policies on using chloroquine combinations for malaria treatment or prevention.Chloroquine-susceptible malaria has definitively returned to Malawi, and other evidence suggests the same trend is occurring throughout the region Any one of the combinations tested in this trial or drugs with similar pharmacodynamic characteristics might be suitable for different indications and transmission settings. The optimal selection of the partner drug(s) should be made based on knowledge of epidemiology including risk of re-infection during the post-treatment period, population pharmacokinetic characteristics, drug-resistance patterns, and other key factors related to implementation including cost, availability and ease of adherence.Checklist S1CONSORT Checklist.(DOC)Click here for additional data file.Protocol S1Trial Protocol.(DOC)Click here for additional data file."}
+{"text": "Malaria is a leading cause of mortality, particularly in sub-Saharan African children. Prompt and efficacious treatment is important as patients may progress within a few hours to severe and possibly fatal disease. Chlorproguanil-dapsone-artesunate (CDA) was a promising artemisinin-based combination therapy (ACT), but its development was prematurely stopped because of safety concerns secondary to its associated risk of haemolytic anaemia in glucose-6-phosphate dehydrogenase (G6PD)-deficient individuals. The objective of the study was to assess whether CDA treatment and G6PD deficiency are risk factors for a post-treatment haemoglobin drop in African children <5\u2009years of age with uncomplicated malaria.This case\u2013control study was performed in the context of a larger multicentre randomized clinical trial comparing safety and efficacy of four different ACT in children with uncomplicated malaria. Children, who after treatment experienced a haemoglobin drop \u22652\u2009g/dl (cases) within the first four days , were compared with those without an Hb drop (controls). Cases and controls were matched for study site, sex, age and baseline haemoglobin measurements. Data were analysed using a conditional logistic regression model.G6PD deficiency prevalence, homo- or hemizygous, was 8.5% (10/117) in cases and 6.8% (16/234) in controls (p\u2009=\u20090.56). The risk of a Hb drop \u22652\u2009g/dl was not associated with either G6PD deficiency (adjusted odds ratio (AOR): 0.81; p\u2009=\u20090.76) or CDA treatment  alone. However, patients having both risk factors tended to have higher odds  of experiencing a Hb drop \u22652\u2009g/dl within the first four days after treatment, however this finding was not statistically significant, mainly because G6PD deficient patients treated with CDA were very few. In non-G6PD deficient individuals, the proportion of cases was similar between treatment groups while in G6PD-deficient individuals, haemolytic anaemia occurred more frequently in children treated with CDA (56%) than in those treated with other ACT (29%), though the difference was not significant (p\u2009=\u20090.49).The use of CDA for treating uncomplicated malaria may increase the risk of haemolytic anaemia in G6PD-deficient children. Plasmodium falciparum uncomplicated malaria [falciparum malaria in non-pregnant adults. Data from the phase III clinical programme conducted in children in sub-Saharan Africa showed that CD achieved significantly higher cure rates compared to sulphadoxine-pyrimethamine (SP) and was well tolerated [Chlorproguanil\u2013dapsone (CD) is a fixed-dose anti-folate combination that was developed by a public-private partnership for the treatment of  malaria . CD receolerated . Importaolerated ,4. Howevolerated ,5, cliniolerated .Dapsone is metabolized in a hydroxylamine metabolite that is susceptible to trigger oxidation damages in red blood cells, leading, if not thwarted by a protection mechanism, to haemolytic anaemia . This isG6PD, a cytoplasmic enzyme expressed in all cells, catalyses the first step of the pentose phosphate pathway and plays an essential role in protecting red blood cells from oxidative stress . The G6Pin vitro studies suggested that G6PD deficiency could confer a protection against P. falciparum infection by inhibiting erythrocyte invasion or intracellular development of the malaria parasite [In a G6PD-deficient individual, oxidative stress can seriously damage red blood cells , resulting in acute haemolysis. When acute haemolysis is triggered by a treatment such as CD, Hb drop can be of about 2\u2009g/dl within one or two days after treatment administration . Unlike parasite . Given tparasite . Howeverparasite . NeverthThis study was part of a large multicentre trial  testing the safety and efficacy of four ACT for uncomplicated malaria in children, namely dihydroartemisinin-piperaquine (DHAPQ), amodiaquine-artesunate (AQ + AS), artemether-lumefantrine (AL), and CDA . DetailP. falciparum mono-infection with density between 2,000 and 200,000/\u03bcl; fever (axillary temperature \u226537.5\u00b0C) or history of fever in the last 24 hours, and Hb \u22657\u2009g/dl. Exclusion criteria were: participation within the last month in any other investigational drug study; known hypersensitivity to at least one of the study drugs; severe malaria [Briefly, CDA was compared with DHAPQ and AL in Rwanda  and in Uganda  and with DHAPQ and AQ + AS in Uganda (Mbarara) and Mozambique (Manhi\u00e7a). Rwandan samples were not included in the series as they were not available at the time of the analysis. Children aged 12\u201359\u2009months old with uncomplicated malaria were randomly allocated to one of the three study arms if they fulfilled the following inclusion criteria: body weight >5\u2009kg;  malaria  or otherTreatment was administered under direct supervision during three consecutive days and according to the patient\u2019s body weight. CD was orally administered at a dose of 2.0\u2009mg/kg of chlorproguanil and 2.5\u2009mg/kg of dapsone once a day during three days, using commercial Lapdap\u00ae paediatric tablets containing each 15\u2009mg of chlorproguanil and 18.75\u2009mg of dapsone. Arsumax\u00ae tablets  containing 50\u2009mg of artesunate were given at the dosage of 4\u2009mg/kg/day during three days. The other formulations of ACT were administered according to the manufacturers\u2019 instructions .All children were kept at the health facility during the three-day dosing period. The mother/guardian was asked to return with the child for scheduled visits on days 3, 7, 14, 21 and 28 post-treatment, or if any symptoms occurred between scheduled visits. Capillary or venous blood was taken at each visit. The follow-up description can be found in the main study methods .Thick and thin blood films were prepared, dried and Giemsa stained; parasite density was estimated by counting the number of asexual parasites in 200 white blood cells (WBC), assuming a standard WBC count of 8,000/\u03bcl. Hb was measured using HemoCue\u00ae device at day 0, before treatment, and using an haematology analyser  at days 1, 2, 3, 7, and 28. Three blood spots were collected on filter paper (Whatman Grade 3) at enrolment and at each visit from day 7 onwards. Each filter paper was dried and individually stored in a plastic bag containing silica gel. All filter papers were subsequently transferred to the Institute of Tropical Medicine  for genotyping.2 1.5\u2009mM, dNTP 0.6\u2009mM, each primer 0.2\u2009\u03bcM, Taq polymerase 0.02U/\u03bcl, DNA 1\u2009\u03bcl, buffer 1x and Milli-Q water. Cycler programme steps were: primary denaturation 5\u2019 at 95\u00b0C, denaturation 1\u2019 at 95\u00b0C, annealing 45\u201d at 64\u00b0C, extension 1\u2019 at 72\u00b0C, 34 cycles, and final extension 5\u2019 at 72\u00b0C. The 202 mutation was detected with the NlaIII restriction enzyme that cleaves DNA only if mutation is present. RFLP conditions were: NlaIII 0.08U/\u03bcl, BSA 1x, buffer, Milli-Q water and 5\u2009\u03bcl of the PCR product. After eight hours of incubation at 37\u00b0C, enzyme was inactivated 20\u2019 at 65\u00b0C. Digested DNA samples were run in a 2% agarose gel by electrophoresis in Tris-Acetate-EDTA (TAE), marked with ethidium bromide and revealed with UV. Results were interpreted as followed: one band (338\u2009bp) corresponds to B or A genotype, two bands  correspond to A- genotype, and three bands  correspond to heterozygotes (BA- or AA- genotype).DNA was extracted from dried blood spot on filter paper with the Chelex method . Considet test. The geometric mean of the parasite density was used as threshold between \u201chigh\u201d and \u201clow\u201d parasitaemia. Mild anaemia was defined as Hb <11\u2009g/dl. Data were analysed using NCSS software . A conditional logistic regression model was built using STATA version 9 . Multi-colinearity was checked and no variance inflation factor was greater than 10. Stepwise multivariate analyses were performed, and all possible interactions up to order 2 were tested, checking at each step how effects and likelihood were modified . Adjusted odds ratios (AORs) and standard errors (SE) were calculated taking into account the interaction term. A p-value <0.05 was considered as significant; all p-values are 2-sided and reported confidence intervals (CI) are 95%CI.A matched case control approach was used. Children with a Hb drop \u22652\u2009g/dl within three days post-treatment were considered as cases and all others as controls. As the number of participants with a Hb drop was low, two matched controls were selected for each case in order to increase the power of the analysis. Controls were matched for study site, sex, age (<10\u2009months difference) and Hb at baseline (<1\u2009g/dl difference). Proportions were compared using chi-square or Fisher exact tests and continuous variables were compared using Student\u2019s\u2009Hb data were not available for at least one of the first four days for 127 patients and 107 Rwandan samples were not available. Among the remaining 571 patients, Hb dropped by \u22652\u2009g/dl in 117 (20.5%). Mean Hb decrease, compared to day 0, was 0.57\u2009g/dl (CI: 0.47-0.60) at day 1, 0.76\u2009g/dl (CI: 0.67-0.85) at day 2 and 0.51\u2009g/dl (CI: 0.43-0.59) at day 3. In most cases , the \u22652\u2009g/dl drop occurred at day 2. Haematological recovery was observed commencing at day 3 onwards, with a mean Hb of 11\u2009g/dl (CI:10.9-11.1) at day 28. In one child Hb dropped from 6.5\u2009g/dl at day 0 to 3.3\u2009g/dl at day 7, requiring a blood transfusion. The child was G6PD heterozygote and treated with CDA.vs 33.8, 32.7 and 31.1 in Manhi\u00e7a, Mbarara and Jinja, respectively, p\u2009=\u20090.0001) The Hb \u22652\u2009g/dl drop was more frequent in children under 28\u2009months (p\u2009=\u20090.026) with a high parasite density (p\u2009=\u20090.034), and without mild anaemia (p\u2009<\u20090.001). Percentage of patients whose Hb dropped by \u22652\u2009g/dl did not differ between treatment groups  have been reported. It is estimated that the A376G-G680T and A376G-T968G genotypes represent less than 5% of type A- deficiencies, though in Senegal they are more frequent than the usual A376G-G202A . There a02A have Pfdhfr I164L mutation, mainly found in Asia while this particular mutation seems rare in sub-Saharan Africa [The available information indicates that CDA in G6PD-deficient patients is associated with a substantial risk of haematological toxicity, due to an oxidant metabolite of dapsone causing haemolytic damages in these patients. Assuming the CDA development would have continued, it would have implied that, before any treatment, patients would have to be tested for G6PD deficiency, e.g. with the cheap and fast Beutler\u2019s fluorescent test  judged nn Africa ,31. For UDA has received travel grants from sanofi Aventis, sigma tau and Novartis. He has also received research funds from sanofi Aventis and sigma tau.UDA and JPVG designed the sub-study. QB/SM/RG, AT/AY and CN/PP/AD/ET were responsible for the study implementation and data collection in Manhi\u00e7a, Jinja/Tororo and Mbarara, respectively. PF, CVO and CVM conceived and performed the G6PD experiments. HvL managed the data. JPVG and CVM analysed the data. ARR supervised data analyses and results reporting. CVM, JPVG and UDA wrote the paper and all authors reviewed the manuscript."}
+{"text": "In vivo efficacy assessments of antimalarials are essential for ensuring effective case management. In Ethiopia, chloroquine (CQ) without primaquine is the first-line treatment for Plasmodium vivax in malarious areas, but artemether-lumefantrine (AL) is also commonly used.P. vivax in Oromia Regional State, Ethiopia. Individuals with P. vivax monoinfection were enrolled. Primary endpoint was day 28 cure rate. In patients with recurrent parasitemia, drug level and genotyping using microsatellite markers were assessed. Using survival analysis, uncorrected patient cure rates at day 28 were 75.7%  66.8\u201382.5) for AL and 90.8% (95% CI 83.6\u201394.9) for CQ. During the 42 days of follow-up, 41.6% (47/113) of patients in the AL arm and 31.8% (34/107) in the CQ arm presented with recurrent P. vivax infection, with the median number of days to recurrence of 28 compared to 35 days in the AL and CQ arm, respectively. Using microsatellite markers to reclassify recurrent parasitemias with a different genotype as non-treatment failures, day 28 cure rates were genotype adjusted to 91.1% (95% CI 84.1\u201395.1) for AL and to 97.2% (91.6\u201399.1) for CQ. Three patients (2.8%) with recurrent parasitemia by day 28 in the CQ arm were noted to have drug levels above 100 ng/ml.In 2009, we conducted a 42-day efficacy study of AL or CQ for P. vivax malaria, but high rates of recurrent parasitemia were noted with both drugs. CQ provided longer post-treatment prophylaxis than AL, resulting in delayed recurrence of parasitemia. Although the current policy of species-specific treatment can be maintained for Ethiopia, the co-administration of primaquine for treatment of P. vivax malaria needs to be urgently considered to prevent relapse infections.In the short term, both AL and CQ were effective and well-tolerated for NCT01052584ClinicalTrials.gov  Plasmodium vivax transmission is more widely distributed globally than P. falciparum, with 2.85 billion people living at risk of its infection P. falciparum and P. vivax substantially contribute to malaria morbidity in Ethiopia, in relative proportions of approximately 60% and 40% P. falciparum resistance to chloroquine (CQ) P. falciparum malaria and of mixed infections with both P. falciparum and P. vivaxP. vivax malaria in Ethiopia, AL is used widely for all cases of mixed infections and clinical malaria where laboratory diagnostics to determine the specific malaria species are not available or not done.With wide-spread P. vivax in 1989 from Papua New Guinea P. vivax has remained rare in Africa. In 1996, Ethiopia published its first report of CQ resistance, with 2% (5/255) of study patients on CQ with persistent parasitemia on day 7 P. vivax, but at levels not exceeding 5% Following the first report of CQ-resistant P. falciparum malaria, mixed infections, and clinically diagnosed malaria, the Ethiopian FMOH scaled up an ambitious plan to provide universal access to prompt malaria diagnosis and treatment \u2013as recommended by the World Health Organization (WHO) P. falciparum from P. vivax infection and expansion of quality malaria microscopy diagnostic capacity, administration of CQ increased in Ethiopia since 2004 in parallel with increased numbers of laboratory confirmed P. vivax mono-infections. Since both AL and CQ without primaquine are widely used according to FMOH guidelines for P. vivax infections in malarious areas, continuously monitoring in vivo drug efficacy to these medications for P. vivax malaria is recommended by WHO After adopting AL as first-line therapy for uncomplicated in vivo efficacy of AL and CQ for treatment of P. vivax for 28 days, and (ii) to assess secondary outcomes of day 42 efficacy, hematologic response, and fever and parasite clearance rates.The objectives of the study reported here were (i) to assess the The protocol of this trial and supporting CONSORT checklist are available as supporting information see .P. falciparum and P. vivax are endemic in the catchment areas of both sites, and the study was conducted during the peak malaria transmission season which occurs between September to December. Detailed study site information has been previously reported along with the results of the P. falciparum treatment outcomes This randomized, open-label study was conducted at two established FMOH health facilities, the Bishoftu Malaria Clinic and the Bulbula Health Center, in Oromia Regional State, Ethiopia, from October 2009 to January 2010. P. vivax mono-infection, with parasitemia density >250 asexual forms per \u00b5l were enrolled. Additional inclusion criteria included age >6 months, weight >5 kg, and axillary temperature \u226537.5\u00b0C or history of fever in the past 48 hours. Only patients living within 20 km of the facilities were enrolled in the study to facilitate follow-up. Exclusion criteria included detection of any Plasmodium sp. infection besides P. vivax mono-infection, any signs or symptoms of severe illness, severe malnutrition, severe anemia (hemoglobin<5 g/dL), known hypersensitivity to AL or CQ, pregnancy, breastfeeding, or presence of another infection or major co-morbid conditions by history.Patients with The study received ethical clearance from the U.S. Centers for Disease Control and Prevention, Columbia University, and the Ethiopian Public Health Association. Written informed consent was obtained from adult patients and guardians of enrolled children. Written assent was also obtained from children 7\u201317 years of age.P. vivax mono-infection based on clinical and parasitological parameters, we conducted a prospective, 42-day, randomized, open-label in vivo efficacy assessment according to the 2003 and 2009 WHO in vivo protocol for measuring antimalarial drug efficacy for areas of low to moderate malaria transmission P. vivax infection was identified by microscopy, the patient was screened for study eligibility criteria. At Bulbula Health Center, all patients attending the outpatient ward were initially clinically evaluated by the health facility staff; those with fever or a history of fever were referred for malaria microscopy testing (as routinely done). Once the patient was noted to have microscopy-confirmed P. vivax infection, the study staff screened the patient for inclusion in the study.To assess AL and CQ therapeutic efficacy for uncomplicated After meeting the inclusion criteria and consenting to study enrollment, all patients underwent hemoglobin testing  and filter paper blood spot collection. All women aged 13\u201349 years underwent a urine pregnancy test and those testing positive were excluded from the study. Giemsa-stained thick and thin blood films were prepared according to standard procedures Patients were enrolled, interviewed and examined by the study clinician. The patients were randomized to standard dosages of oral AL  or CQ  based on weight according to national policy Patients were followed-up for 42 days and asked to return on days 1\u20133, 7, 14, 21, 28, 35, and 42 post-treatment, as well as any other interim day if ill. The study site facilities were open from 0800 to 1800 hrs, and after hours care was also available. Standardized follow-up included documentation of history-taking to elicit symptoms, adverse events, and any concomitant therapy, and physical examination including axillary temperature measurement. All symptoms elicited during follow-up that were not present on day 0 were classified as an adverse event. For all adverse events, severity, duration, outcome and its likely relatedness to the administered drug were assessed by the clinicians. Adverse events were reported for all enrolled patients. Finger pricks for follow-up blood films were taken on scheduled days 2, 3, 7, 14, 21, 28, 35, 42, and at any unscheduled visit. Hemoglobin was measured weekly and filter papers were collected on day 7 for drug level testing and on any day of recurrent parasitemia (after day 3) for CQ level and parasitemia molecular testing. All filter papers were dried and stored in plastic storage bags with desiccant and humidity indicators. The filter papers for drug level and molecular testing were sent to the U.S. Centers for Disease Control and Prevention in Atlanta, USA. All additional sample investigations such as the drug level and molecular testing were performed by individuals who were blinded to the treatment allocation.P. vivax illness requiring hospitalization in presence of parasitemia; or 2) presence of parasitemia and axillary temperature \u226537.5\u00b0C anytime between days 3 and 28/42; or 3) presence of parasitemia on any day between days 7 and 28/42, irrespective of clinical conditions. If no failure was recorded, outcome was classified as a treatment success.Efficacy was assessed by clinical and parasitological outcomes using WHO definitions, with a 42-day follow-up period To refine the identification of recrudescences, three to four drops of blood were collected on filter paper at day 0 prior to treatment, day 7, and on any day of recurrent parasitemia. Blood spots collected on the day of enrollment (day 0) and on the day of failure were used for molecular analysis to determine the genetic identity of the parasites. Blood spots collected on day 7 or day of treatment failure were tested for lumefantrine and CQ drug levels, respectively.Eight of the eleven microsatellite markers described by Imwong et al. eH) and number of alleles per microsatellite locus (A). eH was calculated for each locus as eH\u200a=\u200a[n/(n\u20131)][1\u2013\u2211i2p], where n is the number of isolates sampled and ip is the frequency of the ith allele. The sampling variance for eH was calculated as 2(n\u20131)/n3[2(n\u20132)[\u2211ip3\u2013(\u2211ip2)2]] eHThe genetic variation for each microsatellite locus was measured by calculating the expected heterozygosity , were measured using a high-performance liquid chromatographic method as described by Patchen et al. P. vivax being reported at \u22645%, 10% was chosen as the estimated therapeutic failure rate. Assuming an estimated cure rate of 90% and a 20% loss to follow-up at 42 days, a sample size of 120 subjects per arm was calculated to result in a 95% exact binomial confidence interval from 82.4%\u201395.1%. The study was not powered to estimate differences in efficacy across treatment arms or between sites.Primary efficacy outcome was day 28 cure rates. Secondary outcomes included day 42 cure rates, genotype adjusted minimum cure rates, hematologic outcomes, and fever and parasite clearance. With treatment failure of AL and CQ for 2 test or Fisher\u2019s exact test for categorical variables and the Student\u2019s t-test or Wilcoxon-Mann-Whitney test for continuous variables. A two-sided p-value<0.05 was considered statistically significant.All data were entered into both a Microsoft Access  study database and the Microsoft Excel  standard database developed by WHO P. vivax mono-infections, 242 (217 from Bishoftu and 25 from Bulbula) were enrolled for the in vivo drug efficacy study, with 122 and 120 patients randomized to receive AL or CQ, respectively (P. vivax mono-infection accounted for 83.5% (380/455). At Bulbula Health Center where only suspected malaria cases received microscopy, the slide positivity rate was 30.7%  of which P. vivax mono-infection accounted for 57.3% (292/510). The majority of excluded patients presented late in the afternoon and was therefore not enrolled to ensure appropriate timing of the second AL dose.Of the 4,426 patients tested from the two study sites, 961 were positive for malaria parasites from October 21 to November 30, 2009. The last date of follow-up occurred on January 11, 2010. Of 672 patients with confirmed ectively . At the Although drug assignment was randomized, there was a statistically significant baseline difference in age (p\u200a=\u200a0.03), weight (p\u200a=\u200a0.04), height (p\u200a=\u200a0.05), and ownership of bed nets with fewer patients in the AL drug arm owning one (p\u200a=\u200a0.02) . The othFollow-up was completed for 114 patients to day 28 and 113 to day 42 in the AL arm and 108 patients to day 28 and 107 to day 42 in the CQ arm . Six of Treatment with both AL and CQ resulted in rapid clearance of parasites and reduction of fever. Whereas in the AL arm all patients had cleared their peripheral parasitemia by day 2, 6.0% (7/117) of the patients in the CQ arm had peripheral parasitemia by day 2 and 0.9% (1/115) remained positive on day 3 . The difIn the AL arm, 28 patients (24.3%) presented with recurrent parasitemia by day 28, and 47 (41.6%) by day 42 . In the In the AL arm, the median age for those patients with or without recurrent parasitemia was seven and 17.5 years of age, respectively (p<0.0001). In the CQ arm, the median age for those patients with or without recurrent parasitemia was nine and 20 years of age, respectively (p<0.0001). In both treatment arms, younger age was associated with treatment failure. In the CQ arm, those with recurrent parasitemia had a higher median parasite density (4780 parasites/\u00b5L) compared to those without recurrent parasitemia (2120 parasites/\u00b5L) which was statistically significant (p\u200a=\u200a0.01).In the AL arm, median hemoglobin increased from 13.0 g/dL (standard deviation (SD) 2.3) on day 0 to 13.9 g/dL (SD 1.5) on day 42 . In the P. vivax strain identity using the size polymorphisms in eight microsatellite loci as reported in EH values and several allelic forms existed for all the markers used in this study . Using previously reported cut-off levels identified to predict P. vivax CQ resistance. Of the three recurrent parasitemias with adequate CQ concentrations: one was caused by the identical P. vivax strain (3-year-old boy) and two were caused by different P. vivax strains (7- and 2-year-old boys).Chloroquine and DCQ levels on the day of recurrent parasitemia in the CQ arm ranged from below detection to 485.1 ng/mL. Of those that had recurrent parasitemia by day 28, 30% (3/10) were noted to have CQ+DCQ levels above the minimum effective concentration (i.e. 100 ng/mL). Thus, 2.8% (3/108) of all patients had recurrent parasitemia by day 28 with adequate CQ blood levels, suggesting Both AL and CQ were well tolerated with no serious adverse events reported. Overall, 48.4% (59/122) of the patients in the AL arm and 50.8% (61/120) in the CQ arm reported at least one adverse event. 8.2% (10/122) of the patients in the AL arm and 12.5% (15/120) in the CQ arm presented with oral ulcers, which was the most common complaint, followed by nausea/vomiting and cough .P. vivax treatment response involving 242 patients from two sites in central Ethiopia in late 2009. In both treatment arms, no recurrent parasitemias were seen prior to 21 days and there were very few patients that remained microscopy positive beyond day 2 in contrast to high rates of day 2 and 3 positivity seen in Asia with known high grade CQ resistance P. vivax treatment which noted that patients receiving ACTs clear their parasites more quickly (median parasite clearance time\u200a=\u200a28.8 hrs) compared to CQ-based monotherapy or non-ACT combination therapies (50.4 hrs) We show that both AL and CQ are well-tolerated and remain efficacious for early P. vivax parasitemia findings are higher than some previously reported studies in Ethiopia Although high rates of recurrent parasitemia were noted in both treatment arms, fewer patients receiving CQ presented with recurrent parasitemia by day 28 compared to those receiving AL indicating that CQ may have provided longer post-treatment prophylaxis. Our recurrent P. vivax in vivo studies is not standardized and is not part of the current standard WHO protocol P. vivax parasitemias P. vivax treatment regimens. Recurrent infections with the same genotype (true failure and homologous relapse) were distinguished from those with a different genotype (reinfection or heterologous relapse). Despite the possibility of a recurrent parasitemia arising from a clonal hypnozoite, all identical recurrent parasitemias were classified as treatment failures resulting in a higher, more conservative estimate of treatment failure, but still an improvement over the unadjusted treatment failure rates.Our study also highlights issues related to classifying recurrent parasitemia using molecular methods. Although genotyping to distinguish recurrent parasitemias in P. vivax remains challenging and was of limited benefit in this study.Chloroquine drug level testing on the day of failure as recommended by WHO to identify CQ resistance Approximately half of the patients in both the AL and CQ reported some adverse events, none were severe and most were consistent with ones generally reported There are several limitations of this trial including drug administration procedures, baseline differences, and external validity. Due to logistical reasons, the evening doses of AL were not supervised and the antimalarial treatment administration not blinded. The lower day 28 cure rates seen in the AL arm most likely resulted from its shorter post-prophylactic effect, but poor adherence cannot be discounted as a contributing factor, even though, all patients reported having taken their evening doses. Reassuringly, approximately 70% of the patients with recurrent parasitemia had lumefantrine levels >280 ng/mL as compared to only 51% in a study with supervised artemether-benflumetol administration P. falciparum and P. vivax are sympatric. However, due to a relative short half-life, AL provides less post-exposure prophylaxis against P. vivax recurrent parasitemias. In Indonesia, patients receiving DP compared to AL were half as likely to be anemic and 6.6 times less likely to carry P. vivax gametocytes Some experts have suggested unifying malaria treatments under a single ACT regimen P. vivax infections can lead to severe disease P. vivax illnesses need to be reexamined. In Ethiopia, the addition of primaquine to CQ decreased the cumulative incidence of therapeutic failure at day 28 by a life-table analysis method from 5.76% to 0.75% and the cumulative risk of relapse at day 157 by a life-table method from 61.8% to 26.3% With high recurrent parasitemia rates for both AL and CQ at 42 days following treatment, the addition of primaquine for radical cure should be considered in all areas of Ethiopia as is currently recommended by WHO Table S1Microsatellite diversity for the eight markers for all samples with recurrent parasitemia at both day 0 (pre-treatment) and day of failure.(DOCX)Click here for additional data file.Checklist S1CONSORT checklist.(DOC)Click here for additional data file.Protocol S1Study protocol.(DOC)Click here for additional data file."}
+{"text": "This multicentre study was carried out in Cameroon, Ivory Coast and Senegal to evaluate the non-inferiority of the new paediatric formulation of artesunate/amodiaquine (AS+AQ)(Camoquin-Plus Paediatric\u00ae) in suspension form versus artemether/lumefantrine (AL)(Coartem\u00ae) in the management of African children with uncomplicated falciparum malaria.It was an open randomized trial including children aged between 7 months and 7 years. The endpoints were Adequate Clinical and Parasitological Response (ACPR) at day 28, the clinical and biological tolerability. Statistical analyses were done in Intention To Treat (ITT) and in Per protocol (PP).At the end of the study 481 patients were enrolled in the three countries (249 in the AS+AQ arm and 232 in the AL arm). ACRP in ITT after PCR correction did not show any statistical difference between the two groups with 97.6% for AS+AQ versus 94.8% for AL. In the PP analysis, the corrected ACRP were respectively 98.7% and 96.9% for the two regimens. The clinical tolerance was good without significant difference. Anaemia was significantly higher at D7 in the two groups compared to D0.Plasmodium falciparum malaria in African children.This study demonstrates the non-inferiority of AS+AQ versus AL, its efficacy and tolerance in the management of uncomplicated  Plasmodium falciparum malaria[P. falciparum malaria among children aged between six months and seven years.In spite of all the efforts that have been made over decades, malaria remains a major public health problem in sub-Saharan Africa. Children are still the main victims of the disease with a very high mortality rate. Since 2 malaria. UnfortuThis study was conducted in three countries, Cameroon in central Africa, Ivory Coast and Senegal in West Africa. In Cameroon, the study was performed in the urban health centre of Melen in Yaound\u00e9, the capital city. In Ivory Coast, it was done in El Rapha Health Centre of Abidjan, the capital city, and in Senegal in the health district of Kaolack located 200 km from Dakar, the capital city.P. falciparum parasite density between 1,000 and 100,000/\u03bcl in a low transmission area  and between 2,000 and 200,000/\u03bcl in high transmission areas (Ivory Coast and Cameroon)[Children above 6 months and under 7 years of age, who came to the local health centre, with ameroon), with thameroon) or severThis was a multicentre open, comparative and randomized phase IV trial undertaken in two parallel groups to test non-inferiority between two associations: AS+AQ versus artemether/lumefantrine (AL) in children from six months to seven years. After enrolment, children were weighed and randomized into blocks of 10 to receive one of the two drugs.The formulations of the two associations were the following: AS+AQ in a box with 1 bottle containing AS in powder dosed a 160mg/80ml to suspend and 1 bottle containing AQ in syrup dosed at 50mg/5ml. Children under two years received 10ml of AQ and 20ml of AS per day and children up to two years received 15ml of AQ plus 25 ml of AS per day, in a single administration.AL was presented in fixed tablets containing 20mg of artemether and 120 mg of lumefantrine. Tablets were crushed and mixed with water before administration. No food was given prior the AL administration. It was given twice a day according to the patient\u2019s bodyweight and the manufacturer\u2019s instructions. The two drug regimens covered three days (day 0 to day 2). All the daily doses were administered at the health post, under the direct supervision of the investigators. In case of vomiting within the 30 minutes following the administration, study doses were administered again to the patient. Patients who kept vomiting were withdrawn.Study participants were examined in the study clinic 1, 2, 3, 7, 14, 21, and 28 days after enrolment or at any time if they did not feel well. Quinine treatment was given in case of treatment failure which includes : early treatment failure  and late treatment failure [Finger prick to obtain blood for thick and thin smears were done at Day 1, 2, 3, 7, 14, 21 and Day 28 after inclusion or at any time if the patients did not feel well. Blood smears were stained with Giemsa and 200 leucocytes were counted. Assuming a total leukocytes count of 8,000 per litre, parasite density was determined as the number of asexual parasites \u00d7 8,000/200[Plasmodium falciparum: Merozoite Surface Protein (msp) genes 1 and 2. The primers and schedules described by Faye et al[Four drops of blood were collected on filter paper for each patient at day 0 and at day of recurrent parasitaemia to distinguish recrudescence from new infection. For this, parasite genotyping by nested PCR was conducted to compare two polymorphic genetic markers from aye et al were useAll adverse events were closely monitored during each scheduled visit by interview with patient if possible or guardian and clinical examination by a physician. All signs noted were reported in the case report form. Adverse events were defined as occurring new events or worsening from baseline after administration of treatment. Haematology and biochemical tests were done at the enrolment and at Day 7 to evaluate haemoglobin, creatinine, aspartate amino transferase (ASAT), alanine amino transferase (ALAT) and bilirubine parameters. Biological abnormalities were noted.The number of patients to be included in this study was determined using Epi info software version 6.04d, On the An intention to treat (ITT) and per protocol (PP) analysis were done. ITT included all randomized participants who took at least one full dose and had one post baseline efficacy assessment without major protocol violations as wrong dosage, wrong use of non-assigned drug by mistake, co-infection with other malaria species. Patients with major violations and patients lost during the follow-up or withdrawn  have been considered as failure. The per protocol analysis included patients who received the three doses and who had no major protocol violation up to the day 28. Those lost to follow up, and the withdrawals of consent were excluded from the per protocol analysis.The primary endpoint was the Adequate Clinical and Parasitological Response (ACPR) which is an absence of parasitaemia on Day 28 irrespective of axillary temperature without previously meeting any of the criteria of Early Treatment Failure or Late Clinical Failure. The cumData were analysed by estimations of difference in proportions corresponding to 95% confidence interval. Comparison between groups was made using chi-square test or Fisher exact test for qualitative outcome and student test for quantitative outcome when applicable. Otherwise, non-parametric tests  were used. A p value (two-sided) less than 0.05 was considered statistically significant.The protocol was reviewed and approved by the national ethical committees of each country.In 1,035 patients screened, 481 were included: 249 in AS+AQ arm and 232 in AL arm . Any difference was noted regarding fever and parasitaemia mean Table.Any significant difference was noted between the two groups regarding the hemoglobin, creatinine and liver enzymes means Table.The ACRP at D28 after PCR correction in ITT analysis was 97.6% for AS+AQ versus 94.8% for AL  (p= 0.11). In PP analysis, it was 98.7% in AS+AQ arm versus 96.9% in AL arm . Thus, the non-inferiority of AS+AQ was shown both in intention-to-treat and per-protocol analysis. Regarding treatment failure, after PCR correction, 1.6% (4 patients) in AS+AQ group and 3.9% (9 patients) in AL group were considered as recrudescent  between Plasmodium falciparum malaria among children between six months and 7 years of age. The results obtained show that the AS+AQ association proved to be very effective with a ACPR rate adjusted by 97.2% in ITT and 98.3% in PP. Various studies conducted with artesunate/amodiaquine free combination, showed comparable efficacies with a non-inferiority to AL[This multicentre study allowed an evaluation of the efficacy and tolerance of AS+AQ versus AL in three countries,  in the treatment of uncomplicated ty to AL and othety to AL-18. Thisty to AL. Similarty to AL. Howeverty to AL-23. Thisty to AL.P. falciparum transmission.Fever clearance was fast in both treatment groups, confirming the previous data. A rapidet al showing an increase in anaemia, especially after treatment with the AS+AQ association[The clinical tolerance was good with minor adverse events such as asthenia and vomiting in both treatment groups. Administering a single daily dose of this new formulation seems to be well-tolerated and confirms the previous results. Regardiociation. HoweverP. falciparum malaria in the African child. The single daily dose may help improve compliance when used in daily practice with non-supervised administration.This study shows that the new formulation in a suspension form of the artesunate/amodiaquine association is as effective as artemether/lumefantrine in the treatment of uncomplicated The investigations were financially supported by Pfizer Pharmaceutical Laboratories, West Africa, who also supplied the anti-malarials drugs used for this study.BF wrote the paper; BF, JLN, RCT, KS supervised field work and data collection in Senegal, analysed data of Senegal and analysed from the three countries, reviewed and approved the manuscript; TK, TN collected and analysed data in Cameroon, reviewed and approved the manuscript; EIHM, CPK, CA A collected and analysed data in Ivory Coast, reviewed and approved the manuscript; OG coordinated the study in the three countries, reviewed and approved the manuscript; OF coordinated the study in Senegal, reviewed and approved the manuscript; AS coordinated the study in Cameroon  reviewed and approved the manuscript; MK coordinated  the study in Ivory Coast, reviewed and approved the manuscript. All authors read and approved the final manuscript."}
+{"text": "Malaria continues to be amongst the most frequent infectious diseases imported to Europe. Whilst European treatment guidelines are based on data from studies carried out in endemic areas, there is a paucity of original prospective treatment data. The objective was to summarize data on treatments to harmonize and optimize treatment for uncomplicated malaria in Europe.A prospective observational multicentre study was conducted, assessing tolerance and efficacy of treatment regimens for imported uncomplicated falciparum malaria in adults amongst European centres of tropical and travel medicine.Between December 2003 and 2009, 504 patients were included in 16 centres from five European countries. Eighteen treatment regimens were reported, the top three being atovaquone-proguanil, mefloquine, and artemether-lumefantrine. Treatments significantly differed with respect to the occurrence of treatment changes (p\u2009=\u20090.005) and adverse events (p\u2009=\u20090.001), parasite and fever clearance times (p\u2009<\u20090.001), and hospitalization rates (p\u2009=\u20090.0066) and durations (p\u2009=\u20090.001). Four recrudescences and two progressions to severe disease were observed. Compared to other regimens, quinine alone was associated with more frequent switches to second line treatment, more adverse events and longer inpatient stays. Parasite and fever clearance times were shortest with artemether-mefloquine combination treatment. Vomiting was the most frequent cause of treatment change, occurring in 5.5% of all patients but 9% of the atovaquone-proguanil group.This study highlights the heterogeneity of standards of care within Europe. A consensus discussion at European level is desirable to foster a standardized management of imported falciparum malaria. Plasmodium falciparum, which can lead to complicated disease and death if not treated early. Even in uncomplicated cases, which constitute the majority of cases [With approximately 11,000 reported cases annually, malaria remains the most important tropical disease imported into Europe both in clinical terms and in absolute figures . The majof cases , treatmeof cases .Assuming that efficacy and tolerance are similar in imported malaria, the recommended therapy is based on data predominantly derived from therapeutic studies performed in endemic areas -14. NeveThe combination atovaquone-proguanil, largely used in non-endemic countries, has neither been tested extensively in endemic areas nor in non-endemic areas since conducting large studies appears not to be feasible -18. CompThe overall aim of this multicentre observational study is to summarize data on treatment regimens in Europe, in order to harmonize treatment modalities for uncomplicated falciparum malaria, and to optimize drug treatment strategies amongst European centres of tropical and travel medicine.The centres contributing to this prospective observational multicentre study were self-selected members of the then clinical surveillance networks TropNetEurop and SIMPID. Patients with uncomplicated falciparum malaria were recruited at the participating centres between December 2003 and December 2009. Criteria for study inclusion were parasitologically confirmed uncomplicated falciparum malaria with or without chemoprophylaxis, no previous malaria therapy for the same episode of illness, and informed consent. Exclusion criteria were age less than 18\u2009years, complicated/severe falciparum malaria on inclusion, or refusal of consent. Pregnancy was not an exclusion criterion due to the observational design of the study. Mixed infections were excluded for comparability reasons.The primary objective of the study was to describe the profile of anti-malarial regimens used in Europe with a focus on safety and tolerability. Investigated indicators for safety and tolerability were rate and reasons of treatment change, and type, rate and severity of adverse events. Secondary endpoints were clinical and parasitological cure on days 7 and 28, and duration of hospitalization.Clinical cure was determined as fever clearance time in hours and parasitological cure as parasite clearance time in hours on the grounds of expert microscopy of at least 100 visual fields of a Giemsa-stained thick film.Allocation to the particular therapeutic regimen was done by the individual participating centres based on country- or centre-specific recommendations, practice pattern, and individual patient\u2019s situation in terms of past exposure, prophylaxis intake, co-morbidity, etc. Randomization was not performed. Doses were given on the basis of manufacturers\u2019 recommendations.Data were collected anonymously at each participating centre with an electronic entry and reporting tool linked to the regular network surveillance software. In order to assess a standardized and complete minimal dataset from each centre, the tool was equipped with mandatory entry fields, predefined answer categories and data formats and built-in plausibility checks. Treatment outcome was assessed in a standardized follow-up procedure. If feasible, post-discharge control examinations were performed on day 28. If not feasible, a telephone survey was to be carried out on day 28 to detect possible treatment failures and possible late-onset adverse events of drug therapy. The adverse events reporting section of the study software was structured in order to be able to differentiate between signs attributable to the disease itself and to the drug, and to assess their severity. Completed cases were sent electronically to the TropNetEurop/SIMPID coordination centre using the export function of the study software.Statistical analysis was performed using SAS software . For each study endpoint the heterogeneity of treatment regimens was first assessed with an overall test. In the case of a significant result, each regimen was tested against all others using a Bonferroni-Holm adjustment for multiple comparisons. Categorical data were analysed using chi-square or Fisher\u2019s exact tests. If asymptotic chi-square tests yielded unreliable results and exact tests were unfeasible due to limited computational resources, Monte Carlo estimates for the exact test based on one million replications were performed. Continuous data were analyzed using a Wilcoxon or Kruskal-Wallis test.For stratum-adjusted analysis of categorical and continuous data, The Cochran-Mantel-Haenszel test for general association and Friedman\u2019s test were applied, respectively.An umbrella ethical approval was granted by the site hosting the principal investigator (MPG) at the time point of study initiation. Few centres required, and were granted, additional site-specific approval.Five hundred and four malaria patients were included in 16 different centres from five European countries. The contribution of patients by the various centres was heterogeneous, ranging from one to 141. The three main countries providing patients were, by order of number of patients, France, Italy and Germany, respectively.Plasmodium spp. in endemic areas other than sub-Saharan Africa. Median age was 38\u2009years, with a male:female sex ratio of 1.6. Before travelling, a minority of patients sought medical advice (27%) and/or took an at least partially effective chemoprophylaxis (26%). Median travel duration was one month, with a few patients acquiring malaria with exposure as short as four days. The median delay between onset of symptoms and diagnosis was three days. At time of diagnosis, median temperature was 39.0\u2009\u00b0C and parasitaemia varied between very low density to hyperparasitaemia .Patient characteristics are summarized in Additional file A total of 18 different regimens were observed: eight comprising 98% of the prescribed treatments, the 10 other regimens being based either on artemisinine derivatives alone or associated to various anti-malarials in six cases or on quinine associated mainly to pyrimethamine and sulphadoxine at different dosages. Atovaquone-proguanil, artemether-lumefantrine and quinine were used in four countries. Mefloquine was reported only from German and Italian centres, the association \u03b2-artemether\u2009+ mefloquine only in one centre in Italy, intravenous quinine followed by atovaquone-proguanil mainly in one centre in France (in vomiting patients), the combination quinine-clindamycin mainly in another French centre and the combination quinine-tetracycline mainly in Belgium. In 90% of cases, patients received only one regimen.Overall, most patients were hospitalized , but differences in the rate of hospitalization between both the different treatment groups (p\u2009=\u20090.0056) and the different countries participating in the study (p\u2009<\u20090.0001) were observed. The rate of hospitalization between countries was as follows: 100% in Belgium (one centre), 98.7% in Germany (eight centres), 87.1% in Italy (four centres), 85.7% in the UK (one centre) and 71.4% in France (three centres).Globally, the rate of adverse events (at least one event reported) was 16% (n\u2009=\u200982) with differences between treatment groups (p <0.001)  of the 504 cases; with reasons for change differing between treatment regimens (p\u2009=\u20090.0053). When comparing each regimen to all others, quinine monotherapy was more often associated to change regimen (p 0.0004). Frequency of treatment changes varied from 0 with quinine-atovaquone-proguanil treatment to 24% with quinine monotherapy. Globally, reasons for change were vomiting in 30 cases (57%), adverse events other than vomiting in 10 cases, progression to severe disease in two cases, and various reasons in 11 cases. Vomiting  led to a second-line treatment by intravenous quinine in 24 cases (including nineteen on atovaquone/proguanil and three on mefloquine). Atovaquone-proguanil with a 9% proportion of treatment change due to vomiting differed significantly from the overall average (p\u2009=\u20090.013).Overall median parasite clearance time (PCT) was 72\u2009hours, with significant differences between treatment groups (p <0.001). When comparing one regimen to the others, the shorter median PCT was observed in patients treated by the association artemether-mefloquine  and the longer was with atovaquone-proguanil . In pair wise comparisons adjusted for multiple comparisons, PCT was shorter with artemether-mefloquine compared to atovaquone-proguanil (p\u2009<\u20090.001) and to mefloquine alone (p\u2009=\u20090.026) but not compared to the combination artemether-lumefantrine (p\u2009=\u20090.055).Median fever clearance time (FCT) was 48\u2009hours with significant differences observed between treatment groups (p <0.001). Compared to all other treatments, median FCT was shorter in patients treated with artemether-mefloquine (p\u2009=\u20090.008) or mefloquine alone (p\u2009=\u20090.001) and was longer in patients treated with quinine-tetracycline (p\u2009<\u20090.001). In multivariate analysis, pair-wise comparisons showed a shorter FCT between artemether-lumefantrine and quinine-tetracycline (p\u2009=\u20090.04), mefloquine and atovaquone-proguanil (p\u2009=\u20090.03), mefloquine and quinine-tetracycline (p\u2009<\u20090.001), artemether-mefloquine, and quinine-tetracycline (p\u2009=\u20090.005).Duration of hospitalization differed between regimens (p\u2009=\u20090.0002), even after adjusting for country-specific effects (p <0.0001). Median duration was four days, with 25% hospitalized for more than five days, and 5% for more than seven days (maximum of 18\u2009days). Hospitalization was longer with quinine alone (p\u2009=\u20090.0229) or with quinine-clindamycin (p\u2009=\u20090.0156).The overall cure rate was 74% in the intention-to-treat analysis with a missing data rate due to loss to follow-up of patients of 25%. Excluding those patients lost to follow-up, the cure rate was 99% with four recrudescences occurring in three cases treated with atovaquone-proguanil and in one case treated with artemether-lumefantrine (p\u2009>\u20090.05). Two cases of progression of the disease under treatment were recorded (both with atovaquone-proguanil), with no significant difference between regimens.P. falciparum malaria managed in 16 different centres from five countries. This is to date the largest study on imported malaria in terms of numbers of patients. Patient profiles are similar to those observed in the majority of studies on imported malaria [This prospective observational study included 504 cases of uncomplicated  malaria ,15-26, eEighteen different treatment regimens were used. Atovaquone-proguanil was predominantly used  followed by older drugs such as mefloquine (19%) or quinine alone or in combination with clindamycin or tetracyclines (14%) ,27,28. AA vast majority of patients (81%) were hospitalized, but differences were observed between centres with hospitalization rates from 71% in France to nearly 100% in Belgium or Germany. These differences can be explained by the fact that usages and recommendations differ considerably between countries Table\u2009, and witP. falciparum, which is in the focus of this paper) continues to emerge on a global scale, already also including artemisinines, thus potentially also jeopardizing the favourable primary treatment outcomes as routinely observed in most travellers.The cure rate of 99%  is comparable to that one observed in the literature on imported malaria based on smaller series and equal or higher to studies performed in endemic areas -18,32. HFour recrudescences were reported in three cases of atovaquone-proguanil treatment and in one case of artemether-lumefantrine treatment; both drugs necessitating to be absorbed with a fatty meal due to the lipophily of, respectively, atovaquone and lumefantrine ,34. DetaThe rate of patients lost to follow-up at day 28 was high (25%), especially in certain treatment groups where it reached 45% (quinine-clindamycin) or 55% (artemether-mefloquine). Given that recrudescence occurs usually between day 14 and day 30 after treatment, physicians should organize a \u201crecapture\u201d system for these patients, even limited to a call, in order to identify promptly a possible recrudescence ,19,36.Under treatment, mean PCT and FCT were 72 and 48\u2009hours, respectively, which is consistent with the literature even if direct comparisons are not easy, since methodology to calculate PCT and FCT may differ between studies -20,22,25Adverse events were overall relatively frequent (16%), with quinine exhibiting the least favourable tolerance profile. Although none of the adverse events was classified as severe, they constituted the main cause for switching over to second-line treatment in 10% of patients. Vomiting, mainly due to atovaquone-proguanil but also mefloquine, was the leading cause of change and led frequently to an intravenous treatment (quinine).In the present study, atovaquone-proguanil was by far the most frequent regimen prescribed, providing the opportunity to describe a large cohort of malaria patients treated by this drug in \u201creal-life\u201d conditions. Even if the cure rate was good and statistically comparable to the other regimens, three relapses and two cases of progression to severe malaria were observed, possibly due to poor absorption. In addition, PCT was longer compared to the other regimens. The rate of adverse events was 13%, better than that one of quinine but vomiting appeared to be specifically related to that regimen leading in a high proportion to a second-line treatment. Surprisingly, these digestive side-effects were not clearly reported in studies both performed in endemic areas or in non-endemic areas and were only suggested in one study on imported malaria ,32,39,40Whilst this study constitutes the largest cohort reported on falciparum malaria imported to Europe to date, it has numerous limitations mainly due to its observational design and its overall limited number of observations; and the patient collective being heterogeneous with \u201ccentre effects\u201d. For example, it was impossible to standardize timing of follow-up testing for fever and parasite clearance. Consequently, comparisons between treatment groups or countries, even statistically significant, have to be carefully interpreted. Nevertheless, the methodology used here is the only way to observe a large cohort of imported-malaria patients in routine conditions as data in this specific field of malaria are very limited. To that end, the collection of observational data on the scale of what is presented here should be perpetuated and extended to additional sites involved in the treatment of malaria imported into Europe.Despite limitations in its design, this observational multicentre European study of 504 patients with uncomplicated falciparum malaria offers relevant data on management of imported malaria in Europe. The hospitalization rate appeared high with heterogeneity between centres. The number of treatment regimens was as high as eighteen. Atovaquone-proguanil, the most prescribed drug in this study, appeared to be a valuable option but poor absorption, delay in PCT and FCT and vomiting, frequently inducing a second-line treatment are limiting factors. Due to a poor tolerance profile possibly leading to prolongation of hospitalization, quinine alone should not be recommended any more as a first-line treatment. The study results, by highlighting the current diversity of actual treatments against a backdrop of national and supranational guidelines which favour few established regimens, should stimulate and an intensified consensus discussion at European level, which is desirable in order to more strictly select a limited number of treatments and to foster standardized management of imported malaria.The authors declare they have no conflict of interest.OB contributed to the design of the study, contributed patient data and led writing the paper. NM designed the database, served as the study data manager, analysed the data and contributed to writing of the first draft and final version of the paper. AM contributed to the design of the study, to patient data collection and to the writing of the final paper. All other authors contributed patient data and contributed to the writing of the final paper. MPG conceived and designed the study, served as Principal Investigator and contributed to the writing of the first draft and final version of the paper. All authors read and approved the final manuscript.Main characteristics of 504 malaria patients in different European settings according to their treatment regimen.Click here for file"}
+{"text": "Exchange transfusion (ET) has remained a controversial adjunct therapy for the treatment of severe malaria. In order to assess the relative contribution of ET to parasite clearance in severe malaria, all patients receiving ET as an adjunct treatment to parenteral quinine or to artesunate were compared with patients treated with parenteral treatment with quinine or artesunate but who did not receive ET. ET was executed using a standardized manual isovolumetric exchange protocol.P. falciparum malaria at the Institute for Tropical Diseases of the Harbour Hospital between 1999 and 2011 were included in this retrospective follow-up study. Both a two-stage approach and a log-linear mixed model approach were used to estimate parasite clearance times (PCTs) in patients with imported malaria. Severe malaria was defined according to WHO criteria.All patients in the Rotterdam Malaria Cohort treated for severe A total of 87 patients with severe malaria was included; 61 received intravenous quinine, whereas 26 patients received intravenous artesunate. Thirty-nine patients received ET as an adjunct treatment to either quinine (n\u2009=\u200923) or artesunate (n\u2009=\u200916). Data from 84 of 87 patients were suitable for estimation of parasite clearance rates. PCTs were significantly shorter after administration of artesunate as compared with quinine. In both models, ET did not contribute significantly to overall parasite clearance.Manual exchange transfusion does not significantly contribute to parasite clearance in artesunate-treated individuals. There may be a small effect of ET on parasite clearance under quinine treatment. Institution of ET to promote parasite clearance in settings where artesunate is available is not recommended, at least not with manually executed exchange procedures. Plasmodium falciparum malaria became apparent in two large field trials in Asia and Africa [European travellers returning from malaria-endemic areas are at increased risk of developing severe malaria. If left untreated, the disease can evolve rapidly into a life-threatening disease with a near to 100% case-fatality rate [d Africa ,3, artesd Africa , even thd Africa .P. falciparum malaria in industrialized countries is still associated with a case-fatality rate of up to 10% [In circumstances where optimal anti-malarial and supportive treatment is available, severe p to 10% . In an ep to 10% . Furtherp to 10% . Much ofp to 10% -14.In 1998, ET was added to the standard of care for patients with severe malaria at the Institute for Tropical Diseases of the Harbour Hospital in Rotterdam, with additional selection criteria over the WHO criteria for severe malaria (see below). Given the low case-fatality rate of severe malaria since then, the ET selection criteria and exchange protocol remained virtually unchanged except for replacing quinine by artesunate as the parenteral anti-malarial treatment of choice since 2008. The mainstay of treatment of severe malaria is a rapid reduction of parasite biomass. Since quinine and artesunate both induce parasite elimination but at different rates, the relative contribution of ET to parasite clearance is unknown. In the present study the contribution of ET to parasite clearance was examined by studying patients receiving intravenous quinine or artesunate only as well as patients who received ET as an adjunctive treatment on top of treatment with either intravenous quinine or artesunate.P. falciparum malaria at the Institute for Tropical Diseases of the Harbour Hospital between 1999 and 2011 were included in this retrospective follow-up study. All patients were diagnosed with imported P. falciparum malaria and received intravenous anti-malarial treatment and - when indicated - ET in a well-equipped intensive care unit (ICU). In order to assess the relative contribution of ET to parasite clearance in (severe) malaria, all patients receiving ET as an adjunct treatment to parenteral quinine or to artesunate were compared with patients treated with parenteral treatment with quinine or artesunate but who did not receive ET. In 2008, artesunate became available in the Netherlands and replaced quinine as the treatment of choice for severe malaria. Therefore, four treatment groups could be discerned. Group I (Quinine only) consisted of malaria patients receiving quinine without ET. Group II (Artesunate only) consisted of malaria patients receiving artesunate mono-therapy. Group III (Quinine + ET) consisted of malaria patients receiving ET as an adjunct treatment to parenteral treatment with quinine., whereas group IV (Artesunate + ET) consisted of malaria patients who received artesunate plus ET. In order to properly evaluate the parasite clearance in relation to treatment mode, of each malaria patient, demographic, clinical, and laboratory data were collected using a standardized case report form after de-identification of individual patient data.All patients in the Rotterdam Malaria Cohort treated for severe P. falciparum trophozoites per 100 red blood cells (RBCs) in a thin film or the number of parasites per 100 white blood cells (WBCs) in a thick film. The parasite load  was calculated from these figures using the actual WBC and RBC counts. Other laboratory examinations included haematocrit (Hct), platelet (PLT) count, serum electrolytes and creatinine, liver enzymes and total bilirubin, blood glucose and plasma lactate, as described previously [Malaria was diagnosed following standard procedures that comprise quantitative buffy coat (QBC) analysis, a rapid diagnostic test (RDT) targeting Histidine Rich Protein II , and microscopic examination of thick and thin blood smears prepared from freshly collected blood specimens from finger punctures. Thick blood smears were stained with Field\u2019s stain  and thin smears were fixed with absolute methanol for 3\u00a0minutes and stained with Diff Quick . Parasite density was expressed as the number of eviously . AdditioP. falciparum malaria if they met predefined criteria for severe malaria as published previously [Patients were considered having severe eviously .Patients were considered candidates for ET if they met one of the following additional criteria :\u2022Parasitaemia of more than 10% infected RBCs\u2022Presence of schizonts in peripheral blood\u2022Indication of organ damage \u2022Acidaemia (pH\u2009<\u20097.25)\u2022Diffuse intravascular coagulation\u2022HaemoglobinuriaParasite clearance time (PCT) 50%, PCT 90%, PCT 95%, PCT 99% were defined as the time (in hours) to obtain a 50%, 90%, 95%, 99% reduction in parasite burden after start of the treatment, respectively. T\u2009=\u20090 was defined as the time point at which the first dose of parenteral anti-malarial treatment was given.Quinine dihydrochloride  was administered intravenously with an initial loading dose of 20\u00a0mg/kg  infused over 4\u00a0hours (in 500\u00a0mL of 5% dextrose in water), followed by 10\u00a0mg/kg infused over 8\u00a0hours three times a day  until starting oral therapy.Artesunate  was given intravenously in a dose of 2.4\u00a0mg/kg on T\u2009=\u20090, 12 and 24\u00a0hours later and then daily thereafter until the patient could tolerate oral medication.ET was performed using a manual isovolumetric approach as described previously . The ET Both a two-stage approach  and log-50), 90% (PCT90), 95% (PCT95) and 99% (PCT99) clearance of the parasites was calculated. Treatment mode (artesunate or quinine), exchange transfusion (yes or no) and time were entered as explanatory variables, including the interaction between treatment and time and between exchange transfusion and time. To account for the repeated measurements, intercepts and slopes were allowed to change over patients. Linear mixed effects modeling was done on the logarithmic values of the parasite count using the MIXED function in SPSS.To account for these imitations, the data were also analysed with a population model. With population modelling, such as with linear mixed models, all data from the entire population are analysed simultaneously, while still taking into account the correlation in the data, i.e. grouping the observations from individual patients . It addiIn this study, both methods were used to estimate the role of exchange transfusion in parasite clearance since the WWARN model has recently been promoted as the method of choice for estimating parasite clearance . AlthougAll analyses were done with software SPSS version 18.0 .e.g. comparing quinine\u2009+\u2009ET (group III) with artesunate\u2009+\u2009ET (group IV) and comparing quinine only (group I) with artesunate only (group II) on the other hand) with the use of non-parametric Mann\u2013Whitney test. A p-value of less than 0.05 was considered significant.Demographic data, clinical presentations and outcome were compared between patients who were treated with quinine or artesunate alone and patients who received ET adjunctive to quinine or artesunate using Kruskal-Wallis test for non-parametric comparisons followed by Dunn. Since patients receiving exchange transfusion are more likely to present with more severe disease, statistical comparisons focused on comparisons of groups with comparable disease severity  were classified with severe malaria (one or more WHO criteria), the remaining 31 patients were given parenteral anti-malarial (24 received quinine while seven received artesunate monotherapy) because parasitaemia was between 2-5%, which is the per-protocol treatment policy at the Harbour hospital of Rotterdam. Patients receiving ET presented with more severe disease (including significantly higher parasitaemia) as compared with malaria patients not receiving ET  or artesunate (n\u2009=\u200916). As shown in Table\u00a0The crude parasite count of the 87 malaria patients and the parasite clearance in relation to time and treatment are shown in Figure\u00a0vs 122,567  respectively, p value <0.0001). In contrast, in the model that estimates the parasite clearance rate, the slope of the elimination curve significantly improved when the treatment modality (quinine or artesunate) was added as a random factor but not when adding adjunctive treatment by ET or not.The findings of the mixed effects model evaluating the effect of time (as a fixed effect) and treatment mode, ET and their interaction (random effects) on parasite clearance are summarized in Table\u00a0versus 6.6 (95% CI 6.1-7.2) hours, p value <0.0001, Additional files 90 and PCT95) but not at PCT50 and PCT99 , there were no differences at baseline between artesunate and quinine treated patients and this suggest that a proper comparison is allowed. A prospective comparative trial in industrialized non-endemic countries is deemed not feasible because the number of eligible patients suffering from severe imported P. falciparum malaria is treated with intravenous quinine, there may be a small beneficial effect of manual ET on parasite clearance. However, there is no benefit of ET on parasite clearance in artesunate-treated individuals. Based on these findings manual exchange transfusion is no longer recommended for patients with severe P. falciparum malaria who are being treated with intravenous artesunate.When severe ARKV is PhD fellow, employed by ACE Pharmaceuticals. The other authors declare that they have no competing interests.ARKV, MdMM, RK, JJvH and PJvG collected data. The data were analysed by ARKV, PJdV and PJvG. ARKV and PJvG wrote the first draft of the manuscript. The database of the Rotterdam Malaria Cohort is maintained by RK. All authors were involved in critical revision and approval of the paper.Parasite clearance curves according to the two-stage approach according to WWARN analysis (left panel) and according to the linear mixed model analysis (right panel).Click here for fileParasite clearance times of various modes of parenteral anti-malarial treatment. Data are given as mean .Click here for fileParasite clearance parameters (WWARN model) in relation to mode of parenteral anti-malarial treatment and adjunct treatment. Data are given as mean .Click here for file"}
+{"text": "Chlorproguanil-dapsone (Lapdap), developed as a low-cost antimalarial, was withdrawn in 2008 after concerns about safety in G6PD deficient patients. This trial was conducted in 2004 to evaluate the safety and effectiveness of CD and comparison with artemether-lumefantrine (AL) under conditions of routine use in G6PD normal and G6PD deficient patients with uncomplicated malaria in The Gambia. We also examined the effects of a common genetic variant that affects chlorproguanil metabolism on risk of treatment failure.1238 children aged 6 months to 10 years with uncomplicated malaria were randomized to receive CD or artemether-lumefantrine (AL) and followed for 28 days. The first dose was supervised, subsequent doses given unsupervised at home. G6PD genotype was determined to assess the interaction between treatment and G6PD status in their effects on anaemia. The main endpoints were clinical treatment failure by day 28, incidence of severe anaemia (Hb<5 g/dL), and haemoglobin concentration on day 3.One third of patients treated with AL, and 6% of patients treated with CD, did not complete their course of medication. 18% (109/595) of children treated with CD and 6.1% (36/587) with AL required rescue medication within 4 weeks, risk difference 12% (95%CI 8.9%\u201316%). 23 children developed severe anaemia (17 (2.9%) treated with CD and 6 (1.0%) with AL, risk difference 1.8%, 95%CI 0.3%\u20133.4%, P\u200a=\u200a0.02). Haemoglobin concentration on day 3 was lower among children treated with CD than AL , and within the CD group was lower among those children who had higher parasite density at enrolment. Only 17 out of 1069 children who were typed were G6PD A- deficient, of these 2/9 treated with CD and 1/8 treated with AL developed severe anaemia. 5/9 treated with CD had a fall of 2 g/dL or more in haemoglobin concentration by day 3.AL was well tolerated and highly effective and when given under operational conditions despite poor adherence to the six-dose regimen. There were more cases of severe malaria and anaemia after CD treatment although G6PD deficiency was uncommon.NCT00118794Clinicaltrials.gov  Chlorproguanil-dapsone (CD) was developed as a low-cost treatment for uncomplicated malaria In The Gambia the first line treatment for uncomplicated malaria was changed from chloroquine (CQ) to sulfadoxine-pyrimethamine plus CQ in 2004 following the finding of 28% clinical failure rate in children treated with CQ in 2003 , and artemisinin combination therapy with artemether-lumefantrine (AL) was adopted in 2007. The present trial was conducted in 2004 to evaluate the safety of CD and its effectiveness in operational settings in comparison with AL, and to compare effects of these drugs on anaemia in G6PD normal and G6PD deficient patients with uncomplicated malaria. We also examined the effects of a common genetic variant that affects chlorproguanil metabolism on risk of treatment failure. We aimed to evalue the alternative regimens under conditions as similar as possible to those of routine use, when doses given at home are not supervised by trial staff, since adherence may affect outcome P. falciparum with density between 500\u2013200,000 parasites/\u00b5L, PCV \u226520%, no serious concomitant illness, and a parent or guardian had given written consent. Patients were excluded if there were signs or symptoms of severe malaria . Axillary temperature was measured using a digital thermometer, the patient was weighed, and a finger prick blood sample was collected for measurement of packed cell volume, haemoglobin concentration and malaria microscopy. The study was explained by a field assistant who sought written consent. Patients were then assessed by a nurse or clinician , and were invited to enrol into the study if they had monoinfection with The randomization list was produced with Stata version 8 (Statacorp Texas) using permuted blocks of size 10. Eligible children were allocated to receive three daily doses of CD , or a six-dose course of AL . Patients who were enrolled were issued with an opaque envelope, labelled with the next sequential study number, containing the treatment allocation and an appointment card for follow-up visits pre-printed with the same study number. The children were then referred to a drug dispenser, a nurse who took no part in patient assessment, who opened the randomization envelope and supervised administration of the first treatment dose. Dosages of both medications were weight-determined according to manufacturers' weight cut-off instructions and were dispensed in their original blister packs. For AL, patients received the World Health Organization (WHO) weight-specific blister packs as follows: 10.\u221214.9 kg: 1 tablet per dose;15.0\u221224.9 kg: 2 tablets; 25.0\u221234.9 kg: 3 tablets, and \u2265 35 kg: 4 tablets. The packs contained pictorial instructions on the regimen schedule. CD was supplied in two formulations: Lapdap 15/18.75, tablets containing 15 mg chlorproguanil hydrochloride and 18.75 mg dapsone are for use in infants and young children, white capsule-shaped tablets with a break line on one side for easy dispensing as half tablet dose and marked CD15 on the other side, and Lapdap 80/100 tablets containing 80 mg chlorproguanil hydrochloride and 100 mg dapsone, pink capsule-shaped tablets indicated for the treatment of older children and adults. Both tablet formulations have a break line on one side and are marked with the strength (CD15 or CD80) on the other side. CD was given according to weight. Children under 16 kg received Lapdap 15 tablets . Children 16 kg and over received Lapdap 80 tablets .The drug dispenser explained the number and timing of treatment doses to be given, and the first dose of medication was given by the parent/carer at the health centre under direct observation. Patients were observed for 30 minutes, and children who vomited during the observation period had their doses repeated. If the child vomited both the first and replacement medication then a rescue medication was given . In addition to the study drugs, oral paracetamol (10 mg/kg) was given in the clinic and additional doses were given to parents for administration to the children three times a day at home. Iron tablets (0.5 tablet per day for children under 5 and 1 tablet per day for older children) to be taken daily for 2 weeks, were given to children whose PCV at enrolment was less than 30%.The parent or carer was encouraged to bring the child back to clinic if the condition did not improve or if they had any concerns about the child's health. Subsequent antimalarial treatment doses were to be given at home unsupervised. The parent/carer was requested to return the patient to the clinic on days 14 and 28 after treatment for clinical evaluation and blood sampling for measurement of haemoglobin concentration, PCV, malaria microscopy and parasite genotyping. Children were visited at home on the day after the regimen should have been completed (day 3) to ask about compliance with the treatment regimen and adverse events, and to check for signs of anaemia and haemolysis . Blister packs were inspected to count left over medication, and a finger prick blood sample was collected from the child to measure haemoglobin concentration. Patients with severe anaemia (defined as Hb \u22645 g/dl) or Hb that had fallen by \u22652 g/dl from Day 0 measurement were promptly referred to the clinic for clinical assessment and treatment . Nurses and drivers were placed on 24-hour emergency duty at the health centres/hospital where the study was sited to handle emergencies.P. falciparum negative up to day 28, frequency matched by centre. In a secondary analysis, a subset of these cases, defined as children with parasitological treatment failure by day 14, was used. In addition, for the CD treatment failures, parasites were typed from samples collected on the day of failure, to detect any increase in the proportion of children carrying parasites with the resistant mutations. The same cases and controls were used to investigate the possible association between CYP2C19 genotype and CD treatment failure. This cytochrome P450 enzyme is responsible for activating chlorproguanil. The *2 allele is a frequently-occurring functional CYP2C19 variant characterised by a G/A polymorphism in exon 5 which creates an aberrant splice site. The resulting change in reading frame of the mRNA introduces a premature stop codon which results in a truncated and non functional protein A nested case control study was used to determine whether CD is less effective in children who when screened carried parasites with the triple dihydrofolate reductase (DHFR) mutations encoding isoleucine at position 51 (51I), argentine at position 59 (59R), and asparagines at position 108 (108N). Cases were children treated with CD who had detectable parasitaemia by day 28, and controls were children from the same treatment group who remained At screening and at follow-up visits, two thick blood films were prepared, one was stained with Field Stain and read immediately for malaria diagnosis for enrolment, the second was dried overnight and stained with Giemsa for definitive microscopy. One hundred high power field fields (HPFs) were read before a slide was declared negative. Parasite density was determined assuming one parasite per HPF equivalent to 500 parasites/\u00b5L blood Using DNA extracted from blood spots, the DHFR codon 51, 59 and 108 alleles were genotyped by polymerase chain reaction (PCR) amplification followed by analysis restriction fragment length polymorphism (PCR-RFLP) G6PD genotype was estimated using an ARMS PCR technique to detect the two A-associated mutations in G6PD enzyme, an A to G transition at position 376, encoding the B  to A change and a G to A transition at position 202, encoding the specific A- change CYP2C19 typing was performed using the Taqman real time PCR technique to detect the *2 allele by the presence or absence of the A/G polymorphism The primary endpoint was clinical failure by day 28 and change in haemoglobin between day 0 and day 3. To demonstrate that the treatments were at least as good as each other assuming a non-inferiority limit of 3% with at least 80% power at 0.025 significance level, 1180 patients were required to be randomized, allowing for 10% loss to follow up. Since patients with glucose-6-phosphate dehydrogenase (G6PD) A- deficiency are more susceptible to the haemolytic effects of the dapsone component of CD we were interested in the interaction between G6PD status and treatment drug in their effects on haemoglobin concentration. We estimated that a trial of this size would have 80% power (with a 5% significance level) to detect an interaction between treatment drug and G6PD status if the average fall in Hb by day 3 was 2 g/dl in normal patients in both drug groups, and if the fall is 3.1 g/dl or more in G6PD patients who took CD. Other secondary endpoints were the overall clinical and parasitological failure rate (including detection of parasitaemia after day 3), severe anaemia (Hb\u22645 g/dL) at any time during the 28-day follow-up, and a drop to 2 g/dL or more below the screening value at any time during the 28 days. An analysis plan was written before the treatment code was linked to the study database.95% CI for risk differences were computed using a generalised linear model with an identity link to adjust for centre, in Stata 8.2 . Primary analysis was according to protocol . Changes in Hb concentration from baseline were compared between groups by using analysis of covariance, adjusting for the baseline value as a covariate.The study protocol was written following the principles of the Declaration of Helsinki and was reviewed by the MRC Scientific Coordinating Committee (SCC) and approved by the joint Gambian Government/MRC Ethics Committee. Meetings were held in study villages to explain the aims of the study. At the clinics, the aims and procedures and potential risks were explained to the mother or carer of children who had screened positive for malaria, in their usual language, and their signed consent was sought before the child was enrolled. A Data Safety Monitoring Board was set up for the trial and an experienced physician, who was not part of the trial team, was appointed as local safety monitor to advise on adverse events. The protocol for this trial and supporting CONSORT checklist are available as supporting information; see P.falciparum, 26 for P.malariae and 2 with P.ovale. Of those with P falciparum monoinfection, 1238 were enrolled and randomized to receive AL or CD. Of the remaining 593 who did not satisfy the inclusion criteria, 251 (42%) had low density asexual parasitaemia (<500 parasites/\u00b5L), 90 (15%) had signs and symptoms of severe malaria, 55 (9.3%) were severely anaemic (PCV <20%). 54 (9.1%) patients were not enrolled because parents/guardians did not consent to their participation in the study, 47 had concomitant disease, 42 had very high density asexual parasitaemia . A total of 1122 (90.6%) patients  were seen on day 28. The trial profile is shown in 2698 children presenting with fever, history of fever or other symptoms suggestive of malaria were screened, 1831 (68%) were positive for 2[1 df]\u200a=\u200a0.148, P\u200a=\u200a0.7008) with an allele frequency of 0.168.4 children randomised to AL who received CD in error and 1 child randomised to CD who received AL, were analysed according to the treatment received. 56 patients with no outcome data: 29 withdrew consent to be followed up  and 27 were not contactable due to wrong address or residence across the Gambian border  were excluded from the analysis. There were 44 serious adverse events. 13 children were admitted with severe malaria ; 23 children had severe anaemia (Hb<5), 17 (2.9%) in the CD group and 6 (1.0%) in the AL group  and 8 children were hospitalized with other conditions, not thought to be related to treatment (5 in the CD group and 3 in the AL group). There were no deaths.2Of the 23 severe anaemia cases, 19 occurred on or before day 3 (15 in the CD group and 4 in the AL group). Children were given iron and folate to be taken at home. Although transfusion was recommended for 10/23 children, only 2 of these were transfused, parents of the other 8 refused. All 23 severe anaemia cases were seen again on day 14 or 28 and their Hb found to have improved (range 7.6 g/dL\u201311.7 g/dL). One child who had Hb of 5.3 g/dL, and therefore not severely anaemic according to our definition, was brought back to the clinic on day 2 was transfused. G6PD genotype was determined for 1069/1238 children. The frequency of the A- allele was 3.1% (95%CI 2.3% to 4.1%), the genotypes of boys and girls were in Hardy-Weinberg equilibrium X2.Of the 13 severe malaria cases, 7 occurred in the first 3 days, all in the CD group. G6PD genotype was obtained for 12 of these cases, all were normal.4 children who received AL and 6 who received CD vomitted the first and repeat dose of study medication on day 1 and were given rescue treatment.On day 3, 93 of 601 (15.5%) parents/carers of patients in the CD group and 25 of 606 (4.1%) in the AL group said their children were still unwell, the most commonly reported symptoms were fever, headache, gastrointestinal complains , and cough.109/595 children (18%) in the CD group and 36/587 (6.1%) in the AL group required rescue treatment within 4 weeks, risk difference adjusted for centre 13% (95%CI 8.9% to 16%). By day 14, the corresponding rates were 5.8% and 0.9% .Clinical and parasitological failures at days 14 and 28 were significantly higher in the CD group compared with AL treatment (p<0.001). There was an increase in mean haemoglobin from baseline by day 28, with values significantly higher in the AL group compared with CD treatment. The incidence of anaemia by day 28, defined as haemoglobin <5 g/dl, was higher in the CD group.Mean haemoglobin concentration fell after treatment in both groups, with values in the CD group lower on day 3 than those in the AL group . To determine whether CD treatment increased the risk of anaemia among G6PD deficient patients, the interaction between treatment group and G6PD status in their effects on Hb concentration on day 3 needs to be examined. G6PD genotype was determined for 1069 of 1238 (86%) children. Boys carrying the A- allele, and girls homozygous or heterozygous for the A- allele, were identified and classified by DNA genotyping. Hb concentration on day 3 was lower in G6PD A- than in G6PD wild type children, and was lower in children treated with CD than those who received AL, but the number of A- children was too small to evaluate the strength of interaction. When the analysis was limited to G6PD A- deficient hemizygous boys and to the one homozygous girl identified, (thus excluding heterozygous girls who may not be deficient) similar results were obtained; in this subgroup, Hb was slightly lower in the CD group at baseline (9.3 g/dL on day 0 compared to 10.4 g/dL in the AL group); on day 3 the corresponding mean values were 7.2 g/dL and 8.6 g/dL.Children with higher parasite densities on day 0 were at increased risk of anaemia after treatment. This effect was evident in both drug groups at day 3 and day 14, and on day 3 appeared to be more marked in the CD group  is 0.41/dL, 95%CI 0.10 to 0.71, P\u200a=\u200a0.009), Of children in the AL group whose mothers were interviewed about compliance, 402/600 (67%) took all 5 doses at home, 190 (32%) took some but not all the recommended doses at home, 7 (1.2%) took no medication at home. There was no association of compliance with outcome (data not shown). When blister packs were inspected 186/591 (31%) of those whose packs were seen had left-over medication. Of 599 children in the CD group with compliance information, 562 (94%) took all 3 doses, 12 (2%) took no doses at home, 25 (4%) took only 1 dose at home. The poorer compliance with the AL regimen primarily reflects the larger number of doses. Again no association of compliance with outcome was evident. 51/568 (9%) whose blister packets were inspected had left-over medication. The most common reason given for not giving treatments at home was that the mother forgot or did not understand the regimen.2 6.25 (1df) p\u200a=\u200a0.0124).Of 229 cases, 8 individuals were excluded because they did not receive CD: 6 vomitted study medication and repeat dose and were given rescue medication and 2 assigned to CD received AL in error, leaving 214 cases, and 206 controls. Parasite DHFR genotyping data was obtained from the three polymorphisms  for 181/214 cases and 170 controls frequency matched on centre. 161/181 (89%) cases were positive for the triple DHFR resistant allele  when enrolled compared to 146/170 (86%) among controls (odds ratio 1.3 (95%CI 0.7\u20132.5) P\u200a=\u200a0.404). Considering parasitological failures by day 14, 51/54 (94%) of treatment failures were positive for the triple DHFR mutation compared to 146/170 (86%) among controls, odds ratio adjusted for centre 3.0 . 111 of the parasitological failures by day 28 were typed on both the day of screening and the day of failure. On day 0, 97/111 (87%) were positive for the triple mutation, on the day of failure, 134/139 (96%) were positive, a significant increase  P\u200a=\u200a0.803, and 2.9% and 2.3% respectively were homozygous (odds ratio 0.81 (0.24\u20132.72) P\u200a=\u200a0.731). Similar results were obtained when cases were defined as parasitological failures by day 14.This study, conducted as far as possible under conditions similar to those of routine use of drug treatments by nurses in rural clinics, showed that CD was less effective than AL, there were more early returns to clinic, and higher incidence of severe malaria cases and severe anaemia after treatment, although the frequency of G6PD deficiency was low. In our study only 17/1069 (1.6%) subjects were A- deficient (1 homozygous girl and 16 hemizygous boys). AL, although given without food and with doses other than the first unsupervised, was well tolerated and highly effective among children from 6 months to 10 yrs of age.Assessment of the safety of antimalarials is complicated when potential side effects of the drugs are also symptoms of the illness, and is complicated further when the risk of adverse reactions is confined to a relatively small subgroup of patients. CD was developed by a public-private partnership as a low-cost treatment for uncomplicated falciparum malaria et al.In our study anaemia after treatment was more common with CD, but the low frequency of the G6PD A- genotype limited the power of our study to assess this interaction with genotype. In the normal pattern of G6PD deficiency no evidence of haemolysis is apparent until 48\u201372 hours after substance ingestion In sub-Saharan African populations, the most common G6PD deficiency is due to the A- allele with 12% enzymatic activity. G6PD A- is present at high frequencies in many sub-Saharan African countries  et al.The G6PD ARMS assay used in our study was validated on 100 different G6PD 202 genotypes by an RFLP- based assay Treatment failure rates after CD in our study were high but not dissimilar to those found in other studies, a comparable CD parasitological failure rate, mainly attributed to reinfections, has been reported from Kilifi District Hospital, Kenya, where a randomised trial of CD using a similar study design in children aged 6 months-5 years with uncomplicated malaria resulted in day 28 parasitological failure in 41% (95% CI 33\u201349) The presence of the DHFR triple resistant mutant at day 0 was not strongly associated with CD treatment failure but the high frequency of this mutation, similar to that seen previously We found the frequency of the *2 CYP2C19 allele in Gambians to be similar to that in Europeans. The poor metaboliser phenotype is much more common in Asia (approximately 20%) due to an increased frequency of the *2 allele and the presence of another loss of function allele *3 The parasitological failure rate of CD in this study was high. Its slow action, evidenced by symptom clearance by day 3, could be due to poor chlorproguanil metabolism but we failed to see an association with the CYP2C19*2 allele. There has been relatively little work on the pharmacogenetics of antimalarial drugs in Africa\u2013much of our information on potentially relevant alleles comes from Europe and Asia. It would be useful to define the full range of genetic variants in this population which could help to inform policy decisions.et al.One third of patients treated with AL did not complete their course of medication but despite poor adherence to the recommended regimen, AL had excellent effectiveness up to 28 days after treatment. This is in contrast to a study in Papua New Guinea by Schoepflin Checklist S1CONSORT Checklist.(DOC)Click here for additional data file.Protocol S1Trial Protocol.(DOC)Click here for additional data file."}
+{"text": "Adverse drug reactions (ADRs) contribute to ill-health or life-threatening outcomes of therapy during management of infectious diseases. The exposure to anti-malarial and use of mobile phone technology to report ADRs following drug exposures were investigated in Sagamu - a peri-urban community in Southwest Nigeria.Purchase of medicines was actively monitored for 28 days in three Community Pharmacies (CP) and four Patent and Proprietary Medicine Stores (PPMS) in the community. Information on experience of ADRs was obtained by telephone from 100 volunteers who purchased anti-malarials during the 28-day period.A total of 12,093 purchases were recorded during the period. Antibiotics, analgesics, vitamins and anti-malarials were the most frequently purchased medicines. A total of 1,500 complete courses of anti-malarials were purchased ; of this number, purchases of sulphadoxine-pyrimethamine (SP) and chloroquine (CQ) were highest (39.3 and 25.2% respectiuvely). Other anti-malarials purchased were artesunate monotherapy (AS) - 16.1%, artemether-lumefantrine (AL) 10.0%, amodiaquine (AQ) - 6.6%, quinine (QNN) - 1.9%, halofantrine (HF) - 0.2% and proguanil (PR) - 0.2%. CQ was the cheapest (USD 0.3) and halofantrine the most expensive (USD 7.7). AL was 15.6 times ($4.68) more expensive than CQ. The response to mobile phone monitoring of ADRs was 57% in the first 24 hours (day 1) after purchase and decreased to 33% by day 4. Participants in this monitoring exercise were mostly with low level of education (54%).The findings from this study indicate that ineffective anti-malaria medicines including monotherapies remain widely available and are frequently purchased in the study area. Cost may be a factor in the continued use of ineffective monotherapies. Availability of a toll-free telephone line may facilitate pharmacovigilance and follow up of response to medicines in a resource-poor setting. Unfortunately emergence of Plasmodium falciparum resistance to these drugs in South East Asia and eventual spread to all malaria endemic areas compromised malaria treatment of falciparum malaria in several malaria endemic countries, including Nigeria [Infection with malaria parasites remains one of the leading causes of hospital visits in Nigeria. Half of the estimated population of 150 million experience at least one malarial attack each year . Chloroq Nigeria  Artemisi Nigeria -4. As AC Nigeria ,7. Anti- Nigeria  and thus Nigeria . This st2 (237/Sqm) with an estimated population of 228,382. A large proportion of the population commutes to the city of Lagos daily for work or other commercial activities. Malaria is highly endemic in the area, accounting for most outpatient visits in the health facilities. Transmission occurs all year around with an upsurge in the rainy season - June to September [Sagamu, located 50 km north of Lagos is the seat of the Sagamu Local Government Area, Ogun State in south-west Nigeria. The town is spread over 614 Kmeptember . The comStudy centers were identified and selected from the list of registered pharmacies and Patent and Proprietary Medicine Stores (PPMS) compiled by the Department of Pharmaceutical Services, Ministry of Health, Ogun State. There are 128 PPMS and 13 pharmacies listed in the state's official records. The patient load in these facilities record average of 225 per day; .http://www.surveysystem.com/sscalc.htm. The specific facilities were chosen using computer generated simple randomization procedure.From the 141 drug outlets a sample size of seven (three pharmacies and four PPMS) was determined at confidence interval of 50% and confidence level of 95% using Survey System sample size calculator The study was descriptive and prospective, and involved active follow-up of sales of drugs in the selected centres for a period of four weeks (28 days) to determine the exposure to anti-malarial drug, measured by the amount of drugs purchased within the period. A drug-data chart was designed to capture information on category of drugs purchased. The proportion of anti-malarial of the different drugs purchased at the drug retail outlets, which at the time of study were not tracked by any active surveillance method, and sales per week of the medicines were determined. The anti-malarial drugs were classified to assess the drug use pattern for the different anti-malarial drugs and the rate of purchase between PPMS and the community pharmacies. The retail prices of the different anti-malarial drugs in a complete dosage pack were recorded.The willingness of the buyers of the anti-malarial drugs in this community to participate in pharmacovigilance (PV), through a toll free mobile phone monitoring of adverse drug reactions, was evaluated in a total of 100 subjects who presented at the study sites to purchase anti-malarial drugs, volunteered to participate and provided information via mobile phones over a period of two weeks following purchase of medicine. The mobile phones were used to provide information on any ADRs experienced following ingestion of the anti-malarial drug, particularly ACT.Prior to enrolment in the mobile phone ADR monitoring, the objective was explained to each of subjects after drug purchase was completed by a member of the study team and informed consent obtained.The mobile phone number of the participant was obtained on permission and entered into a registration form. One member of the study team called the mobile number to validate correctness of entry and connectivity. A series of questions were prepared to inquire about any complaint or reactions observed by the subjects to whom the drug is administered, if adult, or a caregiver when subject is a child. Three of the investigators  were involved in the documentation of any complaints or observations following drug use. Each participant is called from mobile phone designated for the exercise by a member of the study team. The participants mobile phones were called at specified times to record time of use of drug and 6-hourly for the first three days and 24 hourly from day 3 to day 14. All complaints or observations were recorded.The pharmacies and medicine stores operate on an average of 13 hours daily (09.00 hr - 22.00 hr). Drugs purchased are recorded daily to monitor sales and to prevent stock-outs. The data on anti-malaria drug sales was collected daily from the outlets in the 28 days and transferred into a computer database. Information provided during mobile phone calls were transferred to forms designed for the purpose.Following compilation of mobile phone interaction with patients or caregivers, each complaint or observation was examined and determined qualified for ADR. A complaint or response to inquiries by one of the research team member on phone is categorized as ADR from a patient if it qualifies to be 'any response to the drug which is noxious and unintended, and that occurs at doses normally used in man for prophylaxis, diagnosis, or therapy of diseases or modification of physiological function'; and an adverse event if 'any untoward medical occurrence present in a patient administered the medicine and which does not necessarily have to have causal relation with the treatment'.Generic names, brand names (if possible), pharmacological classifications, collection centre, date of collection and day of study (0-27 days) were entered into a Database. The data generated were analysed using SPSS version 10 for Windows . Chi-square analysis was used to compare proportions and linear regression analysis was used to obtain relationship between responses and duration of follow up for the monitoring aspect of this study.Ethical approval for the study was obtained from the Joint Ethics Committee of the Olabisi Onabanjo University Teaching Hospital and Obafemi Awolowo College of Health Science, Olabisi Onabanjo University, Sagamu.The study was carried out between May-September 2010. In the seven drug retail outlets, the distribution of drugs purchased in this community in the period of the study is shown in Table Of 10,581 complete treatment dosage drugs purchased at the pharmacies, 10.7% were anti-malarials. Sales of anti-malarials at the PPMS were 23.7% of complete treatment dosages  sold. The weekly sales of anti-malarial and other categories of drugs from these outlets are shown in Table 2 = 13.6, P = 0.0002).The various anti-malarial drugs that were available and frequently purchased from the study drug outlets were sulphadoxine-pyrimethamine (SP), chloroquine (CQ), artesunate (ART), artemether-lumefantrine (AL), amodiaquine (AQ), quinine (QN), herbal preparations (HP), halofantrine (HF), and proguanil (PR). Table On average, artesunate was the most purchased anti-malarial at the pharmacies each week of the study period, but the least purchased at the PPMS. CQ was however the least purchased at the community pharmacies while it was the most purchased at the PPMS. The retail price per dose of anti-malarial drug Table  varied w100 volunteers (84 women and 26 men) participated in the toll free mobile phone monitoring of adverse drug reactions. 54% of the participants had between 6 and 12 years of formal education and 46% of the subjects had more than 12 years of formal education (post-secondary school), 5% of them were healthcare providers (nurses or doctors). The trend of the responses received showed that most people responded within the first twenty-four hours of follow-up. By 96 hours, the level of responses began to decline  is purchased at the PPMS. This implies that people in the study area, prefer PPMS as the first point of contact for anti-malarials than the pharmacies. Since the Alma Ata Declaration recognized the Primary Health Care (PHC) centres as positioned to be the first contact to individual, family and community, the frequency of visit of the people studied on PPMS for anti-malarials may suggest that the primary function of the PHC is not optimized in the study area. The centres are either underutilized or not easily accessible. The number of PPMS (128) and community pharmacies (13) in the community is a strong indicator of the challenges of PHC programme. PPMS appear more accessible given the high purchase rate for anti-malarial drugs recorded in this study and may be considered viable units for intervention programmes for malaria control in endemic areas, where PPMS exist. Negligence of these drug retail outlets may in turn, increase the risk of incorrect dosing, inappropriate treatment, impacting negatively on drug safety .The large volume of analgesics may suggest the probability that most people at the point of purchasing anti-malarials also buy analgesics to relief the symptoms of malaria, which include fever and arthralgia. The observation during the study showed that over 50% of those purchasing anti-malarial drugs also bought analgesics.The frequency of purchase of anti-malarials in communities will differ greatly and may influence serious ADRs, such as allergic and alveolitis. Despite continuing education by the Federal Ministry of Health, SP and CQ use still remains high despite widespread resistance reported in the area of study ; indicatResistance to CQ and failure of mono therapy in many endemic countries led to a widespread promotion of ACT . NeverthThe monitoring of adverse reactions to these drugs should be an important component of the health care system. The challenges of pharmacovigilance in Africa are not unconnected to the poor reporting structures and lack of motivation to report. The present study evaluated the use of mobile phone technology for monitoring ADRs, especially knowing that the exposure to antimalarial is relatively high in the community. Finding from the study showed that mobile phones offer a practical means of reporting adverse reaction to anti-malarial drugs and may be a model of choice in Africa where mobile telephony penetration and coverage is already driving innovation in agriculture and commerce . The entThe aim of this research is to provide \"proof of principle\" of an innovative approach to support pharmacovigilance in Africa. With over 60% of Africans, in the cities and small villages owning mobile phones, it is exIt is noteworthy that retail sectors do serve as source of malaria treatment and care, complementary to health facility , howeverAlthough, there was neither increase in the frequency of known ADRs or new ADRs detected during the study, it is important to ensure continuing monitoring of anti-malarial drugs safety and scale up use of mobile phone technology to support pharmacovigilance of anti-malarial drugs.ADRs: Adverse drug reactions; AL: Artemether-lumefantrine; AQ: Amodiaquine; AS: Artesunate monotherapy; CP: Community Pharmacies; CQ: Chloroquine; HF: Halofantrine; NSAIDs: Non Steroidal Anti-inflammatory Drugs; PHC: Primary Health Care; PPMS: Patent and Proprietary Medicine Stores; PR: Proguanil; QN: Quinine; SP-Sulphadoxine-pyrimethamine; USD: United State DollarsThe authors declare that they have no competing interests.AAA participated in the conception of the study, field collection of data, analysis and writing of the manuscript. BS, AT, MA, OO, MOA, and OAT participated in planning the study, field collection of data, entering of data and analysis, and writing of the manuscript. IAO- participated in the conception of the study, planning of field work and monitoring. OAO- participated in the conception of the study, planning of field work and monitoring. OOA - participated in the conception of the study, planning of field work and monitoring. TTA- participated in the conception of the study, technical analysis, evaluation and writing of script. FAF - participated in the conception of the study, technical analysis, evaluation and writing of script. OATO- participated in the conception of the study, technical analysis, evaluation and writing of script. All authors have read and approved the final manuscript."}
+{"text": "Plasmodium vivax led to questioning of the current malaria treatment guidelines. A therapeutic efficacy study was conducted using artemether-lumefantrine + primaquine against P. vivax to evaluate a treatment alternative for chloroquine.In Guyana, chloroquine + primaquine is used for the treatment of vivax malaria. A worldwide increase of chloroquine resistance in P. vivax infection were treated with artemether-lumefantrine as a six-dose regimen twice a day for three days with additional 0.25 mg/kg/d primaquine at day 0 for 14 days. Clinical and parasitological parameters were followed on days 0,1,2,3,7,14 and 28 in agreement with WHO guidelines. Plasmodium vivax DNA from eight patients was analysed for pvmdr1, molecular marker of resistance.From 2009 to 2010, a non-controlled study in two hospitals in Guyana was conducted. A total 61 patients with pvmdr1 Y976F polymorphism was detected. The treatment regimen was well tolerated.Artemether-lumefantrine cleared 100% of parasites on day 1, but two patients (3%) had recurrence of parasites on day 28, suggesting relapse. No P. vivax when combined with primaquine. Availability of this alternative will be of great importance in case of emerging chloroquine resistance against P. vivax.In Guyana, artemether-lumefantrine represents an adequate treatment option against  Plasmodium vivax, followed by Plasmodium falciparum[P. vivax responsible for three quarters and P. falciparum for one quarter of cases [Approximately 216 million malaria cases were reported world-wide in 2010 [P. falciparum to artemisinin-derivatives. Although in vitro resistance to artemether is reported for P. falciparum from South America [P. vivax is available.Artemisinin-based combination therapy (ACT) has been and still is one of the main pillars of the treatment of malaria and is responsible for the reduction of malaria infections worldwide -7. In reAmerica , so far P. falciparum and 51.3% by P. vivax. The number of malaria cases declined rapidly until 2007, but increased slightly in 2009, with 93% of the population still being at risk of contracting malaria [In Guyana, 42.3% of all malaria cases between 2005 and 2009 were caused by P. falciparum and chloroquine + primaquine for P. vivax infections.Anti-malarial drug policies in South American countries, including Guyana, use ACT for uncomplicated P. vivax was first reported in 1989 from Papua New Guinea [P. vivax cases in 2011 [Chloroquine resistance against  Guinea . Since t Guinea -16. Thaiin 2011 , while iin 2011 . Reportsin 2011 .P. vivax malaria, as a decline in susceptibility towards chloroquine can be predicted for the coming years. In the light of poor access to diagnostic facilities and emerging chloroquine resistance it is important to test the efficacy of artemether-lumefantrine as a common malaria treatment in Guyana. In remote areas, WHO recommends the use of rapid tests for malaria diagnosis. While several of these commercially available tests showed very good performances, their sensitivity and specificity for vivax Malaria is generally lower than for falciparum malaria. Since malaria species diagnosis could be uncertain, the issue of treatment efficacy of a single common treatment for all circulating plasmodium species had to be addressed.This study aims to assess artemether-lumefantrine + primaquine as an alternative treatment option for pvmdr1 single-nucleotide polymorphism at codon Y976F, suspected to be involved in P. vivax resistance to chloroquine [Further the study aims to screen for roquine .P. vivax. Samples have been collected at two sites, the Malaria clinic Georgetown, situated in the administrative district 4 , and at Port Kaituma Hospital in region 1  of Guyana.A prospective multicentre study was conducted in 2009/2010 for artemether-lumefantrine + primaquine against P. vivax mono-infection.Included were all patients older than six month, weighing more than 10 kg and with a parasitological confirmed Patients were excluded from the study, if any of the following occurred: (i) presence of general danger signs in children aged under five years or signs of severe falciparum malaria according to the definitions of WHO , (ii) prP. falciparum infections, according to the national drug policy. They received artemether-lumefantrine  as a six-dose regimen twice a day for three days and additional 0.25 mg/kg/d primaquine  for 14 days. The treatment dose was re-administered, if patients vomited during 30 minutes after administration. Patients repeatedly vomiting after their first dose of study medication were excluded from the study. The three-day artemether-lumefantrine treatment was directly supervised on site, while the intake of primaquine was not directly observed. The study participants were not checked for glucose-6-phosphate dehydrogenase (G6PD) deficiency prior to primaquine administration, as its prevalence in Guyana is assumed to be low; however accurate information on this deficiency in Guyana is scarce. When participants presented symptoms of haemolytic anaemia  during the follow-up visits, the primaquine treatment had to be stopped and they were excluded from the study.All study participants were treated with the treatment regimen used for Treatment outcomes were classified on the basis of the parasitological and clinical outcome according to the latest WHO guidelines , which cClinical examination, including measurement of the axillary temperature and Giemsa staining of thick and thin blood films, was carried out on days 0, 1, 2, 3, 7, 14, 21, and 28. Thick and thin blood smears were prepared from finger sticks. After Giemsa staining, each slide was examined microscopically independently by two experienced technicians at the collection site and again at the Malaria Research Unit, University Lyon 1. The parasitaemia was recorded as number of asexual parasites counted per 200 white blood cells present in the thick smear. The thick-film was considered to be negative, if no parasite had been found in 100 high-power fields. On days 0, 7, 14, 21 and 28 blood was blotted onto filter paper during follow-up and stored for DNA analysis.P. vivax cases.DNA was extracted from blood spots on filter paper with resin-based Instagene Matrix , as described before . Genus-spvmdr1 (Y976F) was performed on the eight P. vivax cases using a LightCycler system (Roche) and fluorescence resonance energy transfer technology. Primers and probes were designed and synthesized by TIB Molbiol . The polymerase chain reaction mixture and assay conditions were used as described in detail elsewhere [Detection of SNPs in sewhere ,26,27.The study protocol was reviewed and approved by the Ethics Committee of the Ministry of Health of Guyana. Informed written consent was provided by all patients or their parents/guardians before inclusion in the study.A total of 561 patients were screened for possible enrolment into the study and 74 (13.2%) patients met the inclusion criteria. The Georgetown hospital contributed 87.8% (n=65) of patients, 12.2% (n=9) were recruited at the Port Kaituma Hospital. Among the patients included, 90.5% were male, 9.5% female, with the majority of all participants (86.5%) being between 15\u201349 years old.The different ethnicities of Guyana were representatively distributed with 47.3% from mixed ethnicities, 24.3% East Indians, 16.2% Amerindians, 10.8% Afroguyanes and 1.4% of other groups. In total, 98% of the study population originated from the three highest malaria endemic regions 1, 7 and 8. The patients presented with the following symptoms in the preceding 24h: (i) Fever 20.3%, (ii) vomiting 1.4%, (iii) diarrhoea 2.8%, (iv) problems with coordination 1.4%, (v) dizziness or fainting 4.1%. However the fever incidence will be underestimated due to frequent self-medication with antipyretic drugs. The mean parasitaemia counts were 3,920 parasites/\u03bcl ranging from a maximum of 29,100 parasites/\u03bcl to a minimum of 30 parasites/\u03bcl  by day 14, decreasing to 59/61 (97%) by day 28. All patients tolerated the treatment well with very few side effects and no serious adverse events observed. No participants had to be excluded for haemolytic anaemia after primaquine administration.P. vivax mono-infection was PCR-confirmed. The others were microscopically confirmed by three independent readers. DNA from the parasites from the eight confirmed cases underwent screening for pvmdr1 single-nucleotide polymorphism at codon Y976F. No mutations were found.In a subset of eight patients, the P. vivax. Chloroquine and primaquine have been used worldwide against P. vivax as first-line drugs since 1946 and 1950 respectively. Reports from South-American countries such as Colombia and Brazil [P. vivax and a change of drug policies from chloroquine to ACT for vivax infections in four countries ) [P. vivax and even less studies looked at artemether-lumefantrine and primaquine.The study assessed artemether-lumefantrine + primaquine as a treatment option against  Brazil ,21 on deP. vivax infections. Drug interactions between the two drugs seem to be unlikely [All patients in the study cleared parasites on day 1, which makes artemether-lumefantrine an efficacious alternative for chloroquine in the treatment of vivax malaria. No severe side effects or adverse events were observed, similar to other studies, which show a good safety profile for artemether-lumefantrine alone -31 and fnlikely .P. vivax was PCR-confirmed at day 28, with negative PCRs on day 7, 14 and 21. It can be assumed that the new parasitaemia derived from relapse, rather than recrudescence, although a new infection or a recrudescence cannot be ruled out completely since there is no evidence that genotyping of P. vivax parasites may help to distinguish these events [P. vivax relapse, but not prevent it. Lumefantrine on the other hand is less likely to induce resistance compared to long half-life drugs as chloroquine or piperaquine [Two patients (2.7%) presented with parasitological failure at day 28. In one of those patients  events . Frequenraquine . In a siraquine , which cP. vivax infections resembling the course of malign P. falciparum infections [With recent reports clearly demonstrating severe ections ,37 the apvmdr1 single-nucleotide polymorphism and no mutations in codon Y976F were found. Low parasitaemia and sub-optimal sample storage conditions might partly explain the poor usefulness of the PCR method with these samples, while it usually works well with many other blood spot samples. No conclusions can be drawn on any resistance associated with pvmdr1 polymorphism. One out of the two patients with treatment failure was tested for Y976F mutation, but was wild type. In total, 90% of the study participants were male. In Guyana, malaria affects many more males than females at a ratio of almost 4:1 . This reflects the fact, that the migrant populations, primarily miners, are by far the most affected group. In addition to that all women aged 12 to 26 were excluded, in order to avoid potential pregnant study participants. 60% of all reported malaria cases occur in the Amerindian population, to which 16.2% of the study participants belong [A drawback to this study is that only eight patients could be screened for  belong .P. vivax infections would establish a common treatment for both circulating Plasmodium species, which are P. falciparum and P. vivax. Artemether-lumefantrine is highly effective against P. falciparum in Guyana, based on a study conducted in 2007/2008 . A common treatment could result in a number of advantages. Reports from Thailand and Indonesia detected up to 23% of P. falciparum infections being wrongly diagnosed as P. vivax, leading to inappropriate treatment with chloroquine [P. vivax more quickly than chloroquine [P. falciparum with artemether-lumefantrine + primaquine would have the advantage of also treating potential dormant P. vivax hypnozoites, which are thought to become activated due to P. falciparum infections, thereby explaining the high number of vivax infections after P. falciparum treatment [A switch of drug policies in Guyana to artemether-lumefantrine + primaquine for roquine ,40. ACT roquine . Treatineatment .P. falciparum in the private sector. Disadvantages of a common treatment will be the higher price for ACT as well as a decreased impetus for laboratories and hospitals in performing a correct plasmodium species diagnostic. The prescription of primaquine to all malaria cases without prior testing for G6PD deficiency presents a potential risk for severe haemolytic anaemia in G6PD-deficient patients, although G6PD deficiency is an infrequent trait in Guyana.Keeping chloroquine in the market, could lead to its administration against P. vivax infections. The treatment efficacy for chloroquine for P. vivax must be under close surveillance in Guyana. Should resistance against P. vivax increase, the tested regimen with artemether-lumefantrine + primaquine will be a more than equal alternative and drug policies should be switched.This study shows excellent efficacy for artemether-lumefantrine in combination with primaquine against The authors declare that they have no competing interests.DE contributed to molecular genetic studies, interpretation of results and the drafting of the manuscript. NC organized the collection of samples in Guyana and helped with the interpretation of results. KK and KC contributed to the study design and the interpretation of results. GB generated a portion of the molecular data. ALB contributed to the interpretation of the data and writing of the manuscript. SP contributed to the study design, interpretation of results and the writing of the manuscript. All authors read and approved the final manuscript."}
+{"text": "Since 2005, Burkina Faso adopted artesunate plus amodiaquine (ASAQ) and artemether-lumefantrine (AL) as first-line treatment for uncomplicated malaria. Despite improvement in that treatment, malaria remains the first cause of morbidity and mortality in the country.falciparum malaria in Burkina Faso five years after their adoption.This study aimed to analyze the therapeutic efficacies of ASAQ and AL for the treatment of uncomplicated P. falciparum malaria, treated with the recommended regimen of AL or ASAQ, were analyzed according to WWARN analytical methods. Patients benefited from a clinical and biological 28-day follow-up and performed on days 2, 3, 7, 14 and 28 to evaluate clinical and parasitological outcomes. Treatment failures have been corrected by PCR.Per-protocol individual data from four randomized clinical trials supported by IRSS-DRO Bobo Dioulasso in 2006, 2008, 2009 and 2010, including 1076 patients with uncomplicated Using WWARN analytical methods, the unadjusted Kaplan-Meier survival estimates are 76.4% (95% CI (72.5-79.8)) in the AL group (N=544) and 87.1% (95% CI (83.9-89.7)) in the ASAQ group (N=532). After PCR correction, AL was less efficacious than ASAQ respectively 95.8% (95% CI (93.6-97.3)) vs 98.2% (95% CI (96.6-99.1)); OR=0,486 (95% CI (0.217-1.089). There was no significant correlation between the occurrence of recrudescent at day 28 end-point and study year in two groups (coefficient<0).AL and ASAQ remain effective as treatment for uncomplicated malaria according to WHO recommendations, though AL was inferior in preventing recrudescent for 28-day follow-up.None declared."}
+{"text": "Plasmodium falciparum population dynamics during the early phase of anti-malarial drug treatment with artemisinin-based combination therapy in children with clinical malaria in a high transmission area in Africa.This study aimed to explore P. falciparum malaria in Bagamoyo District, Tanzania, were enrolled. Participants were hospitalized and received supervised standard treatment with artemether-lumefantrine according to body weight in six doses over 3 days. Blood samples were collected 11 times, i.e. at time of diagnosis (-2 h) and 0, 2, 4, 8, 16, 24, 36, 48, 60 and 72 h after initiation of treatment. Parasite population dynamics were assessed using nested polymerase chain reaction (PCR)-genotyping of merozoite surface protein (msp) 1 and 2.A total of 50 children aged 1-10 years with acute uncomplicated msp1 and msp2, respectively, non-detectable in the pre-treatment samples (-2 and 0 h combined). Some 15/20 (75%) and 14/21 (67%) of these genotypes were identified within 24 h, whereas 17/20 (85%) and 19/21 (90%) within 48 h for msp1 and msp2, respectively. The genotype profile was diverse, and varied considerably over time both within and between patients, molecular markers and their respective families.PCR-analyses from nine sequential blood samples collected after initiation of treatment identified 20 and 21 additional genotypes in 15/50 (30%) and 14/50 (28%) children with PCR analyses from multiple blood samples collected during the early treatment phase revealed a complex picture of parasite sub-populations. This underlines the importance of interpreting PCR-outcomes with caution and suggests that the present use of PCR-adjustment from paired blood samples in anti-malarial drug trials may overestimate assessment of drug efficacy in high transmission areas in Africa.http://www.clinicaltrials.gov with identifier NCT00336375.The study is registered at  Plasmodium falciparum in a stepwise manner from one blood sample collected at time of enrolment (pre-treatment sample) with another blood sample collected at time of parasite recurrence during follow-up after treatment, i.e. paired blood samples  and msp2 , respectively. A total of 16/50 (32%) and 12/50 (24%) patients had different genotype patterns detected in the two pre-treatment samples for msp1 and msp2, respectively. This comprised 19 genotypes in msp1 and 18 in msp2. A total of eight msp1 genotypes  and eight msp2 (3 FC27 and 5 IC) were detected -2 h but absent 0 h, whereas 11  and 10 (3 FC27 and 7 IC) genotypes, respectively, appeared at 0 h without having been identified -2 h.The complexity of infection pre-treatment (-2 and 0 h combined) was high, with 32/50 (64%) and 37/50 (74%) patients carrying multiple genotypes in msp1 genotyping identified a total of 20  genotypes in 15/50 (30%) patients, which were non-detectable pre-treatment (-2 and 0 h combined). Of these 15 patients, 11 had one, 3 had two and 1 had three new genotypes, respectively. Similarly, msp2 genotyping after initiation of treatment identified 21 (6 FC27 and 15 IC) new genotypes in 14/50 (28%) children, of whom seven children each had one and two new genotypes, as compared with the combined pre-treatment outcome. Including these 20 and 21 new genotypes as part of the overall complexity of infection the proportion of children harbouring multiple genotypes increased from 32/50 (64%) and 37/50 (74%) to 36/50 (72%) and 39/50 (78%) for msp1 and msp2, respectively, without affecting the median number of genotypes detected in either of the genetic markers.In the nine sequential blood samples collected after initiation of treatment, msp1 and msp2 genotypes exclusively identified after initiation of treatment. Of these 20 and 21 genotypes, 15 (75%) and 14 (67%) were identified within 24 h, whereas 17 (85%) and 19 (90%) were detected within 48 h for msp1 and msp2, respectively.Figure msp1) and msp2). The genotype profile fluctuated considerably over time, both within and between patients, molecular markers and their respective families making it difficult to categorize or quantify specific genotype patterns. However, some general genotype patterns were identified:PCR outcome for each participant during the entire study are presented in Figures a) Continous and fading; one or several genotypes present pre-treatment, persist/s for some time and finally fade/disappear.b) Continuous and fading with appearance of random genotypes; this pattern is similar to pattern a (above), but with additional random band/s appearing at any time-point during follow-up.c) Intermittent; one or several genotypes present pre-treatment disappear and reappear over time.d) Random; no clear pattern can be discerned.Example of genotype patterns a, b and c (above) is presented in Figure per se, but on a chain of different events preceding the DNA amplification and separation of PCR products, from blood sampling in the field and duration of storage to extraction of parasite DNA, each being associated with potential risk of influencing the final PCR outcome and thus the interpretation of the efficacy of the drugs at test [The introduction of PCR genotyping in clinical drug trials of anti-malarial drugs has significantly improved the distinction between treatment failure (recrudescence) and reinfections among recurrent infections post-treatment. The PCR technique is in theory simple and straight-forward. However, in real life situations the technique is well known to have limitations, particularly to amplify minority clones in complex infections, which is a common feature in children residing in high-transmission areas in Africa, but also due to constraints imposed by the biology of the parasite, with sequestration in the deep vascular system during the later part of the erythrocytic life cycle . However at test ,18. Ther at test .msp1 and msp2 during the early phase of anti-malarial drug treatment in children with acute uncomplicated P. falciparum infections in a high-transmission area in Tanzania, were explored in detail. The results indicate that a single blood sample may not provide a complete picture of all parasite subpopulations present in children with acute uncomplicated malaria in high-transmission areas in Africa. This is in agreement with our previous observations [et al. [In this study, parasite population dynamics, as assessed with PCR genotyping of rvations . However [et al. .et al. previous report [Similarly with M\u00e5rtensson s report , a majors report .The child (ID 2) with the most complex genotype pattern in our trial, presented both with a low grade parasitaemia and fever. Despite fulfilling the study inclusion criteria, this child may have suffered from a self-limiting cause of fever other than malaria. If so, the genotype pattern observed may represent an asymptomatic infection rather than parasite population dynamics in acute uncomplicated malaria.et al. found that individual measurements of the same PCR fragments run in different lanes on the same agarose gel varied from 2 to 16 basepairs [It should be underlined that parasite population dynamics were assessed based on size polymorphisms of PCR fragments after separation with electrophoresis on agarose gels. This method still remains one of the recommended methods in anti-malarial trials to distinguish treatment failures (recrudescence) from new infections  and is sasepairs . To accoThe results from this study indicate that PCR analyses from multiple blood samples collected during the early treatment phase revealed a complex picture of parasite sub-populations. However, it remains open whether the complex genotype pattern observed in our study reflects true parasite population dynamics or limitations of the PCR-technique to detect all minority genotypes present due to e.g. template competition in complex infections. The results underline the importance of interpreting PCR outcomes with caution and suggest that the present use of PCR-adjustment from paired single day blood samples in anti-malarial drug trials may overestimate assessment of drug efficacy in high transmission areas in Africa.We declare that we have no conflict of interest.AMC performed PCR-genotyping, data analyses and interpretation and assisted in writing of the report. BEN, SD and CM, planned and conducted the field trial. LR participated in the interpretation of data and in writing of the report. SA and ZP participated in study planning, coordination the field trial and assisted in writing of the report. JPG, MIV and AB contributed to study design, planning and coordination, interpretation of results, and preparation of the report. AM designed and planned the study, conducted the field trial, performed data analyses and interpreted the results, and wrote the report. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria in Togo, between 2005 and 2009.Malaria remains a major public health problem in Togo. The national malaria control programme in Togo changed the anti-malarial treatment policy from monotherapy to artemisinin combination therapy in 2004. This study reports the results of therapeutic efficacy studies conducted on artemether-lumefantrine and artesunate-amodiaquine for the treatment of uncomplicated P. falciparum, were treated with either artemether-lumefantrine or artesunate-amodiaquine. The primary end-point was the 28-day cure rate, PCR-corrected for reinfection and recrudescence. Studies were conducted according to the standardized WHO protocol for the assessment of the efficacy of anti-malarial treatment. Differences between categorical data were compared using the chi-square test or the Fisher\u2019s exact test where cell counts were \u2264 5. Differences in continuous data were compared using a t-test.Children between 6 and 59 months of age, who were symptomatically infected with A total of 16 studies were conducted in five sentinel sites, with 459, 505 and 332 children included in 2005, 2007 and 2009, respectively. The PCR-corrected 28-day cure rates using the per-protocol analysis were between 96%-100% for artemether-lumefantrine and 94%-100% for artesunate-amodiaquine.Both formulations of artemisinin-based combination therapy were effective over time and no severe adverse events related to the treatment were reported during the studies. Plasmodium falciparum resistance to anti-malarial medicines, the Togo National Malaria Control Programme (NMCP) set up a routine surveillance system, with five sentinel sites covering the distinct epidemiological zones of the country. Between 2001 and 2003, treatment failures with chloroquine and sulphadoxine\u2212pyrimethamine reached 63% and 26%, respectively[Malaria remains a major public health problem in Togo, an area of high transmission, with prevalence highest during the rainy season. Children carry the highest burden: in 2009, 56% of all reported cases and 73% of all reported deaths occurred among children under five years of age. In 2001The studies were conducted in the following sites: 1) Agbalp\u00e9dogan J\u00e9rusalem Medical Centre and Adakpam\u00e9 District Hospital in the capital city of Lom\u00e9; 2) La Providence Medical Centre in Kouv\u00e9; 3) Sodok\u00e9 and Kpangalam \u201cBon Secours\u201d Medical Centres located in Sodok\u00e9; 4) Doufelgou District Hospital in Niamtougou; and 5) Tantigou \u201cYendoub\u00e9\u201d Paediatric Hospital in Dapaong. The last four sentinel sites extend north from Lom\u00e9, in the order listed above, by 92 km, 350 km, 425 km and 620 km, respectively. Studies were conducted during the high transmission season for malaria, between October and December, except for Lom\u00e9, where the study was conducted from August to November.P. falciparum mono-infection with parasite density between 2,000 and 200,000 asexual parasites/mm3. Exclusion criteria included: one or more signs of severe or complicated malaria, mixed infection or infection with another species, malnutrition, concomitant disease, chronic or severe diseases, hypersensitivity or contra-indication to the study drugs and absence of informed consent of the parents. Clinical examination, including measurement of axillary temperature and blood smear for parasite counts, was performed at enrolment and on day 1, 2, 3, 7, 14, 21 and 28. Parasite counts were determined on Giemsa-stained thick films and recorded as the number of parasites per 200 white blood cells at admission and per 1,000 white cells on follow-up days based on a putative count of 6,000 white blood cells per microlitre of blood. The presence of gametocytes was also recorded in the 2007 and 2009 trials. Changes to haemoglobin levels after ACT treatment in 2007 and 2009 were measured using Hemocue haemoglobinometer. Capillary blood was sampled for haemoglobin at day 0, day 14 and day 28. All studies were approved by the Bioethics committee of the Ministry of Health and WHO Ethical Research Committee in 2007. Sample size was calculated according to WHO recommendations[The studies were based on the standardized World Health Organization (WHO) protocol for the assessment of the efficacy of anti-malarial treatment. PatientArtemether-lumefantrine  tablets containing 20 mg of artemether and 120 mg lumefantrine were administered every 12 hours over 3 days. Treatment was given without co-administration of fatty food. Weight-based dosing was applied, with one tablet for children weighing 5-14 kg and two tablets for children weighing 15-24 kg. Artesunate and amodiaquine were administered at an average dose of 4 mg/kg/day and 10 mg/kg/day over 3 days, respectively. Two different presentations of artesunate-amodiaquine were used: in 2005 and 2007, artesunate was purchased from Sanofi Synth\u00e9labo (France), and amodiaquine was supplied by Hoechst Marion Roussel (France). A co-blister of artesunate-amodiaquine manufactured by sanofi aventis (France) was used in 2009. The medicines were administered under supervision and allocated randomly. The purpose of the studies was not to compare the efficacy of the two combinations, but to monitor their efficacy independently.msp1 and msp2) were used. In 2007 and 2009, all three molecular markers  were used. Parasite DNA was extracted from blood spots collected on day 0 and on the day of reappearance of asexual parasitaemia[Treatment outcomes were classified based on an assessment of parasitological and clinical outcomes, according to the methods recommended by WHO in 2003. Unlike sitaemia. Patientt-test. Exact 95% confidence intervals were calculated for the treatment failure rates.All data were entered twice in the WHO Microsoft\u00ae Office Excel spreadsheet. Analysis was conducted with Stata/IC 11.0 . Differences between categorical data were compared using the chi-square test or the Fisher\u2019s exact test where cell counts were \u2264 5. Differences in continuous data were compared using a Patients treated with artesunate-amodiaquine (n\u2009=\u2009651) had a sex ratio of 1.1, a mean age of 2.7 years (standard deviation (SD)\u2009=\u20091.3 years), a mean weight of 12.2 kg (SD\u2009=\u20093.1 kg), and a mean temperature of 38.7 \u00b0C (SD\u2009=\u20091.0 \u00b0C). The geometric mean parasitaemia on day 0 was 23,494/\u03bcl . Patients treated with artemether-lumefantrine (n\u2009=\u2009645) had a sex ratio of 1.3, a mean age of 2.9 years (SD\u2009=\u20091.3 years), a mean weight of 12.5 kg (SD\u2009=\u20093.1 kg), a mean temperature of 38.7 \u00b0C (SD\u2009=\u20091.0 \u00b0C). The geometric mean parasitaemia on day 0 was 21,183/\u03bcl .Differences in patient characteristics on admission were investigated among studies conducted at the same site and on the same treatment between 2005 and 2007 (Niamtougou), and 2005 and 2009 (Dapaong and Kouv\u00e9) Table. StudiesThe overall PCR-corrected treatment failure rates remained low: between 0-4.4% for artemether-lumefantrine, and 0-6% for artesunate-amodiaquine Table. Both trNon-PCR-corrected treatment failure rates among studies conducted in 2005, 2007 and 2009 were 8.4%, 13.6% and 11.5%, respectively. However, the proportion of non-PCR-corrected treatment failures subsequently PCR-corrected as recrudescence decreased from 23.7% in 2005, to 4.6% in 2007 and 2.6% in 2009. There was a corresponding increase in the proportion classified as reinfections by PCR, from 55.3% in 2005, to 80% in 2007 and 84.2% in 2009 (P\u2009=\u20090.004). This increase was observed in all three sites where studies were conducted twice within the four-year period, but the increase was most pronounced in Kouv\u00e9, with 28.6% in 2005 and 71.4% in 2009 (P\u2009=\u20090.04), and in Niamtougou, where the proportion of non-PCR-corrected treatment failures subsequently PCR-corrected as reinfections increased from 24.4% in 2005 to 75.6% in 2007 (P\u2009=\u20090.001). The increase was consistent in both treatment groups. No severe adverse events related to the ACT were reported during the 16 studies.Haemoglobin levels for subjects in both treatment groups increased progressively from day 0 to day 14 and to day 28 Table. SignifiWhile there was some variation in the proportion of patients with gametocytes on day 0, no differences were found among sites or treatment regimens at other time points Table. There wThese studies showed the high therapeutic efficacy of artemether-lumefantrine and artesunate-amodiaquine in Togo between 2005 and 2009. The studies also demonstrated the significant improvement of haemoglobin levels following treatment. In addition, the artemisinin\u2019s gametocytocidal effect was observed by the initial, rapid reduction in gametocytes, and the failure of any new gametocytes to appear over the 28-day period.In Togo, the efficacy of artemether-lumefantrine was high and did not change between 2005 and 2009, despite the absence of co-administration of fat. This is consistent with a review of studies from sub-Saharan Africa, where it was found that the fat content of standard meals or breast milk was adequate for absorption of lumefantrine. In 2009Similarly, the efficacy of artesunate-amodiaquine was high in Togo, and did not change between 2005 and 2009. Two different presentations were used in these studies: loose tablets were used in 2005 and 2007, and co-blistered treatment was used in 2009. Despite different presentations, the treatment outcome did not vary significantly over time. In 2009, artesunate-amodiaquine was the first-line treatment for uncomplicated malaria in 22 African countries. Howeverglurp, which improved the ability to distinguish between reinfection and recrudescence. In 2005, only msp1 and msp 2 were used. When glurp was added in 2007 and 2009, the molecular marker detected 53% and 65% of the reinfections, respectively, demonstrating its high ability to discriminate between a reinfection and a recrudescence. Failure to use all three molecular markers in PCR analyses may result in incorrect conclusions regarding the true efficacy of the ACT. These findings demonstrate the importance of following a standardized protocol to enable the comparison of therapeutic efficacy results across sites and over time.The studies presented here showed an increase in the proportion of reinfections detected in 2007 and 2009 when compared to 2005. The increase in the proportion of reinfections was likely caused by the addition of a third molecular marker, There are very few studies on therapeutic efficacy and drug resistance in Togo. In a literature review, only one publication on the therapeutic efficacy of ACT in Togo was found. Among 8Artemether-lumefantrine and artesunate-amodiaquine are both first-line medicines for the treatment of uncomplicated malaria in Togo, and both have shown high efficacy. WHO only recommends a change in treatment policy if the PCR-corrected treatment failure is higher than 10%, which is not the case in Togo. Nevertheless, routine monitoring with adherence to standardized protocols should be continued in order to detect artemisinin resistance if it emerges, and to monitor changes to the therapeutic effect of the partner drug in an ACT.ETF: Early treatment failure; ACPR: Adequate clinical and parasitological response; ACT: Artemisinin-based combination therapy; LCF: Late clinical failure; LPF: Late parasitological failure; NMCP: National Malaria Control Programme; WHO: World Health Organization.The authors declare that they have no competing interests.MD conceived and designed the study, conducted the research, collected and interpreted the data. YA conducted the research. AB performed the statistical analysis and drafted the manuscript. SK collected the data and conducted the research. HB conducted the research and performed the PCR analysis. YS conceived and designed the study, performed statistical analysis and interpretation. KM conceived and designed the study and conducted the research. All authors read and approved the final manuscript."}
+{"text": "The National Malaria Control Programme in Senegal, introduced since 2006, artemisinin-based combination therapy (ACT administration) for the treatment of uncomplicated malaria cases. In this framework, an anti-malarial pharmacovigilance plan was developed and implemented in all public health services. This study investigated the occurrence of Adverse Drug Events (ADEs) after ACT.The study was conducted between January 2007 and December 2009. It was based on spontaneous reports of ADEs in public health facilities. Data on patient demographic characteristics, dispensing facility, adverse signs and symptoms and causality were collected from a total of 123 patients.The age range of these patients was six months to 93 years with a mean of 25.9 years. Of the reported symptoms, 46.7% were related to the abdomen and the digestive system. Symptoms related to the nervous system, skin and subcutaneous tissue, circulatory and respiratory systems and general symptoms and signs were 7%, 9.7%, 3.5% and 31.3%, respectively. Causality results linked 14.3% of symptoms to Falcimon\u00ae (Artesunate-Amodiaquine) with certainty. Effects were classified as mild and severe in 69.1% and 7.3% of cases respectively while 23.6% were serious. All patients with serious ADEs were hospitalized. One death was reported in a patient who had taken 24 pills at once.These results confirm the need to develop and implement pharmacovigilance systems in malaria endemic countries in order to monitor the safety of anti-malarial treatments. Early diagnosis and prompt treatment with an effective anti-malarial drug remains a priority for the National Malaria Control Programme (NMCP) in Senegal. This strategy was threatened by the spread of chloroquine resistance and the NMCP adopted artemisinin-based combination therapy (ACT) in line with WHO treatment guidelines. In Senegal, two combinations are recommended as first-line treatment for uncomplicated malaria cases: artesunate-amodiaquine (AS-AQ) and artemether-lumefantrine (AL). In 2006, with the financial support of the Global Fund against AIDS, Tuberculosis and Malaria, the AS-AQ combination was scaled up in the Senegalese health care system .ACT is based on a combination of drugs some of which have been recently marketed. Safety profile data for their long-term use in the general population in sub-Saharan Africa are scarce. Several studies have reported safe use of ACT, particularly the AS-AQ combination , but theIn Senegal, a national pharmacovigilance system based at the National Centre of Pharmacovigilance (\u201cCentre Antipoison\u201d) was created in 1998, but it was not scaled up and therefore the reporting was very poor until recently. Since 2009 Senegal has been a member of the WHO Collaborative Programme for International Drug Monitoring, based at the Uppsala Monitoring Centre, Sweden. Because pharmacovigilance should be an essential component for all public health programmes that use medicines, there was need to develop a pharmacovigilance system for antimalarials and in particular for ACT as part of the introduction plan of these new drugs. This brought together the competencies of the Ministry of health, the National Centre of Pharmacovigilance, academic and private sectors and partners such as WHO, UNICEF, USAID and civil society organisations under the coordination of the NMCP to develop and implement a dedicated safety monitoring system of antimalarial drugs in all the public health services in Senegal in 2007.This paper reports Adverse Drug Events (ADEs) collected in health facilities after anti-malarial treatment between January 2007 and December 2009.Senegal is a West African country, with a population of approximately 12,141,673 inhabitants (2010). Malaria is endemic with a high transmission season between July and November. The entire country\u2019s population is at risk of malaria.Malaria case management is integrated into primary health services. Health services are provided nationwide through 75 health districts, which cover 78 district hospitals and 1,018 health posts. Implementation of the pharmacovigilance system involves all public health facilities. Between April 2006 and June 2009, artesunate\u2009+\u2009amodiaquine (AS\u2009+\u2009AQ) in a co-blister was used to treat uncomplicated malaria. In July 2009, co-blisters were replaced by fixed dose artesunate-amodiaquine (AS-AQ fixed dose). Drugs cost 0.6 US$ per course for adults and 0.3 US$ per course for children during the period covered by the study.The treatment was administered once a day for three days using the following drugs:Between April 2006 and June 2009: AS\u2009+\u2009AQ in co-blistered or Falcimon\u00ae . It contains 50 mg of AS per tablet and 153 mg AQ base per tablet. The recommended doses were 4 mg/kg/day for AS and 10 mg/kg/day for AQ. The product was available in three formulations: three AS tablets and three AQ tablets for children 0\u20136 years, six AS tablets and six AQ tablets for children 6\u201313 years and 12 AS tablets and 12 AQ tablets for adults\u2009\u2265\u200914 years.From July to December 2009: AS-AQ fixed-dose co-formulation or ASAQ Winthrop\u00ae (Sanofi-Aventis). There were four different formulations: AS 25 mg and AQ base 67,5 mg tablets for children 2\u201311 months; AS 50 mg and AQ base 135 mg tablets for children 1\u20135 years; and AS 100 mg and AQ base 270 mg tablets for children 6\u201313 years (three tablets) and for adults\u2009\u2265\u200914 years (six tablets).This study was conducted countrywide and patients have been reported by public health facilities located in different regions in Senegal. All patients who received partial or full treatment of ACT and who presented ADEs which had been reported by health providers, were enrolled in the study.This study is a retrospective analysis of ADEs cases related to ACT as self-reported by patients at health facilities during a period of three years.Data were collected by health workers using the standard ADEs reporting form developed by the Ministry of Health. Collected data were:(i) the patient , (ii) all the drugs used by the patients , (iii) the adverse reactions presented by the patient , (iv) and the reporter .When an ADE was diagnosed, the health worker filled the standard form and sent it out to the NMCP focal point for case management and pharmacovigilance through the District Health Officer. All forms were checked and validated, including telephoning the reporter if necessary. In all cases a letter was sent to the reporter to acknowledge receipt of the form.Centre Antipoison) for causality analysis. Afterwards all reported cases and causality results were recorded in the NMCP pharmacovigilance database using Microsoft\u00ae Excel programme.After validation the forms were sent to the National Centre of Pharmacovigilance  Certain: clinical event occurring in a plausible time relationship to drug administration, which cannot be explained by concurrent disease or other drugs of chemicals. The response to withdrawal of the drugs  should be clinically plausible. Finally, the event must be definitive pharmacologically or phenomenologically, using a satisfactory rechallenge procedure if necessary. (2) Probably/Likely: clinical event with a reasonable time sequence to administration of the drug; unlikely to be attributed to concurrent disease or other drugs or chemicals, which follows a clinically reasonable response on withdrawal . Rechallenge information is not required to fulfil this definition. (3) Possible: clinical event, with a reasonable time sequence to administrations of the drug, but which could also be explained by concurrent disease or other drugs or chemicals. Information on drug withdrawal may be lacking or unclear. (4) Unlikely: Clinical event with a temporal relationship to drug administration which makes a causal relationship improbable, and in which other drugs, chemicals or underlying disease provide plausible explanations. (5) Conditional/Unclassified: Clinical event, reported as an adverse reaction, about which more data is essential for a proper assessment, or the additional data is under examination. (6) Unassessable/Unclassifiable: a report suggesting an adverse reaction which cannot be judged because information is insufficient or contradictory, and which cannot be supplemented or verified.Data analysis was done using Stata 11. Age was treated as a categorical variable using strata. In addition age and appearance of ADEs were summarized as means. Other socio-demographic variables and safety indicators were presented as frequencies and percentages. Incidence of ADEs was calculated using the number of treatment distributed per year. Lastly signs and symptoms have been classified on the basis of the International Statistical Classification of Diseases and Related Health Problems, 10th Revision (ICD-10) .This intervention was administered by the Ministry of Health and included in the routine work of health workers according to its malaria policy. Institutional Review Board clearance was not required.Between 1st January 2007 and 31st December 2009, a total of 123 patients with ADEs related to ACT were reported. The mean age of patients was 25.9 years (range 6 months-93 years) with a F/M sex ratio of 1.12  after the beginning of therapy while the duration of ADEs varied from less than 24 hours to 32 days. The events were reported following the use of three different combination therapies: AS\u2009+\u2009AQ co-blister, AS-AQ fixed dose and artesunate-mefloquine (AS-MFQ). Eighty five percent of the notifications were related to the AS\u2009+\u2009AQ coblister, while 14% were related to AS-AQ fixed dose and one reported case was related to AS-MFQ.Gastrointestinal disorders represented 46.7% of symptoms and signs reported, followed by general symptoms and signs 31.3%). Disorders of the skin and cutaneous system, nervous system and circulatory and respiratory systems represented 9.7%, 7% and 3.5% of the effects respectively . All the serious ADEs were hospitalized and there was one death due to misuse of drug. The patient was a 42-year old man who had taken all the 24 tablets of AS-AQ at once together with paracetamol and ciprofloxacin tablets. On the same day he overdosed, he had presented with movement disorders, vertigo, dyspnoea and hiccough and was admitted in a rural district hospital before he died.Using the WHO Causality criteria, 14.3% of ADEs related to AS-AQ co-blister treatment were classified as Certain. The main symptoms were nausea and vomiting, abdominal pains, diarrhoea, extra pyramidal and movement disorders, pruritus, urticaria, dyspnoea and hiccough. 22.8% and 37.2% of ADEs related to this combination were classified as Probable and Possible respectively. Twenty-seven reports were classified as Unlikely, Conditional/Unclassified or Unassessable/Unclassifiable. None of the events linked to use of the AS-AQ fixed dose were classified as Certain. However 23.5% of cases were classified as Probable and 53% as Possible. The main effects reported were nausea and vomiting, pruritus, vertigo, fatigue, tinnitus and confusion. Four reports were classified as Unlikely or Unassessable/Unclassifiable. One ADE related to AS-MFQ was reported. The patient presented with pruritus which was classified as Probably linked to this drug.With the wide use of ACT in endemic areas, safety monitoring of anti-malarial drugs is critical. This study serves to reinforce previous reports advocating for increased pharmacovigilance on adverse drug events of artemisinin-based combination therapy, particularly in Africa -12. WhilMost of the ADEs were reported by nurses in peripheral health posts. Because routine spontaneous reporting relies on vigilance of health workers, all the ADEs may not be fully reported, this could lead to a low reporting rate. Even though data were limited and based on passive reporting, the causality classifications support a link between the adverse events and administration of ACT. Adverse drug events are different from adverse drug reactions. While an adverse event is any undesirable medical occurrence that develops after administration of a drug, regardless of the suspected relationship between the drug product and the event, to classify the event as an adverse drug reaction, a causal relationship must be established .One difficulty in this study was distinguishing between adverse events and common symptoms of malaria . Indeed The apparent incidence of ADEs is far less than the results of a clinical trial conducted in Senegal. That is undoubtedly a reflection of the nature of the reporting scheme, which relies on busy nurses and other health professionals to report adverse events they suspect are related to the anti-malarial medications. In addition, no biological effects have been reported because all the ADEs were collected from peripheral health facilities where there is no laboratory service . The freThis study\u2019s findings confirm the relative safety of use of ACT as reported by previous studies ,14,15. MAlthough endemic countries may have inadequate infrastructure and limited resources for implementing pharmacovigilance, drug safety monitoring mechanisms are imperative. This study demonstrates the feasibility and the need to set up a safety monitoring system for anti-malarial drugs within a national pharmacovigilance system. The latter should be scaled up in health facilities and hospitals and involve all health workers to improve the notification of adverse drug events of ACT. Even though the safety of ACT is established, post-marketing safety monitoring of ADEs is critical and should be developed and implemented in endemic areas. Furthermore because many endemic countries are scaling up malaria case management at community level, and given that people living in malaria areas can receive more than one course of treatment a year, the safety monitoring system of ACT should not only involve health facilities but also be extended at community level.The authors declare that they have no competing interests.ST coordinated this study and devised the study objectives and design. ST and JLN designed the methodology, did the analysis and wrote the manuscript. ID, FBF, MLD, MBD, MN carried out the planning, development of tools, training, implementation and monitoring during the study. NED assisted with data collection, data cleaning and analysis. BF assisted with training and supervision. PG assisted in writing the first draft and in revising the manuscript. OG and MT oversaw this study. All authors read and approved the manuscript."}
+{"text": "Though India has already switched to ACT for treating falciparum malaria, there is need to have multiple options of alternative forms of ACT. A randomized trial was conducted to assess the safety and efficacy of the fixed dose combination of artesunate-amodiaquine (ASAQ) and amodiaquine (AQ) for the treatment of uncomplicated falciparum malaria for the first time in India. The study sites are located in malaria-endemic, chloroquine-resistant areas.Artemisinin-based combination therapy (ACT) has been recommended for the treatment of Plasmodium falciparum mono-infection were randomly allocated to ASAQ and AQ arms. The primary endpoint was 28-day PCR-corrected parasitological cure rate.This was an open label, randomized trial conducted at two sites in India from January 2007 to January 2008. Patients between six months and 60 years of age having Three hundred patients were enrolled at two participating centres, Ranchi, Jharkhand and Rourkela, Odisha. Two patients in AQ arm had early treatment failure while there was no early treatment failure in ASAQ arm. Late treatment failures were seen in 13 and 12 patients in ASAQ and AQ arms, respectively. The PCR-corrected cure rates in intent-to-treat population were 97.51% (94.6-99.1%) in ASAQ and 88.65% (81.3-93.9%) in AQ arms. In per-protocol population, they were 97.47% (94.2-99.2%) and 88.30% (80-94%) in ASAQ and AQ arms respectively. Seven serious adverse events (SAEs) were reported in five patients, of which two were reported as related to the treatment. All SAEs resolved without sequel.P. falciparum malaria. Amodiaquine also showed acceptable efficacy, making it a suitable partner of artesunate. The combination could prove to be a viable option in case India opts for fixed dose combination ACT.The fixed dose combination of ASAQ was found to be efficacious and safe treatment for ISRCTN84408319 Plasmodium falciparum and many malaria endemic countries are using it. The combination of amodiaquine and artesunate, along with four more combinations, is recommended by WHO for malaria control programmes [The World Health Organization (WHO) has recommended artemisinin-based combination therapy (ACT) as the treatment for ogrammes .falciparum malaria [dhfr and dhps mutations [India has switched over to ACT and the National Vector Borne Disease Control Programme recommends the use of artesunate + sulphadoxine-pyrimethamine (AS + SP) for the treatment for  malaria . There iutations  and treautations . FurtherP. falciparum malaria conducted over the past 10 years in Africa showed that amodiaquine (AQ) proved significantly more effective than chloroquine in clearing parasites, with a tendency for faster clinical recovery. This difference was also observed in areas with considerable chloroquine resistance. Further, serious adverse events have not been reported with curative short-term regimen of AQ [The combination artesunate-amodiaquine (ASAQ) has been extensively studied and good efficacy and tolerability has been reported. A systematic review of relevant studies -8 on theen of AQ .falciparum malaria for the first time in India. The study sites were located in malaria-endemic, chloroquine-resistant areas.A randomized trial was conducted to assess the safety and efficacy of the fixed dose combination of ASAQ and AQ alone for treatment of uncomplicated P. falciparum cases of about 96%. Ranchi district has peak malaria season from August to November with relatively low proportion (50%) of falciparum cases. The efficacy of chloroquine was 54 to 57% in Sundargarh and 72% in Ranchi in an earlier study [falciparum malaria in both the study areas during this period.This was an open label, randomized study carried out at Mahadevi Birla Hospital, Ranchi, Jharkhand and Community Welfare Society Hospital, Rourkela, Odisha from January 2007 to January 2008 Figure . Rourkeler study . AS + SPP. falciparum mono-infection  and fever \u226537.5\u00b0C (99.5\u00b0F). Patients with signs of severe malaria, febrile conditions due to diseases other than malaria, known severe disease, history of hypersensitivity to drug(s), positive pregnancy test or lactating women, history of anti-malarial treatment in past 15 days were excluded. Patients were also excluded from the study if they had anaemia (haemoglobin < 7 g/dl), hepatic  or renal impairment (serum creatinine \u2265 1.2 ULN).The study was carried out in patients aged from six months to 60 years, weighing more than 5 kg and having Patients were randomized to one of the two arms in 2:1 ratio; fixed dose combination tablets of ASAQ and AQ tablets alone using a predetermined randomization list. Individual, opaque, sealed and sequentially numbered envelopes were used for randomization. ASAQ was given once daily orally on days 0, 1 and 2 of the study according to age group. Two different strengths of the combination were formulated; paediatric/lower strength (AS: 25 mg/AQ: 67.5 mg) and adult/higher strength (AS: 100 mg/AQ: 270 mg). Children in the six to 11 months age group were given one lower strength tablet, one to five years, two lower strength tablets, six to 13 years one higher strength tablet and those older than 14 years were given two higher strength tablets. AQ tablets (153 mg base) were administered orally, once daily for three days. The number of tablets was decided according to age and weight with the adult dose being four tablets on day 0, 1 and three tablets on day 2.Anti-malarial drug efficacy was assessed by using the WHO protocol with a 28-day follow up . ParticiThe primary endpoint of the study was the PCR -corrected cure rate based on the adequate clinical and parasitological response, which is defined as the absence of parasitaemia, irrespective of patient's body temperature, with the patient not meeting any criteria of early treatment failure or late clinical or parasitological failure as defined by the WHO .The secondary endpoints included the Parasite Reduction Ratio, Parasite Clearance Time, Fever Clearance Time, percent patients without gametocytes on day 28 and proportion of patients with early and late treatment failure and late parasitological failure.msp1 and msp2) and glutamate rich protein (glurp) were used [Blood samples were collected by fingerpick method for parasitological assessment at enrolment, during admission and follow up visits. Thick and thin smears were prepared, Giemsa-stained and examined for parasite density. This was done by counting the number of asexual parasites per 200 leucocytes in thick blood film. Parasite density per \u03bcl was calculated as /(number of leukocytes counted). Two qualified microscopists independently read all of the slides and parasite densities were calculated by averaging the two counts. Two to three drops of blood were collected on a chromatography filter paper no.3  from each patient at inclusion. A second specimen was collected only in the case of reappearance of parasites as evidenced by a positive microscopy slide. Genotyping was carried out at NIMR, Delhi. For differentiating recrudescence from re-infection, three genetic markers, merozoite surface proteins  and biochemical  assessments on day 0, 7, 28 and on the day of recurrent parasitaemia.This was not a head to head comparative study but rather a parallel study in which the AQ alone arm acted as a positive control. Hence, the sample size was estimated using the precision method with a two-sided alpha of 0.05 and a power of 0.8. On the assumption that ASAQ will have a cure rate of 95% and a precision of 5%, the estimated sample size was 73. If a cure rate of 90% was considered with precision of 5%, the sample size increased to 138. Hence, it was planned to recruit about 300 patients; 200 in ASAQ and 100 in AQ arm. Epi Info 6.04b software was used for these calculations.Three patient populations were evaluated. The safety population comprised all patients who were randomized and received at least one dose of study drug. The safety analysis and listing was done on safety population. The intent to treat (ITT) population comprised of all patients who were randomized, received at least one dose of study drug and underwent at least one efficacy assessment. The secondary efficacy analysis was performed on ITT population. Per protocol (PP) population comprised of all patients who completed the study as per the protocol. The primary and secondary efficacy analysis was performed on PP population. Patients who withdrew from the study or did not appear for the scheduled visits on or before day 28 were considered as drop-outs. These patients were part of ITT and safety population and hence their data were reported.The primary endpoint of day 28 cure rate was evaluated in both ITT and PP populations. Cure rates were based on simple proportions. The 95% confidence interval was calculated for both the treatment arms. The secondary efficacy parameter parasite reduction ratio at 48 hr and Parasite and Fever clearance times were analyzed in ITT population using Kaplan Meier analysis. Statistical analyses were done using the software SAS 9.1.3.The study was conducted in accordance with the local laws and regulations including the schedule Y, Indian Good Clinical Practices, Ethical guidelines on biomedical research issued by the Indian Council of Medical Research, International Conference on Harmonization - Good Clinical Practice (ICH-GCP). The protocol was reviewed and approved by the Ethics Committees of the National Institute of Malaria Research, Delhi and Community Welfare Society Hospital, Rourkela. Written informed consent was obtained from participants/guardians. In case of an illiterate patient, his/her thumb impression and signature of an independent witness was obtained.Three hundred and twenty seven patients were screened at the two sites of which, 300 were randomized to the two treatment arms. The ITT population comprised of 298 subjects, 201 in ASAQ and 97 in AQ arm  and 81.44% (72.3%-88.6%) in ASAQ and AQ arms respectively. In PP population, they were 92.42% (87.8-95.7%) in ASAQ arm and 82.98% (72.8-89.9%) in AQ arm (Table There were 27 treatment failures (13 in ASAQ and 14 in AQ arm). The PCR analysis showed that six subjects had new infections (five in ASAQ and one in AQ arm) and 16 had recrudescence (five in ASAQ and 11 in AQ arm). The mean age of failure patients was 21.8 and 13.2 years in ASAQ and AQ arms respectively. The mean initial parasitaemia was 49065 and 37862 respectively in ASAQ and AQ arms. Four samples were inconclusive (three in ASAQ and one in AQ arm), while result could not be obtained in one sample (AQ arm). The patients with indeterminate PCR results were classified as recrudescent infections whereas the new infections were classified as 'cured'.After PCR correction, the cure rates were 97.51% (94.6-99.1%) and 88.65% (81.3-93.9%) in ITT population. PCR corrected cure rates in PP population were 97.47% 94.2-99.2%) in ASAQ and 88.30% (95% CI 80.0-94.0%) in AQ arm . The mean parasite clearance time was 1.5 and 2.1 days in ASAQ and AQ arms respectively.The Kaplan Meier estimate of the proportion of aparasitaemic patients at 48 hours was 93% (187/201) in ASAQ arm and 65% (63/97) in AQ arm . The median Fever Clearance Time was same (one day) in both the treatment arms Figure .There was no early treatment failure in ASAQ arm while two (2.1%) patients had early treatment failure in AQ arm. The proportion of patients with late treatment failures was 6.5% (13/201) in ASAQ and 12.4% (12/97) in AQ arms. The proportion of patients with gametocytes at the end of follow-up (day 28) was 1.5% (03/201) in ASAQ and 2.1% (02/97) in AQ arms. The late clinical failures, which represented patients from day 4 to 28 with parasitaemia and fever were 4.0% (8/201) in ASAQ and 5.2% (5/97) in AQ arms. The late parasitological failures defined as presence of parasitaemia from day 7 to day 28 were 2.5% (05/201) in ASAQ and 7.2% (07/97) in AQ arms. None of these parameters was statistically significant in the two treatment arms (p > 0.05).There were in total 10 protocol deviations and five protocol violations in the study. Of the 10 protocol deviations, nine were related to the study procedures. One protocol violation was related to the study assessment when the patient was not available at the correct time point.The adverse event (AE) profiles for ASAQ and AQ treatments were similar in terms of type and frequency of events and were mostly those expected in malaria patients. More than half the patients (57.42% in ASAQ and 59.18% in AQ) experienced AE Table . The difSerious adverse events (SAEs) were reported in five patients, three (1.5%) in ASAQ and two (2%) in AQ arm. They were leukocytosis, diarrhoea, vomiting, hyperbilirubinaemia and hypersensitivity reaction. In the ASAQ arm, two SAEs were considered drug related by investigators. One was angioedema without signs of anaphylaxis and another, severe diarrhoea requiring hospitalization. The SAEs in the AQ arm were unrelated to the drug. All SAEs resolved without sequelae.The haematological and biochemical values on day 0 are shown in Table There were no significant changes in RBC and WBC counts over 28 days. There was a marginal decrease in the levels of total bilirubin, SGOT, SGPT and creatinine over this period.falciparum malaria in India. The PCR-corrected cure rates were 97.47% (95% CI 94.2 - 99.2%) and 88.3% (95% CI 80.0 - 94.0%) in ASAQ and AQ arms respectively. The cure rates were similar in different age groups. The study also supports the role of AQ as a suitable partner in ASAQ. The cure rates in PP population were similar to that in ITT population. Further, they were similar at both the sites. The combination has the advantage of once daily administration, thus increasing compliance. The results of this study are in line with the results obtained in Cameroon, Madagascar, Mali [The study shows that ASAQ fixed dose combination is efficacious for treatment of ar, Mali , Senegalar, Mali ,14, and ar, Mali . A recenar, Mali dhfr and dhps mutations especially in the north-east [Currently, AS + SP has been recommended by the National Vector Borne Disease Control Programme in India. The combination is efficacious  but trearth-east . Furtherrth-east . Even thrth-east  and alsorth-east . The comAccording to WHO, to qualify as ACT the combination should also have independent anti-malarial activity . The stuFever was the most common AE but was considered unrelated to the study medication and caused by malaria itself. In the related subgroup, nausea and vomiting were the more common AEs, and they may indicate the gastric irritation caused by the study medication.The drug has been registered for marketing in India on the basis of results of this study.falciparum malaria in both the study areas. The study also showed that the partner drug, AQ was effective in the study areas, making it a suitable partner of artesunate. The combination could prove to be one of the viable options in case India opts for fixed-dose combination ACT.Fixed dose combination ASAQ proved to be an efficacious and safe treatment for ACT: Artemisinin based combination therapy; AQ: Amodiaquine; ASAQ: Artesunate amodiaquine; AS + SP: Artesunate + sulphadoxine-pyrimethamine; ITT: Intent to treat; PCR: Polymerase chain reaction; PP: Per protocol; SAE: Serious adverse eventThe authors declare that they have no competing interests.NV, JK and BS conceived the idea. NV was the PI and study coordinator. NV and BS also supervised all the field sites. AD was involved in overall supervision. AA, BHS, TKB, PKT, SS were involved in field work. PS did the molecular analysis. BS2 was involved in quality assurance of microscopy. All authors read and approved the manuscript."}
+{"text": "New-generation antiepileptic drugs (AEDs) tend to replace traditional AEDs as the first-line choice for epilepsy. However, whether this change results in better outcome, especially in China, remains unknown.Two broad spectrum AEDs, the traditional drug of sustained-release formulation of valproate (SRVPA) and the new-generation drug of topiramate, were compared in patients with epilepsy as monotherapy in this multi-centre, observational cohort study from 2000 to 2011. The primary outcome was time to treatment failure. The secondary outcomes included time to first seizure, time to 12-month remission, and time to 24-month remission. Drug tolerability was assessed. Cox proportional hazard models  were used to analyse the relative risks expressed as hazard ratios (HR).Of the 1008 recruited patients, 519 received SRVPA and 489 received topiramate. SRVPA was better than topiramate  in primary outcome, and in time to first seizure . No significant difference was observed between two groups in time to 12-month remission  and time to 24-month remission . 36 patients (6.9%) in SRVPA group and 37 patients (7.6%) in topiramate group presented treatment failure associated with intolerable adverse events, there was no significant difference between the two groups (p\u200a=\u200a0.70).The SRVPA is more suitable than topiramate for Chinese epileptic patients, and our results support the viewpoint that traditional AEDs should be the first-line choice for epilepsy rather than new-generation AEDs. Epilepsy is one of the most common neurological disorders, affecting approximately 50 million people worldwide International League Against Epilepsy (ILEA) proposed that choice of optimal AED for epilepsy should be based on randomized controlled trials (RCTs). Although RCTs could provide less biased results In recent years, about ten new-generation AEDs have been registered in China, and the prescriptions of new drugs are rapidly increasing. Since previous studies have not provided a clear answer as to which generations of AED should be the first choice This study was undertaken in one chartered city and four provinces of China: Chongqing City, Guangdong Province, Sichuan Province, Zhejiang Province and Henan Province. Based on the data of the sixth national census of population from National Bureau of statistics of China, the sample area covered a total population of 362 017 960 people. The beginning date of this census was 1st November, 2010, and the reporting date was 29th April, 2011. Recruitment of this study occurred from August 2000 to March 2010. The last follow-up visit was between March and July 2011. This study was approved by the Ethics Committee of Chongqing Medical University. All patients or their guardians provided written informed consent.Patients with a definite diagnosis of epilepsy, treated with SRVPA or topiramate as monotherapy, between 2 to 75 years old, were enrolled in this study. The exclusion criteria included the following: epileptic syndromes; only acute symptomatic or non-epileptic seizures; a history of psychiatric or mood disorders; clinically significant laboratory abnormalities, including abnormal liver function, abnormal haematological system function, abnormal kidney function, abnormal endocrine system function, or heart disease; and clinician or the patient feeling that the treatment was contraindicated.The information recorded during the first visit included patient demographics, information about previous antiepileptic treatment, history of febrile seizures, birth traumas, epilepsy in first-degree family members, and neurological diseases . A general physical examination and a neurological examination were performed. Laboratory examinations were carried out. Electrocardiogram (ECG) examinations were also performed, if necessary. Surface electroencephalography was performed on each patient to detect significant changes that might contribute to diagnosis. Computed tomography, magnetic resonance imaging and additional examinations, such as thyroid hormones, autoantibodies, and rheoencephalography, were carried out if clinically needed. Clinicians were asked to classify the types of epilepsy, epileptic syndromes, and types of seizures according to the criteria of the ILAE For the patients with a definite diagnosis of epilepsy, physicians prescribed them the AED which they could afford. And only the patients treated by the SRVPA or topiramate were enrolled in this study. The guidelines for the initial drug dose and titration were provided as follows. In children and adolescents (ages 2\u201316), the initial dosage of SRVPA was 10\u201315 mg/kg/day, with weekly increments of 5\u201310 mg/kg/day, and the target dosage was 20\u201330 mg/kg/day; the initial dosage of topiramate was 0.5\u20131 mg/kg/day, with weekly increments of 0.5\u20131 mg/kg/day, and the target dosage was 5\u20139 mg/kg/day. In adults, the starting dosage of topiramate was 25 mg per night, with a weekly increment of 25 mg/day, and the target dosage was 100\u2013250 mg/day; the initial dosage of SRVPA was 500 mg/day, with a weekly increment of 250 mg/day, and the target dosage was 1000\u20132000 mg/day. In general, medications were given with small initial doses, and the doses were slowly increased until the seizures were under control. Efficacy and adverse events were balanced in the adjustment of the AED dosages.The patients were asked to return for subsequent reviews at the second week, the first month, the third month, the sixth month and at successive half-year intervals from the date of initial medication. If clinical attention was necessary, more visits were scheduled between the regularly scheduled appointments. To control recall bias, each patient treated at our centres was asked to keep a medical diary with information on seizure onset, combinations with other drugs, adverse events, and hospital admissions. To control the loss of follow-up bias in the case of patients who did not appear for regular visits, follow-up data were obtained through telephone interviews or with structured questionnaire letters by mail. A patient who was lost of contact for more than one year was defined as a follow-up loss.The primary outcome of this study was time to treatment failure . A patient with poor compliance was defined as having discontinued SRVPA or topiramate treatment by his or her own volition. Secondary clinical outcomes were the time to first seizure, the time from SRVAP or topiramate treatment to achieve 12-month remission of seizures (patients without any type of seizure for at least 12 months), the time to 24-month remission of seizures and the drug tolerability. The incidence of clinically important adverse events (AEs) and the incidence of IAEs directly leading to treatment failure were analysed to investigate the outcome of drug tolerability.Sample size calculations were based on the primary outcome. It was assumed that the treatment failure for SRVPA and topiramate were 25% and 35% after one year, respectively The baseline characteristics of the two groups were compared using Chi-square tests and Fisher's exact tests, except the data represented as mean \u00b1 standard deviation (SD) which were analysed by using student's t-test. For the analysis of the primary and secondary outcomes, intention-to-treat (ITT) analysis was performed . The ITTA total of 1008 patients were enrolled in this study . The stuIn the ITT analysis, a total of 350 treatment failure events were observed over the whole study . There wThe reasons for treatment failure in the SRVPA group and topiramate group are shown in The ITT analysis showed that the time to first seizure was significantly different between the SRVPA group and the topiramate group  . Table 2Overall, 118 patients (22.7%) in the SRVPA group and 122 patients (24.9%) in the topiramate group reported clinically important AEs . A totalThe major finding of this study is that SRVPA was less likely to be associated with treatment failure than topiramate. No differences were found in terms of tolerability, time to 12-month remission, or time to 24-month remission between the two drugs. However, SRVPA was more suitable for Chinese epileptic patients to prevent first seizure than topiramate. These results do not support the viewpoint of starting with new-generation AEDs rather than traditional AEDs.The change of prescription habits should be always based on the studies which could translate their findings into everyday use. However, few studies have answered which generation AEDs should be the first choice of initial monotherapy for Chinese epileptic patients. There was only one study have investigated the effectiveness of three AEDs for generalized onset and unclassified seizures in Chinese epileptic children retrospectively For time to treatment failure, our study showed that the failure rates of topiramate were higher than those of valpriate, which was consistent with the results from the SANAD study (Arm B) Seizure recurrence brings patients a great deal of mental burden. Also, time to seizure remission is one of the indicators to judge medication persistence. Thus, we added time to first seizure and time to 12- or 24-month seizure remission as the secondary outcomes of this study. Our results demonstrated that SRVPA was also better than topiramate in time to first seizure, although no differences were found in time to 12-month or 24-month remission. In addition, our study showed lower IAEs (both IAEs and LE & IAEs) rates than those in previous studies The target dosages of SRVPA and topiramate were 1000\u20132000 mg/day and 100\u2013250 mg/day, respectively. To achieve this dosage, the patients needed to take the drugs twice every day. Indeed, 19.5% of the patients treated with SRVPA and 31.7% of the patients treated with topiramate in our study complained about the inconvenience of taking their medication. Some patients stated that taking drugs twice a day meant doubling the risk of forgetting to take the drugs and doubling the worry that they would be known as epileptic patients by classmates or colleagues. Although there was no difference between the two medications in terms of treatment failure caused by poor compliance, this inconvenience should be noted by clinicians in practice.Cohort studies have been used to examine a variety of outcomes after single exposure and are considered to be the best way to identify potential incidents Loss of follow-up was an important limitation for interpreting our results. During the long period of follow-up, 19\u00b73% and 22\u00b75% of patients were lost in SRVPA and topiramate groups, respectively. However, no difference was found between the two groups regarding the incidence of loss to follow-up, and both the PP analysis results and the ITT analysis results indicated that SRVPA is better than topiramate for primary outcomes.The effectiveness of the new-generational drug of topiramate is not superior to that of SRVPA. The SRVPA, one of the oldest broad-spectrum AEDs, by virtue of its lower long-term treatment failure rates and favourable efficacy profile in preventing occurrence of first seizures after medication, is still the optimal choice for Chinese patients with epilepsy. Lack of efficacy, rather than adverse events, was the most frequent reason for treatment failure, and the inconvenience of drug taking resulted in poor compliance, which was another important reason for treatment failure. In clinical practice, lower starting dosages and slower titration of AEDs will contribute to drug tolerability.Materials S1The Protocol of this study.(DOC)Click here for additional data file.Materials S2The checklist of items for this cohort study based on STROBE statement.(DOC)Click here for additional data file."}
+{"text": "ISRCTN59549825.Folic acid supplementation may potentially alter the efficacy of sulfadoxine-pyrimethamine (SP) treatment in children with malaria. However, there is lack of evidence from randomized controlled trials and effects of folic acid supplementation on clinical efficacy of SP therapy remain moderately understood among children. In a double masked, placebo-controlled trial among preschool children in Pemba Island (Tanzania), iron and folic acid supplementation (Fe/FA) showed an increased risk of hospitalizations and death. In the present paper, we evaluated if folic acid supplementation reduced the efficacy of malaria treatment and thereby contributed to observed adverse effects. During the study, 1648 children had confirmed malarial episodes and received either sulphadoxine-pyrimethamine (SP) treatment and iron folic acid or SP treatment and placebo. These children were evaluated for recovery and incidence of hospitalization during the next 15, 30, and 140 days. Two groups did not differ in malarial episode or hospitalization rate on subsequent 15, 30, and 140 days. Altered efficacy of SP by folic acid was not observed and did not contribute to adverse events in the previous trial. This trial is registered with Controlled-trials.com  With widespread emergence of chloroquine resistant P. falciparum infections, drugs targeting the critically important folate metabolism of malarial parasite have been frequently used [Malaria affects approximately 219 million people each year (range 154\u2013289 million) with an estimated 660,000 deaths. Ninety percent of malaria deaths occur in Africa, where malaria accounts for about one in six of all childhood deaths , or (b) Fe, FA and zinc (Zn) (10\u2009mg/day), or (c) Zn alone or (d) placebo. Children <12 months received half the dose. All the children also got vitamin A as per WHO recommendations. They were visited weekly at home when the supplement was delivered and information on mortality and hospitalization was collected. All hospitalizations were prospectively monitored, and blood samples were collected.The details methods of the study have been reported elsewhere . In the In the substudy, 3171 children in the age group of 1\u201335 months were included as a part of which a detailed physical examination was performed; data related to their height and weight was taken and parental interview was conducted by clinicians and trained health workers. A 3\u2009mL venous blood sample was also obtained from children for detailed haematological analysis , erythrocyte zinc protoporphyrin , and malarial parasite count (thick and air-dried thin films were prepared and stained with Giemsa and read by light microscope with a \u00d7100 oil immersion lens). Repeat assessments were performed at 6 months and 12 months after enrollment. Hospitalizations of the children in any of the five hospitals of Pemba were documented; information included details on illnesses of admitted children, tests for malaria parasites, discharge diagnosis and treatment received, or death. As per the WHO recommendations, for treatment purpose, cases were defined as all confirmed malaria with parasite count of \u2265125 per 200 leucocytes. In these children and children visiting hospitals, a dose of sulfadoxine-pyrimethamine (SP) as a first-line treatment for malaria was provided by the study staff after laboratory confirmed diagnosis of malaria. A study worker read the consent statement to the primary caregiver and signed the form if consent was given. The trial was approved by the Institutional Review Board (IRB) at Johns Hopkins University and the Zanzibar Research Council.For the present study, follow-up morbidity and subsequent 140-day hospitalization data for all 1648 children who were identified as malaria positive by research laboratory and given SP treatment was extracted.Analysis compared children receiving SP and placebo with children receiving SP and Fe/FA or Fe/FA/Zn. Further to assess the effect of folic acid, we combined the data from Fe/FA and Fe/FA/Zn . We estimated the relative risk ) of duration of treated episode of malaria extending beyond 4 days or 7 days and the RR (95% CI) of hospitalization during consequent 15 days, 30 days, or 140 days.For this analysis, out of the 4 groups in the previous trial, we included data from 3 groups with laboratory confirmed malaria: 552 children in placebo group, 550 children in Fe/FA group, and 546 children in Fe/FA/Zn group and compared children receiving SP and placebo with children receiving SP and Fe/FA or Fe/FA/Zn. In all there were 1096 children who received FA along with SP (FA group). Our results suggest that consumption of FA during the SP treatment for confirmed uncomplicated malaria did not increase the risk of prolonging the duration of treated malarial episode nor did it result in relapse of illness with higher hospitalization rates during subsequent 15, 30, or 140 days. Although this analysis cannot definitively address the question of effect of FA supplement on efficacy of SP, they do exclude this effect being responsible for the observed increase in adverse events associated with Fe/FA supplementation in the previous trial [P. falciparum grown in culture converts Guanosine-5\u2032-triphosphate to polyglutamated derivatives, mainly methyltetrahydrofolate pentaglutamate [Biosynthesis of folate in the malarial parasite and the folate dependent reactions that it can carry out are not fully defined . There alutamate . Plasmodlutamate . Howeverlutamate .In the red cell, methyltetrahydrofolate exists highly concentrated almost exclusively as polyglutamate. It has been suggested that plasmodium cannot use this large store of reduced folate because of lack of the enzyme glutamylhydrolase to remove the polyglutamate moieties . ClinicaP. falciparum malaria is complicated by the ability of many strains to use the exogenous folate present in the host, thus obviating the need for de novo synthesis and bypassing sulfadoxine inhibition of dihydropteroate synthetase. Pyrimethamine selectively inhibits the plasmodial enzyme dihydrofolate reductase to markedly different extent depending on its source, with the plasmodial enzyme inhibited by a concentration 3500 times less than the human enzyme [P. falciparum that use exogenous folate efficiently, sulfadoxine inhibition can be restored by pyrimethamine. Thus, the combination of sulfadoxine and pyrimethamine (SP) acts synergistically, inhibiting two key enzymes in the biosynthesis and utilization of folate in P. falciparum and thus the synthesis of DNA and cell growth [Sulfadoxine is a competitive inhibitor of the malarial enzyme dihydropteroate synthetase required for the condensation of p-aminobenzoate (pABA) and pterin in the de novo synthesis of folate. The use of sulfadoxine alone in n enzyme . In stral growth .\u03bcg FA daily for 12 days along with SP therapy to adults and reported no difference in parasite clearance [\u03bcg of FA along with standard SP therapy indicate that folic acid supplementation was associated with increased survivability of parasites beyond 28 days\u2014a parasitological failure [P. falciparum infection, a high blood folate concentration was associated with a 1.5-fold increased risk of late treatment failure [In an earlier study, Tong et al. provided 5\u2009learance . However failure . Similar failure .In spite of the observed parasitological failure, studies that have evaluated both parasitological failure and clinical failure, have not reported any significant difference in clinical failure of therapy or adverse clinical outcomes , 14.\u03bcg/day) in regard to clinical recovery, relapse, or severity during 15 or 30 days after SP treatment. Therefore, it seems unlikely that folic acid supplement could have been responsible for higher rate of adverse events associated with Fe/FA supplementation in the previous trial at least by a mechanism affecting the efficacy of SP treatment.Our study was not designed to evaluate the association between the different folate doses and parasite density or clearance of malarial parasites, and this paper does not address that outcome. The supplement formulation (specific amounts of each nutrient) used for the study was the one which is recommended by Tanzania Food and Nutrition Center with the assumption that it can later be adopted for a national nutritional program. However, we did not find any evidence of reduced efficacy of SP therapy by folic acid (50\u2009The present study indicates that the dose of folic acid used had no effect on SP efficacy and therefore suggests that the negative impact on morbidity observed could not have been due to folic acid affecting antimalarial treatment. The evidence for the need of supplementing folic acid is weak and in areas with high malaria burden and SP being used as first-line treatment withholding folic acid may be advisable."}
+{"text": "Plasmodium falciparum malaria from seven sub-Saharan African countries.The Four Artemisinin-Based Combinations (4ABC) Study Group reports a randomized, non-inferiority trial comparing the efficacy and safety of four ACTs in children with mild  Artemisinin-based combination therapies (ACTs) are the mainstay for the management of uncomplicated malaria cases. However, up-to-date data able to assist sub-Saharan African countries formulating appropriate antimalarial drug policies are scarce.Plasmodium falciparum malaria were treated , actively followed up until day 28, and then passively followed up for the next 6 mo. At day 28, for the PCR-adjusted efficacy, non-inferiority was established for three pair-wise comparisons: DHAPQ (97.3%) versus AL (95.5%) ; DHAPQ (97.6%) versus ASAQ (96.8%) , and ASAQ (97.1%) versus AL (94.4%) . For the PCR-unadjusted efficacy, AL was significantly less efficacious than DHAPQ (72.7% versus 89.5%)  and ASAQ (66.2% versus 80.4%) , while DHAPQ (92.2%) had higher efficacy than ASAQ (80.8%) but non-inferiority could not be excluded . CD+A was significantly less efficacious than the other three treatments. Day 63 results were similar to those observed at day 28.Between 9 July 2007 and 19 June 2009, a randomized, non-inferiority (10% difference threshold in efficacy at day 28) clinical trial was carried out at 12 sites in seven sub-Saharan African countries. Each site compared three of four ACTs, namely amodiaquine-artesunate (ASAQ), dihydroartemisinin-piperaquine (DHAPQ), artemether-lumefantrine (AL), or chlorproguanil-dapsone-artesunate (CD+A). Overall, 4,116 children 6\u201359 mo old with uncomplicated P. falciparum malaria.This large head-to-head comparison of most currently available ACTs in sub-Saharan Africa showed that AL, ASAQ, and DHAPQ had excellent efficacy, up to day 63 post-treatment. The risk of recurrent infections was significantly lower for DHAPQ, followed by ASAQ and then AL, supporting the recent recommendation of considering DHAPQ as a valid option for the treatment of uncomplicated NCT00393679; Pan African Clinical Trials Registry PACTR2009010000911750ClinicalTrials.gov Please see later in the article for the Editors' Summary Plasmodium falciparum is responsible for most of these deaths. During the second half of the 20th century, the main treatments for malaria were inexpensive \u201cmonotherapies\u201d such as chloroquine and sulfadoxine-pyrimethamine. Unfortunately, the malaria parasite quickly developed resistance to many of these monotherapies, and in the 1990 s, there was a widespread upsurge in P. falciparum malaria. To combat this increase, the World Health Organization (WHO) now recommends artemisinin-based combination therapy (ACT) for first-line treatment of P. falciparum malaria in all regions where there is drug-resistant malaria. In ACT, artemisinin derivatives  are used in combination with another antimalarial drug (a partner drug) to reduce the chances of P. falciparum becoming resistant to either drug.Malaria is a global public-health problem. Half the world's population is at risk of this mosquito-borne parasitic disease, which kills a million people (mainly children living in sub-Saharan Africa) every year. Although several parasites cause malaria, P. falciparum malaria. In a randomized trial, groups of randomly chosen patients with a specific disease are given different treatments and then followed to compare the outcomes of these interventions. A non-inferiority trial investigates whether one treatment is not worse than another treatment.WHO currently recommends five ACTs\u2014amodiaquine-artesunate (ASAQ), dihydroartemisinin-piperaquine (DHAPQ), artemether-lumefantrine (AL), artesunate-mefloquine, and artesunate-sulfadoxine-pyrimethamine\u2014for the treatment of malaria. Its treatment guidelines state that the choice of ACT in a country or region should be based on the local level of resistance to the non-artemisinin-based partner drug in the combination. However, data on resistance levels to these partner drugs are scarce or unavailable for many sub-Saharan African countries. To help these countries make an informed choice about their national antimalarial treatment policies, in this randomized, non-inferiority trial, the researchers compare the efficacy and safety of four ACTs in African children with uncomplicated (mild) Each of twelve sites in seven sub-Saharan African countries compared three ACTs out of ASAQ, DHAPQ, AL, and chlorproguanil-dapsone-artesunate (CD+A). Overall, 4,116 young children with uncomplicated malaria were treated with ACT, actively followed up for 28 days (their parents brought them back to the site for pre-arranged check-ups), and passively followed up for six months (parents brought their children back if they developed any illnesses). At each visit, blood samples were examined for the presence of parasites, and a technique called PCR was used to determine which cases of malaria were new infections and which were recurrences of the original infection. The researchers then calculated the percentage of patients with no infection or with a new infection  and the percentage of patients with no infection (the PCR-unadjusted ACPR). For the PCR-adjusted efficacy, three pair-wise comparisons  showed non-inferiority at 28 days. That is, for example, similar percentages of patients given DHAPQ or AL  had either no infection or a new infection. CD+A was less efficacious than the other three treatments. For the PCR-unadjusted efficacy, AL was significantly less efficacious than DHAPQ and ASAQ; DHAPQ had a higher efficacy than ASAQ, but non-inferiority could not be excluded. That is, the difference in efficacy of these two drugs might have happened by chance.P. falciparum malaria.These findings suggest that AL, ASAQ, and DHAPQ are all efficacious for the treatment of uncomplicated malaria in children; CD+A was withdrawn partway through the trial because of side effects, but these findings also suggest that it was less efficacious than the other ACTs. Importantly, the PCR-unadjusted results indicate that the risk of children becoming re-infected with malaria parasites soon after treatment was lowest for DHAPQ, followed by ASAQ, and then AL. Because these findings are based on pooled results from seven sub-Saharan African countries, they are likely to be generalizable and thus of use in setting national antimalarial drug policies throughout the region. AL and ASAQ are already included in the antimalarial drug policies of many sub-Saharan African countries, note the researchers, but these findings support the WHO recommendation that DHAPQ should also be considered for the treatment for uncomplicated http://dx.doi.org/10.1371/journal.pmed.1001119.Please access these websites via the online version of this summary at malaria ; the 2010 World Malaria Report provides details of the current global malaria situation; the WHO Guidelines for the Treatment of Malaria and the report Assessment and Monitoring of Antimalarial Drug Efficacy for the Treatment of Uncomplicated Malaria are availableInformation is available from WHO on malaria (in English and Spanish), including a selection of personal stories about malariaThe US Centers for Disease Control and Prevention provide information on global control of malaria including fact sheets about ACTs and about malaria in AfricaInformation is available from the Roll Back Malaria Partnership on the malaria (in English and Spanish)MedlinePlus provides links to additional information on  The burden of malaria has declined substantially in several areas of sub-Saharan Africa, particularly in the past 3\u20135 y The WHO recommends five ACTs, namely artemether-lumefantrine (AL), amodiaquine-artesunate (ASAQ), mefloquine-artesunate, sulfadoxine-pyrimethamine-artesunate, and, most recently included, dihydroartemisinin-piperaquine (DHAPQ) AL was the first co-formulated ACT to become available and, together with ASAQ, is the most common ACT used in sub-Saharan Africa. DHAPQ has been recently submitted for registration to the European Medicines Agency under the orphan drug legislation Between 9 July 2007 and 19 June 2009, a randomized, open-label, multicenter, non-inferiority clinical trial was carried out at 12 sites located in seven African countries . See protocol  and amenEach site compared three of the four ACTs under investigation, ASAQ, DHAPQ, AL, or CD+A. The decision of which treatments to test at a given site was made by considering the current first-line treatments, the known antimalarial resistance profile, and local malaria endemicity . PatientPlasmodium falciparum mono-infection with asexual parasite densities between 2,000 and 200,000/\u00b5l, fever (axillary temperature \u226537.5\u00b0C) or history of fever in the preceding 24 h, and Hb \u22657.0 g/dl. Patients were not recruited if they met at least one of the following exclusion criteria: participation in any other investigational drug study during the previous 30 d; known hypersensitivity to the study drugs; severe malaria Children 6\u201359 mo old (12\u201359 mo old at sites where CD+A was used) attending the health facilities with suspected uncomplicated malaria were included in the study if they fulfilled all the following inclusion criteria: body weight >5 kg, microscopically confirmed Three treatments were co-formulated , while CD+A consisted of separate tablets of chlorproguanil-dapsone and artesunate. All drugs were administered under direct supervision during three consecutive days, according to the patient's body weight. ASAQ  was given once daily, at the standard dose of 2.8\u20135.5 mg/kg and 7.5\u201315 mg/kg of artesunate and amodiaquine, respectively. Three formulations were used  and given at the dosage of one tablet/day according to body weight . DHAPQ  was given once daily, at the standard dosage of 2.25 mg/kg and 18 mg/kg of dihydroartemisinin and piperaquine, respectively, rounded up to the nearest half tablet. Two formulations were used (20 mg dihydroartemisinin +160 mg piperaquine and 40 mg dihydroartemisinin +320 mg piperaquine). AL  was administered twice a day  according to the following dosage: weight\u200a=\u200a5\u201314 kg: one tablet per dose; weight\u200a=\u200a15\u201324 kg: two tablets per dose; weight\u200a=\u200a25\u201334 kg: three tablets per dose. Chlorproguanil-dapsone  was administered once a day at the dose of 2.0 mg/kg chlorproguanil and 2.5 mg/kg dapsone. The daily dose of the formulation used (15 mg chlorproguanil and 18.75 mg dapsone) was given according to body weight, i.e., 4\u20135.9 kg: 0.5 tablet; 6\u20139.9 kg: 1 tablet; 10\u201313.9 kg: 1.5 tablet; 14\u201315.9 kg: 2 tablets; 16\u201319.9 kg: 2.5 tablets; 20\u201324.9 kg: 3 tablets; 25\u201330.9 kg: 4 tablets. Arsumax  contained 50 mg artesunate per tablet and was administered with chlorproguanil-dapsone according to the following dosage: 5\u20138.3 kg: 0.5 tablet; 8.4\u201316.7 kg: 1 tablet; 16.8\u201320.8 kg: 1.5 tablet; 20.9\u201329.2 kg: 2 tablets. In case of vomiting, a full dose was repeated if this occurred within 30 min, or half a dose if it occurred between 30 min and 1 h. AL was administered concomitantly with a fatty meal (as recommended by the manufacturer), e.g., milk or groundnuts, while for the other three treatments no specific instructions regarding co-administration with food were given. For infants, medicines were crushed, mixed with water, and administered as a slurry. Treatment was administered by a study nurse, with the clinician or other staff following up the patient and assessing the end points blinded to the treatment assignment whenever possible.Treatment was directly observed and children stayed at the health facility at least 1 h to check for any vomiting. At some sites children were kept at the health facility for the whole 3-d dosing period. The parent/guardian was asked to return with the child for scheduled visits on days 3, 7, 14, 21, and 28, or if any symptoms occurred (active follow-up period). Field workers traced patients who missed any visit. In addition, after day 28 and for the next 6 mo, the parent/guardian was encouraged to attend the health facility whenever the child was sick, so as to detect any malaria episode(s) (passive follow-up period). In case of clinical malaria, the child received the same treatment as for the primary episode and was actively followed up for the following 28 d (unpublished data). For each visit, a physical examination was performed by the study clinicians, vital signs were recorded, and axillary temperature was measured with an electronic thermometer. Adverse events (AEs) and serious adverse events (SAEs) were recorded and monitored throughout the study. Rescue treatment for recurrent infections identified during the 28-d follow-up, as well as for severe malaria, was according to local national guidelines.Capillary or venous blood was taken at every visit. Thick and thin blood films were prepared, dried, and Giemsa-stained, and parasite density estimated by counting the number of asexual parasites in 200 white blood cells, assuming a standard white blood cell count of 8,000/\u00b5l. Quality control was performed in blind conditions on 10% of all the slides. Samples for hematology (full blood count) were taken at enrollment and at days 3, 7, 14, and 28, while biochemistry  was performed at enrollment and days 7 and 28. Laboratory tests were also performed at any other visit if judged necessary by the clinician. For PCR analysis, blood samples were collected on filter paper (Whatman 3MM) at enrollment and at any visit after day 7. Each filter paper was dried and individually stored in a plastic bag containing silica gel. All filter papers were subsequently transported to the Institute of Tropical Medicine, Antwerp, Belgium, where centralized genotyping according to international recommendations was conducted Treatment outcomes were classified according to the WHO guidelines The primary end points were PCR-adjusted and unadjusted ACPR at day 28; secondary efficacy outcomes included PCR-adjusted and unadjusted ACPR at day 63  http://clinicaltrial.gov/ct2/show/NCT00393679) and in the Pan African Clinical Trials Registry .The study protocol and successive amendments were approved by the Institutional Review Board of the Institute of Tropical Medicine, Antwerp; the Ethical Committee of the Antwerp University Hospital; the national Ethics Review Committee or Institutional Review Board at each trial site; and the national competent authorities, as appropriate, e.g., Ministry of Health and national drug regulatory authorities. The trial was conducted under the provisions of the Declaration of Helsinki and in accordance with Good Clinical Practices guidelines set up by the WHO and by the International Conference on Harmonization. An independent Data Safety and Monitoring Committee was created prior to the beginning of the trial and regularly reviewed both the safety data and the quality of the information collected. The trial was registered prior to the enrollment of the first patient in the ClinicalTrials.gov registry , per protocol, and two sensitivity analyses. Patients lost to follow-up or withdrawn for reasons unrelated to malaria, as determined by the end point adjudication committee, were excluded from the ITT analysis. Major protocol violators defined prior to analysis  were excluded from the per protocol analysis. All secondary outcomes were analyzed using the ITT approach, with patients having received rescue treatment or lost to follow-up censored at their last visit. Because CD+A was discontinued partway through the study and the lower age limit for inclusion changed at that time, the analyses involving CD+A include only patients randomized prior the discontinuation.The primary hypothesis was that the four treatments are clinically non-inferior to each other as measured by the proportion of children with PCR-adjusted and unadjusted ACPR at day 28. This was tested for each of all possible (six) pair-wise comparisons and the two primary end points, with the limit of non-inferiority on an odds ratio (OR) scale corresponding to a 10% absolute difference in ACPR. Data from sites testing the same two treatments were included and combined into a meta-analysis using the study site as unit.The meta-analysis was performed using ORs rather than risk differences as the latter usually display more heterogeneity than ORs For the secondary outcomes, including ACPR at day 63, differences between treatments were estimated using differences in means, ORs, or hazard-ratios, as appropriate, and the corresponding 95% CIs. Parasite and fever clearance were defined as two consecutive days without parasite or fever, respectively, and analyzed using Cox proportional hazard models stratified by study site. Gametocyte prevalences were assessed by calculating the number of gametocyte carriage days for each patient as an area under the curve. The number of gametocyte carriage days was analyzed using zero-inflated Poisson models with the number of days in active follow-up as offset, and summarized as ORs (for zero gametocyte carriage days) and ratios of the expected number of gametocyte carriage days between groups. This last ratio is expressed as the percentage of the reduction in gametocyte carriage days.Safety was assessed through AE reporting; all individuals having received at least one dose of the treatment were included and analyzed according to the treatment actually received.This study was designed as a non-inferiority trial, and on an expected ACPR at day 28 of at least 90%. Though the primary intention was to carry out a pooled analysis able to provide overall head-to-head comparisons between the different treatments, the sample size was based on the individual sites to also produce locally meaningful results. Therefore, for each individual site, 170 children per arm should be able to show that the difference in efficacy between treatments was less than 10%, at 5% significance level and 90% power, assuming equal true ACPR rates.Overall, 11,030 patients were screened, 6,382 did not meet the inclusion/exclusion criteria, 532 declined to participate, and 4,116 were recruited and randomized to receive the study drugs  . The smaThe safety analysis included all other randomized patients apart from eight patients: two (randomized to DHAPQ) were withdrawn before receiving the study drug  and six were withdrawn because they vomited their first dose twice .All results presented below refer to the ITT analysis unless specified otherwise. As an indication of the overall efficacy of the different treatments, the pooled results are reported below. However, the analysis of each of the six pair-wise comparisons was stratified by site. The pooled  PCR-adjusted ACPR was high, around 95%, for ASAQ, DHAPQ, and AL, both at day 28 and 63, while for CD+A this was around 85%. In addition, CD+A had the lowest PCR-unadjusted ACPR, followed by AL, ASAQ, and then DHAPQ, with the majority of recurrences due to new infections . At day Day 63 results were similar to those observed at day 28. For the PCR-corrected efficacy, DHAPQ, ASAQ, and AL had high (\u226594.6%) and similar efficacy. CD+A, in each pair-wise comparison, had a significantly lower efficacy (<85%) than any of the other three treatments. PCR-unadjusted efficacy was significantly higher for DHAPQ than for ASAQ  and AL , while that for ASAQ was significantly higher than for AL . CD+A had a significantly lower efficacy than the other three treatments . Pair-wiDHAPQ had a PCR-adjusted efficacy similar to that of AL at most sites, with the exception of Pamol (Nigeria), where AL efficacy both at day 28 and 63 tended to be higher, and in Nanoro (Burkina Faso), with a significantly higher efficacy of DHAPQ at day 28 but not at day 63 . PCR-adjParasite clearance was rapid in all treatment groups as the large majority of patients had no detectable infection at day 3, with no major differences in the time of clearance between treatment groups . SimilarThe evolution of gametocyte prevalence by treatment group is shown in Hb increased in all treatment groups and attained the day 0 levels at day 7, except in patients treated with CD+A . In the A total of 4,108 patients were included in this analysis. Up to day 28, 37 patients experienced SAEs, their occurrence being relatively more frequent in patients treated with ASAQ or CD+A than among those treated with DHAPQ or AL . Most SADuring the first 28 days of follow-up, over 60% of patients experienced at least one AE . VomitinThe median levels of alanine aminotransferase and creatinine before treatment, as well as the proportion of patients with values above the normal range , were similar between the four study arms, and this did not change during the follow-ups at day 7 and 28 .This large multicenter trial aimed to collect information that would assist national malaria control programs in sub-Saharan African countries in choosing the most appropriate ACTs for their specific setting. The four ACTs most likely to be considered for this purpose at the time the study was conceived and implemented, namely AL, ASAQ, DHAPQ, and CD+A, were compared at 12 sites distributed over seven countries. The development of CD+A was discontinued partway through the study, following the results of a phase III clinical trial that showed a higher risk for severe and clinically concerning Hb decrease Each site tested three of the four ACTs, i.e., in ten sites both AL and DHAPQ were tested; in six of these the third arm was ASAQ and in the other four, CD+A; two additional sites tested ASAQ, DHAPQ, and CD+A. Following the discontinuation of the CD+A arm, six sites continued testing only two study treatments. Such study design does not allow the use of the pooled results of the four study treatments, as the efficacy of a given ACT tested in one site cannot be directly compared with that of another ACT tested only in another site. Therefore, it is necessary to analyze the six pair-wise comparisons and to include in each comparison only sites in which the two study treatments were actually tested.When considering the pooled data across all sites, non-inferiority could be established for the three pair-wise comparisons without CD+A, indicating that ASAQ, AL, and DHAPQ had similar and high PCR-adjusted efficacy, both at day 28 and 63; efficacy was lower for CD+A. Conversely, the unadjusted efficacy estimates were higher for DHAPQ, followed by ASAQ and then AL, though at day 63 the difference between DHAPQ and ASAQ seemed to disappear; also in this case CD+A had a much lower efficacy than the other three treatments. It is important to mention that beyond day 28, the detection of recurrent infections was passive, i.e., only sick children attending the health facility had a blood slide prepared, so that some asymptomatic infections may not have been detected. Nevertheless, the important difference in the PCR-unadjusted efficacy between day 28 and 63 suggests that during this period a substantial proportion of children had a recurrent malaria infection, mostly a new infection, as indicated by the small difference between the PCR-adjusted efficacy at day 28 and 63.The site-specific estimates mirror the overall results, with some notable and surprising exceptions. DHAPQ had the highest PCR-unadjusted efficacy in several sites known for their intense malaria transmission, e.g., Nanoro (Burkina Faso) and Tororo (Uganda), so that the difference observed can be explained by the long post-treatment prophylactic period related to the slow elimination of piperaquine. This is also the reason why DHAPQ performed better than ASAQ at the only high-transmission site where it was tested, i.e., Nanoro (Burkina Faso). Nevertheless, it is surprising that PCR-unadjusted efficacy for DHAPQ was significantly better than that of AL and ASAQ in Ndola (Zambia), where transmission has decreased dramatically over the past few years, to the extent that the target sample size for this trial could not be attained. Unlike Tororo (Uganda), where the PCR-adjusted efficacies for AL and DHAPQ were similar, in Ndola (Zambia), such efficacy tended to be higher for DHAPQ, and the difference probably did not reach statistical significance because of the small sample size, though AL efficacy was almost 95%. Similarly, at this site the difference between DHAPQ and ASAQ PCR-adjusted efficacy was of borderline significance, though ASAQ efficacy was about 94%. It is unclear what such differences mean, as all treatments, with the exception of CD+A, had a PCR-adjusted efficacy >90% and the trial aimed at showing non-inferiority at a 10% difference threshold. Non-inferiority was demonstrated for the three pair-wise comparisons involving DHAPQ, ASAQ, and AL at most sites, with a few exceptions where the individual sites' 95% CI cross the non-inferiority limit.In west Africa, ASAQ continues to have excellent efficacy, comparable to that of AL In children treated with AL, gametocyte prevalence during follow-up and gametocyte carriage time were significantly lower than in children treated with either DHAPQ or ASAQ. The difference remained significant for gametocyte carriage time even when excluding patients with gametocytes at enrollment. Higher gametocyte carriage after treatment with DHAPQ, when compared to either AL or mefloquine-artesunate, has already been reported in some Hematological recovery up to day 28 post-treatment was similar for all ACTs tested except for CD+A, for which this was significantly slower, with a more marked Hb decrease up to day 7, confirming previous results P. falciparum malaria, as its efficacy is excellent and comparable to the other ACTs, while its long post-treatment prophylaxis could be an additional advantage.In conclusion, this is, to our knowledge, the largest head-to-head comparison of most of the currently available ACTs for falciparum malaria in sub-Saharan Africa. CD+A was suspended partway through the trial, leaving AL, ASAQ, and DHAPQ under investigation. These three ACTs showed excellent efficacy, up to day 63 post-treatment, but the risk of recurrent infections was significantly lower, even in areas of high transmission, for DHAPQ, followed by ASAQ, and then AL. Although the gametocyte carriage rate differed between regimens, with those treated with AL having the lowest carriage rate and those treated with ASAQ having the highest carriage rate, the meaning of these different carriage rates with relation to transmission potential is unclear. The possibility of adding a single dose of primaquine to any of these three ACTs, with the objective of further reducing gametocyte carriage, should be explored Text S1Study protocol.(PDF)Click here for additional data file.Text S2CONSORT checklist.(PDF)Click here for additional data file.Text S3Protocol amendment 1.(PDF)Click here for additional data file.Text S4Protocol amendment 2.(PDF)Click here for additional data file.Text S5Protocol amendment 3.(PDF)Click here for additional data file."}
+{"text": "Safety surveillance of widely used artemisinin-based combination therapy (ACT) is essential, but tolerability data in the over five years age group are largely anecdotal.Plasmodium falciparum malaria patients aged over five years (Study-T), and, ii) an efficacy trial with a secondary objective of collecting tolerability data among 300 children age six to 59\u00a0months (Study-E). In both studies patients were randomized to fixed-dose artesunate-amodiaquine (ASAQ Winthrop\u00ae) or artemether-lumefantrine , respectively. Clinical- and laboratory-adverse events (AEs) were recorded until day 28.Two open-label, randomized trials were conducted in Nimba County, Liberia: i) the main tolerability trial with 1,000 vs 16.3%, p\u2009<\u20090.001), vomiting , nausea , and anaemia  were more frequently recorded in the ASAQ versus AL arm. Study-E: mild or moderate AEs were common, including anaemia, fatigue, vomiting or diarrhoea. The few severe events were asymptomatic blood system disorders and four clinical events .Study-T: most patients experienced at least one AE. Severe AEs were few, primarily asymptomatic blood system disorders or increased liver enzyme values. No treatment or study discontinuation occurred. Mild or moderate fatigue .The protocols were registered with Current Controlled Trials, under the identifier numbers  Plasmodium falciparum malaria. Among the most widely available current ACT are artemether-lumefantrine (AL) and artesunate-amodiaquine (AS\u2009+\u2009AQ). Artemisinin and its derivatives are well tolerated [Artemisinin-based combination therapy (ACT) is a key tool in malaria control and is the World Health Organization\u2019s (WHO) recommended treatment for uncomplicated olerated , and theolerated -4. Serioolerated -7, or seolerated . A dose-olerated . A recenolerated . HemolytIdeally ACT should be provided as fixed-dose combinations (FDC) to improve compliance. The first FDC of AS with AQ, (ASAQ Winthrop\u00ae Sanofi-Aventis) achieved WHO-prequalification in 2008 and is now registered in 32 countries. Phase III studies showed safety, good tolerability and high efficacy ,11,12. GTwo open-label, randomized, controlled, two-arm clinical trials were conducted in the Comprehensive Healthcare Center (CHC) of Saclepea, Nimba County, Northern Liberia. M\u00e9decins sans Fronti\u00e8res (MSF) Switzerland coordinated the CHC in collaboration with the Ministry of Health. Since 2004, AS\u2009+\u2009AQ co-blister has been provided as first-line treatment in this area.P. falciparum malaria compared to artemether-lumefantrine (AL) . Tolerability was assessed based on the frequency and severity of adverse events. Inclusion criteria were: age older than five years; weight equal or higher than 18\u00a0kg; fever (axillary temperature \u2265 37.5\u00b0C), or history of fever in previous 48\u00a0hours; microscopic confirmation of asexual stages of P. falciparum or mixed infection; high probability of attending the follow-up visits; signed informed consent (responsible caregiver). Exclusion criteria were: pregnancy; or severe malaria; severe anaemia (<5\u00a0g/dl haemoglobin (Hb)); or full course of AS\u2009+\u2009AQ or AL treatment or more than two doses of another anti-malarial in the past four weeks; or known hypersensitivity to artemisinin derivates, amodiaquine, or artemether-lumefantrine; or concomitant febrile illness if additional medication is required other than antipyretics.The principal objective of the main study reported here was to describe the tolerability of the fixed dose of ASAQ (Winthrop FDC) in adults and children aged over five years with uncomplicated P. falciparum malaria after 42\u00a0days of follow up  [P. falciparum malaria  and parasite density between 2000\u2013200,000/\u03bcl blood; high probability of attending follow-up; signed informed consent (responsible caregiver). Exclusion criteria were: general danger signs; severe/complicated malaria [The principal objective of the second study was to assess the genotyping-adjusted cure rates of ASAQ compared to AL in 300 children six to 59\u00a0months old with uncomplicated Study-E) . As a se malaria ; severe Between September 2008 and May 2009, suspected malaria patients were pre-screened by an HRP-2 rapid diagnostic test (Paracheck\u00ae), followed by screening with a clinical examination performed by trained physician assistants, malaria blood smear, Hb from capillary blood (HemoCue\u00ae), and urine-pregnancy test . Patients who met the eligibility criteria of the respective trials were randomized to ASAQ or AL (ratio 1:1). In Study-T, randomization was stratified by weight  to balance treatment allocation between adults and children. Allocation lists were computer-generated with a block size of six. The study site was unaware of block size. Treatment allocation was concealed in sealed opaque envelopes to be opened by study nurses in consecutive order at randomization. This was not disclosed to the medical staff performing the clinical assessments. The laboratory team was aware of treatment allocation since samples were taken to measure drug concentrations of the artemisinin partner compounds on day 0 and day 7, which required distinction between treatment arms.http://www.controlled-trials.com/. ISRCTN51688713, ISRCTN40020296).The procedures followed were in accordance with the ethical standards of the Helsinki Declaration. All participants or responsible caretakers (\u226518 years) gave written informed consent. The Liberian Institute for Biomedical Research (LIBR) ethics committee, the Ministry of Health and Social Welfare, Monrovia, Liberia, and the Comit\u00e9 de Protection des Personnes (CPP) Ile de France XI (Saint Germain en Laye), France, approved the studies. The studies were registered at Controlled Trials  was two doses per day, six to 12\u00a0hours between doses, administered with a high-fat cookie or breast-feeding encouraged. Dosage was by weight: ASAQ Winthrop\u00ae 5 to <9\u00a0kg: one tablet/day of both AS 25\u00a0mg/AQ 67.5\u00a0mg; 9 to <18\u00a0kg: one tablet/day of both AS 50\u00a0mg/AQ 135\u00a0mg; 18 to 36\u00a0kg: one tablet/day of both AS 100\u00a0mg/AQ 270\u00a0mg; \u2265 36 kg: two tablets/day of both AS 100\u00a0mg/AQ 270\u00a0mg. Coartem\u00ae tablet strength was 20\u00a0mg artemether/120\u00a0mg lumefantrine: 5 to <15\u00a0kg: one tablet/dose; 15 to <25\u00a0kg: two tablets/dose; 25 to <35\u00a0kg: three tablets/dose; \u226535 kg: four tablets/dose. All doses were administered in the study site followed by 30\u00a0minutes\u2019 observation. If a dose was vomited/spat-out within 30\u00a0minutes, a full dose was re-administered. If the re-administered dose was vomited/spat-out within 30\u00a0minutes, the patient was withdrawn and rescue treatment given .At each visit  a standardized symptoms questionnaire and physical examination were conducted. A \u03b2-human chorionic gonadotropin (\u03b2HCG) urine-pregnancy test was done on day 0 and 28 for females \u226512 years. In Study-T, malaria blood smears were obligatory on days 0, 2 and 28. In Study-E blood smears were done on days 0, 2, 3, 7, 14, 21, 28, 35, 42. On day 0 a serum sample (from venous blood) was stored at \u221220\u00b0C for further analyses. The routine assessment of blood levels of liver enzymes aspartate aminotransferase (AST) and alanine aminotransferase (ALT) was implemented after the start of the tolerability trial (Study-T), following the recommendation of the data safety monitoring committee. Liver function tests (LFTs) were conducted by assessment of blood levels of AST and ALT  on day 0 (fresh serum) and on day 28 (finger-prick capillary blood). Missing day 0 AST values  were measured retrospectively from frozen serum  and added to summary statistics (ALT not sufficiently stable in frozen serum). Baseline total bilirubin (direct) and creatinine were assessed retrospectively from day 0 serum by spectrophotometry . Hb was measured on each visit . A full blood count (FBC) was done on days 0, 7 and 28 , or if the HemoCue\u00ae indicated anaemia.In both studies, day 0 and day 7 blood concentrations of AQ, desethyl-amodiaquine (DEAQ) (ASAQ arm) and lumefantrine (LF) (AL arm) were measured from dried spots of venous blood on filter paper using high performance liquid chromatography with ultraviolet or by tandem-mass spectrometry detection method, respectively ,16. LimiBaseline serum samples from both studies were retrospectively subjected to a screen for viral hepatitis serological markers: a) hepatitis B virus (HBV): hepatitis B surface antigen (HBs Ag) ; b) if Ag HBs-positive a test for the presence of Immunoglobulin-anti-bodies against the hepatitis B core antigen (HBc IgM) was performed ; c) hepatitis C virus (HCV): a test for the presence of immunoglobulin G anti-bodies to HCV(anti HCV IgG) ; and, d) for hepatitis E virus (HEV): the serum of all patients with elevated baseline AST or ALT  were tested for the presence of IgM antibodies to HEV (anti HEV IgM) .versus AL treatment efficacy [Study-T: one thousand participants allowed to detect a minimum of 2.7-3.3% difference between relatively low AE frequencies , and a minimum of 8\u20138.4% differences for relatively high AE frequencies (25-30% in one study arm versus 38.4% in the second arm (80% power and 0.05 significance level) . The Study-E sample size was powered to assess the non-inferiority of ASAQ efficacy .versus baseline grades were displayed in shift tables for all patients for \u201cAST or ALT increased\u201d, or \u201cneutropaenia\u201d. All analyses were performed with STATA 10.1 .Clinical- or laboratory signs and symptoms which occurred or worsened at any time after the first drug intake up to day 28 were recorded as adverse events (AEs) in both studies. AEs were defined according to International Conference on Harmonisation (ICH) guidelines for good clinical practice . For eacIn Study-T 1,000 patients were randomized, 498 allocated to the ASAQ arm and 502 to the AL arm  had grade 2 anaemia at baseline (8.0\u00a0g/dl), and completely recovered during follow up. One serious AE (SAE), spontaneous abortion, was recorded for a 15\u00a0year-old female patient (ASAQ arm) with a positive day 28 \u03b2-HCG urine-pregnancy test (test negative at inclusion) who reported symptoms of vaginal discharge and lower abdominal pain (with onset nine days earlier). A sexually transmitted infection was suspected and treated with standard antibiotics and symptoms subsided. A post-study follow up urine pregnancy test day 40) was negative, and spontaneous abortion diagnosed, declared as - serious AE \u201cpossibly related to study drug\u201d. A retrospective blood \u03b2-HCG test on day 0 serum was positive ), indicating early pregnancy at inclusion. The direct comparison between treatment arms on a set of pre-selected AEs of specific interest revealed significantly higher frequencies of mild or moderate fatigue, vomiting, anaemia and nausea in the ASAQ arm , severe pneumonia (ASAQ), severe vomiting   Table\u00a0. The thrAQ and DEAQ blood concentrations (ASAQ arm) and LF blood concentrations (AL arm) were assessed from day 0 and day 7 blood spots. In Study-T, AQ intake shortly before blood sampling (protocol violation) was indicated for one patient with quantifiable AQ on day 0, for one patient with high DEAQ blood concentration (>200\u00a0ng/ml) on day 0, and for four patients with quantifiable AQ on day 7. Four patients had day 7 DEAQ values <100\u00a0ng/ml, suggesting insufficient intake or malabsorption. One hundred patients in the ASAQ arm had low day 0 DEAQ concentrations (median 3.9\u00a0ng/ml), indicating AQ intake\u2009>\u2009four weeks before inclusion (no deviation). In the AL arm, 19 patients had non-quantifiable LF values on day 7 (<LOQ 200\u00a0ng/ml). Their day 7 values were set to LOQ/2 (100\u00a0ng/ml), since the LOQ was considered low.Study-T: Median day 7 DEAQ and LF concentrations are depicted in Table\u00a0Study-E: very recent AQ intake was indicated for six children who had quantifiable AQ concentrations on day 0 ) and five children with quantifiable AQ on day 7 (range: 2.6-4.2\u00a0ng/ml) (protocol violation). Three children had non-quantifiable DEAQ on day 7 (<2.5\u00a0ng/ml), suggesting malabsorption. In the AL arm, 33 patients had non-quantifiable day 7 LF concentrations , which were set as LOQ/2. The median day 7 DEAQ blood concentration was significantly higher in children with AE fatigue (p\u2009=\u20090.004) than in children without  around 15% HBV prevalence (HBV sAg positive), among these about 4% with a probable (recent) active hepatitis (IgM anti-HBc positive). A total of 4% of included patients were identified as HCV carriers (Study-T). Among children six to 59\u00a0months (Study-E), 12% were HBV carriers (HBV sAg positive), and among these about 5% were also IgM anti-HBc positive. A total of 4% were HCV carriers (Study-E) Table\u00a0.P. falciparum malaria, was assessed in a trial of 1,000 patients over five years old, an age group not frequently studied in the past and able to express symptoms. The findings were complemented with safety data collected in a parallel trial from 300 children under five years. In both studies, ASAQ and AL were very well tolerated and highly efficacious  on Hb and neutrophil counts in both age-groups were minimal, in line with recent data ,21,22. EA comparative analysis on the frequency of patients presenting with adverse events of specific interest was performed in the main tolerability trial (Study-T). The only difference identified between treatment arms were higher frequencies of mild or moderate AEs fatigue, vomiting, nausea, and anaemia recorded in the ASAQ arm. The clinical events were mostly early onset (\u2264day 3), did not affect treatment intake, and were largely resolved before day 28.Mild or moderate fatigue AEs were very frequently reported in both treatment arms, with almost twice as many patients in the ASAQ arm than in the AL arm . Asthenia is considered among common undesirable effects of ASAQ Winthrop (SmPC ASAQ Winthrop\u00ae), similar to what is noted for AL ,24,30,31Among gastrointestinal AEs, mild or moderate vomiting AE was about four times more frequently reported in the ASAQ arm. The incidence of early vomiting immediately after drug intake though was reassuringly low in both study arms. Recent trials conducted in Ghana, Senegal and Ivory Coast, respectively, also reported nausea and vomiting AEs at higher frequency in the AS\u2009+\u2009AQ arm when compared to AL ,30. No dThe bioavailability of the non-artemisinin partner compound of ACTs may also affect tolerability. Among patients over five years (Study-T), the day 7 DEAQ-blood concentration was significantly higher for patients with vomiting AE compared to patients without vomiting AE. Among children under five years (Study-E), the findings were different. Higher day 7-DEAQ values were identified among children with fatigue or hepatotoxicity AE than without, and a higher day 7-LF concentration was found among children with anaemia AE than without. The relevance of these associations is uncertain. Further studies and pooled data analyses are needed to better interpret the relationships of blood concentrations, dosage, and tolerability.Safety and tolerability monitoring of ASAQ FDC and all other ACTs should continue in a standardised manner. Pharmacovigilance networks are not implemented in most settings were ACTs are routinely used. Post-marketing information on safety and tolerability thus relies on individual studies. The need of standardized guidelines for adverse event monitoring of ACTs was recently emphasized ,34. MoreThese findings confirmed the good tolerability of ASAQ FDC (Winthrop\u00ae) and AL (Coartem\u00ae) in patients of all ages in Liberia. Hepatotoxicity, neutropenia or dystonia were of no specific concern in the present studies. Tolerability monitoring of ASAQ FDC should continue, as for all widely used ACT.The authors declare that they have no competing interests.BS: overall trial coordination, participation in study design, protocol and data analysis plan, study documents, writing of the manuscript. PV: field trial coordination, medical coordination, trial team supervision, supervision of data collection. EB: support study design, coordination of data management, data analysis plan, data analysis, revision of the manuscript. CM: field laboratory coordination, laboratory team supervision, - standard operating procedures, - data collection. RS: field coordination during study preparation, writing of administrative and clinical standard operating procedures, preparation of study site, staff training and trial implementation. LP: data management, support data analysis plan, data analysis, revision of the manuscript. MD and JB: coordination and interpretation of the malaria parasite genotyping. TS: support for field trial coordination, team supervision and data collection. YMZ and JJJ: technical support to all study steps, participation in study implementation and training. EC: support study initiation, participation in study design, technical support. AB: baseline biochemistry  for sample subset and baseline creatinine . MB: baseline viral hepatitis serology. VJ: development and conduct of blood concentration analyses of artemisinin-partner compounds. GC: technical advise to protocol development, study preparation, support field training and study conduct, review of study documents, coordination of study monitors and data monitoring committee, revision of the manuscript. JRK: study initiation, technical and scientific advice to study protocol, study preparation and trial conduct, review of study documents, revision of the manuscript. EAA: study initiation, study design and protocol, scientific and medical advise to all steps of the study, revision of study documents, support data analysis plan, revision of the manuscript. PJG: study initiation, study design and protocol, scientific and medical advise to all steps of the study, review of study documents, medical review of adverse events, support data analysis plan, revision of the manuscript. All authors read and approved the manuscript.Mean liver enzyme (AST/ALT) and blood cell parameter values on Days 0, 7 and 28, and change from baseline, respectively, by treatment group \u2013 Safety population, both studies.Click here for file"}
+{"text": "Fascioliasis is an emerging zoonotic disease of considerable veterinary and public health importance. Triclabendazole is the only available drug for treatment. Laboratory studies have documented promising fasciocidal properties of the artemisinins .Fasciola-infected individuals in Egypt. Efficacy was determined by cure rate (CR) and egg reduction rate (ERR) based on multiple Kato-Katz thick smears before and after drug administration. Patients who remained Fasciola-positive following artemether dosing were treated with single 10 mg/kg oral triclabendazole. In case of treatment failure, triclabendazole was re-administered at 20 mg/kg in two divided doses.We carried out two exploratory phase-2 trials to assess the efficacy and safety of oral artemether administered at (i) 6\u00d780 mg over 3 consecutive days, and (ii) 3\u00d7200 mg within 24 h in 36 CRs achieved with 6\u00d780 mg and 3\u00d7200 mg artemether were 35% and 6%, respectively. The corresponding ERRs were 63% and nil, respectively. Artemether was well tolerated. A high efficacy was observed with triclabendazole administered at 10 mg/kg  and 20 mg/kg .Artemether, administered at malaria treatment regimens, shows no or only little effect against fascioliasis, and hence does not represent an alternative to triclabendazole. The role of artemether and other artemisinin derivatives as partner drug in combination chemotherapy remains to be elucidated. Fasciola hepatica and F. gigantica are two liver flukes that parasitize herbivorous large size mammals , as well as humans. A single drug is available to treat infections with Fasciola flukes, namely, triclabendazole. Recently, laboratory studies and clinical trials in sheep and humans suffering from acute fascioliasis have shown that artesunate and artemether  also show activity against fascioliasis. Hence, we were motivated to assess the efficacy and safety of oral artemether in patients with chronic Fasciola infections. The study was carried out in Egypt and artemether administered according to two different malaria treatment regimens. Cure rates observed with 6\u00d780 mg and 3\u00d7200 mg artemether were 35% and 6%, respectively. In addition, high efficacy was observed when triclabendazole, the current drug of choice against human fascioliasis, was administered to patients remaining Fasciola positive following artemether treatment. Concluding, monotherapy with artemether does not represent an alternative to triclabendazole against fascioliasis, but its role in combination chemotherapy regimen remains to be investigated. Fasciola hepatica and F. gigantica, is of considerable veterinary and public health importance Fasciola spp. appear to be emerging or re-emerging in several parts of the world Fasciola infection include cholangitis, cholecystitis, jaundice, and biliary colic Fascioliasis, a zoonotic disease caused by a liver fluke infection of the species Fasciola infections have been reduced in recent years, explained by control measures put forth by the Egyptian governorates, including triclabendazole administration In Egypt, fascioliasis is an important clinical problem, particularly among school-aged children living in rural areas of the Nile Delta F. hepatica when artesunate and artemether were administered at single oral doses  8 weeks postinfection Fasciola spp. were incubated with an artemisinin derivative (50\u2013100 \u00b5g/ml) in vitroF. hepatica infections in sheep There is a need to develop new fasciocidal drugs. Several studies have documented that the artemisinins , which have become the most important antimalarial drugs, particularly when deployed as artemisinin-based combination therapy (ACT) Fasciola spp. infection. The study was carried out in a Fasciola-endemic area of Egypt, where Schistosoma mansoni co-exists, but malaria is absent.The aim of the present study was to assess the efficacy and safety of oral artemether, adhering to two different malaria treatment regimens Ethical clearance was obtained from the Theodor Bilharz Research Institute , the Ministry of Health and Population , and the Ethics Committee of Basel, Switzerland . The trial is registered with Current Controlled Trials (reference no. ISRCTN10372301). Written informed consent was obtained from eligible study participants or parents/legal guardians from individuals aged below 16 years.Fasciola-infected patients. Twenty individuals were assigned to each study, following recommendations for pilot studies of at least 12 patients per treatment The study was designed as an interventional, open-label, non-randomized, proof-of-concept trial, consisting of two separate single-arm studies, to evaluate the efficacy and safety of two artemether regimens in the treatment of asymptomatic Fasciola egg-negative after treatment, who were egg-positive at study enrollment) and egg reduction rate  egg output after treatment divided by the GM of the same individuals before treatment, multiplied by a factor 100) of Fasciola infection, 28 days after the final dosing. Incidence of adverse events, monitored up to 2 days after the final dosing, was used as secondary outcome measure.The primary end points were cure rate  and 50 mg tablets (study 2) was purchased from Kunming Pharmaceutical Cooperation . The following two treatment schemes were investigated: (i) 6\u00d780 mg over 3 consecutive days (study 1) and (ii) 3\u00d7200 mg within 24 h (study 2). Treatment was supervised by a physician with date and precise time of drug administration recorded. Patients were observed for 1 h to ensure retention of medication. In case of vomiting or any treatment-related adverse events, a second dose of artemether was administered.Fasciola egg-negative following artemether administration received 10 mg/kg triclabendazole. The triclabendazole dosage, according to the patients' weight, was calculated in half-tablet increments with a maximum of 2.5 tablets (625 mg). In case of triclabendazole treatment failures (assessed in study 2), patients were provided two doses of 10 mg/kg of triclabendazole given on subsequent days according to manufacturer's instructions.Triclabendazole  was the product of Novartis . Patients who failed to become F. hepatica and/or F. gigantica eggs. Stool collection containers were labeled with patient's name and a unique identifier (ID). Filled containters were transfered to a laboratory for diagnostic work-up. Two additional stool samples were collected on consecutive days among participants who were found with Fasciola eggs in their feces. In addition, a blood sample was collected before drug administration to examine hematologic parameters, liver, and kidney functions.Several weeks before conducting a parasitological baseline survey, the health directorate of Beheira (study 1) and Alexandria governorate (study 2) were informed about the objectives, procedures, and potential risks and benefits. After written informed consent was obtained, participants were asked to provide a stool sample in order to screen for the presence of At enrollment a full clinical examination was carried out to assess participants' general health status. Exclusion criteria were: (i) age below 5 years, (ii) pregnancy, (iii) major systemic illnesses , and (iv) recent history of anthelmintic treatment . Patients meeting our inclusion criteria were treated with artemether, which was administered over 3 consecutive days (study 1) or within 24 h (study 2).Adverse events were monitored on each treatment day and for 24\u201348 h following the final dosing. Participants were asked to report any potential drug-related signs and symptoms using a standardized questionnaire. Full clinical examinations were performed on all participants. Adverse events were graded  and recorded. Therapy was offered to patients presenting with adverse events, as judged by the study physician.Fasciola eggs in their stool following artemether administration were treated with 10 mg/kg triclabendazole. In study 2, stool samples were collected from triclabendazole-treated patients 28 days posttreatment over 3 consecutive days and CRs and ERRs were determined. Those patients who remained Fasciola positive were retreated with a double dose of triclabendazole (20 mg/kg given 24 h apart) Five and 28 days posttreatment, blood samples were collected for clinical chemistry analyses. The final parasitological assessment started on day 28 posttreatment: stool samples were obtained from all study participants over 5 consecutive days. Patients found with Fasciola eggs, all stool samples were processed shortly after collection using the Kato-Katz technique S. mansoni and soil-transmitted helminths  was also determined and recorded for each participant individually. Each slide was examined independently in a blind manner by two microscopists. For quality control, several slides were re-examined by a senior staff. For confirmation of Fasciola and other helminth eggs, at baseline the merthiolate-iodine formaldehyde (MIF) concentration technique For detection and quantification of g for 10\u201315 min. All blood specimens were analyzed on the day of collection.Laboratory investigations of blood included total leukocyte count, hemoglobin, eosinophilic count, alanine transpeptidase (ALT), aspartate transpeptidase (AST), alkaline phosphatase (ALP), gamma glutamyl transpeptidase (GGT), total serum bilirubin, blood urea, and serum creatinine. The blood specimens were collected into gel serum tubes  and EDTA tubes (for hematology variables). Blood specimens collected into gel tubes were centrifuged at 1800\u20132000 Fasciola eggs before treatment and absence of eggs at study end. To determine infection intensity, the number of Fasciola eggs per Kato-Katz thick smear (41.7 mg of stool) was multiplied by a factor 24 to obtain eggs per gram of stool (EPG). Fecal egg counts (FECs) of multiple slides per individual were averaged, using the arithmetric mean. To calculate the reduction in infection intensity, individual egg counts were logarithmically transformed (log (count + 1) and the GM expressed as the antilogarithm of the mean. The ERR was calculated as [1 - GM FEC after treatment divided by GM FEC at admission multiplied by a factor 100]. Although infection intensity thresholds are currently lacking for infections with FasciolaS. mansoni infection t-test and the Kruskal-Wallis tests were employed to compare the clinical parameters before and after treatment.Data were entered using EpiData version 6.04 . CR was calculated as proportion of individuals excreting Fasciola-positive. Two patients were excluded . Twenty patients  were included in study 1 (Of 584 villagers and 51 school-aged children screened in El-Haddad El-Bahary village (study 1), 22 individuals were found  study 1 .Fasciola-positive subjects were identified. Of these, 17 patients    were incFasciola FECs in the two studies were 28.3 EPG and 29.1 EPG  and 8% (3\u00d7200 mg artemether) . Treatment with artemether over 3 consecutive days resulted in ERRs of 63% (67% for light infections and 55% for infections \u2265100 EPG). The individual pretreatment and posttreatment FECs are presented in Data from all patients were included in the analysis, as no patient was lost to follow-up . CRs achieved with 6\u00d780 mg and 3\u00d7200 mg artemether were 35% and 6%, respectively . Fisher'S. mansoni. At treatment follow-up, three out of the five patients in each study were recorded egg-free (CR: 60%).In each of the two studies, five patients were co-infected with Fasciola-positive after treatment with 3\u00d7200 mg artemether were administered a single 10 mg/kg oral dose of triclabendazole. CR and ERR were 69% and 94%, respectively; significantly higher than CR  and ERR (P<0.001) observed following treatment with 3\u00d7200 mg artemether. The infection intensity did not influence the treatment outcome (data not shown). Four out of five patients who were still passing Fasciola eggs following a single triclabendazole dose were provided a double dose of triclabendazole and the respective CR and ERR were 75% and 96%.Sixteen patients who were still found There were no noteworthy effects of artemether on the liver enzymes and renal function parameters, with the exception of a statistically significant increase in GGT 5 days after the final dosing of artemether (6\u00d780 mg) . FollowiThe comparison between pre- and posttreatment values of liver and renal function and hematological parameters showed no significant differences following administration of triclabendazole (10 and 20 mg/kg)  apart frBoth artemether regimens were well tolerated and no participant required special medical follow-up. As summarized in Abdominal pain was more often observed after treatment with triclabendazole  than aftFasciola spp. observed in the two study sites  was similar to previous studies in these areas While the veterinary importance of fascioliasis cannot be overemphasized, this zoonotic disease is also of considerable and growing public health importance, yet it often remains neglected. A major challenge is that treatment is restricted to a single drug, i.e., triclabendazole, which is registered for human use only in Ecuador, Egypt, France, and Venezuela F. hepaticavs. 63%) is striking, yet difficult to explain. Since the half life of artemether is very short (<1 h) in vitro drug sensitivity and pharmacokinetic studies are required to further elucidate this issue. It is interesting to note that the CRs (nil vs. 54%) and ERRs (55% vs. 67%) were higher in patients classified as lightly infected compared to moderate/heavy infections in the 6-dose regimen. A similar trend was observed in a recent study, which assessed the efficacy of an artesunate-sulfalene plus pyrimethamine combination in S. mansoni-infected school-aged children in Kenya: significantly higher CRs were observed in children harboring a light S. mansoni infection compared to moderate and heavy infections Our study failed to extend promising findings obtained with the artemisinins in rats experimentally, and sheep naturally, infected with Fasciola spp. and S. mansoni. Moderate CRs were observed against S. mansoni (60%) regardless of the selected treatment regimen. This finding is in line with previous studies, which documented low-to-moderate efficacies of an artemisinin monotherapy in the treatment of chronic infections with Schistosoma spp. S. mansoni and S. haematobium. Although promising results were obtained in small exploratory trials with the artemisinins against schistosomiasis, larger clinical trials could not confirm these findings, and hence praziquantel remains the drug of choice In the present study, five participants in each of the two studies were co-infected with Fasciola spp. to triclabendazole might play a role in the somewhat lower efficacies observed here compared to previous studies. Participants treated with triclabendazole showed a higher incidence of abdominal pain compared to those treated with artemether, which might be related to the higher efficacy of triclabendazole (dying worms).CRs of 69% and 75% were observed in patients treated with triclabendazole at 10 mg/kg and 20 mg/kg, respectively. The observed efficacy is slightly lower than a calculated overall CR of 83% following 10 mg/kg and reported CRs ranging from 93 to 100% following a double dose of triclabendazole F. hepatica-infected rats F. hepatica-infected rats In conclusion, significantly higher CRs and ERRs were observed with triclabendazole when compared to artemether, the latter administered following two malaria treatment schedules. Hence, triclabendazole remains the drug of choice against fascioliasis. In view of threatening triclabendazole resistance development, concerted efforts are required, including structure-activity relationships with the synthetic peroxides in Checklist S1CONSORT Checklist.(DOC)Click here for additional data file.Protocol S1Trial Protocol.(DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria from chloroquine to artemether-lumefantrine (AL) in 2005. Since then, no information on AL efficacy has been reported. With evidence of resistance to artemisinin derivatives in adjacent Cambodia, there has been a concern as to AL efficacy. Monitoring of AL efficacy would help the Lao Government to make decisions on appropriate malaria treatment.The Lao Government changed the national policy for uncomplicated The efficacy of a three-day, twice daily oral artemether-lumefantrine for the treatment of uncomplicated falciparum malaria in Xepon District, Savannakhet Province, southern Laos was studied over 42\u2009days follow-up. This was part of a trial of thiamin supplementation in falciparum malaria.P. falciparum enrolled in the trial of thiamin treatment, 549  were included in this study. The per protocol 42-day cure rates were 97% (524/541) . By conventional intention-to-treat analysis, the 42-day cure rates adjusted for re-infection, were 97% (532/549) . The proportion of patients who remained parasitaemic at day 1 after treatment was significantly higher in children [33% (116/356)] compared to adults [15% (28/192)] (p\u2009<\u20090.001) and only one adult patient had detectable parasitaemia on day 2. There were no serious adverse events. Potential side effects after treatment were reported more commonly in adults (32%) compared to children (15%) (p\u2009<\u20090.001). Patients with recrudescent infections were significantly younger, had longer mean time to fever clearance, and had longer median time to parasite clearance compared to those who were cured.Of 630 patients with P. falciparum parasites should appear.The current nationally-recommended anti-malarial treatment (artemether-lumefantrine) remains highly efficacious for the treatment of uncomplicated falciparum malaria five years after introduction in Laos. Regular monitoring is required in case artemisinin-resistant ISRCTN85411059. Plasmodium falciparum malaria has declined recently in northern and central Lao PDR (Laos), malaria remains an important cause of morbidity and mortality in southern Laos, with an estimated incidence of 4.7 to 23.5/1,000 population between 2006-2008 10\u2009mg immediately after anti-malarial drugs, followed by 10\u2009mg daily for 7\u2009days followed by 5\u2009mg daily until day 42), or an identical placebo.The clinical trial of thiamin supplementation in falciparum malaria conducted among patients with P. falciparum infection or mixed Plasmodium species infections, with a history of fever, were willing and able to comply with the study protocol for the duration of the 42\u2009days follow up, and had not taken a full course of any anti-malarial drugs in the previous three days. Patients were excluded if they had a history of hypersensitivity to thiamin or artemether-lumefantrine, presented with intercurrent non-malarial illness or any condition, which in the judgement of the investigator would place the subject at undue risk or interfere with the results of the study, and if they had clinically apparent suspected thiamin deficiency (beriberi) had recrudescent infections. One child with P. falciparum reappearance had undetermined PCR result and, considering this child as having a recrudescent infection, the overall 42-day cure rates per protocol, excluding patients who were lost to follow up, withdrew consent, had persistent vomiting, or re-infection, were 97% (524/541) . When the patients with repeated vomiting, who were lost to follow up or were withdrawn (censored at the time last seen), considering the child with undetermined PCR result as recrudescence, and regarding recrudescent infections as failures, the 42-day cure rates (95%CI) by survival analysis (Kaplan-Meier) were 99.0% (98.3 \u2013 99.4%) for children and 99.8% (99.0 \u2013 99.9%) for adults. For the PCR adjusted survival analysis, the cure rates (95%CI) were 99.0% (98.3 \u2013 99.4%) and 99.8% (99.0 \u2013 99.9%) for children and adults, respectively. By conventional ITT analysis, the overall 42-day cure rates, adjusted for re-infection, were 97% (532/549) . Of 17 patients considered as having recrudescent infection (15 children and two adults), 15 (13 children and two adults) had parasite recurrence at or before day 28. Similarly, the 28-day cure rates by ITT analysis were 344/357 (96%) for children and 190/192 (99%) for adults. The median (range) interval to recrudescent falciparum infections was 21 (14\u201342) days. By ITT and per protocol analyses, the 42-day PCR uncorrected cure rates for children and adults were 323/357 (90%) vs 183/192 (95%); (p\u2009=\u20090.04), and 323/352 (92%) vs 183/189 (97%) (p\u2009=\u20090.02), respectively. Of 129 children aged <5\u2009years, six had recrudescent infection and one had undetermined PCR result. If the patient with undetermined PCR result was considered a recrudescent infection, the 42-day PCR corrected cure rates among children\u2009<\u20095\u2009years were 95%. There were 18 episodes of vivax malaria during the follow-up period, of which 17 were in children (p\u2009<\u20090.05). The median (range) interval to P. vivax appearance was 28 (21\u201342) days (TableOf 43 (7.8%) patients with subsequent 0\u2009C] than those who were cured . Patients with recrudescent infections had a significantly longer mean FCT  than those patients without . Patients with recrudescent infections also had a significantly lower haematocrit at days 3, 14, 21, 28, and 35 after starting therapy than those without recrudescence detected . The proportion of the patients reporting a past history of malaria was significantly lower in the patients with recrudescent infection (6%) compared to those without (28%) (p\u2009=\u20090.043). In a multiple logistic regression analysis, admission high temperature , longer fever clearance times , and lower haematocrit at day 28  were significantly associated with treatment failure.Patients with recrudescent infections were significantly younger [median (range) age 5 (2\u201325) years], than those whose infections were treated successfully . Considering the child with undetermined PCR result as recrudescent infection, the proportion of treatment failures adjusted for re-infection by PCR among children was 4.2% (15/357 patients) compared with 1.0% (2/192 patients) among adults (p\u2009=\u20090.042). The admission mean (95%CI) body temperature was significantly higher in patients with recrudescent infections [39.1 (38.4\u201339.8) The possible adverse events following treatment with AL are shown in TableThis large clinical trial shows that artemether-lumefantrine retains excellent efficacy in the treatment of uncomplicated falciparum malaria at one site in southern Laos in both children and adults with 42-days PCR-corrected cure rates of 96% and 99%, respectively. These results are similar to those described in a comparable trial conducted in 2002-3 in nearby Phalanxay District (~60 Km west of Xepon), in which there was a 42-days cure rate of 97% in children and adults. As prevP. vivax appearance rates than adults. A study comparing oral chloroquine and sulphadoxine-pyrimethamine for the treatment of uncomplicated falciparum malaria, before the introduction of AL, found treatment failure rates among Lao children to be 4.9 times higher than those among adults[Patients\u2019 background immunity contributes substantially to the treatment response of uncomplicated malaria in Laos,27. In tg adults. The cong adults,9. Anti-g adults. Anti-mag adults.P. falciparum has not spread to this area, of southern Laos. However, regular monitoring of the clinical efficacy of artemisinin-derivatives in southern Laos, especially closer to the Cambodian border, is required.The evidence, both from a trial of artesunate monotherapy with a mean (95%CI) PCT (hours) of 23.2 (21.2-25.3) and 42-day PCR-corrected cure rate of 100%, and froP. falciparum parasites.The Lao nationally recommended anti-malarial drug (artemether-lumefantrine) remains highly efficacious for the treatment of uncomplicated falciparum malaria five years after its introduction. However, regular monitoring of the efficacy of artemisinin derivatives is essential as early warning of the potential spread of artemisinin-resistant The authors declare that they have no competing interests.MM designed the study, recruited and followed up the patients, analysed data and drafted paper. MK and OC designed the study, recruited and followed up the patients, and revised the paper. MI designed the study, performed the molecular genetic studies, and revised the paper. TP, BH, VV, SP and NJW designed the study and revised the paper. PNN designed the study, analysed data, drafted and revised the paper. All authors read and approved the final manuscript."}
+{"text": "The An open-label, randomized controlled non-inferiority trial compared the genotyping adjusted day 42 cure rates of ASAQ FDC (ASAQ Winthrop\u00ae) to AL (Coartem\u00ae) in 300 children aged six to 59 months with uncomplicated falciparum malaria. Inclusion was between December 2008 and May 2009. Randomization (1:1) was to a three-day observed oral regimen . Day 7 desethylamodiaquine and lumefantrine blood-concentrations were also measured.The day 42 genotyping-adjusted cure rate estimates were 97.3% [95% CI: 91.6-99.1] for ASAQ and 94.2% [88.1-97.2] for AL . The difference in day 42 cure rates was \u22123.1% [upper limit 95% CI: 1.2%]. These results were confirmed by observed proportion of patients cured at day 42 on the per-protocol population. Parasite clearance was 100% (ASAQ) and 99.3% (AL) on day 3. The probability to remain free of re-infection was 0.55 [95% CI: 0.46-0.63] (ASAQ) and 0.66 [0.57-0.73] (AL) (p\u2009=\u20090.017).Both ASAQ and AL were highly efficacious and ASAQ was non-inferior to AL. The proportion of patients with re-infection was high in both arms in this highly endemic setting. In 2010, ASAQ FDC was adopted as the first-line national treatment in Liberia. Continuous efficacy monitoring is recommended.ISRCTN51688713, ISRCTN40020296.The protocols were registered with Current Controlled Trials, under the identifier numbers  Plasmodium falciparum malaria remains a serious public health problem especially for the African continent  were censored at the last visit performed according to schedule for mITT/KM analysis. Pre-mature study discontinuation due AE were excluded from the PP/% analysis , or intake of concomitant medication (except for anti-pyretics) during follow-up if medication is of known anti-malarial activity. Results were reported by treatment group as allocated. Treatment outcomes were classified using standard definitions: adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late parasitological failure (LPF), and late clinical failure (LCF)  (AL) (mITT/KM) Table\u00a0. GenotypBlood concentrations of amodiaquine (AQ), desethylamodiaquine (DEAQ) (ASAQ arm) and lumefantrine (LF) (AL arm) were assessed from day 0 (baseline) and day 7 blood samples. Amodiaquine-intake shortly before study inclusion (protocol violation) was evident for six patients (ASAQ arm) who had quantifiable day 0 AQ levels (range: 3.6 \u2013 34 ng/ml). Three of these had also high DEAQ levels (>200 ng/ml). Seventy-eight patients had low DEAQ blood concentrations on day 0 (range: 5.0 -142 ng/ml), indicating a previous treatment with AQ (no violation). On day 7, three patients had DEAQ values below the LOQ (<2.5 ng/ml), suggestive of sub-optimal drug intake or mal-absorption. All three patients had completed the drug intake according to schedule and had day 42 efficacy outcome ACPR. Five patients also had detectable AQ on day 7, indicative of AQ intake before sample collection (protocol violation) (range: 2.6-4.2 ng/ml). No LF blood concentrations were detected in the AL arm at baseline. On day 7, 33 patients had non-quantifiable LF concentrations . These were set as 100 ng/ml (=LOQ/2) by convention for all blood concentration analyses, since the LOQ was considered very high.P. falciparum recurrences combined were not significantly different  when compared to ACPRs . The median day 7 LF concentration was 310 ng/ml . The recrudescences in the ASAQ arm had day 7 DEAQ blood concentrations of 242, 401 and 688 ng/ml, respectively. Recrudescences in the AL arm had day 7 LF concentrations of 271, 447 and 515 ng/ml, respectively . Comparison of day 7 blood concentrations between patients with day 42 efficacy outcome ACPR versus recrudescence revealed no significant differences for DEAQ  or LF , respectively  and artemether-lumefantrine (AL) among 6\u201359 month old children with uncomplicated falciparum malaria in Nimba County, a highly Pf malaria endemic area in Northern Liberia. The day 42 genotyping-adjusted cure rate estimates of ASAQ and AL reached 97.3% and 94.2% respectively (mITT / KM). Both treatments were well above the WHO-recommended 90% threshold for treatments in use [Since 2003, AS + AQ loose dose has been adopted as first-line treatment for uncomplicated falciparum malaria in Liberia. Published information on the efficacy of AS + AQ or other forms of ACT in the country are currently limited to only one observational study on the efficacy of artemether-lumefantrine in Tubmanburg and Harper region . We repos in use . ASAQ FDs in use .Overall, efficacy results obtained in the present studies were very good for both ACTs. This was also encouraging for the utility of ASAQ FDC in a geographical setting where amodiaquine monotherapy and AS + AQ  were used for several years. Notably however, anecdotal reports suggested that the prescription of AS + AQ loose dose provided the opportunity to disregard the amodiaquine compound, referring to patients\u2019 and providers\u2019 concern on tolerability related to prescriptions of higher dose amodiaquine monotherapy in the past . The preThe proportion of parasitaemic patients on day 3 has been reported as an interesting indicator for monitoring artemisinin resistance . AlthougP. falciparum re-infection rate was high in the present study, emphasizing the burden of malaria among the non-immune\u2009<\u20095 year olds in Nimba County. Re-infection rates were higher in the ASAQ arm than the AL arm, and AL conferred a longer secondary prophylactic effect than ASAQ with a significantly lower median time to re-infection in the ASAQ arm. The current WHO treatment protocol emphasizes the curative effect of the anti-malarial treatment, i.e. a fast elimination of the parasite [The overall parasite . A potenparasite ,31, and In 2001 high resistance to chloroquine and sulphadoxine-pyrymethamine (SP), the respective first- and second-line treatments in the country at the time, were reported in the Harper region, Liberia . A seconASAQ and AL were both highly efficacious treatments for uncomplicated falciparum malaria in Nimba County. These findings support both drugs as treatment options. Since the end of 2010, ASAQ FDC was adopted as national first-line treatment in Liberia. As for all malaria endemic settings, anti-malarial treatment monitoring should continue on a regular basis, ideally including recently developed early indicators of emerging artemisinin resistance.The authors declared no competing interests.BS overall trial coordination, participation in study design and protocol, data analysis plan, study documents, drafting of the manuscript. PV field trial coordination, medical coordination, trial team supervision, supervision of data collection. EB support study design, coordination of data management, data analysis plan, data analysis, revision of the manuscript. CM field laboratory coordination, laboratory standard operating procedures and data collection. RS field coordination during study preparation, writing of administrative and clinical standard operating procedures, preparation of study site, staff training, trial implementation. LP data management, support data analysis plan, data analysis, revision of the manuscript. MD and JB coordination and interpretation of the malaria parasite genotyping. TS support field trial coordination, team supervision and data collection. YMZ and JJJ technical support to all study steps, participation in study implementation and training. EC support study initiation and study design, technical support during study conduct. PH anti-malarials baseline serum analysis. VJ development and conduct of blood concentration analyses of artemisinin-partner compounds. GC technical advise to protocol development and study preparation, support field training and study conduct, review of study documents, coordination of study monitors and data monitoring committee, revision of the manuscript. JR. study initiation, technical and scientific advise to study protocol-, study preparation and conduct, review of study documents, revision of the manuscript. EAA study initiation, study design and protocol, scientific and medical advise to all steps of the study, revision of study documents, support data analysis plan, revision of the manuscript. PJG study initiation, study design and protocol, scientific and medical advise to all steps of the study, review of study documents and medical review of adverse events, support data analysis plan, revision of the manuscript. All authors read and approved the manuscript.Distribution of patients\u2019 age and weight by weight-based drug-dosing group and by study arm (mITT population).Click here for file"}
+{"text": "Plasmodium falciparum drug resistance. Artemether-lumefantrine (AL) and artesunate-amodiaquine (AA) are efficacious regimens that have been widely adopted in sub-Saharan Africa. However, most study designs ignore the effects of these regimens on peripheral parasitaemia in the first 24 hours of therapy. The study protocol was designed to evaluate more closely the early effects and the standard measures of efficacies of these two regimens.Combination treatments, preferably containing an artemisinin derivative, are recommended to improve efficacy and prevent Plasmodium falciparum.In an open label, randomized controlled clinical trial, children aged 12 months to 132 months were randomized to receive AL  or artesunate (4 mg/kg daily) plus amodiaquine (10 mg/kg daily) for three days. Peripheral blood smears were made hourly in the first 4 hours, 8 h, 16 h, 24 h, and daily on days 2-7, and on days 7, 14, 21, 28, 35, and 42 for microscopic identification and quantification of A total of 193 children were randomized to receive either AL (97) or AA (96). In children that received both medications, early response of peripheral parasitaemia showed that 42% of children who received AL and 36.7% of those who received AA had an immediate rise in peripheral parasitaemia (0-4 h after treatment) followed by a rapid fall. The rise in parasitaemia was significant and seems to suggest a mobilization of asexual parasites from the deep tissues to the periphery. Days 3, 7, 14, 28, and 42 cure rates in the per protocol (PP) population were > 90% in both groups of children. Both drug combinations were well tolerated with minimal side effects.The study showed the high efficacy of AL and AA in Nigerian children. In addition the study demonstrated the mobilisation of asexual parasites from the deep to the periphery in the early hours of commencing ACT treatment in a subset of patients in both study groups. It is unclear whether the early parasite dynamics discovered in this study play any role in the development of drug resistance and thus it is important to further evaluate this discovery. It may be useful for studies investigating delay in parasite clearance of artemisinin derivatives as a way of monitoring the development of resistance to artemisinin to assess the early effects of the drugs on the parasites. The World Health Organization (WHO) recommends artemisinin-based combination therapy (ACT) for the treatment of malaria in countries experiencing resistance to anti-malarial drug monotherapy . Artemisin vivo efficacies of ACT be evolved in view of the presence of artemisinins in both arms of studies evaluating ACT.The guidelines for the comparative studies of anti-malarial therapies have largely remained unchanged since the time of widespread use of chloroquine and other monotherapies. The standard endpoints of efficacy studies remain the evaluation of events around the following days of therapy; days 3, 7, 14, 28, and more recently day 42. Most studies on ACT using these standard endpoints have consistently reported similarities in efficacies of ACT with cure rates above 90% . This isnd day of therapy). The early and late efficacy of these two different forms of ACT on peripheral asexual falciparum parasitaemia was assessed.In this study, the comparative efficacy and safety of two different forms of ACT [artesunate-amodiaquine (AA) and artemether-lumefanthrine (AL)] was evaluated using a modified protocol to evaluate two time points of efficacy; early effects (\u2264 72 hours of therapy) and late effects  or water (AA) before administration. Study drugs were administered supervised by a nurse or a physician. The enrolled patients were admitted to hospital ward for three days for observation of drug administration and clinical states. After administration of drugs, patients were observed for 30 min and the dose was re-administered if the patient vomited within the period. Febrile children with axillary temperature \u226538\u00b0C were exposed, tepid-sponged and given oral paracetamol. Children were withdrawn for the following reasons: withdrawal of consent, protocol violation, repeated vomiting or loss to follow-up.P. falciparum under a light microscope at 1,000\u00d7 magnification. Blood spots were also collected on filter paper for molecular analysis at entry and daily for the first 7 days, on days 14, 21, 28, 35 and 42.Study participants were followed up daily on days 0-7 and on days 14, 21, 28, 35 and 42. At each visit children were physically examined and their vital signs recorded. In addition, thick blood films were prepared from finger prick blood samples. Thick blood films were prepared hourly for the first 4 hours, at 8, 16 and 24 hours (day 0) following therapy. Subsequently thick films were made daily on days 1-7, 14, 21, 28, 35 and 42. These were stained with 10% fresh Giemsa stain and examined for the presence of asexual forms of Children who were not brought to the clinic on scheduled days were visited at home for the follow-up procedures. Each child and the parent or guardian was questioned about symptoms observed following commencement of therapy. Children who failed treatment were re-treated with artesunate-mefloquine.Efficacy was comparatively evaluated using two time points. Early drug effects were classified as effects occurring between 0 h and < 72 h of treatment. Late effects were those occurring between > 72 h and day 42 post-treatment.Early treatment failure (ETF), defined as development of danger signs of malaria or severe malaria on post-treatment days 1, 2 or 3, in the presence of parasitaemia, parasitaemia on day 2 higher than day 0 count irrespective of axillary temperature, parasitaemia on day 3 which is > 25% of count on day 0 and parasitaemia on day 3 with axillary temperature 37.5\u00b0C .Late clinical failure (LCF) was defined as development of danger signs or severe malaria after day 3 in the presence of parasitaemia without previously meeting any of the criteria of ETF and presence of parasitaemia and axillary temperature of 37.5\u00b0C on any day from day 4 to day 42, without previously meeting any of the criteria of ETF.Late parasitological failure (LPF) was defined as presence of parasitaemia on any day from day 7 to day 42 and axillary temperature of < 37.5\u00b0C without previously meeting any of the criteria of ETF or LCF.Adequate clinical and parasitological response (ACPR) was defined as absence of parasitaemia on day 42 irrespective of axillary temperature without previously meeting any of the ETF, LCF or LPF.P. falciparum infection in the study were characterized on the basis of the fragment size of alleles of msp-2 after amplification by PCR. A recrudescent infection was defined as the occurrence of the same or a subset of the alleles at each of the families (FC27 or IC1/3D7) of msp-2 in the primary and post-treatment samples. A lack of allelic identity in the two families of msp-2 in matched primary and post-treatment samples indicated a newly acquired infection. Infections were defined as polyclonal if parasites in matched primary and post-treatment samples from the same patient showed more than one allele of FC27 or IC1/3D7 families of msp-2. If an isolate had one allele at each of the families, the clone number was taken to be one.Discrimination between recrudescence and re-infections in treatment failures was done using molecular analysis ,19. IsolAn adverse event (AE) was defined as signs, symptoms or abnormal laboratory finding not present at enrolment, but occurring during follow-up, or being present at day 0 and becoming worse during follow-up despite clearance of parasitaemia. All AEs were monitored and recorded. Assessment was by asking about the progress of presenting symptoms and new symptoms noticed during follow-up, physical examination, and or laboratory evaluation.Data collected were recorded in case record forms and entered into Epi-info version 6.04 database for analysis. SPSS version 16 was also used for analysis. Efficacy analysis of the data was done for the intention to treat (ITT) and per protocol (PP) populations. All children enrolled into the study were considered ITT. Patient who completed the study without violating the study protocol were considered as PP population. Parasite densities are reported as geometric mean parasite densities. The means and standard deviations (\u00b1 SD) of normally distributed data were compared using Student's t-test and analysis of variance (ANOVA). Proportions were compared by chi-square. Numerical values are given as means \u00b1 SD, p-values < 0.05 was taken as statistically significant.A total of 1,888 children were screened for the study and 193 consented after meeting the inclusion criteria. Ninety-seven (97) children were randomized to receive AL and 96 were randomized to receive AA. Ten (10) of the children enrolled (5.2%) were lost to follow-up: four patients from AL group and six (6) from AA group , vomiting (51%) and abdominal pain (35%). Ninety-three of 97 (95.9%) patients treated with AL completed the study while 90/96 (93.7%) of those that receive AA completed the study (per protocol population) , respectively, while mean parasite clearance times (PCT) were 28.6 \u00b1 18.4 hours AL and 24.0 \u00b1 15.4 hours for AL and AA respectively (p = 0.067). Cure rates by day 3 in both groups were 100%  for AL and AA were 29.9 \u00b1 18.4 vs. 28.9 \u00b1 12.7 hours, respectively (p = 0.630). The 24 and 48-h parasite reduction ratios (PRR) were 11 \u00d7 10Analysis of drug effects on hourly changes in peripheral asexual parasitaemia in the first 24 hours following treatment showed that there were two sub-groups; those in whom parasitaemia increased in the first 4 hours after the first dose of the medications and those that did not show any increase in peripheral parasitaemia Figure . Forty-tIn the AL and AA groups, peripheral asexual parasite changes in the first 24 hours and at others time points of the study were similar between the two sub-groups described above. Geometric means of parasitaemia at 0 hour and 1 hour in the two subgroups of AL and AA in those that showed no increase in parasitaemia in the first hour following therapy were 99,028 parasites/\u03bcL and 50,075 parasites/\u03bcL (for AL), and 72,927 parasites/\u03bcL and 33,167 parasites/\u03bcL (for AA) respectively . Geometric means of parasitaemia at 0 hour and 1 hour in the two subgroups of AL and AA in those that showed increase in parasitaemia in the first hour following therapy were 53,001 parasites/\u03bcL and 60,744 parasites/\u03bcL (for AL) and 63,424 parasites/\u03bcL and 106,185 parasites/\u03bcL (for AA) [p-values 0.698 and 0.248]. PRR in the first 24 hours and 48 hours were similar in both groups  population: Day 14, 28, and 42 cure rates for AL and AA were 100%, 97.9%, 92.8%, and 100%, 96.9%, 93.8% respectively. PCR corrected cure rates on day 28, 42 for AL and AA were 97.9%, 94.8%, and 97.9%, 93.8% respectively. Per-protocol (PP) population: Day 14, 28, and 42 cure rates for AL and AA were 100%, 96.8%, 92.5%, and 100%, 97.8%, 92.2% respectively. PCR corrected cure rates on day 28, 42 for AL and AA were 97.8%, 93.5%, and 97.8%, 93.3% respectively  was formally introduced in Nigeria in 2005 following the progressive decline in efficacy of chloroquine and sulphadoxine-pyrimethamine. The two most widely adopted ACT regimens are AL and AA -23, althP falciparum in the Nigerian population has a higher tendency to sequestration, or that a proportion of the children have a tendency to having more of sequestered parasites. These postulates remain to be fully explored.The effects of the study drugs on asexual parasites immediately following treatment uncovered an unexpected finding. Two patterns of asexual parasite responses occurred; children that experienced an acute rise in peripheral parasitaemia hours after commencing treatment and those that did not. This effect was uncovered due to frequent blood sampling for malaria parasite evaluation in the first 24 hours of the study. Daily sampling, as recommended by the WHO, would have missed this important finding because these effects occurred within 24 hours of initiating treatment. Parasitaemia reached a peak at 1 hour in the sub-group of children that had an initial rise in peripheral parasitaemia after commencing therapy. Analysis of demographic and clinical parameters of children in these two sub-groups Table  did not et al [Studies in the past have described increase in parasitaemia 6 to 12 hours after administration of parenteral anti-malarial drugs and have attributed this phenomenon of natural progression of sequestered parasites on account of a lag period between drug administration, absorption and onset of parasitocidal effects of the drugs . The laget al , in a reThe clinical efficacy of artemisinin is known to be characterized by an almost immediate onset and rapid reduction of parasitaemia . HoweverBased on the observations of potential mobilization of parasites from the tissues to the periphery in this study, it is possible that AA and AL may have a property of rapidly releasing parasites from sequestration sites in the deep tissues. It is not clear if the early parasite dynamics discovered in this study play any role in the development of drug resistance and thus it is important to further evaluate this finding. Studies investigating delay in parasite clearance of artemisinin derivatives as a way of monitoring the development of resistance to artemisinin should also assesses the early effects of the drugs on the parasites.Plasmodium falciparum parasitaemia in the treatment of acute uncomplicated malaria in Nigerian children. Therapeutic responses were rapid, and efficacy at day 42 was above 90% for both combinations. Both drugs were well tolerated with minimal side effects.The two forms of ACT studied, AL and AA, had similar efficacies and effects on peripheral asexual The study also revealed that asexual parasite dynamics in the early hours of initiating therapy showed two patterns; those who had an early rise in peripheral parasitaemia immediately after treatment and those who did not. The significance of this finding remains to be further explored and explained.The authors declare that they have no competing interests.HCT, GOG, AS and AMJO, conceived and designed of the study. OSM, TO, HCT, GOG, OAF and AS participated in patients enrolment, samples collection and data analysis. OSM and HCT drafted the manuscript. All the authors read, made suggestions and approved the manuscripts that were submitted for review and publication."}
+{"text": "Tanzania adopted artemether-lumefantrine (AL) as first-line drug for uncomplicated malaria in 2006. Recently, there was an anecdotal report on high malaria recurrence rate following AL treatment in in the (urban and peri-urban), western part of Tanzania. The current report is an exploratory study to carefully and systematically assess AL efficacy in the area.Between June and August 2011, a total of 1,126 patients were screened for malaria, 33 had malaria, of which 20 patients met inclusion criteria and were enrolled and treated with standard dose of AL as recommended in the WHO protocol. Treated patients were followed up for 28 days to assess treatment responses. Before treatment (Day 0) and post-treatment (Day 7) plasma lumefantrine levels were determined to assess prior AL use and ascertain parasites exposure to adequate plasma leveles of lumefantrine, respectively.The cure rate was 100%. All Day 0 plasma lumefantrine were below HPLC detectable level. The median Day 7 lumefantrine concentration was 404, . Six out of 20 patients (30%) were gametocytaemic and all cleared gametocytes by Day 14. One patient showed an increase in gametocytes from four on Day 0 to 68, per 500 WBC on Day 2.Plasmodium falciparum malaria. The elevation of gametocytaemia despite AL treatment needs to be evaluated in a larger study.Artemether lumefantrine is highly efficacious against uncomplicated  The adoption of artemisinin-based combination therapy (ACT) in sub-Saharan Africa as first-line drug for the treatment of uncomplicated malaria  was largin vivo study was carried out between mid-June and mid-August 2011 at Kitete Regional Hospital and St. Philip\u2019s and St Anne\u2019s health facilities serving the urban and peri-urban areas of Tabora municipality. Tabora is a high-burden area and malaria transmission peaks during the rain season, i.e., February to May [An y to May . Childrey to May . The recThe clinical and parasitological responses were assessed on Day 2, 3, 7, 14, 21 and 28 during the 28-day follow-up period. Parasitological responses were assessed microscopically by counting against 200 or 500 white blood cells for asexual and sexual (gametocytes), respectively. Gametocytes were not staged. Admittedly, one major limitation is that microscopy understimates the prevalence of gametocytes. Plasma lumefantrine levels were determined before treatment (Day 0) and post-treatment (Day 7). In the three months, of 1,126 children screened 33 had malaria and only 20 met the inclusion criteria and were recruited  laboratory for lumefantrine level analysis by high performance liquid chromatographic (HPLC) method as described .The study protocol was reviewed and approved by Ethical Review Committee of the Ifakara Health Institute (IHI) and MUHAS. Prior to the trial, a written consent form was signed by the parent or guardian of each participating child.A total number of 1,126 patients were screened from the three health facilities in Tabora region as follows: St Philip\u2019s Health Centre: n= 663; St Anne\u2019s Health Centre: n=345; and, Kitete Regional Hospital: n=118. The baseline characteristics of the study participants are indicated in Table\u2009Twenty-four 24) out of 663 (3.6%), five out of 345 1.4%) and four out of 118 (3.4%) patients in St Phillip\u2019s, St Anne\u2019s Health Centres and Ketete Hospital, respectively, were positive for malaria parasites by microscopy. Comparison of routine (before the study) and expert (during the study) malaria diagnosis was done. The routine and expert malaria microscopy results recorded in the first (50.3% positive n=163) and second half (5.1% positive n=176) of June 2011, respectively, were significantly different (p < 0.005). Only 20 patients met the inclusion criteria and were enrolled. Patients were treated and assessed for treatment outcome without loss of follow up as detailed in Figure\u2009.4% and f4 out of The lumefantrine plasma concentrations measured in patients before drug administration in all 20 enrolled patients were below HPLC detection level. The median Day 7 lumefantrine concentration level was 404.4 ng/ml (range 189.4-894.3 ng/ml). Only four (20%) patients had lumefantrine plasma concentration level < 280 ng/ml, and no patient had lumefantrine plasma concentration level <175 ng/ml. Lumefantrine concentration at day 7 was slightly decreased with unit increase in weight (kg) for any age but this decrease was not statistically significant .Table\u2009Artemisinins are known to be highly potent anti-malarial drugs that are active against immature gametocytes ,14, hencThis study recorded high (100%) efficacy of AL. The reported efficacy level is more or less similar to >98% recorded in Tanzania, prior to introduction of AL , and elsIn this study, none of the patients reported to have taken AL prior to hospital presentation, which was confirmed by lack of lumefantrine traces in the plasma. In addition, all patients had adequate therapautic levels of lumefantrine on Day 7. However, the Day 7 plasma lumefantrine levels showed wide variation between individuals. Indeed, it is known that the efficacy of AL combination is strongly influenced by wide variation in the pharmacokinetics of lumefantrine among individuals . The maxArtemether-lumefantrine showed short fever and parasites clearence time and quick recovery of haemoglobin levels. Only one patient had fever up to Day 1 and neither fever nor parasitaemia was reported on Day 2, contrary to a report in central Ethiopia where fever and parasitaemia persisted to Day 3 . HaemoglThe Day 0 gametocyte prevalence 30%) recorded in this study is higher compared with those reported previously in Tanzania ,30. Furt% recordeACT is effective against immature sequestered gametocytes ,34,35, hThe phenomenon of increase in gametocytaemia in this study and of sexual stage count in Nigeria  despite P. falciparum malaria was high in the study area. All patients were not exposed to AL prior to hospital presentation and levels of lumefantrine on Day 7 were therapeutically adequate but highly variable. The unusual increase in gametocytes following treatment needs be evaluated in a larger study concurrent with improvement capacity to diagnose malaria and handling of non-malarial fevers.The therapeutic efficacy of AL against uncomplicated The authors declare that they have no competing interests.KM, AM and ZP designed the study. KM and DJ supervised the field work. SA and AMK and ZP supervised the clinical part. MOMS supervised the drug level analysis and pharmacokinetics. DJ was involved in drug analysis and analysed data. RH is the in charge of St Philip\u2019s Health Centre. DJ and KM wrote the manuscript. All authors read and approved the final manuscript."}
+{"text": "A \u201creverse pharmacology\u201d approach to developing an anti-malarial phytomedicine was designed and implemented in Mali, resulting in a new standardized herbal anti-malarial after six years of research. The first step was to select a remedy for development, through a retrospective treatment-outcome study. The second step was a dose-escalating clinical trial that showed a dose-response phenomenon and helped select the safest and most efficacious dose. The third step was a randomized controlled trial to compare the phytomedicine to the standard first-line treatment. The last step was to identify active compounds which can be used as markers for standardization and quality control. This example of \u201creverse pharmacology\u201d shows that a standardized phytomedicine can be developed faster and more cheaply than conventional drugs. Even if both approaches are not fully comparable, their efficiency in terms of public health and their complementarity should be thoroughly considered. Malaria elimination efforts will lead to the much wider use of the few currently effective anti-malarial drugs, such as artesunate / amodiaquine, artesunate / sulphadoxine-pyrimethamine (SP), and artemether / lumefantrine. There is already discussion about intermittent presumptive treatment of infants, children, pregnant women, and even mass drug administration in some settings . ResistaIn this context it is important to maximize the lifespan of existing anti-malarials, and to consider all options for the development of new anti-malarials. Traditional medicinal plants have provided the source of the two major families of anti-malarial drugs still in use today, artemisinin and quinine, so many researchers are screening plants for novel chemical entities to develop as \u201clead compounds\u201d for new anti-malarial drugs . HoweverIn contrast the parallel development of standardized phytomedicines can be done faster, more cheaply, and more sustainably for remote areas. They could then be proposed and tested as a complement to existing strategies, for example as first aid in remote areas in case there is some delay until ACT treatment can be started. Their use might also delay the development of resistance to current standard drugs. The concept of \u201creverse pharmacology\u201d was coined in India to develop pharmaceuticals from Ayurvedic medicines, and was also championed by the Chinese in the 1950s , but stiArgemone mexicana decoction, which is in the process of being approved in Mali. The regulatory requirements for herbal medicines are completely different to those for new drugs. It should be emphasized that the primary objective of the project described here was not to develop new drugs, but to improve the utilization of herbal medicines, which are already in use. All the clinical studies described below were reviewed and approved by the Ethics Committee of the Institut National de Recherche en Sant\u00e9 Publique (INRSP) in Mali. This process took six years and cost about 400,000 Euros.In order to develop a standardized phytomedicine, a \u201creverse pharmacology\u201d approach was tested, where clinical evaluation was prioritized from the start. Isolation of compounds was done only at the end of the pathway, mainly for the purposes of quality control, agronomic selection and standardization, if justified by the clinical results. This experience led to the development of a new anti-malarial phytomedicine from a traditional herbal remedy, namely The research project is described here as it actually happened. Some aspects are reviewed in the discussion section, as with the benefit of hindsight some procedures might be improved. The hope is that this paper may help others who are interested in conducting a similar process  through a clear report of what was planned, what was opportunistically added \u2013 and what was obtained.et al developed a new method called a \u201cRetrospective Treatment Outcome Study\u201d (RTO) [The classical way of identifying medicinal plants for further research is through ethnobotanical studies. Yet conventional ethnobotanical studies rarely involve clinicians. They could and should provide much more clinical information if the ultimate goal is to know which one, among numerous treatments for a given ailment, has the best effects . Althougy\u201d (RTO) . This siIn the RTO, the first step was to understand local concepts and terms for diseases. The aim was to maximize the chances that the respondents were giving information about the disease of interest to researchers. For uncomplicated malaria, the definition was \u201cfever with no other obvious cause during the rainy season\u201d and for severe malaria, it was \u201cfever with convulsions or loss of consciousness during the rainy season\u201d. In MaliThe second step was to choose a representative random sample of households in the study area (by cluster sampling), and to ask in each whether anyone has had the disease of interest in the recent past. The timing was at the end of the malaria season . For uncomplicated malaria, a recall time of two weeks was used  . For sevThe third step, if a respondent had had the condition of interest, was to ask in detail what treatments they had taken, in what order, at what stage they had recovered from their illness and if the cure was complete or with sequelae. In this way it was possible to understand what treatments patients were actually using in real life and with what results.Argemone mexicana . None of the existing databases or books can cover all published information on a given topic, and therefore as many sources of information as possible were consulted: firstly freely available online databases [Search terms included the plant species and major chemical compounds known to exist in the plant  without some clinical evidence of effect size and safety.The traditional preparation was given to patients with uncomplicated malaria, who met the following criteria:1. Inclusion criteria:a. Symptoms of malaria (fever) within the last 24 hoursb. Parasitaemia >2000/mcl and <200 000 / mcl on microscopyc. Informed consent of patient or parent2. Exclusion criteria:a. Signs of severe malariab. age <3 monthsc. pregnancyd. other concomitant febrile illnesse. administration of a full course of anti-malarial  within the previous weekf. inability to return for follow-up.Patients were followed up closely on days 1, 2, 3, 7, 14 and 28, and were advised to return immediately at any other time if their condition deteriorated. Monitoring included parameters of efficacy  and safety . The design of the whole study became a sequential follow-up of patients using, in the first group, a dose lower than the one traditionally used , then the bottom and top of the usual dose range  was insufficient, with an ACR in only 35% of patients. When questioned why he had chosen this dose, the healer replied that he thought it was \u201cmore scientific\u201d . He then revealed that in fact he normally told patients how to prepare the remedy, and advised them to drink as much of it as possible. Therefore two other standard doses were agreed: one glass twice a day for 7 days (B), and one glass 4 times a day for the first 4 days, followed by one glass twice a day up to 7 days (C). Increasing the dose from A to B improved the efficacy (the proportion of patients with ACR increased from 35% to 73%) without an increase in adverse effects. However, at the maximal dose, there was no additional benefit (ACR in 65%), and two patients developed a prolonged QTc interval on their ECG. Thus an intermediate dose (B) was chosen as the safest and most effective to take forward into the next stage .A voucher specimen of the plant harvested for making the phytomedicine was deposited in the herbarium of the Department for Traditional Medicine. Thin-layer chromatography of the plant extract (methanol) and of the decoction was used to identify already published constituents. This preliminary study was subsequently confirmed by HPLC and mass spectroscopy. Berberine and sanguinarine were detected in the methanol extract but sanguinarine was not detectable in the decoction. Lyophilized samples of the phytomedicine used were kept for reference, and future phytochemical fingerprinting.Although according to WHO guidelines it would not have been necessary, the opportunity arose to conduct toxicological studies. These were conducted in parallel with the dose-escalating clinical trial. The LD50 of the freeze-dried decoction of the aerial parts was determined twice in two different laboratories which both showed that it was >3000mg/kg, as no rats or mice were adversely affected even by this high dose ,34. TherAs results from all previous stages were encouraging, the aim at this stage was to test the effectiveness of the phytomedicine in the field. In Mali, the objective was to develop a phytomedicine for home-based management of malaria (HMM), with the aims of symptomatic improvement and preventing severe malaria. The vision was that, if effective, the plant could be recommended to communities to be cultivated and prepared locally as a first-line treatment for presumed malaria. Therefore, the inclusion criteria for the RCT reflected this: all patients with presumed malaria  were included.It is not ethical to give placebo or no treatment to the control group, because falciparum malaria is potentially fatal and can progress rapidly, particularly in non-immune patients. The most useful comparator is the nationally recommended first-line treatment. In most countries this is now an artemisinin combination therapy (ACT). In Mali it was artesunate-amodiaquine. Artemisinins are the most effective and rapid anti-malarial drugs ever discovered, so it is not realistic to aim for a herbal treatment to outperform an ACT. Rather the aim should be non-inferiority for the selected appropriate outcome measures, or at least reaching a certain pre-defined level of effectiveness.A. mexicana, versus 95% (95% CI 88.8\u201398.3) on artesunate-amodiaquine. An important secondary outcome measure was incidence of severe malaria, which is the most important outcome in public health terms. Large numbers of patients are needed in order to demonstrate non-inferiority, because severe malaria is a relatively uncommon event. However another approach is to see whether the incidence of severe malaria is kept below a pre-specified level in both groups [A. mexicana decoction); an unequal randomization ratio was chosen in order to collect, with equal means, more information on the less known treatment. The observed age-specific incidence of severe malaria (in children aged 0-5 years) was 1.9% in both groups (Argemone mexicana and ACT) over the first 28 days of follow-up. The follow-up was extended to three months, and over this time the age-specific incidence of severe malaria was 2% per month in the herbal group and 1% per month in the ACT group. With 95% confidence, the age-specific incidence of severe malaria in both groups was <6% per month[The outcome measures are summarized in table h groups . In a prh groups . The aimper month.This is the last step of \u201creverse pharmacology\u201d. A phytomedicine can be developed without isolating an active ingredient, but it is useful to do this for two reasons. First and foremost there needs to be a phytochemical marker for quality control and standardization of the herbal medicine, and also to permit agronomic selection of the best plants. Secondly it is possible that a new modern drug could be developed in parallel by the pharmaceutical industry. However it makes more sense to do this after the clinical safety and effectiveness have already been demonstrated, as chances may be higher that the isolated compound (or a derivative) will also be safe and effective. Much time and money is wasted in developing drugs which turn out to be unsafe or ineffective in humans .Argemone mexicana contains at least three protoberberine alkaloids in similar amounts  with similar anti-malarial activity: berberine, protopine and allocryptopine  [in vitro, the absorption of berberine is poor in some animal models, although it can be improved by P-glycoprotein inhibitors [A. mexicana contains any P-glycoprotein inhibitors, but if it does, their concentration would also be important. The pharmacokinetics of protopine and allocryptopine have not yet been studied in humans, so it is not known which of these is the best marker, or whether there is synergy between them . Unlike berberine, protopine and allocryptopine show a good selectivity for Plasmodium and their cytotoxicities are low [in vivo tests using freeze dried AM decoction were unsuccessful both in mouse and in rat models , the plan is now to study the in vitro antiplasmodial activity of plasma samples from healthy volunteers to identify plant substances or metabolites involved in such activity.Isolating pure active ingredients from a phytomedicine is not straightforward. Most phytomedicines contain several compounds with additive or synergistic activities, or even pro-drugs. ctively) . Whereashibitors . It is n are low . Since pWhile developing new compounds from natural products could be an important source of new anti-malarials in the long term, it is also possible to develop standardized and validated phytomedicines more quickly and cheaply. The scheme used has already saved considerable time and money in developing a new herbal anti-malarial in Mali. Since the studied plant is a pan-tropical weed, results of such a research programme could be applied in many countries, provided there is local quality control of the plants.It is of paramount importance to conduct such research in an ethical manner, and all the clinical trials were submitted and approved by an ethics committee. To be ethical, a non-inferiority trial needs to test a strategy that could be sustained after the end of the research. During the study there must be proper safeguards in place to ensure the safety of the patients, so a medical team was stationed in the village for the whole period to give immediate care when required. The result was to inform the villagers which of their traditional remedies has been clinically shown to be an effective anti-malarial, at what dose, what precautions are necessary in its preparation, and that they should rapidly seek modern medical treatment if they do not improve or if danger signs appear. It is likely that this knowledge is of more benefit in the long term, and so more ethical, than a short-term unsustainable intervention. This hypothesis will be tested in future research on the public health impact of such information.in vitro activity, which can be misleading. In this case the in vitro activity was largely attributable to berberine, which is poorly absorbed, and is probably not (or not directly) responsible for the activity in humans.With the benefit of hindsight, there are some improvements which could be made to the scheme. In the initial selection of the plant, the determining factor should be the observed treatment-effect correlation in the RTO rather than the In the dose-escalating study, it would have been better to start by consulting those familiar with the remedy about the minimum and maximum doses which patients can take, to ensure that the traditional healer was proposing his usual range of doses for the study. Based on this information, two or three different standard dosages of the phytomedicine could be defined in advance in the trial protocol. Of course, the same inclusion criteria must be used throughout the trial, so that the patients in each group are as similar as possible.Artemisia annua infusion is being tested as a backup for situations in which the recommended first-line treatment is not available. These include stock-outs of standard drugs, and remote areas which are not reached by the healthcare infrastructure. Brazil is a low transmission area, and total parasite clearance is considered mandatory. In this context ACPR is the chosen outcome measure.In other contexts the aim of treatment may be different. For example in Brazil, Cochlospermum planchonii root decoction), in Ghana (Cryptolepis sanguinolenta root infusion) [Artemisia annua Anamed leaf infusion). Much of the development work has already been done on these: their safety has been demonstrated and they seem efficacious in preliminary clinical trials. However further work is needed to decide how they would fit into public health strategies for control or elimination of malaria. It is important to develop cheap and reliable tests for quality control and standardization of plant material. Larger scale clinical trials are needed, including children who are most at risk of severe malaria, if they are intended to be future users of a validated and officially recommended phytomedicine. This is not the case in Mali, where it has been proposed to test on a small scale a health policy including Argemone mexicana decoction for the home-based management of malaria in patients aged over five years in high transmission areas, thereby saving ACTs for children aged five years and under [Other phytomedicines for malaria have already been developed and are government-approved in Burkina Faso (nfusion) , and in nd under .There is a range of other promising anti-malarial phytomedicines which could be developed much faster and more cheaply than new chemical compounds, because preliminary work has already provided some information on their safety and efficacy . Such phSustainable public health improvement in remote areas is a key consideration in such a discussion. Innovative Public Private Partnerships could also be considered with companies already expert in the production of standardized phytomedicines.Funding organizations should support the possibility of developing new types of medicines, including phytomedicines, rather than restricting funding only to conventional development of isolated active compounds. The authors declare that they have no competing interests."}
+{"text": "Plasmodium lactate dehydrogenase (pLDH) in treatment monitoring of uncomplicated falciparum malaria was carried out in an endemic setting in Myanmar.Malaria rapid diagnostic tests (RDT) are used for diagnostic purpose in malaria-endemic areas where reliable microscopy is not available. Persistence of the antigenaemia causes over-diagnosis and may limit the usefulness of the RDT in monitoring treatment. In this study, the usefulness of histidine-rich protein-2 (HRP2) and pan-specific or species-specific Plasmodium falciparum mono-infected cases. After direct treatment with an artemether-lumefantrine combination, patients were followed up on day 3, 7, 14, 21, 28 and any other day of recurrent fever. Blood film examination and RDT were carried out on day 0 and all follow-up days.A prospective longitudinal, single-arm, cohort study was done by microscopy to confirm Out of 77 recruited falciparum cases, 63 became adequate clinical and parasitological response (ACPR) cases, and 60.3% of them were still positive for HRP2 up to day 28. Eleven out of 12 treatment failure cases (91.6%) were detected by pan pLDH. The mean duration required to become negative of HRP2 was 20\u00a0days (SD\u2009\u00b1\u20096.03) and that of pan pLDH was six days with or without gametocytes and 3.7\u00a0days without gametocytes.Although treatment monitoring cannot be performed by HRP2, it can be assessed by pan pLDH-based assay after day 3 if a gametocidal drug has been administered and after day 7 if the presence of gametocytes was not excluded. The pan pLDH-based assay was a suitable test to monitor the treatment response of uncomplicated falciparum malaria patients. The World Health Organization (WHO) recommends that malaria case diagnosis and management should be parasite-based and microscopy is still the cornerstone of diagnosis and remains the recommended method for therapeutic monitoring ,2. HowevPlasmodium falciparum), pan-specific or species-specific Plasmodium lactate dehydrogenase (pLDH) or aldolase, which is specific to all the major Plasmodium species [P. falciparum[Plasmodium species that infect humans [Many new technologies, including immunochromatographic tests, appear to overcome the limitation of microscopy . The tes species . The HRPalciparum. The pLDalciparum is produt humans ,7.et al. (1999) study showed that the immunochromatographic assay may be useful in post-treatment monitoring of malaria [Palmer et al. 199 study  malaria ,7 and it malaria , type of malaria , type of malaria  and stra malaria . It is n malaria ,14.This prospective longitudinal, single-arm cohort study was done in Myanmar-Thailand border areas, Myawaddy Township, Kayin State from October to December 2010. The Township health profile showed that malaria was the leading cause of morbidity and mortality in this area from 2007 to 2009 and is defined as a hyper-endemic area . The MyaSample size is calculated according to the anticipated proportion table  and 73 pP. falciparum , axillary temperature \u226537.5\u00b0C or history of fever within previous 24\u00a0hours, ability to swallow oral medication and ability and willingness to comply with the study protocol for the duration of the study and to comply with the study visit schedule. Any person who showed presence of signs and symptoms of severe and complicated falciparum malaria according to current WHO definitions [Plasmodium species, or other species of Plasmodium, presence of severe malnutrition, presence of febrile conditions due to diseases other than malaria, and pregnancy or lactating mothers, were excluded from the study.Patients were recruited to the study were at least six years old, had mono-infection with microscopy confirmed The study was approved by the ethical committee from Department of Medical Research (Lower Myanmar) and University of Medicine 1, Yangon. Written informed consents were taken from all the participants.The participants were screened by peripheral blood smear stained by 10% Giemsa and examined by microscopy. Individuals who met the inclusion criteria were enrolled, tested by HRP2 and panpLDH based RDT, thick and thin films examination for 3% Giemsa and treated on site with artemether-lumefantrine (Coartem\u00ae). The patients involved were monitored for a period of 28\u00a0days according to the scheduled visits.The microscopic blood film examinations were done according to WHO recommendations described in \u201cMethods for surveillance of anti-malarial drug efficacy\u201d . Thick aIn this study, malaria antigen (HRP2 and pan pLDH based) Pf/Pan immunochromatographic test kit  was used. Test procedure was done according to manufacturer\u2019s instruction. Briefly, blood sample (5\u00a0\u03bcl) was taken from finger tip by a capillary pipette, and the open end immersed in the blood drop and then gently released for the pressure to draw blood into the capillary pipette to sample well. Four drops of assay diluents were added to the assay diluent well. The result was read after 15\u00a0minutes; never after 30\u00a0minutes to avoid false result. Therapeutic evaluation of the patients was not interfered by the result of this test. Band intensity of the result was noted as follows: 0 no band (negative), 1+ faint band, but clearly visible (positive), 2+ medium intensity bands, stronger than 1+ but less than control band (positive), 3+ equal or stronger than the control band (positive).The test kits were stored in a temperature and humidity control room at the Quality Control Laboratory for malaria rapid diagnosis test (RDT), Department of Medical Research (Lower Myanmar). In the field, the test kits were stored at the recommended temperature and humidity. Temperature and humidity were recorded three times a day, i e, 6\u00a0am, 12 noon and 6\u00a0pm by using a thermo hygrometer to ensure the recommended temperature and humidity. After the study, 10 test kits from the field were randomly selected and tested with the quality control blood samples that were prepared according to the methods manual for laboratory quality control testing of malaria RDT to check the validity of the assay kits .After the study was completed, data were entered onto a database using double independent data entry by using Microsoft Excel and SPSS software version 16. Sensitivity, specificity, positive predictive value, negative predictive values were calculated as blood film examination was gold standard. Sensitivity is the probability (percentage) that patients with the infection  will have a positive result using the test under evaluation.Plasmodium vivax; 29.4% (102 cases) were P. falciparum; and, 7.2% (25 cases) were mixed infection, i.e, P. falciparum and P. vivax.A total of 844 patients were screened by active case detection and malaria parasites were detected in 40.9% of the screening slides (n\u2009=\u2009346). The 63.2% (219 cases) of the microscopy positive cases were P. falciparum mono-infection, 77 participants were recruited for the study. The screening algorithm of study cases is shown in Figure\u00a0Of the P. falciparum (n\u2009=\u20094) was positive on day of failure. However, long-term persistence of HRP2 was observed before day of treatment failure in all of the cases. Persistence positive of the HRP2 was observed up to the day of failure. It was difficult to differentiate between the persistence of antigen from newly emergent antigen. Their band intensity was increased in two failure cases but was the same as the previous schedule visit in the other two.The result of HRP2 for all the treatment failure patients of In 12 parasite-reappeared cases, there was no persistence positivity of pan pLDH results. Band intensity of the pan pLDH was decreased on subsequent follow-up days, a feature which was parallel to the microscopy result. The reappearance of the band of the pan pLDH was observed in all parasite-reappeared cases regardless of the species, except in only one case in which parasite density was low (31 per \u03bcl of blood)  . However, if P. falciparum gametocytaemia was considered positive for P. falciparum, the sensitivity of the pan pLDH was significantly increased .The pLDH had high specificity (73.97\u201396.87%). The specificity was increased to 87.10\u2013100% if \u03c72\u2009=\u20092.853, p = 0.7227, 95% CI) in all ACPR cases. However, positive results of HRP2 were significantly higher than of microscopy  as shown in Figure\u00a0The results of pan pLDH were parallel to the microscopy result on day 0 and all of the follow-up days  showed 1+ band intensity on day 28. Without gametocytes, only five cases of false positive were observed on day 3. There was no case of more than +1 band intensity in all of the cases, with or without sexual parasite, on and after day 3.Long-term persistence of the false positive was directly correlated with the initial parasite count in ACPR cases (p\u2009=\u20090.000 for both HRP2 and pLDH). Initial parasite count was also correlated with the persistence of higher band intensity of HRP2 on day 28 as shown in Figure\u00a0P. falciparum, the mean time required to become negative result of HRP2 was 21\u00a0days (SD\u2009\u00b1\u20096.41) and of pLDH was 3.7\u00a0days (SD\u2009\u00b1\u20094.11).Among the 63 ACPR cases, a total of 25 cases became HRP2 negative before day 28 and all 63 cases became pLDH negative within 28\u00a0days. Mean time to become negative result of HRP2 was 20\u00a0days (SD\u2009\u00b1\u20096.03) and pLDH was six days (\u00b16.31). If gametocytes were considered positive for P. falciparum gametocytaemia was considered positive for P. falciparum, false positive of pLDH was less than 10%. However, false positive for HRP2 was more than 60% in all of the follow-up visits even if P. falciparum gametocytaemia was considered positive for P. falciparum.Day 3 persistence of asexual parasitaemia was detected in four patients. On day 3, 59 (94%) were HRP2 false positive. On day 7, 62 (98%) were HRP2 false positive. A total of 56 (89%) on day 14, 45 (71%) on day 21, and 38 (60%) on day 28 were still positive without asexual parasitaemia. If P. falciparum has been documented [The therapeutic monitoring and early detection of treatment failure is very important in management of malaria cases in the era of increasing drug resistant malaria. The ability of the test to monitor response to therapy is ideal in malaria-endemic areas, especially where drug resistant cumented .P. falciparum only, it was useless in non-P. falciparum infection [P. falciparum but also non-P. falciparum infection, but it cannot differentiate between the two. However, if the sexual stage parasites were still present on follow-up visits, and the pLDH test became positive, this can cause confusion in early detection of the treatment failure [HRP2 persistence makes early detection of treatment failure difficult. The sudden increase of the band intensity of HRP2 on subsequent follow-up visits is an alert to further investigation to confirm treatment failure. However, without continuous monitoring of cases by the recording of band intensity, a definite conclusion cannot be reached of treatment failure. Moreover, being the HRP2 produced from nfection . The pan failure .P. falciparum gametocytaemia was considered positive for P. falciparum, indicating that persistence gametocytaemia may cause positive result of pLDH. Therefore, it is necessary to follow the National Anti-malarial Treatment Guidelines and WHO recommendations, which mention to add a single dose of primaquine (0.75\u00a0mg/kg) as a gametocidal drug to artemisinin combination therapy for uncomplicated falciparum malaria [There were many studies focusing on the sensitivity of pan pLDH. However, the results were varied: one study in Uganda  found th malaria .P. falciparum gametocytaemia was considered positive for P. falciparum. Other studies showed the median duration for pLDH to become negative was two days for CareStart\u00ae malaria tests and seven days for OptiMAL-IT\u00ae [The result of pan pLDH was parallel to that of microscopy. However, the results of HRP2 cause more false positives due to the long persistence of antigenaemia on subsequent follow-up days. The long-term persistence of HRP2 reduces its usefulness in monitoring the response to treatment. It was documented that during follow up after treatment, 98.2%, 94.6%, 92.0% and 73.5% of effectively treated children were still false-positive by RDT at day 14, 21, 28 and 35, respectively, and this antigenaemia could persist up to 35\u00a0day after treatment . In thisiMAL-IT\u00ae .However, continuous reducing of the band intensity of HRP2 was observed in most of the ACPR patients. There were evidence of association of the band intensity and parasite count , a \u2018plusThe frequent occurrence of false positive results can lead to unnecessary treatment. This can have several negative outcomes, including clinicians inappropriately focusing on malaria, and not identifying the true cause of illness, and unnecessary exposure to anti-malarials. In some cases, the inappropriately treated patient may return with similar symptoms, leading the clinician to falsely report the presence of parasite drug resistance. This could lead to the clinician not trusting the efficacy of the first-line anti-malarial and consequently dispensing the second-line anti-malarial, increasing the cost of treatment and further delaying appropriate treatment . The treP. falciparum infection during the follow-up period. The persistence of pan pLDH was shorter in duration than that of HRP2. The meaningful result of pan pLDH can be observed in monitoring treatment response in uncomplicated falciparum malaria patients after day 3 of artemisinin combination therapy if the gametocidal drug was added according to the recommendation by WHO in 2010. Therefore, pLDH is useful for therapeutic monitoring of uncomplicated falciparum malaria patients.In this study, HRP2 had higher sensitivity than pan pLDH assay. However, specificity was higher in pan pLDH. The persistence of HRP2 up to day 28 in ACPR cases causes false positive results and this is the reason HRP2 is not fit for monitoring treatment response. The pan pLDH can also detect late parasitological failure and non-The authors declare that they have no competing interests.MPK, KKW and MHN conceived and designed the study. MHN conducted the field work. KMN and KMM conducted data validation and management. All authors contributed during writing, and read and approved the manuscript."}
+{"text": "Plasmodium falciparum malaria in Yingjiang County along the China-Myanmar border and investigate genetic polymorphisms in the P. falciparum chloroquine-resistance transporter (pfcrt), multidrug resistance 1 (pfmdr1), dihydrofolate reductase (pfdhfr), dihydropteroate synthase (pfdhps) and ATPase (pfatp6) genes.The aim of this study was to evaluate the clinical outcome after seven-day artesunate monotherapy for uncomplicated P. falciparum mono-infection were included. Patients received anti-malarial treatment with artesunate  by directly observed therapy. After a 28-day follow-up, treatment efficacy and effectiveness were assessed based on clinical and parasitological outcomes. Treatment failure was defined as recrudescence of the original parasite and distinguished with new infection confirmed by PCR. Analysis of gene mutation and amplification were performed by nested polymerase chain reaction.Patients\u2009\u2265\u2009one year of age with fever (axillary temperature \u226537.5\u00b0C) or history of fever and pfcrt mutation in codon 76 was found in all isolates (100%), and mutations in codons 71 and 72 were found in 4.8% of parasite isolates. No mutation of pfmdr1 (codons 86 or 1246) was found. Among all samples, 5.1% were wild type for pfdhfr, whereas the other samples had mutations in four codons , and mutations in pfdhps  were found in all isolates. No samples had mutations in pfatp6 codons 623 or 769, but two new mutations (N683K and R756K) were found in 4.6% and 9.2% of parasite isolates, respectively.Sixty-five patients were enrolled; 10 withdrew from the study, and six were lost to follow-up. All but two patients demonstrated adequate clinical and parasitological response; 12 had detectable parasitaemia on day 3. These two patients were confirmed to be new infection by PCR. The efficacy of artesunate was 95.9%. The Plasmodium falciparum infection was associated with slow parasite clearance and suspected artemisinin resistance at the China-Myanmar border area. The prevalence of pfcrt 76\u2009T and markers for SP resistance are still high. It should be strengthened further on parasite clearance time or clearance half life to confirm the resistance status, and molecular epidemiology should provide complementary information to assess the appropriateness of current policies based on artemisinin derivatives. Plasmodium falciparum tolerance/resistance to artesunate in populations on the Cambodia-Thailand border[Malaria is a severe infectious disease, and drug resistance to anti-malarial drugs is a major public health problem worldwide. In the d border,4.Falciparum malaria is now found in only two provinces in China: Yunnan province, which borders Myanmar, Laos and Vietnam, and Hainan province. Malaria control measures have been actively implemented in this area for more than 30\u2009years with considerable success, and there are no local falciparum malaria cases in Hainan. HoweverP. falciparum is resistant to other anti-malarial medicines to optimize therapeutic effectiveness and delay the emergence of resistance. In 2006, the WHO advocated a complete ban on artemisinin monotherapy for uncomplicated malaria[China was the first country to use artemisinins, but their wide-scale use in China began only in the early 1990s. The standard treatment regimen was 12\u2009mg/kg artesunate or artemether over five days, which continued to be widely used until 2007. Since 2001, the WHO has recommended the use of artemisinin-based combination therapy in all areas where in vivo malaria therapeutic efficacy studies in sentinel sites to monitor P. falciparum and Plasmodium vivax sensitivity to first-line, anti-malarial treatments.Until now, ACT has been the only effective treatment for falciparum malaria. At a time when ACT is being rolled out in endemic malaria areas all over the world and is beginning to have an effect on morbidity and mortality in tropical Africa, the development and spread of ACT resistance would have grave consequences for global malarial control. Because of these concerns, national malaria programmes conduct systematic periodic P. falciparum and P. vivax. The most relevant polymorphisms are presented below. The 76\u2009T allele in the chloroquine-resistance transporter gene (pfcrt) is predictive of chloroquine and amodiaquine treatment failure[pfmdr1) has been linked to chloroquine and amodiaquine resistance, and increased chloroquine inhibitory concentrations in P. falciparum have been linked with pfcrt 76\u2009T[pfdhfr) haplotype N51I/C59R/S108N has been associated with sulphadoxine-pyrimethamine (SP) treatment failure, and the addition of the dihydropteroate synthase (pfdhps) SNPs G437A and K540E produces highly resistant P. falciparum[2+-ATPase ortholog of P. falciparum (pfatp6) was suggested to be involved in the mechanism of action and resistance of the parasite to artemisinins[Several genetic polymorphisms that can provide reliable data about the prevalence of drug resistance have been described in  failure-10. The crt 76\u2009T. The trialciparum-16. The misinins.P. falciparum malaria in Yingjiang County along the China-Myanmar border and to investigate the prevalence of genetic polymorphisms in pfcrt, pfmdr1, pfdhfr, pfdhps and pfatp6.The aim of this study was to evaluate clinical outcome after seven-day artesunate monotherapy for uncomplicated The study was a one-arm prospective evaluation of clinical and parasitological response to directly observe treatment for uncomplicated malaria. The study was conducted in Yingjiang County, which is near Lazan City in Myanmar.\u226537.5\u00b0C) or history of fever in the previous 48 hours were included. The included patients had mono-infection with P. falciparum, parasitaemia levels between 250 and 100,000 asexual parasites/\u03bcl, no history of anti-malarial use in the previous 14\u2009days and no signs of severe malaria or danger signs.Patients over six months of age with fever . Direct observed therapy (DOTS) was given for seven days by the health worker. Tablets of artesunate were obtained from the WHO . In case of vomiting within 30 minutes, the dose was repeated; if vomiting occurred within 30 minutes after the repeated dose, the patients were excluded from the study and referred for treatment at the healthcare facility.Patients were followed up on days 1, 2, 3, 4, 5, 6, 7, 14, 21 and 28 for clinical and laboratory tests, which included axillary temperature measurement and thick blood smear preparation. If a patient could not be located on the day of the visit, the study staff attempted to locate the patient at his or her home. Patients who were not located within one day of the expected visit were considered lost to follow-up and excluded from the study.Sample size was calculated based on a predicted treatment failure rate of 15% because the treatment failure rate of artesunate in the area was unknown. At a confidence level of 95% and a precision of approximately 5%, a minimum of 50 patients were required for the drug to be tested. At least 60 patients should be included to allow up to 20% loss to follow-up and withdrawals during the 28-day follow-up period.Thick blood smears obtained on every visit day were stained with 10% Giemsa for 10 minutes and examined at 1,000\u00d7 magnification by a trained microscopist to confirm the species and parasite density. The parasite density was determined by counting the number of asexual forms and gametocytes in 200 leukocytes. Parasitaemia was estimated by assuming 6,000 leukocytes/\u03bcl. A second microscopist blinded to the first result re-examined the thick blood smear, and in case of discrepancies greater than 50%, the smear was read by a third microscopist. The geometric mean of the two closest results was used as the parasitaemia value for each reading. When the number of asexual parasites was less than 10 per 200 white blood cells in follow-up smears, counting was performed in at least 500 white blood cells . A blood slide was considered to be negative when examination of 1,000 white blood cells revealed no asexual parasites. The presence of gametocytes on an enrolment or follow-up slide was noted, but this information did not contribute to the basic evaluation.msp1msp2 and glurp[Genotype analysis was conducted to differentiate recrudescence (same parasite strain) from newly acquired infection (different parasite strain). This analysis is based on the extensive genetic diversity among the malaria parasite genes and glurp. The genin vivo efficacy monitoring[Treatment efficacy and effectiveness were evaluated among observed patients based on clinical and parasitologic outcomes and study endpoints in accordance with WHO guidelines for nitoring. The outThe study was reviewed and approved by the ethical committee of the Chinese Centre for Disease Control and Prevention (China CDC). This study was also registered at the website under thpfcrtpfmdr1pfdhfrpfdhps and pfatp6. Sequencing reactions were carried out using an ABI PRISM BigDye Terminator v3.1 Cycle Sequencing kit  as specified by the manufacturer. The sequences of the amplicons were aligned with data published in the NCBI database by BLAST analysis.DNA was extracted from dried blood samples on filter papers using a QIAGEN mini kit and stored at \u221220\u00b0C until future use. Nested PCR,14,19,20All of the patients were observed at the Rose Clinic in Lazan City, a small town on the Myanmar side of the border located 24\u00b047\u203221.22\u2033N and 97\u00b033\u203221.05\u2033 E . A total of 302 malaria patients were found, including 153 cases of P. falciparum patients, 17 were female (26.2%) and 48 were male (73.8%). The age distribution was 3.0% under five years, 15.5% between five and 15\u2009years and 81.5% >15\u2009years. There was a mild predominance of young adult males. All of the patients had a fever in the 48 hours preceding presentation to the clinic, and 73.8% (48/65) exhibited an axillary temperature >37.5\u00b0C. The geometric mean parasite density was 69,535 parasites/\u03bcl. The probability of the patients with parasite on day 3 was correlated with the parasite densities on day 0, that is, the more parasite densities the more probability of positive on days 3 TableOf the The patients\u2019 ethnicity was also noted: 84% belonged to the Jingpo ethnic group, which is dominant on both sides of the border in this area; 13% were Bamar (Myanmar majority group); and the remaining 3% belonged to the local Lisu ethnic group. Among the 7% who were Chinese citizens, most were Han, and a few belonged to the Jingpo group.Sixty-five patients were enrolled and received directly observed anti-malarial treatment with artesunate ; 10 withdrew from the study, and six were lost to follow-up. The efficacy of artesunate monotherapy was 95.9%.msp1, msp2 and glurp were genotyped and the results of the pre- and post-parasite strains of the two patients were shown in TableAll but two patients in the observed group demonstrated adequate clinical and parasitological response. The two patients with treatment failure were confirmed to have new infection by PCR. The allelic variants of Twelve patients had detectable parasitaemia on day 3. The geometric mean parasite density on Day 0 in the cases that were positive on day 3 was 89,519 asexual parasites per \u03bcl, indicating a strong association between day 3 positivity and high initial parasite density, as observed in other studies. The lowest day 0 parasitaemia associated with D3 positivity was 33,652 parasites/\u03bcl.The average body temperature and parasite density of the patients decreased dramatically on days 0, 1, 2, 3, 7, 14, 21 and 28 after seven days of artesunate treatment , and mutations in codons 71 and 72 were found in 4.8% of parasite isolates. No mutation in pfmdr1 (codons 86 or 1246) was found.A total of 63 samples were successfully amplified at positions 71, 72 and 76. Mutation in pfdhfr genes from 63 samples were successfully amplified and compared with the wild-type sequence. The frequencies of N51I, C59R, and I164L are shown in FigureThe pfdhps, successful amplification was achieved in 59 samples, and mutations in codons 436, 437, 540 and 581 were found in all of the isolates (100%) including single, double, triple or quadruple mutants. The A437G and K540E mutants were most prevalent in pfdhps. Double mutants in pfdhps were found in 57.6% of samples and included 436/437, 436/540, 437/540, and 437/581  were found in 4.6% and 9.2% of parasite isolates, respectively.No mutation was found in de novo occurrence of artemisinin resistance in China is a serious concern. This study was designed to evaluate clinical outcome after seven-day artesunate monotherapy for uncomplicated P. falciparum malaria in Yingjiang County along the China-Myanmar border and investigate the prevalence of anti-malarial drug resistance markers. A parallel study evaluating DHA-PIP was conducted simultaneously in another county, and the results of that study will be published elsewhere.Artemisinin-based combination therapy is recommended to slow the emergence and spread of drug resistance. This effect is achieved by the use of anti-malarial drugs with different mechanisms of action and the use of a fast-acting highly effective anti-malarial drug, such as DHA, in combination with a long-lasting anti-malarial drug, such as piperaquine. China was the first country to use artemisinin, and this drug was used for malaria treatment in China for almost 20\u2009years (from the 1990s to 2007). In recent years, some reports confirmed artemisinin resistance in Southeast Asia, especially near Thailand-Cambodia border areas, but the status of artemisinin in China was unclear. The spread of artemisinin resistance from Cambodia to China or the P. falciparum.Yunnan province is located in southern China, with a population of approximately 30 million people in 129 counties. The province borders Myanmar, the Lao People\u2019s Democratic Republic and Vietnam. The border with Myanmar is 1,997\u2009km long , ie, confirmed re-infection. Therefore, this patient excluded as treatment failure with PCR correction until day 14. However, a key observation was the high rate of parasite positivity on day 3. A total of 18.5% of the enrolled patients were parasite positive on day 3. The initial parasite density was high, which might partially explain the high day 3 positivity rate, but the response still suggested a lowered susceptibility to artesunate, which was lower than the response rate observed in western Cambodia in recent years. It is impossible to say whether these findings reflect a spread from Cambodia or local factors. Local factors include long-term access to artemisinin (since at least the early or mid-1990s) and highly irregular treatment schedules in an area with continued moderate levels of transmission, as suggested by the predominance of malaria in young adults. Therefore, therapeutic efficacy tests should be continued, and the containment of artemisinin resistance in these areas should be prioritized.The majority of patients included in the study were from Myanmar or had been infected in a region of Myanmar close to the Chinese border. Among the 65 patients with Pfcrt and pfmdr1 genes have been proposed as molecular markers of chloroquine resistance, and they also influence P. falciparum susceptibility to mefloquine, quinine, halofantrine and artemisinin[pfcrt. As might be expected from the previous studies on in vivo and in vitro chloroquine responses in China[P. falciparum examined in the present study frequently contained the pfcrt K76T mutation, especially in Yunnan province. Although China has not used chloroquine to treat P. falciparum for over 30\u2009years, the stable and high prevalence of this mutation may be caused by the continued use of chloroquine as a first-line drug for P. vivax over several decades. Compared with the high prevalence of pfcrt mutations, there was no mutation in codons 86 or 1246 of pfmdr1 in any sample. This result was consistent with other researchers\u2019 findings[P. falciparum in China is still sensitive to mefloquine, quinine, halofantrine and artemisinin.emisinin-23. Accoin China, the isofindings-27. Untipfdhfr and pfdhps mutations on the mechanism of resistance to SP drugs have been well described. Pyrimethamine was used for the radical treatment of P. vivax in combination with primaquine 40\u2009years ago[pfdhps A437G mutation confers resistance to sulphadoxine and is often coupled with the K540E mutant allele.The effects of ears ago. Additioears ago. Pyrimetears ago and sulpears ago. This stears ago. This re2+ ATPase6. The analysis of naturally occurring pfatp6 polymorphisms in field isolates suggested that a polymorphism at codon 769 may be associated with the reduced susceptibility of these isolates to artemether in vitro[pfatp6 and artemisinins[pfatp6 in artemisinin resistance, all of the polymorphisms in this gene should be carefully monitored.The mechanism of action of artemisinin remains controversial. One of its proposed mechanisms involves an interaction with the sarcoplasmic reticulum Ca in vitro. In linemisinins. Howevermisinins. The mutmisinins-36 couldmisinins on emergGreater Mekong Sub-region Programme for Artemisinin Resistance Containment\u201d, which will be launching this year or next year.Slow parasite clearance and suspected artemisinin resistance was observed in Yingjiang County at the China-Myanmar border area, therefore it should be strengthen further on parasite clearance time or clearance half life to confirm the resistance status. And now Yunnan province of China, as part of Greater Mekong Sub-region, has already involved in \u201cAlso, this study provided basic data on artemisinin resistance in this area, which will be helpful for other countries in the Mekong sub-region. Moreover, molecular epidemiology should be continued as routine surveillance and to provide complementary information to assess the appropriateness of current policies based on artemisinin derivatives.The authors hereby certify that no conflict of interest of any kind occurred in the framework of this study.FH was responsible for the molecular genetic studies, participated in the field work, and drafted the manuscript. LT was responsible for the overall study design and was involved in all stages of the study, including field work. HY was involved in the design of the field work. SZ contributed to the molecular genetic studies and data analysis. XS and HL contributed to the field study, patient follow-up and data analysis. All of the authors read and approved the final manuscript."}
+{"text": "In order to compare the efficacy of the fixed-dose formulation ASAQ Plasmodium falciparum malaria were enrolled and randomly allocated into one of the two regimens. The risk of recurrent parasitaemia by day 42, both unadjusted and adjusted by PCR genotyping to distinguish recrudescence from new infection, was analysed.Children aged six and 59\u2009months with uncomplicated Between April 2008 and March 2009, 301 children were included: 156 with ASAQ and 145 with AL. No early treatment failures were reported. Among the 256 patients followed-up at day 42, 32 patients developed late clinical or parasitological failure (9.9% (13/131) in the ASAQ group and 15.2% (19/125) in the AL group). After PCR correction, cure rates were 98.3%  in the ASAQ group and 99.1%  in the AL group . Kaplan-Meier PCR-adjusted cure rates were similar. Both treatment regimens were generally well tolerated.falciparum malaria in this area of Katanga, DRC. However, in a very large country, such as DRC, and because of possible emergence of resistance from other endemic regions, surveillance of efficacy of artemisinin-based combination treatments, including other evaluations of the resistance of ASAQ, need to be done in other provinces.Both ASAQ and AL are highly effective and currently adequate as the first-line treatment of uncomplicated The protocol was registered with the clinicaltrials.gov, open clinical trial registry under the identifier number NCT01567423. Plasmodium falciparum accounts for nearly 95% of all malaria cases in this region [Despite recent progress in access to effective preventive and therapeutic measures to control malaria, it remains one of the leading causes of morbidity and mortality worldwide, and especially in Democratic Republic of Congo (DRC) . In thiss region .In 2005, following increased reports of sulphadoxine-pyrimethamine (SP) resistance , the NatUntil 2007, the combination of artemether-lumefantrine (AL) was the only fixed-dose ACT registered to international standards that was widely available in malaria-endemic countries. A new fixed-dose combination of artesunate and amodiaquine (ASAQ Winthrop\u00ae), developed by the Drugs for Neglected Diseases Initiative (DNDi) in collaboration with Sanofi-Aventis, was pre-qualified by the WHO in 2008.2, repeated evaluations of the ACT efficacy are needed in different regions.Current international guidelines advocate that once a new anti-malarial treatment is introduced in a country, the efficacy of such novel regimens needs to be monitored regularly in order to detect early signs of declining efficacy, which can have implications for policy markers . The fewGiven that the transition in drug policy from a loose to a fixed dose combination would be easier option for uncomplicated malaria treatment, it was decided to assess the efficacy of the new fixed-dose combination of artesunate and amodiaquine (ASAQ) in comparison with the fixed-dose combination of artemeter and lumefantrine (AL), in the province of Katanga, south-eastern DRC.The aims of the study were to compare (i) treatment efficacy of fixed-dose ASAQ and its alternative AL, expressed as product limit estimates of failure from survival analysis and as simple proportions from per protocol analysis, on day 42, and (ii) analysis of adjusted and unadjusted results by genotyping.An open randomized study was conducted to test the hypothesis that the risk of recurrent parasitaemia after 42\u2009days is not inferior in the group receiving ASAQ regimen compared to the group receiving AL regimen. The patients were recruited from the outpatient department of the general reference hospital of Pweto, health district of Pweto, province of Katanga, DRC, between April 2008 and March 2009 Figure .This protocol was registered with ClinicalTrials.gov, under the idenfier number NCT01567423.P. falciparum infections , fever or history of fever in the previous 24 hours, no signs of severe malaria, no reported hypersensitivities of the studied drugs, and no serious concomitant febrile illness [Children aged between six and 59\u2009months and with a body weight\u2009\u2265\u20095\u2009kg were eligible for enrolment if they had The children included were randomized in one of the two treatment arms, in a 1:1 ratio without stratification. The randomization list was generated by a computer in blocks of six. Treatment allocations were kept in sealed and numbered opaque envelopes. Participants were enrolled in the same order in which they were diagnosed.ASAQ  was administered once daily for three days, as follows: one tablet of artesunate 25\u2009mg/amodiaquine 67.5\u2009mg for children between 5 to 8.9\u2009kg, artesunate 50\u2009mg/amodiaquine 135\u2009mg for children between 9 to 17.9\u2009kg. One tablet of artemether 20\u2009mg/lumefantrine 120\u2009mg  was administered twice daily for three days to children with a body weight of 5 to 14.9\u2009kg and two tablets were administered twice daily for three days to children with a body weight of 15 to 24.9\u2009kg. All doses were administered under direct observation for the three days. Full doses of drugs were re-administered if a patient vomited within 30\u2009min after receipt. Patients who vomited more than twice were excluded from the study and were treated with oral quinine 10\u2009mg/kg/8\u2009h for 7\u2009days.e.g. severe malnutrition), intake of anti-malarial treatment in the last seven days, and mixed malaria infection were also excluded from the study.Signs of severe malaria or any other serious health condition , tick and thin smears and haemoglobin measurement were performed on days 2, 3, 7, 14, 21, 28, 35 and 42 or any day in between in the event of illness.Outcomes were classified according to 2009 WHO guidelines as adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) or follow-up interrupted . Follow-P. falciparum genes, such as the merozoite surface protein 1 and 2 genes (msp1 and msp2) and the glutamate-rich protein gene (glurp) were genotyped by PCR, as previously described [In areas of intense transmission, where multiple genotype infections are common , a seconescribed . PCR anaescribed .All ETFs were considered to be due to recrudescence. Patients meeting the criteria for LCF or LPF in whom genotyping was done; according to the results, patients were classified as either (i) resolved by PCR and further categorized as recrudescences or new infections or (ii) unsuccessfully genotyped with the reason recorded .This study underwent ethical review and was approved by Ethical Committee of the School of Public Health, University of Kinshasa, DRC, the ethical review board of M\u00e9decins Sans Fronti\u00e8res and the Comit\u00e9 de Protection des Personnes \u201cIle de France XI\u201d, St Germain en Laye, France. The study was authorized by the relevant health authority. Written informed consent was obtained from the parents or legal guardians of the enrolled children.Based on previous data for AL efficacy -14, effiThe primary outcome was the PCR-adjusted parasitological cure rate up to day 42 of the follow-up period. Two analytical approaches were used to assess efficacy data. First, a per protocol (PP) analysis was performed including only the patients who were followed throughout the protocol, defined follow-up period and in whom a clear treatment outcome can be determined. The risk of failure for each treatment group was calculated as the proportion of patients classified as failure (the numerator) divided by the number of patients in the evaluable population (the denominator). In the second approach, survival analysis was performed and patients with incomplete follow-up who did not reach the primary outcome interest were included in the analysis as non-failures, but censored on the last day of follow-up. The risk of failure was calculated using the Kaplan-Meier product limit formula with data censored for patients who were not classified as failures and with interrupted follow-up. Patients wrongly included, who did not meet study inclusion criteria, were excluded from both analyses.falciparum new infections were censored on the last day of observation. For the adjusted calculations, censored patients also included those with new P. falciparum infections.In the PP analysis (adjusting by genotyping), the evaluable population included only patients classified as ACPR, ETF or LCF/LPF due to recrudescence. In the survival analysis, the evaluable population for adjusted and unadjusted calculations included all patients enrolled in the study, with the exception that LCF/LPF outcomes with unsuccessful genotyping outcomes were excluded from the adjusted calculations. For the unadjusted calculations, patients with follow-up interrupted and non-Other variables were compared using the chi2 test or Fisher\u2019s exact test for variables and Student\u2019s test for continuous variables.Data were double entered and validated using Epidata version 3.1 . All analyses were performed with Stata, version 10 . A p-value\u2009<\u20090.05 was considered statistically significant.Between April 2008 and March 2009, a total of 1,993 patients were screened, 301 children aged between six and 59\u2009months were enrolled in the two treatment arms, 156 with ASAQ and 145 with AL. The mean age of patients at baseline was 27.5\u2009months (range 6\u201359\u2009months) and 152 (50.5%) were females. There were no differences between treatment arms at study inclusion, for all variables studied, including demographic, clinical and laboratory characteristics, confirming adequate randomization , taking another anti-malarial drug (1/14), severe anaemia (1/14) and other violation protocol (4/14). All these 14 patients were excluded from the analyses.The clinical trial profile is presented in Figure Among the 256 patients still followed-up at day 42, adequate clinical and parasitological response was observed in patients: 90.1% (118/131) in the ASAQ group and 84.8% (106/125) in the AL group. Overall, no patient experienced early treatment failure, and 32 patients developed late clinical or parasitological failure (9.9% (13/131) in the ASAQ group and 15.2% (19/125) in the AL group)  of 32 blood samples (Table Table Clearance of fever (defined as a temperature <37.5\u00b0C) was more rapid in patients given ASAQ than in patients given AL, one day after treatment initiation. Among patient who had fever at inclusion, one (1.3%) of 79 ASAQ recipients and eight (11.3%) of 71 AL recipients remained febrile . By day 2, nearly all fevers had resolved and the two regimens did not differ . Rapid clinical improvement was recorded within the two treatment groups.By day 2, parasite clearance did not differ between the two regimens, with positive slide results observed for nine (6.0%) of 150 ASAQ-treated patients and for seven (4.9%) or the 143 AL-treated patients . By day 3 and day 7, only one patient (respectively in the AL group and in the ASAQ group) had not yet cleared all parasites.At baseline, gametocyte carriage rates were low in both regimen groups Table . GametocBoth treatment regimens were generally well tolerated. Twelve patients (ten (6.4%) in the ASAQ group and two (1.4%) in the AL group) vomited within 30\u2009min after the administration of the dose; nine of them  had received a second dose and none was excluded from the study.No deaths occurred during the follow-up period, but six patients (one in the ASAQ group and five in the AL group) developed moderate or severe adverse events associated with clinically suspected severe malaria . All patients were transfused and one received the rescue treatment consisting on intravenous quinine . Four of these six patients  were hospitalized for two to six days. Thus, these events could not be attributed to the intervention drugs.Other mild side-effects were less common and were not related to a specific treatment: lower acute respiratory infections (54.6% in the ASAQ group and 51.3% in the AL group), conjunctivitis (16% in the ASAQ group and 10.8% in the AL group), abdominal pains and diarrhoea (8% in the ASAQ group and 14.9% in the AL group).falciparum malaria and follow-up patients for 42\u2009days show that the efficacy of fixed-dose combination of ASAQ is non-inferior to fixed-dose combination of AL, with regard to the margin prespecified at 7%.The results of this randomized trial of Congolese children in Katanga province with uncomplicated et al. in Equatorial province of DRC [High PCR adjusted cure rates of 98.3% to 98.4%  were seen in patients assigned to ASAQ, compared with rates of 99.1% to 99.2% in patients assigned to AL. These results are comparable to those from previous studies where AS+AQ were routinely used as multiple tablets (loose combination) in other sub-Saharan countries -14, but e of DRC . They are of DRC ,16.As with other malaria efficacy studies conducted in areas of high transmission , the unaP. falciparum usually constitutes an additional confounding factor for the adjusted analysis. Whereas patients with such infections are removed from the PP analyses, they are censored in the survival analysis after contributing a period of observation to the cumulative risk during which treatment failure was not observed [In this study, the adjusted estimates of treatment efficacy derived from the two statistical approaches were not significantly different. New infection with observed . This diSurvival analysis allows for more available data to contribute to the analysis, thus increasing the precision of the derived estimates. It avoids systematic biases introduced by dropping from the analysis patients who do not complete follow-up ,18,19, eDuring the first days of treatment, a rapid decrease of fever, parasitaemia and gametocyte carriage was observed in the two regimen groups. These findings confirmed the results of efficacy studies on artemisinin-containing combination which led to lower gametocyte carriage, improving the cure rates, decreasing the transmission of falciparum malaria and reducing the spread of resistance to non-artemisinin drugs ,21. ParaThis study showed that both combinations were well tolerated, with only 6% of children with drug-induced vomiting. Nevertheless, differents between treatment arms were observed in the first few days following treatment; vomiting was higher in the ASAQ group compared to AL. Only four patients developed severe malaria, and no deaths occurred during the course of the study. It is worth mentioning that the study was not designed nor power to compare tolerabilty between the two combinations. The ASAQ FDC presents the advantage of requiring one intake per day while AL is twice per day . HoweverA limitation of this study was that pharmacology measures were not performed. It would improve the differentiation between true recrudescence to a problem of absorption . The limAccording to WHO guidelines, which recommend a change in anti-malarial treatment policy when the cure rate for the current recommended therapy falls below 90% , this ste.g., shorter follow-up of patients, and validate molecular marker of artemisinin resistance.While emergence of artemisinin resistance in South East Asia is of serious concern ,23, possInitiatives, including development of paediatric formulations, home-based management of malaria and improving public sector procurement and supply chains should make ACT more accessible for sub-Saharan African children ,32.The authors declare that they have no competing interests.The authors accept full responsibility for the overall content of this report. EE was the principal investigator of the study. She designed and coordinated the study, managed and analysed the data, and wrote the manuscript. AL participated to the development of the study protocol, gave a technical support to the investigator and revised the manuscript. BA, as director of the National Malaria Control Programme in DRC, gave support and approval of the study and revised the manuscript. LF participated in the development of the study protocol, gave a technical support for the laboratory issues, coordinated the external quality control of the reading of malaria slides and revised the manuscript. EMS, as the local authority and health staff, gave support and approval of the study and revised the manuscript. MD performed the PCR analyses and gave advice on PCR results interpretation. PPPU participated in the development of the study protocol, gave technical support to the investigators and revised the manuscript. JPG gave technical support to the principal investigator during the protocol writing, analysis and writing of the manuscript."}
+{"text": "In 2008, artemether - lumefantrine (AL) and dihydroartemisinin - piperaquine (DHAP) were added to artesunate - amodiaquine (AS-AQ) as first-line drugs for uncomplicated malaria in Ghana. The introduction of new drugs calls for continuous monitoring of these drugs to provide timely information on trends of their efficacy and safety to enhance timely evidence-based decision making by the National Malaria Control Programme. In this regard, the therapeutic efficacy of AL was monitored from September 2010 to April 2011 in four sentinel sites representing the three main ecological zones of the country.The study was a one-arm prospective evaluation of clinical and parasitological responses to directly observed treatment for uncomplicated malaria among children aged 6\u2009months to 59\u2009months using the 2009 WHO protocol for surveillance of anti-malarial drug efficacy. Children recruited into the study received weight-based 20/120\u2009mg AL at 0, 8, 24, 36, 48, and 60\u2009hrs. Parasitaemia levels were assessed on days 2, 3, 7, 14, 21, 28, and at any time a study child was brought to the clinic with fever.P\u2009=\u20090.017). Fever and parasite clearance were slower among children enrolled in the savannah zone. Gametocytaemia after day-3 post-treatment was rare in all the zones.A total of 175 children were enrolled into the study: 56 in the savanna zone, 78 in the forest zone and 41 in the coastal zone. Per-protocol analysis showed that the overall PCR-corrected cure rates on day 14 and day 28 were 96.5%  and 95.4% , respectively, with statistically significant differences between the ecological zones. The 90.4%\u2009day-28 cure rate observed in the savannah zone  was significantly the lowest compared with 100%  in the forest zone and 93.8%  in the coastal zone (The study has shown that AL remains efficacious in Ghana with significant ecologic zonal differences. The savannah zone may be a potential zone for any emergence of resistant alleles as a result of the slower parasite clearance observed in the zone. Malaria remains one of the major causes of morbidity and mortality worldwide. It is estimated that in 2010 there were approximately 216 million cases of malaria worldwide, of which 81% were in Africa. In the same year, malaria accounted for approximately 655,000 deaths worldwide, of which 91% were in Africa [The current control strategy in Ghana has case management based on prompt recognition and adequate treatment as its main focus. Chloroquine had been the first-line drug for the treatment of uncomplicated malaria in Ghana for decades until 2001, when results from studies conducted in six sentinel sites (between 1998 and 2001) showed treatment failure rates of between 8.6% and 26.8% . At a coThe introduction of new drugs calls for continuous monitoring of these drugs to provide timely information on trends of their efficacy and safety to enhance timely evidence-based decision making by the National Malaria Control Programme (NMCP). In this regard, a surveillance system, coordinated by the Noguchi Memorial Institute for Medical Research (NMIMR), was established in 2005 to provide data on first-line anti-malarial drugs from 10 sentinel sites across the country. This paper covers the therapeutic efficacy of AL across three ecological zones in Ghana during the period September 2010 to April 2011 using the 2009 WHO protocol for surveillance of anti-malarial drug efficacy .The AL study was conducted in four of the 10 sentinel sites representing the three main ecological zones of Ghana. These were Navrongo War Memorial Hospital in the savannah zone; Bekwai Municipal Hospital and Begoro Government Hospital in the forest zone; and Ewim Health Centre in the coastal zone.The Navrongo War Memorial Hospital is located in the Kassena Nakana East district in the Upper East region of Ghana. The Kassena Nankana East district has an estimated population of 109,944 with an annual average rainfall of 950\u2009mm. Malaria in the district is perennial with marked seasonal variation. The peak transmission season coincides with the major rains between June and October. The Bekwai Municipal Hospital is located in the Bekwai Municipality in the Ashanti region of Ghana. The Bekwai Municipality has an estimated population of 118,024. Annual rainfall in the municipality is 1,600 \u2013 1,800\u2009mm with double maxima rainfall in May and October each year. Malaria transmission in the Municipality is intense and perennial. The Begoro Government Hospital is located in the Fanteakwa district in the Eastern region of Ghana. The Fanteakwa district has an estimated population of 108,614. Annual rainfall in the district is 150 \u2013 2,000\u2009mm with double maxima rainfall in June and October each year. Malaria transmission in the district is intense and perennial. The Ewim Health Centre is located within the Cape-Coast Metropolis in the Central region of Ghana. The Cape-Coast metropolis has an estimated population of 169,894. Annual rainfall in the metropolis is 750 \u2013 1,000\u2009mm with double maxima rainfall in June and December each year. Malaria transmission in the metropolis is perennial .Plasmodium falciparum; parasite count ranging between 1,000 and 250,000 per \u03bcl; haemoglobin level\u2009>\u20095\u2009g/dl; absence of signs/symptoms of severe malaria; and parent\u2019s willingness to give their consent.The study population involved all children aged between six and 59\u2009months presenting at the Out-Patient Department (OPD) of a study site clinic with symptoms suggestive of malaria. Once a clinical diagnosis of malaria was made by the study Nurse, samples of blood were obtained from a finger prick to prepare thick and thin smears for malaria microscopy and haemoglobin level determination. Children meeting the inclusion criteria were recruited into the study and followed up for a minimum of 14\u2009days and a maximum of 28\u2009days. Briefly, the inclusion criteria included axillary temperature\u2009\u2265\u200937.5\u00b0C or history of fever during the past 24\u2009hrs; mono-infection with Children recruited into the study received weight-based 20/120\u2009mg AL  at 0, 8, 24, 36, 48, and 60\u2009hrs. All treatments were given under direct observation by a study nurse and children were observed for 30\u2009minutes to ascertain retention of medicine. Children who vomited during the observation period were re-treated with the same dose of medicine and observed for an additional 30\u2009minutes. Children with repeated vomiting were given parenteral therapy with quinine as per national standard treatment guidelines and excluded from the study. All children were allowed use of antipyretics. Children who showed signs/symptoms of severe malaria, had serious adverse events or required blood transfusion were withdrawn from the study.msp1, msp2), and glutamate-rich protein (glurp) used to distinguish between re-infection and recrudescence. Haemoglobin levels were assessed for all study children on days 0, 14, and 28.Outpatient follow-up visits were scheduled for days 1, 2, 3, 7, 14, 21, and 28 after treatment (day of treatment was counted as day-0). At each visit to the clinic, children were examined physically and information on symptoms, axillary temperature, and respiratory rate recorded on a Case Record Form (CRF). Parasitaemia levels  were assessed on days 2, 3, 7, 14, 21, 28, and any day within the 28-day follow-up period that a child is brought to the clinic with fever. Thick and thin smears were stained with 3% Giemsa for 30\u201345\u2009minutes for quantification of asexual parasites against 200 white blood cells using a hand tally counter. Sexual parasite counts were done per 1,000 white blood cells. Parasitaemia levels were expressed per \u03bcl blood assuming white blood cell count of 8,000 per \u03bcl blood. A smear was declared negative when examination of 100 thick-film fields did not show presence of asexual parasites. For quality control purposes, all blood slides were read by two qualified independent microscopists, and discordant readings were re-examined by a third qualified independent microscopist. Discordance was defined as differences between the first and second microscopists regarding presence/absence of asexual or sexual parasites; species diagnosis; and day-0 counts meeting the inclusion criterion of 1,000 \u2013 250,000 per \u03bcl blood. The first or second reading was taken as final depending on whichever agrees with the third reading. Filter paper blots were obtained on day-0 and at recurrence of parasitaemia for PCR genotyping, and merozoite surface proteins 1 and 2 , Late Parasitological Failure (LPF), Late Clinical Failure (LCF), and Adequate Clinical and Parasitological Response (ACPR) [The Institutional Review Board of the Noguchi Memorial Institute for Medical Research, University of Ghana, reviewed and approved the study. Written informed consent was obtained from each parent/guardian at the start of the study. Each parent/guardian was informed of the objectives, methods, anticipated benefits and potential hazards of the study. They were also informed that they were at liberty to withdraw their children from the study at any time without penalty.Of the 360 children screened, 175 (48.6%) met the inclusion criteria and were enrolled into the study: 56 in the savannah zone, 78 in the forest zone, and 41 in the coastal zone. Characteristics of patients at enrolment are presented in Table\u2009Out of 169 evaluable patients assessed on day-14, 163 or 96.5% 95% CI: 92.1, 98.6) showed an adequate clinical and parasitological response. Five of the six treatment failures were early treatment failures that occurred in the savannah zone. The only late parasitological failure observed on day-14 occurred in the coastal zone and was confirmed by PCR   in the savannah zone; 67 out of 70 or 95.7%  in the forest zone; and 30 out of 38 or 79%  in the coastal zone showed adequate clinical and parasitological response   on day-2, the proportion increased to 18.2%  on day-3 before decreasing again to 7.8%  on day-7. Febrile cases were, therefore, recorded on all days within the first follow-up week in the savannah zone. The proportion of children still febrile on day-1 post-treatment was significantly higher in the savannah zone  compared with the forest zone  and the coastal zone   were parasitaemic on day-2. However, no child was parasitaemic on day-3 and day-7 . By day-2 post-treatment, prevalence of gametocytaemia had declined to 0% and remained so in the zone on days 3, 7, and 21. Days 14 and 28 in the savannah zone had only 1 child (1.8%) with gametocytes. Gametocytaemia was absent on all follow-up days among children enrolled in the forest zone. For children enrolled in the coastal zone gametocytaemia was prevalent on day-2 post-treatment at a rate of 10.5% (4/38), halved by day-3 at a rate of 5.3% (2/38), declined to 0% by day-7, and remained same throughout the remaining follow-up days   on day-0 to 11.0\u2009g/dl  on day-28 (4) Table\u2009.Following the inclusion of artemether - lumefantrine combination as one of the first-line drugs for the treatment of uncomplicated malaria in Ghana in 2008 , there wThe study has shown that the overall PCR-corrected cure rate for Ghana is 95.4%  with the savannah zone showing a significantly lower rate of 90.4% . The over 90% cure rate of AL in Ghana is comparable with findings from other studies in Africa within the past decade -20, and p\u2009<\u20090.001). Additionally, 8/55 or 14.5%  of the children enrolled in the savannah zone were parasitaemic on day-2 post-treatment whilst no child was parasitaemic on the same day in the forest and coastal zones. Out of the 8 parasitaemic children 5  had parasite count greater than the day-0 count but remained aparasitaemic on days 3, 7, 14, 21, and 28 post-treatment without any rescue treatment. The rapid rise in asexual parasitaemia after commencement of ACT treatment has been reported in Nigeria and attributed to a possible larger load of sequestered parasites in deep tissues in some patients. In the Nigeria study, parasitaemia levels peaked at one hour after the first dose of AL, and by 16\u2009hours post-treatment all parasites had been cleared among children with initial rise in parasitaemia [Fever and parasite clearance were slower among children enrolled in the savannah zone. Whereas no child was febrile on day-3 and day-7 in the forest and coastal zones, febrile cases were reported on all days among children in the savannah zone during the first follow-up week. The proportion of children still febrile on day-1 post-treatment was significantly highest in the savannah zone  compared with the forest zone  and the coastal zone   and declined to 0% by day-2 post-treatment whilst gametocytaemia was prevalent on day-2 post-treatment in the coastal zone (10.5%), halved by day-3 (5.3%), and declined to 0% by day-7 post-treatment. This finding suggests that ACT remains efficacious in gametocyte clearance in Ghana. The efficacy in gametocyte clearance has the advantage of potentially slowing down the transmission of resistant alleles .Post-treatment mean haemoglobin concentration was significantly higher than pre-treatment concentration in the coastal zone but not the forest and savannah zones. This finding suggests that the therapeutic efficacy of AL resulting from rapid parasite clearance may not necessarily translate into significant post-treatment increases in haemoglobin concentrations during a 28-day follow up period. A couple of studies in Africa have reported non-significant post-treatment increases in haemoglobin concentrations with AL treatment during a 28-day follow-up period of patients treated for acute uncomplicated malaria ,26 whilsThere is evidence to show that AL remains efficacious in Ghana with significant ecologic zonal differences. The savannah zone may be a potential zone for any emergence of resistant alleles as a result of the slower parasite clearance observed in the zone. Pharmacokinetic studies will be useful in future anti-malarial drug resistance surveillance activities in Ghana to better describe treatment failures. Ghana\u2019s policy of multiple first-line therapy is in the right direction, and needs to be supported by all stakeholders as it delays the emergence and slows spread of drug resistance ,29. FurtThe authors declare that they have no competing interests.BA and KK conceived and designed the study. ND and NQ carried out molecular genetic studies to distinguish between re-infection and recrudescence. BA and LQ coordinated the study. BA and KK participated in the data analysis. BA drafted the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium vivax is the main malaria parasite in China, and China is now making efforts to eliminate malaria by 2020. Radical cure of vivax malaria is one of challenges for malaria elimination. The purpose is to evaluate the efficacy and safety of artemisinin-naphthoquine (ANQ) versus chloroquine-primaquine (CQ-PQ) in treatment of vivax malaria in Yunnan Province, China.P. vivax were randomly assigned to receive either a total target dose of ANQ 24.5\u00a0mg/kg (naphthoquine 7\u00a0mg/kg and artemisinin 17.5\u00a0mg/kg), once a day for three days, or a total CQ dose of 24\u00a0mg base/kg, once a day for three days plus a PQ dose of 0.45\u00a0mg base/kg/day, once a day for eight days. Patients were followed up for one year. The difference in efficacy between ANQ and CQ-PQ was compared via Wilson\u2019s test.An open-label randomized and non-inferiority design, eligible patients with monoinfections of By day 42, the number of patients free of recurrence was 125  for ANQ arm and 123  for CQ-PQ, and non-significant (P\u2009=\u20090.4496). By day 365, the number was 101  for ANQ and 106  for CQ-PQ, and non-significant (P\u2009=\u20090.610). So the proportions of patients free of recurrence had no significant difference between ANQ and CQ-PQ groups by day 28, 42 and 365; compared with CQ-PQ, the side effect of ANQ was mild.ANQ is non-inferior to CQ-PQ in terms of patients free of recurrence, and safer than CQ-PQ. Plasmodium vivax is the main malaria parasite in China. Yunnan Province is a vivax and falciparum malaria co-endemic region and one of two provinces with endemic falciparum malaria. China is now making efforts to eliminate malaria by 2020 [P. vivax was first described in 1989 in Papua New Guinea [ by 2020 ,2. Radic by 2020 . The tre by 2020 ,5. Furthw Guinea , the decw Guinea .P. vivax, there might be a compelling rationale for a unified ACT strategy for vivax and falciparum malaria in all co-endemic regions [P. vivax monoinfections in Yunnan Province, China.In general, the use of artemisinin-based combination therapy (ACT) has been limited to patients with falciparum malaria and showed advantages in terms of adherence and safety -11. Howe regions . The use regions , Papua,  regions , Thailan regions  and Ethi regions . These sP. vivax malaria  were enrolled into the study. Patients were not admitted to the study if any the following criteria present: (1) pregnancy, (2) severe malaria, (3) having taken any anti-malarial drug within the past 14\u00a0days, (4) history of hypersensitivity to any of the study drugs, (5) severe dysfunction of kidney, liver and heart, (6) residence living at an altitude lower than 2,000\u00a0m, (7) unable to follow up.From February 2009 to December 2010, patients were recruited into our open-label randomized study at Tengchong County Center for Disease Control and Prevention, and Tengchong County Hospitals at the China-Myanmar border. The local transmission is from June to September in most parts of Tengchong County. Because of in malaria pre-elimination phase and an altitude higher than 2,000\u00a0m, there was only imported malaria and rarely local infection in recent years. Patients older than five years of age who weighed more than 15\u00a0kg and presented with single As soon as confirmed patients were enrolled into the study, they were randomly assigned to receive either ANQ or CQ-PQ regimens. A researcher, who did not have a role in recruitment, put sealed envelopes in blocks of 50 (25 ANQ and CQ-PQ respectively) in a box, and an enrolled patient drew an envelope from it to achieve treatment allocations in equal numbers. When the box was empty, another 50 envelopes were added.Plasmodium vivax, ANQ was given once a day for three days, with a total target dose of 24.5\u00a0mg/kg (naphthoquine 7\u00a0mg/kg and artemisinin 17.5\u00a0mg/kg). CQ was given once a day for three days with a total target dose of 24\u00a0mg base/kg and PQ was offered once a day for eight days with a dose of 0.45\u00a0mg base/kg/day.ANQ was registered by Food and Drug Administration, China as GYZZ H20050270 and licensed to Kunming Pharmaceutical Corp to produce. A tablet of ANQ is composed of 50\u00a0mg naphthoquine and 125\u00a0mg artemisinin. CQ was registered as GYZZ H31020423 and PQ as GYZZ H2005984 in China. CQ-PQ was produced and pre-packed by Shanghai Sino-West Pharmaceutical Corp. Both ANQ and CQ-PQ were administered following the anti-malarial drug policy of China. Based on the hypothesis that a three day dosing could reduce recurrence of P. vivax again, they were retreated with CQ (24\u00a0mg base/kg) for three days and PQ (0.25\u00a0mg base/kg/d) for 14\u00a0days. Patients were observed and interviewed with semi-structured questionnaires in depth for adverse reactions and compliance during administering each dose and follow-up visits on days 1, 2, 3, 7, 14, 21 and 28. Patients were followed up for one year through monthly visits.The researchers visited all patients every 8\u00a0hrs for the first three days. Axillary temperatures were measured every 8\u00a0hrs after treatment until after 48\u00a0hrs of fever clearance. Thick and thin blood smears were taken and examined every 8\u00a0hrs in each active visit, and then day 7, 14, 21 and 28 respectively. The subsequent PQ doses of CQ-PQ groups were given under supervision of patient caretakers one hour after supper and before going to bed from the fourth day. Patients received the remaining PQ doses packed by aluminium-plastic foil and were instructed clearly about their subsequent treatment, emphasizing the importance of taking drugs after food and before going to bed, and taking their medicines even when their symptoms had subsided. The patients were asked to return for treatment in any case that they had dark urine; this was done instead of testing for G6PD deficiency because of shortage of reagent kit and equipment supplies. Patients were visited and interviewed every month; meanwhile blood smears were done during the monthly visits, and asked to return for screening and treatment at any time, if they became ill. If a patient had fever, blood smears were prepared and examined for malaria at any time. Microscopists examining blood films were unaware of treatment allocation. If any patients were positive of Malaria blood films were stained with Giemsa, and both asexual parasites and gametocytaemia were counted per 500 white blood cells. The number of parasites was calculated as per \u03bcl of blood by the level of 8,000 of leukocyte per \u03bcl . ParasitThe primary end point was the proportion of patients free of recurrence at day 42, and the secondary end points included at day 28 and 365, the parasite and fever clearance times, and the adverse events reported by patients during follow-up.Based on calculation recommended by literature , a samplx2 tests. Mean fever and parasite clearance time were compared by covariance.The difference of patients free of recurrence and its two-sided 95% confidence intervals between ANQ and CQ-PQ arms were calculated via Wilson\u2019s test. Proportions were compared by using Yates corrected The study was reviewed and approved by The Academic Board of Yunnan Institute of Parasitic Diseases (YIPD) in China as protocol 200807. Approval was also obtained from YIPD\u2019s ethics committees. The purpose of the study was explained and approval was sought from patients and their caretakers. Informed written consent was obtained from patient or caretakers of child patients. All results were kept confidential and were unlinked to any identifying information.P. vivax. 401 met the enrollment criteria, but 141 did not agree to participate in the study. 260 were randomly assigned to one of treatment groups. 128 (49.2%) and 132 (50.8%) received ANQ and CQ-PQ respectively. Three (1.2%) have not completed the 365\u00a0day follow-up and two (0.8%) withdrew from the study on day 1 . When these patients knew that they might take CQ-PQ for eight days, they were not willing to be involved in the trial. Of 17,619 fever patients screened for malaria, 425 had The proportion of patients with recurrence in the ANQ group were not significant (P\u2009=\u20090.4496) to that in the CQ-PQ arm by day 42 Table\u00a0.All 255 patients cleared parasitaemia by day 3, ANQ treatment group by hour 48 and CQ-PQ by hour 64. 50% parasite, full parasite, gametocyte clearance times of ANQ were shorter than of CQ-PQ. ANQ cleared parasitaemia very rapid, the proportion of patients with parasitaemia at 24\u00a0hrs after therapy was significantly lower than of CQ-PQ. The fever clearance time (FCT) of ANQ was significantly shorter than of CQ-PQ group  was higher than CQ 74.3%) by day 42, but longer fever clearance times (FCT) and parasite clearance times (PCT) than of CQ . ANQ wassitaemia . In Thai4.3% by dan in CQ . All thePlasmodium falciparum. The finding in Papua New Guinea showed that the lower single ANQ dose was associated with relatively frequent recurrence of P. vivax[Artemisinin has a short half-life. A single dose or a two-day dosing of ANQ is usually for treatment of  P. vivax. ConsideP. vivax infections was declining on the Thai-Myanmar border [P. vivax at nine weeks was 79.1% in patients treated with only CQ and 54.9% treated with dihydroartemisinin- piperaquine on the Thai-Myanmar border [P. vivax at 365\u00a0days were respectively 21.05% in patients treated with ANQ and 17.2% treated with CQ-PQ. This indicated that PQ and NP significantly reduced recurrence with P. vivax. This might attribute to two reasons. One is that NP has a long half-life despite ANQ cannot kill the liver stages, whereas CQ-PQ can; the other is that no evidence documented that less than 14\u00a0days PQ can radically cure P. vivax[The efficacy of CQ in the treatment of r border  and in Vr border . The cumANQ had a mild side effect. However, two patients could not complete the treatment because of PQ toxicity. 2064 persons were screened for G6PD deficiency in China-Myanmar border region in July 2009. The result was 2.3% 95%CI, 1.7-3.0%) of prevalence of G6PD deficiency (not published). These showed that using CQ-PQ has an intrinsic problem in the region. Patients of vivax malaria can obtain 8-day CQ-PQ regimen free from public health sector in China. As a reality, malaria patients want to get rid of symptoms as quick as possible. They commonly seek artemether or pyronaridine injection by self-medication or from private clinics because they considered injection working better and faster than oral tablets, and intolerability of the 8-day CQ-PQ regimen. All the update studies showed that ACT was well tolerated with no serious adverse events -3.0% of ,20. TherP. vivax infections and is a good option for Yunnan Province of China if people do not want to take PQ.In terms of efficacy, the three-day ANQ regimen is as effective as the 8-day CQ-PQ, safer and more acceptable than CQ-PQ, but ANQ does not prevent relapse completely. ANQ is a great blood schizonticide for The authors declare that they have no competing interests.HL, H-L Y designed the study and developed the protocol. J-W X and HL analyzed and interpreted the data. HL supervised the clinical trial. HL, J-Z W, R-H N and C-F L conducted the clinical trial, and entered the data. J-WX and HL wrote the first draft of the paper. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria over the last decade, which can to a large extent be attributed to the wide deployment of insecticide-treated bed nets and the introduction of the highly effective artemisinin combination therapies. Effective early asexual stage antimalarial treatment will prevent the transition to early developing gametocytes, but in addition the artemisinin derivatives affect young gametocytes, which augments their transmission-reducing capacity. However, mature gametocytes responsible for transmission of the disease can be present on first presentation and last for several weeks even with artemisinin-based combination therapy (ACT). The 8-aminoquinolone primaquine is the only drug in general use with gametocytocidal activity against these mature forms, and can be an important adjunct to reduce malaria transmission essential for the ultimate aim of malaria elimination. This aim has gained importance with the emergence of partial resistance to artemisinins on the Cambodian-Thai and Thai-Myanmar borders, which has the potential to spread to other places in the region [The world has seen a significant reduction in the burden of e region . Containe region . DeploymClinical Infectious Diseases makes a contribution to establish the risk-benefit profile of primaquine as an adjunct to ACT in the treatment of uncomplicated falciparum malaria in South Sumatra, Western Indonesia [P = .002). In patients without gametocytes on day 3, none of the patients in either study arm developed microscopically detectable gametocytemia. The addition of primaquine was well tolerated in this population. In particular, gastrointestinal complaints were similar in the 2 treatment arms, as were the hemoglobin concentrations on 7 and 42 days after start of treatment. Follow-up was remarkably complete. The authors conclude that in settings where screening for G6PD is feasible, a single gametocytocidal dose of primaquine should be added to ACT for the treatment of falciparum malaria in low-transmission settings. However, as the authors recognize, routine screening for G6PD may not be feasible in most treatment settings.In this context, the well-designed and carefully executed open randomized controlled trial by Sutanto and colleagues published in this issue of ndonesia . Prevalendonesia , but no More practical point-of-care dipstick format tests to detect G6PD deficiency are under development, which will facilitate testing in the future. Prevalence of G6PD deficiency in malaria-endemic areas is usually somewhere between 5% and 20% [For this reason, the World Health Organization (WHO) has recently convened an evidence review group on the use of single-dose primaquine as a gametocytocidal drug in the treatment of falciparum malaria . On the The timing of the primaquine dose in the revised guideline is on the first day of treatment, instead of the third day in the study by Sutanto. The authors base this delay in dosing on a mathematical modeling study showing that postponing the dose increases its transmission blocking effect. An optimal effect is predicted with a single primaquine dose on day 8, related to the fact that in falciparum malaria the peak in gametocyte density is usually delayed by around 1\u20132 weeks after the initial acute attack . HoweverIn conclusion, the study of Sutanto and colleagues clearly demonstrates the efficacy and safety of a single dose of 0.45 mg/kg primaquine on the third day of treatment as a gametocytocide drug in the treatment of uncomplicated falciparum malaria, excluding patients with G6PD deficiency. Dose and timing differ from the recently adopted WHO guideline, recommending a single dose of 0.25 mg/kg primaquine on the first day of treatment in similar settings, without prior testing for G6PD enzyme activity. Further studies will be needed to provide additional evidence for the optimal timing and dose of an efficacious yet safe deployment of primaquine as gametocytocidal and transmission-blocking treatment in populations with a high prevalence of G6PD deficiency."}
+{"text": "Artemether-lumefantrine (AL) was adopted as first-line treatment for uncomplicated malaria in Kenya in 2006. Monitoring drug efficacy at regular intervals is essential to prevent unnecessary morbidity and mortality. The efficacy of AL and dihydroartemisinin-piperaquine (DP) were evaluated for the treatment of uncomplicated malaria in children aged six to 59 months in western Kenya.Plasmodium falciparum mono-infection were enrolled in an in vivo efficacy trial in accordance with World Health Organization (WHO) guidelines. The children were randomized to treatment with a three-day course of AL or DP and efficacy outcomes were measured at 28 and 42 days after treatment initiation.From October 2010 to August 2011, children with fever or history of fever with uncomplicated A total of 137 children were enrolled in each treatment arm. There were no early treatment failures and all children except one had cleared parasites by day 3. Polymerase chain reaction (PCR)-uncorrected adequate clinical and parasitological response rate (ACPR) was 61% in the AL arm and 83% in the DP arm at day 28 (p\u2009=\u20090.001). PCR-corrected ACPR at day 28 was 97% in the AL group and 99% in the DP group, and it was 96% in both arms at day 42.AL and DP remain efficacious for the treatment of uncomplicated malaria among children in western Kenya. The longer half-life of piperaquine relative to lumefantrine may provide a prophylactic effect, accounting for the lower rate of re-infection in the first 28 days after treatment in the DP arm. Plasmodium falciparum has been an ongoing global public health concern since chloroquine resistance emerged in the 1960s [Resistance to anti-malarials by P. falciparum malaria until 1998, despite the presence of high levels of chloroquine resistance since the early 1990s [In Kenya, chloroquine was first-line treatment for uncomplicated ly 1990s . In 1998Artemisinins, the newest class of anti-malarials, have a very short half-life (<8 hours) and rapidly reduce parasite burden; their use in combination with other anti-malarials decreases the chance of resistance emergence . HoweverP. falciparum malaria in Kenya.Dihydroartemisinin-piperaquine (DP), another ACT regimen, has been studied in East Africa as an alternative to AL -10. Its Regular monitoring of anti-malarial efficacy is essential to better inform national malaria policies . This stP. falciparum with high malaria transmission and two seasonal peaks, April to July and November to December. The entomological inoculation rate (EIR) in this area, historically around 300 infectious bites per person per year, has recently been estimated to be ten infectious bites per person per year . Study subjects were recruited from the outpatient paediatric department of SDH, which serves approximately 100 patients per day.This study was conducted between October 2010 and August 2011 at Siaya District Hospital (SDH) in Nyanza Province, western Kenya. This region is holoendemic for P. falciparum mono-infection were enrolled. Additional inclusion criteria were axillary temperature \u226537.5\u00b0C or history of fever in the previous 48 hours, weight \u22655.0 kg, parasitaemia 1-200,000 asexual forms per \u03bcL , residence within 10 km of SDH, and written informed consent by caregiver. Subjects were excluded if any of the following were present: lethargy, convulsions, inability to drink, persistent vomiting, symptoms of severe malaria, severe malnutrition  standards), severe anaemia (haemoglobin (Hb) <5 g/dl), known hypersensitivity to study drugs, presence of febrile illness other than malaria , presence of chronic medical conditions, treatment with any anti-malarial in the previous two weeks, or previous enrolment in any malaria study.Children aged six to 59 months with This study received ethical clearance from the US Centers for Disease Control and Prevention  and the Kenya Medical Research Institute . Written informed consent was obtained from caregivers of enrolled subjects and a long-lasting insecticide-treated bed net (ITN) was provided to enrolled subjects.in vivo trial [This was a 42-day, open-label vo trial . InitialCaregivers of enrolled children were interviewed and children were examined by a study clinician. Children were block randomized in fixed blocks of ten to treatment with AL  or DP . Samples of the AL and DP used in this study were sent to CDC laboratories for quality testing using high-performance liquid chromatography (HPLC) . Both treatments were co-formulated, fixed-dose ACT regimens and were administered under direct observation by study staff at the study clinic (except AL evening doses) for three consecutive days. AL tablets, consisting of 20 mg of artemether and 120 mg lumefantrine, were administered twice daily according to patient weight: 5-14 kg: one tablet per dose; weight 15-24 kg: two tablets per dose; weight 25-34 kg: three tablets per dose. Morning doses were given with milk and directly observed in the study clinic. Caregivers were given evening doses to administer at home with food or milk. DP tablets, consisting of 20 mg dihydroartemisinin and 160 mg of piperaquine phosphate, were administered once daily by study staff according to patient weight: 5-6 kg: one-half tablet daily; 7-9 kg: one tablet daily; 10-14 kg: two tablets on day 0, then one tablet on days 1 and 2; 5-19 kg: two tablets daily. A full dose was re-administered if the patient vomited within 30 min or a half dose if vomiting occurred between 31 and 60 min. Patients with vomiting within 30 min of the second dose were referred for parenteral treatment and withdrawn from the study.Children were followed for 42 days and asked to return on days 1, 2, 3, 7, 14, 21, 28, 35, and 42 following enrolment, as well as any day if ill. The study clinic was open daily during regular hours; study personnel provided after-hours care at SDH. A clinical assessment was performed and blood smears were collected at each study visit. Hb levels were measured on days 0, 7, 14, 28, and 42. A filter paper blood spot was collected on days 0, 3, and 7 and in case of suspected treatment failure for molecular analysis. Adverse events were investigated and addressed.P. falciparum parasitaemia occurring between day 7 and 28 or 42 without fever were classified as late parasitological failure (LPF). Those with fever occurring between day 4 and day 28 or day 42 with parasitaemia were classified as late clinical failure (LCF). If no failure was recorded by day 28 or day 42, the outcome was classified as adequate clinical and parasitological response (ACPR). All treatment failures with uncomplicated malaria were treated with AL and treatment failures with severe malaria were treated with parenteral quinine. Follow-up ended once a study subject met one of the four classification criteria: ETF, LPF, LCP or ACPR.Efficacy was assessed by clinical and parasitological outcomes using WHO definitions . Childremsp2), glutamate-rich protein (glurp), and merozoite surface protein-1 (msp1) was performed by PCR using filter-paper blood spots [msp2, glurp, and msp1) in the pre- and post-treatment samples.To differentiate between recrudescence and re-infection, a genotypic analysis based on merozoite surface protein-2  for continuous variables. A two-sided p-value <0.05 was considered statistically significant.Study forms were scanned into a Microsoft Access 2000  database. Statistical analysis was performed using SAS\u00ae 9.2 . Per protocol (PP) analysis of outcomes excluded those children withdrawn from the study for any reason. Intention-to-treat (ITT) analysis was performed using survival analysis. Kaplan-Meier curves were estimated for both 28 and 42 days of follow up; the log-rank test was used for comparing the curves. For ITT analysis, all withdrawals, losses to follow up and treatment failures were censored on the last day of follow up. Comparisons were made using \u03c7P. falciparum mono-infection. Of these remaining 324 eligible children, 50 (15%) did not meet other inclusion criteria or did not give consent. Two hundred and seventy-four children were enrolled in the study, 137 in each arm. Among those enrolled, 224 were included in the day 42 analysis, 111 in the AL arm and 113 in the DP arm. Baseline characteristics of the children enrolled in the two arms were similar . PCR-uncorrected results showed that the day 42 LCF and LPF were 18% and 37% in the AL arm, respectively; while LCF was 10% and LPF was 36% in the DP arm. The re-infection rate was similar for the two arms at day 42 (p\u2009=\u20090.7). After PCR correction, ACPR for both arms was 96%. Similarly, the children for whom PCR results were missing were excluded from analysis (ten in AL arm and four in DP arm).To ensure the results of the PCR-corrected analysis were not biased by missing samples, sensitivity analyses were performed assuming all missing PCR results were due to re-infection and all were due to recrudescence, resulting in a range of PCR-corrected ACPR. For day 28, the maximum ranges of PCR-corrected ACPR for the treatment arms are 91-97% (95% CI: 85-100%) in the AL arm and 98-99% (95% CI: 96-100%) in the DP arm. For day 42, ACPR would be 87-96% (95% CI: 81-100%) in the AL arm and 92-96% (95% CI: 87-100%) in the DP arm.At day 28, the survival analysis using the ITT definition demonstrated a PCR-uncorrected cure rate of 67% for AL and 85% for DP (p\u2009=\u20090.0004)  and five (4%) children remained parasitaemic on day 2 in the AL and DP arms, respectively. One child in the study remained parasitaemic on day 3 (AL arm). Over 90% of children were afebrile by day 1 in both treatment arms. Mean Hb of children who were not re-infected increased from a baseline of 9.8 g/dL to 11.6 g/dL at day 42, whereas the mean Hb of those re-infected increased from a baseline of 9.9 g/dL to 11.1 g/dL on the last study day (p\u2009=\u20090.9). The change in Hb from baseline to the study endpoint was similar among the two study arms. Drug samples tested for quality assurance contained adequate concentrations of active ingredients.There were three children in the DP arm who vomited the drug twice following enrolment and were referred for alternative treatment. The rates of vomiting for the first dose of medication were similar for AL and DP  and not associated with age.Three enrolled children developed severe malaria more than 28 days after treatment; two in the AL arm and one in the DP arm. These children were hospitalized for parenteral treatment. These outcomes are attributable to re-infection , not poor efficacy of treatment regimens. All children recovered completely; no deaths occurred during the study.P. falciparum malaria in children in western Kenya. Recurrent parasitaemia among children under five years is frequent in this area despite high coverage with ITNs (70% household ownership and 42% usage in children under five years) [P. falciparum. As seen in other African countries, recurrent parasitaemia occurs significantly more frequently in those children treated with AL in the first 28 days [Both AL and DP are efficacious in treating uncomplicated e years)  and is m 28 days ,11. ThisP. falciparum in children in the study area. Lastly, data collected in 2009 in the Mbita region of western Kenya had similar PCR-corrected ACPR in the AL group (98.6% at day 28 and 97.2% at day 42) as what was observed in this evaluation, which further demonstrates the continued efficacy of AL in the region [Data from similar studies conducted in western Kenya from 2005 to 2009 assessing the efficacy of AL and DP had similar results. Data collected in 2005 on children with uncomplicated malaria and treated with AL showed PCR-uncorrected ACPR at day 28 of 71% and at day 42 of 41%, compared to 61% and 44% in this study . Howevere region .There is growing concern about the emergence of artemisinin resistance, signalled by delayed parasite clearance, as observed in Southeast Asia . One recPlasmodium vivax and Plasmodium ovale, lack of access to care resulting in delayed treatment, asymptomatic carriers, and frequent anti-malarial stock-outs [Although the annual EIR in western Kenya has decreased dramatically, from 300 infectious bites per person per year in 1990  to ten iock-outs . This epBoth drug regimens were well tolerated with a low frequency of vomiting. Although the frequency of vomiting the first dose was similar between the two treatment arms, only those in the DP arm (n\u2009=\u20093) vomited the drug twice and required alternative treatment. Increased vomiting in children aged six to 24 months with uncomplicated malaria treated with DP, compared to AL, has been noted previously . VomitinP. falciparum mono-infection. This increased the risk of including false positives; however, only certified microscopists read slides and only nine subjects had parasitaemia <2,000 parasites/\u03bcL. Lastly, contrary to the original protocol, two months after initiation of recruitment, study staff excluded some children with any history of vomiting. The number of children excluded for this reason is unknown, as only persistent vomiting was an original exclusion criterion. Therefore, this study may underestimate the incidence of vomiting associated with the treatment regimens, as well as inadvertently excluding some children with high parasitaemia or high fever.There are a few limitations to this study. First, the evening doses of AL were not directly observed. Although caregivers were asked to confirm administration of evening doses and children who missed doses were withdrawn, the efficacy of AL may be underestimated in this study if some missed doses were not reported. Similar studies have used the same approach . HoweverDP may benefit children in this high-transmission setting because the long half-life of piperaquine is associated with a decreased re-infection rate in the first 28 days after treatment compared to AL. This longer prophylactic effect may allow more time for Hb recovery, thus decreasing the severity of re-infections. The once-a-day dosing is another advantage of this regimen and may improve adherence. However, the higher cost of a treatment dose of DP compared to AL, US$4 and US$1, respectively, may be a barrier to its use as first line. In addition, DP may be more prone to the development of resistance because of the long half-life of piperaquine.P. falciparum malaria in western Kenya. With day 3 parasite clearance rates of nearly 100%, there is no evidence of delayed parasite clearance to indicate emerging artemisinin resistance. Following WHO recommendations, regular monitoring to evaluate anti-malarial efficacy at least every two years should be maintained to confirm the continued efficacy of first-line anti-malarial therapy.The results of this study demonstrate that AL and DP remain efficacious treatment regimens for uncomplicated The authors declare that they have no competing interests.All authors contributed to the design of the study and assisted with data interpretation. KO carried out the molecular genetic studies. MM, MD, LS, SK and SPK conceived of the study, and participated in its design and coordination as well as manuscript production. AA participated in the design and coordination of the study, performed data analysis and drafted the manuscript. PO and CO participated in the coordination of the study and data analysis. JW contributed to the design of the study and assisted in statistical analysis. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria. However, limited efficacy and tolerability data are available on alternative forms of ACT in Vietnam in case there is a reduction in the susceptibility of dihydroartemisinin-piperaquine. A study was conducted to compare the efficacy and tolerability of two fixed-dose formulations of ACT, artemisinin\u2013piperaquine  and artesunate-amodiaquine  for the treatment of P. falciparum malaria in south-central Vietnam.In Vietnam, the artemisinin-based combination therapy (ACT) of dihydroartemisinin-piperaquine is currently used for first-line treatment of uncomplicated A randomized, open-label trial was conducted comparing the efficacy of a two-day regimen of ARPQ (~2.8\u2009mg/kg artemisinin plus ~17.1\u2009mg/kg of piperaquine per day) and a three-day regimen of ASAQ (~4.7\u2009mg/kg of artesunate plus ~12.6\u2009mg/kg of amodiaquine per day) for the treatment of children and adults with uncomplicated falciparum malaria. Primary efficacy endpoint was day 42, PCR-corrected, parasitological cure rate. Secondary endpoints were parasite and fever clearance times and tolerability.vs. 36\u2009h, P<0.001) and fever clearance times were shorter in the ASAQ group . The two forms of ACT were well tolerated with no serious adverse events.Of 128 patients enrolled, 63 were administered ARPQ and 65 ASAQ. Of the patients who completed the 42\u2009days follow-up period or had a recurrence of malaria, 55 were on ARPQ  and 59 were on ASAQ . Recrudescent parasitaemia was PCR-confirmed for one patient in each treatment group, with cure rates at day 42 of 98% (95% CI: 88\u2013100) for both forms of ACT. The median parasite clearance time was significantly slower in the ARPQ group compared with the ASAQ group (48\u2009h P. falciparum malaria. Although the two-day course of ARPQ was equally as effective as the three-day course of ASAQ, parasite and fever clearance times were shorter with ASAQ. Further studies are warranted in different regions of Vietnam to determine the nationwide efficacy of ASAQ.Both forms of ACT were highly efficacious in the treatment of uncomplicated Australian New Zealand Clinical Trials Registry Number, ACTRN12609000816257 Plasmodium falciparum malaria worldwide, WHO recommends artemisinin-based combination therapy (ACT), such as artemether-lumefantrine, artesunate-amodiaquine, artesunate-mefloquine, artesunate-sulphadoxine-pyrimethamine and dihydroartemisinin-piperaquine [In 2010, the World Health Organization (WHO) estimated that half of the world\u2019s population was at risk of malaria, with an estimated 216 million cases and about 655,000 deaths [eraquine . Of theseraquine -8.P. falciparum malaria are of immense concern, as the Cambodia-Thailand border region has been the epicentre of anti-malarial drug resistance [Since the mid-1990s, artesunate plus mefloquine , usually given as separate tablets, has been the treatment of choice for multidrug-resistant falciparum malaria in Southeast Asia, particularly along the north-western border of Thailand, where mefloquine resistance is prevalent  and Cambsistance , which hP. falciparum in Vietnam [In Vietnam, the three-day fixed-dose combination regimen of dihydroartemisinin-piperaquine (Arterakine\u00ae) is now the recommended first-line ACT for the treatment of uncomplicated falciparum malaria . Althoug Vietnam ,17, ther Vietnam .Anopheles dirus and Anopheles maculatus are the main malaria vectors on the forested hill areas near the commune [P. falciparum mono-infection with a parasite density of 200\u2013200,000 asexual parasites/\u03bcL; tympanic temperature \u226538\u00b0C at the time of enrolment or history of fever during the preceding 24\u2009h and willingness to be followed-up for 42\u2009days after starting treatment. Exclusion criteria were as follows: patients with symptoms and/or signs of severe malaria; evidence of another serious medical disease; a history of drug or alcohol abuse; anti-malarial treatment within the preceding seven days; mixed plasmodial infection or pregnant and lactating females. Written informed consent was obtained from each adult patient or from parents or legal guardians of enrolled children. The study was ethically approved by the Vietnam Ministry of Health, Vietnam People\u2019s Army Department of Military Medicine and the Australian Defence Human Research Ethics Committee .The study was conducted at Phuoc Chien Commune, Thuan Bac District, Ninh Thuan Province in south-central Vietnam, located about 300\u2009km north of Ho Chi Minh City. The commune has a population of about 4,000 people, with most belonging to the Ra-glai ethnic group. Malaria transmission is low and occurs perennially with two peaks (May-June and October-November) . Anophel commune . Study pP. falciparum and a three-day regimen of chloroquine (25\u2009mg/kg) for Plasmodium vivax as per national policy.In the open-label, randomized study the patients were sequentially allocated to the two treatment groups: artemisinin-piperaquine  and artesunate-amodiaquine , with the first patient receiving ARPQ, the second patient ASAQ, the third patient ARPQ, and so on. A weight-based regimen of ARPQ (2.8\u2009mg/kg artemisinin plus ~16.7\u2009mg/kg piperaquine per day for two days) and ASAQ (~4.4\u2009mg/kg artesunate plus ~12.0\u2009mg/kg amodiaquine per day for three days) based on a 45\u2009kg adult was rounded up or down to the nearest half tablet. Both forms of ACT were administered under direct observation and the drugs were given with condensed milk. If a patient vomited the drugs within 30\u2009min, a full dose was readministered. The rescue medication was a seven-day regimen of artesunate (4\u2009mg/kg on the first day and then 2\u2009mg/kg daily for six days) to be implemented if either parasitaemia had not declined by at least 75% within 48\u2009h of commencing treatment with ARPQ or ASAQ or parasites were still present on day 7 after starting treatment. Patients with parasite reappearance were treated with the seven-day regimen of artesunate for Although artesunate plus amodiaquine given as separate tablets has been reported to be highly efficacious (cure rate 98%), the efficacy of the two-day regimen of ARPQ, with a 42\u2009day follow-up period is unknown. WHO recommends that regardless of the treatment rate of failure, a minimum sample size of 50 patients in clinical trials is required in order for a study to be representative . Thus, tClinical assessment and parasite density counts were performed on days 0, 1, 2, 3, 7, 14, 21, 28, 35 and 42 or on any day of recurrent malaria infection. Thick and thin blood films were collected before and every 12\u2009h after commencement of treatment until blood films were negative for three consecutive examinations. The Giemsa-stained blood films were examined by two microscopists and parasitaemia was quantified by examination of thick film fields  against 200 leukocytes, assuming a total leukocytes count of 8,000/\u03bcL. Parasite clearance time was the time in hours from starting treatment until the asexual parasite count fell below detectable levels in thick blood films. Patient\u2019s tympanic body temperature was measured immediately before and then every 12\u2009h after starting treatment until their temperature was <38\u00b0C for two consecutive days. Fever clearance time was the time in hours from starting treatment until the patient\u2019s tympanic body temperature remained <38\u00b0C for more than 48\u2009h.P. falciparumP. vivaxPlasmodium ovale and Plasmodium malariae using specific PCR primers [P. falciparum DNA (day 0 and day of recurrence of parasitaemia) for allelic variation at three loci . If at least one allele of the recurrent parasitaemia was different to the before treatment parasitaemia, the infection was considered a reinfection.Blood spots on filter paper  were collected at the same time as blood smears. PCR parasite genotyping was done to confirm plasmodial species using the QIAamp DNA mini kit  for DNA extraction, single round PCR detection and identification of the four species,  primers . In ordeHow do you feel since you took the last tablets?\u2019 Since malaria infection, particularly during the acute phase can cause adverse effects such as nausea, abdominal pain, headache and dizziness, which often makes it difficult to distinguish disease effects from drug effects [Drug tolerability was assessed clinically. An adverse event was defined as any sign or symptom that occurred or became severe during the study regardless as to whether it was related to the medication. Adverse events were recorded by the physician at each drug administration and patients where asked to respond to the non-leading question \u2018 effects , no causg for 5\u2009min. The separated plasma was transferred to Nunc cryopreservation tubes and stored at \u221225\u00b0C. The plasma samples were later transferred on dry-ice to the Military Institute of Hygiene and Epidemiology in Hanoi and then shipped to the Australian Army Malaria Institute in Brisbane, where they were stored at \u221280\u00b0C until analysis.Adult patients were invited to provide a blood sample for drug analysis on day 7 after commencement of treatment. Blood (5\u2009mL) was obtained by venipuncture, transferred to heparinized tubes and centrifuged at 1,400\u2009The reference compounds of piperaquine, amodiaquine, desethylamodiaquine and amodiaquine anologue were donated by WWARN . Plasma t-butyl ether. The contents were mixed for 20\u2009min, centrifuged  and the organic phase transferred to a new polypropylene tube. The organic phase was evaporated at 40\u00b0C using instrument grade air and the residue was reconstituted with 150\u2009\u03bcL of mobile phase of which 50\u2009\u03bcL was injected onto the HPLC. The retention times were about 6.5\u2009min for desethylamodiaquine, 8.5\u2009min for amodiaquine and 11.0\u2009min for the internal standard. The interday precision was 9.6% at 5\u2009ng/mL, 2.4% at 50\u2009ng/mL and 0.4% at 500\u2009ng/mL for amodiaquine. Corresponding interday precision for desethylamodiaquine were 4.0%, 6.2% and 6.0%. The extraction efficiencies were 76% and 71% for amodiaquine and desethylamodiaquine, respectively. The LLOQ for amodiaquine and desethylamodiaquine was 5\u2009ng/mL.Plasma amodiaquine and its principal biologically active metabolite, desethylamodiaquine concentrations were measured by HPLC using a Agilent 1100 Series consisting of Pump Model 1100-G1310A, Autosample Model 1100-G131A and Absorbance UV/VIS Detector Model G1314A set at 342\u2009nm. The column used was a Zorbax SB-CN cartridge  with Zorbax SB-CN  guard column. The mobile phase consisted of acetonitrile: 1\u2009M sodium perchlorate: 0.1\u2009M phosphate buffer pH 2  with a flow rate of 1\u2009mL/min. Samples were prepared for analysis as follows: to a 10\u2009mL polypropylene tube were added plasma (500\u2009\u03bcL), amodiaquine analogue  and 1\u2009mL of acetonitrile. The contents were vortexed for 30\u2009sec followed by the addition of 2\u2009mL ammonia (specific gravity 0.88) and 5\u2009mL methyl t-test and the Mann\u2013Whitney U test, respectively. The primary efficacy endpoint was PCR-adjusted 42-day cure rates after starting treatment. Secondary endpoints were parasite and fever clearance times and the occurrence of adverse events. Demographic and efficacy data were assessed by means of a per-protocol analysis, with recipients of rescue treatment counted as failures and new infections as cured. A Kaplan-Meier survival analysis with a log-rank test for statistical significance was applied. Differences in proportions were compared using the \u03c72 or Fisher\u2019s exact test. Statistical significance was defined as a P<0.05.All data were analysed using SigmaStat  and STATA 10.0 . Descriptive statistics were used to summarize baseline values and demographic data. Normally distributed and nonnormally distributed data were compared using the Student\u2019s\u2009P\u2009=\u20090.84). For both treatments, the geometrical mean parasitaemia was about two-fold higher in children compared with adults, but the differences were not significant . At the time of admission, 64% (35/55) of patients on ARPQ and 59% (35/59) on ASAQ were febrile.The study was conducted between May 2008 and December 2009, with 128 patients enrolled consisting of 63 patients on ARPQ and 65 patients on ASAQ  of 2.8\u2009mg/kg (2.5 to 3.1) artemisinin and 17.0\u2009mg/kg (15.0 to 18.8) piperaquine. Corresponding values for children were 2.9\u2009mg/kg (2.2 to 3.2) artemisinin and 17.3\u2009mg/kg (13.4 to 19.2) piperaquine. For the three-day regimen of ASAQ, adults were administered daily 4.4\u2009mg/kg (4.1 to 4.8) artesunate and 11.8\u2009mg/kg (11.0 to 13.0) amodiaquine. Corresponding values for children were 5.0\u2009mg/kg (4.2 to 5.3) artesunate and 13.5\u2009mg/kg (11.3 to 14.4) amodiaquine.vs. 36\u2009h, P<0.001) and parasite clearance tended to be faster in adults than in children . The median fever clearance time was slower in the ARPQ group than in the ASAQ group but the difference was not significant .Treatment outcomes as per-protocol analysis for the two treatment groups are summarized in Table\u2009For each treatment group only one patient had a recrudescence by day 42 of follow-up. The patient in the ARPQ group was a 14-year old male and at admission was febrile with a parasitaemia of 7,096 parasites/\u03bcL. His parasite clearance time was 48\u2009h and his daily dose of artemisinin and piperaquine were 3.3\u2009mg/kg and 19.7\u2009mg/kg, respectively, which was slightly higher than the corresponding median doses of 2.8\u2009mg/kg and 17.1\u2009mg/kg for the cured children. He was diagnosed with a recrudescence on day 12 after starting treatment. In contrast to the ARPQ failure, the patient in the ASAQ group was a 28-year old male and at admission was febrile with a parasitaemia of 23,092 parasites/\u03bcL. His parasite clearance time was 36\u2009h and his daily dose of artesunate and amodiaquine were 3.6\u2009mg/kg and 9.8\u2009mg/kg, respectively, which was less than the corresponding median doses of 4.4\u2009mg/kg and 11.9\u2009mg/kg for the cured adults. He was diagnosed with a recrudescence on day 33 after commencing treatment.P\u2009=\u20090.96). Seven patients in the ARPQ group and nine patients in the ASAQ group had reinfections over the 42-day follow-up period. When combining recrudescences and reinfections, the cure rates were 87% (95% CI: 71%-91%) for ARPQ and 85% (95% CI: 73%-92%) for ASAQ (P\u2009=\u20090.70). During the first 28\u2009days of follow-up, no patient on ARPQ had a reinfection, whereas two patients on ASAQ were diagnosed with new infections of P. falciparum on days 21 and 28. For each treatment group, three patients with reinfections were diagnosed with P. vivax malaria. Although not used in the analysis, of the protocol violations, one patient in each treatment group had a parasitaemia greater than 200,000 parasites/\u03bcL and both were successfully treated. Furthermore, the three patients with mixed infections of P. falciparum and P. vixax malaria at admission (two on ARPQ and one on ASAQ), based on subsequent PCR analysis, none had a recurrence of malaria during the follow-up period.The PCR-adjusted cure rates were 98% for the ARPQ group (95% CI: 88%-100%) and 98% (95% CI: 88%-100%) for the ASAQ group (None of the patients had a serious adverse event and both treatments were well tolerated. The most common adverse events reported before and after treatment are shown in Table\u2009Seventy-two percent (18/25) of adults on ARPQ and 61% (17/28) of adults on ASAQ provided a blood sample at day 7 after starting treatment. The median day 7 plasma concentrations of piperaquine and desethylamodiaquine were 42\u2009ng/mL  and 54\u2009ng/mL , respectively. No amodiaquine was present at day 7.P. falciparum malaria. The efficacy findings are in accord with an earlier study, at the same study site in south-central Vietnam, in which artesunate and amodiaquine were coadministered as separate tablets to patients [vs. 16,776 parasites/\u03bcL). The median parasite clearance times, however, were significantly longer in patients given ASAQ compared with patients administered separate tablets  despite the ASAQ group receiving a slightly higher daily dose of the individual drugs (~4.7\u2009mg/kg artesunate plus ~12.6\u2009mg/kg amodiaquine vs. ~4.4. mg/kg artesunate plus ~10.6\u2009mg/kg amodiaquine). Artesunate and amodiaquine given either as a fixed-dose combination or separate tablets was well tolerated in the Vietnamese patients and is in accord with the two drugs administered as different formulations (i.e. fixed vs. separate) for the treatment of P. falciparum in African children [The present study is the first to report on the efficacy and tolerability of ASAQ (Coarsucam\u2122) in Vietnam. ASAQ was found to be highly efficacious (PCR-adjusted 42-day cure rate of 98%) in the treatment of uncomplicated patients . When cochildren . The comchildren .vs. three-day regimen). In 2004, a comparative randomized efficacy study of two-day regimens of ARPQ (one dose daily per day) and dihydroartemisinin-piperaquine (Artekin\u00ae twice daily over two days) was carried out at Phuoc Chien Commune in 103 patients, with a 28-day cure rate of 100% for both forms of ACT [Unlike the more commonly used fixed-dose combinations of ACT such as artemether-lumefantrine and dihydroartemisinin-piperaquine, there is limited data on the efficacy of ARPQ. The fixed-dose combination was developed by the Chinese as an alternative to dihydroartemisinin-piperaquine (Artekin\u00ae or Duo-Cotecxin\u00ae), with the manufacturer proposing the potential benefits of ARPQ\u2019s cheaper cost of production and improve compliance  in treating Thai patients with uncomplicated P. falciparum malaria, with a follow-up period of 28\u2009days.Similar to the study carried out in 2004, the shorter two-day regimen of ARPQ was highly efficacious, with a PCR-adjusted cure rate of 98%. This compares favourably with an earlier study at the same study site using three-day regimens of dihydroartemisinin-piperaquine (Arterakine\u00ae) and artesunate plus amodiaquine given as separate tablets . As withod et al. showed tPresently, no marker exists that defines a consistent method for identifying reduced susceptibility of parasites to forms of ACT, particularly to the artemisinin component. Currently, prolongation of the parasite clearance time has been used as an important early warning sign of reduced artemisinin susceptibility. Most patients cleared their infections within 48\u2009h of commencing their treatment with either ARPQ or ASAQ. Of the patients, 16.7% (9/54) on ARPQ and 3.4% (2/58) on ASAQ took up to 60\u2009h to clear their infections. However, up to 72\u2009h was required to clear parasites in one patient in each treatment group. Both patients were children of 12\u2009years of age, with an admission parasitaemia of about 55,000 parasites/\u03bcL of blood. They were not administered a sub-optimal dose of the ACT, as they received a dose of each component above the median mg/kg body weight for the treatment group.4 and by themselves they need to be present at parasitological blood concentrations during at least three asexual blood stages to eliminate all parasites [Although a two-day regimen may improve compliance such shorter dose regimens are not recommended by WHO . The raparasites . A limitarasites , which lPlasma concentrations of piperaquine and desethylamodiaquine on day 7 have been found to correlate with drug exposure (i.e. area under the plasma drug concentration curve) and have been identified as major determinants of therapeutic response to dihydroartemisinin-piperaquine  and ASAQP. falciparum malaria in children and adults in south-central Vietnam. However, when comparing the two treatment regimens the three-day course of ASAQ resulted in a faster parasite and shorter fever clearance times than the two-day course of ARPQ. These findings warrant further investigation of the efficacy of ASAQ in other regions of Vietnam to ascertain the potential value of the ACT as an alternative option to dihydroartemisinin-piperaquine in case the first-line ACT starts to fail in Vietnam.ARPQ and ASAQ were highly and equally efficacious in the treatment of uncomplicated The authors declare that they have no conflict of interest.NXT, TNT, BD, GDS and MDE designed the study and developed the protocol. NCP and HHQ executed and coordinated the study. NCP, MC and MDE analysed and interpreted the data. MDE wrote the first draft of the paper. All authors read and approved the final manuscript."}
+{"text": "Emerging antimalarial drug resistance in mobile populations remains a significant public health concern. We compared two regimens of dihydroartemisinin-piperaquine in military and civilians on the Thai-Cambodian border to evaluate national treatment policy.Plasmodium falciparum or vivax malaria were randomized 1:1 to receive either regimen  and followed weekly for up to 6 months. The primary endpoint was malaria recurrence by day 42. Volunteers with vivax infection received primaquine at study discharge with six months follow-up.Efficacy and safety of two and three-day regimens of dihydroartemisinin-piperaquine were compared as a nested open-label evaluation within a malaria cohort study in 222 otherwise healthy volunteers . The first 80 volunteers with slide-confirmed vivax, 15 falciparum, and 5 mixed) were randomized to dihydroartemisinin-piperaquine. Intention-to-treat all-species efficacy at Day 42 was 85% for the two-day regimen (95% CI 69\u201394) and 90% for the three-day regimen (95% CI 75\u201397). PCR-adjusted falciparum efficacy was 75% in both groups with nearly half (45%) still parasitemic at Day 3. Plasma piperaquine levels were comparable to prior published reports, but on the day of recrudescence were below measurable in vitro piperaquine IC50 levels in all falciparum treatment failures.Eighty patients (60 50 levels in excess of plasma piperaquine levels seen only in treatment failures raises concern for clinically significant piperaquine resistance in Cambodia. These findings warrant improved monitoring of clinical outcomes and follow-up, given few available alternative drugs.In the brief period since introduction of dihydroartemisinin-piperaquine, there is early evidence suggesting declining efficacy relative to previous reports. Parasite ICNCT01280162ClinicalTrials.gov  Plasmodium falciparum along the western Cambodian-Thai border is now resistant to both components of multiple artemisinin combination treatments (ACTs), emphasizing the importance of ongoing monitoring of antimalarial therapeutic efficacy and resistance in this region Recent public health efforts have made considerable progress in reducing the morbidity and mortality of malaria in Southeast Asia. These advances are threatened by the emergence of documented artemisinin resistance in four countries including Cambodia, Myanmar, Thailand, and Vietnam falciparum although recent reports suggest declining efficacy in western Cambodia Dihydroartemisinin-piperaquine is a fast-acting, potent artemisinin derivative paired with a long-acting 4-aminoquinoline falciparum infection and chloroquine monotherapy for vivax infection. However, military practice guidelines employed various two and three-day regimens of dihydroartemisinin-piperaquine under the brand names Artekin, Artequick, and Duocotexin, in order to improve compliance in austere settings. In anticipation of the Cambodian national malaria control (CNM) program\u2019s plan to replace artesunate plus mefloquine with dihydroartemisinin-piperaquine for all species, we conducted a treatment study nested within a malaria cohort study in order to provide evidence-based treatment policy recommendations. The primary objective aimed to compare the efficacy of two and three-day regimens of dihydroartemisinin-piperaquine in a dynamic setting of emerging multi-drug resistance.At the time of this study, Cambodian national treatment guidelines recommended artesunate plus mefloquine combination therapy for The study protocol was approved by specific institutional review boards at participating institution including Walter Reed Army Institute of Research (IRB#00000794), National Ethics Committee for Health Research in Cambodia (IRB#00003143), and University of North Carolina (IRB#00002074). This trial was registered prior to initiation  and adhered to CONSORT guidelines. All participants provided written informed consent for participation, collection of samples, and subsequent analyses.falciparum and vivax malaria. The study was conducted in part to identify potential sites to conduct antimalarial chemoprophylaxis studies. Between September 2010 and March 2011, otherwise healthy civilian and military personnel aged 18 to 65 years were enrolled into the cohort study at two sites in Oddar Meanchay province near the northern Thai border. Exclusion criteria included allergic reaction or other contraindication to dihydroartemisinin or piperaquine, pregnancy, lactation, or abnormal EKG including a QTc interval greater than 500 milliseconds. Volunteers developing slide-confirmed asexual falciparum and/or vivax parasites were randomized to receive a directly observed two or three-day course of dihydroartemisinin-piperaquine and assessed weekly for malaria for a minimum of 42 days. Volunteers with recurrent malaria received first-line therapy per national guidelines, which at the time of the study included artesunate-mefloquine for P. falciparum and chloroquine for P. vivax. Upon cohort study discharge, volunteers who had developed vivax infection were treated with commercially-obtained primaquine phosphate. Those with normal glucose-6-phosphate dehydrogenase (G6PD) activity received 30mg of oral primaquine daily for 14 days whereas G6PD-deficient volunteers received 45mg of oral primaquine weekly for eight weeks. To evaluate primaquine for vivax radical cure and relapse prevention, volunteers had passive follow-up via monthly clinic visit or telephone interview for 6 months after primaquine administration.This was a two-arm, randomized, open-label trial nested within an active observational cohort study to compare the efficacy and safety of two versus three-day regimens of dihydroartemisinin-piperaquine in The primary efficacy endpoint was the first occurrence of any-species treatment failure by 42 days following dihydroartemisinin-piperaquine administration. Treatment failure was defined as any-species blood-stage recurrence before Day 42 to include mixed infection Blocked randomization  assigned consecutive volunteers with positive malaria smear to receive either a two-day or three-day regimen, with treatment allocation masked from study staff and volunteers in sealed envelopes. A block size of 2 was chosen to maximize the comparability of treatment groups between enrollment sites, since malaria attack rates were not known a priori, and was variability was anticipated. Duocotecxin provided by Zhejiang Holley Nanhu Pharmaceutical Co., Ltd was procured by CNM. The two-day regimen (180mg dihydroartemisinin and 1440mg piperaquine given at time of diagnosis and then at 24 hours +/\u22121 hour) and three-day regimen  both passed British Pharmacopeia 2004 content and uniformity standards conducted by AFRIMS Pharmacology Lab Plasmodium infection, and if symptom-free, monthly blood smears. Volunteers with clinical suspicion of malaria at routine follow-up, or at any time during the study, had blood smears. Two blinded microscopists examined Giemsa-stained thick and thin smears; a third blinded microscopist determined the final result for discordant readings. Parasite densities were calculated as a parasite count per 200 WBCs (thick smear) or per 5000 RBCs (thin smear). A total of 200 oil immersion fields were examined on the thick film before it was considered negative.After enrollment, cohort volunteers had a baseline history and physical examination, 12-lead electrocardiogram (ECG), and laboratory evaluation . Copy number increased relative to Day 0 in all three cases of secondary recurrence  and did not increase significantly with recurrent parasitemia  baseline hematocrit for the G6PD-deficient group was 41.2 (3.7) mg/dL , similar to the mean follow-up hematocrit within 1 month of initiating therapy [40.4 (4.9) mg/dL ]. Only two G6PD-deficient volunteers had>10% reduction from baseline hematocrit, dropping 18% and 14% below baseline at 19 and nine days, respectively, and both resolved within one month with iron and multivitamin supplementation. Six months of passive follow-up was completed in person or by phone for 96% (69/72) of volunteers, the majority of whom remained in the transmission area. Twenty-six volunteers had recurrences  with no difference between standard and weekly primaquine regimens (p\u200a=\u200a0.19)   , with a , 50\u201372) .falciparum resistance patterns. The majority of published studies demonstrated dihydroartemisinin-piperaquine treatment failure rates of 0\u20135% in both Africa and Southeast Asia falciparum efficacy seen (as low as 75% at Day 42). Prolonged parasite clearance times with 45% Day 3 positivity were observed despite relatively low initial parasitemia and DOT. Dihydroartemisinin-piperaquine was efficacious against blood-stage vivax with treatment failure as low as 3% at Day 42. A recent report indicated falciparum treatment failure rates as high as 25% in western Cambodia, worsening over three years (2008\u20132010), but 100% efficacy in Preah Vihear province falciparum malaria here, our findings suggest that the decline in efficacy of dihydroartemisinin-piperaquine may have already spread to northern Cambodia as early as 2010.In an open-label, randomized trial of two and three-day dihydroartemisinin-piperaquine regimens, we could not distinguish a difference in efficacy for treatment of all-species malaria. We did find that 42-day treatment failure rates (7.5% and 10% respectively) were higher than previously reported; however, the small numbers of treatment failures limits the ability to draw conclusions on emerging ex vivo IC50 levels in excess of plasma piperaquine levels at the time of recrudescence, providing evidence for clinical piperaquine resistance. This is concerning given dihydroartemisinin-piperaquine had been regularly used in this population for only three years prior. Possible reasons for the rapid decline in dihydroartemisinin-piperaquine efficacy include private sector availability, self-treatment, poor compliance, and the long terminal half-life of piperaquine potentially exposing surviving parasites to subtherapeutic drug levels max for the 3-day course in the fed state, and a higher Cmax as expected for the 2-day course. While calculated AUC was larger for the 2-day course, the AUC in the 3-day course was likely underestimated due to the limited time points available. Short courses of artemisinins alone as part of an ACT regimen are not intended by themselves to be curative, but to rapidly clear initial parasite burden and shorten the febrile period. Because clear pharmacokinetic-pharmacodynamic relationships have not been established for either P. falciparum or P. vivax, the relative contributions of artemisinin levels to therapeutic outcome of ACTs have yet to be established, and were not measured More importantly, treatment failures had Day 0 parasite falciparum, permits neither definitive conclusions regarding dihydroartemisinin-piperaquine efficacy nor detailed associations between molecular markers of drug resistance and treatment failures. However, the high proportion of falciparum failures, and association with drug levels below parasite piperaquine IC50s is concerning. Increased Pfmdr1 copy number was not associated with first recurrence, supporting prior correlations between in vitro piperaquine and chloroquine sensitivity and Pfmdr1 copy number amplification in Thai-Burmese falciparum isolates Pfmdr1 184F mutation, associated with mefloquine selective pressure in western Cambodia pfcrt CVIET haplotype associated with reduced susceptibility to chloroquine and piperaquine pvmdr1 amplification in vivax infections, previously associated with decreased chloroquine susceptibility, also suggests possible chloroquine resistance Limited sample size, particularly for While piperaquine was relatively well tolerated with few treatment-emergent adverse events beyond those associated with malaria itself, there was a safety signal suggested by the degree of QT interval prolongation. This signal may have been underestimated as EKGs in our study were obtained at 24-hour trough but not peak piperaquine concentrations, which typically occur at 4\u20136 hours post-dose. A food effect cannot be ruled out as a contributing factor, given that DHA-piperaquine was administered with a low-fat (17g) snack. While substantial cardiac safety signals from DHA-piperaquine were not reported in a prior regulated multi-center clinical trial Plasmodium transmission is geographically variable, military personnel may act as a reservoir for malaria transmission to the general population vivax may be impossible. A key positive finding was that primaquine administered at cohort discharge was safe and well-tolerated even in G6PD deficient individuals, despite a 39% vivax six-month recurrence rate. Despite this, the very small number of G6PD-deficient patients treated with primaquine, and diagnostics limited to qualitative fluorescent spot testing does not warrant widespread use of primaquine without testing in this population. Further investigations into primaquine safety in G6PD-deficient individuals are currently underway.This is the first study to be carried out in a predominantly military population in Cambodia since 1977 Evidence of reduced dihydroartemisinin-piperaquine efficacy shortly after introduction in Cambodia along with clinical evidence for piperaquine resistance are concerning. Frequent recurrences following effective therapy and a potentially large asymptomatic carrier pool will pose substantial challenges to the possibility of malaria elimination in Cambodia. Novel antimalarial development and improved elimination strategies are urgently needed. In the interim, improvements in case management and patient follow-up offer the best hope for containing drug-resistant malaria.Figure S1st recurrence and 2nd recurrence.Pfmdr1 copy number from baseline infection, 1 Pfmdr1 copy number for initial falciparum cases  at baseline infection (blue), 1st recurrence (red) and 2nd recurrence (green).(TIF)Click here for additional data file.Figure S2Symptoms on Day 1 or 2 not reported at baseline according to treatment arm. Symptoms reported on Day 1 or 2 in patients without symptoms on admission according to treatment arm .(TIF)Click here for additional data file.Figure S3Reported malaria episodes after administration of primaquine in G6PD-deficient versus G6PD-normal individuals. Over 6-month follow-up in person or by telephone, there was no difference in malaria recurrence between G6PD-deficient (n\u200a=\u200a6) and G6PD-normal volunteers (n\u200a=\u200a20) (p\u200a=\u200a0\u00b719). One vivax recurrence in Month 4 occurred in the same individual who relapsed in Month 1. During Months 5 and 6, there were 3 volunteers with previous vivax recurrences who also had falciparum episodes, and 1 volunteer had falciparum infection in Month 6 with no vivax recurrence after primaquine administration.(TIF)Click here for additional data file.Figure S4Ex vivo drug susceptibility at day 0 and day of recurrence for falciparum infection. Black symbols represent IC50 of parasites from ACPR patients with mean values displayed below in black, while blue symbols/text represent IC50 and mean IC50 at baseline respectively of parasites from recurrences. Mean differences in IC50 between paired samples are displayed as \u201c\u0394\u200a=\u200a\u201d.(TIF)Click here for additional data file.Table S1Pvmdr1 copy number in initial and recurrent vivax parasitemia.(DOCX)Click here for additional data file.Checklist S1CONSORT Checklist.(DOC)Click here for additional data file.Protocol S1Trial Protocol.(PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria, respectively. The baseline efficacy of AS\u2009+\u2009AQ was evaluated from February to August 2005 in patients living in Brazzaville, the capital city of the Republic of Congo.Congo-Brazzaville adopted artemisinin-based combination therapy (ACT) in 2006. Artesunate-amodiaquine (AS\u2009+\u2009AQ) and artemether-lumefantrine are the first-line and second-line anti-malarial drugs to treat uncomplicated Plasmodium falciparum recrudescent isolates from day 7 to day 28 were compared to pretreatment isolates by polymerase chain reaction (PCR) to distinguish between re-infection and recrudescence.One hundred and ninety-seven patients  were recruited in a non-randomized study, treated under supervision with AS\u2009+\u2009AQ, and were followed up for 28 days in accordance with the 2003 World Health Organization protocol. The overall efficacy of AS\u2009+\u2009AQ after PCR correction on day 28 was 94.4%. An adequate clinical and parasitological response was observed in 94.3% and 94.4% of children aged \u22645 years old and those aged >5 years old (including adults), respectively. The main reported adverse events were dizziness, vomiting, diarrhoea, pruritus, headache, anorexia, and abdominal pain.This study has shown the high efficacy of AS\u2009+\u2009AQ in Congolese patients of all ages with acute uncomplicated falciparum malaria and serves as the baseline efficacy and tolerance of this ACT in Brazzaville. In 2006, there were 247 million malaria cases in 109 endemic countries, of which 212 million cases were registered in Africa. Mortality due to malaria was estimated to be 881,000 deaths, of which 91% occurred in Africa, mostly in children aged <5 years old. The Republic of Congo reported 1,331,668 malaria cases and 4,566 deaths associated with malaria in 2006, while in 2009, 92,855 cases and 116 deaths due to malaria were reported [The end of the 20th Century was marked by a considerable increase in global malaria burden due to resistance of the parasites to anti-malarial drugs, in particular to chloroquine -6. HowevBefore implementation of ACT in the Republic of Congo, malaria was the third cause of medical consultation . In the Plasmodium falciparum isolates collected and analysed in these studies carried K76T substitution in P. falciparum chloroquine resistance transporter (pfcrt) gene, which is associated with chloroquine resistance. Sulphadoxine-pyrimethamine, the former alternative drug to treat chloroquine-resistant malaria, had also shown low efficacy, with day-28 failure rate of 30% [dhfr) and dihydropteroate synthase (dhps) genes in clinical P. falciparum isolates has demonstrated a high prevalence of \u2018quadruple\u2019 mutations consisting of dhfr triple mutation  and dhps mutation A437G [In Congo, chloroquine, the former first-line anti-malarial drug, was shown to have a very low efficacy, with a failure rate ranging from 43 to 62.5% on day 14 and 96% on day 28 ,18. All e of 30% . Analysion A437G ,21.\u00ae) efficacy in 2006 in Brazzaville are presented.In 2006, the Republic of Congo adopted artesunate\u2009+\u2009amodiaquine (AS\u2009+\u2009AQ) and artemether-lumefantrine as first-line and second-line drugs, respectively, for the treatment of acute uncomplicated malaria. Only one study had evaluated the efficacy of these combinations in a rural area in Congo . AlthougThe total population of the Republic of Congo was 3,697,490 inhabitants in 2007, and 1,373,382 of these persons (37%) reside in Brazzaville, the capital city . BrazzavP. falciparum \u22652,000 asexual parasites/\u03bcL, (ii) axillary temperature >38\u00b0C, (iii) absence of danger signs in young children  or signs of severe and complicated malaria in older children and adults, (iv) a pack cell volume (PCV) >15%, (v) absence of other febrile illnesses, (vi) written informed consent signed by the patient (for adults) or the parents or legal guardian (for minors), and, (vii) easy accessibility of their residence for home visits [All febrile patients were referred to the health centres\u2019 laboratory for malaria parasite screening. Giemsa-stained thick blood films were prepared from finger-prick capillary blood and examined under the microscope. Symptomatic patients with at least 2,000 asexual parasites/\u03bcL were examined by a physician. Patients with the following criteria were included in the study: (i) mono-infection with e visits . Before \u00ae, Sanofi Aventis, Paris, France) containing 50 mg of artesunate and 153 mg of amodiaquine base per tablet was administered to patients under supervision. Patients received the standard daily dose of 10 mg base/kg body weight of amodiaquine and 4 mg/kg body weight of artesunate for three days. Based on the calculation using the body weight of patients, quarter, half, or three-quarter tablet was administered. For young children, the tablet was crushed and suspended in sugar-containing syrup. The patient was observed for 30 min. If vomiting occurred within 30 min following the initial drug administration, the treatment was repeated. The patient was excluded from the study if he or she vomited twice during the observation period.The co-blistered pack of AS\u2009+\u2009AQ  reviewed the study protocol and consent forms in French, English and local languages.Plasmodium falciparum gametocytes were counted against 1,000 WBC. PCV was obtained by micro haematocrit centrifugation.Thick blood films were stained with 10% Giemsa for 15 min. Asexual parasites were counted against 200 white blood cells (WBCs) and expressed as the number of asexual parasites/\u03bcL of blood, assuming a WBC count of 8,000/\u03bcL of blood. In case of hyperparasitaemia, the parasite count was determined when 500 asexual parasites were counted even if 200 WBCs were not reached. Parasite density was determined by two independent technicians. Genomic DNA was extracted from blood samples collected on filter paper using QiaAmp DNA mini kit  according to the manufacturer\u2019s instruction. DNA was recovered in 100 \u03bcL of elution buffer. All extracted samples of parasite DNA were stored at \u221220\u00b0C until use.msp-1) and the central region of merozoite surface protein-2 (msp-2), were used as markers for genotyping, as described previously [msp-1 and specific primers for FC27 and 3D7 allelic families of msp-2. Allelic specific positive controls and DNA-free negative controls were included in each set of reactions. Five microlitres of PCR products were loaded on 2% agarose gel, stained with SYBR Green, separated by electrophoresis, and visualized under ultraviolet transillumination.For patients with treatment failure, the isolates collected at inclusion and recrudescent isolates were genotyped in parallel using nested PCR technique. The highly polymorphic loci, block 2 of merozoite surface protein-1  DNA ladder marker . The size polymorphism in each allelic family was analysed, assuming that one band represents one amplified PCR fragment derived from a single copy of P. falciparum isolates present before treatment (day 0) or by P. falciparum infections that occurred after treatment. First, paired pre-treatment and post-treatment samples were genotyped using msp-2 locus. If different band profile was found, it was concluded that the re-appearance of parasites on or after day 7 was due to a new infection. If similar profile of bands was observed, these samples were further analysed for a second locus, ie, msp-1. The outcome was defined as recrudescence if the paired samples (day 0 and recrudescent sample) displayed identical alleles. The outcome was defined as new infection if the recrudescent sample had only newly identified alleles. If the recrudescent sample showed new alleles and alleles identified on day 0, this sample was recorded as mixed infections or unclassified.It was assumed that after a patient was initially treated for malaria, a subsequent episode was caused by either The responses were classified before PCR correction and with PCR correction . The PCRSigns and symptoms that developed after drug intake were observed and reported during clinical examination and questioning the patients or, in case of children, their parents or guardians, during each visit. Adverse events can also be an exacerbation of disease symptoms.All patients consulting one of the two health centres and satisfying the inclusion criteria were enrolled after written informed consent, regardless of their age. Two age groups were constituted: \u22645 years old and >5 years old. Moreover, the residence of the patients was classified as urban  or suburban  area.Clinical and parasitological data were entered into pre-programmed Excel spreadsheet provided by the Department of Global Malaria Programme, WHO  with the possibility to analyse PCR-uncorrected and PCR-corrected responses. Proportions and 95% confidence interval (95% CI) were calculated using Epi-info 6.04 .From February to August 2005, 1,087 febrile patients were screened for malaria in the urban Tenrikyo health centre, where 259 (23.8%) had a positive smear. During the same period, 161 patients were screened in the suburban Madibou health centre, where 76 (47%) patients were smear-positive. Of 1,087 patients, 264 (24.3%) received an anti-malarial treatment at home or in another health service. Monotherapies with classical anti-malarial drugs represented 89.8% of drug intake, of which >50% were chloroquine. Artemisinin derivatives accounted for 10% of drug intake  were children \u22645 years old and 101 (51.8%) were aged >5 years old (p >0.5)  or protocol violation : two on day 1, three on day 2, three on day 3, three on day 7, one on day 14, and three on day 21). One 12-year-old patient with 30,600 asexual parasites/\u03bcL on day 0 presented asthenia on day 1. The attending physician considered that there was clinical aggravation and referred the patient for hospitalization. The outcome of this patient was considered as ETF. On day 1, eight of 197 (4%) patients were still febrile (axillary temperature \u226537.5\u00b0C). On day 2 and day 3, only two of 189 (1%) patients and one of 187 (0.5%) patients were still febrile, respectively. Parasite density decreased from the geometric mean (95% CI) 30,800 asexual parasites/\u03bcL  on day 0 to 235 asexual parasites/\u03bcL  on day 2. On day 3, only one patient had a positive smear at a low parasite density .On day 0, six of 197 (3%) patients were gametocyte carriers. On day 2, day 3, day 7, and day 14, there were 16 of 190 (8.4%), 12 of 189 (6.3%), 11 of 187 (6%), and six of 185 (3%) gametocyte carriers, respectively. The highest mean gametocyte density, observed on day 2, day 3, and day 7, varied from 10 to 11 gametocytes/\u03bcL.On day 28, the PCR-uncorrected cure rate  was 83% 151 of 182). In children \u22645 years old and children >5 years old and adults, ACPR were observed in 76.7% (66 of 86 patients) and 88.5% (85 of 96 patients), respectively (p\u2009=\u20090.07) . In children \u22645 years old and those aged >5 years, the proportions of ACPR were 94.3% (66 of 70 patients) and 94.4% (85 of 90 patients), respectively (p\u2009=\u20090.5) , vomiting (14.6%), diarrhoea (9%), pruritus (9%), headache (7%), anorexia (1.5%), and abdominal pain (1.5%). Most of these signs and symptoms disappeared on day 3. In a few cases however, headache and pruritus persisted until day 7. On day 1, two patients were excluded due to repeated vomiting.The present study was conducted to provide supporting evidence for the clinical efficacy of AS\u2009+\u2009AQ, which was adopted in the new anti-malarial drug policy in central Africa, including Congo, to replace ineffective, classical anti-malarial treatment based on chloroquine and sulphadoxine-pyrimethamine . Despitevs >5 years old, including adults).This is the first clinical study on ACT conducted in Brazzaville, where 37% of Congolese population resides. A single earlier clinical study on AS\u2009+\u2009AQ and artemether-lumefantrine was conducted in a rural area . HoweverThe results of the present study show that AS\u2009+\u2009AQ reduces fever very rapidly. More than 95% of the patients were afebrile on day 1, ie, after the first dose of treatment. The geometric mean initial parasitaemia decreased rapidly after only two doses, and 99% of the patients had negative smears on day 2. There were only a few gametocyte carriers during the follow-up period. Other clinical studies in Africa have already highlighted these properties of AS\u2009+\u2009AQ on fever, parasitaemia and gametocytes -31.The overall efficacy of AS\u2009+\u2009AQ after PCR correction, represented by the ACPR rate, was 94.4%. The results of the present study are in agreement with those of the earlier study in a rural zone in Congo, in which a cure rate of 98.5% was reported (n\u2009=\u200966) . If the \u00ae). The reported efficacy was excellent in one randomized study conducted in Angola with 100% of ACPR [Studies conducted in Central Africa before or during the same period as the present study used the non-fixed formulation of AS\u2009+\u2009AQ , which is available today, reduces the total number of tablets for the three-day treatment regimen and improves patient compliance. Since 2008, all these dosages have been available free of charge in the Congolese public health sector as first-line anti-malarial.The present study demonstrated AS\u2009+\u2009AQ efficacy in Brazzaville using non-coformulated ACT. The present results serve as a database for further clinical evaluation to determine therapeutic efficacy and tolerance of fixed AS-AQ formulation in the capital city of Congo.95% CI: 95% confidence intervals; ACPR: Adequate clinical and parasitological response; ACT: Artemisinin-based combination therapy; ETF: Early treatment failure; LCF: Late clinical failure; LTF: Late treatment failure; PCR: Polymerase chain reaction; SCRIHS: WHO Secretariat Committee on Research Involving Human Subjects; WHO: World Health Organization.The authors declare that they have no competing interests.MN, PIM, PNC, and DL performed the clinical study. MN, LKB, FN and PB performed and validated the data analysis and wrote the manuscript. All authors read and approved the final manuscript."}
+{"text": "P. falciparum malaria across diverse transmission settings in Africa. Longitudinal follow-up of patients in these trials supported previous findings that DP had a longer post-treatment prophylactic effect than AL, reducing the risk of reinfection and conferring additional health benefits to patients, particularly in areas with moderate to high malaria transmission.Recent multi-centre trials showed that dihydroartemisinin-piperaquine (DP) was as efficacious and safe as artemether-lumefantrine (AL) for treatment of young children with uncomplicated We developed a Markov model to assess the cost-effectiveness of DP versus AL for first-line treatment of uncomplicated malaria in young children from the provider perspective, taking into consideration the post-treatment prophylactic effects of the drugs as reported by a recent multi-centre trial in Africa and using the maximum manufacturer drug prices for artemisinin-based combination therapies set by the Global Fund in 2013. We estimated the price per course of treatment threshold above which DP would cease to be a cost-saving alternative to AL as a first-line antimalarial drug.First-line treatment with DP compared to AL averted 0.03 DALYs (95% CI: 0.006\u20130.07) and 0.001 deaths (95% CI: 0.00\u20130.002) and saved $0.96 (95% CI: 0.33\u20132.46) per child over one year. The results of the threshold analysis showed that DP remained cost-saving over AL for any DP cost below $1.23 per course of treatment.P. falciparum malaria in young children. A paediatric dispersible formulation of DP is under development and should facilitate a targeted deployment of this antimalarial drug. The use of DP as first-line antimalarial drug in paediatric malaria patients in moderate to high transmission areas of Africa merits serious consideration by health policymakers.DP is superior to AL from both the clinical and economic perspectives for treatment of uncomplicated  Plasmodium falciparum malaria is critical in preventing the progression of acute infections to severe disease and reducing the risk of further morbidity, disability and premature mortality from the disease P. falciparum malaria worldwide Despite a rapid scaling up of malaria control efforts and recent reports of decreasing transmission intensities in African countries, malaria remains an important cause of morbidity and mortality, particularly in young children Recent multi-centre trials showed that dihydroartemisinin-piperaquine (DP), a newer fixed-dose co-formulated ACT, was as efficacious and safe as AL for treatment of young children with uncomplicated malaria across diverse transmission settings in Africa Significant financial resources have been put into scaling up the access to ACTs through global subsidies on manufacturing prices of drugs Cost-effectiveness analysis was performed using a Markov model  that defined a set of mutually exclusive health states, simulating the progression of malarial disease and the risk of recurrent malaria in children younger than five years of age, receiving DP or AL for first-line treatment of uncomplicated malaria . The MarThe Markov model had 11 health states: State 1: Healthy; State 2: Uncomplicated malaria; State 3: Severe malaria; States 4\u201310: Post-treatment from week one to week seven; and State 11: Dead . The posPublished estimates were used for the probability of developing severe malaria following oral antimalarial treatment Children entered the Markov model in the \u201chealthy\u201d state. They were subjected to a hazard rate for unscomplicated malarial disease estimated from the trial data and flowed through the subsequent health states in weekly cycles. We assumed that all children with uncomplicated malaria would be treated promptly with either DP or AL, and all children with severe malaria would receive inpatient care. Children, who were treated orally with antimalarial drugs, would either recover and enter the \u201cpost-treatment\u201d states, or develop severe malaria. Depending on the antimalarial treatment choice, children in the post-treatment states were subjected to a weekly hazard rate for recurrent malaria during the first seven weeks following treatment . ChildreHealth outcomes were measured in terms of disability adjusted life years (DALYs) averted. DALYs combine years of life lost because of premature mortality with years of life lived with disability. We used an average life expectancy of 57.25 years for children aged 1\u20134 years on the basis of the life tables for African men and women for the WHO sub-region with high child and high adult mortality We considered the costs of oral antimalarial drugs for treating uncomplicated cases and the costs of inpatient care for severe cases over one year , but excWe used published estimates of inpatient care costs at primary level hospitals from a recent Kenyan costing study To assess the uncertainty in key model parameters and the robustness of the results to model assumptions, we performed a probabilistic sensitivity analysis using a Monte Carlo simulation technique with 10,000 iterations. For each iteration, a value for each input variable was selected randomly from its distribution given in First-line treatment with DP was the economically dominant treatment strategy over AL with a 90% probability , resultiP. falciparum malaria that considered the post-treatment prophylactic effects of antimalarial drugs \u2013 an important secondary benefit of antimalarial treatment, particularly in areas with moderate to high malaria transmission in Africa. DP was both clinically superior and less costly compared to AL for first-line treatment of uncomplicated malaria in young children. It proved to be the economically dominant treatment strategy in such transmission settings. The threshold analysis showed that first-line treatment with DP would remain cost saving over AL for any DP cost below $1.23 per course of treatment.This is the first economic analysis of first-line treatment options for uncomplicated P. falciparum malaria per child when treated with DP and AL, respectively. A recent study predicted an annual incidence rate of 1.7 (95% CI: 1.4\u20133.9) cases of uncomplicated malaria per child for this age group living in such transmission settings in Africa Our analysis is based on the primary data from a cohort of children who participated in a multi-centre trial on the efficacy of ACTs conducted in areas with moderate to high malaria transmission in Africa We assumed full access to treatment with AL and DP and same level of patient compliance to both therapies to evaluate the costs and benefits conferred by the post-treatment prophylactic effects of the drugs. While both drugs are administered over three days, DP has a simple, once daily dosing regimen whereas AL should be given twice daily and ideally with a fatty meal. Therefore, DP has the potential to improve patient compliance and treatment effectiveness and hence overall management of pediatric patients. However, such an advantage of treatment with DP could not be evaluated in a clinical trial with an observed drug intake A limitation of our analysis is the lack of data on the long-term post-treatment prophylactic effect of DP compared to AL. Our model incorporated data from a drug efficacy trial with limited (63 days) longitudinal follow-up of patients Maximum manufacturer prices are negotiated by the Global Fund for several ACTs, including AL and DP. However, these prices do not necessarily reflect the first-line buyer prices; co-payment amounts go up to 98% of manufacturer sales prices for some African countries P. falciparum endemicity will affect the efficacy and the post-treatment prophylactic effect of these drugs. Decision makers should contextualise costs and assess patient compliance, pattern of clinical malaria, and other key parameters in their own settings to arrive at more locally representative results.It should be mentioned that a change in national malaria treatment policies may require substantial resources P. falciparum (and P.vivax) malaria P. falciparum malaria in children compared to the conventional treatment. From a patient's perspective, the differentiation between recrudescence and re-infection is insignificant. PCR-correction discounts the post-treatment prophylactic effect of a drug by excluding new infections that occur during the follow-up period. Large-scale clinical trials in African settings have consistently shown that DP was more efficacious than AL using PCR-uncorrected cure rates P. falciparum malaria in children in areas with moderate to high transmission.There is limited evidence on the cost-effectiveness of ACTs for treatment of uncomplicated malaria in children. A recent study from Papua New Guinea compared the cost-effectiveness of three different ACTs against the conventional treatment with chloroquine plus SP for treatment of A main concern with DP and other long-acting antimalarial drugs is that residual drug levels play a critical role in the emergence and spread of drug resistance P. falciparum malaria in young children. A paediatric dispersible formulation of DP suitable for use by children between the ages of six months and five years is under development There is an urgent need to improve the management of malaria in African children. Decision makers who aim to provide optimal treatment strategies to the populations at risk need to consider various aspects of antimalarial drugs \u2013 including safety, tolerability, dosing schedule, level of drug resistance, post-treatment prophylactic effect, and cost \u2013 and malaria endemicity. Our study demonstrates the superiority of DP over AL from the clinical and economic perspectives for treatment of uncomplicated"}
+{"text": "Plasmodium falciparum malaria.Children are most vulnerable to malaria. A pyronaridine-artesunate pediatric granule formulation is being developed for the treatment of uncomplicated P. falciparum malaria. Patients were randomized (2:1) to pyronaridine-artesunate granules (60/20 mg) once daily or artemether-lumefantrine crushed tablets (20/120 mg) twice daily, both dosed by bodyweight, orally (liquid suspension) for three days.This phase III, multi-center, comparative, open-label, parallel-group, controlled clinical trial included patients aged \u226412 years, bodyweight \u22655 to <25 kg, with a reported history of fever at inclusion or in the previous 24 h and microscopically-confirmed uncomplicated P\u2009<\u2009.0001). Pyronaridine-artesunate was non-inferior to artemether-lumefantrine: treatment difference -1.8% (95% CI -4.3 to 1.6). The incidence of drug-related adverse events was 37.2% (132/355) with pyronaridine-artesunate, 44.4% (80/180) with artemether-lumefantrine. Clinical biochemistry results showed similar mean changes versus baseline in the two treatment groups. From day 3 until study completion, one patient in each treatment group had peak alanine aminotransferase (ALT) >3 times the upper limit of normal (ULN) and peak total bilirubin >2xULN (i.e. within the Hy\u2019s law definition).Of 535 patients randomized, 355 received pyronaridine-artesunate and 180 received artemether-lumefantrine. Day-28 adequate clinical and parasitological response (ACPR), corrected for re-infection using polymerase chain reaction (PCR) genotyping (per-protocol population) was 97.1%  for pyronaridine-artesunate; 98.8%  for artemether-lumefantrine. The primary endpoint was achieved: pyronaridine-artesunate PCR-corrected day-28 ACPR was statistically significantly >90% (The pyronaridine-artesunate pediatric granule formulation was efficacious and was non-inferior to artemether-lumefantrine. The adverse event profile was similar for the two comparators. Pyronaridine-artesunate should be considered for inclusion in paediatric malaria treatment programmes.ClinicalTrials.gov: identifier NCT00541385 Plasmodium falciparum malaria kills approximately 850,000 children annually, most are under five years old [P. falciparum malaria [ears old ,2. The W malaria . However malaria . ClearlyP. falciparum and Plasmodium vivax malaria [P. falciparum malaria, day-28 adequate clinical and parasitological response (ACPR) rates in the per-protocol population were 99.5% (780/784) and 99.2% (743/749) when corrected for re-infection with polymerase chain reaction genotyping (PCR-corrected) [Pyronaridine-artesunate (3:1 ratio) is an ACT being developed for the treatment of uncomplicated  malaria -7. In twrrected) . Pyronarrrected) . The advrrected) .P. falciparum malaria. The primary efficacy outcome of this trial was to demonstrate >90% efficacy of pyronaridine-artesunate granules in children with P. falciparum mono-infection evaluated using PCR-corrected day-28 ACPR in the per-protocol population [Children are most vulnerable to malaria ,2,8-10 apulation . A seconThe protocol was approved by each study center\u2019s independent ethics committee and the study was conducted in accordance with the Declaration of Helsinki (Tokyo 2004), Good Clinical Practice, and applicable regulations. Informed written or witnessed oral consent was obtained from all patients\u2019 parents/guardians; assent was required from children able to understand the study.This multi-center, comparative, randomized, open-label, parallel-group clinical trial followed WHO guidelines . Study dP. falciparum mono-infection . If applicable, a negative pregnancy test was required. Subjects were excluded if they had: signs and symptoms of severe/complicated malaria [Plasmodium infection; severe vomiting (>3 times in the previous 24 h); severe diarrhoea (\u22653 watery stools per day); other clinically significant disorders, including hepatitis or HIV infection; other febrile conditions; hepatic/renal impairment; electrolyte imbalance; anemia (hemoglobin <8 g/dL); hypersensitivity/allergy to study drugs; anti-malarial therapy in the previous two weeks, an investigational drug within four weeks, or any drug metabolized by cytochrome enzyme CYP2D6; participated previously in pyronaridine-artesunate clinical studies.Eligible subjects were of either sex, \u226412 years old, with a bodyweight \u22655 kg and <25 kg , a reported history of fever at inclusion or within the previous 24 h, and microscopically-confirmed uncomplicated  malaria ; mixed PThe sponsor provided a computer-generated randomization schedule. Patients were randomized 2:1 to pyronaridine-artesunate or artemether-lumefantrine. Randomization numbers were assigned in ascending order. Individually numbered treatment packs of similar appearance were masked on allocation. Clinical assessments and drug administration were performed by different clinical personnel. Drugs were given open-label. The study sponsor remained blinded to treatment allocation.Study drugs were given orally for 3 days , dosed according to bodyweight. All doses were directly observed. Pyronaridine-artesunate was given once daily: \u22655 to <9 kg, one sachet; 9 to <17 kg, two sachets; 17 to <25 kg, three sachets (dose range 6.7/2.2 to 13.3/4.4 mg/kg/dose). Artemether-lumefantrine was given twice daily: \u22655 to <15 kg, one tablet; 15 to <25 kg, two tablets (dose range 1.3/8.0 to 4.0/24.0 mg/kg/dose); the second day-0 dose was 8 h after the first dose, the first day-1 dose was 24 h after the first day-0 dose, with all subsequent doses 12 h apart.Oral suspensions were prepared immediately before dosing. Pyronaridine-artesunate granules were stirred into 50 mL of water, milk, or soup. Artemether-lumefantrine tablets were crushed to coarse particles, added to 50 mL of water and shaken to a uniform suspension. Residual drug was given by adding 100 mL of water to the dosing cup. Artemether-lumefantrine was given with food or milk as per local guidelines. Vomiting within 30 minutes following the first drug dose resulted in re-dosing. Vomiting after repeat dosing or any subsequent dose resulted in study withdrawal and treatment with rescue medication .At screening, a medical history was taken and a physical examination performed. Eligible patients were hospitalized from day 0 to day 3, with follow-up at days 7, 14, 21, 28, 35, and 42. Temperature was taken at screening, every 8 h over \u226572 h following the first dose or until two normal readings between 7 and 25 h apart, then at each visit or as clinically indicated. All study sites were provided with equipment which was used solely for the rapid analysis of biochemical and hematological samples obtained from the subjects evaluated for inclusion in the trial or samples obtained during the trial. Venous blood samples were taken for clinical biochemistry and hematology at screening, days 3, 7, 28, and 42; urinalysis was performed at screening. Electrocardiographs (ECG) were done at screening and day 2, and if indicated at days 7, 14, and 28.Parasitological assessments were conducted according to WHO guidelines . Venous P. falciparum genes msp1, msp2, and glurp, recrudescence was defined as at least one matching allelic band in all markers between baseline and post-day-7 samples [Following parasite reappearance, recrudescence and re-infection were distinguished by PCR genotyping performed at a central laboratory . Using  samples ,15. TherThe primary efficacy endpoint was pyronaridine-artesunate day-28 PCR-corrected ACPR >90%. The main secondary efficacy endpoint was non-inferiority of pyronaridine-artesunate to artemether-lumefantrine for day-28 PCR-corrected ACPR . TreatmeOther secondary efficacy outcomes were: day-28 crude (non-PCR corrected) ACPR; day-42 PCR-corrected and crude ACPR; parasite clearance time ; fever clearance time ; and the proportion of patients with parasite clearance or fever clearance on days 1, 2, and 3. Exploratory efficacy outcomes were: gametocyte density and proportion of patients with gametocytes; and gametocyte clearance time (defined as for parasite clearance time).Safety outcomes were: adverse events, categorized using MedDRA ; laboratory abnormalities graded using the Division of Microbiology and Infectious Diseases Toxicity Scale ; and ECG abnormalities.A 95% cure rate was assumed for both treatments. For the primary efficacy endpoint, 320 evaluable patients in the pyronaridine-artesunate group provided 91% power to reject the null hypothesis (i.e. day-28 cure rate \u226490%) using a 1-sided exact binomial test with a nominal significance level of 2.5%. For the main secondary efficacy endpoint, 480 evaluable patients randomized 2:1 provided >99% power to demonstrate non-inferiority of pyronaridine-artesunate versus artemether-lumefantrine with a non-inferiority limit of 10%. Allowing for a 10% drop out rate, target recruitment was 534 subjects .The intent-to-treat population included all randomized subjects who received any study medication and was the same as the safety population. The per-protocol population included patients that received a full course of study medication, had a known day-28 primary endpoint, and did not violate the protocol so as to impair evaluation of the primary endpoint.The primary efficacy endpoint was evaluated in the per-protocol population using the exact binomial test (significance limit\u2009\u2264\u2009.025). The associated exact (Pearson\u2013Clopper) 2-sided 95% confidence interval (CI) was presented. The analysis was repeated for the intent-to-treat population. The primary efficacy endpoint was summarized by center, age category , gender, previous episode of malaria (yes/no), and actual pyronaridine-artesunate dose .The main secondary efficacy endpoint was evaluated in the per-protocol population. Pyronaridine-artesunate was non-inferior to artemether-lumefantrine if the lower limit of the 2-sided 95% CI (Newcombe\u2013Wilson score method without continuity correction) for the difference between treatments was not lower than \u221210%. If pyronaridine-artesunate was non-inferior, superiority was tested using 2-sided Chi-square test (significance limit\u2009<\u2009.05). No multiplicity testing adjustment was required. The analysis was repeated for the intent-to-treat population, day-28 crude ACPR and day-42 PCR-corrected and crude ACPR.All other analyses were presented for the intent-to-treat population. A post-hoc Kaplan\u2013Meier analysis of recrudescence rate and re-infection rate was conducted and treatments compared using the log-rank test. Patients who did not have the event (recrudescence or re-infection) reported were censored at the last available parasite assessment date. In addition, patients with major protocol deviations were censored at the time of the deviations if they did not have the event before that time.Parasite, fever and gametocyte clearance times were summarized using Kaplan\u2013Meier estimates and treatment groups compared using the log-rank test. Patients without parasite or fever clearance within 72 h after the first drug dose were censored at that time point. The proportion of subjects with parasite, fever, and gametocyte clearance on days 1, 2 and 3 was calculated using Kaplan\u2013Meier estimates. Statistical analysis was performed using SAS Version 9.1.3.Between November 2007 and September 2008, 355 patients were randomized to pyronaridine-artesunate, 180 to artemether-lumefantrine ; statistically significantly >90% . The efficacy of pyronaridine-artesunate for children aged <1 year was 81.8% , for children >1\u2013\u2009<\u20095 years was 95.7%  and for children 5\u201312 years was 98.9%   showed no difference between the two treatment groups for recrudescence rate . Complete gametocyte clearance was achieved between day 41 and day 44 in the pyronaridine-artesunate group and between days 31 and 40 in the artemether-lumefantrine group. In the pyronaridine-artesunate group a total 26.8% (95/354) patients had post-baseline gametocytes versus 24.7% (44/178) with artemether-lumefantrine. New occurrences of gametocytes in patients that had none at baseline occurred in 15.0% (53/354) of patients in the pyronaridine-artesunate group and 11.2% (20/178) in the artemether-lumefantrine group.Additional file P. ovale (three in the pyronaridine-artesunate group); all occurred on or after day 28. There were two cases of P. malariae, occurring at day 35 and day 42 (both in the artemether-lumefantrine group). There were no cases of P. vivax infection. All cases of non-falciparum malaria were treated as per local guidelines.During follow-up, there were five cases of Mean total drug exposure was 27.7/9.2 mg/kg (average daily dose 9.2/3.1 mg/kg) for pyronaridine-artesunate and 11.5/68.9 mg/kg (average daily dose 3.8/23.0 mg/kg) for artemether-lumefantrine.Adverse events of any cause were experienced by 285/355 (80.3%) patients in the pyronaridine-artesunate, and 143/180 (79.4%) patients in the artemether-lumefantrine group  occurred in a 2-year-old male, resulting in study withdrawal on day 0 (11 h 20 mins after inclusion) after receiving one dose of pyronaridine-artesunate. He was treated with intramuscular artemether (day 1 and days 6\u201310) and intravenous quinine (days 1\u20135) and recovered well. The investigator noted this adverse event as unrelated to study treatment. In both treatment groups, 1.7% of patients had adverse events leading to drug discontinuation and study withdrawal: six patients receiving pyronaridine-artesunate  and three receiving artemether-lumefantrine (three with vomiting).Hematology results showed similar mean changes from baseline in the two treatment groups consistent with effective anti-malarial therapy. Mean hemoglobin concentrations decreased by \u20130.60 to \u20130.68 g/dL on day 3 and recovered by Day 28  in the per-protocol population. This is consistent with the high efficacy rates reported with pyronaridine-artesunate tablets in Phase III trials conducted in children and adults with P. falciparum malaria [In children with uncomplicated  malaria . StatistPyronaridine-artesunate was highly efficacious across all seven study centers and was equally effective across all age groups  [P. falciparum may have contributed to a more sustained prophylactic effect.Pyronaridine-artesunate efficacy in children was non-inferior to that of artemether-lumefantrine; consistent with previous findings in children and adults . There w\u2009=\u2009.007) . This miP\u2009=\u2009.02). This was seen previously [As expected, both treatments reduced parasitemia rapidly. Parasite clearance time was shorter with pyronaridine-artesunate versus artemether-lumefantrine  patients [P. vivax[The incidence of peak ALT >3xULN plus peak total bilirubin >2xULN was 0.3% 1/355) in the pyronaridine-artesunate group and 0.6% (1/180) in the artemether-lumefantrine group. ALT elevations with increased bilirubin have been observed with pyronaridine-artesunate tablets at a similar incidence in two Phase III studies in 5 in the [P. vivax.Meta-analysis has suggested that pediatric ACT formulations have lower rates of drug-related gastrointestinal adverse events versus tablets . Drug-reP. falciparum malaria. Considering these data and those of the other Phase III trials [P. falciparum malaria.Pyronaridine-artesunate pediatric granules were efficacious and well tolerated in this study of children under 12 years of age with uncomplicated I trials ,6, pyronIB-F and SD are employees of the Medicines for Malaria Venture, C-SS is an employee of Shin Poong Pharmaceutical Co. Ltd. There was no conflict of interest for KK, OKD, LKP, ATO, KMB, JK, AKT, JKHT, MR, PMdeS, ABT, AO, MDGB, FQ, or LF.IB-F, SD, C-SS and LF made substantial contributions to the concept and design of the study. KK, OKD, LKP, ATO, KMB, JK, AKT, JKHT, MR, PMdeS, ABT, AO, MDGB, FQ and LF were involved in the acquisition of data. All authors contributed to the analysis and interpretation of data. All authors critically reviewed the paper and read and approved the final manuscript.In addition to the named authors, the following co-investigators contributed to this study: Bakary Sidib\u00e9, Abdoulaye Djimd\u00e9 ; Berenger A. A. Ako, Aristide M'Lanhoro Coulibaly (Ivory Coast); Moses Omwoyo, Jacqueleen Wanjiru (Kenya); Nsengi Ntamabyaliro, Raoul Mpoyi Ngambua (Democratic Republic of Congo); Sabine B\u00e9lard, Florian Kurth (Gabon); D\u00e9sir\u00e9 Kargougou, David T. Kangoye (Burkina Faso); Jennifer Rabang (Philippines).Presentation: This study was presented in part at the 5th MIM Pan-African Malaria Conference, Nairobi, Kenya 2\u20136 November, 2009. Kassoum Kayentao et al. Phase III pivotal trial of pyronaridine artesunate versus artemether lumefantrine in paediatric patients with acute uncomplicated Plasmodium falciparum malaria. Abstract MIM16689330.PCR-corrected day-28 adequate clinical and parasitological response rates in the per-protocol population by country and patient age.Click here for fileNumber of patients in the intent-to-treat population with parasite, fever and gametocyte clearance, median clearance times and the proportion of patients with clearance at days 1, 2, and 3.Click here for fileKey laboratory variables: baseline values, changes from baseline at days 3 and 7 and 28, and incidence of post-baseline grade 3 or 4 toxicity values for hepatic enzymes and total bilirubin.Click here for file"}
+{"text": "The choice of appropriate artemisinin-based combination therapy depends on several factors . To assess whether the combination dihydroartemisinin-piperaquine (DP) could be an alternative to artemether-lumefantrine (AL), the efficacy and the tolerability of the two products for the treatment of uncomplicated falciparum malaria in sub-Saharan Africa have been compared.A multicentric open randomized controlled clinical trial of three-day treatment of DP against AL for the treatment of two parallel groups of patients aged two years and above and suffering from uncomplicated falciparum malaria was carried out in Cameroon, C\u00f4te d'Ivoire and Senegal. Within each group, patients were randomly assigned supervised treatment. DP was given once a day for three days and AL twice a day for three days. Follow-up visits were performed on day 1 to 4 and on day 7, 14, 21, 28 to evaluate clinical and parasitological results. The primary endpoint was the recovery rate by day 28.Plasmodium falciparum. In the DP group, after PCR genotyping, one of the two recurrences was classified as a new infection and the other as recrudescence. In AL group, two recurrences were classified after correction by PCR as recrudescence. All cases of recrudescence were classified as Late Parasitological Failure (LPF). In each group, a rapid recovery from fever and parasitaemia was noticed. More than 90% of patients did no longer present fever or parasitaemia 48 hours after treatment. Both drugs were well tolerated. Indeed, no serious adverse events were reported during the follow-up period. Most of the adverse events which developed were moderate and did not result in the treatment being stopped in either treatment group.Of 384 patients enrolled, 197 were assigned DP and 187 AL. The recovery rates adjusted by genotyping, 99.5% in the DP group and 98.9% in the AL group, were not statistically different (p = 0.538). No Early Therapeutic Failure (ETF) was observed. At day 28, two patients in the DP group and five in AL group had recurrent parasitaemia with Dihydroartemisinin-piperaquine was as effective and well-tolerated as artemether-lumefantrine in the treatment of uncomplicated falciparum malaria. In addition, dihydroartemisinin-piperaquine, a single daily dose, could be an advantage over artemether-lumefantrine in Africa because of better treatment observance. Plasmodium falciparum is a serious concern for public health and development in Africa. The most part of the continent is facing increasing resistance of this parasite to chloroquine and sulphadoxine-pyrimethamine, the widely available and cheap anti-malarial drugs. In order to overcome this resistance problem, several African countries have recently adopted artemisinin-based combination therapy (ACT) as first-line treatment for uncomplicated malaria [Malaria caused by  malaria ,2. The a malaria -5. Howev malaria  are a diDihydroartemisinin-piperaquine (DP) is a new ACT administered as single daily dose that has proved to be well tolerated and highly effective against uncomplicated falciparum malaria in southeast Asia -12 and iThis study was carried out from November 2006 to May 2008 in three sub-Saharan countries: Senegal and C\u00f4te d'Ivoire are in western Africa, Cameroon is in central Africa. In Cameroon, the study took place at the University Hospital Centre. In C\u00f4te d'Ivoire, it took place at the health care centre of Bocabo, which is located in Abobo in the north of Abidjan . In Senegal, the assessment took place at the health care centre of Darou Marnane in the sanitary district of Touba, which is located in the centre of the country at 200 km of Dakar.P. falciparum mono-infection with a parasitaemia from 2,000 to 200,000/\u03bcl of blood, in Cameroon and C\u00f4te d'Ivoire, and from 1,000 to 100,000/\u03bcl of blood in S\u00e9n\u00e9gal; 4) provision of the written and informed consent by the patient or his legal guardian for children; 5) no history of previous serious side-effects to the drugs used in the trial; 6) no evidence of a concomitant febrile illness; 7) no danger signs or evidence of severe malaria; 8) no treatment with 4-amino quinoleines, sulphadoxine-pyrimethamine, mefloquine or halofantrine in the previous seven days, or with quinine, artemisinin or cyclins in the previous three days, 9) no pregnancy or nursing; 10) no ongoing anti-malarial treatment.The study population consisted of outpatients visiting health care centres for uncomplicated falciparum malaria-like symptoms. Patients were enrolled if they meet the following selection criteria: 1) at least two years old; 2) fever with axillary temperature > or = 37.5\u00b0C; 3) The number of patients to be enrolled was determined by Epi Info 2000 software. The estimated expected recovery rate with AL was 98%, with a maximum acceptable difference of 5% to conclude that DP was non-inferior and a power of 85%. The minimum number of people to be included in each arm was calculated from these assumptions to be 180 patients. Assuming a 5% loss to follow up the overall final target sample size of 380 participants was estimated.This study was a randomized, controlled and open therapeutical trial of DP against AL as the reference treatment. The three criteria of judgment were clinical efficacy, biological efficacy and tolerance. The study protocol was first approved in each country by the National Ethical Committee according to protocols and standards operating procedures of Good Clinical Practices of the ICH harmonized Triplicate Guide Lines for Good Clinical Practice made in 1996 and the Helsinki Declaration on human being research. This approval was critical for the study start.For each patient who met inclusion criteria, the protocol was read and explained to him/her or the legal guardian (for children) who in case of acceptance had to sign the written informed sheet afterward to authorize recruitment of the child in the study. The patient or guardian was given a copy of the informed consent and patient information sheet. For patients who gave their consent, baseline examinations and laboratory investigations were carried immediately free of charge. Those of patients who met inclusion criteria at baseline (day 1) were randomly assigned to one of the two treatment groups following a randomization list. In each study site computer generated randomization codes were prepared by an independent individual. These codes were enclosed in sequentially numbered opaque sealed envelopes, each of which contained the treatment allocation. The envelopes were assigned in sequential order to participants after inclusion.\u00ae )  once daily for three consecutive days. Each tablet of DP contains 40 mg of dihydroartemisinin and 320 mg of piperaquine. DP treatments varied as follows: patients between 5-9 kg received half a tablet per dose, those between 10-14 kg 3/4 tablet, 15-19 kg 1 tablet per dose, 20-24 kg 1 tablet + 1/4 per dose, 25-29 kg 1 tablet + 1/2 per dose, 30-34 kg 1 tablet + 3/4 per dose, 35-39 kg 2 tablets per dose, 40-44 kg 2 tablets + 1/4 per dose, 45-49 kg 2 tablets + 1/2 per dose and patients \u2265 50 kg 3 tablets per dose. The second patient group was allocated artemether-lumefantrine (AL) (Coartem\u00ae ) tablets , each tablet containing 20 mg artemether and 120 mg lumefantrine. Patients received treatment dose according to the following scheme: i.e. patients between 5-14 kg received one tablet per dose, those between 15-24 kg two tablets, those between 25-34 kg received three tablets per dose and those with body weight \u2265 to 35 kg received four tablets per dose. Doses were given at T0h, T8h, T24h, T36h, T48h and T60h. All treatments were given under direct supervision of the study co-investigators. If the patient vomited within thirty minutes after drug administration, the whole dose was re-administered. If the vomiting persisted, the patient was excluded from the study and referred to the health centre doctor for management according to the current national policy. The dose could not be administered again if vomiting occurred more than 60 minutes after administration.Treatments in the two groups were allocated according to body weight. First patient group was allocated dihydroartemisinin-piperaquine (DP)  genotyping was performed as described by Faye [In order to distinguish re-infection (RI) from recrudescence (RE), merozoite surface protein 1 and 2  after follow-up for 28 days. Efficacy was evaluated using a modified intention to treat analysis, which included the 379 patients, randomized and not lost to follow-up. The secondary end points were the incidence of early clinical failure (ECF), late parasitological failure (LPF), late clinical failure (LCF), change in gametocyte carrier status, fever and parasites clearance and adverse clinical and laboratory events.All data were recorded and checked using Epi data version 3.1 and analysed with SPSS for windows (version 12.0). Patient's characteristics of the two groups at inclusion were compared using Pearson's Chi-2 test, independent samples t test or Mann-Whitney test. The modified intention to treat analysis was performed using Kaplan-Meier survival analysis with log rank testing the treatment failures distribution function. The cases of protocol violation and withdrawn consent were censored at the time they left the study. The distributions of fever and parasite clearance were compared using Pearson's Chi-2 test. Differences of haemoglobin and biochemical parameters values within individuals between D1 and D4 were computed. Changes in haemoglobin concentrations and in biochemical parameters were compared using the paired t test. The level of significance for statistical tests was set at 0.05.A total of 384 patients were included in the study. 197 patients were randomized to DP and 187 to AL. In the DP group, six patients were excluded after enrollment. Of these, four patients were lost during follow-up and one case of protocol violation  and one case of withdrawal of consent were noticed. In the AL group, four patients were excluded during the follow-up: one patient was lost sight. There were one case of protocol violation and two cases of withdrawal of the consent.Finally, 191 patients and 183 patients were successfully followed up, respectively in DP and AL groups . These recovery rates adjusted by genotyping, 99.5% in the DP group and 98.9% in the AL group, were not statistically different (p = 0.538). There was no significant study site effect on these estimates.P falciparum. In the DP group, after PCR genotyping, one of the two recurrences was classified as a new infection and the other as recrudescence. In AL group, two recurrences were classified after correction by PCR as recrudescence. All cases of recrudescence were classified as Late Parasitological Failure (LPF).No Early Therapeutic Failure (ETF) was observed. At day 28, 2 patients in the DP group and 5 in AL group had recurrent parasitaemia with In the two groups, almost 94% of patients had cleared parasitaemia within 48 hours after enrollment; there was no statistical difference between the two groups (p = 0.866) Figure . Fever cAmong the 384 patients followed, 35 (23 in DP group and 12 in AL group) had adverse events (9.1%). There was no severe adverse event in the two groups and no treatment was interrupted due to adverse events. Most of adverse events were of moderate severity. The most commonly reported adverse events in both treatment groups were abdominal pain, dizziness, diarrhoea, vomiting, pruritus and nausea  than DP group (0.14/g/dl). The decrease was significant in AL group (p = 0.001) but not in DP group (p = 0.221). In DP group from the beginning of the treatment to day 4, there was a decrease of the mean of AST and a small increase of ALT mean, while in the AL group, AST and ALT means increased. However, these variations were not significantly different. The decrease of the mean of creatinin from the beginning of the treatment to day 4 was not significant in the DP group but was significant in the AL group. In the two groups, the bilirubin decrease was significant . Thus, even if anti-malarial drugs are effective and well-tolerated, it is very important to continue to use prevention tools, such as long-lasting insecticide-treated nets to fight malaria.The haemoglobin rate decrease was significant in the AL group, but not in the DP group. This result suggests that convalescence is obtained faster in the DP group. However, as these data were obtained only four days after enrollment, it would be more significant to assess the haemoglobin rate decrease over 28 days. As in several studies which reported good tolerance of DP ,26 and AP. falciparum malaria in endemic regions. Its once daily dose is even more a significant advantage because it could contribute to improve the patient's treatment compliance.In conclusion, DP presented good and equivalent efficacy and tolerability profile as AL. Therefore, DP is a good alternative for the first-line treatment of uncomplicated DP: combination Dihydroartemisinin - Piperaquine phosphate; AL: Artemether - Lumefantrine.The authors declare that they have no competing interests.OF, SE, KM supervised the clinical studies. KM was the principal investigator. SAO, WY and KM analysed the data. All authors read and approved the final manuscript."}
+{"text": "A three-arm randomized trial conducted among infants in Papua New Guinea estimates the preventive effect against malaria episodes of intermittent preventive treatment, in an area where children are exposed to both falciparum and vivax malaria. Plasmodium falciparum (Pf) transmission. It remains unclear whether IPTi is an appropriate prevention strategy in non-African settings or those co-endemic for P. vivax (Pv).Intermittent preventive treatment in infants (IPTi) has been shown in randomized trials to reduce malaria-related morbidity in African infants living in areas of high n\u200a=\u200a374), SP plus artesunate (AS) , or placebo (n\u200a=\u200a373), given at 3, 6, 9 and 12 mo. Both participants and study teams were blinded to treatment allocation. The primary end point was protective efficacy (PE) against all episodes of clinical malaria from 3 to 15 mo of age. Analysis was by modified intention to treat. The PE (compared to placebo) against clinical malaria episodes  was 29%  in children receiving SP-AQ and 12%  in those receiving SP-AS. Efficacy was higher against Pf than Pv. In the SP-AQ group, Pf incidence was 35%  and Pv incidence was 23%  lower than in the placebo group. IPTi with SP-AS protected only against Pf episodes , not against Pv episodes . Number of observed adverse events/serious adverse events did not differ between treatment arms (p>0.55). None of the serious adverse events were thought to be treatment-related, and the vomiting rate was low in both treatment groups (1.4%\u20132.0%). No rebound in malaria morbidity was observed for 6 mo following the intervention.In this study, 1,121 Papua New Guinean infants were enrolled into a three-arm placebo-controlled randomized trial and assigned to sulfadoxine-pyrimethamine (SP) (25 mg/kg and 1.25 mg/kg) plus amodiaquine (AQ)  in many of the tropical and subtropical regions of the world where malaria is endemic . Malaria is transmitted to people through the bites of night-flying mosquitoes. It can be prevented by controlling the mosquitoes that spread the parasite and by sleeping under insecticide-treated nets to avoid mosquito bites. Prompt treatment of malaria with antimalarial drugs can also reduce malaria transmission. In addition, intermittent preventative treatment (IPT)\u2014the treatment of symptom-free individuals with full therapeutic courses of antimalarial drugs at fixed intervals regardless of their infection status\u2014has been shown to reduce malaria-related morbidity among pregnant women in malaria-endemic areas and among African infants living in areas of high P. falciparum transmission.Malaria is a major global public health problem. Half the world's population is at risk of this parasitic disease, which kills about one million people (mainly young children in sub-Saharan Africa) every year. Most of these deaths are caused by P. falciparum is the predominant parasite and P. vivax is uncommon, it is not known whether IPTi would be an appropriate prevention strategy in non-African settings or in regions where both P. falciparum and P. vivax are endemic. In this randomized placebo-controlled trial, the researchers investigate the efficacy of IPTi in infants living in an area of Papua New Guinea where P. falciparum and P. vivax are both highly endemic. In a randomized placebo-controlled trial, the effects of an intervention and of a placebo (dummy) intervention are compared in groups of individuals chosen through the play of chance.The World Health Organization recently recommended that, in Africa, IPT should be given during infancy  at the same time as routine immunizations. Because the studies on which this recommendation is based were all carried out in sub-Saharan Africa, in populations where P. falciparum and P. vivax combined) was 29% for SP-AQ, but SP-AS was not associated with a statistically significant reduction in all malaria episodes as compared to placebo. For the two species of malaria separately, the incidence of P. falciparum malaria was 35% lower among the children receiving SP-AQ than among the children receiving placebo, whereas the incidence of P. vivax was reduced by 23%; IPTi with SP-AS provided protection only against P. falciparum malaria (protective efficacy 31%). Importantly, the number of adverse events (possible drug side effects) was similar in all the treatment arms, none of the severe adverse events were treatment-related, and there was no rebound in malaria-related morbidity for six months following the end of the intervention.The researchers assigned more than 1,000 infants to receive sulfadoxine/pyrimethamine (SP) plus amodiaquine (AQ) , SP plus artesunate (AS) , or placebo at 3, 6, 9, and 12 months old. They recorded the number of malaria episodes that occurred among the children between the ages of 3 and 15 months. Then, by comparing the number of episodes occurring among the children receiving SP-AS or SP-AQ with the number occurring among the children receiving placebo, the researchers calculated the protective efficacy of the two drug combinations over the study period. The protective efficacy of IPTi against all clinical malaria episodes  can effectively and safely prevent malaria in a non-African population living in a region where http://dx.doi.org/10.1371/journal.pmed.1001195.Please access these web sites via the online version of this summary at malaria ; the 2011 World Malaria Report provides details of the current global malaria situation; and the WHO policy recommendation on IPTi for P. falciparum malaria control in Africa is availableInformation is available from the World Health Organization on malaria (in English and Spanish), including a selection of personal stories about malariaThe US Centers for Disease Control and Prevention provide information on global control of malaria, including a fact sheet about malaria in children and information on malaria in Papua New GuineaInformation is available from the Roll Back Malaria Partnership on the IPTi Consortium was established to evaluate IPTi and inform public health policy makingThe P. vivax malariaThe Malaria Vaccine Initiative has a fact sheet on Vivaxmalaria.com provides information about P. vivaxmalaria (in English and Spanish)MedlinePlus provides links to additional information on  Malaria and anemia are major causes of morbidity and mortality in children in tropical countries In the past 10 y, eight randomized controlled trials have investigated IPTi in different African countries, with various drug regimens. Sulfadoxine-pyrimethamine (SP), dispensed as a single dose three or four times during the first year of life, is the most studied drug and has been associated with a protective efficacy (PE) against clinical malaria episodes of 30% (ranging from 20% to 59%) in populations where SP resistance is not extensive. It also reduced the risk of anemia by 21% in these trials Plasmodium falciparum (Pf) is the predominant parasite and P. vivax (Pv) is uncommon because of the high prevalence of Duffy negativity Pv is a major cause of malaria morbidity, including severe illness and death in young children in particular. Therefore, although IPTi may have significant benefits in these regions, to date there are no data of which we are aware on IPTi efficacy in non-African settings, or on the use of IPTi for malaria due to P. vivax. Results from Africa cannot be easily extrapolated to other settings because of differences in the biology of Pf and PvPv to relapse from long-lasting liver stages, and also the ability of Pv to quickly acquire resistance to SP To date, all reported IPTi studies have been carried out exclusively in sub-Saharan Africa in populations where Pf and Pv specific) and anemia in a Pf and Pv co-endemic setting, we undertook a three-arm randomized placebo-controlled trial of IPTi in an area of Papua New Guinea (PNG) that is highly endemic for both Pf and Pv. We tested two different drug regimens, combining a long-acting drug (SP) with either 3 d of amodiaquine (AQ)  or 3 d of artesunate (AS) , given in conjunction with routine immunization activities. These two drug regimens were chosen for two reasons: (1) they were the first and second line treatments in PNG at the time of the commencement of the study, facilitating their possible implementation into PNG standard practice as treatments for IPTi, and (2) the comparison allows differentiation between the importance of potentiallyimproved clearance of existing infections (using SP-AS) versus better prevention of new infections post-treatment (using the long-acting combination of SP-AQ).In order to determine the efficacy of IPTi in reducing the burden of malaria  given as IPTi at routine vaccination time points  and at 12 mo in conjunction with routine vitamin A supplementation as part of the World Health Organization's Expanded Programme on Immunization (EPI) in PNG. Participants, field teams, and investigators were all blinded with respect to treatment allocation. The efficacy of the intervention was assessed for a 12-mo period from enrollment to a primary assessment time point at 15 mo of age, with a 6-mo extended follow-up (15\u201321 mo) to assess potential rebound.2 coastal area 30\u201360 km north of Madang town. The region receives over 3,000 mm of rainfall annually, with a short dry season (June to October), and is considered to have hyper-endemic malaria The study was carried out between 6 June 2006 and 14 May 2010 on the north coast of PNG in the Mugil area of Sumkar District, Madang Province. The study site included 20 villages situated in an \u223c400-kmOriginally, the trial was started as a two-site study, with a second site in Wosera (East Sepik Province). However, an interim analysis performed mid-study found that the incidence of malaria was dramatically different between the two sites, restricting the comparability of the two sites. Consequently, enrollments were ceased in Wosera in May 2008 and, in line with a revised sample size calculation, enrollments at the Madang site were increased to 334 children per arm. The data from the Wosera cohort were subsequently used only for an overall safety analysis .http://www.clinicaltrials.gov (number NCT00285662) and formed part of the IPTi Consortium  The study was carried out in accordance with Good Clinical Practice (ICH GCP E6) guidelines and externally monitored by two independent monitors and the Data Safety Monitoring Board. The study was approved by the PNG Medical Research Advisory Committee (MRAC number 05.20). The trial was registered on Children were enrolled if they fulfilled the following criteria: (1) permanent resident of the area, (2) aged 3 (\u00b11) mo old, (3) no disability, (4) no chronic illness, (5) no known allergy to study drugs, (6) Hb>5 g/dl, and (7) no severe malnutrition . Study drugs and placebo were identical in appearance and color, but as drugs were administered with sweet syrup no attempt was made to match the moderately bitter taste of AQ. The quality of all drugs and corresponding placebo was independently verified. Study drugs and placebo were assigned to the respective treatment codes by an independent technician. The entire study team and principal investigators remained fully blinded for the entire study period.The three different study interventions were (1) SP-AQ, a single dose of SP (25 mg/kg sulfadoxine and 1.25 mg/kg pyrimethamine) combined with 3 d of AQ (10 mg/kg) plus 3 d of AS placebo; (2) SP-AS, a single dose of SP plus 3 d of AS (4 mg/kg) plus 3 d of AQ placebo; (3) matching placebos for all three drugs. The tablets were split  according to the weight (in kilograms) of each child. The two treatment arms corresponded to the first and second line treatments of the PNG national standard guidelines for malaria at that time.The intervention was delivered four times during the first year of life alongside the EPI at 3, 6, 9, and 12 mo of age . The druThe parents or guardians of potential study participants were contacted during monthly routine EPI clinics run jointly by study staff and health center staff when the child was 1\u20132 mo of age. The study was explained in detail to the parents through both individual and community awareness meetings. An information brochure and consent form in English and in the local language (Pidgin) was given to the parents to take home for further discussion.A child who met the inclusion criteria was formally enrolled during the next clinic visit, when the child was 3 mo of age (\u00b11 mo) and after at least one parent gave written consent. Upon enrollment, a concise medical history including bed net use, possible disabilities, and presence of acute illness was performed. A brief physical examination was done, including measurement of weight. Throughout the study period, a passive case detection system was maintained at the Mugil health center and three outlying clinics where study participants received treatment free of charge. Each illness episode was assessed by study staff using a standard case report form. In the case of history of fever within the past 48 h or an axillary temperature >37.5\u00b0C, a rapid malaria diagnostic test (ICT Combo) was performed, two blood smears and 250 ml of whole blood were collected, and Hb level was measured using a portable HemoCue 201 machine. Only infants positive for malaria on the rapid diagnostic test were treated with artemether/lumefantrine (Coartem). The children with moderate to severe anemia (Hb<8 g/dl) received iron supplementation . All other illnesses were treated according to the standard treatment guidelines of PNG. Children presenting with any danger signs or symptoms were referred to the health center for admission and treatment. Study participants who attended after hours were treated by health center staff based on presumptive diagnosis. In such cases, study staff recorded the reason for admission and (when possible) performed a finger prick to check parasitemia and Hb the next day.All illness episodes were considered adverse events (AEs) and were graded by the study clinicians according to severity from grade 1 (less severe) to grade 3 or as a serious adverse event . Hospital admissions that did not fulfill the criteria of SAE were considered grade 3. The study clinicians assessed all SAEs and causality of AEs. SAEs and AEs possibly or certainly related to the intervention were reported to the Data Safety Monitoring Board, which assessed grade and causality using IPTi Consortium guidelines.Plasmodium species, if the species was detected independently by at least two microscopists. Additionally, if the final slide readings were discrepant compared to PCR results (positivity and species), a final confirmatory read by a senior microscopist was performed. Parasite densities were recorded as the number of parasites per 200 white blood cells and converted to the number of parasites/microliter assuming 8,000 white blood cells/microliter All blood slides were read by two expert microscopists. In case of discrepancies, a third read was performed. Thick blood films were examined by light microscopy for 200 thick-film fields (under 100\u00d7 oil immersion lens) before being declared infection-negative. Slides were scored as light-microscopy-positive for an individual Blood samples were collected from finger pricks in tubes coated with ethylenediaminetetraacetic acid and/or in plain tubes (15-mo visit only). Upon arrival in the laboratory, samples were separated into plasma or serum and cell pellet. Plasma and serum were stored at \u221280\u00b0C until further use. DNA was extracted from cell pellets using the QiaAMP 96 extraction kit (Qiagen) and the 96-well Genomic DNA Extraction Kit (Favorgen).The presence of each of the four human malarial species was assessed in all blood samples using a semi-quantitative post-PCR ligase detection reaction/fluorescent microsphere assay (LDR-FMA) Pf and 0.5 for Pv), a power of 90% and alpha\u200a=\u200a0.025, a sample size of 250 children per arm was estimated to be necessary to find at least a 28% reduction in the incidence of all malaria episodes, a 30% reduction in the incidence of Pv, and a 35%\u201337% reduction in the incidence of Pf episodes. Assuming a drop out of participants of 20% in conjunction with a fall in incidence of malaria of 30% to 50% due to long-lasting insecticide-treated net introduction through the national program in November 2008, a final adjusted sample size of 366 children per arm, or 1,100 children in total, was calculated.Based on a predicted incidence rate of 1.2 episodes/child/year  and according to protocol (ATP). The mITT population includes all randomized children in Mugil that received at least one dose of IPTi or placebo . Following the mITT principle, participants were analyzed according to the preventive treatment they were assigned to at randomization. In the ATP analysis, the population includes all randomized participants who received at least three IPTi treatments (each given within 4 wk after the scheduled treatment time point) and were under passive surveillance for the entire 12-mo period.Plasmodium species from 3 to 15 mo of age. Secondary outcomes included (1) the specific incidence of symptomatic malaria due to Pf or Pv from 3 to 15 mo of age, (2) the incidence of symptomatic malaria  from 15 to 21 mo , (3) the incidence of moderate-to-severe anemia (Hb<8 g/dl) and severe anemia (Hb<5 g/dl) from 3 to 15 mo and 15 to 21 mo, and (4) the prevalence of malaria parasitemia at 15 and 21 mo.The primary objective of the trial was to evaluate the protective effect of the two IPTi regimens. The primary outcome was measured as the incidence of symptomatic malaria due to any Symptomatic malaria was defined as history of fever or axillary temperature \u226537.5\u00b0C and a positive blood smear or a positive rapid diagnostic test, confirmed by positive PCR. The incidences were compared between groups using negative binomial regression to take into account possible extra-Poisson variation due to frailty at the individual level. Analyses were adjusted by sex, number of IPTi treatments received, season of enrollment (wet versus dry), average bed net use, and site (grouped by 12 recruitment zones). The time at risk was calculated starting on the date of enrollment until the date defined according to the analysis, or until the participant was withdrawn from the study. An arbitrary period of 28 d was excluded after each malaria episode. The same methodology was used for evaluating of the incidence of moderate-to-severe (Hb<8 g/dl) and severe anemia (Hb<5 g/dl). Efficacy results are presented as either incidence rate ratio (IRR) or PE (PE\u200a=\u200a1\u2212IRR).The risk of malaria within 35 d following the administration of each of the treatment doses was investigated by calculating incidence rates and hazard ratios using a Cox regression model. Differences in prevalence of infection at 15 and 21 mo were evaluated using logistic regression.The analyses were performed in Stata version 11 (StataCorp).From 6 June 2006 to 14 May 2010, 1,125 3-mo-old infants were enrolled, randomized, and followed-up in the trial in the Mugil area of Madang Province . Four inThe baseline characteristics of all study infants were similar across the three treatment arms . No signPf and Pv, respectively. In mITT analyses, the incidence of all cases of malaria was 29%  lower in the SP-AQ group as compared to placebo, but not significantly reduced in the SP-AS group as compared to placebo . The PE was higher against Pf than Pv in both groups  and Pv incidence by 23%  in comparison to placebo. In contrast, the SP-AS group showed a significant reduction only for incidence of Pf , not for incidence of Pv .The incidence of clinical malaria between the first dose at 3 mo and 15 mo in the placebo arm was 1.07 per person-year at risk (PYAR) overall, and 0.28 and 0.82 for h groups . In the p<0.001), with reductions of 49%  and 31%  for Pf and Pv malaria episodes, respectively. Administration of SP-AS resulted in a 23% reduction in all malaria episodes , with a statistically significant reduction for Pf  but not for Pv .Estimates of protection were higher when only children with three or more IPTi treatments were considered  and Pv  Pf  but not against higher density Pv. The results for the mITT cohort are comparable.Comparable effects were seen when more specific definitions for p\u200a=\u200a0.002) in the SP-AQ group and 34%  in the SP-AS group. Few cases of severe anemia (<5 g/dl) were observed during the study. In the ATP analysis, there were seven cases in the placebo group, one in the SP-AQ group, and two in the SP-AS group (p\u200a=\u200a0.04). Results for the mITT population were of similar magnitude, although in general the estimates obtained were lower and not statistically significant (p\u200a=\u200a0.09\u20130.13)  see .Adjustment for gender, village of residency, season of recruitment, and use of bed nets did not significantly alter estimates of PE .p<0.001) with SP-AQ and 63%  with SP-AS for all malaria infection  than against Pv , whereas SP-AS was more efficacious against Pf  than against Pv . Both SP-AQ  and SP-AS  were associated with a significant reduction in the incidence of Pf malaria between IPTi doses . However, no effect on the incidence of Pv malaria was observed for either treatment group beyond the first 35 d .A total of 6,165 AEs and 275 SAEs were observed during the intervention period, with no significant difference between treatment arms . None ofp\u200a=\u200a0.011). An additional two deaths (one each in the SP-AS and placebo groups) were observed during post-intervention follow-up. The likely diagnoses for the nine deaths were as follows: one severe Pv malaria  with severe anemia and possible lower respiratory tract infection (severe respiratory distress and cough), three lower respiratory tract infections, two severe dehydrations, one meningitis, and two unknown diagnoses.In total, ten deaths occurred among study participants. One was excluded from the analysis because he/she did not receive any IPTi treatment dose. Out of the nine study deaths, seven occurred during the intervention period . Of these, six occurred in the placebo group, one in the SP-AS group, and none in the SP-AQ group . Similarly, no significant differences in incidence of Pf malaria, Pv malaria, and anemia were observed.The incidence of clinical malaria between 15 and 21 mo was 1.09 PYAR overall and 0.43 and 0.70 for Pf and Pv increased from 1.5% and 8.5% at 6 mo to 3.0% and 15.8% at 15 mo and 4.4% and 20.3% at 21 mo, respectively, when detected by microscopy  by light microscopy but not by LDR-FMA. No significant differences in prevalence were observed between treatment arms at 21 mo or any other time point for either Pf or Pv. Infections with P. malariae and P. ovale were rare by microscopy  and LDR-FMA .The prevalence rates of both croscopy . Using mwith age . At 15 mPv malaria. IPTi with a combination of SP plus 3 d of AQ resulted in a 29% decrease in all malaria episodes and a 25% decrease in episodes of moderate-to-severe anemia. PE was higher in children who received at least three doses of IPTi . In line with local resistance patterns and SP-AS pharmacokinetic properties, SP-AS was found to be inferior to SP-AQ. Overall, the efficacy of IPTi with SP-AQ in PNG infants is comparable to the 30% efficacy observed in other studies of African infants receiving IPTi with SP Pf and Pv malaria. Both IPTi treatments were safe and well tolerated. Therefore, these findings provide essential proof-of-principle evidence for the safety and efficacy of IPTi outside Africa, which has important implications for public health policy in non-African settings.This study in PNG infants provides the first evidence of which we are aware for the efficacy of IPTi with SP-based regimens for the prevention of malaria and anemia in a non-African population, including data on the efficacy against Pf than Pv malaria , and was more effective than SP-AS for Pf and Pv malaria. This was not unexpected. Two recent African clinical trials of IPTi have investigated alternative drug regimens, including at least one long half-life antimalarial such as mefloquine (125 mg single dose), a combination of SP (single dose) plus 3 d of AS, or AQ plus AS. These treatments had protective efficacies ranging from 26% to 38% Pf for the first 35 d (72%\u201382%) as well as a moderate inter-dose effect, resulting in an overall efficacy of IPTi against Pf that was towards the higher end of those found in African studies Overall, IPTi with SP-AQ was more effective in preventing Pv episodes in the first 35 d following treatment, indicating a similar post-treatment prophylactic effect, but the overall efficacy of IPTi against Pv malaria was lower than for Pf. SP-AS, however, showed significantly less post-treatment prophylactic effect (PE\u200a=\u200a59%) and no effect against Pv malaria overall. The most probable reasons for this are the ability of Pv to relapse from long-lasting liver stages Pv. Pv strains from New Guinea are thought to relapse very frequently Pv blood-stage infections are therefore re-established soon after drug treatment. In a concurrently conducted drug trial, 49% of PNG children 0.5\u20135 y treated with SP-chloroquine (3 d) for Pv malaria had recurrent Pv parasitemia, and an additional 18% had a recurrent Pv clinical episode by day 42, while 26% and 6% of children treated for Pf malaria had recurrent Pf parasitemia and clinical episodes, respectively SP-AQ also prevented 83% of Pf and Pv malaria is consistent with the pharmacokinetic properties of the drugs used and previously observed drug resistance patterns. PCR-corrected day 28 in vivo failure rates from studies conducted concurrently or just prior to the trial were 10.3% for SP-AQ Pf, and 3.6% for SP-AQ Pv. As SP has never been used as monotherapy in PNG, there are no estimates of SP in vivo resistance for either Pf or Pv. However, molecular markers of SP resistance have been analyzed for both species in the study population. In Pf, the quintuple Pf dihydrofolate reductase (pfdhfr) 51\u201359\u2013108/Pf dihydropteroate synthase (pfdhps) 436\u2013547 mutant genotype is associated with a high degree of drug resistance Pv, the 57L-58R-61M-117T pvdhfr quadruple mutation (irrespective of pvdhps mutations) has been associated with increased risk of treatment failure pvdhfr mutation in PNG  976F quintuple mutant genotype has been associated with parasitological failure pvmdr1 976F mutation was present in \u223c70% of Pv isolates tested in 2006/2007, the predominantly late occurrence of recurrent parasitemia  following treatment with SP-chloroquine indicates that 4-aminoquinolines retained good clinical and at least partial parasitological efficacy Although chloroquine-resistant Pf is likely to be largely due to the effect of SP. The high level of preventative efficacy for 35 d is consistent with the terminal elimination half-life (t1/2\u03b2) of 15.6 and 9.1 d for pyrimethamine and sulfadoxine, respectively, in PNG infants Pv, the success of IPTi against Pv malaria relies largely on the efficacy of the partner drug. Whereas Pv parasites are efficiently cleared by SP-AS, the short half-life of AS provides no post-treatment prophylactic effect, and recurrent Pv infections are very common Pv but not Pf adds further evidence to the observation that while SP can retain a considerable effect at low-to-moderate levels of resistance Given the high level of AQ resistance and the very short half-life of AS Besides reducing the burden of malaria episodes, IPTi also reduced the risk of moderate-to-severe anemia  and severe anemia (51%\u201387%). These effects are in line with previous findings on the prevention of anemia in Africa No difference was observed between the groups in number of SAEs . Surprisingly, a significantly lower death rate was observed in the two treatment arms compared to the placebo arm. Even though this finding is very encouraging, it must be interpreted with caution. The present study was not powered to observe an effect of IPTi on mortality, and in the meta-analysis of six African studies, no effect of IPTi with SP on mortality was detected Pf and Pv and lead to a rebound in malaria risk. These data are very reassuring in regard to a possible use of these drug combinations in PNG and add to the safety data from African IPTi studies IPTi with SP-AQ or SP-AS was very safe. More than 4,000 doses of SP and 6,000 doses of AQ and AS were given, and no study-drug-related SAEs were observed, while AEs such as vomiting were rare in all groups. As reported in earlier studies, no significant increase in risk of malaria or anemia was recorded in the 6 mo after completion of the 12 mo of IPTi, indicating that IPTi does not impair the acquisition of immunity to both The use of SP as prophylaxis has been criticized because of the potential risk of Stevens-Johnson syndrome, a severe skin reaction The size of the difference in IPTi efficacy between the ATP and mITT analyses indicates that if the intervention is implemented, it will be important to achieve high coverage and high adherence in order to achieve good effectiveness.In both the SP-AQ and SP-AS groups only the first dose was given as directly observed treatment. Parents were then counseled on the importance of giving the remaining two doses at home. With this approach a very high rate of compliance (>90%) was achieved. As a result, SP and AQ day 4 drug levels were found to be comparable  to those found in pharmacological studies with directly observed drug administration Pf and Pv. Additionally, this study provides an essential proof of principle that IPTi is an appropriate strategy for the prevention of Pv malaria, if an effective, long half-life drug is used. Policy makers should therefore consider IPTi in areas outside Africa or in countries in the Horn of Africa, where the burden of Pv malaria is high. Which areas are suited for IPTi introduction will depend on transmission level and choice of drug, both of which will affect the cost-effectiveness of the intervention Pv, in particular.This study provides evidence of the efficacy of IPTi for the prevention of malaria and anemia in a region highly endemic for both Pf [SP] or Pv [AQ]) is an appropriate drug choice for IPTi, and its introduction into the national standard treatment guidelines should be considered. The replacement of AQ and SP by artemether-lumefantrine as the national first line treatment will reduce the selection pressure for resistance against AQ and SP. Therefore, the efficacy of the two drugs in IPTi may be retained over time. Appropriate monitoring of prophylactic efficacy of AQ and SP, as well as evaluating new regimens, such as dihydroartemisinin-piperaquine, should accompany IPTi introduction in PNG.In the PNG context, the combination of SP-AQ Click here for additional data file.Table S2Baseline characteristics of study participants\u2014ATP cohort.(DOCX)Click here for additional data file."}
+{"text": "This case study describes how a public-private partnership initiated to develop a new anti-malarial combination, ASAQ Winthrop, has evolved over time to address issues posed by its effective deployment in the field.i created the FACT project to develop two fixed-dose combinations, artesunate-amodiaquine and artesunate-mefloquine, to meet the WHO anti-malarial treatment recommendations and international regulatory agencies approval standards. In 2002, Sanofi-aventis had started a development programme for a fixed-dose combination of artesunate and amodiaquine, to replace its co-blister combination. DNDi and sanofi-aventis joined forces in 2004, with the objective of developing within the shortest possible time frame a non-patented, affordable and easy to use fixed-dose combination of artesunate and amodiaquine adapted to the needs of patients, in particular, those of children. The partners developed Coarsucam\u00ae/Artesunate Amodiaquine Winthrop\u00ae (\"ASAQ Winthrop\") which was prequalified by the WHO in 2008. Additional partnerships have since been established by DNDi and sanofi-aventis to ensure: 1) the adoption of this new medicine by malaria-endemic countries, 2) its appropriate usage through a broad range of information tools, and 3) the monitoring of its safety and efficacy in the field through an innovative Risk Management Plan.In 2002, DNDi and sanofi-aventis has enabled the development and pre-qualification of ASAQ Winthrop in a short timeframe. As a result of the multiple collaborations established by the two partners, as of late 2010, ASAQ Winthrop was registered in 30 sub-Saharan African countries and in India, with over 80 million treatments distributed in 21 countries. To date, 10 clinical studies, involving 3432 patients with ASAQ Winthrop were completed to document efficacy and safety issues identified in the Risk Management Plan.The partnership between DNDThe speed at which ASAQ Winthrop was adopted in the field shows that this drug fits the needs of patients and health authorities. It also demonstrates the power of partnerships that combine different sets of strengths and skills, and that evolve to include additional actors to meet new global health challenges for poverty-related diseases. Due to the high efficacy of artesunate-based combination therapy (ACT) in the treatment of uncomplicated malaria in Africa and Asia, a technical consultation organized by the WHO in 2001 recommended their use to provide effective treatment against malaria and to slow down the spread of drug resistance. In 2006, WHO further stated its preference for ACT in fixed-dose combinations to overcome the issues related with non-fixed dose options [initiative (DNDi) and sanofi-aventis to develop together a fixed-dose combination of artesunate and amodiaquine, one of the forms of ACT recommended by the WHO, which at that time only existed as a non-fixed combination of the two drugs. This case study aims at retracing the main features of this partnership, and describes how it was progressively enlarged to include new partners to maximize the launch and distribution of the new medicine. The lessons from this experience may help in the development of other need-driven drugs.A partnership was initiated in 2004 between the Drugs for Neglected Diseases i Working Group, which led to the creation of the DNDi foundation in 2003, the Fixed-Dose Artesunate-based Combination Therapies (FACT) Project was initiated. Its aim was to develop two fixed-dose combinations, namely artesunate-amodiaquine and artesunate-mefloquine, in the shortest possible timeframe to meet the WHO treatment recommendations and international quality standards. The FACT project consisted of DNDi, Farmanguinhos/Fiocruz (Brazil), Tropival of the Bordeaux II Victor-Segalen University (France), Oxford University (UK), Universiti Sains , Mahidol University (Thailand), the Special Programme for Research and Training in Tropical Diseases WHO/TDR (Switzerland), and the Centre National de Recherche et de Formation sur le Paludisme (Burkina Faso). In line with DNDi's mission of collaboration based on relative strengths, the FACT project capitalized on the skills and know-how of a broad range of partners in both developing and developed countries. Initially, the FACT project was financially supported by the European Union (INCO-Dev programme) and M\u00e9decins Sans Fronti\u00e8res, then by government agencies from France, the Netherlands, Spain and the UK. At the end of 2004, the formulation of the \"bilayer\" tablet had been developed with good stability characteristics and a packaging that guaranteed stability in tropical field conditions [In 2002, under the auspices of the DNDnditions Meanwhile, in 2004, sanofi-aventis had registered and launched a co-blistered, non-fixed form of artesunate and amodiaquine (Arsucam\u00ae) in sub-Saharan Africa. Recognizing the advantages of fixed-dose combinations to improve patient compliance, dosing accuracy and avoiding the risks related with monotherapy, the company had also initiated, in 2002, a programme to develop a fixed-dose combination of artesunate and amodiaquine.i and sanofi-aventis decided to join forces to form a public-private partnership with the objective of developing a non-patented fixed-dose combination of artesunate and amodiaquine that would be made available at prices lower than those available at the time, i.e. less than US$1 for an adult treatment and less than US$0.50 for a child's treatment in the non-profit public sector. Sanofi-aventis did not seek any patent protection. This decision was made to be consistent with DNDi's intellectual property policy to develop drugs as public goods when possible, and with both partners' determination to make the new medicine widely available to those patients in greatest need. Sanofi-aventis agreed to pay DNDi 3% of the net private sector turnover over a period of seven years. DNDi decided to use this payment to support the Risk Management Plan, which is described later in this paper.In December 2004, DNDi's key contribution at the initial stage of the partnership was brought by the University Victor-Segalen of Bordeaux and then by Ellipse Pharma (France), which developed the \"bi-layer\" formulation with the objective of keeping separated the artesunate and the amodiaquine components, and defined the key features of the manufacturing process of the fixed-dose combination. The process allowed for a reduction in the size and number of tablets, as well product stability in tropical climates [DNDclimates ,6.i enabled making the most of each partner's skills and resources, and a steering committee provided arbitrage in case of disagreement. Project teams composition evolved over time as required by the various steps of the development . Meetings between the two parties were jointly managed by one representative of each partner. The roles and responsibilities of each partner, and, therefore, the rules for decision-making, were laid out early on. DNDi was responsible for the actual formulation of ASAQ Winthrop, early Phase I data, as well as for one pivotal comparative study. Sanofi-aventis was in charge of the manufacturing process at industrial scale, managing the second pivotal clinical study, building the registration file, ensuring registration in endemic countries, obtaining WHO pre-qualification and launch and marketing of the new drug. Decisions which could not be reached by consensus between the two parties were discussed at steering committee meetings which occurred approximately four times a year. When consensus could still not be reached, the final decision was made by the party responsible for carrying out the task under consideration. Collaborative work between the two partners led to the following outcomes:Once collaboration was started, regular joint projects teams between sanofi-aventis and DNDi was of the opinion that the age-based dosing was more practical in developing countries. Sanofi-aventis, however, was in favour of developing a weight-based formulation in keeping with major regulatory agencies regulations. Both partners came to consensus thanks to an innovative body of work that modeled actual weight-for-age data from malaria endemic African countries [The development of the ASAQ Winthrop tablets strengths was focused towards optimizing safe and effective drug levels across all age groups, with a specific emphasis on the needs of children who are the first victims of malaria. The formulation of ASAQ Winthrop enables the drug to be dissolved in water, making it easier to administer to infants and small children. In most endemic countries, dosing recommendations based on age are simpler to use in the field than those based on bodyweight. DNDountries . This da- artesunate: 2 to 10 mg/kg (5 fold), reflecting its tolerability and, therefore, wide- therapeutic index.- amodiaquine: 7.5 to 15 mg/kg (2 fold). There was less flexibility with amodiaquine, therefore, amodiaquine determined the final dosing and age categories.In addition, two conventions were adopted:- select age groups with an approximate doubling in median bodyweight, and- double the drug dose per age category.These conventions led to selecting 4 dosage forms based on age and weight ranges, for infants , toddlers , children  and adults . Figure i to cover all tasks required to license ASAQ Winthrop: industrial pharmaceutical development ; pre-clinical pharmaco-toxicology studies to document the safety profile of artesunate plus amodiaquine in animals; clinical ; regulatory , as well as marketing and medical activities to prepare the drug's launch and marketing in endemic countries and design post-launch clinical studies to monitor its safety and efficacy in the field.Between 2004 and 2007, a joint project team was set up between sanofi-aventis and DNDThe registration dossier included clinical data on 1,003 patients treated with ASAQ Winthrop from two randomized controlled studies: one managed by DNDi in Burkina Faso comparing ASAQ Winthrop to a non-fixed combination of artesunate and amodiaquine , and ano\u00ae, for private markets (1 blister per box), and Artesunate Amodiaquine Winthrop\u00ae for public markets (25 blisters per box). Winthrop is the name of sanofi-aventis generic range of medicines. The shortened name \"ASAQ Winthrop\" in this article refers to both brand names and is used to highlight the fact that this product is a specific fixed-dose combination of artesunate and amodiaquine. The registration dossier was submitted in December 2005 and approval was granted in Morocco on February 1, 2007, making ASAQ Winthrop the first new anti-malarial treatment developed and delivered by a public-private partnership [\u00ae), at market prices that include profit margins. Specifically, ASAQ Winthrop is available in the public sector at prices below $1 for adults and $0.50 for children, while Coarsucam\u00ae is sold for approximately $2-$3 to wholesalers for the private markets. The profit margins made though sales in the private sector ensure that the mechanisms that enable very low \"no profit-no loss\" prices in the public sector can be sustained over the long-term.Many discussions were held over the best way to register ASAQ Winthrop to make it available as quickly as possible in endemic countries, while demonstrating its adherence to high-quality standards. It was decided to seek initial registration from Morocco, the country where the drug is manufactured, to enable registration in endemic countries, but to also apply for WHO prequalification as further evidence of ASAQ Winthrop's quality standards, and to enable purchasing by international agencies, The Casablanca production plant was chosen in line with sanofi-aventis' commitment to develop its existing industrial assets in developing and emerging countries. A major financial investment was made to ensure appropriate production capacity level, as well as adherence of the plant to Good Manufacturing Practice standards. To allow for differential pricing between the public and private markets, two brand names were selected: Coarsucamtnership . Followii wanted to ensure that appropriate post-marketing data was available as quickly as possible on ASAQ Winthrop safety and effectiveness in the field. This was deemed necessary because of the relatively poor image of amodiaquine safety, in spite of its re-installment in the 1990s as a safe and effective drug for the treatment of malaria [Early in their collaboration, sanofi-aventis and DND- Several randomized controlled studies in sub-Saharan African countries that provide safety and efficacy data on ASAQ Winthrop, compared with other forms of ACT, usually in the treatment of single malaria episodes in patients with documented malaria.- Two cohort studies in Senegal and in Uganda: in each study, cohorts of 200 patients were randomly assigned to receive either ASAQ Winthrop or artemether-lumefantrine for each malaria attack occurring during the two-year study period. The Uganda study focuses on children less than five years of age. The Senegal study includes both adults and children.- A Phase IV field programme to assess ASAQ Winthrop \"real life\" safety and effectiveness over two years in a C\u00f4te d'Ivoire health district that uses ASAQ Winthrop as first-line treatment, through two approaches. First, an active monitoring of clinical tolerability and compliance for all malaria patients attending district health centres for treatment, performed by community health care workers who visit treated patients at home to collect information. In this study, patients diagnosed with malaria are prescribed ASAQ Winthrop to take at home under non-supervised conditions as is standard medical practice currently in C\u00f4te d'Ivoire. Blood smears are collected to enable post-treatment laboratory diagnosis of malaria, to identify which patients definitely had malaria. This approach will enable getting a \"real-life\" view of the drug's safety and efficacy in patients treated after clinical diagnosis as frequently happens in Africa, and to assess possible differences between patients with and without biologically-proven malaria. The World Malaria Report 2009  and the As more data on ASAQ Winthrop, generated through studies managed by either one of the two partners or by other institutions become available, they will be pooled into a common database and analysed by an academic investigation team to provide comprehensive information on the drug's efficacy and safety.The WHO Department of Medicines Policy and Standards expressed a strong interest in this programme, and requested a formalized Risk Management Plan (RMP). This was done in 2009, and a series of documents was submitted following the format required by European Medicines Agency for RMP. Table i partnership to set up the C\u00f4te d'Ivoire field programme, the most ambitious component of the RMP, and is funding most of the costs thereof. This collaboration was seen by MMV as a way to develop and test methodologies that could be relevant to design RMP for future anti-malarials.In 2009, the Medicines for Malaria Venture (MMV) joined the sanofi-aventis/DNDi have engaged in collaborations with a number of partners, including experts, national malaria programmes, WHO and other international organizations, research institutes, funding agencies, and NGOs. DNDi has, for instance, coordinated a clinical study with its founding partner, the Indian Council of Medical Research, to facilitate the adoption of a new anti-malarial policy in India; this study allowed registration of ASAQ Winthrop in India. DNDi has also convened an independent panel of experts, the FACT Implementation Advisory Group, to provide independent advice and critical guidance about issues related to implementation and rational use of ASAQ Winthrop, ensuring equitable access.The partnership that led to the development and registration of ASAQ Winthrop had to evolve to ensure the new drug's successful launch and adoption by endemic countries. First among the many partners needed at this crucial stage are the endemic countries' National Malaria Control Programmes (NMCP) that are charged with the design and implementation of national action plans to control malaria. In working to facilitate the implementation and availability of ASAQ Winthrop, both sanofi-aventis and DNDSanofi-aventis has focused its efforts on supporting the introduction of the new treatment in public health systems through a series of information tools to assist with the change in prescription and dispensing habits. In partnership with African NMCP and malaria experts, a variety of information and educational tools were designed, not only about the rationale and instructions on how to use a fixed-dose ACT, but more broadly about the comprehensive management of the disease, that includes malaria prevention, diagnosis and treatment. Specific supports are designed for each \"constituency\" involved in the fight against malaria, such as textbooks, slide kits, CDs, educational flipcharts, games, etc. Information for physicians was focused on malaria diagnosis and treatment, while those aimed at communities focused primarily on malaria prevention. These tools are non-promotional and aim at providing the most comprehensive and up-to-date information in a way that is adapted to everyone's role.Two of these tools deserve highlighting since they illustrate how widely different techniques can be used to convey overall similar messages to different audiences. Firstly, a \"train the trainers\" module is proposed to update countries' NMCP \"senior trainers\" on malaria management. These persons will then be responsible for cascading the information through the national public system networks so that best practices are widely used. A comprehensive training module on malaria diagnosis, treatment and prevention is delivered by African university professors through a three-day workshop organized with approximately 15 NMCP senior trainers. In addition to the technical training provided by these experts, a day is spent with a communication specialist to improve the educational techniques and teaching skills of the trainers. Together with supporting materials provided at the end of the training session, the NMCP senior trainers are thus better equipped to organize effective training sessions for their colleagues in the field.Another initiative called \"Schoolchildren against Malaria\" was designed, aimed at young children approximately 10 years of age. In this programme, schoolchildren are taught about the basics of malaria, with a focus on preventative measures at the community level, and are asked by their teachers to design a theatre play on this theme.To create some emulation, a competition is organized among schools, and the final contest for the best play is organized as a festive event. Through this approach, children not only learn about malaria but that they will also act as information relays for their families and friends. Between 2008 and 2010, a total of approximately 200,000 children were involved in this initiative in C\u00f4te d'Ivoire, Ghana and Burkina Faso. This approach, which builds on the African tradition of oral culture and theatre, will be expanded to additional countries in the coming years. Educational tools offered to countries and NGOs that want to introduce ASAQ Winthrop are not protected by patents or copyrights so that they can be used, copied and adapted as broadly as needed, according to each country's needs and priorities.As is the case for HIV/AIDS and tuberculosis, most of the malaria drugs made available in public health systems are bought through international agencies . With the advent of the Affordable Medicines Facility-malaria (AMFm) initiative, major adaptations to the packaging of eligible medicines are requested by the funders, to enable, on a country-by-country basis, the identification of drugs purchased through this facility . Althougi launched ASAQ Winthrop, their partnership was perceived as an unlikely marriage. Nevertheless, one of the world's largest pharmaceutical companies and a not-for-profit R&D organization successfully brought their respective strengths together in order to respond to an urgent public health need. Through their efforts, the organizations managed to provide patients with a new anti-malarial and prove that public and private organizations can successfully work together to achieve a common objective.This paper intends to illustrate how a two-sided product development partnership initiated to develop a new anti-malarial drug has evolved to include additional partners to address the challenges posed by the launch of this new drug, its effective introduction in national programmes, and the need to ensure a steady supply. What lessons can be drawn from this experience? In early 2007, when sanofi-aventis and DNDi possessed all of these attributes. ASAQ Winthrop represents a good example of needs-driven innovation. While it does not represent an innovative new molecule developed from the benchtop , it represents a very significant innovation for patients in resource-poor settings. In a disease where drug resistance represents a major problem, a fixed-dose combination with a simple dosing regimen provides a clear improvement over non-fixed combination of the two separate drugs. The fixed-dose combination ensures that patients take both drugs simultaneously and in the correct proportions. The additional advantage of tablets that can be dissolved in water was specifically developed for children and it spared the need to develop a distinct pediatric formulation, which would have required additional time and funds.A successful partnership requires a common desire, complementary knowledge, and shared responsibilities. The ASAQ Winthrop partnership between sanofi-aventis and DNDThe availability of a new medicine does not imply access in the field. As such, the partnership made two bold commitments. First, the product would receive no patent protection. Second, the partners set a target price of one 1 USD per treatment for adults and 0.5 USD cents for children. Before ASAQ Winthrop's introduction to the marketplace, the price for most ACTs in public markets was approximately 2.50 USD for an adult treatment. After ASAQ Winthrop's introduction, the global reference price for ACTs on public markets decreased to approximately 1 USD.raisons d'\u00eatre will not always agree and will have different views and approaches towards achieving a common goal. This has, on more than one occasion, led to debates, and delayed decisions. The ASAQ Winthrop partnership, sanofi-aventis and DNDi learned how to utilize their respective skills to have the greatest possible impact. For instance, sanofi-aventis provided its expertise in regulatory practices that was critical in obtaining registration in countries and prequalification by the WHO, and DNDi stressed the importance of providing instructions on packages that patients with low literacy could understand. In addition, the two organizations engaged complementary networks of contact organizations and found that dialogue and actions were often times easier and more fruitful when the partners worked together rather than in isolation. Lastly, the launch of ASAQ Winthrop in 2007 drew wide media attention in both specialized and lay press. It is possible that the global media exposure of the \"$1 malaria treatment\" played a role in establishing the new reference price for anti-malarials. Both organizations now understand the importance of creative and innovative contracts between public-private partnerships, and that joining forces in a common development team can allow for programme acceleration.Clearly, two organizations with different i from sales on the private markets. The lessons learned from this experience could prove invaluable in assessing future drugs' efficacy and safety in the field. Partnering with NMCPs to design and implement informational tools and techniques proved to be a very rich, and ongoing, experience. Each country has its own priorities in terms of information needs, and offering a variety of tools enables targeting specific audiences with adapted tools and techniques. While tools intended for health professionals usually only require minor country-specific adaptations, those intended for communities often require translation into local languages, as well as the adaptation of pictures and drawings, so that local populations effectively relate with the messages. Similarly, collaboration with international purchasing agencies requires adaptation to each organization's requirements and working habits, to establish long-term effective relationships. Lastly, the importance of the contribution of public and philanthropic funders at all stages of the partnership cannot be overemphasized.Both organizations decided to extend their successful collaboration to ensure the deployment of ASAQ Winthrop and monitor its long-term safety and efficacy. This was formalized through the Risk Management Plan, that is partly supported by Medicines for Malaria Venture and by the royalties that sanofi-aventis pays to DNDi or academic teams, have been completed, involving 3432 patients and 9986 malaria episodes treated with ASAQ Winthrop, in 16 African countries, as well as in India and in Colombia  Project; RMP: Risk Management Plan; WHO: World Health Organization; NMCP: National Malaria Control Programme; NGO: Non-Governmental Organization.ACT: artemisinin-based combination therapy; ASAQ Winthrop: Coarsucaminitiative.FB and RS are employed by sanofi-aventis, JRK and BP are employed by the Drugs for Neglected Diseases FB wrote the initial manuscript, JRK, RS and BP made substantial contributions and helped to finalize the manuscript. All authors read and approved the final manuscript."}
+{"text": "Artemisinin-based combination therapy is currently recommended by the World Health Organization as first-line treatment of uncomplicated malaria. Recommendations were adapted in 2010 regarding rescue treatment in case of treatment failure. Instead of quinine monotherapy, it should be combined with an antibiotic with antimalarial properties; alternatively, another artemisinin-based combination therapy may be used. However, for informing these policy changes, no clear evidence is yet available. The need to provide the policy makers with hard data on the appropriate rescue therapy is obvious. We hypothesize that the efficacy of the same artemisinin-based combination therapy used as rescue treatment is as efficacious as quinine + clindamycin or an alternative artemisinin-based combination therapy, without the risk of selecting drug resistant strains.We embed a randomized, open label, three-arm clinical trial in a longitudinal cohort design following up children with uncomplicated malaria until they are malaria parasite free for 4 weeks. The study is conducted in both the Democratic Republic of Congo and Uganda and performed in three steps. In the first step, the pre-randomized controlled trial (RCT) phase, children aged 12 to 59 months with uncomplicated malaria are treated with the recommended first-line drug and constitute a cohort that is passively followed up for 42 days. If the patients experience an uncomplicated malaria episode between days 14 and 42 of follow-up, they are randomized either to quinine + clindamycin, or an alternative artemisinin-based combination therapy, or the same first-line artemisinin-based combination therapy to be followed up for 28 additional days. If between days 14 and 28 the patients experience a recurrent parasitemia, they are retreated with the recommended first-line regimen and actively followed up for another 28 additional days . The same methodology is followed for each subsequent failure. In any case, all patients without an infection at day 28 are classified as treatment successes and reach a study endpoint. The RCT phase allows the comparison of the safety and efficacy of three rescue treatments. The prolonged follow-up of all children until they are 28 days parasite-free allows us to assess epidemiological-, host- and parasite-related predictors for repeated malaria infection.NCT01374581 and PACTR201203000351114 Malaria remains one of the great infectious killers in Africa. An estimated 300 to 500 million cases occur each year, causing 1.5 to 2.7 million deaths, primarily in children under the age of 5 years [Following the World Health Organization (WHO) guidelines, most African countries have already opted for artemisinin-based combination therapy (ACT). Several clinical trials on artesunate-amodiaquine (ASAQ), an ACT, completed in Africa have shown an efficacy >90% -6. FurthPlasmodium falciparum in vitro sensitivity to quinine was high and had not changed over the past decade [P. falciparum, it remains an important drug for severe malaria, although the current trend is to replace this with intravenous artesunate, which in some settings has already occurred whilst in others it is currently happening. Therefore, quinine should be protected from resistance by rational use, as its effectiveness in uncomplicated malaria is lower than ACT [In DRC and Uganda, quinine is the rescue treatment for malaria. It is cheap, widely available and generally considered to be effective but is not popular due to its side effects. Quinine has a very short half-life; therefore, repeated dosing is required. In an efficacy study of quinine and artemisinin for uncomplicated malaria in Vietnam, recrudescence rates were 16% after 7 days of quinine monotherapy . In studt decade . Althougt decade . As a ret decade , which ct decade . Furtherthan ACT .P. falciparum multidrug resistance (pfmdr)1 will be analyzed in order to evaluate the potential risk of selecting drug-resistant strains. The longitudinal, continued follow-up of all treatment failures will additionally enable host-, environmental- and parasite-related predictors of recurrent, recrudescent or reinfections to be identified.This study aims to assess the possible impact of using the same first-line treatment as rescue treatment. Furthermore, as part of the study protocol, mutations in the Considering the facts that (i) over >75% of treatment failure to ASAQ or AL are new infections, (ii) parasite density is low in case of recrudescence occurring from day 14 onwards, and (iii) in real-life situations patients are retreated with the same first-line drug, there is a need to assess the role of the first-line treatment as rescue treatment. This efficacy will be compared to quinine + clindamycin and another ACT treatment in line with the WHO guideline . We hypoP. falciparum malaria within 42 days of treatment with an ACT (ASAQ in DRC or AL in Uganda), the PCR adjusted efficacy at 28 days after retreatment with the same artemisinin-based combination therapy is at least 90%; and (2) to estimate the relative efficacy of retreatment with the same ACT compared to treatment with quinine + clindamycin and treatment with an alternative ACT (ASAQ after first-line AL treatment or AL after first-line ASAQ treatment).The primary objectives are: (1) to show that, in children aged 12 to 59 months with recurrent uncomplicated P. falciparum malaria episode occurring 2 weeks after the administration of the first-line treatment, with and without PCR adjustment; (3) to evaluate and compare the 42-day clinical efficacy of AL (Uganda) and ASAQ (DRC) for the first-line treatment of uncomplicated P. falciparum malaria, with and without PCR adjustment; and (4) to evaluate and compare the efficacy of the different rescue treatment regimens in terms of fever clearance time, asexual parasite clearance time, gametocytemia at days 7, 14, 21 and 28, and hemoglobin changes between day 0 and days 14 and 28.Secondary objectives are: (1) to evaluate the PCR-unadjusted efficacy at 28 days of retreatment with the same ACT and to compare it to treatment with quinine + clindamycin and treatment with another ACT (ASAQ after first-line AL treatment or AL after first-line ASAQ treatment); (2) to evaluate and compare the efficacy of AL, ASAQ and quinine + clindamycin as rescue treatment for a recurrent P. falciparum pfmdr1 and other alleles following therapy with quinine + clindamycin, AL and ASAQ; and (2) to assess epidemiological-, parasitological- and host-related predictors (including genetic factors) for recurrent malaria infections (adjacent studies will be developed in separate nested-study protocols).Additional objectives are: (1) to evaluate transcriptional changes and selection of To evaluate the safety and tolerability of AL, ASAQ and quinine + clindamycin when used as rescue treatments.P. falciparum. Quinine belongs to the aryl amino alcohol group of drugs. It is a cinchona alkaloid that has a rapid schizonticidal action on the intra-erythrocytic parasites and is also gametocytocidal for P. vivax and P. malariae but not for P. falciparum.Quinine is an alkaloid from the bark of the cinchona tree that still constitutes one of the major components of the antimalarial pharmacopeia, as it has for over three centuries. Quinine is still widely used for the treatment of severe malaria and serves in oral formulation as a reserve drug for uncomplicated malaria, mostly in combination with a tetracycline or doxycycline or clindamycin. It is effective against all species of malaria including chloroquine-resistant strains of In DRC, quinine + clindamycin is the second-line treatment for malaria. In Uganda, quinine is the rescue treatment, but a combination with clindamycin might be expected in the near future. Quinine is cheap, widely available and generally considered to be effective though it is not popular due to the unwanted side effects. Quinamax\u00ae is a formulation developed by Sanofi-Aventis  which comprises four alkoloids: quinine, quinidine, cinchonin and cinchonidine. In this study, we use dry tablets (125 mg) with no dosage adaptation for children below 9 kg (which explains why children below 12 months are excluded from our study).Clindamycin is a lincosamide antibiotic derivative of lincomycin. It is very soluble in water. It inhibits the early stages of protein synthesis by a mechanism similar to that of the macrolides. It may be administered by mouth as capsules containing the hydrochloride or as oral liquid preparations containing the palmitate hydrochloride. Clindamycin is used twice daily for at least 5 days at 10 mg/kg for children aged 11 years and under. Clindamycin used in combination with quinine is safe but limited data have so far been gathered . In thisPlasmodium infections. The combination is expected to confer mutual protection against resistance and prevent recrudescence after artemether therapy. The components of this combination were originally studied and developed in China by the Academy of Military Medical Sciences . The fixed combination has been registered in China since 1992 and has undergone further development when Novartis  signed a collaborative agreement in 1994 with the Academy of Military Medical Sciences and CITITEC, the technology arm of the China International Trust and Investment Corporation . Studies for the international registration started in 1995. AL from Novartis, marketed under the trademarks Riamet\u00ae and Coartem\u00ae, was registered in Switzerland in 1999. It is prequalified by WHO and has since received marketing authorization in several endemic and non-endemic countries. A recent review showed that the drug combination is highly efficacious against sensitive and multidrug resistant falciparum malaria as it offers the advantage of rapid clearance of parasites by artemether and the slower elimination of residual parasites by lumefantrine [This is a fixed-dose combination of artemether (a semi-synthetic artemisinin derivative) and lumefantrine . The registered indications and branding for AL cover treatment of uncomplicated malaria caused by mono or mixed fantrine .ASAQ  is safe, easy to use and efficacious, and the second most used ACT worldwide ,6. DRC, This is a bi-centre, phase IIIb, randomized, open label, three-arm trial. It is performed in three steps Figure\u00a0 and therAll patients are treated with the first-line treatment - ASAQ for DRC and AL for Uganda. Before treatment, a blood sample is collected on filter paper  for subsequent parasite genotyping. A serum sample is also collected and frozen for further immunological assessment (adjacent study). Patients are passively followed up for the next 42 days. During this period, no systematic screening for malaria infection is done. Instead, blood samples are collected only for patients attending the health facilities with suspected clinical malaria. In case of confirmed failure before day 14, patients are treated with quinine + clindamycin and excluded from the follow-up as they have reached one of the endpoints. Patients experiencing a clinical failure (fever and parasitemia with any parasite density) between days 14 and 42 are eligible for the second phase. The day of failure corresponds to day 0 of the following phase. Patients are assessed as summarized in Additional file This step constitutes the core of the study. After the second informed consent , patients are randomly assigned to ASAQ, AL or quinine + clindamycin. Rescue treatment allocation is concealed until the recruitment of the patient in the RCT phase. The randomization list was generated prior to the beginning of the study by the study statistician. At the study site, treatment allocation and administration of medications are performed by the study physician or nurse. Patients are assessed as summarized in Additional file Parents/guardians are encouraged to return to the clinic for follow-up assessments on days 3, 7, 14, 21 and 28, and on any unscheduled day if the child is not well.All patients included in the RCT phase and presenting a clinical or parasitological failure from day 14 onwards are retreated with the country\u2019s first-line treatment (AL or ASAQ). We expect about 50 patients per site to participate in this phase. All patients are followed up exactly as during the RCT and study procedures are also identical. In case of confirmed failure before day 14, patients are treated with the first-line ACT and excluded from further follow-up. The same procedures are followed for each repetitive treatment failure occurring between 14 and 28 days after each treatment course. The liver function and hematologic parameters are monitored in participants exposed to three ASAQ courses or more.In order to be eligible, patients should satisfy the following inclusion criteria:1. Males and females aged between 12 months and 59 months inclusively. This criterion applies only for the recruitment in the first follow-up. For the subsequent follow-up, children included in the first follow-up are eligible, regardless of their age.2. Body weight of 9 kg and above.P. falciparum or mixed infection containing P. falciparum .3. Microscopically confirmed, mono-infection of 4. Fever (tympanic temperature \u2265 38.0\u00b0C) or history of fever in the previous 24 hours.5. Hemoglobin value \u2265 6.0 g/dl.6. Signed  informed consent by the parents or guardians. Note the first informed consent will be asked at recruitment in the pre-RCT phase and it will cover the first 42 days follow-up. The second informed consent will be asked for at enrolment for the randomized trial and will cover the remaining period of the study.7. Parents\u2019 or guardians\u2019 willingness and ability to comply with the study protocol for the duration of the study.Patients with at least one of the following criteria will be excluded:1. Participation in any other investigational drug study  during the previous 30 days.2. Known hypersensitivity and previous serious adverse events (SAEs) related to the study drugs.3. Severe malaria  or dange4. Presence of intercurrent illness or any condition  which would place the subject at undue risk or interfere with the results of the study, including known glucose-6-phosphate dehydrogenase deficiency.5. Patients who are taking drug which may prolong the QT interval on the electrocardiogram .6. Severe malnutrition .Pneumocistis carinii pneumonia in children born to HIV+ women.7. Ongoing prophylaxis with drugs having antimalarial activity such as cotrimoxazole for the prevention of 1. Have been enrolled in the first phaseP. falciparum infection with clinical symptoms (this could be either mono or mixed infections).2. Recurrent 3. Parents\u2019 or guardians\u2019 willingness and ability to comply with the study protocol for the duration of the study.4. Signed (or thumb-printed whenever parents/guardians are illiterate) (second) informed consent by the parents or guardians. Note, the informed consent will cover the whole period of the study, including additional active follow-ups.Patients with any of the following criteria will not be admitted to the study:1. Known hypersensitivity or serious drug-related adverse event (AE) to the study drugs.2. Severe malaria.3. Danger signs: not able to drink or breast-feed, vomiting (more than twice in 24 hours), recent history of convulsions (more than once in 24 hours), unconscious state, unable to sit or stand.4. Treatment failure within 14 days in the first study phase.5. Body weight below 9 kg.The primary endpoints are: 1) PCR-adjusted efficacy at 28 days: the proportion of children with PCR adequate clinical and parasitological response (ACPR) at day 28; and (2) all early failures before day 7 plus the recurrent parasitemias detected later and classified by genotyping as recrudescence. The treatment failure is defined according to the WHO criteria  as the s PCR-adjuThe ACPR is defined as absence of parasitemia at the end of the follow-up period (day 28), irrespective of tympanic temperature without previously meeting any of the criteria of ETF or LTF. In the PCR-adjusted analyses, patients with late asexual parasite reappearance (with or without fever) will be considered ACPR if the PCR analysis shows a new infection rather than a recrudescence.1. PCR-unadjusted efficacy at 28 days - the proportion of children without (PCR not adjusted) treatment failure; all treatment failures detected during the active follow-up, regardless of genotyping.2. Clinical efficacy at 42 days - all clinical treatment failures detected during the 42 days follow-up for the first-line treatment, with and without PCR adjustment. As no active monitoring of parasitemia after day 3 is planned, this includes ETF and late clinical failure (LCF) following WHO criteria.3. Fever clearance time - defined as the time (in days) from the time of randomization to the first two consecutive measurements on 2 different days of tympanic temperature below 38.0\u00b0C.4. Asexual parasite clearance time - defined as the time (in days) from time of randomization to 2 consecutive negative blood slides (collected at different days). The time to the event will be taken as the time to the first negative slide.5. Gametocytemia  at days 7, 14, 21 and 28 after treatment.6. Hemoglobin changes between day 0 and days 14 and 28.Tertiary efficacy endpoints will be defined in the substudy protocols.Subjects will be monitored throughout the study for possible development of AEs. All AEs will be recorded on the specific form in the case report form (CRF). Vital signs and hematology will be monitored and changes in relevant laboratory parameters will be assessed.PCR plays a key role as it determines if recrudescence will be eliminated after retreatment with the first-line ACT Figure\u00a0. Blood smsp1, msp2 and glurp genes in the P. falciparum genome. PCR amplification of DNA from a single parasite clone results in a single amplification product. For the three genes, each PCR amplification product of a different size is considered to originate from a different clone of P. falciparum and reflects a different genotype. For the samples collected from the same patient at day 0 and on the day of recurrent parasitemia, the length polymorphism of msp1, msp2 and glurp will be determined (that is the number of bands in each PCR reaction and their respective size). Results will be interpreted as follows: recrudescence , at least one identical length polymorphism is found in the sample collected at day 0 and on the day of recurrent parasitemia); new infection ; indeterminate (samples that failed to produce a result due to an inability to amplify DNA at day 0 and/or on the day of recurrent parasitemia).Genotyping of the recurrent infections will be done by characterizing Frozen serum samples are stored on site at -70\u00b0C or at the sponsors premises until transferred for analysis if a research protocol adjacent to this study has been accepted by the relevant Ethics Committee.Assuming the true PCR-corrected efficacy at 28 days of retreatment with the same ACT is 95%, a sample size of 248 patients  is needed to show with 80% power that the lower 95% confidence interval of the efficacy of retreatment with the same ACT should be 90%.Based on the non-inferiority design, recruiting the same number of patients on the alternative ACT  and half on quinine + clindamycin will allow the estimation of the relative risk in efficacy between the treatment groups to be within 5% assuming a similar efficacy of 95% for all three treatment groups .To allow for up to 15% drop-outs or non-evaluable patients, we will recruit 357 patients in each site. The 2:2:1 allocation ratio is chosen to reduce the number of patients randomized to the more demanding quinine + clindamycin treatment regimen. In addition, this allows greater precision for the documentation of the efficacy and safety of ACT regimens as a second line, rather than the standard rescue treatment quinine + clindamycin. However, the inclusion of the quinine + clindamycin arm is necessary as a benchmark for assessing the efficacy of the ACT as a second line and an internal consistency check of the study.In DRC, the most recent study results  on the cIn Uganda, most recent study results on the clinical efficacy of AL in Mbarara in 2005 show a PCR-unadjusted clinical failure rate of 23% after 28 days of follow-up . EmpiricThe recruitment will continue in each country until the required sample size in the RCT phase is reached. If recruitment rates in the RCT are lower than expected, the study may be extended to additional research sites.The study treatment is administered under the direct supervision of a study nurse. If a patient fails to return to the clinic in a timely manner for their daily dose of study drug, they are visited at home and brought to the clinic the same day. If a patient misses any dose of study drugs, they are not excluded from the study. The study nurse records the date and time that study drugs are administered. Study drugs are given to young children as specified by the manufacturer of each product. After drug administration, patients are kept for 60 minutes in the clinic. A dose is repeated in full if vomiting occurs within 30 minutes of administration, and in half if vomiting occurs between 30 and 60 minutes. If vomiting persists beyond two additional doses, the patient is withdrawn from the study and treatment changed.Any antimalarial or antibiotic with antimalarial activity  are disallowed. A list of drugs having antiplasmodium activity was provided to the study clinicians.During the trial, patients can be prescribed drugs such as paracetamol and antibiotics with no known antimalarial activity . The dose, route, time and duration of any concomitant medical treatment is recorded in the CRF.Parents or guardians are discouraged from obtaining drugs from any other source such as private pharmacies, markets or clinics. Parents/guardians are encouraged to bring their children to the study clinic if they are unwell or if they are worried about their health.All patients who develop severe malaria during follow-up are treated following National Guidelines. Patients randomized to ACT and experiencing treatment failure before 14 days are treated with quinine 10 mg/kg orally three times a day in combination with clindamycin 10 mg/kg twice daily for at least 5 days. Patients who fail with quinine + clindamycin therapy are treated with the first-line therapy (ASAQ or AL).Patients will be excluded from further assessment if there is withdrawal of informed consent. SAEs related to the study drug are also a reason for withdrawal from the study. Intake of disallowed drugs leads to withdrawal of the patient from active follow-up. Participants also may be withdrawn if the study sponsor, government or regulatory authorities, or site Ethics Committee terminate the study prior to its planned end date. Every reasonable effort will be made to complete a final evaluation of participants who terminate from the study prior to the planned termination time period and study staff will record the reason(s) for all withdrawals from the study in participants\u2019 study records. At the time of withdrawal, all study procedures outlined for the end of study visit are completed. The primary reason for a patient\u2019s withdrawal from the study is to be recorded in the source documents.A protocol violation occurs when an event happens that does not allow for accurate interpretation of response to treatment. Protocol violations will be defined in the statistical analysis plan.Safety and tolerability of the treatments will be evaluated by recording AEs and grading laboratory, and vital sign evaluations.At each visit, the investigator will ascertain the occurrence of any AE since the last visit. Any event must be recorded on the CRF.An AE is any untoward medical occurrence in a patient or clinical investigation subject administered a pharmaceutical product and which does not necessarily have a causal relationship with this treatment. An AE can therefore be any unfavorable and unintended sign , symptom or disease temporally associated with the use of a medicinal product, whether or not considered related to the medicinal product.Examples of an AE include:1. Significant or unexpected worsening or exacerbation of the condition under study.2. Exacerbation of a chronic or intermittent pre-existing condition including either an increase in frequency and/or intensity of the condition.3. New conditions detected or diagnosed after investigational product administration even though it may have been present prior to the start of the study.4. Signs, symptoms, or the clinical sequelae of a suspected interaction.5. Signs, symptoms, or the clinical sequelae of a suspected overdose of either investigational product or a concurrent medication (overdose as such should not be reported as an AE/SAE).6. Significant failure of expected pharmacological or biological action.Examples of an AE do not include:1. Situations where an untoward medical occurrence did not occur .2. Anticipated day-to-day fluctuations of pre-existing chronic disease(s) or condition(s) present or detected at the start of the study that do not worsen.The severity of a clinical AE is to be scored according to the following scale:1. Mild: awareness of sign or symptom, but easily tolerated2. Moderate: discomfort enough to cause interference with usual activity3. Severe: incapacitating with inability to work or perform usual activity4. Life-threatening: patient at risk of death at the time of the eventThe investigator is obliged to assess the relationship between the investigational product and the occurrence of each AE/SAE. The investigator will use clinical judgment to determine the relationship. Alternative causes, such as natural history of the underlying diseases, concomitant therapy, other risk factors, and the temporal relationship of the event to the investigational product will be considered and investigated. The investigator will also consult the drug information and the Data Safety and Monitoring Board (DSMB) as needed in the determination of their assessment.There may be situations when an SAE has occurred and the investigator has minimal information to include in the initial report to the Trial Management Group (TMG). However, it is very important that the investigator always make an assessment of causality for every event prior to transmission of the SAE report to the TMG. The investigator may change their opinion of causality in light of follow-up information, amending the SAE CRF accordingly.The relationship of an AE to study drug is to be assessed according to the following definitions:1. Definitely unrelated: should be reserved for those events which occur prior to test drug administration  or for those events which cannot be even remotely related to study participation .2. Unlikely: there is no reasonable temporal association between the study drug and the suspected event and the event could have been produced by the subject's clinical state or other modes of therapy administered to the subject.3. Possible: the suspected AE may or may not follow a reasonable temporal sequence from study drug administration but seems to be the type of reaction that cannot be dismissed as unlikely. The event could have been produced or mimicked by the subject's clinical state or by other modes of therapy concomitantly administered to the subject.4. Probable: the suspected AE follows a reasonable temporal sequence from study drug administration, abates upon discontinuation of the drug, and cannot be reasonably explained by the known characteristics of the subject's clinical state.5. Definitely related: should be reserved for those events which have no uncertainty in their relationship to test drug administration. This means that a rechallenge was positive.The outcome of each AE must be assessed according to the following classification:1. Completely recovered: the patient has fully recovered with no observable residual effects.2. Not yet completely recovered: improvement in the patient\u2019s condition has occurred, but the patient still has some residual effects.3. Deterioration: the patient\u2019s overall condition has worsened.4. Permanent damage: the AE has resulted in a permanent impairment.5. Death: the patient died due to the AE.6. Ongoing: the AE has not resolved and remains the same as at onset.7. Unknown: the outcome of the AE is not known because the patient did not return for follow-up (lost to follow-up).An SAE or reaction is any untoward medical occurrence that at any dose fulfils at least one of the following criteria:\u2022Results in death.\u2022Is life-threatening. \u2022Requires hospitalization (other than for drug administration) or prolongation of existing hospitalization.  at the hospital or emergency ward for observation and/or treatment that would not have been appropriate in the physician\u2019s office or outpatient setting. Complications that occur during hospitalization are AEs. If a complication prolongs hospitalization or fulfills any other serious criteria, the event is an SAE. When in doubt as to whether 'hospitalization' occurred or was necessary, the event should be considered an SAE. Hospitalization for elective treatment of a pre-existing condition that did not worsen from baseline is not considered an AE, nor hospitalization for non-medical reasons .)\u2022Results in persistent or significant disability/incapacity.  which may interfere or prevent everyday life functions but do not constitute a substantial disruption.)Medical or scientific judgment should be exercised in deciding whether reporting is appropriate in other situations, such as important medical events that may not be immediately life-threatening or result in death or hospitalization but may jeopardize the subject or may require medical or surgical intervention to prevent one of the other outcomes listed in the above definition. These should also be considered serious. Examples of such events are invasive or malignant cancers, intensive treatment in an emergency room or at home for allergic bronchospasm, blood dyscrasias or convulsions that do not result in hospitalization, or development of drug dependency or drug abuse.All SAEs, whether or not deemed drug-related or expected, must be reported immediately or within 24 hours (one working day), using the Serious Adverse Event Notification Form, by telefax to +32-2652875 or email to international.health@ua.ac.be . The fax should state \u201cUrgent Serious Adverse Event\u201d and the study code on the cover page. All other AEs not fulfilling the criteria of immediate reporting must be recorded on the CRF. This AE information will be collected on a regular basis during the clinical trial.For all AEs identified, an AE form is completed. For each possible AE identified and considered as serious, an SAE event notification form is completed.A severity grading scale, based on toxicity grading scales developed by the WHO and the National Institutes of Health, Division of Microbiology and Infectious Diseases, is used to grade severity of all symptoms, physical examination findings, and hemoglobin results . Any new event, or an event present at baseline that is increasing in severity, will be considered as an AE.For AEs presenting during the study, a patient still experiencing an AE at the end of a follow-up (that is at day 42 or 28) will be managed as follows:\u2013 If the AE has been detected and reported before the last visit and\u25cbit is mild (grade 1), the patient will be managed according to good medical practice and the active follow-up will be stopped. The end date for the AE will be recorded as day 28.\u25cbits grade is more than 1, the patient will be followed until the AE resolves, improves, or stabilizes.\u2013 If the AE is new, the AE will be reported and the patient will be followed until the AE resolves, improves, or stabilizes.For patients classified as clinical treatment failures ), formal study follow-up ends when a patient is classified as a treatment failure (ETF or LTF/LPF), and patients should be treated and managed according to good medical practice. Additional follow-up for AEs in patients classified as treatment failures is not typically indicated, unless the AE is serious, or is felt to be probably or definitely related to the study medications.For patients with SAEs, any patient who experiences an SAE should be followed until it resolves or improves (< grade 1). Although formal study follow-up is typically terminated when a patient is classified as a treatment failure, any patient with severe malaria/danger signs should be followed up to ensure their SAE has resolved/improved.Blood samples are properly labeled with patients' initials, site number, study number, the study day and the date the sample is taken. Hematology assessments are performed locally at sites. All laboratory results are reported in Standard International Units or in conventional units. Blood samples collected on filter paper for PCR genotyping will be analyzed at the Institute of Tropical Medicine, Antwerp, Belgium.All data and observations must be initially documented in the CRF or copied from source documents to the CRF . For this study we will use DATAfax .The Sponsor will share monitoring and quality assurance with the Amsterdam Institute for Global Health and Development based in Uganda. An agreement was signed to this effect. The task of the monitor is to verify the best conduct of the study through frequent contacts by phone and in person with the Principal Investigator and site staff, in accordance with the Standard Operating Procedures and Good Clinical Practice, with the purposes of facilitating the work and attaining the objectives of the study. These visits enable the monitor to maintain current, personal knowledge of the study through review of the records, comparison with source documents, observation and discussion of the conduct of the study with the investigator. Each site will be visited at least five times during the conduct of the trial, including a pre-study visit and a close-out visit. At each visit, the monitor carries out at least 30% source data verification on the first phase of the study (pre-RCT phase) and 30% source data verification on the second phase of the study (RCT and post-RCT phase); however, these percentages might be increased in case of serious or systematic findings during the monitoring visits. The investigator shall maintain source documents for each patient in the study, consisting of case and visit notes  containing demographic and medical information, laboratory data, and the results of any other tests or assessments. The data on the CRF shall be either the source or traceable to the source documents in the patient\u2019s file. The investigator shall keep one copy of the original informed consent form signed by the parent/guardian (the other will remain with the parent/guardian). The investigator shall give the monitor, auditors, inspectors, Institutional Review Board, and Ethics Committee full access to all relevant source documents to confirm their consistency with the CRF entries.The Infectious Diseases Institute in Uganda is in charge of centralized data management. Clinical data, as explained above, are collected and recorded on appropriate paper CRF. Laboratory data, as requested in the protocol, are registered at the laboratory and recorded onto the CRF. All data are then processed from the CRFs into an electronic database. During the conduct of the study, data are verified and reviewed to produce and maintain high-quality data. All unresolved issues are queried and resolved on a regular basis. Data transfer and handling is done with appropriate security measures and with regard to rights, safety and well-being of trial subjects. A report on data management processes will be produced at study completion. The report will include a full field listing and description of the file structure of the electronic data:\u2013 Reference ranges and units for laboratory data\u2013 A brief description of all programs run on the data\u2013 Level of errors found at each stage of checking the data\u2013 General comments on data quality and significant problems encountered with the data\u2013 A detailed list of any unresolved data queries\u2013 A statement of any queries/errors which have not been corrected on the database\u2013 A statement of the storage location of the electronic databaseThe statistician will review the database prior to finalization and report on any problem encountered during the analysis.A detailed analysis plan will be drawn up prior to the analysis. The statistical analysis of the study will be performed by the DRC Principal Investigator (PI), Dr Hypolite Muhindo Mavoko, in consultation with statisticians at the Institute of Tropical Medicine and/or University of Antwerp, who will also review the analysis plans and analysis results.Children in the treatment groups in each country will be described separately with respect to baseline characteristics. The clinical importance of any imbalance will be noted, although statistical tests of significance will not be undertaken.The primary hypothesis of the study is that the PCR-adjusted efficacy at 28 days of retreatment with the same ACT is at least 90%. This hypothesis will be tested by constructing a two-sided 95% confidence interval for the proportion of children without PCR-adjusted rescue treatment failure. If the 95% confidence interval lies entirely above 90% the hypothesis is accepted.The confidence interval will be constructed using Wilson's score method, pooling the data from the two countries .In addition, the relative efficacy of the retreatment with the same ACT compared to treatment with quinine + clindamycin and treatment with another ACT will be assessed using log-binomial models  with fixFor the efficacy analysis, both an intention-to-treat and a per-protocol approach will be adopted, with the intention-to-treat analysis being the primary approach. Rules for inclusion/exclusion of children from the per-protocol population will be specified in advance.Statistical methods for secondary analyses will be described in the data analysis plan.Every effort will be made to minimize the amount of missing data in the trial. Whenever possible, information on the reason for missing data will be obtained. Sensitivity analyses will be undertaken to assess the robustness of the conclusions to the missing data.All non-serious and SAEs will be grouped according to a pre-specified side-effect coding system and tabulated for each treatment group. In contrast to the primary efficacy analysis, actual treatment groups will be assessed.t tests or non-parametric tests. A P value of <0.05 will be considered statistically significant. Estimates of the risk of failure for all primary outcomes will be made using the Kaplan-Meier product limit formula. Patients excluded after enrollment will be censored at the time of their last assessment. Additionally, for genotyping adjusted outcomes, patients with recurrent malaria or recurrent parasitemia due to new infections will be censored. Stratified analysis shall be done for outcomes in patients with recurrent symptomatic  and recurrent asymptomatic (LPF) malaria. The number (and percentage) of patients experiencing each AE will be compared between the treatment groups. No formal statistical testing will be undertaken.The number (and percentage) of patients experiencing any AE, SAE, or drug-related SAE will be compared between treatment groups using Fisher's exact test. Safety will be analyzed using the all-patients-treated approach. Data analysis will be primarily performed using STATA statistical software packages . Descriptive statistics will be used to summarize baseline characteristics of study patients. Efficacy and safety data will be evaluated using a modified intention-to-treat analysis and will only include patients who meet all selection criteria. Categorical variables will be compared between the treatment groups using odds ratio, chi-square tests or Fisher\u2019s exact tests, and continuous variables will be compared using The term \"investigator\" as used in this protocol and on the CRFs refers to the PI or a member of the staff that the investigator designates to perform a certain duty under this protocol. The investigator is ultimately responsible for the conduct of all aspects of the study. For all other relevant investigator responsibilities, see .This protocol has been approved by the Ethical Committee of the University of Antwerp (Reference: UA A11-02), the Uganda National Council for Science and Technology (Reference: HS 1108) and the Ethic Committee of the School of Public Health, University of Kinshasa (Reference: ESP/CE/012B/2012).All interviews are conducted in the native language of the patients by the study personnel. Consent forms in the local language are provided to the parents or guardians for their review. The parents or guardians are asked to sign (or thumb-print whenever the parents/guardians are illiterate) consent to participate in a research study. The informed consent describes the purpose of the study, the procedures to be followed, and the risks and benefits of participation. If a parent or guardian is unable to read or write, an impartial witness takes part in the informed consent discussion and signs the consent form. Parents or guardians are informed that participation in the study is completely voluntary and that they may withdraw their child from the study at any time without any negative consequences.All study documents are provided by the investigators and their appointed staff. None of this material may be disclosed to any party not directly involved in the study without written permission from the investigator. Publication policy will be addressed in a separate agreement.Clinical protocol amendments are alterations to a legal document  and have the same legal status and must pass through the appropriate steps before being implemented. In general, any change must be prior approved by the Independent Ethics Committee to be effective. Administrative changes need only notification to the Independent Ethics Committee without approval. Any subsequent amendments shall be made available with a new protocol with the amendments incorporated and on separate sheets and must pass through the approval process.A no-fault liability insurance has been taken by the Sponsor and covers both trial sites.The relevant study documents are those documents which individually and collectively permit assessment of the conduct of the trial, the quality of the data produced and the compliance with Good Clinical Practice standards and applicable regulatory requirements. The on-site file of the PI contains all the essential documents as listed in the Sponsor\u2019s SOP . A copy of all source data and CRFs must always be kept on site.The PI is responsible for ensuring a secure and appropriate location for their file and any other trial-related documentation present at site, as well as for ensuring that only site staff that are competent and delegated to work for the trial has access to the files. All the relevant study documentation present at all partners involved should be retained for a minimum of 5 years and according to the applicable National Legislation. The Sponsor should always be informed prior to destruction of the files. After completion of the study, the investigator\u2019s file will remain available for internal audits and/or inspections of regulatory authorities for a period of 20 years, unless otherwise requested by national authorities.An independent DSMB with three members  has been created before field activities started. The DSMB was established for the purpose of providing independent advice on the quality of the data produced and the efficacy and safety of the treatments tested, so contributing to safeguarding the interests of the trial participants. More specifically the DSMB:\u2022assesses the quality of data, including completeness;\u2022monitors recruitment figures and loss to follow-up;\u2022monitors compliance with the protocol by participants and investigators;\u2022monitors evidence for treatment differences in the main efficacy and safety outcome measures, thus recommending action when/whether the main trial question has been answered;\u2022monitors evidence for treatment harm ;\u2022recommends whether the trial should continue to recruit or follow-up;\u2022recommends any major changes to the protocol where necessary ;\u2022advises on and/or endorsing any major protocol modifications suggested by the Trial Steering Committee ;\u2022monitors planned sample size assumptions;\u2022suggests additional data analyses;\u2022assesses the impact and relevance of any external evidence provided;\u2022monitors compliance with previous DSMB recommendations;\u2022considers the ethical implications of any recommendations made by the DSMB.In this respect, the DSMB has the possibility to monitor the safety of the treatment on a continuous basis and possibly stop the study if any major safety concern appears. A DSMB charter, where the relevant terms of reference are clearly defined, has been signed by each member of the Committee.The investigators agreed to conduct the present study in full agreement with the principles of the \u201cDeclaration of Helsinki\u201d and subsequent relevant amendments. All research activities are run in accordance with the standards and codes of conduct accepted by the International Conference on Harmonisation guidelines.AL and ASAQ have been successfully used for the treatment of falciparum malaria in Phase III studies and are both widely used to treat uncomplicated malaria in Africa. The combinations are well tolerated. In DRC, quinine + clindamycin is the second-line treatment for malaria. Uganda has not yet adjusted a treatment policy. The treatment is cheap, widely available and generally considered to be effective.Blood is collected for the hematology, biochemistry, blood slides, and later genotyping and other tests that the clinician may request. The amount collected for hemotology and biochemistry is around 3 ml (venous) while for the other tests the amount is a few drops collected by fingerprick. Residual serum will is separated, stored and transported at -70\u00b0C to be assayed by ELISA and fluorescence-activated cell sorting for malarial antibody quantification .Active recruitment.Plasmodium falciparum multidrug resistance; PI: Principal Investigator; RCT: Randomized clinical trial; SAE: Serious adverse event; TMG: Trial Management Group; WHO: World Health Organization.ACPR: Adequate clinical and parasitological response; ACT: Artemisinin-based combination therapy; AE: Adverse event; AL: Artemether-lumefantrine; ASAQ: Artesunate-amodiaquine; CRF: Case report form; DSMB: Data Safety and Monitoring Board; DRC: Democratic Republic of Congo; ELISA: Enzyme-linked immunosorbent assay; ETF: Early treatment failure; LCF: Late clinical failure; LPF: Late parasitological failure; LTF: Late treatment failure; PCR: Polymerase chain reaction; pfmdr: The authors declare that they have no competing interests.HMM and JPVG have written and designed the study protocol. CN, HT, UDA, MPG and PL have commented and delivered expert opinion on the protocol. HMM and CN are site principal investigators, respectively, in DRC and Uganda. JPVG is coordinating the trial on behalf of the sponsor. All authors read and approved the final manuscript.Follow-up chart for the pre-randomized controlled trial phase.Click here for fileFollow-up chart for the randomized controlled trial phase .Click here for file"}
+{"text": "The treatment of falciparum malaria poses unique challenges in settings where malaria transmission intensity is high because recurrent infections are common. These could be new infections, recrudescences, or a combination of the two. Though several African countries continue to use quinine as the second line treatment for patients with recurrent infections, there is little information on its efficacy when used for rescue therapy. Moreover, such practice goes against the World Health Organisation (WHO) recommendation to use combination therapy for uncomplicated malaria.We conducted a nested, randomized, open label, three-arm clinical trial of rescue therapy in children 6\u201359 months old with recurrent malaria infection during 28 days post treatment with artemisinin combination treatment (ACT). Patients were randomly assigned to receive either quinine, artemether-lumefantrine (AL) or dihydroartemisinin-piperaquine (DHAPQ), and actively followed up for 28 days..6 %) were assigned an efficacy outcome and 218 (99.1 %) were assessed for safety. The risk of recurrent infection was significantly higher in patients treated with quinine  and AL , compared to DHAPQ . Recrudescence tended to be lower in the DHAPQ  than in the quinine  or AL  group, though it was not statistically significant. No serious adverse events were reported.Among 220 patients enrolled, 217 (98Recurrent infections observed after the administration of an ACT can be successfully treated with an alternative ACT rather than with quinine.ISRCTN99046537Current Controlled Trials  Most malaria endemic countries have deployed artemisinin combination therapy (ACTs) as first line regimens for the treatment of uncomplicated falciparum malaria This nested study was carried out in Tororo, Uganda, one of the sites participating in a large multi-centre trial (12 sites spread over 7 countries) on the safety and efficacy of four ACTs . Details of the multi-centre trial are published elsewhere The nested study was conducted between December 2007 and April 2009 at Nagongera Health Centre, in Tororo district, Uganda, an area of intense perennial malaria transmission  Patients from the main trial were included in this nested study if they had late treatment failure (recurrent infection with or without fever) between day 4 and day 28 of the two active follow ups. The participant selection criteria in the main study have been reported elsewhere .25 mg/kg and 18 mg/kg per dose of DHA and PQP, respectively, rounded up to the nearest \u00bc tablet. Participants in the quinine arm received a seven day course of quinine sulphate as 10 mg/kg body weight per dose three times daily. Quinine sulphate was provided as 300 mg tablets ; the quality of the drug was certified by the Uganda National Drug Authority. We used a pill cutter to ensure the tablet fractions were as close to the nearest \u00bc tablet as possible. The administration of all doses was directly observed. Patients were kept for 30 minutes after treatment and the dose was re-administered if vomiting occurred. Patients who vomited persistently were given parenteral quinine. All patients were provided with a 3-day supply of paracetamol for the treatment of febrile symptoms. Patients with haemoglobin <10\u00b70 g/dL were treated with ferrous sulphate for 14 days and given mebendazole if they were over one year of age and had not been treated in the previous 6 months. All patients who developed severe/complicated malaria during active follow-up were treated with parenteral quinine. Patients randomised to ACT who failed therapy were treated with quinine 10 mg/kg orally three times a day for 7 days. Patients who failed quinine therapy were treated with artesunate (2 mg/kg once a day) and clindamycin (10 mg/kg twice a day) for 7 days.At enrolment, patient's symptoms were evaluated. Temperature (axillary) and weight were measured and a focused physical examination was performed. A thick and thin blood smear for parasitaemia and a blood sample on filter paper (Whatman 3MM) for later molecular analysis were collected before treatment. Patients were randomly assigned to receive either quinine or one of the two ACTs, (AL or DHAPQ). For the latter, to avoid giving the same ACT administered for the primary episode as rescue treatment, patients having received AL as primary treatment were treated with DHAPQ as rescue treatment and vice versa; patients having received CD+A were randomized to one of the two ACTs . A randoAt enrolment, children's parents or guardians were asked about use of other medications and presence of common symptoms. Axillary temperature and weight were recorded, and a physical examination performed. A blood sample for thick and thin blood smears, hemoglobin assessment and later molecular analysis (on filter paper) was collected by fingerprick. Similarly, at follow up visits scheduled for Days 1, 2, 3, 7, 14, 21 and 28, a standardized history was collected and physical examination performed. At each visit, including unscheduled ones, blood for thick blood smears and later genotyping was collected (by finger prick). Hemoglobin measurement was repeated on day 28 or the day of late clinical failure. Patients were encouraged to attend the clinic at any time if ill. Patients not attending the clinic at the scheduled visits were actively followed up at home.Thick and thin blood smears were stained with 2% Giemsa for 30 minutes. Parasite density was determined by reading the thick blood smear and counting the number of asexual parasites per 200 white blood cells (WBCs), assuming a WBC count of 8,000/\u00b5l. Slides were considered negative if no parasite was detected after reading 100 high-powered fields. Presence of gametocytes was also recorded. Thin blood smears were reviewed for non-falciparum infections. Two microscopists independently read all slides and parasite densities were calculated by averaging the two counts. Readings with discordant results  were re-examined by a third microscopist; the parasite density was calculated by averaging the two closest densities while the final parasite species was determined by the two concordant reads. Hemoglobin measurements were made using a portable spectrophotometer .Molecular genotyping was carried out at the Institute of Tropical Medicine, Antwerp, Belgium, on samples collected from patients with late treatment failure to discriminate between a recrudescent and a new infection. Parasite deoxyribonucleic acid (DNA) was isolated from filter paper blood samples collected at enrolment and on the day of recurrent parasitemia using chelex extraction. Genotyping was done according to international recommendations The objectives of the study were to compare the safety and efficacy of quinine with that of two other ACT, i.e. AL or DHAPQ, when used as rescue treatment for late treatment failure  following a primary episode of uncomplicated malaria.Treatment outcomes were classified according to the WHO guidelines for areas of intense transmission as adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF), and late parasitological failure (LPF) Day 28 treatment outcomes were adjusted to distinguish recrudescent and new infections using molecular genotyping based on GLURP, MSP2 and MSP1 polymorphisms. Recrudescence was defined by a paired sample having at least one identical allele present on each of the three loci. A new infection was classified as infection with all alleles different in at least one locus. When no amplification was observed for either of the paired samples, the outcome was classified as indeterminate.The primary efficacy endpoint was the clinical and parasitological outcome at day 28, unadjusted and adjusted by genotyping. Secondary efficacy endpoints included, fever and parasite clearance, gametocytaemia  at day 7, 14, 21 and 28, haemoglobin changes between Day 0 and Day 28 or the day of treatment failure, and incidence of adverse events.Safety outcomes included risk of adverse events. At each follow-up visit, any new or worsening event and laboratory results were assessed. An adverse event was defined as any untoward medical occurrence, irrespective of its relationship to the study medications . All events were graded by severity  and relationship to study treatment was classified as none, unlikely, possible, probable, or definite using WHO  and National Institute of Health  guidelines. A serious adverse event was defined as any event that resulted in inpatient hospitalization, death, life threatening experience, persistent/significant disability, or specific medical/surgical intervention to prevent serious outcome.The study was designed to test the hypothesis that treatment with AL or DHAPQ would change the risk of recurrent parasitaemia (unadjusted by genotyping) after 28 days of treatment compared to Quinine. The main study included 510 children (170 per arm) with uncomplicated malaria. It was expected that about half of them (230) between days 4 and 28 would need rescue treatment as this is an area of high transmission Plasmodium falciparum infections detected on the basis of genotyping were also censored. Comparisons of treatment efficacy were made using a Cox proportional hazards model. We applied a Bonferroni correction to mitigate the probability of observing a significant difference by chance. With three comparisons, a p-value less than 0.017 (0.05/3) was considered significant.Efficacy and safety data were evaluated using an intention-to-treat analysis, including all patients with falciparum malaria randomized to one of the study treatments. Data were double-entered , verified, and analyzed using Stata version 10\u00b70 . Parasite densities were normalized using logarithmic transformation. Risks of treatment failure were estimated using the Kaplan-Meier product limit formula. Data were censored for patients who did not complete follow-up or were reinfected with non-falciparum species. For the polymerase chain reaction (PCR)-adjusted ACPR, new P. ovale infection and one was lost to follow up (p<0.0001) and AL (p<0.0002) had a significantly higher risk of recurrent infection as compared to DHAPQ . The mean haemoglobin increase between Day 0 and Day 28 was similar in the three groups (The proportion of children with anaemia (haemoglobin of <10\u00b70 g/dL as per the Integrated Management of Childhood Illness guidelines) decreased in all treatment groups between Day 0 and Day 28  were highly efficacious for the treatment of recurrent falciparum malaria. AL has shown good efficacy in repeated treatments of recurrent The cumulative risk of recurrent parasitaemia was higher after the second week of follow up. Similar findings have been observed in areas of intense malaria transmission in Uganda Historically, effective treatment \u2013 a vital component of an effective malaria control strategy- was hampered by the serial development of resistance to the most commonly used drugs. In the early 2000 s, the World Health Organization (WHO) recommended the adoption of ACTs for the treatment of non-complicated malaria. Most African countries now recommend ACTs as the first line regimen for treating uncomplicated malaria. Unfortunately, this effective treatment option is now under threat as recent reports from South East Asia suggest emerging resistance to the artemisinin derivatives This is the first study assessing the appropriate regimen for rescue therapy after ACT treatment in children. It is apparent that in areas of intense transmission, the risk of recurrent infection after quinine treatment, the current recommended second line regimen for Uganda and many other African countries, was unacceptably high compared to DHAPQ, which was highly efficacious and operationally preferable to quinine because of a less intensive dosing schedule. Therefore, in such settings DHAPQ could be an ideal second line regimen. In areas of less intense transmission, where the risk of recurrent infections is low, either DHAPQ or AL could be preferred to quinine because of their shorter dosing schedule and better tolerability, hence probably a better compliance. Unfortunately, more than half of the national malaria control programmes in sub-Saharan Africa still recommend monotherapy with oral quinine as second line treatment. Further, in routine practice the use of oral quinine as a first line treatment seems to be widespread This study was conducted in a high transmission area and we believe our findings can be generalisable to other settings in Africa with similar malaria transmission intensityThe desired sample size of 260 on which power calculations were based could not be attained, affecting the power of the study. In addition, the number of patients in the AL group was disproportionately small (n\u200a=\u200a35), presumably because failure within 28 days of DHAPQ in the main study was rare.Randomization was dependent on previous treatment, as a result different groups had different randomization processes. All doses of the study regimens were given under direct supervision hence patients who received quinine had a longer interaction with the health care system. This could have enhanced the likelihood of detection of recurrent infection in the quinine group when compared with the other groups. However, all parents were encouraged to bring their children to the clinic whenever they were unwell.P.falciparum malaria. Several highly efficacious ACTs are now available and can be used as second-line treatments.Checklist S1Completed CONSORT checklist.(DOC)Click here for additional data file.Protocol S1Study protocol.(DOC)Click here for additional data file.File S1InGenius Gel Documentation and Analysis system Standard Operating Procedure (SOP).(DOC)Click here for additional data file.File S2Makerere Institutional Review Board/Ethics Committee approval document.(PDF)Click here for additional data file.File S3Uganda National Council of Science and Technology approval document.(PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria in northern KwaZulu-Natal since 2001, its efficacy has not been assessed since 2002. The objectives of this study were to quantify the proportion of patients treated for uncomplicated P. falciparum malaria with artemether-lumefantrine who failed treatment after 28 days, and to determine the prevalence of molecular markers associated with artemether-lumefantrine and chloroquine resistance.Recent malaria epidemics in KwaZulu-Natal indicate that effective anti-malarial therapy is essential for malaria control. Although artemether-lumefantrine has been used as first-line treatment for uncomplicated P. falciparum malaria by rapid diagnostic test, had blood taken for malaria blood films and P. falciparum DNA polymerase chain reaction (PCR). Following diagnosis, patients were treated with artemether-lumefantrine (Coartem\u00ae) and invited to return to the health facility after 28 days for repeat blood film and PCR. All PCR P. falciparum positive samples were analysed for molecular markers of lumefantrine and chloroquine resistance.An observational cohort of 49 symptomatic patients, diagnosed with uncomplicated P. falciparum by PCR. At follow-up, 14 were PCR-negative for malaria, one was lost to follow-up and one blood specimen had insufficient blood for a PCR analysis. All 16 with PCR-confirmed malaria carried a single copy of the multi-drug resistant (mdr1) gene, and the wild type asparagine allele mdr1 codon 86 (mdr1 86N). Ten of the 16 samples carried the wild type haplotype (CVMNK) at codons 72-76 of the chloroquine resistance transporter gene (pfcrt); three samples carried the resistant CVIET allele; one carried both the resistant and wild type, and in two samples the allele could not be analysed.Of 49 patients recruited on the basis of a positive rapid diagnostic test, only 16 were confirmed to have mdr1 gene copy number variation detected in this study suggests lumefantrine resistance has yet to emerge in KwaZulu-Natal. In addition, data from this investigation implies the possible re-emergence of chloroquine-sensitive parasites. Results from this study must be viewed with caution, given the extremely small sample size. A larger study is needed to accurately determine therapeutic efficacy of artemether-lumefantrine and resistance marker prevalence. The high proportion of rapid diagnostic test false-positive results requires further investigation.The absence of  Plasmodium falciparum malaria in northern KwaZulu-Natal since it was introduced in response to these drug-resistant epidemics in 2001 [The World Health Organization (WHO) has recommended that drug efficacy be regularly assessed ,2. Failu in 2001 ,5. StudiP. falciparum malaria in KwaZulu-Natal [Chloroquine resistance was first detected in KwaZulu-Natal in 1985 , and hadlu-Natal ,8,9. SP lu-Natal ,5,10.P. falciparum parasites in the region shown to have developed at least 61%  resistance to SP in a clinical efficacy study, rendering the drug ineffective in northern KwaZulu-Natal [P. falciparum malaria, together with the reintroduction of DDT insecticide for indoor residual house spraying in 2001, dramatically reduced malaria incidence in the area [Only in 2000, were the area ,5. It hathe area . MalariaIt has been estimated that in 2000, at the height of the epidemic, the malaria incidence amongst the exposed population in northern KwaZulu-Natal was 5,972 per 100,000 . It shouP. falciparum malaria because of the rapid reduction in parasite load caused by artemisisin or its derivative; the consequent reduced likelihood of resistance emerging to the partner drug; the reduction in gametocyte carriage, and rapid clinical response [P. falciparum malaria [Artemisinin-based combination therapy (ACT) is advocated for the treatment of uncomplicated response . ACT is  malaria , and AL  malaria ,12. Stud malaria ,13,14. SArtemether-lumefantrine is a combination of two drugs, artemether and lumefantrine, manufactured in tablet form as Coartem\u00ae by Novartis. Each tablet contains 20 mg artemether and 120 mg lumefantrine . The twoCoartem\u00ae is taken as a six-dose oral regimen over three days. The dosage depends mainly upon the weight of the patient. The dosage for persons aged 12 years or more, or younger children weighing 35 kg and above, is four tablets as a single dose at the time of initial diagnosis, four tablets after eight hours, and then four tablets twice daily on each of the following two days . It is rIn the 2009, WHO anti-malarial drug efficacy testing guide , inadequAlthough there has been no anecdotal evidence of resistance to AL in KwaZulu-Natal since its implementation, artemisinin resistance, characterised by slow clearing of parasite has been confirmed in South East Asia , and sugP. falciparum recrudescence and re-infection seven days or more after treatment in areas of both low to moderate, and high, transmission [Requiring patients to return a clinic six or seven times in one month for assessment requires considerable resources and the risk of drop-out from the study is high. A single follow-up assessment at 28 days was therefore chosen which required a patient to return only once. PCR is recommended by the WHO to distinguish between smission .P. falciparum multidrug resistant (mdr)1 gene copy number [mdr186N allele [mdr1 gene has been linked with lumefantrine resistance in Southeast Asia [mdr1 gene modulate lumefantrine efficacy [According to the 2002 WHO report , moleculy number , and theN allele ,30. Multast Asia , while mefficacy .Storage of blood samples on filter paper for future testing as new molecular markers become available is recommended .The study was approved by the University of KwaZulu-Natal Biomedical Research Ethics Committee, and by the Health Research Committee of the KwaZulu-Natal Department of Health.P. falciparum malaria in Umkhanyakude Health District, northern KwaZulu-Natal, using the P. falciparum malaria rapid diagnostic test  detection rapid card test manufactured by Premier Medical Corporation Limited, Kachigam, Daman (UT) 396215, India).The study population included symptomatic persons presenting to health facilities, diagnosed with uncomplicated Symptomatic patients aged from five years to 69 years, self presenting to health facilities, diagnosed with uncomplicated malaria in Umkhanyakude Health District between January and May 2012, were invited to participate in the study.Patients with the following danger signs or symptoms of severe malaria: unable to drink; vomiting everything; a convulsion during previous seven days; lethargic or decreased level of consciousness; unable to stand or sit , were exisiZulu detailing the purpose of the study, which was also explained verbally. Patient queries were answered, after which they were invited to provide written consent.At recruitment patients were provided with an information sheet in English and Finger-prick blood spots blotted on to Guthrie 903 filter paper cards , and blood samples were collected from all participants for molecular analysis and malaria microscopy. The patient was then asked to return in four weeks for repeat malaria film microscopy and blood spot collection, with the offer of ZAR50 (US$5.79) in travelling expenses upon return. RDT was not performed at follow-up due to persistence of histidine-rich protein, HRP-2, in patients for as long as 28 days after parasite clearance ,33. BlooThick and thin blood films were prepared according to the National Health Laboratory Service standard operating procedure for processing specimens for malaria parasites . Slides P. falciparum parasites [Parasite DNA was extracted from all blood spots using the QIAamp DNA Mini Kit . The extracted DNA was then subjected qPCR and nested PCR analysis to confirm the presence of arasites ,36.P. falciparum positive by PCR, were subjected to mutational analysis to detect the prevalence of molecular markers linked with resistance to lumefantrine, (mdr1 gene copy number amplification) and chloroquine  gene) [P. falciparum PCR and blood film.All samples confirmed as odon 86,  and codot) gene) . At the A total of 49 patients with a diagnosis of malaria based on a rapid diagnostic test were enrolled in the study. Two patients did not have their age recorded. The age range of the remaining 47 patients was 2 \u2013 69 years; median 15 years, and mean 21.1 years. The largest group comprised those less than 10 years of age  of those with PCR-confirmed malaria were aged less than 10 years. Results for the P. falciparum PCR positive patients are summarized in Table\u2009Only 33% 16/49) patients were confirmed to have 6/49 patiFever (auxiliary temperature \u226537.5\u00b0C) was recorded in 64% (27/42) of all patients who were initially diagnosed with malaria and 77% (11/14) of the PCR malaria confirmed cases, while 57% (16/28) of those PCR negative had a fever. Temperature was not recorded for two malaria PCR positive patients.Six of the 16 PCR malaria confirmed cases reported having travelled to Mozambique within the previous month.P. falciparum positive by microscopy with a parasitaemia of 0.25%. All other 14 blood films were microscopy negative for P. falciparum.Due to liaison difficulties with a local laboratory, blood films were only obtained in 15/49 recruited patients. Of these, four were PCR-confirmed malaria samples, of which one was P. falciparum positive samples collected at enrolment had a single copy of the mdr1 gene and carried the wild type asparagine allele at codon 86 of the mdr1 gene (mdr1 86N) (Table\u2009crt 72-76 genotyped. Ten samples carried the wild chloroquine sensitive haplotype (CVMNK), three the pure mutant haplotype (CVIET) associated with chloroquine treatment failure, while one carried both wild and mutant alleles. Two samples proved inadequate for crt genotyping.All 16 PCR P. falciparum PCR positive at enrolment. Malaria control personnel tracked down and obtained blood specimens for follow-up PCR from a further nine non-returning PCR-confirmed patients, at varying intervals from four weeks following recruitment. Of the initial PCR positive cohort, 14 were found to be negative at follow up, one sample contained insufficient blood for testing and one patient was lost to follow-up.Only 14% (7/49) of the patients returned for follow-up, of which six were P. falciparum malaria in northern KwaZulu-Natal. The study was hampered by a scarcity of diagnosed malaria cases. Of 49 patients enrolled in the study on the basis of a positive P. falciparum rapid diagnostic test, only 16 were subsequently confirmed to have P. falciparum malaria by PCR analysis.The primary aim of this study was to assess the therapeutic efficacy of AL, the current first-line treatment of uncomplicated mdr1 gene copy number associated with lumefantrine resistance in South East Asia was not detected in this study [mdr1 copy number was observed following two years of AL deployment, and supports the hypothesis that mdr1 amplification is rare in Africa [Amplification of is study . This reis study , where nn Africa .mdr1N86Y mutation associated with chloroquine resistance [crt 72-76 wild type haplotype (CVMNK) in the study area suggests AL deployment removed chloroquine drug pressure, allowing chloroquine sensitive parasites to re-emerge as seen in Malawi [The sistance  was compn Malawi  and Mozan Malawi . This reP. falciparum, implying sustained AL efficacy, 11 years after it was initially rolled out in KwaZulu-Natal. On a cautionary note, the high mdr186N allele prevalence is a cause for some concern. It has been suggested that increases in mdr186N prevalence is the first step towards lumefantrine tolerance [mdr186N allele in KwaZulu-Natal. In contrast, the removal of chloroquine drug pressure probably selected for this allele in Mozambique [All follow-up samples were PCR negative for olerance . Sustainzambique . Given tDespite the extremely small sample, some valuable data was produced by this study.In a low malaria-incidence setting, blood spot samples proved to be a good a source for molecular analysis. Collection of the filter paper samples was relatively easy and inexpensive. Once collected and correctly stored the samples were resilient to delays in collection and transport. PCR is capable of detecting malaria parasites at a density of four parasites per microlitre, whereas thick film microscopy is only reliable at a density of 500 parasites per microlitre, meaning PCR is more than 100 times more sensitive than microscopy for the diagnosis of malaria parasitaemia ,42. PCR in vivo sensitivity studies, is much less expensive and time consuming, and is a reasonable method of surveillance for emerging drug resistance [Monitoring molecular markers of drug resistance, while a less rigorous method of assessing drug efficacy than sistance . The WHOsistance . Moleculsistance .The incidence of malaria in the study area remained low during the study period and cases were geographically scattered, presenting to several different clinics and hospitals in Umkhanyakude District. Management and control of the patient records and specimens was difficult. The cooperation of healthcare workers from several health facilities was required and consistency of the enrolment procedure was difficult to achieve. Blood was sent by clinic nurses to the local hospital laboratory who declined to perform blood films on many patients.Of the 49 recruited patients only 16 were PCR confirmed malaria. The high proportion of false positive RDT results was probably mainly due to incorrect use of the test, indicating a lack of familiarity, and the need for more training. It is possible that false positive RDT results have erroneously inflated the notified malaria cases in the district for some time, and is deserving of further investigation.The study aimed to obtain a sample size of 50, which is the minimum recommended by the WHO regardless of rates of failure anticipated, in order to be representative . HoweverDespite the financial incentive offered to recompense for travelling expenses, most patients had to be tracked down by malaria control personnel at varying time intervals after treatment.P. falciparum parasitaemia by day 28, it would not be possible to distinguish between early treatment failure, late clinical failure, and late parasitological failure. The finding of persistence of parasitaemia by day 28 would provide a motivation for a further study following the WHO protocol [Use of a single follow-up visit on day 28, rather than follow-up visits on days 1,2,3,7,14 and 28, as recommended by the WHO ,32, meanDetermining drug efficacy, particularly as malaria transmission approaches zero, is critical, as the last remaining parasites are most likely the most resistant . Since tfalciparum positive by PCR. Preliminary investigations appear to indicate that incorrect use of RDT was the principal reason for the high proportion of false-positive results. Since definitive diagnosis is a fundamental tenant of the elimination agenda, further investigation into the cause of the false-positive RDT results is indicated, and corrective measures put in place to prevent misdiagnosis.Although 49 patients were recruited into the study based on RDT results, only 16 were confirmed mdr1 copy number amplification found in this study. However rigorous regular lumefantrine resistance monitoring is recommended given the high prevalence of the mdr186N allele associated with lumefantrine tolerance and widespread use of AL in southern Africa.Despite the small sample size, all samples were malaria negative at Day 28, or longer, suggesting sustained AL efficacy in KwaZulu-Natal. Support for this is provided by the absence of crt gene: Chloroquine resistance transporter gene; DNA: Deoxyribonucleic acid; HRP: Histidine rich protein; mdr gene: Multidrug resistance gene; pfcrt gene: Plasmodium falciparum chloroquine resistance transporter gene; PCR: Polymerase chain reaction; RDT: Rapid diagnostic test; SP: Sulphadoxine-pyrimethamine; WHO: World Health Organization.ACT: Artemisinin-based combination therapy; AL: Artemether-lumefantrine; The authors declare that they have no competing interests.All authors have reviewed the final draft and agreed to its submission. CHVW initiated the research project, wrote the protocol, co-ordinated the research, collated the data, and composed most of the research report. JR contributed to study design, performed the PCR and molecular analysis, and assisted with editing of the manuscript. ER participated in the study design and supervised Malaria Control Personnel for the follow-up of non-returning patients. EI co-ordinated the recruitment of patients at local clinics, the collection of samples, and was responsible for overseeing clinical care of recruited patients. HR co-ordinated local recruitment of patients, collection of samples, and was responsible for overseeing clinical care of recruited patients. KG reviewed the study protocol, co-ordinated local recruitment of patients, and was responsible for overseeing clinical care of recruited patients. SK assisted with the development of the protocol and the write-up of the report of the research."}
+{"text": "Trypanosoma cruzi infection in vivo.Current chemotherapy for Chagas disease is unsatisfactory due to its limited efficacy, particularly in the chronic phase, with frequent side effects that can lead to treatment discontinuation. Combined therapy is envisioned as an ideal approach since it may improve treatment efficacy whilst decreasing toxicity and the likelihood of resistance development. We evaluated the efficacy of posaconazole in combination with benznidazole on Benznidazole and posaconazole were administered individually or in combination in an experimental acute murine infection model. Using a rapid treatment protocol for 7 days, the combined treatments were more efficacious in reducing parasitemia levels than the drugs given alone, with the effects most evident in combinations of sub-optimal doses of the drugs. Subsequently, the curative action of these drug combinations was investigated, using the same infection model and 25, 50, 75 or 100 mg/kg/day (mpk) of benznidazole in combination with 5, 10 or 20 mpk of posaconazole, given alone or concomitantly for 20 days. The effects of the combination treatments on parasitological cures were higher than the sum of such effects when the drugs were administered separately at the same doses, indicating synergistic activity. Finally, sequential therapy experiments were carried out with benznidazole or posaconazole over a short interval (10 days), followed by the second drug administered for the same period of time. It was found that the sequence of benznidazole (100 mpk) followed by posaconazole (20 mpk) provided cure rates comparable to those obtained with the full (20 days) treatments with either drug alone, and no cure was observed for the short treatments with drugs given alone.Our data demonstrate the importance of investigating the potential beneficial effects of combination treatments with marketed compounds, and showed that combinations of benznidazole with posaconazole have a positive interaction in murine models of Chagas disease. Trypanosoma cruzi acute infections in mice, to support the potential clinical evaluation of such combination therapy for Chagas disease. The curative action of benznidazole/posaconazole combinations was explored in an established acute infection model with the Y strain in which benznidazole and posaconazole treatments induced a 70% and 80% cure rate, respectively, when administered alone at optimal doses. When tested in combination, a 80% to 90% cure rate was detected in mice receiving 25, 50 or 75 mpk of benznidazole, plus 5 or 10 mpk of posaconazole, while treatment with the sub-optimal doses of the drugs given alone induced only 0\u201343% cures, indicating synergistic effects. Finally, sequential short (10 days) treatments with benznidazole (100 mpk) followed by posaconazole (20 mpk) led to an 80% cure rate, comparable with full-length treatments with either drug given alone, while no cures were observed for short treatments with single drugs. Our results demonstrate that it is possible to achieve the same or better therapeutic effect using lower dosages of posaconazole and benznidazole in combination, decreasing treatment costs and potential toxicity.In this study, we investigated the efficacy of posaconazole in combination with benznidazole against  Trypanosoma cruzi kinetoplast DNA extracted from mummified Andean humans demonstrated that Chagas disease was highly prevalent in the Southern Andean coastal area 9,000 years ago T. cruzi from Mexico through Central and South America, with 28 million remaining at risk of infection The use of a DNA probe targeted against Current specific chemotherapy for Chagas disease, based on nitroheterocyclic compounds, is unsatisfactory. Long-term longitudinal studies using benznidazole or nifurtimox have shown that the earlier diagnosis is made and specific treatment is started, the better are the chances of a complete parasitological cure. In congenital Chagas disease, early treatment of newborns with benznidazole or nifurtimox is effective in 66% to 100% of the cases No consistently effective treatment exists for the established chronic forms of the disease, which is currently the most common clinical presentation in both endemic and non-endemic areas. Some studies showed improved clinical and serological evolution of patients treated with benznidazole, as compared with untreated chronic patients, but with variable and limited results in some settings T. cruzi agents. Their mechanism of action is based on the essential T. cruzi requirement for endogenous sterols for survival and proliferation and its inability to use the abundant supply of cholesterol available in mammalian hosts in vitro and in vivo activity of these novel azole derivatives against T. cruzi and the absence of cross-resistance with currently available drugs, the response to drug treatment varies strongly among different parasitic strains http://clinicaltrials.gov/show/NCT01377480 and http://clinicaltrials.gov/show/NCT01489228).Ergosterol biosynthesis inhibitors (EBI) are currently among the most promising anti-T. cruzi effects in vitro and in vivoT. cruzi efficacy of combinations of benznidazole and posaconazole, used in concomitant or sequential therapy, in an experimental murine model of acute Chagas disease to support the potential clinical evaluation of such combination therapies.Combination therapy can be a valuable way to improve treatment efficacy due its capacity to reduce dose levels and toxicity and to prevent the potential development of resistance to anti-infective drugs T. cruzi Y (DTU II) and VL-10 (DTU II) T. cruzi collection at Chagas Disease Laboratory, Federal University of Ouro Preto (UFOP).The original isolates N,N-bis(2-chloroethyl)-1,3,2-oxazaphosphinan-2-amine 2-oxide .The following drugs were commercially purchased from, or provided by their respective pharmaceutical companies: (i) benznidazole \u2013 2-nitro-imidazole-; (ii) posaconazole \u2013 (2)-4-[4-[4-[4-[[(2Rcis)-5--tetrahydro-5-furan-3-yl]methoxy]phenyl]-2,4-dihydro2-[(S)-1-ethyl-2(S)-hydroxypropyl]-3H-1,2,4-triazol-3-one  and (iii) cyclophosphamide  were obtained from the Animal Facility at the Federal University of Ouro Preto, Minas Gerais, Brazil, and maintained in a temperature-controlled room with access to water and food Infected animals were divided into groups of six and received drugs  alone or in different combinations. All compounds were suspended in distilled water using 2% methyl-cellulose (Sigma), and each animal received 0.2 mL of drug suspension daily by gavage for 7 consecutive days. For the treatment with each drug alone, three doses were selected: (i) the dose able to induce a parasitological cure in a longer treatment course, as previously determined by Filardi & Brener The drug combinations consisted of benznidazole plus posaconazole at the following dosages: 25, 50 or 100 mpk of benznidazole in combination with 5, 10 or 20 mpk of posaconazole. Treatment efficacy was assessed based on three parameters: parasite clearance, pre-patent parasitemia interval, and mortality. Parasitemia and mortality were checked every day during and until ten days after treatment and every other day afterwards until 30 days after treatment.T. cruzi  A set of experiments was designed to determine the capacity of benznidazole and posaconazole, given alone or in combination  treatments to induce parasitological cure in mice infected with the Y strain of th day post-inoculation.The concomitant drug combinations consisted of benznidazole plus posaconazole at the following doses: 25, 50, 75 or 100 mpk of benznidazole in combination with 5, 10 or 20 mpk of posaconazole. Results were compared to those reached with benznidazole and posaconazole monotherapies and with infected and non-infected control groups. The drugs were administered for 20 consecutive days upon detection of parasitemia, which occurs on the 4T. cruzi strain A second set of experiments was designed to determine the efficacy of benznidazole plus posaconazole combination therapy to induce survival and parasitological cure in mice infected with VL-10, a benznidazole-resistant The sequential drug therapies consisted of benznidazole (100 mpk) administered for 10 days followed by posaconazole (20 mpk) for another 10 days and the inverse sequence. Results were compared to those reached with benznidazole and posaconazole given alone for 10 and 20 days and with infected and non-infected control groups.et alst (before the CyI) and 6th month after treatment. Animals showing negative results in the two tests were considered cured.The parasitological cure was determined following the methodology standardized by Caldas th day post-treatment to determine the natural reactivation of infection. Animals that did not present reactivation of parasitemia after treatment were submitted to CyI, which consisted of three cycles of 50 mg of cyclophosphamide/kg of body weight, for four consecutive days, with intervals of three days between each cycle. The parasitemia of these animals was evaluated during the CyI cycles, and for 10 days following immunosuppression. Animals did not present reactivation of parasitemia after CyI were bled 180 days after treatment for PCR assay.Fresh blood examination - parasitemia of the animals was evaluated during and up to the 30et al.5\u2032-AAATAATGTACGGG(T/G)GAGATGCATGA-3\u2032 and S36 5\u2032-GGGTTCGATTGGGGTTGGTGT-3\u2032PCR assay. Mice were bled from the orbital venous sinus and 200 \u00b5L of blood were collected 30 and 180 days after the treatment ended. PCR was performed only on samples from animals with negative parasitemia in fresh blood examination. DNA extraction and PCR were performed according to Gomes All procedures and experimental protocols were conducted in accordance with the guidelines issued by the Brazilian College of Animal Experimentation (COBEA) and approved by the Ethics Committee in Animal Research at UFOP (number 2009/16 and 2011/76).To evaluate variations in the levels of parasitemia among animals treated with each drug alone or in combination, the data were converted using a logarithmic transformation and tested using the Analysis of Variance, with the group comparison performed by using Tukey's Multiple Comparison Test. Differences were considered significant when p<0.05.T. cruzi Y strain were assessed. All untreated animals presented high levels of parasitemia, which peaked on the 8th day post-infection, and mortality occurred on average at 15 days post-infection  and compared them with the results obtained with the same doses of each individual drug given alone. At the doses tested, each monotherapy pair produced statistically-indistinguishable parasitemia levels and the combinations being consistently either statistically significant, or near significantly, different from each monotherapy, suggesting a positive drug interaction. This was most evident in combinations of sub-optimal doses of the drugs (benznidazole at 25 mpk with posaconazole 5 mpk and benznidazole at 50 mpk with posaconazole 10 mpk), which reduced the parasitemia levels to values significantly lower than those obtained with the drugs when given alone, even at the optimal doses, and led to 83.3 to 100% survival  for 20 days . As expep<0.001)  benznidazole-resistant strain VL-10 p<0.001) . The resBased on the encouraging results obtained with the drugs used in concomitant combination, we investigated the effects of the drugs when given in sequential treatments in the same experimental model. Interestingly, the efficacy of the treatments was related to the order of drug administration: it was found that treatment with benznidazole at 100 mpk for 10 days followed by posaconazole at 20 mpk for another 10 days induced 80% of parasitological cure, indistinguishable from the effects of the drugs given alone at the same doses over the full 20-days treatment, but when the order of treatment was reversed (posaconazole at 20 mpk for 10 days followed by benznidazole at 100 mpk for another 10 days) the percent of cures dropped to 30% . On the T. cruzi, there is still a need to develop safe, efficient and affordable new specific treatments for Chagas disease, particularly in its chronic stage Although encouraging advances have been made in the control of vectorial and transfusional transmission of Recent studies have shown that compounds such as the sterol C14\u03b1 demethylase inhibitor posaconazole provide a high percentage of parasitological cures in several animal models of acute and chronic Chagas disease in vivo activity of posaconazole and benznidazole, which confer almost complete protection against death to infected mice when used at doses of 20 mpk and 100 mpk, respectively, are in agreement with the activity originally reported by Filardi and Brener T. cruzi activity of these drugs. The first therapeutic scheme used in this study (7 consecutive days) allowed a fast comparative analysis of the anti-T. cruzi activity for each compound, making it amenable for the testing of large numbers of experimental groups, an important strategy for the evaluation of the anti-T. cruzi effects of drugs in combination. The data presented in High levels of When a rigorous evaluation of the curative efficacy of the drugs used alone or in combination was done 30 days after treatment by immunosuppressing the treated animals and monitoring the reactivation of the Y-strain infections , it was T. cruzi infections in humans, such as those of congenital or reactivated Chagas disease patients.Similar conclusions were reached from the analysis of the outcomes of combined treatments against the benznidazole-resistant VL-10 strain . These rT. cruzi activity in vitro and in vivo when used in combination with posaconazole. Ara\u00fajo et al. T. cruzi CYP51 but has no curative activity in experimental or human Chagas disease, has synergistic activity when combined with benznidazole in a murine model of acute disease. More recently, different studies have demonstrated the increased efficacy of different class of compounds when used in combination with nifurtimox Studies using experimental models have demonstrated the efficacy of combination therapy using different pharmacological classes of compounds in Chagas disease. Benaim et al. Taken together, the results of the present study demonstrate that combinations of benznidazole and posaconazole, at sub-optimal doses or using shorter treatments, have equivalent or superior efficacy than the drugs given at their optimal doses and full treatment length in a murine model of acute Chagas disease, indicating the positive interaction of both concomitant and sequential treatments. Since both drugs are commercially available, their use in combination should be considered for evaluation in the treatment of Chagas disease patients, aiming to reduce the doses and/or the length of the treatment, hence reducing its potential toxicity (benznidazole) or cost (posaconazole)."}
+{"text": "A comprehensive appreciation of its effects on haematology Individual-patient data analysis conducted on a database from seven randomized controlled trials conducted in sub-Saharan African comparing AS&AQ to reference treatments in uncomplicated falciparum malaria patients of all ages. Haematologic values  were analysed as both continuous and categorical variables for their occurrence, (severity grade 1-4) and changes during follow-up. Risks and trends were calculated using multivariate logistic random effect models.p = 0.001). Multivariate analysis showed that the risk of anaemia, thrombocytopaenia, and leucopaenia decreased with follow-up time, while neutropaenia increased; the risk of anaemia and thrombocytopaenia increased with higher baseline parasitaemia and parasitological reappearance. White cells total count was not a good surrogate for neutropaenia. No systematic significant difference between treatments was detected. Older patients were at lower risks.4,502 patients (72% < 5 years old), from 13 sites in nine countries with 28-day follow-up were treated with AS&AQ (45%) or a comparator . Pre-treatment leucopaenia (3%) and neutropaenia (6%) were infrequent; anaemia was common (39%). The treatment-emergent adverse events incidence (TEAE = condition not present or less severe pre-treatment) was 11% for neutropaenia, 6% for thrombocytopaenia with AS&AQ and not different from treatment groups; anaemia was higher with AS&AQ (20%) or other forms of ACT (22%) than in non-artemisinin groups (4%, The effects of AS&AQ on haematologic parameters were not different from those of other anti-malarial treatments used in sub-Saharan Africa. This analysis provides the basis for a broader evaluation of haematology following anti-malarial treatment. Continuing monitoring of haematologic safety on larger databases is required. Plasmodium falciparum malaria [Artemisinin-based combinations (ACT)-the treatment of choice for uncomplicated  malaria -are gene malaria . It has  malaria ; other s malaria . Neutrop malaria -8, and a malaria .Artesunate combined with amodiaquine (AS&AQ) is the second most widely used ACT, adopted as first-line treatment in 18 countries. Over time, AS&AQ has been available in a non-fixed formulation , and more recently, as a fixed-dose, WHO-prequalified, co-formulation (ASAQ).Although serious adverse events following ACT seem to be uncommon, very few trials compared the haematologic variations between treatments. Two randomized controlled trials (RCT) conducted in sub-Saharan Africa reported no difference in neutropaenia between artesunate-amodiaquine and artemether-lumefantrine ,11.Safety in general, and laboratory data in particular, are underreported in malaria trials; risk should be assessed comparatively; databases should be large enough and representative of the spectrum composition of patients and conditions. Limited information can be derived from aggregated data, and individual patient data are best suited for such assessment. The widespread use of these anti-malarial combinations calls for a comprehensive synthesis of available individual patient's haematologic data.P. falciparum. Details on these studies are provided elsewhere [Data on age, parasitaemia, haematologic parameters , neutrophil and lymphocyte counts, haemoglobin or haematocrit and platelet counts), treatment and treatment outcome were extracted from a database of randomized controlled trials (RCT) including AS&AQ groups conducted in sub-Saharan Africa with 28-day follow-up . Data were censored when patients dropped out or had recurrent lsewhere .The criteria for selection of the seven RCT were based on the presence of haematologic data n = 4,502) and comparators including monotherapies: amodiaquine (AQ) or artesunate (AS) alone or other artemisinin combination therapy: artemether-lumefantrine (AL), artesunate plus sulphadoxine/pyrimethamine (AS+SP), dihydroartemisinin-piperaquine (DP) or non-artemisinin containing combinations (AQ+SP) 02 and co-17 or fixed-dose co-formulations (ASAQ). The loose AS+AQ was dosed based on body weight, while the co-formulated ASAQ was based on age and weight range.The proportion of patients treated with AS&AQ was 45%  of which 31%  were treated with a fixed-dose ASAQ combination , and 69%  were treated with loose combinations: in Gabon, Kenya and Uganda was AS  and AQ ; in Zanzibar: AS  and AQ ; in Rwanda .The loose combination target dose was AS 12 mg/kg over three days and AQ 30 mg/kg over 3 days, except in Uganda where AQ was given at 25 mg/kg (10 mg/kg on Days 0 and 1 then 5 mg/kg on Day 2). The fixed-dose combinations of ASAQ  were available as two-and three-strength products given by age. The fixed-dose combination was given either once or twice a day. For the two-strength fixed-dose combination ASAQ , the dosing categories were: (i) 0-1 months: 1/2 paediatric; (ii) 2-11 months: 1 paediatric; (iii) 1-6 years: 2 paediatric; (iv) 7-13 years: 1 adult; and (v) \u2265 14 years: 2 adults. For the three-strength ASAQ, age- and weight-based doses were administered once-a-day for three days: one tablet/day for children up to 13 years of age (\u2264 35 kg) or two tablets/day for adolescents aged 14 years and above and adults (\u2265 36 kg). Doses available were: infants (two to 11 months) received AQ 25 mg/AS 67.5 mg; young children (one to 4 years) received AQ 50 mg/AS 135 mg; children (six to 13 years) received one tablet/day of AQ 100 mg/AS 270 mg, and adults (14 years or more) received two tablets (AQ 100 mg/AS 270 mg) per day.Patients treated with other forms of ACT accounted for 27%  of the total: 728 with AL (20 mg artemether/120 mg lumefantrine given according to weight as one (5-14 kg), two (15-24 kg), three (25-34 kg), and four 4 (\u2265 35 kg) tablets given twice daily for three days); 251 with DP (around 2.3 mg/kg/day dihydroartemisinin and 18.4 mg/kg for three days); and 249 with AS+SP  as a single dose).Patients treated with a non-artemisinin containing combination accounted for 12% : AQ+SP (AQ 10 mg/kg/day for three days and SP 25 mg/kg of sulphadoxine and 1.25 mg/kg/of pyrimethamine administered in a co-formulated tablet (SP) as a single dose).AQ only (10% of patients) was given at 10 mg/kg/day for three days; AS only (6% of patients) was given at a total dose of 12 mg/kg over five days.The grading of all paediatric haematological values was derived from international standards -20.9/L; mild/moderate was 3 \u00d7 109/L to 2 \u00d7 109/L, and severe/very severe leucopaenia was < 2 \u00d7 109/L (grade 3: 1.9 \u00d7 109/L to 1 \u00d7 109/L and 4: < 1 \u00d7 109/L).Leucopaenia was defined as white blood cell counts (WCC) < 3 \u00d7 109/L; mild/moderate neutropaenia ranged from 1.19-0.40 \u00d7 109/L (grade 1: 1.19 \u00d7 109/L to 0.75 \u00d7 109/L and 2: 0.74 \u00d7 109/L to 0.40 \u00d7 109/L), and severe/very severe neutropaenia was < 0.40 \u00d7 109/L (grade 3: 0.39 \u00d7 109/L to 0.25 \u00d7 109/L and 4: < 0.25 \u00d7 109/L).Neutropaenia was defined as neutrophil counts < 1.20 \u00d7 10For anaemia the cut off was set at haematocrit < 30% or haemoglobin < 10 g/dL and was categorized as: mild/moderate from 9.9-8.0 g/dL of haemoglobin (grade 1: 9.9 g/dl to 9.0 g/dL and 2: 8.9 g/dL to 8 g/dL), and severe/very severe < 8 g/dL of haemoglobin (grade 3: 8 g/dL to 5 g/dL and 4: < 5 g/dL).9/L; mild/moderate ranged 150-50 \u00d7 109/L (grade 1: 149 \u00d7 109/L to 75 \u00d7 109/L and 2: 74 \u00d7 109/L to 50 \u00d7 109/L), and severe/very severe thrombocytopaenia < 50 \u00d7 109/L (grade 3: 49 \u00d7 109/L to 20 \u00d7 109/L and 4: < 25 \u00d7 109/L).Thrombocytopaenia was defined as platelets count < 150 \u00d7 109/L, for neutropaenia as \u22651.2 \u00d7 109/L, for anaemia as haematocrit becoming \u226530% or haemoglobin \u226510 g/dL, for thrombocytopaenia as platelets \u2265150 \u00d7 109/L.Recovery from an abnormal condition was defined for leucopaenia as WCC becoming \u22653 \u00d7 10A haematological adverse event (AE) was defined as the occurrence of an abnormal value  after treatment start independent of the pre-treatment value. In this analysis, all visits were considered, thus a subject could have multiple AEs.A treatment-emergent AE (TEAE) expresses the worsening of the condition-i.e. any occurrence of an abnormal value at any follow-up visit (days 7 through 28) as compared to baseline in patients who either had a normal condition pre-treatment or an abnormal value that was of lower grade than post-treatment. Drug-event relationship could not be attributed in the present analysis.Haematologic changes were analysed between Days 0-7, 0-14, and 0-28 using t-student paired analysis and presented as relative difference (Day 0 as the reference). In each RCT, the patients' paired differences were compared between treatment groups using the Mann-Whitney rank test. Categorical data were compared using a chi-square test or a Fischer exact test or a Mantel-Haenszel chi-square test stratified by site and the comparison presented by odds ratio (OR), as appropriate.During the 28-day follow-up, the timing of post-treatment haematologic assessments varied across studies .For relative differences, confidence intervals (CI) were calculated using the normal distribution. All CIs were calculated at 95% (95%CI) and comparisons considered significant when All the studies had been approved by the relevant ethics and institution review committees ,10,13-17The main baseline characteristics of the patients enrolled are displayed in Table During follow-up , abnormal values occurred in 2%  for leucopaenia, 8%  for neutropaenia, 40%  for anaemia, 19%  for thrombocytopaenia.Of all the above events, the proportion of those that were severe (grade 3 and 4) was 8% (19/224) for leucopaenia, 3% (18/621) for neutropaenia, 28%  for anaemia, and 15% (100/685) for thrombocytopaenia.p = 0.023 stratified by site, accounted for by a higher frequency with AL in Zanzibar) while no difference was detected for severe neutropaenia, severe anaemia, and severe thrombocytopaenia .Overall, the proportion of patients with severe leucopaenia was lower on AS&AQ than other treatments  (Table p = 0.475) or non-artemisinin treatments (p = 0.642), and for thrombocytopaenia compared to other artemisinin treatments (p = 0.734). In contrast with the incidence of TEAE for anaemia that was higher in AS&AQ or other artemisinin groups compared to non-artemisinin treatments .No difference was observed for neutropaenia between AS&AQ compared to other artemisinin treatments (Two RCTs comparing AS&AQ (n = 464) to AQ alone n = 457) conducted at five sites 7 conduct showed np = 0.882). In each group one patient was leucopaenic on admission and recovered; one in the AQ group became leucopaenic and none in the AS&AQ.i. no significant variation in WCC through Day 28 in both groups, and no difference in variations between AS&AQ and AQ  for AS&AQ and on Day 7 for AQ alone (-21%), but no difference between the two groups; there was one neutropaenic patient in the AQ group who recovered and none in the AS&AQ group. Only one patient from the AS&AQ group became neutropaenic.p = 0.037). The proportions of patients becoming anaemic were not different between AQ  and AS&AQ .iii. a significant increase in haemoglobin in both groups on Days 14 and 28  with no significant difference between the two groups. The proportion of patients recovering from anaemia by Day 28 was higher with AQ  compared to AS+AQ ; 15% (4/27) and 40% (2/5), respectively became thrombocytopaenic (p = 0.291).ii. an increase in platelets in both groups without difference between the two groups; 92% (11/12) in the AS+AQ, and 75% (3/4) of the patients in the AS group recovered (p = 0.031). The proportion of patients recovering from anaemia was slightly higher in AS+AQ  but not significantly different from AS . The risk of becoming anaemic was higher with AS alone  than AS+AQ .iii. a significant transient decrease in haemoglobin by Day 7 with AS+AQ and AS alike; by Day 28, the gain in haemoglobin was greater in patients treated with AS+AQ  than AS alone .In Mali , AS+AQ (Two RCTs comparing AS&AQ n = 1070) to AL (n = 762) in seven sites 70 to AL  showed:vs 0%, 95%CI -5%-4%, p = 0.017); however, Day 28 WCC were not different between the two groups . The proportion of patients recovering from leucopaenia on admission was not different . In AS&AQ < 1% (2/753), and 1% (5/461) of the patients in AL group became leucopaenic (p = 0.092).i. no change in WCC through Day 28 with AL and a significant decrease with AS&AQ on Days 14 and 28 which was significantly greater than AL on Day 28 ; however, Day 28 neutrophil counts were not different . The proportion of patients recovering from pre-treatment neutropaenia was similar in the two groups , as well as the proportion of patients becoming neutropaenic .ii. a significant decrease in neutrophil counts with both drugs at each time point  which was significantly greater with AS&AQ than AL on Days 14 and 28 .p = 0.001), and a smaller increase by Day 14 (p = 0.049), but by Day 28 no significant difference in haemoglobin levels and the proportions of patients recovering from anaemia  or becoming anaemic .iv. by Day 7, a more marked decrease in haemoglobin in the AS&AQ compared to the AL group (v. a significant increase in platelet counts of about 90% was observed in both groups. All patients recovered in both groups with no patients developing thrombocytopaenia.One RCT in Rwanda  comparinp = 0.24). The proportion of patients recovering from leucopaenia was higher in the AS+AQ  than in the DP group . However by Day 14, the prevalence rates of leucopaenic patients were not different between groups .i. WCC decreased significantly in the DP group but not in the AS+AQ group, but the difference between the two groups was statistically non-significant (p = 0.060); at Day 14 the prevalence rates were also not different in the AS+AQ group  compared to the DP group . A significant decrease in neutrophils was observed in both groups. No difference was found between the two groups in the fall in neutrophil counts (p = 0.800) or the proportions of patients recovering (p = 0.720).ii. the prevalence of neutropaenia on admission was 15% (38/252) in the AS+AQ group and 10% (24/251) in the DP group (p = 0.040). The proportion of patients recovering from anaemia was not different between the two groups (p = 0.45), but the risk of becoming anaemic with DP  was higher (but not significantly different) than with AS+AQ .iii. Patients on AS+AQ had a faster haemoglobin recovery  by Day 14 than those treated with DP  was compared to AQ+SP (n = 527) in Rwanda and Uganda ,15 wherep = 0.001, LM test) requiring the use of random effects. All the following analysis used multivariate logistic regression with random intercept on individuals including all potential risk factors. During the drug-free, post-treatment follow-up, it was found that:Large inter-individual differences in haematologic outcomes were detected (p = 0.001), and was lower in older patients  and higher in patients with higher baseline parasitaemia . No difference was detected between AS&AQ and comparator treatments  and treatment outcome  (p = 0.11). The risk of severe (grade 3 and 4) leucopaenia was lower in older patients (p = 0.031) and higher in patients treated from AL  compared to AS&AQ.i. the risk of leucopaenia decreased ; older patients were at lower risks . No difference was detected between AS&AQ and either AQ (p = 0.16), AL (p = 0.56), AQ+SP (p = 0.98), or DP (p = 0.08), or with respect to treatment outcome (p = 0.67) or baseline parasitaemia (p = 0.56). No risk of severe neutropaenia (grade 3 and 4) was detected.ii. the risk of neutropaenia increased ; patients with higher baseline parasitaemia  and those experiencing recurrent parasitaemia  were at higher risk, while older patients were at lower risk . No difference was detected between AS&AQ and either AL (p = 0.67), AS + SP (p = 0.21), DP (p = 0.69), AQ+SP (p = 0.22), AQ (p = 0.48), or AS (p = 0.08). No risk of severe anaemia (grade 3 and 4) was detected.iii. the risk of anaemia decreased ; older patients  were at lower risks compared to younger patients; patients with higher baseline parasitaemia , treated in Uganda with AL  or AQ+SP  were at higher risks for thrombocytopaenia by Day 14. In Mali, the risks of patients treated with AS (p = 0.64) or AS+SP (p = 0.47) were not significantly different from AS&AQ. Patients with a parasitological recurrence were at higher risks for thrombocytopaenia . There were no data for AQ and DP. Similarly, the risk of severe thrombocytopaenia dropped significantly during follow-up (p = 0.025); younger patients (p = 0.001) and patients treated with AS (p = 0.002) were at significant higher risks compared to AS&AQ; no difference was detected for the other treatments.iv. the risk of thrombocytopaenia decreased  therapies. Studies were identified through a systematic review of the literature conducted in 2008 and contacting investigators who may be willing to contribute their data [Safety in general and laboratory data in particular are under-reported in malaria, and limited information can be derived from aggregated data meta-analyses. This study obviates some of these shortcomings by collecting and analysing individual data on a substantial number of patients  from RCTs of loose or fixed-dose artesunate-amodiaquine combinations  eir data . A few aeir data ,22,23 coIn contrast with meta-analysis of aggregated study reports, individual patient data permit analysis of haematologic parameters as both continuous and categorical (e.g. using common toxicity grades) variables, as well as multivariate analyses including covariates such as individuals, study site, age, baseline parasitaemia, treatment outcome, and treatment group. As a result, conclusions can be drawn as to the contributions to haematologic changes (both toxicity and recovery) of either components of the combination (artesunate and amodiaquine) both individually and together.Overall, there appears to be no obvious, specific haematologic risk systematically associated with artesunate-amodiaquine as compared to single-agent and other combination (with or without an artemisinin) therapies.P. falciparum [Knowledge of the haematologic changes occurring during acute malaria and recovery is incomplete, limiting our ability to analyse and understand drug-induced changes. A recent analysis from this group of data in African children under five years of age treated for lciparum  showed tDifferently from the above-cited study, the present one had patients of all ages  and no lymphocyte counts. Six of the seven trials of this analysis are in common with the previous analysis . BurkinaIn the present study including also about one-third of adults, acute malaria (pre-treatment) was associated with a very low risk of leucopaenia (3%) and neutropaenia (6%); instead, anaemia (about 60%) and thrombocytopaenia (about one-third) were common. Also, older patients were at lower risk for leucopaenia, neutropaenia, anaemia, and thrombocytopaenia, contrary to findings in the narrower group of children under five years of age, for whom no age-effect was apparent except for anaemia .Post-treatment, the risk of leucopaenia, anaemia and thrombocytopaenia decreased, while the risk of neutropaenia increased over time. When interpreting this finding, it should be noted that: (i) the risk post-treatment includes the prevalence of all the events occurring throughout the 28-day follow-up period and (ii) the time trends  are minimal in particular for leucopaenia and neutropaenia. The respective adjusted risk (AOR) 95%CIs were 0.94-0.98 and 1.02-1.04.Leucopaenia was infrequent both at baseline and post-treatment. White blood cells total counts without differential counts appear to be uninformative as they will not capture larger variations in neutrophil counts that are partly compensated by opposite variations in lymphocytes . BaseliBaseline neutrophil counts were overall normal , but wiAge appeared to protect against neutropaenia (here like in ), but heAnaemia was frequent before treatment  ,16 and dvs amodiaquine alone and one vs amodiaquine plus SP). No clear indication results from these studies. In two studies [Artemisinin compounds have been shown to induce reticulocytopaenia both in experimental and clinical conditions potentially by suppressing erythroblasts and that malaria itself could protect against reticulocytopaenia . None of studies ,17 the pp = 0.040) and AL (p = 0.013) compared to the AS&AQ, while no difference between groups was detected in Mali.Only two studies recorded platelets in Mali  and in UDeviation of laboratory values from normality is graded according to widely accepted severity criteria ; howeverThe size of this database (relatively large), the spectrum representation (locations and ages) and the analyses done (multivariate) are all positive features and advantages over single-site papers or meta-analyses of aggregate data. However, a note of caution is needed as to how to interpret these results. The absence of a signal does not mean that there is no risk, or certainty about no excess risk (over other treatments). Rare events will require a very large sample size that was not available here. The sample size is further reduced for individual variables ; also, all variables were not assessed uniformly at the same time. There is also the issue of special risk groups, such as HIV-coinfected subjects  and pACT: Artemisinin Combination Therapy; AE: adverse event; AL: Artemether-Lumefantrine; AQ: amodiaquine; AS: artesunate; DP: Dihydroartemisinin-piperaquine; IPD: Individual Patient Data; LM: Lagrange multiplier test; Max: maximum; Min: minimum; RCT: Randomized Controlled Trial; SD: standard deviation; SP: Sulphadoxine-Pyrimethamine; TEAE: treatment emergent adverse event; WHO: World Health Organization.The authors declare that they have no competing interests.JZ and PO designed the analysis, interpreted the data and prepared the manuscript. All authors read and approved the final manuscript.P. Olliaro is a staff member of the WHO; the authors alone are responsible for the views expressed in this publication and they do not necessarily represent the decisions, policy or views of the WHO.vs comparator groupsHaematologic paired analysis, RCTs, AS&AQ .Click here for file"}
+{"text": "Plasmodium falciparum malaria among adolescents and adults in Calabar, South-east Nigeria.The use of anti-malarial drug combinations with artemisinin, or with one of its derivatives, is now widely recommended to overcome drug resistance in falciparum malaria. Fixed-dose combination of artemisinin and naphthoquine is a new generation artemisinin combination therapy (ACT) offered as a single dose therapy. The aim of the study was to assess the therapeutic efficacy, safety and tolerability of three dosage schedules of fixed-dose combination of artemisinin (125 mg) and naphthoquine (50 mg) for treating uncomplicated P. falciparum malaria were enrolled and randomly assigned to three dosage schedules: (A) 700 mg (four tablets) single dose; (B) 700 mg 12-hourly x two doses; and (C) 1,400 mg (eight tablets) single dose. Patients were observed for 28 days, with clinical, parasitological, and haematological assessments.A total of 121 patients aged \u226515 years with uncomplicated A total of 108 patients completed the study. The overall 28-day cure rate was 88.9%. Day 28-cure rates of the three dosage schedules were 85.3%, 93.1% and 88.9% for Group A, B and C respectively. Adverse events were few and mild, the commonest being weakness and headache; there was no serious adverse event.Concerns for emergence of parasite resistance due to the use of artemisinin-naphthoquine as single dose regimen is likely to compromise the usefulness of this potentially important combination treatment. A robust multi-centre trial is recommended to evaluate a three-day regimen with potentials to achieve high cure rates while minimizing the risk of emergence of resistant parasite strains. Plasmodium falciparum resistance to chloroquine[Malaria remains a major public health problem in Nigeria accounting for as much as 30% childhood deaths and 11% maternal mortality in Nigeria[oroquine and sulporoquine.Nigeria adopted artemisinin based combination therapy (ACT) as the treatment of choice for uncomplicated plasmodium falciparum malaria. The goal of combination therapy is to increase effectiveness of available antimalarial drugs and delay the emergence and spread of drug resistance,6. The sA fixed dose combination of artemisinin-naphthoquine (Arco\u2122) developed by Kunming Pharmaceuticals Corporation (KPC), China was tested in this study. The drug is administered orally as a single dose treatment for uncomplicated falciparum malaria. In addition to the high cure rate of 97% recorded in two earlier reports by Chinese investigators, this trThe aim of this study was to assess the therapeutic efficacy, safety and tolerability of fixed dose combination of artemisinin-naphthoquine for the treatment of uncomplicated falciparum malaria among adolescents and adults in Nigeria. Separate trials in younger children (< 5 years and 5\u201314 years) are planned for the same study area.th March 2006 to 19th August 2006 in a Primary Health Centre (PHC) in Ikot Ansa, Calabar of Cross River State. The health centre is located in a semi-urban community with holoendemic P. falciparum transmission. The level of Plasmodium falciparum resistance to chloroquine and sulphadoxine-pyrimethamine is high[The study was conducted from 6 is high.P. falciparum malaria with a diagnosis confirmed by a positive blood smear with asexual forms of P. falciparum; (2) residence in the study area throughout duration of follow up; (3) age \u226515 years; (4) history of fever within the past 24 hours or axillary temperature \u226537.5\u00b0C; (5) parasite density of 500\u2013200,000 asexual parasites/\u03bcl; (6) absence of signs of severe malaria as defined by World Health Organization\u2019s criteria[The study was an open-label, non-comparative therapeutic efficacy clinical trial carried out following the standard WHO protocol for in vivo efficacy. Patientcriteria) or any d) of 10% point, the calculated sample size for each arm was 35. An adjustment of 20% for follow-up losses and withdrawals was made as recommended in the WHO protocol for studies with follow-up periods > 14 days. The adjusted sample size per arm was 42 participants. The aim was to enrol a minimum of 42 patients per arm or retain at least 35 at completion of follow-up.The sample size was determined based on the WHO standard protocol for non-comparative therapeutic efficacy studies. AssuminThe study was approved by the national regulatory authority in Nigeria. Ethical approval was received from the Ethics Committee of University of Calabar Teaching Hospital prior to onset of study. A written informed consent was obtained from each patient  prior to enrolment.Patients were randomly assigned to one of three dosage scheduled of fixed-dose combination of artemisinin-naphthoquine as follows: Group A received four tablets as a single dose; Group B received four tablets twice at 12 hours interval ; Group C received eight tablets as a single dose. The allocation sequence was generated by simple balloting and concealed in serially numbered, opaque, sealed envelopes. This was opened by the study nurse at the point of drug administration following completion of consent and all clinical procedures. All the treatments were given in the health facility under direct observation of the study personnel. Drugs were given with water and after patient had a meal. Tepid sponging, exposure and administration of paracetamol were used to reduce high fever. Patients were observed for about 30 minutes after administration of drug and treatment was repeated once if patient vomited within 30 minutes of drug administration. Patients that vomited a second time were excluded from the trial and treated as severe illness. All patients were treated as outpatient.Clinical examination including axillary temperature measurement, body weight and height were recorded on day of enrolment (day 0). During each follow-up visit, clinical examination including temperature was performed. Parasitological examination (thin and thick blood films) were performed on enrolment using Giemsa and performed on each follow up visits. Parasite density was enumerated using thick film as described by Shute. Blood fFollow-up visits were scheduled on days 1, 2, 3, 7, 14, 21 and 28. Packed cell volume (PCV) was estimated on day zero and repeated on days 14 and 28 for all patients using blood sample collected in a heparinized capillary tube, spun for 5 minutes at 5,000 rpm with micro-haematocrit centrifuge and read with micro-haematocrit reader. Participants that did not return on schedule for follow up were visited at home the same day by study field staff.Outcomes were defined based on the WHO guidelines for assessing therapeutic efficacy. The 28-Clinical and laboratory procedures were subjected to internal quality control. There was no external monitor for this study. However, the trial was audited by officials from the regulatory body in Nigeria; the National Agency for Food and Drug Administration and Control (NAFDAC). Standard operating procedures based on the principles of good clinical practice were prepared for measurement of height, weight, temperature, collection of blood samples and calculation of parasite counts. Malaria parasites were counted by two microscopists independently and the average count was taken as the final count. Where there was a marked difference in count (>25%) between study team microscopists, such slides were sent to an external field microscopist to confirm the count. The external microscopist also randomly selected and reviewed 10% of all the slides read by study team microscopists. Haematology and biochemical tests were conducted in the laboratories of the University of Calabar Teaching Hospital with quality control measures in place.Data generated were recorded in a log book, and individual patients\u2019 case report files were double \u2013 entered and analysed first in EPI Info Version 6 and later using SPSS (version 11.0.1). Arithmetic and geometric means as well as standard deviation were calculated for baseline characteristics. One-way ANOVA was used to test for statistical significance and p-values less than 0.05 were considered to be statistically significant. Both intention-to-treat and per protocol analyses were presented for the main outcomes.A total of 992 persons were screened; 348 35.1%) were males and 644 (64.9%) females. One hundred and twenty-one (121) patients with uncomplicated malaria who fulfilled the inclusion criteria were enrolled: 36 to Group A; 35 to Group B and 50 to Group C . TableThe combination was well tolerated by the study participants. Biochemistry and haematological parameters did not deviate significantly from normal values (data not shown).This study showed an overall 28-day cure rate of 79.3% for the fixed dose combination of artemisinin-naphthoquine. It was not possible to perform polymerase chain reaction (PCR) to confirm the genetic characteristics of the species reported as treatment failures in this study to determine whether these were due to recrudescence or actually new infections. This cure rate is lower than those reported in other efficacy studies on the same drug, most of which were between 97% and 98%,16. TreaSingle dose artemisinin-naphthoquine has also been shown to have an efficacy rate comparable to that of artemether-lumefantrine. This stThe recommended artemisinin-naphthoquine dose for those aged 15 years and above is a single blister  comprising eight tablets (125/50 mg). The exploratory dose-finding assessment performed in this study showed that the treatment arm that received the total recommended dose in two split doses of four tablets (500/200 mg) per dose had a higher 28-day cure rate than those that received the recommended single dose regimen without any difference in tolerability. This observation is limited by the lack of pharmacokinetics data to adequately investigate bioequivalence.P. falciparum malaria and, therefore, is able to sustain and complete elimination long after the artemisinin component has waned to below therapeutic levels[P. falciparum. Recent evidence from clinical evaluation of a large cohort of children treated with artemether-lumefantrine and artesunate-amodiaquine showed that these artemisinin-based combination treatments are still highly efficacious in Nigeria[P falciparum resistant to the artemisinin compound in south-east Asia calls for greater attention to rationale use of ACT regimens[The unique advantage of artemisinin-naphthoquine is its simple dosage schedule, a characteristic which is more likely to be associated with better patient adherence to treatment than ACT regimens that required multiple dosing schedules,20. The c levels. While tc levels,21. This Nigeria. Howeverregimens.P. falciparum resistance to the component drugs of this potentially beneficial combination regimen (arteminisin-naphthoquine), it would be advisable to use a once daily regimen for a period long enough to ensure complete elimination of all susceptible parasites in each treatment course. In this study, the use of a single-dose regimen of half the recommended dose was explored and revealed a fairly good 28-day cure rate but lower than the rate obtained with the recommended dose. It is likely that administration of single daily dose of 500/200 mg for three days will achieve high cure rates with much lower risk of recrudescence than would be the case with the current practice of single dose of 1,000/400 mg give only for one day. A well designed trial to test this dosage option would be of immense public health importance to avert the likely deterioration of this combination regimen from widespread use as single dose regimen.In order to prevent or delay the emergence and spread of One of the key elements in any drug development and evaluation is the issue of safety of the population for which the drug is intended. Artemisinin-naphthoquine was well tolerated and no serious adverse event was recorded in the study. The commonest adverse events were weakness and headache which were reported more in the group that took the recommended dosage. Since both symptoms could also be caused by malaria, it is not certain whether these symptoms were due to the drug alone or caused by the illness itself. This is in keeping with findings from earlier studies in China that showed that this drug is safe and well tolerated,10,16. LP. falciparum is recommended. A well-designed, adequately powered multicentre randomized controlled trial that meets all GCP requirements will be required to determine how this potentially important combination can be put to the best public health use.The artemisinin-naphthoquine combination is a potentially important ACT, which deserves further clinical evaluation. While the single dose regimen currently recommended by the developers has a promise for enhanced patient adherence to treatment, it raises serious concerns about the likely emergence of resistant parasite strains to the component drugs. The use of the formulations in reduced strength over a three-day period to minimize the risk of emergence of resistant mutants of The authors declare that they have no competing interests.ENE and MMM contributed to design of the study; FO, CO and MMM coordinated the study, performed data analysis and drafted the paper; EE, FO and EE performed clinical assessment; CO, OO and VA performed laboratory tests, AA supervised laboratory team, EBE contributed to data analysis and drafting of paper. All authors read and approved the final manuscript."}
+{"text": "A total of 197 subjects were recruited into the study and administered Malartin for 3 days and SP as a single dose on day 0. Only 174 of the subjects were successfully followed up on days 3, 7, and 14. The overall success rate of the drug combination was 92.53%. Parasite density decreased during the follow-up period in different age groups, sexes, and social classes. The prevalence of anaemia decreased from 22.99% at enrolment to 9.77% on day 14, and the difference was significant (P < 0.05) on all days of followup. The drug combination did not give rise to any serious side effects.Artemisinin derivatives are now the most potent and rapidly acting antimalarials. The aim of this study was to assess the  Malaria continues to be a debilitating disease with public health consequences on an ailing population that is already poor . Many maPlasmodium falciparum is the main infecting species and its infections can be very fatal unless promptly treated. It is responsible for the majority of deaths due to malaria and the situation has been worsened by the development of resistance by this parasite to most antimalarial drugs [P. falciparum in endemic areas, since resistance may be delayed or prevented by using combinations of drugs with independent modes of action and different biochemical targets in the parasite. Artemisinin or one of its derivatives is usually used with one or more other longer acting partner drug(s) in a treatment regimen that can protect against recrudescent infections. The rapid onset of action of artemisinins reduces the biomass of the parasite within a short period of time and the remaining parasites are eliminated by the longer-acting partner drug [al drugs . Resistaal drugs . Drug real drugs . Hence ial drugs  now reconer drug . CamerooP. falciparum in Dibanda, Southwest Cameroon.The artesunate, Malartin, (Kakwa Biofarm) was recently introduced into the Cameroon market and its efficacy alone and in combination with amodiaquine has been reported elsewhere . Since iP. falciparum transmission in the area.This work was carried out in Dibanda village, southwest Cameroon. The centre for recruiting and treating patients was the Cameroon Christian Medical Foundation (CAMEF). Dibanda has a population of about 3500 people and is located on the eastern slope of Mount Cameroon, where the dense equatorial forest has been cleared. Weather records from the Cameroon Development Corporation indicate a mean relative humidity of 83.1%, temperature range of 18\u00b0C to 35\u00b0C and an annual rainfall of 4000\u2009mm . There iP. falciparum malaria, parasitaemia \u22651000 asexual parasites/\u03bcL blood, absence of concomitant illness, and written informed consent by the patient or legal guardian in the case of minors. Those excluded from the study were all participants who took antimalarial drugs in the previous 48 hours, pregnant and lactating women, those with severe malaria or any danger signs as defined by the WHO or mixed infections of plasmodia, and those who refused to participate in the study. Voluntary withdrawal, development of severe manifestations following therapy and overt toxicity (hypersensitivity) to the drugs were considered reasons enough for withdrawing patients from the study. They were then treated with oral quinine (25\u2009mg body weight per day for 5 days). An ethical clearance for this work was obtained from the Ethical Review Committee of the Delegation of Public Health, South West Region, Cameroon. Overall, 576 patients were screened, amongst whom 197 were eligible for inclusion in the study.The study was carried out between December 2007 and August, 2008. Patients who presented with signs and symptoms of malaria at CAMEF were referred to members of the research team after consultation. The reason for the study was explained to them before asking for their written informed consent. Participants were eligible for inclusion into the study if they were \u22651 year, diagnosed with monoinfections of uncomplicated \u03bcL blood [P. falciparum [Before blood collection, demographic data such as age, gender, social class , weight, prior use of antimalarial drugs, and clinical manifestations were recorded. Blood samples were obtained through a finger prick for the preparation of thick and thin blood films and determination of packed cell volume (PCV). Blood films were Giemsa-stained according to the method of Cheesbrough  and obse\u03bcL blood . This walciparum . In ordeg for 5 minutes and PCV values were read using a PCV reader. Patients were considered to be anaemic if PCV values were <31%.Blood-filled capillary tubes were spun at 12,000\u2009Each patient was treated with 4\u2009mg/Kg body weight/day of malartin on days 0, 1, and 2 and 25\u2009mg/kg body weight of sulphadoxine and 1.25\u2009mg/Kg body weight of pyrimethamine in a single dose [The blood of the patients was collected on all days of followup to determine their parasite density and PCV levels. Study participants were considered to have attained a study endpoint whenever there was onset of symptoms requiring alternative drugs and/or hospitalization, receipt of another antimalarial from outside the study team, or voluntary withdrawal. Participants still parasitaemic on day 14 were referred for alternative treatment.Any signs and symptoms that first occurred or became more severe after treatment were considered to be the adverse effects of treatment and were recorded during the course of followup.Treatment outcome was classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), or adequate clinical and parasitological response (ACPR), according to the WHO  criteria(i) To be categorized as an ETF, a patient had to show danger signs or severe malaria on days 1, 2, or 3, in the presence of parasitaemia; a day 2 parasitaemia that was greater than the day 0 (irrespective of axillary temperature); a parasitaemia on day 3, with fever (an axillary temperature of at least 37.5\u00b0C); or a day 3 parasitaemia that was at least 25% of the day 0.(ii) To qualify as LCF, the patient had to show danger signs or severe malaria, in the presence of parasitaemia, on any day after day 4 and before day 14, without previously meeting any of the ETF criteria; or fever in the presence of parasitaemia on any day between day 4 and day 14, without previously meeting any of the ETF criteria.(iii) Any patient found parasitaemic between day 4 and day 14, but with a temperature of <37.5%, without previously meeting any of the criteria of ETF or LCF, was considered as an LPF.(iv) All other patients who were aparasitaemic on day 14, irrespective of axillary temperature, were considered ACPR.t-test or analysis of variance (ANOVA) was used to test the differences between the treatment means. The chi square test or Fisher's exact \u03c72 was used for comparisons of the cure rates. The level of significance was set at P < 0.05.The data were analyzed using Microsoft Excel 2002 and SPSS version 10. Students' Overall, 576 people aged 1 to 78 years were screened and a total of 197 participants satisfied the enrolment criteria and were included in the study. Twenty-three patients were lost in the course of followup. A total of 174 participants were successfully followed up . The median age was 11.4 . OverallThe adequate clinical and parasitological response (ACPR) rate on day 14 was 92.53% (161/174). The early treatment failure (ETF) rate was 0.60% (01/174), while the late clinical failure (LCF) and late parasitological failure (LPF) rates were 03.45% (06/174) and 03.45% (06/174), respectively. Overall, the clinical treatment failure rate was 07.50% .\u03c72 = 4.826, P = 0.03). GMPD decreased steadily in all age groups from day 0 to day 14 following treatment . There was no significant difference in GMPD across the different social classes during followup after treatment with Malartin and SP .At enrolment, 22.99% (40/174) of the participants were anaemic and had a higher GMPD when compared with the nonanaemic subjects . This GMP < 0.05) on all days of followup .77.01% (134/174) of participants were not anaemic at enrolment. Anaemia was generally more prevalent in children (<15 years) than in adults (\u226515 years), and the difference was significant (followup . The proP > 0.05). Overall, anaemia was more prevalent in the middle class, and generally decreased gradually in all the different social classes after treatment with Malartin and Fansidar. On day 14, the highest prevalence (10.30%) was recorded in the poor class. There was no significant difference in the prevalence of anaemia across the different social classes during followup after treatment with Malartin and Fansidar .It was observed that after administering malartin and SP to the subjects, their body temperatures reduced rapidly and fever clearance on day 3 was 97.00%. Variation of body temperature during followup was done by comparing the mean temperature on day 0 with that of the rest of the followup days (data not shown). The mean body temperature on day 0 (37.91 \u00b1 0.93) was significantly higher than that of day 14 (37 \u00b1 0.08) , fever (22.50%), and dizziness (17.50%) were the most prevalent side effects. These side effects were generally minor and self-limiting, and most of them disappeared by day 7.P. falciparum has been reported to have a complex genome, and is responsible for over 90% of the malaria cases transmitted in southwest Cameroon [This study addresses drug-resistant malaria, which has become one of the most important problems in malaria control in recent years especially as it has been extremely difficult to develop an effective antimalarial vaccine and presently there is no single method of control that can completely eliminate malaria. According to WHO , the figCameroon , 10. IncCameroon . An optiCameroon .P. falciparum infection. The observed day 14 ACPR of 92.53% is similar to results of previous studies in Cameroon, where ACPR with a combination of artesunate and Fansidar on day 14 was 93.00% [In this study, the ACPR at day 14 was 92.53% following treatment with Malartin and SP. This study therefore demonstrated that a combination of Malartin and SP is good for the treatment of uncomplicated s 93.00% ; 93.80% s 93.00%  and 91.7s 93.00% .dhfr) to SP. SP has been widely used in Cameroon as a monotherapy, and is still available in the saturated pool of antimalarials in the country. A study by Mbacham et al. [P. falciparum, and is known to be rapidly schizonticidal, and by suppressing cytokines and the production of tumour necrosis factor (TNF) helps shorten fever-clearance times [A clinical treatment failure rate of 07.50% was recorded in this study and this may be attributed to the presence of resistant genes , drug interaction and poor or erratic absorption. Perhaps such factors might have increased the likelihood of parasites being exposed to suboptimal drug levels .P. falciparum. This phenomenon suggests that there is a synergy between drug treatment and acquired immunity. In high-transmission areas especially in Africa, the generally accepted objective of malaria treatment is not so much the clearance of parasitaemia but the resolution of clinical symptoms and acute febrile illness as measured by the adequate clinical response and early and late treatment failures [The progressive decrease in GMPD in our study indicated that the drugs were effective in reducing the parasite load; there were still some low levels of parasitaemia on day 14 in a few patients. Perhaps this could be due to reinfection, since Dibanda is a highly endemic area for malaria. According to Enevold et al. , in malaP. falciparum was the major cause of anaemia in the patients, and it is therefore important to monitor the development of drug resistance in this area in order to maintain the therapeutic lifespan of effective drugs for as long as possible. Repeated episodes of malaria due to reinfection or failure to clear parasites as a result of antimalarial drug resistance may result in life-threatening anaemia [During followup there was also an amelioration in PCV levels. This agrees with other studies in Cameroon, Sudan, Senegal, and Kenya where PCV levels increased following treatment with ACTs , 16, 21. anaemia .In our study, 78.20% of the subjects belonged to the poor class. Anaemia was found to be more prevalent in the middle class patients, and interestingly, there was no significant difference in GMPD across the different social classes during followup. This could be due to our relatively small sample size of the rich and middle classes. This finding could be investigated further with a much larger sample size of the middle and rich classes. It is also worth noting that a possible selection bias might have occurred in the study especially at enrolment of the patients. This is evident as the study subjects were not very representative of the patient population at large. Maybe, for instance, the presence of the medical team attracted those patients who had already suffered a treatment failure with drugs they got from drug stores where patients do not need any medical prescription to buy drugs and self-medication is common. On the other hand, the presence of the investigators and the availability of free diagnostic and treatment facilities during the study period might have attracted to the clinic patients who would otherwise not have come for treatment and this is typical of malaria endemic areas.Overall, 22.99% (40/174) of the subjects experienced side effects. This is not surprising as artesunate therapy is not known to give rise to any serious side effects , 16, 21.In future, antimalarial therapy may be expanded by combining chemotherapy with vaccines (or other drugs) specifically designed to inhibit transmission of malaria. These \u201ctransmission blocking\u201d vaccines or drugs could reduce the potential for onward transmission of gametocytes carrying resistant genes, even if a relatively large number of parasites survive initial treatment. This could work through use of drugs or vaccines with a high degree of specific antigametocidal activity, drugs that nonspecifically reduce the likelihood of gametocytes developing, or drugs or vaccines that interfere with sexual reproduction and infection of the parasite within the mosquitoes when taken up with a blood meal .This study has shown that malartin and SP are effective and safe against uncomplicated falciparum malaria. This drug combination is therefore a better alternative for the patients that react to malartin/amodiaquine combination in Cameroon. One of the cornerstones of the current approach to malaria control remains the provision of prompt, effective malaria treatment. Finally, malaria control measures, be they directed at the vector or malaria parasites in the human host through mass drug therapy or susceptible hosts by way of chemoprophylaxis, must be coordinated by integrating available technology with an understanding of the epidemiology of the local malaria situation, which includes a consideration of the behaviour of both humans and the vector so as to be able to make the most appropriate and effective interventions."}
+{"text": "Refugee treatment guidelines recommended standard three-day chloroquine treatment (25 mg/kg) for first episodes and extended five-day treatment (40 mg/kg) for recrudescent infections, based on the assumption that a five-day course would more likely achieve a cure. An 142 falciparum patients were recruited into CQ25 or CQ40 treatment arms and followed up to 60 days with regular blood smears. The primary outcome was parasitological cure without recrudescence. Treatment failures were retreated with CQ40. PCR genotyping of 270 samples, from the same and nearby sites, was used to support interpretation of outcomes.pfcrt 76T was found in all isolates analysed, while pfmdr1 86Y and 184Y were found in 18% and 37% of isolates respectively.84% of CQ25 versus 51% of CQ40 patients experienced parasite recrudescence during follow-up . Cure rates were significantly improved with CQ40, particularly among adults. Fever clearance time, parasite clearance time, and proportions gametocytaemic post-treatment were similar between treatment groups. Second-line CQ40 treatment resulted in higher failure rates than first-line CQ40 treatment. CQ-resistance marker CQ is not suitable for first-line falciparum treatment in Afghan refugee communities. The extended-dose CQ regimen can overcome 39% of resistant infections that would recrudesce under the standard regimen, but the high failure rate after directly observed treatment demonstrates its use is inappropriate. Plasmodium falciparum and the remainder to Plasmodium vivax [During the extended Afghan conflict, waves of refugees totalling almost three million entered northwest Pakistan and more than one million remain ,2. Malarum vivax . Chloroqum vivax . It remain vivo resistance survey had been undertaken in refugee camps, despite CQ-resistant falciparum parasites spreading widely in Pakistan in the 1990s [The United Nations High Commissioner for Refugees (UNHCR), following national guidelines, adopted a three-day CQ treatment course  in refugee settlements. However, it became apparent during the 1990s that CQ was failing ,5. Basiche 1990s -6.As there was no evidence to support claims of poor adherence or the efficacy of extended-dose CQ, an open-label randomized clinical trial was conducted to determine whether supervised CQ treatment administered at 40 mg/kg over five days (CQ40) was more effective than 25 mg/kg over three days (CQ25) for curing infections completely without recrudescence . The triThe primary trial outcome was the proportion of individuals in each treatment arm that showed clinical and parasitological cure with no recrudescence. Sample size was calculated to detect a difference of 15% in cure rate between CQ25 and CQ40 treatment arms with 95% confidence and 90% precision, assuming a 20% loss to follow-up. The surveys were conducted during winter months to select only recrudescent episodes. Mosquito densities and malaria transmission drop during December and January, providing little opportunity for trial participants to receive further infective bites within the 60-day follow-up period ,9. Thus,in vivo studies . In Adizai camp, 21 patients were recruited per treatment group and followed for only 28 days. CQ25 patients received standard three-day treatment . CQ40 patients received extended 5-day treatment (CQ 40 mg/kg as 10 mg/kg/day on Days 0-2 and 5 mg/kg/day on Days 3-4). Dosages were measured in 1/4 CQ tablets of 37.5 mg each to give an average dosage (range) of 26.2  mg/kg for the CQ25 arm and 42.1  mg/kg for the CQ40 arm. All treatment was directly observed for 30 minutes post-treatment.Two trials, completed in 1998, were conducted in Baghicha, Kagan and Adizai refugee camps .If parasites reappeared during the follow-up period, patients received CQ40 second-line rescue treatment as per MoH and UNHCR guidelines. If parasites reappeared a further time, patients received single-dose sulphadoxine-pyrimethamine  or mefloquine (25 mg base/kg) treatment. SP and mefloquine tablets were manufactured by Roche. Chloroquine was manufactured by Aventis and supplied by the World Health Organization-Special Programme for Research and Training in Tropical Diseases (TDR). Samples of in vivo selection criteria for low to moderate transmission settings were randomized to either CQ25 or CQ40 groups using randomized lists [Participants were recruited through passive case detection at BHUs and active case detection in communities. Individuals with symptomatic falciparum malaria who met WHO ed lists . Exclusied lists .Local health supervisors collected demographic and clinical information at enrolment, including weight, temperature, and symptoms. Supervisors directly observed treatment according to dosing schedules, prepared blood smears, and recorded temperature and clinical symptoms daily for the first five days, then every third day until day 28. Patients in Kagan and Baghicha were additionally observed every four days until day 60.Thick and thin blood smears were stained with 3.5% Giemsa solution and all slides read on day of collection by a BHU-based microscopist. Trophozoites and gametocytes were counted against 200 white blood cells (WBC) from thick blood smears, assuming a WBC count of 8,000/\u03bcl. A smear was declared negative if no parasites were seen after examining 100 fields. Slides were re-examined for accuracy of diagnosis and recounted by an independent senior microscopist, blinded to patient, follow-up day, original result, and outcome. Differences in count were on average no greater than 5%. Finger-prick blood samples (~200 \u03bcL) were dried on Whatman filter paper prior to treatment (Day 0) and sent to LSHTM for genetic analysis.in vivo criteria [Trial outcomes were treatment failure rates, fever clearance times (FCT), parasite clearance times (PCT), and number of recrudescences. Therapeutic responses were early treatment failure (ETF), late treatment failure (LTF) and adequate clinical and parasitological response (ACPR) using standard WHO criteria . Parasitcriteria ,12.\u00aeExcel, with range and consistency checks to reduce transposition error, and analysed using Stata\u00ae11.0. Analysis was conducted on an intention-to-treat basis. Data from the three study sites was combined for the first 28 days to calculate therapeutic outcomes and analyse subsequent malaria episodes and Kaplan-Meier survival estimates [2) tests for proportions and Mann-Whitney U tests for continuous data. Logistic regression was used to calculate odds ratios (OR) of treatment success at weekly intervals and differences between treatment outcomes. A priori confounders  were adjusted for in multivariate analysis.Data was double-entered in Microsoftstimates . Data reP. falciparum chloroquine-resistance transporter gene (pfcrt)at codons 72-76 and P. falciparum multidrug resistance protein-1 (pfmdr1) at codons 86 and 184 [pfcrt leading to a lysine to threonine change at codon 76 (K76T) while pfmdr1 N86Y and Y184F are thought to have a modulatory effect [pfcrt 76T and pfmdr1 86Y alleles may serve as predictive markers for CQ resistance in non-immune individuals living in low-transmission areas, while combined pfcrt 76T and pfmdr1 86Y may be useful molecular markers for resistance to additional drugs, such as amodiaquine (AQ) [PCR genotyping could not be conducted on patient data to determine recrudescences as samples were lost in transit. However, the authors were able to analyse 90 blood samples from falciparum cases in Adizai (the same camp) and 180 from Jalalabad, Afghanistan, collected shortly afterwards for a clinical trial to characterize resistance genotypes (Rowland unpublished). Parasite DNA was extracted from 270 blood spots collected on filter paper pre-treatment (Day 0) as described elsewhere . PCR-seq and 184 . CQ resiy effect ,16. The ine (AQ) -19.Figure in vivo system of early and late treatment failure or adequate clinical and parasitological response, and (b) the parasitological response system of S, RI, RII, RIII [No participants were lost to follow up by day 28. Table II, RIII ,20. FeveCQ40 patients had fewer recrudescences than did CQ25 patients during the first 28 days. Among CQ40 patients, only one recrudescent episode occurred before Day 7. Among CQ25 patients, 86% of recrudescence occurred between days 7 and 28 post-treatment. The parasitological failure rate was negatively associated with age, with failure highest among under-fives and lowest among over-fifteens  was present in 100% of samples successfully analysed from Adizai (63) and Jalabad (179). Pfmdr1 86Y was found in 14% (12/88) of Adizai and 22% (33/151) of Jalalabad samples. The pfmdr1 184Y allele was found in 27% (22/82) of Adizai samples and 46% (69/151) of Jalalabad samples.Table Chloroquine failure rates were higher than anticipated, and since administration was directly observed, failure was due to resistance rather than poor adherence. Analysis showed that with 51% failure in CQ40 and 84% failure in CQ25, chloroquine is no longer suitable for falciparum malaria treatment among Afghan refugees, either as first or second-line with short or extended regimens, and usage should stop. Second-line CQ40 achieved a higher failure rate than did first-line, demonstrating lack of suitability for this purpose.in vivo trials conducted in the area that did include genotyping indicated fewer than 5% would be new infections [While the authors were unable to make use of PCR genotyping to distinguish recrudescent from new infections, other fections . The mospfcrt 76T mutation in 100% of isolates analysed is consistent with a low degree of heterogeneity in the parasite population, as also shown by Khatoon et al in isolates from nearby Bannu district NWFP [pfcrt 76T allele is strongly associated with CQ and amodiaquine (AQ) resistance in falciparum isolates from Asia, Papua New Guinea, Africa, and South America [Pfmdr1 86Y and 184Y alleles, which are also associated with CQ and AQ resistance, were present in only a minority of our isolates from Adizai camp or from Bannu district [pfmdr1 alleles were not strongly selected among treatment failures [pfcrt codon 72-76 haplotype SVMNT present in Pakistan is sufficient by itself (i.e. without pfmdr1 86Y and 184Y) to cause high-level CQ and AQ resistance [pfcrt codon 72-76 CVIET appears to be the predominant haplotype, AQ remains relatively effective [in vivo resistance, the CQ-resistant variant CVIET haplotype was present with pfmdr1 86Y and 184Y alleles which presumably added to the resistance there [The finding of the ict NWFP . The pfc America ,15. Pfmdfailures . These fsistance ,22. By cffective . In the ce there ,23-26.et al have, in parallel, undertaken clinical studies with high-dose CQ in Guinea-Bissau [pfcrt 76T were successfully treated compared to only 34% with 25 mg/kg [The rate of parasitological failure was higher after second-line than after first-line CQ40 treatment. Recrudescent infections presumably started with higher proportions of resistant parasites than did initial infections. However, this cannot explain why the initial CQ40 course seemed to eliminate around 39% of resistant infections, as indicated by the improved cure rates over 60 days following initial five-day (51% recrudescence) as compared to three-day treatment (84% recrudescence). Among this 39%, any resistant parasites must have been removed by the additional two days of treatment and did not reappear over the subsequent 60 days. Ursing a-Bissau ,28. They25 mg/kg . This wapfcrt 76T was highly prevalent in the Pakistan samples, pfcrt 76T prevalence in the Guinea-Bissau population, discussed above, remained stable at a much lower 25% between the years 1990 and 2005 [pfcrt 76T between continents are likely due to differences in the fitness of resistance alleles, as the pfcrt 72-76 SVMNT resistance haplotype dominant in India, Iran, Pakistan and Afghanistan is not associated with re-emergence of CQ sensitivity or fluctuations in seasonal prevalence shown by the CVIET haplotype in some parts of Africa [pfcrt 76T frequency would remain high. In the African settings, sensitive parasites may have found a niche in the many untreated infections, where their greater fitness would allow them to compete better than any co-infecting resistant parasites. It has been more difficult for CVIET-carrying parasites to gain the same high prevalence in Africa as SVMNT-carrying parasites have achieved in parts of Asia.While and 2005 ,29. Thesf Africa ,28,30-32The present trial did not measure adherence, as it was designed to assess efficacy rather than effectiveness. Consequently, it cannot challenge the initial assumption that refugees fail to adhere to either three-day or five-day courses. However, recent research demonstrates that with appropriate instructions - as in a trial of unsupervised 14-day primaquine treatment - Afghan refugees will adhere to much longer treatment regimens than the five-day course described here ,33. WhatIn Afghanistan, where many refugees have returned, most malaria cases are still treated with CQ, without parasitological diagnosis by either microscopy or rapid diagnostic test . This maAlthough combination therapy using artesunate-SP has been adopted as policy for treatment of confirmed falciparum malaria in Pakistan and Afghanistan, implementation is patchy and uptake slow . While fThe authors declare that they have no competing interests.NH analysed and interpreted data and drafted the manuscript. ND was responsible for data collection, PCR analysis, and critically reviewing the manuscript. SS assisted with initial analysis and critically reviewed the manuscript. RH and KB provided critical analysis of genotyping results and critically reviewed the manuscript. MR conceived and designed the study, and revised the manuscript critically for intellectual content. All authors approved the final version for publication."}
+{"text": "Plasmodium falciparum malaria over the past decade.Anopheles vector control measures, including insecticide-treated bednets and indoor residual spraying, ACTs have helped achieve important reductions in malaria mortality and morbidity.11Artemisinin-based combination therapies (ACTs) have been adopted globally as first-line drugs for the treatment of t1/2P or slope half-life).in vivo parasite clearance rates with a heritable component; parasite genome-wide mapping studies are narrowing the search to several candidate loci.14Resistance to artemisinin is unlike any other resistance seen to date, and it is inferred by observations of delayed parasite clearance after treatment. An early report from Cambodia used the median parasite clearance time (defined as the time to parasite-negative blood smears post-initiation of treatment), which was found to be substantially slower in Pailin, western Cambodia, compared with Wang Pha, Thailand .American Journal of Tropical Medicine and Hygiene, the work by Maiga and othersP. falciparum malaria in a prospective trial; they were treated with a curative 7-day regimen of oral artesunate, and their clinical and parasitological responses were followed. No significant differences between the two study periods were found in terms of the percentage of patients that had cleared 95% of their parasitemias or completely cleared their infections within 24 hours . All patients were blood smear negative by 72 hours and; 100% cure rates were observed 28 days post-treatment. Equal median fever clearance times (\u223c24 hours) were recorded in both trials. Overall, the data clearly illustrate that artesunate has not become less effective 6 years after the initiation of widespread use of ACTs in Mali.In this issue of the This study by Maiga and colleaguesIn vitro studies to date suggest that resistance is both hard to select for and when obtained, can be unstable.17\u2013in vitro and in vivo data is also paramount, and there is a recognized need to implement a coordinated global resistance surveillance network.Why would artemisinin resistance begin to emerge first in Cambodia and not high-transmission settings in Africa? The recent decline in artemisinin efficacy in Cambodia has often been attributed to prior decades of artesunate monotherapy that, due to the short drug half-life, can readily lead to parasite recrudescence and thereby potentially enrich for resistant parasites."}
+{"text": "Successful implementation of malaria treatment policy depends on the prescription practices for patients with malaria. This paper describes prescription patterns and assesses factors associated with co-prescription of antibiotics and artemether-lumefantrine (AL) for patients presenting with fever in rural Tanzania.From June 2009 to September 2011, a cohort event monitoring program was conducted among all patients treated at 8 selected health facilities in Ifakara and Rufiji Health and Demographic Surveillance System (HDSS). It included all patients presenting with fever and prescribed with AL. Logistic regression was used to model the predictors on the outcome variable which is co-prescription of AL and antibiotics on a single clinical visit.A cohort of 11,648 was recruited and followed up with 92% presenting with fever. Presumptive treatment was used in 56% of patients treated with AL. On average 2.4 (1 \u2013 7) drugs was prescribed per encounter, indicating co-prescription of AL with other drugs. Children under five had higher odds of AL and antibiotics co-prescription  than those aged more than five years. Patients testing negative had higher odds  of AL and antibiotics co-prescription. Patients receiving treatment from dispensaries had higher odds  of AL and antibiotics co-prescription than those served in health centres even though the deference was not statistically significant.Regardless the fact that Malaria is declining but due to lack of laboratories and mRDT in most health facilities in the rural areas, clinicians are still treating malaria presumptively. This leads them to prescribe more drugs to treat all possibilities. Malaria remains a public health problem in Africa. It is estimated to claim about 1 million deaths and over 400 million malaria cases worldwide each year, with 90% of these deaths occurring in sub Saharan Africa . AssessmFever has been used as a major clinical symptom for malaria , Now repMicroscopic examination of Giemsa-stained blood films remains a cornerstone of malaria diagnosis throughout Tanzania, but is only available at hospitals and some health centers. Historically, more than 5,000 of the lowest-level facilities  had no laboratory diagnostic capacity, leaving health care workers at more than 90% of facilities to diagnose malaria on the basis of clinical signs and symptoms alone. According to the recent WHO guidelines, all suspected malaria cases should be parasitological confirmed prior to treatment, including children under five. NMCP\u2019s policy has changed from presumptive treatment to confirmatory parasitological diagnosis. The NMCP objective is to increase the percentage of laboratory-confirmed malaria cases in public health facilities from a baseline of 20% to 80%. It is clear from numerous assessments that the quality of malaria microscopy is very poor at almost all levels of the health system. Phased rollout of RDTs began in April 2009, starting in areas of low/moderate transmission and expanded to areas of stable/high transmission. Currently, laboratory confirmation is happening in only 20% of the suspected cases and there is no system for laboratory quality assurance and quality control . Over-diSeveral factors influences prescribing behaviour of clinicians, therefore, to improve prescription behaviour, it is necessary to understand predictors of those behaviours ,18. ThisThe INDEPTH Effectiveness and Safety Studies of Anti-malarial Drugs in Africa (INESS) is an exciting new platform that aims to enable African researchers to carry out large Phase IV trials . INDEPTHThe study was conducted in 8 selected health facilities located in the Rufiji and Ifakara HDSS areas from May 2010 to December 2011. The Ifakara HDSS, situated 320\u00a0km south- west of Dar es Salaam, has been in operation since 1996. It covers part of Kilombero and Ulanga districts with a total population of 99,000 people, served by 14 health facilities . Out of The study design was a cohort event monitoring which was observational, longitudinal and prospective. All patients prescribed AL from the 8 selected health facilities within HDSS area were recruited. Patients were asked to come to the health facility on day 3 and day 7 for clinical evaluation and assessing their prognosis including if they have experienced any of the side effects. They could come at any day as well when they experienced any adverse event or if they have any doubts. If patients did not come at the health facility on scheduled days, they were immediately followed up on the following day at their respective households by a trained field worker, and for some patients follow ups were conducted by using phones. Phone was only used for those with mobile phones and did not turn to the health facility on scheduled days. Patients were declared lost to follow up if three attempts have been made to trace him/her at his/her households and five times by using mobile phone. Information on demographic, complaining symptoms, laboratory investigations, past medical history, past medical history, medication used and all events were recorded at recruitment and during follow ups.All patients attended 8 selected health facilities in Ifakara and Rufiji HDSS areas that were prescribed with artemether-lumefantrine for malaria treatment regardless of their demographic characteristics.IHI/IRB/No.A67-2009.The INESS platform and its modules passed through and were reviewed and approved by the Tanzanian National Institutes of Medical Research and IHI\u2019s Ethical Review Boards with reference number At recruitment and after obtaining the informed consent for participation in the study, data collection was done using a standardized questionnaire developed in English and translated into Kiswahili. Iinformation on demographic, complaining symptoms, laboratory investigations, past medical history, present medical history, medication used, history of drug reactions and all events were collected. A trained clinician interviewed patients as they come for treatment and once they were prescribed with antimalarial a clinician filled in a clinical questionnaire. Follow up information was collected by trained field workers using a standardized questionnaire on day 3 and 7.All questionnaires used to collect information at recruitment and at follow up were taken for manual editing, validation and data entry which was done using the Epidata 3.1 . Data enA total of 11,648 patients who were prescribed with AL were recruited from eight government health facilities in Ifakara HDSS and Rufiji HDSS. More than half (55%) were female, median age was 6.4\u00a0years (Inter Quartile Range: 2 \u2013 19) and a quarter of the patients had used medicine before getting to the health facility  strategy . The IMCet al.,27 suggeFindings from this study shows that presumptive treatment was done for all age groups; under five and those above 5\u00a0years. Similar results of presumptive treatment for malaria patients above 5\u00a0years was fund in study done in Kenya . PresumpThe mean number of drugs prescribed was 2.4 drugs per patient per encounter, which is above the WHO guidelines on rational use of drugs with reference values of (1.6-1.8) drugs per encounter . Since aAnalgesics were commonly prescribed class of drugs as more than 50% of patients prescribed AL were co-prescribed it with Analgesics. This is a common practice to clinicians and reported in other countries such as Sudan  and YemeThe study indicates that antibiotics were co-prescribed for 20% of encounters which is less compared to 30.8% observed in Ghana . Antibioet al. in [Furthermore this study assessed the predictors of antibiotics co-prescription. Predictors found to be associated with the risk of being co-prescribed of other drugs with AL were age group and type of diagnosis . Children aged less than five years were more likely to be co-prescribed with antibiotics than those aged 5\u00a0years and more Table\u00a0. Similart al. in . This mit al. in .Patients with fever who had negative results on malaria, due to lack of laboratory services, clinicians tend to deal with all possibilities by giving antimalarial if fever was due to malaria and antibiotics if fever was caused by bacterial infection or analgesics for fever itself. A similar observation was done in Zanzibar as those with clinical diagnosis were more likely to be co-prescribed with antibiotics as compared to those with malaria rapid diagnosis tests (mRDT). Those who were not tested were presumptively treated and IMCI guidelines were used for children under five years . For patFever is still the main complain regardless malaria decline. Presumptive treatment is still practised. When a child is having fever and tested malaria negative clinicians tends to give more drugs including atimalarial to cure for all possibilities.More training and supervision of clinicians\u2019 prescription pattern especially to children to avoid concomitant use of antibiotics.Authorities should make sure that facilities are equipped with mRDT or laboratory for better malaria diagnosis and minimise the un-necessary prescription of antibiotics.Study was conducted for outpatients who were treated with artemether-lumefatrine and can never be generalized for in-patients treated with any antimalarial drug or outpatients not treated with artemether-lumefatrine. The study was non-interventional and did not assess whether prescription was appropriate for reported symptom and according to diagnosis.The authors declare that they have no competing interests.MN wrote the first draft, MN, MS and DK data analysis and reviewed manuscript, MA, RK, and IM reviewed the manuscript, AD and SA supervised the writing and contributed to the discussion. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2458/13/1097/prepub"}
+{"text": "Increasing affordability of artemisinin combination therapy (ACT) in the African retail sector could be critical to expanding access to effective malaria treatment, but must be balanced by efforts to protect the efficacy of these drugs. Previous research estimates ACT adherence rates among public sector patients, but adherence among retail sector purchasers could differ substantially. This study aimed to estimate adherence rates to subsidized, over-the-counter ACT in rural Uganda.An intervention study was conducted with four licensed drug shops in Eastern Uganda in December 2009. Artemether-lumefantrine (AL) was made available for sale at a 95% subsidy over-the counter. Customers completed a brief survey at the time of purchase and then were randomly assigned to one of three study arms: no follow-up, follow-up after two days or follow-up after three days. Surveyors recorded the number of pills remaining through blister pack observation or through self-report if the pack was unavailable. The purpose of the three-day follow-up arm was to capture non-adherence in the sense of an incomplete treatment course (\"under-dosing\"). The purpose of the two-day follow-up arm was to capture whether participants completed the full course too soon (\"over-dosing\").Of the 106 patients in the two-day follow-up sample, 14 (13.2%) had finished the entire treatment course by the second day. Of the 152 patients in the three-day follow-up sample, 49 (32.2%) were definitely non-adherent, three (2%) were probably non-adherent and 100 (65.8%) were probably adherent. Among the 52 who were non-adherent, 31 (59.6%) had more than a full day of treatment remaining.Overall, adherence to subsidized ACT purchased over-the-counter was found to be moderate. Further, a non-trivial fraction of those who complete treatment are taking the full course too quickly. Strategies to increase adherence in the retail sector are needed in the context of increasing availability and affordability of ACT in this sector. Plasmodium falciparum malaria in nearly all malaria endemic countries and is the most effective treatment for the disease [Artemisinin Combination Therapy (ACT) is the WHO recommended first-line therapy for  disease . Several disease -4. The d disease ,6.Volumes of ACT purchased and distributed through private and retail sector channels are increasing substantially ,8, thougNon-adherence to recommended treatment regimens, resulting in sub-therapeutic drug concentrations is a key driver of parasite resistance ,20-22. NAs financing and other initiatives aiming to increase access to ACT reach the substantial share of malaria patients who seek treatment in the retail sector, it is increasingly crucial to have estimates of adherence rates among this population. Understanding ACT adherence in the retail sector is necessary for developing appropriate strategies to prevent the emergence of artemisinin resistance. This study estimates adherence rates and explores associated characteristics among patients purchasing over-the-counter ACT in Uganda. It is the first study to use follow-up methods to estimate adherence rates to subsidized, over-the-counter ACT.The study was conducted in Soroti District, a rural area in Northeast Uganda, over three weeks between November and December of 2009. Soroti has a very high rate of malaria transmission, with more than 100 infective bites per person per year . As commBrief surveys were conducted at 23 retail outlets (\"drug shops\") in Soroti, with information recorded on location, shop owner and attendant qualifications and years in business, customer traffic, days/hours open and most common anti-malarials sold. Four shops, all licensed and registered with Uganda's National Drug Authority, were selected from this sample to participate in the study. These shops were licensed \"Class C\" outlets, meaning that they can solely dispense over-the-counter medicines. Drug shops in Uganda are distinct from pharmacies, which can sell prescription medicines as well, but are typically located in urban areas. At the time of this study, ACT was in the process of being approved for over-the-counter dispensing in Uganda and we received special permission from the Ugandan Ministry of Health to allow our participating shops to dispense ACT. The shops that were included in the study were chosen because they were far apart from one another, staffed by well-qualified attendants, and were open a sufficient amount of time with enough customer traffic for us to obtain our patient sample in a three week time-frame. All four shops were managed by nursing assistants, were open all seven days a week (except one shop which was closed on Sunday) and were open from the early morning until mid-evening. The number of malaria patients visiting the shop per day ranged from 5 in one shop to 20 at the shop with the highest patient traffic. All shops cited Quinine as their most commonly sold anti-malarial. All four shops were located in the area of their village known as the trading center, which generally constitutes a grouping of a small number of retail outlets often including at least one drug shop. None of the shops were in a very remote location or in a busy town center.Patients or their caregivers who purchased subsidized AL from the participating shops--either because they requested it or because the shop attendant recommended it--were recruited to enroll in the study immediately following purchase. Pregnant women and infants under four months of age were excluded from the study since AL is not approved for this patient population. Patients were recruited until enrollment reached 395, a sample size allowing us to detect a projected adherence rate in the three-day follow-up group of 70% with a 10% margin of error, a design effect of 2 and 20% loss to follow up.Shop attendants participated in a one-day training session led by a Uganda Ministry of Health-trained professional on storage, administration, and appropriate use of AL. With the information and materials obtained during the training, attendants were instructed to follow their normal dispensing practices. Shop attendants were trained to refer any patients who either did not recover after completion of anti-malarial treatment or who were exhibiting signs of severe malaria to the nearest public health facility.\u00ae manufactured by Novartis, for sale in the shops. The Coartem treatment package was the standard packaging commonly available in the private sector, which is a simple orange and white box with black and red text on it. Unlike some packaging provided in the public sector, this packaging included no drawings or visual cues on appropriate dosing other than a paper insert with written English instructions printed in black and white. The price of Coartem to consumers was fixed at 200, 400, 600 or 800 Ugandan Shillings ($0.10 - $0.40 at 2009 exchange rates) depending on the age/weight band. Shops were permitted to keep 50 Ugandan Shillings on every sale. Prices were equivalent to those in the CAPSS ACT subsidy pilot programme [Following the training, the study team provided the selected drug shops with a controlled supply of AL, brand name Coartemrogramme  and rougrogramme . A full rogramme .All eligible customers who purchased AL and gave consent to be surveyed at the shop were enrolled. The study team conducted a brief survey at the shop to collect symptom and background information (including the village and location of the household) and to record observations regarding the interaction between the shop attendant and the customer. If the customer was purchasing AL for another family member or neighbour, the household information for both the customer and the patient was obtained. Neither customers nor patients were informed of the intent to follow-up or were asked to retain their blister pack. All questionnaires and informed consent documents were translated into Ateso, the predominant language in Soroti. Interviews and informed consent processes were conducted in either English or Ateso, according to the language preferences of the participant.A pseudo-random number generator in STATA Version 11  was used to create a list that assigned a follow-up assignment to each patient, based on the order in which they were enrolled. The project manager had one list numbered 1 to 100 for each shop, with a pre-filled follow-up assignment for each number. Surveyors enrolling patients at the shop (and the patients themselves) were blinded to the eventual follow-up assignment. Participants were randomly assigned to either no follow-up, follow-up at day two or follow-up at day three. The aim of the three-day follow-up was to assess whether patients delayed or discontinued treatment. The aim of the two-day follow-up was to assess whether patients finished their entire treatment course too early. The aim of the no follow-up arm was to introduce additional uncertainty about the intent to follow up. The window of time following enrollment for the two-day follow-up arm was between 41 and 55 hours, while for the three-day follow-up arm it was between 72 and 96 hours.At the household follow-up visit, patients or their caregivers were again asked for consent to be surveyed. Follow-up questionnaires included sections on demographic and health background information and household assets in order to characterize socioeconomic status. Additional sections focused on the patient's medicine-taking experience and behaviour, during which time the surveyor asked to see the blister pack and recorded the number of tablets remaining in the pack. If the pack was unavailable or the participant declined to show the pack, the surveyor asked the participant how many pills were remaining. Finally, surveyors reviewed appropriate adherence practices with the participant. If participants were still feeling ill at the time of the household visit, enumerators were instructed to recommend to patients that they visit the nearest public health facility as soon as possible.Adherence is defined with regard to completion of the entire age-appropriate treatment course at the three-day follow-up visit, based on blister pack observation or, in its absence, self-report. A patient was considered \"probably adherent\" if the blister pack was available for inspection and there were no pills left or if no blister pack was available but the patient reported that all the pills were taken. A patient was considered \"definitely non-adherent\" if the blister pack was available for inspection with pills remaining in the pack and \"probably non-adherent\" if no blister pack was available for inspection but the patient reported that the treatment course was not completed.The study gained approval from the Uganda National Council for Science and Technology (UNCST), Uganda's Office of the President, and the Harvard School of Public Health Institutional Review Board.Survey data were double entered in CSPro Version 3.1  and crosschecked for discrepancies until the error rate calculated for all sections was below 0.5%. The investigators cleaned and analysed data using STATA Version 11 software . Analysis of variables associated with adherence is conducted in two ways. First, running probit regressions of the dichotomous variable \"probably adherent\" (as defined above) on each of the variables listed in Tables Variables potentially associated with adherence are presented in Tables Of the 395 enrolled customers, 39 (9.9%) were assigned to no follow-up, 159 (40.3%) were assigned to two-day follow-up and 197 (49.9%) were assigned to three-day follow-up. Loss to follow-up was balanced across groups at 27 (17%) and 32 (16.2%) in the two and three-day follow-up groups, respectively. Since the timing of the survey and blister pack observation is critical for accurate adherence data, those found beyond the appropriate time were excluded from the analysis. This includes 26 (16.4%) patients in the two-day follow-up group and 13 (6.6%) in the three-day follow-up group, leaving a sample of 106 patients in the two-day group and 152 in the three-day group. Among those found on time 91.5% (n = 97) and 73.7% (n = 112) showed their blister pack for inspection in the two-day and three-day groups, respectively. The most common reasons for not showing the blister pack in the three-day group were that it was thrown away (58.7%) or lost (30.4%).The randomization successfully achieved balance across observable characteristics Table --the twoAlthough this study was conducted prior to the AMFm, familiarity with Coartem was high in this population, possibly because of experience with the drug through the public sector. When asked at the follow-up visit why the patient or caregiver purchased Coartem half of respondents answered that they believed it was the most effective anti-malarial and a quarter responded that they had taken it before and liked it. The majority of patients/caregivers, when asked which anti-malarial they prefer if money were no concern stated Coartem  and again the most commonly stated reasons for this preference were because it was believed to be most effective or because they had taken it before and liked it. Nearly all patients said their condition had improved and they could work or play at follow-up, but just over 20% said they were still experiencing some fever. Respondents' expectations of the likelihood of malaria in the next month were very high--roughly 50% chance of infection in the next month for adults and nearly 75% chance for babies.Nearly all patients received the correct dose for their age group and paid the correct price Table . Those wJust over 13% (14/106) of patients had finished all of their medicine at the time of the home visit in the two-day follow-up group. The mean number of doses left on day two was 1.67 .Out of 152 patients in the three-day follow-up group, 100 (65.8%) were \"probably adherent\", whereas 49 32.2%) were \"definitely non-adherent\" and 3 (2.0%) were \"probably non-adherent.\" Among the non-adherent group (n = 52), the average number of doses left was 2.27 . 31 (59.6%) patients in the non-adherent group had more than a full day of treatment remaining  and had .63 fewer doses remaining  than those whose condition had not improved. Those still experiencing fever had .45 more doses left  than those whose fever had resolved. Higher expected likelihood of malaria in the next 30 days is significantly associated with lower levels of treatment completion and more doses left for children ages 6-15 .The data are mixed on the relationship between shop attendant instructions and adherence behavior. There is no significant relationship between treatment completion or the number of pills left and any of the variables measuring instructions given at the shop based on observation. However, we also asked patients (or their caregivers) at the time of follow-up whether or not they were provided with instructions and whether or not those instructions were clear. Based on these self-reported measures, having received instructions is associated with a 28% increase in treatment completion  and .81 fewer doses remaining . Further, among patients who received written instructions, those reporting that the instructions were clear were 16.5% more likely to complete treatment  and had .64 doses fewer remaining  than those reporting that the instructions were unclear.A substantial amount of research has been conducted on adherence to ACT obtained through the public sector with generally encouraging results ,29,33,34Follow-up home visits from the two-day follow-up group revealed that 13% of patients had already finished their medicine. This is a non-trivial fraction of patients, but it is low enough to infer that, for the most part, people are aware that the treatment should not be completed by day two. Combined with the fact that we observed shop attendants giving dosing instructions in the majority of cases, this suggests that non-adherence is less likely due to a general non-comprehension of dosing than to a non-comprehension of the importance of finishing the treatment course specifically, a message that should perhaps be emphasized in drug shop training or other interventions to increase adherence. Adherence rates are found to be higher among patients whose condition has improved and who are no longer experiencing fever, possibly suggesting that people stop treatment when they feel it is not helping. It is also possible that this association between adherence and illness resolution is simply due to the fact that finishing the entire course leads people to feel better. Shop attendants in this study very rarely provided instructions in case the illness got worse or did not resolve and it is possible that this lack of detailed instruction contributed to the decision to discontinue treatment. Further, it seems likely that verbal and written instructions in the retail sector need to be made more understandable since: 1) non-adherence is significantly more likely among those reporting that they did not receive clear instructions, 2) the packaging insert was in English, while the majority of patients could not read English, and 3) education was significantly correlated with adherence. Patients who anticipate frequent malaria infections were found to be less likely to adhere, suggestive that the adherence decision could be related to the desire to keep pills for the next malaria episode.The setting and characteristics of the study could affect the generalizability of these results to retail sector ACT purchasers in other countries and contexts. ACT was very heavily subsidized. To the extent that price influences adherence , it may not be appropriate to infer adherence rates from this study to areas in which ACT is unsubsidized or is subsidized but has a substantially higher retail price. While we show in Table The sample sizes used for the analysis were smaller than ideal  because of the loss to follow-up and the fact that people were found beyond the appropriate time frame. This attrition was largely due to the fact that patients were not made aware of the intention to come to their household for a follow-up visit and were not asked for directions to the household, a trade-off made to reduce the degree to which patients adhered to treatment because they knew they were under observation. Finally, blister packs were observed for roughly three-quarters of the sample. Those who did not show blister packs could have thrown them away because they had finished treatment or could have been reluctant to show us the packs because pills were left over. If the latter is the case, our estimates of adherence are too high.The retail sector is the most common source of anti-malarials in general, and increasingly a common source for ACT. Nearly 150 million subsidized courses of ACT have been purchased through the AMFm for retail sector distribution, an unprecedented scale and speed of a public intervention to finance distribution of a drug in the private sector in the developing world. At the end of 2012, informed by the results of an independent evaluation, the Global Fund Board will make a critical policy decision on whether to continue with this initiative or potentially expand it. The first phase of the AMFm has included some interventions which could increase ACT adherence, such as additional ACT packaging requirements and training of private retail staff. ACT manufacturers participating in the AMFm were required to create separate packaging by age/weight band, to clearly mark the individual dose sub-units and to provide symbolic representations of key instructions. As our adherence estimates pertain to a sample of patients who received ACT from trained shop attendants who administered the correct (age-appropriate) dose in nearly every case, these findings suggest that additional measures may be needed to achieve high adherence rates. Any such measures will need to be highly cost-effective and operationally simple given the constraints on global health resources and the scale of private sector ACT distribution. Options could include enhanced simple information or instructions on ACT packaging and mobile phone based reminders to patients, both of which are being examined through research being implemented in connection with the AMFm . ResultsOS and AM are members of, and JC receives research support from, the Clinton Health Access Initiative, which is actively supporting efforts to expand funding for and implementation of ACT treatment in the private sector.JC, JA and OS conceptualized and designed the study. JC was Principal Investigator and AM and JA managed study implementation. JC, EY and AM conducted data analysis. All authors contributed to the manuscript. All authors read and approved the final manuscript.Table S1. Dosage Given by Patient Age.Click here for file"}
+{"text": "Plasmodium falciparum malaria in most areas of the world, where malaria is endemic, including Sudan. However, few published data are available on the use of ACT for treatment of P. vivax malaria.Artemisinin-based combination therapy (ACT) is the treatment of choice for uncomplicated P. vivax malaria received artemether-lumefantrine (AL) tablets (containing 20mg artemether and 120 mg lumefantrine) and were monitored for 28 days.This study was conducted at a health centre in Kassala, eastern Sudan, from October to December 2011. Patients with uncomplicated Out of the 43 cases enrolled in this study, 38 completed the 28-day follow-up. Their mean age was 25.1 years (SD: 1.5). On day 3 following AL treatment, all of the patients were afebrile and aparasitaemic. By day 28, all 38 patients exhibited adequate clinical and parasitological responses to AL treatment. The cure rate was 100% and 88.4% for the per protocol analysis andfor the intention to treat analysis, respectively. Mild adverse effects  that resolved spontaneously were observed in four (10.5%) of the patients.P. vivax malaria in the study in eastern Sudan.AL combination therapy was fully effective for treatment of Trial. Gov: NCT01625871 Plasmodium vivax infection is a major global health problem. This species of parasite has the broadest geographic distribution of the five malaria species known to infect humans [P. vivax annually [P. vivax is mainly endemic in Southeast Asia and Latin America [t humans . There aannually ,3. Altho America -8.Plasmodium falciparum malaria in Sudan [P. falciparum malaria, respectively.Malaria is a important health problem in Sudan, and in 2002, an estimated 9 million disease episodes and 44,000 deaths from the disease occurred [in Sudan ,11 has lP. vivax infection should prompt urgent treatment with effective anti-malarial medication [P. falciparum occurring [P. vivax in most endemic countries. However, there is growing evidence that the efficacy of chloroquine against P. vivax is declining in many areas, especially Southeast Asia [P. vivax and P. falciparum is preferred, especially in a country like Sudan where chloroquine is no longer registered or available [P. falciparum has led to a closer examination of their role in the management of P. vivax malaria. AL is one such combination therapy that is widely used to treat uncomplicated P. falciparum malaria, and is the first-line treatment for P. vivax malaria according to the national malaria control programme in Sudan. However, there is no published data on the efficacy of AL for the treatment of P. vivax malaria in Sudan. Therefore, the efficacy of AL for the treatment of uncomplicated P. vivax malaria was investigated in Sudan. The study was conducted at a health centre in eastern Sudan, an area characterized by unstable malaria transmission [P. vivax malarial disease has been observed in the area [Early diagnosis and effective treatment with an appropriate drug is one of the main components of the World Health Organization\u2019s strategy to reduce malaria related mortality . Episodedication . While mccurring , chloroqast Asia -16. Hencvailable . The farsmission . Recentlthe area . The stuP. vivax mono-infection and willing to participate in the study were enrolled. Those individuals with severe malnutrition, pregnancy, or with a history of allergy and/or intolerance to the drugs were excluded. Informed consent was obtained from all of the patients, or in the case of children, their guardians. Structured questionnaires were used together with socio-demographic characteristics, medical histories (including the duration of the illness) and physical findings.This observational  clinical trial was conducted at the Fatima Eldeaig Health Centre in Kassala, eastern Sudan  and took place from October to December 2011 /\u03bcL of blood as the multiplier. Smears were considered negatives if no parasites could be seen in 100 high-power microscopic fields. Blood films were double checked (blind) by two independent examiners, and discordant results were agreed on after evaluating the evidence of each examiner. Parasite densities differing by more than 10% between the different microscopists were evaluated by a third microscopist, whose result was deemed to be final.AL  was administered over a three-day period. AL tablets contained 20 mg of artemether and 120 mg of lumefantrine. Patients were weighed and those corresponding to 5\u201314, 15\u201324, 25\u2013 34 or >35 kg were each given six doses of AL , each dose consisted of one, two, three or four tablets, respectively. The morning dose was given at the health Centre under the direct supervision of a medical officer. Young children who could not swallow tablets were given suspensions created by crushing and dissolving their tablets in water. All patients were observed for vomiting for 1 h after each treatment. If vomiting occurred within 30 min of a dose, a second full dose was administered. If vomiting occurred 31\u201360 min after a dose, however, only half a dose was given. If vomiting occurred after a repeated  dose, the patient was given a quinine infusion and excluded from the study. These instructions were given to all of the patients (or guardian in the case of children) because the evening dose was given un-supervised. Patients also received unsupervised primaquine tablets (15 mg) daily for 14 days.Patients were asked to return on days 1, 2, 3, 7, 14, 21 and 28 of the study, or at any other time if they felt unwell. During each visit, the participants\u2019 temperatures were measured and blood smears were prepared and checked as described above. Treatment outcomes were classified according to WHO guidelines for evaluation of the therapeutic efficacy of antimalarial drugs in the treatment of uncomplicated malaria . During Data were entered into a computer database and analysed using SPSS software . Mean and SD were calculated for the variables presented. The intention-to-treat analysis included all enrolled patients who met the inclusion criteria and took at least one full dose of AL. Patients lost to follow-up or withdrawn from the study were considered to be treatment failures. The per protocol (PP) analysis of outcomes included data for patients who had completed the follow-up. Those patients lost to the follow-up or were withdrawn because of protocol violations were excluded from the PP analysis.The current study received ethical approval from the Health Research Board at the Ministry of Health in Kassala state, eastern Sudan. The trial was registered at Trial. Gov: NCT01625871.P. falciparum, uncomplicated P. vivax and mixed P. falciparum and P. vivax infections, respectively  completed the 28 days follow-up, but the remaining 5 were lost to follow- up. The base line characteristics of the study participants are shown in Table P. vivax).During the study period, 1,185 febrile patients presented to the health centre, of which 251 had uncomplicated diagnoses of malaria confirmed. Out of these 251 patients, 204, 43, and four were uncomplicated On day 1 all patients were afebrile, but 2 (5.3%) of them were parasitaemic. By day 3, all patients were afebrile and aparasitaemic. All of the patients 38 (100%) responded to treatment with AL and had ACPR. The cure rate was 100% and 88.4% for the per protocol analysis and for the intention to treat analysis, respectively. Mild adverse effects  that resolved spontaneously were observed in four (10.5%) patients.P. vivax in eastern Sudan. Consistent with a recent study, the present study showed that P. vivax is a health problem and that its ratio to P. falciparum has increased [P. falciparum malaria in eastern Sudan as well as other regions of the country [P. vivax malaria [P. vivax was first confirmed in Ethiopia [P. vivax malaria in the present study is consistent with other studies that showed a rapid clearance of P. vivax parasitaemia and fever and an overall high cure rate for AL treatment of P. vivax malaria [The current study showed full efficacy  of the AL combination for treatment of ncreased . Previou country ,21. Inte malaria . The hig malaria . In thisEthiopia . The ful malaria ,23.P. vivax infection according to WHO guidelines and it is still effective in most malaria endemic areas [P. vivax parasites have been observed in South East Asia [P. vivax malaria in Africa [P. vivax and P. falciparum is attractive, especially in settings where two malaria parasite species are co-endemic. ACT is highly effective against uncomplicated P. falciparum malaria and, as a consequence of high levels of chloroquine resistance, is now widely adopted as the first-line therapy in most malaria endemic countries, including Sudan [P. vivax and P. falciparum malaria [P. falciparum infection being treated with chloroquine.Chloroquine remains the drug of choice for treating ic areas ,25. Unfoast Asia -28. In an Africa ,22,29. Tng Sudan . Indeed, malaria . A policP. vivax malaria, this study provides useful insight into AL treatment of P. vivax infection. Secondly, the 28-day follow-up of patients treated with AL is another limitation; a longer follow-up period could be advantageous as it would provide more information about AL treatment outcomes. Thirdly, in this study, patients were treated with primaquine without prior assessment of their G6PD enzyme levels, because G6PD enzyme tests were not available.The limitations of this study concern its non-randomized character and small sample size; however, with a lack of published data about the efficacy of AL against P. vivax malaria using AL in this area of Sudan.Although this study did not detect any treatment failures during treatment of uncomplicated The authors declare that they have no competing interests.TM, GIG, IA designed the study, AAA, IRM and MB conducted the clinical work. All of the authors drafted and approved the manuscript."}
+{"text": "P = 0.002) and RR = 0.80 (P = 0.001), respectively. The treatment efficacy was similar and above 95% in all arms. Although all drugs were highly efficacious and well tolerated, AS+AQ and AS+SP were associated with less episodes of malaria.Artemisinin-based combination therapies (ACTs) are the first-line treatment of uncomplicated malaria. The public health benefit and safety of repeated administration of a given ACT are poorly studied. We conducted a randomized trial comparing artemether-lumefantrine, artesunate plus amodiaquine (AS+AQ) and artesunate plus sulfadoxine-pyrimethamine (AS+SP) in patients 6 months of age and older with uncomplicated malaria in Mali from July 2005 to July 2007. The patient received the same initial treatment of each subsequent uncomplicated malaria episode except for treatment failures where quinine was used. Overall, 780 patients were included. Patients in the AS+AQ and AS+SP arms had significantly less risk of having malaria episodes; risk ratio (RR) = 0.84 ( Plasmodium falciparum to monotherapies, and to improve treatment outcome, WHO recommends that artemisinin-based combination therapies (ACTs) be used for the treatment of uncomplicated P. falciparum malaria.2According to the World health Organization (WHO) 2010 report, in the 106 malaria-endemic countries, 225 million cases of malaria led to an estimate of 781,000 deaths in 2009.The choice of the ACTs in sub-Saharan African countries was mostly based on the efficacy and safety data of a single malaria episode treatment that does not take into account the long-term impact on malaria incidence and safety, which may have a given ACT when used over and over to treat consecutive malaria episodes.In addition, as the first generation ACTs are being rolled out, studies show that they may rapidly increase the prevalence of molecular markers associated with resistance to the partner drugs, i.e., lumefantrine, amodiaquine, or sulfadoxine-pyrimethamine.7Although data are available for the use of ACTs during single-episode treatment,17To address these gaps this study aimed to evaluate the public health impact in terms of incidence of clinical episodes and the efficacy and safety of artemether-lumefantrine (AL), artesunate plus amodiaquine (AS+AQ), and artesunate plus sulfadoxine-pyrimethamine (AS+SP) when used repeatedly for consecutive clinical malaria episodes during as long as 24 months.Plasmodium falciparum is hyperendemic with seasonal peaks that occur during the rainy season (June\u2013November).The study was conducted from July 2005 to July 2007 in Bougoula-Hameau, a peri-urban village of \u223c5,000 inhabitants located in the South of Mali. This was an open-label, randomized clinical trial comparing three different ACTs: AL, AS+AQ, and AS+SP.Plasmodium sp. infection with a parasite density between 2,000 and 200,000 asexual forms per microliter of blood for the first episode. The first subject was enrolled in July 2005 and the enrollment completed with the last subject enrolled 30 October 2006, although the last malaria episode follow-up ended in July 2007. Subjects with the following characteristics were not included in the study: the presence of severe or complicated malariaWhen malaria was suspected finger-prick blood was taken for thick blood films. After ensuring that inclusion criteria are met, a written informed consent was obtained. Patients were included if they were at least 6 months of age, weighed \u2265 5 kg, resided in the study village, were able to receive oral treatment, had an axillary temperature \u2265 37.5\u00b0C, and had Eligible subjects were randomized to receive either AL or AS+AQ or AS+SP, and followed up for 28 days after the treatment initiation with one of the three ACTs, according to the 2003 WHOPlasmodium sp. infection , they were re-treated with that same treatment regimen (except in case of parasite recurrence cases occurring within each 28-day treatment follow-up period where quinine was used). Patients were closely followed both clinically and biologically to record any adverse event. Patients with > 200,000 parasites/\u03bcL or who had treatment failure during the 28-day follow-up period or who developed severe malaria were treated with quinine and/or hospitalized if necessary. The study team was present 24 hours a day, 7 days a week at the study site during the study period.Once subjects were assigned to a given group, in the event of subsequent uncomplicated malaria episodes defined as the presence of malaria signs or symptoms plus Enrolled patients were asked to return for clinical and/or biological follow-up on Days 1, 2, 3, 7, 14, 21, 28, or on days of recurrent illness. Patients or guardians were asked about any medicine consumption outside the research facility since the last clinic visit. At each visit, a physical examination, including axillary temperature, was performed and blood was taken for thick smears. Patients were asked to return to the clinic any time if they became ill.Thick blood films and blood onto filter paper samples were made on Days 0, 2, 3, 7, 14, 21, 28, and on any days of recurrent illness. Parasitemia were measured by counting the number of asexual parasites against 300 leukocytes on the thick blood film, and then reported to a microliter of blood based on a count of 7,500 leukocytes/\u03bcL.Blood blotted onto filter-paper samples were obtained for molecular analyses of drug resistance markers and to distinguish recrudescent from re-infections.Hematology (hemogramme) and biochemistry  analysis were performed on Day 0, 7, and also on any other day if deemed clinically necessary to assess the biological toxicity of the study drugs. Normal values were based on laboratory parameters obtained from Malian population (our unpublished data). The values outside the normal ranges were considered abnormal and graded.MSP1, MSP2, and CA1 polymorphisms were analyzed using standard methods with polymerase chain reaction (PCR) to discriminate true recrudescent parasites from new infections.7Plasmodium sp. infection, which is defined as a positive blood smear with parasite density < 200, 000 parasites/\u03bcL) with the same study treatment arm. Quinine was chosen because it was the second-line drug for malaria treatment.Severe malaria cases and parasite recurrence cases occurring within each 28-day treatment follow-up period were treated with quinine (8 bmg/kg three times daily for 7 days) and re-enrolled (after complete remission) for the subsequent uncomplicated malaria , given once daily for 3 days to the following scheme: for patients < 10 kg, 1/2 tablet AS + 1/2 tablet AQ; 10\u201320 kg, 1 tablet AS+ 1 tablet AQ; 21\u201340 kg, 2 tablets AS+ 2 tablets AQ; > 40 kg, 4 tablets AS+ 4 tablets AQ.The AS+SP was Arsumax  + sulfadoxine\u2013pyrimethamine , given once daily for 3 days to the following scheme: for patients \u2264 10 kg, 1/2 tablet AS+ 1/2 tablet SP; 11\u201320 kg, 1 tablet AS+ 1 tablet SP; 21\u201340 kg, 2 tablets AS+ 2 tablets SP; > 40 kg, 4 tablets AS+ 3 tablets SP. The SP is given only the first day, whereas AS is given over 3 days.The AL was Coartem , given twice daily for 3 days to the following scheme: for patients < 15 kg, 1 tablet; 15\u201324 kg, 2 tablets; 25\u201334 kg, 3 tablets; \u2265 35 kg, 4 tablets.The study protocol was reviewed and approved by the Ethical Committee of the Faculty of Medicine, Pharmacy and Dentistry, University of Bamako, Mali. Community permission was obtained before the start of the study as described by Diallo and others.Calculation of sample size was based on the assumptions that patients experienced an average of two clinical episodes per person per year if treated by AL and living in the study areas and that treatment with AS+AQ or AS+SP would reduce this value by 35%, which is 1.3 clinical episodes per person per year. With an error risk alpha set at 0.05, a power of 90%, 174 subjects were needed in each arm. If one takes into account loss of follow-up and data attrition, 260 subjects were needed in each arm .Data were double entered and validated using Microsoft Access  and then analyzed using Stata software version 10.0 .2 or Fisher's test if appropriate was performed to compare categorical variables. Normally distributed continuous variables were compared with analysis of variance. Nonparametric tests were used when appropriate.For univariate analysis, \u03c7Parasite recurrence occurring within each 28-day treatment follow-up was counted as a new malaria episode because it was associated with full malaria rescue treatment regimen. However, following 14 days after each malaria rescue treatment with quinine, the subject was not considered at risk of malaria, therefore that time was deducted in the subject's total length of time. Negative binomial regression analysis adjusted for repeated events for the same subject was used to estimate relative risk between study drugs.The incidence rate is defined as number of malaria episodes in each study arm divided by the time length in days of each participant (person-time) and then expressed in a year (by dividing by 365.25 days).An intention-to-treat (ITT) analysis for drug efficacy was based on all subjects randomized to their respective treatment arm.A per-protocol (PP) analysis for drug efficacy was also performed and excluded missing  data and major protocol violation cases.For safety data, all subjects who received at least one dose starting from their first episode of malaria were included into the safety analysis.The trial profile is summarized in P. falciparum, and the prevalence of gametocyte carriage were all similar in all three treatment arms. Only two cases of Plasmodium ovale and no case of Plasmodium malariae monospecific infection was detected, whereas 8% (N = 780) of P. falciparum + P. malariae and 0.8% (N = 780) of P. falciparum + P. ovale mixed infection were found (data not shown).At enrollment , the meaP = 0.002) and RR = 0.80 (P = 0.001), respectively. Therefore, compared with AL the risk of malaria incidence reduction was 16% and 20% for AS+AQ and AS+SP, respectively. Using the age group \u2265 15 years of age as reference (P = 0.002), RR = 2.44 (P = 0.001), and RR = 2.76 (P = 0.001), respectively.In multivariate modeling , patienteference , patientN = 665), 78.5% (N = 619), and 89.1% (N = 678) in AL, AS+AQ, and AS+SP arms, respectively, P < 0.001).Non-PCR-corrected ACPR was significantly different between the study arms , followed by abdominal pain and headaches  were documented and included 2 severe cases of anemia ; 2 cases of seizure ; 1 case of severe malaria (in AL arm); 1 case of coma caused by hypoglycemia (in AS+SP); 1 case of presumed pneumopathy (in AS+SP arm); 2 cases of death in AS+SP arm (one caused by pneumonia and the other to hypoglycemia). The occurrence of SAE was comparable in the three arms. All of the above non-fatal SAE resolved without sequelae. The two cases of death happened in the AS+SP arm, although a causal relationship with the study product was not established. All SAEs were assessed as not related to the study drugs. There was no significant laboratory abnormalities  related to any of the study treatment drugs .in vitro studies conducted during the same period in Mali did not show any lumefantrine resistance.22Over 24 months of follow-up of this study show that the incidence of uncomplicated malaria was significantly lower in the AS+SP and AS+AQ arms than in the AL arm. This could be explained by the lowest re-infection rate seen in the AS+SP and AS+AQ arms and maybe because of the longer half life of the partner drugsOur results are in contrast with the result of a study conducted in Ghanaian children2004 where ACPRs rates of 100% and 99.1% were found with AS+SP and AS+AQ, respectively.24The high PCR-corrected 28-day follow-up treatment efficacy results are consistent with reports from the same area inP < 0.001). This high reinfection rate with AL is in contrast with studies in UgandaThe re-infection rate was higher in the AL arm as compared with AS+AQ and AS+SP arms (Our participants received quinine as a second-line treatment. However, it is unclear whether quinine, the same ACT, or a different ACT would be the optimal treatment. Given that nearly all recurrent parasitemia were caused by new infections, it might have been reasonable to retreat with the same ACT regimen, rather than with quinine.The repeated treatment using the same ACT in this study showed a good safety profile. This finding is consistent with other studies that also reported a good safety profile of repeated treatment of ACT,Limitations of this study include the consideration of asymptomatic parasite carriers from Day 7 during each 28-day treatment follow-up as treatment failures (as in the WHO treatment assessment guidelinesWe show that ACTs that had very high and similar efficacy and safety by the standard 28-day drug efficacy test could have a different impact on malaria incidence over a longer period. In this era of high efficacy of ACTs, the choice of a first-line treatment policy should go beyond short-term efficacy and safety to include longer follow-ups and repetitive treatments with the same regimen. Additional studies will be required to assess the overall impact of the large-scale deployment of ACTs on malaria in the field."}
+{"text": "P. falciparum/P. vivax infections are common in southeast Asia. When patients with P. falciparum malaria are treated and followed for several weeks, a significant proportion will develop P. vivax malaria. In a combined analysis of 243 patients recruited to two malaria treatment trials in western Cambodia, 20/43 (47%) of those with P. falciparum gametocytes on admission developed P. vivax malaria by Day 28 of follow-up. The presence of Pf gametocytes on an initial blood smear was associated with a 3.5-fold greater rate of vivax parasitemia post-treatment . The increased rate of post-treatment P. vivax infection persisted when correlates of exposure and immunity such as a history of malaria, male gender, and age were controlled for . Polymerase chain reaction (PCR) confirmed that only a low proportion of subjects (5/55 or 9.1%) who developed vivax during follow-up had detectable Pv parasites in the peripheral blood at baseline. Molecular detection of falciparum gametocytes by reverse transcriptase PCR in a subset of patients strengthened the observed association, while PCR detection of Pv parasitemia at follow-up was similar to microscopy results. These findings suggest that the majority of vivax infections arising after treatment of falciparum malaria originate from relapsing liver-stage parasites. In settings such as western Cambodia, the presence of both sexual and asexual forms of P. falciparum on blood smear at presentation with acute falciparum malaria serves as a marker for possible occult P. vivax coinfection and subsequent relapse. These patients may benefit from empiric treatment with an 8-aminoquinolone such as primaquine.Mixed  P. falciparum and P. vivax in South and Southeast Asia, Oceania, and parts of South America, mixed infections of the two species were rarely reported in the past, with cross-sectional prevalence rates reported at <5% P. falciparum/P. vivaxP. falciparum mono-infection developed P. vivax infection within 28 days of treatment with short half-life antimalarials such as artesunate P. vivax malaria post-treatment suggested a relapse from a previously unappreciated vivax infection rather than new infection.Despite prevalence of both P. vivax infection post-treatment, especially when follow-up is continued beyond the period when the antimalarials used are expected to maintain efficacious drug levels in the bloodstream P. falciparum monoinfection by peripheral smear developed P. vivax malaria during the 63-day follow-up period all patients with microscopically confirmed malaria where both species are endemic P. vivax relapse.Subsequent antimalarial trials conducted in Southeast Asia and Papua New Guinea have continued to demonstrate high rates of P. falciparum malaria were randomized in a 2\u22361 ratio to receive 7 days of artesunate (4 mg/kg/day) or quinine (30 mg/kg/day) plus tetracycline (25 mg/kg/day) for 7 days. In ARC2, patients with uncomplicated falciparum malaria were randomized to receive one of three artesunate dosing regimens: 2, 4, or 6 mg/kg/day for 7 days. In both studies, enrollment was limited to otherwise healthy adults with P. falciparum mono-infection as determined by light microscopy and with no history of antimalarial use in the 30 days prior to enrollment. The only difference in inclusion criteria between the two studies was a minimum parasite density of 1,000 parasites/\u00b5l in ARC2 compared to 100 parasites/\u00b5l in ARC1. Written informed consent was obtained from all study participants prior to enrollment. Subjects received directly observed therapy over 7 days with thick and thin blood smears prepared every 12 hours in ARC1 and every 2\u20136 hours in ARC2 until parasite clearance was achieved. Both studies recorded temperature every 4 hours, with fever clearance defined as the first time the patient became afebrile with absence of fever for the next 24 hours. In ARC1, patients remained in a study ward for 21 days to prevent reinfection, with weekly clinical and blood smear assessments on days 7, 14 and 21, and then returned for one final outpatient follow-up on Day 28. In ARC2, subjects remained on the study ward for the 7 days of therapy, then returned weekly until Day 42 for outpatient follow-up. Further details of these studies have been previously published Between 2006\u20132009, two open-label clinical trials were conducted at the same site in Tasanh, western Cambodia to investigate reports of emerging artemisinin resistance. In ARC1, patients with uncomplicated Giemsa-stained slides were examined by two microscopists blinded to each other's results and to the treatment status of the study subject. Counts were reported as the number of asexual parasites and gametocytes per 200 white blood cells seen on the thick smear. If the thick film exceeded 500 parasites per 200 WBCs, parasites were counted per 2000 RBCs (ARC1) or per 5000 RBCs (ARC2). At least 200 oil immersion fields were examined on the thick film before a blood smear was considered negative. The final count was determined by taking the geometric mean of the two microscopists' counts. Discordant results were resolved by a third reference microscopist.t value of 37 cycles was used as the threshold cutoff for positivity in the Pv assay. Using this Ct value, it was previously determined that the assay has a limit of detection for P. vivax of 0.5\u20135 parasites/\u00b5l.Molecular identification of parasite species was performed on all admission blood samples (Day 0) from both studies and on weekly blood samples available in the ARC2 patients to assess for subpatent mixed infection at baseline and occult parasitemia during follow-up. This was done using real-time PCR assays targeting the 18srRNA gene . Briefly, DNA extracted from 200 \u00b5Ls of EDTA blood using the QIAamp\u00ae DNA blood mini kit was diluted down to a working concentration of 3 ng/\u00b5l. Primers and FAM-labeled MGB probes previously adapted from published protocols were used for a pan-species Plasmodium assay, a falciparum-specific assay, and a vivax-specific assay Reverse transcriptase PCR (RT-PCR) based on the mature gametocyte marker Pfs25 was used to assess for subpatent carriage of falciparum gametocytes in admission blood samples from the patients enrolled in ARC2. RNA purified from 2.5 mLs of whole blood using the Qiagen PAXgene Blood RNA kit was diluted down to a working concentration of 50 ng/\u00b5l. cDNA synthesis and nested PCR were then carried out with the Qiagen Omniscript\u00ae RT kit and Qiagen HotStarTaq\u00ae PCR kit and primers following a previously published protocol P. vivax infection rates and risk factors for the development of post-treatment vivax malaria. Data was entered into Microsoft Excel and analyzed with STATA 10.1  and SPSS 18 . Clinical characteristics of the patients enrolled in ARC1 vs. ARC2 as well as those with and without gametocytes on admission were compared using Pearson Chi-Square, Mann-Whitney U, Fisher's exact test, or student's t-tests as appropriate. Kaplan Meier analysis was used to visually compare the incidence of Pv infection over time. Pv incidence rate (Pv IR) was selected as the measure of frequency to account for the different lengths of person follow-up between the ARC trials. Pv IR was calculated as the number of Pv episodes among any patient over the number of days until a Pv episode or the end of follow-up. Pv incidence rates were analyzed by ARC study, study treatment, and clinical variables of interest. Patients who did not complete study follow-up were not included in the analysis.Microscopic and clinical data from the two studies were combined and retrospectively analyzed to determine A multivariate analysis was conducted to estimate the unbiased association of gametocytemia on admission and post-treatment vivax infection. Covariates for the model were selected based on the univariate analysis and through prior knowledge regarding the association between gametocytemia and Pv relapse. A Poisson model with robust standard errors was constructed and the selection of final covariates was determined through a backwards elimination process to capture joint effects. Confounding was assessed by examining the change in estimate for the exposure when each covariate was individually removed. If the removal of a covariate changed the estimate by at least 10% it was considered an important confounder and the variable was retained. Covariates that were examined included gametocyte carriage at admit, study (ARC1 vs. ARC2), presence of anemia, age, male gender, parasite density, history of malaria, parasite clearance time, fever clearance time, white blood cell count, and duration of symptoms.Ethics approval for this study was obtained from the ethics committees of the Walter Reed Army Institute of Research  and the National Ethics Committee for Health Research .P. falciparum malaria were enrolled in the ARC1 and ARC2 trials, respectively. Four and 7 patients were withdrawn or lost to follow-up in the respective studies, leaving a total of 243 patients who completed 28 or 42-day follow-up that were included in the analysis.Between October 2006\u2013February 2007 and August 2008\u2013July 2009, 111 and 143 patients with Clinical characteristics of the two study populations differed in several ways . Due to Falciparum gametocyte carriage at admission was similar in both studies . In the Molecular detection of Pf gametocytes by RT-PCR for those patients enrolled in ARC2 showed good concordance with microscopy . All butIn the combined data, risk factors associated with baseline patent gametocytemia by univariate analysis included longer duration of symptoms, a greater number of malaria episodes in the previous 12 months, lower baseline hematocrit, lower baseline temperature, and a higher baseline platelet count . GametocP. vivax infection during follow-up, 32 by Day 28 in ARC1, and 27 by Day 42 in ARC2  had detectable vivax parasitemia by PCR at Day 0, suggesting that the majority did not have mixed Pf/Pv blood-stage infection at presentation that was missed by microscopy (59/243 (24%) patients developed  in ARC2 . Of thescroscopy . When PCcroscopy .The vivax incidence rate (IR) was calculated to account for the different durations of follow-up between the two studies. In ARC1, Pv IR was 331 per 1000 patient months of follow-up. This was more than double the ARC2 Pv IR of 148 per 1000 patient months of follow-up . Just coOn combined univariate analysis, the most prominent risk factors for the development of post-treatment vivax infection were the presence of falciparum gametocytes on peripheral smear and a shorter fever clearance time . While tP. vivax within 28 days compared to 27/200 (14%) patients without admission Pf gametocytemia who also relapsed. Those with smear-detectable baseline gametocytemia demonstrated a 3.5-fold increased incidence rate of vivax malaria post-treatment  (P. vivax at admission (n\u200a=\u200a16) or no baseline samples available for analysis (n\u200a=\u200a14) were excluded , as well as when Pv parasitemia by PCR was used as the marker of relapse rather than smear positivity. Finally, the association remained statistically significant regardless of trial and drug treatment allocation (data not shown).In total, of the 43/243 (18%) patients who carried smear-detectable falciparum gametocytes at admission, nearly half  had a malaria relapse with p<0.001) . If usedOther variables associated with post-treatment vivax infection in the univariate analysis included a history of malaria in the previous 12 months, male gender, a lower baseline parasitemia, and a higher baseline platelet count.In the multivariate analysis, each of the major risk factors identified in univariate analysis as well as other potential confounding variables, including those that could lead to greater gametocyte carriage, were adjusted for as covariates. After removing the bias from these confounding factors, baseline gametocytemia was associated with a 3.0-fold increased rate of relapse with vivax malaria  . Male geP. falciparum and P. vivax to show that falciparum gametocytes seen at presentation in patients with P. falciparum malaria is associated with subsequent relapse with P. vivax malaria. The finding is robust despite differences in the two study populations, including a higher Pv incidence rate in ARC1 which may reflect changing malaria epidemiology in this region. When baseline subpatent gametocytemia was detected using Pfs25 RT-PCR, the association was strengthened. Gametocyte carriage continued to be an independent risk factor even after controlling for potential confounding variables related to exposure and immunity. Prior inadequate drug treatment is another potential confounder that may have contributed to high gametocyte carriage rates in those who relapsed. However, antimalarial use in the previous 30 days was an exclusion criterion in both studies, and an ex vivo anti P. falciparum bioassay, conducted on all baseline ARC2 samples, failed to detect anti-falciparum activity in plasma collected from any of the patients who developed post-treatment vivax infection This analysis uses clinical, microscopic, and molecular data from two malaria treatment trials conducted in an area co-endemic for The low proportion of patients with PCR-detectable Pv at baseline and the excellent efficacy of the study drugs against Pv blood stages P. vivax infection in patients with occult mixed infection. We offer two possible explanations for why this might be the case. First, the presence of Pf gametocytes may indicate increased malaria exposure in general and resultant higher levels of acquired immunity. In this case, patients carrying gametocytes may be more likely to have acquired a Pv infection in the past and to harbor hypnozoites in their liver when they are infected with Pf. The theory that repeated exposure may lead to heightened acquired immunity and subsequent gametocytemia is supported by previous in vitro experiments whereby falciparum parasites in culture displayed increased levels of gametocytogenesis after exposure to immune Pf sera or anti-Pf antibodies Taken together, these findings suggest that falciparum gametocytes on a blood smear at presentation may be a marker for liver-stage P. falciparum/P. malariae infections. In a review of Boyd's early malariotherapy studies in which neurosyphilis patients were experimentally inoculated with malaria sporozoites, McKenzie et al. found that those inoculated with P. falciparum following P. malariae inoculation developed 3\u20134 fold higher falciparum gametocyte densities and gametocytes that appeared 4 days earlier than in their counterparts with single species P. falciparum infection P. falciparum/P. malariae infection demonstrated higher gametocyte densities over 28 days with an area under the curve (AUC) more than twice that of those with P. falciparum mono-infection A second explanation is that competition from a second malaria species may boost falciparum gametocytogenesis as an evolutionary adaptation. In this scenario, competition from a second co-circulating species or immune stress induced by the presence of another species may induce falciparum parasites to divert from the asexual to the sexual life cycle, accelerating transmission to new hosts. This theory is supported by observations from mixed P. falciparum and P. vivax sporozoites and followed daily blood smears The natural history of mixed infections is characterized by a pattern of alternating dominance of the two species, where often only one species is patent despite an underlying mixed infection. This pattern was evident when Boyd simultaneously inoculated two malaria-na\u00efve patients with Alternatively, these results could lead to the conclusion that falciparum gametocytes directly trigger reactivation of vivax hypnozoites in the liver. However, in the Myanmar ACT study, a single dose of primaquine, which was very effective at decreasing post-treatment gametocyte carriage, did not seem to have an effect on the post-treatment Pv incidence rate Plasmodium infections as well.A recent study by Douglas et al., based on 15 years of clinical trial data from the Thai-Burmese border, reports a similar association in falciparum-infected patients who relapsed with vivax malaria. Though the reported effect size was smaller , those with Pf gametocytemia at enrollment still represented the single group with the highest risk of Pv recurrence (41% by day 63) according to baseline risk factors P. vivax coinfection in gametocytemic individuals further highlights the potential public health gains that could be achieved with more widespread G6PD testing P. vivax malaria, arguably the harder species to eliminate because of its propensity for relapse.Our study adds to the literature by providing insights into the pathophysiology of relapse in patients with occult Pf/Pv infection in southeast Asia and identifying an easily distinguished marker for patients with falciparum malaria who are at risk for vivax relapse. Recognizing the high likelihood of occult P. falciparum gametocytes on blood smear at presentation is associated with subsequent P. vivax relapse; this has important implications for malaria treatment and control strategies in settings where mixed P. falciparum/P. vivax infections are common.In summary, in Cambodian patients with acute falciparum malaria, the presence of Table S1Comparison of microscopy and PCR detection of falaciparum gametocytes at baseline in ARC2 patients.(TIFF)Click here for additional data file.Table S2Risk Factors for gametocyte carriage at admit. Values are reported as median and interquartile ranges unless otherwise specified. *Data only available for ARC2 patients . **Antimalarial activity of sera against P. falciparum lab strains in a previously published ex-vivo bioassay, used as a surrogate measure of prior use of antimalarial drugs (TIFF)Click here for additional data file.Table S3Comparison of microscopy and PCR detection of post-treatment Pv parasitemia in ARC2 patients.(TIFF)Click here for additional data file."}
+{"text": "The current guidelines for treatment of malaria include paracetamol to children with fever. No convincing evidence for the beneficial effects of this practice exists. Studies show that time to parasite clearance is significantly longer in children treated with paracetamol, which questions the policy. Whether this is of clinical importance has not been investigated.Plasmodium falciparum monoinfection and \u226520 parasites per 200 leucocytes at the Bandim Health Centre, Guinea-Bissau were randomized to receive paracetamol or placebo together with chloroquine for three days in a double blind randomized study. Temperature and symptoms were recorded twice daily during treatment and on day 3. The participants were interviewed and a malaria film taken once weekly until day 35. The data is in the form of grouped failure-times, the outcome of interest being time until parasitaemia during follow-up. Mantel-Haenszel weighted odds ratios are given. Other differences between and within the two groups have been tested using the Chi-square test and Mann-Whitney U test.Children with In the evening of the day of inclusion, the temperature was slightly, but statistically insignificant, higher in the placebo group and significantly more children complained of headache. At no other time was a significant difference in temperature or symptoms detected. However, 6 children from the placebo-group as compared to two children from the paracetamol-group were admitted to hospital with high fever and convulsions by day 3. No differences in the cumulative percentages of children with adequate clinical and parasitological response were found in the intention-to-treat analysis or in the per-protocol analysis.Fewer children had early treatment failure and the mean temperature was slightly lower in the afternoon on day 0 in the paracetamol group. However, the cumulative adequate clinical and parasitological cure rates were not significantly different during the period of study. It is doubtful whether adding paracetamol to the treatment of uncomplicated malaria in children is beneficial.NCT00137566. Fever is a common symptom of many childhood illnesses. Although the disease process may be harmful, there is no evidence that fever is harmful in itself. In fact, fever may be beneficial by enhancing the host response to infection ,2. Thereet al found a longer parasite clearance time when paracetamol was used [P. falciparum malaria were randomized to receive mechanical antipyretics  either alone or in combination with paracetamol [The role of fever in malaria is unclear. Tumour necrosis factor (TNF) is an important mediator of fever in malaria ,9, and ewas used . This fiacetamol . Time toacetamol .The current WHO guidelines on the management of fever recommend the use of paracetamol for children with a temperature of 38.5\u00b0C or above  and the The study was performed in the area of Bandim on the outskirts of Bissau, Guinea-Bissau. Parents from Bandim attending the Bandim Health Centre with children weighting more than 7.5 kg and having fever or other symptoms compatible with malaria and stating that the children had not taken any antimalarial drug during the previous week were informed of the study. Included children had a thick film examined for malaria. Children with convulsions, severe vomiting, severe anaemia, a severe concurrent infection or who for other reasons were considered in need of hospital care were not eligible.P. falciparum and 20 or more parasites per 200 leukocytes (= 800/\u03bcl assuming a leukocyte count of 8000/\u03bcl) were enrolled from June 2004 to July 2006. Children were allocated to one of six 5 kg-interval weight groups or to a group of children with a body-weight above 37.5 kg. For each weight group, numbered boxes had been prepared at random with either paracetamol tablets or undistinguishable placebo tablets corresponding to approximately 50 mg paracetamol per kg bodyweight per day for three days. The children were given study numbers consecutively within each weight-group. The first dose of paracetamol/placebo from the tablet-box corresponding to the study number of the child was given by an experienced nurse at the health centre. For logistic reasons  the mothers were then given the tablet-box containing tablets until the end of day 2 and were carefully instructed on how to give the tablets. A health worker visited the house on day 3 and asked to see the tablet-box and counted the number of remaining tablets. The randomisation numbers were kept separately at the Department of Paediatrics in Kolding, Denmark. An interim analysis was made by one of the researchers not involved in the recruitment of the patients when a total of 40 and 140 children had completed the follow-up to ensure that none of the groups had an unacceptable high rate of recurrent parasitaemia using the results of previous in-vivo chloroquine studies as guidelines [Children with mono-infection with idelines -18.\u00ae dihydrochloride  per kg bodyweight was given intramuscularly and the child was referred to hospital.Following recommendations of the National Malaria Programme at the time of the study, 25 mg/kg bodyweight of chloroquine was split into daily doses of 10, 10 and 5 mg/kg and given days 0, 1 and 2 for treatment of malaria. An experienced nurse ascertained that the tablets were swallowed and recorded the time the dose was given. If the child did not turn up as planned, a project health worker went to the house to ensure the medication. If the child vomited within 30 minutes after receiving the tablets, the dose was repeated. If the child vomited again during the next 30 minutes, a dose of 10 mg QuinimaxTablets with 160 mg chloroquine-phosphate (= 100 mg chloroquine base) were obtained from Recip SB, 12054 \u00c5rsta, Sweden. Paracetamol tablets of 500 mg and undistinguishable placebo tablets were obtained from GlaxoSmithKline, Dungarvan, Irland.The children or their mothers were asked for any symptoms twice daily on day 0, day 1 and day 2 and in the evening on day 3. At the same time, the axillary temperature was measured using an electronic thermometer.Following treatment, the children were visited once weekly until day 35 for thick and thin films. The mothers were interviewed about any medication given since the last visit, and the health worker assessed the condition of the child. During the study period, the parents were asked to bring the child to the health centre in case of any illness to ensure early detection of clinical malaria. All treatments were free during the study-period. Recrudescent infections were treated according to the national recommendations which were then sulphadoxine/pyrimethamine or, for severe cases, quinine.Before inclusion, a thick and a thin film were examined. Thick films were also examined on day 3, 7, 14, 21, 28 and 35 after initiation of the treatment and whenever a child sought medical attention at the Bandim Health Centre due to fever or other symptoms suggestive of malaria. To ensure the quality of the microscopy, 10% of the thick films were re-examined by an experienced laboratory technician. The parasites were counted per 200 leukocytes. If abundant, the number of leukocytes was counted per 500 parasites.pfmsp1) and merozoite surface protein 2 (pfmsp2) were amplified by PCR as described previously [Pfmsp1 and pfmsp2 were amplified from DNA collected on the day of inclusion and the day of failure. Recrudescent infections were defined as reappearance of at least one band from both pfmsp1 and pfmsp2 during the follow-up or the re-appearance of a band in the successfully amplified gene if either pfmsp1 or 2 failed. Re-infections were defined as those where no band re-occurred. Whenever a child received antimalarial medication elsewhere, the blood sample drawn on the last visit was analysed.Each time a thick smear was made, approximately 100 \u03bcL of blood was also put onto filter paper, dried and then stored in a separate sealed plastic bag. Approximately 50 \u03bcl of blood was cut from the filter papers and DNA was extracted using an ABI Prism 6100 Nucleic Acid Prep Station  according to the manufacturer's recommendations. Extracted DNA was frozen at -20\u00b0C until amplification by PCR. Merozoite surface protein 1 (eviously . PCR proEarly treatment failure (ETF) was defined as 1) development of severe malaria or appearance of danger signs on day 1 to 3; 2) positive malaria film and temperature \u2265 37.5\u00b0C on day 3; or 3) parasitaemia on day 3 \u2265 25% of the parasitaemia on day 0. Late clinical failure (LCF) was defined as either; 1) fever and a positive malaria film; 2) a history of fever and clinical symptoms of malaria as well as a positive malaria film. Late parasitological failure (LPF) was defined as reappearing parasiteamia on day 7 or later for children without ETF or LCF. Adequate clinical and parasitological response (ACPR) was defined as absence of parasitaemia without the child meeting any of the criteria of ETF, LCF or LPF.The primary outcome was the cumulative PCR-uncorrected ACPR rate until day 35 and PCR-corrected failure rates on day 28 and 35. Secondary outcomes were the ETF rates, the symptoms as reported by the mothers, the temperature measured during the treatment with paracetamol/placebo; and possible clinical adverse effects of the medication given.An intention-to-treat analysis was performed to answer the question as to whether the treatments were valuable for the children ,21. TherA per protocol analysis was done to evaluate the efficacy of the treatments. Children admitted to hospital on the day of inclusion were then excluded as they were considered to be so sick that they had been included in violation of the inclusion criteria. Children treated outside the study during follow up without having re-parasitaemia confirmed by a positive malaria film and/or a positive PCR-analysis were considered withdrawal of consent and excluded on the day of retreatment.The data is in the form of grouped failure-times, the outcome of interest being time until parasitaemia during follow-up. Mantel-Haenszel weighted odds ratios are given. Loss to follow-up between two analysis times has been treated by censoring at the beginning of the time interval. Other differences between and within the two groups have been tested using the Chi-square test and Mann-Whitney U test.The protocol stated that any study group should be terminated if parasitaemia reappeared in 50% or more of at least 40 children. Ethical clearance was obtained from Direc\u00e7\u00e3o de Higiene e Epidemiologia, Minist\u00e9rio da Sa\u00fade Publica in Guinea-Bissau (030/DHE/2004) and the Central Ethical Committee in Denmark (2004-7041-11). Children and/or their parents were informed of the study orally as the literacy rate is low. Written information in Creole was given on request. The information was standardised and in accordance with the principles of the Helsinki declaration. This trial was registered at ClinicalTrials.gov  with theA total of 14.861 children were screened at the Bandim Health Centre and 3.569 had a malaria film examined due to symptoms compatible with malaria. Of these, 354 had 20 or more parasites per 200 leucocytes. Six children were not included as they were leaving the study area and 5 were not included for unknown reasons. Five children or their caretakers refused to participate. The remaining 338 children who fulfilled the inclusion criteria were randomised into one of the two study groups. The number of children lost to follow-up and the number of children with reappearing parasitaemia are shown in Figure In the paracetamol and the placebo groups 59% (99/167) and 59% (100/170) of the children, respectively claimed to have taken paracetamol during the past 12 hours prior to inclusion. During the three days of treatment, only 1 (0.3%), 14 (4.1%) and 17 (5.0%) children received paracetamol or other anti-pyretic apart from what had been prescribed on inclusion. On day 3, after ending the treatment prescribed at the health centre, a total of 34 children (10.1%) were given paracetamol. There were no differences between the two study groups on any of the days.When asked to show the tablet-box on day 3, 39% (62/161) and 37% (60/161) in the paracetamol and the placebo groups respectively, still had tablets left with a median number of 1.5 (range 0.5 - 6.0) vs. 2.0 (range: 0.5 - 6.0). The rest showed empty boxes. Assuming that the children had taken the tablets removed from the boxes, the median dose during the 3 days were 136 mg paracetamol/kg (range: 0 - 188) and 143 mg placebo/kg (range: 0 - 188), respectively (p = 0.34).On inclusion, there were no differences between the groups in the symptoms and signs reported by the mothers. The most frequent symptoms and signs were that the child felt/looked ill , had fever , had eaten less , had headache , vomited , had diarrhoea (14%. 49/338), and drank less . In the evening of day 0, more children in the placebo group complained of headache than in the paracetamol group . For all other complaints, no statistical significant differences were found during the first three days (data not shown). On day one, three children in the paracetamol group and eight children in the placebo group complained of itching (p = 0.13), but continued the anti malarial treatment. Due to vomiting, 11 children in the paracetamol group and two children in the placebo group had the first dose of chloroquine repeated. The second dose was repeated in four and three children respectively. On day 3, parasites were detected in 19 and 21 of the children in the paracetamol and the placebo group, respectively ).The temperature was measured in the morning and evening of day 0, day 1 and day 2 and in the evening on day 3 ). Two children in the paracetamol group were admitted to hospital after the end of the treatment, one at day 7 and one at day 14, both due to high fever. One child in the placebo group was admitted at day 5 due to anaemia.Pfmsp 1 and 2 amplification failed in 11 samples collected on the day of treatment failure, three of these were treated outside the study. No differences in the cumulative percentages of children with ACPR were found, in the intention-to-treat analysis or in the per-protocol analysis )  children were admitted to hospital due to convulsions and high fever in the paracetamol group, though the finding was not statistically significant. Treatment of children suffering from uncomplicated malaria with paracetamol in the doses recommended did not influence the overall cure-rate, however the clinical beneficial effects are doubtful.The authors declare that they have no competing interests.PK participated in the design of the study, performed the statistical analyses and drafted the manuscript. JU performed the PCR analyses and interpreted the results, participated in preparing the manuscript. AR participated in designing the study and the daily supervision, participated in preparing the manuscript. LR participated in designing the study, participated in preparing the manuscript.All authors read and approved the final manuscript."}
+{"text": "Background.\u2003A high prevalence of chloroquine-resistant Plasmodium vivax in Indonesia has shifted first-line treatment to artemisinin-based combination therapies, combined with primaquine (PQ) for radical cure. Which combination is most effective and safe remains to be established.Methods.\u2003We conducted a prospective open-label randomized comparison of 14 days of PQ (0.25 mg base/kg) plus either artesunate-amodiaquine (AAQ + PQ) or dihydroartemisinin-piperaquine (DHP + PQ) for the treatment of uncomplicated monoinfection P. vivax malaria in North Sumatera, Indonesia. Patients were randomized and treatments were given without prior testing for G6PD status. The primary outcome was parasitological failure at day 42. Patients were followed up to 1 year.Results.\u2003Between December 2010 and April 2012, 331 patients were included. After treatment with AAQ + PQ, recurrent infection occurred in 0 of 167 patients within 42 days and in 15 of 130  within a year. With DHP + PQ, this was 1 of 164  and 13 of 143 , respectively (P > .2). Intravascular hemolysis occurred in 5 patients, of which 3 males were hemizygous for the G6PD-Mahidol mutation. Minor adverse events were more frequent with AAQ + PQ.Conclusions.\u2003In North Sumatera, Indonesia, AAQ and DHP, both combined with PQ, were effective for blood-stage parasite clearance of uncomplicated P. vivax malaria. Both treatments were safe, but DHP + PQ was better tolerated.Clinical Trials Registration.\u2003NCT01288820. Plasmodium vivax infection worldwide, of whom half live in Southeast Asia [Plasmodium falciparum malaria, P. vivax can cause relapse infections emerging from dormant hypnozoite forms in the liver. Strains in tropical regions such as Sumatera are characterized by frequent (>30%) and early (around 1 month) relapses [Approximately 2.6 billion people are at risk of acquiring ast Asia . In contrequent >0% and earelapses . The comrelapses , but whiPlasmodium vivax resistance to chloroquine is prominent in many parts of Indonesia, ranging from 43% in Sumatera island to >80% in Papua [in Papua \u20138, In 20in Papua , 10. HowP. vivax monoinfection in nonpregnant adults and children aged >1 year presenting at a rural clinic in Tanjung Leidong village, Labuhan Batu, North Sumatera, Indonesia. Routine G6PD testing is not available here. Clinical malaria incidence is 400\u2013500 per year among a population of 32 837 (in 2010), equally divided between P. vivax and P. falciparum infections .We performed a prospective, open-label, randomized study comparing AAQ + PQ and DHP + PQ for the treatment of uncomplicated symptomatic P. vivax monoinfection (\u2265250/\u00b5L) were eligible. Exclusion criteria included any feature of severe malaria [Patients with fever (or recent fever <48 hours) and microscopically confirmed  malaria , severe The study was approved by the Ethics Committee of the National Institute of Health Research and Development, Indonesian Ministry of Health, Jakarta, Indonesia; Faculty of Tropical Medicine, Mahidol University, Thailand; and the Oxford Tropical Research Ethics Committee, Oxford University, United Kingdom.Parasite density was assessed per 200 white blood cells on a Giemsa-stained thick film, and assumed to be absent if not detected in 200 high-power fields. Gametocytes were counted per 1000 white blood cells. Parasite species was confirmed in thin smear, and 10% of slides were cross-checked at the Faculty of Tropical Medicine, Mahidol University. Other investigations included hemoglobin measurement (Hemocue201+), hemoglobin-methemoglobinemia by pulse oximetry , and G6PD genotyping from a filter paper blood spot (Whatman 3M). Genotyping by polymerase chain reaction\u2013restriction fragment-length polymorphism (PCR-RFLP) enabled identification of 3 common mutations  . In patiPatients were not screened for G6PD status before the start of therapy and were managed as outpatients, both current practice in Sumatera. All patients were followed daily for 14 days and then weekly until 42 days, followed by monthly visits up to a year, or in between in case of symptoms. Hemoglobin levels were assessed on days 0, 2, and 7, and then weekly. During PQ therapy, methemoglobinemia was monitored daily. PQ therapy was discontinued in case of macroscopic hemoglobinuria, a drop in hemoglobin >2 g/dL, or when methemoglobin increased to >20% of total hemoglobin. At the end of the study, all patients were invited to test for G6PD status using a NADPH qualitative spot test .Patients randomized to AAQ  received artesunate 12 mg/kg and amodiaquine 30 mg/kg divided over 3 days. Patients randomized to DHP , received dihydroartemisinin 6.75 mg/kg and piperaquine 54 mg/kg in divided doses over 3 days. All patients also received PQ (Phapros Inc) in a dose of 0.25 mg base/kg (or 15 mg for >40 kg) for 14 days started on the first day. All treatment doses were given directly observed and together with some biscuits . If the patient vomited within 30 minutes, the dose was repeated. Recurrent vivax malaria infections occurring in the first 42 days of follow-up were treated with quinine/doxycycline following Indonesian guidelines; episodes occurring after this point were treated with the same regimen as the initial treatment. All patients were provided with insecticide-treated bednets.Patients were randomized by an independent statistician in blocks of 10, with each treatment allocation concealed in an opaque, sealed envelope, opened only after enrollment.P. vivax infection during 1-year follow-up, fever and parasitemia clearance times, gametocyte carriage rates and clearance times, hematological recovery, and safety and tolerability of treatments.Patient outcomes, including early treatment failure, late treatment failure, and adequate clinical and parasitological response, were classified according to World Health Organization guidelines . The priIncluding a 10% anticipated loss, a sample size of 165 patients per study arm was calculated to detect a difference in 42-day cure rate of 90% with AAQ + PQ vs 98% with DHP + PQ with 95% confidence and 80% power.U test, Student t test, \u03c72 test, and Fisher exact test where appropriate. Efficacy at 42 days and after 1 year of follow-up were assessed by Kaplan\u2013Meier survival analysis with log-rank test for statistical significance.Data were anonymized and double entered into a secured database (OpenClinica). Analysis was done using Stata software (StataCorp). The primary intention-to-treat analysis included all randomized patients and per-protocol analysis of all patients who completed 42 days of follow-up. Comparisons between groups were made by Mann\u2013Whitney Between December 2010 and April 2012, 3168 patients were screened, of whom 331 were enrolled in the study. A total of 167 patients were treated with AAQ + PQ and 164 with DHP + PQ . Per-protocol analysis of patients with complete 42-day follow-up showed cure rates of 100%  with AAQ + PQ and 99.3%  with DHP + PQ (P = .31). Parasite clearance was within 48 hours in both treatment arms, except for 1 patient with early treatment failure after DHP + PQ (who received rescue treatment) and another 2 patients after DHP + PQ who cleared parasites after >72 hours; neither showed recurrent infection during follow-up. No late treatment failures until day 42 were found in either treatment group. During 1-year follow-up, recurrent infections were observed in 15 of 130 (11.5%) patients after AAQ + PQ (of whom 2 had a second recurrent P. vivax infection) and 13 of 143 (9.1%) after DHP + PQ . The earliest recurrence after treatment with AAQ + PQ was at day 54 compared to 83 days after DHP + PQ. After 1 year, the mean day of recurrence was day 165  for patients treated with AAQ + PQ and day 203  for those treated with DHP + PQ (P = .23). Among 28 patients with recurrent infections, 24 had monoinfection with P. vivax, 2 had monoinfection with P. falciparum, and 2 had mixed infection . Cumulative risk of recurrence for the total group during the 1-year follow-up period was 17.5 per 100 person-years.Intention-to-treat survival analysis showed an adequate parasitological cure rate at 42 days of 91%  with AAQ + PQ and 94%  with DHP + PQ . In patients presenting with gametocytemia, 55 of 67 (82%) of patients treated with AAQ + PQ and 63 of 74 (85%) with DHP + PQ cleared gametocytemia within day 1 (P = .63), and all patients cleared gametocytemia by day 2. At day 42, the mean hemoglobin was 11.9 g/dL  with DHP + PQ vs 11.9 g/dL  with AAQ + PQ (P = .91). Hemoglobin levels did not differ between treatment arms at any time point.On admission, 92 of 167 (55.1%) patients in the AAQ + PQ arm and 96 of 164 (58.5%) in the DHP + PQ arm had fever (\u226537.5\u00b0C). All patients treated with DHP + PQ cleared their fever within 1 day, compared to 89 of 92 (97%) with AAQ + PQ (P = .22).In patients treated with AAQ + PQ, 3 had a drop in hemoglobin level >2 g/dL , of whom 2 developed cola-colored urine temporarily without other complications. One patient had an increased methemoglobin level of 20.3%, after which PQ was discontinued. One patient developed a generalized urticarial rash half an hour after the first dose of AAQ + PQ. This patient recovered after treatment with an antihistamine and was subsequently treated with quinine/doxycycline. In patients treated with DHP + PQ, 1 male and 1 female patient had a drop in hemoglobin level >2 g/dL  and 2 had increased methemoglobin levels to 20.2% and 21.6% respectively, after which PQ was discontinued. None of the patients with intravascular hemolysis needed blood transfusion, and hemoglobin levels returned to normal (>10 g/dL) after a median of 14  days. An increase of >10% in methemoglobin level occurred in 17 of 167 (10.2%) patients treated with AAQ + PQ compared to 24 of 164 (14.6%) treated with DHP + PQ  patients were screened for G6PD status by fluorescence spot test. Two males and 5 females (2.6%) were G6PD deficient according to the screening test. The median reduction in hemoglobin levels in these patients was 1.4 g/dL . Gene sequencing showed that 1 male patient was hemizygous for the Mahidol variant and another male carried the 1311C\u2192T intron 11 nt93T\u2192C mutation. One of the 5 females was heterozygous for the C 1311 T/C intron 11 nt 93 T/C and intron 2 nt 8 C/A mutations, whereas the other 4 had wild-type genotype  adverse events than AAQ + PQ, as has also been reported in other studies , 24. In This study has several limitations: 12% of patients were lost for follow-up at day 42, related to poor accessibility of some areas in rural northern Sumatera, and 22% were not tested for G6PD status at the end of the study, so our prevalence estimate may be imprecise. Patients with hemolysis were not formally assessed for changes in renal function, but no patient reported anuria or developed symptoms of renal failure during follow-up.The number of G6PD-deficient patients in the current study was low, and because enzyme activity can vary considerably even within specific genotypes, assessment of the hemolysis risk after low-dose PQ within specific genotypes requires larger studies. Further prevalence studies on the genetic variants of G6PD and their corresponding phenotypes in various parts of Indonesia will be required to generalize our current findings to other parts of Indonesia.P. vivax in North Sumatera. DHP + PQ was better tolerated and had a longer posttherapeutic prophylactic effect.In conclusion, radical treatment with AAQ or DHP, both combined with low-dose PQ (0.25 mg/kg for 14 days), without prior testing for G6PD deficiency proved a safe and efficacious treatment for uncomplicated"}
+{"text": "Plasmodium vivax malaria in people living in endemic countries.This study aimed to synthesize available evidence on the efficacy of dihydroartemisinin-piperaquine (DHP) in treating uncomplicated I2: 0%) or artemether-lumefentrine (AL) . On the basis of GRADE criteria, further research is likely to have an important impact on our confidence in the estimate of effect and may change the estimate.This is a meta-analysis of randomized controlled trials (RCT). We searched relevant studies in electronic databases up to May 2013. RCTs comparing efficacy of (DHP) with other artemisinin-based combination therapy (ACT), non-ACT or placebo were selected. The primary endpoint was efficacy expressed as PCR-corrected parasitological failure. Efficacy was pooled by hazard ratio (HR) and 95% CI, if studies reported time-to-event outcomes by the Kaplan-Meier method or data available for calculation of HR Nine RCTs with 14 datasets were included in the quantitative analysis. Overall, most of the studies were of high quality. Only a few studies compared with the same antimalarial drugs and reported the outcomes of the same follow-up duration, which created some difficulties in pooling of outcome data. We found the superiority of DHP over chloroquine (CQ)  are taken together. However, DHP is not active against the hypnozoite stage of P. vivax. DHP has the potential to become an alternative antimalarial drug for the treatment uncomplicated P. vivax malaria. This should be substantiated by future RCTs with other ACTs. Additional work is required to establish how best to combine this treatment with appropriate antirelapse therapy (primaquine or other drugs under development).Findings document that DHP is more efficacious than CQ and AL in treating uncomplicated  Plasmodium vivax accounts for up to 50% of malaria cases with prevalence rates between 1% and 6% of the population in South and South East Asia, where the majority of P. vivax malaria occurs. In Central and South America and Eastern and Southern Africa, it accounts for 71-81% and 10% of malaria cases, respectively [P. vivax malaria has a reputation of being a benign infection, severe and fatal complications also occur [P. vivax was first documented in 1989 among Australians repatriated from Papua New Guinea [P. vivax drug sensitivity. Early diagnosis followed by prompt and effective treatment remains a cornerstone for the reduction of malaria-related morbidity and mortality [P. vivax are needed [P. falciparum malaria [According to a recent estimate, ectively . Althougso occur  such as w Guinea . Since te needed . In the  malaria . The art malaria . The abi malaria ,14 resul malaria ,16. This malaria ,14,17.falciparum malaria [P. falciparum per se [falciparum malaria, there has been a surge of published RCTs undertaken in endemic countries to compare DHP with other antimalarial agents for the treatment of vivax malaria. As the epidemiology of malaria is complex and heterogeneous, with variations over small areas [A previous review of 14 trials solely from the Asian region has reported that DHP is safe and highly effective for treatment of uncomplicated  malaria . A Cochr malaria  assessinm per se . Since tll areas  as well ll areas , informaP. vivax malaria in people living in malaria-endemic countries.Taken as a whole, the objective of the present review was to synthesize the available evidence assessing the efficacy of DHP in treating uncomplicated http://www.clinicaltrials.gov, http://www.controlled-trials.com, andhttp://www.nci.nih.gov/clinicaltrials. Furthermore, we manually searched the reference sections of the selected studies and relevant reviews to look for any additional studies which were not found in the initial search. Searches were limited to English language and those with human participants. The search terms we used were malaria, vivax, treatment, dihydroartemisinin, piperaquine, dihydroartemisinin-piperaquine, Arteken, efficacy, treatment success, treatment failure, safety, tolerability, resistance. We determined the inclusion criteria following the PICO format; (1) Participants (P): those having confirmed P. vivax  mono-infection at enrollment, regardless of age and pregnancy status; (2) Interventions (I): RCTs in which participants in one arm should use fixed-dose coformulated DHP; (3) Comparisons (C): the efficacy of DHP with ACT antimalarial(s), non-ACT antimalarial(s), or placebo, (4)\u00a0Outcomes (O): the proportion of patients with parasitaemia and provided the effect estimates  relative risk (RR), hazards ratio (HR) or odds ratio (OR) and their corresponding 95% confidence interval (CI). If a selected study included more than one comparator, each comparison was regarded as a separate study. We included studies with participants having mixed infection  for subgroup analysis. Studies on economic evaluation, mathematical modeling or pharmacokinetics were not included.We conducted a literature search in MEDLINE, EMBASE, CINHAL, the Cochrane Library and the database of abstract of Reviews and Effectiveness from January 1989 to May 2013. For ongoing and unpublished trials, we also looked at the websites such as In the present review, outcomes were defined as follows.1) Polymerase chain reaction (PCR) confirmed parasitological failure by day 28 after starting treatment ;2) PCR-confirmed parasitological failure by day 42 after starting treatment ; 3) PCR-confirmed parasitological failure for more than 42 days after starting treatment .1) Safety outcomes (incidence of adverse events); 2) Resolution of fever (i.e. time to fever clearance (FCT)) and 3) time to parasite clearance (PCT).An adverse event (AE) was defined as any unfavorable, unintended sign, symptom, syndrome or disease that develops or worsens with the use of a medicinal product, regardless of whether it is related to the actual medicinal product. A serious AE was defined as any untoward medical occurrence that at any dose; resulted in death; was life threatening; requiring hospitalization or prolongation of hospitalization; resulted in a persistent or significant disability or incapacity; or caused a congenital anomaly or birth defect .P. vivax infection, duration of follow up, outcomes for each included articles, using a piloted data abstraction form. Power calculation for the required sample size was also assessed, if available. If articles contained information on the same or overlapping study population, we included the study with the most complete information.Two investigators read all the titles and abstracts collected through the electronic search and filtered article(s) potentially eligible for the present study. The two investigators collected information on baseline characteristics of study design, participants, characteristics of the experimental drug, confirmation of The two investigators independently assessed risk for bias, following the procedures suggested by the Cochrane Risk of Bias tool . The domWe performed meta-analysis when 2 or more individual studies were suitable for pooling on the basis of similarity. Parasitological efficacy was compared with the rate of parasitological failure between DHP and the comparator drug. The parasitological efficacy was pooled by HR and corresponding 95% CI, if studies reported time-to-event outcomes by the Kaplan-Meier method or data available for calculation of HR, using formula described by Parmer  and TierI2 test. I2 value greater than 50% represented substantial heterogeneity [We assessed heterogeneity by chi-square test and the ogeneity . Meta-anogeneity .The protocol of the present study is available in PROSPERO (CRD: CRD 42013004625) . The metP. vivax infection detected at enrollment [P. vivax and/or mixed infections was not provided [rollment , 2) diffprovided ,41, 3) sprovided ,43, 4) oprovided  or 5), tprovided .Baseline characteristics of the included studies are presented in In the present review, DHP was compared with artesunate-mefloquine (MAS3) in two trials ,37, withDHP versus CQ: At day > 42- 63, two studies (n = 1028) [I2: 75%) in treating uncomplicated P. vivax malaria (I2: 74%). Of note is the substantial statistical heterogeneity. The results with HR indicated the higher cumulative risk of recurrence in the CQ group  ( = 1028) ,35 showe malaria . As an aI2: 0%) .DHP versus AL: At day 42, two studies (n = 837) [I2: 93%). We reanalyzed the data, using the fixed-effect model and DHP showed better efficacy than AL ; due to substantial heterogeneity, it is not ideal for the pooled estimate (I2: 39%) (n = 837) ,36 showeestimate . The res2: 39%) .We planned to stratify analyses by brand of DHP and age groups. Due to limitations in the data, we are unable to perform this analysis.Inadequate data restricted the ability to conduct a pooled analysis of FCT and PCT. An individual study  showed sP. vivax infection, pooled estimates of AE incidences were not attainable. An individual study [Due to difficulties in collecting the symptoms reported or exclusively relating to al study  showed tP. falciparum) showed a comparable efficacy between DHP and MAS3 [As a subgroup analysis, two separate studies with mixed infection . Although we planned to reanalyze the effect estimates by excluding individual studies from the meta-analysis, this was not possible due to inadequate studies. According to the GRADE criteria to interpret results, further research is likely to have an important impact on our confidence in the estimate of effect and may change the estimate  and exposure to therapeutic drug levels over many parasite cycles is an important determinant of response ,33. The stration . As all stration ,15. ThisP. vivax is dominant and there is high prevalence of G6PD deficiency such as in certain ethnic groups. For example, Afghani and Asian patients can suffer significant haemolysis due to some antimalarials. The rapid development of resistance to SP when the drug is employed on the national or regional scale is attributable to the requirement [The relatively shorter mean PCT of DHP compared with CQ-SP would result in a relatively faster initial symptomatic response, increasing the confidence of parents/guardians of children as well as adult patients regarding the new drug. This is highly important from a clinical viewpoint. Moreover, the SP component of a drug is potentially important in settings where uirement .P. vivax may consist of a mixture of relapses from dormant liver stage (hypnozoites), recrudescences of the erythrocytic infection (due to inadequate drug levels or resistance), and reinfections acquired from additional inoculations. It is not possible with current methodologies to distinguish reliably between these possibilities [P. vivax infections is uncertain [P. vivax in antimalarial drug trials has not been carried out as more than half of the parasites that caused the relapse had a different genotype from those that caused the primary infection [Of note is that recurrences of bilities . The intncertain . Therefonfection ,48. A hinfection , and trunfection .P. vivax [P. vivax in Southeast Asians has traditionally been reported to be 6 weeks [P. vivax cannot be maintained to confine in vitro testing of drugs to assays on fresh isolates. Therefore, it is difficult to document unequivocal cases of treatment failure in areas where resistance is emerging, but any P. vivax infection that occurs within 28 days after the start of CQ treatment, whether recrudescence, relapse, or new infection, has grown through residual CQ concentrations in blood. If these concentrations are adequate, then, by definition, the infection is resistant [Initial parasite clearance was significantly faster after DHP in the present study and this was consistent with the pharmacodynamic properties of ACTs observed in P. vivax . The rapP. vivax  suggest P. vivax . As such 6 weeks . There a 6 weeks -49, and esistant . Minimumesistant  was presesistant . The latesistant . Therefoesistant .P. falciparum and P. vivax were common [P. vivax infection in patients with mixed infections at baseline have merits. The delay in relapse and reinfections conferred by DHP gave patients a lengthened period without symptomatic malaria, allowing for a greater time for haematological recovery [Outside Africa, mixed infections of e common ,36. The recovery ,36 and arecovery ,36,38. IP. vivax accounts for over half of all malaria transmitted outside Africa [Of human malaria infections, e Africa . As almoP. falciparum infection [P. vivax infection. Future studies of well designed with adequate samples assessing efficacy of DHP in patients having P. vivax infection are recommended. Studies in endemic countries where P. vivax is proportionally dominant would be of great value.For highly effective treatments, it is more appropriate to show that a treatment is non-inferior or not worse than the standard treatment, i.e. that the difference in failure rate is not higher than a pre-specified non-inferiority margin . Along tnfection -20. Due P. vivax the numbers of patients who experience one or more, two or more, three or more relapses are exponential [Without radical treatment for onential . AdditioOur review has strengths. Clinically important differences in the effect of treatment may be obscured if the proportions of survivors or recovered individuals in the treatment group are simply compared to that of the control group at a single point in time, such as at the conclusion of the trial. Time-to-event analysis is, therefore, a potentially more powerful and informative method of analysis ,55. The Despite this, limitations also exist in the present study. Treatment failure attributable to \u2018genuine resistance\u2019  was not P. vivax malaria. Future RCTs with other ACTs are recommended. Additional work is required to establish how best to combine this treatment with appropriate antirelapse therapy (PQ or other drugs under development).Findings suggest that DHP is better than CQ and AL in the treatment of uncomplicated Checklist S1PRISMA checklist.(RTF)Click here for additional data file.Figure S1Summary evidence of the effect estimation on the basis of GRADE criteria.(TIF)Click here for additional data file.Table S1Baseline characteristics of the included studies.(RTF)Click here for additional data file."}
+{"text": "Plasmodium falciparum.Malaria in Zambia remains a public health and developmental challenge, affecting mostly children under five and pregnant women. In 2002, the first-line treatment for uncomplicated malaria was changed to artemether-lumefantrine (AL) that has proved to be highly efficacious against multidrug resistant P. falciparum malaria in Ndola, Zambia.The study objective was to determine whether dihydroartemisinin-piperaquine (DHA/PQP) had similar efficacy, safety and tolerability as AL for the treatment of children with uncomplicated P. falciparum were enrolled, randomized to AL (101) or DHA/PQP (203) and followed up for 42 days. Outcome of treatment was defined according to the standard WHO classification, i.e. early treatment failure (ETF), late clinical failure  and adequate clinical and parasitological response (ACPR). Recurrent infections were genotyped to distinguish between recrudescence and new infection.Between 2005 and 2006, 304 children (6-59 months old) with uncomplicated No ETF was observed. At day 28, PCR-uncorrected ACPR was 92% in the DHA/PQP and 74% in the AL arm . Most failure were new infections and PCR-corrected ACPR was similar in the two study arms . Similar results were observed for day 42, i.e. higher PCR-uncorrected ACPR for DHA/PQP, mainly due to the difference observed up to day 28, while the PCR-corrected ACPR was similar: DHA/PQP: 93% (179/192), AL: 93% (84/90), . Except for cough, more frequent in the DHA/PQP arm (p = 0.04), there were no differences between treatment arms in the occurrence of adverse events. Two serious adverse events were probably associated to AL treatment.DHA/PQP was as efficacious, safe and well tolerated in treatment of uncomplicated malaria as AL, though in the latter group more new infections during the follow up were observed. DHA/PQP seems a potential candidate to be used as an alternative first-line or rescue treatment in Zambia.ISRCTN16263443, at http://www.controlled-trials.com/isrctn In 1998, in the Western Province, parasitological resistance (RII-RIII) to CQ (7-day follow up) was 60% and that to SP 26% (14-day follow up), a figure higher than previously reported [Plasmodium falciparum malaria [P. falciparum malaria in children was carried out in Ndola. This was part of a phase III multicentre study done, besides Zambia, in four other African countries  [In Zambia, chloroquine (CQ) has been for a long time the first-line treatment of uncomplicated malaria . Reduced in 1978  and CQ r in 1978 . In 1996 in 1978 . Between Uganda) . More deThe study (randomized and open-label) was conducted between September 2005 and May 2006 in Ndola, Zambia, where four peri-urban health centres  were identified for the recruitment of the patients. The study was sponsored by the Sigma-Tau, Industrie Farmaceutiche Riunite, Italy, and funded by Medicines for Malaria Venture (MMV). In Zambia, in 2006, malaria prevalence in children under five was on average 22% . In NdolP. falciparum mono-infection with asexual parasite densities between 2,000 and 200,000/\u03bcl; fever (axillary temperature \u226537.5\u00b0C) or history of fever in the preceding 24 h. Patients were not recruited if they met at least one of the following exclusion criteria: severe malaria or other danger signs; acute malnutrition  [Children 6-59 months old attending the health facilities with suspected malaria were included if they fulfilled the following inclusion criteria: body weight >5 kg; microscopically confirmed ference) ,14 or anPatients were individually randomized according to a 2:1 (DHA/PQP:AL) scheme so as to have more patients in the DHA/PQP arm to provide better estimates for its cure rates and more cases for the integrated safety data base. A total of 304 study participants were enrolled. A randomization list was generated by an independent off site contract research organization (CRO), with each treatment allocation concealed in opaque sealed envelopes that were opened only after the patient's recruitment.Both drugs were co-formulated, fixed-dose ACTs and they were administered under direct supervision during three consecutive days, according to the patient's body weight. AL  was administered twice a day  according to the following dosage: weight 5-14 kg: one tablet per dose; weight 15-24 kg: two tablets per dose; weight 25-34 kg: three tablets per dose. DHA/PQP  was given once daily, at the standard dosage of 2.25 mg/kg and 18 mg/kg per dose of DHA and PQP, respectively, rounded up to the nearest half tablet. To facilitate the correct dosing of DHA/PQP, two formulations were used (DHA 20 mg + PPQ 160 mg and DHA 40 mg + PPQ 320 mg). In case of vomiting, a full dose was repeated if this occurred within the first half an hour or half a dose if it occurred between 30 minutes and 1 h. In agreement with the instruction of the manufacturer, AL was administered concomitantly with milk to facilitate the absorption of lumefantrine, while DHA/PQP was administered only with some water. For infants, drugs were crushed, mixed with water and administered as slurry. In order to minimize bias, treatment allocation was concealed until recruitment of the patient was completed. Both patient allocation to the different analysis populations and assessment of the primary end-point were made by staff blinded to the treatment assignment and before availability of the PCR results.http://www.controlled-trials.com/isrctn.The study was approved by a local institutional ethics committee  and the ethical and scientific committee of the Institute of Tropical Medicine, Antwerp, Belgium. The trial was conducted under the provisions of the Declaration of Helsinki (2002) and in accordance with Good Clinical Practices guidelines set up by the International Conference on Harmonization. A Study Steering Committee, a Data Monitoring Committee and a Clinical Development Committee were created prior to the beginning of the trial, and worked independently to harmonize and monitor the study. The trial was registered prior to the enrolment of the first patient in the International Standard Randomized Controlled Trials Register, number ISRCTN16263443, at All children were kept at the health facility for the three-day dosing period. The mother/guardian was asked to return with the child for scheduled visits on days 7, 14, 21, 28, 35 and 42 post-treatment, or if any symptoms occurred. Field workers traced patients missing any visit. For each visit, a physical examination was performed by the study clinicians, vital signs were recorded and axillary temperature measured with an electronic thermometer. Adverse events and serious adverse events were recorded and monitored throughout the study.The study was monitored by an external CRO who carried out visits on a monthly basis. Rescue treatment for recurrent parasitaemia was quinine 10 mg/kg orally three times a day for 7 days. All participants received a free insecticide-treated bed net at recruitment.Capillary or venous blood was taken at every visit. Thick and thin blood films were prepared, dried and Giemsa-stained, and parasite density estimated by counting the number of asexual parasites in 200 white blood cells (WBC), assuming a standard WBC count of 8,000/\u03bcl. In addition, quality control was performed on 20% of all the slides at a central laboratory. Samples for haematology (full blood count) and biochemistry  were taken at enrolment, at days 3, 28 and 42, and at any other visit if judged necessary by the clinician. For PCR analysis, three blood spots were collected on filter paper (Whatmann 3 MM) at enrolment and at any visit after day 7. Each filter paper was dried and individually stored in a plastic bag containing silica gel. All filter papers were subsequently transferred to the Institute of Tropical Medicine  where centralized genotyping was conducted. Purification of DNA was conducted as previously described . Three pTreatment outcome was established according to standard WHO classification : Early TThe primary endpoint was the PCR-corrected adequate clinical and parasitological response (ACPR) at day 28; secondary efficacy outcomes included PCR-corrected cure rates at day 42, PCR-uncorrected cure rates at days 28 and 42; parasite and fever clearance times, presence and clearance of gametocytes, and haemoglobin (Hb) recovery from baseline to day 28. All standard safety outcomes such as incidence of adverse events, changes from baseline on haematology and clinical chemistry parameters and vital sign variation during the study were also evaluated. The treatment outcome was analysed as per protocol analysis based purely on the standard definitions of early/late clinical and parasitological failure  Table 11. All caSerious Adverse Events (SAE) were defined using the ICH GCP guidelines as any untoward medical occurrence that at any dose resulted in death, was life threatening, required hospitalization or prolongation of existing hospitalization, resulted in persistent or significant disability/incapacity.This is a sub-analysis of a multicentre study designed as a non-inferiority trial. This study, being part of a multicenter clinical trial, was not powered to compare treatments within Zambia. All randomized patients fulfilling the protocol eligibility criteria, having taken at least 80% of the study medication, having completed the day-28 assessment and having an evaluable PCR in case of recurrent parasitaemia were included. All drop-outs and all patients with missing or non-interpretable PCR results were excluded from the PP population. Data was entered using SAS programme. Statistical analysis was done using Epi Info software version 3.4.3 and STATA statistical analysis software package . An univariate analysis was done to characterize the sample of patients in the study. To determine the risk of treatment failure, proportions were compared using the \u03c72 or Fisher's exact test. A two sided p-value of \u22640.05 was defined as statistical significant. In the proportional hazard regression, all patients were censored at the time of the last recorded visit and the ITT definition was used to define failure or ACPR  and peripheral parasitaemia by microscopy. Nevertheless, 429 were not included because the parasite density was outside the required range (49.0%), refused to take part in the study (30.3%), had taken an anti-malarial before screening (5.8%), were coming outside the study site (3.5%) or other 11.4%)  and LPF  (Table vs. 6.7% (6/90) for DHA/PQP and AL, respectively, and not statistically different between the two study arms .No ETF was observed. At day 14, the ACPR for both arms was 100%. At day 28, the percentage of patients with recurrent infection was significantly lower in the DHA/PQP  as compared to the AL group 25.6%, 23/90) . This difference was seen for both the LCF , while the proportion of LPF did not differ between the two study arms . As for the estimation at day 28, most recurrent infections were new infections, significantly less in the DHA/PQP  than in the AL group  (Table vs. 11.1%). Only four recrudescences were observed after day 28, all of them in the DHA/PQP group. The PCR-corrected ACPR was similar between the two study groups: DHA/PQP: 93.2% (179/192), AL: 93.3% (84/90), .At day 42, the percentage of patients with a recurrent infection was lower in the DHA/PQP  than the AL group  . Such difference was mainly due to a lower proportion of LCF in the DHA/PQP group compared to AL group . About 60% of patients had fever at baseline while at day 2 more than 97% of patients were afebrile in both treatment groups. Gametocyte prevalence at recruitment was similar in both study arms . However, gametocyte carriage measured as rate of person-gametocyte-weeks was significantly higher in the DHA-PQP group than in the AL group, both for the ITT  and the PP  populations.In patients with no recurrent parasitemia, hemoglobin was increased at day 28 in both DHA-PPQ arm  and AL  and was different between treatment groups (p = 0.047). In patients with no recurrent parasitemia, hemoglobin was increased at day 42 in both DHA-PPQ arm  and AL  and was not different between treatment groups (p = 0.45).At day 28 the survival analysis, using the ITT definition, showed a hazard Ratio (HR) (DHA/PQP/AL) of 0.35 for PCR-uncorrected treatment failure  Figure . The surSeven serious adverse events (SAE) were observed: three of them for patients (two in DHA/PQP and one in AL) having prolonged hospitalization due to late fever clearance; in addition, in the DHA/PQP group, one patient developed severe malaria and another had persistent fever, weakness and anorexia; both were treated with quinine. In the AL group, one patient developed severe anaemia on day 3, possibly associated with study drug, and was treated with quinine and blood transfusion; the other patients had jaundice on day 3, possibly associated to study drug. All patients recovered completely. No deaths were observed during the study.Both treatments were generally well tolerated and most adverse events were associated with the disease during the initial clinical episode or to the recurrent parasitemia during the follow-up. Cough, attributed to respiratory tract infections, was significantly more frequent in the DHA/PQP arm than the AL arm while diarrhoea was of borderline significance Table .Both DHA/PQP and AL were highly efficacious in treating uncomplicated malaria in Zambian children, even if statistically significant differences in favour of DHA/PQP were observed for the uncorrected ACPR . In addition, the rate of recurrent infections, particularly within the first 28 days of follow up, was significantly higher in children treated with AL. One of the strengths of this study is that it was done in an area of meso-endemic malaria, in children under five, a high risk group, and had a prolonged follow-up period of 42 days not to miss late recrudescences and to reflect the terminal half-lives of the drugs . One of P. falciparum multi-drug resistant genes, which were weakly and indirectly associated with a decreased in vitro susceptibility to lumefantrine [P. falciparum [In this study, patients treated with DHA/PQP had fewer re-infections, an added advantage as it allows patient haematological recovery before a new infection sets in, especially in high transmission areas. The withdrawal and lost to follow-up rate (7%) were within the WHO recommended limits . There afantrine , indicatlciparum . Anotherlciparum  Similarllciparum . The diflciparum ,22. Indelciparum . Even inlciparum . Such thBoth drugs are co-formulated but AL should be administered twice a day for three days whilst DHA/PQP is given once a day for three days. Furthermore, AL should be administered with a fatty meal for an effective absorption of lumefantrine. In Zambia, where 80% of the population is under the poverty level, the staple diet is based on maize , it is dIn summary, DHA/PQP may be a good alternative to AL , but theConsidering the above challenges, the direct and indirect costs, and the lessons learnt from the current treatment policy change, DHA/PQP could be employed as rescue and/or alternative treatment to AL or as second-line treatment for patients re-attending the health facility with malaria a few weeks after receiving the first line treatment. This would be a major improvement as compared to the current policy of administering quinine to any suspected case of failure, a policy against the WHO recommendations to use an alternative ACT for the second-line treatment . It woulThe authors declare that they have no competing interests.All authors contributed to the design of the study and assisted with data interpretation. MN, UDA and MM coordinated the study and supervised the enrolment and follow-up of patients. JPVG participated in data entry, collection and analysis of data. All authors participated in the preparation of the manuscript and approved the final version."}
+{"text": "In vivo efficacy assessments of the first-line treatments for Plasmodium falciparum malaria are essential for ensuring effective case management. In Ethiopia, artemether-lumefantrine (AL) has been the first-line treatment for uncomplicated P. falciparum malaria since 2004.P. falciparum in individuals >6 months of age at two sites in Oromia State, Ethiopia. Eligible patients who had documented P. falciparum mono-infection were enrolled and followed according to the standard 2009 World Health Organization in vivo drug efficacy monitoring protocol. The primary and secondary endpoints were PCR uncorrected and corrected cure rates, as measured by adequate clinical and parasitological response on days 28 and 42, respectively.Between October and November 2009, we conducted a 42-day, single arm, open label study of AL for P. falciparum patients (6.7%) presented with Plasmodium vivax infection during follow-up and were excluded from the per protocol analysis. Only one patient had persistent parasitaemia at day 3. No serious adverse events were reported, with cough and nausea/vomiting being the most common adverse events.Of 4426 patients tested, 120 with confirmed falciparum malaria were enrolled and treated with AL. Follow-up was completed for 112 patients at day 28 and 104 patients at day 42. There was one late parasitological failure, which was classified as undetermined after genotyping. Uncorrected cure rates at both day 28 and 42 for the per protocol analysis were 99.1% (95% CI 95.1-100.0); corrected cure rates at both day 28 and 42 were 100.0%. Uncorrected cure rates at day 28 and 42 for the intention to treat analysis were 93.3% (95% CI 87.2-97.1) and 86.6% (95% CI 79.1-92.1), respectively, while the corrected cure rates at day 28 and 42 were 94.1% (95% CI 88.2-97.6) and 87.3% (95% CI 79.9-92.7), respectively. Using survival analysis, the unadjusted cure rate was 99.1% and 100.0% adjusted by genotyping for day 28 and 42, respectively. Eight P. vivax possibly from relapse or new infection was observed.AL remains a highly effective and well-tolerated treatment for uncomplicated falciparum malaria in the study setting after several years of universal access to AL. A high rate of parasitaemia with NCT01052584 Plasmodium falciparum malaria ,49P. vivEven with high efficacy of AL in Ethiopia, the effectiveness of this drug will not be optimally realized if patients are not accessing the drug in a timely manner or if patients do not adhere to the treatment regimen. The Malaria Indicator Survey from 2007 in Ethiopia noted that only 3.9% of children with fever were treated with an anti-malarial within 24 hours and of those treated, more received CQ than AL . In an eP. falciparum infection in Ethiopia. The routine monitoring of first-line regimens is crucial for effective malaria control and national policy decision-making. In addition, some simple alterations of the current WHO protocol and reporting conventions could enhance a globally coordinated vigilance for early signals of declining efficacy of artemisinins.Even after several years of free universal access to ACT, AL remains a highly efficacious treatment for uncomplicated The authors declare that they have no competing interests.JH and BHA developed the protocol, supervised the study, conducted the analysis and drafted the manuscript. SPK and SF conceived of the study, assisted with the study design and protocol development, and helped draft the manuscript. ZM, SGT, DH, KG, MK, DJ, JLM, and RR participated in the study design, the study coordination and critically reviewed the draft manuscript. HN and MG carried out the drug level testing. SGB and LM supervised and conducted the laboratory analysis. TT supervised the study and assisted with data management. All authors read and approved the final manuscript."}
+{"text": "Parasitaemia on Day 3 has been proposed as a useful alert of potential artemisinin resistance, however, the normal variation of parasite clearance observed in artemisinin-based combination therapy clinical trials is poorly documented.The trends in early parasitological response following treatment with an artemisinin anti-malarial regimen were reviewed. A PubMed literature search identified all studies using an artemisinin regimen for uncomplicated falciparum malaria published between January 2000 and December 2011. Data from clinical studies were extracted for analysis using a standardized questionnaire.vs 0%, p=0.007), but increased in Asia . In 95% of studies the proportion of patients with peripheral parasitaemia was less than 6% at 72 hours.In total 65,078 patients were enrolled into 213 clinical trials with 413 treatment arms containing either an artemisinin derivative alone (n=26) or in combination with a partner drug (n=387). The proportion of patients remaining parasitaemic at 24, 48 and 72 hours was documented in 115 (28%), 167 (40%) and 153 (37%) treatment arms, respectively. Excluding resistance studies in Cambodia, the median proportion of patients still parasitaemic was 53.8%  on Day 1, 6%  on Day 2 and 0  on Day 3. Comparing studies from 2000 to 2005 and 2006 to 2011, the median proportion of patients reported to remain parasitaemic at 72 hours decreased in Africa (1.2% These results highlight the normal distribution of early parasitological responses following ACT, and the influence that heterogeneity in study design, host and parasite factors have in confounding a surveillance system based on Day 3 parasite positivity. Greater understanding of factors influencing parasite clearance is crucial, but will require analysis of pooled data from individual patient records. Plasmodium falciparum malaria recommended by the World Health Organization (WHO)  for PPRday2 was 2.72 [1.90-3.92] for regimens with less than two days of artemisinin derivative therapy (four arms with 588 patients) and was 6.19 [3.43-11.17] for PPRday3 for treatments with shorter than three days  compared to regimens with longer courses of artemisinin derivatives. In Asia, all treatments included at least three days of artemisinin derivatives, except for 182 patients in two study arms who received only two days. Further analyses of PPRday2 were restricted to patients who received at least two days of artemisinin derivatives and of PPRday3 to patients who received at least three days of artemisinin derivatives.In African studies, the median age of the patient was associated with a reduced risk of parasitaemia at Day 1 (OR 0.97 [95% CI 0.95 - 0.99]), but not on subsequent days Table\u00a0. IncreasIn Asian studies, there were no significant differences in initial parasite clearance between treatment regimens at 48 or 72 hours Table\u00a0. HoweverIn African studies, univariate factors were confirmed in multivariate analysis of parasitaemia at Day 1 and Day 2 Table\u00a0. After cIn the multivariate analysis of the Asian studies, baseline parasitaemia, age and treatment regimen also remained significantly associated with delayed parasite clearance Table\u00a0. After cIn the studies conducted in Asia, there was a temporal trend for an increased risk of parasite positivity at Day 1 (AOR = 0.90 [95% CI 0.81-0.99] per year), but not at other time points. In African studies, the reverse was observed, with a linear trend for decreased positivity at all days, but only statistically significant on Day 1 and Day 3 .Temporal trends in parasitological response were assessed in studies conducted from 2000 to 2005 and compared with 2006 to 2011. In African studies, the median reported proportion of patients remaining parasitaemic during follow up decreased at Day 1 (64% to 43%), Day 2 (10% to 4.6%) and Day 3 (1.2% to 0%). However these changes, adjusted for confounding factors, were not statistically significant in the logistic regression model. In contrast, in Asian studies the median proportion of patients parasitaemic at Day 3 rose from 0.4% in 2000 to 2005 to 6.5% in 2006 to 2011. However after adjusting for confounding factors, and exclusion of Cambodian studies, these temporal trends were no longer statistically significant.The parasite prevalence rates and 95% confidence intervals are presented in the forest plots at Day 3 Figures\u00a0 and 7 anAmong 153 treatment arms reporting parasite positivity rate at Day 3, nine (5.9%) recorded a PPRday3 of 10% or higher. Of them, six treatment regimens were evaluated in Western Cambodia and one study was of a single dose of artesunate  of these data points were estimated from figures presented to depict parasitological response rather than the text itself. From the data available, 91% of patients had cleared their parasitaemia within two days and 97.9% within three days, however there was marked heterogeneity in clearance rates between studies and regions . Heterogeneity between studies I2 : AL = 92.3%; AS+AQ = 92.9%; AS+MQ = 91.7%; AS+SP = 95.6%; PIP+DHA = 60.9%.Click here for fileForest plots of Day 2 parasite positivity rates in Asia. Estimates and 95% CI are shown by treatment, sorted by year in descending order (most recent first). Heterogeneity between studies I2 : AL = 90.3%; AS+AQ = 93.8%; AS+MQ = 95.3%; AS+SP = 42.1%; PIP+DHA = 96.0%.Click here for fileStudies with Day 3 parasitaemia positivity rates \u226510%.Click here for file"}
+{"text": "The emergence of artemisinin resistance has raised concerns that the most potent antimalarial drug may be under threat. The currently recommended daily dose of artesunate (AS) is 4 mg/kg, and is administered for 3 days together with a partner antimalarial drug. This study investigated the impact of different AS doses on clinical and parasitological responses in malaria patients from an area of known artemisinin resistance in western Cambodia.P. falciparum malaria were randomized into one of three 7-day AS monotherapy regimens: 2, 4 or 6 mg/kg/day . Clinical, parasitological, pharmacokinetic and in vitro drug sensitivity data was collected over a 7-day inpatient period and during weekly follow-up to 42 days.Adult patients with uncomplicated 9/L) by Day 14 and resulted in the arm being halted early.143 patients were enrolled . Cure rates were high in all treatment groups at 42 days despite almost half the patients remaining parasitemic on Day 3. There was no impact of increasing AS dose on median parasite clearance times, median parasite clearance rates or on the proportion of patients remaining parasitemic on Day 3. However at the lowest dose used (2 mg/kg/d) patients with parasitemia >10,000/\u00b5L had longer median (IQR) parasite clearance times than those with parasitemia <10,000/\u00b5L (63 (48\u201375) vs. 84 (66\u201396) hours, p<0.0001). 19% of patients in the high-dose arm developed neutropenia (absolute neutrophil count <1.0\u00d710There is no pharmacodynamic benefit of increasing the daily dose of AS (4mg/kg) currently recommended for short-course combination treatment of uncomplicated malaria, even in regions with emerging artemisinin resistance, as long as the partner drug retains high efficacy.NCT00722150.ClinicalTrials.gov  The key features of the phenotype appear to be prolonged parasite clearance times and slower than expected parasite clearance rates in patients with adequate plasma drug concentrations The emergence of artemisinin resistant malaria along the Thai-Cambodian border has provoked global alarm that the most valuable and effective antimalarial drug is in danger of being lost, triggering a campaign to identify and eradicate resistant parasite strains Experimental seven-day AS monotherapy regimens, although impractical and inadvisable for routine, unsupervised use, are valuable research tools and have yielded important scientific information about potentially resistant parasites without the confounding influence of the partner drug and without compromising antimalarial treatment efficacy; a daily dose of 4 mg/kg in a 7-day AS monotherapy regimen was found to have 28-day efficacy of 93% in a previous study conducted in 2007 in Tasanh, western Cambodia This study formed part of the ARC3  Project, in which 4 sites  were selected to evaluate 7-day artesunate (AS) monotherapy regimens using harmonized protocols and common endpoints The protocol for this trial and supporting CONSORT checklist are available as supporting information; see This was a randomized, open-label comparison of 3 regimens of AS monotherapy given as a single oral dose of 2, 4 or 6 mg/kg/day  for 7 days  in otherwise healthy adult patients with acute falciparum malaria. AS2 was selected as the lower limit of what was considered an effective dose to probe for clinical resistance. The ratio of enrollment into the 3 groups was 2\u22361\u22362 into AS2, AS4 and AS6 respectively; AS4 was intended to serve as a control and act as a bridge to an earlier study performed at the same site in 2006/2007 Tasanh Health Center is located in Battambang Province in western Cambodia, close to the Thailand border and due south of Pailin ; this isP. falciparum monoinfection as determined by microscopy with a parasite density of 1000 to 200,000 asexual parasites/\u00b5L; (2) fever/history of fever within 48 hours; (3) age 18\u201365 years; (4) gave written informed consent to participate; (5) otherwise healthy outpatients. Exclusion criteria included the following: (1) pregnancy or unwillingness to use effective contraception if female and of child-bearing age; (2) history of intolerance or hypersensitivity to AS or other artemisinin derivatives; (3) history of any malaria drug therapy within 30 days; (4) history of other significant illness; (5) signs or symptoms indicating a requirement for parenteral antimalarial therapy; (6) signs or symptoms of severe malaria Consecutive patients were recruited into the study if they fulfilled the following criteria: (1) acute symptomatic Randomization was done by an independent statistician using computer-allocated blocks of 10 and was not stratified; individual treatment allocations were contained inside consecutively numbered sealed envelopes, which were opened sequentially by a study investigator or clinical research coordinator (CRC) after the decision to enroll a subject had been made by the study team. AS doses  were calculated individually based on the patient's weight at enrollment and the dose rounded up to the nearest \u00bc tablet (12.5 mg). Doses were administered with water and separated from other concomitant medications wherever possible. All treatment was directly observed. The full dose was repeated if vomiting occurred within 30 minutes, and half-dose given if vomiting occurred within 30\u201360 minutes. Before the study commenced 10 randomly selected tablets of study drug were analyzed in-house for content and weight uniformity according to standard guidelines in vitro drug sensitivity, molecular markers of drug resistance and to distinguish later recrudescence from re-infection by parasite genotyping. Malaria smears were prepared up to 8 times on Day 0, and then 4 times a day until 2 consecutive smears were negative for asexual parasites, then daily until discharge from the ward and then weekly on days 14, 21, 28, 35, and 42, and again if symptoms consistent with malaria appeared during follow-up. Plasma samples for AS and DHA concentrations were collected on Day 0 , and Day 6  doses of AS. Additional blood for parasite genotyping and culture was drawn at the time of treatment failure.Subjects remained as inpatients for the first 7 days then returned for follow-up on days 14, 21, 28, 35 and 42. Before initiating AS therapy blood was drawn to test for 100) was defined as the time from the start of treatment until the first time the blood smear became negative for asexual parasites and remained negative at 2 consecutive measurements. PCT90 and PCT50 were times for the parasitemia to reduce to 90 and 50% of baseline value. Parasite reduction ratios (PRR) were calculated as 100 minus percentage reduction from baseline level at 24, 48 and 72 hours. The slope of the log10 transformed parasite clearance curves was used as a measure of parasite clearance rate and calculated using TableCurve 2D . Fever clearance time (FCT) was defined as the time from baseline until the start of the period in which the tympanic temperature remained below 38\u00b0C for at least 24 hours.Outcomes at 28 and 42 days, including early treatment failure (ETF), late treatment failure (LTF) and adequate clinical and parasitological response (ACPR) were classified according to standard definitions th and 7th AS doses and on Day 14.Because the highest dose arm (AS6) involved a total oral AS dose of 42 mg/kg, which is higher than other previously published studies, a number of safety features were built into the study design . These iGiemsa-stained thick and thin blood smears were examined by two microscopists blinded to each other's results and to the treatment status of the study subject. Parasite densities were calculated based on a count of parasites per 200 WBCs (thick film) or per 5000 RBCs (thin film). At least 200 oil immersion fields were examined on the thick film before a blood smear was considered negative. The final count was determined by taking the geometric mean of the two microscopists counts. In case of a difference in results  between the two microscopists, the blood smear was re-examined by a third microscopist independent of the earlier findings and the third reading accepted as the final result.max and Tmax were estimated by inspection of data. Pharmacokinetic (PK) parameters area under the plasma concentration-time curve (AUC) and t1/2 were calculated for AS and it's major metabolite dihydroartemisinin (DHA) by non-compartmental analysis using PK Solutions Software (Summit Research Services).Whole blood was collected into chilled sodium heparin tubes, centrifuged immediately, plasma separated and frozen at approximately \u221220\u00b0C or below. Samples were transferred to Bangkok for analysis by LC-MS P. falciparum (W2) indicating prior use of antimalarial drugs was performed using a previously described ex vivo bioassay method Plasma samples from blood collected into sodium heparin were frozen at \u221220\u00b0C and transported to AFRIMS for analysis. Determination of antimalarial activity in DHA equivalents against laboratory strains of CBC samples were collected into EDTA tubes and analyzed using a Beckman Coulter\u00ae AcT5diff analyzer. Plasma ALT samples were measured using a Reflotron\u00ae Plus analyzer (Roche Diagnostics).ex vivo assay for susceptibility to DHA, AS, mefloquine (MQ), quinine (QN), chloroquine (CQ), and lumefantrine (LUM) 50 without the confounding effect of baseline parasitemia. Parasite culture and drug sensitivity assays were performed as previously described P. falciparum clone was used as a reference and for quality control of drug-coated plates.Fresh samples, without prior freezing or pre-culturing, were assayed in the histidine-rich protein 2 (HRP2) drug sensitivity P. falciparum merozoite surface proteins MSP-1  gene ID 813575) and MSP-2 (NCBI gene ID 812660), and the glutamate-rich surface protein (GLURP) (NCBI gene ID 810501) were compared in individual samples from baseline and time of failure according to standard methodology To distinguish recrudescence from new infection polymorphisms in genes encoding the P. vivax parasitemia during follow-up, were compared between treatment groups using the Chi-squared and Fishers exact tests. Values of normally distributed data were expressed as means (95%CI) and non-normally distributed data as geometric means (95% CI) or medians (IQR), as appropriate. Means were compared using ANOVA; otherwise the non-parametric Kruskall-Wallis test was used. Comparison of failure rates between groups and the proportion of patients remaining parasitemic over time, were assessed by modified intention-to-treat (ITT) analysis using Kaplan-Meier methods and differences between groups compared using the Logrank test (WHO 2009). Data were analyzed using Stata 10.0  and reported in accordance with CONSORT methodology Sample size estimation assumed a cure rate of 80% in AS2 and 95% in AS6 and was calculated as 60 patients in each of the two arms with >80% chance of detecting a significant difference in cure rate by uncorrected Chi-squared test at a two-sided 0.1 significance level. Proportions of patients, both adjusted and non-adjusted for new infections and for From August 2008 until July 2009 161 patients were screened and 143 enrolled into the study . SeventyEight months after the study started a Cohort Halting Rule was triggered when a fifth subject in AS6 developed neutropenia; safety analysis demonstrated significantly lower geometric mean absolute neutrophil counts (ANC) on Days 6 and 14 in patients in AS6 and led to permanent suspension of the arm; this finding has been reported in detail elsewhere P. vivax infection during the course of the study ; removal of these cases from outcome analyses did not affect the results  was classified as ETF after a clinical deterioration on Day 1, which required transfer to another center for continued care. The remaining 7 cases had fever plus parasitemia on Day 3, though all were much improved clinically from presentation . Six werP. falciparum parasites reappeared during 42 days of follow-up in 8 cases, of whom 5 were confirmed as recrudescence by PCR genotyping  vs. 84 (66\u201396) hours, p<0.0001); there was a similar but smaller effect seen in AS4 (p\u200a=\u200a0.049), but not in AS6 (p\u200a=\u200a0.65). This dose-effect was not seen for PCT90 or PCT50. Comparison of the slope of the log10 transformed parasite clearance curves showed no significant difference between treatment groups, and multiple regression analysis using slope as the dependent variable showed no significant association with parasitemia, history of previous malaria, IC50 DHA or treatment allocation. However comparison of the parasite clearance slopes between patients with a subsequent per protocol failure and those cured at 42 days showed a small but significant difference . Seventy-two hours after AS treatment commenced almost 50% of patients still had asexual parasites on a peripheral blood smear .ITT analysis showed no differences between groups in median PCT72 hours . However100  (100 \u226596 h the rapid clearance group had significantly lower median baseline parasitemia and fewer per protocol failures (p<0.001 and p\u200a=\u200a0.004 respectively); moreover the median slope of the log10 transformed parasite clearance curves was steeper in these rapid clearers than in patients with PCT100 \u226596 hours . Plasma concentrations of AS and DHA could not explain the difference in clearance between these two groups. Compared to patients with PCT100 >48 h patients with PCT100 \u226448 h were more likely to have a history of previous malaria . For patients remaining parasitemic at Day 3 or with PCT100 \u226596 h the odds ratios (95% CI) of per protocol failure were 5.9 (1.2\u201329.3) and 4.6 (1.3\u201316.1) respectively. For patients remaining parasitemic at Day 3 or with PCT100 \u226596 h the odds ratios (95% CI) of PCR-adjusted recrudescence within 42 days were 4.4 (0.5\u201341.8) and 16.9 (1.6\u2013176.5) respectively.Comparison of parasitological responses between recrudescent subjects and those successfully cured at 42 days showed significant differences in PCT100 . Howeverm Pailin . Only 24 (\u226448 h) . CompareP. vivax relapse rate during follow-up .Significantly more patients remained parasitemic at 72 hours in the second compared to the first 6 months of the study , despite an even distribution of treatment allocation in each period. Patients in the second half of the study were less likely to give a history of a diagnosis of malaria within the previous 12 months (55 vs 77% p\u200a=\u200a0.005). They also had a lower occurrence of gametocytes on a baseline peripheral blood smear  and a reduced max and AUC0-8h for AS and DHA increased with AS dose but there was wide variation in individual plasma levels. PK values (Cmax and AUC0-8h for AS and DHA) for PCR-confirmed recrudescence cases were all above the 25th percentile of the median of the respective parameters compared to the 64 subjects cured with AS 2 mg/kg/day. In aggregate the 8 patients classified as ETF had comparable drug levels to those cured  profiles were obtained following the first AS dose in all 143 subjects and also around the final dose in the 136 subjects completing 7 days of monotherapy. As expected median Cse cured ; howeverhed data , half-li50 data are shown in 50 values for DHA were 8.19 (IQR 5.13\u201314.0) nM and for AS were 5.66 (3.57\u20137.81) nM, more than double the values measured at the same site using the same methodology 3 years previously 1. Individual IC50 values for DHA and AS did not correlate well with most parasitological parameters, and did not differentiate between cured and recrudescent patients (50 for patients remaining parasitemic at 72 hours was 9.60 (6.82\u201314.7) nM, significantly higher than the median IC50 of 6.26 (4.22\u201313.4) nM for subjects clearing parasites within 72 hours (p\u200a=\u200a0.013). There was no significant difference observed between AS IC50 for patients who were and were not parasitemic at 72 hours .ICpatients . In contThis is the largest reported clinical trial to characterize clinical and parasitological responses to AS monotherapy in a region of artemisinin resistant malaria and evaluate a public health strategy to overcome these slow responses. Although impractical and inadvisable for routine, unsupervised use, AS monotherapy regimens are important research tools and allow detailed analysis of the dose response to AS without the confounding influence of a partner drug. Despite high treatment efficacy almost 50% of patients in this study population remained parasitemic 72 h after commencing AS, suggesting an alarming trend towards artemisinin resistance in the parasite population P. falciparum strains may require higher doses of AS to achieve the same clinical and pharmacodynamic effects as fully sensitive strains. However, because per protocol cure rates in Tasanh were high in all dosing groups, a finding in keeping with some other recent studies of AS monotherapy, we did not demonstrate improved 42-day efficacy using high-dose treatment. Even if the high-dose arm had recruited its planned allocation of subjects we would not have been able to demonstrate a dose-dependent effect between groups in terms of clinical outcome. This contrasts with the 30% recrudescence rate and prolonged fever clearance seen in Pailin; there, the high failure rate might in part be explained by the much higher median parasitemia at baseline (due to differing entry criteria), and lower apparent drug concentrations  in a polyclonal infection may require more parasite lifecycles before complete killing occurs, and that PCT50 and PCT90 values reflect killing of the more sensitive clones. Only 17% patients in this study were rapid parasite clearers (PCT100 \u226448 h); these patients had a significantly lower median parasitemia and a higher reported history of previous malaria episodes indicating some degree of prior immunity; none failed treatment during 42d. The influence of both parasitemia and immunity on parasite clearance times has been well described previously We did not demonstrate any improvement in pharmacodynamic responses when higher doses of AS were given. Within AS2 however we did observe that parasitemia >10,000/\u00b5L is associated with significantly prolonged median PCT50 for DHA and AS between failures and cures was surprising, but is in keeping with previous reports from this region P. falciparum infections are known to be polyclonal, the wild-type, non-artemisinin resistant clone may have a selective advantage in the environment over the artemisinin resistant clone. Although the degree of polyclonality of malaria infections in Tasanh has not yet been demonstrated, PCR genotyping in the subset of patients from this study with reappearance of P. falciparum parasites indicated that 7 of 9 (78%) had polyclonal infections at baseline. Moreover analysis of both Thai and African isolates by Druihle et al revealed 33 and 34 clones respectively in primary specimens in vitro responses to antimalarial drugs. Our in vitro technique involves culture of fresh isolates in the field (\u201cex-vivo\u201d) to mitigate losing clones, perhaps artemisinin resistant clones, which may not survive cryopreservation. Even so, this technique still does not allow easy differentiation of small clonal populations from within a polyclonal primary sample. The finding of lower median IC50 DHA and AS in recrudescent isolates at baseline might be explained by the very low parasitemias seen at recrudescence. HRP2 is produced by parasites and there is a strong relationship between parasite count and HRP2 level; in high parasitemias this is corrected for by dilution of the fresh sample prior to plating, but the converse is not possible with current methodologies. Additional work is needed to standardize in vitro methodologies as well as optimize strategies for isolating resistant clones from polyclonal infections.The lack of a clear difference in median ICth patient had a mixed infection on Day 35 but the P. falciparum component was a new infection. This indicates that prolonged courses of AS were still effective in curing the malaria infection despite slow clinical and parasitological responses. Two of these ETF cases had sub-optimal plasma drug concentrations despite receiving weight-based dosing and directly observed therapy. The huge variability seen in individual pharmacokinetic profiles in malaria patients, even in the highly controlled environment of a clinical research trial, emphasizes the importance of controlling unregulated use of poor quality drugs, which likely plays a significant role in inducing resistance in low transmission settings A crucial observation was that 5 of 6 cases meeting ETF criteria of fever and parasitemia on Day 3 who continued AS monotherapy went on to clear parasites and remained cured to Day 42 without a requirement for rescue treatment; the 6In summary this study demonstrates that the parasitological response to antimalarial therapy is a complex interaction of factors including parasitemia, immune status, plasma drug concentrations and innate parasite resistance to the antimalarial drug. Increased doses of AS did not impact on clinical or pharmacodynamic outcomes in this setting of emerging artemisinin resistant malaria and were toxic at the highest cumulative dose evaluated. Since completion of this clinical trial, Tasanh has been at the center of a high profile containment program; only time will tell if these efforts have been successful or whether artemisinin resistant clones have already spread. Artemisinin resistance remains a significant threat to the malaria-endemic world and this study highlights that simple dose escalation is not a viable strategy to overcome it. Continued coordinated public health strategies are necessary to keep this threat at bay.Checklist S1CONSORT Checklist(DOC)Click here for additional data file.Protocol S1Trial Protocol(DOC)Click here for additional data file."}
+{"text": "Plasmodium vivax malaria cases in Ethiopia. However, emergence of chloroquine resistant strains of the parasite has challenged the current efficacy of the drug. Therefore, the aim of this study was to assess the effectiveness of chloroquine against P. vivax strains in one of the malaria endemic areas of Ethiopia, namely Halaba district, located in South Nations and Nationalities Peoples Region (SNNPR) of South EthiopiaChloroquine is an anti-malarial drug being used to treat Among 87 malaria patients enrolled in the study, only 80 of them completed the 28-days follow-up. Seven of them dropped from the study for different reasons. Among those study participants that completed their follow-up, 69 were classified under the category of adequate clinical and parasitological response (ACPR). However, the remaining 11 cases were considered as under treatment failure mainly due to recurrence of parasitemia on day 7 (four patients), day 14 (six patients), and day 21 (one patient). The age of all cases of treatment failures was found to be less than 20 years. The load of parasitemia of patients with treatment failure on day of admission (4709.4/\u03bcl) was higher than day of recurrence (372.37/\u03bcl). Parasite reduction ratio (PRR) of treatment failure cases was 12.6/\u03bcl.P. vivax (CRPv) strains to malaria endemic areas of Ethiopia. It is recommended that all concerned bodies should act aggressively before further expansion of the current drug resistant malaria.This report revealed the rise in treatment failure (13% [95% CI = 0.074 - 0.217]) as compared to earlier reports from Ethiopia. It signals the spreading of chloroquine resistant  Plasmodium parasites. For more than 50 years, chloroquine (CQ) was the drug of choice for effective treatment of uncomplicated malaria of all species [Plasmodium falciparium was first documented in 1986 from the border areas of Ethiopia [\u00ae) (AL) in place of SP as a first line treatment in 2004 [The efficacies of anti-malarial drugs have been challenged by the emergence of drug resistant strains of  species . ResistaEthiopia  several Ethiopia , followeEthiopia -7. The hP. vivax was not known in Ethiopia until recent years. It has been used as the first line drug for many years for treatment of P. vivax infection. However, after the first report from Debreziet, Ethiopia [P. falciparium and P. vivax) in the study area are 30 and 70%, respectively [P. falciparium from the same district [P. vivax malaria and the presence of antimalarial drug resistant P. falciparium in the study area could be an indicator for the possible emergence of CRPv strains. In agreement with the hypothesis, the outcome of the study showed the highest ever reported decline in the efficacy of chloroquine drug against P. vivax. Except for these few reports, however, the nationwide prevalence of chloroquine resistant P. vivax remains poorly studied. This necessitates country wide survey for the current status of chloroquine resistant P. vivax malaria in other malaria endemic areas of the country.Even though, significant numbers of clinical resistance have been reported for a long time in most parts of Asia, mainly from Indonesia -12, chloEthiopia , gradualEthiopia , Serbo, Ethiopia , and NazEthiopia . The preectively . In addidistrict . The higWeinadega') and low-land ('Kola'), which accounts for 86% and 14%, respectively . The annual rainfall is estimated to be 857-1085 mm, while the mean annual temperature varies from 17 to 20\u00b0C with a mean value of 18\u00b0C. The most common health problem in the district is malaria [Anopheles arabiensis is a chief vector of the malaria parasite in the region [The clinical trial was conducted at Halaba Kulito Health Center, located in Halaba town about 313 km south of Addis Ababa, the capital city of Ethiopia  [Individuals seeking treatment for malaria infection at Halaba Kulito Health Center during the study period and having t by WHO , 87 pati P (1-P)  and 20% P. vivax mono-infection with parasite density above 250/\u03bcl, history of fever during 48 hours prior to time of recruitment, ability and willingness to participate in the study based on information given to parent or guardian, access to health facility and informed consent [The criteria used for inclusion of the study participants were; age >6 months, positive for  consent .Presence of clinical conditions that requires hospitalization, presence of severe malnutrition, pregnancy, significant concomitant febrile illness which would interfere with follow-up, chronic infectious diseases other than malaria, known allergy and/or intolerance to drug (s) being tested were the criteria used for exclusion of the patients .in-vivo follow-up days, patients who failed to respond to chloroquine of therapeutic dose were re-treated with quinine (10 mg base/Kg), the second line drug in Ethiopia.Patients confirmed to fulfill the inclusion criteria, and volunteer to participate in the study were treated with chloroquine under supervision of investigators and health professionals working in the health center. Each patient was treated with 25 mg base/kg of chloroquine for three consecutive days  followedThe status of chloroquine phosphate , used to treat all patients was checked for its quality before administered to patients. Accordingly, five tests, including weight variation, disintegration, dissolution, identification and chloroquine hydrochloride injection assay were made, following the standard procedures recommended by British Pharmacopoeia, ,23, usinP. vivax asexual stages were counted against 200 white blood cells (WBC), assuming the median total WBC count of 8,000/\u03bcl.Parasites were identified by microscopic observation of the parasite's morphology using duplicate thick and thin blood smears taken on day of enrolment (Day 0). In addition, blood slides were done at subsequent visits (scheduled and non-scheduled). After fixing the thin film in methanol, both the thin and thick smears were stained with Giemsa  for 30 min and the thick films were examined under oil immersion objective for malaria parasites. o/P2 (where Po is parasite count on day 0 and P2 parasite count on day 2).Parasite reduction ratio (PRR) on day of admission and day 2 (after 48 hours) was evaluated as suggested by White  using thP. vivax positive and negative slides were re-examined by senior laboratory technician at Jimma University.Each slide was carefully examined by experienced laboratory technicians of Halaba Health Center. All The study population were classified into three categories; early treatment failure (in the case of persistence of parasitemia up to day7), late treatment failure (in the presence of parasite recurrence within 28 days follow-up), and non-treatment failure (the absence of parasitemia in 28-days follow-up) .in-vivo therapeutic efficacy test was double entered and analyzed using SPSS software . Kaplan-Meier survival probability analysis was used to evaluate treatment outcome of the study participants during follow-up period. Data of patient having mixed infection with P. falciparum and vomiting was excluded from the analysis. While data of those patients lost to follow-up were included in the analysis considering them as non-treatment failure cases. In non-normally distributed data (age), median was used to measure the central tendency. Parasite count and parasite reduction ratio were analyzed using geometric mean. In all analysis, significance level was considered at 95% confidence interval.Data collected from The study protocol was reviewed and approved by Ethical Review Committee of Jimma University, College of Natural Sciences. Written informed consent was obtained from each patient or their guardians for patient younger than 18 years.P. vivax mono-infection. Only 87 of the 119, who fulfilled inclusion criteria set by WHO [During the enrollment period from January 4 to 10, 2009 (six consecutive days), a total of 311 blood films of patients suspected to have malaria infection were inspected at Halaba Kulito Health Center. A total of 198 cases were positive for malaria infection and 119 cases were confirmed to have The socio-demographic and some clinical features of the study subjects are as shown below Table . MajoritP. falciparium (1 patient) , vomiting (1 patient), and mixed infection with Since CQ is a fast acting antimalarial drug, parasitaemia clearance times for almost all patients involved in the 28 days in-vivo study were 48 hours except four patients whose parasitaemia persisted for seven days. Relative PRR of the study participants was 10.7/\u03bcl. Similarly, gametocyte was detected for a week in 15 patients and completely cleared on day 14. The study participants were relatively not febrile on day of admission (36.7\u00b0C). However, the body temperature dropped with follow-up days. Likewise, patients with treatment failure on day of recurrence were not febrile except three cases . Vomiting and diarrhea, the common malaria symptoms were encountered in 11% and 19% of the cases on day of admission, respectively, and disappeared on day7.In this study, about 13% (11/85) treatment failure was observed Table . Among tP. vivax malaria infection in the country. However, some reports on chloroquine resistant P. vivax malaria (CRPv) are coming from different malaria endemic areas of Ethiopia [Chloroquine has been in use both for treatment and prophylaxis in health centers and the communities of Ethiopia. It is an anti-malaria drug recommended by the Ministry of Health of Ethiopia for treatment of Ethiopia -16. In sEthiopia ,26,27.P. vivaxAs stated by Baird , a subjeP. vivax malaria after anti-malarial drug treatment might not necessarily be an indication of the emergence of CRPv strains. There could be other factors contributing to the failure such as malabsorpition of the drug, relapse, poor drug quality, re-infection, low level of drug in blood, and recrudence of parasitemia [Treatment failure of asitemia . In thisasitemia .Besides, for most of the early treatment failures (four), the original parasite persisted up to day 7 though their numbers were less than the day of admission. Under such a situation, it is impossible to think of the treatment failures to be due to re-infection or relapse as a new parasite needs a longer incubation period (ranges from ten to seventeen days). In addition, Baird and Hoffman  stated tTreatment failures associated with drug resistance may also be due to poor drug quality . HoweverIn-vivo therapeutic efficacy studies remain the 'gold standard' method for assessing antimalarial drug efficacy. However, due to the difficulty in classifying true treatment failure, as recurrences of parasitemia  or due to malabsorption of drug by patients, supplementing it with other confirmatory tests is important. For instance measurement of CQ and its major metabolite DCQ in blood of patients with treatment failures could clearly reveal the presence of malabsorption or recurrences of resistant parasites.P. vivax strains in malaria endemic area of Ethiopia. Thus, besides taking the right measures by the concerned bodies to control further expansion of resistance against chloquine, more study on the degree of dissemination of the resistance pattern within the P.vivax strains in other malaria endemic regions of the country is recommended to have clear picture of the country wide distribution of the problem.The treatment failure reported in this study (13%) was relatively the highest as compared to earlier few reports from Ethiopia and Africa. This report is a good indicator of the emergence and spread of chloroquine resistant Plasmodium vivax; DACA: Drug Administration and Control Authority of Ethiopia; PRR: Parasite Reduction Ratio; SNNPR: South Nations and Nationalities Peoples Region; WBC: White Blood Cells; WHO: World Health OrganizationACPR: Adequate Clinical and Parasitological Response; CQ: Chloroquine; CRPv: Chloroquine Resistant The authors declare that they have no competing interests.TK was fully involved in all phases of the study, including data collection and monitoring both in the field and laboratory, data analysis, interpretation, and write-up of the manuscript; KB was involved in data collection and field monitory, statistical analysis of data and critical revision of the manuscript for publication; KG sketched map of the study site and involved in data collection. All authors read and approved the final version of the manuscript."}
+{"text": "Plasmodium chabaudi malaria parasites selected for resistance to artesunate (an artemisinin derivative) through a step-wise increase in drug dose evolved slower clearance rates extremely rapidly. In single infections, these slower clearance rates, similar to those seen in the field, provided fitness advantages to the parasite through increased overall density, recrudescence after treatment and increased transmission potential. In mixed infections, removal of susceptible parasites by drug treatment led to substantial increases in the densities and transmission potential of resistant parasites (competitive release). Our results demonstrate the double-edged sword for resistance management: in our initial selection experiments, no parasites survived aggressive chemotherapy, but after selection, the fitness advantage for resistant parasites was greatest at high drug doses. Aggressive treatment of mixed infections resulted in resistant parasites dominating the pool of gametocytes, without providing additional health benefits to hosts. Slower clearance rates can evolve rapidly and can provide a strong fitness advantage during drug treatment in both single and mixed strain infections.The evolution of drug resistance, a key challenge for our ability to treat and control infections, depends on two processes: de-novo resistance mutations, and the selection for and spread of resistant mutants within a population. Understanding the factors influencing the rates of these two processes is essential for maximizing the useful lifespan of drugs and, therefore, effective disease control. For malaria parasites, artemisinin-based drugs are the frontline weapons in the fight against disease, but reports from the field of slower parasite clearance rates during drug treatment are generating concern that the useful lifespan of these drugs may be limited. Whether slower clearance rates represent true resistance, and how this provides a selective advantage for parasites is uncertain. Here, we show that  st century. In the case of malaria parasites, this is particularly apparent, as the introduction of each drug has been followed by the rapid development and spread of resistant parasites. Without a constant supply of new drugs to replace those that are no longer effective, it is important to understand the processes that lead to the selection and spread of resistance though a parasite population, so that the useful lifespan of current drugs can be maximized. Here, we use a rodent malaria model system to try to select for reduced susceptibility to the current frontline malaria drug, artemisinin. We then examine the growth and transmission potential of resistant parasites in single infections and in competition with susceptible parasites (mixed infections) in drug-treated hosts. We show that parasites selected for reduced susceptibility to drugs have increased fitness in both situations. Our results also indicate that the consequences of different treatment regimes on the rate of spread of resistance should be evaluated and taken into account during regime choice.The evolution of drug resistance is a major challenge facing medicine in the 21 The evolution of drug resistance in pathogens is a major public health concern, as it has the potential to undermine many of the health advances achieved in the last century. In addition to the individual health impacts of treatment failure, drug resistance has substantial economic costs, including research into alternative treatments and new drug development Changes in the parasite response to artemisinins are unlike resistance observed to other anti-malarial drugs. Reduced clearance rates have been observed de novo and on the rate of spread of resistant parasites within a population, which is in large part a function of the strength of selection. The traditional view has been that aggressive chemotherapy, involving high doses applied for sufficiently long to eliminate parasites, best minimises the evolution of resistance because it reduces the probability of a de novo resistant mutant arising de novo resistance, but providing the strongest selection for resistant mutants already present in an infection The useful lifespan of a drug depends both on the probability that resistance arises Plasmodium falciparum gametocytes (the parasite stage infective to mosquitoes) take 7 days to mature in vitro tests or molecular markers of resistance are not currently available The spread of the artemisinin slow-clearance phenotype in parasites in South-East Asia suggests a fitness benefit for the parasite. However, as Plasmodium chabaudi to separately examine both sides of the double-edged sword in resistance management. First, we exposed malaria parasites to various starting doses of the antimalarial artesunate, thereby testing the ability of high drug doses to prevent the origin of resistance. Second, we used one of our selected lines to examine the fitness implications of slower clearance rates and of competitive interactions between resistant and susceptible parasites under different drug pressures. This work was done across three experiments . Initial infections were effectively cleared (no detectible parasites by microscopy for at least 7 days post treatment) with high doses of artesunate . But, for a lower dose treatment (8 mg/kg twice daily for 4 days), sufficient parasites survived drug treatment in 2 out of 5 infections to establish new infections; these parasites were used to establish our 2 selection lines (th passage) for 11 passages before reaching our experimental, selected lines: AS116P(art) & AS117P(art)). A control line (AS109P(s)) was passaged in parallel through mice without exposure to drugs. We continued selection on all lines for a further 12 passages, stepping up to 32 mg/kg and then 64 mg/kg, to generate at the end parasites in treated lines which survived a dose of 64 mg/kg  or AS117P(art)) or our control line (AS109P(s)). From day 6 (corresponding to peak parasite density), infections were treated with 4, 16, 32 or 64 mg/kg of artesunate twice a day for 5 days, or left untreated (5\u20137 mice per parasite and treatment combination). We used two measures of resistance: (1) drug efficacy during treatment, measured as the slope of the parasite clearance curve In order to test for resistance in our drug-selected lines, we infected mice with 1021\u200a=\u200a0.32, p\u200a=\u200a0.58; 24\u200a=\u200a3.91, p\u200a=\u200a0.42; Experiment 1 was conducted over two experimental blocks. In block A, both of our two replicate selection lines (AS117P(art) and AS116P(art)) were used in addition to the control line (AS109P(s)). There was no significant difference between the two selected lines in either clearance rate under drug pressure : AS116P(art) vs. AS117P(art) \u03c721,35\u200a=\u200a8.78, p\u200a=\u200a0.003; 22,33\u200a=\u200a4.94, p\u200a=\u200a0.085), nor an interaction between drug dose and parasite line . The change in clearance rates and half-lives in our selected line were similar to the resistance phenotype seen in human malaria infections P. falciparum parasites in culture 1,36\u200a=\u200a8.55, p\u200a=\u200a0.006; 2,35\u200a=\u200a3.05, p\u200a=\u200a0.061), and there was no significant interaction between drug dose and parasite line \u200a=\u200a8.23(\u00b10.55 SEM)hrs; AS117P(art)\u200a=\u200a10.18(\u00b10.51 SEM)hrs; \u03c71,30\u200a=\u200a\u221231.79, p<0.001). All mice were given a second drug treatment (32 mg/kg) each day at 4pm.Decreased drug sensitivity of ring stage parasites has been implicated in slower clearance rates under artemisinin treatment 21,19\u200a=\u200a8.45, p\u200a=\u200a0.009; effect size\u200a=\u200a1.63 hrs) but this was not significantly effected by time of day of the first drug treatment . Although time of treatment had no affect on the half-life of resistant and susceptible lines, it did seem to effect whether or not the resistant line declined immediately following treatment or whether there was a delay of a day between the initiation of drug treatment and a decline in parasite densities. For most cases we observed no lag at all: when the control line received early treatment, 0 out of 5 infections had a lag phase, when the control line received the late treatment, 1 out of 5 infections had a lag phase and when the resistant line received early treatment, 1 out of 5 infections had a lag phase. However, when the resistant line received the late treatment, 4 out of 5 infections had a lag phase. This meant that 2 days after the start of treatment there were more parasites when the resistant parasite received late treatment as compared to early treatment (late treatment\u200a=\u200a5.49\u00d7105 parasites per \u00b5L (\u00b19.3\u00d7104 SEM); Early treatment\u200a=\u200a3.30\u00d7105 per \u00b5L (\u00b16.8\u00d7104 SEM))In support of our earlier experiments, resistant parasites had a significantly longer half-life than our control line . After the completion of drug treatment, both the resistant line and control line infections recrudesced. This recrudescence occurred earlier and was larger for the resistant line, both for asexual parasite densities  day 7 to day 11 post infection, corresponding to the period of drug treatment; (ii) day 12 to day 28 post infection, corresponding to the post drug treatment peak in parasite density. Slower clearance rates in our resistant line  resulted3 or \u223c20 resistant parasites injected alone or in a mixed inoculum with 106 susceptible competitors. Infections were then left untreated (control group), treated with a low dose of artesunate (4 mg/kg) or treated with a moderate dose of artesunate (16 mg/kg). Drug treatment was given twice a day for 3 days (days 6\u20138 post infection). This treatment was shorter in duration than in our experiments characterising the resistance phenotype (experiments 1\u20132), since those experiments were explicitly testing the limits of the resistance phenotype and resistant parasites were at a much lower density at the time of treatment in mixed infections (due to a combination of lower inoculums and competitive suppression). The dynamics of resistant parasites in mixed infections with a susceptible competitor were unaffected by number of parasites in the initial inoculum , and so these treatments were grouped together for further analysis.The effect of drug treatment on our selected lines within mixed infections was examined in experiment 3 by initiating infections with either 1021,12\u200a=\u200a1.15, p\u200a=\u200a0.28; treated vs. untreated \u03c721,13\u200a=\u200a2.39, p\u200a=\u200a0.12; 21,12\u200a=\u200a9.69, p\u200a=\u200a0.002; The asexual stage density of drug-selected parasites in the absence of competition was unaffected by drug treatment or dose  and this depended on the drug dose . Across the whole infection, the highest density of asexual stage resistant parasites occurred in infections treated with the higher drug dose, and the lowest density occurred in the untreated infections , where again the highest densities of resistant parasites were seen in the infections treated with the highest drug dose . Similarly, the highest drug treatment led to the greatest relative abundance of resistant line gametocytes .As expected, the parasite dynamics for susceptible parasites in mixed infections were significantly affected by drug treatment and dose, with the highest drug treatment reducing the density of susceptible parasites to the greatest extent. This was the case for both asexual densities , but there was no additional reduction with a higher dose treatment .High dose treatment of mixed infections increased the cumulative number of gametocytes produced by the resistant line nearly 7-fold (as compared to densities in low dose treatment) . The pro21,28\u200a=\u200a17.37, p<0.0001; weight \u03c721,27\u200a=\u200a5.83, p\u200a=\u200a0.016). However, the higher-dose drug treatment did not result in any additional improvement to health outcomes as compared to the lower drug dose , which delayed the peak in parasite density. This meant that untreated infections were still in growth phase when drugs were administered to the treatment group; in this case, parasite densities continued to rise in the presence of drugs. This observation demonstrates that our selected line was largely resistant to artesunate, and it agrees with previous data suggesting that clearance rates estimates can be highly dependent on the within-host environment For the majority of our experimental infections, drug treatment was given at the peak of parasite density, when symptoms become apparent. This timing was chosen because people normally begin treatment only after the onset of symptoms. At that point in a periment . These iP. falciparum infection dynamics from areas with variable clearance rates suggest that susceptibility to artemisinin may be parasite stage specific, with resistance developing predominantly in the ring stage parasite The mode of action for artemisinin-based drugs, and the mechanism by which parasites lose susceptibility, remains unclear. Models of The majority of malaria infections consist of multiple genotypes, so that when resistance is rare, as it necessarily is when it first arises, the fitness benefits and costs of resistance will largely depend on interactions with susceptible competitors. Aggressive drug treatment of malaria infections with pyrimethamine can result in \u2018competitive release\u2019 of resistant parasites by removing susceptible competitors, while resistant parasites remain unaffected de novo resistance from arising. In contrast, parasites exposed to a step-wise increase in drug doses displayed significant increases in resistance, surviving 8 times the drug dose they were exposed to at the start of the selection regime  used in the selection regimes and the susceptible AJ strain parasites used in the competition experiment (experiment 3) were originally collected from thicket rats (Thamnomys rutilans) in the Congo Both the parental AS 6 susceptible parasites (AS13P) and treated with 8, 16, or 32 mg/kg artesunate on days 2\u20136 post-infection, or with 64 mg/kg artesunate on days 6\u201310 post infection (5 mice per dose). Drug treatment was given twice a day as an IP injection of artesunate (Sigma-Aldrich) dissolved in sterile water with the dose adjusted for the weight of the mouse at approximately 11am and 4pm. Pilot work suggested that twice-daily treatments were more effective. P. chabaudi has a 24 hour life-cycle, meaning that twice daily treatment is equivalent to daily treatment of P. falciparum, which has a 48 hour life cycle.Infections were initiated in 20 mice via intraperitoneal (IP) injection of 10Previous studies selecting for resistance to artemisinins have varied in their precise selection regimes but have, in general, treated early in the infection 6 parasites to new mice, starting parallel selection lines . Both lines were passaged eight times at this dose to arrive at the experimental clones (AS116P(art) and AS117P(art)). Subsequently, we continued our selection for another 7 passages at 16 mg/kg, 3 passages at 32 mg/kg, followed by two passages at 64 mg/kg, to derive the final selected clones AS148P(art) and AS149P(art).Thin blood smears were taken from all infections from two days after the final drug treatment and examined for the presence of parasites. In the 64 mg/kg group, blood was additionally passaged on to na\u00efve mice at two-day intervals (days 15\u201325 post infection) to test if parasites were present below the detection threshold. No parasites were detected in mice treated with the three higher doses, and no infections established in na\u00efve mice. In the 8 mg/kg group, three out of five mice had microscopically-detectable recrudescence, two of which reached sufficient densities to passage 106 parasites and then the mouse with the highest parasitemia to two new mice at 106 at day 7 post infection. After twelve passages, this led to experimental control clone AS109P(s). Note that this control clone was passaged one more time than the experimental line. At each passage stage, parasites were frozen down for storage and all parasites are maintained on liquid nitrogen at The Pennsylvania State University.From the same ancestral clone as the drug-selected lines (AS13P), an untreated control line was evolved. Two mice were inoculated with 106 parasites of AS116P(art), AS117P(art) or AS109P(s) (the control line). Infections were either left untreated or treated with artesunate  twice a day for 5 days (\u223c11am and \u223c4pm on days 6\u201310 post infection). Infections were monitored daily from day 3\u201321 post infection. During sampling, mouse weight and red blood cell density , and so these clearance slopes were used to calculate the parasite half-life during drug treatment for each infection \u00d724)).Parasite clearance rate was calculated by fitting the slope of the linear decline in parasite density over time  during the period of drug treatment 6 parasites of either the drug selected line (AS117P(art)) or the control line (AS109P(s)) treated with 32 mg/kg twice daily for 5 days (days 6\u201310 post infection). Half of our mice received the first treatment of the day at 9am and half at 1pm (5 mice per line per treatment time). All mice were given a second dose of drug each day at 4pm. Thin blood smears were taken from all mice at 8.45am and 12.45pm (15 minutes prior to drug treatment) in order to check the stage of parasites exposed to drugs. Slides were fixed in methanol, and stained with Giemsa. To establish the proportion of early stage rings present in infections, slides from day 5 post-infection were examined under the microscope and a minimum of 100 parasites per infection (10 mice per parasite line) staged for each time point. Staging was carried out on day 5 infections to avoid the possibility of drugs differentially killing some parasite stages.In experiment 2 infections were initiated with 10Infections were monitored daily from day 3 to day 21 post infection and then three times a week until day 54 post infection. In addition to the measurements made in experiment 1, 10 \u00b5L of blood a day was taken to estimate gametocytes by quantitative PCR 3 or \u223c20 AS117P(art) parasites inoculated at the same time as 106 parasites of a susceptible competitor (P.chabaudi strain AJ). Single infection controls were initiated with 103 AS117P(art) parasites. Infections were then either left untreated, treated with a low dose of artesunate (4 mg/kg twice a day for 3 days), or treated with a high dose of artesunate (16 mg/kg twice a day for 3 days). Infections were monitored daily for mouse weight, red blood cell density, asexual stage parasites and gametocytes from day 3\u201321 post infection. Further monitoring of infections occurred three times a week until day 41. Genotype-specific qrtPCR allowed us to monitor parasite densities from our drug-selected parasite line and a susceptible competitor line independently  over the full experiment Mixed infections were initiated with either 10http://www.R-project.org). Parasite half-lives (log transformed) and the cumulative density in the week post treatment were analysed using general linear models (Gaussian error structure). The proportion of ring stage parasites was analysed with a general linear model with a binomial error structure. Asexual densities (log10 transformed), gametocyte densities (log10 transformed) and the proportion of resistant parasites over time (arcsine square root transformed) were analysed using linear mixed effect models with mouse (nested within block for experiment 1) as random effects. As is common with repeated-measure parasite data 2 distributions, until the minimal adequate model was reached. Degrees of freedom correspond to the difference in the number of terms in the model.All statistical tests were carried out using R version 2.14.1 .Figure S1Comparison of parasite clearance curves for the two replicate selection lines AS117P(art) and AS116P(art). Mean clearance curves for AS117P(art) (dark red) and AS116P(art) (orange). Dashed lines show the standard error around the mean. Mean clearance rate taken from across 3 drug doses . Data from Experiment 1 block A.(TIFF)Click here for additional data file.Figure S2Comparison of parasite dynamics for the two replicate selection lines AS117P(art) and AS116P(art). Parasite dynamics for AS117P(art) (dark red) and AS116P(art) (orange) in untreated infections and in infections treated with 4, 16, or 32 mg/kg of Artesunate. Shaded area indicates the period of drug treatment. Data from Experiment 1 block A.(TIFF)Click here for additional data file.Figure S3Drug treatment and within-host competition: cumulative parasite densities from the start of drug treatment. Cumulative total parasite density (A\u2013B) and cumulative total gametocyte density (C\u2013D) after the start of drug treatment (day 6\u201341 post infection). Drug selected line (AS117P(art)) is shown in red (A & C) and susceptible competitor (AJ) in blue (B & D). Density of the drug-selected line significantly increases with drug dose for both asexuals  and gametocytes . For the susceptible competitor there is a non-significant negative relationship with drug dose for asexual density  and a significant negative relationship for gametocytes . Data are taken from experiment 3 and show summary statistics for the same patterns shown in (TIFF)Click here for additional data file."}
+{"text": "Plasmodium falciparum resistance to artemisinins, the first line treatment for malaria worldwide, has been reported in western Cambodia. Resistance is characterized by significantly delayed clearance of parasites following artemisinin treatment. Artemisinin resistance has not previously been reported in Myanmar, which has the highest falciparum malaria burden among Southeast Asian countries.P. falciparum malaria in Kawthaung, southern Myanmar. Parasite density was measured every 12 hours until two consecutive negative smears were obtained. Participants were followed weekly at the study clinic for three additional weeks. Co-primary endpoints included parasite clearance time , parasite clearance half-life (the time required for parasitemia to decrease by 50% based on the linear portion of the parasite clearance slope), and detectable parasitemia 72 hours after commencement of artesunate treatment. Drug pharmacokinetics were measured to rule out delayed clearance due to suboptimal drug levels.A non-randomized, single-arm, open-label clinical trial of artesunate monotherapy (4 mg/kg daily for seven days) was conducted in adults with acute blood-smear positive The median (range) parasite clearance half-life and time were 4.8 (2.1\u20139.7) and 60 (24\u201396) hours, respectively. The frequency distributions of parasite clearance half-life and time were bimodal, with very slow parasite clearance characteristic of the slowest-clearing Cambodian parasites  in approximately 1/3 of infections. Fourteen of 52 participants (26.9%) had a measurable parasitemia 72 hours after initiating artesunate treatment. Parasite clearance was not associated with drug pharmacokinetics.P. falciparum infections in southern Myanmar displayed markedly delayed clearance following artemisinin treatment, suggesting either emergence of artemisinin resistance in southern Myanmar or spread to this location from its site of origin in western Cambodia. Resistance containment efforts are underway in Myanmar.A subset of Australian New Zealand Clinical Trials Registry ACTRN12610000896077  Plasmodium falciparum malaria worldwide because of their high clinical efficacy, rapid parasite clearance and fast clinical recovery. ACT efficacy is now threatened by the recent confirmation of P. falciparum resistance to artemisinins in western Cambodia Artemisinin-based combination therapies (ACTs) are the first-line treatment for P. falciparum is defined clinically as delayed parasite clearance following artesunate monotherapy. No molecular marker of artemisinin resistance has been validated, and correlation between in vivo clinical outcomes and in vitro measures of parasite susceptibility is poor P. falciparum that is fully susceptible to artemisinins Artemisinin-resistant P. falciparum without confounding by the effect of a partner drug.A World Health Organization (WHO)-led surveillance program to monitor ACT efficacy was initiated in 2006 in Myanmar. Progressive increases in the proportion of patients with persistent parasitemia 72 hours after starting treatment were observed in Myanmar starting in 2009 The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Checklist S1 and Protocol S1.This was a non-randomized, single-arm, open-label clinical trial to assess parasite clearance rate following directly-observed 7-day oral artesunate (4 mg/kg) daily monotherapy for the treatment of adult participants with uncomplicated blood smear positive P. falciparum malaria. The study was approved by ethical review committees of the Department of Medical Research (Lower Myanmar), the Myanmar Ministry of Health, and the WHO.P. falciparum, asexual parasite density 10,000\u2013100,000/\u00b5l, fever defined as axillary temperature >37.5\u00b0C or history of subjective fever in the last 24 hours, ability to tolerate oral intake, provision of written informed consent, and agreement to comply with the study protocol, including hospitalization for seven days. Exclusion criteria included severe malaria The study was conducted in Palm Tree Hospital in Kawthaung, a town located at southern tip of Myanmar bordering Thailand . This stOnce-daily oral doses of artesunate 4 mg/kg/day, procured by WHO from Guilin Pharmaceutical Co. Ltd.  were administered under direct observation with 8 oz. of milk (to standardize diet before drug administration which may impact drug absorption) on days 0\u20136. Treatment was repeated in case of vomiting within 30\u201360 minutes after drug administration. Parasite density was assessed by blood smear every 12 hours by two independent microscopists, using the average of the two readings, until two consecutive negative smears were obtained. The exact time of artesunate administration, and the scheduled and actual time of blood collection for smear examination were recorded for each participant. At each finger-prick for blood smear, dried blood spots were also collected on Whatman 3 MM filter paper. Filter papers were air-dried, labeled and stored in individual plastic bags with desiccant until analyzed.Participants were evaluated daily for solicited and unsolicited adverse events. Blood was collected for pharmacokinetic analysis of artesunate and its major metabolite dihydroartemisinin (DHA) in pre-chilled blood collection tubes containing fluoride-potassium oxalate, immediately before, and 0.25, 0.5, 0.75, 1.0, 1.25, 1.5, 3, 4, 6 and 8 hours after the first dose of artesunate. Within 15 minutes of collection, blood was centrifuged at 4\u00b0C at 2000\u00d7g for 7 minutes, and plasma was collected and stored in liquid nitrogen until analysis.Participants were discharged from the hospital on day 7 and monitored as outpatients on days 14, 21 and 28. Hemoglobin was measured on days 14 and 28. Finger-prick blood was collected for malaria smear and filter paper blood spots on days 7, 14, 21 and at any time the participant felt unwell.P. falciparum or P. vivax was treated, following the National Malaria Treatment Guidelines, with unsupervised standard WHO-recommended regimens of co-formulated artemether-lumefantrine  80/480 mg every 12 hour for three days or chloroquine 25 mg base/kg once a day for three days followed by primaquine 0.25 mg/kg once a day for 14 days, respectively.Treatment outcomes were classified following WHO-recommended methods for monitoring antimalarial drug efficacy P. vivax infection was identified by nested PCR of 18ssrRNA using previously described methods Microscopy: Giemsa-stained thick and thin blood smears were prepared for parasite density determination and speciation, respectively. Parasites were counted against white blood cells (WBC) and parasite density per \u00b5l was calculated by dividing the number of asexual parasites by the number of WBC counted  and multiplying by an assumed WBC count of 6,000/\u00b5l. Smears were considered negative when no asexual parasites were found after counting 1,000 WBC. Molecular analysis of recurrent infections: DNA was extracted from dried blood spots following manufacturer\u2019s instructions , and amplified and analyzed as previously described to distinguish whether post-treatment recurrent infections were new infections or recrudescences Pharmacokinetic analysis: Plasma concentrations of artesunate and DHA were measured by liquid chromatography-tandem mass spectroscopy after solid phase extraction using published validated methods Parasite clearance half-life, parasite clearance time, and the proportion with persistent parasitemia at 72 hours following start of treatment were co-primary endpoints. Because prior studies only provided information on the latter endpoint, parasitemia at 72 hours was used for sample size determination. Assuming <5% 72-hour parasitemia is in areas with no resistance Data were double-entered into the Microsoft Excel database and analyzed using Stata , and R . R and SigmaPlot  were used to produce figure illustrations. Parasite clearance half-lives were estimated from serial parasite counts using the publicly available WorldWide Antimalarial Resistance Network (WWARN) Parasite Clearance Estimator Pharmacokinetics of artesunate and DHA were assessed by standard non-compartmental modeling using WinNonlin Professional , and described as maximal concentration (Cmax), time to Cmax, area under the concentration-time curve (AUC0-inf), and terminal elimination half-life. Total apparent oral clearance and apparent volume of distribution were estimated with assumed equivalent bioavailability (F) since there was no intravenous comparator. Pharmacokinetic data were skewed in distribution and summarized using medians and inter-quartile ranges. Data were also compared between participants with and without 72-hour persistent parasitemia, and between fast and slow clearers  Of a total of 68 potential candidates screened, 15 were excluded from enrollment (due to inability to comply with study protocol or evidence of mixed infections), and one from data analysis (enrollment parasitemia below the inclusion criterion); 52 were included in the final analysis . ParticiP. falciparum on day 21; three mixed P. falciparum and P. vivax on day 28; and 22 P. vivax on days 21\u201328. All of these recurrent infections responded to treatment with artemether-lumefantrine  or chloroquine and primaquine (P. vivax mono-infection). All recurrent P. falciparum cases were classified as new infections by PCR genotyping, yielding a PCR-corrected cure rate of 100%. Retrospective PCR analysis showed that at enrollment 24 of 52 (46.2%) participants were co-infected with sub-patent P. vivax in addition to P. falciparum. All but two smear-positive P. vivax infections found during follow up were also PCR-positive for P. vivax. Four additional cases of P. vivax were identified by PCR only.Enrollment parasite densities were moderate . The median (range) time to fever clearance was 3 days (2\u20137 days) and time to parasite clearance was 60 hours (24\u201396 hours) . Of 52 emax) (Parasite clearance curves (log-parasitemia versus time plots) were produced for each participant, and the parasite clearance half-life and the time required to reach 50%, 90% and 95% of the initial value  are summarized in max)  and totamax)  or artesThe time-plasma concentration-time plots of artesunate and DHA are displayed in P. falciparum to artemisinins in Myanmar, in a subset of the parasite population in southern Myanmar near the Thailand border. Although this is the first report of artemisinin resistance originating within Myanmar, a progressive decline in parasite response, over-time from 2001 to 2010, to various artesunate-based regimens has been reported in patients with hyperparasitemia in Thailand along the northwestern border of Thailand with Myanmar de novo in Myanmar.This study provides evidence of decreased sensitivity of We also observed a high rate of persistent parasitemia 72 hours after artesunate monotherapy, despite the 100% PCR-corrected 28-day clinical efficacy. While the 100% cure rate we observed indicates that artemisinins retain good efficacy and ACTs can still be used in Myanmar at present, this apparently high efficacy of artemisinin in our study should be interpreted with caution. In our study, the participants were followed only 28 days, instead of 42 days, and we may have missed cases of late treatment failure. In addition, the study was not adequately powered to detect the true efficacy of artesunate. Of note, we do not endorse the use of artesunate monotherapy for any purpose other than in clinical trials designed to assess the independent ability of artemisinins, to clear P. falciparum without confounding by the effect of a partner drug.One limitation of our study was that the frequency of blood smear examination was every 12 hours, while most other recent studies (except the Bangladesh study) sampled parasitemia every 6\u20138 hours. This relatively infrequent observation carries apparent potential for over-estimation of the time required to clear parasites in some cases and misinterpretation of the observed bimodal distribution of the rate of clearance. Nevertheless, our data can be directly compared with the Bangladesh data, and our findings are in striking contrast to those from Bangladesh, where almost all infections cleared in fewer than 36 hours P. vivax infections is consistent with other studies reporting that P. vivax was a common cause of recurrent parasitemia following treatment of acute falciparum malaria in this region P. falciparum, relapse of vivax infection  may have occurred after the falciparum infection subsided and artesunate was eliminated from the body. With growing recognition of the public health significance of P. vivax, arguments have been made for presumptive radical treatment of P. vivax using primaquine after all P. falciparum treatments in settings where mixed infections are common P. vivax and for its antigametocyticidal effect on P. falciparum.Our observation of highly frequent P. falciparum parasites to artemisinins in Myanmar, adjacent to its southern border with Thailand. Myanmar has a population of approximately 58 million people with diverse ethnic backgrounds, more than half of whom occupy remote rural areas where malaria is highly endemic, including areas bordering Thailand, Laos, Bangladesh, India, and China. Although Myanmar has only 4% of Southeast Asia\u2019s population, it has 20% of the region\u2019s malaria burden This study provides credible evidence of reduced susceptibility of Figure S1Map of Myanmar. The red star indicates the location of study site, Kawthaung.(TIF)Click here for additional data file.Figure S2Lack of association between parasite clearance rate and initial parasite density.(TIFF)Click here for additional data file.Figure S3Lack of association between parasite clearance rate and maximal concentration of dihydroartemisinin.(TIFF)Click here for additional data file.Figure S4Lack of association between parasite clearance rate and total exposure of dihydroartemisinin.(TIFF)Click here for additional data file.Figure S5Plasma concentration-time curve of artesunate (Panel A) and DHA (Panel B) in participants with (dashed line) and without (solid line) persistent parasitemia 72 hours after artesunate treatment.(TIFF)Click here for additional data file.Protocol S1Trial protocol.(DOC)Click here for additional data file.Checklist S1CONSORT checklist.(DOC)Click here for additional data file."}
+{"text": "The effectiveness of intermittent preventive treatment of malaria in pregnancy (IPTp) may be compromised by the spread of resistance to sulphadoxine/pyrimethamine (SP) across Africa. But little informtion exists on alternative drugs for IPTp or alternative strategies for the prevention of malaria in pregnancy. Therefore, we have investigated whether screening with a rapid diagnostic test and treatment of those who are positive (IST) at routine antenatal clinic attendances is as effective and as safe as SP-IPTp in pregnant women.During antenatal clinic sessions in six health facilities in Ghana held between March 2007 and September 2007, 3333 pregnant women who satisfied inclusion criteria were randomised into three intervention arms (1) standard SP-IPTp, (2) IST and treatment with SP or (3) IST and treatment with amodiaquine+artesunate (AQ+AS). All women received a long-lasting insecticide treated net. Study women had a maximum of three scheduled follow-up visits following enrolment. Haemoglobin concentration and peripheral parasitaemia were assessed between 36 and 40 weeks of gestation. Birth weight was measured at delivery or within 72 hours for babies delivered at home. Parasite prevalence at enrolment in primigravidae and in multigravidae was 29.6% and 10.2% respectively. At 36\u201340 weeks of gestation the prevalence of asymptomatic parasitaemia was 12.1% in study women overall and was very similar in all treatment groups. The risk of third trimester severe anaemia or low birth weight did not differ significantly between the three treatment groups regardless of gravidity. IST with AQ+AS or SP was not inferior to SP-IPTp in reducing the risk of low birth weight ; third trimester severe anaemia .The results of this study suggest that in an area of moderately high malaria transmission, IST with SP or AS+AQ may be a safe and effective strategy for the control of malaria in pregnancy. However, it is important that these encouraging findings are confirmed in other geographical areas and that the impact of IST on placental malaria is investigated.NCT00432367]ClinicalTrials.gov NCT00432367 [ Plasmodium falciparum infection in pregnancy causes maternal anaemia and low birth weight associated with parasitisation of the placenta Intermittent preventive treatment linked to antenatal care will remain an effective and sustainable strategy for the prevention of malaria in pregnancy provided that the antimalarial drug used is efficacious, safe, tolerable, cheap, and easy to administer, preferably as a single dose In the absence of an effective drug to replace SP for IPTp alternative strategies for the prevention of malaria in pregnancy need to be evaluated. Insecticide treated bed nets (ITNs) used during pregnancy are beneficial to both mother and her newborn baby Screening for malaria infection using a malaria rapid diagnostic test (RDT) at scheduled antenatal clinic visits and treatment of women who are positive with an effective antimalarial drug (IST), combined with effective vector control, provides a potential, alternative strategy to SP-IPTp. The WHO's current recommendation of four scheduled antenatal clinic visits, one at booking and three subsequent visits 4 to 8 weeks apart, provides a potential framework for implementation of an IST strategy. This strategy may be considered in parts of Africa where resistance to SP is increasing at alarming rates and in situations of low exposure to malaria as in The Gambia and Zanzibar where the incidence of malaria has gone down Intermittent screening and treatment combined with vector control may be an especially attractive option in such communities where continuing use of IPTp with SP will result in a large proportion of pregnant women receiving SP unnecessarily. This is also the case for communities where the incidence of malaria is highly seasonal. In these areas, many women receive IPTp during months of the year when the risk of malaria is minimal.Whether a screening and treatment strategy would prove to be as effective as SP-IPTp is not known. Therefore, we have undertaken a randomised, controlled trial in an area of Ghana with moderate malaria transmission to determine whether IST using SP or amodiaquine+artesunate (AQ+AS) is as effective in preventing maternal anaemia and low birth weight as SP-IPTp. The costs and acceptability to women and providers of such a strategy have also been assessed.The protocol for this trial and supporting CONSORT checklist are available as supporting information; see P. falciparum. The entomological inoculation rate in the neighbouring area of Kintampo is about 250 infectious bites per year. HIV prevalence in Ashanti region is reported to be 3.0% and 2.2% in the general population and in pregnant women respectively .The study was conducted in the Ejisu-Juaben and Afigya-Sekyere East districts of the Ashanti Region of Ghana from March 2007 to September 2008. Malaria transmission in this area is perennial but with a peak in the rainy season. The predominant parasite is Enrolment was undertaken at the antenatal clinic of three district hospitals and three health centres in the study area. The study population comprised pregnant women of all parities who presented at the antenatal clinics with a gestational age of 16 to 24 weeks at their first booking. Women who were temporary residents, had had a prior dose of SP-IPTp, had a haemoglobin concentration <5 g/dl, gave a history of sensitivity to SP, amodiaquine or an artemisinin, had an illness requiring hospital admission or declined to join the trial were excluded .The study protocol was approved by the ethics committee of the London School of Hygiene and Tropical Medicine and the Committee on Human Research and Publications Ethics of the School of Medical Sciences, Kumasi \u2013 Ghana. A Data, Safety and Monitoring Board (DSMB) approved the protocol, standard operating procedures and analysis plan.Sample size was calculated on the assumption that the prevalence of moderately severe anaemia (Hb<8 g/dl) in the third trimester of pregnancy and of low birth weight in women in the SP-IPTp arm of the study would be at least 12% and 6% respectively, figures based on findings from a study undertaken in The Gambia After written informed consent had been obtained, eligible women of all parities were randomised to one of the three treatment groups described above. Women who declined to participate were given SP-IPTp according to the national guidelines. The following process was followed to randomise women into the study arms. A list of random numbers was computer generated as identification numbers, randomly allocated to treatment groups and grouped in blocks of 15 by an IT specialist who did not participate further in the study. The list of identification numbers with their corresponding treatment groups was printed and cut into slips. Fifteen slips each with an identification number and an allocated treatment group were sealed in an envelope. During enrolment an eligible pregnant woman was asked by the recruitment team to pick a slip from the sealed envelope. The allocated treatment group printed on each slip indicated which treatment arm the women belonged to and the corresponding identification number was used to identify her. This constituted entering the study, and treatment group allocation was binding on the recruiting team and the woman from this point. Another envelope was opened only when the contents of the previous one had been exhausted. The PI and all other project staff were blinded to the randomisation process and treatment allocation.At enrolment, a finger prick blood sample was obtained for measurement of haemoglobin concentration, preparation of thin and thick blood films for malaria parasite counts and preparation of a filter paper blood spot. During the last 3 months of the trial, women in treatment group 1 who had a positive blood film and received SP-IPTp were seen again on day 14 and day 28 after treatment and a further blood film and blood spot obtained to check on the response to treatment with SP. HIV screening was offered to all pregnant women study women as part of the routine antenatal services recommended in Ghana with an option for treatment but the results of HIV screening were not available to the investigators. Women in the SP-IPTp arm received an initial dose of SP (1500 mg sulphadoxine/75 mg pyrimethamine) as a single dose. Pregnant women in treatment groups 2 and 3 were screened for malaria infection with an OptiMAL\u00ae dipstick, a lactate dehydrogenase (LDH) based RDT. The OptiMAL\u00ae dipsticks were purchased from DiaMed AG, Cressier, Switzerland who supplied and organised transportation to the study site in batches as and when needed,. The test kits were kept at room temperature always at the study site. The viability of the kits were tested monthly throughout the study period using positive test controls obtained from the manufacturer. The tests were performed and interpreted by the study team following the manufacturer's instructions. Women in treatment group 2 were treated with a single dose of SP (1500 mg sulphadoxine/75 mg pyrimethamine) if the RDT was positive. Women in treatment group 3 were treated with AQ+AS (AQ-300 mg + AS-100 mg twice daily for 3 days) if the RDT was positive. All treatments with SP and the first dose of AQ+AS treatment were administered by a member of the study team but doses 2 and 3 of AQ+AS were given to study women to take at home unsupervised. Women in groups 2 and 3 received no antimalarial treatment or IPT if their RDT results were negative. All women received a daily supplement of ferrous sulphate (200 mg) and folic acid (4 mg) tablets as part of the routine antenatal care services provided in Ghana throughout the study period. SP was purchased from Kinapharma Ltd, Ghana and AQ+AS was provided by the Ministry of Health, Ghana. All study women received a long-lasting insecticide treated bed net (LLIN) and instructions on how to use this at enrolment.Study women were asked to attend for follow-up antenatal care and IPTp or screening at 24, 32 and 36 weeks of gestation. At the 24 and 32 week visits, women in treatment group 1 received SP-IPTp whilst women in treatment groups 2 and 3 were screened with the RDT and, if positive, treated with SP or AS+AQ according to the allocation arm. As part of safety assessment, all women were visited at home by a trained community health worker to record any complaints that the women might have seven days after each scheduled antenatal visit.Blood samples were obtained from all study women between 36 and 40 weeks of gestation (before delivery) for determination of haemoglobin concentration and for preparation of thin and thick blood films. However, the smears were read retrospectively and so the results were not available for the point of care. Any study woman who presented with a history of fever or other features suggestive of malaria between scheduled antenatal visits was screened for malaria using an RDT and treated with quinine (30 mg/kg in divided doses daily for 5 days) regardless of treatment group.Information on the outcome of pregnancy was obtained for 2706 of the 3333 study women (81.2%), 2144 of whom delivered at a health centre or hospital and 562 at home. If a woman delivered at a health facility, birth weight was recorded by a midwife who was unaware of the treatment group of the woman who she was attending. Women who delivered at home were traced through a network of trained community health workers within 72 hours of delivery and the infant's weight was measured at home. The occurrence of miscarriages, still births, neonatal deaths and the presence of congenital abnormalities was recorded by midwives. If any congenital abnormality was suspected, a full examination, including a neurological assessment, was undertaken by a qualified medical doctor. Field teams visited all women and babies at approximately 6 weeks post delivery to obtain reports of any neonatal adverse events and a blood film was obtained from mothers at this time. We could not determine the prevalence of placenta malaria and efficacy of SP in postpartum study women as proposed due respectively to inadequate funding and difficulty in finding postpartum women with parasitaemia.An experienced microscopist, unaware of treatment group assignment, read all the blood films and quantified parasitaemia against 200 leucocytes in thick blood smears. A thick blood film was declared negative only after examination of 100 high power fields (HPF). An independent expert microscopist from the Noguchi Memorial Institute of Medical Research read ten percent of all blood slides obtained at enrolment and on follow up visits for quality assurance. Agreement between the study microscopist and the reference microscopist was 91.7% and a kappa of 0.83. Haemoglobin (Hb) concentration was measured using Hb 301 Hemocues . Paired samples from women who failed treatment with SP were tested for molecular markers to differentiate reinfections from recrudescences using genetic markers as described previously Stata version 10  was used for data analyses. The primary objective of the study was to demonstrate that the risk of third trimester moderately severe anaemia (Hb<8 g/dl) in the IST groups was no more than 5% greater than in the SP-IPTp arm. Secondary objectives were to demonstrate that the risks of low birth weight (BW<2500 g), spontaneous abortions, intrauterine deaths/stillbirths, neonatal and maternal mortality were not more than 4% higher in women in the IST groups than in women who received SP-IPTp. The principal analysis of primary and secondary outcomes was per protocol but an \"intention-to-treat\" analysis was also undertaken according to a statistical analysis plan approved by the Data and Safety Monitoring Board. In the per protocol analysis, only data from women who remained within their randomization group, received two courses of SP-IPTp (Group 1) or were screened twice using an RDT at scheduled visits (groups 2 and 3) and in whom the primary outcome had been recorded were considered for analysis. In the intent-to-treat analysis, women were included if they had received an initial treatment of IPTp or had had an initial screening test done and provided that an outcome had been recorded.The proportion of the per protocol and intention-to-treat populations experiencing each primary and secondary outcome for the treatment groups, and the associated 2-sided 95% CI for the difference, was estimated using the generalized linear model. To declare non-inferiority with a significance level of 0.05%, the upper boundary of the 2-sided 95% CI for the estimated treatment effect (risk difference) had to be below the pre-defined non-inferiority margins (\u0394) of 5% and 4% for third trimester severe anaemia and low birth weight respectively. We controlled for gestational age at enrolment, gravidity, baseline parasitaemia and anaemia using binomial regression.A total of 3333 of the 3472 potentially eligible pregnant women who were screened (96%) were enrolled into one of the three treatment groups . At the P. falciparum was the most prevalent malaria species. P. malariae and P.ovale were also detected but only few women (<3%) carried these species. Additional blood samples were obtained 14 and 28 days after administration of a first dose of SPI-IPTp from 71 women who were parasitaemic on presentation. Eleven  were positive by day 28. PCR corrected parasitological failure on day 28 was 5.6% (95% CI 1.6%-13.8%). The overall prevalence of HIV in pregnant women who agreed to screening in the study clinics during the period of the trial was approximately 1.5%.Baseline characteristics of women in the three treatment groups were very similar at enrolment . In all At 36\u201340 weeks of gestation (before delivery) the prevalence of severe anaemia (Hb<8 g/dl) and moderate (8<\u200a=\u200aHb<11 g/dl) anaemia was respectively 1.7% and 45.9% overall and was similar in all treatment groups; similarly asymptomatic parasitaemia was 12.0% in study women overall and was very similar in all treatment groups. The mean Hb concentration of all women between 36 and 40 weeks of gestation was 11.0 g/dl and was similar in all the groups. Generally, there was a significant increase in mean Hb concentration of 0.29 g/dl at 36\u201340 weeks gestation over the baseline Hb concentration . The incThe risk of anaemia (severe or moderate) did not differ significantly between the three treatment groups across gravidity . It was Similarly, the risk of low birth weight did not differ significantly between the three treatment groups regardless of gravidity; . The upper boundaries of the 2-sided 95% CI for the risk differences estimated between women in the SP-IPTp group and the IST lie below the non-inferiority margin of 4% set for low birth weight . In a unOne hundred and forty-seven episodes of illness associated with malaria parasitaemia were recorded among study women between scheduled antenatal clinic visits; 44, 51 and 52 of these were in SP-IPTp, IST with SP, and IST with AS+ AQ groups respectively. The differences in the illness episodes are not statistically significant.In all 2211 RDT tests were done in women in treatment groups 2 and 3 respectively of which 23% and 23% were positive and led to treatment. The risk of third trimester severe anaemia and low birth weight in those who were RDT negative throughout pregnancy and did not receive antimalarial treatment were no higher than in the SP-IPT arm. But the risk of parasitaemia at 36\u201340 weeks was higher in the SP-IPT arm than in the screening and treatment arms . Supplementary This study has shown that IST, using either SP or AS + AQ, was not inferior to IPTp with SP in preventing maternal anaemia and low birth weight, according to the non-inferiority criteria that were set prior to the trial, in women who used an LLIN in an area of moderately high malaria transmission in Ghana. There are a number of possible reasons why IST performed as well as SP-IPTp in our study. Firstly, it is possible that malaria is not an important cause of anaemia or low birth weight in the study area, in which case it would be difficult to detect an impact of different control strategies on the prevalence of these complications of malaria in pregnancy unless a much larger trial than the one we did was undertaken. We think that this is an unlikely explanation for our findings for several reasons. The study area is one of derived forest with perennial malaria transmission, although with a pronounced seasonal peak. The entomological inoculation rate in the neighbouring area of Kintampo was recently estimated to be 267 infective bites per year Demonstration of efficacy is only one aspect of the evaluation of a potentially new control tool; other important aspects are the ease with which it could be implemented, its acceptability and its cost. These aspects of IST have been evaluated in the current study and compared with those of SP-IPTp. The results of these studies will be presented in detail elsewhere. However, preliminary analysis of the findings studies indicates that IST can be introduced into a busy, antenatal clinic without disrupting its function. The approach appears acceptable to women who are used to receiving IPT with SP provided that the rationale for the intervention is explained to them Available evidence suggests that RDTs detect circulating parasite antigen and so compared to microscopy are better at detecting sub microscopic parasitaemia and may be a reliable indicator of placental infections No difference was found between the prevalence of peripheral blood parasitaemia between the treatment groups at 36\u201340 weeks of pregnancy or at six weeks post partum but the possibility that IST with SP or AS+AQ is not as effective as IPT with SP in preventing infection of the placenta cannot be excluded. This study had only limited financial resources which precluded analysis of the impact of each of the study interventions on the prevalence of placental malaria as measured by histology. How important such a difference might be in the absence of any apparent clinical adverse effects is uncertain but this needs to be investigated. Other weaknesses of this study are the absence of linked data on HIV positivity, although it is known that the overall prevalence of HIV positivity in the study population is relatively low, approximately 1.5% in the study clinics.We are confident that the major finding of this study that IST is not inferior to SP-IPTp in the prevention of anaemia in pregnancy and low birth weight is sound and that IST is a potentially promising strategy for the control of malaria in pregnancy, especially in areas where the incidence of malaria is decreasing or where malaria is very seasonal. However, it is important that these encouraging findings including birth outcomes and maternal morbidity findings are confirmed in other geographical areas including parts of eastern and southern Africa, where SP is now failing since we have no alternative drug yet for SP and because the next generation of potential drugs are less likely to fulfill the same favourable profile of SP. As part of these it is also important that the impact of IST on placental malaria is investigated. A large multicentre trial of IST  has started in four countries in Africa where malaria transmission is markedly seasonal will address these issues.Checklist S1(0.06 MB DOC)Click here for additional data file.Protocol S1(0.16 MB DOC)Click here for additional data file.Table S1Comparison of SP-IPTp with IST in women who were RDT negative throughout pregnancy and did not receive an antimalarial.(0.06 MB DOC)Click here for additional data file.Table S2Comparison of pregnancy outcomes for singleton births.(0.03 MB DOC)Click here for additional data file.Table S3Comparison of number of women who experienced adverse events within seven days of drug administration.(0.04 MB DOC)Click here for additional data file.Table S4Comparison of key outcomes in women enrolled in SP-IPTp or IST groups.(0.06 MB DOC)Click here for additional data file.Table S5Factors associated with third trimester severe anaemia in study women(0.05 MB DOC)Click here for additional data file.Table S6Factors associated with low birth weight of babies delivered by study women(0.05 MB DOC)Click here for additional data file."}
+{"text": "Antimalarial resistance has led to a global policy of artemisinin-based combination therapy. Despite growing resistance chloroquine (CQ) remained until recently the official first-line treatment for falciparum malaria in Pakistan, with sulfadoxine-pyrimethamine (SP) second-line. Co-treatment with the gametocytocidal primaquine (PQ) is recommended for transmission control in South Asia. The relative effect of artesunate (AS) or primaquine, as partner drugs, on clinical outcomes and gametocyte carriage in this setting were unknown.A single-blinded, randomized trial among Afghan refugees in Pakistan compared six treatment arms: CQ; CQ+(single-dose)PQ; CQ+(3 d)AS; SP; SP+(single-dose)PQ, and SP+(3 d)AS. The objectives were to compare treatment failure rates and effect on gametocyte carriage, of CQ or SP monotherapy against the respective combinations (PQ or AS). Outcomes included trophozoite and gametocyte clearance (read by light microscopy), and clinical and parasitological failure.A total of 308 (87%) patients completed the trial. Failure rates by day 28 were: CQ 55/68 (81%); CQ+AS 19/67 (28%), SP 4/41 (9.8%), SP+AS 1/41 (2.4%). The addition of PQ to CQ or SP did not affect failure rates (CQ+PQ 49/67 (73%) failed; SP+PQ 5/33 (16%) failed). AS was superior to PQ at clearing gametocytes; gametocytes were seen on d7 in 85% of CQ, 40% of CQ+PQ, 21% of CQ+AS, 91% of SP, 76% of SP+PQ and 23% of SP+AS treated patients. PQ was more effective at clearing older gametocyte infections whereas AS was more effective at preventing emergence of mature gametocytes, except in cases that recrudesced.CQ is no longer appropriate by itself or in combination. These findings influenced the replacement of CQ with SP+AS for first-line treatment of uncomplicated falciparum malaria in the WHO Eastern Mediterranean Region. The threat of SP resistance remains as SP monotherapy is still common. Three day AS was superior to single-dose PQ for reducing gametocyte carriage.ClinicalTrials.govbold> In South Asia, the standard policy has been to co-treat with a single dose of primaquine to reduce gametocyte carriage. Primaquine is known to have poor efficacy as a direct treatment but has repeatedly been shown to be highly gametocytocidal A direct comparison of artesunate and primaquine on treatment failure and gametocyte carriage has not been conducted before. Many Afghan refugee communities in Pakistan have a history of falciparum malaria and are prone to outbreaks The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Checklist S1 and Protocol S1.The study recruited patients from five Afghan refugee villages within an 80 km radius of Peshawar, Khyber Province (formerly North West Frontier Province), Pakistan. The 5 villages were established in the early 1980s and each has a history of falciparum transmission. Three  are situated on the banks of the Kabul river, and two (Mohammed Khoja and Kotki) are situated to the south of Peshawar in Kohat district. Malaria transmission is seasonal with vivax malaria occurring from March to November and falciparum from July to December.Participants were Afghan refugees who were permanently resident in the Pakistan villages. A minority might have acquired their infections in Afghanistan, but admission criteria required 4\u20136 weeks of follow-up which would have excluded the more regular travellers.The study was conducted through three sites with well-managed clinics run by Non-Governmental Organisations (NGO) and government agencies. Site 1 was Adizai village which also served the population of nearby Naguman. Site 2 was Yakka Ghund village. Site 3 was Kotki which also serviced Mohammed Khoja village. Health staff from the NGO HealthNet TPO were seconded to each clinic to manage the recruitment and follow-up of patients.in vivo survey across all refugee camps along the length of the 1000 km North-South axis of Khyber Province showed little heterogeneity in resistance frequency to chloroquine and SP The study took place over three malaria seasons from 2000 to 2003. In the first season only site 1 was used, in the second season site 2 was added, and in the third season site 3 was added. The stepped inclusion of sites resulted from unexpectedly low recruitment rates at the first study site, and is summarized in Study design and procedures. Individuals presenting with clinical symptoms and diagnosed microscopically with falciparum malaria were referred to trial staff for further assessment. Inclusion criteria were: over two years of age, not pregnant, P. falciparum mono-infection, more than 1 asexual parasite per 10 fields, no other serious disease, resident in the refugee village for the full period of follow-up, no verbal report of antimalarial use during the last 21 days, and no signs of severe malaria.1) (25 mg/kg) over 3 days; CQ (25 mg/kg) over 3 days plus primaquine (PQ) (0.5 mg/kg) on the last day of treatment; CQ (25 mg/kg) over 3 days plus artesunate (AS) (4 mg/kg per day) over 3 days; sulfadoxine (25 mg/kg) and pyrimethamine (1.25 mg/kg) (SP) on day 0; SP (25:1.25 mg/kg) plus PQ (0.5 mg/kg) on day 0; SP (25:1.25 mg/kg) on day 0 plus AS (4 mg/kg per day) over 3 days. The primaquine co-treatment regimens were based on recommendations for use of primaquine as a gametocytocide treatment for falciparum malaria Individuals meeting the inclusion criteria and giving informed consent were allocated using randomization tables stratified by age and sex to the following treatment arms: chloroquine . The assignment of treatment group to patient number was concealed until after enrolment. The random allocation sequence was generated by two of the investigators  neither of whom enrolled or assessed patients.\u00ae, Roche), chloroquine , primaquine (IDA), artesunate . Although chloroquine resistance was known to exist in the study area, chloroquine was the first line treatment policy of the Government of Pakistan and the UN refugee agency (UNHCR) at the time and was therefore included.The brands and manufacturers/suppliers were: SP  the appearance of the different drugs, and (ii) the different times of follow up for the SP arms compared to the chloroquine arms.Patients were given directly observed treatment , monitored for 30 minutes and re-dosed if vomiting occurred. Health workers then recorded the other day 0 biomedical parameters.Patients were asked to return on each day of treatment  and on days 3, 7, 14, 21 and 28 post treatment. Patients in the three SP arms were also followed up on days 35 and 42 for detection of late recrudescence in lieu of mefloquine monotherapy which was unavailable in the study area. Those who developed severe malaria, severe anaemia or other complications were referred to Khyber Hospital, Peshawar for treatment.Patients found to be parasitaemic on any day after day 3 were treated with SP  or with SP and mefloquine  for those whose initial treatment was SP based. SP resistant parasites were sensitive to the mefloquine component of Fansimef, which was used Laboratory tests. Thick and thin blood smears were stained with 2% Giemsa solution. All slides were read on the day of collection by a microscopist based at each site. Differential diagnosis of vivax and falciparum (trophozoites and gametocytes) was by examination of thick and thin smears according to standard microscopy methods Outcome measures. Patients completed the trial if treatment was administered fully and all follow-up appointments conducted, or if they failed treatment on any day of follow-up. The primary endpoint of the trial was any clinical or parasitological failure up to day 28, although a subset of patients in the SP treatment arms was also followed until day 42. Patient outcomes were classified under the WHO parasitological classification system of sensitive (S) or resistant  infections with those whose outcomes were classified as S (sensitive) being treatment successes and those with outcomes of RI, RII or RIII being classified as treatment failures Other parameters examined were time to fever resolution (with fever defined as axillary temperature \u226537.5\u00b0C), asexual parasite clearance, gametocyte clearance and gametocyte carriage on or after day 7.Molecular characterisation. PCR genotyping was conducted on a subset of samples to distinguish recrudescent from new infections using the protocol described by Brockman et al P. falciparum genes msp-1, msp-2 and glurp were genotyped according to polymorphisms present at variable loci on day 0 and day of failure. Approximately 50% of cases were available for genotyping.PCR corrected outcomes were applied as a secondary analysis using redefined outcomes based on the number and frequency of true recrudescent infections and reclassifying cases with new infections as treatment successes, excluding those with indeterminate outcomes or negative PCR results. PCR testing was performed at the Shoklo Malaria Research Unit, Thailand.pfcrt, pfmdr1, pfdhfr and pfdhps genes pfcrt codon 76, pfmdr1 codons 86 and 184, pfdhfr codons 16, 51, 59 and 108 and pfdhps codons 436, 437, 581 and 613 We also conducted an analysis at the London School of Hygiene and Tropical Medicine (LSHTM), UK, for known resistance-associated mutations to chloroquine and antifolate drugs. Samples collected at enrolment were tested for known mutations in the The sample size required to detect a difference in treatment failure or gametocyte carriage with 80% power and 95% confidence (two-sided significance level for type 1 error of 0.05) were calculated using the following predictions. In the chloroquine arms (i) the estimated frequency of failure in chloroquine monotherapy arm was 30% and in each of the combination arms (CQ+PQ or CQ+AS) it was 10% or less; (ii) the estimated proportion of gametocyte positive individuals after 7 days in the chloroquine arm was 50% and in each of the combination arms it was 25%. In the SP arms (i) the estimated frequency of failure in the SP arm was 10% and in each of the combination arms (SP+PQ or SP+AS) it was 1%; (ii) the estimated frequency of gametocyte positive patients after 7 days in the SP arm was 50% and in each combination arm it was 25%. The estimated samples sizes were 64 per arm in the chloroquine arms and 121 per arm in the SP arms. For gametocyte carriage the required sample sizes were 65 per arm. The target sample sizes for the study were 65 for the chloroquine arms and 121 for the SP arms. To allow for 15% loss to follow up the targets for recruitment were 76 for the chloroquine arms and 142 for the SP arms.The primary aims of the study were to evaluate: 1) the relative efficacy of the combination drugs in providing parasite clearance with no recrudescence compared to monotherapy, and 2) the effect on gametocyte clearance. The secondary aim was to determine the proportion of individuals in each arm with classifiable treatment failure.2 test. Treatment failure was also analysed using the definitions of ACPR and ETF or LTF, and parasitological classifications using the S-R scale in addition to the PCR corrected analysis The primary outcome was the proportion of individuals in each treatment group classified as a treatment failure during the 28 days follow-up compared to the respective monotherapy. The odds of treatment failure between each of the study groups at day 28 were estimated after adjusting for potential confounders  using logistic regression analysis or Mantel-Haenszel \u03c7For assessing potential effects on gametocyte carriage, the primary outcome was the presence or absence of gametocytes on day 7 with a secondary analysis examining the presence or absence of gametocytes and the geometric mean gametocyte density on each day of follow-up.All data were entered in Microsoft Excel (1997) by one data clerk and the database checked patient by patient against paper records by a second data clerk. Analysis was performed using STATA 10.0 .The study protocol was approved by the ethics committees of the Pakistan Medical Research Council and LSHTM, UK. All patients or their guardians gave written informed consent to participate. The trial was registered under clinicaltrials.gov [NCT00959517].A total of 355 cases of microscopically confirmed falciparum malaria were enrolled into the study between July 2000 and December 2002. Characteristics of patients recruited into the 3 chloroquine arms were broadly similar, as were the three SP arms. The SP+AS arm showed slightly higher asexual parasite densities on enrolment than other groups .Of 355 patients enrolled, 38 (10.7%) were either withdrawn or lost to follow up leaving 317 for possible inclusion . Of thesClinical Outcomes. The 28-day failure rate in the CQ and CQ+PQ arms was 81% and 73% respectively  were associated with treatment outcome so adjustments were unnecessary in the final regression analysis. Crude odds ratios are presented.Clinical and parasitological outcomes are shown in Trophozoite clearance time (days until negative smear) was lowest for the artesunate combination arms. Addition of primaquine to either chloroquine or SP did not appear to affect clearance times. Clearance times for each arm (median (interquartile range)) were: CQ, 3 days (2\u20137); CQ+PQ, 3 days (2\u20137); CQ+AS, 2 days (1\u20132); SP, 2 days (2\u20133); SP+PQ, 2 days (1.5\u20133); SP+AS, 1 day (1\u20132).Of the 133 failures, 79 (47%) matched pairs were available for PCR evaluation of MSP-1, MSP-2 and GLURP. Matched pairs were collected on day 0 and on the day of failure. Outcomes of the PCR are shown in In the subset of patients followed-up for 42 days, recrudescence between day 28 and 42 was rare: only 8/317 (2.5%) of individuals classified as adequate clinical and parasitological response (ACPR) or sensitive (S) at day 28 went on to fail by day 42 . Failure rates at day 42 were therefore similar to day 28. The failures between 28-day and 42-day rates were not corrected by PCR and hence could have resulted from new infections.Molecular outcomes. Molecular analysis of drug resistance-associated alleles was conducted on samples taken on day 0  were seen frequently; 108 N was found in all but one of 74 typable samples (98.6%) and 59R occurred in 66/76 (86.8%) samples. However, 51I was only detected in 5/75 (6.7%) samples. Therefore pfdhfr \u2018double mutants\u2019 were common (at codons 108+59) but \u2018triple mutants\u2019 were rare. Interestingly, we observed one sample with the pfdhfr mutations 16V+108T. These are reportedly selected by the antifolate drug cycloguanil, rather than pyrimethamine dhps gene were seen. These PCR results broadly match what would be expected from the clinical outcomes. The pfdhr/pfdhps mutation profile suggests that the parasites would be killed by a full therapeutic dose of SP, but that some tolerance to lower doses existed.on day 0 . The majThe addition of artesunate or primaquine to chloroquine or SP reduced gametocyte carriage on day 7 . CombiniMost patients did not have gametocytaemia on day 0 (291/355 (82.0%) had no gametocytes on day 0). However, the presence or absence of sexual stage parasites on day 0 was an important explanatory variable in the secondary analysis; patients who presented with gametocytes on day 0 were more likely to be gametocytaemic on day 7 than individuals who were not gametocytaemic on day 0 regardless of treatment group . We therefore conducted further analysis comparing those who presented with gametocytes on day 0 and those who did not.did have patent gametocytaemia on day 0, 8/9 were still gametocytaemic on day 7 after treatment. This was not significantly different from the proportion gametocytaemic on day 7 (11/12) after treatment with chloroquine monotherapy (Fisher's exact test p\u200a=\u200a1.0). By contrast, in the chloroquine arms of the patients who did not have patent gametocytaemia on day 0, only 11% (7/63) were gametocytaemic on day 7 when treated with CQ+AS as compared to 47/56 (84%) among those treated with chloroquine monotherapy (\u03c72 p<0.001). Similar trends were evident in the SP arms treated with artesunate; prevalence of gametocytaemia on day 7 was significantly lower in the SP+AS group that was not gametocytaemic on day 0 than in the group that was (Fisher's exact test p<0.001). This indicates that artesunate is less active against older gametocytes than against those newly emerged or immature forms that are not yet emerged. Primaquine showed no such trend: the proportion of patients gametocytaemic on day 7 was not significantly different for patients who were or were not gametocytaemic on day 0. Primaquine therefore appears to be active against gametocytes of all ages.The artesunate treatments were more effective at preventing the emergence of gametocytaemia between day 0 and 7 than in removing established gametocytaemia already present on day 0 . For exaPrimaquine was more effective at controlling gametocytaemia when combined with CQ than with SP, regardless of whether individuals were gametocytaemic at the start of treatment  . By contUsing the presence of gametocytes on day 7 as the secondary endpoint, adjusted for the presence of gametocytes at day 0, both primaquine and artesunate with chloroquine or SP were more effective than their respective monotherapies at reducing the presence of gametocytes .Compared to monotherapy with chloroquine or SP, co-treatment with artesunate was negatively associated with having gametocytes on day 28 whether the accompanying drug was chloroquine or SP . The preThe present study was designed to inform decision-making at several levels: to guide UNHCR on appropriate treatment in the Afghan refugee communities in Pakistan, to build evidence for national treatment policy in the Pakistan Directorate of Malaria Control and in the Afghanistan Ministry of Public Health, to guide the WHO Eastern Mediterranean Region Office (EMRO) on regional treatment recommendations, and to build evidence for international epidemic response guidelines. The findings of this study, when presented at an inter-country meeting of national malaria control managers coordinated by WHO/EMRO in 2004, contributed to a shift in policy from chloroquine monotherapy to ACT in South and West Asia SP monotherapy was effective in the study population, with treatment failure rates remaining in the 5\u201310% range, similar to that seen in other studies in the region The complete failure of chloroquine  convinced policy makers of the need to redefine treatment practices in South Asia. The new data demonstrated higher levels of resistance than in previous studies dating back to the 1980s and 1990s A constraint on the study design was that not all 6 arms could be included at each of the three study sites. Site is therefore a potential confounding covariate that cannot be fully controlled for in the data analysis. Differences in transmission could, for example, affect the risk of re-infection between sites but this was corrected for by the PCR analysis. At each site in any one year each triplet of treatment arms  were examined at the same time, and it is comparisons within these triplets which are of greatest interest. The study did not complete its target sample size for the SP arms because of low recruitment rates.PCR corrected outcomes were not attainable for all failures, and higher numbers of patients were reinfected at one site where only the chloroquine arms were being tested. Low rates of transmission at the other sites make it unlikely that failing cases were due to reinfection. The secondary analysis with PCR-corrected outcomes did not change the overall conclusions on the suitability of the combinations for revised treatment policy.Treatment with either the clinically effective SP monotherapy or the failing chloroquine monotherapy resulted in persistence of gametocytaemia into the second and third weeks in the majority of individuals. This effect was more evident for SP than for chloroquine, SP being well known for high rates of gametocyte carriage post treatment The effect of artemisinin derivatives on reducing gametocyte carriage is already documented Primaquine is more rapidly excreted than artesunate and this may account for the fact that the single dose of primaquine used in this study had a lower impact on gametocyte carriage than the artesunate regimens. The usefulness of primaquine as a gametocytocidal treatment may be improved by administering it after the ACT course In our study, a key factor in the clearance of gametocytes was the presence or absence of gametocytes on day 0. Those who presented with gametocytes were more likely to have gametocytes on day 7, an effect which was independent of the treatment given. Although the majority of patients (80%) presented without gametocytaemia, artesunate appeared to be less active against these older parasites whereas primaquine appeared to be effective against all ages. The proportion presenting with gametocytes may vary according to background transmission levels. If gametocytes persist after treatment with ACTs, the effect on transmission in areas where a high proportion of cases present with gametocytes may prove suboptimal and justify the simultaneous use of primaquine.Primaquine is more rapidly excreted than artesunate and this may account for the fact that the single dose of primaquine used in this study had a lower impact on gametocyte carriage than the artesunate regimens. The usefulness of primaquine as a gametocytocidal treatment may be improved by administering it after the ACT For treatment policy to have a major impact on transmission several criteria need to be met: i) transmission is low to moderate; ii) the majority of people use public health rather than private facilities, or effective private sector interventions ensure adherence to policy; iii) public health facilities correctly prescribe the approved regimen, and iv) patients take the full course. These conditions are largely met in the refugee villages of Pakistan. The region is characterized by low endemicity and low immunity, well supported health care facilities are available in the Afghan refugee villages, and diagnosis of malaria to species level is maintained to a high standard The operational data from this region and elsewhere suggest that drugs showing faster gametocyte clearance in clinical trials do help to reduce transmission The effect of drugs on gametocyte carriage cannot be fully determined if only standard light microscopy (LM) is used. Research in the last decade shows that LM gives starkly different indications of gametocyte prevalence and densities compared to other techniques Gametocytes that are readily identified in blood following treatment may or may not be viable. Gametocytes may not be infective to mosquitoes either because they are recently emerged Click here for additional data file.Checklist S1CONSORT Checklist.(DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria in Tanzania, when provided by community health workers (CHWs) and administered unsupervised by parents or guardians at home.Home-management of malaria (HMM) strategy improves early access of anti-malarial medicines to high-risk groups in remote areas of sub-Saharan Africa. However, limited data are available on the effectiveness of using artemisinin-based combination therapy (ACT) within the HMM strategy. The aim of this study was to assess the effectiveness of artemether-lumefantrine (AL), presently the most favoured ACT in Africa, in under-five children with uncomplicated P. falciparum parasitaemia were definitely enrolled and reviewed weekly by the CHWs during 42 days. Primary outcome measure was PCR corrected parasitological cure rate by day 42, as estimated by Kaplan-Meier survival analysis. This trial is registered with ClinicalTrials.gov, number NCT00454961.An open label, single arm prospective study was conducted in two rural villages with high malaria transmission in Kibaha District, Tanzania. Children presenting to CHWs with uncomplicated fever and a positive rapid malaria diagnostic test (RDT) were provisionally enrolled and provided AL for unsupervised treatment at home. Patients with microscopy confirmed A total of 244 febrile children were enrolled between March-August 2007. Two patients were lost to follow up on day 14, and one patient withdrew consent on day 21. Some 141/241 (58.5%) patients had recurrent infection during follow-up, of whom 14 had recrudescence. The PCR corrected cure rate by day 42 was 93.0% (95% CI 88.3%-95.9%). The median lumefantrine concentration was statistically significantly lower in patients with recrudescence  compared with reinfections , or no parasite reappearance .Provision of AL by CHWs for unsupervised malaria treatment at home was highly effective, which provides evidence base for scaling-up implementation of HMM with AL in Tanzania. Malaria still causes significant morbidity and mortality, primarily among under-five children in sub-Saharan Africa. Access Plasmodium falciparum malaria [Due to widespread resistance to chloroquine and sulphadoxine/pyrimethamine, most malaria-endemic countries in Africa and Asia have adopted the WHO recommendation of introducing artemisinin-based combination therapy (ACT) as first-line treatment for uncomplicated  malaria . Artemet malaria . Althoug malaria -14. More malaria -17. Thes malaria ,19. Anot malaria .To improve targeting of ACT to malaria infected patients parasitological confirmation is essential at community level in remote areas of sub-Saharan Africa, where a majority of fever patients seek care. Antigen-based rapid malaria diagnostic tests (RDTs) may represent an important tool to improve the diagnostic efficiency if incorporated in the HMM strategy, considering that they are easy to use and interpret and do neither require access to skilled technicians nor electricity. Previous studies have shown that RDTs can be accurately used by CHWs ,22. ReceP. falciparum malaria in under-five children, during an extended follow-up period of 42 days, adherence to treatment by measuring lumefantrine concentrations on day 7 after initiation of treatment, and possible selection of genetic markers associated with AL tolerance/resistance.This study reports data on polymerase chain reaction (PCR) corrected effectiveness of AL, when provided at community level by CHWs and used unsupervised by parents or guardians at home for treatment of uncomplicated Plasmodium falciparum is the dominant parasite species. There is one dispensary in each village and most referrals are sent to Mlandizi Health Centre, situated at a distance of 9 to 10 kms from the selected villages. AL was the first-line treatment for uncomplicated malaria at the time of the trial.The study was conducted between March-August 2007 in two neighbouring villages, Ngeta and Mwanabwito, in rural Kibaha District, located about 50 km west of Dar es Salaam, Tanzania. These two villages, with a total population of 4,500 people, were among five villages involved in a previous assessment of RDT use by CHWs to improve targeting of ACT at the community level. The stu\u00ae, according to the manufacturer's instructions  and simplified pictorial instructions (RDT job aid); prepare thick blood smears and blood spots on filter paper (Whatman 3MM) from finger prick blood samples; and filling of case record forms. Each CHW was provided with a weighing scale, a digital thermometer, pre-packed AL tablets, paracetamol tablets, a dosing chart for AL, RDTs, RDT job aid and a storage iron box. The CHWs were responsible for referral of patients and storage of the study drugs and other supplies for the study. During the study period each CHW was given a monthly allowance equivalent to 25 USD.After consultations with dispensary staff and village leaders, six CHWs were selected among the existing 40 CHWs in the two villages. The selection of study CHWs was based on gender balance, residential area and ability to keep records. A three-day training workshop was conducted at each village dispensary. The CHWs were trained on how to recognize symptoms of both uncomplicated and severe malaria, as well as other febrile childhood non-malaria diseases, such as pneumonia (cough with fast breathing) and acute diarrhoea, as well as in administration of AL and how to educate caregivers on the use of AL. The CHWs were also trained on how to perform and interpret RDT, i.e. Paracheck PfP. falciparum malaria confirmed by RDT and later by microscopy; able to ingest tablets orally; able to attend stipulated follow-up visits; provision of written informed consent by a parent or guardian, and absence of any general danger signs of severe disease . Patients with negative RDT test results and/or signs of severe disease were to be immediately referred to the village dispensary for further management.Children presenting to CHWs with symptoms suggestive of uncomplicated malaria were screened for study eligibility. Patients were enrolled if they met the following inclusion criteria: age of 3-59 month; body weight of \u22655 kg; fever (\u2265 37.5\u00b0C axillary) or a history of fever in the preceding 24 hours; P. falciparum infection with an extended follow-up of 42 days. Patients fulfilling the inclusion criteria were provisionally enrolled based on a positive RDT result and provided AL treatment in standard doses according to the body weight: one tablet per dose for patients weighing 5-14.9 kg (yellow blister pack) and two per dose for 15-24 kg (blue blister pack). Every blister pack had pictures showing how the drug should be given. The first dose was given under supervision of the CHWs. If patients vomited the first dose within 30 min; administration of the full dose was repeated. The other five doses were given by the parents/caregivers at home. Standardized verbal instructions on dose, frequency and advice to combine treatment with fatty meals or breast milk were given by CHWs. Paracetamol was provided to all febrile patients. Parents or guardians were instructed to report to the CHWs or the village dispensary if the child's condition had not improved after 48 hours or if worsened at any time. Anti-malarial medicines and other drugs in the study were provided by the research project free-of-charge.This was an open label, single arm prospective study, assessing PCR corrected parasitological cure rate of a six-dose treatment with unsupervised intake of AL provided by CHWs to under-five children with uncomplicated Blood smears were stained with Giemsa and read at the Department of Parasitology, MUHAS. The results were provided to the CHWs and patients within three days of sampling. Children with positive RDTs but negative blood smear results were excluded from the study and referred to the dispensary. Children with microscopically confirmed malaria were definitely enrolled. Parents or guardians were requested to bring their children back to the CHWs on days 7, 14, 21, 28, 35 and 42 or on any day that they felt ill. At each visit, children were assessed for symptoms, possible adverse events, and body temperature. Finger prick blood samples were collected for microscopy (thick blood smears) and PCR analysis (on Whatman filter paper), respectively, at day of enrolment and at each follow-up visit. In addition, the research team leader collected blood samples (100 \u03bcl taken by capillary tube) on pre-treated filter papers on day 7, which were used later for determination of lumefantrine blood concentrations. If patients did not return for scheduled follow-up visits, they were actively followed-up in their homes the next day.Patients with recurrent infections within 14 days after initiation of AL treatment and in children who developed symptoms or signs of severe malaria during follow-up were withdrawn from the study, and referred to the village dispensary or Mlandizi health centre for rescue treatment with quinine. In children with clinical failure  after day 14 or parasitological failure day 42 after the initial therapy were retreated with AL in accordance with national guidelines .All thick blood smears were picked up from CHWs by the research team within 24 hours after collection and stained with 5% Giemsa for 20 minutes at MUHAS. Two qualified microscopists independently read all thick blood smears. The parasite density was estimated by counting the number of asexual parasites per 200  white blood cells and calculating parasites per \u03bcL, assuming a white blood cell count of 8,000 per \u03bcL. A smear was declared negative if no asexual parasites were seen after examining 200 high power fields. Disagreement in readings (positive versus negative) or an at least two fold difference in parasite density, a third decisive microscopy reading, again blinded was to be done.P. falciparum genes merozoite surface protein-2 (msp2), glutamate-rich protein (glurp) and msp1 using standard protocols [Distinction between recrudescence and reinfection events was performed by the use of stepwise genotyping of the rotocols . RecrudeP. falciparum chloroquine resistance transporter gene (pfcrt) K76T and P. falciparum multidrug resistance gene 1 (pfmdr1) N86Y, previously associated with quinoline resistance, were analysed according to established ApoI restriction enzyme-based PCR restriction fragment length polymorphism (PCR-RFLP) protocols [Single nucleotide polymorphisms (SNPs) in the rotocols .Capillary blood samples  were applied on filter papers pre-treated with 0.75 M tartaric acid. These blood samples were dried, put in small zipped plastic bags and frozen at -20\u00b0C the day after sampling. After completion of the field trial they were transported to the Bioanalytics and Pharmacokinetics laboratory of Dalarna University, Sweden, where lumefantrine blood concentrations were measured by solid-phase extraction and liquid chromatography .pfcrt and pfmdr1; and adverse events defined as signs and symptoms that occurred or increased in severity after treatment started.The day-42 PCR corrected parasitological cure rate was the primary efficacy outcome, i.e. proportion of patients with clearance of asexual parasitaemia within seven days of initiation of treatment, without recrudescence within 42 days after initiation of treatment, and without use of rescue medication for clinical signs of malaria. Secondary outcomes included PCR corrected parasitological cure rates on days 14 and 28 after treatment; risk of reinfections after treatment; day 7 blood lumefantrine concentrations; selection of genetic markers (SNPs) in Assuming that AL treatment provided by CHWs would result in less than 85% PCR corrected parasitological cure rate by day 42. According to the WHO protocol , with an2 test or Fisher's exact test as appropriate. All patients with microscopically confirmed P. falciparum malaria who had taken at least one full dose of the study medication and had at least one post-baseline efficacy assessment were included in the analyses. Cure rates and cumulative proportion of recurrent parasitaemia were estimated by Kaplan-Meier survival analysis. Data were censored for patients who were lost to follow-up or for patients with reinfections, and indeterminate or missing PCR results at the last day seen.Data were double-entered and validated using EpiData software, version 3.02  and analysis was performed using Stata 10.0 . Proportions were compared with XThe study was approved by the National Institute for Medical Research in Tanzania and Regional Ethics Committee Stockholm, Sweden, and was registered with identifier NCT00454961. Before P. falciparum positive by microscopy  were provisionally enrolled based on a positive RDT, of whom 250 (83.3%) were y Figure . Of thesy Figure . Three cy Figure ; one dueNo early treatment failure was reported. On day 7, all patients were afebrile, but 7/244 (2.9%) were still parasitaemic. PCR analysis of these seven patients revealed four reinfections, two recrudescences and one indeterminate result. By day 42, some 141/241 (58.5%) patients had recurrent parasitaemia  patients had blood lumefantrine concentrations measured on day 7 after initiation of treatment. The median (range) lumefantrine concentration on day 7 was 205 ng/mL (0-1887). The median lumefantrine concentration was significantly lower in patients with recrudescence  than in those with reinfections , or no parasite reappearance  , while the pfcrt K76T SNP did not show any pattern of selection upon AL administration.Proportions of parasites carrying AL was well tolerated. No deaths occurred. Two serious adverse events were reported, both were due to severe malaria on days 14 and 42. In total 162 adverse events (mild or moderate in severity) were reported during follow-up, the most common being fever (34%), cough (34%) and diarrhoea (12%). These events were considered unrelated to AL treatment.P. falciparum malaria in Tanzanian children below five years of age. This study is the first to assess the PCR corrected cure rate achieved with ACT in the context of HMM after an extended follow-up to 42 days. A previous multicentre study showed high effectiveness of ACT used in HMM [The results from this study showed that intake of unsupervised AL used in HMM was highly effective and well-tolerated for the treatment of acute uncomplicated d in HMM . Howeverd in HMM -14.However, a concern is the limited post-treatment prophylactic effect of AL in this high malaria transmission area, which resulted in more than half of the patients having recurrent infections within the 42-day follow-up period. A great majority of all recurrent infections were due to reinfections, when assessed with stepwise genotyping using three polymorphic genetic markers. Young children with little or no immunity to malaria may have a shorter post-treatment prophylactic effect compared to older children and adults . SimilarAdherence to AL treatment in this trial was assessed by measuring day 7 lumefantrine levels, a potentially more reliable method of measuring adherence to treatment than the use of questionnaires, which has a potential for recall bias ,39. The pfmdr1 N86 allele in recurrent infections after AL treatment [pfcrt K76T SNP - in contrast to a recent study conducted in Bagamoyo District, Tanzania [pfcrt K76 allele in recurrent infections after AL treatment was not observed. This might be due to lack of statistical power of the present study, taking in account the high baseline prevalence of the pfcrt K76 (> 80%). It can also reflect that although the potential of pfcrt K76T to modulate is by now well supported in vitro [in vivo importance might be less general than the observed for pfmdr1 86N, being more dependent on the overall genetic makeup of the studied parasite populations.In high transmission areas, residual low/sub-therapeutic concentrations of the long acting partner drug in ACT create strong selective pressure for development and spread of tolerant/resistant parasites ,45. In treatment -50. ThisTanzania , a signiin vitro ,51, its Selecting study sites that were geographically accessible within three hours by car from Dar es Salaam might have introduced a selection bias. However, the selected villages were not different from those not included in the study in terms of number and quality of CHWs. Moreover, the weekly follow-up and supervision by researchers may have influenced the behaviour of CHWs and caregivers and resulted in improved adherence, however, supervision and monitoring of the community activities is one of the key components of the HMM strategy.in vivo selection of genetic markers associated with drug resistance confirms that AL is vulnerable to selection of resistance-related polymorphisms in areas of high malaria transmission. It is crucial that the HMM strategy incorporates components for monitoring adherence to ACT treatment and drug resistance.In conclusion, this study showed high PCR corrected effectiveness of AL used in HMM during an extended follow-up of 42 days, which provides evidence for scaling-up implementation of HMM strategy with AL in Tanzania. However, the risk of recurrent infections after AL treatment was high, why integration of ACT with preventive interventions is critical for improved malaria control. The The authors declare that they have no competing interests.BEN, ZP and AM conceived and designed the study and contributed to implementation of field study. BEN and ZP supervised the field work. MM, AMC, PEF and JPG performed molecular analyses, DB and YB performed drug level analysis. BEN and MGP analysed data. BEN and AM wrote the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium parasite transmission to mosquito vectors. Here, we report a mechanistic, within-host mathematical model that uses pharmacokinetic (PK) and pharmacodynamic (PD) data to simulate the effects of artemisinin-based combination therapies (ACTs) on Plasmodium falciparum transmission. To contextualize this model, we created a set of global maps of the fold reductions that would be necessary to reduce the malaria RC (i.e. its basic reproductive number under control) to below 1 and thus interrupt transmission. This modeling was applied to low-transmission settings, defined as having a R0<10 based on 2010 data. Our modeling predicts that treating 93\u201398% of symptomatic infections with an ACT within five days of fever onset would interrupt malaria transmission for \u223c91% of the at-risk population of Southeast Asia and \u223c74% of the global at-risk population, and lead these populations towards malaria elimination. This level of treatment coverage corresponds to an estimated 81\u201385% of all infected individuals in these settings. At this coverage level with ACTs, the addition of the gametocytocidal agent primaquine affords no major gains in transmission reduction. Indeed, we estimate that it would require switching \u223c180 people from ACTs to ACTs plus primaquine to achieve the same transmission reduction as switching a single individual from untreated to treated with ACTs. Our model thus predicts that the addition of gametocytocidal drugs to treatment regimens provides very small population-wide benefits and that the focus of control efforts in Southeast Asia should be on increasing prompt ACT coverage. Prospects for elimination in much of Sub-Saharan Africa appear far less favorable currently, due to high rates of infection and less frequent and less rapid treatment.Achieving a theoretical foundation for malaria elimination will require a detailed understanding of the quantitative relationships between patient treatment-seeking behavior, treatment coverage, and the effects of curative therapies that also block  We utilize a within-host mathematical model of malaria transmission to predict the effects of antimalarial treatment across the globe. We predict that areas containing 91% of the at-risk population of Southeast Asia can achieve elimination if at least 93\u201398% of symptomatic individuals are promptly treated with effective artemisinin-based combination therapies (ACTs), based on assessments of treatment and transmission levels as of 2010. The benefit of attaining this level of coverage far outperforms that of adding additional gametocyte-specific transmission-blocking drugs to current ACTs. We advocate for elimination programs in Southeast Asia to focus on maximizing ACT coverage. Plasmodium falciparum, the most virulent of the Plasmodium species that cause malaria in humans, is responsible for hundreds of millions of cases per year P. falciparum also presents unique challenges P. falciparum stages differ markedly in their levels of metabolic activity, within-host locations, and susceptibilities to antimalarials. Mathematical modeling can help guide elimination efforts by providing quantitative predictions to assess the feasibility of different intervention and control strategies Public health and malaria infection experts are increasingly promoting the goal of malaria elimination in areas of low transmission ACTs and other antimalarial drugs reduce transmission in three ways: by killing the disease-causing asexual blood stages and thus preventing continued production of the intra-erythrocytic sexual gametocyte forms; by killing existing gametocytes and reducing or preventing onward transmission to the mosquito (thereby reducing parasite oocyst numbers in mosquito midguts); and by post-treatment drug prophylaxis wherein residual drug levels can protect against new infections P. falciparum infection and transmission, and overlay these findings onto geospatial maps of malaria endemicity in order to predict the benefits of extended coverage of infected individuals and incorporation of transmission-blocking agents into current ACT regimens. In low-transmission settings we predict that if at least 93\u201398% of all symptomatic infections, corresponding to an estimated 81\u201385% of all infected individuals, were treated within five days of first fever, then the RC could be reduced to below one and malaria would progress towards elimination in regions harboring over \u223c91% of at-risk populations in Southeast Asia and \u223c74% of the global at-risk population. Our findings suggest that increasing treatment coverage with ACTs would be more effective than adding additional transmission-blocking agents in driving towards the goal of malaria elimination in Southeast Asia.Here, we report outputs from our within-host model of P. falciparum infection P. falciparum to induce a fever and clear the syphilis infection We used our recently developed within-host model of the progression of 10 parasitized red blood cells (PRBC) per \u00b5L, while P. falciparum; treated individuals are denoted by \u2018Treated\u2019 or \u2018T\u2019. As with the untreated cases, 10 PRBC/\u00b5L, while in vitro and field data to parameterize the various components of drug activity and predict the effects of various antimalarial therapies in real-world settings.As described in detail below, we then incorporated the effects of drug treatment into our within-host model to illustrate how this modeled treatment affects malaria transmission. Text S1. Notably, our study assumed that individuals were fully compliant with treatment and that parasites were sensitive to the various drug combinations. In future work we hope to examine how differences in patient compliance and parasite drug susceptibility impact transmission.To model the effects of different drugs on asexual parasite densities, we first modeled the within-host concentrations of the partner drugs of two ACTs, artesunate+mefloquine (AM) and artemether+lumefantrine (AL). AM is a frequently used first-line therapy in parts of Southeast Asia   illustrates the results of our PK simulations for LMF, MFQ, and CQ, respectively. The black lines indicate the median (LMF) or mean  of the population concentrations, while the blue lines indicate simulated individual concentration profiles. Model outputs revealed wide variations in concentrations within a population, due to differences in rates of drug uptake and clearance. The number of concentrations depicted in each panel corresponds to the number of patients in each of the studies that provided data for model fitting. Of note, LMF plasma concentrations achieved considerably higher levels than the other two agents.in vitro and field data (see the SI). The asexual activities of drugs were quantified as 48-hour parasite reduction ratios (PRRs), i.e., the fold decreases in parasite numbers every 48 hr, corresponding to one cycle of intra-erythrocytic development and reinvasion.  illustrates the drug concentrations translated into these PRRs over time. To calculate the asexual parasite densities in our model under the effects of drugs for a generic day t, we took the densities from day t-1, applied our within-host model to calculate densities on day t including both the effects of parasite growth and host immune responses, then multiplied densities by the square root of the mean 48-hour PRR for drug concentrations during day t. The resulting density was then used to calculate densities at t+1, and so on until the end of the simulation.To simulate the effects of these drug concentrations against asexual parasites, we first calculated hourly plasma and/or blood concentration levels from our PK modeling. We then translated the hourly drug concentrations into asexual activities, assuming that the dose-response relationships could be modeled as Hill functions. These Hill functions were parameterized from SI.Regardless of the drug regimens simulated here, asexual parasite densities fell rapidly when an individual was treated. The PRRs nevertheless showed substantial differences between drugs in later time periods following treatment. For example, the LMF PPRs rapidly declined within 5\u201315 days of treatment, whereas CQ took longer to decline while showing more heterogeneity. MFQ was also heterogeneous but always showed lower PPRs even at peak plasma concentrations. The maximal PRRs, and the differential rates of absorption, clearance, and volumes of drug distribution are described more fully in the While effective drug treatment rapidly clears asexual blood stage parasites, even successful regimens differ markedly in their effects on gametocytes. As gametocytes develop, they become metabolically more inert, thus reducing the number of drug targets available. Specifically, gametocytes mature over the course of \u223c10\u201315 days through Stages I\u2013V, with each stage differing in metabolic activity and drug susceptibility in vitro studies that have measured the stage-specific effects of drugs on gametocytes To parameterize the modeled effects of drugs on gametocytes, we first conducted a literature review to examine the types of datasets that could inform the model. There are few Figure S1 illustrates the prevalence of gametocytes from field studies of patients treated with a variety of antimalarials Figure S1 was likely due to a variety of factors, including the levels of acquired immunity in the population, exact timing of treatment, age of treated individuals, differences in parasite biology, and drug treatment regimens.These data were used to calibrate the gametocyte component of our drug effects model. To generate a set of model outputs to compare against field data, we simulated treating individuals with a three-day course of AM and varied the assumed killing properties of the two component drugs. We assumed that treatment started relatively early in the infection, i.e., 5 days after first fever, in agreement with field studies from Thailand and Indonesia Figures S1, S2): the modeled schizonticidal treatment data were compared to field trials with SP; mild to moderate gametocytocidal outputs were compared to data from field trials of CQ, CQ+SP or AQ+SP; strong gametocytocidal outputs were compared to ACT clinical trials data, and the modeled triple-combination data were compared to field data for ACTs+PQ. For each drug activity type we then chose the sets of simulations that most closely resembled the mean, maximum, and minimum of observed responses to represent the effects of each class of drugs against gametocytes. We also included some intermediate sets of simulations for the sake of comparison. Figure S2 illustrates the model outputs that best approximated the mean and observed variation in the field data; all model means were from 1,000 runs for each parameterization. We modeled the entire range of observed variation in post-treatment gametocytemias to allow for sensitivity and robustness analyses in our results. This \u2018ensemble modeling\u2019 approach has been used previously to model the effects of vaccines on malaria transmission P. falciparum dynamics Once the model outputs had been generated, these were compared to field studies of drug treatments with similar activity , the total mean fold-reductions for ACTs were 45.3 and 8.1, respectively. For ACT+PQ, total mean effect sizes were predicted to be 92.2 (JE) and 8.9 (CG), respectively. The reason for the large differences between these two transformations is how they incorporate pre-treatment infectivity. Thus, pretreatment infectivity plays a major role in the total effect size. For JE, gametocytes were assumed to be non-infectious early in the course of an infection, thus pretreatment infectivity was almost nonexistent and the effect size was determined by post-treatment infectivity. In contrast, the CG model assumed that gametocytes were infectious upon emergence, thus pretreatment infectivity was relatively large compared to post-treatment infectivity.Purely schizonticidal treatments  were predicted to reduce post-treatment transmission 6.2 to 5.7 fold . Including pretreatment infectivity, the mean effect size of a pure schizonticide was 6.2 and 3.9 . For CQ, which is moderately gametocytocidal against stage I\u2013II gametocytes, total effect sizes were estimated to be 15.6 and 6.0 for the JE and CG parameterizations, respectively. These findings highlight the substantial benefit of drugs with more potent gametocytocidal activity in reducing transmission.Text S1 and Figure S3). For our ACT model infectivity, at day 7 the feeding studies indicated an infectiousness of approximately 2\u20133.5%, whereas the model predicted 5\u20138% under the JE parameterization . This small difference between model and field studies nearly disappeared by day 14 (possibly because of a dose-response effect of LMF on mosquito stage development). To incorporate oocidal activity, we thus assumed that ACTs (AL or AM) reduced onward infectivity by 50% compared to the mean values in The fold reductions in When examining the effects of ACTs plus PQ, our adjustment for the oocidal effects of PQ was different than that for LMF or MFQ, because PQ is active against mosquito stages for only a few days after treatment, but reduces infectivity almost completely during its period of activity At 100% coverage , the effect size of ACTs is 87.3 , if R0<1, the disease will disappear within this region (unless there is significant importation).To contextualize the fold-reductions in transmission theoretically achievable with various treatments, we developed a set of maps of the fold-reductions in malaria transmission necessary to achieve elimination in low-transmission settings. These maps were derived from the worldwide maps of the basic reproductive number of malaria, R0 values described in 2).In brief, the R0. One such malaria invariant is the net infectivity of infected humans to mosquitoes, assuming no acquired immunity developed over the course of repeated infections These worldwide maps of predicted parasite rates varied in intensity from pixel to pixel, given different magnitudes of various malaria covariates. By utilizing empirical and theoretical relationships, the maps were combined with aspects of malaria that remain constant over time and space to calculate R0R0 in RC to below 1. Our maps of the transmission reductions include estimates of uncertainty inherited from the RC posterior densities at each pixel. RC fall within the given regions with 75% confidence.To achieve elimination in a given area, the fold-reduction in transmission under control must be greater than or equal to the R0<10, which excludes the more endemic areas of Africa and aligns with the transmission levels prevalent in Southeast Asia. In terms of biting intensity, an R0 of 10 translates to a yearly entomological inoculation rate of approximately 3 infectious bites per person per year, which would result in approximately 1.5 infections every year Because our within-host model simulates the progression of infections for individuals with no prior history of malaria infection, our modeling conclusions are most relevant for areas of low transmission where individuals have accumulated little acquired immunity from prior infections. We thus restricted our analyses to areas with RC<1) assuming a five-fold reduction in transmission. Areas with R0>10 are masked, as these regions have such high transmission that our modeling predictions are less relevant. Fold reductions were organized into 6 bins for clarity. Figures S4 and S5 present maps of the probabilities of interrupting transmission assuming two- or ten-fold reductions in transmission, respectively. These maps are discussed below in the context of the reductions achievable with antimalarial drugs.P. falciparum transmission, using a within-host model of malaria infection R0 values less than 10).This study calculates the effects of antimalarial therapies on From our model outputs, we can generate three major conclusions. First, the infectivity of individuals before treatment plays a crucial role in determining effect size. If treatment is delayed more than only a few days after the onset of fever, and gametocytes are infectious during this period, then the effect sizes achievable even with first-line ACT therapies plus the gametocytocidal agent PQ are limited. Second, if we account for the effects that the partner drugs LMF and MFQ exert upon mosquito stages of the parasite life-cycle, then there is little difference in the benefits of ACTs versus ACTs+PQ in terms of transmission reductions. Both regimens are extremely effective at stopping onward transmission, with many fold greater benefits versus purely schizonticidal treatments that act only upon asexual blood stage parasites. Third, the proportion of individuals receiving treatment has a major impact on reductions in transmission . Higher transmission regions are more difficult to model, given the complex interactions of immunity, superinfection, and control. As can be seen from the map, many areas of Africa have such intense transmission that R0 exceeds 10, and we cannot say how transmission might be affected by the use of drugs in such areas.We can put these fold-reductions in context using our maps of transmission reductions necessary for elimination. However, examining the map, one can visualize many regions of Africa, including the Sahel, most of East Africa, and parts of Southern Africa, where elimination would appear possible with a five-fold reduction in transmission. Further, much of India and Southeast Asia have low enough transmission that elimination would be possible at this level of control. Prospects for elimination in Myanmar and southern Thailand, however, do not appear to be favorable. Given these maps and quantifications of populations at risk, we can apply our modeling results to determine the percentage of the population that needs to be treated promptly with antimalarials to interrupt transmission in various areas. Combining the maps and the within-host modeling results based on our JE parameterization, we thus estimate that promptly treating \u223c81% of the total infected population with ACTs and/or ACTs+PQ would interrupt transmission in areas covering 91% of the population in Southeast Asia. These coverage rates are for the infected population as a whole, regardless of whether individuals are symptomatic or not. In a study conducted in a region of western Thailand with low and seasonal transmission, most infections (87%) were found to be symptomatic Figures S4 and S5 illustrate the probabilities assuming control interventions with two-fold and ten-fold reductions, respectively. At the two-fold level, much of central Thailand can interrupt transmission, but there are significant portions of Myanmar, Cambodia, Southern Laos, and Southern Thailand where elimination is not likely. At the ten-fold level, there are small pockets in Southern Thailand as well as large areas in Myanmar where interruption is still not likely.If we take the former proportion (87%) as the percentage of the infected population that is symptomatic, then 93% of the symptomatic population would need to be treated with ACTs and/or ACTs+PQ to achieve interruption in the areas of These estimates may be somewhat optimistic because we are using only the JE density-to-infectivity relationship when calculating the infectivity of treated individuals. As we are focusing on low transmission areas, this assumption seems reasonable . However, if we take the mean of the JE and CG relationships for treated individuals, then we predict that it would require treating 85.5% of the total population, or 98.3% of symptomatic patients with ACTs to achieve a five-fold reduction . Further, we find that it is not possible to achieve a six-fold reduction only treating symptomatic individuals with either the ACTs or the ACTs+PQ modeled here, assuming the mean of the JE and CG relationships for treated individuals. Thus, the assumed density-to-infectivity relationship has a large effect on the calculated effectiveness of control programs.2), incorporating uncertainty analysis. These average reductions needed to interrupt malaria transmission are not at the per-village or per-household level. Hotspots of transmission will need to be identified and treated in order to achieve elimination in a given region. The uncertainty for each pixel takes into account this heterogeneity to some degree, but nevertheless caution is advised before using these maps for local-scale planning. We would suggest our maps be used to guide elimination planning at a regional or national level; for elimination planning at a district or city level more intensive surveillance will likely be needed.We note that our maps predict the levels of control necessary to interrupt transmission at the per-pixel level (5 kmRC of the region drops to below 1). Once an area has eliminated malaria, the costs of maintaining elimination may be less than those needed to achieve elimination in the first place, though more research is needed on strategies to maintain elimination in previously endemic areas If we consider the timelines to elimination, the narrower the margin by which the effect size exceeds the threshold for elimination, the longer elimination will take, as population-wide transmission will decay more slowly In areas where antimalarials are predicted to be insufficient to achieve elimination, other interventions may be included in control efforts to increase the effect size of the combined control effort. The combined effect size is simply the product of both component interventions. For example, if the coverage level with antimalarials reduces transmission by three-fold and distribution of bed nets reduces transmission another three-fold, the combined effects are a nine-fold reduction in transmission (as long as there are no antagonistic interactions between the two efforts). Thus, high fold-reductions can be achieved by bundling interventions. While we do not compute the effect sizes of other interventions here, the results in this paper can be combined with other modeling efforts for the purposes of an integrated elimination effort.P. falciparum asexual, sexual, and mosquito stages might suffice to interrupt transmission throughout most of Southeast Asia, especially if complimented by insecticide-treated bed net distribution to reduce population infectivity. We also note that the addition of a single dose of a purely gametocytocidal drug such as PQ to ACTs can reduce onward transmission slightly. However, the focus of control efforts should be on maintaining a high level of treatment coverage. Based on our modeling, PQ and similarly gametocytocidal therapies added to ACTs do not appear to be a magic bullet ensuring elimination and add only nominally to the transmission reductions achievable with ACTs that act against the various parasite stages at feasible levels of coverage.Given these conclusions, serious efforts to eliminate malaria will require extensive planning and sustained support RC in Africa to levels that might make elimination an achievable goal.Efforts are ongoing to utilize our model to predict the effects of possible emerging artemisinin resistance, which threatens existing ACT control strategies P. falciparum to induce a fever in order to clear the syphilis Dataset S1 (see SI). We note that our modeling uses discrete-time difference equations rather than a continuous time model, to calculate both asexual densities and gametocyte densities over time. We chose the former, as the calculation of gametocyte densities from asexual densities is difficult with a continuous-time model because gametocyte densities are a function of weighted cumulative asexual densities and are highly stochastic. For a thorough description, we refer to Our recently reported within-host mathematical model RC to below 1 in low-transmission settings.We also note that an insightful report by Kay and Hastings SI.PK modeling was used to simulate the concentrations of antimalarial drugs after uptake. The concentrations of CQ and its active metabolite, monodesethyl-chloroquine (mdCQ), were simulated using a non-compartmental model parameterized with data from Papua New Guinean children a is set to 0, b is set to 1, c is what we term the EC50, d is the Hill slope, and x is the plasma concentration of the drug. The Hill function dose-response curve type was chosen because this function type was utilized by both of the references that provided in vivo EC50 values for MFQ in vitro studies using CQ and LMF used Hill dose-response relationships to model the effects of drugs against asexual blood stage parasites. Because each drug has a characteristic maximum inhibitory effect, this dose-response function was scaled by the maximum parasite reduction ratio (PRR) for each drug. To determine the effect on asexual parasites of a drug concentration on a given day t of modeling, the asexual parasite densities from day t-1 were used as inputs into the within-host model. The predicted densities on day t were then calculated, incorporating the effects of host immunity and parasite growth. The mean of t was then subtracted from the within-host simulations after appropriate log transformation to calculate the end of day asexual parasite densities, incorporating the effects of host immune responses, parasite growth, and drug concentrations. These densities were then used to calculate the asexual parasite densities on day t+1, and so on until the end of the simulation time. A full description of the PD modeling against asexual blood stages is provided in the SI.The dose-response effect of antimalarials against asexual parasites was assumed to follow the commonly used \u2018Hill function\u2019 in vitro IC50 against asexual blood stage parasites scaled by a factor of five SI). To determine the stage-specific gametocytocidal effects of drugs, the within-host malaria model was first run assuming treatment with a purely schizonticidal combination therapy. The post-treatment gametocyte clearance curves from these simulations were then compared to clearance curves from field studies using SP SI.The dose-response effect of antimalarials against gametocytes was assumed to follow a binary model, where antimalarials act against gametocytes only if drug concentrations are above a certain drug-specific threshold. This binary model was adopted because of the current paucity of dose-response data against gametocytes, as compared to asexual blood stage parasites where the many data sets permit the use of Hill slopes to define dose-response relationships. The threshold for gametocyte activity was chosen to be the Once the ensembles of gametocyte densities after treatment had been generated for various drug combinations, we used two different gametocyte density-to-infectivity relationships (\u2018Jeffery-Eyles\u2019 and \u2018Carter & Graves\u2019) to translate the daily gametocyte densities into predicted human-to-mosquito infectivities To quantify the uncertainty associated with our predictions, we utilized an ensemble modeling approach Dataset S1Text of source code for model .(RTF)Click here for additional data file.Figure S1Post-treatment gametocyte prevalences from field studies. The graphs show gametocyte prevalences of field populations after antimalarial treatment. Gametocyte positivity was assessed using microscopy (threshold \u223c5\u201310 gametocytes per \u00b5L blood). The notation (\u2018\u2212 Day 0\u2019) indicates that only individuals who were gametocyte negative at admission were included in the study. Field notes include the location of study and subset of population treated. All study curves represent mean population values linearly interpolated from measured prevalences. (A) The percentage of individuals positive for gametocytes after treatment with sulfadoxine-pyrimethamine (SP) B) Gametocyte prevalences after treatment with chloroquine (CQ) or amodiaquine (AQ) (sometimes in combination with SP) C) Gametocyte prevalences after treatment with various artemisinin-based combination therapies (ACTs): , , ,  D) Gametocyte prevalences after treatment with ACTs plus primaquine (PQ) (PDF)Click here for additional data file.Figure S2Comparison of modeled post-treatment gametocyte prevalences to field study data. The post-treatment gametocyte prevalences from Figure S1 were averaged to create a set of target data to parameterize the modeled effects of antimalarials on transmission. (A) The mean of the field data after treatment with sulfadoxine-pyrimethamine (SP) B) Gametocyte prevalences after treatment with chloroquine (CQ) or amodiaquine (AQ) (sometimes in combination with SP) C) Gametocyte prevalences after treatment with various artemisinin-based combination therapies (ACTs) D) The same data as in (C) are illustrated with the exception that two ACT field studies were removed from the field data curves because of potentially confounding drug resistance effects  Gametocyte prevalences after treatment with ACTs plus primaquine (PQ) s CQ+AS, . (E) Gam(PDF)Click here for additional data file.Figure S3Infectivity to mosquitoes after treatment. These graphs illustrate the probability that a human will infect a mosquito following antimalarial treatment. Infectivity is defined as the probability that a mosquito bite will produce oocysts. Field study data are indicated with markers; model outputs are indicated by curves. All model outputs represent the mean of 1,000 runs; treatment was assumed to begin 5 days after first fever. Field markers represent the mean from a set of mosquito feedings. Model output curve coloring is taken from Figure S1. Two different gametocyte density-to-infectivity relationships were used to model infectivity: Jeffery-Eyles  and Carter & Graves (CG) A) Field data post-treatment with SP (sulfadoxine-pyrimethamine) or SP plus amodiaquine (AQ) B) Field-measured infectivity after treatment with CQ, SP, or CQ+SP C) Field-measured infectivity D) Field-measured infectivity E) Field-measured infectivity after treatment with ACTs F) Field-measured infectivity after treatment with ACTs, excluding individuals positive for gametocytes (PDF)Click here for additional data file.Figure S4Maps of the predicted probabilities that a two-fold effect size will interrupt malaria transmission. The upper map shows the predicted probabilities that a two-fold reduction in transmission (\u2018two-fold effect size\u2019) would interrupt malaria transmission over a given pixel. Map pixel size is 5 km2. In order to interrupt malaria transmission in a given area, the basic reproductive number for malaria under control (RC) needs to be reduced below 1. Probabilities for each pixel are calculated according to Bayesian posterior estimates of uncertainty (45). Probabilities have been binned into six categories for clarity. Areas with high transmission (R0>10) are masked because our model results are applicable to regions of relatively lower transmission. Note that local conditions (within a given pixel) may be more or less favorable to transmission than the per-pixel averages shown here, and so these maps are most applicable for regional or country-level planning, rather than local-level control efforts. Microenvironments or \u2018hotspots\u2019 might require additional interventions and/or greater treatment coverage than the pixel average R0>10) and mapping assumptions as for the upper map. Areas that appear to be uniform may have small-scale heterogeneities in transmission that are beyond the scale of this map.(PDF)Click here for additional data file.Figure S5Maps of the predicted probabilities that a ten-fold effect size will interrupt malaria transmission. The upper and lower maps are illustrated as per Figure S4, except that Figure S5 shows the predicted probabilities that a ten-fold reduction in transmission (\u2018ten-fold effect size\u2019) would interrupt malaria transmission over a given pixel (size is 5 km2).(PDF)Click here for additional data file.Text S1Pharmacokinetic and pharmacodynamic equations and distributions .(PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum, including chloroquine-resistant strains. Studies in various animal models have shown pyronaridine to be effective against strains resistant to other anti-malarials, including chloroquine. Resistance to pyronaridine appears to emerge slowly and is further retarded when pyronaridine is used in combination with other anti-malarials, in particular, artesunate. Pyronaridine toxicity is generally less than that of chloroquine, though evidence of embryotoxicity in rodents suggests use with caution in pregnancy. Clinical pharmacokinetic data for pyronaridine indicates an elimination T1/2 of 13.2 and 9.6\u2009days, respectively, in adults and children with acute uncomplicated falciparum and vivax malaria in artemisinin-combination therapy. Clinical data for mono or combined pyronaridine therapy show excellent anti-malarial effects against P. falciparum and studies of combination therapy also show promise against Plasmodium vivax. Pyronaridine has been developed as a fixed dose combination therapy, in a 3:1 ratio, with artesunate for the treatment of acute uncomplicated P. falciparum malaria and blood stage P. vivax malaria with the name of Pyramax\u00ae and has received Positive Opinion by European Medicines Agency under the Article 58 procedure.Pyronaridine was synthesized in 1970 at the Institute of Chinese Parasitic Disease and has been used in China for over 30\u2009years for the treatment of malaria. Pyronaridine has high potency against  T,4. T3,4]versus the base (Rm 0.773). Pyronaridine has been found to be highly lipophilic at pH 7.4 (logD 0.34); lipophilicity was reduced at pH 5  nM) then trophozoites (14.0 [13.4\u221214.7] nM)  nM) . Pyronarectively .ex vivo study of serum from Saimiri monkeys given 30\u2009mg/kg pyronaridine gave an IC50 of 7 \u00b1 5\u2009ng/ml and an IC90 of 11 \u00b1 9\u2009ng/ml against the multi-drug resistant K1 P. falciparum strain  pyronaridine by P. falciparum-infected erythrocytes, uptake was similar for both chloroquine-sensitive and chloroquine-resistant strains (T9.96 and K1), being five times less for the comparator chloroquine  tail current with an ICry study ,71.No effects were observed on the central nervous system or on pain threshold at doses up to 1,000\u2009mg/kg. A significant analgesia was noted in the acetic acid writhing test at doses of \u2265300\u2009mg/kg. No effects were noted using the hot-plate method. Decreases in body temperature were noted in the mouse (\u2265300\u2009mg/kg), but not in the dog (doses up to 60\u2009mg/kg). A transient increase in respiratory rate was observed at a dose of 500\u2009mg/kg 2\u2009h post-dose. No effect was noted on gut motility, however small, but significant decreases, in gastric acidity and volume of secretion were observed at doses of 100\u2009mg/kg and above in the rat. Significant decreases in urine volume accompanied by increases in density and electrolyte (sodium) concentration were observed with 500\u2009mg/kg pyronaridine .The pharmacokinetics of pyronaridine have been examined in the rat, rabbit, dog and rhesus monkey. In the rat and dog following intravenous administration of pyronaridine, the blood concentrations of pyronaridine declined in a multi-exponential manner with an apparent terminal half-life of 2 to 4\u2009days. Intramuscular pyronaridine in the rabbit and rhesus monkey reached Tmax within 0.75-1.5\u2009h post-dose with an apparent half-life of 2 to 3\u2009days ,93.in vitro and in vivo models were examined. In brief in vitro metabolism showed evidence of 8\u201312 metabolites. Subsequent profiling showed that all human in citro metabolites were present in rat and dog systems. Following oral administration of pyronaridine no significant metabolism was observed with the major dose related compenent found to be unchanged pyronaridine, although in the rat after IP administration metabolites were observed in urine and faeces. Incubations with recombinant human CYP450 isoforms indicated that pyronaridine could be metabolised by CYP1A2, CYP2D6 and CYP3A4  methods were developed using the anti-malarials amodiaquine or quinine as internal standards ,95. Mostnd blood ,96. Bothnd blood ,96.1/2 56.0 \u00b1 7.0\u2009h, 56.0 \u00b1 7.0\u2009h; Cmax 768 \u00b1 17.0\u2009ng/ml, 1,514 \u00b1 376\u2009ng/ml; and Tmax 1.38 \u00b1 0.22\u2009h, 1.62 \u00b1 0.37. Using the same model as for intragastric administration, intramuscular pyronaridine (6\u2009mg/kg) pharmacokinetics showed complete and rapid absorption with a ka of 33.54 \u00b1 21.81\u2009h-1 and a Tmax of 0.75 \u00b1 0.44\u2009h. Intragastric pyronaridine was 34.6% bioavailable compared with IM administration, with a ka of 2.4 \u00b1 1.26\u2009h-1 pyronaridine in rat, dog and human liver microsomes showed all human in vitro metabolites to be present in both rat and dog systems, however no structural identification was attempted because of the low metabolic turnover  pyronaridine to rat and dog, profiling of plasma, urine, faecal and liver extracts indicated a single major dose-related component, which was confirmed to be unchanged pyronaridine. 14\u2009C] pyronaridine was the only component in any sample that represented >5% of the dose administered, thus indicating that pyronaridine undergoes no significant metabolism in rat and dog  pyronaridine tetraphosphate at 60\u2009mg/kg, produced an overall recovery of 93% over the 14\u2009day collection period. A mean of 83% of the dose was excreted in faeces, with 2.6% excreted in urine, 6.3% recovered from the carcass at 14\u2009days and the remainder in cage washings [14\u2009C] pyronaridine tetraphosphate at 9\u2009mg/kg, produced an overall recovery of 44% over the 14\u2009day collection period. A mean of 36% of the dose was excreted in faeces, with 5.5% excreted in urine. The shortfall in recovery was investigated and a significant proportion of the dose was found to remain associated with the liver. The liver removed from a single animal at 6\u2009months post-dose contained 14.3% of administered radioactivity. Other tissues contained considerably less, with the kidney being the next highest with 0.3% of administered radioactivity at 6\u2009months post-dose [Excretion balance investigations in the rat, following a single oral dose of washings . Excretiost-dose . Furtherost-dose . Profiliost-dose .(0\u221212)) was 662.9\u2009ng.h/ml [1/2 was 241\u2009h, the AUC(0\u2212\u221e) 51,700\u2009ng.h.ml, clearance (CLT) 1.90\u2009ml/min/kg and volume of distribution 41.2\u2009L/kg [0\u2212\u221e 29,400 \u00b1 13,100\u2009ng.h/mL. Pyronaridine after repeat dosing was eliminated from plasma with a mean half-life of 194.8 \u00b1 47.8\u2009h [Some data are available regarding the clinical pharmacokinetics of pyronaridine derived from HPLC plasma assays in volunteers and patients ,95,101. P. falciparum or P. vivax malaria using a spectrofluorometric assay. [1/2\u03b1 was 1.0 \u00b1 0.3\u2009h and T1/2\u03b2 was 63 \u00b1 5\u2009h. Distribution in tissues was extensive; Vc was 11 \u00b1 11\u2009L/kg, Vd(SS) 72 \u00b1 33\u2009L/kg and CLT 0.9 \u00b1 0.35\u2009L/kg.h. The relative bioavailability of two oral formulations (600\u2009mg) was 19 \u00b1 7% for enteric-coated tablets (n\u2009=\u20093) and 32 \u00b1 7% for capsules (n\u2009=\u20093) [1/2\u03b2 65 \u00b1 6, 63 \u00b1 6\u2009h; and CLT 0.796 \u00b1 0.002, 0.71 \u00b1 0.14\u2009kg/L.h [1/2\u03b2 was underestimated in this study.As pyronaridine concentrates in erythrocytes ,32,38,93c assay. . Intramu (n\u2009=\u20093) . Pyronar4\u2009kg/L.h . Since b-1, respectively. The corresponding inter-individual variability estimates for CL/F, V2/F, V3/F, Q/F and Ka were 53.6%, 103%, 29%, 28.8% and 67.5%, respectively. The elimination half-lives of pyronaridine in healthy adult subjects, adult malaria subjects were estimated to be 11.3 and 13.2\u2009days, respectively.The population pharmacokinetics of pyronaridine in healthy and malaria infected subjects with uncomplicated falciparum and vivax malaria after the administration of oral pyronaridine/artesunate (3:1 ratio) have been published as an abstract . Pyronarmax and AUC rose from 85.7\u2009ng/ml to 338.5\u2009ng/ml and 17,623 to 35,360\u2009ng/ml*hr, respectively. Tmax was between 2.4 and 3.2\u2009h and the elimination half-life ranged between 6.6 and 9.0\u2009days. Based upon population pharmacokinetic modeling, the elimination half-life of pyronaridine in pediatric malaria subjects was estimated to be 9.6\u2009days (103). Artesunate is rapidly converted to dihydroartemisinin in vivo. Mean Cmax and AUCINF levels for artesunate increased dose dependently from 92.8\u2009ng/ml to 287.0\u2009ng/ml and 104.3 to 232.3\u2009ng/ml*hr, respectively. Tmax was within a range of 0.5 to 1.0\u2009h and t1/2 was between 0.5 to 1.2\u2009h. Dihydroartemisinin showed a linear increase in Cmax from 479.1 to 1,185.9\u2009ng/ml and AUC values from 1,054.5 to 2,961.0\u2009ng/ml*hr. The time to maximal drug concentrations was between 1.3 and 1.7\u2009h after drug administration. Dihydroartemisinin showed a relatively short half-life of 0.9 to 1.2\u2009h in the respective treatment groups. Pharmacokinetic parameters of the granule paediatric co-formulation were compared with the respective tablet formulation of the same dose strength (3\u2009mg/kg artesunate and 9\u2009mg/kg pyronaridine). There were no statistically significant differences in pharmacokinetic parameters except for a higher Cmax of pyronaridine in the patient group receiving the paediatric drug formulation .Pharmacokinetic characteristics of pyronaridine, artesunate, and dihydroartemisinin were evaluated in a clinical study with tablet  and granule (9\u2009+\u20093\u2009mg/kg) formulations in Gabonese children aged two to 14\u2009years . PyronarP. falciparum; no cases of recrudescence were seen in 10 patients after 30\u2009days of follow up [Pyronaridine was approved for human use in China in 1980 at a total oral dose of 1,200\u2009mg (or 24\u2009mg/kg) divided into two doses on Day 0 and one dose on each of the following one or two days. Intramuscular or IV pyronaridine was given as 300\u2009mg (or 6\u2009mg/kg) divided into two doses eight hours apart. The main clinical studies of oral, IM and IV pyronaridine monotherapy conducted in China have been reviewed in English by Fu and Xiao and Shao and are summarized in Additional file ollow up ,106. Plaollow up . AnecdotOutside China, two clinical trials of oral pyronaridine for the treatment of falciparum malaria have been reported in adults: one randomized trial in Cameroon in comparison with chloroquine and one in Thailand comparing two doses of pyronaridine . All patients receiving pyronaridine had a negative parasite count on or before Day 3 or positive on Day 3, but <25% of pre-treatment density plus negative there after until Day 14 compared with 18/41 (43.9%) of chloroquine-treated patients. Four cases of chloroquine failure required treatment with an alternative anti-malarial on or before Day 3. There was no significant difference in parasite clearance or fever clearance times between treatment groups for those patients that had a successful clinical and parasitological response  and 73-fold more active than chloroquine against 39 chloroquine-resistant isolates (IC50 354 nM) [The efficacy of pyronaridine monotherapy in uncomplicated falciparum malaria was compared with that of chloroquine in a randomized open-label study conducted in Cameroon  .et al. evaluated the efficacy of two dosing regimens of pyronaridine in a non-randomized, open-label study of 101 adult Thai patients with uncomplicated falciparum malaria . Recrudescence occurred between Days 13 and 28 (median 24\u2009days) in the 1,200\u2009mg group and between Days 11 and 28 (median 17\u2009days) in the 1,800\u2009mg group. Sixteen patients in the 1,200\u2009mg group and 11 in the 1,800\u2009mg group had at least one parasite count greater than their initial count [50 for 10 patients that were cured was 15.69 \u00b1 3.82 nM compared with 22.98 \u00b1 12.1 nM in 10 patients who had recrudescence. The authors suggested that there was a relationship between treatment result and parasite sensitivity to pyronaridine. However, there was no evidence of the development of resistance after unsuccessful treatment with pyronaridine: the initial pyronaridine mean IC50 was 23.39 \u00b1 12.71 nM versus 22.96 \u00b1 11.54 nM after recrudescence in P. falciparum isolates from nine patients for whom paired data were available [Looareesuwan al count . There wal count . The timvailable .P. falciparum, even in the mid-1990s, presented a difficult challenge for new therapy [50s obtained in the two clinical studies, pyronaridine was 3.3-fold more active against Cameroon isolates than against those from Thai patients that were cured and 4.8-fold more active against those from Thai patients who had recrudescence [In contrast to the Cameroon study, recrudescences were seen in Thailand; fever clearance and parasite clearance times were also extended by 47.9\u221250.5\u2009h and 7.6\u22129.9\u2009h, respectively ,107. The therapy . The 88% therapy . In Came therapy . When codescence ,107.P\u2009=\u20090.001) and parasitological response at Day 14 (P\u2009=\u20090.0001). In the chloroquine group, 5/40 (12.5%) patients required alternative anti-malarial therapy on or before Day 3. There was no significant difference in fever or parasite clearance times between the two therapies in those patients that had a successful clinical and parasitological response  for patients treated with pyronaridine and 5.69 nM  for those who received chloroquine. For patients in the pyronaridine group, 23/40 (57.5%) of the isolates tested were chloroquine-resistant (chloroquine IC50 >100 nM). Clinical and parasitological efficacy of pyronaridine was demonstrable against all of these chloroquine-resistant isolates.One study of oral pyronaridine for the treatment of uncomplicated falciparum malaria in children has been conducted in Cameroon  was conducted in Hainan province, China, where chloroquine resistant versus pyronaridine monotherapy . Time to parasite clearance was significantly faster with the combination (23.8 \u00b1 10.1\u2009h) than with pyronaridine alone . Gametocyte carriage was 20.0% for the combination, 16.7% for DHA alone and 60.9% for pyronaridine alone, which was significantly higher than for the combination (P\u2009<\u20090.01) [versus the monotherapy doses.A double-blind study in China investigated the efficacy of pyronaridine  or dihydroartemisinin  alone or in combination  in uncomplicated falciparum malaria  . These eP. falciparum malaria and for the blood stages of P. vivax malaria in adult and paediatric patients. A conference report is available for a double-blind, multicentre, randomized Phase II study including 477 adults in Africa and SE Asia [Pyronaridine-artesunate has been developed as a fixed-dose combination therapy by Shin Poong Pharmaceutical Ltd, South Korea, with the tetraphosphate pyronaridine salt in a 3:1 ratio with artesunate as an oral once daily treatment for three days, for uncomplicated  SE Asia .max and AUC values and a comparable relative bioavailability of the granule co-formulation. At all dose levels efficacy of pyronaridine artesunate combination therapy was 100% in the per protocol analysis on Day 28 after PCR correction [One Phase II study, conducted in Gabon, has been reported . Two fixrrection .P. falciparum malaria versus artemether-lumefantrine in one study and mefloquine\u2009+\u2009artesunate in the other study, one study in adults and children in P. vivax malaria versus chloroquine [P. falciparum malaria with the granule formulation versus artemether-lumefantrine. Overall, in the clinical programme, a total of 2,815 patients were treated with pyronaridine-artesunate, including 1,528 adults (\u2265 18\u2009years), 401 patients 12\u201318\u2009years and 886 children (< 12\u2009years) [A Phase III programme with fixed-dose pyronaridine-artesunate included four phase III pivotal studies; two studies in adults and children in oroquine  and one 2\u2009years) . Pyronar2\u2009years) ,120.P. falciparum malaria in children and adults [P. falciparum isolates and 100% cure rates from the previous pyronaridine-artesunate clinical study [versus lumefantrine-artemether. Kaplan\u2013Meier analysis confirmed an important difference in re-infection rate between treatment groups through Day 28 and 42. Parasite clearance was more rapid with pyronaridine-artesunate versus lumefantrine-artemether (p\u2009<\u20090\u00b7001), with the greatest difference seen before Day 2. There was no difference in fever clearance time between the treatment groups.A pivotal, multicentre, randomized, comparative, parallel group, double-blind, double-dummy study compared the efficacy and safety of once a day pyronaridine-artesunate with twice a day lumefantrine-artemether in uncomplicated d adults . A totalal study . Day-28 P. falciparum malaria in children and adults. Overall, 1,271 patients were included in the study, 81% from Asia, 19% from Africa. Day-28 PCR-corrected ACPR in the per protocol population were 99.2% for pyronaridine-artesunate and 98.1% for mefloquine\u2009+\u2009artesunate. Pyronaridine-artesunate was non-inferior to mefloquine\u2009+\u2009artesunate; treatment difference 1.1% . ACPR were\u2009\u2265\u200995.7% across individual study centers for both study treatments. In 211 patients in the region of Cambodia where extended parasite clearance times are reported, parasite clearance time for both treatments was prolonged compared with other countries ; median time to parasite clearance was twice as long (64 versus 31\u201332 hours) [A further pivotal, multicentre, randomized, comparative, parallel group, open-label study compared pyronaridine-artesunate and a loose combination of mefloquine\u2009+\u2009artesunate in uncomplicated 2 hours) .versus artemether-lumefantrine tablet (20:120\u2009mg) in 535 patients (mean 5.0\u2009years (0 to 12\u2009years); mean 16.6\u2009kg (6\u201324.9\u2009kg)) with acute uncomplicated P. falciparum malaria. Patients from seven sites in Africa and South East Asia were randomized to receive a three-day course of either pyronaridine-artesunate once a day or artemether-lumefantrine twice a day [A phase III comparative, double-blind, double dummy, randomized, non-inferiority, multicentre clinical study was conducted in paediatric patients to assess the efficacy and safety of a granule fixed dose formulation of oral pyronaridine-artesunate (60:20\u2009mg) ce a day . In the ce a day .P. ovale (n\u2009=\u200910) and/or P. malariae infection (n =16) in Cameroon [Pyronaridine efficacy was investigated in 22 patients with Cameroon . Mean feCameroon . No gameCameroon .P. vivax  primaquine ,124. It P. vivax malaria [P. vivax malaria. The primary objective of this clinical study was to compare the efficacy and safety of the fixed combination of pyronaridine-artesunate with that of standard chloroquine therapy in adults and children with acute, uncomplicated P. vivax malaria. This trial included five centres across Cambodia, Thailand, India, and Indonesia. In a double-dummy design, patients (aged >3\u2013\u226460 years) with microscopically confirmed P. vivax mono-infection were randomized (1:1) to receive pyronaridine-artesunate (target dose 7.2:2.4\u2009mg/kg to 13.8:4.6\u2009mg/kg) or chloroquine (standard dose) once daily for three days. For patients who completed the study up to Day 28 and who had normal glucose-6-phosphate dehydrogenase activity, a 14-day course of primaquine (15\u2009mg/day) was administered starting on Day 28, to complete their radical cure. Each treatment group included 228 randomized patients. Outcomes for the primary endpoint, Day-14 cure rate in the per-protocol population, were 99.5%, with pyronaridine-artesunate and 100% with chloroquine. Pyronaridine was non-inferior to chloroquine: treatment difference \u22120.5% , i.e. the lower limit of the two-sided 95% CI for the treatment difference was greater than \u221210%. Pyronaridine-artesunate cure rates were non-inferior to chloroquine for Days 21, 28, 35 and 42. Parasite clearance time was shorter with pyronaridine-artesunate (median 23.0\u2009h) versus chloroquine , as was fever clearance time . Kaplan-Meier estimates of post-baseline P. falciparum infection incidence until Day 42 were 2.5% with pyronaridine-artesunate, 6.1% with chloroquine . Post-baseline P. vivax or P. falciparum infection incidence until Day 42 was 6.8% and 12.4%, respectively  [The fixed dose pyronaridine-artesunate combination tablet has been tested as an oral once daily treatment over three days for blood stage of  malaria . A pivotnk test) .Integrated analysis of efficacy were performed on pooled data from the four Phase III pivotal studies; the Phase III clinical efficacy database included 1,701 patients from Asia and 1,833 patients from Africa .P. falciparum studies, non-inferiority of pyronaridine-artesunate versus the comparator  was demonstrated as primary endpoint for PCR-corrected ACPR on Day 28 (per protocol and ITT populations). The percentage of subjects with gametocytes gradually decreased to zero or near zero over time in the pyronaridine-artesunate, mefloquine\u2009+\u2009artesunate and artemether-lumefantrine treatment groups.In each of the three Phase III P. vivax malaria, non-inferiority of pyronaridine-artesunate compared with chloroquine was demonstrated with respect to the crude cure rate on Day 14 (per protocol population). Non-inferiority was also demonstrated for both PCR-corrected and crude ACPR at all time points (for both PP and ITT populations).In the study in subjects with P. falciparum and P. vivax adults and children malaria patients and similar efficacy was observed in Asia and Africa. Pyronaridine-artesunate showed high cure rates, similar to those of the current standard of care therapies, along with rapid clearance of parasitaemia and most malaria-related symptoms, coupled with prevention of recrudescence. The relatively long half-life of pyronaridine could explain the observed prophylactic effect up to D42 [Pyronaridine-artesunate showed high levels of efficacy in p to D42 .versus 56% for chloroquine [versus an enteric-coated tablet at 12\u2009mg/kg ; 18.8% (6/32) and 28.1% (9/32) patients experienced an adverse event, respectively.Studies in China found pyronaridine to be generally well tolerated; around 38% of patients experienced adverse events oroquine . Adverseoroquine . There wGastrointestinal effects were mostly absent after IM administration, though there was some local injection-site irritation without necrosis. Intravenous administration was mostly without adverse effects, though nausea, palpitation, diarrhoea and abdominal pain were noted in a few patients . At leasP. malariae or P. ovale infection also reported pruritis with pyronaridine in 3/20 (15.0%) patients, and 9/20 (45.0%) had mild gastrointestinal symptoms [There were some differences in the adverse event profile for pyronaridine monotherapy between trials conducted in Cameroon and Thailand . Between Days 0 and 7 there were significant (P\u2009<\u20090.05) decreases in mean neutrophils, total bilirubin, conjugated bilirubin and blood urea and significant increases in mean lymphocytes, eosinophils, reticulocytes and platelets in both the pyronaridine and chloroquine-treated groups [versus baseline [In a study conducted in Cameroon, the most common treatment emergent adverse events with pyronaridine in children (n\u2009=\u200941) were abdominal pain (22.0%), diarrhoea (12.2%) and headache 9.8%)  or DHA  alone or in combination  there were no serious adverse events in any group . Mild heP. falciparum malaria investigated the safety of fixed dose pyronaridine artesunate at three doses: 6\u2009+\u20092 (n\u2009=\u2009160), 9\u2009+\u20093 (n\u2009=\u2009157) or 12\u2009+\u20094\u2009mg/kg (n\u2009=\u2009159) and limited data have been reported in abstract form [A Phase II study in adults with act form .P. falciparum malaria in children and adults were compared [In the Phase III pivotal, multicentre, randomized, comparative, parallel group, double-blind, double-dummy study in a total of 1,272 patients the efficacy and safety of once a day pyronaridine-artesunate with twice a day lumefantrine-artemether in uncomplicated compared . Safety In the pivotal, multi-centre, randomized, comparative, parallel group, open-label study comparing pyronaridine-artesunate and mefloquine\u2009+\u2009artesunate, 1,271 patients were included. Similar proportions of patients experienced at least one adverse event in both treatment groups; 45.9% for pyronaridine-artesunate and 44.9% for mefloquine\u2009+\u2009artesunate . AdverseP. vivax malaria in a pivotal Phase III multi-centre, randomized, double-blind, double-dummy, parallel-group comparative, non-inferiority trial [versus the chloroquine group (0%). All adverse events in the pyronaridine-artesunate group and the majority  in the chloroquine group were of mild-to-moderate severity. Adverse events deemed study drug related by the investigator occurred in 27/228 (11.8%) patients in the pyronaridine-artesunate group and 23 (10.1%) in the chloroquine group. There were no deaths during the study. Two patients, both in the pyronaridine-artesunate group, had serious adverse events ; neither was considered drug related by the investigator. Two patients, both in the chloroquine group, had adverse events leading to drug discontinuation and study withdrawal ; all considered possibly related to drug treatment by the investigator. No clinically meaningful difference in change from baseline in haemoglobin was observed between patients with or without phenotypic G6PD deficiency. Other haematology laboratory findings  were of similar magnitude in both treatment groups. Biochemistry laboratory observations were generally similar in both treatment groups, with the exception of alanine transaminase (ALT) and aspartate transaminase (AST). From Day 3 to the end of the study, 3/228 (1.3%) patients in the pyronaridine-artesunate group had peak ALT\u2009>\u20095x the upper limit of normal (ULN); two (0.9%) with peak ALT\u2009>\u200910\u2009\u00d7\u2009ULN. Total bilirubin values were within normal limits for both of these subjects throughout the study. No patients in the chloroquine group had ALT\u2009>\u20095xULN. Post-treatment AST was\u2009>\u20095\u2009\u00d7\u2009ULN in one patient in the chloroquine group and\u2009>\u200910\u2009\u00d7\u2009ULN in one patient from the pyronaridine-artesunate group; total bilirubin was within normal limits. Values for AST and ALT were close to or within normal limits by Day 28. No patient had peak ALT or AST\u2009>\u20093\u2009\u00d7\u2009ULN plus total bilirubin\u2009>\u20092\u2009\u00d7\u2009ULN during the study.The fixed dose pyronaridine-artesunate combination tablet has been tested as an oral once daily treatment for three days for blood stage of ty trial . In the One study, conducted in Gabon, included children aged two to 14\u2009years old with the tablet formulation of fixed-dose pyronaridine-artesunate at three different ratios 6:2, 9:3 and 12:4\u2009mg/kg or a paediatric granule formulation of fixed-dose pyronaridine-artesunate at 9:3\u2009mg/kg; both formulations were given once daily for three days . In thisIn an integrated safety analysis of the pyronaridine-artesunate Phase III studies, 57.2% of pyronaridine-artesunate patients reported at least one adverse event after baseline . PyronarThe hepatic data were reviewed by an independent data monitoring committee (IDMC) that consisted of six members, including three international experts in Drug Induced Liver Injury (DILI). The committee concluded that a three-day treatment with pyronaridine-artesunate can cause transient rises in ALT concentration in a small subset of patients. However, the early onset (Day 3\u20137), dose response trend, and rapid resolution were all consistent with direct low-level toxicity. The IDMC commented that with hepatotoxic drugs, serious idiosyncratic hepatotoxicity typically begins after weeks or months of treatment, like described with amodiaquine, another Mannich base . These fP. falciparum and blood stage P. vivax malaria with a fixed-dose combination tablet and granule formulation for paediatric administration. MMV is a not-for-profit organization, operating as a public\u2212private partnership seeking to discover, develop and deliver new anti-malarial drugs. A complete suite of toxicology and safety pharmacology studies and a full clinical programme has been undertaken. Pivotal Phase III comparative clinical trials were completed in 2009, to include over 3,000 children, adolescents and adults across sub-Saharan Africa and South East Asia in more than 18 countries. Positive Opinion was received by the European Medicines Agency under the Article 58 procedure in March 2012. Shin Poong received approval from the Korean Federal Drug Agency in August 2011 and this will be followed by national regulatory submissions and WHO procedures.Pyronaridine tetraphosphate-artesunate (PYRAMAX\u00ae) is being co-developed by the Medicines for Malaria Venture (MMV) and Shin Poong Pharm Co Ltd (Republic of Korea) for the treatment of acute uncomplicated in vitro. Also, studies in vivo animal models indicate a synergistic effect between pyronaridine and artemisinins against parasites resistant to one or both components, restoring efficacy against these strains. Consequently, pyronaridine represents an ideal candidate for combination therapy with artemisinin derivatives, such as artesunate. Pyronaridine also appears to be well tolerated in clinical studies [P. falciparum and P. vivax malaria in adult, children and infant populations [Pyronaridine has shown value as a therapeutic partner to use in artemisinin combination therapy. Pyronaridine brings high efficacy, including against chloroquine and amodiaquine-resistant strains, and the reassurance of many years of successful use in China as monotherapy and in combination with other anti-malarials, without the development of widespread drug resistance. To date no serious toxicities have been reported for pyronaridine.. Notably, pyronaridine in combination with artemisinins appears to reduce the development of resistance  studies ,111,120.ulations ,121.ACT, Artemisinin-based combination therapy; Bid, Twice daily; ECG, Electrocardiogram; ED50, The amount of material required to produce a specified effect in 50% of a population; IC50 IC90, IC99 dose at which 50, 90 or 99% of parasites are inhibited, Respectively; IG, Intragastric; IM, Intramuscular; IP, Intraperitoneal; IV, Intravenous; LD50, Lethal dose of 50% of test population; MLD, Minimal lethal dose; QD, Once daily; SC, Subcutaneous; ALT, Alanine aminotransferase; AST, Aspartate aminotransferase.; Eq/g Pyronaridine tetraphosphate, 910.03\u2009g.mol-1 ; Pyronaridine, 518.05\u2009g.mol-1 ; Log D, 1.7 at pH 7.4.Dr CS Shin is former Head of Pyramax Development at Shin Poong Pharmaceuticals (retired). Other authors declare that they have no competing interests.SLC, SD, SAB, LF and CSS drafted the manuscript. All authors read and approved the final manuscript.Comparative activity of pyronaridine against chloroquine-susceptible and -resistant P. falciparumfield isolates from SE Asia and Africa.Click here for fileComparative IC50 for drug-resistant and -sensitive strains of P. falciparum.Click here for fileAcute toxicity studies with pyronaridine: summary of main findings.Click here for fileSub-acute toxicity studies with pyronaridine: summary of main findings.Click here for fileEfficacy of pyronaridine monotherapy in patients with P. falciparum malaria: studies conducted in China.Click here for fileOral monotherapy with pyronaridine in the treatment of falciparum malaria: International studies.Click here for fileEfficacy of oral pyronaridine combination therapy in patients with P. falciparum malaria.Click here for fileClinical studies conducted in China of pyronaridine alone and in combination with primaquine in patients with P. vivax malaria.Click here for fileTreatment-emergent adverse events with pyronaridine oral monotherapy in the treatment of falciparum malaria.Click here for fileTreatment-emergent adverse events with fixed-dose pyronaridine-artesunate in the treatment of falciparum malaria in children.Click here for file"}
+{"text": "Malaria remains a major public health problem in developing countries. Then in these countries prompt access to effective antimalarial treatment such as Artemisinin based-Combination Therapies (ACT) proves to be an essential tool for controlling the disease. In Senegal, since 2006 a nationwide scaling up program of ACT is being implemented. In this context it has become relevant to monitor ACT efficacy and provide recommendations for the Senegalese national malaria control program.An open randomized trial was conducted during two malaria transmission seasons (2011 and 2012) to assess the efficacy and safety of three combinations: dihydro-artemisinin-piperaquine (DHAPQ), artemether-lumefantrine (AL) and artesunate-amodiaquine (ASAQ). The primary end point of the study was represented by a PCR adjusted adequate clinical and parasitological response (ACPR) at day 28. Secondary end points included: (i) a ACPR at days 35 and 42, (ii) a parasite and fever clearance time, (iii) ACTs safety and tolerability. The 2003 WHO\u2019s protocol for antimalarial drug evaluation was used to assess each outcome.Overall, 534 patients were randomized selected to receive, either ASAQ (n =\u2009180), AL (n =\u2009178) or DHAPQ (n =\u2009176). The PCR adjusted ACPR at day 28 was 99.41% for the group ASAQ, while that was 100% in the AL and DHAPQ groups (p =\u20090.37). The therapeutic efficacy was evaluated at 99.37% in the ASAQ arm versus 100% in AL and DHAPQ arm at day 35 (p =\u20090.37). At day 42, the ACPR was 99.27% in the ASAQ group versus 100% for both AL and DHAPQ groups, (p =\u20090.36). No serious adverse event was noted during the study period. Also a similar safety profile was noted in the 3 study groups.In the context of scaling up of ACTs in Senegal, ASAQ, AL and DHAPQ are highly effective and safe antimalarial drugs. However, it\u2019s remains important to continue to monitor their efficacy.201305000552290.PACTR  P. falciparum resistance to the monotherapies such as chloroquine, sulfadoxine, pyrimethamine, has been a major obstacle for malaria control in sub-saharan countries. Then to deal with these resistance issues, the WHO recommended Artemisinin based-Combination Therapies (ACT) for the management of uncomplicated malaria cases [Plasmodium falciparum by acting on gametocytes and reducing the chances of development of drug resistance [Despite increasing efforts to control malaria, the disease is still a public health problem. According to the World Health Organization (WHO), there are 216 million new malaria infection cases per year in the world. Also, the disease causes 655 000 deaths per year in the world. More than 81% of the new infection cases and 90% of the deaths occur in Africa mainly in children under five years [sistance -5.In Senegal, the National Malaria Control Program (NMCP) initiated in 2006 a nationwide scaling up program of ACT . SeveralRecent studies demonstrated a decline in ACTs efficacy as well as artesunate monotherapy in the Asian region ,8. This Plasmodium falciparum malaria in Senegal.In Senegal, notification of adverse events has been a great challenge for the National Malaria Control Programme although a pharmacovigilance system for monitoring ACT drug related adverse events has been established for several years . Thus thPlasmodium falciparum is the predominant species and transmission is mainly due to Anopheles gambiae s.l. The enrollment of patients started in 2011 and was completed in 2012.The study was carried out during two malaria transmission seasons in two health centers: (i) Deggo which is located 20\u00a0km form Dakar, the capital city and (ii) Keur Soce located 200\u00a0km of South from Dakar. In the areas around the health posts, malaria is highly seasonal during the rainy season (July to October) with a peak of transmission from September to December. Plasmodium falciparum malaria: Artesunate-Amodiaquine (ASAQ), Artemether-Lumefantrine (AL) and Dihydroartemisinin-Piperaquine (DHAPQ). Randomization was permuted in blocks of 9. The primary endpoint was the PCR adjusted adequate clinical and parasitological response (ACPR) at day 28. Secondary end points included: (i) PCR adjusted ACPR at day-35 and day-42, (ii) the parasite clearance time, (iii) the fever clearance time and (iv) ACT tolerability and safety.The study was designed as an open randomized trial comparing three ACTs for the treatment of uncomplicated The study was conducted as part of a national surveillance program aimed at monitoring ACTs efficacy under routine conditions.Plasmodium falciparum malaria with parasite density ranged from 1000 to 100,000 trophozoites/\u03bcl. Ability to take oral medication and written informed consent were required as part of the inclusion criteria. Patients presenting with mono-infection by another species or mixed infectation, severe vomiting, severe malnutrition, severe signs of malaria , a positive pregnancy test and patients who had a history of allergy to study drugs or did not given informed consent were excluded from the study.Subjects were enrolled if their age was above 6\u00a0months and they presented with uncomplicated After inclusion, all patients were weighed and randomized to receive one of three study drugs for three days. The drugs were administered under the direct supervision of the medical staff. In case of vomiting within the 30\u00a0minutes following the first administration, the same dose was administrated again.Participants who vomited a second time were excluded from the study and received intravenous quinine treatment in accordance with the national malaria control program guidelines (25\u00a0mg/kg/day for seven days 3 times daily). The dosages of the study drugs were as follows:Artesunate-Amodiaquine (ASAQ): the tablet contains 4\u00a0mg/kg/day Artesunate (AS) plus 10\u00a0mg/kg/day Amodiaquine (AQ). The drug was given once a day. The dosage was adjusted according to the weight: one tablet per day containing 25\u00a0mg/67.5\u00a0mg (4.5 - 9\u00a0kg), one tablet per day containing 50\u00a0mg/135\u00a0mg (9 - 18\u00a0kg), one tablet per day containing 100\u00a0mg/270\u00a0mg (18 - 36\u00a0kg) and two tablets per day containing 100\u00a0mg/270\u00a0mg if weight was more than 36\u00a0kg.Artemether-Lumefantrine (AL): the drug was not given with additional fat. The tablet contains 20\u00a0mg of Artemether plus 120\u00a0mg of Lumefantrine. The drug was given 2 times a day. The dosage was adjusted according to the weight: two tablets per day (5 \u2013 14\u00a0kg); four tablets per day (15 \u2013 24\u00a0kg); six tablets per day (25 \u2013 45\u00a0kg) and eight tablets per day if weight was more than 45\u00a0kg.Dihydroartemisinin-Piperaquine (DHAPQ): the tablet contains 40\u00a0mg of Dihydroartemisinin (DHA) plus 320\u00a0mg of Piperaquine (PQ). The drug was given once a day. The dosage was adjusted according to the age: 3 tablets per day if age was more than 16\u00a0years; 2 tablets per day if age was between 11 and 16\u00a0years and 1.5 tablets if the age was between 6 and 11\u00a0years.After inclusion, patients were followed at day 0 (day of inclusion), 1, 2, 3, 7, 14, 21 and 28. A random sub-sample of study participants was followed up to day 35 and day 42 to assess the long term protective effect of each drug after curative doses. A clinical and biological assessment was performed for all patients. The 2003 WHO\u2019s protocol for antimalarial drug efficacy evaluation was used .A clinical examination and an interview were performed before the inclusion. After inclusion and first dose administration, all patients were examined during the first 4\u00a0days. At each follow up visit, a clinical examination and interview to evaluate the patient\u2019s clinical conditions as well as the occurrence of adverse events were done. Patients were seen by the medical team at any time if they did not feel well.A blood sample was collected for thick and thin smears for all study patients. Both tests were used to determine the parasite density and the plasmodium species at the day 0, 1, 2, 3, 7, 14, 21, and 28. Both tests were repeated at day 35, 42 and other days of follow up to evaluate parasite clearance times.To distinguish recrudescence form new infection, blood was collected on filter paper at day 0 and at day of parasite reappearance.The haemoglobin level was determined at day 0 and day 7 using Sysmex XS 1000i automate. Creatinine, urea, bilirubin, aspartate amino-transferase (ASAT) and alanine-aminotransferase (ALAT) were also examined at day 0 and day 7 using automate A15 of Biosystems laboratories.Finger prick blood was used to collect blood samples. Thick and thin smears were stained with Giemsa. The parasite density was evaluated by counting the number of asexual parasites per 200 white blood cells and calculated per \u03bcl: number of parasites \u00d7 8000/200 assuming a white blood cell count of 8000 cells per \u03bcl. Thick and thin smears were negative after 100 field microscopics reading.Haematological and biochemical parameters were performed at enrolment and day 7 to determine the haemoglobin level, the concentration of urea, creatinine, bilirubine, asparta-amino-transferase (AST) and alanine-amino-transferase (ALAT).P falciparum genes (Merozo\u00efte Surface Protein): MSP1 and MSP2[The PCR was used to distinguish recrudescence from new infection in case of treatment failure. Nested PCR was conducted to compare the genetic polymorphism of  and MSP2. RecrudeThis was defined as a development of danger signs or severe malaria on days 1\u20133 in the presence of parasitaemia, a patient with parasitaemia on day 2 higher than the day 0 count irrespective of axillary temperature; parasitaemia on day 3 with axillary temperature \u2265\u200937.5\u00b0C and parasitaemia on day 3 that is \u2265\u200925% of count on day 0.This was defined as a presence of parasitaemia on any day from day 7 to day 28 and axillary temperature <\u200937.5\u00b0C, without previously meeting any of the criteria of early treatment failure or late clinical failure.The ACPR was defined as an absence of parasitaemia on day 28 irrespective of axillary temperature without previously meeting any of the criteria of early treatment failure, late clinical failure or late parasitological failure [Based on an expected therapeutic effect of AL not low than 95%  assumingData collected were entered into Excel software and the analysis was done with Stata IC 12 software. Intention to treat and per protocol analysis were performed.The intention to treat included all randomized subjects who took at last one full dose and had one post-baseline efficacy without major protocol deviation. The per protocol analysis include all subjects who received the three dose and had no major protocol deviation.Data were analysed by estimation of difference in proportion according to a 95% confidence interval. Groups were compared using Chi Square test or Fisher exact test for categorical variables and Student\u2019s t-test for continuous variables when these tests were applicable. Otherwise, non- parametric tests  were used.The cumulative incidence of failure rate was calculated in each group and compared using Kaplan Meier method. Changes in biological parameters from day 0 to day 7 were calculated and compared between treatment arms using Bonferroni test.p value <\u20090.05 two side).Statistical significance for all tests was set at 0.05 . Written informed consent was obtained from each participant or their parent or guardian for particiapants with an age below 18\u00a0years. The study was registered at the Pan African Clinical Trial Registry: registration number: PACTR201305000552290.This study was conducted according to the Declaration of Helsinki and existing national legal and regulatory requirements. The protocol was reviewed and approved by the Senegalese National Ethical Committee , 75.8% (135/178) and 76.14% (134/176) respectively in ASAQ, AL and DHAPQ group.The median parasitemia was 13132.5 trophozoites/\u03bcl in ASAQ arm, 26192.5 trophozoites/\u03bcl in AL arm and 21347.5 trophozoites/\u03bcl in DHAPQ arm   and 77% (n =\u2009411) of all patients seen at day 35 and day 42 respectively. Again very good cure rates were observed with no significant difference detected between the groups , 75.8% (135/178) and 76.14% (134/176) respectively in ASAQ, AL and DHAPQ group. The difference was not significative at inclusion between the groups (p =\u20090.96). After first dose administration, 3.8% (7/180) patients in ASAQ group and 3.4% (6/176) patients in DHAPQ group were found with fever. The proportion of patients with fever was more important in the AL group 8.4% (15/178) with no significant difference (p =\u20090.06). The fever clearance was noted at day 2 after the first dose treatment.-3). At day 2, this proportion was 3.3% (6/180), 6.7% (12/178) and 2.3% (4/176). The difference was not significative at day 2 after first dose administration (p =\u20090.08) between the three treatments groups. Complete parasite clearance was obtained at day 3 after inclusion.The three treatments showed rapid parasite clearance time. The parasitemia at inclusion was 13132.5 trophozoites/\u03bcl in ASAQ group, 26192.5 trophozoites/\u03bcl in AL group and 21347.5 trophozoites/\u03bcl in DHAPQ group. At day 1 after first dose administration, the parasitemia decreased to 1139.8 trophozoites/\u03bcl in ASAQ group, 583.03 trophozoites/\u03bcl in AL group and 277.73 trophozoites/\u03bcl in DHAPQ group. The proportion of patients remaining parasitemic at day 1 was 75% (135/180), 66.8% (119/178) and 48.8% (86/176) respectively in ASAQ, AL and DHAPQ group  versus 10.79% (19/176) in DHAPQ group and 7.86% (14/178) in AL group (p =\u20090.31). Vomiting was more frequent in DHAPQ group 5.68% (10/176) compared to AL group 2.45% (4/178) and ASAQ group 1.66% (3/180) (p =\u20090.06). Labial herpes was more frequent in AL group 3.37% (6/178).Biological tolerance was good during the three treatments. No major biological disorder was observed.The mean of haemoglobin was lower at day 7 in the three treatments groups compared to hemoglobin mean at day 0. The difference between the three treatment groups at day 7 was not significative (p =\u20090.19). Anemia was most frequent at day 7 in three groups compared to inclusion. This was more important in AL group (70.79%). The difference was not significative between the three groups (p =\u20090.31).An improvement of liver function was observed between day 0 and day 7. The number of patients with normal transaminase was higher at day 7 compared to the enrollment.Regarding the mean of creatinine, no significant variation was observed between day 0 and day 7 in the three treatment arms. A significant decrease of bilirubin level was noted at day 7 in the three treatment arms Table\u00a0.At the pair analysis, the haemoglobin mean decreased from day 0 to day 7 with no statistical difference between AL and DHAPQ treatment groups (p =\u20091). From day 0 to day 7, this mean decreased to 1.1\u00a0g/dl versus 0.52\u00a0g/dl respectively in AL and ASAQ groups. Haemoglobin level decreased to 1.4\u00a0g/dl from day 0 to day 7 in the DHAPQ group versus 0.52\u00a0g/dl in the ASAQ group (p <\u20090.001). Overall the ALAT mean decreased from day 0 to day 7 in the three treatments groups. No statistical difference was noted between ASAQ (2.57 UI/L) and AL (3.65 UI/L) groups (p =\u20091) and between AL (16.2 UI/L) and DHAPQ (2.58 UI/L) groups (p =\u20091). From day 0 to day 7, the mean ALAT decreased to 3.65 UI/L in AL group versus 2.58 UI/L in DHAPQ group (p =\u20090.48). The mean ASAT decreased to 6.4 UI/L in ASAQ group versus 9.5 UI/L in AL group from day 0 to day 7 (p =\u20090.54). Same tendency was noted in DHAPQ (8.7 UI/L) and AL (9.5 UI/L) groups from day 0 to day 7 (p =\u20091). There was no significant difference in ASAT level from day 0 to day 7 in ASAQ and DHAPQ arms (p =\u20090.94). From day 0 to day 7, the mean production of creatinin decreased to 0.18\u00a0mg/l in ASAQ group versus 0.41\u00a0mg/l in DHAPQ group (p =\u20091). In AL group this mean decreased to 1.39\u00a0mg/l from day 0 to day 7 versus 0.18\u00a0mg/l in ASAQ group (p =\u20090.77). Regarding AL group versus DHAPQ group, the mean creatinin decreased respectively to 1.39\u00a0mg/L and 0.41\u00a0mg/l from day 0 to day 7 (p =\u20091).-3). There was not significative between AL and DHAPQ groups (p =\u20090.46)  Table\u00a0.Overall, the pair analysis showed an improvement of liver and kidney function from day 0 and day 7.Plasmodium falciparum malaria. These findings are consistent with results reported from other trials. In Senegal, Tine et al. in 2010 reported a cure rate of 96.7% at day 28 for AL in a clinical trial assessing the efficacy and tolerability of new formulation of Artesunate-Mefloquine [Faye et al. obtained a cure rate of 97% for AL in a multicentric study  from September 2007 to November 2008 [This study was undertaken to assess the efficacy and the safety of the three artemisinin combination therapies commonly used in Senegal. The study showed that AL, DHAPQ and ASAQ are highly effective for the treatment of uncomplicated floquine . Faye etber 2008 .Menan et al. from, in a multicentric study including Cameroon, Ivory Coast and Senegal September 2008 to February 2009, obtained a PCR adjusted ACPR at day 28 of 99% in AL group [Makanga et al. obtained a therapeutic efficacy for AL above 95% for the treatment of uncomplicated malaria [Faye et al. in 2008 in a multicentric study  evaluating the non-inferiority of the new paediatric formulation of Artesunate/Amodiaquine versus Artemether/Lumefantrine for malaria treatment reported a therapeutic efficacy at day 28 at 98.7% when ASAQ was given to children under 5\u00a0years of age [AL group . Makanga malaria . Regardis of age .Ndiaye et al., in randomized trial assessing the efficacy of a fixed dose of ASAQ combination from March to December 2006, showed an adjusted ACPR at day 28 more than 98% [than 98% .NDounga et al. in 2005 in Brazaville, obtained 94.4% of cure rate at day 28 when ASAQ combination was given to children [Zwang et al. in a multi-centre analysis of the efficacy of ASAQ combination showed good ACPR after PCR correction. For DHAPQ combination similar results were obtained [Yavo et al. from November 2006 to May 2008 in multi-centre study assessing the efficacy of DHAPQ obtained 99.5% of ACPR after PCR correction [children . Zwang errection .Ashley et al. between July 2002 and April 2003 showed a cure rate of 98.3% in the DHAPQ group [Grande et al. and by Zwang et al[In Tha\u00efland, PQ group . Many stPQ group -25. The ang et al,27.Tine et al., Faye et al. in Senegal and Gbotosho et al. in Nigeria [After the administration of the three ACT used in this study, it resulted in a rapid decrease of fever and parasite. The fever and parasite clearance was obtained respectively at day 2 and day 3 after the initial dose. Similar results were obtained by  Nigeria ,14,28.The three antimalarial treatments were well tolerated with a similar safety profile. Indeed, no serious adverse event was noted. The main adverse events were minor and they were represented by vomiting, abdominal pain, herpes labialis, dizziness and diarrhea. However abdominal pains were more frequent in patients treated with ASAQ, vomiting was more frequent in patients treated with DHAPQ and labial herpes was more frequent in AL group. Previous study demonstrated that these ACTs are well tolerated -15.Olliaro et al. observed a decrease of haemoglobin level at day 7 after malaria treatment [Price et al. and by Zwang et al.[No major biological disorder was observed in our study. Anemia was most frequent at day 7 in three groups compared at day 0. Similar results were noted by others study. reatment . Same trng et al.,31.In Senegal, notification of adverse events has been a great challenge for the National Malaria Control Programme although a pharmacovigilance system to monitor ACT drug related adverse events has been established since 2006 . This stAL was not given with fat diet in our study. This could explain the decrease of the efficacy. However many studies have reported good efficacy when AL was given with fat. Thus Ashley et al. in pharmacokinetic study of AL in 2002 showed that the high-fat allowed having a good absorption of the drug. This had resulted in an increase of therapeutic efficacy .Mayxay M et al., in the efficacy study of AL in Southern Laos between June and November from 2008\u20132010 showed good efficacy of AL with a cure rate more than 95% when AL was given with fatty food [tty food .Many studies reported an increase of haemoglobin level from day 7 to 28 after treatment with ACT ,31. In tDHAPQ, AL and ASAQ are highly effective and safe antimalarial drugs. These ACTs remain useful antimalarial interventions for effective malaria control. It is however important to continue to monitor their efficacy in the context of scaling up of ACTs in Senegal.The authors declare that they have no competing interest.KS, RCT, BF, DS, JLN, OG conceived and designed the study. KS and MN monitored the data collection. KF and LAN collected data in the site. KS and RCT analysed the data. MN, ACL, AA were responsible for the PCR analysis. KS and AA wrote the first draft of the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2334/13/598/prepub"}
+{"text": "Plasmodium vivax infection can be achieved, thus preventing potential relapses and the emergence of new cases outside the Amazon region in Brazil. Surveillance for therapeutic failure in non-endemic areas is advantageous, as it is unlikely that recurrence of the disease can be attributed to a new malaria infection in these regions.Malaria is a potentially severe disease widely distributed in tropical and subtropical regions worldwide. Clinically, the progression of the disease can be life-threatening if it is not promptly diagnosed and properly treated. Through treatment, the radical cure of P. vivax and mixed  malaria was conducted at a travel medicine centre between 2005 and 2011 in Rio de Janeiro and a descriptive analysis of the potential factors related to recurrence of P. vivax malaria was performed. Groups with different therapeutic responses were compared using survival analysis based on the length of time to recurrence and a set of independent variables thought to be associated with recurrence.An observational study of 53 cases of P. vivax malaria were observed. The overall median time to relapse, obtained by the Kaplan-Meier method, was 108\u2009days, and the survival analysis demonstrated an association between non-weight-adjusted primaquine dosing and the occurrence of relapse (p\u2009<\u20090.03). Primaquine total dose at 3.6\u2009mg/kg gave improved results in preventing relapses.Twenty-one relapses (39.6%) of P. vivax malaria relapse. Indeed, the total dose of primaquine associated with non-occurrence of relapses was higher than recommended by Brazilian guidelines.A known challenge to individual cure and environmental control of malaria is the possibility of an inappropriate, non-weight-based primaquine dosing, which should be considered a potential cause of  Plasmodium vivax is responsible for 84% of the cases registered, 99.8% of which occur in the Brazilian Amazon [Malaria is a potentially severe disease widely distributed in tropical and subtropical regions worldwide. Because of present-day ease of travel and the magnitude of migratory movements, it can be considered a global problem. Death can occur in non-immune individuals if the disease is not diagnosed in time and treated appropriately. Most malaria cases in the Americas occur in Brazil, where n Amazon -3.P. vivax or Plasmodium ovale latent in the liver) [P. vivax malaria is known for its benign course, there are reports of more complicated cases in the Brazilian Amazon [Relapse, a common feature of this type of malaria, is defined as the reappearance of the disease and parasitaemia after initial eradication of blood forms. It is caused by the survival of hypnozoites (dormant forms of e liver) . Althougn Amazon ,6 and eln Amazon ,8.P. vivax by treatment can be due to several factors. Although other factors \u2013 such as individual pharmacokinetic variations , adherence to the treatment, drug interactions, adverse drug events, dose adjustment to body weight, and treatment duration \u2013 are directly related to the infection\u2019s response to anti-malarial treatment, primaquine tolerance is a very important issue that few studies have addressed [Failure in deactivating the hepatic hypnozoite forms of ddressed ,10.in vitro method for assessing the hypnozoiticidal effects of treatment, patient follow-up \u2013 particularly outside the transmission areas \u2013 remains the best way to monitor the susceptibility of P. vivax to primaquine.Detection of treatment failure or relapse may prevent malaria\u2019s introduction into unaffected areas where the vector remains a regional menace. As there is no standardized P. vivax malaria relapse at a sentinel unit in a non-Amazonian region and the factors potentially associated with this event.This paper aims to describe the frequency of This descriptive study was conducted from January 2005 to October 2011 at a specialized post-travel care clinic for adolescents and adults (\u226512\u2009years old) at the Instituto de Pesquisa Cl\u00ednica Evandro Chagas (IPEC) in Rio de Janeiro, a state located outside the Amazon region and where malaria transmission does not occur.Eighty-nine patients treated for vivax malaria during the period were studied. Those who had returned to an area of malaria transmission after starting treatment or who had less than 28\u2009days of follow-up were excluded. Accordingly, the analysis contemplated 53 patients (59.6%). These were followed up for 28 to 408\u2009days (median\u2009=\u200962\u2009days).vivax, and the other six (11.3%), with P. vivax and Plasmodium falciparum . Males predominated (75.5%), and ages ranged from 14 to 63\u2009years (median\u2009=\u200929\u2009years). Most of the travellers came from South America: Brazilian Amazon (71.6%), French Guiana (13.2%), Venezuela (3.8%), Guyana (1.9%) and Suriname (1.9%); the rest were from Angola (3.8%) and Indonesia (3.8%).Of the 53 patients studied, 47 (88.7%) were infected only with P. Plasmodium vivax infections were treated with chloroquine  and primaquine. Mixed infections caused by P. falciparum and P. vivax were treated with mefloquine  and primaquine, a therapeutic regimen still used by the Brazilian Malaria Therapy Guidelines until 2009, or with artesunate\u2009+\u2009mefloquine (200\u2009mg\u2009+\u2009400\u2009mg for three days) and primaquine [imaquine ,12. Primimaquine ; or (2) The project was approved by the IPEC Research Ethics Committee, Fiocruz (No. 0031.0.009.000-10), and the information obtained was kept strictly secret and confidential.Plasmodium asexual and sexual stages, according to standard procedures [Thin and thick blood smears were stained with Giemsa and analysed by light microscopy using an immersion oil lens (x100 objective magnification) to identify the parasite species and determine the density of ocedures . MalariaData were collected from the clinic and IPEC pharmacovigilance databases. The variables considered were (a) therapeutic response, classified as relapse or non-relapse; (b) gender; (c) age; (d) adverse drug events; (e) primaquine treatment duration; (f) whether or not the primaquine dose was weight-adjusted; (g) and primaquine total dose. Relapse was considered as the recurrence of parasitaemia following parasitological remission after 28\u2009days from starting treatment for vivax malaria. Primaquine treatment duration was considered an indirect factor related to adherence to treatment, since total dose was the same, regardless of regimen.P. vivax malaria. Groups with different therapeutic responses were compared using survival analysis. Survival time was defined as the number of days elapsed between starting treatment and disease recurrence. The event of interest was relapse of vivax malaria. Cases with no evidence of relapse during the study period were censored at the last day of follow-up. The Kaplan-Meier method was applied to calculate the survival functions, which are defined as the probability of relapse not occurring. The log-rank test was used to compare survival functions among categories of each variable. First the hazard ratios (HR) with 95% confidence intervals were calculated, and then the Cox proportional hazards model. Analysis of Schoenfeld and Martingale residuals was used to assess the Cox method assumptions for covariates of the adjusted model. The smoothing function (spline) was applied for the non-linear continuous variable \u201cprimaquine total dose\u201d in order to show the effect of dose on relapses graphically. Spline coefficient values of less than zero indicate occurrence of protection against relapse. The significance level for all hypothesis testing was set at p\u2009\u2264\u20090.05. Data were analysed using R software, version 2.14.2.Descriptive analysis was performed on factors potentially related to relapse of P. vivax malaria were observed. Most (n\u2009=\u200914) occurred in patients from the Brazilian Amazon; the other patients were from French Guiana (n\u2009=\u20094), Venezuela (n\u2009=\u20092) and Indonesia (n\u2009=\u20091). These relapses occurred in 45.7% of patients treated with primaquine short regimen (N\u2009=\u200935) and in 28.6% of patients treated with primaquine long regimen (N\u2009=\u200914) (p\u2009>\u20090.05). Thirty-two cases were censored in survival analysis, because relapse did not occur during the study period (Table\u2009Twenty-one relapses 39.6%) of Most n\u2009=\u20094 occurreMost n\u2009=\u20094 occurreuela n\u2009=\u2009 and IndoMost n\u2009=\u20094 occurreuela n\u2009=\u2009 and Indo9.6% of Muela n\u2009=\u2009 and IndoP. vivax malaria and that failure to adjust primaquine dosing was the most important factor in the occurrence of relapse. Still, the primaquine total dose related to non-occurrence of relapse was higher than recommended by the Brazilian guidelines. Relapse is considered to be the parasitological recurrence of P. vivax after 28\u2009days of treatment in patients who do not return to an endemic area [The current study, performed in the city of Rio de Janeiro, where malaria transmission does not occur, showed that approximately 40% of patients treated within the last six years had relapses of P. vivax malaria showed a relapse rate of 25% over seven years. However, Duarte et al[et al[A retrospective study in S\u00e3o Paulo, Brazil  among 1,rte et al in Cuiab al[et al in MelboP. vivax infection acquired on a trip to Indonesia, where the prevalence of primaquine-tolerant P. vivax is higher than elsewhere [P. vivax in the South Pacific, Southeast Asia, Western Pacific, Oceania and Central America [Relapses can occur months or even years after the time of initial infection, with a maximum recorded time of four years -20. In tlsewhere . Althouglsewhere , the dem America ,10,21,22 America , from whBesides drug resistance, other factors, such as adherence to the treatment, treatment duration, quality of anti-malarials, adverse drug events and dose adjustment to body weight may be related to the infection\u2019s response to anti-malarial treatment.Adherence to anti-malarial treatment remains an important key to malaria control ,25. DireAnti-malarials\u2019 quality ensures their therapeutic effectiveness in eliminating parasites from the blood and achieving malaria treatment success. Studies performed in Brazil indicate that poor quality anti-malarial drugs and inappropriate storage conditions may contribute to the development of parasite resistance ,31. In tMost adverse drug reactions were mild and involved disorders of the gastrointestinal system \u2013 nausea, vomiting and diarrhoea, which could theoretically have reduced drug absorption. However, in the applied model, relapses were not associated with the occurrence of adverse drug reactions in the study.Primaquine is currently the only drug available for treatment of the latent liver forms of the parasite. The literature suggests that a 14-day course of primaquine may be more effective than a 7-day course, and this superiority may be attributable to significant accumulation of the drug\u2019s active metabolite after longer administration -35. On tConversely, the results clearly indicate an association between non-weight-based primaquine dosing and the occurrence of relapses (p\u2009<\u20090.03). These results agree with studies that suggested dosing adjustments for patients weighting over 70\u2009kg at a time when obesity was not yet a global problem -40.Although it is already known that weight-based dosing of primaquine is required to prevent relapses, it is important that an appropriate weight-based primaquine total dose be determined and confirmed to guide clinical decisions. In this study, a total dose of 3.2\u2009mg/kg indicates protection against relapses. However, the probability of relapse occurring still exists given the upper limit of the confidence interval. Therefore, in order to treat the patients in this study more effectively, a more appropriate dose would be higher (between 3.6 and 4.1\u2009mg/kg) than the one recommended by the Brazilian Ministry Health . TheAlthough small sample size may be considered a limitation on the statistical analysis, an association was found between adjusted primaquine dose and protection against relapse. Also, follow-up of infected patients in non-endemic areas is a good methodological approach to define a true relapse, because no confounding re-infection is possible in such areas.et al[The results reported here corroborate the conclusion of Fernando et al that theThe study suggests that the total dose of primaquine above which relapses do not occur is 3.6\u2009mg/kg. This conclusion is in agreement with WHO and other international guidelines, but is higher than the total dose recommended by Brazilian Ministry of Health Guidelines. Given the ease of travel, the evidence of growing primaquine tolerance around the world and the possibility of its introduction into the Amazon basin, it would be advisable to review the recommendations of the Brazilian Malaria Control Programme. Finally, it may be argued that malaria control programmes should use public health educational messaging campaigns to alert travel clinicians to the fact that risk of therapeutic failure can be reduced by tailoring drug therapies individually to each patient.The authors have no conflicts of interest to disclose.RSP: responsible for conception and design of the work, data collection, data analysis, data interpretation and drafting the manuscript. PB: responsible for conception and design of the work, data analysis, data interpretation, literature review and reviewing the manuscript. LG: responsible for conception and design of the work, data analysis, data interpretation, literature review and reviewing the manuscript. DC: responsible for data analysis and interpretation. APC: responsible for data collection and helped to review the text. CTDR: helped in the design of the work and reviewed the text up to the final version to be published. All authors read and approved the final manuscript."}
+{"text": "Priority setting for artemisinin-based antimalarial drugs has become an integral part of malaria treatment policy change in malaria-endemic countries. Although these drugs are more efficacious, they are also more costly than the failing drugs. When Tanzania changed its National Malaria Treatment Policy in 2006, priority setting was an inevitable challenge. Artemether-lumefantrine was prioritised as the first-line drug for the management of uncomplicated malaria to be available at a subsidized price at public and faith-based healthcare facilities.This paper describes the priority-setting process, which involved the selection of a new first-line antimalarial drug in the implementation of artemisinin-based combination therapy policy. These descriptions were further evaluated against the four conditions of the accountability for reasonableness framework. According to this framework, fair decisions must satisfy a set of publicity, relevance, appeals, and revision and enforcement conditions.In-depth interviews were held with key informants using pretested interview guides, supplemented with a review of the treatment guideline. Purposeful sampling was used in order to explore the perceptions of people with different backgrounds and perspectives. The analysis followed an editing organising style.Publicity: The selection decision of artemether-lumefantrine but not the rationale behind it was publicised through radio, television, and newspaper channels in the national language, Swahili. Relevance: The decision was grounded on evidences of clinical efficacy, safety, affordability, and formulation profile. Stakeholders were not adequately involved. There was neither an appeals mechanism to challenge the decision nor enforcement mechanisms to guarantee fairness of the decision outcomes.The priority-setting decision to use artemether-lumefantrine as the first-line antimalarial drug failed to satisfy the four conditions of the accountability for reasonableness framework. In our understanding, this is the first study to evaluate priority-setting decisions for new drugs in Tanzania against the accountability for reasonableness framework. In addition to the demand for enhanced stakeholder involvement, publicity, and transparency, the study also calls for the institution of formal appeals, revision, and regulatory mechanisms in the future change of malaria treatment policies. Plasmodium falciparum malaria-endemic countries, most being in sub-Saharan Africa, had changed their treatment policies to artemisinin-based combination therapies [Malaria case management with pharmaceuticals is a major challenge in many malaria-endemic countries. Old but cheap and effective medicines are increasingly facing resistance. By 2009, nearly all herapies ,2. On thherapies ,4. When herapies . The guiherapies . Limitedherapies . Among tTanzania has a fairly well-distributed healthcare system, within which 66% of all the healthcare facilities are owned by the government . About 8Rational malaria treatment policy formulation presents challenges to many endemic countries. At times, donors may favour an approach that conflicts with the national preferences ,14. StudThe accountability for reasonableness framework (AFR) Table  is a proDistrict and regional health planners have applauded the framework for its ability to enhance participation, transparency, and the development of relevant criteria for decision making . Where iManaging access to pharmaceuticals is a microcosm of the limit-setting problems of healthcare systems as a whole. If we are right that accountability for reasonableness is a solution to the legitimacy problem in health systems as a whole, then it should be possible to illustrate what such accountability would mean in practical terms... --.This is the first study to apply the AFR framework to evaluate priority-setting decisions that involve selection of drugs in Tanzania. The selection process for drugs to be listed on the national essential medicine list also involves priority setting. We argue that evaluation of these decisions against the AFR framework is one method of improving the process of drug selection. This study therefore reports on the evaluation of the priority-setting decision for the implementation of artemisinin-based combination therapy policy against the four conditions of the AFR framework.This was a qualitative study in which in-depth interviews were held with key informants. At least two participants were selected from the following list of institutions: Muhimbili National Hospital (MNH), National Malaria Control Programme (NMCP), WHO office in Tanzania, Ministry of Health (MoH), Tanzania Food & Drug Authority (TFDA), Medical Stores Department (MSD), and Muhimbili University of Health and Allied Sciences (MUHAS). Participants were experts on malaria research, administration of public health institutions, management of patients, and regulation and management of pharmaceuticals.The purposive sampling method was used to select the key informants from the acknowledgements list of the treatment guideline. A sample of 15 participants was selected based on Guest et al.'s recommendations, which imply that, in a piece of research that is narrow in scope and has a homogenous sample, data saturation is achieved after 12 interviews when thematic analysis is employed .Data were collected by in-depth interviews held with key informants using pretested interview guides and reviews of the treatment guideline between May and July 2010. A digital audio recorder was used to record the conversations. The in-depth interview guide consisted of questions corresponding to the AFR framework . Each inInterviews were transcribed and then analysed using the ATLAS.ti , a qualitative data analysis (QDA) software, using an editing organising-style method . First tTo ensure the validity of the data, participants were assured of the aim of the study and their names were anonymised. The in-depth interviews were held in places where interviewees felt comfortable, and the conversations were recorded by using a digital audio recorder. Interviews were transcribed by the interviewer shortly after each session. Telephone numbers of the interviewees were taken so that if any information was missing, they could be called for clarification. Transcripts were read and re-read several times so that the researcher could become familiarised with their contents before analysis. Coding of both the transcripts and the treatment guideline were done separately, and the emerging descriptions were triangulated during memo writing.Ethical clearance (Ref. No. MU/PGS/SAEC/Vol. III/185) was provided by the MUHAS Ethical Review Committee. Participants were asked for verbal informed consent and for permission to be recorded prior to the interviews.A majority of the participants reported that the decision about the selection and use of ALu was disseminated to the public by the use of radio, television, and newspaper channels in the national language, Swahili. Meetings with other stakeholders, such as pharmacists, were also held as a means of disseminating this information. They also reported that all the public healthcare facilities from the dispensary to the regional hospital level received the treatment guidelines, which contained the decision and rationales for the prioritisation of ALu.The decision about the selection of ALu, but not the reasons behind it, was communicated through advocacy campaigns by National Malaria Control Program (NMCP) and Ministry of Health (MoH) using different media channels in Swahili, the national language. The rationales were written in the treatment guidelines, which are not easily accessible to the public, patients, or other stakeholders who were not directly involved with the task force .Participants reported that the problem of parasite resistance to sulphadoxine-pyrimethamine was the major factor that caused the change of the National Malaria Treatment Policy. At the time of this policy change, three choices of artemisinin-based combination drugs recommended by WHO for management of uncomplicated malaria were available. These drugs included sulphadoxine/pyrimethamine-artesunate (SP-AS), amodiaqine-artesunate (AQ-AS), and ALu. ALu had the following advantages over the other drugs: (1) it was the only fixed combination drug among the three, (2) neither of its components had documented reports of parasite resistance, (3) it was safer, as amodiaquine's safety profile had already generated concern among healthcare providers and the general public and (4) it was the most efficacious drug.Participants reported that affordability was not discussed because of the agreement between the government and the Global Fund to provide the drug at a subsidized price at the public and faith-based healthcare facilities. This was facilitated by the agreement between WHO and the company that manufactures ALu, which made the drug available at cost price for use in the public sector for malaria-endemic developing countries. However, some participants raised concerns about the sustainability of this financing approach.... as a country you cannot just sit and wait for somebody to carry the burden of maintaining your population's health, imagine if Global Fund does not keep on sustaining this program!--Representative from MSDParticipants pointed out that, based on Global Fund requirements, the subsidized ALu could only be made available at public and faith-based primary to tertiary healthcare facilities. One participant who works at a tertiary hospital expressed concerns about the low availability of ALu in the primary healthcare facilities:I receive patients from the dispensaries who have been given sulphadoxine-pyrimethamine and amodiaquine and they sometimes ask why they are still prescribing these drugs when the first-line drug is artemether-lumefantrine!--Physician from MNHOne participant was concerned about the urban-poor populations who have to make a choice between long waiting times and unreliable public health services versus reliable but unaffordable services in the private sector. Participants from National Malaria Control Program (NMCP) said that other strategies were being considered to make sure ALu was also available at a subsidized price in the private-for-profit healthcare facilities.Participants said that fixed-combination antimalarial drugs have better compliance, hence are more effective compared to non-fixed combinations. However, artemether-lumefantrine's six-dose regimen of four tablets to be taken twice a day with a fatty meal for three successive days was a major concern.There are obvious improvements, admissions due to severe malaria nowadays are few and this could be explained by the effectiveness of artemether-lumefantrine. As a tertiary hospital we used to have a lot of patients coming in coma due to cerebral malaria; this is not research data but my own observation.--Physician from MNHAbout its cost-effectiveness, the majority of participants reported that there were no research data at the time of the policy change that indicated the cost -effectiveness of ALu.The task force committee was composed of 25 members; 21 were medical doctors and four were pharmacists. The medical doctors were paediatricians, obstetricians/gynaecologists, parasitologists, and pharmacologists from Muhimbili National Hospital (MNH) and Muhimbili University of Health and Allied Sciences (MUHAS). Others were working with the WHO country office, National Malaria Control Program (NMCP) and Integrated Management of Childhood Illnesses (IMCI) programs within Ministry of Health (MoH). The pharmacists were working with Medical Store Department (MSD) in the procurement and distribution of drugs, Tanzania Food and Drug Authority (TFDA) in food and drug regulations, and the pharmaceutical services unit at the Ministry of Health. All the committee members were from institutions based in the city of Dar es Salaam. Participants from NMCP who coordinated the policy-change process reported that they did not have written documents on procedures to select the committee members but that they 'knew' the main stakeholders. This approach was criticised by one of the participants:Sometimes I feel as if it is not adequate because if I had not contacted those people with my findings, I might not have been invited. So there might be other people who are in the field but because they had not made contact with Ministry of Health officials, they did not get the opportunity to be part of the task force committee.--Researcher from MUHASParticipants reported that patients and the public were indirectly involved through representation and consultations. It was also argued that the task force committee members were potential malaria patients, and therefore, they were themselves representatives of patients and the public. The idea of direct patient and general public involvement in the committee was opposed by some participants who labelled it as inappropriate because policy making is an expert's field.Participants reported that many consultations were held before and even after the assembly of the task force committee to get input and technical advice about policy change. Participants from National Malaria Control Program (NMCP) said that the first draft of the guideline was reviewed by experts from the National Institute for Medical Research, Malaria Consortium-Kampala, WHO Afro, M\u00e9decins sans Fronti\u00e8res, Ifakara Health Institute, and Eduardo Mondlane-Maputo (Mozambique). They also reported that the draft guidelines were shared by all district and regional medical officers as well as partner countries for technical advice. Councils gave input during the zonal training sessions that were conducted countrywide.The reasons for the prioritisation of ALu appeal to stakeholders because they were based on scientific evidence of clinical efficacy, safety, and formulation profiles. Financing mechanisms were already in place to ensure that the drug would be available and affordable at the public and faith-based healthcare facilities.The study found that there were no transparent procedures to engage the stakeholders. Some stakeholders were involved because of their positions in the relevant institutions, while others were involved through informal links with officials in the Ministry of Health (MoH) or National Malaria Control Program (NMCP). The committee lacked professional, institutional, and countrywide representation. It was largely made up of medical doctors and a few pharmacists; neither nurses nor medical lab personnel were included. All the physicians, specialists, and academicians within the task force committee came from one hospital, i.e. Muhimbili National Hospital (MNH) and one university, Muhimbili University of Health and Allied Sciences (MUHAS). There were other similar institutions in the country that could have provided members to increase representation.Participants said that it is not easy for anybody to appeal against the policy; however, if established facts and evidence on toxicities and level of resistance were to be provided, the policy decision could be revised.I think the best way for people to appeal is to provide scientific data in the scientific forum; it is impossible for an individual to come up and say this is not the right drug. From the community side, I think the only mechanism they have is to express their concerns in the newspapers and other media; otherwise, I don't see any other channels at the moment.--Participant from TFDAPharmacovigilance forms, which are used as post-marketing surveillance tools in monitoring drug safety, were mentioned as effective tools to generate evidence on the safety profile of ALu. However, one participant disagreed with the reliability of this approach.Most of our clinics are crowded, before you can take a minute or two to fill in the pharmacovigilance form, 20 people are already shouting at you, so it is one avenue but is not satisfactory.--Physician from MNHThere is no mechanism in place to appeal against the decision to prioritiseALu.There was no enforcement mechanism. The demand for enforcement was seen as new and was considered unnecessary by some participants. Other participants said that an independent enforcement body is necessary during a treatment policy change.I am not sure if there was a watchdog. I think it is really very important because sometimes people say the drug companies are pushing for policy change.--Pharmacologist from MUHASDespite growing concerns about the influence of pharmaceutical companies on change of treatment policies, we did not find any enforcement mechanism to ensure the other three conditions were met.This study evaluated the process of changing the Tanzania National Malaria Treatment Policy against the AFR framework. The process resembled other priority-setting decisions, in that it involved making choices among competing alternatives in the context of limited resources, conflicting evidence, diverse stakeholder involvement, and the vagaries of political cycles . The infWhen evaluated against the AFR, this study found that the publicity condition was not fully satisfied. The study found that the decision to prioritiseALu was made public, but not the reasons behind it. Publicity requires that the decision and its rationales, such as the ones for coverage of new drugs like ALu, be publicly accessible to all stakeholders, including the patients . SimilarThe study found that the rationale but not the stakeholder involvement part of the relevance condition was satisfied. ALu was the only artemisinin-based antimalarial drug that was in a fixed-combination formulation. It was also the most efficacious drug and neither of its two components had any documented reports of parasite resistance . Cost-efThe study found that stakeholders were not adequately involved and represented. Appropriately conducted stakeholder participation not only facilitates and widens discussions in the decision-making process but also enhances publicity . The tasEven though we argue in favour of wide professional and institutional representation, this should not aim to replace patients and the general public. There is evidence that health professionals' views can differ considerably from those of their patients . HoweverThe appeals and revision condition was not satisfied. To our knowledge, there are no formal appeals mechanisms in the Tanzanian healthcare system for policy decisions relating to access to new drugs or healthcare. Pharmacovigilance systems in developing countries are weak, and hence cannot be considered an efficient appeal mechanism to inform policy makers of a drug causing harm to its users. The lack of reliable appeal mechanisms suggest that good arguments and evidences against the original decision and which aim at improving faulty policy decisions can have no clear way back onto the decision table. A study by Maluka et al. reported a lack of clear formal channels for appeals at the district level in Tanzania .The enforcement condition was not satisfied. This condition recognises that public or private regulation may be necessary if self-regulation proves unsuccessful or inadequate. There are growing concerns about the influence of pharmaceutical companies, which lobby and push for national treatment policy changes, especially in developing countries ,16. HencThe qualitative research method is the best approach to study complex phenomena when a researcher wants to capture the views of individual persons, including their feelings, understandings, interpretations, beliefs, and ideologies. Through the use of this method, the participants shared their stories and conflicting views beyond what the researchers expected to hear or what is available in the literature.The study used the AFR as an evaluation framework; however, there are objections to its use as an approach to achieve justice and fairness ,42. MoreThis study applied the AFR framework to evaluate the priority-setting decision for new antimalarial drugs at the national level. The study found that the priority-setting decision in favour of ALu failed to satisfy the four conditions of fair process prescribed in the ethical framework of accountability for reasonableness. The findings of this study are consistent with the reports of other studies that evaluated priority-setting decisions against AFR at the district level. The study emphasises the need to involve a wide range of stakeholders in priority-setting decisions. It further highlights the need to make priority-setting decisions and their rationales public and to establish mechanisms for appeal, revision, and regulation in order to enhance fairness and legitimacy of the decision outcomes.The authors declare that they have no competing interests.ATM conceived and designed the study, collected the data, performed the analysis, and prepared the draft of the manuscript. EAK fine-tuned the study design and coordinated the data collection and analysis. All authors read and approved the final manuscript."}
+{"text": "P. falciparum malaria in South-East Asia highlights the need for continued global surveillance of the efficacy of artemisinin-based combination therapies.The emergence of artemisinin-resistant P. falciparum malaria in a randomized controlled trial of dihydroartemisinin-piperaquine vs. artemether-lumefantrine from 2005 to 2008. (ISRCTN88705995)On the Kenyan coast we studied the treatment responses in 474 children 6\u201359 months old with uncomplicated P<0.001) and from 81% to 95%  in the DHA-PPQ and AM-LM groups, respectively. In parallel, Kaplan-Meier estimated risks of apparent recrudescent infection by day 84 increased from 7% to 14% (P\u200a=\u200a0.1) and from 6% to 15% (P\u200a=\u200a0.05) with DHA-PPQ and AM-LM, respectively. Coinciding with decreasing transmission in the study area, clinical tolerance to parasitemia (defined as absence of fever) declined between 2005\u20132006 and 2007\u20132008 . Neither in vitro sensitivity of parasites to DHA nor levels of antibodies against parasite extract accounted for parasite clearance rates or changes thereof.The proportion of patients with residual parasitemia on day 1 rose from 55% in 2005\u20132006 to 87% in 2007\u20132008 (odds ratio, 5.4, 95%CI, 2.7\u201311.1; The significant, albeit small, decline through time of parasitological response rates to treatment with ACTs may be due to the emergence of parasites with reduced drug sensitivity, to the coincident reduction in population-level clinical immunity, or both. Maintaining the efficacy of artemisinin-based therapy in Africa would benefit from a better understanding of the mechanisms underlying reduced parasite clearance rates.ISRCTN88705995Controlled-Trials.com  Over the last few decades the global spread of parasite resistance to key antimalarial drugs such as chloroquine and pyrimethamine has been a challenge for malaria control programs based primarily on prompt and effective treatment in vivo response to two ACT regimens, namely dihydroartemisinin-piperaquine (DHA-PPQ) and artemether-lumefantrine (AM-LM) over time. The study was conducted from 2005 to 2008, coinciding with the introduction of artemether-lumefantrine (Coartem\u2122) as the exclusive first-line treatment for all presumptive cases of uncomplicated P. falciparum malaria in Kilifi District, Coast Province, Kenya in 2006.Using data from a randomized controlled clinical trial, we performed a post-hoc analysis of the The study was conducted at the Pingilikani study site P. falciparum malaria in Kilifi, Kenya . The primary efficacy endpoint was the 28-day cure rate adjusted for reinfection . Assuming a cure rate of 95% with AM-LM and a 5% drop-out rate, we calculated that 250 patients per arm would provide 80% power to test a 5% non-inferiority margin with a 97.5% one-sided confidence interval.This is a detailed analysis of treatment response rates according to year of enrollment in a non-inferiority randomized controlled trial that evaluated the efficacy of DHA-PPQ vs. AM-LM in the treatment of children with uncomplicated P. falciparum malaria who met the following selection criteria: reported or documented fever \u226537.5\u00b0C, P. falciparum mono-infection, microscopically determined peripheral asexual parasite density of 2,000\u2013200,000/\u00b5L, body weight >5 kg and signed informed consent by parent or legal guardian. We excluded patients with known allergies, severe malaria or danger signs We enrolled pediatric outpatients aged 6\u201359 months with uncomplicated Study drugs were administered orally with food or drinks under direct supervision. For children <2 years old tablets were crushed, mixed with 50 ml water, and administered as slurry. DHA-PPQ was given 24-hourly for 3 days at single target doses of 2.25 mg of DHA/kg body weight and 18 mg of PPQ/kg . Doses were rounded up to the nearest half tablet. AM-LM was administered as whole tablets according to the manufacturer's instructions in 6 doses over 3 days  at mean target doses of 2 mg of AM/kg and 12 mg of LM/kg . Participants who vomited or rejected the study drug within 30 min received a second full dose, and those who vomited or rejected the study drug after 30 min but within 1 h received a second half dose. Vomiting or rejecting the second dose led to withdrawal from the study and administration of rescue medication. Stability tests that were performed by the drug manufacturers upon request confirmed that titer and degradation products of study drugs were still within stringent regulatory specifications in 2009, >10 months after completion of the study.During the 3-day treatment phase, patients were admitted to the KEMRI research ward in Pingilikani to ensure strict adhesion to dosing intervals. Patients were seen by study clinicians on days 0, 1, 2 and 3 and then for weekly follow-up visits until day 63 and finally on day 84 for collecting medical history, vital signs, malaria blood smears, and adverse events. Giemsa-stained malaria slides were read blinded by the same microscopist throughout the study according to KEMRI standard operating procedures.MSP2 gene (PFB0300c) MSP1 and GLURP) or semi-automated capillary electrophoresis-based analysis of fluorescence-labeled MSP2 PCR products MDR1 locus (PFE1150w) were determined by quantitative real-time PCR using the \u03b2-tubulin gene (PF10_0084) as internal control and 3D7 (1 copy) and Dd2 (2 copies) reference strains for calibration To distinguish recrudescent from new infections and to determine the multiplicity of infection (MOI) index, matched pairs of parasite isolates obtained at baseline and recurrence were compared using RFLP-based genotyping analysis of repeat length polymorphisms in the Serum samples collected on day 7 were analyzed for LM by solid-phase extraction and liquid chromatography (LC) with UV detection as described previously in vitro culture according to standard protocols in vitro growth of parasite isolates by 50% (IC50) compared to unexposed controls using regression analysis of the dose-response curves from duplicate 3H-hypoxanthine uptake 72-hour exposure experiments Parasite isolates were adapted to We used an established ELISA protocol to measure concentrations of antibodies against parasite schizont extract (A4 strain) D1) as the log10 quotient of baseline and day 1 parasitemias (after setting parasitemias below the microscopic detection threshold on day 1 to 10/\u00b5L). We also analyzed the probability that parasites were detected by microscopy or not on day 1 by binomial logistic regression for the influences of period of enrolment (2005\u20132006 vs. 2007\u20132008), log10 baseline parasitemia, treatment (AM-LM vs. DHA-PPQ), dose per body weight, patient age, number of previous malaria episodes and anti-schizont antibody levels fitting each of these separately in univariable analyses and then combined in one multivariable analysis if they remained significant at the P\u22640.2 level. Similarly, we analyzed percentage reductions of parasite densities from baseline calculated by dividing baseline densities with densities at day 1 or 2 multiplied by 100. Parasite and fever clearance times were estimated by parametric survival analysis of the time from baseline to the first of two consecutive negative blood smears or temperature measurements <37.5\u00b0C, respectively. Risk of recrudescent primary or secondary (re-) infections was assessed by the Kaplan-Meier (KM) method for survival data. For these analyses, patients who did not meet a study endpoint (either absence or recurrence of parasitemia from day 7 to day 84) were censored at the last visit before dropout .As a measure of drug efficacy, we computed the day 1 parasite reduction ratio (PRR dropout . Cox proP>0.3) (P<0.05), weighed about 1 kg more (P<0.05), had higher parasitemias (P\u200a=\u200a0.03) and axillary temperatures (P<0.001), increased number of parasite clones as determined by MSP2 allele typing (P<0.01) and higher hemoglobin concentration (P\u200a=\u200a0.03 in AM-LM group only). Other characteristics, notably platelet concentrations Between September 2005 and April 2008 we enrolled 474 patients. Enrollment was temporarily suspended between July 2006 and March 2007. 450 patients received a full treatment course . RepeateP>0.3) . Early tP>0.3) . Table 1vs. 48 hours, respectively; P<0.001) resulting also in prompter clearance of fever . By day 28, Kaplan-Meier (KM)-estimated PCR-adjusted rates of recrudescent primary infections were 1%  and 1%  in the DHA-PPQ and AM-LM groups, respectively . By day 84 we noted considerably higher PCR-adjusted recrudescence rates with both DHA-PPQ  and AM-LM  . By day 84 reinfections occurred in 39%  and 42%  of children treated with DHA-PPQ and AM-LM groups, respectively (P\u200a=\u200a0.7), with no difference in median time to reinfection  in the DHA-PPQ group and from 81% to 95%  in the AM-LM group. Median day 1 parasite reduction ratio in the DHA-PPQ group dropped by 78% (4.6-fold lower) and in the AM-LM group by 69% (3.2-fold lower) between the 2005\u20132006 and 2007\u20132008 enrollment periods (P<0.001 by logistic regression) and the same effect was observed within age groups (P<0.002) and a more than two fold risk of apparent recrudescent primary infections by day 84 (P\u200a=\u200a0.01 when pooling data across treatment groups). Reinfection rates by day 84, which could have confounded PCR-based classification of recrudescent primary infections P>0.2; When comparing treatment responses in patients enrolled in 2005\u20132006 e groups . These ce groups ; P<0.002A total of 105 and 101 serum samples obtained on day 7 from patients who received either DHA-PPQ or AM-LM were analyzed for piperaquine and lumefantrine, respectively. Day 7 serum concentrations did not differ between the two study periods  . Since sP<0.001). This apparent loss in population-level clinical immunity could not be explained by lower numbers of previous recorded exposure in study participants  or serological indices of exposure   .D1 (OR>1.9 and P<0.001)  . Anti-scanalysis .D1) was analyzed for time differences with baseline parasitemia as a continuous covariate. When this was done, the differences between time periods remained, particularly for those patients with low baseline parasitemia (P\u200a=\u200a0.06). Thus the least parasitized children experienced the greatest decrease in efficacy through time. The risk of parasitemia on day 1 was also significantly higher in children enrolled in 2007\u20132008 after adjusting for patient age . Similar results were obtained for AM-LM , PPQ (n\u200a=\u200a53) and LM (n\u200a=\u200a60) were not associated with PPRD1 . Paradoxically, median DHA IC50 values dropped from 2.2 nM in 2005\u20132006 (n\u200a=\u200a39) to 0.8 nM in 2007\u20132008 (n\u200a=\u200a20) .Baseline ICP<0.001; . In vitrMDR1 per parasite genome  were found to harbor infections with >1 copy of e genome .P. falciparum infections to treatment with ACTs after three years of their use in a randomized controlled clinical trial and less than two years after the introduction of AM-LM as first-line treatment in the Coast Province of Kenya. The parasite prevalence rate on day 1, which is primarily determined by the fast acting artemisinin component of ACTs P. falciparum infection reported from Western Cambodia  in vivo artemisinin resistance Here we report a significant decline of early response rates of P<0.05), had higher body temperatures , and had 1.3-fold higher baseline parasite densities (P<0.05). These differences are most likely to be a consequence of decreased transmission in the study area that occurred mainly before 2005 D1) (D1. Both patient age as a demographic correlate of progressively acquired immunity and specific antibody responses have shown consistent associations with risk of recrudescent primary infections What factors could plausibly have caused the observed drop in early treatment responses measured at peak drug exposure? On average children enrolled in 2007\u20132008 were 6 months older (2005 D1)  thus raiin vitro evidence of reduced artemisinin sensitivity in isolates obtained from patients with reduced in vivo response rates that could have suggested a gradual selection of \u2018less-sensitive\u2019 parasites. One explanation is that either unstable or in vivo-confined phenotypes may have escaped detection when measuring in vitro growth response rates in culture-adapted isolates. Alternatively, our in vitro test which covers >1 intraerythrocytic replication cycle may have been insensitive to changes in stage-specific artemisinin activity causing prolonged circulation of young ring-form parasites Is it possible to quantify the extent, if any, to which changes in parasite sensitivity to the drugs could have contributed to the observed progressive reduction in responsiveness to treatment ? We failen masse to cause the apparently high prevalence of reduced sensitivity observed here. On the other hand, if tolerance is induced, rather than genetically encoded, for example through epigenetic mechanisms, its rise could be very rapid. Currently, we lack efficient molecular tools for mapping the spread of parasites with altered responsiveness and we do not know whether the observed small changes in responsiveness to treatment, which are reminiscent of long term observations from the Thai-Myanmar border per se, whether due to declining clinical immunity or changes in parasite sensitivity, could potentially impact on the clinical benefit of artemisinins in the treatment of malaria The observation of declining parasitological response to ACTs in the context of slowly declining malaria transmission intensity in Kilifi and minimal access to monotherapy raises a number of issues. First, immunity appears to play a complex role in parasite clearance during treatment. Second, it seems unlikely that parasites with reduced sensitivity to ACTs have spread from Southeast Asia to East Africa Figure S1P. falciparum schizont antibody responses  measured at baseline in children enrolled in 2005\u20132006 vs. 2007\u20132008. Solid lines represent medians and dashed lines indicate binomial 95% confidence intervals.Dot plots of anti-(EPS)Click here for additional data file.Figure S2Fitted curves of the risk of residual parasitemia on day 1 in patients treated with DHA-PPQ (A\u2013C) and AM-LM (D\u2013F). The risk of day 1 parasitemia was fitted using logistic regression for baseline parasite density , patient age , and body-weight adjusted doses of DHA (C) and AM (F). Black lines represent predicted risk in patients enrolled in 2005\u20132006 and gray lines represent predicted risk in patients enrolled in 2007\u20132008.(EPS)Click here for additional data file.Figure S350) of culture-adapted P. falciparum isolates collected from patients in 2005\u20132006 and 2007\u20132008. IC50 responses were determined for dihydroartemisinin (DHA)  , piperaqin (DHA)  and lumein (DHA) .(EPS)Click here for additional data file.Table S1Relationship between pharmacokinetic and pharmacodynamic parameters over time.(DOC)Click here for additional data file.Protocol S1Trial Protocol.(DOC)Click here for additional data file.Checklist S1CONSORT Checklist.(DOC)Click here for additional data file."}
+{"text": "Drug resistance of falciparum malaria is a global problem. Sulphadoxine/pyrimethamine-resistant and mefloquine-resistant strains of falciparum malaria have spread in Southeast Asia at lightning speed in 1980s-1990s, and the Cambodia-Thailand border is one of the malaria epidemic areas with the most severe forms of multi-drug resistant falciparum malaria.Artemisinin-piperaquine (AP), dihydroartemisinin-piperaquine phosphate (DHP) and artemether-lumefantrine (AL) were used to treat 110, 55 and 55 uncomplicated malaria patients, respectively. The total dosage for adults is 1,750 mg  of AP, 2,880 mg  of DHP, and 3,360 mg  of AL. The 28-day cure rate, parasite clearance time, fever clearance time, and drug tolerance of patients to the three drugs were compared. All of the above methods were consistent with the current national guidelines.The mean parasite clearance time was similar in all three groups , and there was no remarkable difference between them; the fever clearance time was also similar . After following up for 28-days, the cure rate was 95.1%(97/102), 98.2%(54/55) and 82.4%(42/51); and the recrudescence cases was 4.9%(5/102), 1.8%(1/55) and 17.6%(9/51), respectively. Therefore, the statistical data showed that 28-day cure rate in AP and DHP groups was superior to AL group obviously.The patients had good tolerance to all the three drugs, and some side effects  could be found in every group and they were self-limited; patients in control groups also had good tolerance to DHP and AL, there was no remarkable difference in the three groups.AP, DHP and AL all remained efficacious treatments for the treatment of falciparum malaria in Cambodia-Thailand border area. However, in this particular setting, the AP regimen turned out to be favourable in terms of efficacy and effectiveness, simplicity of administration, cost and compliance.Chinese Clinical Trial Register under identifier 2005L01041.The trial was registered at  Malaria kills around one million people each year, although is a preventable and treatable infectious disease . BecauseDrug resistant falciparum malaria is a global problem. Sulphadoxine/pyrimethamine and mefloquine resistant strains of falciparum parasites have spread quickly throughout Southeast Asia in 1980s-1990s, and the Cambodia-Thailand border is one of the endemic regions with the most severe multi-drug resistant falciparum malaria -6. WHO hDihydroartemisinin-piperaquine phosphate in a fixed-dose preparation of dihydroartemisinin 40 mg and piperaquine phosphate 320 mg has been shown to be highly effective in treating falciparum malaria with an efficacy of over 90% in China, Vietnam, and Cambodia -12. ArteArtemisinin-piperaquine was recently developed by Chinese scientists and has shown good efficacy and safety in a two-day treatment course . Piperaqin vivo [The pharmacodynamics studies of artemisinin and its derivatives have shown that artemisinin (ART) and its derivatives are characterized by a quick absorption, wide distribution and quick excretion in vivo . Clinicain vivo ,17.In this trial, the effectiveness and safety of the three drugs for the treatment of uncomplicated falciparum malaria were compared and evaluated for developing an optimal anti-malarial regimen.The trial was conducted in Pursat Referral Hospital, Pursat province, Cambodia. Pursat is located in the western part of the country bordering Thailand. It is located between the Tonle Sap lake and the northern end of the Cardamom Mountains. The patients for this trial came from the forest area in the Cambodia and Thailand boarder, where multi-drug resistant falciparum malaria is widespread.The implementation of this study is according to the \"Agreement of Cambodia-China Fast Malaria Control Co-operation\" and \"National Ethics Committee for Health Research (No.03G/03NECHR)\" signed by Guangzhou University of Chinese Medicine, China, and the Ministry of Health, Kingdom of Cambodia.It was a prospective, open labeled, randomized controlled trial conducted on patients with microscopically confirmed symptomatic of falciparum malaria, between seven and 65 years of age. All patients with symptoms of malaria, such as fever, and who had not been administrated any anti-malarial drug, including sulphonamide, tetracycline or trimethoprim, in the previous week, were enrolled . A standInclusion criteria: (1) patients with clinical symptoms of malaria, such as fever (axillary temperature\u226537.5\u00b0C), (2) age ranged from seven to 65 years, (3) peripheral blood smears showed that falciparum malaria asexual parasite were 1,000~100,000/\u03bcl, (4) no anti-malarial had been taken within seven days prior to the onset of symptoms or admission, including sulphonamide, tetracycline or trimethoprim.Exclusion criteria: (1) pregnant women;(2) lactating mothers; (3) children under seven years of age, senior persons elder than 65 years of age; (4) patients with severe vomiting or diarrhoea, or other signs of severe malaria; (5) febrile diseases other than falciparum malaria; (6) exiting before the 28-day follow up and/or lack of follow-up data; (7) patients identified as re-infection by PCR were also excluded from the final cure rate and recrudescence rate calculation.Patients were allocated to three treatment arms based on a pre-generated randomization list made in blocks of 20 that was produced and held independently of the field teams by a statistician. The individual allocations were kept in sealed, opaque envelopes and opened only after enrolment by the field team. Patients and clinical field workers were not blinded to the treatment arm after allocation. Microscopists were blinded to treatment allocation at follow-up examinations. Adverse events were assessed by symptom enquiry at every visit.AP tablets containing 62.5 mg artemisinin (ART) and 375 mg piperaquine (PQ) per tablet were provided by Artepharm Co., Ltd., China (batch No. was 20030820). DHP tablets containing 40 mg dihydroartemisinin (DHA) and 375 mg piperaquine phosphate (PQP) were provided by Guangzhou Holleykin Pharmaceutical Co., Ltd. (batch No. was 20030301). AL tablets containing 20 mg artemether and 120 mg lumefantrine were obtained from Beijng Novartis Pharm Ltd. (batch No. was 20030822). The regimens of AP, DHP and AL were listed in Additional file At enrollment, a full physical examination, including a neurological examination and a medical history, was performed. Thick and thin blood smears were examined by microscopy, and routine haematology was performed. Drug administration was observed by study physicians. If vomiting occurred within 1 h of dosing, the medications were re-administrated. The patients were kept in the hospital for 7 days or until parasites were cleared and were followed-up weekly till D28 from the start of treatment or whenever he/she felt sick within 28 days. At each follow-up, blood smear was done via a finger prick. Blood spots were collected from those with reappearance of asexual parasitaemia. All information was recorded on a standard case record form.A total of 245 cases of uncomplicated malaria were enrolled for this study. The patients were separated into three random groups with 122 cases in AP group, 61 cases in DHP group and 62 cases in AL groups. And complete data from 110, 55 and 55 patients, respectively, were obtained and analyzed.Patients were observed in the hospital for 7 days and followed up at D14, D21 and D28 for any symptoms or malaria parasites changes. Temperature was taken every six hours and once daily after it returned to normal(37.5\u00b0C) for 24 hours. Ward rounds were done twice daily in the morning and afternoon, and side effects were inquired and recorded according to the \"Side Effects Record Form\".Thick and thin blood smears were taken at 7 AM and 5 PM from D0 to D4 to calculate the number of asexual form per 200 WBCs (White Blood Cells). If the number of parasites < 1/200 WBC, 200 fields were examined to identify parasite clearance. Three consecutive negative blood smears (from D5 to D7) indicated clearance of parasite. For follow-ups, blood smears were taken at D14, D21 and D28 to investigate recrudescence. If a patient had fever, he or she was advised to go back to the hospital for further examination. Blood spots collected D0 and positive blood smears during 28-day follow up were kept for qualitative PCR analysis. Blood spots of patients whose blood smears were positive within 28-day follow up were sent to the Oxford University Clinical Research Unit, Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam, for PCR qualitative analysis to differentiate re-infection and recrudescence.WBC and RBC (Red Blood Cell) counts and electrocardiogram examination were done at D0 and D7. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), direct bilirubin (DBIL), creatinine (CR) and blood urea nitrogen(BUN)were also examined at D0 and D7. Abnormal items found at D7 were re-examined every week until the patient recovered.The following four main efficacy parameters were used: Early Treatment Failure (ETF); Late Clinical Failure (LCF); Late Parasitological Failure (LPF); Adequate Clinical and Parasitological Response (ACPR) . The occThe patients in the trial continued to live in the area of transmission, therefore, re-infection during the four weeks of follow-up was possible. Then, PCR-genotyping were used to distinguish recrudescence from re-infection. Blood samples were taken at baseline (D0) and on the day of reappearance of parasitaemia . The nesStatistic analysis was conducted using SAS 6.12. Data were presented as means \u00b1 standard deviation (SD) or as the geometric mean and range. All statistical tests were two-tailed, and a significance level of 0.05 was used.P. falciparum, with treatments for AP, DHP and AL, then research on the therapeutic efficacy and safety. The clinical trials registry number was 2005L01041.The study was approved by the National Ethics Committee for Health Research of the Ministry of Health, Phnom Penh, Kingdom of Cambodia, and the Ethics Committee of Guangzhou University of Chinese Medicine, Guangzhou, China. This ethics approval of the trial describes recruitment of patients with uncomplicated infection with From 22 July, 2005 to 18 October, 2005, 220 patients completed in the study with 110 receiving AP, 55 receiving DHP and 55 receiving AL  of the three groups on D28 were 97/110 (88.2%), 54/55 (98.2) and 42/55 (76.4%), respectively , 54/55(98.2%), 42/51 (82.4%) for AP, DHP and AL, respectively .In this study, three different forms of ACTs  were compared in the multi-drug resistant falciparum malaria epidemic areas in the Cambodia-Thailand border.The data from this report demonstrated that the AP regimen turned out to be favourable in terms of efficacy and effectiveness, simplicity of administration, cost and compliance in this particular setting, and it might play an important role for malaria control in the near future.The authors declare that they have no competing interests.JS organized the program and drafted the manuscript; SD participated in and supervised the program; BT participated in the projects and did the field work. S Seila did the field work and was in charge of patients inclusion; YX did the field work and the clinical observation; S Sokunthea and FO checked the blood smear and did the biochemistry analysis; CZ and QW collected the data; CD collected and analyzed the cases; and GL participated in the design of this study and coordination and helped to draft the manuscript. All authors read and approved the final manuscript.Table S1 - Dosing schedules for artemisinin-piperaquine (AP), dihydroartemisinin-piperaquine phosphate (DHP) and artemether-lumefantrine (AL). *It is better to take this drug together with milk and food.Click here for file"}
+{"text": "Treatment options for Visceral Leishmaniasis (VL) in East Africa are far from satisfactory due to cost, toxicity, prolonged treatment duration or emergence of parasite resistance. Hence there is a need to explore alternative treatment protocols such as miltefosine alone or in combinations including miltefosine, sodium stibogluconate (SSG) or liposomal amphotericin B. The aim of this trial is to identify regimen(s) which are sufficiently promising for future trials in East Africa.A phase II randomized, parallel arm, open-labelled trial is being conducted to assess the efficacy of each of the three regimens: liposomal amphotericin B with SSG, Liposomal amphotericin B with miltefosine and miltefosine alone. The primary endpoint is cure at day 28 with secondary endpoint at day 210 (6 months). Initial cure is a single composite measure based on parasitologic evaluation  and clinical assessment. Repeated interim analyses have been planned after recruitment of 15 patients in each arm with a maximum sample size of 63 for each. These will follow group-sequential methods (the triangular test) to identify when a regimen is inadequate (<75% efficacy) or adequate (>90% efficacy). We describe a method to ensure consistency of the sequential analysis of day 28 cure with the non-sequential analysis of day 210 cure.A regimen with adequate efficacy would be a candidate for treatment of VL with reasonable costs. The design allows repeated testing throughout the trial recruitment period while maintaining good statistical properties (Type I & II error rates) and reducing the expected sample sizes.NCT01067443ClinicalTrials.gov:  Visceral leishmaniasis (VL), or kala-azar, is the most serious of all Leishmania infections and is fatal if left untreated . A compl\u00ae), toxicity  or prolonged treatment duration , resulting in the additional concern of compliance and possible emergence of parasite resistance. Paramomycin, both in monotherapy and in combination with SSG has just recently completed development in the East Africa. However its use in monotherapy does not appear realistic in the region [\u00ae and miltefosine which could provide new treatment option with reasonable costs due to reduction in needed dosages and treatment duration and less risk of resistance emerging [The monotherapeutic treatment options available in East Africa are far from satisfactory due to cost  in collaboration with the trial sites and is sponsored by the Drugs for Neglected Diseases initiative (http://www.dndi.org).The areas of the two trial sites are endemic for VL. They are Dooka Hospital, Gedarif State, Sudan and Kimalel Hospital, Baringo district, Kenya. Addition of another Sudanese site will be considered. Data are brought to the Data Centre based at the study's Coordination Centre in Nairobi, Kenya. The trial is being conducted by the Leishmania East Africa Platform  based on clinical evaluation, with parasitology done only if clinically indicated according to a standardised clinical assessment.\u2022 Adverse events and serious adverse events occurring in the three study arms up to day 60.\u2022 Description of the pharmacodynamic properties of all the three arms.\u00ae.\u2022 Description of the pharmacokinetic properties of Miltefosine alone and in combination with Liposomal amphotericin BRandomization sequences were generated, stratified by site in pre-determined block sizes by the Data Centre. Randomization codes have been concealed from investigators at the trial sites using sealed sequentially numbered, opaque envelopes. A copy of the randomization list is securely stored at the Data Centre. The envelopes are numbered sequentially externally. Inside each one there is a randomization sheet with the same sequential number as well as the treatment allocation. These randomization sheets are filed in the source documents which the monitor verifies at each site visit to confirm that only one envelope was opened for each patient to ensure integrity of the randomization process.\u00ae is being given as a single dose on day 1 at a dose of 10 mg/kg body weight infused in 5% dextrose running for 1-2 hours.Liposomal amphotericin BMiltefosine is being given orally at a dose of 2.5 mg/kg body weight daily, up to a maximum of 150 mg, for 28 days when used alone.\u00ae 10 mg/kg.Miltefosine is being given orally at a dose of 2.5 mg/kg body weight daily, up to a maximum of 150 mg, for 10 days starting on day 2 following a single dose of Liposomal amphotericin B\u00ae.SSG is being given IV/IM at a once daily dose of 20 mg/kg body weight daily for 10 days starting on day 2 following a single dose of Liposomal amphotericin BTherefore the dose regimens is as follows\u00ae one dose of 10 mg/kg body weight (IV) on day 1 followed by 10 days of SSG at 20 mg/kg body weight (IV/IM) from days 2-11.\u2022 Liposomal amphotericin B\u00ae one dose of 10 mg/kg body weight (IV) on day 1 followed by 10 days of miltefosine at 2.5 mg/kg body weight  from days 2-11.\u2022 Liposomal amphotericin B\u2022 Monotherapy course of miltefosine at 2.5 mg/kg body weight  from days 1-28.All failure with compliance will be documented on the trial medication pages of the CRF.Patients with clinical signs of VL  and diagnosis confirmed by visualization of parasites in tissue samples on microscopy, aged between 7 and 60 years inclusive, signed written informed consent, negative HIV status.3/mm3, platelets <40,000/mm3, abnormal liver function tests (ALT and AST) of more than three times the upper limit of the normal range, serum creatinine outside the normal range for age and gender, major surgical intervention within two weeks prior to enrolment. These tests are standardised but it is not feasible to standardise routine clinical laboratory assessments as different machines and reagents are being used. However both internal and external QC will be carried out at both sites on a regular basis and training offered to site staff at the beginning and during the trial to ensure that the data collected are reliable and comparable.Patients who have received any anti-leishmanial drugs in the last 6 months/relapse cases, negative lymph node/bone marrow or spleen smears, severe protein and or caloric malnutrition, previous history of hypersensitivity reaction to SSG or Amphotericin B, suffering from concomitant severe infection such as TB or other serious underlying disease which would preclude evaluation of patients response to study medication, suffering from other conditions associated with splenomegaly such as schistosomiasis, previous history of cardiac arrhythmia or an abnormal ECG, female of child bearing age (pregnant or lactating), haemoglobin <5 mg/dL, WBC <1 \u00d7 10All patients are offered counselling and screening for HIV (voluntary counselling and testing programme (VCT). This is done at the same time as consent is obtained for inclusion in the trial. Patients who decline VCT or are found to be HIV positive are not eligible to participate in this trial but receive appropriate treatment, according to national treatment guidelines. Additionally, they are referred onwards for treatment, surveillance and follow up according to the national protocol for HIV positive patients.Standardised consent forms, adapted to the local context and translated into local languages and approved by ethics committees are being used. Signatures or thumbprints are obtained for consent, with witnesses in the latter case (illiterate subjects) for adults. In the case of children, the investigator gets their assent once permission of either the child's parent or guardian has been obtained.Patients who do not meet inclusion criteria or who do not give consent are offered free treatment outside the trial, according to national treatment guidelines.0), and the lowest efficacy considered adequate as 90% (denoted pa). The null and alternative hypotheses are H0: p \u2264 p0 and H1: p >pa respectively.The study is designed and will be analyzed according to group-sequential methods, specifically the triangular test -12. FollThe triangular stopping boundary, as shown in Figure V  and Z (number of observed minus expected treatment successes) and plotting them on Figure Each analysis consists of calculating the quantities n = 15 (indicating adequate efficacy) that arm will be continued regardless until n = 30. However, if the lower boundary is crossed (indicating inadequate efficacy) at n = 15, then it will be stopped.The above procedure is subject to a constraint imposed for the PK component of the trial, which requires at least 30 patients in arms 2 and 3, subject to efficacy not being inadequate. Therefore, if the upper boundary is crossed at C* will be calculated as V* is the terminal value of V from the data sample, and the \u03b8a parameter is the log-odds ratio \u03b8 = loge(p(1- p0)/p0(1- p)) for p = pa. Then, the tables in Whitehead's Appendix A [p. The latter are provided in terms of factors by which to multiply \u03b8a. Finally, these three values of \u03b8 (i.e. point estimate and upper and lower confidence limits) will be transformed to corresponding sample values of p by p = (p0e\u03b8)/(1+ p0(e\u03b8-1)). Analysis will be done using STATA [Crossing a boundary of the region implies that the percentage of patients cured is either greater than 90% (if the upper boundary is crossed) or no more than 75% (if the lower boundary is crossed). The maximum likelihood estimate of the efficacy, i.e. the number cured divided by the number of patients, is a biased estimator due to the sequential nature of the trial. To take this into account, the analysis will follow Bellissant et al . First, pendix A  will be ng STATA .The sequential analysis described above relates only to efficacy on day 28, not day 210. However, because the latter is likely to be highly correlated with the former, ignoring the sequential design in the latter analysis could give inconsistent results. For example, if the status of all patients is the same at day 210 as at day 28, then using different methods for analysis would give different efficacy estimates from the same data. This is because the day 28 estimate takes into account the sequential design which, in general, would differ from conventional analysis done on the same data at day 210.p210) will be considered as the sum of two probabilities, as follows:Therefore, the day 210 efficacy will be estimated by considering it in terms of events over two consecutive time periods: up to day 28, and from day 28 to day 210. More specifically, the day 210 efficacy (i.e. percentage cured or i) probability of being cured at day 28 and remaining cured at day 210.ii) probability of not being cured at day 28 but becoming cured at day 210.p210:Each of these probabilities is the product of two terms and enables a point estimate of where:r is the percentage of those people cured at day 28 who remain cured at day 210.s is the percentage of those people not cured at day 28 who become cured at day 210.p210. Using the 'delta method' [p28, r and s; these three quantities being statistically independent. The sampling variance of p28 will be estimated as the square of its standard error, which in turn will be estimated as the width of its 95% confidence interval divided by 2 \u00d7 1.96, using a normal approximation. The sampling variances of r and s will be estimated by considering them as standard binomial proportions.The confidence interval will be estimated via the sampling variance of  method' , this cap28 and p210 will be equal (because r = 1 and s = 0). The confidence intervals will, however, generally be unequal. This is because treating the progress between day 28 and day 210 as a separate variable implies additional sampling variation. For example, r has sampling variation even when equal to 1. Moreover, sampling variation is considered on the probability scale, not the log-odds scale as for p28. This also means that the calculated confidence interval could exceed the interval 0-1. In this case, it will be truncated at the limits of that interval.The above procedure ensures that, for example, if no patients change cure status between day 28 and day 210 then the point estimates of When the analysis results in an arm being stopped, the final cure rate (day 210) will also be evaluated . If the point estimate of the day 210 cure rate is found to be <90%, then the arm (arm 1 & 2) will not be considered for further study (e.g phase 3).Within each arm, the percentage cured at each time point will be presented by site. Chi-squared or Fisher's exact tests will be used to test for differences between sites in terms of primary and secondary efficacy endpoints at the 5% level of significance.Assessments are timed at days 0, 3, 7, 14, 21, 28, 60 and 210  and treatment failure at final cure if it occurs on or prior to final cure (day 210).Rescue treatment to be given includes:- Liposomal amphotericin B 30 mg/kg IV split into multiple doses (according to country protocol: Sudan - 3 mg/kg/day for 10 days)- SSG 20 mg/kg IM for 30-60+ days: for patients not responding to initial rescue treatment or for patients requiring treatment for severe PKDL.A complication of Visceral Leishmaniasis, particularly in Sudan is post-Kala-azar Dermal Leishmaniasis (PKDL). PKDL is characterised by skin lesions which normally occur in the months following treatment in people who have recovered from VL. Patients are being monitored closely for PKDL at days 0, 28, 60 and 210. Diagnosis will be made clinically, based on the typical appearance and distribution of the rash.Concomitant medication necessary for the health of the patient is permitted during the course of the study. Details of all concomitant medication taken during the study are recorded in the CRF with indication, daily dose, route and dates of administration.The effects of VL disease and geographical differences on the pharmacokinetics (PK) of miltefosine remain largely unknown, with most available PK data coming from a relatively healthy European patient group with cutaneous leishmaniasis (CL) . A thoroBioanalysis of miltefosine will be done on plasma samples with a volume of 50 to 250 \u03bcL taken during both treatment and follow-up  marker [Leishmania parasites, hence the decreased duration of treatment. In this trial the blood parasite counts will be used as a PD marker of the parasite clearance rate and thus response to treatment.To assess treatment response during and after treatment, this study will apply repeated measurements of blood parasite loads using a real time reverse transcriptase polymerase chain reaction (qPCR) for ) marker ,20. DiffIn the arms receiving miltefosine, the outcome and parasite clearance measured by qPCR will be linked to miltefosine PK. Modelling of miltefosine PK/PD in our patients will enable us to establish variability in pharmacokinetics in relation to outcome, which is an essential component in the development of new treatment regimens.All trial site staff; physicians, nurses, laboratory technicians and pharmacists received training on the study protocol, study specific procedures and International conference on harmonisation-good clinical practise (ICH-GCP) guidelines ,22. AddiSuitably qualified Clinical Monitors trained in GCP regularly visit trial sites to monitor all aspects of the trial including; informed consent procedures, drug accountability, source data verification, adverse event reporting, sample handling, analysis and secure data storage.\u00a9 database software and emailed to trial sites, copied to clinical monitors. The sites will print and make resolutions. At the next monitoring visit, monitors will verify resolutions and deliver to the Data Centre. Data corrections will be programmed in Stata to complete the data cleaning audit trail. At analysis, any unusual values detected would be verified with the investigator at the sites before final analysis is done to confirm correctness/completeness. This will also be captured in the data management report prior to database lock.Data are to be recorded on 3-part no carbon required (NCR) case report forms (CRFs) by site investigators, transcribed from hospital source data. Unique patient identifiers, assigned at the time of randomisation, are linked to unique hospital record numbers. During monitoring visits, CRF data are cross-checked against hospital source data as much as possible by the monitor. The top sheet for each page of the CRF is brought to the central Data Centre for double-entry into GCP compliant open-access database software OpenClinica, version 2.7  with ranDNDi, as sponsor, will render all necessary assistance to investigators to ensure timely publication of results in an international peer-reviewed, for the benefit of patients and to inform national decision-making on VL treatment guidelines. Ancillary studies will acknowledge those involved by name where appropriate.Trial site records will contain names and residential information for each patient to allow follow-up to take place. Only the unique numeric identifier assigned to patients will be extracted from patient records and transferred to the Data Centre. Patient data will be kept securely at trial sites under the responsibility of the site investigator.During the course of the trial, site audits will be undertaken to assess compliance with ICH-GCP guidelines. Specific issues to be assessed include adherence to the protocol and standard operational procedures (SOPs), consent, laboratory practise, documentation and record keeping. All areas of non-compliance will be addressed by the site investigators or trial coordination centre depending on the findings.On approaching the end of planned recruitment, the trial coordination office will send written instructions by email to each site regarding the date to cease recruitment. All patients will be followed up as per the protocol, with the data being collected and cleaned. Once the data lock has been completed, site close out visits will be performed by the coordination team and clinical monitors. A decision for premature termination will be taken in consultation and agreement with the sponsor, investigators and the DSMB. All relevant ethics committees and regulatory authorities will also be informed of the reason for termination. Trial master files and completed CRFs will be archived by the coordination centre in Nairobi for 15 years.After recruiting 15 patients per arm, a brief interim analysis report presenting the efficacy, including the triangular analysis will be provided to the Data Safety Monitoring Board (DSMB) whose composition is based on WHO-TDR guidelines . The DSMAt the end of the trial, the clinical study report will be circulated to Principal Investigators, DSMB, Ethics Committees and Ministries of Health.Ethical approval has been obtained from National and local Ethics Committees in Kenya and Sudan prior to the start of the trial in each Country. Ethical approval has also been granted by LSHTM Ethics Committee, the AMC Medical Ethics Committee issued a 'declaration of no objection'.The DNDi Coordination Team, based mainly at the Coordination Centre, DNDi Africa, Kenya Medical Research Institute, Nairobi, are responsible for collation and submission of protocol amendments, organisation of training for trial staff, monitoring and supervision of trial conduct, day to day management of the trial, harvesting data collected at each site, trial monitoring visits and data management, all to GCP standards.Due to limitations of current treatments, and the risk of drug resistance developing, there is an urgent need to develop short course combination treatments against VL in the East Africa region. The aim of this trial is to evaluate potential combinations that can then be evaluated in a large multi centre phase-III clinical trial in the region. The current trial also aims to collect additional data to assist in the registration of miltefosine in the region. For this reason, three separate arms with no between-arm comparisons were chosen and a PK/PD component focusing on the miltefosine treatments included. Due to potential geographical differences in drug response within the region, the trial aims to evaluate treatment in two sites: one from the northern part (Sudan) and one from the Southern part (Kenya) of the East African foci . To makeLEAP: Leishmaniasis East Africa Platform; IM: Intramuscular; IV: Intravenous; PK: Pharmacokinetics; PD: Pharmacodynamics; VL: Visceral Leishmaniasis; RCT: Randomised controlled trial; SSG: Sodium stibogluconate; PKDL: Post kala-azar dermal leishmaniasis; DSMB: Data safety monitoring board; CRF: Case report form; ICH: International Conference on Harmonisation; GCP: Good Clinical Practise; SOP: Standard Operating Procedure; NCR: No Carbon Required; VCT: Voluntary Counselling and Testing.DNDi, as sponsor, is funding the trial, costs relating to maintenance of the LEAP platform and open-access publication of LEAP Study Group manuscripts. MW, RO, SE and MB are current employees of DNDi.All authors have read and approved the final manuscript.LEAP Study Group Investigators , statisticians from LSHTM  and representatives from the sponsor, DNDi,  designed the study. RO, NA and TE drafted this submission.All other LEAP study group authors played an important role in finalising the study protocol."}
+{"text": "Several studies have demonstrated the efficacy of artemisinin-combination therapy (ACT) across malaria zones of the world. Fixed dose ACT with shorter courses and fewer tablets may be key determinants to ease of administration and compliance.Plasmodium falciparum malaria were recruited in Ibadan, south-western Nigeria. A total of 250 children each were randomly assigned to receive three doses of artesunate/sulphamethoxypyrazine/pyrimethamine (AS + SMP) (12 hourly doses over 24 hours) or three doses of artesunate/amodiaquine (AS + AQ) (daily doses over 48 hours). Efficacy and safety of the two drugs were assessed using a 28-day follow-up and the primary outcome was PCR- corrected parasitological cure rate and clinical response.Children aged one year to 13 years presenting with uncomplicated There were two (0.4%) early treatment failures, one in each treatment arm. The PCR corrected cure rates for day 28 was 97.9% in the AS + AQ arm and 95.6% in the AS + SMP arm (p = 0.15). The re-infection rate was 1.7% in the AS + AQ arm and 5.7% in the AS + SMP arm (p = 0.021). The fever clearance time was similar in the two treatment groups: 1 - 2 days for both AS + SMP and AS + AQ (p = 0.271). The parasite clearance time was also similar in the two treatment groups with 1 - 7 days for AS + SMP and 1 - 4 days for AS + AQ (p = 0.941). The proportion of children with gametocytes over the follow-up period was similar in both treatment groups. Serious Adverse Events were not reported in any of the patients and in all children, laboratory values  remained within normal levels during the follow-up period but the packed cell volume was significantly lower in the AS + SMP group.Plasmodium falciparum malaria in children in Nigeria. Both drugs also proved to be safe. Therefore, AS + SMP could be an alternative to currently recommended first-line ACT with continuous resistance surveillance.This study demonstrates that AS + SMP FDC given as three doses over 24 hours  has similar efficacy as AS + AQ FDC given as three doses over 48 hours  for the treatment of uncomplicated  In Nigeria, nearly 110 million of clinical cases of malaria are diagnosed per year, translating into about 50% of the adult population experiencing at least one malaria episode per year, while young children can have up to two to four attacks of malaria annually [National drug efficacy trials conducted in 2002 in Nigeria demonstrated that the first line treatments then employed, chloroquine and sulphadoxine/pyrimethamine (SP) were no longer adequate and in 2005, the highly efficacious artemisinin-based combination therapy (ACT) was introduced in the country . ArtemetAlthough the use of ACT for the treatment of uncomplicated malaria has been introduced several years ago, its utilization in the field is still below expected levels. The reasons explaining this situation include the poor availability and/or relatively high cost of ACT on the African market. One of the key elements of the RBM strategy is that malaria patients should have access to appropriate and adequate treatment within 24 hours of the onset of symptoms ,2. An anIn view of the ideal profile to be found in an anti-malarial drug, the combination of artesunate and sulphamethoxypyrazine/pyrimethamine (AS + SMP) represents an interesting treatment option in case first-line drugs are not readily available. In fact, this product is easy to use, safe, and has previously demonstrated high efficacy in several endemic areas whether it is taken under a 24- or 48- hour regimen -5. PreviIn order to evaluate the safety and efficacy of the 24-hour therapy of AS + SMP FDC in south-west Nigeria, a cohort of children suffering from uncomplicated malaria were treated either with this ACT, or with a 48-hour therapy of AS + AQ FDC.Plasmodium falciparum, is endemic in the region with a six-month transmission season (May-October) reaching its peak in August [Anopheles gambiae s.l. range from 18 to 145 infective bites per person per year [This study was conducted at the University College Hospital and at the Oni Memorial Children's Hospital both located in the urban areas of Ibadan, Oyo State, Nigeria. The intensity of malaria transmission varies considerably throughout Nigeria. Ibadan is located in the forest savannah woodland zone with average rainfall of 975 - 1474 mm/year . Malarian August . The oveper year . Specifiper year .This study was approved by the Joint Ethics Committee of University of Ibadan and University College Hospital, Ibadan, Nigeria. The study was conducted in accordance with all requirements of the ICH Guidelines for Good Clinical Practice as well as the requirements of the Declaration of Helsinki. Clinical trial approval was obtained from the National Agency for Food and Drug Administration and Control in Nigeria (NAFDAC). Written informed consent was obtained from all parents or guardians of eligible children prior to enrolment.P. falciparum, with parasitaemia in the range of 2,000-200,000 asexual parasites per microlitre of blood; and no general danger signs or signs of severe and complicated falciparum malaria as per WHO guidelines [Children presenting to each of the two study sites were screened for eligibility and invited to participate in the study if they met the following inclusion criteria: aged between 1 and 13 years; body weight between 6 and 40 kg; history of fever in the previous 24 hours or measured fever (axillary temperature >37.5\u00b0C); mono-infection with idelines .A randomized, controlled, open-label trial design was used. Assuming a proportion of 94% patients cured (with PCR-corrected cure rate at day 28 as primary endpoint) with AS + SMP, an equal proportion of patients cured with AS + AQ, a sample of 250 patients (taking into account 10-15% loss to follow-up) was required in each treatment arm to detect the 6% difference in parasitological cure rates with a power of 80% (\u03b1 = 0.025 one-sided). Sample size calculations were done using nQuery Advisor 5.0. The randomization code was computer-generated with stratification per treatment centre, from which treatment groups were assigned.At enrolment, a physical examination was performed, weight and axillary temperatures recorded and a medical history was obtained from parents/guardians including presenting symptoms and current medication. A finger prick blood sample was obtained for thin and thick blood smears and blotted on filter paper for parasite genotyping. A blood sample was also obtained for assessment of haematological and biochemical parameters.\u00ae, Dafra Pharma ltd., Kenya), crushed mixed with clean water and administered at 0, 24 and 48 hours. The second group received artesunate/sulphamethoxypyrazine/pyrimethamine 100/250/12.5 mg tablets , crushed, mixed with water and administered at 0, 12 and 24 hours. Treatment doses were given based on the age of the patient: children under 7 years received 1/2 tablet per dose (50/153.1 mg As/AQ and 50/125/6.25 mg As/SM/P) while children aged 7 years and older received a full tablet per dose. They were then observed for 1 hour after drug administration for vomiting. If vomiting occurred within 30 min after administration, the full treatment dose was re-administered. If vomiting occurred between 30-60 min after administration, half the treatment dose was re-administered.Eligible children were then assigned into one of the two treatment groups according to the randomization code. The treatments were given in the clinic by the recruiting physician. One group received three doses of artesunate/amodiaquine 100/206.2 mg tablets  was done on each day of follow-up , or on any other day if the child was feeling sick. Patients were called back on these days or visited at home. During each visit, a brief clinical history was obtained to assess new complaints and possible side effects and a physical examination was performed. The haemogram and biochemical analysis  were done on days 0, 7 and 14 to detect possible significant abnormalities. Filter paper blood samples for parasite genotyping were obtained on day 28 or earlier if presented with repeat of the symptoms. Patients were excluded from the study if they withdrew consent, left the study area or reported taking anti-malarial medication during the follow-up period.Response to drug treatment was assessed using modified WHO clinical classification system: all patients were not febrile at presentation, hence a temperature < 37.5\u00b0C was not an exclusion criterion and patients were followed up for 28 days in this area of intense transmission. The clinical classification system consisted of adequate clinical and parasitological response (ACPR), late parasitological failure (LPF), late clinical failure (LCF) and early treatment failure (ETF). The cure rates on day 28 were adjusted on the basis of the PCR genotyping results of paired samples for patients with recurrent parasitaemia after day 14 of starting treatment.Plasmodium parasite species. All qualitative discordant slides and slides with a parasite density difference \u226550% were read by a third microscopist. For quality control 10% of randomly selected slides were reread by an independent microscopist, not involved in the study.A finger prick blood sample was taken to prepare thick and thin blood smears on days 0, 1, 2, 3, 7, 14, 21, 28 and on any other (unscheduled) visit. The slides were air dried, stained with 10% Giemsa for 20 min and read independently by two technicians. Parasite density (the number of parasites and gametocytes per \u03bcl) was calculated by counting parasites against 200 leukocytes and assuming a leukocyte count of 6,000/\u03bcl of blood. A slide was considered negative after reading at least 200 power fields. Thin blood smears were specifically used to identify P. falciparum infections at enrollment, and during follow-up if the patients were parasitaemic on or before D28 were determined using PCR techniques. Analysis of genetic polymorphisms was performed on paired primary and post-treatment parasites samples obtained from the two treatment groups. Paired primary and post-treatment parasites were analysed using parasite loci that exhibit repeated numbers of polymorphisms to distinguish true treatment failures from new infections. Briefly, block 2 of MSP-1 (merozoite surface proteins-1), and the Block 3 of MSP-2 (merozoite surface protein-2) and region II of GLURP were amplified by two rounds of PCR using primers and amplification conditions [Filter paper blood spots were collected on days 0, 28 or any other day of recurrent parasitaemia. Genotypes of the parasite population in each sample collected from patients with microscopically confirmed nditions ,13. Ten nditions . PatientThe case report forms were double checked to ensure complete and accurate data collection. Dual data entry was done by two clerks after which data was compared, corrected and validated by the data manager. Data analysis was done using Epi Info version 6 and SPSS version 11. The primary analysis was a non-inferiority analysis  based on PCR-corrected adequate clinical and parasitological response (ACPR) on day 28 as primary efficacy endpoint . SecondaOut of 3,500 children screened for malaria, 500 were enrolled into the trial; 250 were assigned to the AS + AQ treatment arm and 250 to the AS + SMP arm Figure . Thirty There were two early treatment failures, one in each treatment arm (0.4%). The PCR corrected cure rates for day 28 was 97.9% in the AS + AQ arm and 95.6% in the AS + SMP arm p = 0.15). The re-infection rate was 1.7% in the AS + AQ arm and 5.7% in the AS + SMP arm (Table 5. The reThe median fever clearance time was similar in the two treatment groups: 1 day for both AS + SMP and AS + AQ p = 0.271). The median parasite clearance time was also similar in the two treatment groups with a range of 1 - 7 days for AS + SMP and 1 - 3 days for AS + AQ (p = 0.941). The proportions of children with gametocytes over the follow-up period were similar in both treatment groups. Sixteen children in each treatment arm had gametocytes on day 0 (7.0% for AS + SMP and 6.7% for AS + AQ) (Table 71. The mSerious adverse events were not reported in any of the patients. There was no significant difference in the proportions of patients reporting adverse events in the two treatment groups. Five patients (2.2%) treated with AS + AQ reported one or more of the following adverse events: vomiting (n = 1), excessive sleepiness (n = 2), abdominal pain (n = 1), and weakness (n = 3). Three patients (1.3%) treated with AS + SMP reported vomiting, one of these patients also reported nausea as adverse event. None of these patients needed to be admitted as a result of these events. For all children, laboratory values  remained within normal levels during the follow-up period Table .P. falciparum malaria in children in an endemic area of south-western Nigeria. The study was based on a 28-day follow-up period.This study is documenting the efficacy and tolerability of AS + SMP compared to the recommended artesunate-amodiaquine (AS + AQ) ACT for the treatment of uncomplicated Many African countries have adopted artemisinin based combination therapies (ACTs) as first-line therapy against uncomplicated malaria, particularly artemether-lumefantrine (AL) and AS + AQ. However based on field experience, there is a need to assess other forms of ACT in order to identify a suitable alternative to recommended treatments in case those are not readily available to the patients who urgently need to be treated. The 28-day cure rates observed in this study were 95.6% in the AS + SMP treatment arm compared to 97.9% in the AS + AQ treatment arm (p = 0.151) suggesting that the therapeutic efficacy of both drugs were similar. These findings are in agreement with previously reported studies on AS + SMP from other endemic and non-endemic malaria areas -7. The oet al reported that, despite slower clearance of sexual parasitaemia in children treated with AQ + SMP compared with those treated with AL, treatment with the former was not associated with increased gametocyte carriage [Gametocyte carriage before and following treatment were similar in the two treatment arms, which is expected to be the result of the action of artesunate . Interescarriage .et al in 1989 [Both AS + SMP and AS + AQ treatments were well tolerated and the frequency of reported adverse events was similar in the two treatment arms. In reality, the reported adverse events were difficult to distinguish from signs and symptoms of malaria. It is notable that the reported frequency of those with itching treated with AS + AQ was low, similar to what was previously reported by Sowunmi  in 1989 . Serious in 1989 . This woPlasmodium falciparum malaria in children in Nigeria. Both drugs also proved to be safe though AS + SMP has a higher re-infection rate. AS + SMP represents a good alternative to recommended first line treatments in areas where these are not available, or only offered at high costs. A continuous surveillance on the development of drug resistance is however essential as a result of potential cross resistance between SP and SMP.In summary, this study demonstrates that AS + SMP FDC given as three doses over 24 hours  has similar efficacy as AS + AQ FDC given as three doses over 48 hours  in terms of fever and parasite clearance for the treatment of uncomplicated FHJ is an employee of the drug company Dafra Pharma. Dafra Pharma supported the study. Dafra Pharma developed and supplied the fixed dose medication artesunate/sulfamethoxypyrazine/pyrimethamine. The authors and the sponsor were involved in study design, trial monitoring and interpretation of the date and writing of the report. No honoraria were paid for running the trial to any of the co-authors.IAA contributed to the design of the study, project implementation  and wrote the manuscript. AGF and AS contributed to the design of the study, the performance of the research, and participated in the writing of the manuscript. FHJ contributed in the design of the study and the writing of the manuscript. All authors have read and approved the final manuscript.Kaplan Meier curve for rates of re-infection in the treatment arms AS+SMP and AS+AQ.Click here for file"}
+{"text": "Ric Price and colleagues pool individual patient data from efficacy trials of dihydroartemisinin-piperaquine shared with WWARN  to examine the potential for underdosing in young children.Please see later in the article for the Editors' Summary Dihydroartemisinin-piperaquine (DP) is increasingly recommended for antimalarial treatment in many endemic countries; however, concerns have been raised over its potential under dosing in young children. We investigated the influence of different dosing schedules on DP's clinical efficacy.n\u200a=\u200a7,072 patients) were included in the analysis. After correcting for reinfection by parasite genotyping, Kaplan\u2013Meier survival estimates were 97.7% (95% CI 97.3%\u201398.1%) at day 42 and 97.2% (95% CI 96.7%\u201397.7%) at day 63. Overall 28.6%  of children aged 1 to 5 years received a total dose of piperaquine below 48 mg/kg (the lower limit recommended by WHO); this risk was 2.3\u20132.9-fold greater compared to that in the other age groups and was associated with reduced efficacy at day 63 . After adjusting for confounding factors, the mg/kg dose of piperaquine was found to be a significant predictor for recrudescence, the risk increasing by 13% (95% CI 5.0%\u201321%) for every 5 mg/kg decrease in dose; p\u200a=\u200a0.002. In a multivariable model increasing the target minimum total dose of piperaquine in children aged 1 to 5 years old from 48 mg/kg to 59 mg/kg would halve the risk of treatment failure and cure at least 95% of patients; such an increment was not associated with gastrointestinal toxicity in the ten studies in which this could be assessed.A systematic search of the literature was conducted to identify all studies published between 1960 and February 2013, in which patients were enrolled and treated with DP. Principal investigators were approached and invited to share individual patient data with the WorldWide Antimalarial Resistance Network (WWARN). Data were pooled using a standardised methodology. Univariable and multivariable risk factors for parasite recrudescence were identified using a Cox's regression model with shared frailty across the study sites. Twenty-four published and two unpublished studies  now recommends artemisinin-based combination therapy (ACT) for first-line treatment of P. falciparum malaria wherever malaria is endemic . In ACT, artemisinin derivatives  are used in combination with a slower acting, more slowly eliminated partner drug that prevents recrudescent infections .Half of the world's population is at risk of malaria, a mosquito-borne parasitic disease that kills a million people (mainly young children in sub-Saharan Africa) every year. During the second half of the 20th century, the main treatments for malaria were \u201cmonotherapies\u201d such as chloroquine and sulfadoxine-pyrimethamine. Unfortunately, parasitic resistance to these drugs rapidly spread and, in the 1990s, there was an upsurge in the illness and death caused by P. falciparum becoming resistant to either the artemisinin derivative or the partner drug, but sub-optimal dosing can result in incomplete elimination of the initial parasitic infection and/or recrudescence. Both these situations drive the selection of parasites with reduced drug susceptibility. Unfortunately, current dosing strategies are usually based on weight or age bands that were determined during early drug development. Inevitably, some patients at the margins of these bands receive inappropriate doses\u2014either too high or too low. Moreover, the doses recommended for children are extrapolated from adult doses and may not be optimal because children are not merely small adults\u2014their drug responses often differ from those of adults. Here, researchers from the WorldWide Antimalarial Resistance Network (WWARN) investigate the influence of different dosing schedules on the clinical efficacy of dihydroartemisinin-piperaquine  by undertaking a pooled analysis of individual patient data collected during ACT clinical studies.ACT reduces the chances of The researchers identified all the studies published between 1960 and February 2013 in which patients were treated with DP and obtained individual patient data for almost 70% of study participants  from the principal investigators. These data were pooled and statistical models were used to identify risk factors for parasite recrudescence (parasite genotyping was used to correct for re-infection with a new parasite). Overall, DP treatment was successful  in 97.7% and 97.2% of patients at day 42 and 63, respectively, after treatment. However, 28.6% of children aged 1\u20135 years received a total dose of piperaquine over the 3-day DP dosing schedule of below 48 mg/kg body weight (the lower limit of dosing recommended by WHO). Children aged 1\u20135 years had a 2.9-fold higher risk of receiving a dose of piperaquine below 48 mg/kg than children aged 5\u201312 years. Moreover, the piperaquine dose was a significant predictor of recrudescence. For every 5 mg/kg decrease in dose, the risk of recrudescence increased by 13%. Finally, the researchers estimated that increasing the target dose of piperaquine in children aged 1\u20135 years to 59 mg/kg would halve the risk of treatment failure and cure at least 95% of young children.These findings suggest that DP treatment of malaria has a good efficacy in a wide range of settings but indicate that young children are at a higher risk of treatment failure than the overall population and that treatment failure, particularly in young children, is associated with a lower dose of piperaquine. Although the accuracy of these findings may be limited by some aspects of this study , they nevertheless suggest that further detailed dose optimization studies in young children are essential to prolong the useful therapeutic life of DP. Moreover, if global elimination of malaria is to be achieved, similar pooled analyses to assess the efficacy of other drug combinations should be undertaken to ensure that ACT remains therapeutically useful for as long as possible.http://dx.doi.org/10.1371/journal.pmed.1001564.Please access these websites via the online version of this summary at PLoS MedicinePerspective by Paul GarnerThis study is further discussed in a malaria ; the 2012 World Malaria Report provides details of the current global malaria situation; the 2010 WHO Guidelines for the Treatment of Malaria are availableInformation is available from the World Health Organization on malaria (in English and Spanish), including a selection of personal stories about malariaThe US Centers for Disease Control and Prevention provide information on global control of malaria including fact sheets about ACTInformation is available from the Roll Back Malaria Partnership on the malaria (in English and Spanish)MedlinePlus provides links to additional information on WorldWide Antimalarial Resistance Network is availableInformation on the  Children under the age of 5 years are particularly vulnerable to failing treatment and developing severe disease, usually attributed to their lower immunity and premunition compared to older patients Plasmodium falciparum and P. vivax infections falciparum malaria The combination of dihydroartemisinin-piperaquine (DP) has been assessed in clinical trials for almost a decade, and shown to be highly efficacious against both The Worldwide Antimalarial Resistance Network (WWARN) brings together malaria researchers from across the world. This collaborative resource provides a unique opportunity to conduct a series of individual patient data meta-analyses in which the dosing strategies of the recommended ACTs can be reviewed, to define the spectrum of treatment doses actually administered, and the degree to which these dose variations impact on the therapeutic efficacy P. falciparum , for a minimum of 28 days. Studies were included only if information was available on the treatment dose administered, the age and weight of the patient, and if genotyping was performed to distinguish between new infections and recrudescent infections. Both randomized and non-randomized studies were included. Data were anonymised, uploaded to the WWARN repository, and standardised using a methodology described in the WWARN Data Management and Statistical Analysis Plan .Where possible, the dose of DHA and PIP administered was calculated from the individual number of tablets administered to each patient daily. Where such data were not available, back-calculations were made, based on the dosing strategy presented in the study protocol, assuming correct adherence to the protocol. Only patients completing a full 3-day treatment regimen and included in the original analysis, were included in the meta-analysis.Study sites were categorised into three strata according to known epidemiology: low, moderate, and high transmission settings. The classification of transmission intensity was based on the author's classification of the site as reported in the study publication. Where no transmission information was reported, then the transmission intensity was defined based on the triangulation of information available from: (i) the study protocol(s), (ii) observed reinfection rate, and (iii) transmission estimates obtained from the Malaria Atlas Project a priori statistical plan P. falciparum recrudescence at the end of study follow-up. Secondary endpoints included the new infections of P. falciparum, parasitological clearance rates, and gametocyte carriage. The incidence risk of these endpoints at day 28, day 42, and day 63 was computed using survival analysis (Kaplan\u2013Meier [K-M] estimates). Definitions of outcome status and censoring are detailed in the WWARN Clinical Module DMSAP v1.2 All statistical analyses were carried out using R , on the basis of an 1)\u00d7(1\u2212PAR2)\u00d7\u2026\u00d7(1\u2212PARn)]. In the final multivariable model, the predicted effect of increasing the mg/kg dose of PIP was calculated for every five unit increase, starting from 30 mg/kg. The 95th percentile of predicted risk was computed for each mg/kg dosage of PIP using each patient individual covariates evaluated at average random effect. Although the primary analysis was focussed on risk factors affecting the efficacy of the DP, the relationship between drug dose and gastrointestinal side effects (vomiting and diarrhoea) was also explored using logistic regression with random effects fitted for the individual study and study sites.The population attributable risks (PARs) for treatment failure were calculated based on the prevalence of the risk factor in the population and its associated relative risk (adjusted hazard ratio) n\u200a=\u200a375) did not meet the inclusion criteria and 634 (8.2%) patients from the rest of the studies were excluded for protocol violations. In total 7,072 patients from 26 studies representing 70% of the targeted published literature on this treatment regimen, were included in the final analysis (n\u200a=\u200a471), age category in one study (n\u200a=\u200a124), and on weight bands in the remaining 23 studies . Six studies  followed up patients for 28 days, 12 studies  for 42 days, one study (n\u200a=\u200a58) for 56 days, and seven studies  for 63 or more days. Three different combinations of DHA and PIP were used in the different studies; 51.4%  of patients were treated with Artekin or Duo-Cotecxin, 47.8%  with Eurartesim (Sigma Tau Industrie Farmaceutiche Riunite), and 0.8% (n\u200a=\u200a55) of the patients were treated with Artecan . Drug intake was reported in all studies, with full supervision in 24 (92.3%) of the studies, partial supervision in one (3.8%), and a combination of full supervision and no supervision in one study (3.8%). Overall 2,628 (37.2%) patients were recruited in areas of high malaria transmission, 1,194 (16.9%) in areas of moderate transmission, and 3,250 (46%) in low transmission areas. Parasite genotyping was carried out in 25 studies: with 17 studies  using three markers ; three studies (n\u200a=\u200a528) using two markers ; three studies (n\u200a=\u200a578) using MSP1, MSP2, and microsatellites; one study (n\u200a=\u200a116) using only microsatellites; and in one study, the genotyping method was not stated (n\u200a=\u200a41). Genotyping was not carried out in one study (n\u200a=\u200a58) as there were no recurrent infections.In total, 23 principal investigators (PIs) were approached, of whom 16 submitted data from 29 studies contributing 8,081 patients for the pooled analysis. These data were derived from 77.7%  of patients reported in 77.1% (27/35) of the targeted published studies and an additional 183 patients from two unpublished studies . Three oanalysis , of whomanalysis . Dosing p<0.001) and had a higher median baseline parasitemia . In the 256 patients from South America the median age was 23.5 years (IQR: 13\u201339) with a median baseline parasitemia of 6,274.5 \u00b5l\u22121 .The baseline characteristics of patients included in the analysis are documented in p\u200a=\u200a0.002]), children from 5 up to 12 years  and 12 years or older ; p<0.005 for all comparisons. Patients from African sites were at greater risk of being exposed to a total dose of DHA dose below 6 mg/kg, compared to those from Asia , but, this was no longer significant after adjusting for the age and weight of the patient .The median total dose of DHA administered was 6.8 mg/kg  . Overallp<0.001]), children from 5 up to 12 years , and patients 12 years or older . These comparisons remained statistically significant after adjusting for body weight . Patients from African sites were at greater risk of being exposed to a total dose of PIP below 48 mg/kg, compared to those from Asia , however like DHA, this was no longer significant after adjusting for age. There was no significant difference in the risk of under dosing PIP between patients from South America and Asia.The median total dose of PIP administered was 53.3 mg/kg , but this varied significantly between age groups . Overallp\u200a=\u200a0.002]).The overall speed of parasite clearance was rapid , with thp\u200a=\u200a0.022) per unit increase in mg/kg of DHA and 0.97  per unit increase in mg/kg PIP dose. These ORs remained statistically significant after controlling for age, baseline parasitemia, and transmission setting, AOR\u200a=\u200a0.80  and 0.97  per unit increase in mg/kg DHA and PIP, respectively. The dose of DP was not a significant predictor of parasite positivity on day 1 or day 2.The dose of DP was a significant predictor of parasite positivity on day 3, with an OR of 0.81 , nine (26.5%) patients failed before day 28, 16 (47%) patients failed between day 28 and day 42, and nine (26.5%) patients recurred after day 42. Overall PCR-corrected K-M survival estimates were 98.8% (95% CI 98.5\u201399%) at day 28, 97.7% (95% CI 97.3\u201398.1%) at day 42, and 97.2% (95% CI 96.7\u201397.7%) at day 63  patients had recurrent parasitemia detected during follow-up, of whom 136  were confirmed by PCR as true recrudescence. In seven studies with a follow-up duration of 63 days , baseline parasitemia  , presence of gametocytes at baseline , being from 1 up to 5 years of age , admission haemoglobin , and body weight  (p\u200a=\u200a0.002] for every 5 mg/kg increase in PIP dose). In total 3%  of patients exposed to a dose of PIP less than 48 mg/kg experienced a recrudescent infection compared to 1.5%  of those who received a dose greater than 48 mg/kg kg .Six univariate factors on admission were associated with recrudescent parasitemia by day 42: the mg/kg PIP dose  . In a mup\u200a=\u200a0.002) for the risk of recrudescence. The risk remained significant after adjusting for baseline parasitemia, and the mg/kg dose of PIP . The population attributable risks of recrudescence are presented in Overall, the risk of recrudescence was greatest in the 1 up to 5 year age group, rising to 5.6% (95% CI 3.8\u20137.4) by day 63  and 6. Cn\u200a=\u200a3,429, 98 recrudescent failures), five risk factors on admission were associated with recrudescence failure by day 42; age (years) , body weight , enrolment parasitemia  , admission haemoglobin , and mg/kg PIP dose . Since weight and age were collinear only weight was included in the multivariable analysis. In the multivariable model, the dose of PIP remained a significant risk factor . The median total dose of PIP was 48.0 mg/kg (IQR: 42.8\u201353.3 mg/kg) in young children with recrudescence compared to 53.3 mg/kg (IQR: 45.7\u201364.0 mg/kg) in those who were cured (p<0.001). In the multivariable model every 5 unit increase in mg/kg dose of PIP was associated with a 13% (95% CI 3\u201322%) decrease in risk of recrudesecence; p\u200a=\u200a0.013. In a predicted risk model, generated from patients' individual covariates, a dose of 59 mg/kg PIP was sufficient to ensure a day 42 cure rate above 95% (p\u200a=\u200a0.008]), and this accounted for 39.3% (PAR) of all recrudescent infections. By day 42 the risk of recrudescence in patients receiving a PIP dose below this threshold was 5.5% (95% CI 4.2\u20136.7) compared to 2.1% (95%: 1.1\u20133.0) in patients receiving a higher dose, p<0.001 (In children aged from 1 up to 5 years of age (bove 95% . Childre p<0.001 .p<0.001 (p\u200a=\u200a0.076]); however, this was significant after adjusting for age, baseline gametocytemia, and parasitemia . The dose of DHA did not correlate with the risk of gametocyte reappearing.The overall gametocyte carriage decreased from 12.3% at enrolment to 8.3% on day 7 a reduction of 4.0% (95% CI 2.8%\u20135.2%) and fell further thereafter; p<0.001 . ExposurIn total 3.2%  of patients with full documentation of tablet administration did not complete a full course of DP. In four (7.4%) cases this was due to adverse events . Information regarding acute vomiting of medication was available for ten studies with 8.8%  of the patients vomiting within an hour of drug administration, this proportion being greatest in infants and children from 1 up to 5 years of age . Data onn\u200a=\u200a2,873), but in none of the patients in eight other studies . In the remaining 13 studies, severe adverse events were not reported or the data were unavailable. These severe events included one patient with diarrheal disease, considered unrelated to treatment who died, three patients with severe anaemia, three with recurrent vomiting, two with pyomyositis, two with acute pyelonephritis, two with recurrent P. falciparum infections, and one patient each with one of the following: abnormal skin, a swollen arm, anorexia, aspiration pneumonia, cerebritis, convulsions, high fever, and hyperparasitemia P. falciparum infection. Five patients were younger than 1 year, five were from 1 up to 5 years of age, and eleven were older than 5 years. There was no evidence of these SAEs being related with the mg/kg dose of PIP.A total of 21 severe adverse events (SAEs) were reported from five studies  was adopted as first line treatment for malaria reveal a cautionary tale. Early evidence of incipient resistance was present long before SP treatment failure became manifest at high levels. It is highly likely that the sub-optimal dosing in young children hastened the demise of SP as a useful antimalarial Our study has a number of limitations. Although the clinical data used in the analysis constitute almost 70% of the relevant published literature on this treatment regimen, eight studies  and 170 patients from targeted studies were not available. No specific reasons were given by the investigators for being unable to join the study group, but the majority of these were from Asia. Comparison of the more complete dataset from Africa showed no regional differences in our analysis suggesting that a systematic attrition bias was unlikely. Another limitation of the study is that only in 23%  of patients, could drug doses be calculated from the actual number of tablets administered, the total dose in the remainder being extrapolated from the number of tablets predicted to have been administered according to age and weight criteria defined in the study protocol, assuming complete adherence by the attending clinical staff. Although errors in drug administration may have occurred these were generally identified and the patients censored from analysis. Reassuringly, a sensitivity analysis of a subgroup of patients in whom the exact number of tablets was recorded generated identical parameters to the models of the complete dataset. Another limitation arose from DP being a fixed dose combination, making it impossible to determine whether the observed treatment effects were attributable to the dose of PIP, DHA, or a combination of both. The initial reduction in parasite biomass is widely regarded as being determined by the artemisinin derivative because of its significantly higher potency and faster action, whereas the parasite biomass remaining after 3 days of artemisinin exposure is dependent on its elimination by the intrinsically less active partner drug with longer half-life. This is not always the case since several longer acting partner drugs have been shown to contribute to the initial parasite clearance following ACTs Our analysis highlights the power of pooled analyses from diverse clinical settings to assess geospatial and temporal trends in antimalarial efficacy. These observations provide critical information for national and international policymakers. Our study confirms that DP is an important addition to the malaria pharmacopoeia; however although its overall efficacy is high, young children are vulnerable to receiving an inadequate dose of PIP and this is associated with an increased risk of recrudescence, prolonged parasite positivity, and greater gametocyte carriage. Together these constitute a potential threat to the useful therapeutic life of one of our most valuable ACTs and suggest that further dose optimisation studies in young children are warranted, including detailed pharmacokinetic evaluation and safety monitoring to ensure the tolerability of any proposed increase in dose. Preservation of the longest possible therapeutic life for our antimalarial armamentarium must be one of the highest priorities for achieving the global elimination of malaria.Text S1References of all DP clinical trials, their study designs, and dosing schedules.(XLSX)Click here for additional data file.Text S2Maps showing locations of published DP clinical efficacy studies and the studies included in the pooled analysis.(PDF)Click here for additional data file.Text S3WWARN clinical data and management statistical analytical plan.(PDF)Click here for additional data file.Text S4Transmission classification.(XLSX)Click here for additional data file.Text S5WWARN DP data and management statistical analytical plan.(PDF)Click here for additional data file.Text S6Authors and contributions.(XLSX)Click here for additional data file."}
+{"text": "P. falciparum malaria. No ACTs are indicated in infants <4.5 kg. Coartem\u00ae , with an available pediatric formulation, has a large clinical trial and postmarketing safety experience in infants \u22655 kg. Published reports on malaria in younger infants are scanty, leaving a significant knowledge gap about the pattern and outcome of malaria in this sub-population. The aim of the current study was to collect data on the clinical features, treatment and outcomes of uncomplicated P. falciparum malaria in infants <5kg in five countries within Sub-Saharan Africa.WHO recommends artemisinin-based combination therapy (ACT) as first-line therapy for infants >5 kg of body weight (BW) with uncomplicated P. falciparum malaria. Details of clinical features, patient age group (<1 month or >1 month), treatment type received, and clinical outcomes were collected, and a descriptive analysis of data was carried out. A total of 907 cases were identified for inclusion.Hospital charts ranging from 2006-2011 from eight hospitals in B\u00e9nin, Burkina Faso, the Democratic Republic of Congo, Nigeria, and Togo were retrospectively reviewed for inpatient and outpatient neonates and infants weighing <5 kg with parasitologically confirmed diagnosis of uncomplicated The annual disease incidence ranged from 12 to 120 cases across hospitals and calendar years. There was a higher proportion of infants in the younger age group in Burkina Faso, Nigeria, and Togo, but in B\u00e9nin and the DRC the opposite was true. In all countries, the most frequent reason for seeking care was fever. Other reasons included dyspnea, cough or vomiting. Parasite density was generally low (<10% of the infants presented with parasitemia >5000/\u00b5L). In all countries other than Togo, quinine was the most common treatment . The second most common treatment overall was an ACT (22% of cases). The vast majority of patients recovered from their illness following treatment.P. falciparum malaria in infants <5 kg. Study results are expected in 2014.Malaria in infants weighing <5 kg exists in Sub-Saharan Africa and calls for appropriate treatment. A prospective, multicenter, open-label study is currently planned to assess clinical efficacy, safety and pharmacokinetics of AL dispersible tablet in the treatment of acute uncomplicated"}
+{"text": "In Jimma Zone, Ethiopia, the first-line treatment of uncomplicated falciparum malaria has been changed from sulphadoxine-pyrimethamine (SP) to artemether-lumefantrine (AL) in 2006. The objective of this study was to assess the effectiveness of AL in Jimma Zone two to three years after its broad introduction.Plasmodium falciparum mono-infection were consecutively enrolled. Follow-up visits were at day 2, 3, 7, 28 and 42 or any other day if symptoms reoccurred. Primary and secondary endpoints were PCR-corrected and uncorrected cure rates (molecular differentiation between recrudescence and re-infection) on days 28 and 42. Other secondary endpoints were gametocytaemia at day 7 and day 28, parasitaemia at day 2 and 3, and re-infection rates at day 28 and day 42.An open-label, single-arm, 42-day study of AL against falciparum malaria was conducted in four areas with moderate transmission in Jimma Zone between November 2008 and January 2009 and between August and December 2009. Patients (one-81\u2009years) with uncomplicated Plasmodium vivax were observed. Microscopically detected gametocytaemia was reduced by 80% between day 0 and day 7.Of 348 enrolled patients, 313 and 301 completed follow-up at day 28 and at day 42, respectively. No early treatment failure occurred. For per protocol analysis, PCR-uncorrected cure rates at day 28 and 42 were 99.1% (95% CI 98.0-100.0) and 91.1% (95% CI 87.9-94.3), respectively. PCR-corrected cure rates at day 28 and 42 were 99.4% (95% CI 98.5-100.0) and 94.7% (95% CI 92.2-97.2), respectively. PCR-corrected cure rate at day 42 for children \u22645\u2009years was 90.6% (95% CI 82.4-98.7) only. Adverse events were in general mild to moderate. Incidence of new infections was 3.4% during 42\u2009days, no new infections with In general, AL was effective and well tolerated in Jimma Zone, Ethiopia. However, the PCR-corrected recrudescence rate per-protocol at day 42 for children \u22645\u2009years was 9.4%. Therefore, further development should be monitored on a regular basis as recommended by WHO. Plasmodium falciparum against previous anti-malarials, such as chloroquine and sulphadoxine-pyrimethamine (SP), most African countries, including Ethiopia, have changed their national policy in recent years towards first-line treatment with artemisinin-based combination therapy (ACT) . Parasite densities differing by more than 10% between microscopists were controlled by a third microscopist, whose result was final.DNA from dried blood spots on Whatman filter paper (3MM) was extracted by Chelex method as described elsewhere . DNA froPlasmodium falciparum isolates were obtained at day 0 and any day of possible treatment failure up to day 42 of follow-up. PCR products of the paired samples were compared. If any pre-treatment alleles differed from post-treatment alleles, a new infection was considered; if no new allele in pre- and post-treatment samples occurred, a recrudescence was assumed  - and at day 3 (two patients) - mean half-life t3 of 21.4 hours [range 11.9-30.9]. All parasites were cleared at day 7. For those without parasitaemia at day 2 or later, mean half-life of <2.9-4.8 hours can be assumed, considering 1,000-100,000 parasite/\u03bcL at admission. Values are only an approximation (\u00b18 hours) as it was not possible to take the blood exactly 48 or 72 hours later (Table\u2009Six patients (1.9%) showed a prolonged duration till clearance with parasites still present at day 2 (four patients) - mean half-life t range 7.-13.9 - aMean age of late responders (>48\u2009hrs) was 11\u2009years [range 1.3-22.0]. They were from all clusters; two were \u22645\u2009years, underweight and stunted. Initial parasitaemia at day 0 was below 10,000/\u03bcL  in all six patients. Criteria for an early treatment failure, defined by WHO , were noHowever, there was evidence for a strong association of prolonged parasite presence (>48\u2009hrs) with recrudescence , meaning a recrudescence was 9.5 times more likely when parasites were cleared after day 2  at day 0. Mean geometric parasitaemia of 286 patients with ACPR was 9,431/\u03bcL . There was no evidence of a real difference between these groups (P\u2009=\u20090.80).Gametocyte prevalence was 9.5% by microscopy. Eighty percent reduction could be observed between day 0 and day 7  with gametocytaemia at day 28 was significant   of the 286 patients with ACPR being positive for Pfs25-mRNA could still be detected in 22 (70%) of the samples by QT-NASBA. That means that microscopy revealed less than one of seven of the prevalence of gametocytes in these 32 patients. As storage duration and conditions of filter-paper samples as well as blood spot sizes were not standardized, molecular quantification of gametocytes was not possible.In 32 randomly chosen microscopically negative samples from day 0, the gametocyte-specific Most patients had fever, headache, shivering and nausea at admission. Abdominal pain and diarrhoea were less frequent  was reported.Of all 348 patients, 91.3% responded clinically and parasitically adequately to the treatment until day 42. Adverse events were expectedly mild to moderate with low prevalence. Abdominal pain and diarrhoea were the most frequent symptoms.The PCR-corrected failure rate showed incidence rates of 3.8% for new infections and 5.1% for recrudescences at day 42. No early treatment failure occurred. Compared to a trial in 2006, these data show a slight increase in recurrent parasitaemia as none had been detected until day 42 .Other Ethiopian studies detected also high cure rates but most had a follow-up period of 28\u2009days only -23. In tUnfortunately, there are several limitations concerning this study. Drug levels were not tested and only the first drug intake was observed. As absorption is fat-dependent, fatty food was provided and patients were precisely instructed how and when to take the rest of the medication. However, intake was not directly controlled and absorption individually varies, also impaired by symptoms like diarrhoea . TherefoAnother limitation is the method of genotyping. Differentiation of recrudescences and re-infections by PCR is unprecise, especially in low to moderate transmission areas with a limited diversity of strains . SimilarArtemether has a very short half-life of about one hour. Lumefantrine has a half-life of three to six days and is able to clear the remaining parasites and to prevent recurrent parasitaemia ,40. The Six patients showed prolonged presence of parasites detectable still at day 2 and day 3, the calculated half-life time was prolonged. As stated above, this could have been caused by low drug levels. Another theory is the hypothesis of a dormant non-responding parasite stage causing delayed susceptibility to drugs ,42. ThisThe low number of recrudescences and prolonged parasite presence (>48\u2009hrs) impaired multivariate analysis. Risk factors for these outcomes could not be determined. There was strong evidence for an association of both outcomes but the result has to be considered with caution as the sample size is low. However, an association of recrudescence and prolonged parasite presence (72\u2009hrs) was described recently . ParasitThis association was observed in the underlying study. Recurrent parasitaemia occurred in two patients with parasites at day 2 (P\u2009=\u20090.01). Prevalence of patients with parasites at day 3 was 0.6% (2/348) only. Even if adequate drug levels could be assumed, tolerance to artemether would not be suspected. Artemisinin resistance seems highly unlikely if prevalence of patients with parasites at day 3 is <3% .Further analysis indicated a correlation of (prolonged) presence of gametocytes with prolonged parasitaemia and recrudescences. Sample size was low as mentioned but associations seemed plausible. Gametocytes have more opportunities to develop if parasite presence is prolonged and sub-microscopic level as with recrudescences seem to suffice for gametocyte production.Interestingly, gametocytaemia was more frequent in season 2008 than 2009. The broad introduction of ACT in this region might have influenced the gametocyte carriage in the population. ACT clears gametocytes better than S/P, the previous first-line drug. ACT is supposed to destroy the early stages of gametocytes ,43. The A random selection of samples showed very high levels of gametocytaemia with QT-NASBA technique compared to microscopy. Therefore, higher submicroscopic levels have to be assumed on recruitment as well as after treatment. Recent studies showed a difference of factor 3\u20136 between microscopic and submicroscopic gametocyte prevalence ,45. LessFurther, an additional drug for gametocyte clearance should be considered in the area. A single dose of primaquine, best given at day 8, might be sufficient for gametocyte clearance . UnfortuP. falciparum mono-infection occurred in only half of the malaria cases in the study area. Prevalence of mixed infections with P. vivax and infections with P. vivax alone was high and has increased over the last years [[P. vivax only. However, there was surprisingly no re-infection with P. vivax observed. One explanation for this and the very few re-infections with P. falciparum might be that new bed nets were given free of charge by the government to every patient visiting the health centres.Interestingly, t years [, pers coPlasmodium vivax infections diagnosed by PCR and afterwards excluded from analysis showed ACPR in all cases, no relapses occurred. Data on P. vivax and AL are unavailable in this area. Chloroquine is still first-line treatment of P. vivax. A recent study from another area from Ethiopia observed treatment with AL and CQ in vivax malaria. Recurrent parasitaemia rates were 19% and 8%, respectively, although, the evening dose of AL was not observed [observed . Studiesobserved ,49. Howeobserved  and mighAL showed good effectiveness in Jimma. The overall PCR-corrected cure rate per protocol was 94.7%, resulting in a recrudescence rate of below 10%. The PCR-corrected per-protocol failure rate for children \u22645\u2009years was 9.4%. Low drug levels due to malabsorption and dosing problems in children could be responsible. To monitor closely the further development of drug susceptibility in this area, a continuous surveillance should be established and controlled ACT studies with measurement of drug levels and observed drug intake should be conducted.The authors declare that they have no competing interests.TE coordinated the clinical study in Ethiopia, participated in the molecular genetic studies and helped draft the manuscript. NA, KB and SF participated in and carried out the clinical studies. AW and MP established the QT-NASBA technology, carried out the gametocyte analysis and revised the manuscript. TL participated in study design and coordination and drafted the manuscript. NB-R designed the study protocol, was PI of the study, carried out the molecular genetic studies as well as the sequence alignments, conducted the statistical analysis and drafted the manuscript. All authors read and approved the final manuscript.Outcome at day 42 stratified in age groups. Description: The table shows the outcome at day 42 stratified in the age groups\u2009>\u20095\u2009years and\u2009\u2264\u20095\u2009years. The cure rates are in general lower for the under five years old children but the difference is not significant.Click here for fileAssociation with outcome recrudescence . Description: The data show a multi-variate analysis with the outcome recrudescences as dependent variable and independent variables like parasitaemia at day 0, gametocytaemia over time, and parasite clearance, controlled for age and gender. Delayed clearance and gametocytaemie at day 28 were associated with recrudescences. There was evidence for a strong association (P\u2009\u2264\u20090.01) but confidence intervals were wide.Click here for fileClearance of microscopically detected gametocytes over time. Description: Clearance of microscopically detected gametocytes during follow-up in both age groups and overall. Clearance in older patients seemed to be faster than in children below 6\u2009years of age.Click here for fileSymptoms at recruitment (n\u2009=\u2009348). Description: The data show all symptoms presented at admission with absolute and relative frequencies and duration. Fever, headache and shivering were the leading symptoms.Click here for fileAdverse events (AE) during follow-up. Description: The table presents all adverse events reported until day 7. Abdominal pain was the most reported AE. Prevalence was overall low.Click here for file"}
+{"text": "The impact of this combination on anti-malarial resistance-associated molecular markers has not been investigated. In this study, an evaluation of the efficacy and prevalence of drug resistance alleles (pfcrt 72\u201376 and pfmdr1 copy number) and direct sequencing of pfmdr1, pfdhfr and pfdhps.A 28-day follow-up efficacy trial of AS/SP was conducted in eastern Sudan during the 2012 transmission season. Blood smears were collected from patients on days 0, 1, 2, 3, 7, 14, 21 and 28. Blood spots on filter paper were obtained pre-treatment and on the day the patient was parasite positive by microscopy. Genotyping of alleles was performed by qPCR (pfcrt-CVMNK was 30.2% and pfmdr1-N86 was 40.3%. The pfmdr1 haplotype NFD occurred in 32.8% of pre-treatment samples and was significantly higher than previous reports (Fisher\u2019s exact p\u2009=\u20090.0001). The pfdhfr-51I/108N combination occurred in all sequenced isolates and 59R was observed in a single individual. pfdhps substitutions 436A, 437G, 540E, 581G and 613S were observed at 7.8, 77.3, 76.9%, 33.8% and 0.0%, respectively. Treatment failures were associated with the pfdhps haplotype SGEGA at these five codons .Sixty-three patients out of 68 (93%) completed the 28-day follow-up, adequate clinical, and parasitological response occurred in 90.5% and 85.3% of the patients in the per-protocol and intent-to-treat analyses, respectively. PCR corrected per-protocol efficacy was 93.7%. The enrolment prevalence of pfdhfr and pfdhps alleles in vivo is required to inform future treatment guidelines.The decrease of CQR associated genotypes reflects the formal policy of complete removal of CQ in Sudan. However, the frequency of markers associated with SP failure is increasing in this study area and may be contributing to the treatment efficacy falling below 90%. Further monitoring of AS/SP efficacy and of post-treatment selection of  Malaria continues to affect over two million people worldwide with more than 600,000 reported deaths in 2010, most of these in sub-Saharan Africa [Plasmodium falciparum causes over 95% of malaria infections and drug resistance to CQ and SP is well established [P. falciparum malaria in 2004 [in vivo efficacy of SP in eastern Sudan was reported at 82% [Malaria in Sudan causes morbidities and mortality among all age groups and was responsible for more than 95,000 hospital admissions in 2011 . Plasmodablished -7. The wablished . Artesund at 82% , similard at 82% ,13 may rd at 82% ,15 has ppfcrt gene at positions 72 \u2013 76  are well-established markers for resistance to CQ [pfcrt-76T increased from 54% in the early 1990\u2019s [pfcrt-76K allele has returned to high prevalence in parts of Africa where CQ was discontinued, such as Malawi [Specific haplotypes of the ce to CQ ,17. In ey 1990\u2019s ,19 to moy 1990\u2019s ,21. Howes Malawi ,23, Kenys Malawi ,25, Senes Malawi  Tanzanias Malawi ,28 and Ms Malawi .Pfmdr1 haplotypes N86, 184F and 1246D have been selected by artemether-lumefantrine in eastern Sudan [pfmdr1 which is associated with decreased sensitivity to artemisinins in South East Asia [pfmdr1 may continue to be under the influence of ACT currently employed in Sudan.rn Sudan  as well rn Sudan -34. Thesrn Sudan . Amplifiast Asia ,37 has rast Asia  and in nast Asia ,39. Therdhfr) [pfdhps gene  encoding the target enzyme for sulphadoxine are also predictors of treatment failure in vivo[pfdhps-540E allele is more than 50% [pfdhfr CICNI haplotype at codons 50, 51, 59, 108 and 164 has been reported at 90% [pfdhps haplotype SGEAA (436/437/540/581/613) was higher than 80% in the current study area prior to the introduction of AS/SP [Resistance to pyrimethamine is due to a set of sequential mutations in the dihydrofolate reductase gene (dhfr) ,41. The dhfr) ,43. Mutae in vivo. It has than 50% . In eastd at 90% . The pfdof AS/SP . These tpfcrt, pfmdr1, pfdhfr and pfdhps) associated with drug resistance following the wide scale deployment of AS/SP in eastern Sudan is investigated and an updated clinical efficacy of AS/SP is reported.In this study, the prevalence of molecular markers in four genes  comprising 4 mg/kg artesunate on days 0\u20132 as a single daily dose and a single dose of SP  on day 0. For children the dose was adjusted by weight and the tablets were dissolved in water for oral administration. A full dose of the drug was repeated in case of vomiting after 60 minutes and half the dose was repeated if the patient vomited between 30 to 60 minutes after drug administration.Patients were asked to return to the health centre on days 1, 2, 3, 7, 14, 21 and 28 or if they remained unwell, on each visit axillary temperature was measured and a blood smear was taken. Patients were asked about any adverse effects of the drugs such as nausea, vomiting, abdominal pain, itching and rashes: these symptoms were considered drug induced if they were not reported on enrolment.P. falciparum throughout the follow up period were considered to have achieved adequate clinical and parasitological response (ACPR). Treatment failure was classified as early treatment failure (ETF), late parasitological failure (LPF) or late clinical failure (LCF) according to the WHO guidelines [\u00ae) as per the Sudanese National Malaria Control Programme guidelines.Patients who remained microscopy negative for asexual parasitaemia of idelines . Those w\u00ae extraction method [pfcrt loci at codons 72 \u2013 76 were detected by a Taqman assay as described previously [Parasite DNA was extracted from dried filter paper in 96-deep-well plates by the Chelexn method  with modn method . The pfceviously  and 0.1 pfmdr1 encompassing polymorphisms at positions N86Y, Y184F, C1034S, D1042N and D1246Y were amplified with previously reported primers [pfdhfr N51I, C59R, S108N, I164L, pfdhps A436, A437G, K540E, A581G and A613S were amplified as previously described [Regions of escribed  and direpfmdr1 gene was investigated by a qPCR assay as previously described [\u00ae Universal Mastermix. DNA from clones 3D7 and Dd2 were employed as controls for single and multiple copy number, respectively. Each sample and control was run in triplicate.Amplification of the escribed  employinMsp1 and msp2 polymorphic markers were amplified as described previously [eviously  to confiThe study received ethical clearance from the Health Research Board at Ministry of Health, Kassala State. Written informed consent was obtained from each patient or child guardian.p-value\u2009\u2264\u20090.05.The intention-to-treat analysis included all enrolled patients who met the inclusion criteria and took at least one full dose of AS/SP. Patients lost to follow-up or withdrawn from the study were considered to be treatment failures. The per protocol (PP) analysis included data for patients who had completed the follow-up. Those patients lost to the follow-up or who were withdrawn because of protocol violations were excluded from the PP analysis. Clinical data were entered in MS-Excel and analysed in SPSS software . The means  were calculated for all patients. The difference in frequency of haplotypes observed in 2003 , 2006 3, and in P. falciparum infection. Sixty-eight patients fulfilled the enrolment criteria and consented to participate in the study, sixty-three (92.6%) of whom completed 28-days of follow up. Patient age, weight, temperature, parasite density and gender are summarized in Table\u00a0A total of 453 patients who came to the clinic with fever and reporting any malaria-like symptoms were screened for On day 1; 12.7% (8/63) patients were febrile and 20.6% (13/63) patients were parasitaemic. However, all patients cleared their parasitaemia by day 3 with one patient remaining febrile (temperature 38\u00b0C). Gametocytaemia was observed in 3.2% (2/63) of patients at recruitment. Adverse effects of the drugs such as nausea, headache, cough, dizziness, abdominal pain and diarrhoea were reported in 34.9% (22/63) of patients between day 19 and 28. In the intent-to-treat analysis where all patients who were enrolled were included in the analysis, the efficacy of AS/SP was 85.3%. While in the per-protocol analysis the efficacy was 90.5%. Two patients failed treatment at day 21, one failed treatment at day 26 and two failed treatment at day 28. The PCR corrected efficacy was 93.6% where 4/5 of treatment failures were classified as recrudescing parasites. Factors such as pre-treatment parasite density, axillary temperature and age did not differ significantly between those who failed treatment and those who were successfully treated.pfcrt haplotype CVMNK has significantly recovered from 10% in 2003 [p\u2009<\u20090.001) in this study. . The N86 allele has also increased significantly from 14% [p\u2009=\u20090.0059). The prevalence of the wild type Y184 was 15.6% but this was not significantly different from 2006 (Fisher\u2019s exact p\u2009=\u20090.16), while there was a significant decrease in mutant 1246Y (Fisher\u2019s exact p\u2009=\u20090.018)  .The wild type  in 2003  to 30.2%from 14%  to 40.3%pfdhfr gene a total of 59 samples were successfully sequenced for positions 50 to 164. All of which carried the 51I and 108N mutant alleles. This is a significant increase from previous years (Fisher\u2019s exact p\u2009=\u20090.041). One sample (1.7%) harboured the mutant 59R and all of the samples were wild-type at position 164. Thus the predominant haplotype for pfdhfr in eastern Sudan is 51I/C59/108N.For the pfdhps positions 430 to 613. A novel polymorphism at position L516F was detected in one sample. The prevalence of the 581G allele significantly increased since the 2003 study from 14% to 33.8% (Fisher\u2019s exact p\u2009=\u20090.0005), while the 437G decreased slightly from 89% to 77% (Fisher\u2019s exact p\u2009=\u20090.0134) [p\u2009=\u20090.0009) and the 437G (p\u2009=\u20090.0018). The prevalence of S436A, A437G, L516F, K540E and A613S are listed in Table\u00a0A total of 65 pre-treatment samples were successfully genotyped at the Pfdhps sequencing was successful for all five treatment failures, collected at day 21 (2), day 26 (1) and day 28 (2). The haplotype across the five polymorphic codons 436, 437, 540, 581 and 613 are shown for these two groups in Figure\u00a0pfdhps haplotype SGEGA, at these five polymorphic positions, was significantly more common in parasites detected after treatment  whether these parasites were recrudescing or re-infecting parasites. Further, the presence of this pfdhps genotype in the pre-treatment infection was a significant risk factor for later treatment failure  the wide confidence intervals indicate that these observations need detailed exploration in larger studies.Plasmodium falciparum carrying mutations associated with drug resistance may be less fit to survive in the absence of drug pressure. The continuous use of CQ monotherapy for decades has lead to almost saturation of the P. falciparum population in eastern Sudan carrying the mutant pfcrt allele CVIET [pfcrt where transmission is higher and perennial in central Sudan [pfdhfr/pfdhps mutants (51I/108N/437G/540E) in the late 1990\u2019s and early 2000\u2019s [in vivo per-protocol PCR corrected efficacy of AS/SP over 28 days is 90.5%. All patients cleared their microscopic parasitaemia by day 3. Approximately 10% of patients exhibited recrudescing parasites as confirmed by msp1 and msp2 genotyping during the study. It is important to emphasize the relatively lower transmission season in eastern Sudan compared to other parts of East Africa. Therefore, to definitely establish whether these post-treatment infections were true recrudescence or re-infections requires genotyping at more polymorphic loci.le CVIET . Lower dal Sudan . Moreovey 2000\u2019s ,20,54. Ty 2000\u2019s -57. The y 2000\u2019s . Therefoy 2000\u2019s . The basy 2000\u2019s ,60-62, wA limitation to our data is the lack of parallel PCR data for days 3 and 14. Sub-microscopic parasites at these time points have been detected in previous studies from eastern Sudan ,30. DeteIn the present study, the prevalence of molecular markers eight years after the adoption of ACT is compared to previous reports from the same study area of eastern Sudan. In this area IPTp is recommended for all pregnant women, as in other parts of Sudan.pfdhfr allele (51I/108N) has reached 100%, which clearly demonstrates the effect of drug pressure on this locus. The observation that contrasts with other African countries is the slow evolution of pfdhfr in this region where the double mutant 51\u2013108 allele has widely spread while the wild type C59 has been maintained. However it is in agreement with a recent report from Yemen [in vivo efficacy of SP in this region. The pfdhfr 59R has been previously reported at low frequency in Khartoum, central Sudan in 1996/1997 [pfdhfr-51I/59R/108N haplotype currently predominates [pfdhfr were stable over four consecutive seasons [pfdhfr allele 51I/108N may have been imported into this area earlier and has been maintained within a limited parasite population in a low transmission area of eastern Sudan. However, it would be of interest to examine pfdhfr sequences and microsatellite markers from other regions of Sudan with different transmission intensities to test this hypothesis.The prevalence of a double mutant om Yemen . The 59R996/1997  and Nuba996/1997 . Howeverominates ,69. Thes seasons . Thus, tpfdhps-437G (Fisher\u2019s exact p\u2009=\u20090.0134) is observed compared to previous reports [pfdhps allele, SGEGA, is significantly associated with treatment failure in patients receiving AS/SP for clinical malaria. However of 23 pretreatment infections harbouring SGEGA, only four recrudesced. The increase of SGEGA has been previously reported from Tanzania in cross sectional surveys following several years of SP use [Interestingly in this study, a small decrease in  reports . The 581 reports  emphasizf SP use . The potpfcrt-76T/pfmdr1-86Y. Wild type genotypes at these two loci have significantly recovered, which provides supporting evidence that CQ use has been successfully reduced [pfmdr1 genotype previously reported from eastern Sudan [pfmdr1amplification has failed to be established and spread in Africa, possibly due to lack of the use of MQ across the continent.As previously reported from other African countries ,24,28, t reduced . Interesrn Sudan  was not rn Sudan  and may pfdhps-540E is above 50% is also recommended [pfdhps haplotype identified here, is required to provide informed treatment guidelines in the near future. In addition the emergence of slow parasite clearance in South East Asia warrants close monitoring of the efficacy of ACT and molecular markers associated with resistance to partner drugs.In Sudan SP is recommended for iPTp and the availability of SP as an over-the-counter drug maintains the pressure on the parasite population. The efficacy of AS/SP is reaching the current threshold for anti-malarial policy change of 90%. Continuous use of SP as a partner to artesunate may be the cause of declined AS/P efficacy. In addition discontinuation of SP in areas where the population prevalence of ommended ,71. MoniThe authors declare that they have no competing interests.in vivo study, analysed the clinical data and drafted the manuscript. SA participated in the molecular genetic studies and analyses. CJS designed the molecular study, performed the statistical association analysis, contributed to the manuscript and critically revised it, IA conceived of the study, participated in its design and coordination and drafted the manuscript. All authors read and approved the final manuscript.NBG carried out the molecular genetic studies, the sequence alignment, analysed the results and drafted the manuscript. TMA carried out the"}
+{"text": "Plasmodium falciparum infection from residual antigens from a previous infection is crucial in endemic areas where residents are repeatedly exposed to malaria. The efficiency of two RDTs based on histidine-rich protein 2 (HRP2) and lactate dehydrogenase (LDH) antigens were studied and compared with two microscopy techniques (Giemsa and acridine orange-stained blood smears) and real-time polymerase chain reaction (PCR) for assessment of initial clearance and detection of recurrent P. falciparum infections after artemisinin-based combination therapy (ACT) in a moderately high endemic area of rural Tanzania.Rapid diagnostic test (RDT) is an important tool for parasite-based malaria diagnosis. High specificity of RDTs to distinguish an active P. falciparum malaria infection were followed up on nine occasions, i.e., day 1, 2, 3, 7, 14, 21, 28, 35 and 42, after initiation of artemether-lumefantrine treatment. At each visit capillary blood samples was collected for the HRP2 and LDH-based RDTs, Giemsa and acridine orange-stained blood smears for microscopy and real-time PCR. Assessment of clearance times and detection of recurrent P. falciparum infections were done for all diagnostic methods.In this exploratory study 53 children\u2009<\u2009five years with uncomplicated The median clearance times were 28 (range seven to >42) and seven (two to 14) days for HRP2 and LDH-based RDTs, two (one to seven) and two (one to 14) days for Giemsa and acridine orange-stained blood smear and two (one to 28) days for real-time PCR. RDT specificity against Giemsa-stained blood smear microscopy was 21% for HRP2 on day 14, reaching 87% on day 42, and \u226596% from day 14 to 42 for LDH. There was no significant correlation between parasite density at enrolment and duration of HRP2 positivity . Recurrent malaria infections occurred in ten (19%) children. The HRP2 and LDH-based RDTs did not detect eight and two of the recurrent infections, respectively.The LDH-based RDT was superior to HRP2-based for monitoring of treatment outcome and detection of recurrent infections after ACT in this moderately high transmission setting. The results may have implications for the choice of RDT devices in similar transmission settings for improved malaria case management.NCT01843764Clinicaltrials.gov,  Plasmodium falciparum diagnosis is generally recommended by the World Health Organization (WHO) to target artemisinin-based combination therapy (ACT) to patients with confirmed malaria infections [Parasite-based fections . This isfections ,3. The afections .P. falciparum infection. However, a concern with HRP2-based RDTs is the presence of residual antigenaemia resulting in persistent positive test results during several weeks after a successful treatment [Malaria RDTs are based on detection of parasite antigens. The main antigen targeted is histidine-rich protein 2 (HRP2), which has proven to be a highly sensitive and stable marker for identification of reatment ,6. This reatment ,8.P. falciparum LDH (PfLDH) and Plasmodium vivax (PvLDH), or pan-Plasmodium (pLDH), detecting all five human malaria species [P. falciparum, but they are more specific since LDH is rapidly cleared from the blood following a successful anti-malarial treatment. Consequently, LDH-based RDTs do not remain positive after parasite clearance [PfLDH-based RDTs have shown sensitivities and heat stabilities similar to HRP2-based RDTs [Another antigen used in RDTs is parasite-specific lactate dehydrogenase (LDH), either species specific, i.e.,  species . LDH-baslearance ,10. Intesed RDTs -14.P. falciparum infections of HRP2 and LDH-based RDTs during 42\u00a0days after initiation of artemether-lumefantrine treatment in children with uncomplicated malaria in a moderately high endemic area of rural Tanzania. Furthermore, since no previous study has included clearance by polymerase chain reaction (PCR) as a comparator, this was done to allow a more comprehensive evaluation of RDT for monitoring of anti-malarial treatment outcome. The entire assessment herein reported thus included two RDTs (HRP2 and LDH-based) for antigen detection, compared with two microscopy techniques (Giemsa-stained thick blood smears and acridine orange-stained thin blood smears) for whole parasite detection and real-time PCR for detection of parasite DNA.Previous RDT studies in this particular field have mainly focused on post-treatment clearance and have primarily followed patients until the tests have become negative ,13. ThusThis health facility-based study was conducted during the peak seasons for malaria transmission, in June to September 2009 and July to October 2010 at Mlandizi health centre, Kibaha district and during March to May 2011 in Fukayosi dispensary, Bagamoyo district, both located in Coast Region, Tanzania. Artemether-lumefantrine was introduced as first-line treatment for uncomplicated malaria in 2006 in the study area, whereas RDT had not yet been implemented for parasite-based malaria diagnosis. At the time of the trial malaria transmission was considered to be moderately high in both study sites.The Mlandizi health centre provides basic in- and outpatient care for a population of approximately 33,000. Laboratory services, including malaria microscopy, are available during office hours. The monthly blood smear positivity rate among febrile children\u2009<\u2009five years of age was 33% (range 20-48%) and 15% (range 11-21%) during the sampling period in 2009 and 2010, respectively. There was a period of artemether-lumefantrine stock-out in Mlandizi in 2009. During this period patient recruitment was stopped. There was an on-going insecticide-treated bed net campaign in the area markedly reducing the incidence of malaria among children\u2009<\u2009five years between 2009 and 2010.Fukayosi dispensary provides basic outpatient care for a population of about 7,000. Malaria microscopy service is available seven days a week. Malaria blood smear positivity rate was 19% (74/384) among febrile children\u2009<\u2009five years of age during the conduct of the trial.Participating laboratory staff and study nurses at the two study sites received one day\u2019s training in performance and interpretation of both RDTs before the start of the study.P. falciparum mono-infection with a parasite density of 2,000-250,000/\u03bcL, and willing/able to comply with the 42\u00a0days follow-up were eligible to participate in the study. Children with a history of anti-malarial drug intake within two weeks or symptoms/signs of severe disease were excluded. Written informed consent was obtained from a parent/guardian of all enrolled children.Children between six and 59\u00a0months presenting at the study sites with fever, i.e., measured axillary temperature of \u226537.5\u00b0C or a history of fever during the preceding 24\u00a0hours, and a positive screening blood slide for P. falciparum mono-infection solely on day 0. A case record form was completed at enrolment by a clinical officer, with clinical and demographic information, including age, sex and information on use of insecticide-treated bed nets. Body temperature, symptoms and prescription of drugs were recorded in the case record forms on all visits.At enrolment, i.e., day 0, a complementary finger-prick capillary blood sample was taken for two thick and thin smears, and two RDTs. In addition, approximately 50\u00a0\u03bcL of blood was spotted on a filter paper. All enrolled children were treated with artemether-lumefantrine (Coartem\u00ae) in standard doses based on body weight, according to national treatment guidelines . Only thP. falciparum the child was retreated with artemether-lumefantrine.Whenever fever and/or any other symptoms/signs of disease re-occurred during follow-up, a Giemsa-stained blood slide was to be read directly at the health centre and if positive for The only incentive given to the study participants was bus fares to cover travel costs during follow up visits.f , detecting P. falciparum-specific HRP2 antigen (hereafter referred to as HRP2) and CareStart\u2122 Malaria (G0151), , detecting P. falciparum-specific LDH antigen (hereafter referred to as LDH), were performed and interpreted on site according to the manufacturer\u2019s instructions. ParaHit was, at the time of the study, approved by the Tanzanian National Malaria Control Programme and was the most deployed RDT. The single Pf CareStart test was chosen based on the heat stability and performance of the CareStart pan-LDH test in the WHO product testing 2009, where it was among the best performing LDH-based tests for P. falciparum detection [P. falciparum and one control band.Two RDTs, ParaHIT \u00aeetection . Both RDA laboratory technician or study nurse, blinded to any Giemsa-stained blood smear result, performed, interpreted and recorded the RDT results. Very faint bands at the test line position were to be defined as positive. Band intensity was not recorded. In case the control line did not appear, the result was considered invalid and the test was repeated. The RDT kits were stored at <30\u00b0C prior to use, the temperature being recorded daily.P. falciparum mono-infection. One of the two thick smears was examined by two independent, experienced microscopists, who were unaware of the RDT results, at MUHAS. A total of 200 microscopic fields (\u00d7100 magnification) were examined before a smear was considered negative. Asexual parasite densities were calculated by counting parasites against 200 white blood cells (WBC), assuming 8,000 WBC/\u03bcL of blood. If less than 10 parasites were detected per 200 WBC, estimates were made against 500 WBC [versus negative result between the readers, were subjected to a third blinded reading at Karolinska Institutet (KI), Sweden. In addition, all blood slides from children showing a negative thick blood smear and a positive PCR at the same time point, as well as a random sample of 10% of all blood slides, plus the blood slides from the time point of each participant with the last CareStart positive, and the first CareStart negative results, were subjected to microscopy reading at KI for quality control. The mean of the two most concordant counts were used to calculate the final parasite density [versus negative results between the first two readers and the third, the third reading at KI was defined as decisive.One of the two thin blood smears collected on day 0 and the thick blood smears from all sampling points were stained with 5% Giemsa for 20\u00a0minutes at the health facilities, after which they were transported once weekly to Muhimbili University of Health and Allied Sciences (MUHAS) in Dar es Salaam. The day 0 Giemsa-stained thin smear was examined for confirmation of  500 WBC . Gametoc density . In caseThe thin blood smears from all sampling points were subjected to acridine orange staining and reading at Muhimbili University Hospital. The thin smears were fixed in methanol. A solution of 0.01% acridine orange in phosphate buffer (pH\u00a07.2) with 5% glycerine was then applied to the smears, which were read in a fluorescence microscope at \u00d740 magnification . The resPlasmodium DNA by an 18S rDNA probe based real-time PCR assay [P. falciparum-specific nested PCR [P. falciparum controls  as well as negative controls were included in each 96-well PCR plate.Approximately 50\u00a0\u03bcL of blood seeded on a filter paper (Whatman 3MM\u00ae) from all sampling points were collected for molecular analysis. The filter papers were dried and put in individual plastic bags and transported to KI. The filter papers were stored at <30\u00b0C until processed. Three 3-mm punches (approximately 10\u201315\u00a0\u03bcL) from each filter paper sample were extracted with a modified version of the ABI 6100 Nucleic Acid Prep Station protocol  as previously described . DNA wasCR assay . A cut-osted PCR . Two posmerozoite surface protein (msp) 1, msp 2 and glutamate rich protein (glurp) was performed according to standard protocols to differentiate re-infection (new infection) from recrudescence (treatment failure). For each marker, recrudescence was defined as the presence of at least one matching allelic band and re-infections were defined as the absence of any matching allelic band in samples at enrolment (day 0) and at day of recurrent infection [Filter-paper blood spots from patients with recurrent PCR positivity during follow-up were re-extracted using the Chelex-100 method . Stepwisnfection ,25.Clearance time was defined as the first sampling day after initiation of treatment when a test result was negative. There were two exceptions to this. First, when a RDT result turned negative for one sampling day, followed by a positive RDT result again the following sampling day, the negative result was ignored if the PCR and/or BS results did not indicate presence of a recurrent infection. Second, when one negative PCR result was followed by a positive PCR result the following sampling day up to day 7, the negative result was ignored. From day 14 and onwards the clearance time was calculated from the first day with negative result.P. falciparum DNA (PCR) confirmed by a positive Giemsa-stained blood smear microscopy and/or LDH during follow-up after the initial infection had been cleared.Recurrent infection was defined as detection of The primary outcomes were clearance time and detection of recurrent infection with the five diagnostic tests. Secondary outcomes included specificity of the two RDTs against Giemsa-stained blood smear microscopy (gold standard), identification of PCR-adjusted re-infection/recrudescence among the recurrent infections and correlation between parasite density at enrolment and persistence of HRP2. In the calculation of correlation between day 0 parasite densities and duration of HRP2 positivity, seven children were included who had cleared HRP2 positivity before they were lost to follow-up. These seven children were not included in any other analysis.The study was considered exploratory, which precludes a power calculation. A sample of \u226550 children was predefined.Data were entered in Microsoft Excel\u00ae and analysed using STATA 12\u00ae software. Categorical variables were compared using Fisher\u2019s exact test. Pearson linear correlations were calculated in SPSS. Sensitivity and specificity of acridine orange against Giemsa-stained blood smear microscopy (gold standard) and both microscopic methods against PCR (gold standard) was calculated. Statistical significance was stated at the 5% level and 95% confidence intervals (CI) are presented.The study was conducted in accordance with the Declaration of Helsinki and Good Clinical and Laboratory Practices. It was approved by the Directorate of Research and Publications, MUHAS (Ref.No.MU//RP/AEC/Vol.XIII/142) and the Regional Ethics Committee, Stockholm, Sweden.http://www.clinicaltrials.gov with study identifier NCT01843764.The study is registered at The flow of patients through the trial is outlined in Figure\u00a0All children were positive with both RDTs at day of inclusion. Baseline characteristics of the 53 children included in the analysis are presented in Table\u00a0The calculations for HRP2 clearance are based on the 43 children without recurrent infection during follow-up, since eight of the ten children with parasite recurrence remained HRP2-positive from enrolment up to the time of recurrent infection. All other tests, i.e., LDH, Giemsa and acridine orange-stained blood smears as well as PCR cleared before parasite recurrence. Consequently, clearance calculations for these tests are based on data from all 53 children. The median clearance times for the five diagnostic tests are presented in Figure\u00a0The geometric mean clearance time for HRP2 was 26.4 (95% CI 23.0-30.3) days. One patient cleared HRP2 by day 7 and three remained positive up to day 42 (last day of follow-up). The false positivity rates for HRP2 against PCR on days 14, 21, 28, 35 and 42 were 80% (32/40), 64% (27/42), 43% (18/42), 24% (10/42) and 7% (3/41), respectively. All PCR positive results were also HRP2-positive throughout the study. There was no significant correlation between parasite density at enrolment and duration of HRP2 positivity  . Four children (8%) remained positive by microscopy until day 3. Two children were microscopy positive day 2 and 3, respectively, with parasite densities of 16 and 32/\u03bcL each. However, PCR was negative for both samples at these time points. There was no significant association between clearance times of Giemsa-stained blood smears and HRP2 (p\u2009=\u20090.50).Acridine orange-stained blood smears had a mean clearance time of 2.1\u00a0days (95% CI 1.9-2.3). The mean clearance time for PCR was 2.9\u00a0days (95% CI 2.3-3.6). Persistent PCR positivity up to day 7, 14 and 21 was observed in one, one and two children, respectively. In two of these four children, solely gametocytes were detected by Giemsa-stained blood smear microscopy at two and three sampling points, respectively, during the time of persistent PCR positivity.P. falciparum infection during the 42\u00a0day follow-up as assessed by PCR, Giemsa-stained blood smear and/or LDH. One recurrent infection was defined as recrudescence, six as re-infections, whereas three were undetermined by PCR genotyping  compared with LDH (seven days). Due to persistent HRP2 positivity from the initial infection, only two out of the ten children with recurrent P. falciparum infections during follow-up were identified by HRP2, whereas LDH was able to recognize eight at the time of parasite recurrence. Acridine orange blood smear microscopy did not provide any additional information compared with the other tests used in this study.The efficiency of two Long clearance times for HRP2-based RDTs after ACT treatment have previously been shown ,26,27, wSimilarly with the present findings, a relatively rapid clearance of LDH has previously been documented ,12. It hExpert blood smear microscopy remains gold standard for estimation of parasite clearance in clinical trials of anti-malarial drugs. Blood smear positivity by day 3, i.e., 72\u00a0hours after initiation of ACT treatment, was observed in 4/53 (8%) of the patients. This is a relatively uncommon finding among African children treated with ACT for uncomplicated malaria ,31. The et al. identified a wide range of HRP2 concentrations at equal parasite densities both in panels of cultures and of field isolates [No correlation was observed between parasite density at enrolment and duration of HRP2 positivity. This is in contrast to several previous publications that report a strong positive correlation ,6,26,27.isolates . Varyingisolates  and antiet al. found in a study performed in Thailand that 40/92 (43%) patients experienced recurrent infection during a 28-day follow-up [In the moderately high transmission area where the study was conducted, ten (19%) children had recurrent infection detected between days 14 and 35 during follow-up. All recurrent infections were detected by PCR, whereas both the LDH-based RDT and Giemsa-stained blood smear detected eight, HRP2 identified two and acridine orange smear microscopy identified only one of these infections at the time of parasite recurrence. Few other studies have looked at the efficiency of RDTs for detection of recurrent infections during follow-up after anti-malarial treatment. Maxay The usefulness of RDTs for monitoring of anti-malarial treatment, i.e., clearance time as well as identification of treatment failure/re-infection, is highly dependent on test specificity. HRP2-based RDTs appear not to be a sufficient tool for this. LDH, with a limited but still longer clearance time as compared with blood smear microscopy, may be useful but for detection of prolonged parasite clearance as a sign of emerging artemisinin tolerance/resistance, it is probably not sufficient.et al. showed that in an area of high transmission the specificity increased towards the end of the rainy seasons, and in older age groups probably due to increased HRP2 antibody levels in the population. In low endemic areas the specificity of HRP2 among febrile patients generally increases due to the low malaria incidence and thus the low risk of detecting remaining antigenaemia after cleared infections. LDH, on the other hand, which has shown a comparatively lower sensitivity for detection of P. falciparum infections with densities <200/\u03bcL, may in areas of low transmission where parasite densities generally are lower especially among asymptomatic cases [Sensitivity and specificity of the HRP2 and LDH antigens for case detection are highly dependent on malaria endemicity ,28. In hic cases , be a leP. falciparum isolates [Other limitations with HRP2-based RDTs are the genetic diversity, including the deletions recently described among African isolates ,36, as wisolates . To dateisolates ,39. All Previous RDT studies in this particular field have mainly followed up the study participants until the RDTs have become negative ,13. ConvA general limitation when assessing performance of parasite based diagnostic tests that indeed detect different things such as antigens (RDT), DNA (PCR) or whole parasites (microscopy) is that the results are not fully comparable. Furthermore, this is a small study with a limited number of patients with recurrent infections detected during follow-up.The LDH-based RDT was superior to HRP2-based for monitoring of treatment outcome and detection of recurrent infections after ACT in this moderately high transmission setting. The results may have implications for the choice of RDT devices in similar transmission settings for improved malaria case management.The authors declare that they have no competing interests.BAS, MM, AM, AB, and ZP conceived and designed the study. MM, BAS, ZP, and BN carried out and supervised the field work. BAS and UM performed the laboratory analyses. BAS, UM, MM, AB, and AM analysed the data and drafted the manuscript. MP gave important intellectual input in the statistical analysis. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum in malaria patients with haemoglobin E trait, but it did not increase parasite inhibition in an in vitro study using haemoglobin AS erythrocytes. The current study describes the efficacy of artemisinin derivatives on P. falciparum clearance in patients with glucose-6-phosphate dehydrogenase deficiency (G6PD), a haemoglobin enzyme deficiency, not yet studied in the same context, but nonetheless is a common in malaria endemic areas, associated with host protection against uncomplicated and severe malaria. The impact of G6PD deficiency on parasite clearance with ACT treatment was compared between G6PD-deficient patients and G6PD-normal group.Artemisinin-based combination therapy (ACT) is currently the most effective medicine for the treatment of uncomplicated malaria. Artemisinin has previously been shown to increase the clearance of P. falciparum malaria residing in Kambila, Mali were analysed. Study participants were randomly assigned to receive either artemether-lumefantrine (Coartem\u00ae) or artesunate plus mefloquine (Artequin\u2122). A restriction-fragment length polymorphism analysis of PCR-amplified DNA samples was used to identify the (A-) allele of the gene mutation responsible for G6PD deficiency (G6PD*A-). 470 blood samples were thus analysed and of these, DNA was extracted from 315 samples using the QIAamp kit for PCR to identify the G6PD*A- gene.Blood samples from children and adults participants (1 to 70 years old) with uncomplicated The DNA amplified from 315 samples using PCR showed that G6PD*A- deficiency was present in 56 participants (17.8%). The distribution of the specific deficiency was 1%, 7% and, 9.8% respectively for homozygous, hemizygous, and heterozygous genotypes. Before treatment, the median parasitaemia and other baseline characteristics  between G6PD deficiency  and G6PD-normal participants were comparable (p > 0.05). After treatment, parasite clearance did not change significantly whether the participants were G6PD deficient or G6PD normal on day 1  and on day 2 .P. falciparum in the treatment of uncomplicated malaria using ACT.The presence of G6PD deficiency does not appear to significantly influence the clearance of  Plasmodium falciparum to older anti-malarial drugs, artemisinin, a natural product found in the leafy portions of Artemisia annua (qinghao) and its derivatives, have emerged as alternative drugs for the treatment of falciparum malaria [P. falciparum. The use of artemisinin-based combination therapy (ACT) is associated with a rapid clearance of the parasite and a low probability of drug-resistant parasite emergence [et al in Nigeria showed a 28-day cure rate of about 95% with artemether-lumefantrine (AL) [et al also reported day 28 cure rates of 95.2% and 92.0% for dihydroartemisinin/piperaquine (Artekin) and ASAQ, respectively, in Rwanda [In a race to combat the ever increasing resistance of  malaria . Artemismergence . ACT is ine (AL)  and 93% ine (AL) . Karema P. falciparum susceptibility to anti-malarial drugs to correlate with abnormal haemoglobins. A clinical study in Thailand first suggested that haemoglobin E trait  interacts with artemisinin derivatives to enhance P. falciparum clearance in malaria patients with haemoglobin E trait compared to patients treated with other anti-malarials [P. falciparum in \u03b1-thalassemia [in vitro [in vitro study has demonstrated no evidence of elevated artemisinin activity on P. falciparum in haemoglobin AS erythrocytes [P. falciparum in abnormal haemoglobin carriers make it imperative to assess the efficacy of ACT in G6PD deficient patients in Mali, where this haemoglobinopathy is common. As far as the literature is concerned, no such studies have been done to study the efficacy of artemisinin against P. falciparum in G6PD deficient patients. G6PD variants correlates with historical distribution of malaria [A-) encoding G6PD with 10%-50% of abnormal enzyme activity is widespread in Africa [P. falciparum and treatment efficacy in uncomplicated malaria as assessed by WHO 28-day protocol using ACT. The study hypothesis was that treatment with ACT may induce faster parasite clearance in G6PD deficient patients compared to G6PD normal patients.Studies have also shown alarials . Anotherlassemia . Other sin vitro . More rehrocytes . These c malaria  and the n Africa . The aim\u00ae) or artesunate plus mefloquine (Artequin\u2122). The G6PD mutant gene was identified by nested PCR performed on stored blood samples (filter paper dot). The study hypothesis was based on fact that P. falciparum in G6PD deficient patients would clear faster after ACT initiation compared to normal patients. A nested case control design was used to compare G6PD deficient to G6PD-normal subjects with a ratio of one case for four controls.The study participants were selected from a randomized study conducted in Kambila village. The study participants were randomized to receive artemisinin-based combination therapy artemether-lumefantrine (CoartemThe study participants were selected from children and adults (1 to 70 years old) patients who came to the health centre for care during the study period. Only patients with uncomplicated malaria who gave their consent after explanation of study objectives and understood were enrolled into the study. Children were enrolled after obtaining a written parental or guardian fully informed consent. Blood samples were collected from both children and adult patients and were analysed for G6PD status, and parasite clearance since it has been shown that parasite clearance is faster in adults than children .P. falciparum is the predominant infecting species, accounting for more than 95% of malaria cases [Study was conducted during a malaria transmission season from August 2004 through January 2005, in Kambila, Mali. Kambila is a peri-urban village of Kati located about 25 km from Bamako with a population of approximately 1,500 inhabitants. Malaria in this area is hyper-endemic and transmission is highly seasonal. ia cases . Kambilaia cases .The sample size was computed based on 70% of parasite clearance observed 24-hours after artesunate/amodiaquine treatment initiation in a general population at Bougoula-Hameau, Sikasso site, a similar malaria hyper-endemic area of Mali. Assuming an error risk alpha set at 0.05, a power of 80%, for one case matched with four controls and fixing a detectable risk estimate at 3.0 (Odds ratio), we will need 55 samples in the cases group (G6PD deficient) and 220 in the control group (non G6PD deficient), rounded up to 56 cases and 259 controls.P. falciparum parasite density of between 2,000 and 200,000/\u03bcl; 3) had an auxiliary temperature \u2265 37.5\u00b0C; 4) were a resident of the study site; and 5) could take medication orally.Participants were included if they fulfilled the following criteria: 1) weighed \u2265 10 kg; 2) had a et al [Persons were excluded if they had symptoms or signs of severe malaria , had a set al . WithdraThe protocol was reviewed and approved by the ethical committee of the Faculty of Medicine, Pharmacy and Odonto-Stomatology of the University of Bamako before starting the randomized clinical trial. Approval letter from these three Faculty ethical of committees was also obtained before conducting the molecular analysis for the G6PD deficiency study.Giemsa-stained thick blood smears were read by experienced laboratory technicians who were blinded to treatment allocations. Parasite densities were calculated by counting the number of asexual parasites until 300 leukocytes were observed and then converting that to parasites per microlitre of blood, assuming an average leukocyte count of 7,500/\u03bcl. For quality control, 10% of the slides were selected at random and re-read by another lab technician who was unaware of the results of the first reading.Two methods were used\u00a0for DNA extraction on blood spotted filter paper:(1). An initial extraction was done using methanol. If no DNA could be extracted from blood samples or if the results of the PCR were conflicting, DNA was extracted using the QIAamp kit . (2). IfA_) allele of the gene responsible for G6PD deficiency was determined by restriction fragment length polymorphism analysis of PCR-amplified DNA samples. Under conditions intended to eliminate the risk of cross-contamination and with appropriate water-only negative controls, exon 4 of G6PD was amplified using a nested PCR protocol: 1 \u03bcg of genomic DNA was first amplified using primers 5'-GTCTTCTGGGTCAGGGAT-3' (forward) and 5'-GGAGAAAGCTCTCTCTCC-3' (reverse). Denaturation at 94\u00b0C for 2 min was followed by 45 cycles of denaturation at 94\u00b0C for 30 s, annealing at 60\u00b0C for 30 s, extension at 72\u00b0C for 60 s, and final extension at 72\u00b0C for 4 min. Nested amplification was performed using primers 5'-CCTGTTCCCTCTGCCACA-3' (forward) and 5'-GGGGGTCTCAAGAAGTAC-3' (reverse). Denaturation at 94\u00b0C for 2 min was followed by 35 cycles of denaturation at 94\u00b0C for 30 s, annealing at 60\u00b0C for 60 s, extension at 72\u00b0C for 30 s, and a final extension at 72\u00b0C for 4 min. Amplification products were recovered with separate pipettes in laboratory areas apart from the PCR-preparation bench and digested with the restriction endonuclease NlaIII [The  at each follow-up visit.The primary study endpoint was the time to parasitaemia disappearance from day 1 after treatment initiation. Parasitaemia clearance was defined as the parasitaemia disappearance at day 1, day 2 and up to day 28 after treatment. Proportion of gametocytes carriers at day 3\u00a0was defined as the percent of subjects carrying gametocytes that day regardless of their gametocyte carriage on day 0. The efficacy of ACT was assessed as an adequate clinical and parasitological response at day 28 according to WHO 2003 guideline [P value (two-sided) < 0.05 was considered as statistically significant.Data were double-entered, validated using Microsoft Access , and analysed with STATA version 10.0 . Chi-square test or Fisher exact test were used to compare proportions between groups. Mann-Whitney test or Kruskal Wallis test were used as appropriate to assess differences of median parasitaemia between groups for continuous variables. Logistic regression was used to control confounding factors, the odds ratio and exact 95% CIs were calculated. A The DNA amplified from 315 samples out of the 470 samples using PCR showed that G6PD*A- deficiency was present in 56 participants (17.8%). The distribution of the specific deficiency was 1%, 7% and, 9.8% respectively for homozygous, hemizygous, and heterozygous genotypes. The mean haemoglobin and the mean body temperature were compared and no difference was seen between G6PD deficient genotype and normal genotype (p > 0.05) . The risk estimate of . The risFrom day 0 to day 3 the comparison of median parasite density and parasite clearance with respect to the different G6PD genotypes including the normal group didn't show any significant difference between groups Figure .Data were analysed by age groups (age groups as indicated in table The purpose of this study was to determine the combined effect of G6PD deficiency and ACT treatment (artemether-lumefantrine and artesunate plus mefloquine) on parasite clearance in patients with uncomplicated falciparum malaria. Data were analysed by age groups to assess age-specific effect on parasite clearance in patients with G6PD deficiency, since studies have shown that parasite clearance was faster in adults than in children ,21. The The overall prevalence of G6PD deficiency was 17.8% is comparable to other authors finding reported elsewhere in Africa where the prevalence is from 0% to 25% . The preParasite clearance between the different genotypes at day 0 was not statistically significant as median parasitaemia between abnormal and normal G6PD genotypes was not different. The study results have shown that hemizygous and heterozygous had a tendency of elevated median parasitaemia. Similar observation with geometric mean parasitaemia was also reported elsewhere .A comparison of the median parasitaemia at day 0 between normal G6PD patients and G6PD deficient patients in general showed no difference . The study results did not confirm findings by Nkuo-Akenji et al., 2004 who showed a significant difference in the mean parasite density in G6PD enzyme deficient group compared to the G6PD normal group. That study found a mean parasite density of 3.7  for enzyme deficient individuals and 4.4  for enzyme active individuals (p < 0.05) .The study results have shown that G6PD deficiency had no significant association with parasite clearance and median parasitaemia from day 0 to day 3 Figure . The detOther factors such as the molecular biology methods used to determine G6PD deficiency used in our study may also undermine the role of G6PD deficiency in parasite clearance. Enzyme assay appears to better reflect the biological effects of the G6PD enzyme more than genotyping. This was suggested in a recent study which showed that among female G6PD- deficient patients, but not male patients, whose genotype was also confirmed by enzymatic methods to have low enzyme activity as shown by enzyme assay were significantly protected from uncomplicated malaria, while patients with higher levels of enzyme activity were not protected against uncomplicated malaria .No significant association between ACT efficacy and G6PD deficiency was seen in our data Table . Althougin vitro studies.The possibility of reduced susceptibility of parasites to ACT in G6PD deficient patients could not also be ruled out since in a study with \u03b1-thalassemia patients, it was hypothesized that the infected variant erythrocytes could not accumulate as much drug as infected normal erythrocytes 7, 9). Furthermore, the authors had shown that the reduction in artemisinin accumulation in infected variant erythrocytes was due partly to competition with uninfected erythrocytes for the drug, and partly to the lower drug accumulation inside the parasite , 9. Furt. TherefoP = .006) [In contrast another study reported that among hemoglobin E patients treated with artemisinin derivatives, the presence of hemoglobin E trait was associated with an estimated 2.9-fold increase in parasite clearance rate (95% CI, 1.4-6.3; The present study looked at the role of G6PD deficiency in gametocyte carriage and found that the proportion of gametocytes carried was similar in G6PD-deficient and G6PD-normal individuals after treatment at day 3. Similar percentage of gametocyte carriage has also been shown with ACT in the general population in Mali ,28.P. falciparum using ACT in G6PD-deficient patients with uncomplicated malaria may require both enzymatic assay and genetic analysis of the mutation to detect and characterize G6PD deficiency. In clinical trials, whilst enzymatic assays seem to more closely approximate biologic function and correlate with protection, genetic analysis may uncover novel variants causing disease and thereby provide important insights for treatment of uncomplicated malaria.The clearance of The likely explanation for a lack of difference in parasite clearance time between those with and without G6PD deficiency is simply that clearance was rapid in all groups due to the rapid activity of artemisinin derivative, and that, if there was any difference, it could not be discriminated with once-daily assessment.in vitro study and in vivo with several daily assessments may be help to understand the interaction between parasite clearance, ACT and G6PD deficiency.The study findings suggest that G6PD deficiency did not increase or reduce the parasite clearance and the efficacy of ACT. The biological assays that detect G6PD enzyme activity during malaria treatment may reflect the level of G6PD deficiency and, therefore, may more accurately assess parasite clearance in the presence of treatment rather than using PCR methodology to determine G6PD deficiency as in this study. Other confounding factors such as other host erythrocyte variants and host relative immunity may also play a crucial role in G6PD activity. An The authors declare that they have no competing interests.IS, MAT, AD, AKK, AW, JK-M and OKD participated in study design. IS, AKK and MTA contributed in data management and analysis. AKK, SD, and AG did the molecular analysis in determining the status of G6PD deficiency. All authors participated in the preparation of the manuscript and approved the final version."}
+{"text": "PD assessment is facilitated in malaria because serial parasite densities are readily assessed by microscopy, and at low densities by quantitative PCR, so that initial therapeutic responses can be quantitated accurately. If the in vivo MIC could be characterized early in phase 2 studies, it would provide a sound basis for the choice of dose in all target populations in subsequent combination treatments. Population PK assessments in phase 2b and phase 3 studies which characterize PK differences between different age groups, clinical disease states, and human populations can then be combined with the PK-PD observations to provide a sound evidence base for dose recommendations in different target groups.Antimalarial drugs have usually been first deployed in areas of malaria endemicity at doses which were too low, particularly for high-risk groups such as young children and pregnant women. This may accelerate the emergence and spread of resistance, thereby shortening the useful life of the drug, but it is an inevitable consequence of the current imprecise method of dose finding. An alternative approach to dose finding is suggested in which phase 2 studies concentrate initially on pharmacokinetic-pharmacodynamic (PK-PD) characterization and  Plasmodium vivax and P. ovale infections, persistent liver-stage parasites (hypnozoites) cause later relapses, despite cure of the blood-stage infection, which complicates therapeutic assessment. These infections require additional treatment with 8-aminoquinolines . Relapses are often genetically heterologous and cannot be distinguished reliably from recrudescences or new infections. This necessitates a different approach for assessment of treatment efficacy in the relapsing malarias\u2014which is yet to be agreed upon.The primary objective of treating severe malaria is to save life. Other considerations such as preventing recrudescence or minor toxicity are secondary. In uncomplicated malaria, the main objective of antimalarial drug treatment is cure of the infection. Speed of response is also important, as this reflects the rate at which the disease is controlled and the corresponding reduction in the risk of progression to severe malaria. Less-serious adverse effects therefore become a more important factor in determining dose. The therapeutic response in malaria is determined by the concentration profile (pharmacokinetics [PK]) of active antimalarial drug or drugs in the blood , their intrinsic pharmacodynamic (PD) properties, the susceptibility of the infecting parasites to the drug(s), the number of asexual malaria parasites in the blood, and the activity of host-defense mechanisms. Ideally, antimalarial treatment should be 100% effective in everyone, but this may not be possible without producing toxicity or recommending a long course of treatment with consequent poor adherence. It is now recommended that all antimalarial treatments for uncomplicated malaria should aim at a >95% cure rate for the blood-stage infection . In PlasP. vivax infections), mefloquine, halofantrine, artemisinin derivatives, artemether-lumefantrine, and dihydroartemisinin-piperaquine  were all deployed initially at doses which were too low in some or all age groups. Pyrimethamine and sulfadoxine doses for children were extrapolated from experience in Caucasian and Asian adults. Their pharmacokinetic properties were not studied in younger age groups before widespread deployment in Africa, where children are the main target group , should be used. The same should apply to vivax malaria, although chloroquine and primaquine can be considered a combination. When drugs are first developed, there is a limited window of opportunity to define the dose-response (or concentration-effect) relationship for the single new compound, but this opportunity must be taken . It is necessary first to consider the factors which affect the pharmacokinetic properties of antimalarials in malaria and then to consider antimalarial pharmacodynamics and how PK-PD relationships should be assessed.Optimizing drug dosing requires characterization of the pharmacokinetic and pharmacodynamic properties of the drug in the target populations. There are four main determinants of the therapeutic response: antimalarial pharmacokinetics , parasite susceptibility , host defense , and parasite burden. In addition, mixed infections can be a factor. For antimalarials, sistance  and theysistance , but thebe taken . Once th patient . SuggestThe pharmacokinetic (PK) properties of antimalarial drugs are often altered in patients with malaria compared with healthy subjects. The PK properties therefore change as the patient recovers. PK properties are also often significantly different in important patient subgroups such as young children and pregnant women . SeveralMalaria is often worst in remote rural areas. The recent development of simple methodologies such as drug measurement from capillary blood filter paper samples , 26 willIn drug development, where a new compound has not been used previously, there is little information on distributions of PK variables and so the important but difficult issue is to determine how much PK-PD information is enough to decide upon a dosage recommendation. For safety reasons, the PK information is usually gathered in the following standard sequence: experimental animals, healthy normal volunteers, adult patients with uncomplicated malaria, children, and, much later, infants and pregnant women.P. falciparum but having weak activity against its asexual stages  of sensitive P. falciparum, but they do not kill the mature P. falciparum gametocytes (stage V). The artemisinins have a broader range of effect on developing P. falciparum sexual stages, as they also kill stage IV and younger stage V gametocytes. Atovaquone and the antifols kill preerythrocytic stages and have sporontocidal activity in the mosquito (interfering with oocyst formation and therefore blocking transmission). Apart from the 8-aminoquinolines, none of these drugs have significant effects on P. vivax or P. ovale hypnozoites. Even within the asexual cycle there are differences in antimalarial activity in relation to parasite development. None of the currently used drugs have significant effects on very young ring stages or mature schizonts, and all have their greatest effects on mature trophozoites in the middle of the asexual cycle  for that drug in that infection, C is the concentration of drug in blood or plasma, EC50 is the blood or plasma concentration resulting in 50% of the maximum effect, and n is a parameter defining the steepness of the dose-response relationship. For most drugs, maximum effects are probably achieved initially. The evidence for this is the lack of a relationship between peak concentrations and parasite clearance  . For drugs in current use, maximum PRRs range from approximately 10-fold to approximately 10,000-fold reductions in parasitemia per asexual cycle. The mean values and their variance in vivo have not been established for several important antimalarial drugs in current use (notably lumefantrine and piperaquine), and for others, where monotherapies have been evaluated, the estimates are often imprecise. There is no evidence for saturation of parasite clearance, but, obviously, the higher the initial biomass, the longer it takes to eliminate all the parasites from the body  . So whilthe body . Consequex vivo depends on the susceptibility of the infecting parasites and the PD readout . Furthermore, each in vitro method assesses a slightly different section of the asexual life cycle, which may result in important differences between methods in the results for drugs with ring-stage activity. It is not clear exactly how the effects of these static drug concentrations in a small volume of dilute blood in the laboratory correspond with in vivo effects  is recorded. In the absence of in vivo information on the concentration-effect relationship, for predictive modeling purposes the slopes of the linear segments of the in vitro and in vivo sigmoid concentration-effect relationships have been assumed to be similar  and in vivo PK-PD responses in patients with malaria could be characterized, then this would facilitate dose finding.For antimalarial effects, the shape and position of the concentration-effect curve studied  effects , 19, 35. similar , 35, butormation . Human mP. falciparum or P. vivax received different dose regimens, serum levels were measured, and therapeutic responses were assessed. This research provided dose-response or concentration-effect relationships and led to the mepacrine loading-dose regimen, characterization of the comparative antimalarial effects of the four main Cinchona alkaloids , and development of the standard dosing regimen for chloroquine . This was still the era of malaria therapy, and the war had focused military attention on malaria. Such volunteer studies are no longer possible today. Since that time, PK-PD relationships have been inferred mainly from clinical studies of antimalarial treatment  or the related area under the plasma concentration-time curve (AUC) that is the best correlate of bacterial killing . With some adjustments, these PK measures can be applied to antimalarial effects , bacterial killing is dependent on the duration for which the antibiotic exceeds the MIC for the bacterial population (\u201ctime above MIC\u201d). For other antibiotics (where concentrations achieved with current regimens remain on the steep part of the concentration-effect relationship), it is the maximum concentration achieved  of several days or weeks. In order to cure the blood-stage infection in a nonimmune patient, antimalarial concentrations in blood (free plasma concentrations) must exceed the MIC for the infecting parasites until the last parasite is killed  are necessary. Maximum PRR values range from approximately 10,000/cycle for artemisinins against sensitive parasites down to \u223c10/cycle for antimalarial antibiotics (pk represents the parasitemia elimination rate constant derived from the slope of the linear decline in Loge parasite counts (per hour) following the start of treatment.Assuming that the parasites are exposed to the antimalarial drug at a sensitive stage, what duration of exposure in a single asexual life cycle is necessary for maximum effect? For sensitive parasites, it appears that up to 4 h of exposure is required , althougibiotics . All othIn reality, the other factors are seldom equal. High parasitemia levels reflect the particular patient's inability to control the particular infection, and so the host contribution to parasite clearance may be substantially less in these patients than in most patients with lower levels of parasitemia . As a co7-\u221e), is increasingly being measured (7-\u221e are linearly related. For artemisinin combination treatments (ACTs), the initial therapeutic response is determined mainly by the artemisinin component which is given for 3 days, covering two asexual parasite cycles. The partner drug is then responsible for removing the residual parasites  which remain in the third cycle . This assertion is supported by detailed parasite clearance studies  and is cmeasured . Nearly rd cycle , 50 7-\u221e), is fections .in vivo MIC is the critical determinant of the dose and duration of antimalarial treatment, but it has not been well defined for any antimalarial drug. The MIC is the blood or plasma concentration giving a parasite multiplication rate of 1. As drug levels fall below the MIC, then, in the absence of an effective immune response, parasitemia starts to rise again. A recrudescent infection can be detected by microscopy when parasite densities reach approximately 50/\u03bcl. Thus, determination of the MIC requires treatment failure. Model-derived estimates have been proposed for quinine in uncomplicated falciparum malaria , as they should be , then ve0%, as thFor an antimalarial treatment to be reliably  effective, it should, by definition, provide >95% efficacy in a nonimmune population. In areas of high transmission, this refers to the younger children, who bear the brunt of the disease, as older children and adults have acquired higher levels of immunity which contribute substantially to therapeutic responses. Self-cure becomes increasingly likely with increasing immunity . CalibraIdeally, antimalarial treatment should cure everyone thus treated, so drug regimens must be sufficient to cure infections with all prevalent parasites in those patients with the highest parasite burdens. As treatment failure is usually necessary for selection and spread of resistance to occur, it is important that dose regimens are sufficient to ensure very high cure rates , 3, 22. Plasmodium falciparum (A dose (either as a single drug or a combination) that achieves cure in 95% of treated patients might be considered most cost-effective and is a minimum requirement for a new treatment. However, it may be prudent in terms of overall benefit to aim for a higher dose range and thus a higher cure rate if possible. For some drugs , resistance develops stepwise in a predictable sequence. Emergence of resistance should be slowed if the chosen dose regimen is also effective against parasites with low-level resistance , 41. Thelciparum . In consWhile these arguments favor higher dosing as a resistance prevention strategy, there is an alternative view. It has been argued, on the basis of artificially balanced mixed infection studies in rodents, that low doses should be deployed intentionally in order to reduce the fitness disadvantage that drug-sensitive parasites have in the presence of antimalarial drugs and to exploit the competition between different genotypes that limits individual growth . Drug-sein vitro antimalarial activity, dihydroartemisinin  is equivalent in molar terms to artesunate , but in weight terms there is a 35% difference in activities. Artemether is less active in vitro than artesunate, although after oral administration approximately two-thirds of the total antimalarial activity is provided by its metabolite dihydroartemisinin  against well-characterized parasites in acute malaria infections and optimum doses chosen. An evidence-based dose regimen can then be used in phase 3. If possible, the two drugs should be combined at doses which are individually curative. Clearly, for currently recommended first-line ACTs, and for atovaquone-proguanil treatments, this is not the case. In ACTs, the artemisinin component is given for 3 days . The slowly eliminated partner drug doses have been chosen at doses which should provide high cure rates in sensitive infections. For piperaquine, amodiaquine, and lumefantrine, this requires three-day dosing in order to load sufficiently to provide therapeutic concentrations  for long enough to eliminate all the parasites. In combinations in which one component is eliminated much more rapidly than the other (such as ACTs), it would seem prudent to aim initially, based on PK-PD studies, for doses of the more slowly eliminated partner drug which are curative. PK-PD studies can then be conducted with the combination to characterize any synergy (or antagonism) with the combination and thus characterize whether the combination dose predicted from the individual drug-dose-finding studies requires adjustment.Before evaluation of the PK-PD properties of a combination, the PK-PD relationships for the individual drugs should be determined In cases of severe malaria, antimalarial drug treatments must aim to kill the infecting parasites as quickly as possible  , 16, 68.P. falciparum in Africa, to the more rapid time course of disease, and even to differences in pathological processes, recent studies suggest a more prosaic explanation, i.e., misdiagnosis in a significant proportion of African children considered to have severe malaria , rates of resolution of renal impairment in adults, or attenuation of the fall in hematocrit, are good correlates of mortality benefits remains to be established. In general, the laboratory variables  have performed better than the clinical measures in clinical trials. In the randomized controlled comparisons of artesunate or artemether versus quinine, which are by far the largest trials ever performed in severe malaria, serial disease severity measures were often worse in the patients receiving artemisinin derivatives despite the substantially lower mortality associated with these treatments  parasites. The ideal treatment would provide a reliable single-dose cure, but this requires providing reliably PRR values close to >103/cycle for 9 days (5 cycles) or >104/cycle for 7 days (covering 4 cycles) against all prevalent parasites.Several antimalarial drugs are in the late stages of development or have entered clinical trials. Arterolane (Rbx11160) is registered in India, and OZ439 and the spiroindolone KAE609 are in phase 2 studies. As most currently available antimalarials are well tolerated, these new drugs will have to match this good acceptability and toxicity profile while being highly efficacious. There is now a critical window of opportunity early in clinical development, before the new drugs are combined with partner antimalarials, in which to identify optimum doses . Dose fiin vivo in humans (a proof of concept). The initial dose will have been chosen on the basis of the doses used in the phase 1 studies, which in turn will have been extrapolated from animal and in vitro studies. If this is a critical decision point (\u201cgo/no go\u201d), then a predicted \u201chigh\u201d dose will probably be chosen for the \u201cconfirmation of antimalarial activity in vivo in humans study\u201d to avoid a false-negative result and unwarranted discontinuation of the drug's development. Volunteer patients in phase 2 studies need to be fully informed of the experimental nature of the study they may participate in, the risks involved, and the absolute need to stay under medical supervision until they have cleared parasitemia and are clinically well. Physicians and nurses need to know the risks and to have low thresholds for providing known effective rescue treatment in cases of clinical deterioration.Optimized dosing should limit the emergence and spread of resistance and maximize cost-benefit . The iniin vivo in humans\u201d phase 2 studies which are safer than first use in patients with symptomatic falciparum malaria.P. vivax malaria could be studied initially. This would provide in vivo data and useful information on PK-PD relationships in disease. P. vivax malaria also has the advantage that it does not sequester, and so, unlike P. falciparum infections, peripheral blood parasite counts correlate closely with the infecting parasite burden, making initial interpretation of parasitological responses more straightforward.Instead of falciparum malaria, the less dangerous The first volunteers could be adults with asymptomatic falciparum malaria. However, asymptomatic infections are generally highly sensitive to any intervention because of the effective immune responses which controlled the infection in the first place. This approach is therefore likely to overestimate the drug effect substantially. Furthermore, parasitemia levels are always low in asymptomatic adults, providing relatively little information on the kinetics of parasite clearance. Finally, the pharmacokinetic properties of the new drug would not reflect acute illness effects.in vivo in humans.\u201dVolunteers can be artificially infected with a known drug-sensitive malarial \u201cstrain\u201d and the evaluation conducted entirely at parasitemia levels below the pyrogenic density using very sensitive quantitative PCR (qPCR) methods , 77\u201379. Initial phase 2 antimalarial studies with new antimalarial compounds should always be performed in nonpregnant adults. Falciparum malaria is inevitably the primary focus, but it is a dangerous disease. Despite the extensive experience with artificially induced malaria in humans, ethics review boards and investigators may be reluctant to take the risk of conducting experiments with an untried medicine in symptomatic patients with falciparum malaria even in intensively supported facilities. There are three alternative approaches to conducting initial \u201cconfirmation of antimalarial activity in vivo in humans is satisfactorily obtained, the conventional approach to dose finding is to reduce doses steadily in small groups of patients until there is clear evidence of a reduced antimalarial effect. This has meant comparing recrudescence rates at different doses\u2014a costly and time-consuming process. Relatively large numbers are needed for adequate precision, so estimates are inevitably imprecise (see below). Even then, the lowest dose providing \u201csatisfactory cure rates\u201d applies to a particular set of patients  with a particular set of infecting parasites.Whichever approach is chosen, frequent antimalarial blood concentration measurements should be made in an attempt to characterize the PK-PD responses, as this would help inform design of the definitive dose-finding studies. Once confirmation of antimalarial activity P. vivax infections and 7-day regimens for P. falciparum and yet ACTs are prescribed in 3-day regimens. These provide maximum parasite killing effects during two asexual cycles but still leave up to 100,000 parasites for the partner drug to remove. All finally recommended treatments should ideally provide antimalarial concentrations which exceed the MPC for \u22654 parasite life cycles (this is necessary to cure infections of >1012 parasites with a PRR of 103 per cycle)  and the overall curative efficacy  , 23. Ther cycle) , 33.With new or experimental treatments, clear clinical and parasitological criteria for treatment failure and patient rescue with an effective antimalarial treatment should be predefined . Frequent sampling for antimalarial drug concentration profiling is needed so that confident assessments of key PK variables can be made. If the PD readout is slowing of parasite clearance , then, as described earlier, a second drug such as mefloquine or atovaquone-proguanil may be given simultaneously with or shortly after administration of the first drug to ensure cure . Althougde novo resistance (12 parasites in the body), which is considerably higher than that in the entry criteria for dose-finding studies. If there is no other PK-PD characterization, then treating different groups of patients with different doses and recording cure rates is likely to be an imprecise and rather expensive method of dose optimization which results in dose recommendations which are too low.For slowly eliminated drugs, the issue is to determine how much drug in total is needed to cure reliably; for rapidly eliminated drugs, it is to determine for how many days the drug should be given. If there is capacity-limited (saturable) absorption of a drug , then dose spacing is also a consideration. In a conventional approach to dose finding, the doses need to be reduced early in the phase 2 studies to determine which concentration profiles are necessary for cure. This inevitably means observing recrudescences\u2014although these may be detected and treated before they become symptomatic if sensitive qPCR parasitemia monitoring is conducted. As new antimalarial drugs should achieve at least 95% efficacy and should no longer be recommended when cure rates fall below 90%, obtaining precision around such high proportions of efficacy requires very large sample sizes . Dose fisistance . Dose reLarge variations in antimalarial blood concentrations may confound estimates.For drugs with large distribution volumes and low clearance, loading may require multiple doses, so dose-response relationships estimated from initial concentration profiles may be different from those estimated after loading.Large interpatient variations in parasite clearance result in imprecise estimates.Dangerous complications may arise in undertreated patients.These approaches identify only the MPC, or another PK correlate, but not the MIC.Drug absorption lag times may confound estimates.If a second drug is given concomitantly for safety reasons, then individual drug effects cannot be separated reliably.(i) If the parasite clearance rate is used as an endpoint:Large sample sizes are required as groups, not individuals, are compared.Dangerous complications may arise in undertreated patients.PK correlates may be inaccurate; this approach does not identify the MIC or MPC.Patients with high parasite burdens have the highest risk of treatment failure, and yet they are systematically excluded from evaluations of drugs in development.(ii) If the cure rate is used as an endpoint:in vitro susceptibility of the infecting parasites and the therapeutic response is not well characterized.In both cases, the relationship between the in vivo MIC as the primary objective of phase 2 studies. If the in vivo MIC can be defined, then comparison of in vivo MIC  estimates with in vitro parasite susceptibility measurements from the same malaria infections would allow calibration of the large body of in vitro data on drug susceptibility that can be generated rapidly from parasite isolates already obtained from across the world. The inhibitory concentration in vitro which corresponds with the MIC and MPC can be identified and the corresponding values for the most resistant parasite isolates extrapolated. These assessments of variance in parasite susceptibility (PD) can then be melded with estimates of PK variance from population pharmacokinetic studies conducted in different patient populations to predict the dosing regimens necessary to ensure cure if the most resistant natural isolates infected the patients with lowest drug exposures.An alternative approach to dose finding is to define the in vitro susceptibility tests performed with a large number of P. falciparum  isolates across different geographic regions  exceed the MIC in malaria, parasite numbers decline, so a generally efficacious regimen is one that maintains blood concentrations above the MIC in all patients until complete elimination of even the most drug-resistant naturally occurring infection. The MIC is a useful theoretical concept, but its utility in dose finding and other aspects of therapeutic assessment remains to be defined. It is not known how variable it is. As the MIC represents both drug and host effects, we do not know yet how much it is affected by the host response even in nonimmune patients. Host contributions to parasite clearance in malaria reflect the duration and severity of the initial infection and so are likely to be lowest early in acute infections in previously unexposed individuals.Defining the in vivo MIC against fluctuating drug concentrations, either for a rapidly eliminated drug such as an artemisinin derivative or for a drug for which distribution determines initial concentration profiles and antimalarial effects (such as chloroquine or piperaquine), is more difficult or may not be possible  and do not have prominent distribution phases. This is because the important confounders such as variable stage susceptibility within the asexual cycle, or second-cycle effects, operate within a narrow range of slowly declining blood concentrations so that the estimate is more precise . Estimatpossible .in vivo concentration-effect relationship would be to give a single relatively low dose of the new antimalarial compound to volunteer patients during the initial dose-finding studies and then to follow the blood concentrations and the parasite densities frequently using microscopy and then sensitive validated quantitative PCR methods  and parasite densities at a time when the multiplication rate is 1. This would usually precede by days or weeks a subsequent recrudescence. One approach to characterize the  methods , 79\u201381.  methods . It is a methods . A major rapidly . As prooWhen the parasite multiplication factor falls to 1 at a total body parasite biomass of less than approximately 100,000 parasites, then parasite densities cannot be quantitated, and so the MIC cannot be measured directly  . The dosIt is now generally agreed that in order to prevent the emergence and spread of resistance, antimalarial drugs should be deployed in combinations. How then should the individual components be evaluated, when they will never be deployed as single drugs? The default position often quoted is that each component should be evaluated independently, and the curative dose established, before putting the drugs together in combination. This is not always necessary for rapidly eliminated drugs. Indeed, the results of such an approach may be misleading. For example, artemisinin combination treatments (ACTs) all comprise a noncurative dose regimen of artesunate, artemether, or dihydroartemisinin. The value of assessing cure rates with these drugs (further complicated by drug-specific parasite dormancy) in determining dose regimens in ACTs is dubious. Furthermore, even if drugs have well-matched pharmacokinetic properties and have no PK or PD interactions, the doses of each component in a combination required to produce >95% cure are nearly always less than those of the drugs used alone; i.e., if the drugs have unlinked distribution and elimination pathways and different mechanisms of action, then in >95% of the occasions when concentration profiles of one drug are insufficient to cure, the other drug will be curative. The exact magnitude of the combination advantage depends on the PK-PD distributions for each drug. The alternative PK-PD approach detailed above with assessment of pharmacokinetic interactions and pharmacodynamic synergy and antagonism should obviate costly and time-consuming assessment of individual drug cure rates at different doses.Investing in characterization of antimalarial drug pharmacokinetic-pharmacodynamic relationships will probably improve current antimalarial dose regimens and will certainly increase the likelihood of choosing an optimum regimen for newly introduced antimalarials."}
+{"text": "Drug resistance to anti-malarials is a major public health problem worldwide. This study aimed at establishing the efficacy of artemether-lumefantrine (ACT) in Igombe-Mwanza, north-western Tanzania after a few years of ACT use, and establish the prevalence of mutations in key targets for artemisinin, chloroquine and sulphadoxine/pyrimetamine (SP) drugs.Pfcrt, Pfmdr1, Pfdhfr, Pfdhps and Pfatp6 were detected using PCR-RFLP methods established previously.A prospective single cohort study was conducted at Igombe health centre using artemether-lumefantrine combination therapy between February 2010 and March 2011. The follow-up period was 28 days and outcome measures were according to WHO guidelines. Blood was collected on Whatman filter paper for DNA analysis. DNA extraction was done using TRIS-EDTA method, and mutations in pfcrt 76 and pfmdr1 86 positions being 87.8% and 93.7% respectively. Mutant parasites predominated at pfdhfr gene at the main three positions 108, 51 and 59 with prevalence of 94.8%, 75.3% and 82.5% respectively. Post-treatment parasites had more wild types of pfdhps at position 437 and 540 than pre-treatment parasites. No mutation was seen in pfatp6 769 in re-infecting or recrudescing parasites.A total of 103 patients completed the 28 days follow-up. The mean haemoglobin was 8.9 g/dl (range 5.0 to 14.5 g/dl) and mean parasite density was 5,608 parasites/\u03bcl. Average parasite clearance time was 34.7 hours and all patients cleared the parasites by day 3. There was no early treatment failure in this study. Late clinical failure was seen in three (2.9%) patients and late parasitological failure (LPF) was seen in two (1.9%). PCR-corrected LPF was 1% and adequate clinical and parasitological response was 96%. The majority of parasites have wild type alleles on pfcrt 76 K and pfmdr1 86 N was high in the study area while markers for SP resistance is still high. Artemether-lumefantrine may be selecting for wild type alleles on both positions (437 and 540) of pfdhps.The efficacy of artemether-lumefantrine for treatment of uncomplicated malaria is still high in the study area although the rate of re-infection is higher than previously reported. Parasite clearance after 48 hours was lower compared to previous studies. The prevalence of wild type allele  Plasmodium parasites, especially Plasmodium falciparum which causes more than 90% of infection in sub-Saharan Africa. Although there are reports of decreasing paediatric malaria infection [et al, 2009 reports that analysis of studies in East Africa shows that the parasites were being controlled less well by the artemisinin component of ACT in 2007/2008 studies than in 2005/2006 [pfcrt 76 K and pfmdr1 86 N) with a concomitant reduction in susceptibility to lumefantrine [pfcrt has been reported in Malawi with restored sensitivity to CQ [Drug resistance to anti-malarials is a major public health problem worldwide . In 2006nfection ,6 and bunfection ; malarianfection . Data frnfection . Clinicanfection -11. Hunt005/2006 . As thes005/2006 -16. Treafantrine . Even bety to CQ ,19. It iThis was an interventional prospective single cohort study conducted at Igombe health centre in the vicinity of Mwanza city in Tanzania. In this area malaria is mesoendemic and the catchment area for Igombe health centre is a semi-urban, with a population of around 40,000 inhabitants.Patients with fever, aged between six and 60 months who attended the clinic during the study periods (between February 2010 and July 2010 and between October 2010 and March 2011) were screened for malaria parasites. Detailed medical history, clinical examination and both thick and thin blood films were done after obtaining an informed consent from the parents or guardian. Recruitment was based on inclusion criteria set by WHO [\u00ae, Novartis, Basel, Switzerland) was used to treat recruited patients. A first dose was given as a direct observed therapy (DOT) and the next doses was supplied to the parents/caretakers for giving to patients at eight hours from the time of the first dose and morning and evenings of successive two days. For patients weighing from 5 kg to less than 14 kg a single tablet (20 mg/120 mg artemether/lumefantrine) was given, those from 15 kg to less than 24 kg received two tablets, those with 25 to less than 34 kg received three tablets. Patients with fever and axillary body temperature \u2265 38.5\u00b0C were given paracetamol and those with anaemia (Hb between 5 and 9 g/dl) were given haematinics. If a patient vomited the treatment drug within 30 minutes the dose was repeated and those who vomited more than once were excluded from the study and changed to parenteral quinine. Patients were followed up on days 1, 2, 3, 7, 14, 21 and 28. Patients were reminded of their visiting dates by mobile phones if the parents owned one, or through the 10 cell-leaders' phones; those who did not turn up on the scheduled days were visited by a member of the research team. A patient was withdrawn from the study if the follow-up was not complete and could not be traced the following day and these included patients who travelled to other places. Also use of other anti-malarial drugs or non-compliance especially to the second dose at eight hours led to withdrawal of patients from the study.A six-dose regimen of artemether-lumefantrine  of one of the following during days 4 to 28: danger signs, severe malaria or an axillary temperature of 37.5\u00b0C or higher in the presence of parasitaemia.LPF was defined as presence of parasitaemia on any day from day 7 to day 28 without signs of severe disease or fever and not previously meeting criteria of ETF or LCFACPR was defined as absence of parasitaemia on day 28 irrespective of axillary temperature without previously meeting any of the criteria of ETF, LTF or LPFThe study was approved by the Joint Weill-Bugando ethical clearance committee and informed consent was obtained from the parents/guardians of all patients.On day of recruitment finger prick was done aseptically and blood was spotted on Whatman no 1 filter paper, thick and thin blood smears were done and stained with Giemsa. A venopunture was done and 3 ml of blood was taken in EDTA vacutainer for full blood count (including haemoglobin estimation) using Cell Dyn 3700 machine (Abbot Laboratories USA). On follow-up only, finger prick blood was collected and spotted on filter papers.pfcrt position 76 were determined using primers and RFLP performed as explained elsewhere [Pfmdr1 gene was also amplified by nested PCR and RFLP done according to protocols published before [pfdhfr, outer PCR was done using primer pfdhfr1 and pfdhfr2 and nested PCR was done using primer pfdhfr3 and pfdhfr4 for position 108 and pfdhfr5 and pfdhfr6 for nested reaction on position 51 and 59  were found positive for malaria parasites but only 108 met the inclusion criteria and were recruited  and females were 51 (49.5%) and the average age was 38.7 months. The mean haemoglobin was 8.9 g/dl (range 5.0 to 14.5 g/dl), and mean parasite density was 5,608 parasites/\u03bcl .Average parasite clearance time was 34.7 hours, 70 (68%) cleared parasites within the first 24 hours and cumulatively 91 (88.4%) patients had cleared the parasites at 48 hours. The remaining 12 (11.7%) patients had positive blood slide at day 2 and they were all cleared of parasites by day 3  patients, LPF was seen in two (1.9%) and ACPR was achieved in 98 (95.1%) patients before correction with PCR. PCR corrected LPF was 1% and ACPR was 96% Table .msp2 patterns.A high number of subpatent infections were detected by PCR (while looking at the drug resistance markers) but not microscopy on both day 14 and day 28 raising the prevalence to 15 (14.6%) and 13 (12.6%) respectively. All subpatent infections were due to re-infections as shown by different pfcrt gene (success rate 87.4%) and among these, the majority 80 (88.9%) were wild type and only 10 (11.1%) were mutants. On follow-up samples, after treatment with AL, 12 samples were successfully amplified on day 14 and 28, of which a majority, eight (66.7%), were wild type alleles and four (33.3%) were mutant alleles. The difference between pre-treatment samples and re-infections could not be shown to be significant   and pure mutants were only six (6.3%). Among the 19 successfully amplified post-treatment samples, the wild type alleles were still in majority at 11 (57.9%) but the proportion of mutants had increased to 42.1%, which is a significant change . At position 108, 92 (94.8%) were mutant while four (4.3%) were wild type and one (1.1%) was a mixed infection.For At position 51, the majority 73 (75.3%) were mutant while 12 (12.4%) were wild type and the other 12 (12.4%) were mixed with more mutants.pfdhfr gene in pre-treatment samples was high with 59 samples (64.1%) while 27 (29.4%) had double mutation with various combinations as illustrated in Table At position 59, 80 (82.5%) were mutant and nine (9.3%) were wild type, while eight (8.2%) were mixed with more mutants. The prevalence of triple mutants in In post-treatment samples, mutants were predominant but at a lower percentage than at day 0: 21/29 (74.2%), 20/29 (69%) and 15/29 (51.7%) on position 108, 51 and 59 respectively. In the post-treatment samples the triple mutants were 13/29 (44.8%) and double mutants were 7/29 (24.1%)  were mutants while 15 (15.6%) were wild type and 10 (10.4%) were mixed (with mutants predominating). Double mutants in pfdhps were 65 (68.4%) out of 95 samples, which amplified at both positions. With the exception of one sample, all mutants at position 540 were also mutants at position 437. There were more wild types at both positions in pfdhps in the follow-up samples at day 14 and 28 after treatment with ACT. Prevalence of wild types were 10/17 (58.8%) and 13/21 (61.9%) at positions 437 and 540 respectively.A total of 95 samples were successfully amplified at position 540 and 66 (69.5%) were mutants, while 15 (15.8%) were wild type and 14 (14.7%) were mixed with more mutants. At pfatp6 at position 263 in either pre-treatment or post-treatment samples. At position 769, mutations were screened only in re-infection and recrudescing parasites and no mutation was detected in these follow-up samples.There was no mutation in et al, showed similar high frequencies of LPF as shown here [et al [The efficacy of AL in this study area was high at 95.1% (PCR uncorrected) and 96% PCR corrected. This is similar to what has been found in other places of Tanzania and Sub-Saharan Africa in general where cure rates ranged from 96 to 100 in children -11,28. Town here . The meee [et al , presente [et al . In thispfcrt 76 K and pfmdr186N) is high in this study area compared to other studies done previously in Tanzania. This is a good indicator of return of CQ-sensitive parasites as was shown in Malawi study where Laufer et al, found good in vivo sensitivity to CQ following a study that revealed decline in mutants (pfcrt76T) at this position [pfcrt mutations was reported to be high more than five years after discontinued use of CQ [pfcrt and pfmdr1 in subpatent re-infections found in follow-up samples. This contrasts with most earlier studies, but a similar tendency for pfcrt is observed in Ngasala et al, where at least no significant difference in pfcrt frequencies were found in re-infections [pfcrt and pfmdr1 mutations indicates that it will be very difficult to completely reverse CQ resistance once the resistance has been established and reusing CQ may lead to rapid emergence of CQ resistance. Any attempt to reuse CQ should only be done under proper controls where the in vitro susceptibility can be monitored.The prevalence of parasites with wild type alleles  of pfdhpsThe efficacy of artemether-lumefantrine for treatment of uncomplicated malaria was still high in the study area although the rate of re-infection was high. There was a high number of patients who got subpatent malaria infection after treatment with ACT, which was diagnosed by PCR later at day 14 or 28. The prevalence of The authors declare that they have no competing interests.EK, SJ and GS carried out the molecular genetic studies and drafted the manuscript. MM, JK and EK carried out field study, patients follow-up and drafted the manuscript. GK, FK and GS conceived the study, and participated in its design and coordination. All authors read and approved the final manuscript."}
+{"text": "The area along the Thai-Cambodian border is considered an epicenter of anti-malarial drug resistance. Recently, parasite resistance to artemisinin-based therapies has been reported in the area. The artemisinin resistance containment project was initiated in November 2008, with the aim to limit resistant parasites and eliminate malaria in this region. This study describes the response to artemisinin-based therapy among falciparum malaria patients in the area, using data from the malaria surveillance programmed under the containment project.Plasmodium falciparum-positive patients during January 2009 to December 2011 were obtained from the electronic malaria information system (eMIS) Web-based reporting system. All P. falciparum cases were followed for 42\u2009days, as the routine case follow-up protocol. The demographic characteristics of the patients were described. Statistical analysis was performed to determine the cure rate of the current standard anti-malarial drug regimen--mefloquine-artesunate combination therapy (MAS). The proportion of patients who remained parasite-positive at each follow-up day was calculated. In addition, factors related to the delayed parasite clearance on day-3 post-treatment, were explored.The study was conducted in seven provinces of Thailand along the Thai-Cambodian border. Data of P. falciparum-positive cases were reported during the study period. Almost 70% of falciparum cases received MAS therapy . The majority of cases were males, aged between 31 and 50\u2009years. The overall MAS cure rate was >90% over the three-year period. Almost all patients were able to clear the parasite within 7 to 14\u2009days post-treatment. Approximately 14% of patients undergoing MAS remained parasite-positive on day-3. Delayed parasite clearance was not significantly associated with patient gender, age, or citizenship. However, delayed parasite clearance varied across the study area.A total of 1,709 Anti-malarial drug-resistant parasites should be closely monitored in the area along the Thai-Cambodian border. Although the MAS cure rate in this study area was above 90%, an increasing trend of treatment failure has been reported in neighboring parts. Effective malaria surveillance is an important component to monitor drug-resistance in the malaria containment project. Plasmodium falciparum infections in the Greater Mekong Sub-region (GMS) . Among patients managed with MAS in this study, about 30% and 15% still had parasitaemia on day-2 and day-3, respectively. Almost all patients were parasite-free on day-7, with about 2% showing parasite re-appearance during 28-day follow-up. In Pailin, western Cambodia, percentages of parasitaemic patients on day-2 and day-3 undergoing artesunate monotherapy or mefloquine-artesunate treatment were as high as 73% and 55%, respectively [7]. A recent study with high-dose artesunate monotherapy conducted in Battambang Province, western Cambodia, also showed, after treatment, a relatively high proportion of parasitaemic patients on day-2 and day-3  [18]. The proportion of parasite-positive patients observed in the study area is relatively high compared to other locations in western Thailand and in Vietnam .Experience over the past decades has shown that artemisinin can rapidly reduce parasite load, clearing them from blood generally after a two-day intake of treatment [17]. However, in case of suspected in-vitro studies indicated that artemisinin resistance is suspected by exploring parasite clearance time rather than any sudden change in therapeutic cure rate [Unlike the resistance patterns of other anti-malarial drugs, the results of clinical and ure rate ,9. On thure rate . In thisSome limitations have to be considered in this study. The pattern of parasite clearance was presented as the cumulative proportion of patients who remained parasitaemic by day after treatment. Parasite clearance curve that shows reduction of parasite load by exploring additional specific periods of time may provide more information on parasite stage-specific response to artemisinins . HoweverAbout 32% of overall falciparum malaria in this area did not receive MAS treatment. These were a combination of patients who received other second-line drugs, and others who had missing information on anti-malarial drugs received. Since there are no commercial anti-malarial drugs available in Thailand, those missing drug information were likely to have received their treatment in hospitals where standardized reporting forms were not used and computerized, as in peripheral malaria units. Unlike malaria patients who were originally detected at malaria clinics/posts under BVBD supervision, where patients\u2019 details would be directly recorded into eMIS, details of cases at local hospitals were captured later into eMIS after admissions or treatment. Malaria treatment provided at hospital could be either first-line regimen (MAS), or second-line regimens . Further improvement in the reporting system is needed to assist data sharing between hospitals and public health sectors.In this study, the follow-up rate on day-42 was relatively high (50%), considering constraints linked to field operation. About 90% of patients who received complete follow-up on day-7 were followed up on day-42. This allowed the study to estimate a 42-day cure rate, which is recommended to be long enough to monitor MAS resistance due to a long elimination half-life of Mefloquine [Plasmodium falciparum resistance to anti-malarial drugs has always been a major threat in the region, and globally, in malaria control and elimination [P. falciparum resistance is critical in this region.mination . In areamination ,14. AlthContainment of parasites developing resistance to anti-malarial drugs is one of the major goals supported by countries and partners in the Mekong region, as well as globally, to progress from malaria control towards elimination. Malaria surveillance, with a focus on monitoring resistance, is critical to achieving such targets. The electronic Malaria Information System (eMIS) has been effectively set up and implemented to increase the performance of the existing national surveillance system. However, case detection at operational level and data sharing between units and organizations remains a day-to-day programmatic challenge, as maintaining the system requires cross-checking computerized information, then taking appropriate decisions.The authors declare that they have no competing interests. Development of the eMIS was supported by the malaria containment initiative of the WHO and the Bill & Melinda Gates Foundation.SL, PS(1), WS, JK designed the study. SL, JK performed statistical analysis. PS(1), AK, SS, TP, AS worked on the design of eMIS and the applications module, and monitored and maintained the module\u2019s implementation, and extracted data for analysis. PS(1), WS, SV were responsible for managing and supervising the overall malaria control programmer\u2019s activities. AK, JK, DC were in charge of monitoring the progress of eMIS applications. SL, JK, WS, DC, PS(2) drafted the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria in Southeast Asia but has not been tested in Cambodia.Cambodia stopped using co-blistered, non-fixed, artesunate-mefloquine (ASMQ) in 2008 when treatment failure rates approximated 20%. Fixed dose combination (FDC) ASMQ is efficacious against acute uncomplicated, drug resistant P. falciparum. Daily administered FDC ASMQ for three days was dosed by age. Genotyping of isolates at day 0 and day of recrudescence by polymerase chain reaction (PCR) classified post-treatment recurrent falciparum parasitaemia. Ex vivo drug sensitivity testing ([3H] hypoxanthine method) was performed on baseline parasites and reported as the drug concentration inhibiting 50% parasite growth vs no drug (IC50).A 42-day WHO therapeutic efficacy study (TES) was conducted in 2010 in Oral, Kampong Speu province, south-west Cambodia, in patients with acute uncomplicated 50s ranged from 11.5-238.9 (median 58.6) nM.Recruited patients numbered 45; five aged <15 years. On day 3, five of 45 [11.1 (3.7-24.05)] % patients were still parasite-positive; one of whom later failed treatment on day 21. There were 5/45 (11.1%) late treatment failures on day 21, 28 and 35; all were PCR diagnosed recrudescent infections. The day 0 MQ IC50s. Efficacy testing of FDC ASMQ should continue in Cambodia and be considered for reintroduction if efficacy returns.This TES demonstrated reasonable efficacy in an area of possible reduced artemisinin sensitivity and high MQ IC Plasmodium falciparum was non-fixed artesunate-mefloquine (NF-ASMQ). Prior to this, there had been a long history of anti-malarial drug resistance, starting with chloroquine hypoxanthine method) and a higher mean IC50 was reported in resistant infections (90 nM) vs sensitive (56 nM) infections and 8.4 nM (3.6-27.5) [1\u2013143.4], respectively.This standard WHO TES was condNovember. The stueference) from thThe study was approved by the National Ethic Committee for Health Research of the Ministry of Health, Cambodia.P. falciparum between 1,000 and 200,000 asexual parasites/\u03bcL; and, (iv) were aged two years and above.Patients who attended the Oral Health Centre were enrolled if they met all of the following inclusion criteria: (i) informed consent was signed; (ii) they had a documented fever or a history of fever within the previous 48 hours; (iii) microscopic Exclusion criteria included: (i) pregnant or breast feeding women; (ii) unable to complete the follow-up; (iii) allergic to AS or MQ; (iv) a neuropsychiatric contraindication to MQ, namely, epilepsy, history of a severe psychiatric illness, e g, bipolar affective disorder, psychosis, anxiety neurosis; (v) presence of any danger signs or any sign of signs of severe malaria; (vi) sePatients who were malaria slide-positive were asked if they were interested to join a research study. After informed consent was signed, they were assessed to see if they met the criteria for study enrolment. This involved a medical history, physical and blood examinations for a repeat malaria slide (Giemsa-stained and read in the field), haematocrit, total white cell count and blood glucose.Parasite counts were determined on Giemsa-stained thick films and recorded as the number of parasites per 200 white blood cells. Two qualified microscopists read independently all malaria slides and parasite densities were recorded as the average of these two counts. Slides were to be reread if the parasitaemia difference was >50%. The parasitaemia/\u03bcL was determined by multiplying the parasite count/200 white cells by 40. A slide was declared negative after reading 100 thick film fields.msp 1, msp 2) and the glutamate rich protein (glurp) hypoxanthine isotopic method hypoxanthine was inhibited, compared to the drug-free control wells. These IC50s are determined by non-linear regression using the ICEstimator % patients were still parasite-positive; one of whom later failed treatment on day 21. Their day 0 day 3 parasite counts (N/\u03bcL) were: 13,067-270, 14,074-953, 30,718-1872, 46083\u2013434, and 108,013-134.The PCT ranged from two to seven days for a median of three days. All febrile patients at presentation (n\u2009=\u200939) were afebrile by day 2 for a median FCT of 24 hours.Gametocytes were not detected on days 0 to 21 and 35 and 42; 4/44 (9.1%) patients had gametocytes on day 28.ASMQ was well tolerated acutely. There were no patients with drug-induced vomiting who needed rescue treatment. Over the first three days, seven patients reported adverse events, five of whom had received >24 mg/kg of mefloquine. Two (4.4%) patients complained of dizziness, five (11.1%) complained of nausea, and five (11.1%) complained of palpitations. Of these, one complained of all three symptoms and three complained of nausea and palpitations. None required treatment.ex vivo testing. These 18 patients had similar baseline characteristics and received similar drug doses as the 27 who did not undergo ex vivo testing. Excluding the untested patients with parasite counts\u2009\u2264\u20095,000/\u03bcL, the median baseline parasite counts were not significantly different similar (p\u2009=\u20090.37): 39,815 (n\u2009=\u200918) vs. 20,103/\u03bcL (n\u2009=\u200919). The median IC50s (including IQR and range) are presented in Table\u00a050 values varied from four-fold (piperaquine) to approximately 60-fold (CQ). Of the 16 ASMQ treated patients with IC50 data, only one was classed as a treatment failure: 55.3 nM. The IC50 values in patients with ACPR ranged 11.5 to 238.92 for a median of 58.74 nM. The results were almost the same when stratifying by day 3 positivity.Among the 45 isolates collected, 18 met the criteria for in vitro IC50 cut off values for distinguishing resistant from sensitive parasites : (i) 14/16 MQ tested parasites had evidence of in vitro resistance: median (range) IC50\u2009=\u200965 nM (35\u2013239); (ii) 5/13 (quinine) IC50\u2009=\u2009666 nM (603\u2013725); and, (iii) 13/16 (CQ) IC50\u2009=\u2009209 nM (138\u2013302).Using WHO suggestedin vitro in vitro data suggesting reduced MQ sensitivity and where the day 3 positivity rate exceeded the original 3% cut-off.in vivo investigation in which the parasite clearance half-life should be measured. The current in vitro methods are useful to track trends over time but are insensitive for detecting artemisinin resistance. However, a new in vitro test, the RSA [Malaria control programmes should be aware that the day 3 positivity rate is an indicator of the possibility that parasites may be artemisinin resistant and thatl Assay), is showl Assay).Pfmdr-1 copy number was not counted. This molecular marker of reduced MQ sensitivity[Pfmdr-1 copy number declined markedly from 33% in 2005 to 5% in 2007[The study had limitations. The number of patients enrolled in the TES was quite small, 45, due to the low malaria burden and this reduces the strength of the study conclusions. Most patients recruited were adult males, consistent with the malaria epidemiology in other areas of Cambodia. Follow-up extended to day 42 so the true failure rate may have been a little higher had follow-up extended to 63 days, the recommended follow-up time for long half drugs such as mefloquine. These lsitivity is anothin vivo and in vitro status of FDC-ASMQ in Oral. The 11% failure rate is not discouraging and CNM should continue to monitor FDC-ASMQ efficacy, preferably to day 63, in conjunction with in vitro data and Pfmdr-1 copy number to see if FDC-ASMQ can be recommended in the future for treating symptomatic patients. FDC-ASMQ should also be assessed in PCR positive asymptomatic individuals for a potential role in malaria elimination.To conclude, this TES has benchmarked in the The authors have declared that there are no competing interests.In vitro drug sensitivity testing was done by DM and previous in vitro data were given by PL and FA. The study was coordinated and monitored by VN and JRK. WRJT, RL and DM analysed the data and wrote the first draft of the paper. All authors have seen and approved the final manuscript.The study PI was LR. LR and JRK adapted the WHO TES protocol. LR, RS and DS performed the study."}
+{"text": "The World Health Organization has urged all member states to deploy artemisinin-based combination therapy and progressively withdraw oral artemisinin monotherapies from the market due to their high recrudescence rates and to reduce the risk of drug resistance. Prescription practices by physicians and the availability of oral artemisinin monotherapies with pharmacists directly affect the pattern of their use. Thus, treatment practices for malaria, with special reference to artemisinin monotherapy prescription, in selected states of India were evaluated.Structured, tested questionnaires were used to conduct convenience surveys of physicians and pharmacists in eleven purposively selected districts across six states in 2008. In addition, exit interviews of patients with a diagnosis of uncomplicated malaria or a prescription for an anti-malarial drug were also performed. Logistic regression was used to determine patient clinical care, and institutional factors associated with artemisinin monotherapy prescription.Five hundred and eleven physicians from 196 health facilities, 530 pharmacists, and 1, 832 patients were interviewed. Artemisinin monotherapy was available in 72.6% of pharmacies and was prescribed by physicians for uncomplicated malaria in all study states. Exit interviews among patients confirmed the high rate of use of artemisinin monotherapy with 14.8% receiving such a prescription. Case management, i.e. method of diagnosis and overall treatment, varied by state and public or private sector. Treatment in the private sector  was the strongest predictor of artemisinin monotherapy prescription when accounting for other factors. Use of the combination therapy recommended by the national drug policy, artesunate + sulphadoxine-pyrimethamine, was minimal (4.9%), with the exception of one state.Plasmodium falciparum malaria and was deployed at full scale.Artemisinin monotherapy use was widespread across India in 2008. The accessible sale of oral artemisinin monotherapy in retail market and an inadequate supply of recommended drugs in the public sector health facilities promoted its prescription. This study resulted in notifications to all state drug controllers in India to withdraw the oral artemisinin formulations from the market. In 2010, artesunate + sulphadoxine-pyrimethamine became the universal first-line treatment for confirmed  Plasmodium falciparum malaria in Southeast Asia for more than 10 years and is now recommended as the first-line treatment throughout the world [Artemisinin-based combination therapy (ACT) has been the mainstay treatment for he world . The Worhe world .P. falciparum and P. vivax treatment in areas where therapeutic studies had not been conducted or where treatment failure was less than 10%. However, systematic analyses of chloroquine resistance suggested the phenomenon was widespread [P. falciparum cases [In India, the national treatment policy prior to 2008 recommended chloroquine for despread , not focum cases . PresentThe success of a new treatment policy depends on the adherence of health providers to the guideline and the compliance of patients with the recommended treatment. Equally, information on current provider and patient practices is needed to inform and improve future treatment policy. Recent reports note changes in pattern of drug prescription and utilization in public and private health facilities following changes in national treatment policy ,7. HowevP. falciparum control representing 55.1% of the reported burden in the country [P. vivax only. In each state, except Delhi, two districts (third level administrative unit of one to three million population) were selected based on the proportion of P. falciparum among all malaria (\u226550%) in the district from surveillance data and the introduction of ACT by NVBDCP.Six states were selected: Assam, Delhi, Goa, Gujarat, Jharkhand, and Orissa. Out of these, Assam, Jharkhand, and Orissa were priority states for et al., unpublished data) among treatments. With 90% power and an alpha of 0.05, at least 1, 733 patients would be needed to detect a 20% prevalence in such a setting.A cross-section of conveniently selected health facilities in each site were surveyed during August to November 2008. Different levels of health facilities including district hospitals, primary health centres (PHC), subcentres, and private hospitals or clinics were interviewed. All physicians present in the selected health facility were interviewed. Surveys were also conducted amongst all pharmacists around a 5 km radius of selected health facilities. Finally, exit interviews were conducted in all patients leaving the selected health facility that day with either a diagnosis  of malaria or to whom anti-malarial medicines were prescribed. An earlier study in Jharkhand described 17% artemisinin monotherapy  for diagnosis, and the prescription and frequency of use of different anti-malarials. Patients were interviewed about their symptoms, the diagnostic workup received along with its result, and drugs prescribed (if any). In the case of minor children, guardians/parents were interviewed. Paper records of the visit, tests, and prescription were also examined, if available. Pharmacists were interviewed and asked about the availability and sale of all anti-malarials in their stock.A single data entry staff recorded results using Microsoft Excel 2003 and each entry was cross-checked by a senior investigator. Range checks were used to detect implausible values. Data analysis was conducted with STATA v10. Questionnaire responses were summated, physician characteristics were compared by public and private sector status, and diagnostic and treatment indicators were compared by state. Logistic regression was used to determine adjusted odds ratios for facility and clinical factors associated with artemisinin monotherapy. All univariate variables were included in the multivariable model except in the case of collinear predictors where the model with the better fit (lower AIC criterion) was retained.Verbal consent was obtained from all interviewees, data was de-identified, and the Scientific Advisory Committee of the National Institute of Malaria Research approved the study design and questionnaires. The study was a public health program evaluation conducted at the request of NVBDCP and thus IRB exempt.Five hundred and eleven physicians and 1, 832 patients were interviewed in 98 private and 98 public sector health facilities (Table Most physicians 62.2%) reported using both slides and RDKs for diagnosis. A higher proportion of private sector physicians used RDKs exclusively (9.2%) compared to those in the public sector (1.6%). Private sector physicians 'always' prescribed more artemisinins alone, artemisinin plus other anti-malarials, and less artesunate + sulphadoxine-pyrimethamine  versus those in the public sector   followed by patients without anti-malarial prescription (n = 23).Exit interviews results show a wide range of anti-malarial groups and treatments, as well as primaquine and antibiotics, were prescribed to 93.8% of patients . Patients who were prescribed artemisinin monotherapy versus other regimens were compared by facility, patient, and management risks , followed by RDK (18.5%), and 7.3% were tested with both RDK and microscopy together  while the highest proportion of shops having sold injectable artemisinin derivatives were in Assam and Jharkhand .Artemisisin monotherapy was widely available in Indian pharmacies and frequently prescribed by according to both patient and physician interviews. Overall case management varied between the public and private sector as well as region.P. falciparum patients and P. falciparum areas. After that however, use was highest among patients who did not receive a blood test and were treated clinically. Dodoo et al. reported similar results from Ghana where the prescription of artesunate monotherapy was highest in blood smear positive patients (32.9%) followed by patients without blood tests (21%), underscoring the need for discouraging presumptive treatment [The inappropriate use of anti-malarials in the past has contributed to decreased drug sensitivities today . Recent reatment .P. falciparum alone. Radical treatment, anti-relapse therapy for P. vivax or anti-gametocidal therapy for P. falciparum, using primaquine is also an important goal for control. At the time of the study no guidelines for the use of primaquine with ACT in uncomplicated P. falciparum malaria were available. For confirmed vivax malaria most patients seen in the public sector received a prescription for primaquine but only half of those in the private sector did. Whether the dose and duration of treatment, as well as counselling for side-effects, were appropriate is unknown. Finally, the co-administration of antibiotics in uncomplicated malaria cases was frequent and is a cause for concern.Proper case management, guided by a laboratory diagnosis, reduces the overconsumption of all anti-malarials, including artemisinin monotherapy. Presumptive treatment in India is discouraged and over 90% of patients in both sectors received a blood test and only a small proportion of patients testing negative or unknown received artemisinin monotherapy. However, many smear or RDK negative patients still received another anti-malarial suggesting a lack of confidence in the diagnosis and/or a lack of concern about the overuse of anti-malarials. In the case of patients tested with RDK alone and microscopy is not available, treatment for vivax malaria may be reasonable as all kits in the public sector were monovalent and detect P. falciparum cases in limited areas: demonstrated chloroquine resistance, malaria deaths, high proportion of P. falciparum. A total of 117 districts were eligible and represented the majority of the reported falciparum malaria burden of the country. ACT blister packs were deployed through NVBDCP for 15 years or older age groups and efforts to produce blister packs for other age groups were ongoing. In this study all of the sampled districts were targeted for ACT except for Delhi. However, the use of ACT for confirmed P. falciparum cases was low with the exception of Goa. International attention has focused on identifying sustainable resources for purchasing the needed drugs for widespread deployment through public sector health systems [This study was conducted in 2008 about 1 year after ACT became the recommended treatment of  systems . In IndiThis study was conducted in 2008 relatively soon after the first widespread introduction of ACT in country. Even blister packs for different age groups were unavailable necessitating supply of loose tablets of artesunate to be made available in the PHC which may have increased monotherapy use. As the study was conducted for public health needs, important areas were purposively selected rather than random sites and the results may not be population representative. Physician and pharmacist responses may have been biased to appear desirable, however, confidentiality was stressed and participation was 100%.Following this study, strict regulation on industry as well as retail outlets was imposed by the Drug Controller General of India leading to a total ban on the sale of oral artemisinin monotherapy since July 2009. Creating awareness among health providers to change clinical practices remains the greatest challenge, particularly in the large private sector. Fixed dose combinations, which prevent the accidental or intentional use of artemisinin monotherapy, are the best available option for future policy. Ensuring access to effective ACT in both public and private sectors, while progressively removing oral artemisinin monotherapies from the market, is essential to provide effective treatment and prevent the development of artemisinin resistance.The authors declare that they have no competing interests.NM, NV were responsible for the design of the project proposal and monitored the study progress. NM, SKS, HCS, KP, MKD, HK and PG carried out the surveys at respective sites. NM VKK, and NKS compiled and analysed the data. NM and ARA wrote the first draft and NV, NKS, APD and YKG corrected the draft and all authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum msp1, msp2 and glurp genes has become well established both to describe variability in alleles within a population of parasites, as well as classify treatment outcome in cases of recurrent disease. The primary objective was to assess the emergence of minority parasite clones during seven days of artesunate (AS) treatment in a location with established artemisinin resistance. An additional objective was to investigate whether the classification of clinical outcomes remained valid when additional genotyping was performed.Despite widespread coverage of the emergence of artemisinin resistance, relatively little is known about the parasite populations responsible. The use of PCR genotyping around the highly polymorphic msp1 and msp2) and the glutamate-rich protein (glurp) was performed at baseline, daily during seven days of treatment, and again at failure. Allelic variants were analysed with respect to the size of polymorphisms using Quantity One software to enable identification of polyclonal infections.Blood for parasite genotyping was collected from 143 adult patients presenting with uncomplicated falciparum malaria during a clinical trial of AS monotherapy in Western Cambodia. Nested allelic type-specific amplification of the genes encoding the merozoite surface proteins 1 and 2 . Emergence of minority alleles during treatment was detected in only one of twenty-three cases defined as being artemisinin resistant. Moreover, daily genotyping did not alter the final outcome classification in any recurrent cases.Considerable variation of The parasites responsible for artemisinin-resistant malaria in a clinical trial in Western Cambodia comprise the dominant clones of acute malaria infections rather than minority clones emerging during treatment. Additional genotyping during therapy was not beneficial. Disproportionately high rates of polyclonal infections in cases of recurrence suggest complex infections lead to poor treatment outcomes. Current research objectives should be broadened to include identification and follow-up of recurrent polyclonal infections so as to define their role as potential agents of emerging resistance. With the emergence of artemisinin resistance in Western Cambodia and along its international border with Thailand as well as further afield in Vietnam and along Thailand\u2019s western border with Myanmar, there is a public health mandate to characterize the parasite populations responsible for poor clinical outcomes, as well as better understand the dynamics of how these parasite clones co-exist -3. Gene Plasmodium falciparum parasites, has become well established both to describe variability in alleles within a population of parasites, as well as to distinguish recrudescence from new infection in cases of recurrent disease [msp1 and msp2) and the glutamate-rich protein (glurp) [msp2 gene can be similarly grouped into two allelic families, FC27 and IC3D7 [glurp gene [The use of polymerase chain reaction (PCR) genotyping around a limited number of polymorphic genes, which are known to be highly variable in  disease -8. This  (glurp) . This me (glurp) . The usend IC3D7 ,12. Onlyurp gene .P. falciparum malaria. The trial was conducted in a rural district of Western Cambodia, close to the international border with Thailand and where reports of AS treatment failures had already emerged [msp1, msp2 and glurp in this Cambodian study population, and (2) to assess whether conventional classification of recrudescence/re-infection based on samples collected only at baseline and day of failure (Df) remained valid once interim sampling was performed. There were two working hypotheses. Firstly, that AS-resistant patients were infected with novel subpopulations of relatively drug-resistant parasites not identified at baseline; as the majority of more sensitive parasites were killed, the resistant parasites would persist and become detectable during a seven-day course of AS therapy. Secondly, that new falciparum infections during follow-up had persistence of these intrinsically more resistant parasite subpopulations. Hence, these cases should be reclassified as recrudescent, making the true failure rate higher with longitudinal genotyping compared to the conventional time points of baseline and Df.The present analysis was conducted as part of a clinical trial of artesunate (AS) monotherapy designed to probe for and characterize clinical artemisinin resistance in adult patients with uncomplicated  emerged . The maiP. falciparum malaria were randomized to receive one of three AS monotherapy regimens given as a single daily oral dose: 2, 4 or 6\u00a0mg/kg/day  for seven days. Inclusion criteria were age 18 to 65\u00a0years, fever or history of fever within 48\u00a0hours of presentation, mono-infection with P. falciparum as determined on Giemsa-stained thick and thin blood films by light microscopy, and parasite density 1,000-200,000 asexual parasites/ml blood. Exclusion criteria were signs and symptoms of severe malaria and documented use of anti-malarial drugs within the preceding 30\u00a0days. All patients remained on the study ward for the first week until completion of AS administration and returned for weekly outpatient review until Day 42. Patient outcomes for the main treatment study were classified according to clinical and parasitological responses [The study was part of a randomized clinical trial conducted during 2008-2009 at Tasanh Health Center located in Battambang Province in Western Cambodia, a region of low malaria transmission, with just one third of the study population reporting one or more previous episodes of malaria. Full details of the study are described elsewhere . Brieflyesponses . PrimaryP. falciparum throughout the follow up period (n\u2009=\u200914); (Group C) \u201cRecurrence\u201d . Groups B and C when combined were termed \u201cartemisinin resistant\u201d. Group D constituted the remaining patients not included in A, B or C. For the purposes of the present analysis, enrolled patients developing Plasmodium vivax mono-infection during follow-up and confirmed by vivax-specific PCR were considered cured of P. falciparum infection.For the longitudinal genotyping analysis conducted over the first seven days (Days 0 to 6), three subgroups of patients were selected and defined as follows: (Group A) \u201cartemisinin sensitive\u201d  (n\u2009=\u200918); (Group B) \u201cslow parasite clearance\u201d (parasite clearance time >96\u00a0hours by microscopy and remained cured of f) if malaria recurrence occurred. All samples were transferred to cryovials, stored in liquid nitrogen at the study site for transfer to Bangkok, and then kept frozen at -80\u00b0C until used for DNA extraction.Prior to initiation of AS therapy (Day 0), blood for parasite density, species confirmation and genotyping was collected into EDTA. Further samples were collected daily on Days 1 to 6 and again on the day of treatment failure (Dmsp1 (block 2), msp2 (block 3), and glurp (R2 repeat region) were amplified by nested PCR. In the primary reaction, the oligonucleotide primers corresponded to conserved sequences within msp1 (block 2), msp2 (block 3) and glurp, and in the nested reaction, separate primer pairs targeted the respective allelic types of msp1  and msp2 (FC27 and IC3D7) for amplification [msp1, msp2 and glurp PCR products were loaded on 2% agarose gels, stained with ethidium bromide, separated by electrophoresis and visualized under UV trans-illumination . Samples from an individual patient were run in adjacent lanes. If there was no amplification for any allelic family, the PCR was repeated with three times the quantity of template DNA. If no amplification was detected after this second reaction, amplification was classified as unsuccessful. Analyses of number of genotypes and size polymorphism were digitalized using Quantity One\u00ae software . A minimum of ten base pairs size difference was required to define an additional genotype.Genomic DNA was extracted from 200\u00a0\u03bcl whole blood per patient using the Qiagen DNA extraction kit  according to manufacturer\u2019s instructions. The polymorphic regions of the merozoite surface proteins fication . For conmsp1, msp2 and glurp at baseline and at Df was used to classify parasitological outcomes for LTF cases. Genotyping was done stepwise with an initial msp2 determination, followed by msp1 genotyping of the paired blood samples found to have at least one identical msp2 allele before and after treatment. Based on analysis of the 3D7 strain, alleles were considered the same if molecular weights were within ten base pairs (bp) [Comparison of genotyping patterns of irs (bp) . It was irs (bp) . If a suAllelic variants were analysed only with respect to the size of polymorphisms given the high level of discrimination afforded by the Quantity One\u00ae software computer analysis.Continuous data were expressed as medians with interquartile ranges or, in the case of parasite counts, as geometric means with 95% confidence intervals (95% CI). Continuous data from patient groups were compared using the non-parametric Kruskall-Wallis test. Numerical data were expressed as proportions and compared using Chi-square or Fisher\u2019s exact tests as appropriate. All statistical tests were performed at the 5% significance level and corresponding 95% confidence intervals were estimated. Statistical analyses were performed using SPSS version 12.0  and Stata version 11 .msp1 125, msp2 137 and glurp 143). Four patients were subsequently determined not to meet all treatment study entry criteria, but genotype samples from these cases were retained for this molecular analysis. Some 139 patients were randomized to one of three oral AS monotherapy regimens; three were subsequently lost to follow-up while the remaining 136 were followed until a study endpoint was met. One case re-presented with recurrence 56\u00a0days after enrolment into the treatment study and was included in this analysis; another developed neutropenia on D3 and was withdrawn from the treatment study but retained for safety follow up and developed malaria recurrence at D42. Baseline characteristics and clinical/parasitological outcomes for patients are given in Table\u00a0f).PCR genotyping was successful for one or more genes in baseline samples from 143 patients who had undergone screening procedures prior to study enrolment  than msp1  and glurp  , but not in the AS-resistant subgroup.Some 13 distinct patterns of genotype (haplotypes) were observed at baseline Tables\u00a0. Recurremsp1, msp2 or glurp alleles present  msp2 and 8% (12/143) glurp samples, respectively. The maximum number of clones seen for a single gene was six (one case with six clear bands of msp2-FC27 in addition to two clones of msp1-K1); this case remained cured at 42\u00a0days but had a prolonged parasite clearance time of 102\u00a0hours. All other polyclonal infections had just two or three clones. In the three subgroups of artemisinin sensitive, slow clearers and recurrences, multiple P. falciparum alleles of msp-1, msp-2 and glurp, were detected in 5/18, 3/14 and 7/9 patients  . Five of nine cases were polyclonal at Df.Overall, 45/143 (31%) of the study population had evidence of a polyclonal infection at baseline with two or more Parasite genotype was evaluated daily on Days 0-6 of AS therapy for individual recurrence cases Figure\u00a0. In all msp2-3D7) observed on Days 3 and 5, when asexual parasitaemia was still positive by light microscopy; this could represent a minority clone potentially relatively resistant to AS treatment. However, this allele was not detected again at Df. In the remaining eight recurrence cases, while fewer of the alleles present at baseline were detected as parasitaemia cleared, no new alleles emerged.Only one case of recurrence had a new allele  no minority alleles emerged during seven days of treatment.f samples, eight of nine (89%) Plasmodium falciparum recurrences were determined to be recrudescent and one of nine (11%) was a new infection. Longitudinal genotyping on days 1-6 revealed that in six of nine cases no new minority parasite populations were detected during treatment, thus supporting the classification of recrudescence, although not all alleles present at baseline were detected at the time of recurrence. In one case (participant 140) described above, minority alleles were observed on Days 3 and 5 of AS treatment but not on Df. and thus the classification remained as recrudescence. One case (88) was also classified as recrudescent based on the appearance of at least one allele present in the baseline sample, as well as novel alleles at Df. Case 112 was classified as a new infection based on the appearance of novel alleles at the time of recurrence, none of which were observed during the first week. Thus in all nine cases of malaria recurrence, additional genotyping during AS treatment on days 1-6 did not alter classification of the final parasitological outcome.Based on conventional genotyping methodology using only baseline and Df, so did not appear to be associated with subsequent treatment failure. Two of the 23 cases had novel alleles present at Df but these had not been detected at all during AS therapy despite the sensitivity of the molecular methods used. In this Cambodian setting, where most enrolled patients had persistence of asexual parasitaemia beyond 72\u00a0hours, the parasites responsible for slow parasite clearance and recurrent malaria are the dominant or co-dominant clones in an acute infection. Further, because they are also present in fast parasite clearers, they are well established in the Tasanh parasite population. These data also support recent reports of highly conserved, yet genotypically very distinct, populations of parasites from a number of sites in Cambodia including this study site [This study aimed to detect emerging drug-resistant minority parasite clones during slowly clearing malaria infections that could account for cases of unrecognized treatment failure in a setting of clinical AS resistance. In fact, of the 23 cases in the artemisinin-resistant subgroup in whom daily parasite genotyping was undertaken for seven days, including nine cases with recurrent disease, a minority clone was detected in only one; moreover this clone was not present at Dudy site .P. falciparum parasites was revealed in baseline samples from the symptomatic patient population. In keeping with surveys from West Africa and Thailand, these Cambodian isolates showed greater polymorphism of the msp2 allelic family (IC3D7 and FC27) compared to msp1 and glurp[msp1, msp2 and glurp in a relatively isolated area of Western Cambodia are similar to those from directly across the border in Trat Province, Thailand [In this setting of low malaria transmission and emerging drug resistance, considerable heterogeneity within the and glurp,16. ThesThailand . Comparamsp1, msp2 and glurp within a single sample, was found in just 31% of subjects overall at baseline . This suggests a relatively low diversity of the P. falciparum parasite population in this localized area of Western Cambodia. A study from Kampala, Uganda, also a relatively low transmission area, found 45/98 (46%) samples were polyclonal for msp2 at baseline [msp2 clones detected [msp1 32% of 45 symptomatic and 64% of 45 asymptomatic infections in a hyperendemic region of Gabon had polyclonality of msp1[Evidence of polyclonal infection, in the form of the presence of two or more alleles from detected . For mspy of msp1. An intrIn the present study, six clones were demonstrated in one baseline sample, although only two or three clones were detected in the remaining 44 patients with polyclonal infections. In general, studies from high transmission areas have shown that asymptomatic malaria patients harbour more parasite clones than those with symptomatic disease . The samP. falciparum parasitaemia (Df), which occurred between 21 and 58\u00a0days after commencing AS. Genotyping patterns from seven of nine treatment failure cases showed multiple parasite alleles of msp1 and/or msp2 at baseline, although only a minority were then detected during the following days of AS treatment. This may be due to greater sensitivity of some parasite strains to AS treatment, as well as to declining parasitaemia in the small volume of sampled blood, or to a combination of both. An alternative explanation for the disappearance of some parasite strains, which later re-appear at the time of treatment failure, is parasite dormancy triggered by the presence of AS [Additional daily samples were genotyped during seven days of AS treatment (Days 0 to 6) as well as on the day of recurrent ce of AS ,25.P. falciparum malaria in France revealed 19/20 (95%) cases harboured between two and five clones, with some cases having up to three dominant clones in addition to other minority clones [msp1 and msp2 detected at baseline [However, repeated genotyping of symptomatic travellers returning from Africa before and during 96\u00a0hours of quinine treatment for y clones . Moreovebaseline . This laP. falciparum populations undergoing selection or modification in the presence of drug. Parasite DNA is detectable in peripheral blood for less than 48\u00a0hours following the death of a parasite in response to drug or immune mechanisms and before being removed from the circulation by circulating phagocytes and spleen macrophages [versus 10 parasites/\u03bcL [f. Therefore the quantification results are unlikely to be influenced significantly by the presence of circulating gametocytes. Using current PCR strategies, gametocytes can only be detected differentially by reverse transcription-PCR (RT-PCR) [A striking feature in patients undergoing longitudinal sampling over seven days was the presence of parasite DNA up to seven days after the initiation of AS therapy; this may represent relatively drug-resistant rophages . Therefosites/\u03bcL ), an add(RT-PCR) . The abif has been made previously and may be particularly valuable in high transmission settings [P. falciparum genotyping recommends possible use of polyacrylamide gels, capillary electrophoresis, or use of dedicated fragment sizing software (used here), to increase discriminatory power and reduce inconsistencies between clinical trial methodologies [In this study, conventional genotyping assigned the outcome of six of nine samples as recrudescence; additional genotyping on Days 1-6 did not alter this assessment. In two further cases, while the final classification was not altered by additional genotyping the presence of additional bands does illustrate some of the dilemmas inherent in this genotyping technique. Mixed genotype results may profoundly impact estimates of drug resistance if they form a significant proportion of treatment failure cases in clinical trials, and depending on how they are interpreted . If circsettings ,31. Convdologies .P. falciparum would have enabled potential misclassification of outcome to be examined in more detail. Additional emergent minority populations may have been detected if sampling had occurred more frequently than once daily. Band size estimates, while measured by automated reader with as much precision as possible, may also have led to minor inaccuracies. Moreover band size variability between the same alleles can potentially occur, leading to false categorization as different alleles. Analysis of samples in this study was limited to once per day so minority parasite clones with only a sporadic and low-level presence in the peripheral circulation compared to the dominant clone(s) could have been missed.There are several limitations to this analysis. More cases of treatment failure, in particular of new infections with msp1, msp2 and glurp in malaria patients from a localized area of Western Cambodia revealed considerable variation in msp2 alleles but well-conserved msp1 and glurp. Longitudinal genotyping of cases with recurrent malaria during seven days of anti-malarial treatment showed homogeneous genotype dynamics and did not alter the outcome classification compared to conventional genotyping at baseline and Df; however it did serve to illustrate some of the inherent complexities when interpreting the parasitological basis of recurrent disease. In contrast to Africa, where malaria transmission rates are higher and artemisinin resistance does not currently appear problematic, lower polyclonality of infection was observed in malaria patients in Cambodia. The exception was the group of recurrent malaria infections in which polyclonality was much higher. The potential role of polyclonal infections in contributing to the spread of artemisinin resistance deserves further investigation.Limited genotyping of The authors declare that they have no competing interests.DB conceived the study; PG, SS, SW, KP, and ST performed the molecular genetic studies; YS, CL, SS, SD, MF, DS, DB conducted the clinical trial; PG, CL and DB analysed the data; PG, CL, ST, MS, CMC, DW, MF, JM, DS, and DB wrote the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria. Therefore, an extended follow-up of adult patients treated for severe imported malaria was started in August 2011 at the University Medical Center Hamburg-Eppendorf. Until January 2012, three patients with hyperparasitaemia (range: 14-21%) were included for analysis. In all three patients, delayed haemolysis was detected in the second week after the first dose of intravenous artesunate. Reticulocyte production index remained inadequately low in the 7 \u2013 14 days following the first dose of artesunate despite rapid parasite clearance. Post-treatment haemolysis after parenteral artesunate may be of clinical relevance in particular in imported severe malaria characterized by high parasite levels. Extended follow-up of at least 30 days including controls of haematological parameters after artesunate treatment seems to be indicated. Further investigations are needed to assess frequency and pathophysiological background of this complication.Parenteral artesunate has been shown to be a superior treatment option compared to parenteral quinine in adults and children with severe malaria. Little evidence, however, is available on long-term safety. Recently, cases of late-onset haemolysis after parenteral treatment with artesunate have been reported in European travellers with imported  Approximately 10,000 to 12,000 malaria cases imported to Europe are reported to the World Health Organization (WHO) annually .Parenteral artesunate has been shown to be a superior treatment option compared to parenteral quinine in adults and children with severe malaria in endemic countries ,5. In a et al.[Regarding short-term safety, hypoglycaemia occurred significantly less frequently in patients treated with artesunate. In children but not in adults, however, the incidence of neurological sequelae at the time of discharge was higher after treatment with artesunate compared to quinine which might be due to the fact that more severely diseased children survived. There was no difference discernible on long-term neurologic sequelae between artesunate and quinine arms 28 days after discharge in the African multicentre trial by Dondorp et al.,6. Littlet al.. The aetAn extensive follow-up of patients treated for severe malaria to detect any uncommon findings and complications after parenteral artesunate was implemented at the University Medical Centre Hamburg-Eppendorf. From the beginning of this follow-up programme in August 2011 to January 2012, three patients with hyperparasitaemia were treated with parenteral artesunate, all of which developed late-onset haemolysis.The first patient was a 19-year old German female without migrational background presenting without any co-morbidities or prior medication who stayed in Uganda for three weeks. She did not take any malaria prophylaxis. Four days after returning to Germany she developed fever and headaches. At the emergency department of a community hospital, she was diagnosed with uncomplicated malaria and showed 2% parasitaemia. She was transferred to a tertiary care centre specializing in tropical medicine for treatment of uncomplicated malaria. At the first evaluation in the emergency department (day 0), she did not fulfill any of the WHO criteria for severe malaria , and in The second patient was a 54-year old German male without migrational background returning from a one week-trip to Gambia. No malaria prophylaxis was taken. Prior medical history included surgically resected colorectal carcinoma two years earlier and arterial hypertension. His current medication consisted of metoprolol, hydrochlothiazide and felodipine for hypertension. Eight days after returning to Germany, he developed fever up to 41\u00b0C. The primary care physician diagnosed community acquired pneumonia and prescribed doxycycline for 7 days. After completing this course of antibiotic treatment, the symptoms still persisted and the patient became icteric. He presented himself to the emergency ward of a community hospital. Malaria was diagnosed and the patient was referred to a tertiary care hospital specializing in tropical medicine. Upon presentation at the tertiary care hospital (day 0), he appeared somnolent and showed hypotension of 100/50 mmHg with a pulse rate of 120 bpm. Hyperparasitaemia of 21% was diagnosed in Giemsa-stained microscopy. Furthermore, hyperbilirubinaemia  and acute renal failure  were present. He was transferred to the intensive care unit and intravenous artesunate  was given in a total of four doses of 160 mg on day 0, after 12, 24 and 48 hours equivalent to a total dose of 9 mg/kg body weight. For haemodynamic support, he had to receive norepinephrine intermittently. On day 1, he received one unit of packed thrombocytes for thrombocytopenia of 7,000/\u03bcl . Two units of packed red blood cells were transfused for anaemia at an Hb of 7.7 g/dl  on day 3. Anti-malarial treatment was continued with oral atovaquone/proguanil on day 3 (three doses of 1000 mg atovaquone and 250 mg proguanil each) and on day 5 the patient was transferred to a regular ward. Parasite levels declined rapidly from 21% initially to 12% on day 1 and 1.5% on day 3. On day 7, blood film was negative. Two weeks after initial presentation at the tertiary care hospital, the Hb reached its lowest level of 5.7 g/dl  with a concurrent rise in LDH, which peaked on day 21  without any co-morbidities or prior medication. He returned from a business trip to Senegal and Gambia where he stayed for one week without malaria prophylaxis. Ten days after returning to Germany he developed multiple fever episodes up to 39\u00b0C and chills without any other complaints. He presented to a community hospital emergency department where the following laboratory findings were obtained: creatinine 3.6 mg/dl , urea 43 mg/dl  and total bilirubin 7.1 mg/dl . Giemsa-stained microscopy revealed a parasitaemia of 20% and the patient was transferred to a tertiary care hospital specializing in tropical medicine for treatment of severe malaria. Surprisingly, the patient did not present any clinical complaints or symptoms upon first evaluation in the tertiary care hospital (day 0). Because of the laboratory diagnosis of severe malaria (hyperparasitaemia), a course of intravenous artesunate was provided in four doses of 240 mg on day 0, after 12, 24 and 48 hours, equivalent to a total dose of 9 mg/kg body weight which was switched to oral atovaquone/proguanil on day 3 (three doses of 1000 mg atovaquone and 250 mg proguanil each). Parasite levels declined rapidly from 20% initially to less than 1% on day 3. On day 9, the patient was discharged on his own explicit wish in good health condition. The patient\u2019s initial Hb of 14.7 mg/dl decreased rapidly during the first day of hospitalization and was stabilized at 10.7 g/dl at discharge  mg/dl; Figure This case series describes three cases of late-onset haemolysis after parenteral artesunate for severe malaria in non-immune travellers returning from sub-Saharan Africa. In all three patients, an initial stabilization of Hb- and a decline of elevated LDH-levels in the first week after treatment initiation was seen. In the second week, Hb levels continued to decline while around day 14 a second rise in LDH levels was observed. At this time point haptoglobin was undetectable and total bilirubin rose again. Re-occurrence of biochemical markers compatible with delayed haemolytic anaemia 1\u20132 weeks after starting parenteral artesunate contrasted the rapid clinical improvement seen early after administering the first dose. Blood stage parasites were cleared around day 4\u20135 and all patients remained parasite-free during follow-up. One patient required blood transfusions. In all cases, haemolysis resolved slowly and Hb levels had not returned to normal levels 30 days after treatment initiation.et al.[These findings are in line with a recent report by Zoller et al.. Of 25 pAs this case report is only descriptive and no control group (e.g. treated with quinine) exists, it is not possible to state with absolute certainty that the observed haemolysis is drug-related rather than disease-specific. The literature on longer-term follow-up of haematological parameters after severe malaria in general as well as in relation to specific antimalarial treatment is scarce. One report of 192 children treated with quinine showed a  drop in Hb after treatment and stabilization between days five and 21 . The lacHaemolytic anaemia is a characteristic finding in acute malaria. The aetiology of malarial anaemia is multifactorial and includes destruction of infected and uninfected erythrocytes as well as impaired erythropoiesis. Both, parasite toxicity as well as host immune mechanisms are causally involved . Impaireet al. also had high parasite levels upon presentation (4 - 30%) [An explanation of a second peak of haemolysis might lie in the reduced life span of \u201cpitted\u201d erythrocytes. The number of these once-infected erythrocytes rises significantly after treatment with artesunate but not with quinine ,18. This - 30%) .Another potential explanation of late-onset haemolysis includes a delayed (re-)activation of pro-inflammatory reactions possibly triggered by the rapid and massive destruction of malaria parasite by artesunate and an increased presentation of parasitic antigens.et al.[The fact that the patients described in this report received lower cumulative doses of artesunate than those in the publication by Zoller et al. argues aet al., Coombs\u2019 test was negative in all three patients in whom this test was performed [One (patient 2) of the two patients receiving immuno-haematological testing developed a positive Coombs\u2019 test with IgG of anti-E specificity. This patient was the only one to receive packed red blood cell transfusions repeatedly after the first dose of artesunate. A delayed haemolytic transfusion reaction cannot be completely excluded in this patient, although it seems very unlikely. Delayed haemolytic transfusion reactions generally occur following alloimmunization after a previous transfusion. The time of onset is usually two to eleven days after the transfusion. The extent of haemolysis is mostly mild without clinical implications as only the transfused erythrocytes are being destroyed ,20. The rformed . All in Two of the patients received drugs with anti-malarial activity prior to the first dose of artesunate. Patient 1 received 750 mg of mefloquine while patient 2 received a seven-day course of doxycycline for suspected bacterial infection. It is unclear whether these medications may have contributed to delayed haemolysis.After treating three hyperparasitaemic patients with parenteral artesunate, malaria parasites were cleared within a few days and the patients\u2019 clinical condition improved rapidly. Post-treatment haemolysis, however, seems to be a relevant complication in non-immune travellers with imported malaria. Risk factors and pathophysiology are unknown. To gain statistically significant results for patients with imported severe malaria, data from cases at multiple centres will have to be accumulated in a standardized manner. Whether this complication also occurs in children with severe malaria in endemic regions is currently unknown. A regular follow-up of at least one month after treatment with parenteral artesunate including controls of haematological parameters seems to be indicated.Written informed consent was obtained from the patients for publication of this Case report. A copy of the written consent is available for review by the Editor-in-Chief of the journal.The authors declare having no competing interests.TR, JPC, DW and SS took part in the patients\u2019 care. TR drafted the manuscript with contributions of JPC and GDB. All authors read and approved the final manuscript."}
+{"text": "Previous investigations indicate that methotrexate, an old anticancer drug, could be used at low doses to treat malaria. A phase I evaluation was conducted to assess the safety and pharmacokinetic profile of this drug in healthy adult male Kenyan volunteers.Twenty five healthy adult volunteers were recruited and admitted to receive a 5 mg dose of methotrexate/day/5 days. Pharmacokinetics blood sampling was carried out at 2, 4, 6, 12 and 24 hours following each dose. Nausea, vomiting, oral ulcers and other adverse events were solicited during follow up of 42 days.max) was 160-200 nM and after 6 hours, the effective concentration (Ceff) was <150 nM.The mean age of participants was 23.9 \u00b1 3.3 years. Adherence to protocol was 100%. No grade 3 solicited adverse events were observed. However, one case of transiently elevated liver enzymes, and one serious adverse event (not related to the product) were reported. The maximum concentration (Ceff of 250-400-nM required to clear malaria infection in vivo. Further dose finding and safety studies are necessary to confirm suitability of this drug as an anti-malarial agent.Low-dose methotraxate had an acceptable safety profile. However, methotrexate blood levels did not reach the desirable C Plasmodium falciparum parasite resistant to artemisinin derivatives has been reported in South Asia, and there is concern that this would spread to Africa, which would compromise the current ACT strategy and the use of artesunate in severe malaria [Chemotherapy remains one of the most important tools for the management and control of malaria. The World Health Organization has recommended the use of artemisinin combination therapy (ACTs) as first-line treatments of uncomplicated malaria; ACT is also used for the treatment of malaria in pregnancy during the second and third trimesters . Artemet malaria .Quinine has become the second-line of treatment of uncomplicated malaria in many countries . HoweverP. falciparum strains (parasites harboring wild type or mutant dhfr respectively), including those carrying the Ileu-164-Leu dhfr codon, with IC50 < 85 nM (inhibitory concentration that kill 50% of parasitaemia) [Several investigators have demonstrated that the anti-cancer methotrexate (MTX) and trimetrexate (TMX), which are inhibitors of dihydrofolate reductase (DHFR), are potent against both pyrimethamine (PM) sensitive and PM-resistant itaemia) .in vivo concentrations >250 nM, which can clear malaria parasites in vivo (MTX IC99 of 200-400 nM) [P. falciparum and Plasmodium vivax in adults [MTX is used at high dose, up to 5 to 24 g per adult per week (130-300 mg/kg) for several weeks for the treatment of cancer. This dose can yield serum concentrations of >1000 \u03bcM, a concentration range associated with MTX life-threatening toxicity . By cont-400 nM) -17. In an adults ,19. Takefalciparum malaria. This trial was registered at ClinTrials.gov (NCT# 00791531). As started earlier, a dose as low as 2.5 mg/day for 3 to 5 days was found to clear malaria infection, and the use of MTX in arthritis indicates that 7.5-30 mg per week was relatively safe. Thus, a dose of 5 mg/day/5 days  was chosen for investigation since it did not exceed 30 mg per treatment (the weekly dose used in arthritis), and was double the 2.5 mg/day .This is a report of a clinical trial of LD-MTX in healthy adult Kenyans to assess its safety, tolerability and pharmacokinetics, as a step towards its development for the treatment of uncomplicated rd March 2009 and 31st August 2009 at the Kenya Medical Research Institute (KEMRI), Nairobi. After obtaining all the required ethical approvals, the study was advertised using posters and word of mouth with messages from the approved subject information sheets. The study nurses obtained informed consent from the study participants, after which recruitment into the study began. The participants underwent pre-HIV test counselling, full physical examination and a baseline blood sample was taken. They were advised to report to the study clinic the following day, ready for admission in the study ward for 5 days, if they met the eligibility criteria. The day of admission was considered as day 0 for each volunteer. Only eligible volunteers were assigned a unique study identification (ID) number, which was linked to a pre-generated random allocation to the pharmacokinetics (PK) sampling group.The study was conducted between 3This study was reviewed and approved by the KEMRI Scientific Steering Committee, KEMRI/National Ethical Review Committee (ERC), Pharmacy and Poisons Board, Kenya and the Oxford Tropical Research Ethics Committee (OXTREC number 5608). The study was conducted in compliance with ICH-GCP guidelines and the Declaration of Helsinki, 2008 ,22.The study population was a group of 25 healthy adult male volunteers screened from communities living around the KEMRI facility in Nairobi. A summary of the inclusion and exclusion criteria applied to this study is shown in Table MTX was purchased from Wyeth Pharmaceuticals (South Africa) as 2.5 mg tablets, with a shelf-life of three years. The drug was stored in a dry, locked cabinet at a temperature below 30\u00b0C and it was used according to the manufacturer's specifications. At each dosing time point, two tablets were administered with water in the morning before breakfast every day for five days.max), half-life (T1/2) and area under the curve (AUC).The 25 study ID numbers were randomly pre-allocated (using a computer-generated simple randomization list) to five groups of five participants in each group. This allocation ensured that on each day, only one group of five participants was scheduled to give blood sample (2 ml) at 2, 4, 6, 12 and 24 hours after each daily MTX dose administration for PK study. Blood samples were collected in EDTA tubes and stored at -20\u00b0C. Samples were later transported to the KEMRI-Wellcome Trust Programme, Kilifi for analysis. Plasma concentrations of MTX were measured using Abbott TDx FLx fluorescence polarisation immunoassay analyser . The MTX II reagent, calibrator and control kits, and diluent buffer were supplied by the manufacturer. The inter-assay coefficients of variation over the range of low (0.07 \u03bcmol/L), medium (0.4 \u03bcmol/L) and high (0.8 \u03bcmol/L) concentrations of the quality control standards were 9.23, 3.95 and 3.32% , respectively. Pharmacokinetic parameters calculated were maximum concentration ; serious adverse events (SAE) and suspected unexpected serious adverse reactions (SUSARs) were used according to standard ICH-GCP . SAE's, Standardized clinical laboratory methods were employed on blood samples collected in EDTA and heparinized tubes for haematology/biochemistry and PK respectively. A, BECKMAN Coulter\u2122  haematology analyzer was used with manufacturer reagents to estimate total white cell count, red cell count and haemoglobin levels. A Selectra  chemistry analyser with reagents from Randox\u2122 (Ireland) was used to measure creatinine, alanine aminotransferase (ALT), aspertate aminotransferase (AST), alkaline phosphatase (ALP) and total bilirubin.No formal sample size calculation was done, since this was a descriptive phase I safety study. However, because of PK considerations requiring population sampling for at least five time-points, the sample size of 25 was adopted to allow for 5 concentration-time data points for each blood sampling time-point.Because the trial involved healthy volunteers with no medical records, specific participant files were used to capture all information as source. Data was then transcribed on to case record files, which were checked by the monitors at 100% verification against the source documents. The case records where then reviewed for completeness by the data manager before entry on to the study database.OpenClinica\u2122 , running on Microsoft Windows Server 2003, Apache Tomcat\u2122  and PostgreSQL\u2122  was used for setting up and managing the clinical trial data.Standard statistical analyses, based on the review of the individual values of clinically significance variables and descriptive statistics  were conducted using the data analysis and statistical programme, STATA (version 11); StataCorp LP, Texas, USA. Non-parametric methods were used to assess for any statistically significant changes of the clinical laboratory safety parameters.At total of 60 adult male volunteers were screened, out of which 25 with mean age of 23.9 \u00b1 3.3 years were enrolled in the study. Other baseline demographic characteristics collected included height, weight and body mass index (BMI), and are summarized in Table Among the solicited adverse events, only nausea was observed in 1/25 (4%) participant between days 3 and 7. Neither oral ulcers nor vomiting were reported at any time during the trial period. For other unsolicited symptoms, the following were reported: dizziness 2/25 (8%) and diarrhoea 1/25 (4%), and headache 3/25 (12%), and they all occurred between day 0 and 2. No other non-serious AEs were reported at follow up times till day 28 when one volunteer reported headache, diarrhoea and abdominal pains, and another volunteer complained of headache on day 42. All these events resolved without sequelae.One serious adverse event was reported on day 21. The volunteer was brought to the study clinic and admitted in the ward in a semi-comatose state, with cough and fever. The diagnosis was right-sided lobar pneumonia with X-ray evidence of lung consolidation, which was treated with parenteral crystalline penicillin. He fully recovered and was discharged after three days without any further complications. This serious adverse event was deemed to be unrelated to the study medication.For the haematological profiles, there was in general, a trend towards a mild decrease in haemoglobin, red cell count and total white blood cell count, during the first week following drug administration. However, these levels normalised by week 2-3, except those of the total white cell count, which, although remained within normal ranges, showed a statistically significant declining trend (P = 0.025). However, all these events were without any clinically significant outcome  and total bilirubin levels tended to increase during the first week, and by the fourth week, the levels returned to normal values Figure . Howevermax ranged from 160-200 nM, and these levels were achieved within the first hour. Thereafter, concentrations rapidly declined and the effective concentration (Ceff) was <150 nM after 6 hours, following drug administration  on day 0 were 170 and 959 nmoles hours/L, respectively, compared with 145 and 936 nmoles hours/L, respectively, on day 4. These results suggest that MTX does not accumulate in the body following repeated administration of the LD-MTX over 5 days. Drug levels for each participant are given in Additional file PK analysis of LD-MTX (5 mg/day for 5 days) showed that Cn Figure . These lLD-MTX, at doses between 7.5-30 mg/adult per week for several years has been extensively used in the western world for the treatment inflammatory diseases including RA, JIA and psoriasis -14. At tThe use of MTX is known to mildly increase liver enzymes, and the levels always normalise within a few weeks after drug discontinuation ,24. ThusLD-MTX is increasingly being used in Africa , includiLD-MTX is also being used in the treatment of various disease conditions including: inflammatory bowel disease ; urticar50 around 40-85 nM were reported these values correspond to IC99 around 200-400 nM [eff of MTX >400 nM should achieved to clear malaria infection. However our PK analysis shows that the achieved mean Cmax was around 200 nM only, and after 6 hours, MTX concentrations declined below 150 nM. These concentrations are not high enough to clear malaria infection; concentrations above 400 nM for 6-12 hours would need to be achieved. This implies that further studies should be carried out to evaluate the dose range that will yield adequate plasma concentrations and still be well tolerated. One of the approaches would be to reduce the days of treatment from 5 to 3 only, and increase the dose to 7.5, 10 or 12.5 mg per day. The total dose for each treatment course should however remain around 22.5-37 mg, which is close to the range of doses administered weekly in the treatment of inflammatory diseases (7.5-35 mg per adult). At such doses, it is expected that, the drug would remain safe since its toxicity is mainly the result of chronic use, while in the case of malaria treatment, it would be used for three days only. Another approach could be to start with a loading dose of 10 to 15 mg for the first day, and subsequently use lower doses to 5 or 7.5 mg, a strategy similarly used with quinine and artemether/lumefantrine in the treatment of severe and uncomplicated malaria, respectively [In previous work, MTX IC0-400 nM ,37, thusIn conclusion, 5 mg of MTX/day for 5 days is safe and well tolerated. However, MTX blood levels achieved are not high enough to clear malaria infection. Therefore, other dose ranging clinical investigation need to be carried out to establish the optimal doses that are safe and can yield adequate MTX concentrations in the body.The authors declare that they have no competing interests.RC, AA, MB, RJ, TL, AN, OO, TN and KM were involved in the conception of the project, study design, protocol development, and overall organization of the study. HL, AA, HL, PN, GM, AS, EA were involved in the implementation of trial, data collection, coordination, and monitoring of the study. EW was the manager who created the database and managed data. RC, JJ, AN, TL, HL, CK and MB were involved in data analysis and interpretation of results. RC and AA developed and coordinated manuscript writing. GK and SNM contributed in the study design, drug level measurement and pharmacokinetic data analysis.All the authors read and approved the final manuscript.This study was supported by the European Developing Countries Clinical Trials Partnership (EDCTP) and the Wellcome Trust WT077092 (to Prof Kevin Marsh for activity of the KEMRI/Wellcome Trust programme).Mean and standard deviation (SD) of plasma methotrexate concentrations (1000 \u00d7 nmol/L). As explained in Material and Methods, the 25 participants were randomly selected in group of five, for a total of 5 groups. Each day, only one group was scheduled to give blood sample after each daily MTX dose administration for pharmacokinetic analysis (PK). Tables A, B, C, D, E represent data for participants selected for day 0 (the first day of the study), day1, day 2, day 3, and day 4 respectively. All participants were followed up on 7, 14, 28 and 42 days, and blooded collected for PK analysis on each of these aforementioned day. ND stands for \"not determined\".Click here for file"}
+{"text": "Plasmodium falciparum malaria worldwide. Artemisinin resistance has now been reported in Southeast Asia with a clinical phenotype manifested by slow parasite clearance. Although there are no reliable reports of artemisinin resistance in Africa, there is a need to better understand the dynamics of parasite clearance in African children treated with ACT in order to better detect the emergence of artemisinin resistance.Artemisinin-based combination therapy (ACT) is widely recommended as first-line therapy for uncomplicated P. falciparum malaria over a 14-month period, daily blood smears were performed for three days following the initiation of therapy. Associations between pre-treatment variables of interest and persistent parasitaemia were estimated using multivariate, generalized, estimating equations with adjustment for repeated measures in the same patient.Data from a cohort of Ugandan children four to five years old, enrolled in a longitudinal, randomized, clinical trial comparing two leading ACT, artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP), were analysed. For all episodes of uncomplicated vs DP , having a temperature \u226538.0\u00b0C vs\u2009<\u200937.0\u00b0C  and having a parasite density >20,000/\u03bcL vs <4,000/\u03bcL . Independent risk factors for having persistent parasitaemia on day 2 included elevated temperature, higher parasite density, and being HIV infected.A total of 202 children were included, resulting in 416 episodes of malaria treated with AL and 354 episodes treated with DP. The prevalence of parasitaemia on days 1, 2, and 3 following initiation of therapy was 67.6, 5.6 and 0% in those treated with AL, and 52.2, 5.7 and 0.3% in those treated with DP. Independent risk factors for persistent parasitaemia on day 1 included treatment with AL Among Ugandan children, parasite clearance following treatment with AL or DP was excellent with only one of 752 patients tested having a positive blood slide three days after initiation of therapy. The type of ACT given, pre-treatment temperature, pre-treatment parasite density and HIV status were associated with differences in persistent parasitaemia, one or two days following therapy.NCT00527800.Current Controlled Trials Identifier Plasmodium falciparum malaria by the World Health Organization and has been adopted as first-line therapy in most malaria-endemic countries[Artemisinin-based combination therapy (ACT) is currently recommended for the treatment of uncomplicated ountries. Marked ountries. Howeverountries-6 poses ountries-9. Thus in vitro correlates have been inconsistent[in vivo studies to measure early clearance of peripheral parasitaemia by microscopy. The use of sampling multiple times a day at measured time points to estimate the rate of parasite clearance has been proposed as an accurate and reliable method for the early detection of artemisinin resistance[in vivo drug efficacy studies among outpatients. An alternative approach is to measure the proportion of patients with detectable parasitaemia one, two or three days after the initiation of therapy[The phenotype of artemisinin resistance is characterized by a significant delay in parasite clearance following initiation of therapy-6. As thnsistent, surveilsistance. However therapy,13. AlthIn this study the proportion of patients with detectable parasitaemia one, two and three days after initiation of therapy were measured in a cohort of Ugandan children randomized to two different ACT regimens. The objective of the study was to identify factors associated with early parasite clearance in a setting where artemisinin resistance has not yet occurred. This information will be important for the interpretation of future surveillance studies aimed at identifying early signs of artemisinin resistance in African populations.This study was part of a larger open-label, randomized trial conducted at Tororo, an area in Eastern Uganda with high malaria transmission intensity. The metSubjects were followed for all of their medical problems at a dedicated study clinic open seven days a week and parents/guardians were encouraged to bring their children to the study clinic whenever they were ill. HIV-infected children were prescribed daily trimethoprim-sulphamethoxazole (TS) prophylaxis. Medications with anti-malarial activity were avoided for the treatment of non-malarial illnesses. Study participants were followed until they reached five years of age or met one of the following criteria for early study withdrawal: 1) movement out of the study area, 2) inability to be located for >60 consecutive days, 3) withdrawal of informed consent, 4) inability to adhere to the study schedule and procedures, or, 5) inability to tolerate the drugs used for malaria treatment.Children who presented to the study clinic with a documented fever (tympanic temperature \u226538.0\u02daC) or history of fever in the previous 24 hours had blood obtained by finger prick for a thick smear. If the thick smear was positive, the patient was diagnosed with malaria regardless of parasite density. Children who were aged\u2009\u2265\u2009four months and weighing \u22655\u00a0kg were randomized to receive either artemether-lumefantrine (AL) or dihydroartemisinin-piperaquine (DP) at the time they got their first episode of uncomplicated malaria. Study participants received the same treatment regimen for all subsequent episodes of uncomplicated malaria. Study drugs were given according to weight-based guidelines for fractions of tablets as follows: AL , administered as one (5\u201314\u00a0kg) or two (15\u201324\u00a0kg) tablets given twice daily for three days; and DP , targeting a total dose of 6.4 and 51.2\u00a0mg/kg of dihydroartemisinin and piperaquine, respectively, given in three equally divided daily doses to the nearest one-quarter tablet. Study drugs were crushed, mixed with water and administered to the patient. Patients were given a glass of milk after each dose. The first daily dose of study drugs was directly observed at the study clinic. After each dose, children were observed for 30 minutes, and the dose was re-administered if vomiting occurred. For children treated with AL, the second daily dose was packaged and the parent or guardian was given verbal instructions for proper administration of the medication at home with clear emphasis on when the evening dose should be given. Episodes of complicated malaria and treatment failures occurring within 14 days of initiating treatment were treated with quinine. All children with malaria were followed up on days 1, 2, 3, 7, 14, 21, and 28 following enrolment.Thick and thin blood smears were stained with 2% Giemsa for 30 minutes and read by trained laboratory technologists who were not involved in direct patient care. Parasite densities were calculated from thick blood smears by counting the number of asexual parasites per 200 leukocytes , assuming a leukocyte count of 8,000/\u03bcl. A blood smear was considered negative when the examination of 100 high power fields did not reveal asexual parasites. For quality control, all slides were read by a second reader. An independent third reader settled any discrepancies between the first and second readings. Laboratory technicians were blinded to the study participants\u2019 treatment assignments. Thin smears were used to determine the parasite species.P. falciparum malaria diagnosed between October 2011 and December 2012 when routine blood smears were added on day 1 of malaria follow-up. During this period, all children remaining in the cohort study were between 47 and 60 months of age. The primary outcome of interest was the proportion of patients who remained parasitaemic by microscopy one, two and three days following initiation of therapy. Secondary outcomes included persistence of fever  during the first three days following initiation of therapy and 28-day standardized WHO treatment outcomes unadjusted by genotyping. The treatment outcomes were classified as: adequate clinical and parasitological response ; early treatment failure ; late clinical failure (first positive blood smear after day 3 in the presence of fever); and late parasitological failure (first positive blood smear after day 3 in the absence of fever). Risk factors of interest included the ACT regimen (AL vs DP); mg/kg dosing of anti-malarials; treatment episode number; pre-treatment temperature, parasite density and haemoglobin; age; gender; and HIV status. We explored the relationship between pre-treatment parasite density and proportion with parasitaemia and it was nonlinear. We thus created a categorical variable for parasite density to create the most parsimonious model that best fit the distribution of the data. Associations between risk factors of interest and early persistent parasitaemia were estimated using generalized estimating equations with robust standard errors and adjustment for repeated measures in the same patient. Risk factors associated with the outcome of interest with a p-value of\u2009<\u20090.05 in univariate analyses were included in the final multivariate model, with the exception of age and gender which were also included.Data were double entered into an Access database and analysed using Stata version 11 . This study included only data from episodes of uncomplicated Informed consent was obtained from the parents or legal guardian of all study participants. The study protocol was approved by the Uganda National Council of Science and Technology and the institutional review boards of the University of California, San Francisco, Makerere University, the University of Washington, and the Centers for Disease Control and Prevention.P. falciparum, 416 episodes were treated with AL and 354 episodes were treated with DP. The mean age, temperature, and haemoglobin level was similar in both treatment arms . A total of 38 (4.9%) episodes occurred in HIV-infected children and were equally distributed between the two treatment arms.A total of 787 episodes of malaria were treated in 202 study participants during the study period of October 2011 to December 2012. Three episodes of complicated malaria treated with quinine and 14 episodes of malaria caused by non-falciparum species were excluded from the analysis . On day two following initiation of anti-malarial treatment, a total of 43 of 764 (5.6%) episodes with blood smears done had persistent asexual parasitaemia by microscopy and the prevalence was similar between the two ACT treatment arms. On day three following initiation of anti-malarial treatment, only one of 752 (0.1%) episodes with blood smears done had persistent parasitaemia  episodes with blood smears done had persistent asexual parasitaemia by microscopy and the prevalence was higher in those treated with AL compared to DP . There was no significant difference between the two ACT treatment arms in the prevalence of fever on day two and three following initiation of anti-malarial therapy  ; pre-treatment temperature \u226538.0 compared to <37.0 ; and pre-treatment parasite density >20,000/\u03bcL  and 4,000-20,000/\u03bcL  compared to <4,000/\u03bcL [In this study a higher pre-treatment temperature was also associated with persistent parasitaemia on days one and two. This finding has been reported in another study from Kenya and may =\u20090.003). Age has=\u20090.003).Although early parasite clearance is predominantly a function of artemisinin activity, different formulations of the artemisinin component and partner drugs used in various ACT may also influence early parasite clearance. In this study, treatment with AL was associated with a higher risk of persistent parasitaemia on day one compared to treatment with DP. This finding appears to have some clinical significance as persistent fever on day one was also higher in patients treated with AL compared to DP. Similar findings of a higher risk of persistent parasitaemia on day one in AL compared to DP have also been reported in a study from Kenya as well There were several limitations to this study. As mentioned earlier, blood smear samples were obtained once daily, therefore, it was not possible to accurately estimate the parasite clearance rate as a continuous variable. This study also included patients across a narrow age range (47\u201360\u00a0months) and over a relatively short period of calendar time, thus it was not possible to evaluate age as a potential risk factor or temporal trends in early parasite clearance. Finally, only DP was fully administered as directly observed therapy, as it was given once daily when a child presented to the study clinic. In contrast, AL was given twice daily and only one of the two daily doses was directly observed. If the second dose of AL was not appropriately administered at home, differences in parasite clearance between AL and DP may have been overestimated.In summary, results from this study provide reassuring evidence that among children living in an area of high malaria endemicity in Uganda, parasite clearance following treatment with AL and DP was excellent. To provide a \"baseline\" assessment of the dynamics of early parasite clearance in this study population, several factors were associated with persistent parasitaemia at days one and two after the initiation of therapy, including pre-treatment parasite density, pre-treatment temperature, which ACT was given, and HIV status. In order to accurately detect temporal changes in early parasite clearance associated with the emergence of artemisinin resistance in Africa, analyses should control for these risk factors and potentially others when comparing data across studies. Finally, studies with richer parasite density sampling may be required to model the optimum parasite density sampling approaches  to robustly track changes in early parasite clearance in Africa.The authors declare that they have no competing interests.MKM, AK, MRK, and GD conceived and designed the study. MKM, AK, EO, FO and EA participated in data collection. MKM, AK, MRK, and GD preformed the data analysis. All authors participated in the writing of the manuscript. All authors read and approved the final manuscript."}
+{"text": "Background. Plasmodium falciparum malaria, as well as certain antimalarial drugs, is associated with hearing impairment in adults. There is little information, however, on the extent, if any, of this effect in children, and the evidence linking artemisinin combination therapies (ACTs) with hearing is inconclusive. Methods. Audiometry was conducted in children with uncomplicated malaria treated with artesunate-amodiaquine (n = 37), artemether-lumefantrine (n = 35), or amodiaquine (n = 8) in Accra, Ghana. Audiometry was repeated 3, 7, and 28 days later and after 9 months. Audiometric thresholds were compared with those of a control group of children (n = 57) from the same area. Findings. During the acute stage, hearing threshold levels of treated children were significantly elevated compared with controls (P < 0.001). The threshold elevations persisted up to 28 days, but no differences in hearing thresholds were evident between treated children and controls after 9 months. The hearing thresholds of children treated with the two ACT regimens were comparable but lower than those of amodiaquine-treated children during acute illness. Interpretation. Malaria is the likely cause of the elevated hearing threshold levels during the acute illness, a finding that has implications for learning and development in areas of intense transmission, as well as for evaluating potential ototoxicity of new antimalarial drugs. Plasmodium falciparum malaria is associated with varying degrees of neurological involvement, depending on the severity. Few studies have, however, investigated the effect of the disease on hearing specifically in those with uncomplicated malaria. Furthermore, several antimalarial drugs, including quinine [Acute  quinine , 2, chlo quinine , and mef quinine , have be quinine \u20139. Altho quinine , conclud quinine \u201315, the  quinine , as well quinine . In addi quinine , but fewWe have, in this study, conducted serial audiometric measurements with a follow-up time ranging between 9 and 15 months, in children with uncomplicated malaria treated with artesunate-amodiaquine, artemether-lumefantrine, or amodiaquine monotherapy. These measurements have been done not only to evaluate artemisinin-based and non-artemisinin-based antimalarial treatments, but also to compare artemisinin combination therapy (ACT) regimens that have been associated with differential propensities for neurotoxicity in animal studies. The audiometric measurements were also compared with those of age-matched children from the same area. The study design also permits comparison of audiograms obtained during acute illness with audiograms done at different postrecovery time points, allowing assessment of the potential effect of the acute malarial disease and recovery on the dynamics of hearing threshold changes over the study period.The audiometric assessment was a subcomponent of a clinical trial that was done to evaluate the efficacy and safety of artemisinin-based combination therapies for uncomplicated malaria in Ghana. Approval for the study was granted by the Ethics and Protocol Review Committee of the University of Ghana Medical School. The clinical trial was initiated at a time when chloroquine was still the official first-line treatment for uncomplicated malaria in Ghana. The full description of the study site and trial results have been previously reported . BrieflyOtoscopic examination was done in all children as a part of the initial screening. Children with conditions such as serous otitis media, active or inactive chronic suppurative otitis media, impacted cerumen, eardrum perforation, tympanic membrane scarring, or other clinically evident outer and middle ear abnormalities, were excluded. Children with a past medical history or clinically obvious symptoms and signs of sickle cell anaemia, renal or liver disease, malnutrition, craniofacial abnormalities, or dysmorphism, as well as children, when they were newborns, were reported to have been admitted into neonatal intensive care for any cause or those with a history of head trauma were excluded. Additionally, children with a past medical history suggesting possible birth asphyxia, neonatal jaundice, and meningitis, and those who were known to have taken aminoglycoside antibiotics, loop diuretics, or herbal medications within the past three months were excluded.Air conduction threshold was obtained for each ear separately in a quiet (nonairtight) room, using a portable audiometer , with noise-attenuated TDH-39 headsets and earphones. The audiometer was calibrated according to the manufacturer's recommendations by an external expert. The level of ambient noise measured in the examination room was <50\u2009dB A . The unmasked psychoacoustical hearing thresholds were established at 0.125, 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 6, and 8\u2009kHz, for tonal stimuli, using a standard two-step-down, one-step-up method, as per the modified Hughson-Westlake procedure . BrieflyVenous blood was collected into EDTA and heparinised tubes on days 0, 3, 7, 14, and 28 and on any day of recurrent symptomatic parasitaemia for routine haematological and biochemical investigations. Parasite counts were determined in Giemsa-stained blood films relative to 200 white blood cells (WBCs) and the measured WBC count. Total WBCs and differential counts were measured by an automated haematology analyzer .P values <0.05 were considered significant. Data are presented for selected wavelengths for clarity. Inclusion of additional wavelengths in the analysis did not alter the conclusions. Statistics were done, using SigmaPlot 11 .Continuous normally distributed data were described by the mean and standard deviation or standard error and non normally distributed data by the median and range. For the latter data, all statistics were performed on ranks. Percentages were given for categorical data, which were compared using the Chi-square test with Yates' correction, or Fischer's exact test, as appropriate. One-way analysis of variance, with Holm-Sidak post hoc pairwise testing, or two-way repeated measures analysis of variance was used to compare variables between the treatment groups and controls or to test for intra-individual and within-group differences, respectively. For some comparisons, a delta-value was calculated by subtracting the values measured at two time points in the same individual. Multiple linear regression analysis was used to evaluate the effect of selected subject characteristics on the audiometric thresholds. n = 37; artemether-lumefantrine, n = 35; amodiaquine, n = 8) on days 0, 3, 7, and 28; however, for each day of repeated testing, various children did not turn up, explaining the variable numbers of comparison in each analysis below. In all repeated comparisons, only patients tested at both time points were included in the data analysis. At the end of the follow-up period, audiometry was done in 58 of the originally recruited 80 treated children  and in 57 healthy controls. Audiometric analysis was performed in 80 malaria patients . Hearing threshold levels were similar for the three treatment groups, each of which was significantly different from the control group , hearing thresholds of all the acutely ill subjects were significantly elevated compared with the hearing thresholds of the control subjects . There were significant improvements (decreases) in hearing threshold levels on day 28 compared with day 0, at specific frequencies .There were no differences in hearing threshold levels from day 0 to day 3 or from day 3 to day 7 , except for measurements for the right ear at 125\u2009Hz and 250\u2009Hz , lower than the hearing thresholds of controls (d 250\u2009Hz .P = 0.7). On day 7, the hearing threshold levels in the amodiaquine monotherapy arm were higher, whereas in the two ACT arms they remained at the level of day 0. Thus, the hearing threshold levels on day 7 in the monotherapy arm were significantly higher than the ACT arms, especially at frequencies <1000\u2009Hz .At the final followup (9\u201315 months), there were no differences in the mean hearing threshold levels between the amodiaquine monotherapy and the artesunate-amodiaquine or artemether-lumefantrine arms  were the most consistent predictors of elevated hearing threshold measured on the exit (follow-up) audiogram (data not shown).In this study, serial audiometric measurements were done before treatment and at prespecified times after treatment of uncomplicated malaria, with ACT regimens that have been associated with different propensities for toxicity in animal studies, as well as in subjects treated with a nonartemisinin (amodiaquine) antimalarial drug. The results showed elevated hearing thresholds in all three groups of children at presentation and through the acute illness stage but complete reversibility after a sufficiently long follow-up period. Although the data shows progressive improvement (decrease) of hearing threshold levels during the initial days of treatment, especially in the two ACT groups, there were some residual elevation 28 days after treatment. The pattern of hearing threshold elevations, which was more pronounced at the lower frequencies, is consistent with that of the malaria-attributed hearing impairment that has been recently demonstrated in animals .It has also been shown, by fundoscopic examinations, that uncomplicated malaria may be associated with some neurological effects , a findiThe data did not show any differences between hearing threshold levels of children treated with artesunate-amodiaquine or artemether-lumefantrine. However, treatment with ACT-based antimalarials seemed to halt the persistence or progression of the threshold elevations during acute illness, whereas amodiaquine monotherapy treatment did not show such an effect. Although the number of subjects in the amodiaquine monotherapy group was limited, precluding definitive conclusions on differences between ACT and non-ACT treatment groups, this apparent difference between the ACT and non-ACT antimalarial regimens could be due to the rapid parasite clearance and faster fever resolution in the ACT groups . The quiThe findings of this study therefore suggest that there were minimal, if any, detectable effect of the two ACT regimens, at the administered doses, on hearing as measured by audiometry. Furthermore, since hearing threshold change is considered a better indicator of potential drug-induced ototoxicity than hearing threshold levels , the comThe complete reversibility of the elevated threshold elevations in all the three groups at the nine-month follow-up audiogram suggests acute malaria to be the likely cause of these changes, which is consistent with results from studies in adults , 13, 15.It has been hypothesized that malaria could contribute to hearing impairment by impairing labyrinth artery microcirculation  and it hThese findings taken together imply that not only does hearing impairment occur as part of the natural history of uncomplicated malaria, but also if audiometry is used to evaluate drug induced ototoxicity of newly introduced antimalarial drugs, the effect of the disease could confound any hearing threshold elevations if testing of different individuals is done at different posttreatment times.Audiometric thresholds measured in children with uncomplicated malaria treated with different antimalarial regimens showed reversible hearing threshold elevations in all treated groups, implying that these changes were disease- rather than drug-related. This has potential implications for learning, development, and behaviour of children repeatedly exposed to malaria in endemic areas and also has implications for evaluating potential ototoxicity of newly introduced antimalarial drugs."}
+{"text": "Plasmodium falciparum to commonly used anti-malarial drugs, especially chloroquine, is being increasingly documented in India. By 2007, the first-line treatment for uncomplicated malaria has been revised to recommend artemisinin-based combination therapy (ACT) for all confirmed P. falciparum cases.Resistance in P. falciparum malaria in India.The objective of this study was to compare the efficacy, safety and tolerability between dihydroartemisinin-piperaquine (DP) and artesunate plus mefloquine (A\u2009+\u2009M) drug combinations in the treatment of uncomplicated P. falciparum malaria were enrolled, randomized to DP (101) or A\u2009+\u2009M (49) and followed up for 63\u2009days as part of an open-label, non-inferiority, randomized, phase III multicenter trial in Asia.Between 2006 and 2007, 150 patients with acute uncomplicated The heterogeneity analysis showed no statistically significant difference between India and the other countries involved in the phase III study, for both the PCR-corrected and uncorrected cure rates. As shown at the whole study level, both forms of ACT were highly efficacious in India. In fact, in the per protocol population, the 63-day cure rates were 100% for A\u2009+\u2009M and 98.8% for DP. The DP combination exerted a significant post-treatment prophylactic effect, and compared with A\u2009+\u2009M a significant reduction in the incidence of new infections for DP was observed (respectively 17.1% versus 7.5% of patients experienced new infection within follow up). Parasite and fever clearance was rapid in both treatment arms (median time to parasite clearance of one day for both groups). Both DP and A\u2009+\u2009M were well tolerated, with the majority of adverse events of mild or moderate severity. The frequencies of individual adverse events were generally similar between treatments, although the incidence of post treatment adverse events was slightly higher in patients who received A\u2009+\u2009M with respect to those treated with DP.P. falciparum malaria and could potentially be considered for the first-line treatment of uncomplicated falciparum malaria in India.DP is a new ACT displaying high efficacy and safety in the treatment of uncomplicated Current Controlled Trials ISRCTN 81306618 Plasmodium falciparum and Plasmodium vivax), mosquito vectors, ecological conditions, and socio-economic factors. The Annual Parasite Incidence (API), an indicator of the disease incidence, is reportedly less than two in most parts of India. However, endemic regions with an API greater than five are spread in the states of Rajasthan, Gujarat, Karnataka, Goa, Southern Madhya Pradesh, Chhattisgarh, Jharkhand, Orissa and the North-Eastern states [Malaria continues to be a major public health challenge in South East Asia, where 30% of the global population is estimated to be at risk of malaria. India alone contributes about 70% of reported cases . More thP. falciparum to the commonly used anti-malarial drugs, especially chloroquine, has been progressively reported in India [P. falciparum cases [P. falciparum malaria. However, more investigations are needed for new fixed-dose combinations to facilitate the National Programme in adopting the most effective, easy to administer, feasible, safe and cost-effective strategy.Resistance of in India . Assam win India . As a coum cases . By 2010P. falciparum malaria. Under the WHO system of efficacy assessment, more than 95% of patients remain free of malaria 28\u2009days after A\u2009+\u2009M treatment [Combination of artesunate and mefloquine (A\u2009+\u2009M) has been used widely for many years in South-East Asian countries, such as Thailand and Cambodia, and remains highly effective for uncomplicated reatment . HoweverP. falciparum and P. vivax malaria have been conducted in Asian countries. The overall 28-day cure rates (Kaplan-Meier estimates) for the DP treatment consistently exceeded 97% in China, Myanmar, Cambodia, Laos, Thailand and Vietnam [Dihydroartemisinin-piperaquine (DP) is a potential alternative; it is a fixed-dose formulation to be taken once a day over a full treatment course of three days. In the last decade, several clinical trials evaluating the DP combination for the treatment of uncomplicated P. falciparum malaria was carried out in India and two other Asian countries (Laos and Thailand) [Recently, a DP formulation produced under Good Manufacturing Practices (GMP) has been developed by Sigma-Tau , in collaboration with Medicines for Malaria Venture (MMV). During years 2005\u20132007, a phase III multicenter study comparing this novel formulation with artesunate plus mefloquine for the treatment of acute uncomplicated hailand) . In the Three Indian centres Figure  includinThe Indian study was conducted over one malaria season, with patients screened from November 2006 to February 2007. The selected centres are situated in areas of perennial transmission (Assam), perennial transmission with a seasonal peak from June to September (Goa), and transmission active in post-monsoon months (Karnataka). Chloroquine (CQ) resistance was present at all study sites, with site records showing 28-day treatment failure rates of 32% in Assam (in 2002), 54% in Goa (in 2007) and 67% in Mangalore City (in 2005). ACT represents the standard treatment in Goa and Mangalore  and Assam. For this reason, the study protocol was amended in April 2006, requiring the prohibition of chloroquine during the study.P. falciparum mono- or mixed-infection  were eligible for the study. Local regulations were followed in India that required all recruited patients to be aged 18\u2009years and over. Key exclusion criteria were: severe malaria, treatment with mefloquine in the 60\u2009days prior to screening, treatment with DP in the three months prior to screening and >4% parasitized red blood cells. Pregnant or lactating women were not eligible for the study.Patient inclusion and exclusion criteria have been reported elsewhere . BrieflyP. falciparum during follow-up [At presentation, venous blood samples were taken for parasite count, haematology, biochemistry, and 3 blood spots were collected onto 3MM filter paper  for genotyping by polymerase chain reaction (PCR) in the event of reappearance of ollow-up . Urine wIf the study inclusion criteria were met, the patients were randomly assigned to receive either A) A\u2009+\u2009M: artesunate , 4\u2009mg/kg/day for 3\u2009days (Days 0\u20132) plus mefloquine  15\u2009mg base/kg on day 1 and 10\u2009mg base/kg on day 2, or B) DP: Dihydroartemisinin-piperaquine  2.1/16.8\u2009mg/kg in a single daily dose for 3\u2009days. Each tablet contained 40\u2009mg of DHA and 320\u2009mg of piperaquine. The Food and Drug Department, Ministry of Health, India approved the use of both A\u2009+\u2009M and DP in this trial.The treatment choice was kept in a sealed opaque envelope, which was opened only after the decision to recruit had been made. An unequal randomization, 2:1 (DP:A\u2009+\u2009M), was used to provide more precise estimates of DP cure rates, as well as to provide more patients for the safety database of DP. Axillary temperature was measured every six hours. Study drug administration was observed directly by the study physician. Study medications were given as tablets or fractions of tablets, according to body weight, given orally with a glass of water. Patients were observed for one hour to ensure that the medications were not vomited or regurgitated.P. falciparum recurrence. Venous blood samples were taken for haematologic and biochemical analysis on days 28 and 63 if the results were abnormal on day 28 and were also performed on the day of P. falciparum recurrence.Patients were observed daily until parasites cleared, then weekly for 63\u2009days from the start of treatment, or at other times if he/she felt unwell. At each visit finger prick blood was taken for a malaria smear and haematocrit. Blood spots were collected onto filter papers from those patients with reappearance of asexual parasitaemia for parasite genotyping . Twelve-Patients with treatment failure were withdrawn from the study, treated, and followed up as per local practice. They did not have to undergo efficacy evaluation thereafter. Patients requiring rescue therapy were treated with artesunate 2\u2009mg/kg/day orally for seven days plus doxycycline if there was no contraindication. Any patient diagnosed with severe malaria or danger signs during follow-up was referred for treatment with parenteral artesunate and supportive measures at the local facility or hospital. Those patients who received additional courses of therapy were also followed-up for 63\u2009days. Potential adverse events were recorded.Written informed consent was obtained from all participants. Ethical clearance for the study was granted by the following ethics committees: Institutional Ethics Committee, National Institute of Malaria Research (ICMR), Delhi, India; Goa Medical College and Hospitals Local Ethics Committee, Goa, India; Institutional Ethics Committee, Kasturba Medical College, Mangalore, India. The trial registration number is ISRCTN 81306618. The trial was monitored by MDS Pharma Services.P. falciparum trophozoites per 1000 red blood cells x haematocrit x 125.6). Polymerase chain reaction amplification was performed on paired samples for parasite genotyping to distinguish between reinfection and recrudescence. To this aim, three spots of blood were collected onto 3MM filter paper . Filter papers were dried and individually stored in a plastic bag containing silica gel. All filter papers were subsequently transferred to the Institute of Tropical Medicine  where centralized genotyping was conducted; deoxyribonucleic acid was purified as described elsewhere [Parasite counts in thick and thin films stained with Giemsa were obtained daily until parasite clearance and then weekly from day 7 up to day 63 . At screlsewhere . Three pThe primary endpoint of the study was the PCR-corrected cure rate on day 63, based on the adequate clinical and parasitological response (ACPR), which was defined as the absence of parasitaemia irrespective of the patient\u2019s body temperature, with the patient not meeting any of the criteria of early treatment failure or late clinical or parasitological failure . This deSecondary endpoints included PCR-corrected adequate clinical and parasitological response on days 28 and 42, PCR-uncorrected adequate clinical and parasitological response (PCR-uncorrected), proportion of patients with treatment failure, proportion of aparasitaemic patients, proportion of afebrile patients, number of new infections, gametocyte carriage and the safety profile of the two treatments including adverse events and electrocardiograph parameters.The Breslow-Day heterogeneity test or a logistic regression model  were useThe two most extreme populations, i.e. Intention to Treat (ITT) and Per Protocol (PP) populations, were presented in this work, in line with the presentation of the whole study results . The ITTThe analysis of the PCR-corrected and uncorrected cure rates at day 63, 42 and 28 was based on simple proportions and 95% (two-sided) confidence intervals (CIs) were computed on the difference between the treatment proportions. Survival analysis techniques were applied for analysing time to parasite clearance and estimating the rate of new infections and true treatment failure, as established by the WHO . The latIn the survival analysis performed for new infections (ITT population), lost to follow-up, informative withdrawals, recrudescences, patients with missing PCR and all the successes were censored. In the survival analysis performed for true treatment failures (PP population), patients withdrawing the study, with a new infection, with missing PCR and all the successes were censored.Rates of person-fever-days and person-gametocyte-weeks were calculated as the number of weeks in which fever was present or blood slides were positive for gametocytes, respectively, divided by the number of follow-up weeks and expressed per 1,000 person-weeks.The ITT population was used for all safety assessments. Adverse events were coded using the MedDRA dictionary (MedDRA V8.1). Proportions of patients experiencing at least one adverse event were compared between treatments using the Pearson Chi square test.A total of 154 Indian patients were screened, with 101 patients randomized to the DP arm and 49 patients randomized to the A\u2009+\u2009M arm Figure . Three cAt enrolment, the DP and A\u2009+\u2009M groups had similar demographic and clinical characteristics except that the geometric mean parasitaemia was higher in the DP group compared to A\u2009+\u2009M arm Table . All patTwenty-nine patients (21 patients in the DP arm and 8 in the A\u2009+\u2009M arm) were excluded from the PP population because lost to follow-up before or at Day 63 (see above) or had major protocol violations. Major protocol violations occurred only in the DP arm: PCR missing or indeterminate or not done before or at day 63 (3 patients); subjects not taking at least 80% of the study drug medication (1 patient); subjects having haemoglobin value below the lower limit (< 5\u2009g/dL) at day 0 or missing at day 0 and day 7 (2 patients) and no availability of the Day 63 assessment (2 patients).One early treatment failure (ETF) was observed in the DP arm due to an adverse event  occurring after the first drug administration. Due to this event, the study drug was discontinued and his malaria was successfully treated with primaquine plus quinine. No late treatment failure (LTF) was observed in both ITT and PP populations for the A\u2009+\u2009M study group at the end of follow up (day 63). For the DP arm, two late clinical failures were observed in the PP population due to recurrent parasitaemia occurring in two patients at day 39 and day 45, respectively.In the analysis of the PCR-corrected cure rate at day 63, DP was found to be similar to A\u2009+\u2009M  and 69.4% (A\u2009+\u2009M) with a difference of 4.9% (95% CI for DP-A\u2009+\u2009M: -10.6% to 20.3%) while in the PP populations the cure rates were 91.3% (DP) and 82.9% (A\u2009+\u2009M) with a difference of 8.4% (95% CI for DP-A\u2009+\u2009M: -4.8% to 21.4%). With respect to the overall phase III findings, the Indian results were better in terms of absolute values .As for the other time-points (day 42 and day 28) analogous findings were observed in all study populations (data not shown).Plasmodium species different from P. falciparum.The treatment difference on uncorrrected cure rates was mainly due to new infections Figure , which aParasite clearance was rapid in both treatment groups Table . ProportFever clearance was also fast in both arms Table  and propThe gametocyte carriage was assessed at weekly visits, from day 7 onwards. The time to clearance of gametocytes is reported in Table Blood haemoglobin (Hb) was evaluated weekly. In the ITT population, 5 (5%) patients in DP and 4 (8%) patients in A\u2009+\u2009M presented with an Hb level\u2009<\u200970\u2009g/L. Three of these patients (two in the DP group and one in the A\u2009+\u2009M group) worsened at day 7 and a SAE was communicated by the investigator (see below); all the other patients stabilized or improved their anaemic status by the following visit. There was no apparent difference between treatment groups. In the ITT population, the mean Hb fell from 105.8\u2009g/l (DP) and 104.5\u2009g/L (A\u2009+\u2009M) on day 0 to 103.2\u2009g/l (DP) and 100.6\u2009g/l (A\u2009+\u2009M) on day 7, before increasing steadily thereafter  was slightly higher in patients who received A\u2009+\u2009M with respect to those treated with DP Table . In partSix serious adverse events (SAE) were observed during the study until day 63 of follow up: five in the DP arm and one in the A\u2009+\u2009M group. All patients recovered completed by the end of the study.In the DP group, five SAE occurred in five patients. One patient  developed malaria of moderate severity on day 39 which was considered not to be related to the study drug by the investigator. His malaria was treated with oral quinine (1800\u2009mg/day) for six days. He made a full recovery. One subject  had Wolff-Parkinson-White (WPW) syndrome diagnosed on day 2. The investigator thought this was possibly related to DP. However, since WPW is a congenital cardiac conduction anomaly it cannot be caused by DP. One patient , with no history of convulsions, suffered a grand mal convulsion three hours after the first DP dose. The event was treated with i.v. phenytoin 900\u2009mg/day and diazepam 10\u2009mg/day. DP was discontinued and his malaria was treated with primaquine 45\u2009mg/day orally and quinine 600\u2009mg/day orally. He recovered completely. In the investigator\u2019s opinion the convulsion was possibly related to DP. Two patients developed worsening anaemia that was considered unlikely to be related to DP by the investigators. In particular, the first patient  was anaemic at enrolment (Hb 45\u2009g/dL). The anaemia worsened by day 7 (Hb 40\u2009g/dL) which was considered a life-threatening SAE. Ferric ammonium citrate 320\u2009mg/day was started on day 7 and continued up to study completion. The second patient  was anaemic at enrolment (Hb 50\u2009g/dL). The anaemia worsened by day 7 (Hb 40\u2009g/dL) which was considered a life-threatening SAE by the investigator. Ferric ammonium citrate 320\u2009mg/day was started on day 7 and continued until day 35.In the A\u2009+\u2009M group, one SAE (severe anaemia) occurred in one patient. The subject  was anaemic at enrolment (Hb 50\u2009g/dL) and became severely anaemic by day 8 (Hb 45\u2009g/dL). She was treated with ferric ammonium citrate 320\u2009mg/day and recovered fully. The investigator thought it unlikely that the anaemia was related to the investigational drug.No deaths were observed during the study.Malaria is one of major public health problems in India. Malaria in India contributes significantly to the world malaria burden. The National Vector Borne Disease Control Program of India reported around 1.6 million cases and 1100 malaria deaths in 2009 . Some exversus A\u2009+\u2009M was conducted in 1150 Asian patients with acute uncomplicated P. falciparum malaria [In 2005\u20132007 a phase III, multi country trial of oral DP  malaria . This st malaria .One of the strengths of this study is that it was done in settings of different transmission and had a prolonged follow-up period of 63\u2009days not to miss late reinfections and to reflect the terminal half-lives of the drugs ,18. One P. falciparum malaria in India. These products are only available as co-blistered formulations and are only prescribed for adults above 15\u2009years of age [P. falciparum parasite is now exhibiting resistance to anti-folates, though for the time being most parasites are sensitive to the drug. The reported resistance strains have been shown to be so because of mutations in candidate genes dihydrofolate reductase (dhfr) and dihydropteroate synthetase (dhps)[In 2010, artesunate plus sulphadoxine-pyrimethamine combination became the first-line treatment for confirmed s of age . As DP ise (dhps). The useOne of the limitations of this study is the lack of children below 18\u2009years of age. The burden in children in India remains  significThe DP combination exerted a significant post-treatment prophylactic effect in this study. Significant reductions in the incidence of new infections for DP compared with A\u2009+\u2009M were seen. Higher uncorrected cure rates were observed in the DP arm than with A\u2009+\u2009M, as the uncorrected cure rate endpoint includes new infections as well as recurrence of infection. This effect is thought to be caused by residual levels of drug remaining in the body because of the longer half-life of piperaquine. The post-treatment prophylaxis was better for DP reflecting the differential terminal half-life of piperaquine and mefloquine . The post-treatment prophylactic effect is of particular importance in regions with a high risk of new infection. This can reduce the risk of anaemia and allow patients more time for haematological recovery between infections.Mefloquine has been linked with adverse events of gastrointestinal and central nervous system origin . In thisP. falciparum malaria in India. The combination of DP provided some protection against new infections post treatment. DP can play an important role in controlling and eliminating malaria in India, possibly in the context of multiple first-line treatments of uncomplicated falciparum malaria, which will reduce drug pressure on existing combinations.In conclusion, the fixed dose dihydroartemisinin-piperaquine in this study emerged as a highly efficacious treatment for Nicola Gargano, Silva Tommasini, Antonella Bacchieri and Marco Corsi were from Sigma-Tau developing the product. None of the other authors has any conflict of interest.NG, ST, AB, and MC: Development and manufacturing of drug, DU: Manuscript preparation and overall coordination of study, PCB, BHK and ND: Medical supervision and patient care during clinical trial at Down town, Wenlock Government District and Goa Medical College hospitals respectively, VD, SKG and AK enrolment of patients, follow up and coordination at respective study sites, BS: parasitological assessment and quality assurance, NV: Overall coordination, protocol development, review and editing of manuscript. All authors read and approved the final manuscript."}
+{"text": "New treatment strategies are needed for artemisinin-resistant falciparum malaria. This randomized trial shows that neither increasing nor splitting the standard once-daily artesunate dose reverses the markedly reduced parasite clearance rate in patients with artemisinin-resistant falciparum malaria. Background.\u2003The emergence of Plasmodium falciparum resistance to artemisinins on the Cambodian and Myanmar-Thai borders poses severe threats to malaria control. We investigated whether increasing or splitting the dose of the short-half-life drug artesunate improves parasite clearance in falciparum malaria in the 2 regions.Methods.\u2003In Pailin, western Cambodia (from 2008 to 2010), and Wang Pha, northwestern Thailand (2009\u20132010), patients with uncomplicated falciparum malaria were randomized to oral artesunate 6 mg/kg/d as a once-daily or twice-daily dose for 7 days, or artesunate 8 mg/kg/d as a once-daily or twice-daily dose for 3 days, followed by mefloquine. Parasite clearance and recrudescence for up to 63 days of follow-up were assessed.Results.\u2003A total of 159 patients were enrolled. Overall median (interquartile range [IQR]) parasitemia half-life  was 6.03 (4.89\u20137.28) hours in Pailin versus 3.42 (2.20\u20134.85) hours in Wang Pha (P = .0001). Splitting or increasing the artesunate dose did not shorten half-life in either site. Pharmacokinetic profiles of artesunate and dihydroartemisinin were similar between sites and did not correlate with half-life. Recrudescent infections occurred in 4 of 79 patients in Pailin and 5 of 80 in Wang Pha and was not different between treatment arms (P = .68).Conclusions.\u2003Increasing the artesunate treatment dose up to 8 mg/kg/d or splitting the dose does not improve parasite clearance in either artemisinin resistant or more sensitive infections with P. falciparum.Clinical Trials Registration.\u2003ISRCTN15351875. Plasmodium falciparum on the Cambodia-Thailand border and more recently on the Myanmar-Thailand border jeopardizes the renewed global efforts of control and elimination of malaria [The emergence of partial artemisinin resistance in  malaria \u20134. This  malaria . IncreasA unique property of the artemisinin drugs is their broad-stage specificity including young ring-stage asexual parasites . There iThe current study explored whether increasing or splitting the once-daily AS dose increased efficacy, defined as parasite half-life , in patiIn 2 open-label, randomized clinical trials, we compared different AS dosing regimens in patients with uncomplicated falciparum malaria presenting in Pailin, western Cambodia (2008\u20132010) and in Wang Pha, northwestern Thailand (2009\u20132010), areas with low and seasonal transmission. Studies were conducted according to Good Clinical Practices, and monitored independently . A data safety monitoring committee (DSMC) assured safety of the participants. Ethical approval was obtained from the Ministry of Health in Cambodia, the Ethics Committee of the Faculty of Tropical Medicine of Mahidol University in Thailand, the Oxford Tropical Medicine Ethical Committee, and the World Health Organization Research Ethics Review Committee.Patients with acute uncomplicated falciparum malaria, monoinfection, and asexual stage parasitemia between 10 000 parasites/\u03bcL and 175 000 parasites/\u03bcL assessed by microscopy were eligible provided that written informed consent was obtained from all patients or their guardian (for children). In Pailin, patients of 6 years and older were studied, whereas in Wang Pha only adult patients (\u226518 years) were studied. Pregnant or lactating women were excluded as well as patients with any signs of severe infection . AntimalPatients were randomly allocated to 1 of 4 treatment arms: (1) AS alone in a dose of 6 mg/kg/d for 7 days (AS7); (2) the same total dose, but given as a split twice-daily dose (AS7_split); (3) AS in a dose of 8 mg/kg/d for 3 days, followed by mefloquine in a dose of 15 mg/kg on day 3 and 10 mg/kg on day 4 (8MAS3); (4) the same total dose, but AS given as a split twice-daily dose (8MAS3_split). Arms AS7 and AS7_split were suspended after an association with neutropenia was reported in a separate study . The 7-dRandomization was computer generated in blocks of 20. Opaque sealed envelopes were opened sequentially by a study investigator and contained unique study numbers referring to treatment allocations. AS  and mefloquine  doses were calculated according to body weight. Mefloquine was rounded to the nearest quarter tablet and AS to the nearest half tablet in the single daily dose arm and to the nearest quarter tablet in the split-dose arm. Drug administration was directly supervised. The full dose was repeated if vomiting occurred within 30 minutes after drug administration, and half of the dose was given if within 30\u201360 minutes.P. falciparum infections were treated with quinine combined with doxycycline or clindamycin (children). Adverse events were recorded on a standard form.Patients were hospitalized for 7 days and then followed weekly until day 63 after enrollment. A detailed history and physical examination were performed and blood samples were taken for baseline biochemistry and hematology. Parasitemia was assessed at 0, 2, 4, 6, 8, and 12 hours after enrollment and then every 6 hours until 2 consecutive slides were negative. Quality checks were performed on 10% of randomly selected slides. Plasma concentrations of AS and dihydroartemisinin (DHA) were assessed at 0 (predose), 0.25, 0.5, 1, 1.5, 2, 3, 4, 5, 6, and 8 hours after the first drug administration and after the first dose of the last treatment day (day 3 or 7 depending on treatment arm). Recurrent e transformed parasite clearance curves [msp-1, msp-2, and glurp [The primary endpoint was the parasitemia half-life  defined by the slope of the loge curves . The pare curves . Reinfecnd glurp .Whole blood samples were collected in prechilled fluoride-oxalate tubes and immediately centrifuged at 4\u00b0C. Plasma was stored in liquid nitrogen until analysis at the Mahidol Oxford Research Unit Pharmacology Laboratory in Bangkok. AS and DHA were extracted from plasma samples using solid phase extraction and quantified by high-performance liquid chromatography with tandem mass spectrometry . Individt test or analysis of variance, otherwise Mann-Whitney U or Kruskal-Wallis tests were used. For categorical variables, proportions were examined using \u03c72 or Fisher exact test. Rates of gametocyte carriage were calculated as total carriage time per person-weeks of follow-up. Efficacy rates were assessed by survival analysis using the Kaplan-Meier method. The rate of increase in reticulocyte counts (up to day 7) and recovery (from day 7 to day 63) was assessed using a random effects model. Analysis was performed using Stata software, version 11.2 (StataCorp).A sample size of 40 patients per arm in split versus single daily dosing of AS in artemisinin-resistant malaria was calculated to be sufficient with \u03b1 = .05 and \u03b2 = .80 to detect a shortening in half-life of 1.2 hours from a mean of 6.0  hours as observed with once-daily dosing in recent studies in Pailin [Supplementary Table 1. Because of the difference in inclusion criteria, in Pailin 22 of 79 (28%) of patients were \u226418 years.A total of 79 patients in Pailin and 80 in Wang Pha were studied between March 2008 and December 2010 . Within sites, there was no difference in half-life between patients receiving 6 or 8 mg/kg per day: median 6.0  versus 6.1  hours (P = .90) in Pailin and 3.2  versus 3.4  hours (P = .90) in Wang Pha .Overall, the median half-life was 6.03  hours in Pailin versus 3.42  hours in Wang Pha (P = .0001) and correlated with admission parasitemia in both sites . Corresponding to this, at 48 hours 66 of 78 (85%) patients remained parasite positive (PPR) in Pailin versus 32 of 77 (42%) in Wang Pha (P < .001). At 72 hours the PPR was 41 of 78 (53%) in Pailin compared to 17 of 77 (22%) in Wang Pha (P < .001).Other measures of parasite clearance  confirmed the markedly slower parasite clearance in Pailin compared to Wang Pha (Table P = .0005). Thirteen of 79 (16%) patients in Pailin and 17 of 80 (21%) in Wang Pha had gametocytemia at some point during the study (P = .54). In gametocytemic patients, the median duration of gametocyte carriage was 4  days in Pailin and 3  days in Wang Pha (P = .72). Gametocyte carriage in patients presenting with gametocytemia was overall shorter in patients treated with a split dose of AS  compared to a single daily dose .The mean duration of patent gametocytemia per person-weeks of follow-up was 0.018  person-weeks in Pailin and 0.029  in Wang Pha (P = .53). Late parasitologic failure, defined as polymerase chain reaction (PCR)\u2013confirmed recrudescence of P. falciparum infection \u22657 days after the start of treatment [FCT was shorter in patients in Wang Pha compared to Pailin, whereas there was no difference in FCT between single and split daily-dose arms in either site Table . ETF defreatment , occurreP = .54 and P = .87, respectively). Adjusted for baseline hematocrit level, recovery of reticulocyte counts after day 6 was faster in Pailin  compared to Wang Pha  and counts were maximal at day 14 in both sites.Nadir in hematocrit level was reached at day 7 in both sites, with a mean daily decrease of 0.45% per day  in Pailin and 0.50%  in Wang Pha. This initial drop in hematocrit level was not correlated significantly with reticulocyte counts. Reticulocyte counts on admission and days 6\u20137 were available from 36 patients in Pailin and 33 in Wang Pha. Compared to admission, there was a median fractional reduction after 5 days of 66.7%  in Pailin and 66.7%  in Wang Pha. In both sites this was not different between patients receiving 6 mg/kg/d versus 8 mg/kg/d AS (Supplementary Table 2). In Pailin, a 29-year-old man treated with a total of 24 mg/kg of AS (8MAS3) developed transient afebrile neutropenia with a nadir of 0.78 \u00d7 103/\u03bcL on day 14, which normalized on day 17. In total, 2 patients in Wang Pha and 1 patient in Pailin developed severe malaria shortly after enrollment and received treatment with intravenous AS and quinine. Of these, a 30-year-old woman in Pailin unfortunately died on the fourth day of hospitalization. DSMC evaluation of the case concluded that severe malaria was the likely direct cause of death.The number of patients reporting minor adverse events were more frequent in Pailin (max) and exposure (AUC0-\u221e) to both drugs  and DHA (P = .0001). There was no correlation between half-life and the individual exposure or maximum concentration of AS or DHA or both drugs combined. Pharmacokinetic parameters were otherwise similar to that previously reported in the same populations [The pharmacokinetic profiles of AS and DHA were comparable between sites Table . Howevers Figure . Compareulations .Table 3This study evaluated the use of an increased AS dose and an increased dosing frequency  in uncomplicated falciparum malaria with varying degrees of resistance to artemisinins. The common dose of AS is 4 mg/kg/d as a once-daily dose for 3 days as part of an ACT. This study showed that increasing the AS dose up to 8 mg/kg/d did not affect parasite clearance parameters, strongly suggesting a reduction in the maximum obtainable effect , 16. The3/\u03bcL) was observed in 19% of patients [3/\u03bcL, with no signs of concomitant infection [Further increase of the total dose of AS is limited by toxicity. A study from western Cambodia has recently shown that AS in a dose of 6 mg/kg/d for 7 days resulted in transient neutropenia with a nadir at 2 weeks after treatment. Severe neutropenia  in Wang Pha, northwestern Thailand . In the Recrudescence rates, which strongly depend on parasite sensitivity to the partner drug, were not different between treatment arms and sites. Despite marked slower parasite clearance in Pailin, the AS-mefloquine combination was borderline efficacious in both sites with PCR-adjusted cure rates at day 63 of 93.3% in Pailin and 90.4% in Wang Pha. In the current study with a relatively small sample size, there was no clear relationship between gametocyte carriage and asexual stage resistance to AS. A larger study is under way to address this important determinant of transmissibility , 22.max) of AS and DHA in Wang Pha compared to Pailin, but individual parasite clearance times were not correlated with Cmax. Total drug exposure was not different between sites. Differences in half-life between western Cambodia and northwestern Thailand can thus not be explained by differences in pharmacokinetics. There was a proportional increase in exposure with increased dosing of AS, which supports dose-independent absorption and clearance of AS and DHA. Doubling the frequency of dosing proportionately increased the duration of plasma parasiticidal drug concentrations, but this did not significantly accelerate parasite half-life. The marked decrease in total exposure for AS and DHA on day 7 compared to admission is likely to be a disease-related effect resulting from an increase in relative bioavailability during the acute phase of the disease. This has previously been reported in pregnant women on the Thai-Myanmar border [In agreement with previous studies, considerable interindividual variation was observed in the pharmacokinetic profiles for AS and the active metabolite DHA, but there were no major differences between sites. There was a higher maximum concentration . Supplementary materials consist of data provided by the author that are published to benefit the reader. The posted materials are not copyedited. The contents of all supplementary data are the sole responsibility of the authors. Questions or messages regarding errors should be addressed to the author."}
+{"text": "Pharmacokinetic (PK) data on amodiaquine (AQ) and artesunate (AS) are limited in children, an important risk group for malaria. The aim of this study was to evaluate the PK properties of a newly developed and registered fixed dose combination (FDC) of artesunate and amodiaquine.A prospective population pharmacokinetic study of AS and AQ was conducted in children aged six months to five years. Participants were randomized to receive the new artesunate and amodiaquine FDC or the same drugs given in separate tablets. Children were divided into two groups of 70 (35 in each treatment arm) to evaluate the pharmacokinetic properties of AS and AQ, respectively. Population pharmacokinetic models for dihydroartemisinin (DHA) and desethylamodiaquine (DeAq), the principal pharmacologically active metabolites of AS and AQ, respectively, and total artemisinin anti-malarial activity, defined as the sum of the molar equivalent plasma concentrations of DHA and artesunate, were constructed using the non-linear mixed effects approach. Relative bioavailability between products was compared by estimating the ratios (and 95% CI) between the areas under the plasma concentration-time curves (AUC).The two regimens had similar PK properties in young children with acute malaria. The ratio of loose formulation to fixed co-formulation AUCs, was estimated as 1.043 (95% CI: 0.956 to 1.138) for DeAq. For DHA and total anti-malarial activity AUCs were estimated to be the same. Artesunate was rapidly absorbed, hydrolysed to DHA, and eliminated. Plasma concentrations were significantly higher following the first dose, when patients were acutely ill, than after subsequent doses when patients were usually afebrile and clinically improved. Amodiaquine was converted rapidly to DeAq, which was then eliminated with an estimated median (range) elimination half-life of 9 (7 to 12) days. Efficacy was similar in the two treatments groups, with cure rates of 0.946 (95% CI: 0.840\u20130.982) in the AS+AQ group and 0.892 (95% CI: 0.787 \u2013 0.947) in the AS/AQ group. Four out of five patients with PCR confirmed recrudescences received AQ doses < 10 mg/kg. Both regimens were well tolerated. No child developed severe, post treatment neutropaenia . There was no evidence of AQ dose related hepatotoxicity, but one patient developed an asymptomatic rise in liver enzymes that was resolving by Day-28.The bioavailability of the co-formulated AS-AQ FDC was similar to that of the separate tablets for desethylamodiaquine, DHA and the total anti-malarial activity. These data support the use this new AS-AQ FDC in children with acute uncomplicated falciparum malaria. Plasmodium falciparum in tropical countries has resulted in an increase in childhood morbidity and mortality, especially in African under fives, the most vulnerable group affected by falciparum malaria  \u03bcmol/L) and median serum creatinine fell from 42.3 (14.1 \u2013 71.6) to 34.8 (18.1 \u2013 105.7) \u03bcmol/L (median change: -7.3 [-34.7\u201334.0] \u03bcmol/L). Median serum alanine aminotransferase (ALT) values changed little over time (23.9 [6.6\u2013174.8] to 25.2 [12.5\u2013759.9] IU/L) with the median (range) of 0.8 (-149.6 \u2013 724.5) IU/L.One FDC group patient whose post Day-2 DeAQ concentration was 616.8 ng/ml developed CTC grade 3 (> 5 \u2013 \u2264 20 \u00d7 ULN) liver enzymes during follow up : AST = 64 IU/L (D0), 294 (D14), 352 IU/L (D21) and 222 IU/L (D28). Corresponding ALT values 23, 389 and 227 IU/L. The patient's bilirubin levels were normal during the whole follow-up period. All other patients with DeAq levels > 600 ng/mL had normal biochemical results.Severe neutropaenia (defined as a neutrophil count of <1000/\u03bcL) rates on Day-0 and Day-28 were 1/132 (0.8%) and 0/84 (0.0%), respectively. Leukopaenia  rates on Day-0 and Day-28 were 2/135 (1.5%) and 0/85 (0.0%), respectively.Measured concentrations of desethylamodiaquine, DHA, and AS in all patients are presented in Figures Positive desethylamodiaquine levels, but negative corresponding AQ levels, were detected in 28 patients (14 on each treatment arm) before the first dose, suggesting previous AQ administration. These levels were modelled assuming first order elimination of the previous dose, and subtraction from the study drug profile.One child, who received a total dose of 39 mg/kg, had a very high plasma concentration of desethylamodiaquine recorded 4 hours after the last dose (1308 ng/mL). However, the plasma concentration on Day-7 (142.6 ng/mL) was within the range for other patients. This patient had a low desethylamodiaquine concentration before the first dose (18.5 ng/mL), which did not explain the subsequent very high level.In total 60 samples (31 in the AS+AQ group and 29 in the AS/AQ group) had detectable levels of AQ, all taken at approximately 4 hours (median [range] = 4 [3.9\u20134.8]) after the third dose, at similar times in the two treatment groups . These levels were not significantly different between treatment arms  median (range): 15.5 (3.8 \u2013 48.9) and 14.1 (5.6 \u2013 54.4) ng/mL, respectively.The pharmacokinetics of desethylamodiaquine was best described by a two compartment open model with first order absorption and elimination. The best fit was obtained with concentration modelled on a log scale and homoscedastic error model. The final estimates of the pharmacokinetic parameters are presented in Table There were no differences in the distribution of the posterior individual estimates of total oral clearance (CL/F) between the two treatment arms: mean  0.61  L/kg/h in the loose formulation arm and 0.63  L/kg/h in the fixed formulation arm; difference 0.024 (95%CI: -0.077 to 0.030) L/kg/h; normal distribution for the estimates could be assumed. This reflected very similar AUCs in the two treatment groups, normalized for an oral dose of 34 mg/kg; median (range) of 57.76 (40.00 \u2013 82.32) ng/mL.h with the loose formulation arm and 54.75 (40.73 \u2013 73.64) ng/mL.h with the fixed formulation. The ratio of the loose formulation AUC to the fixed formulation AUC was estimated as 1.043 (95% CI: 0.956 to 1.138).However, the actual dose received was approximately 10% higher in the loose combination arm so the observed AUCs were also higher: median (range) of 62.5 (24.4 \u2013 95.3) ng/mL.h in the loose formulation group and 48.7 (39.0 \u2013 92.3) ng/mL.h in the fixed formulation group (p < 0.001).In total fourteen patients in the group of patients with measured desethylamodiaquine levels had recurrent parasitaemia: nine in the fixed dose combination group  and five in the loose combination group . Based on pharmacokinetic models these patients had relatively low predicted concentrations of desethylamodiaquine Figure . Predictth, 7th, 28th and 67th percentile, respectively  measurements on Day-14 (median [range]: 25.4 [11.7\u201384.2] ng/mL) were below the median value for the non-recurrent cases , including the only PCR confirmed recrudescence in this group. Four out of five patients with PCR confirmed recrudescence received doses below 10 mg/kg  and their estimated clearance was at the 54% measure-7), but modelling clearance as a fixed parameter which differed with dosing periods improved the model significantly (p < 0.001 for both). This was justified by previous studies which have shown disease related changes in the pharmacokinetics of the AS derivatives [The pharmacokinetics of DHA and the total anti-malarial activity were best described by a one compartment model with volume of distribution fitted as a random effect, on the original scale and homoscedastic error model Table . There wivatives ,19. SomeSince no differences in clearance for both DHA and anti-malarial activity were found between treatment arms (p = 0.73 and p = 0.71 respectively), the corresponding AUCs were estimated as 4.29 (95% CI = 3.48 \u2013 5.59) ng/mL.h and 4.43 (95% CI = 3.61 \u2013 5.75) ng/mL.h after the first dose for all patients, assuming a 3.7 mg/kg dose of AS. After the third dose the clearance approximately doubled so the AUCs were approximately halved: 1.96 (95% CI = 1.47 \u2013 2.93) ng/mL.h and 2.14 (95% CI = 1.59 \u2013 3.25) ng/mL.h, for the same dose.For diseases requiring treatment with a combination of drugs, fixed combinations (FDC) are increasingly considered the optimum therapeutic approach. It is essential, therefore, to confirm that the bioavailability of any newly developed FDC is similar to that of the previously established and validated loose drug combination in the disease being treated.This population pharmacokinetic evaluation of two AS-AQ regimens \u2013 one comprising loose tablets, and the other a newly developed fixed dose co-formulation \u2013 indicates that the two formulations have similar pharmacokinetic properties in young children with acute malaria, the main target group for this treatment. Amodiaquine was converted rapidly to desethylamodiaquine presumably by hepatic metabolism in a \"first pass\" and so it is the metabolite, which contributes most of the anti-malarial activity ,21. The In this study, measured concentrations of desethylamodiaquine had more symmetrical distributions and narrower ranges in the fixed dose combination but this does not reach statistical significance. This is in line with the observations made in the comparative pharmacokinetic study in healthy volunteers in which it appears that the FDC generates less variable results .in vivo and most of the anti-malarial activity derives from the metabolite. The pharmacokinetic parameter estimates of DHA and the total anti-malarial activity, defined as the sum of the molar equivalent plasma levels of DHA and AS, indicated rapid absorption and elimination. As noted in previous pharmacokinetic studies [Artesunate is converted rapidly to dihydroartemisinin (DHA)  studies  plasma lAssociations were identified between outcome and desethylamodiaquine levels on Day-7 and 14. Predicted levels at Day-7 and Day-14 were significantly lower (p < 0.001) in patients with recurrent parasitaemia. For measured concentrations, the association was less clear, probably because of the low number of patients with plasma levels available, but all patients with PCR confirmed recrudescent parasitaemia or with recurrent parasitaemia and missing PCR results had Day-7 concentrations below 75 ng/mL giving 100% cure rate in patients with Day-7 concentrations above 75 ng/mL. This cure rate was estimated as 97% based on the estimated Day-7 concentrations.rd dose, only one had evidence of hepatocellular injury that was probably AQ related and was resolving by Day-28. Transient, AQ induced hepatocellular liver toxicity is well described and is probably a type II immune reaction.There was no indication of a relationship between high blood concentrations of either drug and adverse effects. Of the 10 patients with DeAq blood levels higher than 600 ng/ml post 3Although rates of absorption were not examined in detail because of the sparse sampling, there were no obvious major differences in the observed concentrations collected during the absorption phase for the two drug formulations. The elimination kinetics estimated from sparse data may have missed a slower elimination phase and, therefore, underestimated the terminal elimination phase, but this is unlikely to contribute significantly to the therapeutic response. Importantly, the estimated bioavailability from the co-formulated product was similar to that of the separate tablets. Thus, if there is a longer terminal phase of elimination, it is unlikely to start at different concentrations with the two formulations.These data support the pharmacological equivalence of the new fixed dose AS/AQ formulation with the previously established separate products. The likely improved adherence associated with this relatively well tolerated, simply administered, stable , inexpensive (cost below 0.5/1 USD for child/adults for 3-day treatment) FDC should be a significant operational advantage.ACT: Artemisinin Combination Therapy; AQ: Amodiaquine; AS: Artesunate; AS/AQ: Artesunate and Amodiaquine in a fixed dose combination; AS+AQ: Artesunate and Amodiaquine in a loose dose combination; DHA: Dihydroartemisinin; DeAQ: Desethylamodiaquine; FDC: Fixed-dose combination.NJW is co-chairman of the WHO Global Malaria Programme technical expert group on the prevention and treatment of malaria. This clinical study was funded by the Research Directorate General of the European Commission (Contract N\u00b0 ICA4- CT-2002610046) and the Drugs for Neglected Diseases initiative.KS analysed the data and drafted the manuscript, NJW contributed to the design of the study, data analysis, and manuscript writing. JRK was the project manager for the development of the ASAQ FDC and for the specific study, report and publication. WT was the clinical manager of the study, was part of the protocol development team and reviewed the data and the first draft of the manuscript. SS was the Principal Investigator, was also part of the protocol development team, managed the clinical team in Burkina Faso and reviewed the first draft of the manuscript. JAS designed the PK schedule and contributed to the manuscript writing. CCM analysed the clinical data and was responsible for the clinical study report."}
+{"text": "The widespread use of artesunate-amodiaquine (ASAQ) for treating uncomplicated malaria makes it important to gather and analyse information on its tolerability.An individual-patient tolerability analysis was conducted using data from eight randomized controlled clinical trials conducted at 17 sites in nine sub-Saharan countries comparing ASAQ to other anti-malarial treatments. All patients who received at least one dose of the study drug were included in the analysis. Differences in adverse event (AE) and treatment emergent adverse event (TEAE) were analysed by Day 28.Of the 6,179 patients enrolled (74% <5\u2009years of age), 50%  received ASAQ, 20%  another ACT, and 30%  a non-ACT (combination or single-agent) treatment. Overall, 8,542 AEs were recorded. The proportion of patients experiencing at least one gastro-intestinal AE on ASAQ was 43% (and higher than that with artemether-lumefantrine and dihydroartemisinin-piperaquine at two sites), and was 23% for any other AEs (not different from other treatments). Specifically, the risk of diarrhoea, vomiting, cough and weakness was lower with artemether-lumefantrine; artemether-lumefantrine and dihydroartemisinin-piperaquine carried a higher risk of pruritus, chloroquine-SP had a higher risk of nausea. Parasitological recurrence increased the risk of occurrence of any AE. No other difference was detected. Comparing AE to TEAE in patients who had pre-treatment occurrence and grades of intensity recorded, AEs were significantly more related to the pre-treatment prevalence of the symptom ; AEs overestimated TEAEs by a factor ranging from none to five-fold. The overall incidence of serious AEs (SAEs) with ASAQ was nine per 1,000  and mortality was one per 1,000 ; both were similar to other treatments.ASAQ was comparatively well-tolerated. Safety information is important, and must be collected and analysed in a standardized way. TEAEs are a more objective measure of treatment-induced toxicity. Plasmodium falciparum malaria [Artemisinin-containing combination therapy (ACT) is the World Health Organization (WHO) recommended first-line treatment of acute uncomplicated malaria . One sucTM FDC had been ordered by 2012 for the private sector of the seven African countries  included in the AMFm  pilot phase (from 2010 to February 2012)[Worldwide, ASAQ (in its various formulations) is the second most widely used ACT. The volumes of ASAQ FDC have increased from less than one million treatment courses in 2007 to 41 million in 2010 . Over 23ry 2012).Although ACT is regarded as highly effective and generally well tolerated, safety remains largely under-reported. Synthesizing available information using individual patient data for randomized controlled trials (RCTs) can help better define treatments in terms of risk-profiling and risk-management. There is also a real need to develop a standardized system for generating and analysing tolerability data from anti-malarial drug efficacy studies; while an excellent standardized system for efficacy analyses exists, there is no equivalent for safety and tolerability.For this study, relevant information was retrieved from randomized controlled trials (RCTs) identified through a systematic review for which individual patient data were made available. This analysis included a majority of children under five years old from sub-Saharan Africa since they represent 86% of the death toll due to malaria in the world .The analysis of tolerability was by intent-to-treat (ITT), including all participants who were randomized to the study medications and took at least one dose and followed-up to Day 28. The primary study endpoint in all the RCTs included in the analysis was efficacy. Follow-up ceased at the time of parasitological failure (either primary or recurrence), protocol violation, loss to follow-up, and no tolerability data were recorded thereafter. Signs or symptoms were recorded at enrolment (Day 0) and tolerability outcomes were recorded post-treatment from Day 1 to Day 28 according to study procedures.The studies were identified through a systematic review of clinical trials and personal contacts, regardless of language or publication status . Published studies were identified through electronic searches up to April 2007 of MEDLINE, EMBASE, LILACS, the Cochrane Infectious Diseases Group's trials register and the Cochrane Central Register of Controlled Trials (CENTRAL) using the search terms: malaria, amodiaquine, artesunate, and artemisinin. To be included, studies were to be RCTs conducted in Africa comparing any formulation of ASAQ to any other single or combination treatment of uncomplicated falciparum malaria with follow-up of at least 28\u2009days .For the studies meeting these criteria, investigators were contacted to provide individual patient data and datasets received were examined for inclusion.Trials were all randomized comparative: one was double-blinded , four weThe tolerability outcome measure was any treatment adverse event (AE) and treatment-emergent adverse event (TEAE) occurring within 28\u2009days of starting treatment. In each of the studies included, signs or symptoms were actively screened during follow-up -14.An AE was defined as any sign or symptom occurring after the start of treatment (first drug intake), irrespective of whether that sign or symptom was present at baseline or not, of its severity and drug-event relationship.A TEAE was defined as the worsening of the condition - ie any occurrence of an abnormal condition as compared to baseline in patients who either had a normal condition pre-treatment or an abnormal condition that was of lower intensity than that recorded post-treatment . Intensity was graded nought to four  - this analysis was only possible in the sub-group of patients from the sites who graded signs and symptoms; in two other studies ,9 gradesSerious adverse events (SAEs) were defined as events that were fatal, life threatening or required admission to hospital, irrespective of drug-effect relationship. Death was also reported separately. The incidence of AEs and TEAE were calculated for each study and treatment arm after standardizing AE terminology (i) translating French into English language (ii) including redness, swelling, burning, itching, rash skin symptom (iii) grouping weakness with fatigue and asthenia.The disposition of the two components of the ASAQ combination is very different; AS is rapidly absorbed and eliminated , while the estimated median (range) elimination half-life of desethyl-AQ (AQ active metabolite) is around nine (seven to 12) days. The same is also true for the other ACT. Therefore most of the AEs will likely happen before Day 28.Data were standardized for an ITT analysis. Incidences (proportions of patients experiencing an event) were calculated for each treatment group and each sign or symptom and defined as the number of patients reporting the event divided by the number of patients initially at risk (exposed). As all studies did not provide information on the exact day of the event, it was not possible to calculate the overall time-to-event expressed as person-day to calculate incidence rate ratios, and AEs and TEAEs were analysed as a binary variable occurring per patient. The risks of experiencing AEs/TEAEs with ASAQ compared to other drugs were measured by using multivariate logistic regression. A random intercept for each site was included when the Lagrange multiplier (LM) test  was signAn additional analysis comparing AE and TEAE was included using the sub-group of patients with sign or symptom recorded and graded both pre- and post-treatment. Categorical data were compared using the chi-square or the Fisher exact test. The non-normally distributed correlation used the Spearman test and compared by the Fisher test. Confidence intervals were calculated at 95% (95%CI), and comparisons considered significant when P\u2009<\u20090.05. Data were analysed using Stata v10 (Stata Corp.).The majority of the patients were treated with individually formulated (loose) AS and AQ. The target dose was AS 12\u2009mg/kg over three days and AQ 30\u2009mg/kg over three days except in Uganda where AQ was given at 25\u2009mg/kg . The loose combinations ASAQ were given based on body weight, while in two studies the fixed combination (FDC) ASAQ was given based on age and weight ,14. The ACTs: AL (20\u2009mg artemether/120\u2009mg lumefantrine given according to weight as one (5\u201314\u2009kg), two (15\u201324\u2009kg), three (25\u201334\u2009kg), and four (\u2265 35\u2009kg) tablets given twice daily co-administrated with fat for 3\u2009days); DP (2.3\u2009mg/kg/day dihydroartemisinin and 18.4\u2009mg/kg piperaquine for three days); AS\u2009+\u2009SP ;Non-ACTs: AQ\u2009+\u2009SP (AQ 10\u2009mg/kg/day for three days and SP 25\u2009mg/kg of sulphadoxine and 1.25\u2009mg/kg/of pyrimethamine administered in a co-formulated tablet SP as a single dose); CQ (25\u2009mg/kg chloroquine over three days) and SP; AQ mono-therapy (10\u2009mg/kg/day for three days); AS mono-therapy (AS 12\u2009mg/kg over five days).All studies have been approved by the relevant ethics and institution review committees -14.A total of 6,179 patients was enrolled in eight studies conducted at 17 sites in nine countries (Table (i) Two multi-country studies n\u2009=\u20091,879): one comparing ASAQ FDC  to AL (n\u2009=\u2009312) in Mendong-Cameroon; Tsiroanomandidy-Madagascar, Bancoumana-Mali, Mlomp-Senegal [9: one co(ii) Two single centre study were conducted in Pouytenga-Burkina Faso comparing ASAQ fixed dose  to the loose ASAQ combination (n\u2009=\u2009375)  and in B(iii) Two studies conducted in a single country with multiple sites in Rwanda-Mashesha, Rukara, and Kicukiro comparing loose ASAQ (n\u2009=\u2009252) to AQ\u2009+\u2009SP (n\u2009=\u2009258) and DP (n\u2009=\u2009252) ; and in (iv) Two different studies in Uganda used the same protocol in three common sites: Apac, Jinja and Tororo comparing loose ASAQ (n\u2009=\u2009761) to AL (n\u2009=\u2009204), AQ\u2009+\u2009SP (n\u2009=\u2009550), CQ\u2009+\u2009SP (n\u2009=\u2009519),14.Individually, the sites enrolled between 174 and 949 patients treated with either ASAQ, which constituted 50%  of the total patient population, or a comparator: another ACT  or non-ACT 30%, 1,849/6,179)  and DP  than with ASAQ, resulting from differences seen in two sites only; the occurrence of GI AEs was significantly related to the risk of parasitological recurrence ; older patients were at lower risks for GI . For other AEs, no difference between treatment groups was detected. The risk was significantly related to the risk of parasitological recurrence , age , and duration of follow-up .There was no difference between the loose and fixed dose ASAQ combination; no other risk was detected.In patients treated with ASAQ Table , the incp\u2009=\u20090.59). In Mali [In Rwanda , 11.5% oIn Mali , there wIn Mali , six patIn Mali , 13% 12p\u2009=\u20090.59.In Mali .Overall , multivariate analysis for each GI AE showed that compared to ASAQ Figure , patientOther risk differences were related to age: older patients were at lower risk for vomiting, anorexia and diarrhoea, and at higher risk for abdominal pain ; the risk of anorexia  was higher in patients with a parasitological recurrence as well as the risk of diarrhoea , which increased over time .The incidence of headache on ASAQ was 9%  increasing with age  and higher in patients treated with CQ\u2009+\u2009SP  than ASAQ. Dizziness was infrequent (1%) in all groups. One case (in 848) of mild tinnitus was reported with loose ASAQ on the day of recrudescence (Day 21). Patients treated with CQ\u2009+\u2009SP  were at higher risks for this hearing disturbance compared to ASAQ . Romberg test and nystagmus were negative in all the patients screened . The incidence of pruritus was 19% (193/1636) with ASAQ; the risk was comparatively higher with AQ\u2009+\u2009SP  and AL . The incidence of skin conditions  was 16% (118/759) on ASAQ (not different from other groups).The incidence of weakness with ASAQ was 20% 539/2,749), and the risk was higher in patients with a parasitological failure  and lower in patients treated with AL, resulting from a significant incidence difference found in Madagascar  . No difference between treatments was detected, but older patients were at lower risks , and the risk increased during the follow-up . Jaundice was infrequent (<1%) in all groups.In this sub-group, 3,943 TEAEs or 6,294 AEs occurred in 3,334 patients who had received at least one treatment dose Figure . Some 1,Using multivariate analysis, compared to ASAQ, patients treated with DP were at lower risk for GI AEs or TEAEs , while patients treated with AQ were at lower risk for GI TEAEs  but not AEs (p\u2009=\u20090.927). For any other AEs or TEAEs, no difference between treatments was observed. Older patients were at significant lower risks for gastro-intestinal AE and TEAE. Between-treatment comparisons for each TEAE are presented using forest plots in Figure The incidence of AEs and TEAEs following ASAQ treatment is presented in Table The comparison between prevalence of signs/symptoms on admission and AE and TEAE intended to quantify the difference in appreciation of drug tolerability when reporting TEAEs or AEs (i.e. whether accounting for pre-treatment intensity of signs/symptoms).Table For GI events, the prevalence of diarrhoea at enrolment was 20%; the incidence of AEs and TEAEs was 25% and 17% respectively, and the difference between the two was +47% . For nausea (18% at enrolment), the difference between AE and TEAE was +386%.For neurological effects, headache was common at enrolment (46%) and decreased post-treatment . Both dizziness and tinnitus were infrequent at enrolment and after treatment. For dermatological symptoms, the prevalence of pruritus was 26% pre-treatment, and the incidence of AEs and TEAEs was 25% and 19% (difference +36%), meaning that 6% of the patients had the same or decreasing symptom intensity during follow-up.Regarding other signs or symptoms, cough was the most frequent at enrolment (69%); 16%  of patients had subsequently decreasing or stable intensity. Weakness was also very frequent at enrolment (59%) with 26% of patients with a decreasing or stable intensity post-treatment. Muscle pain was quite frequent at enrolment (32%), and there was no significant difference between TEAE (8%) and AE incidence (10%); most of the patients developing muscle pain after treatment did not have that symptom at enrolment, while others recovered. Jaundice was infrequent .Overall six deaths occurred in the studies, corresponding to a mortality of one per 1,000  with ASAQ, as well as with comparators . None of them was attributed to the study treatment. In Senegal  two patiTwo male patients in Pouytenga-Burkina Faso, recruited for uncomplicated malaria and treated with the loose and FDC ASAQ respectively, died of severe cerebral malaria occurring after the first day of treatment . One patIn Uganda , two patNo deaths occurred in the studies included in the studies conducted in Gabon and Senegal  Mali 1, Rwanda The incidence of other SAEs (except deaths) was nine per 1,000  with ASAQ and six per 1,000  with comparators (p\u2009=\u20090.124).In Burkina-Faso , SAEs ocIn Rwanda  one seizIn Uganda  20 SAEs In Uganda  SAEs occIn Mali , two SAEIn a multi-centre study , four paIn Senegal and Gabon , three pIn Zanzibar , nine paWhile not exhaustive, this is so far the largest dataset of individual-patient tolerability data compiled on an ACT (ASAQ); it includes a sizeable number of patients  with tolerability outcomes , and is representative of the spectrum composition of malaria patients (it comprises mostly children under five years of age (74%) from areas of moderate to high intensity of malaria transmission of nine sub-Saharan African countries). In more than half of these patients , events were recorded pre-treatment and graded in terms of intensity, which allowed identifying and analysing treatment emergent adverse events (TEAEs).Both efficacy and tolerability information is essential to guide treatment policy. However, while millions of ACT treatments are given every year, and thousands of patients are enrolled in trials with ACT, little tolerability information is available. Tolerability data are often cursorily presented in papers and difficult to standardize and summarize. The availability of individual patient data and the use of standardized methods for analysis (including ) made itOver 3,100 of these patients received ASAQ formulated as different products  by various manufacturers. Therefore, the conclusions reported here do not relate to an individual drug product, but rather the collective tolerability profile of ASAQ. Overall, the risk of experiencing an AE was not different in patients exposed to ASAQ or other forms of ACT and non-ACT, with some exceptions.This analysis also provides important information on a number of safety-related issues.Risks varied across the different trial sites; clinical safety appreciation differs from site to site and probably from investigator to investigator. Also, the type of data and the way they are collected varies. While standard criteria exist , it woulOne particular example is vomiting. Vomiting is a symptom of malaria, but can also be induced or made worse by treatment. In studies where treatment is given under direct observation it is possible to monitor the patients for the first hour post-dosing in case drug is vomited; this also allows re-administering the dose (in toto or in part) when so required, and distinguishing between early and late vomiting . Only onMultivariate logistic regression analysis, as well as the comparison of TEAEs and AEs, pointed to the importance of looking to the considerable background noise generated by malaria itself (pretreatment and recurrence) as well as other diseases and conditions. For instance, the risk of GI events in general  and other AEs (weakness) was higher in patients with recurrent malaria; cough  increased with follow-up time; older age was associated with a lower risk of most AEs.It is important to distinguish the signs/symptoms of malaria from those that may be caused by the treatment. Assessing the drug-event relationship is highly subjective and prone to bias. Recording the occurrence and severity of (a defined set of) events before treatment is administered allows a better appreciation of the real contribution of a treatment to a patient\u2019s status. When this is done , it is possible to report TEAE - ie the occurrence post-treatment of a sign/symptom that was not present before treatment, or its worsening with treatment. This applies also to RCTs, where randomization is expected to even out the risks of events at enrolment, as one is interested not only on the relative (between-treatment) but also the absolute risks for toxicity. These analyses showed that the risks, when considering AEs over TEAEs, ranged from no difference  to double (vomiting), triple , five-fold  differences. The relationship with prevalence of signs/symptom at enrolment was greater for AEs compared to TEAEs (meaning that the definition of AE is less independent of the sign/symptoms related to the disease itself).Traditionally, the main concern in malaria has been efficacy, which has driven treatment recommendations (and a living databases of efficacy has been created ). SafetyIn conclusion, this paper provides both (i) a comparative evaluation of the safety risks following various ACT on a large, representative sample of malaria patients; and (ii) a proposal for improved methods to assess safety risks.ACT: Artemisinin Combination Therapy; AE: adverse event; AL: Artemether-Lumefantrine; AQ: amodiaquine; AS: artesunate; CQ: chloroquine; DP: Dihydroartemisinin-piperaquine; LM: Lagrange multiplier test; Max: maximum; Min: minimum; RCT: Randomized Controlled Trial; SP: sulphadoxine-pyrimethamine; TEAE: treatment emergent adverse event; WHO: World Health Organization.The authors declare that they have no competing interests.JZ and PO designed the analysis, interpreted the data and prepared the manuscript. All authors read and approved the final manuscript.P. Olliaro is a staff member of the WHO; the authors alone are responsible for the views expressed in this publication and they do not necessarily represent the decisions, policy or views of the WHO."}
+{"text": "Plasmodium falciparum malaria. In these studies, the categorical criterion proposed by the World Health Organization (WHO) to assess the relative effectiveness of anti-malarial drugs was repeatedly evaluated on Days 14, 21 and 28 after treatment initiation. The aim of the present study was to compare the effects of different treatments on this repeated ordinal outcome, hence using the fully available information.Artemisinin-based combination therapies (ACT) are widely used in African countries, including Cameroon. Between 2005 and 2007, five randomized studies comparing different treatment arms among artesunate-amodiaquine and other ACT were conducted in Cameroonian children aged two to 60 months who had uncomplicated via the WinBUGS software. After selecting the best model using Deviance Information Criterion (DIC), mixed treatment comparisons were based on the estimated treatment effects.The quantitative synthesis was based on individual patient data. Due to the incomplete block design concerning treatment arms between different trials, a mixed treatment comparison (MTC) meta-analysis approach was adopted. The repeated ordinal outcome was modelled through a latent variable, as a proportional odds mixed model with trial, period and treatment arms as covariates. The model was further complexified to account for the variance heterogeneity, and the individual log-residual variance was modelled as a linear mixed model, as well. The effects of individual covariates at inclusion, such as parasitaemia, fever, gender and weight, were also tested. Model parameters were estimated using a Bayesian approach Modeling the residual variance improved the model ability to adjust the data. The results showed that, compared to artesunate-amodiaquine (ASAQ), dihydroartemisinin-piperaquine (DHPP) was significantly more efficacious. Artesunate-chlorproguanil-dapsone (ASCD) was less efficacious than artesunate-sulphadoxine-pyrimethamine (ASSP), artemether-lumefantrine (AMLM) and DHPP, the difference with the latter being significant. No difference in efficacy was found between ASAQ and AMLM.P. falciparum malaria in Cameroonian children.Bayesian mixed treatment comparisons of a network of connected randomized trials with repeated measurements of the primary categorical outcome allowed to take into account both the individual- and between- studies sources of heterogeneity. The results of the present study complete the previous quantitative review based on a binary outcome at a fixed time point, suggesting that DHPP represents an alternative for the treatment of uncomplicated  Plasmodium falciparum particularly affects both the young children and pregnant women. Eighty-five percent of the deaths concern children less than five years of age ). DHPP was more efficacious than ASCD . As compared to AMLM, the combination ASCD was less efficacious ; ASCD was found less efficacious than ASSP , though both differences were not significant. All the other direct and indirect comparisons did not differ significantly as well.ASAQ and DHPP treatments differed significantly. DHPP was globally more efficacious than ASAQ  as it was not connected to the other ones, whereas the four remaining studies  were analysed with a mixed ordinal logistic model incorporating a between-subject heterogeneity variance model. This approach can be considered as an extension of the multi-treatment approach of Jansen et al., which wl.[et al., where tRegarding the categorical outcome, LCF is symptomatic, whereas LPF is not. It remains possible that a patient with LPF become symptomatic beyond day 28. However, the protocol was designed to separate the two endpoints when performing a 28-day treatment evaluation. The clinical implications of LPF and LCF on day 28 seem to be quite different. Moreover, the more recent WHO document  maintainet al.[In the present study, analyses were based on the observed treatment responses between day 14 and day 28 (due to the absence of ETF), and the contribution of each observed category was evaluated. Pooling randomized clinical trials raises the issue of heterogeneity between studies. However, all studies included in this analysis were based on the same population of children, within the same age range and in the same geographic area. These studies had the same design and were run by the same investigators and field workers over the years. Mixed treatment comparison (MTC) meta-analysis faces several limits leading to the possibility of biased estimates. Comparing treatment arms using indirect comparisons apparently exposes to the loss of the benefits of randomization. However, it is partially preserved using adjusted comparisons with possibly less biased differences towards positive results, according to Song et al.. A studyet al., or a loet al.. AnalyseFrom the clinical standpoint, it is worth noting that the present data set concerned the use of highly efficacious anti-malarial combination drugs, thus explaining the absence of the ETF category. AMLM is already an alternative to ASAQ in Cameroon. Both ASAQ and AMLM treatments are recommended by the WHO, based on several published trials, comparing different subsets of the treatments listed in the present analysis -38. The et al.[DHPP showed a higher efficacy as compared to the reference treatment ASAQ in all tested models, whereas ASCD appeared less efficacious than ASAQ. AMLM did not differ significantly in efficacy from ASAQ. It should be remembered that the present analysis discarded study #2, as it was unconnected to others, increasing the power for comparison between the remaining treatment arms. The final result is in agreement with the meta-analysis conducted by Sinclair et al., which wACT, Artemisinin-based combination therapy; AMLM, Artemether - lumefantrine; ASAQ, Artesunate - amodiaquine; ASCD, Artesunate - chlorproguanil - dapsone; ASSP, Artesunate-sulphadoxine-pyrimethamine; DHPP, Dihydroartemisinin - piperaquine; ASMQ, Artesunate - mefloquine; AQSP, Amodiaquine - sulphadoxine - pyrimethamine; PP, Per protocol; WHO, World Health Organization.The authors declare that they have no competing interests.SWY developed the analysis plan and carried out both the statistical analyses and software implementations under the close supervision of AS and JCT. She also drafted the manuscript. LKB was responsible for the overall data collection and supervision of clinical trials. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum and treated with therapeutic doses of AS, AE, and AL. Course of infection was studied in the infected and treated groups up to day 42. Peak level of parasitaemia (38%) was observed on day 11 in infected group. Haematological indices indicated significant (P < 0.05) decrease in RBC, WBC, haemoglobin, packed cell volume, mean cell volume, and platelet counts in infected mice. But all the parameters were restored to normal values, and significant (P < 0.05) changes were observed in all drug-treated groups. Insignificant changes were observed for MCHC (P > 0.05) in all drug-treated groups. Percent of peak parasitaemia was much reduced in AL- (3.2% on day 3) treated group in comparison with AE- (2.4% on day 4) and AS- (4% on day 2) treated groups. Parasites were completely cleared on day 6 in AS group, day 5 in AE group, and day 4 in AL group. Hence, our results strongly support that combination therapy has high efficacy rates than monotherapy. No adverse effects were observed on haematological parameters when animals were treated with therapeutic dosages.To find out the efficacy and effect of artemisinin derivatives on haematological indices, C57BL/6J mice were challenged with  Plasmodium falciparum and occurrence of systemic complications. Most of the systemic complications from malaria are mainly because of hyperparasitaemia [Despite advances in knowledge, malaria continues to cause significant morbidity and mortality worldwide. Over 40% of the world population lives in malaria-endemic areas and it is very high 20%) in severe malaria (parasitaemia > 5%). Today malaria is the most important problem for which an estimated 300\u2013500 million cases were recorded and 1.5\u20132.7 million deaths occur each year [0% in sevBlood is the most easily accessible diagnostic tissue. Variations in haematological parameters are influenced by any disease condition which affects the haemopoietic physiology. This is likely to happen with an endemic disease such as malaria that affects the host homeostasis . The tarHaematological changes associated with malaria infection are well recognized, but specific changes may differ from the level of malaria endemicity, background haemoglobinopathy, nutritional status, demographic factors, and immunity to malaria . Hence, P. falciparum. Hence it is widely used for the treatment of malaria [\u03b1, \u03b2 arteether (30\u2009:\u200970 mixture of enantiomers) is a fast-acting blood schizonticide completed multicentric clinical trials in P. falciparum endemic areas and was found to be effective and marketed in 1997 as E Mal. Arteether has a higher safety margin compared to other artemisinin derivatives, has longer half-life, and produces high cure rates when administered in a short 3-dose (i.m.) regimen [Artemisinin(s) monotherapy and combination therapy (ACTs) are currently used as first line treatment for uncomplicated malaria. Artesunate is a semisynthetic derivative of artemisinin consisting of sodium succinyl salt of dihydroartemisinin, a potent blood schizonticide highly effective against multidrug resistant strains of  malaria . \u03b1, \u03b2 ar regimen . Artemetoral), arteether (i.m.), and artemether + lumefantrine  on haematological parameters. The present investigation was also aimed to study the course of infection in falciparum-infected and drug-treated mice to know the efficacy of Artemisinin derivatives based on parasite clearance time (PCT). Thus the aim of our study was to investigate the different hematological changes with P. falciparum malaria and to define the possible role of Plasmodium species in the pathogenesis related to haematological changes.The present study was aimed to reveal the effect of 3 frequently used artemisinin derivatives, namely, artesunate , National Institute of Nutrition (NIN), Hyderabad, India, and allowed to acclimatize for 15 days. Animals were fed with standard feed daily and water was given  Animals . P. falciparum was collected from an infected person from Government General Hospital in Guntur, Andhra Pradesh. Patient is tested with SD BIOLINE Malaria rapid test (P.f/p.v) and blood smear examination by 10 to 15 years experienced microbiologist who confirmed P. falciparum with high parasitaemia. At the time of blood collection the patient does not receive any preantimalarial treatment even paracetamol. The blood containing P. falciparum was extracted from peripheral vein of hand of the infected individual using 5\u2009mL sterile syringe. Immediately blood was transferred to EDTA vacutainer ; kept in thermocol ice box, and transferred to the laboratory. The obtained antigen was inoculated into the mice within half an hour of collection.For the present experiment the species of P. falciparum-infected erythrocytes was diluted with PBS. After obtaining P. f. antigen, the parasites were maintained experimentally in three C57BL/6J male mice, and the level of parasitaemia was monitored after the next day of inoculation by smear preparation. After the achievement of high level of parasitaemia (stock blood), blood samples were collected and diluted in normal saline at the ratio of 75% parasitized blood and 25% PBS. The diluted parasitized blood was then inoculated into different experimental mice groups on day \u201c0\u201d via intraperitoneal (i.p.) route because malaria parasite penetrates the peritoneal wall into the blood stream within one minute of inoculation. Mice of control group were inoculated with distilled water on day \u201c0\u201d and maintained as control.The collected blood was washed several times in phosphate buffered saline  by centrifugation at 1000\u2009rpm/15\u2009min. The washed erythrocytes were suspended in PBS and packed by centrifugation, and supernatant was removed by a pipette. The sediment with Artesunate: through oral gavage in 4 (double divided dose), 2, 2, 2, and 2\u2009mg/kg body weight for five days. Arteether: intramuscularly in 3\u2009mg/kg body weight for three days.Artemether + Lumefantrine: through oral gavage in 3.5\u2009mg/kg body weight for three days.A total number of 40 C57BL/6J male mice were distributed into 5 groups, namely, control (CON), infected (INF), drug treated with artesunate (DT AS), drug treated with arteether (DT AE) and drug treated with artemether + lumefantrine (DT AL) of 8 animals each. For all drug-treated groups, therapeutic dosages according to WHO recommendation were administered. Doses were calculated according to the average body weight of the mice (approximately 30\u2009gm/mouse). For monotherapy, artesunate (AS) tablets  from Zydus Cadila Health Care Limited, India, and arteether (AE)  from Themis Chemicals Limited, Mumbai, India, were obtained. For combination therapy, artemether + lumefantrine (AL) (Lumerax-20 DT) from Ipca Laboratories Limited, India, was obtained.Course of infection was studied in all the 12-week C57BL/6J experimental mice. Parasitaemia was monitored daily up to day \u201c42\u201d by making peripheral blood smears. Comparison of parasitaemia between infected mice and drug-treated mice revealed the efficacy of each drug used for the study. Thin blood films were made on the prelabelled slide with free flowing whole blood directly from the mouse tail snips after the first drop was wiped off with cotton wool. The blood films were stained with JSB I and JSB II for the detection of malarial parasites and for estimation of parasitaemia. After staining, slides were washed with tap water to remove excess stain and allowed to drain in a vertical position and to air dry. A field was selected where the RBCs are in an evenly distributed monolayer and observed under 100x oil immersion objective. A minimum of 1000\u2009RBCs were counted from 10 fields under microscope, and the number of infected RBCs will be recorded. The percent of parasitaemia was determined by enumerating the number of infected RBCs per total number of RBCs counted (5):Parasite clearance time (PCT) was defined as the time from the start of treatment until first negative blood smear for asexual stages which remained negative for an additional 24 hours. For our study we followed 42-day followup for all treatment groups. Thus PCT was observed in all the experimental groups. \u00b5L of blood was aspirated into a needle divided and distributed to the various chambers for sample analysis.At the end of the series of experiments , 6 animals in all the drug-treated groups were sacrificed using chloroform anaesthesia. Six animals in infected and six animals in control group were sacrificed on day 28. Two animals in all the groups kept for 42 days, and one animal is used to calculate parasitaemia on each day. Blood samples were collected by cardiac puncture into EDTA vacutainer to see the frequency of haematological abnormalities especially anaemia, thrombocytopenia, and reduced blood counts in malaria. Whole blood was immediately analyzed for complete blood picture (CBP), that is, red blood cell (RBC) count, haemoglobin (HGB), hematocrit (HCT), mean cell volume (MCV), mean cell haemoglobin (MCH), mean cell haemoglobin concentration (MCHC), white blood cell (WBC) count, and platelet (PLT) count using the fully automated ABX Pentra 60 + Analyser . Briefly, 53\u2009t-test with MINITAB 11.12.32. Bit statistical package and graphs were drawn from MS Excel 2010. The values were given as mean \u00b1 SD and are statistically significant at t > 2.306, P < 0.05* (significant), P < 0.001** (more significant), and P < 0.0001*** (highly significant). P value more than 0.05 was considered as statistically not significant (NS).The means, standard deviations of normally distributed data were compared between control versus infected group and infected versus drug-treated groups using Student's  P. falciparum erythrocytic stages were developed in the blood of C57BL/6J mouse model after inoculation of 0.3\u2009mL. of P. f antigen intramuscularly to the experimental mice. Various stages of parasite are seen in the blood films stained with JSB-I and JSB-II solutions. The blood films were scanned under high power objective (\u00d740) and examined closely under oil immersion objective (\u00d7100). The various erythrocytic stages of P. falciparum were seen such as trophozoites (immature and mature), schizonts, and gametocyte. Appearance of ring forms at the edge of RBC is the characteristic feature of Plasmodium falciparum species  group, the parasites started developing in the peripheral blood from day \u201c1\u201d onwards and reached to peak level on 11th day with 38% parasitaemia. Then the parasitaemia gradually decreased and completely disappeared by day 28  on successive 3 days. After Arteether treatment, the peak level of infection was observed on day \u201c2\u201d with 4.2% parasitaemia. Then the parasites were completely disappeared by the 5th day  was administered orally for the next three days. With the combination therapy, peak level of infection was observed on day \u201c3\u201d with 3.2% parasitemia, and no parasites were observed on day \u201c4\u201d till the 42nd day (After giving 42nd day . The ParP < 0.0001), RBC is 5.9 \u00b1 0.201 (P < 0.0001), PCV is 2892 \u00b1 1.46 (P < 0.0001), MCV is 81.07 \u00b1 2.19 (<0.0001), MCH is 27.67 \u00b1 1.17 (0.0001), PLT is 516 \u00b1 23.4 (<0.0001), WBC is 6957 \u00b1 102 (<0.0001), neutrophils is 28.3 \u00b1 0.724 (P < 0.0001), lymphocytes is 69.70 \u00b1 2.00 (P < 0.0001), and eosinophils is 1.0 \u00b1 0.669 (P < 0.014). And these values were significantly decreased when compared to the control values , respectively. MCHC  and monocytes  have not shown significant change in the infected group when compared to control values , respectively. P < 0.0001), RBC (P < 0.0001), PCV (P < 0.01), MCV (P < 0.01), PLT (P < 0.0001), WBC (P < 0.001), neutrophils (P < 0.0001), lymphocytes (P < 0.0001), and eosinophil (P < 0.05) in artesunate-treated group when compared with control. MCH (P > 0.05), MCHC (P > 0.05), and monocytes (P > 0.05) did not show statistically significant difference between INF and DT AS groups.P < 0.0001), RBC (P < 0.0001), PCV (P < 0.05), PLT (P < 0.0001), WBC (P < 0.001), neutrophils (P < 0.0001), lymphocytes (P < 0.0001), and eosinophil (P < 0.05) values were significantly increased in DT AE group when compared to INF group. MCV (P > 0.05), MCH (P > 0.05), MCHC (P > 0.05), and monocytes (P > 0.05) have not shown significant change in DT AE group when compared to INF group. P < 0.0001), RBC (P < 0.0001), PCV (P < 0.01), MCV (P < 0.001), PLT (P < 0.0001), WBC (P < 0.001), neutrophils (P < 0.0001), lymphocytes (P < 0.01), and eosinophil (P < 0.05) in artemether + lumefantrine-treated group have significantly increased when compared to the infected group. Insignificant changes were observed for MCH (P > 0.05), MCHC (P > 0.05), and monocytes (P > 0.05) between INF and DT AL groups.falciparum malaria is associated with large parasite burdens. The peripheral blood parasite count is the prognostic indicator that correlates with the total parasite biomass and thus the severity of falciparum malaria. Haematological abnormalities are considered a hallmark of malaria and reported to be the most pronounced in P. falciparum infection, probably as a result of the higher levels of parasitaemia [falciparum-infected C57BL/6J mice using an experimental model.Severe sitaemia . This stfalciparum-infected mice when compared to control animals. The reduction was more evident in falciparum infection. It is thought to result from a combination of haemolysis of parasitized red blood cells, accelerated removal of both parasitized and innocently unparasitized red cells, depressed as well as ineffective erythropoiesis with dyserythropoietic changes, and anaemia of chronic disease [In this study we observed that there is a marked reduction in RBC count, WBC count, haemoglobin (HGB), packed cell volume (PCV), mean cell volume (MCV), and platelet counts in  disease . Other f disease . Malaria disease .The low PCV value with a correspondent high density of malaria parasite was suggested to be due to excessive destruction of red blood cells by the malaria parasites. As noted in this study, it was established that the more malaria parasites in the blood circulation cause more destruction of the red blood cells as demonstrated by the low PCV. The low PCV may necessitate the need for blood transfusion which in turn has a high risk of transmitting viral hepatitis, HIV, and other associated risks. The significant reduction in PCV level indicates a relationship between malaria parasite and anaemia .P > 0.05) and monocyte (P > 0.05) values in infected mice when compared to control mice. MCHC levels were not significantly changed which is consistent with the earlier reports [P < 0.0001). Leucopenia appears to be a common finding in both nonimmune patients with falciparum malaria and semi-immune children living in malaria-endemic regions [Statistically insignificant changes were observed in MCHC  than control animals. Thrombocytopenia seems to be due mainly to a reduced platelet life span and splenic pooling. The reduced platelet life span may be caused by binding of malaria antigen onto platelets followed by antibody-mediated phagocytosis [It is a general consensus that thrombocytopenia is very common in  malaria , 28 and ocytosis  or to plocytosis . The relocytosis .Plasmodium parasite (as monotherapies); combination therapies consisting of artemisinin(s) and other antimalarial drugs have been demonstrated to have better parasite clearance and efficacies [++, these drugs generate carbon centered free radicals that could damage the RBC membrane or cytoskeleton and thereby increase the rigidity of the infected RBC. Artesunate, by acting on young ring forms, attenuated the reduction in deformability parasite, prevented their development to more rigid mature trophozoites, and thereby attenuated the reduction in deformability associated with continued parasite growth. Artesunate induces changes either in the parasite or in the RBC directly and led to increased antigenicity and thus increased opsonization. Terminal half-lives of the orally administered drugs are usually less than 2\u2009h. We observed that once daily administration with artemisinin derivatives provides equivalent cure rates to more frequent administration which is consistent with prior studies [P > 0.05), MCHC (P > 0.05), and monocytes (P > 0.05) for artesunate-treated group.Artemisinin derivatives are most effective against ficacies \u201334. Antificacies . Artemis studies , 37. In P > 0.05), MCH (P > 0.05), MCHC (P > 0.05), and monocytes (P > 0.05) for arteether-treated group. No adverse effects were observed on hematological parameters when animals-treated with \u03b1, \u03b2 arteether which are consistent with prior studies [Alpha beta arteether is an ethyl derivative of artemisinin which is an efficient schizonticidal drug in mild malaria. The clinical efficacy of arteether is characterized by an almost immediate onset and rapid reduction in parasitaemia, with complete clearance in most cases within 48 hours. But in our study we observed delayed parasite clearance time in AE-treated groups when compared to previous studies. In arteether group parasites cleared on day 5. We observed positive blood smears for one additional day after the completion of 3-day drug course. The peak level of parasitaemia (4.2% on day 2) was increased when compared to AS-treated group (4 on day 2). Insignificant changes were observed in MCV  0 and monocytes (P > 0.05) for artemether + lumefantrine-treated group.AE showed lower efficacy than AL and AS. The levels of efficacy are similar to the findings of the recent studies on ACTs in Africa which revealed that AL has higher efficacy rates when compared to other antimalarials \u201343. In ofalciparum positive blood smear in all treatment groups on day 3 (72\u2009h) was a good predictor for treatment failure and considered as a simple screening measure for artemisinin resistance. Considering the higher efficacy rates of artemether +lumefantrine (AL) as compared with artesunate (AS) and arteether (AE), we conclude that AL is clinically more effective than AS and AE. No adverse effects were observed on haematological parameters when animals were treated with artemisinin derivatives. Artemisinin resistance is characterized by prolongation in clearance times which we observed in the present study. The result of"}
+{"text": "Plasmodium falciparum and Plasmodium vivax malaria.Pyronaridine-artesunate (PA) is indicated for the treatment of acute uncomplicated P. falciparum (five studies) or P. vivax (one study) malaria. Efficacy against P. falciparum was evaluated across three Phase III clinical trials.Individual patient data on safety outcomes were integrated from six randomized clinical trials conducted in Africa and Asia in patients with microscopically confirmed versus 51.5%  for comparators, most commonly : headache , cough  and anaemia . Serious averse events were uncommon for all treatments (0\u20130.7%). Transient increases in alanine aminotransferase and aspartate aminotransferase were observed with PA but did not lead to any clinical sequelae. For P. falciparum malaria, day-28 PCR-corrected adequate clinical and parasitological response with PA was 93.6%  in the intent-to-treat population and 98.5%  in the per-protocol population. Median parasite clearance time was 24.1\u00a0h with PA, 31.9\u00a0h with MQ\u2009+\u2009AS, and 24.0\u00a0h with AL. Median fever clearance time was 15.5\u00a0h with PA, 15.8\u00a0h with MQ\u2009+\u2009AS, and 14.0\u00a0h with AL. By day 42, P. falciparum gametocytes had declined to near zero for all treatments.The safety population included 2,815 patients randomized to PA, 1,254 to comparators: mefloquine\u2009+\u2009artesunate (MQ\u2009+\u2009AS), artemether-lumefantrine (AL), or chloroquine. All treatments were generally well tolerated. Adverse events occurred in 57.2%  of patients with PA Pyronaridine-artesunate was well tolerated with no safety concerns with the exception of mostly mild transient rises in transaminases. Efficacy was high and met the requirements for use as first-line therapy. Pyronaridine-artesunate should be considered for inclusion in malaria treatment programmes.Clinicaltrials.gov: NCT00331136; NCT00403260; NCT00422084; NCT00440999; NCT00541385; NCT01594931 Plasmodium falciparum infection in children under five years of age,55. In p Figure\u00a0. There aChildren are the most important target group for anti-malarial therapy, having reduced immunity and consequently poorer outcomes. Despite this, until recently paediatric ACT formulations have been generally lacking. PaediatP. falciparum clinical trials. However, the efficacy of PA in P. vivax has also been evaluated in one Phase III study in adults and children[P. vivax malaria and PA efficacy in areas of chloroquine-resistant P. vivax.For the efficacy analysis, this report is concerned with the integrated analysis of the PA Phase III children. In summchildren. AdditioP. falciparum was consistently high regardless of geographical region, patient age, gender, or degree of parasitaemia. Against P. vivax, PA had efficacy at least as good as CQ but with more rapid parasite and fever clearance and a lower parasite re-emergence rate. Importantly, PA has been developed to have a specific paediatric granule formulation available, shown to have equivalent pharmacokinetics and efficacy to the tablet formulation[Overall, PA was well tolerated with a similar adverse event profile to comparators. Although PA was associated with transient increases in transaminases in a relatively small proportion of patients, there was no indication of a risk of progressive liver injury. Pyronaridine-artesunate efficacy against mulation. InitialSD, and IB-F are employees of MMV; JCC is a former employee of MMV; SA-B and RMM are contractors employed by MMV; C-SS is a former employee of Shin Poong Pharmaceutical Co. Ltd.; LF is an advisor to MMV.All authors were involved in study design and conduct, contributed to the paper and approved the final version for submission."}
+{"text": "Recent reports indicate that first cases of genuine artemisinin resistance have already emerged along the Thai-Cambodian border. The main objective of this trial was to track the potential emergence of artemisinin resistance in Bangladesh, which in terms of drug resistance forms a gateway to the Indian subcontinent.We conducted an open-label, randomized, controlled 42-day clinical trial in Southeastern Bangladesh to investigate the potential spread of clinical artemisinin resistance from Southeast Asia. A total of 126 uncomplicated falciparum malaria patients were randomized to one of 3 treatment arms (artesunate monotherapy with 2 or 4 mg/kg/day once daily or quinine plus doxycycline TID for 7 days). Only cases fulfilling a stringent set of criteria were considered as being artemisinin-resistant.ex vivo results indicate significantly higher susceptibility  to artemisinins as compared to SE-Asia.The 28-day and 42-day cure rates in the artesunate monotherapy (2 and 4 mg/kg) and quinine/doxycyline arms were 97.8% , 100% (95% CI: 91.1\u2013100%), and 100% (95% CI: 83.4\u2013100%), respectively. One case of re-infection was seen in the artesunate high dose arm, and a single case of recrudescence was observed in the low dose group on day 26. No differences in median parasite and fever clearance times were found between the 2 artesunate arms (29.8 h and 17.9 h vs. 29.5 h and 19.1 h). Not a single case fulfilled our criteria of artemisinin resistance. Parasite clearance times were considerably shorter and There is currently no indication that artemisinin resistance has reached Bangladesh. However, the fact that resistance has recently been reported from nearby Myanmar indicates an urgent need for close monitoring of artemisinin resistance in the region.NCT00639873.ClinicalTrials.gov  P. falciparum to artemisinin derivates with individual cases of genuine resistance to artesunate have been reported from the Thai-Cambodian and more recently the Thai-Myanmar border suggesting a westward spread of resistance st century and potentially endanger all recently initiated malaria elimination efforts. Historical evidence from traditional antimalarials suggests that drug resistance has a tendency to spread westwards from its origins in Southeast Asia Virtually all malaria-endemic countries have adjusted their treatment guidelines to spreading resistance to practically all traditional antimalarials and now recommend artemisinin-based combination therapies (ACTs) as first-line therapy for the treatment of uncomplicated falciparum malaria This study was part of the Artemisinin Resistance Confirmation, Characterization and Containment Project (ARC3), an effort coordinated by the World Health Organization (WHO) and funded by the Bill & Melinda Gates Foundation, to define the problem, extent, and spread of artemisinin resistance in South and Southeast Asia. The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Checklist S1 and Protocol S1. Registration information: Clinicaltrials.gov Identifier NCT00639873.Written informed consent was obtained from all study participants or their legal representatives, and the study was approved by the ethical review boards of the Medical University of Vienna, the Research Ethics Review Committee of the World Health Organization and the Ethical Review Committee of the International Centre for Diarrhoeal Disease Research, Bangladesh .Plasmodium falciparum mono-infections with a parasite density of 1,000\u2013100,000 asexual parasites/\u03bcL as determinate by microscopy on a screening slide. All female patients between the age of 12 and 65 were required to have a negative human chorionic gonadotropin (hCG) urine pregnancy test . All females of childbearing potential were required to use an acceptable contraception throughout the duration of their participation in the study. Patients who presented with one of the following criteria were excluded from study participation: sings or symptoms of severe malaria (as defined by WHO criteria) This study was conducted at the MARIB  field site in Bandarban in Southeastern Bangladesh. Study participants were male or nonpregnant female patients aged 8 to 65 years, who presented with acute symptomatic The study was a single-center, open label, randomized, controlled clinical trial specifically designed to detect the potential emergence of artemisinin resistance. Patients were hospitalized for 7 days (the duration of study drug administration), or until complete fever and parasite clearance, whichever took longer. Participants were asked to return for follow-up visits on days 14, 21, 28, 35, and 42 and whenever signs and symptoms consistent with malaria reappeared. Insecticide-treated mosquito nets were provided to all patients to limit the chances of reinfection after discharge from the hospital. Patients who failed initial therapy in the artesunate groups were treated with 7 days of oral quinine , patients in group 3 who failed initial therapy received 3 days Coartem according to the national treatment guidelines.Enrollment target was 100 evaluable subjects (i.e. study participants reaching primary study endpoint). Subjects were enrolled in the 3 groups at a ratio of 2\u22362\u22361. Any dropouts  were replaced by continuing randomization until a total number of 100 evaluable subjects were reached.Subjects were allocated unique identification codes and were randomly assigned to one of the three treatment groups using block randomization in blocks of 10 by reference to a statistical series based on random sampling numbers drawn up for each patient by the study staff; the details of the series were unknown to any of the investigators or to the coordinator and were contained in a set of sealed envelopes.Artesunate  was provided by WHO, Geneva, Switzerland, quinine sulphate BP  was purchased from Jayson Pharmaceuticals Ldt., Dhaka, Bangladesh and doxycyline hyclate  was procured from local suppliers.Patients in all 3 treatment groups received oral antimalarial therapy for 7 days. Artesunate monotherapy was administrated over 7 days either as 2 mg/kg body weight once daily in the low dose arm or as 4 mg/kg body weight once daily in the higher dose arm. Patients in the control group received oral quinine (10 mg/kg body weight 3 times daily) plus doxycyline (2 mg/kg twice daily) over 7 days. A study physician observed study drug administration and the full dose was repeated if vomiting occurred within one hour after drug administration. Concomitant medication was provided for the symptomatic treatment of fever, headache, myalgia, nausea and/or vomiting.\u00ae blood collection system  on day 0 and 3 or whenever clinically warranted. For DNA fingerprinting, RNA, and in vitro drug sensitivity assays blood samples were collected on day 0 and in case of recurrence of parasitemia. Plasma samples for determining drug levels were collected from all patients on days 0 and 6.After enrollment a full medical history, the results of the physical examination, vital signs, adverse events, clinical signs and symptoms as well as medication history were recorded daily until day 7 and whenever patients returned for follow-up. A urine pregnancy test was performed on admission day for all females between the age of 12 and 50. Venous blood samples for complete blood count and blood chemistry were collected using the VacuettePeripheral malaria blood smears were performed from finger prick blood at admission and after this twice daily until two consecutive slides were confirmed to be negative as well as at every follow-up visit. Giemsa-stained thick and thin blood smears were examined by a microscopist who was blinded to treatment, clinical status of the study subject and to any other results. All slides that were positive on any follow-up day were reexamined by a second microscopist. In case of a difference in the result (positive/negative or species) between the two microscopists, the blood smear was re-examined by a third senior microscopist and this reading was accepted as final result. Parasite density was determined based on the count of parasites per 200 white blood cells (thick film) or 2000 red blood cells (thin film). A total of 200 immersions oil field were screened before a blood film was considered as negative as well as to rule out mixed infections.Outcome measures for the clinical study were based on the criteria set forth by the WHO Blood samples for DNA fingerprinting were collected before drug intake and in case of reemergence of parasites to distinguish between reinfection and recrudescence. Gene loci  of these samples were compared by polymerase chain reaction (PCR) P. falciparum clone was used as a reference and for quality control of drug-coated culture plates.All fresh parasite samples obtained on day 0 and on the day of recrudescence were tested in a histidine-rich protein 2 (HRP2) drug susceptibility assay for their sensitivity to dihydroartemisinin (DHA), artesunate, and artemisinin. The cultures and enzyme-linked immunosorbent assays (ELISA) were carried out as previously described Venous blood samples were collected from all patients in arm 1 and 2 into heparinized tubes just before and 60, 120, and 180 (\u00b115 minutes) after artesunate intake on day 0 and day 6. The samples were centrifuged immediately and plasma stored at \u221220\u00b0C before transferring to \u221280\u00b0C within a maximum of 3 weeks. DHA levels were determined by LC-MS as described elsewhere in vitro drug sensitivity tests on clinical isolates suggesting elevated inhibitory concentrations for artemisinins and/or genetic makers indicating reduced drug susceptibility The definition of artemisinin resistance has been a controversial issue ever since the discovery of the first cases of resistance. We therefore used a conservative approach in defining resistance. Only cases fulfilling all of the following criteria were defined as being artemisinin-resistant: recrudescence during follow up or failing to clear parasites after 7 days of directly observed monotherapy with 2 or 4 mg/kg/day of artesunate in the absence of any signs of vomiting after drug administration or malabsorption and with an enrollment parasite density of <100,000 asexual parasites/\u00b5L; prolonged PCT indicating poor clinical response; exclusion of re-infection by PCR; pharmacokinetic parameters confirming adequate (greater than mean in cures -1SD) drug levels for DHA; in vitro drug inhibitory concentrations. Continuous data were compared by t-test/ANOVA. Data not showing normal distribution were compared by Mann-Whitney U/Kruskal-Wallis ANOVA. Proportions were compared using Fisher's Exact/Chi2 test.Cure rates were estimated using Kaplan-Meier survival analysis to calculate the proportion of aparasitemic patients at each point in time. Nonlinear regression analysis based on a polynomial regression model was used to calculate individual P. vivax infections between day 21 and 42 . All dropouts, except for the one who was withdrawn, occurred after completing the full treatment course and after clearing parasites and fever. Forty-two subjects were evaluable for primary study end point in Arm 1, 44 in Arm 2, and 20 in the control group. All three arms were well matched in regard to sex, age, weight, height, and parasite density on enrollment , 100% (95% CI: 91.1\u2013100%) for Arm 2, and 100% (95% CI: 83.4\u2013100%) for Arm 3 . Of the The median parasite clearance times (PCTs) were similar in the artesunate lower and higher dose arms with 29.8  h and 29.5 (IQR: 19.8\u201332.4) h, respectively. With 43.1 (IQR: 33.6\u201354.8) h the PCT in the control group was significantly (P<0.001) slower. At the first measurement more than 24 hours after initiation of treatment 13.7% (95% CI: 6.15\u201326.9) of the patients in Arm 1, 16.0% (95% CI: 7.6\u201329.7) in Arm 2 and 70.8% (95% CI: 48.8\u201386.6) in the control group were still parasitemic. All patients in the artesunate arms had cleared their parasitemia after 48 hours, whereas 16,7% (95% CI: 5.5\u201338.2) in the control group were still positive. After 72 hours all patients tested negative for asexual malaria parasites. The median 50% parasite reduction times were 9.3 (IQR: 6.5\u201318.4), 8.5 (IQR: 7.1\u201317.5), and 29.5 (IQR: 7.1\u201331.0) h, for Arms 1\u20133, respectively. The corresponding mean 90% parasite reduction times were 17.6 (IQR: 8.9\u201319.7), 17.4 (IQR: 8.3\u201319.5), and 30.0 (IQR: 18.2\u201331.6) h.The median parasite reduction ratios at 12, 24, 36, and 48 h  were 0.93 (IQR: 0\u20132.98), and 0.0 (IQR: 0.0\u20130.0) in Arm1; 0.78 (IQR: 0.0\u20133.08), and 0.0 (IQR: 0\u20130\u20130.0) in Arm 2; 70.37 (IQR: 5.71\u2013187.84), 0.8 (IQR: 0.0\u20137.14), 0.0 (IQR: 0.0\u20130.57) and 0.0 (0.0\u20130.0) in Arm 3. The ratios in the artesunate groups were lower than in the quinine group after 12 and 24 hours (P<0.001). There was no difference in the parasite reduction ratio between the artesunate subgroups.With a median FCT of 17.9 (IQR: 4.6\u201320.7), 19.1 (IQR: 6.6\u201323.6), and 29.3 (IQR: 16.0\u201343.1) h, for Arms 1\u20133, respectively, the artesunate arms showed a significantly (P\u200a=\u200a0.001) faster clinical response than the control group.in vitro drug susceptibility to DHA, artemisinin and artesunate. The geometric mean 50% inhibitory concentration (IC50) for DHA was 1.10 nM , 1.37 nM  for artesunate and 3.74  nM for artemisinin, respectively. No correlation was observed between PCT and IC50s of DHA, artemisinin or artesunate in the artesunate arms. The IC50 of DHA, artesunate and artemisinin for the reference clone 3D7 was 0.78 nM (95% CI: 0.71\u20130.85), 1.6 nM (95% CI: 1.32\u20131.94) and 5.27 nM (95% CI: 4.57\u20136.08), respectively. The patient classified as recrudescence had an IC50 of 1.28, 1.74 and 5.53 nM for DHA, artesunate and artemisinin, respectively.Out of a total of 106 parasite samples obtained on the day of study admission 83  were successfully tested for their Plasma DHA levels measured after drug intake of the first dose and last dose were used to determine whether there had been adequate drug exposure in patients treated with artesunate. The total mean drug exposure estimated by the area under curve (\u00b1SD) of DHA in the low dose group after the first and last dose was 1,365\u00b1665 ng/mLxh and 592\u00b1228 ng/mLxh, and in the high dose group 2,936\u00b11,576 ng/mLxh and 1,463\u00b1736 ng/mLxh, respectively. Adequate DHA concentrations were defined as the mean \u00b11 SD. The area under the curve estimated for the recrudescent patient in the low dose group was considered adequate at 1,384 ng/mLxh after the first and 1,077 ng/mLxh last dose, respectively.No serious adverse events were observed. All treatment regimes were generally well tolerated. Out of 78 recorded adverse events, 48  were classified as drug-related. No difference in the proportion of patients with/without adverse events was observed between the three groups (P\u200a=\u200a0.91). The most common adverse event in all 3 arms was gastrointestinal disorder.in vitro artemisinin resistance First indications of high failure rates associated with the extensive use of ACTs in Thailand were reported in a paper published in 2006 suggesting the possibility of poor clinical response to ACTs along the Thai-Cambodian border and coincide with the first reports of the experimental induction of in vivo-ex vivo approach was conducted in Southeastern Bangladesh in close proximity to the borders of India and Myanmar and was specifically designed to answer the question of whether clinical artemisinin resistance has already spread to the region. As compared to Southeast Asia the major difference in terms of exposure of malaria parasites to artemisinins is the fact that in Bangladesh ACTs have only been introduced very recently and that the local parasite populations have therefore not been exposed to artemisinins on any significant scale. The parasite phenotype seen in Bangladesh is therefore likely to be representative of Asian P. falciparum populations before the introduction of artemisinins. Historical data suggest that in terms of the spread of resistance from Southeast Asia the malaria-endemic regions of Bangladesh may represent an important gateway to the Indian Subcontinent.This study using a systematic de novo emerging artemisinin resistance. Although a single case of recrudescence was observed, this case did not fulfill our stringent criteria for artemisinin resistance ex vivo data confirm earlier published data suggesting a continuous and significant decrease in ex vivo artemisinin susceptibility from Bangladesh throughout western to eastern Thailand and Cambodia 50 for artemisinins as compared to our earlier studies in the same region suggesting the absence of a temporal trend towards reduced sensitivity Our data clearly indicate that in this region there is currently no evidence for spreading or The discussion of the role and importance of poor clinical response and failures with 7 days of high-dose artemisinins along the Thai-Cambodian border has largely been focusing on the question of whether innately resistant clones occur naturally as a fixed subset of parasite populations from different locales. Our data from an area in Asia where artemisinins have as yet not been deployed on any significant scale would suggest that this is not the case.ex vivo drug sensitivity therefore play a crucial role in detecting early stages of compromised drug sensitivity ex vivo drug sensitivity between Bangladesh and Cambodia seems to closely match the major differences in treatment response In its early stages reduced drug susceptibility is not necessarily reflected in an increased proportion of treatment failures. Clinical response parameters (particularly parasite clearance) and intrinsic The obvious advantage of artemisinins is their capacity to quickly reduce the initial parasite burden making them highly successful even in combination treatment courses as short as 3 days . However, with parasite clearance times reaching 100 hours or more in selected patients in Cambodia Both, the prolongation in parasite clearance and persisting parasitemia 72 hours after initiation of treatment are strong indicators for reduced susceptibility or artemisinin resistance In summary, there is currently no indication of compromised drug sensitivity to artemisinin derivatives in the region. The rapid reduction in parasite densities as well as the high efficacy seen with 7 days artesunate monotherapy in Bangladesh suggests that the phenomenon seen in Cambodia and along the eastern borders of Myanmar is likely to be based on the selection of parasite populations with reduced sensitivity under drug pressure rather than due to innately resistant clones occurring naturally as a fixed subset of parasite populations. So far artemisinins have not been used on any significant scale in Bangladesh but the situation is likely to change in the future, either due to resistance spreading from Southeast Asia or due to de novo emergence of resistance under drug pressure. These data therefore also provide an important baseline for future assessments of temporal and geospatial trends in artemisinin resistance in the region.Checklist S1CONSORT Checklist.(DOC)Click here for additional data file.Protocol S1Trial Protocol.(PDF)Click here for additional data file."}
+{"text": "Sulphadoxine-pyrimethamine, in combination with artesunate or amodiaquine, is recommended for the treatment of uncomplicated malaria and is being evaluated for intermittent preventive treatment. Yet, limited data is available on pharmacokinetic interactions between these drugs.falciparum malaria, received either one dose of sulphadoxine-pyrimethamine alone (SP), one dose of SP plus three daily doses of amodiaquine (SP+AQ) or one dose of SP plus 3 daily doses of artesunate (SP+AS). Exactly 100 \u03bcl of capillary blood was collected onto filter paper before drug administration at day 0 and at days 1, 3, 7, 14, 21 and 28 after drug administration for analysis of sulphadoxine and pyrimethamine pharmacokinetic parameters.In a randomized controlled trial, children aged 6-59 months with uncomplicated p = 0.001) and thus elimination half-life . This study confirmed the lower SP concentrations previously reported for young children when compared with adult malaria patients.Fourty, 38 and 31 patients in the SP, SP+AQ and SP+AS arms, respectively were included in this study. The concentrations on day 7 (that are associated with therapeutic efficacy) were similar between the SP, SP+AQ and SP+AS treatment arms for sulphadoxine  and for pyrimethamine . There were statistically significant differences between the pyrimethamine volumes of distribution (4.65 [3.93-6.40], 4.00 [3.03-5.43] and 5.60 [4.40-7.20] L/kg; falciparum malaria treatment in young Malian children.Despite slight differences in pyrimethamine volumes of distribution and elimination half-life, these data show similar exposure to SP over the critical initial seven days of treatment and support the current use of SP in combination with either AQ or AS for uncomplicated  Plasmodium falciparum malaria [Malaria remains a major public health problem, particularly in sub-Saharan Africa, where it claims nearly 750,000 lives of children under the age of five years . To impr malaria . However malaria -5. SP mo malaria -11.dhfr and dhps mutation levels, particularly dhps540 which best predicts the in vivo SP resistance [Resistance to SP is wide spread in Africa, with high level of resistance in east Africa compared to west Africa. That is confirmed by the level of sistance .Despite some level of resistance to SP and AQ, the combination of the two drugs is effective on falciparum malaria in much of West Africa.And, despite these widespread use of SP, the pharmacokinetic parameters of SP when used in combination with AS or AQ is poorly documented, particularly in young children. Altered pharmacokinetic parameters contribute to the increased risk of young children failing SP treatment. After adjusting for dosage, median sulphadoxine and pyrimethamine areas under the concentration time curves in children aged 2-5 years old are approximately half those in adults . This stPlasmodium falciparum malaria is both endemic and seasonal with parasitaemia prevalence rates ranging from 40-50% in the dry season (October-May) and 70-85% in the rainy season (June-September)[P. falciparum mono-specific parasitaemia between 2,000 and 200,000 parasites/ul (parasite density was measured by microscopy after staining thick smear with Giemsa). Exclusion criteria included 1) features of severe malaria , 2) danger signs such as prostration, 3) a history of allergy or other severe adverse reaction to the study drugs and 4) more than two episodes of vomiting per day. Children with any of the above exclusion characteristics or who were considered too ill for inclusion in the study, or who declined to participate in the study, received standard and appropriate treatment with chloroquine as then recommended by the National Malaria Control Programme.This was an open label clinical trial using the WHO 2003 protocol  conducteeptember),16. InclEnrolled children were randomized into three treatment arms  by computer-generated randomization. Directly observed treatment was administered based on weight to the nearest half tablet that would provide the following minimum dose(s): 25 mg/kg of sulphadoxine and 1.25 mg/kg of pyrimethamine as a single dose on day 0; with or without AQ 10 mg/kg/day over three days or artesunate 4 mg/kg/day over three days. All subjects were observed for 60 minutes to monitor for adverse reactions and to make sure that the medicine was not vomited. If vomiting occurred within 30 minutes, the full dose was re-administered. If vomiting occurred between 30 and 60 minutes a half-dose was re-administered. All acute concomitant illnesses were treated for free by the study team, with concomitant medication which had no known SP drug interactions. Parental written informed consent was obtained before any protocol specific procedure was performed. The study was conducted in accordance with the Helsinki Declaration of 1975 (as revised in 1983). The protocol was approved by the ethical committee of the Faculty of Medicine, Pharmacy and Odonto-Stomatology, Bamako, Mali.\u00ae tablet for AQ, Fansidar\u00ae tablet for SP and Artesunate tablet for AS)were bought at a private pharmacy in Bamako, Mali. The quality of these drugs were authenticated according to the United States Pharmacopoeia and National Formula (USP-NF) 2004 methods using dissolution testing apparatus in combination with High Performance Liquid Chromatography (17). More than 75% of the amount of the three drugs  of the study were dissolved in the time indicated for each drug.Study drugs , air dried at room temperature and stored in plastic folders with solid desiccant until assayed in March 2008. Concentrations of sulphadoxine and pyrimethamine were determined by a validated method using liquid chromatography-mass spectrometry/mass spectrometry, with the lower limits of quantification set at 10 \u03bcg/mL for sulphadoxine and 10 ng/mL for pyrimethamine. The upper limits of quantification were set at 200 \u03bcg/mL for sulphadoxine and 1 \u03bcg/mL for pyrimethamine. The coefficient of variation for sulphadoxine was 10.3% at 65 \u03bcg/mL, and pyrimethamine had a coefficient of variation of 13.8% at165 ng/mL as previously described [The area under the capillary blood concentration-time curve to infinity (AUC), terminal elimination half-life (t1/2), apparent volume of distribution (VD) and apparent clearance (Cl) of pyrimethamine and sulphadoxine were determined using a non compartmental model in WinNonLin Professional 3.3 (Pharsight). As there was no intravenous comparator arm in this study, equivalent bioavailability was assumed for comparisons of apparent volumes of distribution and clearance, which were expressed as Vd/f and Cl/f, where f is the fraction of drug absorbed (unknown).As most pharmacokinetic parameters were not normally distributed, these were summarized using medians and inter-quartile ranges and compared between the three treatment arms using the Kruskal-Wallis equality-of-populations rank test. Categorical data were compared using the Chi-squared test.msp2, the ACPR rates were above 97% in all three groups.Between 2004 and 2006, we conducted a clinical trial comparing the therapeutic efficacy of SP, SP+AQ and SP+AS. A total of 455 children were included and 441 (97%) were successfully followed for 28 days. The baseline characteristics in the three treatment arms were comparable for mean age, mean weight and gender (P > 0.05). Without PCR correction the 28-day uncorrected adequate clinical and parasitological response (ACPR) rates were above 95% for each of the treatment arms. Early treatment failure (ETF) was only found in the SP-monotherapy arm in three patients . After PCR correction by Among the study population children with complete pharmacokinetic samples from day 0 to day 28 were included in this pharmacokinetic analysis i.e. 41 children in SP arm, 40 in SP+AQ and 33 in SP+AS. The ages of the children enrolled ranged between 10 and 60 months. Treatment groups were well matched for the baseline characteristics of age, gender, weight, SP mg/kg dosage, temperature and hemoglobin . The two children retreated with the full dose for vomiting within 30 minutes had SP concentrations on day 1 that were almost double the median (133 and 137 \u03bcg/ml for sulphadoxine and 513 and 661 ng/ml for pyrimethamine). There were no serious adverse events during this study.P. falciparum malaria in the context of combination therapies, and confirmed the lower SP concentrations previously reported for this age group when compared with adult malaria patients [This study reports on SP pharmacokinetic parameters in young African children with uncomplicated patients . There wpatients  and is rAlthough adding amodiaquine to SP had no effect on sulphadoxine pharmacokinetic parameters, a significant decrease in the apparent volume of distribution and elimination half-life of pyrimethamine in the SP+AQ arm was observed. However, this did not result in a significant change in pyrimethamine concentrations in the blood in SP+AQ arm over the critical initial seven days of treatment nor the AUC. The absence of a substantial PK interaction in our study population supports the effectiveness of SP+AQ in IPTc ,10 and iDay-7 concentrations of sulphadoxine and pyrimethamine are a good surrogate measure of their respective total whole blood AUCs  and Day The vast majority of our patients achieved maximum concentrations of both sulphadoxine and pyrimethamine on day 1. However, the exact Cmax and Tmax could not be determined precisely because of limited sampling frequency. The concentration of artesunate could not be quantified because of the sampling methodology used, and the concentrations and pharmacokinetic parameters of amodiaquine (when administered with SP) will be reported separately.The two children who were retreated with the full dose after vomiting within 30 minutes of SP administration had approximately double the median SP concentrations on day 1, suggesting that further data is needed to inform optimal dosing in patients with early vomiting.dhfr/dhps mutations accumulate the SP concentrations achieved may be insufficient to ensure an adequate clinical and parasitological response rate as recently described in Malawi [In this study conducted in Mali in 2004-2006, SP or its combination with AS or AQ remained highly effective with more than a 95% adequate clinical and parasitological response rate. Although this level of efficacy of SP when administered alone or in combination with either AS or AQ is quite high in comparison with data from Eastern or Southern Africa , these rn Malawi .dhfr and dhps mutations that typically occurs in areas with large-scale deployment of SP.This pharmacokinetic data supports the current use of SP in combination with AS to treat young Malian children with uncomplicated falciparum malaria, and the use of SP + AQ for IPTc. Ongoing monitoring is essential, particularly given the lower pyrimethamine concentrations and the accumulation of The authors declare that they have no competing interests.MMT contributed to the study design, field studies, pharmacokinetic analyses, data analysis and drafted the manuscript. AHB, CPOS, HM and ZIT, conducted the field studies, collected the samples, and assisted with manuscript preparation. PS, AF and AE developed and or conducted pharmacokinetic assay. KIB contributed to data analysis and manuscript writing. OKD contributed to study design and manuscript writing. AD contributed to the design of the study, oversaw the field and laboratory studies, and to the writing and final approval of the manuscript. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum malaria worldwide but decreased artemisinin susceptibility, phenotypically characterized as slow parasite clearance time (PCT), has now been reported in Southeast Asia. This makes it all too important to measure the dynamics of parasite clearance in African patients treated with ACT over time, to understand trends and detect changes early enough to interveneArtemisinin-based combination therapy (ACT) is the recommended first-line therapy for uncomplicated vs comparators conducted between 1999 and 2009 were analysed for parasite clearance on modified intent-to-treat (ITT) basis.Individual patient data from 27 clinical trials of artesunate-amodiaquine (ASAQ) P. falciparum malaria at 44 sites in 20 sub-Saharan African countries were included in the analysis; 51%  vs 49%  were treated with ASAQ and comparator treatments, respectively. Seventy-seven per cent (77%) were children under six years of age. The proportion of the patients treated with ASAQ with persistent parasitaemia on Day 2 was 8.6%, and 1.5% on Day 3. Risk factor for not clearing parasites on Day 2 and Day 3 calculated by multivariate logistic regression with random effect on site and controlling for treatment were: high parasitaemia before treatment was (adjusted risk ratios (AOR) 2.12, 95% CI 1.91-2.35, AOR 2.43, 95% CI 1.98-3.00, respectively); non-ACT treatment . Anaemia (p=0.001) was an additional factor for Day 2 and young age (p=0.005) for Day 3.Overall 15,017 patients treated for uncomplicated In patients treated with ASAQ in studies who had complete parasitaemia data every 24 hours up to Day 3 and additionally Day 7, the parasite reduction ratio was 93.9% by Day 1 and 99.9% by Day 2. Using the median parasitaemia before treatment  and a fitted model, the predicted PCT (pPCT = 3.614*ln (p0) \u2013 6.135, r\u00b2 = 0.94) in ASAQ recipients was 31 hours.Plasmodium falciparum clearance continues to be achieved in Sub-Saharan African patients treated with ACT, and in particular with ASAQ. The prediction formula for parasite clearance time could be a pragmatic tool for studies with binary outcomes and once-daily sampling, both for research and monitoring purposes.Within the period covered by these studies, rapid  Plasmodium falciparum response to artemisinin is reported in Southeast Asia, and may spread to other, more intensely malaria-endemic areas. Artemisinin compounds contribute rapid parasite killing and faster clearance to artemisinin-containing combination therapy (ACT). Artemisinin tolerance/resistance manifests itself with slower parasite clearance, while ACT remain generally effective both clinically and parasitologically [in vitro assay (Ring Stage Assay) has also focused on the very early phases of the ring-stage parasite [in vivo resistance [Delayed ogically -4. Genomogically , and a Sogically . An in vparasite . Howeversistance ; the prosistance  is practsistance , requiriApplying these and other measures both retrospectively and prospectively to commonly-used ACT will help understand trends and detect changes early enough to prompt effective responses to contain the spread of resistance. Artesunate-amodiaquine (ASAQ) is the second most commonly used ACT in the world (the first-line treatment in 22 out of 44 sub-Saharan African countries); more than 200 million treatments of ASAQ Winthrop have been distributed in Africa since the medication became available in 2007 -13. WhilGreater understanding of factors influencing parasite clearance is crucial, and requires the analysis of pooled data from individual patient records. Therefore, this study collected and analysed a large sample  of individual patient data (of whom approximately three-quarters were children under six years of age) from clinical trials conducted in sub-Saharan Africa between 1999 and 2009.Studies such as this will help provide the foundations for further analyses of parasite clearance trends across sub-Saharan malaria-endemic countries.The database was constructed from studies identified through a systematic review of clinical trials and personal contacts. To be included, efficacy or tolerability monitoring studies had to have been conducted in sub-Saharan Africa including any formulation of ASAQ to any other single or combination treatment of uncomplicated falciparum malaria (non-severe or non hyperparasitaemic) with follow-up of at least 28\u00a0days. For the studies meeting these criteria, investigators were contacted to provide individual patient data and the datasets received were examined for inclusion .Out of the 27 studies included (conducted between 1999 and 2009), three were multi-country studies -19, two Table\u00a0The curve of the proportions of patients remaining parasitaemic during follow-up was fitted using logistic regression to give an estimate for clearance time. The fitted model for the logit of the proportion of parasitaemic patients over time and expressed in hours from the logistic regression:In order to simplify the calculations of the predicted parasite clearance time ((pPCT), for a patient or a group of patients using the median time parasite clearance) the patient population was classed into ten pre-treatment parasite densities categories. A logistic model was fitted for each group and the corresponding predicted PCT was calculated, and once the results of predicted PCT for the ten groups were obtained, a simple logarithmic model was fitted with the aggregated results.The risks, presented as adjusted risks ratios (AOR), were assessed by logistic multivariate analysis with random effect on the site in an attempt to account for potential statistical heterogeneity while controlling for age (continuous), parasitaemia before treatment (log-transformed), anaemia (binary), and year (continuous). Categorical data were compared using the Chi-square or the Fisher exact test as appropriate. The Spearman test was used to analyse the relationship between clearance reduction and pre-treatment parasitaemia. Confidence intervals (CI) were calculated at 95% and statistical significance was set at p-value <0.05.All studies had been approved by the relevant ethics and institution review committees as reported in the individual papers.The majority of the patients were treated with individually formulated AS and AQ. The target dose was AS 12\u00a0mg/kg over three days and AQ 30\u00a0mg/kg over three days except in Uganda where AQ was given at 25\u00a0mg/kg . The loose combinations of ASAQ were dosed based on body weight, while in four studies the fixed dose combination (FDC) ASAQ was based on age and weight range -22. The ASAQ FDC (Coarsucam\u2122 Winthrop\u00ae Sanofi Aventis); AS 25\u00a0mg/AQ 67.5\u00a0mg one tab/day for three days in children 5\u20138.9\u00a0kg; AS 50\u00a0mg/AQ 135\u00a0mg one tab/day for three days in children 9\u201317.9\u00a0kg; AS 100\u00a0mg/AQ 270\u00a0mg one tab/day for three days in children 18\u201335.9\u00a0kg.i. ACT: artemether-lumefantrine (AL)(20\u00a0mg artemether/120\u00a0mg lumefantrine) given according to weight as one (5\u201314\u00a0kg), two (15\u201324\u00a0kg), three (25\u201334\u00a0kg), and four (\u226535\u00a0kg) tablets given twice daily co-administrated with fat for three days; Coartem\u2122, Novartis); dihydroartemisinin-piperaquine (DP) was given once daily over three days, at the standard dosage of 2.25\u00a0mg/kg and 18\u00a0mg/kg of dihydroartemisinin-piperaquine, respectively, rounded up to the nearest half tablet ; AS\u2009+\u2009sulphadoxine- pyrimethamine (SP);ii. Non-ACT: AQ\u2009+\u2009SP (AQ 10\u00a0mg/kg/day for three days and SP 25\u00a0mg/kg of sulphadoxine and 1.25\u00a0mg/kg of pyrimethamine administered in a co-formulated tablet (SP) as a single dose); chloroquine (CQ)(25\u00a0mg/kg over three days) and SP; AQ monotherapy (10\u00a0mg/kg/day for three days); AS monotherapy (AS 12\u00a0mg/kg over five days).P. falciparum malaria at 44 sites in 20 sub-Saharan African countries were included in the analysis  on comparator treatments; 31%  of the patients had complete parasitaemia data every 24\u00a0hours up to Day 3 and then Day 7 . Overall, the proportion of children under six years of age was 77%, and the overall geometric mean parasitaemia pre-treatment was 16,964/\u03bcL  under ASAQ treatment in comparative and non-comparative trials was analysed in 44 sub-Saharan African sites over the period 1999\u20132009.The proportion of patients on ASAQ who were still parasitaemic on Day 2 was 8.6%  and ranged from 1.0% in Burkina Faso-Nanoro (2008) and Congo-Kindamba (2004) to 57.1% in DRC-Boende in 2003 , ranging from 0% in many various sites across sub-Saharan Africa to 55.9% in DRC-Boende. Using multivariate logistic analysis with random effects on sites, younger patients  and patients with higher parasitaemia at enrolment  were at higher risk of remaining parasitaemic on Day 3. Compared to ASAQ, the risk of being parasitaemic on Day 3 was higher for patients treated with a non-ACT: AQ\u2009+\u2009SP , AQ , CQ\u2009+\u2009SP , while no significant difference was detected with other ACT   compared to non-ACT , the parasite reduction ratio (parasitaemia before treatment/parasitaemia on day (n)) was 93.9% by Day 1 and 99.9% by Day 2 Table\u00a0. The parThe number of patients included in RCT at 17 sites with complete parasitaemia record (every 24\u00a0hours from Day 0 to Day 3 plus Day 7) was 4,848 of whom 2,355 were treated with ASAQ and 2,493 with a comparator drug.Using logistic regression, the predicted time of parasite clearance was overall\u2009\u2248\u200931\u00a0hours for a median baseline parasitaemia of 27,125/\u03bcL, ranging from\u2009\u2248\u200919\u00a0hours for patients with parasitaemia before treatment <2,500/\u03bcL to\u2009\u2248\u200937\u00a0hours for patients with parasitaemia >100,000/\u03bcL  of 0.88 for both adjustments  of uncomplicated malaria treatments  from 44 sites and 20 sub-Saharan African countries with different levels of endemicity, covering the decade 1999\u20132009.Plasmodium falciparum clearance continues to be achieved in sub-Saharan African patients treated with ACT, and in particular with ASAQ, although direct within-site comparisons are not available; it has therefore benchmarking value as reference for future studies. This study also presents a prediction formula for PCT as a pragmatic tool adapted for studies with binary data (not quantitating parasite densities) and once-daily sampling, such as those that would be done at the typical study site in peripheral settings and in routine surveillance by control programmes. This prediction could apply to groups of patients as well as to the individual patient.This analysis shows that rapid 2\u2009=\u20090.94). There was also a (weaker) correlation between parasitaemia before treatment and the parasite reduction ratio (PRR) at 24\u00a0h.These results confirm the correlation between pre-treatment parasitaemia and PCT , and offMonitoring treatment response and detecting artemisinin resistance as early as possible have become a major issue in malaria control. The rate at which treatment clears parasites within the first few days is at present the most useful practical test for ACT, as early response to treatment relies predominantly on the parasite response to artemisinin, independent of whether parasites are later cleared for good through the combination of the longer-lived companion drug and the host\u2019s immune response.The present analysis focused on initial response to ACT treatment, as a proxy for artemisinin resistance and failure, and aimed to identify variables that were independently associated with persistent parasitaemia on Days 2 and 3 in settings where artemisinin resistance has probably not yet emerged.Within the period covered by these studies, there was no indication of delayed response to ACT. ACT cleared parasites rapidly leaving very few patients with a low parasitaemia on Day 3 (first day after the completion of the three-day ACT regimen). In particular, with ASAQ the parasite reduction ratio was 93.9% by Day 1 and 99.9% by Day 2; only 1.5% of the patients were still positive on Day 3 (against a proposed threshold of 3% indicating delayed response ). These Plasmodium falciparum in the DRC. No subsequent studies are available to confirm or belie this original study. Understanding the dynamics of parasite clearance could help detect early signs of artemisinin resistance and distinguish biologically meaningful changes in early parasite clearance over time from changes that may be due to the role of effect modifiers like higher pre-treatment parasitaemia, young age and anaemia by applying multivariate analysis.Anaemia was an additional risk factor for failing to clear parasites by Day 2; younger patient\u2019s age for Day 3. The only exception was the study conducted in DRC-Boende  which had a high proportion of parasite clearance failures with both ASAQ and AL. These findings are difficult to explain and should be taken with caution, but one cannot exclude that there might also have been a focus of resistant falciparum with artesunate for seven days and where there was no evidence of delayed parasitaemia, the median parasitaemia at enrolment (P0) was 27,070\u00a0\u03bcL and the observed median time of parasite clearance was 32\u00a0hours. Applying the suggested method using the logarithmic correlation found in the present paper (pPCT\u2009=\u20093.614*ln(P0) \u2013 6.135) gave very similar results .Consistently with previous results  and the P. falciparum will likely require a combination of more sophisticated studies and routine monitoring. Between parasite clearance estimators [This simple method was tested to predict a \u201ctypical\u201d PCT in situations when resistance has not emerged. However, this estimation of the predicted time to parasite clearance is not without its limitations. The calculations may lack precision with studies sampling once a day and at variable intervals from treatment intake . It has been suggested that parasite clearance rate can only be accurately estimated if sampling is at least every six hours ; it is ttimators  and Day timators , simple The authors declare that they have no competing interests.JZ and PO designed the analysis, interpreted the data and prepared the manuscript. All authors read and approved the final manuscript.Proportion by site of patients still parasitaemic on Day 1, Day 2, Day 3, and parasite clearance failure, ASAQ groups.Click here for file"}
+{"text": "Malaria prophylaxis is recommended for persons with sickle cell disease (SCD), but the value of this has been questioned. The aim of this study was to find out whether intermittent preventive treatment (IPT) with a fixed-dose combination of mefloquine-artesunate (MQAS) or sulfadoxine-pyrimethamine plus amodiaquine (SPAQ) was more effective than daily proguanil for malaria prevention in subjects with SCD.Patients with SCD were randomized to receive daily treatment with proguanil or IPT with either MQAS or SPAQ once every 2 months at routine clinic visits. Patients were followed up for 14 months.A total of 270 patients with SCD were studied, with 90 in each group. Adherence to the IPT regimens was excellent, but 57% of patients took <75% of their daily doses of proguanil. IPT was well tolerated; the most common side effects were vomiting and abdominal pain. Protective efficacy against malaria, compared with daily proguanil, was 61%  for MQAS and 36% (40%\u201370%) for SPAQ. There were fewer outpatient illness episodes in children who received IPT than those who received proguanil.IPT with MQAS administered to patients with SCD during routine clinic visits was well tolerated and more effective in preventing malaria than daily prophylaxis with proguanil.NCT01319448 and ISRCTN46158146. Plasmodium falciparum infection is thought to be lower in children with sickle cell disease (SCD) than in normal children [P. falciparum parasitemia and incidence of P. falciparum infection in patients with HbSS, compared with children with HbAA. However, in the Kenyan study, mortality in children with SCD hospitalized with malaria was substantially higher than that among children without SCD [P. falciparum infection can be fatal [dhfr) and dihydropteroate synthase (dhps) has been little studied but appears to be high [Although the incidence of children , the conchildren  and Tanzchildren  in whichhout SCD . Malariabe fatal , 5. Malabe fatal , but thebe fatal , and onebe fatal . Studiesbe fatal ; a studybe fatal ; and a sbe fatal . Severalbe fatal \u201314. The  be high .Adherence to a daily or weekly prophylactic regimen is likely to be poor and difficult to sustain over long periods. The national antimalarial policy in Nigeria for protection of patients with SCD against malaria at the time that this trial was conducted was daily proguanil, and proguanil prophylaxis continues to be used widely in Nigeria, although more-recent guidelines (from 2011) state that, because proguanil may not be effective, patients with SCD should rely on long-lasting insecticide-treated bed nets and, if they have malaria, prompt treatment.Chemoprevention with drugs being given under supervision by health workers is increasingly being used to prevent malaria in countries of endemicity. The World Health Organization now recommends intermittent preventive treatment (IPT) with SP for prevention of malaria in pregnant women, administered during antenatal clinic visits; IPT with SP in infants (IPTi), given when children come for routine vaccinations; and, in areas of seasonal transmission, seasonal malaria chemoprevention with SP plus amodiaquine (SPAQ) for children <5 years of age, delivered monthly at home by a community health worker. Patients with SCD who are stable are recommended to visit the clinic regularly, providing an opportunity to administer chemoprevention under supervision, but for this approach to be effective, a long-acting antimalarial drug combination is needed.SPAQ is effective when administered monthly, but protection wanes rapidly after about 4 weeks . MQ-arteIn Nigeria, patients with SCD who are stable are recommended to attend the clinic once every 2 months. The aim of this trial was, therefore, to compare the efficacy, safety, and tolerability of either IPT with MQAS given over 3 days or SPAQ (a single dose of SP plus AQ over 3 days) administered under supervision at routine bimonthly (once every 2 months) clinic visits to the efficacy, safety, and tolerability of the standard regimen of daily proguanil in patients with SCD.This study was undertaken at the SCD clinic of the Outpatient Unit of the University of Ilorin Teaching Hospital, Kwara state, Nigeria. Patients aged \u22656 months and weighing \u22655 kg with a documented genotype of SS or SC were invited to participate. Those who had an acute illness or any additional chronic disease, had a known allergy to any of the study drugs, or had been treated during the 2 weeks prior to recruitment with SP, AQ, or MQ were excluded. Signed consent was sought from the participants or their parents after the aims and procedures of the study had been explained.A list of randomly permuted blocks of 9, generated using Stata, version 11.1, was used to randomly assign eligible patients to receive bimonthly IPT with MQAS or SPAQ or standard daily treatment with proguanil. Participants were asked to return to the clinic once every 2 months for the next year to receive their treatment and, in the case of the proguanil group, to be given the next supply of proguanil tablets. Treatment allocations were kept inside opaque sealed envelopes. When a participant was enrolled, they were assigned the next envelope in numerical sequence. Their name and the date were written on the envelope, which was then opened to determine treatment allocation. Participants were issued identification cards bearing the participant's name, study number, and phone numbers of the investigators and were contacted by phone the day before each clinic visit to remind them to attend.P. falciparum and analysis of molecular markers of drug resistance. Nested PCR was used to amplify fragments at loci of 3 genes in which mutations are associated with pyrimethamine resistance , sulfadoxine resistance , and chloroquine resistance  [Participants were asked to record any illness in a diary, and at each clinic visit patients were asked about symptoms experienced since their last visit. They were then given a clinical examination, a venous blood sample was collected for hematological and biochemical measurements, a finger-prick blood sample was obtained for making thick and thin blood smears for microscopy, and a blood spot was collected onto filter paper for polymerase chain reaction (PCR) detection of  gene 1) , 20.For patients assigned to receive bimonthly IPT, the first dose of the 3-day regimen was administered by study staff, and the remaining doses were given to the participant or their caregiver to administer at home on each of the next 2 days. For those in the proguanil group, a dose of proguanil was administered, unused tablets were counted, and a supply of tablets for the next 2 months was given to the participant or their caregiver. All patients were given folic acid tablets and vitamin C tablets, to be taken daily, the standard of care for SCD in Nigeria. Study participants were asked to return 3 days after the clinic visit to be interviewed about adverse events over the previous 3 days and, for those in the IPT groups, to check adherence to the home doses by tablet counts. Those who were not able to be present for interview were contacted by phone, and details were cross-checked at their next visit. Participants were asked to come to the hospital if they were unwell at any time, where they were managed as outpatients or admitted to the pediatric ward. If a patient had suspected malaria, a rapid diagnostic test was performed to determine treatment.Supplementary Materials. The primary end point, for which the trial was powered, was the occurrence of any adverse event. Secondary end points included the occurrence of vomiting, adherence to the regimen, the incidence of malaria, the incidence of illnesses treated in outpatient settings, and the incidence of illnesses treated in inpatient settings. Statistical analysis was based on intention to treat and included all patients who were randomized. Statistical methods are described in the Supplementary Materials.The study was open label; although participants and study physicians unblinded to treatment, laboratory staff were unaware of treatment allocations. Laboratory methods are described in the Any study subject who had malaria  was treated with artemether/lumefantrine over 3 days. If this malaria treatment occurred at a scheduled clinic visit, the patient was not given the trial medications for that visit.Ethical approval for the study was obtained from the ethics committee of the University of Ilorin Teaching Hospital and the London School of Hygiene and Tropical Medicine. An independent data safety monitoring board provided oversight for the trial.Supplementary Table 1). Twenty-three of 270 samples (9%) were positive for P. falciparum by PCR at baseline, of which 9 (39%) carried the mdr1 mutation N86Y, 20 (87%) carried the dhfr triple mutations S108N, N51I, and C59R, and 18 (78%) carried the dhps double mutations A437G and K540E. The flow of participants through the study is shown in Figure Patients were recruited from 26 September 2011 to 12 April 2012, and follow-up for the last patients ended in April 2013. A total of 318 patients were screened; 270 who met inclusion criteria were randomly assigned in a 1:1:1 ratio to receive MQAS once every 2 months, SPAQ once every 2 months, or daily proguanil. Baseline characteristics were similar in the 3 groups . Pill counts at the clinic suggested that all patients took both home doses of MQAS or SPAQ, although this could not be verified. Adherence to daily proguanil doses, also based on tablet counts at the clinic, was poor, with 57% of patients using <75% of daily doses. Adherence was better in children aged <10 years than in older children (Supplementary Table 4).Doses of study drugs administered are shown in Supplementary Table 6). Admissions were more frequent among female participants than male participants, in all 3 groups. Less severe illnesses treated in outpatient settings were most frequent in the proguanil group, especially among females. The protective efficacy against episodes of outpatient illness , compared with the proguanil group, was 23%  for MQAS and 25%  for SPAQ . An adverse event during the 3 days following treatment was reported by 24%  of patients who received MQAS, 14%  who received SPAQ, and 5.4%  who received proguanil . A similar proportion of male and female participants reported side effects on day 3. Among participants who received MQAS, the risk of vomiting was substantially higher for those who received MQ doses in excess of 35 mg/kg .When participants were asked about side effects on day 3 after each bimonthly visit, vomiting, body pain, and abdominal pain were the most frequently reported symptoms Table  and wereWhen participants were asked about side effects and illness symptoms recorded in their diary in the 2 months prior to a routine clinic visit, 18% in the proguanil group reported an adverse event, compared with 13% in each of the other groups Table . FemalesStaphylococcus aureus, a girl aged 6 years who died at home with a suspected cerebrovascular accident, a boy aged 10 years admitted with severe anemia and sepsis, and a boy aged 1 year with a febrile illness who died at home). Three deaths were in the proguanil group . There were no deaths in the SPAQ group.Seven participants died during the study . Four of these had received MQAS .Thirty-eight episodes of clinical malaria were recorded (15 inpatients and 23 outpatients). Incidence was lower in the MQAS group, with a relative protective efficacy of 61% , compared with the proguanil group. The relative efficacy of SPAQ versus that of proguanil was 36%  for this regimen to give a high degree of protection.IPT with MQAS administered when patients with SCD came for routine clinic visits was well-tolerated, mild adverse events, the most common of which were vomiting and abdominal pain, did not deter patients from returning for their clinic visits. MQAS given once every 2 months was more effective in preventing malaria than daily prophylaxis with proguanil. MQAS reduced the prevalence of parasitemia at routine clinic assessments and reduced all-cause illness episodes treated as outpatients but did not reduce the burden of severe disease requiring hospital admission. Adherence to daily proguanil prophylaxis was suboptimal, and P. falciparum malaria undertaken in Nigeria, Agomo et al [MQ can cause neurological and psychiatric side effects and should not be used in patients with psychiatric illness or epilepsy . Howevermo et al  reportedIllnesses treated in outpatient settings and reports of illness symptoms recorded in patient diaries were more common among female participants than among male participants who took daily proguanil. Some other studies have reported that girls with SCD can have more-frequent pain episodes than boys . Sex-speThe lack of an effect against hospital admissions in the MQAS group, despite a high efficacy against malaria, is consistent with the low prevalence of malaria in the study population. There was limited scope to demonstrate an impact on all-cause admissions, but it is possible a greater benefit would be seen in areas with more intense malaria transmission. In the 2010 malaria indicator survey in Nigeria, the national prevalence of malaria in children aged <5 years in urban areas was 23%, compared with 48% in rural areas .Supplementary Materials). Our trial was done in an area with year-round transmission. In areas of highly seasonal transmission, chemoprevention could be limited to the transmission period, as is now recommended for children aged <5 years in the Sahel subregion [Malaria prevention in patients with SCD using IPT with long-acting antimalarials has a number of advantages. It is recommended that stable patients should attend a clinic every 2 months, and the IPT strategy can take advantage of these routine visits. The first dose of the 3-day regimen can be administered by clinic staff or directly observed, and adherence in taking the further 2 doses is likely to be higher than that of a regimen that requires daily treatment. The cost of the MQAS regimen compares favorably with that of the proguanil regimen , and the estimates of efficacy were correspondingly imprecise. We did not investigate the pharmacokinetics of MQ used for bimonthly prophylaxis, and the incidence of malaria was too low to be able to estimate duration of protection directly, but such studies would be useful.Haemophilus influenzae type b vaccination. Nigeria introduced pentavalent vaccine, which included H. influenzae type b vaccine, in 2012 and plans to introduce pneumococcal vaccine, and the impact of these vaccines on survival among patients with SCD needs to be evaluated. A recent survey of SCD management practices in Nigeria [A high burden of severe disease was observed, with a rate of 0.6 episodes requiring admission per person per year and a mortality rate of 25 episodes per 1000 persons per year. Additional strategies are needed to reduce the burden of severe disease in patients with SCD in Africa. In the United States, an improvement in survival among patients with SCD has been attributed to introduction of pneumococcal and  Nigeria  found thSupplementary materials are available at The Journal of Infectious Diseases online . Supplementary materials consist of data provided by the author that are published to benefit the reader. The posted materials are not copyedited. The contents of all supplementary data are the sole responsibility of the authors. Questions or messages regarding errors should be addressed to the author."}
+{"text": "The Bangladeshi national treatment guidelines for uncomplicated malaria follow WHO recommendations but without G6PD testing prior to primaquine administration. A prospective observational study was conducted to assess the efficacy of the current antimalarial policy.P. falciparum infections, or with chloroquine (days 0\u20132) plus 14 days primaquine  for P. vivax infections. Hb was measured on days 0, 2 and 9 in all patients and also on days 16 and 30 in patients with P. vivax infection. Participants were followed for 30 days. The study was registered with the clinical trials website (NCT02389374).Patients with uncomplicated malaria, confirmed by microscopy, attending a health care facility in the Chittagong Hill Tracts, Bangladesh, were treated with artemether-lumefantrine (days 0\u20132) plus single dose primaquine (0.75mg/kg on day2) for P. falciparum, 30% P. vivax and 6% mixed infections). Median parasite clearance times were 22.0  hours for P. falciparum, 20.0 (IQR: 9.5\u201322.7) hours for P. vivax and 16.6 (IQR: 10.0\u201346.0) hours for mixed infections. All participants were afebrile within 48 hours, two patients with P. falciparum infection remained parasitemic at 48 hours. No patient had recurrent parasitaemia within 30 days. Adjusted male median G6PD activity was 7.82U/gHb. One male participant (1/174) had severe G6PD deficiency (<10% activity), five participants (5/174) had mild G6PD deficiency (10\u201360% activity). The Hb nadir occurred on day 2 prior to primaquine treatment in P. falciparum and P. vivax infected patients; mean fractional fall in Hb was -8.8% (95%CI -6.7% to -11.0%) and -7.4% (95%CI: -4.5 to -10.4%) respectively.Between September 2014 and February 2015 a total of 181 patients were enrolled (64% The current antimalarial policy remains effective. The prevalence of G6PD deficiency was low. Main contribution to haemolysis in G6PD normal individuals was attributable to acute malaria rather than primaquine administration.NCT02389374ClinicalTrials.gov  Plasmodium falciparum infection, and chloroquine (CQ) plus a 14 day regimen of primaquine (14DPQ) for those with P. vivax infection. G6PD testing is not done on a routine basis before primaquine treatment. Patients co-infected with P. falciparum and P. vivax are treated with AL plus 14DPQ.In Bangladesh approximately 14 million people are at risk of malaria infection, with 13 out of 64 districts reporting clinical cases in 2012 ..45].P. falciparum mono-infection, all participants receiving 14DPQ where classified as G6PD normal. In the remaining patients receiving 14DPQ only one had an Hb drop of greater than 25% (to 10.5gHb/dL) after starting PQ radical cure.Malaria, irrespective of species had a far greater impact on Hb concentrations than the subsequent administration of PQ. While no study participant required a blood transfusion or had apparent haemoglobinuria, one patient (1.5%) with severe G6PD deficiency was excluded from PQ radical cure. Interestingly all other participants suffering from mild G6PD deficiency suffered from a The national treatment guidelines recommend a dose of 0.75mg/kg for SDPQ whereas the current WHO treatment guidelines consider a single dose of 0.25 mg/kg PQ safe even in G6PD deficient patients . This stThe current national malaria treatment guidelines in Bangladesh appear to be efficacious, although the delay in parasite clearance in two patients treated with AL warrants further investigation. The current Bangladeshi recommendations for single dose PQ are higher as suggested by the WHO; reducing the total dose to 0.25 mg/kg should be considered in the absence of routine G6PD testing . The curS1 Fig(DOCX)Click here for additional data file.S1 File(DOCX)Click here for additional data file.S1 Table(DOCX)Click here for additional data file."}
+{"text": "In the Republic of Congo, previous epidemiological studies have only been conducted in the south of the country where it is most accessible. Nationally representative data on the efficacy of new anti-malarial tools are lacking in thecountry. As an initial step to close the gap, clinical efficacy of two artemisinin-based combinations, artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL), was assessed in Owando, a city in equatorial flooded forest in northern Republic of Congo.Plasmodium falciparum parasites/\u00b5l of blood were clinically examined, included after informed consent, and followed up for 28\u00a0days, according to the 2009 World Health Organization protocol. Patients were randomly assigned to co-formulated ASAQ (Coarsucam\u00ae) or AL (Coartem\u00ae) treatment groups. Plasmodium falciparum recrudescent isolates were compared to pre-treatment isolates by polymerase chain reaction (PCR) using msp1, msp2, and glurp genes to distinguish between re-infection and recrudescence.Under 12\u00a0years old febrile children attending public health facilities were screened for malaria parasites using lactate dehydrogenase (LDH)-based rapid diagnostic test (RDT) for malaria and microscopic examination of thick blood films. Patients with at least 1,000 asexual Between November 2012 and February 2013, 857 under 12\u00a0years old febrile children were screened, of whom 198 (23.1\u00a0%) had positive RDT and 167 (19.5\u00a0%) positive thick films. ASAQ and AL efficacies were 92.7 and 94.2\u00a0% before PCR correction, respectively. After genotyping, the overall efficacy was 100\u00a0% for ASAQ and 98.0\u00a0% for AL.The data reported here represent partially the burden of malaria in 0\u201311\u00a0years old febrile children examined in public health centres of Owando city and serve as reference for further studies. Both artemisinin-based combinations were highly efficacious in patients under 12\u00a0years old with acute uncomplicated malaria. ASAQ was associated with more adverse events, which may reduce compliance in unsupervised treatment.Trial registration: ACTRN12612000940875 Despite international commitments that have led to the global malaria strategy from which the first results are expected within the Millennium Development Goals in 2015 , the decAfter several decades of intensive use of chloroquine and sulfadoxine-pyrimethamine (SP) to treat uncomplicated malaria, these two classical anti-malarial drugs had become less effective \u20139, resulTo encourage the use of these new treatments by health personnel, the Congolese government decreed in 2008 free malaria treatment for all children less than 15\u00a0years old. ITNs have also been distributed free of charge to pregnant women and children under 5\u00a0years of age. This mass distribution was later extended to the entire population and performed regularly. Pregnant women also receive three doses of SP for intermittent preventive treatment of malaria. These measures have contributed to the reduction of malaria in pregnant women and children . HoweverThe present study was conducted to assess the therapeutic efficacy of ASAQ and AL from November 2012 to February 2013 in the city of Owando, located in equatorial flooded forest in northern Republic of Congo. All febrile children aged less than 12\u00a0years spontaneously consulting at one of two public health centres were screened for the presence of malaria parasites using rapid diagnostic test (RDT) and microscopy. The objectives of this study were to determine the burden of malaria among febrile children less than 12\u00a0years old consulting spontaneously at the health centres during the study period and to evaluate the tolerance and effectiveness of these two forms of ACT.P. falciparum [Owando  is the main administrative city of Cuvette Department located 550\u00a0km to the north of Brazzaville, the capital city of the Republic of Congo, and 70\u00a0km to the south of the Equator, along the Kouyou River, one of the tributaries of the Congo River , without any concomitant febrile illness or any signs of danger or severe and complicated malaria, were included according to the 2009 WHO protocol for the assessment of anti-malarial drug efficacy after informed consent [Basic clinical information  of patients from 0 to 11\u00a0years was recorded in individual clinical forms. In the laboratory, fingerpick capillary blood was collected for RDT for malaria , preparation of thick films, haematocrit measurement, and storage of parasite DNA. Giemsa-stained thick films were examined under the microscope to determine the parasite density. Children with parasite density equal to or greater than 1000 asexual parasites/\u00b5l of blood were examined by a medical doctor. Children with fever (axillary temperature\u00a0\u226537.5\u00a0\u00b0C), sufficient parasite load  received 1 tablet per dose for 5\u201314\u00a0kg body weight, 2 tablets per dose for 15\u201324\u00a0kg body weight, 3 tablets per dose for 25\u201334\u00a0kg body weight, and 4 tablets per dose for\u00a0\u226535\u00a0kg body weight. Each patient received a total of 6 doses . Patients treated with artesunate-amodiaquine  received one tablet containing 25\u00a0mg of AS/67.5\u00a0mg of AQ for\u00a0<9\u00a0kg body weight, one tablet of 50\u00a0mg of AS/135\u00a0mg AQ for 9 to <18\u00a0kg, one tablet of 100\u00a0mg of AS/270\u00a0mg AQ for 18 to <36\u00a0kg, and two tablets of 100\u00a0mg of AS/270\u00a0mg AQ for\u00a0\u226536\u00a0kg of body weight. ASAQ was administered once daily for 3\u00a0days. Both drugs were provided by Drug Resistance and Containment, Global Malaria Programme of the World Health Organization (WHO), Geneva, Switzerland.This was an open-label study. Eligible patients were allocated to one of the two treatment groups after randomization in blocks of 10 according to a pre-established list of treatment groups. Blocks 1, 3, 5, 7, and 9 were affected to ASAQ; patients assigned to blocks 2, 4, 6, 8, and 10 were treated with AL. Patients assigned to AL group  analysis. The filter papers were stored in airtight plastic bags with desiccant and stored at room temperature.The patients were followed on days 1, 2, 3, 7, 14, 21, and 28, as in the previous studies on the efficacy of ASAQ and AL in the Republic of Congo. Clinical examination and measurement of axillary temperature were performed during each visit, and any adverse events or unauthorized concomitant therapies were recorded. As recommended in the 2009 WHO protocol , blood fmsa1, NCBI Gene ID: 813575, PlasmoDBlocus tag: PF3D7_0930300), merozoite surface antigen gene 2 , and the fragment size of glutamine-rich protein  were compared, as recommended by the WHO [In case of treatment failure, parasites before treatment (day 0 pre-treatment sample) were compared to parasites detected on the day of treatment failure. Parasite DNA was extracted from filter papers using EZNA blood DNA kit  according to the manufacturer\u2019s recommendations. The allelic families and fragment size of merozoite surface antigen gene 1  to determine the percentage of febrile children attending public health facilities with malaria parasites detected with either microscopy or RDT. The performance of microscopy, as compared to RDT, was assessed by calculating the sensitivity (Se), specificity (Sp), positive predictive value (PPV), and negative predictive value (NPV). Cohen\u2019s kappa coefficient (\u03ba) was determined to measure the degree of agreement between the two diagnostic tests . The 95\u00a0The sample size was calculated based on an expected failure rate of 5, 10\u00a0% precision, and a confidence level of 95\u00a0%, with an estimated loss of 20\u00a0% due to exclusions, withdrawals, and loss-to-follow-up. At least 60 patients per treatment group were recruited .Clinical and parasitological data were analysed using the pre-programmed Excel spreadsheet provided by the Global Malaria Programme, WHO . Patients who were excluded or lost to follow-up during the 28-day period were excluded from further analysis. Per protocol analysis is recommended by the WHO protocol and allows comparison of data with those of other sentinel sites in the country, mainly Brazzaville. Treatment responses were classified before and after PCR correction, as described in previous studies , 16.t test. The percentage of early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), and adequate clinical and parasitological response (ACPR) was calculated. Treatment failure rate was defined as the number of patients responding with ETF, LCF, or LPF divided by the total number of included patients who completed the 28-day follow-up. The 95\u00a0% CI of the outcomes  was determined using the exact binomial method. Treatment outcomes before and after PCR correction were presented on Kaplan\u2013Meier survival curves and compared using the LogRank test. The percentages of patients responding with ACPR and the frequencies of adverse effects associated with each ACT were compared using the Fisher\u2019s exact test.The baseline characteristics of patients were compared using the Student\u2019s \u00ae were 100 and 95.5\u00a0%, respectively. The kappa coefficient (\u03ba) was 0.88, i.e. there was an excellent concordance between RDT and microscopy results.The study was conducted from November 2012 to February 2013. A total of 857 febrile children aged from 0 to 11\u00a0years were screened for malaria parasites  or AL (61 children) treatment group  patients treated with ASAQ still presented low parasitaemia. On day 3, all patients had negative smears.In AL-treated group, three patients were excluded after enrollment: two patients were not infected with malaria parasites on day 0 after controlling their thick films; 1 patient had repeated vomiting and was unable to swallow oral medication on day 1. Six AL-treated patients were lost to follow-up. On day 1, 11 of 56 (19.6\u00a0%) patients (per protocol population) were still febrile. On day 2 and day 3, all patients were afebrile. On day 2 and day 3, 7 (12.5\u00a0%) and 1 (1.8\u00a0%) patients had positive thick films, respectively. On day 7, all patients had negative microscopy results.Per protocol analysis of the ASAQ treatment group showed four cases of treatment failure: 1 of 55 (1.8\u00a0%) patients presented LCF and 3 (5.5\u00a0%), LPF Table\u00a0, 5.TableASAQ treatment was associated with asthenia, vomiting, pruritus, abdominal pain, and headache in a few patients, whereas few AL-treated children reported asthenia and vomiting. These adverse events occurred between day 1 and day 3 Table\u00a0. AstheniIn 2006, the Republic of Congo adopted a new drug policy for malaria, which includes the treatment of uncomplicated malaria with ASAQ and AL, treatment of severe and complicated malaria with parenteral quinine, and recommendations for collective and individual preventive measures . Long-laRDTs are important tools that contribute to fight effectively against malaria. The ease of use of these devices allows untrained medical personnel to use them after a short training. Despite many advantages, the use of RDTs in Congolese health facilities is limited by its high cost and/or lack of adequate supply to meet the country\u2019s demands. In public health centres with a microscope, the performance of microscopy to establish malaria diagnosis costs 500 FCFA (US $1.00). The local laboratory suppliers sell a box containing 25 RDTs for 25,000 FCFA, i.e. US $2.00 per RDT. At this price, RDT is not competitive in public health facilities.[In Uganda, the treatment of clinical malaria episodes was cost-effective when diagnosis was established with RDT, as compared to microscopic examination of thick blood films , while i, 28\u201331. \u201334.Under these conditions, microscopy is expected to continue to occupy an important place in the diagnosis of malaria in the context of declining malaria prevalence, with a decrease in the number of clinical malaria cases, and for determining parasite density, when required, as long as RDT is not subsidized like drugs. Whether malaria diagnosis is confirmed by microscopy or RDT, a strong commitment of sub-Saharan African states would be essential to implement the national and regional anti-malarial treatment guidelines .\u00ae RDT is known for its good performance [This is the first epidemiological study conducted on malaria in the northern part of the country reporting malaria percentage in 0\u201311\u00a0years old febrile children attending health facilities spontaneously and drug efficacy. A reliable diagnosis was established in individual patients using both microscopy and RDT. Advantage Mal Cardformance , 34, andformance . The comThe choice of age group 0\u201311\u00a0years constitutes one of the weaknesses of this study. An enrollment of all symptomatic malaria-infected patients (children and adults) would have generated more representative data in the health centres. The choice of 0\u201311\u00a0years old is justified by a limited quantity of available RDT. However, the 0\u201311\u00a0years age group is pertinent and of interest because it includes children under 5\u00a0years old who are considered as one of the target age groups, together with pregnant women, in the fight against malaria. The longitudinal study in Brazzaville from 2003 to 2007  had shown that in a suburban area of Brazzaville, 46.6\u00a0% of febrile children under 5\u00a0years old and 63.5\u00a0% of febrile patients aged between 5 and 10\u00a0years had clinical malaria , showed Despite the unavailability of inoculation rate data, the city of Owando, located in a forest with shallows that are often flooded and scattered dwellings, may be considered as an area of high transmission encountered in Congolese forest regions. However, based on the results of the present study, the percentage of malaria-associated clinical episodes  among patients 0\u201311\u00a0years attending Owando health centres appeared to be unusually low. Further studies at different times of the year are required to characterize malaria transmission in Owando.P. falciparum lactate dehydrogenase (PfLDH) can be used for rapid screening of patients for inclusion, and parasite density can be determined by microscopy [In therapeutic efficacy studies, RDT which detects croscopy . In the croscopy . These Acroscopy , 16. Advcroscopy \u201316 may bcroscopy . HoweverThis is the first study that provides data on clinical malaria episodes in patients aged between 0 and 11\u00a0years attending public health facilities in Owando, located in northern Republic of Congo, and clinical efficacy of two ACT (ASAQ and AL) for the treatment of uncomplicated falciparum malaria. Data presented here show a relatively low proportion of clinical episodes of malaria among febrile children attending public health facilities and high efficiency of both ACT recommended by the Congolese National Malaria Control Unit for the treatment of laboratory-confirmed uncomplicated malaria in health facilities."}
+{"text": "In Brazil, 99.7\u00a0% of malaria cases occur in the Amazon region. Although the number of cases is decreasing, the country accounted for almost 60\u00a0% of cases in the Americas Region, in 2013. Novel approaches for malaria treatment open the possibility of eliminating the disease, but suboptimal dispensing and lack of adherence influence treatment outcomes. The aim of this paper is to show the results on dispensing practices, non-adherence and determinants of non-adherence to treatment of non-complicated malaria.Plasmodium vivax and Plasmodium falciparum transmission in the Brazilian Amazon and based on the theoretical framework of the Mafalda Project, which included investigation of dispensing and adherence. The World Health Organization Rapid Evaluation Method has been used to estimate sample size. Individuals over 15\u00a0years of age with malaria were approached at health facilities and invited to participate through informed consent. Data was collected in chart review forms focusing on diagnosis, Plasmodium type, prescribing, and dispensing . Follow-up household interviews complemented data collection at health facility. Non-adherence was measured during the implementation phase, by self-reports and pill-counts. Analysis was descriptive and statistical tests were carried out. Determinants of non-adherence and quality of dispensing were assessed according to the literature.The study was conducted in six high-risk municipalities with P. vivax but inadequate for P. falciparum medicines. Non-adherent patients were 12.1\u00a0% according to self-reports and 21.8\u00a0% according to pill-counts. Results point to greater non-adherence among all P. falciparum patients and among malaria non-na\u00eeve patients. More patients informed understanding adverse effects than \u2018how to use\u2019 anti-malarials.The study involved 165 patients. Dispensing was done according to the national guidelines. Labelling was adequate for P. falciparum diagnosis and those in their second or more malaria episode. New taxonomies and concepts on adherence stress the importance of focusing on the individual patient. Interventions targeted to and tailored for malaria patients must be addressed by health policy and implemented by managers and clinicians.Non-adherent patients were mostly those with a  Plasmodium vivax and Plasmodium falciparum co-exist, and to a lesser extent, Plasmodium malariae is present.Despite the decrease in the number of malaria cases worldwide in the past decade, this disease remains a major public health problem. Brazil accounted for almost 60\u00a0% of cases in the Americas Region, with 178,613 cases reported in the year 2013. Transmission still occurs in 808 municipalities of the country [P. vivax malaria, for instance, the recommended treatment is the combination of chloroquine plus primaquine and this protocol remains unchanged.Following the recommendations of the Amsterdam conference in 1992, Brazil bases malaria control on early diagnosis and adequate treatment, while also favouring prevention strategies . For P. P. falciparum malaria treatment, from quinine plus doxycycline plus primaquine to artemisinin combination therapy (ACT). According to NMCP, this treatment change resulted in a decrease in total number of P. falciparum cases, from 24.9\u00a0% of all registered malaria cases in 2006, to <16.2\u00a0% in 2011 [In 2006, the Brazilian National Malaria Control Programme (NMCP) introduced changes in Lack of treatment adherence is frequent in malaria , due to Adherence has been measured by a series of direct and indirect methods, and is usually expressed as a percentage of total number of doses, according to dosing regimen , 8, 9. CP. vivax and P. falciparum transmission in high-risk municipalities of the Brazilian Amazon.The main goal of this study is to show the results on dispensing practices, non-adherence to treatment during the implementation phase and determinants of non-adherence to treatment of non-complicated malaria in settings with P. vivax and P. falciparum in high-risk municipalities of the Brazilian Amazon: organization of services, prescribing, dispensing and adherence to treatment\u201d). Initially, in order to subsidize the project, a comprehensive review of pharmaceutical services for malaria was carried out and published [The methods employed in this work were based on the theoretical framework of the \u201cMafalda\u201d Project  guidelines  and adapPlasmodium spp, treatment characteristics (including prescribing), dispensing , information given to patient by health worker  and compliance to national guideline. For the household survey, another data collection instrument, for objective data, was applied, according to treatment regime.Questionnaires, observation forms, interviews forms and chart review forms were used during field work, according to the dimension that was being investigated . SpecifiThe investigation was carried out in six malaria high-risk municipalities of the Brazilian Amazon, selected according to Annual Parasitic Index (API) and population. Up to four high-coverage primary health care facilities were selected to guarantee number of eligible individuals. Individuals of both genders, \u226515\u00a0years of age, excluding pregnant women, with parasitological confirmed mild malaria were followed throughout the study. More detailed information on the field study and participants may be found elsewhere .P. falciparum or to P. vivax treatment, on the second or on the fifth day of treatment (D2 or D5), respectively.All procedures for investigation of organization of services and prescribing were complemented by those designed to investigate dispensing and adherence. For information regarding these dimensions, patients were approached in a two-step process. Data collection during consultation at health facility and observation of dispensing practices was complemented by household interviews. Patients that had been recruited for the first part of the investigation  were asked if household follow-up visit was welcomed, which indicated their inclusion in the next part of the investigation. Data was collected, according to Analysis was based on the theoretical framework developed for the Mafalda Project . DispensP. falciparum) or D5 (P. vivax). Household visits occurred during the last day of treatment. Non-adherence was measured during the implementation phase, by self-reports (adherence accepted as no missed doses during treatment period) and pill counts (adherence accepted as quantity received as proxy of quantity consumed), both of which are expressed by percentages.Adherence was approached considering the assumption that all patients initiated treatment on D0 and discontinued treatment at the end of D2  Ethics in Research Committee gave study approval .P. vivax, all receiving first-line treatment  had received first-line treatment at the time of the study  had been diagnosed with P. vivax as well as for P. falciparum. Among 165 patients, 134 patients (81.2\u00a0%) received verbal instruction at dispensing; 112 (67.9\u00a0%) patients informed understanding of how to use the anti-malarials and 137 (83\u00a0%) of their possible adverse effects.Observation of dispensing for these patients in health facilities first resulted in findings related to dispensing requirements. Dispensing of first-line regimens followed indication, and was done in accordance to the national guideline, for P. vivax regimens. At the time of data collection, first-line regimen for P. falciparum included quinine sulfate, doxycycline and primaquine. However, labels for these medicines  were all inadequate. Labels presented problems among the 15 patients receiving alternative P. falciparum treatment, of which 14 received artemether\u2013lumefantrine and primaquine. Inadequately labelled medicine was also dispensed to one patient receiving mefloquine and primaquine for P. falciparum . Non-adherence during implementation measured by self-reports revealed 144 adherent patients (there was one missing measure for among P. vivax patients). Twenty patients (12.2\u00a0%), non-adherent in self-reports, informed they had stopped using at least one anti-malarial during treatment (Table\u00a0One hundred and sixty-five patients were visited for household interviews during investigation of adherence (134 kappa) between these two methods of measuring adherence was 0.74.Pill counts were conducted for 165 patients, and the quantity that they had in their possession, in relation to day of treatment, was accurate in 129 (78.2\u00a0%) and inaccurate for 36 (21.8\u00a0%), patients designated as non-adherent . All other variables were non-significant.Table\u00a0P. falciparum patients, of the 12 non-adherent by pill-counts, 11 were on the first-line regimen and one was on artemether\u2013lumefantrine and primaquine; of the eight non-adherent patients by self-reports, seven were on first-line treatment and one was on the alternative regimen with artemether\u2013lumefantrine and primaquine. The distribution of non-adherence determinants by malaria type was not possible because of sample size.Regarding Various possible reasons for non-adherence were given by individuals: forgetfulness (8 interviewees), occurrence of adverse effects (10), unwillingness to take medicine (6), alcohol consumption (1), \u2018felt cured\u2019 (3), other (20). Patients sometimes informed one, two or more reasons for non-adherence.P. vivax and P. falciparum in high-risk municipalities of the Brazilian Amazon: organization of services, prescribing, dispensing and adherence to treatment\u201d) was developed in order to provide data of the situation of pharmaceutical services for malaria in municipalities at high risk for the disease in the Amazon [The Mafalda Project  mentioned understanding adverse effects. These possible malaria treatment outcomes may be acutely felt by patients , and areP. falciparum patients and among malaria non-naive patients. For the first group, reasons may be associated to change in treatment regimens, from a three-medicine 5-day treatment to a single-medicine (ACT) 3-day treatment. Many patients experienced various episodes of P. falciparum malaria and were already accustomed to the treatment regimen. With the lack of adequate instructions and labels, switches might be confusing. Another possibility is the acuteness of adverse effects with the traditional P. falciparum regimen , leading patients to discontinue treatment as soon as they feel a little better.Results point to greater non-adherence among all More than suboptimal dosing history (implementation of treatment regimen), early discontinuation (or short persistence) is the largest single factor for a decrease in adherence . Three pP. falciparum and P. vivax.Adherence is a crucial step for any pharmacological treatment. Acute-phase, complex treatments, such as anti-malarial treatment, oblige prescriber-patient collaboration mainly as to the initiation and implementation steps of adherence . IntervePlasmodium falciparum and non-na\u00efve patients constitute possible target groups for adherence interventions in malaria treatment [P. falciparum cases [P. falciparum malaria before emergence of resistance to ACT [reatment . Brazil um cases  and the e to ACT . Howevere to ACT  must be e to ACT  and idene to ACT , 28.Studies on adherence cover many types of definitions of adherence and measures , 9. SeveP. falciparum, elimination). This may be the case with malaria, mainly because of complex treatment regimens [Concepts on adherence have traditionally followed a stepwise process. The WHO  has propregimens , 30.Through controlled studies, new concepts associated with adherence have emerged which subvert previous understanding of how adherence should be measured \u201310, 28. P. falciparum patients in respect to differences in treatment regimens.As the sample in this study was designed for a traditional measure of adherence, numbers produced an overall idea of adherent and non-adherent patients, in respect to treatment implementation. Determinants for non-adherence were consistent with the literature . HoweverP. falciparum and P. vivax patients in the Brazilian Amazon. Non-adherent patients were mostly those with a P. falciparum diagnosis and those in their second or more malaria episode. In face of new taxonomies and concepts on adherence and because the emergence of resistance to ACT and other anti-malarials are of utmost importance to public health, interventions targeted to and tailored for malaria patients must be addressed by health policy and implemented by managers and clinicians.This study produced an overview of non-adherence measured in the implementation phase and of determinants associated with non-adherence among"}
+{"text": "Plasmodium falciparum malaria. However, data on effectiveness and safety of AL in patients in non-endemic settings are limited.Data from clinical studies show that artemether-lumefantrine (AL) is effective and well tolerated in adults and children with uncomplicated P.\u00a0falciparum malaria in the USA, as reported to the National Malaria Surveillance System at the Centers for Disease Control and Prevention. Descriptive analyses included demographics, baseline characteristics, clinical effectiveness, and safety. From May 2010 to April 2015, demographics and baseline characteristics were collected for 203 patients and safety data for 108 patients. Treatment effectiveness data at day 7 were collected for 117 patients and at day 28 for 98 patients.A 5-year surveillance plan included all AL-treated adult and paediatric patients with confirmed or suspected 2. All patients with effectiveness data had confirmed (n\u00a0=\u00a0116) or suspected (n\u00a0=\u00a01) malaria. The overall cure rate for patients treated with AL was 91.5\u00a0% (95\u00a0% CI 84.8\u201395.8\u00a0%) at day 7 and 96.9\u00a0% (95\u00a0% CI 91.3\u201399.4\u00a0%) at day 28. Adverse events were reported in four (3.7\u00a0%) patients, and there were no new or unexpected safety signals.The majority of patients were male (58.6\u00a0%), Black (62.6\u00a0%), non-Hispanic (92.6\u00a0%), and likely malaria non-immune (80.8\u00a0%). The median age was 32\u00a0(range 1\u201388) years and the median body mass index was 25.5 (range 13.8\u201342.4) kg/mP.\u00a0falciparum malaria.AL was effective and well tolerated in the treatment of likely non-immune patients with  Both components are blood schizonticides with complementary pharmacokinetic profiles and dissimilar modes of action, thus providing synergistic anti-malarial activity , likely malaria immune status, malaria species , and status of malaria diagnosis .Data analysis was not based on a specific a priori statistical hypothesis and was simply descriptive. Summary statistics were presented for quantitative variables, and counts and percentages were calculated for categorical data. For the day 7 and day 28 cure rates, the proportions of cured patients and two-sided 95\u00a0% confidence intervals were calculated using exact Pearson-Clopper limits [A total of 203 AL-treated cases were reported from May 2010 to April 2015. Figure\u00a0P.\u00a0falciparum or another species. For the day 28 cure rate, effectiveness data were available for 98 patients with confirmed malaria. Effectiveness results were analysed with missing effectiveness data excluded.Table\u00a0At day 7, the overall cure rate was 91.5\u00a0% (107/117). Out of the ten patients who did not resolve their malaria by day 7, seven had severe malaria at baseline and should not have received AL to begin with, one had an adverse event of nausea that was considered by the treating physician to be related to AL and was switched to atovaquone-proguanil, one was rehospitalized because of haemolytic anaemia thought to be due to persistent malaria and was discharged on quinine and doxycycline, and one did not complete the treatment course due to a supply shortage at the treating hospital. At day 28, clinical effectiveness data were available for 98 patients and the overall cure rate was 96.9\u00a0% (95/98). Three patients experienced a recurrence of signs or symptoms of malaria within 28\u00a0days. Of the 19 patients who were assessed at day 7 but not at day 28, data were missing for nine and were not collected for the ten patients who did not resolve at day 7 and were mainly switched to other anti-malarials. The day 7 (91.5\u00a0%) cure rate is not a true representation of the effectiveness of AL as at least seven patients should not have received AL for treatment of their malaria. Notably, although 22 other patients with severe malaria resolved upon receiving AL without further treatment, AL is indicated only for the treatment of uncomplicated falciparum malaria.Plasmodium\u00a0vivax. AL cleared the original infection, but one month later, after re-appearance of symptoms, the patient was treated with atovaquone-proguanil, doxycycline and primaquine, and the vivax co-infection was cleared.Adverse events (AEs) were reported in four 3.7\u00a0%) patients , and the lag time between occurrence of a case and its reporting to CDC can vary from a few days to 1\u00a0year or longer. Data available from 150 cases showed that they were reported to CDC on average 84 (range 1\u2013733) days after the onset of signs and symptoms. As a result, there are possibly additional cases that may have been applicable to the timeframe of this project but were excluded due to a delay in reporting.The delay in case reporting and retrospective data collection, small sample size, and incomplete data mainly because of loss to follow-up are all inherent limitations of this surveillance project. Other limitations include the non-comparative and non-randomized methods of data collection, unsupervised treatment, and clinical effectiveness and safety outcomes that were solely determined by the healthcare provider. Therefore, effectiveness data from this surveillance could hardly be considered as robust as those from some effectiveness studies conducted in endemic countries , 19. DesAL was effective and well tolerated in the treatment of likely non-immune patients in the USA with falciparum malaria."}
+{"text": "This review compared two of the ACTs\u2014Dihydroartemisinin-Piperaquine (DP) and Artemether-Lumefantrine (AL) to provide evidence which one has the ability to offer superior posttreatment prophylaxis at 28 and 42 days posttreatment. Four databases  were searched on June 2, 2013 and a total of seven randomized controlled trials conducted in sub-Sahara Africa were included. Results involving 2, 340 participants indicates that reduction in risk for recurrent new falciparum infections (RNIs) was 79% at day 28 in favour of DP , and at day 42 was 44% favouring DP . No significant difference was seen in treatment failure rates between the two drugs at days 28 and 42. It is concluded that use of DP offers superior posttreatment prophylaxis compared to AL in the study areas. Hence DP can help reduce malaria cases in such areas more than AL.Malaria contributes significantly to the global disease burden. The World Health Organization recommended the use of artemisinin-based combination therapies (ACTs) for treatment of uncomplicated  Plasmodium which are transmitted to susceptible persons through the bite of infected female Anopheles mosquitoes.Malaria has attracted global attention as one of the world's leading major diseases due to its high morbidity and mortality. The World Health Organization (WHO) estimated that about 3.3 billion people are at risk of malaria because they live in malaria endemic areas  and abouThe burden of malaria is greatest in sub-Sahara Africa where approximately 80% of the global malaria cases as well as 90% of fatalities occur . Childre Plasmodium falciparum had developed resistance to chloroquine which was the most affordable and readily available antimalaria drug in Africa . This implies that there was an average of 55% reduction in risk for TTF at day 28 in favour of DP treatment; largest plausible reduction possible was 80%; however, the upper boundary of the 95% CI includes the number of no effect (1) and a risk for harm of 1.2%.Result on total treatment failure (TTF) due to recrudescence was obtained from six studies and involved a total of 3,172 patients. Of this, 1,861 participants received DP, out of which 107 (5.7%) experienced treatment failure at day 28. On the other hand, 1,311 received AL, of which 80 (6.1%) experienced treatment failure. The pooled RR yielded 0.453, 95% CI: , which means that for every 63 patients treated with DP, one case of treatment failure was prevented, which would have occurred if such patients had received AL treatment.Extent of heterogeneity was 47%  and includes 14% harm of failure. Test for heterogeneity in random effect model shows high statistical heterogeneity of 71% across the studies  = 3, I2 = 71%, P = 0.02)\u2014see Data on TTF at day 42 was extracted from four studies involving 2, 662 participants, out of which treatment failure was reported in 274 participants representing 10.3%. A total of 1,598 participants were those treated with DP and 161 of them (10.1%) experienced treatment failure. Those who received AL treatment were 1,064 participants out of which 113(10.6%) experienced failure, an indication that DP was associated with a marginal reduction in treatment failure compared to AL. The pooled estimate for the RR was 0.560 with 95% CI of . This indicates that risk associated with a patient getting new infections at day 28 was significantly reduced averagely by 79% in favour of DP treatment. The lowest plausible reduction was 69% and highest was 86%; the difference was statistically significant (P < 0.001)\u2014Results from four studies involving a total of 2,340 patients showed that 104 of the patients (representing 4.4%) acquired new falciparum parasites. Those treated with DP were 1,598, out of which 218 (14%) acquired new falciparum infections. On the other hand, a total of 1,064 received AL, of which 237 (22%) experienced new infections. The pooled RR was 0.557, 95% CI [0.342 to 0.908]. This means that the risk of a patient acquiring new falciparum infection at 42 day following treatment was averagely reduced by 44% in favour of DP compared to AL\u2014see P = 0.02. There was a substantial heterogeneity (extent was 83%) associated with this result .Results extracted from four studies involving a total of 2,662 patients indicated that 455 patients representing 17% experienced new infections with falciparum infections (RNIs) and treatment failure. The null hypothesis which was tested was that there is no difference between the two drugs in reducing risk of treatment failure and recurrent new falciparum infections at days 28 and 42. However, the overall evidence gathered suggests that patients who were treated with DP experienced less total treatment failure (TTF) and less recurrent new falciparum infections (RNIs) than those who received AL.This review compared two ACTs\u2014Dihydroartemisinin-Piperaquine (DP) and Artemether-Lumefantrine (AL)\u2014to determine which has the greatest effect in reducing recurrent newMajority of studies included in this review were conducted in East Africa and only few were carried out in West Africa and therefore findings and conclusion are more applicable to countries in the East Africa region. The evidence synthesized indicates that both DP and AL are effective in preventing treatment failure at days 28 and 42 after treatment. However, DP reduced treatment failure higher than AL at 28 and 42 days but the difference in magnitude of failure reduction between the two drugs is clinically marginal and not statistically significant. This finding concurs with that of earlier review conducted on studies done in Asia, America, and Africa , in whicIt has been speculated that DP could offer a greater posttreatment prophylaxis (PTP) than other ACTs due to its longer half-life by which it could exert longer efficacy against newly infecting parasites and reduce risk for development of both clinical malaria and resistance parasite strains . However falciparum infections (RNIs) in this review demonstrates that DP offered greater and significant PTP for patients against RNI than AL. Average percentage reduction in risk for RNI was up to 79% at day 28 in favour of DP and up to 44% at day 42 in favour of DP. There is, however, substantially significant statistical heterogeneity (extent was 83%) associated with pooled result on RNI at day 42 in this review. The possible source of this heterogeneity is attributed to the variability in malaria infection transmission rate of the various study sites; some of the studies were conducted in settings where rate of malaria transmission intensity was very high while other studies were conducted in areas of relatively low malaria transmission intensity. Thus, study settings with very high transmission intensity are likely to be associated with higher rates of Anopheles mosquito bites which will result in recurrent new infections more than settings with low transmission intensities. Even though participants were provided with insecticide treated nets, it cannot be ascertained whether patients actually slept in the nets to prevent mosquito bites after treatment and this could also affect the proportion of patient who experienced recurrent new infections.Result on prevention of recurrent new falciparum parasites to DP  and 44% in favour of DP at day 42 . It is, therefore, concluded that treatment of uncomplicated falciparum malaria using Dihydroartemisinin-Piperaquine (DP) in high transmission areas in sub-Sahara Africa  could result in an average reduction in risk for recurrent new falciparum infections of up to 79% and 44% within 28 and 42 days, respectively, compared to Artemether-Lumefantrine (AL). And this implies that use of DP can help reduce burden of malaria in such areas more than AL. However, both DP and AL have similar effectiveness in preventing treatment failure though DP has marginal benefit over AL.On the other hand, DP offers a significant posttreatment prophylaxis against recurrent new falciparum strains. More studies should be conducted in other parts of the sub-Sahara Africa such as West Africa to determine stronger evidence which will be more applicable to countries in that area.It is recommended that the antimalaria drug policy in countries especially in East Africa should be streamlined to include use of Dihydroartemisinin-Piperaquine alongside Artemether-Lumefantrine and other ACTs in countries where it is found to be effective but this must be done bearing in mind the potential risk for development of resistantIt has been identified that the current study protocol by the WHO regarding antimalaria drug efficacy research does not incorporate specific outcome measure on recurrent new infections. It is, therefore, recommended that measurement of rate of recurrent new infections should be incorporated into future guidelines, and future trial investigators should make direct assessment on it."}
+{"text": "Plasmodium vivax is prevalent in eastern Malaysia. Artesunate-mefloquine is an efficacious artemisinin combination therapy (ACT) for all malaria species. Wider CQ-efficacy surveillance is needed in vivax-endemic regions with earlier replacement with ACT when treatment failure is detected.High-grade chloroquine (CQ)-resistant  Background.\u2003Chloroquine (CQ)-resistant Plasmodium vivax is increasingly reported throughout southeast Asia. The efficacy of CQ and alternative artemisinin combination therapies (ACTs) for vivax malaria in Malaysia is unknown.Methods.\u2003A randomized, controlled trial of CQ vs artesunate-mefloquine (AS-MQ) for uncomplicated vivax malaria was conducted in 3 district hospitals in Sabah, Malaysia. Primaquine was administered on day 28. The primary outcome was the cumulative risk of treatment failure by day 28 by Kaplan\u2013Meier analysis.Results.\u2003From 2012 to 2014, 103 adults and children were enrolled. Treatment failure by day 28 was 61.1%  after CQ and 0%  following AS-MQ (P < .001), of which 8.2%  were early treatment failures. All patients with treatment failure had therapeutic plasma CQ concentrations at day 7. Compared with CQ, AS-MQ was associated with faster parasite clearance  and fever clearance  and with lower risk of anemia at day 28 . Gametocytes were present at day 28 in 23.8% (10/42) of patients following CQ vs none with AS-MQ (P < .001). AS-MQ resulted in lower bed occupancy: 4037 vs 6510 days/1000 patients . One patient developed severe anemia not regarded as related to their AS-MQ treatment.Conclusions.\u2003High-grade CQ-resistant P. vivax is prevalent in eastern Malaysia. AS-MQ is an efficacious ACT for all malaria species. Wider CQ-efficacy surveillance is needed in vivax-endemic regions with earlier replacement with ACT when treatment failure is detected.Clinical Trials Registration. NCT01708876. Plasmodium vivax is estimated at 2.48 billion people [Plasmodium vivax causes substantial morbidity due to recurrent infections [P. vivax in Sabah, Malaysia, was consistent with low endemicity and multiple subpopulations, resulting in vulnerability to epidemic expansions from new or introduced parasite strains [The global population at risk of malaria due to infection with n people , with sofections , which afections , and abifections \u20138. Whilefections , 10, trafections . A recen strains .P. vivax in the presence of therapeutic CQ concentrations has been documented in a returned traveller from Sabah in 1996 [P. vivax, including high-grade CQ-resistance in Indonesia and Papua New Guinea and low-grade CQ-resistance in Philippines, Thailand, and Vietnam [P. vivax, ongoing clinical monitoring of antimalarial efficacy is vital to guide effective P. vivax treatment policy [Current Malaysian Ministry of Health malaria treatment guidelines recommend chloroquine (CQ) and primaquine for uncomplicated vivax malaria . CQ-resi in 1996  and has  in 1996 , 16, but Vietnam . The aret policy , particuPlasmodium falciparum and P. vivax [Plasmodium knowlesi is also prevalent [P. vivax, P. knowlesi and P. falciparum [P. falciparum.There is growing support for artemisinin-based combination therapy (ACT) as a unified first-line treatment in areas co-endemic for P. vivax . In Sabarevalent , with frlciparum . A unifilciparum , 21; furP. falciparum and P. knowlesi malaria in Malaysia [P. vivax and determine whether the fixed combination of AS-MQ was superior to CQ for blood-stage treatment of uncomplicated vivax malaria in Malaysia.Artesunate-mefloquine (AS-MQ) is 1 of 2 ACTs recommended for Malaysia . Our aimThis study was approved by the relevant human medical research ethics committees of the Malaysian Ministry of Health, and Menzies School of Health Research, Australia. Approval for drug analysis was obtained from the Australian Defence Human Research Ethics Committee (ADHREC 717-13).P. vivax monoinfection, negative for P. falciparum by rapid diagnostic test (histidine-rich protein-2), and fever (\u226537.5\u00b0C) or history of fever in the last 48 hours. Written informed consent was obtained from the patient or their guardian. Exclusion criteria were severe malaria or warning signs according to modified World Health Organization (WHO) 2010 criteria [This 2-arm, randomized, open-label trial was conducted at 3 hospitals in Sabah: Kudat, Kota Marudu, and Pitas District. Included were patients with acute, uncomplicated vivax malaria presenting to the study hospitals, aged >1 year and weighing >10 kg, with microscopic diagnosis of  fever \u22653.5\u00b0C or hAS-MQ  was administered as a fixed-dose combination with 3 oral formulations . Doses were administered at enrollment and 24 and 48 hours after . CQ diphosphate , consisting of 155 mg base tablets, was administered at enrollment and 6, 24, and 48 hours after . Primaquine  as 15-mg tablets was commenced on day 28 (per WHO treatment surveillance guidelines in order not to underestimate the true risk of CQ resistance with concomitant primaquine ) to gluce parasite-time profile [P. vivax gametocytes at day 28; the risk of adverse events; and the length of hospitalization. A total of 66 patients in each arm were required to detect an absolute difference of 15% in day 28 treatment failure rates between the study arms, with 80% power and 95% confidence, assuming a CQ efficacy of 85% and 10% loss to follow-up.The primary endpoint was the cumulative risk of treatment failure by day 28 as defined by WHO . Seconda profile ; the ris profile ; the fraPatient allocation codes were computer generated in blocks of 20 for each study arm using STATA (version 12) by an independent statistician, separately sealed in an opaque envelope, and opened by a study nurse after patient enrollment. The primary endpoint and other parasitological measurements were determined by microscopists blinded to patient treatment allocation.Polymerase chain reaction (PCR) confirmation of species  and G6PDPlasmodium vivax genotyping was performed with 9 previously described short tandem repeat markers: Pv3.27, msp1F3, MS1, MS5, MS8, MS10, MS12, MS16, and MS20  [aterial) .Supplementary Material). CQ resistance was defined as failure to clear peripheral P. vivax parasitemia on blood smear or recurrent parasitemia within 42 days in the presence of plasma blood concentration of CQ + DCQ in excess of the minimal effective concentration (MEC > 15 ng/mL CQ + DCQ), corresponding to a whole blood concentration of 100 ng/mL [Heparinized blood samples were collected on day 7 , 30 and 00 ng/mL , 31. Patt test or Wilcoxon-Mann\u2013Whitney test for continuous variables and using \u03c72 or Fisher exact test for categorical variables. Microscopic asexual parasitemia and gametocytemia were calculated from thick blood smears [e parasite counts vs time per the parasite clearance methodology of the World Wide Antimalarial Resistance Network [Data were double-entered into Epidata (version 3.1) and analyzed with STATA (version 12). Primary analysis was intention-to-treat, with incidence risk of treatment failure at days 28 and 42 calculated using the Kaplan\u2013Meier method and compared using the Mantel-Haenszel log-rank test. Intergroup differences were compared using the Student d smears . Best-fi Network .P. vivax monoinfection, of whom 58.4% (111/190) were enrolled in the study  of malaria patients were diagnosed with In view of the high risk of treatment failure in the CQ treatment arm, an independent interim analysis was undertaken following which the study was stopped due to the large difference in the primary outcome between treatment arms.There were no differences in baseline patient demographics between treatment groups (Table P < .001), and this was reflected by the slope of the log10 normalized parasite clearance curve . Clearance was faster after AS-MQ than CQ for both ring-stage  and the trophozoite-stages . Fever clearance time correlated with PCT  and was faster with AS-MQ  compared with CQ . The median number of days spent in the hospital, including readmission for recurrent parasitemia, was higher in the CQ arm  vs the AS-MQ arm , the cumulative bed occupancy being 6510 days/1000 persons and 4037 days/1000 persons after CQ and AS-MQ treatment, respectively .Parasite clearance times (PCT) were significantly faster with AS-MQ  compared with CQ ; P < .001; Figure P < .001). Of the 26 patients with CQ treatment failure treated with AS-MQ, 76.9% (20) could be followed up for an additional 28-day period from the date of administration, none of whom had further recurrence.By day 28 the cumulative risk of treatment failure was 61.1%  in the CQ arm compared with 0% (0/46) in the AS-MQ arm . There were no differences in any of the other secondary hematological outcomes. One adult patient in the AS-MQ arm developed severe anemia  of patients treated with CQ were anemic compared with 24.4% (11/45) treated with AS-MQ  on day Blood samples were available from day 0 and the day of failure for 77% (17/22) of patients with LTF. PCR amplification/genotyping was successful in all samples analyzed. Comparison of the genotype profiles between day 0 and recurrent infections revealed that all 17 recurrences were homologous. The PCR-adjusted cure rate was therefore identical to the clinical/parasitological cure rate.P = .0013) .Blood samples were obtained from 22 (88%) of the 25 patients with treatment failure by day 28 who completed a full course of CQ. Of the 16 patients with no treatment failure, blood samples were obtained from 15 (94%). All patients had therapeutic plasma CQ concentrations at day 7 post-treatment. Of those with recurrent parasitemia, 45% (9/20) had composite plasma CQ and DCQ concentrations greater than the MEC of 15 ng/mL on the day of recurrence, while those below this cutoff presented later .One serious adverse event was reported in the AS-MQ arm: severe anemia 3 days after commencing treatment, not thought to be related to the study medication. Other adverse events between treatment groups were comparable, including the proportion with headache (91%), dizziness (77%), vomiting (51%), or hearing changes potentially associated with MQ use, or in grouped-system analyses  of AS compared with CQ [Patients treated with AS-MQ exhibited a faster early therapeutic response, with significantly shorter parasite and fever clearance times compared with CQ. This benefit was apparent even in those patients with no subsequent recurrence. The higher rates of both ring and trophozoite clearance with AS-MQ are also consistent with previous in vitro drug susceptibility testing, demonstrating relatively lower inhibitory concentrations . Based on our study's bed-occupancy data, this gives a minimum cost-saving difference for hospital inpatient days of $163 218 per 1000 patients treated with ACT, which far exceeds the higher medication cost of $1750 per 1000 patients.P. vivax is challenging [P. vivax in the peripheral blood smear within 28 days of CQ treatment is highly indicative of resistance. However, recurrent infections can be due to inadequate drug absorption or reinfection in the presence of low blood CQ concentrations following rapid drug metabolism and elimination. All patients treated with CQ in our study had observed administration with correct dosage based on body weight and confirmed therapeutic plasma CQ concentrations at day 7. Furthermore, all recurrent infections were homologous, which is in keeping with them being true recrudescences, although the possibility of homologous CQ-sensitive relapses cannot be excluded in those recurring in the presence of CQ + DCQ concentrations below the MEC. At the time of recurrence, 9 of 20 patients assessed had plasma CQ + DCQ concentrations in excess of 15 ng/mL , demonstrating parasite growth in the presence of adequate drug concentrations. By definition, these parasites are CQ resistant.The interpretation of CQ efficacy in llenging . ConfirmP. vivax recurrences between day 28 and day 42.Limitations of the study included the relatively small number of patients enrolled in each arm. Furthermore, using the standard commercially sourced medications, it was not possible to blind research staff to treatment allocation after the initial randomization. Importantly, however, the primary and most secondary outcome measures were determined by microscopy, with study microscopists blinded to treatment allocation. Primaquine administration was delayed to day 28 since its blood-stage activity limits can confound the interpretation of CQ efficacy . HoweverP. vivax in Malaysia. It demonstrates hitherto undetected high-grade CQ resistance and associated high risk of clinical failure to CQ treatment. Despite increasing reports of CQ resistance, CQ remains the first-line therapy for P. vivax in most vivax-endemic countries. The identification of high levels of CQ resistance in Malaysia highlights that CQ efficacy cannot be assumed and supports the need for ongoing evaluation of its efficacy in vivax-endemic regions [P. vivax in the study area. Additional clinical and public health advantages of AS-MQ treatment included faster parasite and fever clearance, earlier hospital discharge, lower bed occupancy rates, decreased anemia and morbidity, and lower transmission risk. Our findings support a switch to an ACT for vivax malaria in this region as recommended by the WHO if >10% resistance is documented [Plasmodium spp. would provide excellent efficacy for the artemisinin-sensitive P. falciparum and P. knowlesi and CQ-resistant P. vivax found in co-endemic areas such as Malaysia. Further therapeutic efficacy trials of alternative ACTs for uncomplicated P. vivax malaria addressing both safety and efficacy are warranted.This prospective study represents the first antimalarial efficacy study of  regions , 22. AS-Supplementary materials are available at http://cid.oxfordjournals.org. Consisting of data provided by the author to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the author, so questions or comments should be addressed to the author."}
+{"text": "Plasmodium vivax malaria in most endemic countries, but resistance is increasing. Monitoring of antimalarial efficacy is essential, but in P vivax infections the assessment of treatment efficacy is confounded by relapse from the dormant liver stages. We systematically reviewed P vivax malaria treatment efficacy studies to establish the global extent of chloroquine resistance.Chloroquine is the first-line treatment for P vivax malaria. We excluded studies that did not include supervised schizonticidal treatment without primaquine. We determined rates of chloroquine resistance according to P vivax malaria recurrence rates by day 28 whole-blood chloroquine concentrations at the time of recurrence and study enrolment criteria.We searched Medline, Web of Science, Embase, and the Cochrane Database of Systematic Reviews to identify studies published in English between Jan 1, 1960, and April 30, 2014, which investigated antimalarial treatment efficacy in P vivax. Clearance of parasitaemia assessed by microscopy in 95% of patients by day 2, or all patients by day 3, was 100% predictive of chloroquine sensitivity.We identified 129 eligible clinical trials involving 21\u2008694 patients at 179 study sites and 26 case reports describing 54 patients. Chloroquine resistance was present in 58 (53%) of 113 assessable study sites, spread across most countries that are endemic for P vivax, which is now present across most countries endemic for P vivax. Improved methods for monitoring of drug resistance are needed to inform antimalarial policy in these regions.Heterogeneity of study design and analysis has confounded global surveillance of chloroquine-resistant Wellcome Trust (UK). Plasmodium vivax is a major cause of morbidity, causing 72\u2013390 million clinical cases of malaria worldwide each year.Plasmodium falciparum malaria, P vivax forms dormant liver stages (hypnozoites), which cause relapses of infection weeks to months after the initial attack. In some areas, recurrent infections can occur as often as every 3 weeks, with relapses the main cause of vivax illness. Acute febrile episodes are associated with anaemia, intrauterine growth retardation,7P falciparum, which is the main cause of malaria mortality. Substantial progress has been made in reduction of the global burden of malaria, much of which has been attributed to increases in access to health-care services, early diagnosis, treatment with highly effective antimalarial drug regimens, and deployment of insecticide-treated bednets. One of the greatest threats to control and elimination efforts is the emergence and spread of antimalarial drug resistance. This recurring problem has plagued malaria therapeutics for more than 60 years. In the early 1950s, chloroquine became established as the best antimalarial drug for all human malarias, but within a decade resistance had emerged. Chloroquine-resistant P falciparum spread throughout malaria-endemic countries killing millions of people. Antimalarial drug resistance is measured by the prevalence and severity of treatment failure. In P falciparum malaria recrudescent infection is defined as the re-emergence of a genetically identical parasite in the peripheral blood after its initial treatment with a usually curative drug regimen (thereby distinguishing it from a newly acquired infection). P vivax clinical efficacy studies are more difficult to interpret, because recurrent infections can arise from recrudescence, reinfection, or relapses (arising from the dormant liver stages).Policy makers and malaria researchers have generally focused on P vivax malaria in most endemic countries. When given with primaquine , the combination is highly effective against both the acute illness and in prevention of relapses from hypnozoites. Chloroquine-resistant P vivax was first reported in 1989, almost 30 years after chloroquine-resistant P falciparum was first noted.P vivax has almost certainly contributed to the delayed recognition of this emerging problem.P vivax is prevalent (such as Indonesia and Oceania), partly effective drug treatments and consequent recurrent infections are an important contributing factor to severe anaemia from P vivax malaria.P vivax drug trials that summarises the geographical extent and level of evidence for reduced P vivax susceptibility to chloroquine and estimate risk of recurrence by day 28.Chloroquine is the first-line treatment for P vivax were also screened for relevant studies in English.Our analysis adhered to the recommendations of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines.We systematically extracted data on study characteristics, recurrence, and side-effects from the articles and entered into an EpiInfo database (version 3.5.1). We extracted the study method, including location of study site, inclusion and exclusion criteria, clinical setting, randomisation and masking, and the chloroquine dose regimen , and the co-administration of primaquine; these data are available online.P vivax parasitaemia at day 28. Results of previous studies have shown that the prolonged elimination of chloroquine provides blood concentrations that prevent recurrence of chloroquine-sensitive P vivax for about 35 days. Hence, no recurrent parasitaemia should be noted within 28 days of treatment in patients taking a complete treatment course with adequate absorption.P vivax parasitaemia by day 28 is therefore a useful measure of chloroquine resistance, whether recurrence is caused by recrudescence, relapse, or a new infection. Secondary outcome measures included the proportion of patients with parasitaemia on days 1, 2, and 3; the day of the first recurrence; and the proportion of patients with early treatment failure, defined according to WHO criteria for P falciparum.17We extracted estimates of chloroquine efficacy and present them separately for each treatment group at each study location (termed site estimates). The primary outcome measure was the risk of recurrent We separated chloroquine resistance a priori into four categories according to the primary outcome. Category 1 resistance constituted greater than 10% recurrences by day 28 (with a lower 95% CI of >5%), irrespective of confirmation of adequate blood chloroquine concentration; occasional breakthrough recurrences do occur within 28 days of chloroquine treatment, but a risk greater than 10% in a large enough sample is very suggestive of resistance. Category 2 resistance included confirmation of recurrences within 28 days, in the presence of whole-blood chloroquine concentrations greater than 100 nm;P vivax during 28 days of follow-up, with the denominator being the number of patients followed until the study endpoint. We estimated the risk of recurrence for each chloroquine group, stratified by study location. We estimated the effect of primaquine by comparing recurrence rates in randomised studies in which patients were treated with and without concomitant primaquine. We estimated the comparative risk of recurrence by day 28 as Mantel-Haenszel odds ratios with Review Manager version 5.2. We analysed all data with SPSS v19.0 for Windows.We restricted analysis of drug efficacy to patients treated with a chloroquine regimen. We calculated the risk of recurrence per protocol with the number of patients with recurrent The funder had no role in study design, data collection, data analysis, data interpretation, or writing of the report. RNP had full access to all the data in the study, and had final responsibility for the decision to submit for publication.P vivax, of which we excluded 68 . 14 studies were designed as antirelapse studies to assess the radical curative efficacy of primaquine or tafenoquine, but also enrolled a chloroquine-only comparative treatment group (nine studies) or were designed to estimate both schizonticidal and antirelapse efficacy (six studies). In 89 (69%) of 129 studies, patients were recruited from outpatient clinics; however, in 19 (15%) studies, patients were enrolled from hospitals, and in 21 (16%) studies, patients were recruited from cross-sectional surveys. Patients recruited from surveys had lower baseline parasitaemia  than did those recruited from hospital , or outpatient clinics . We assessed 230 treatment groups at 179 study sites, enrolling 21\u2008694 patients. Chloroquine was included in 144 treatment groups , sulfadoxine with pyrimethamine in nine treatment groups , dihydroartemisinin with piperaquine in nine treatment groups , artemether with lumefantrine in seven treatment groups , mefloquine in six treatment groups , and halofantrine in six treatment groups .We identified 198 clinical trials enrolling patients with luded 68 . Of the We noted heterogeneity in the design of the clinical studies . 55 publThe standard dose was a 3 day regimen of chloroquine , although in eight treatment groups (583 patients) the same total dose was spread over four doses, and in five treatment groups (1595 patients) only a single dose of 7\u00b75\u201310 mg/kg of chloroquine was given. In four treatment groups, chloroquine was given with sulfadoxine and pyrimethamine, and in one group given with a 7 day course of doxycycline. Primaquine was given in 73 (51%) chloroquine treatment groups: a very high dose  in two groups, a high dose  in eight groups, a low dose  in 49 groups, and a very low dose  in nine groups.P vivax recurrence by day 28 than did patients receiving chloroquine alone  of 112 site estimates, 35 (61%) of these sites fulfilled the predefined criteria for category 1 resistance, eight (14%) for category 2 resistance, and 14 (25%) for category 3 resistance. The median risk of recurrence in these chloroquine-resistant sites was 16\u00b77% (IQR 9\u00b78\u201331\u00b74). The earliest treatment failure occurred at a median of 14 days (range 3\u201328), and the interval was negatively correlated with both delayed parasite clearance , and the risk of recurrent infections at day 28 . These correlations remained significant after controlling for the place from which patients were recruited .Estimates of chloroquine efficacy could be derived for 177 sites from 144 treatment groups. In 65 (37%) of these sites chloroquine resistance could not be categorised for the following reasons: early administration of primaquine (37 sites); no documentation of parasitological data at day 28 (15 sites); sample size of fewer than ten patients (four sites); good response documented in asymptomatic patients (four sites); and administration of only a single dose of chloroquine (five sites). Overall P vivax was categorised as being chloroquine-resistant . Clearance of parasitaemia (assessed by microscopy) in all patients by day 3, or in 95% of patients by day 2, was 100% predictive of chloroquine sensitivity in the study population as defined by day 28 recurrence.Measures of early parasite clearance were documented in 99 (56%) of 177 site estimates, although the actual proportions of patients who had parasitaemia on days 1, 2, and 3 could be deduced for only 60 (61%) of these site estimates. Sites where esistant  had a siesistant . The difP vivax (P vivax after treatment (seven with blood-chloroquine concentrations exceeding 100 ng/mL at the time of recurrence). 20 cases had confirmed parasite growth in blood chloroquine concentrations exceeding 100 ng/mL . Highly resistant parasites continue to grow despite high blood concentrations of the drug, which results in early treatment failure. Our review identified 20 clinical trials and case reports in which this result was documented . The length of the interval from the start of treatment to parasite recrudescence depends on the pharmacology of the initial treatment regimen, the degree of drug resistance, and the level of host immunity.cumented .P vivax is on the island of New Guinea, where studies have consistently shown high-grade resistance manifested by early clinical deterioration requiring hospitalisation, by delayed parasite clearance, and by early recurrent parasitaemia.P vivax, especially that of severe anaemia in young children.23The epicentre for chloroquine-resistant P vivax clinical trials are done in malaria-endemic regions. Patients might therefore acquire new infections during follow-up. This risk is proportional to the duration of follow-up and the intensity of transmission. Relapses also occur, and again the risk is linked to duration of follow-up, host immunity, and the geographical location of the study. The risk of relapse varies substantially, from 50\u201380% in equatorial regions to 5\u201310% in temperate areas, where the first relapse occurs many months after the initial infection.P vivax drug efficacy. However, genotyping cannot discriminate between a recrudescence of the blood-stage infection or a relapse with a homologous strain. Furthermore, as relapse can also be with a heterologous parasite strain, reliable distinction of the causes of recurrent infection within an endemic area is not possible at present.P vivax therefore are a combination of blood-stage schizonticidal activity and post-treatment prophylaxis suppressing early relapse or reinfection. The unadjusted risk of P vivax recurrence needs to be interpreted in light of the half-life of the treatment regimen, concomitant use of primaquine, the timing of recurrence, available molecular data, schizonticidal drug concentrations, entomological inoculation rates, and the epidemiology of relapse in the study area.Most P vivax clinical studies reported were supported by pharmacokinetic analysis.Recurrent parasitaemia within 28 days of chloroquine treatment does not necessarily imply chloroquine resistance. Sensitive parasites that have been exposed to insufficient treatment because of an incorrect weight-adjusted dose, poor quality drug, poor gastrointestinal absorption, or unusual pharmacokinetics can also recrudesce early. The definitive diagnosis of chloroquine resistance therefore requires documentation of adequate drug exposure at the time of recurrence, to confirm the growth of parasites in concentrations of drug above the minimum inhibitory concentration.P vivax; delayed parasite clearance  and early recurrence were strongly correlated (rs=\u20130\u00b772). Two studies have explored this correlation with individual patient's data. Results of the first, in Papua New Guinea, showed that a P vivax parasite reduction ratio of less than 7\u00b74 predicted subsequent treatment failure with 70% sensitivity and 63% specificity.P vivax recurrence by day 28.P vivax.In falciparum malaria, the speed of parasite clearance correlates with the risk and timing of recrudescence and the infection's transmission potential.P vivax in different endemic settings, and the optimum sampling strategy to define it.A major advantage of parasite clearance rate as a marker for resistance is that it is not confounded by relapse or reinfection. In this Review, clearance of parasitaemia (by microscopy) in all patients by day 3 or in 95% of patients by day 2 was 100% predictive of chloroquine sensitivity in the study population, as defined by the day 28 recurrence rate. Further studies are needed to investigate early parasite clearance for monitoring of P vivax malaria. Primaquine is usually started with the initial dose of chloroquine. Primaquine has activity against both blood and liver stages, including against chloroquine-resistant strains.Most national guidelines recommend a combination of chloroquine for rapid blood-stage activity and a 14 day course of primaquine for the radical cure of the hypnozoite stages in P vivax, the absence of recurrences from such studies should not be held as evidence of drug sensitivity.Drug-resistant plasmodia often occur in areas of high drug pressure and low parasite prevalence. When incidence is low, recruitment of patients to studies might be slow. Some investigators have used active recruitment, seeking out patients with parasitaemia in cross-sectional surveys of target populations. However, patients identified in this manner have significantly lower peripheral parasitaemias compared with those of symptomatic patients presenting to a health-care facility. Since the initial parasitaemia before treatment is a major determinant of therapeutic efficacy, studies enrolling patients through active case detection are likely to underestimate resistance. Individuals with low-grade parasitaemias probably have sufficient immunity to suppress symptomatic disease, thereby reducing the risk of recrudescence even from drug-resistant parasites. Antimalarial efficacy studies should aim to document the clinical response in symptomatic patients. Although a high risk of recurrence in patients recruited from surveys can draw attention to areas of potential drug-resistant P vivax were categorised according to the risk of recurrence at day 28, the 95% CI on this estimate, the documentation of blood drug concentrations, the enrolment of patients with symptomatic disease, and the early co-administration of primaquine. Our definitions were conservative, with data excluded from almost 40% of studies that we thought could be confounded. Almost half of the informative studies suggested evidence of reduced chloroquine efficacy. 15 of these studies and five case reports documented evidence of early treatment failure (Studies of chloroquine efficacy for  failure .P vivax suggests that chloroquine resistance has emerged or spread in many P vivax-endemic countries (P vivax from Thailand,The geographical extent of chloroquine-resistant ountries . These iP vivax endemic countries. However, in practice, clinicians are often reluctant to prescribe primaquine because of fears of haemolysis.P vivax.P falciparum and P vivax has many advantages. The rationale for the use of artemisinin-based combination therapies for all malaria infections is compelling in areas where chloroquine resistance is established.The widespread emergence and spread of chloroquine resistance should cause substantial concern. The combination of chloroquine plus primaquine retains good efficacy in areas with moderate levels of chloroquine resistance, and is the default in most P vivax continues to inflict a substantial burden on the malaria-endemic world with the morbidity and mortality related to its propensity to cause recurrent infections. Frequent recurrences of vivax malaria result from the use of partially effective blood-stage treatment and our poor ability to achieve radical cure safely.P vivax malaria is essential. Enhanced monitoring will require standardised methods and novel approaches for more precise quantification of drug efficacy. A tendency exists to assume that present antimalarial treatment regimens continue to retain efficacy long after evidence of decreasing antimalarial activity has begun to emerge. Inadequate surveillance methods compounded by complacency have delayed the detection and containment of chloroquine-resistant P falciparum, with devastating public health consequences. If a repetition of history is to be avoided, then the threat of chloroquine-resistant P vivax needs to be acknowledged and greater resources applied to develop standardised and validated methods for its detection, characterisation, and containment."}
+{"text": "Plasmodium vivax has recently shown resistance to chloroquine in some places. This study compared the efficacy and safety of fixed dose combination (FDC) of arterolane maleate and piperaquine phosphate (PQP) with chloroquine in the treatment of uncomplicated vivax malaria.Chloroquine has been the treatmentof choice for acute vivax malaria for more than 60\u00a0years. Malaria caused by P. vivax along with fever or fever in the previous 48\u00a0h were included. The 317 eligible patients were randomly assigned to receive FDC of arterolane maleate and PQP (n\u00a0=\u00a0159) or chloroquine (n\u00a0=\u00a0158) for 3\u00a0days. Primaquine was given as an anti-relapse measure on day 3 and continued for 14 consecutive days. Primary efficacy analysis included assessment of the proportion of aparasitaemic and afebrile patients at 72\u00a0h. Safety endpoints were analysis of adverse events, vital signs, laboratory data, and abnormalities on electrocardiograph. Patients participated in the study for at least 42\u00a0days.Patients aged 13\u201365\u00a0years with confirmed mono-infection of 1/2 of 3.2\u00a0h. Piperaquine is also well absorbed after oral administration with a t1/2 of about 228.33\u00a0h.In per protocol population, the proportion of aparasitaemic and afebrile patients at 72\u00a0h was 100\u00a0% (140/140) in the FDC of arterolane maleate and PQP group, and 99.3\u00a0% (145/146) in the chloroquine group . In intent to treat population, the corresponding value was reported to be 96.9\u00a0% (154/159) in the FDC of arterolane maleate and PQP group and 98.7\u00a0% (156/158) in the chloroquine group . The median parasite clearance time was 24\u00a0h in FDC of arterolane maleate and PQP group and 26\u00a0h in chloroquine group . Similarly, median fever clearance time was 24\u00a0h in both the groups . In PP population, day 28 cure rates were 100\u00a0% in both the groups ). Incidence of adverse events was 82.4\u00a0% in the FDC of arterolane maleate and PQP group and 85.4\u00a0% in the chloroquine group. Most of the adverse events were mild to moderate in intensity. The commonly reported clinical adverse events in the FDC of arterolane maleate and PQP versus chloroquine group were vomiting (5.0 vs 5.1\u00a0%), headache (1.3 vs 3.2\u00a0%) and prolonged QT (1.9 vs 3.2\u00a0%). No deaths were reported. The pharmacokinetic analysis indicates that arterolane maleate is well absorbed and has a relatively short tThe study showed that FDC of arterolane maleate and PQP effectively cured vivax malaria and attained acceptable level of cure up to day 28. Both the groups showed similar safety profile.Trial Registration Clinical Trial Registry India: CTRI/2011/11/002129The online version of this article (doi:10.1186/s12936-016-1084-1) contains supplementary material, which is available to authorized users. Plasmodium vivax, the second most important species causing human malaria, accounts for about 40\u00a0% of malaria cases worldwide. It is prevalent in endemic areas in the Middle East, Asia, Oceania, Central and South America. In most areas where P. vivax is prevalent, malaria transmission rates are low, and the affected populations, therefore, achieve little immunity to this parasite. Consequently, people of all ages are at risk  patients. Secondary efficacy endpoint analysis . Whereas, the results were comparable in ITT population with a total of 109  patients out of 159 were considered to be cured on day 28 after administration of FDC of arterolane maleate and PQP and 115  patients out of 158 were considered to be cured on day 28 after receiving chloroquine (Table\u00a0The median PCT was estimated to be 24\u00a0h (18.0\u201331.0\u00a0h) in FDC of arterolane maleate and PQP group whereas PCT of 26\u00a0h (18.0\u201336.0\u00a0h) was noted in chloroquine group . Both the study treatments demonstrated identical PCT   and a mean t1/2 of 3.20\u00a0h (ranging from 0.48 to 8.74\u00a0h) was observed for arterolane. Similarly, the mean Cmax and mean t1/2 for piperaquine were 317.82\u00a0ng/mL (ranging from 89.13 to 817.14) and 228.33\u00a0h (ranging from 10.26 to 757.03\u00a0h), respectively.A mean Cmax of 464.17\u00a0ng/mL (ranging from 181.57 to 2467.27\u00a0ng/mL) and mean t1/2 of 172.47\u00a0h (ranging from 12.92 to 757.32\u00a0h), while desethylchloroquine exhibited a mean Cmax of 146.33\u00a0ng/mL (ranging from 51.70 to 790.32\u00a0ng/mL) and a mean t1/2 of 209.06\u00a0h (ranging from 17.32 to 525.77\u00a0h).Chloroquine showed a mean CMean pharmacokinetic parameters along with descriptive statistics for FDC of arterolane maleate and PQP and chloroquine are summarized in Table\u00a0max of arterolane and chloroquine is shown in Figs.\u00a0max of arterolane and chloroquine. Exposure (AUC last) of arterolane and chloroquine was also plotted against PCT (hours). From the plot, it appears that arterolane exposure levels (AUC last) of less than ~2.5\u00a0\u00b5g.h/ml  of less than ~80\u00a0\u00b5g.h/ml  in FDC of arterolane maleate and PQP and 85.4\u00a0% (135/158) in chloroquine group  from FDC of arterolane maleate and PQP group and five patients (3.2\u00a0%) from chloroquine group. The mean QTc interval after Fridericia\u2019s correction was 395.3\u00a0ms on day 0 and 401.1\u00a0ms on day 2 after treatment with FDC of arterolane maleate and PQP, whereas, the mean QTc interval after Fridericia\u2019s correction was 395.7\u00a0ms on day 0 and 406.3\u00a0ms on day 2 after treatment with chloroquine . All events of prolongation of QTc interval were judged by the investigators to be mild to moderate in severity. No event was judged to be clinically significant. There were no instances of Torsade de Pointes, sudden death, ventricular tachycardia, ventricular fibrillation and flutter, syncope or seizure in any of the patients which could be identified as a direct outcome of significant prolongation of QTc interval.Amongst the laboratory parameters, concentration of haemoglobin, platelet count, reticulocyte count, eosinophil count, serum potassium and sodium, ALT and blood glucose values reported on day 0, 3, 28 and 42 were evaluated. Mean concentration/counts were similar after the treatment with arterolane maleate and PQP or chloroquine at these time points.Plasmodium vivax is the dominant species of malaria in many areas outside Africa [P. falciparum and P. vivax to chloroquine, amodiaquine and sulfadoxine-pyrimethamine, only limited treatment options are available [e Africa . Due to vailable .In this study, 100\u00a0% of patients treated with FDC of arterolane maleate and PQP were aparasitaemic and afebrile at day 3 compared to 99.3\u00a0% of patients on chloroquine treatment. The efficacy rate of more than 95\u00a0% was reported at 72\u00a0h in the both treatment groups.P. vivax in many places, has been adopted as the first-line treatment for falciparum malaria, may also be used for vivax malaria. Also, in areas with chloroquine-resistant P. vivax, ACT  is recommended for the treatment of vivax malaria [It is recommended by WHO that in areas where ACT, except artesunate plus sulfadoxine-pyrimethamine which is not effective against  malaria . The art malaria .Arterolane maleate is the short-acting component in the FDC of arterolane maleate and PQP, which contributes significantly in achieving early parasite clearance. The time to parasite clearance and fever clearance seen with FDC of arterolane maleate and PQP in this study are on expected lines since the clinical efficacy of the artemisinin derivatives is characterized by an almost immediate onset of activity and rapid reduction of parasitaemia, with complete clearance in most cases within 48\u00a0h . Median Day 28 cure rates were 100\u00a0% in PP population in both the treatment groups. In ITT population analysis cure rates were reported to be close to 70\u00a0% because most of the patients were lost to follow-up, which was considered as failure in the statistical analysis. The possible reason for loss of patients to follow up was migration as the study was conducted in urban settings.P. vivax infections would establish a common treatment for both circulating Plasmodium species, P. falciparum and P. vivax. FDC of arterolane maleate and PQP is highly effective against P. falciparum based on previously conducted studies [P. falciparum infections being wrongly diagnosed as P. vivax, leading to inappropriate treatment with chloroquine. Treating P. falciparum with FDC of arterolane maleate and PQP and primaquine would have the advantage of also treating potential dormant P. vivax hypnozoites, which are thought to become activated due to P. falciparum infections, thereby explaining the high number of vivax infections after P. falciparum treatment [Effectiveness of FDC of arterolane maleate and PQP for  studies , 23. Repreatment .In this study, no deaths were reported and all the four serious adverse events were mild to moderate in intensity and recovered without sequelae. As these patients were hospitalized, the adverse events were considered to be serious in nature. Overall, the incidence of adverse events was similar to incidence reported in patients treated with PQP combinations with DHA in various other studies , 13.The proportion of patients with QTc prolongation of >60\u00a0ms from baseline in patients treated with FDC of arterolane maleate and PQP were 1.9\u00a0% compared to 3.2\u00a0% in chloroquine group. Evaluation of QT interval changes is problematic in malaria because of systematic differences between the acute febrile admission before anti-malarial drugs are given and early convalescence. At presentation, patients are usually anxious, fasting and febrile with increased autonomic tone and a raised heart rate. This is in contrast to the relaxed, fed, supine, afebrile state 3\u00a0days later when most anti-malarial treatments finish and anti-malarial concentrations are at the highest. It has been argued that this systematic reduction in sympathetic activity with recovery leads to a consistent increase in the QT interval which has been mistakenly ascribed to anti-malarial drug effects. It has been reported in literature that QTc prolongation of >60\u00a0ms is within the bounds of normal daily variation in the QTc (up to 75\u2013100\u00a0ms) , 26.Amongst the laboratory parameters, concentration of haemoglobin, platelet count, reticulocyte count, eosinophil count, serum potassium and sodium, ALT and blood glucose were evaluated. Mean concentration/counts reported during laboratory assessments were similar after treatment with FDC of arterolane maleate and PQP or chloroquine. By proportion, the incidence of increased transaminase (ALT and AST) was similar in both the treatment groups. This observation combined with the fact that the FDC of arterolane maleate and PQP is only administered once daily for 3\u00a0days suggests that there is a very low risk of liver injury with FDC of arterolane maleate and PQP.max and AUC for chloroquine and desethylchloroquine corroborated well with literature-based information [Pharmacokinetic results through non-compartmental analysis indicate that the mean rate and extent of absorption for arterolane maleate and PQP were apparently similar to those observed in patients treated for falciparum malaria. Similarly, the Cormation . PCT forP. vivax. Once-a-day treatment with three doses of FDC of arterolane maleate and PQP efficacy and safety profile similar to chloroquine and no reappearance of vivax malaria noted until day 42 makes this combination a promising candidate, which could lead to better treatment compliance and patient adherence.Arterolane  in combination with PQP exhibits a rapid onset of action, potent activity against all erythrocytic stages of The findings from this study show that FDC of arterolane maleate and PQP effectively cure vivax malaria and attain acceptable level of cure up to day 28. Both the groups have shown similar safety profile."}
+{"text": "The World Health Organization recommends that regular efficacy monitoring should be undertaken by all malaria endemic countries that have deployed artemisinin combination therapy (ACT). Although ACT is still efficacious for treatment of uncomplicated malaria, artemisinin resistance has been reported in South East Asia suggesting that surveillance needs to be intensified by all malaria endemic countries. This study assessed the efficacy and safety of artemether-lumefantrine (AL) for the treatment of uncomplicated falciparum malaria in Muheza district of north-eastern Tanzania, an area where the transmission has significantly declined in recent years.Eighty eight children (aged 6 months to 10 years) with uncomplicated falciparum malaria were recruited into the study. The patients were treated with standard doses of AL and followed up for 28 days. The primary end point was parasitological cure on day 28 while the secondary end points included: improvement in haemoglobin levels and occurrence, and severity of adverse events.A total of 163 febrile patients were screened, out of which 88 patients (56 under-fives and 32 aged \u22655 years) were enrolled and 79 (89.8%) completed the 28 days of follow-up. There were no cases of early treatment failure whilst 40 (78.4%) under-fives and 21(75.0%) older children had adequate clinical and parasitological response (ACPR) before PCR correction. Late clinical failure was seen in 5.6% (n\u2009=\u200951) and 3.6% (n\u2009=\u200928) of the under-fives and older children respectively; while 15.7% and 21.6% had late parasitological failure in the two groups respectively. After PCR correction, ACPR was 100% in both groups. Reported adverse events included cough (49.7%), fever (20.2%), abdominal pain (10.1%), diarrhoea (1.3%), headache (1.3%) and skin rashes (1.3%).This study showed that AL was safe, well-tolerated and efficacious for treatment of uncomplicated falciparum malaria. Since Muheza has historically been a hotspot of drug resistance , surveillance needs to be continued to detect future changes in parasite sensitivity to ACT. Despite the current reported declining trends, malaria has remained an important public health problem and the main cause of morbidity and mortality in Tanzania and elsewhere in sub-Saharan Africa (SSA). Globally, an estimated 3.4 billion people were at risk of malaria and there were an estimated 627, 000 malaria deaths in 2012. Of the reported deaths, 90% occurred in SSA and majority involved children less than five years of age), a fixed combination of 20 mg of artemether and 120 mg lumefantrine in a tablet. The drugs were administered according to the recommended doses based on the weight of patient. One tablet was given to children weighing 5-14 kg; two tablets to children with 15\u201324 kg and three tablets to children weighing 25\u201335 kg. A full course of AL consisted of six doses given twice daily (8 hourly apart for day 0 and 12 hourly apart for days 1 and 2). Patients were observed for 30 minutes to ensure that they did not vomit the study drugs. When vomiting occurred, a repeat dose was given after vomiting stopped. Any patient who persistently vomited the study medication, treatment was discontinued and such patient was withdrawn from the study, and treated with parenteral quinine according to the national guidelines for management of complicated and severe malaria. Paracetamol was given to all patients with body temperature greater than or equal to 38\u00b0C. All AL doses were administered orally under direct observation of a study nurse.Patients enrolled in the study were treated with AL  when patient felt unwell. Parents/guardians were informed and encouraged to bring the children to the clinic whenever they were unwell without waiting for their scheduled visits. Patients who could not come for their scheduled visit by mid day were visited at home by a member of the study team and asked to come to the health centre. Patients who travelled to other places and could not be traced for scheduled follow-up were withdrawn from the study. During the visits, both clinical and parasitological assessments were performed; and follow-up samples  were also collected.Human white blood cells (WBCs) were depleted using CF11 cellulose columns and paraThe primary end point was parasitological cure on day 28 as per WHO protocol of 2009 and secoEthical clearance for this study was obtained from the National Medical Research Coordination Committee of the National Institute for Medical Research (NIMR-MRCC). Permission to conduct the study in Muheza was sought in writing from the relevant regional and district medical authorities. Oral and written informed consent was obtained from parents or guardians of all patients before they were screened for possible inclusion into the study.The data was entered (by double entry) into a Microsoft Access database followed by validation and cleaning. Data analysis was done using STATA for Windows, version 11 . Descriptive statistics as percentages, mean, median, standard deviation, and range were applied as appropriate. Treatment outcome was analysed based on per protocol method and Kaplan\u2013Meier analysis. Patients were stratified into two groups of children aged \u22655 years and under-fives; and baseline characteristics, primary and secondary outcomes were compared between the two groups. Continuous variables such as parasite density, age and haemoglobin concentrations between the two groups were compared using t-test  or Mann\u2013Whitney U test . For all statistical tests, p-value was set at 0.05.P. falciparum mono-infections. Of the P. falciparum mono-infected patients, one had parasite density below 250 asexual parasites per microlitre and was therefore not enrolled while 88 patients fulfilled the inclusion criteria and were enrolled into the study. During follow-up, a total of nine participants were lost to follow-up as they travelled out of the study area, leaving 79 patients for further analysis of treatment outcome  patients had malaria parasites by microscopy and 89 (54.6%) had 2\u2009=\u20095.47, p\u2009=\u20090.019). The overall mean body temperature at enrolment was 38.3\u2009\u00b1\u20091.3\u00b0C while the mean haemoglobin level was 10.4\u2009\u00b1\u20091.8 g/dl. The parasite density ranged from 256 to 200,000 with a geometric mean of 18,551 asexual parasites/\u03bcl. All baseline characteristics (except sex and age) were similar in the two age groups (TableForty six (52%) of the enrolled patients were males and there were significantly more males among children aged \u22655 years compared to under-fives (\u03a7There were no cases of ETF while LCF was observed in three (5.6%) and one (3.6%) cases among under-fives and those aged \u22655 years, respectively Table. Eight .Table 2A total of 68 77.3%) patients had malaria parasites on day 1 post-treatment while 17(19.5%) had parasites on day 2 and only one patient (1.4%) had parasites on day 3. Twenty three (26.1%) and 2 (2.3%) patients had fever on day 1 and 2 respectively. Two patients (2.3%) still had fever on day 3 post-treatment although the axillary temperature was low (\u226437.9\u00b0C). There was no significant difference in parasite and fever clearance between under-fives and children aged\u2009\u2265\u20095 years  in both under-fives and children aged \u22655 years. The haematological recovery in children aged \u22655 years was statistically significant (p\u2009=\u20090.006) while Hb recovery in under-fives was not statistically significant (p\u2009=\u20090. 305)  to day 28  reported were cough (49.4%), fever (20.2%), abdominal pain (10.1%), diarrhoea 1.3%), headache (1.3%) and skin rashes (1.3%). However, most of the AEs occurred in under-fives compared to older children aged \u22655 years (only 6% of the under-fives had no AEs compared to about 36% of the older children). There were no serious adverse events and all the AEs were not related to the medications  with SP. BecauseP. falciparum malaria with rapid fever and parasite clearance. Although the clinical efficacy was low , PCR corrected outcome was higher in all children irrespective of age group (ACPR\u2009=\u2009100%). These findings are consistent with other AL therapeutic efficacy studies conducted in Tanzania and elsewhere in SSA where PCR corrected ACPR was 96 - 100%[et al., unpublished data). In the current study, a total of 88 children were recruited in six weeks only, suggesting that local heterogeneity and micro-epidemiological factors could be the key drivers of malaria transmission in Muheza.The present study showed that the standard six-dose of AL was efficacious for treatment of uncomplicated 6 - 100%\u201336. The 6 - 100% and thisRapid parasite clearance was observed whereby only one patient (1.1%) had parasitaemia on day 3 post-treatment. Studies conducted elsewhere showed similar findings whereby <1.1% the children had parasite on day three post-treatment. ArtemetThe mean temperature of children recruited in the study was high  and consistent with the reports of studies conducted elsewhere in Tanzania and EthiThe progressive haematological recovery post-treatment suggests that malaria could be the major contributing factors to the low haemoglobin levels at recruitment especially in older children. Insignificant haematological recovery irrespective of parasite clearance in the younger children suggests that other factors, such as geohelminths, may also play a key role in the chronicity of anaemia as reported in other studies.In the present study, adverse events were reported in 66 (75%) of the recruited children but they were mild and not associated with the treatment. Cough was the most frequent adverse event in both age groups, which is in line with previous studies which showed that respiratory infections are common in African children with malaria. SimilarDue to declining malaria transmission in Muheza district and other parts of Tanzania, 25, theHowever, there were more female children among under-fives compared to older children aged \u22655 years. The difference in the proportion of female and male in the present study may simply be related to the difference in health-seeking behaviour rather than actual difference in disease prevalence as it has been reported elsewhere. AlthougP. falciparum malaria. With the recent reports of declining trends in malaria prevalence in most parts of Tanzania including north-eastern Tanzania, anti-malarial drug pressure in the community has decreased and this may possibly limit the occurrence and spread of parasite tolerance/resistance to ACT. Despite such promising reports, as the world strives to achieve malaria elimination, further studies are needed to monitor efficacy of AL and other ACT in order to provide evidence for timely review of malaria treatment policy in the country. Since Muheza has historically been a hotspot of drug resistance , surveillance needs to be continued at this and other sites in the country to detect future changes in parasite sensitivity to ACT.The efficacy of AL for treatment of uncomplicated malaria was still high in the study area despite higher rates of re-infection. Thus, despite the reported cases of late clinical failure, these findings showed that AL was safe, well tolerated and effective for treatment of uncomplicated"}
+{"text": "Plasmodium falciparum malaria. In endemic regions, it has proven more effective in treating the disease, and even in reducing its transmission. Nonetheless, there is a scarcity of studies carried out in non-endemic areas on imported uncomplicated malaria.Artemisinin-based combination therapy (ACT) has been adopted by the World Health Organization as a first-line treatment for uncomplicated P. falciparum malaria between 2004 and 2015. The objective was to compare the parasite clearance period and the average hospital length of stay for patients treated with ACT vs those receiving other treatment regimens.This is a retrospective, observational study performed on patients diagnosed and admitted with uncomplicated Eighty-five patients were included in the study. Fifty-one received ACT treatment (dihydroartemisinin\u2013piperaquine) and thirty-four patients were treated with quinine sulfate+doxycycline or atovaquone/proguanil. The parasite clearance period was shorter in the group of patients treated with ACT compared to those receiving other treatment types: 24\u00a0h (IQR 24) vs 48\u00a0h (IQR 48), p\u00a0<\u00a00.01. The average hospital stay was also shorter in the ACT group with respect to the second group: 2.67\u00a0days (IQR 1.08) vs 3.96\u00a0days (IQR 2.87), p\u00a0<\u00a00.001. A mild case of hepatitis was registered in the group treated with ACT.P. falciparum reduced the days spent hospitalized as well as producing a more rapid parasite clearance compared to classic treatment. In spite of being treated with safe medications, one has to be alert to possible adverse effects such as hepatitis and delayed haemolytic anaemia.ACT treatment of admitted hospital patients with imported uncomplicated malaria from The online version of this article (doi:10.1186/s12936-016-1408-1) contains supplementary material, which is available to authorized users. P. falciparum [Malaria is the most important parasitic disease worldwide. It has been calculated that each year is it responsible for new episodes in around 214 million patients, causing the death of approximately 438,000 the majority of which are children under 5\u00a0years of age and pregnant women . Most imP. falciparum malaria treatment, whether complicated (severe) or uncomplicated, both in endemic and non-endemic countries [Malaria treatment has changed substantially over recent years . Artesunountries \u201320.P. falciparum, there are few studies comparing the differences in non-endemic regions [Artemisinin treatment leads to a reduction in average hospital stays, more rapid parasite clearance in comparison with any other anti-malarial , 12, 17;P. falciparum to a hospital in a non-endemic area.This study aims to analyse the impact that the use of ACT has had compared to two classic treatments (quinine sulfate and doxycycline/clindamycin or atovaquone-proguanil) to treat patients admitted with uncomplicated malaria from P. falciparum [\u00ae) using lateral flow assays. The malarial PCR (semi-nested multiplex PCR) was used only in cases where a more precise confirmation was needed of the species, or where there was a suspicion of mixed parasitic infection. A clinical history and complete physical examination were carried out on all patients, gathering epidemiological and clinical data. On admission, screening was performed for the most prevalent imported pathologies; this included HIV, HBV\u00a0and\u00a0HCV\u00a0blood tests. Parasitaemia monitoring was carried out via a daily smear until a negative result was obtained. After leaving hospital, the patients were checked with a blood test and a blood smear at 7 and 28\u00a0days.A retrospective observational study was carried out on all patients admitted with malaria to Hospital Poniente  from January 2004 to December 2015. The hospital serves a population of approximately 250,000 people, in an area with a large African immigrant population, many of whom work in horticultural greenhouses. The study included patients over 14\u00a0years of age whose reason for hospital admission was exclusively the diagnosis of uncomplicated malaria from lciparum . Pregnan\u00ae 320\u00a0mg/40\u00a0mg, Sigma Tau) was the ACT chosen; this medication is currently the commercial form available in Spain.Uncomplicated malaria is defined following the recommendations for the management of imported malaria in Europe . With reVariables were analysed corresponding to epidemiological, clinical and analytical characteristics as well as the parasite clearance period and the average hospital stay. The level of parasitaemia was divided into patients with\u00a0<1\u00a0%, between 1 and 2\u00a0%, and\u00a0>2\u20135\u00a0%. Any possible adverse secondary effects associated with the treatment were also recorded.Data were analysed using the statistical software package SPSS v17. A descriptive analysis was carried out on the quantitative variables using the mean and the standard deviation, as well as the range. Qualitative variables were described using absolute frequencies and percentages. To observe the possible differences between the group of patients treated with quinine and doxycycline/clindamycin or atovaquone\u2013proguanil and ACT, the Pearson\u2019s Chi square test or Fisher\u2019s exact test was used when the variables were qualitative. When the variables for comparison were quantitative, the mean differences were estimated using the Student\u2019s t test for independent samples if they complied with the normal hypothesis and the Mann\u2013Whitney U test when they did not.A multivariate linear regression model was performed using days of hospitalization as the dependent variable and the treatment group, the haemoglobin (Hb) and platelet levels (at the time of diagnosis) and the level of parasitaemia as the independent variables. The goodness-of-fit was calculated along with the determination coefficient. For all tests, the fixed level of significance was 0.05. This study was approved by the local ethic committee  with code 12/2016.A total of 85 patients diagnosed with uncomplicated malaria were included in the study. The pharmaceuticals used for malaria treatment were quinine sulfate+doxycycline or atovaquone/proguanil until 2012. In the same year, dihydroartemisinin\u2013piperaquine was introduced, remaining as the first-line treatment since then. Fifty-one patients were treated with dihydroartemisinin\u2013piperaquine and 34 with quinine sulfate+doxycycline (n\u00a0=\u00a033) or atovaquone/proguanil (n\u00a0=\u00a01). The dataset supporting the conclusions of this study is included as Additional file The general patient characteristics, both together and by groups, are shown in Table\u00a0The global parasite clearance time was 24\u00a0h (IQR 24) and the average hospital stay was 2.88\u00a0days IQR 1.88). In the subgroup analysis, the parasite clearance time was significantly less in the patient group treated with ACT rather than the classic treatment: 24\u00a0h (IQR 24) vs 48\u00a0h (IQR 48), p\u00a0<\u00a00.01. The average stay was also less in the artemisinin group with respect to the group treated with quinine+doxycycline/atovaquone-proguanil: 2.67\u00a0days (IQR 1.08) vs 3.96\u00a0days (IQR 2.87), p\u00a0<\u00a00.001  vs 12.65\u00a0g/dL\u00a0\u00b1\u00a01.42 (range 10\u201315.5), p\u00a0=\u00a00.03; day 28: 14.32\u00a0g/dL\u00a0\u00b1\u00a01.2 (range 11.7\u201316.8) vs 13.45\u00a0g/dL\u00a0\u00b1\u00a01.38 (range 9.1\u201314.8), p\u00a0=\u00a00.02.In the lineal regression model Table\u00a0, the varWith regard to any reported adverse effects, in the group treated with dihydroartemisinin\u2013piperaquine, there was one mild case of hepatitis, in the only non-sub-Saharan patient; this was self-limiting but, nevertheless, led to more days spent in hospital (6\u00a0days).ACT is a very effective and safe treatment in the management of imported uncomplicated malaria. Besides producing more rapid parasite clearance, its use in patients admitted with uncomplicated malaria reduces the period of hospitalization compared to classic treatments that were used previously, resulting in healthcare savings.P. falciparum is the species most frequently involved, and also the one which most results in morbidity and/or mortality in travellers (tourists and expatriates), as well as in foreigners who return home to visit friends and family [The management of imported malaria is still a challenge for many healthcare professionals in non-endemic zones. d family \u20135. The ud family \u201320. Quind family , 18\u201320.Most studies concerning imported malaria have been carried out on patients with complicated malaria , 12 withP. falciparum, the general recommendation is hospital admittance and oral treatment for at least 24\u201348\u00a0h [In the case of uncomplicated malaria caused by  24\u201348\u00a0h , with th 24\u201348\u00a0h  conclude 24\u201348\u00a0h . There w 24\u201348\u00a0h , 22. In The overall average hospital stay for patients in our study was 2.88\u00a0days. The fact that most of the patients are immigrants has probably contributed to a slightly longer stay to that recommended. The language barrier that exists with many of these patients, as well as the problems concerning frequent follow-up visits to the healthcare centres, often for economic or work-related reasons, means that it is common their doctors feel more comfortable keeping them admitted to hospital until they are sure of good treatment tolerance and response. The rapid parasite clearance and the markedly shorter treatment regimen using ACT are probably the factors that most influence the length of stay.P. falciparum [Plasmodium resistance.Atovaquone\u2013proguanil is another medication recommended as a first-line treatment for uncomplicated malaria from lciparum \u201320; howelciparum . FurtherWith regard to possible adverse reactions, the compounds derived from artemisinin have shown good tolerance and have a good safety profile , 24. TheThe most severe adverse effect of artemisinin derivatives however is delayed haemolytic anaemia \u201328. ThisWith regard to pregnancy, there is insufficient information about the safety and efficacy of many anti-malarials, especially in the first trimester. Consequently, the only drugs considered safe are quinine, chloroquine, clindamycin and proguanil. Artemisinin-derived drugs are safe and efficacious in the second and third trimesters, in that they have been demonstrated not to be teratogenic , 30.The limitations present in this study come from the fact that it is a retrospective work.Furthermore, given the characteristics of the population that is the object of the study, the results may only be strictly extrapolated to African immigrants treated with two treatment regimens, one which has already passed to second-line use (quinine sulfate\u2013doxycycline) and the other, an ACT (dihydroartemisinin\u2013piperaquine), which is a first-line treatment. New prospective studies will need to be carried out to evaluate if any differences exist in terms of safety, efficacy and healthcare costs between the ACT-based treatments and, for example, atovaquone\u2013proguanil, which is also a fixed combination over the same period . Moreover, with atovaquone\u2013proguanil, ample experience has been gathered in non-endemic regions and it has a proven safety record.The results of this work reinforce the results of the few studies published to date regarding ACT treatments for patients with uncomplicated malaria in non-endemic regions. They are drugs with a good safety record, clearing parasitaemia more rapidly than other anti-malarials, and are probably more cost effective by reducing the average length of stay in hospital. Nonetheless, despite being safe drugs, one should be alert to the possibility of adverse effects such as hepatitis and delayed haemolytic anaemia."}
+{"text": "Standard treatment of cutaneous leishmaniasis (CL) in Suriname entails three injections of pentamidine isethionate (PI) 4 mg/kg per injection in 7 days (7 day regimen). Compliance to treatment is low and may contribute to increasing therapy failure. A 3 day regimen, including 2 injections of 7 mg/kg in 3 days may increase compliance.In a randomized, single-blinded non-inferiority trial conducted in Suriname, 84 CL patients received the 7 day regimen and 79 CL patients received the 3 day regimen. Primary objective was the proportion of patients clinically cured at 6 weeks follow-up. Secondary objectives were clinical cure at 12 weeks follow-up; parasitological cure at 6 and 12 weeks; adverse and drug related toxicity events recorded one week after the end of treatment and health related quality of life. The non-inferiority margin was set at 15%, 1 sided test, \u03b1 = 0.1.At 6 weeks follow-up 31 (39%) patients in the 3 day regimen and 41 (49%) patients in the 7 day regimen were clinically cured. Intention to treat (ITT) analyses showed that the difference in proportion clinically cured was -9.6% : -22.3% to 3.2%). Per protocol (PP) analysis showed that the difference in proportion clinically cured was 0.2% (90% CI: -14.6% to 15.2%). ITT analysis showed that the difference in proportion parasitological cured at 6 weeks was -15.2% (90% CI:-28.0% to -2.5%). PP analyses showed similar results. Non-inferiority could not be concluded for all adverse and toxicological events.We cannot conclude that the 3 day regimen is non-inferior to the 7 day regimen regarding proportion clinically and parasitological cured. Therefore there is no evidence to change the current standard practice of the 7 day regimen for the treatment of CL in Suriname. Leishmania, is treated in Suriname with pentamidine isethionate (PI). Standard treatment consists of 3 intramuscular injections of 4 mg per kg bodyweight given in 7 days. Compliance to treatment is low which may lead to treatment failure. A shorter treatment course of PI, 3 days, may aid in solving the compliance problem. In this study CL patients were included in two study arms, the control arm, PI 4 mg/kg/injection, three injections in 7 days (84 patients) and the intervention arm, PI 7 mg/kg/injection, two injections in 3 days (79 patients). Based on the study results it cannot be concluded that that the 3 day regimen is at least as good (non-inferior) to the 7 day regimen, as far as clinical and parasitological cure, toxic and adverse events. Based on these findings there is no evidence to change the standard treatment of 3 injections of 4 mg/kg PI in Suriname.Cutaneous Leishmaniasis (CL), caused by the single cellular protozoan parasite of the genus  Phlebotomus and Lutzomyia transmit the protozoan Leishmania parasites.[With a global yearly incidence around 1.5 million cases, Cutaneous leishmaniasis (CL) is the most common clinical presentation of Leishmania infection.[arasites.L. (V.) guyanensis. Pentamidine isethionate (PI) is the preferred treatment.[In Suriname CL is endemic and, mainly caused by reatment. The stanVan der Meide et al. found that only 50% of the patients received the complete three injections therapy. Many patA shorter treatment regimen might improve compliance based on our beliefs that noncompliance is caused by the costs involved in seeking help for CL and the consequential income loss. A treatment regime of either one or two intramuscular injections with 7 mg/kg PI salt in 3 days, evaluated in a retrospective French Guyanese study, indicated similar cure rates for the non-severe cases. Lost to In previous studies the 7 mg/kg PI salt regimens have never been evaluated as opposed to the 4 mg/kg regimen in controlled trial settings. Therefore, we have conducted a randomized single-blinded non-inferiority trial to assess if a 3 day injection regimen consisting of 7 mg/kg PI is not worse (non-inferior) with respect to clinical and parasitological cure rate, adverse events and health related quality of life (HRQL) compared to the standard 7 day PI regimen for the treatment of CL patients visiting the dermatological outpatient clinic in Paramaribo, Suriname.This study was approved by the Medical Ethical Commission of the Ministry of Health Suriname (VG 006-2009). Written informed consent has been obtained from all participants.rd 2010\u2014April 30th 2013 at the outpatient clinic of the Dermatology Service in Paramaribo. The study was approved by the ethical review commission of the Suriname Ministry of Health and registered at the Dutch Trial Register (NTR 2076).We conducted a randomized single-blinded non-inferiority trial from January 3Eligible individuals were \u2265 16 years with laboratory confirmed CL (histopathology and/or Giemsa smear of biopsy) who could be contacted by phone. Exclusion criteria were CL patients treated in the previous six months, pregnancy or lactation, unable to attend one of the study visits, medical history of diabetes mellitus, cardiac-, renal- and hepatic disease, abnormal baseline values for amylase, aspartate aminotransferase and alanine aminotransferase, creatinine, glucose, hemoglobin, leucocytes, thrombocytes and patients with known allergy to PI.After written informed consent was provided, patients were randomized for the intervention regime of two intramuscular injections of 7 mg/kg PI salt on days 1 and 3 (3 day regimen) or the control regime of three intramuscular injections of 4 mg/kg on days 1, 4 and 7 (7 day regimen). Injections were administered by the dermatologist aware of the intervention allocation. Randomization was performed by a computerized balanced block randomization scheme that was stratified on disease severity based upon the presence or absence of clinical loco regional lymphadenitis.The primary objective was to establish if the 3 day regimen is non-inferior to the 7 day regimen regarding the primary endpoint clinical cure six weeks after end of treatment.The secondary endpoints were clinical cure at 12 weeks, parasitological cure at 6 and 12 weeks, adverse and drug related toxicity events one week after the end of treatment and HRLQ differences before treatment and at 6 weeks follow-up visit.Clinical evaluation was conducted at enrollment, during the treatment visits and during the follow-up visits 1, 6 and 12 weeks after treatment. Lesions were measured, recorded and photographed. If more than one lesion existed, the largest lesion was followed up. Lymph tracks and regional lymph node stations were palpated. Questionnaires for HRQL . Therapy failure was observed in case of incomplete re-epithelization and/or inflammatory signs. Two independent blinded dermatologists with at least four years of diagnostic CL experience determined clinical cure (using standardized photographs of lesions). In case of disagreement a third blinded dermatologist passed the final judgment.Parasitological cure was defined as a log3 decrease in parasite load in the follow up lesion at visits 6 and 12 weeks compared to baseline. The SKINDEX-29 questionnaire was used to measure the HRLQ. With theWith a structured questionnaire, patient reported adverse events were identified throughout treatment and follow-up. Drug related toxicity; drop in diastolic blood pressure > 10mm Hg measured during and after injection, and abnormalities in laboratory values were recorded.Seventy patients per group were required to be 80% sure that the lower limit of a 90% two sided confidence interval will exclude a difference in favor of the standard 3 day regimen of more than 15% (non-inferiority margin), assuming a 85% clinical cure for both groups. The 15% Differences in proportions were presented with the 90% Confidence Interval (CI). The 3 day regimen was considered non-inferior if the left-side of the 90% CI did not exceed the non-inferior margin of minus 15% except for the difference in proportions of adverse and toxicological events were the right-side of the 90% CI should not exceed the 15%.2 tests were appropriate.Data were analyzed as intention to treat (ITT) and per protocol (PP). For ITT analysis, subjects withdrawn from the study, lost to follow-up, or not attending the evaluation visit, were either considered therapeutic failures or, having an adverse event or abnormal clinical laboratory value and they were excluded from the PP analysis. Furthermore, patients not receiving additional treatment at follow up visit 6 according to protocol were excluded from the PP analyses at follow up visit 12 weeks. The safety data set included individuals who had received at least one injection. To estimate if disease-specific HRLQ scores changed before and six weeks after intervention and to see if the change in HRLQ over time was different between the two groups, a generalized estimation equation with independent correlation structure and a robust estimation of the standard errors were conducted. Age and gender were included in this model. The data set included only those individuals without missing data. HRLQ between groups was compared using the Mann Whitney test and \u03c7Except for HRLQ, we conducted sensitivity analyses based on the ITT population in which it was assumed that individuals without data on the outcome of interest were considered to be either cured  or not having any adverse events Statistical analyses were performed using STATA SE12.1.rd 2010 to April 30th 2013, 185 patients with suspected CL were assessed for eligibility. A total of 163 eligible patients were randomized   and in tThe proportion parasitological cured was 44.3% in the 3 day regimen compared to 59.5% in the 7 day regimen; a difference of minus 15.2% (90% CI:-28.0% to -2.5%) in the ITT analysis and minus 6.4% (90% CI: -20.3% to 7.5%) in the PP analysis. For the ITT population, not only the left side of the 90% CI exceeded the non-inferior margin, also the right side of the 90% CI was lower than 0% (-2.5%) and even the right side of the 95% CI was slightly less than 0% (-0.01%) indicating that the 3 day regimen was not only non-inferior but could even be worse compared to the 7 day regimen.For both clinical and parasitological cure results at 12 weeks were similar to the 6 weeks treatment outcome. Furthermore, with respect to the difference in proportion parasitological cured the 3 day regimen was not only non-inferior but also worse compared to the 7 day regimen. The 2 groups did not differ on any of the SKINDEX-29 domains before injections and at 6 weeks follow-up . Before In general PI as study medication was well tolerated. No serious toxicity occurred and none of the reported side effects required discontinuation of treatment in any patient. Pain at the injection site was seen most frequently, 77.1% of patients in the 7 day regimen and 81.0% in the 3 day regimen . Non-infFor hemoglobin, creatinine and amylase levels we could not prove non-inferiority of the 3 day regimen over the 7 day regimen based on laboratory findings. Creatine levels were even significantly higher in the 3 day group . In the st injection 8.9% (90% CI: -3.0%\u201314.8%) more patients in the 3 day regimen had a drop in the diastolic BP > 10mmHG, 10.1% (8/79) compared to 1.2% (1/83) in the 7 day regimen.During the first injection, a drop in the diastolic BP of > 10 mmHg was recorded in 2.5% (2/79) in the 3 day regimen patients and 2.4% (2/83) in the 7 day regimen patients: difference 0.1% (90% CI: -3.9% to 4.1%). Thirty minutes after the 1Sensitivity analyses comparing missing data as successes showed that the 3 day regimen was non-inferior to the 7 day regimen: 5.5% (90% CI: -6.4% to 17.3%) and minus 1.4% (90% CI: -12.1% to 9.3%) at 6 weeks outcome for clinical and parasitological cure respectively. Twelve weeks after end of treatment non-inferiority was shown for clinical cure  but not for parasitological cure .Sensitivity analyses of the safety data set revealed that for adverse events the results were similarly not non-inferior to the ITT and PP analyses  Tables.We conducted a non-inferiority trial because we intended to show that the new 3 day regimen was not worse compared to the standard 7 day regimen. Our results showed that the 3 day regimen is not non inferior to the standard treatment  regarding clinical cure at 6 weeks. Therefore, we cannot recommend the 3 day regimen over the 7 day regimen to treat CL in Suriname. Although the 90% CI for the ITT analyses are slightly lower than the non-inferiority margin of 15%, consistency between PP and ITT analyses is an important requisition in non-inferiority trials before conclusions regarding non-inferiority can be drawn.,15 The oNonetheless sensitivity analyses  showed non-inferiority, except for parasitological cure at 12 weeks. Seven lost to follow-up patients in the 7 day regimen and six in the 3 day regimen could be reached by telephone. All confirmed that their lesions had healed. However, we also know from clinical practice that non-cured patients sometimes sought help in traditional medicine.Based on an average found in previous studies in Suriname we assumed a cure rate of 85% for both groups.,4 In theThe safety analyses should be considered as explorative data analyses because the power of the study was based on the primary endpoint and the issue of multiplicity arising from numerous comparisons may arise. The latter may explain why taste change occurred more frequently in the 3 day regimen while for the remaining safety analyses no differences were found. The absence of non-inferiority for several adverse events and laboratory values may be due to the higher concentration of PI per injection in the 3 versus the 7 day regimen.guyanensis and other spiecies like L. amazonensis, L. braziliensis and L. naff\u00ef have only been reported sporadically,[guyanensis.One of the limitations of this study is that we cannot mention the causative species affecting our participants. Since the far majority of CL in Suriname is caused by L. adically,,18,19 weTwo studies using the Dermatology Quality of Life Scale found lower HRQL in patients with active CL compared with healed CL patients., 21 We aA priori we hypothesized that the 3 day regimen would have resulted in a decreased number of individuals lost to follow up for the control visits. However, in the 3 day regimen the percentage lost to follow-up at 6 weeks was even higher with 33% versus 18% in the 7 day regimen. Although not statistically significant, this could be attributed to more side effects in the 3 day regimen. Another explanation could be a higher portion of healed patients in the 3 day regimen without a need for return visits. Unfortunately we were unable to conduct active follow-up as most patients returned to the hinterlands and could not be contacted. The high number of lost to follow-up may have biased the study results, especially because the percentage of individual\u2019s lost to follow-up where higher in the 3 day regimen. Because the sensitivity analyses showed non inferiority of the 3 day regimen there is indeed indication that the high percentage of loss to follow-up has biased the study results. Furthermore, it is possible that the power of our study was too low to detect non-inferiority because the clinical and parasitological cure rates were lower than assumed in the power calculation (85%). Too low power in a non-inferiority trial indicates that there is more chance to falsely conclude that the intervention is not non-inferior while the intervention is non-inferior. A retrospective study performed in Suriname in 2001 in which patients were treated with pentamidine from 1979 till 2000 found a cure rate of 90% after a follow-up of 3\u20136 months.[Since we knew from standard clinical practice that compliance is low in CL patients, those who doubted if they could attend all follow-up visits were not eligible. As a result, our study may suffer from a lower external validity because the study population might not be completely representative for the CL patient population in Suriname.In future studies serious attention should be given to reduce the number of lost to follow-up patients, for example via financial compensation for study related expenses. Moreover, novel oral CL treatment like miltefosine instead of painful PI injections could improve compliance.In conclusion, we cannot conclude that two injections of 7 mg/kg PI within three days is non-inferior to the standard regimen of three injections of 4 mg/kg PI within 7 days, for treatment of CL in Suriname. Therefore, there is no evidence to change the current standard practice of the 7 day regimen treatment for CL in Suriname.S1 ChecklistRandomized single-blinded non-inferiority trial of 7 mg/kg pentamidine isethionate versus 4 mg/kg pentamidine isethionate for cutaneous leishmaniaisis in Suriname(PDF)Click here for additional data file.S1 TableAdverse events 1 week after treatment according to treatment group of cutaneous leishmaniasis patients in Suriname from 2010\u20132013.(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file."}
+{"text": "Prompt diagnosis and effective treatment are considered the cornerstones of malaria control and artemisinin-based combination therapy (ACT) is currently the main anti-malarial drugs used for case management. After deployment of ACT due to widespread parasite resistance to the cheap and widely used anti-malarial drugs, chloroquine and sulphadoxine/pyrimethamine, the World Health Organization recommends regular surveillance to monitor the efficacy of the new drugs. The present paper assessed the implementation of anti-malarial efficacy testing for monitoring the therapeutic efficacy of ACT for treatment of uncomplicated malaria in Tanzania before and after policy changes in 2006.A literature search was performed for published clinical trials conducted in Tanzania from 2001 to 2014. It focused on studies which assessed at least one form of ACT for treatment of uncomplicated falciparum malaria in children less than 10\u00a0years and reported efficacy and safety of the tested anti-malarials. References were imported into the Endnote library and duplicates removed. An electronic matrix was developed in Microsoft Excel followed by full text review with predetermined criteria. Studies were independently assessed and information related to ACT efficacy and safety extracted.P. falciparum infections in Tanzania was high with PCR-corrected cure rates on day 28 of 91-100% and 88-93.8%, respectively. The highest day-3 parasite positivity rate was 1.4%. Adverse events ranged from mild to serious but were not directly attributed to the drugs.Nine papers were selected from 125 papers screened. The efficacy of both artemether-lumefantrine (AL) and artesunate-amodiaquine (AS\u2009+\u2009AQ) against uncomplicated ACT is efficacious and safe for treatment of uncomplicated malaria in Tanzania. However, few trials were conducted in Tanzania before and after policy changes in 2006 and thus more surveillance should be urgently undertaken to detect future changes in parasite sensitivity to ACT. Plasmodium falciparum to chloroquine (CQ) and the rapid spread of resistance to sulphadoxine-pyrimethamine (SP) prompted the introduction of artemisinin combination therapy (ACT). In 2001, a World Health Organization (WHO) expert panel recommended use of ACT for treatment of uncomplicated falciparum malaria in all endemic countries [Malaria is by far the most important parasitic disease in Tanzania and in other tropical countries, causing loss of life and morbidity with more than three billion people at risk globally . Prompt ountries .P. falciparum [The combination therapy involves simultaneous use of two or more blood schizonticidal drugs with independent modes of action and different biochemical targets in the parasite, a mechanism which delays development of parasite resistance ,7. ACT clciparum .The current recommended combinations are artemether-lumefantrine (AL), artesunate-amodiaquine (AS\u2009+\u2009AQ), artesunate-mefloquine (AS\u2009+\u2009MQ), dihydroartemisinin-piperaquine (DHA\u2009+\u2009PQ), and artesunate-sulphadoxine/pyrimethamine (AS\u2009+\u2009SP) . ArtemisParasite resistance to anti-malarials is of great concern in the efforts to control malaria worldwide. The parasites develop resistance by initially becoming tolerant to the drugs before they become fully resistant. The resistant parasites have an ability to survive under therapeutic levels of anti-malarial drugs which would otherwise kill both sensitive and tolerant parasites . ParasitIn vivo response of patients to treatments provides more information to clinicians and policy makers and is considered the gold standard for assessing anti-malarial efficacy. However, therapeutic efficacy must be interpreted as an interaction between the host factors , the parasite factors  and the drug factors  . In vitrWHO recommends regular efficacy testing for monitoring the efficacy of anti-malarials ,8,23. InP. falciparum resistance to SP was reported [Tanzania changed its malaria treatment policy from CQ to SP monotherapy as the first-line drug for the treatment of uncomplicated malaria in 2001 . Howeverreported ,27 and treported  to introreported  which wareported . Whereasreported , Tanzanireported .Unfortunately, artemisinin-resistant field isolates have been reported recently in four countries of Southeast Asia  and threatens the current progress in controlling the disease ,2,19. ThThe present paper reviewed the implementation of in vivo efficacy testing in Tanzania before and after deployment of ACT in order to monitor the efficacy of ACT for the treatment of uncomplicated malaria. The paper compares the cure rates, parasite clearance and fever clearance times and safety data reported in clinical trials involving ACT in Tanzania that were published between 2001 and 2014. It provides updates on country-specific performance of ACT after its wide-scale deployment for treating uncomplicated falciparum malaria.Published literature was searched and it involved papers published from January 2001 to August 2014. English language articles indexed in PubMed were searched using search terms: \u2018Tanzania AND malaria AND artemether-lumefantrine\u2019, \u2018Tanzania AND malaria AND artesunate-amodiaquine\u2019, \u2018Tanzania AND malaria AND artesunate-mefloquine\u2019, \u2018Tanzania AND malaria AND dihydroartemisinin-piperaquine\u2019 and \u2018Tanzania AND malaria AND artesunate-sulphadoxine-pyrimethamine\u2019. PubMed was used for primary search but in addition, Google Scholar, the Worldwide Antimalarial Resistance Network (WWARN) standardized analyses of ACT efficacy data repository and the African Journals Online (AJOL) were used to confirm that no study was missed. Inclusion criteria were clinical trials conducted in Tanzania between 2001 and 2014 and involved at least one ACT for treatment of uncomplicated falciparum malaria. The studies should have reported the efficacy and/or safety of the tested drugs. The starting year  was purposely chosen because that was the year when WHO advocated use of ACT for treating uncomplicated falciparum malaria . ReferenLiterature search yielded 126 records, and 21 of these were duplicate records which were removed. The titles and abstracts of the remaining records (105) were screened based on the inclusion criteria and 11 articles qualified for a full-text review. From the review, two additional papers were from multicentre trials across Africa, which partly included Tanzania, and these were also removed. Only nine papers were left and fully reviewed as summarized in Figure\u00a0Before and after the official adoption of ACT for treatment of uncomplicated malaria in Tanzania in 2007 , nine clFor the trials that tested AL, the reported PCR-corrected cure rates ranged from 91 to 100% Table\u00a0. The higAmong the five supervised trials that reported fever clearance, more than 80 and 79% of the patients cleared fever by day 1 post-treatment with AL and AS\u2009+\u2009AQ, respectively. None of the patients had fever on day 3  in a patient during the study regardless of whether it was related to the treatment. Adverse events were judged according to their causal association with ACT  and severity  . Cough wP. falciparum artemisinin-resistant field isolates in Southeast Asia [Following the recent reports of emergence of ast Asia ,2,19 andast Asia . InformaApparently, due to limited resources, especially funding, NMCPs in most endemic countries have not been able to implement regular anti-malarial drug efficacy monitoring at sentinel sites and there has been a strong call for regional networks to facilitate the implementation . The forThe findings of this review showed that nine clinical trials have been conducted to monitor the efficacy of ACT before and after Tanzania adopted ACT for treatment of uncomplicated malaria. Of these, only one study  was condThe present review has shown that the efficacy of AL, which is the first-line anti-malarial drug for treatment of uncomplicated falciparum malaria in Tanzania, was high even after unsupervised treatment. The PCR-corrected cure rate on day 28 was >91% and this is in line with findings from other studies in eastern Africa ,44. The The cure rates of AQ\u2009+\u2009AS in the present review compares well with those reported elsewhere in East Africa whereby day-28 adequate clinical and parasitological response (ACPR) in children treated with AS\u2009+\u2009AQ was 90.2% in Kenya , 90.3% iThere is a concern about the limited post-treatment prophylactic effects of both AL and AS\u2009+\u2009AQ in high transmission areas. In fact in one trial, more than half of the recruited patients had recurrent infections within the 42-day follow-up period after treatment with AL. However, the majority of recurrent infections were due to re-infections which suggests that the partner drug cannot give prolonged protection despite high therapeutic efficacy . SimilarIn most of the studies, a great majority of the recurrent infections were due to re-infections, when assessed with a step-wise PCR genotyping protocol, which signifies that the drugs are still efficacious and the high rates of re-infections could only be attributed to high malaria transmission. In terms of clinical practice, the high re-infection rates are of great concern among clinicians. Clinicians should be clearly guided on what to expect and how to handle such cases with recurrent infections within a period of three to eight weeks post-treatment. The observed high re-infection rates after ACT treatment underscores the importance of integrating treatment with vector control interventions, including use of long-lasting insecticide-treated nets so as to effectively block malaria transmission and prevent recurrent infections .The study which tested AZ\u2009+\u2009AS showed that the drug had low efficacy (28\u00a0days ACPR\u2009=\u200968%) and could not be considered a potential anti-malarial drug in Tanzania and other malaria-endemic countries . It is pMeasurement and reporting of parasite clearance on day 3 after treatment with ACT is particularly important, as this is one of the first signals of emergence of parasite tolerance/resistance to artemisinin . In the Artemisinins are known to be highly potent anti-malarial drugs that are active against immature gametocytes and are useful in the reduction of malaria transmission and elimination/eradication agenda . In clinIt is well established that the efficacy of AL combination is strongly influenced by variations in the pharmacokinetics of lumefantrine among individuals . The maxThe present review showed no unexpected adverse events and overall, AL, AS\u2009+\u2009AQ and AZ\u2009+\u2009A were well tolerated. Admittedly, the few studies that reported safety profile in the present review  would not enable a firm comparison of safety of different anti-malarials. However, other studies in Africa have shown that certain mild or moderate adverse events, such as vomiting and anaemia, were more frequent in patients treated with AS\u2009+\u2009AQ than those treated with AL ,71. ThisP. falciparum malaria in the country.This review was meant to assess the implementation of the WHO recommendations of undertaking regular monitoring of antimalarial efficacy studies and also provide Tanzania-specific current efficacy and safety profile of ACT in the treatment of uncomplicated falciparum malaria. The review was limited to peer-reviewed publications, thus unpublished data were not included. However the review highlighted the levels of implementation of TETs in Tanzania and provides an overall country-specific performance of ACT after their wide-scale deployment as first-line anti-malarials for treating uncomplicated P. falciparum after treatment with artemisinins, the demand for tracking parasite sensitivity to artemisinin and its derivatives has become more important. More accurate estimates of parasite clearance measurements through frequent parasite counts (at least twice daily) to assess delayed parasite clearance should be adopted in future therapeutic efficacy testing studies [Following the emergence of artemisinin resistance in Southeast Asia, manifested as delayed clearance of  studies ,60. HoweGiven the recent documentation of K13-propeller polymorphism as a molecular marker for monitoring resistance of artemisinin and its derivatives  and despP. falciparum infections in Tanzania. These findings support continued use of AL and AS\u2009+\u2009AQ for the treatment of uncomplicated malaria in Tanzania mainland and Zanzibar, respectively. Although currently there is no evidence of artemisinin resistance in Africa, regular monitoring and surveillance, as recommended by the WHO-supported GPARC must be implemented so that the emergence of artemisinin resistance in African can be timely detected, reported and contained. More surveillance and monitoring of anti-malarial efficacy and safety should be performed to detect future changes in parasite sensitivity to ACT.The present review has shown that few studies have been conducted in Tanzania to monitor the efficacy and safety of ACT and majority of these were not done under the NMCP framework. However, the findings revealed that the efficacy AL and AS\u2009+\u2009AQ was reasonably high and the drugs were safe when used for treatment of uncomplicated"}
+{"text": "Malaria and HIV infections are both highly prevalent in sub-Saharan Africa, with HIV-infected patients being at higher risks of acquiring malaria. The majority of antiretroviral (ART) and anti-malarial drugs are metabolized by the CYP450 system, creating a chance of drug-drug interaction upon co-administration. Limited data are available on the effectiveness of the artemether-lumefantrine combination (AL) when co-administered with non-nucleoside reverse transcriptase inhibitors (NNRTIs). The aim of this study was to compare anti-malarial treatment responses between HIV-1 infected patients on either nevirapine- or efavirenz-based treatment and those not yet on ART (control-arm) with uncomplicated falciparum malaria, treated with AL.This was a prospective, non-randomized, open-label study conducted in Bagamoyo district, with three arms of HIV-infected adults: efavirenz-based treatment arm (EFV-arm) n\u2009=\u200966, nevirapine-based treatment arm (NVP-arm) n\u2009=\u2009128, and control-arm n\u2009=\u200975, with uncomplicated malaria. All patients were treated with AL and followed up for 28\u00a0days. The primary outcome measure was an adequate clinical and parasitological response (ACPR) after treatment with AL by day 28.Day 28 ACPR was 97.6%, 82.5% and 94.5% for the NVP-arm, EFV-arm and control-arm, respectively. No early treatment or late parasitological failure was reported. The cumulative risk of recurrent parasitaemia was >19-fold higher in the EFV-arm than in the control-arm . The cumulative risk of recurrent parasitaemia in the NVP-arm was not significantly higher than in the control-arm . The median (IQR) day 7 plasma concentrations of lumefantrine for the three arms were: 1,125\u00a0ng/m (638.8-1913), 300.4\u00a0ng/ml (220.8-343.1) and 970\u00a0ng/ml (562.1-1729) for the NVP-arm, the EFV-arm and the control-arm, respectively (P\u2009<\u20090.001). In all three arms, the reported adverse events were mostly mild.After 28\u00a0days of follow-up, AL was statistically safe and effective in the treatment of uncomplicated malaria in the NVP-arm. The results of this study also provide an indication of the possible impact of EFV on the performance of AL and the likelihood of it affecting uncomplicated falciparum malaria treatment outcome. Malaria and HIV-1 are the most common infections in sub-Saharan Africa and to a lesser extent in other developing countries ,3. WorldThe World Health Organization (WHO) recommends the use of artemisinin-based combination therapy (ACT) as first-line treatment for uncomplicated falciparum malaria in all malaria-endemic countries . ArtemetArtemether is metabolized to dihydroartemisinin (DHA) via cytochrome P450 (CYP) CYP3A4, CYP2B6 and possibly CYP2A6 . LumefanThe treatment guidelines for uncomplicated malaria set by the National Malaria Control Programmes in sub-Saharan Africa do not discriminate between the dosing regimen of AL in non-HIV infected patients and those undergoing ART.To date, most of the reported studies describe the effect of HIV-1 infection on malaria treatment outcome in the absence of antiretroviral drug treatment -7,12-20.This study was conducted between May 2010 and August 2012 at an HIV clinic at Bagamoyo District Hospital. Bagamoyo district is an area of moderate malaria transmission. In this district, the peak time for malaria transmission peak time is usually around April to August. The study was approved by the Muhimbili University of Health and Allied Sciences (MUHAS) research and ethics committee and was conducted according to Good Clinical Practice. To ensure confidentiality, patients were identified by special identification numbers.This study involved HIV-1 infected adults receiving ART as 200\u00a0mg NVP twice daily or 600\u00a0mg EFV at night for more than two months and those not yet on ART. Enrolled in the study were 128 patients in the nevirapine-based treatment arm (NVP-arm), 66 patients in the efavirenz-based treatment arm (EFV-arm) and 75 patients not yet on ART (control-arm).Plasmodium falciparum with no signs of complicated (severe) malaria; no history of an allergic reaction or serious side effects to AL or treatment with anti-malarial drugs for at least four weeks prior to enrollment; no evidence of chronic diseases, such as renal or liver failure; not on anti-tuberculosis drugs for at least three months prior to enrollment; not pregnant or a nursing mother; easy accessibility to the health-care facility (travel time\u2009<\u20092\u00a0hours) and willingness to attend for the stipulated follow-up visits. Before enrollment, written informed consent was obtained from all patients. All patient information was recorded in a case report form (CRF). The enrolled patients were encouraged to take their ART and AL as prescribed.All HIV-1 infected patients presenting at the HIV clinic for routine medical care, with fever and other symptoms suggestive of uncomplicated falciparum malaria such as chills, sweats, headaches, muscle aches, nausea, vomiting, diarrhoea, body weakness, poor appetite, pallor and enlarged spleen were screened for eligibility. Patients were only enrolled into the study after meeting the inclusion criteria: Aged\u2009\u2265\u200918\u00a0years; reported fever within the last 24\u00a0hours and/or an axillary temperature\u2009\u2265\u200937.5\u00b0C and with any of the above-mentioned symptoms of uncomplicated falciparum malaria; haemoglobin\u2009\u2265\u20097\u00a0g/dl and\u2009\u2265\u200935\u00a0kg body weight; microscopically-confirmed This was a prospective, non-randomized, open-label, parallel and three-arm study. Patients were followed up for 28\u00a0days. Patients meeting the inclusion criteria were enrolled and took the full dose (three-day course) of AL  at 0, 8, 24, 36, 48 and 60\u00a0hrs. The first and fifth doses of AL were taken by direct observed therapy (DOT) with 250\u00a0ml of milk (3.5% fat). The other four doses were taken at home. All patients were given verbal instructions on dosing intervals and on the importance of combining treatment with fatty meals. Additionally, patients were supplied with 10 extra 250\u00a0ml packets of milk (3.5% fat) to be taken with the rest of the doses at home. For the first and fifth doses, which were given by DOT and paracetamol was administered to all febrile patients.Patients involved in this study were counseled to abstain from using alcohol, tea, caffeine and any drugs which may induce CYP3A4, such as griseofulvin, prednisolone, phenytoin, carbamezapine and phernobarbital. Non-prescription drugs, herbal medicines, oral contraceptives, grapefruits or grapefruit juice were also prohibited during the study.At enrollment all patients gave a finger-prick blood sample for thick smear and for haemoglobin (Hb) estimation. Blood slides for malaria parasites were all read at Ifakara Health Institute-Bagamoyo Research and Training Centre (IHI-BRTC). Venous blood was collected at pre-determined times for quantification of lumefantrine plasma concentrations. Patients\u2019 baseline CD4+ cell count was obtained from their records, the timeline being within 3\u00a0months prior to study enrollment.Laboratory and clinical assessments were conducted on days 2, 3, 7, 14, 21 and 28 or on any day of recurrent illness. A reminder was sent to all patients by a study nurse via telephone about their medication and study visit schedules. Patients were encouraged to return to the study site any time they felt ill. Patients who failed to return on the scheduled day were visited and assessed at home. If the study nurse failed to locate a patient\u2019s house, they were classified as lost to follow-up. Any additional medications taken during the study period were documented in the CRF.The time to recurrent parasitaemia or the risk of recurrent parasitaemia (RP) was defined as the number of days between taking the first dose of AL and the day of microscopically detecting malaria parasites in the thick blood film. The time at risk ended whenever one of the following conditions occurred: RP, loss to follow-up, withdrawal, or end of follow-up period .P. falciparum during the 28\u00a0days of follow-up were treated with either quinine tablets or injection as described in the malaria treatment guidelines (2006) of Tanzania [Patients with microscopically-confirmed Tanzania .All thick blood smears were stained with 10% Giemsa stain for 30\u00a0minutes. Parasite density was estimated by counting the number of asexual parasites per 200  white blood cells (WBC) on a thick smear. Parasite density per \u03bcl was calculated by assuming a WBC count of 8,000 per \u03bcl . All thiBlood samples from patients (4\u00a0ml) were collected in heparinized vacutainer tubes and centrifuged (\u00d72000\u00a0g for 10\u00a0min) immediately to obtain plasma. Aliquots of plasma were transferred into 1\u00a0ml cryo-tubes, and stored at \u221280\u00b0C at the Ifakara Health Institute\u2013BRTC until transfer to MUHAS for analysis. All patients\u2019 plasma samples were analyzed at the MUHAS-Sida bio-analytical laboratory in Dar es Salaam. Lumefantrine concentrations were quantified using an HPLC method with UV detection as previously reported . The coeAn adequate clinical and parasitological response in patients at 28\u00a0days after anti-malarial treatment was the primary study objective. The WHO guidelines for assessment and monitoring of anti-malarial drug efficacy for the treatment of uncomplicated falciparum malaria were used in the evaluation of the time to RP after treatment with AL . AccordiThis study was considered to be exploratory. In total, 269 HIV-1 infected adult patients with uncomplicated falciparum malaria were enrolled with a minimum sample size of at least 50 patients for each arm . The datA total of 1,528 patients presenting at the HIV clinic with fever and other symptoms suggestive of malaria infection were screened. Among the screened patients, 316 (21%) had positive thick blood smears for malaria parasite. In total 269 (85%) patients met the inclusion criteria and were enrolled .In the current study, about 48% of the enrolled patients had CD4 cell counts of\u2009<\u2009350 cells/\u03bcl. Patients with high parasitaemia had low CD4 cell counts and there was a strong association between CD4 cell counts and parasitaemia (P\u2009<\u20090.001).No early treatment failure or late parasitological failure was observed in the three arms. Overall, after day 28 of follow-up, 97.6% , 82.5%  and 94.5%  of patients in the NVP-arm, EFV-arm and control-arm, respectively, had no recurrent parasitaemia, thus meeting the WHO criteria for ACPR. The differences in the treatment outcome in the three arms were highly statistically significant (P\u2009<\u20090.001)  patients had lumefantrine plasma concentrations measured on day 7 after initiation of treatment. The median (IQR) day 7 plasma concentrations of lumefantrine for the three arms were: 1,125\u00a0ng/ml (638.8-1913), 300.4\u00a0ng/ml (220.8-343.1) and 970\u00a0ng/ml (562.1-1729) for the NVP and EFV-based treatment arms and for the control-arm, respectively. The difference in day 7 lumefantrine plasma concentrations between the EFV-arm and the control-arm was statistically significant (P\u2009<\u20090.001), while there was no statistically significance difference in the day 7 lumefantrine concentration between the NVP-arm and the control-arm (P\u2009=\u20090.063). In all three arms, the median lumefantrine concentration was significantly lower in patients with RP as compared to those with no RP  as opposed to a subject with poor immunity. However, in patients with poor immunity, the initial rate of parasite clearance is determined by the intrinsic activity of the drug, the susceptibility of infecting parasites and the drug levels achieved . The patPossible DDIs between ACT and NNRTIs in HIV-1 patients without malaria have been reported in a few studies -34. ThisP. falciparum malaria for a first-line drug in non-HIV patients should be at least 90% and preferably >95% [P. falciparum treated with AL is >95% [In the present study, all patients from the three arms were treated with artemether-lumefantrine and followed up for 28\u00a0days to determine malaria treatment response (ACPR). Based on WHO recommendations, the cure rates for bly >95% . Various is >95% . Under t is >95% , this stThe observed results in the EFV-arm may be due to possible DDIs between EFV and lumefantrine during the course of treatment as a result of the induction of common metabolic enzyme CYP3A4, leading to increased clearance of lumefantrine and artemether -30,42. AThese results are in agreement with findings from previous studies, which indicated a possible malaria treatment failure in patients treated with AL and EFV ,45 and bLumefantrine absorption varies considerably among individuals with its bioavailability being improved by fatty meal intake. As this study was unsupervised, poor adherence to treatment or inadequate fatty meal intake might have contributed to the observed sub-optimal day 7 lumefantrine plasma concentrations in the three arms, thus increasing the risk of recurrent parasitaemia. However, an AL cure rate of >96% was recently reported irrespective of whether it was given under supervision, with food or unsupervised .Differences in immune status among the studied population might have also contributed to the observed differences in the ACPR between the three arms. Patients in the EFV-arm had lower mean CD4 cell counts compared with those in the NVP and control-arms, although the differences did not reach statistical significance (P\u2009=\u20090.629). This is in line with previous reports ,17. Studet al. reported high risks of recrudescence in patients with high BPC and low plasma concentration of lumefantrine [Malaria parasite density was slightly higher among patients in both the NVP-arm and the EFV-arm than in the control-arm, although this difference was not statistically significant. Lower CD4 cell counts in the EFV and NVP-arms than in the control-arm might also have contributed to higher parasite density. This finding is in agreement with other studies, which documented higher parasite density with decreased immunity among HIV-1 infected patients ,17,46,47fantrine .P. falciparum parasitaemia, while the presented fever was caused by opportunistic infections. Reports from other studies have indicated that the febrile illness that occurs more frequently in HIV/AIDS patients, especially those with low CD4 cell counts, even in the presence of malaria infection, may be caused not by malaria but by other opportunistic infections or may be due to adverse drug reactions, thus complicating the diagnosis of malaria [P. falciparum parasitaemia among ART-treated patients and healthy people, respectively [While the results presented and discussed above are of clinical importance in improving the quality of life for HIV-1 infected patients on ART with uncomplicated falciparum malaria and treated with AL, nevertheless, they are not without limitations. In the present study, the enrolled patients were carefully screened for fever history, the presence or absence of other obvious causes of fever and the researchers carefully followed up guidelines for malaria screening and diagnosis ,36, neve malaria ,52-54. Tectively ,56.P. falciparum infection is reported to be equal or superior to routine microscopy (but inferior to expert microscopy) and do not generate all information provided by microscopy [Decreasing the risk of false negative malaria, especially in patients with low malaria parasitaemia, using more sensitive diagnostic methods such as the malaria rapid diagnostic test (MRDT) and quantitative real-time polymerase chain reaction (PCR), could have complimented the thick blood smear film method. However, the MRDT and PCR are not without limitations as it is in the case of thick blood film. The accuracy of RDT for the diagnosis of uncomplicated croscopy . The thicroscopy ,59.P. falciparum deficiency (Pfhrp2 gene deletions) and due to cross-reactions between Plasmodium species. The sensitivity of MRDT has been reported to be reduced to\u2009<\u200975% with\u2009<\u20091000 parasites/\u03bcl [Thick blood films have been reported to give conflicting results in various studies -62. Howesites/\u03bcl ,64,65. Fsites/\u03bcl ,62,64,66sites/\u03bcl . This stsites/\u03bcl ,61,64,68In the present study, malaria parasites were not genotyped to differentiate recrudescence from re-infection ; in areaIn conclusion, the findings of this study suggest that AL is safe and effective in the treatment of uncomplicated falciparum malaria in patients receiving NVP-based ART compared to those receiving EFV-based ART. The results of this study also provide an indication of the possible impact of EFV on the performance of AL and the likelihood of it affecting malaria treatment outcome. Surveillance on the effectiveness and efficacy of AL in HIV infected patients, particularly those on EFV-based ART, should be performed in order to elucidate the role of the latter in the treatment outcome of uncomplicated falciparum malaria.The authors declare that they have no competing interests.OMSM and SP conceived the study, participated in the study design, coordination, data analysis and manuscript writing. BM participated in study design, data collection and analysis and in the preparation and writing of the manuscript. BN participated in the data analysis as well as manuscript preparation. All authors participated in reading and approving the final manuscript."}
+{"text": "Plasmodium vivax depends upon co-administered blood schizontocidal therapy in radical cure. We assessed primaquine (PQ) as hypnozoitocide when administered with dihydroartemisinin-piperaquine  or artesunate-pyronaridine  to affirm its good tolerability and efficacy. A third arm, artesunate followed by primaquine, was not intended as therapy for practice, but addressed a hypothesis concerning primaquine efficacy without co-administration of blood schizontocide.Safety and efficacy of primaquine against repeated attacks of P. vivax recurrences in the 12\u00a0months following therapy were classified as relapses. A historic relapse control derived from a cohort of soldiers who served in the same area of Papua was applied to estimate risk of relapse among randomized treatment groups. Those were: 1) AS followed 2d later by PQ (0.5\u00a0mg/kg daily for 14d); 2) co-formulated AS-PYR concurrent with the same regimen of PQ; or 3) co-formulated DHA-PP concurrent with the same regimen of PQ.During March to July 2013, an open-label, randomized trial enrolled Indonesian soldiers with vivax malaria at Sragen, Central Java, after six months duty in malarious Papua, Indonesia. No malaria transmission occurred at the study site and Among 532 soldiers, 219 had vivax malaria during the four months following repatriation to Java; 180 of these were otherwise healthy and G6PD-normal and enrolled in the trial. Subjects in all treatment groups tolerated the therapies well without untoward events and cleared parasitemia within three days. First relapse appeared at day 39 post-enrollment, and the last at day 270. Therapeutic efficacy of PQ against relapse by incidence density analysis was 92\u00a0% (95 %CI\u2009=\u200983\u201397\u00a0%), 94\u00a0%(95 %CI\u2009=\u200986\u201397\u00a0%), and 95\u00a0%(95 %CI\u2009=\u200988\u201398\u00a0%) when combined with AS, AS-PYR, or DHA-PP, respectively.P. vivax relapse when administered concurrently with DHA-PP or AS-PYR. These offer alternative partner drugs for radical cure with primaquine. The AS arm demonstrated efficacy with a total dose of 7\u00a0mg/kg PQ without concurrently administered blood schizontocide, another option when primaquine therapy is removed in time from the treatment of the acute malaria or applied presumptively without an attack.This trial offers evidence of good tolerability and efficacy of PQ against ISRCTN82366390, assigned 20 March 2013.Current Controlled Trials  Plasmodium vivax threatens nearly three billion people and sickens tens of millions annually [Plasmodium falciparum, P. vivax places latent stages called hypnozoites in the liver after inoculation of mosquito-borne sporozoites. Hypnozoites later awaken and provoke renewed clinical attacks, each causing debilitating illness with risk of threatening complications and onward transmission.annually . Impropeannually , 3. UnliP. vivax relapse behaviors resemble those in the Chesson strain from New Guinea [P. vivax attacks within two months [P. vivax occurs [In tropical Southeast Asia, w Guinea , 5. \u201cChew Guinea , 7. Amono months . Relapsex occurs , 9. TherP. vivax malaria. Primaquine remains the only option against relapse, although another 8-aminoquinoline, the investigational drug tafenoquine, approaches licensure [P. vivax often occurs, especially in Southeast Asia [P. vivax dominate across the Indonesian archipelago, and chloroquine was abandoned in favor of artemisinin-combined therapies (ACT) therapies over a decade ago [Radical cure of vivax malaria includes blood schizontocidal and hypnozoitocidal therapies. In 1952 the United States licensed the 8-aminoquinoline primaquine combined with the 4-aminoquinoline chloroquine for radical cure of icensure . Newer bast Asia . Resistacade ago .P. vivax malaria cite good evidence of efficacy [P. vivax in Indonesia is the ACT dihydroartemisinin-piperaquine (DHA-PP) [P. vivax [Authoritative recommendations for the use of ACTs for treatment of acute efficacy , 14. Howefficacy . Frontli(DHA-PP) . We repo(DHA-PP) . Later s(DHA-PP) . The cur(DHA-PP) . Both ofP. vivax \u201321.The trial included a third treatment arm, artesunate  followedP. vivax strains dominating eastern Indonesia [We recruited Indonesian soldiers exposed to malaria in eastern Indonesia after returning to a site in Central Java free of malaria transmission . In addindonesia .P. vivax malaria after returning from malarious Papua . These soldiers received treatment according to national guidelines [The battalion identified for study returned to Java by ship, disembarking at Tanjung Mas at Semarang on 28 February 2013. Routine microscopic screening for malaria at portside revealed 67 of 536 soldiers had malaria  occupies 942\u00a0kmP. vivax were screened for eligibility by obtaining a medical history, and conducting physical and laboratory examinations that included electro-cardiogram , complete blood counts , full blood chemistry panel , and G6PD deficiency screening . Eligible subjects had uncomplicated vivax malaria, were not under treatment for another illness, had not recently consumed antimalarials, and showed normal ECG, blood laboratory values, and G6PD activity by fluorescent spot testing .Consenting soldiers with confirmed The trial statistician block-allocated treatment assignments by varying blocking number at random . A plainGroups received one of the following treatments under directly observed, signature-affirmed supervision by a member of the research team:AS\u2009+\u2009PQ \u2013 200\u00a0mg AS  was administered, followed by a daily dose of 100\u00a0mg daily for six days. After a 48-hour pause for AS washout, PQ was administered with a daily dose of two tablets, each containing 15\u00a0mg PQ base for 14\u00a0days. As in all other groups, Sanofi Pharmaceuticals  provided the PQ administered.AS-PYR\u2009+\u2009PQ \u2013 single daily dose of three tablets, each containing 60\u00a0mg AS and 180\u00a0mg of PYR provided as Pyramax\u00ae from Shin Poong Pharmaceuticals, Seoul, South Korea, for three days given concurrently with a daily dose of 30\u00a0mg PQ base for 14\u00a0days.DHA-PP\u2009+\u2009PQ \u2013 a single daily dose of three tablets, each containing 40\u00a0mg DHA and 320\u00a0mg of PP base provided as Eurartesim\u00ae from Sigma-Tau Industrie Farmaceutiche Riunite S.p.A., Pomezia, Rome, Italy, for three days given concurrently with a daily dose of 30\u00a0mg PQ base for 14\u00a0days.Subjects weighing >70\u00a0kg were treated as above except instead receiving 45-mg daily dose PQ (three tablets) for 14\u00a0days, five daily AS tablets instead of four, and four rather than three tablets of either AS, AS-PYR or DHA-PP daily. All subjects were offered a non-fatty carbohydrate snack prior to dosing to mitigate stomach upset caused by PQ . SubjectP. vivax received DHA-PP\u2009+\u2009PQ (0.5\u00a0mg/kg/day\u2009\u00d7\u200914\u00a0days) and were released from study after 28\u00a0days. Subjects not relapsing were followed for one year.Physical complaints and methemoglobin  were recorded daily during therapy. Routine blood film examinations occurred on days 3, 7, 14, 21, 28, 35, 42, 56, 63, 70, 84, 126, 140, 180, and 365 post-enrollment. Subjects reporting to the clinic with illness were examined the same day. Subjects positive for P. vivax in the year following radical cure. Patients reaching that endpoint contributed time at risk up to that event or withdrawal. Microscopic diagnoses of P. vivax were confirmed by another microscopist on site, and later by a blinded third microscopic read in Jakarta. Blood blots were collected onto Whatman FTA Classic\u2122 filter paper for PCR confirmation of the diagnosis, and DNA extracted (QIAmp DNA Blood Mini\u2122) before nested PCR Plasmodium genus-specific primers (Nest 1) and then with species-specific primers for Nest 2 as detailed elsewhere [Primary endpoint was incidence density (events/person-year) of first relapse by lsewhere . SecondaWe powered the trial for precision in independent estimates of PQ efficacy when combined with AS, AS-PYR, or DHA-PP. The precision estimate was derived iteratively through analysis of binomial distribution of variance around a predicted 98\u00a0% efficacy . A samplAS-Lumajang) against incidence density (ID) of recurrent P. vivax post-therapy among groups as follows:The primary endpoint of incidence density informed the estimate of efficacy derived from a relapse control. That control adjusts for geographic variability in relapse attack rates , 29. We The 95\u00a0% confidence interval (CI) for each of the three estimates of efficacy was calculated from the binomial distribution.P. vivax attacks divided by the sum of person-years at risk at end of study. Therapeutic efficacy expressed as proportion of subjects remaining free of recurrence after six months was included in this trial analysis for sake of reference to other hypnozoitocidal trials where reinfection possibly confounded recurrence rates [Incidence density of relapse represents the preferred mathematical treatment for events occurring over prolonged periods , 31. Likce rates .Badan Pengawasan Obat dan Makanan, Jakarta; ref.no. PN.01.06.313.03.13.998) approved and monitored this trial.Institutional ethics review boards of the Faculty of Medicine, University of Indonesia (ref.no. 13/H2.F1/ethic/2013) and the Centre for Tropical Medicine, Nuffield Department of Medicine, University of Oxford (ref.no.179-12) reviewed and approved a protocol detailing this trial. Indonesian food and drug regulatory authorities (P. vivax (confirmed by PCR) ranging from 16\u20139,488/\u03bcL . Fever (>37.5\u00a0\u00b0C) occurred in 22\u201330\u00a0% among groups (P\u2009=\u20090.72). No significant differences appeared between treatment groups in baseline vital signs or laboratory testing, except weight (P\u2009=\u20090.05). These same characteristics in the historic relapse control cohort are shown at the far right of Table\u00a0Figure\u00a0P. vivax after complaining of illness, but a minority was discovered during routine blood survey of the battalion (53/180). Illness was mild in most cases and dominated by headache, fever, chills, or muscle ache. A few subjects did suffer moderate illness with nausea or vomiting (12/180).Table\u00a0Physical complaints resolved within several days. Nausea, vomiting, and headache occurred in fewer than 3 of 60 subjects among treatment arms. Adverse events of grade 3 or higher occurred among five, three, and one subject receiving AS\u2009+\u2009PQ, AS-PYR\u2009+\u2009PQ, and DHA-PP\u2009+\u2009PQ, respectively. None were definitely related to drug, and two were probably related to drug (vomiting with AS-PYR\u2009+\u2009PQ). Seven subjects were hospitalized during the study and each was classified as having experienced a serious adverse event unrelated to acute malaria or study drugs .Most blood chemistry and cell counts were normal at enrollment and did not deviate during follow-up. However, lymphocytes, platelets, granulocytes, total bilirubin, and hemoglobin were abnormal but returned to normal Fig.\u00a0: depressLiver and other blood chemistry values changed after initiating therapy Fig.\u00a0. AspartaFigure\u00a0All subjects cleared asexual parasitemia by day 3. Most subjects were gametocytemic at enrollment (121/160), representing 60\u00a0%, 62\u00a0%, and 70\u00a0% rates among AS\u2009+\u2009PQ, AS\u2009+\u2009PYR, and DHA-PP groups, respectively (P\u2009=\u20090.342). Gametocytemia occurred more frequently among symptomatic  compared to asymptomatic  subjects; OR\u2009=\u20094.4 . The initial geometric mean (range) per \u03bcL of 128(16\u20131232), 80(16\u20131200), and 64(16\u20131200) among those respective groups (P\u2009=\u20090.287) vanished within 24\u00a0hr of initiating therapy except in one, one, and two subjects. All were free of microscopically patent gametocytemia by day 3. Table\u00a0This randomized trial of 0.5\u00a0mg/kg PQ daily for 14\u00a0days concurrent with AS-PYR or DHA-PP for radical cure of vivax malaria demonstrated good safety, tolerability, and efficacy for these options. All subjects cleared parasitemia by day 3 and none recurred before day 39, affirming good efficacy of AS-PYR and DHA-PP against the asexual blood stage parasites \u201321. InitDrug-related changes occurred among subjects. The significant elevation of AST/ALT in a minority of subjects receiving AS-PYR (5/60) has been reported in other studies . QTc eloThe AS\u2009+\u2009PQ treatment in the current trial tested the hypothesis that PQ requires concurrent blood schizontocidal therapy for good efficacy against hypnozoites. In 1955, Alving et al.  demonstrP. vivax strains from tropical Southeast Asia and New Guinea to the lower dose [PQ is prescribed at a total dose of either 3.5 or 7.0\u00a0mg/kg over 14\u00a0days, depending upon geographic origin of infection . The Newwer dose , 35, 36.wer dose , and cytwer dose , 39. We In addition to the primary incidence density endpoint analysis, Table\u00a0This trial provided robust estimates of PQ efficacy against relapse when combined with modern therapies. The findings of good safety, tolerability and efficacy in this trial reassures, but does not address the serious problem of PQ effectiveness. PQ often fails because it is: 1) unavailable or prohibited ; 2) rareP. vivax receiving AS-PYR\u2009+\u2009PQ or DHA-PP\u2009+\u2009PQ enjoyed good tolerability, safety and efficacy of these regimens. AS-PYR and DHA-PP may thus be considered validated options to chloroquine for radical cure of P. vivax with PQ. Further, PQ at 7\u00a0mg/kg total dose does not require concurrent administration of blood schizontocide for good efficacy. PQ may thus follow therapy with untried blood schizontocides for either therapy or chemoprophylaxis with the expectation of good efficacy.Patients with"}
+{"text": "In malaria endemic areas, schoolchildren usually have asymptomatic malaria infections and consequently remain untreated. Therefore, intermittent preventive treatment with sulfadoxine-pyrimethamine in schoolchildren would be a plausible strategy in malaria stable transmission areas to prevent anaemia and malnutrition. However, in contrast to infancy and pregnancy, antimalaria intermittent preventive treatment in children has been barely investigated. As the implementation of intermittent preventive treatment may be challenged by sulfadoxine-pyrimethamine resistance, sulfadoxine-pyrimethamine combined with piperaquine may be a better alternative than sulfadoxine-pyrimethamine monotherapy. A clinical trial is being conducted to assess the efficacy and safety of intermittent preventive treatments versus controls in Democratic Republic of Congo (DRCongo) schoolchildren and their impact on sulfadoxine-pyrimethamine resistance.A phase IIIb, randomised, controlled trial will enroll asymptomatic schoolchildren. For interventions, sulfadoxine-pyrimethamine is compared to sulfadoxine-pyrimethamine plus piperaquine and to a control group. The two treatments are given four-monthly from baseline for a year as a single dose for sulfadoxine-pyrimethamine and two doses at 24-hour intervals for piperaquine. All participants receive praziquantel and albendazole as mass-treatment for helminthiasis at enrolment. The primary endpoint is haemoglobin concentration change at 12 months follow-up. Secondary endpoints are malaria parasite load and malaria prevalence, at baseline and at month 12. Malaria and helminthiasis incidence will be monitored throughout the study. Statistical analysis will use multilevel modelling due to repeated measurements and clustering effect of participants.The very few studies on intermittent preventive treatment in schoolchildren in malaria stable transmission areas have contradictory results. This randomised controlled trial is unique in comparing efficacy and safety of a prophylactic combination therapy to monotherapy or a control group after 12 months follow-up. Resistance markers for sulfadoxine-pyrimethamine (including break through parasitaemias) will also be recorded. Its uniqueness lies also in the fact that we use piperaquine, a long acting antimalarial, in combination with sulfadoxine-pyrimethamine. Artemisinin derivatives have been excluded as it is part of the treatment policies in virtually all malaria endemic countries. Our findings may, therefore, contribute to the public health of youngsters who fail to thrive and grow due to multiple morbidities.NCT01722539; PACTR201211000449323 Plasmodium falciparum [Malaria is a major parasitic disease in developing countries and particularly in Sub Saharan Africa (SSA). Malaria is caused by five plasmodium parasites, the most severe being lciparum . More thlciparum ; 90% of lciparum ,4. The Dlciparum .Plasmodium triggers the development of acquired immunity, the premunition that somewhat protects older children and adults against clinical malaria and reduces the risk of death in school-age children [Plasmodium infections, if untreated, persist and maintain malaria-induced inflammation that is commonly associated with iron deficiency anaemia through impaired intestinal iron absorption and iron release from hepatocytes, and impairment of the recycling of iron derived from parasitized red blood cells phagocytosis [Severe anaemia is one of the leading causes of death due to malaria disease in childhood in intense and stable malaria transmission areas . In endechildren ,8. Asympocytosis . In low ocytosis . Also maocytosis .Intermittent preventive treatment (IPT) with sulfadoxine-pyrimethamine (SP) is a safe and efficacious control tool for malaria that combined or not with insecticidal treated nets, showed substantial protection against malaria in vulnerable populations -13. IPT IPT markedly reduces clinical malaria incidence and overall child mortality, and improves haemoglobin concentration and the nutritional status of children <5-years-old ,15. AddiP. falciparum.Favourable drugs for use as IPT should balance long half-life against efficacy, safety, tolerability and potentiality for cross-resistance selection . Use of Other long-acting drugs available are mefloquine, amodiaquine, and piperaquine. However, due to safety concerns, mefloquine might not be optimal for IPT. Amodiaquine is not suitable for IPT due to its three-day treatment regimen and is used as a first line treatment in several countries including DRCongo. Piperaquine has been extensively used for mass prophylaxis and treatment since 1978 in China and other malaria endemic countries of Asia .P. falciparum life cycle [pfDHFR and pfDHPS gene mutations as a consequence of an increasing proportion of individuals unnecessarily exposed to drugs [Pyrimethamine binds to dihydrofolate reductase (DHFR) enzyme and sulfadoxine to dihydropteroate synthase (DHPS) and, thereby, the two SP components inhibit the fe cycle . Howeverto drugs . Of noteto drugs .et al. and Gready [To minimise the risk of SP resistance, SP could be combined with piperaquine (SP-PQ) as recommended by Cairns d Gready ,24. AlsoConsidering the facts that in our study area: (1) IPT of malaria provides substantial protection against anaemia and malaria in school children; (2) the level of SP resistance has no significant impact on the prophylactic efficacy ; and (3)This study primarily aims to assess the effect of antimalarial IPT on anaemia one year after initial intervention in school-aged children of stable malaria transmission and hyper endemic areas.Secondarily, in the same study population, the purpose of the study is to:1. Determine the prevalence of SP resistance markers at baseline and at month 12 follow-up;.2. Compare the efficacy and safety of SP versus SP-PQ.3. Assess the impact of antimalarial IPTsc on malaria incidence and severity.4. Evaluate the impact of IPTsc on the nutritional status of schoolchildren.5. Evaluate the effect of antimalarial IPTsc on school performance.6. Identify the potential environmental and host-related predictors of malaria (re)infections.The study is conducted in the Mokali health area of the Biyela health zone, in Kinshasa province. The health zone of Biyela has recently been identified as highly endemic for malaria and prevalent for schistosomiasis and soil transmitted helminths (STH) (unpublished observations of Lutumba P). Biyela is urban\u2013rural and its population is estimated at 182,421 inhabitants. Biyela has nine subzones of which two are considered rural. Three rivers cross the health zone: Mango, Mabanga and Mokali. Of the nine health areas of Biyela, Mokali is the most populous with a population estimated at 27,455 inhabitants. Biyela has one regional reference hospital that is located in the Mokali health area. The participants in the study are recruited at EP Boyambi and EP Likabo, two of the nearest primary schools of Mokali to the regional health centre. EP Boyambi and EP Likabo were built by the Catholic Church and each school has 14 teachers and 12 classes (two classes per each school year). The estimated number of schoolchildren was 650 per school at the beginning of the year.In order to be eligible, children should satisfy all of the following criteria:1. Male and female primary school children.2. Anticipated local residence for the study duration.3. Signed or thumb-printed informed consent by the parents or guardians and witnessed by an impartial witness (whenever parents/guardians are illiterate).4. Informed consent for children 12-years-old or older.Participants with at least one of the following criteria are excluded:1. Children of the sixth primary school year.2. Participation in any other investigational drug study  during the previous 30 days.3. Known or suspected hypersensitivity or serious adverse events (AE) to the study drugs.4. Malaria symptoms at baseline irrespective of the severity [see Additional file 5. Febrile conditions caused by diseases other than malaria at the first visit.6. Decompensated anaemia.7. Illness or conditions, such as hematologic, cardiac, renal, hepatic diseases, which in the judgement of the investigators would place the subject at undue risk or interfere with the results of the study, including known glucose-6-phosphate dehydrogenase deficiency and sickle cell.8. Body weight <14 kg.9. Children with major chronic infectious diseases .Patients will be excluded from further assessment if any of the following criteria are met:1. Withdrawal of informed consent.2. Severe AE related to the study drug.3. Termination of the study prior to its planned end date by the sponsor, regulatory authorities, or Ethics Committee.Every reasonable effort is made to complete a final evaluation of participants who are withdrawn from the study prior to the planned termination time period. The study staff records the reason(s) for all withdrawals from the study in the participants\u2019 study records. At the time of withdrawal, all study procedures outlined for the End of Study visit should be completed. Any withdrawal or loss to follow-up are documented and notified to the site coordinator as soon as possible.The primary endpoint is the change in mean Hb concentration at month 12 of follow-up and anaemia prevalence one year after the initial preventive treatment.1. Change in mean Hb concentration at month 4 and 8 of follow-up.2. Prevalence of asymptomatic and clinical malaria at baseline and one year after enrolment.pfDHFR and pfDHPS gene mutations at baseline and at month 12 follow-up.3. Prevalence of 4. Clinical (severe) malaria incidence and parasitaemia at month 4, 8 and 12.5. Percentages of acute and severe malnutrition at month 0, 4, 8, 12 through weight/height and height/age z-scores, and skin folds.6. School achievement at the end of follow-up and school attendance.7. Prevalence and risk of environmental and host-related predictors for malaria (re)infections.Subjects are monitored throughout the study for possible development of AE. All AEs are recorded on the specific form in the Case Report Form (CRF). Any changes in relevant laboratory parameters are also assessed. For all AEs, a relative risk (RR) and also the frequency, severity and seriousness will be measured.The study is an unblinded, randomised controlled trial, enrolling asymptomatic school children, either gender. At baseline, month four, and month eight, IPT with SP or SP-PQ is given. SP is administered as a single treatment dose. However, PQ is given as two doses with a 24-hour interval. IPT is administered at four month intervals in line with the long half-lives of the drugs and for compliance. Each treatment arm will be compared to the control group of untreated participants. Due to the prevalence of STH and schistosomiasis in the study area all participants are treated with albendazole or praziquantel according to the WHO guidelines . Before Within each school, each schoolchild is randomly assigned to one of the two intervention groups or the control group as follows: IPT-SP, IPT-SP-PQ and no-treatment. Each study arm consists of one-third of all individuals.To estimate the statistical power of our studies, we used an effect size of 0.56 g/dl mean Hb level improvement by the antimalarial IPT compared to the control treatment. This effect size was based upon a previous study.A previous study estimated the mean Hb concentration of 13.01 g/dl respectively in schoolchildren with IPT and 12.45 g/dl without . The samGeneral information on each school regarding composition, water sources, sanitation, health services, and current and previous treatments are collected by questionnaire. The social, economic and demographic patterns are recorded, physical/clinical and complementary examinations are performed, and study treatments and possible co-medications are given according to the study treatment allocation and the participant\u2019s health status . Any child with a health problem is advised to consult doctors at the regional hospital of Biyela that is the health centre chosen for the study. Parents and guardians are advised to visit the health centre immediately when a child is unwell at home.Clinical malaria cases observed during school visits are documented and referred to the Biyela regional hospital. At the end of follow-up all participants will receive one dose of SP, albendazole and praziquantel and will be passively monitored for three months [See Additional file A child attending the health centre is examined and treated in line with national guidelines. All relevant medical records  of the patient are recorded on the CRF [see Additional file In vitro, sulfadoxine and pyrimethamine are active against the asexual erythrocytic stages of P. falciparum. SP may also be effective against strains of P. falciparum resistant to chloroquine. Fansidar is approved and commercialised for the treatment of uncomplicated malaria and the prevention of malaria.Sulfadoxine\u2013Pyrimethamine is an established drug used for uncomplicated malaria in children and adults and for IPT in pregnant women. The safety and efficacy of SP has been widely established in infants and children in clinical trials. For the current study the Fansidar brand of SP is used. Fansidar is supplied as scored tablets, containing 500 mg sulfadoxine and 25 mg pyrimethamine unit doses. Sulfadoxine and pyrimethamine are folic acid antagonists. Sulfadoxine inhibits the activity of dihydropteroate synthase whereas pyrimethamine inhibits dihydrofolate reductase. Owing to its slow elimination profile, levels of SP sufficient to inhibit parasite growth persist for 60 days after treatment, providing a period of post-treatment prophylaxis during which successful development of a blood-stage infection is prevented. After administration of one tablet, peak plasma levels for pyrimethamine (approximately 0.2 mg/L) and for sulfadoxine (approximately 60 mg/L) are reached after about four hours. The volume of distribution for sulfadoxine and pyrimethamine is 0.14 L/kg and 2.3 L/kg, respectively. Plasma protein binding is about 90% for both pyrimethamine and sulfadoxine. A relatively long elimination half-life is characteristic of both components. The mean values for plasma elimination half-lives are up to 11 days for sulfadoxine and 5 days for pyrimethamine, mainly via the kidneys .Fansidar is given as a single oral dose of \u00bd tablet of 500 mg sulfadoxine - 25 mg pyrimethamine per 10 kg of weigh: 1 tablet for weight less than 20 kg, 1.5 tablets for 20 to 29 kg, and 2 tablets for children who weigh 30 kg or more.P. falciparum malaria [P. berghei K-173, its CT100 of 42 hours and the long elimination half-life [PQ is not approved as a single entity. A fixed dose combination of PQ with dihydroartemisinin (DHA) was approved by the European Medicines Agency and marketed as Eurartesim for the treatment of uncomplicated malaria in children and infants of six months or older. The exact mechanism of action of PQ is unknown. PQ likely mirrors its closest structural analogue chloroquine. Chloroquine binds to toxic haeme (derived from the patient\u2019s haemoglobin) within the parasite and prevents its detoxification. The Cmax and the AUC0-24 of PQ is 179 nm/ml and 1.679 ng/ml*h, respectively. The elimination half-life of PQ is around 20 days for paediatric patients. Due to its slow elimination, PQ may accumulate in plasma after multiple doses. In healthy volunteers, PQ exposure is increased approximately three-fold when administered with a high fat/high calorie meal. In combination curative treatment with DHA, PQ is safe and efficacious at 60 to 73.9 mg/kg dose in three daily doses against uncomplicated  malaria ,30. Howealf-life , PQ 320 Albendazole is a benzimidazole anthelmintic agent highly effective against a wide range of intestinal helminths and, at higher doses, against tissue helminth infections. Albendazole exhibits larvicidal, ovicidal and vermicidal activity, and is thought to act via inhibition of tubulin polymerization. This causes a cascade of metabolic disruption, including energy depletion, which immobilizes and then kills the susceptible helminth. Albendazole is poorly absorbed from the gastrointestinal tract due to its low aqueous solubility. Oral bioavailability appears to be enhanced when albendazole is co-administered with a fatty meal (estimated fat content 40 g) as evidenced by higher (up to five-fold on average) plasma concentrations of albendazole sulfoxide as compared to the fasted state. Plasma concentrations of albendazole sulfoxide increase in a dose-proportional manner over the therapeutic dose range following ingestion of a fatty meal (fat content 43.1 g). The mean apparent terminal elimination half-life of albendazole sulfoxide typically ranges from 8 to 12 hours in normal subjects. Albendazole sulfoxide is 70% bound to plasma protein and is widely distributed throughout the body. Following oral administration, albendazole has not been detected in human urine. Biliary elimination presumably accounts for a portion of the elimination .Albendazole is used in children of 1 to 2 years at one oral 200 mg dose and in adults and children older than 2 years at 400 mg in one oral dose. Albendazole will be given at enrolment and four-month intervals in accordance with the WHO guideline .Praziquantel is a trematodicide used for oral treatment of infections due to schistosome and liver fluke. Praziquantel induces a rapid contraction of schistosomes by a specific effect on the permeability of the cell membrane. The drug further causes vacuolization and disintegration of the schistosome tegument. After oral administration praziquantel is rapidly absorbed (80%), subjected to a first pass effect, metabolized and eliminated by the kidneys. Maximal serum concentration is achieved one to three hours after dosing. The half-life of praziquantel in serum is 0.8 to 1.5 hours. However, in patients with moderate-to-severe hepatic dysfunction (Child-Pugh class B and C), praziquantel half-life, C max, and AUC increased progressively with the degree of hepatic impairment. In patients with schistosomiasis the half-life is 2.99 hour \u00b1 1.28, the T max is 1.48 hour \u00b1 0.74, the C max is 0.83 \u03bcg/mL \u00b1 0.52 and the AUC is 3.02 \u03bcg/mL hour \u00b1 0.59 (34). Praziquantel will be given at 40 mg/kg at enrolment and at the twelve-month follow-up .The administration of paracetamol (acetaminophen) is allowed if the patient\u2019s condition warrants it and should be recorded in the appropriate section of the CRF. Drugs with antimalarial activity  should be reported as concomitant medications. All concomitant medications taken by the patient during the study, from the date of signature of the informed consent are recorded in the appropriate section of the CRF.Safety and tolerability of the treatments are evaluated by recording AEs and grading, laboratory, and vital signs evaluations . All AEsThe investigators assess the drug-event relationship of each AE/SAE by using clinical judgment. Alternative causes, such as natural history of the underlying conditions or diseases, concomitant therapy, other risk factors and the temporal relationship of the event to the investigational product are considered and investigated. The investigator also consults the drug information and the Data and Safety Monitoring Board as needed in the determination of his/her assessment. For every AE, it is very important that the investigator always make an assessment of causality prior to transmission to the Trial Management Group even if the investigator disposes of minimal information to include in the initial report to the Trial Management Group. The investigator may change his/her opinion of causality in the light of follow-up information, amending the SAE case report form accordingly.The outcome of each AE must be assessed according to the following classification: completely recovered, not yet completely recovered, deterioration, permanent damage, death, ongoing, or unknown.All serious AEs, whether or not deemed drug-related, or expected, must be reported immediately or within 24 hours (one working day), using the Serious Adverse Event Notification Form by telefax or email to the sponsor. The report should state \u201cUrgent Serious Adverse Event\u201d and highlight the study code on the cover page. All other AEs not fulfilling the criteria of immediate reporting are recorded on the Case Report Form. This AE information is collected on a regular basis during the clinical trial.The effect of the type of intervention  will all be analysed using linear mixed model analysis. In each analysis, Hb concentration is modeled versus time, testing whether the change of Hb concentration over time differs between the two types of intervention. In each case, a linear mixed model is fitted with Hb concentration as outcome variable, and fixed effects including the type of intervention (indicator variable), time and the interaction between them. In these models, the effect of interest is the interaction between intervention and time, which formally tests the null hypothesis that the change in Hb concentration with time is the same in all interventions. Significance of the interaction term is tested using the F-test with a Kenwardroger correction for the number of degrees of freedom. As the study includes several  measurements within the same individual, and since the individuals cluster within schools, the dependency between individuals will be accounted for by including random effect terms into the model. More particularly, the model will include random effects (intercept and slope) for individual, nested within the random effect of school. An additional advantage of this technique is that it gives unbiased results in case individuals are lost to follow-up during the study.Analysis of the data will be performed using the nlme package from the statistical software R or the proc mixed procedure from the software package SAS. All statistical analyses will be performed in close collaboration with StatUa, the Center for Statistics at the University of Antwerp.The study site maintains the study drugs under adequate security. Study drugs sent to the study centre are verified by the investigator or designee; the amount sent and the exact supplies received are documented by signing and dating the appropriate shipping documents. Study drugs are dispensed as per the randomization list to each patient who meets the enrolment criteria. The investigators or designees record the patient\u2019s number, patient\u2019s initials and date dispensed on the Drug Accountability Form. The investigators maintain an accurate running accountability of the study drugs. At the end of the study, final disposal of the unused drugs will be decided by the sponsor after final reconciliation has been made.All data and observations are documented and entered in the CRF. For this study, the CRF allows identification of the study site and patient; recording of the selection and inclusion of patients in the study; recording of all data collected at each different study visit; recording of possible AE and any suspension of the study. Participants are identified by their initials and a study identification number on the CRF. When recording data on the CRF and entering the data in the database, confidentiality and security of the forms are ensured at all steps. All corrections are made on CRFs by striking through the incorrect entry with a single line and entering the correct information adjacent to it. The correction is initialed and dated by the investigator. Additional records are kept in the clinical and laboratory record books at the core facility in Kinshasa. The investigators verify data at each study visit.Children fulfilling the inclusion/exclusion criteria will be assigned a sequential study number and will be treated according to a predetermined randomization list of blocks of varying size per school. Within each block each schoolchild will randomly be assigned to one of the two intervention groups or the control group in 1:1:1 balance. Sealed envelopes labeled with the school unique code and containing the list with the treatment allocated to the participants will be provided according to the above mentioned list and opened at the moment the children are recruited.The study treatment is administered under the direct supervision of a study nurse. Study drugs are given to the children consistent with the investigational brochure of each product. After drug administration, children will be kept for thirty minutes in the class. In case vomiting occurs within 30 minutes of administration a dose is repeated in full. This event is documented in the CRF. If vomiting persists beyond two additional doses, the child is withdrawn from the study.Upon arrival at a school, an inventory is performed and a drug receipt log filled out and signed by the staff. Accountability for the drug at all school sites is the responsibility of the principal investigator. The investigators ensure that the drug is used only in accordance with this protocol. Accountability records include dates, quantities, lot numbers, expiration dates, and patient numbers.To ensure the rights, safety and well-being of the human subjects enrolled in this trial and to validate the Investigator\u2019s adherence to the protocol, all applicable regulations of the International Conference on Harmonisation (ICH) and the Good Clinical Practices (GCP) guidelines are strictly applied. The quality assurance of records and data are guaranteed. The study clinicians are trained about the study protocol prior to the onset of the trial. The study clinicians complete case records at each hospital visit. The medical records of the clinicians are transferred to a CRF by the investigators. At school visits, the investigators record data on the CRF. The investigators carry out 100% Source Data Verification on the data collected at the regional hospital centre during the passive follow-up to guarantee the best conduct of the study. To ensure accuracy of the data, a CRF completed by an investigator is contra-checked by another investigator. The consistence of data of each participant is checked by the investigators at each follow-up visit by cross-checking the reports of the school teachers and of parents/guardians, and the medical records at the regional hospital centre. The investigators, therefore, have frequent contacts with the clinicians and the school teachers.All data is processed from the CRF into an electronic database by two different persons after each follow-up visit. Then the two files are merged. Unmerging information is verified and corrected and added to the final database. During the conduct of the study, data are verified and reviewed to produce and maintain high quality data. All unresolved issues are queried and resolved before locking the database. Data transfer and handling are done with appropriate security measures and with regard to rights, safety and well-being of trial subjects. Two back-up files of the database are stored on compact discs after each data entry session. For quality control, check programs are written into the database to limit the entry of incorrect data and ensure entry of data into required fields.The Investigators agreed to conduct the present study in full agreement with the principles of the 'Declaration of Helsinki\u2019 and subsequent relevant amendments. All research activities will be conducted in accordance with the standards and codes of conduct accepted by the ICH guidelines. The study is approved by the Ethical Committees of the University of Antwerp, Belgium and of the School of Public Health, University of Kinshasa, DRCongo. Approval was also obtained from the Ministry of Health of DRCongo. Prior to the start of the project, special permission was obtained from the DRCongo Ministry of Education and the local health and education authorities of Biyela. Written informed consent was obtained from the guardians for all children before entering the study. An assent was obtained, as well, from children who were 12-years-old or older. In the control arm no treatment is given as no foreseeable serious risk can be anticipated and no standard of care yet exists in schoolchildren. All participants are followed-up, examined and treated free of charge if malaria or other illnesses occur during the period of the study.P. falciparum parasitaemia, finger pricks are done at each visit for hemocontrol and microscopy. A total of 3 ml of venous blood is collected at baseline and month 12 for genotyping and other tests that the clinician may request for the management of illnesses. Residual plasma is separated, stored and transported at -70\u00b0C to be assayed by ELISA and fluorescence activated cell sorting (FACS) for malarial cytokines and antibody quantification and other malaria-related parameters . The human genome will not be examined.To measure Hb level and et al.[et al. [et al. [Little research has been performed on IPTsc in malaria stable transmission. Rohner et al. found no.[et al.  found a  [et al.  reportedOur clinical trial is unique in investigating the impact of antimalaria IPT in schoolchildren of hyper endemic areas for longer follow-up, resistance markers for SP at baseline and after 12 months (including break through parasitaemias). Our study is also unique in using piperaquine in combination with SP. Artemisinin derivative has been excluded as it is part of the treatment policies in virtually all malaria endemic countries .The findings of the clinical trial are, therefore, expected to make a major contribution to the public health of youngsters who fail to thrive and grow due to multiple morbidities.Recruiting.Written informed consent was obtained from the participants\u2019 parents or guardians for the conduct of the study and the publication of this manuscript. As part of the clinical trial protocol, an information sheet for participants and an informed consent form were reviewed and approved by the Ethical Committee of the University of Antwerp, Belgium, the Ethical Committee of the School of Public Health, University of Kinshasa, DRCongo and the Ministry of Health of DRCongo. A copy of the information sheet and the written consent form in French, English and in the local language are available for review by the Editor-in-Chief of the Trials Journal.AE: Adverse events; CRF: Case report form; DHA: Dihydroartemisinin; DHFR: Dihydrofolate reductase; DHPS: Dihydropteroate synthase; ELISA: Enzyme-linked immunosorbent assay; FACS: Fluorescence activated cell sorting; GCP: Good Clinical Practice; H/A: Height for age; ICH: International Conference on Harmonisation; IPT: Intermittent preventive treatment; IPTc: Intermittent preventive treatment in children; IPTi: Intermittent preventive treatment in infancy; IPTp: Intermittent preventive treatment in pregnancy; IPTsc: Intermittent preventive treatment in schoolchildren; LIC: Low income countries; MIC: Minimal inhibitory concentration; PCR: Polymerase chain reaction; PQ: Piperaquine; RCT: Randomised controlled trial; RR: Relative risk; SAE: Serious adverse events; SP: Sulfadoxine-pyrimethamine; SP-PQ: Sulfadoxine-pyrimethamine plus piperaquine; SSA: Sub Saharan Africa; STH: Soil transmitted helminths; W/H: Weight for height.The research is funded by the Flemish Interuniversity Council (VLIR-UOS) and the Research Foundation - Flanders (FWO) of Belgium. The authors declare that they have no competing interests.JYD and JPVg conceptualised the idea. JYD wrote the study protocol and the trial essential documents. JM contributed to the essential documents writing. JYD and JM did the approval procedures of the protocol by ethical committees and regulatory authorities. JYD and JM implement the study protocol in the field. PL and JPVg reviewed the protocol and supervise the conduct of the trial. JPVg carries the sponsorship of the study. All authors read and approved the submitted manuscript.JYD is a Medical Doctor, epidemiologist and drug regulatory consultant. He is affiliated to the International Health Unit, Department of Epidemiology and Social Medicine, Faculty of Medicine and Health Sciences, University of Antwerp. JM is a Medical Doctor, currently MSc student. He is affiliated to the Parasitology Department, Faculty of Medicine, University of Kinshasa. PL is a Medical Doctor, epidemiologist, PhD. He is professor at the Parasitology Department, Faculty of Medicine, University of Kinshasa. JPVg is a Medical Doctor, epidemiologist, PhD. He has been involved for a decade in RCTs in Low Income Countries. He is heading the International Health Unit, Department of Epidemiology and Social Medicine, Faculty of Medicine and Health Sciences, University of Antwerp.WHO criteria for malaria diagnosis (WHO 2010).Click here for filePatient flow at school visit.Click here for filePatient flow at hospital visit.Click here for file"}
+{"text": "Plasmodium falciparum infection in Afghanistan in 2003.Combination therapy with artesunate plus sulfadoxine-pyrimethamine (SP) was adopted as recommended treatment for P. falciparum were undertaken in sentinel sites in Afghanistan from 2007 to 2014, accompanied by relevant molecular studies. The first study was a randomized trial of AS\u00a0+\u00a0SP versus dihydroartemisinin-piperaquine, while two subsequent studies were standard therapeutic efficacy studies of AS\u00a0+\u00a0SP.A series of prospective clinical studies examining the efficacy of artesunate plus sulfadoxine-pyrimethamine (AS\u00a0+\u00a0SP) against dhfr indicated fixation of the S108\u00a0N mutation and a prevalence of the C59R mutation of approximately 95\u00a0% across all sites. Other mutations in dhfr and dhps remained rare or absent entirely, although five isolates from the first trial carried the dhps triple mutant SGEGA haplotype. In the last study undertaken in 2012\u20132014 the K13 artemisinin resistance marker was examined; only two of 60 successfully sequenced samples carried a K13-propeller mutation.Three hundred and three patients were enrolled across four provinces in the north and east of the country. Curative efficacy was high in all the trials, with an adequate clinical and parasitological response (ACPR) of more than 95\u00a0% in all groups and trial stages. Genotyping for drug-resistance alleles at P. falciparum in Afghanistan. The molecular data indicate that despite a substantial fall in incidence, resistance has not developed to artemisinins, or intensified to the ACT partner drug components.These data confirm maintained efficacy 10\u00a0years after introduction of artesunate plus SP as combination treatment of Trial Registrationhttp://www.clinicaltrials.gov/ct NCT00682578, NCT01115439 and NCT01707199 Plasmodium falciparum transmission occurs at its most northerly latitudes in Afghanistan, where the distribution of cases is determined by the physical geography of the country, malaria being confined to lower lying areas with sufficient rainfall for mosquito survival. These consist of the northern plains [n plains , the Jaln plains . The cenn plains , 4. The n plains . FollowiP. falciparum was evident in Afghanistan and Pakistan by the 1990s [pfcrt haplotype [P. falciparum infection in a number of south Asian countries, becoming first-line treatment in Afghanistan (2003), Iran (2006), Pakistan (2007) and India (2007).Chloroquine-resistant he 1990s \u20139 and thhe 1990s  consisteaplotype . Howeveraplotype  and nortaplotype , findingP. falciparum in sentinel sites in Afghanistan from 2007 to 2014, accompanied by relevant molecular studies. These confirm maintained efficacy of the combination against P. falciparum with no evidence for increase in prevalence of molecular markers of artemisinin and SP resistance.This paper describes a series of therapeutic efficacy studies undertaken to examine the efficacy of AS\u00a0+\u00a0SP against Studies were undertaken at, or within 20\u00a0km of, four provincial control centres, two located to the north and two to the south and east of the Hindu Kush mountains. Jalalabad, capital of Nangarhar province, is a referral centre for the whole eastern region and lies adjacent to the Pakistani border where population movement in both directions takes place. The population is multi-ethnic with Pashtoon being the dominant group. The province as a whole is generally semiarid but where possible rice is cultivated. Asadabad is the capital city of Kunar province, a largely mountainous area bordering Pakistan, with which there is uncontrolled movement via unofficial borders. Taloqan is the capital of the north-east province of Takhar with a population that is mainly Tajik with significant proportions of Uzbek and Pashtoon. Rice is cultivated on the plain and malaria is a well-known health problem . MaimanaP. falciparum. Additional study inclusion criteria were the ability to swallow oral medication, ability and willingness to comply with the study protocol for the duration of the study and written informed consent (from a parent or guardian in the case of children under 18\u00a0years of age). In trial 1 there was no lower limit of asexual parasitaemia and patients with parasitemia of more than 100,000/\u00b5L were excluded. In trials 2 and 3 only patients with 500\u2013150,000/\u00b5L asexual forms were included.Trial 1 was a prospective, open label, parallel group randomized trial with a 1:1 allocation ratio of AS\u00a0+\u00a0SP to dihydroartemisinin-piperaquine (DP) undertaken in all four centres. Trials 2 and 3 were therapeutic single-arm efficacy studies of AS\u00a0+\u00a0SP undertaken in Jalalabad (Nangarhar province), and Asadabad (Kunar province) respectively. In all trials eligible participants were adults and children over 6\u00a0months old presenting with symptomatic, uncomplicated, microscopically confirmed monoinfection with asexual stages of In all trials exclusion criteria included the presence of general danger signs in children aged under 5\u00a0years or signs of severe falciparum malaria according to the definitions of WHO , mixed mStandard case record forms were used to record demographic information, details of symptoms and their duration, and previous antimalarial medications. Clinical examination findings and vital signs were documented, including axillary temperature measured with a digital thermometer. Blood samples were obtained for haemoglobin measurement and Giemsa-staining of thick and thin blood films to confirm parasite species and parasitaemia by a standard approximation method (40\u00a0\u00d7\u00a0number of parasites per 200 white blood cells on the thick film). Slide examination results were considered to be negative after examination of 200 oil fields of the thick film. 20\u2013100\u00a0\u00b5l baseline blood was collected on filter papers for subsequent confirmation of species by PCR and testing of drug resistance markers.For patients receiving AS\u00a0+\u00a0SP, each drug was dosed to the nearest half tablet, determined from a dosing chart  and broaPatients receiving DP  received Duo-Cotecxin\u00ae  in which each co-formulated tablet contained 40\u00a0mg dihydroartemisinin and 320\u00a0mg piperaquine phosphate. The target total dose of 6/48\u00a0mg/kg was made up of three divided doses given on a daily basis. According to the body weight, individual doses were rounded to the nearest quarter tablet.All doses of medicine were administered under the supervision of a qualified member of the staff designated by the principal investigator. The study patients were observed for 30\u00a0min after medicine administration for adverse reactions and/or vomiting. Patients who vomited during this observation period were re-treated with the same dose of medicine and observed for an additional 30\u00a0min. Patients vomiting again were to be given parenteral  therapy with quinine sulphate (10\u00a0mg base per kg body weight three times per day), along with once daily tetracycline (clindamycin in children) according to national malaria treatment guidelines, and withdrawn from further study.P. vivax). Patients failing to attend follow-up visits received a visit at home and were asked to re-attend the centre for scheduled assessments. Transportation was provided for all follow-up visits for all subjects. All enrolled patients were given one long-lasting insecticide-treated bed net.Subjects were seen on days 0\u20133 inclusive and weekly to 56\u00a0days  or 42\u00a0days . Patients were also requested to attend if they felt unwell on any other day during follow-up. At each follow up visit a clinical assessment was carried out and the patient was asked about adverse events by the use of a standard symptom questionnaire. Thick and thin blood slides were examined and haemoglobin was measured. Other tests such as peripheral white blood cell count and urine examination were undertaken as indicated to rule out other conditions and investigate possible adverse effects of drugs. Patients were censored from the analysis if there was stated withdrawal of consent at any stage, persistent vomiting during the acute phase  or occurrence during the follow-up of concomitant disease that would interfere with a clear classification of the treatment outcome  for 7\u00a0days with standard follow up.Patients with recurrence of The primary outcome was the proportion of patients with PCR-corrected adequate clinical and parasitological response (ACPR) at day 42 . SecondaIn trial 1 the aim was to examine whether DP was as effective as AS\u00a0+\u00a0SP via a non-inferiority design, given the relatively high predicted efficacy of AS\u00a0+\u00a0SP ; the nonTrials 2 and 3 were single arm monitoring studies and a sample size of 100 was chosen in each case consistent with WHO guidelines for therapeutic efficacy studies .Patients were allocated to the two treatment arms based on a randomization list created using Stata 9.0 statistical software  with a 1:1 allocation using a block size of 20 that was produced and held independently of the field teams by a statistician. The individual treatment allocations were kept in sequentially numbered, opaque, sealed envelopes and opened by the relevant field worker only after participant enrolment by the field team. Patients and clinical field workers were not blinded to the treatment arm after allocation. Laboratory workers assessing blood films and genotyping were not informed of the treatment arm.Data were entered into Microsoft Access or the WHO Study Information Sheet for calculation of baseline characteristics. Subsequent analyses were conducted using STATA  and GraphPad Prism . The Student\u2019s t test, Mann\u2013Whitney U and Chi squared (or Fisher\u2019s exact) tests were used for comparison of baseline variables between the two treatment arms of trial 1 and Wilson\u2019s test was used for calculation of confidence intervals for ACPR.Day 7 blood samples were collected for drug measurements in Nangarhar and Kunar provinces in year 3 of trial 1, during the period September to December 2009. Approximately 100\u00a0\u00b5l capillary blood was collected onto Whatman 3MM filter papers and stored individually with silica gel for transportation to the Department of Clinical Pharmacology at MORU. Piperaquine concentrations (DP arm), adjusted for estimated sample volume, were determined by liquid chromatography and tandem mass spectrometry as described previously , 18. SulP. falciparum according to established methods. In order to differentiate a recrudescence from a reinfection, a genotype analysis was conducted based on the extensive genetic diversity in the polymorphic genes msp1, msp2 and glurp. The genotypic profiles of pre- and post-treatment strains were compared by standard methods [Blood samples were collected on filter paper (Whatman 3MM) at enrollment and at any visit where parasites were observed after day 7. Each filter paper was dried and stored individually in a plastic bag containing silica gel. All filter papers were subsequently transported to the Faculty of Tropical Medicine, Mahidol University. Parasite DNA from dried blood spots was extracted via the QIAmp DNA Mini kit using the standard protocol and stored at \u221220\u00a0\u00b0C until use. Nested PCR was performed to detect pfdhr, pfdhps, pfcrt and pfmdr1 single nucleotide polymorphisms (SNPs) and pfmdr1 copy number assessment using previously described techniques [pfdhr and pfdhps alone were examined and in trial 3 the artemisinin resistance marker K13 propeller [PCR\u2013RFLP and sequencing was undertaken for baseline samples in trial 1 examining chniques , 22; thechniques . In triaropeller  was sequropeller . HeterozAll trials were approved by the Oxford Tropical Research Ethics Committee, Oxford University, UK  and the Institutional Review Board of the Afghanistan National Public Health Institute, Ministry of Public Health, Afghanistan. In addition trial 1 was approved by the Ethics Committee of the Faculty of Tropical Medicine, Mahidol University, Thailand and trials 2 and 3 were approved by the Ethical Review Committee of the WHO.P. falciparum transmission seasons, with three withdrawing before completion of treatment. Given evidence of falling transmission, it was decided not to continue recruitment into a third season. Overall follow-up rates to day 42 were more than 90\u00a0%  or AS\u00a0+\u00a0SP (N\u00a0=\u00a061); one patient in the DP arm withdrew after two doses of treatment   and at day 2 in 3 of 297 patients (1\u00a0%) with no patients remaining positive at day 3. In the randomized trial of DP vs. AS\u00a0+\u00a0SP, there were no differences between the groups in terms of the proportion of patients with gametocytes or fever (temperature more than 37\u00a0\u00b0C) after treatment  for piperaquine were 98.5\u00a0\u00b1\u00a073.3\u00a0ng/mL (n\u00a0=\u00a020 patients), although in two cases levels were less than the limit of detection (0.75\u00a0ng/mL). There was a non-significant positive correlation between piperaquine concentration and the body weight-normalized total dose . Day 7 sulphadoxine levels were below the limit of detection (0.5\u00a0\u00b5g/mL) in nine of the 17 patients studied (spanning a range of ages), a finding that is challenging to correlate with the high measured efficacy.dhfr indicated fixation of the S108\u00a0N mutation and that approximately 95\u00a0% of parasites carried the C59R mutation .P. falciparum infection in Afghanistan, provide very reassuring evidence of maintained efficacy with rates of ACPR well over 95\u00a0% in all studies. Accompanying assessments of molecular markers explain these clinical responses. As in the rest of Asia and much of sub-Saharan Africa, two polymorphisms in dhfr (59R and 108\u00a0N) remain at or near fixation, but no other dhfr mutations are found in Afghanistan. A previous report generated concerns over the prevalence of the dhps triple mutation 437G/540E/581G in Nangarhar and Kunar provinces  [Prospective trials of the clinical efficacy of AS\u00a0+\u00a0SP, spanning the decade following its introduction as first-line recommended treatment for s paper)  but the P. falciparum does not yet appear to have been significantly compromised by molecular changes in any of these locations. Only in northeast India is there evidence for a high prevalence of \u2018quintuple\u2019 mutation haplotypes based on these two genes [Compared to this, neighbouring countries show relatively more concerning resistance marker profiles. Surveys from Pakistan within the last decade show significant numbers of parasites mutated at DHPS 437G \u201328 as wewo genes  in assocThis is the first study to examine the K13 artemisinin resistance marker in Central Asia. Only two samples (less than 5\u00a0%) contained a mutation in the propeller domain; Y493C and S700L. Neither mutation has been previously reported, although a different mutation at 493, Y493H, is common and causally linked to artemisinin resistance in Cambodia , 34. Thedhps gene or increasing mutations in dhfr, is to some degree reminiscent of the return of in vitro mefloquine sensitivity following addition of artesunate to mefloquine at the Thai\u2013Myanmar border in the 1990s [The maintained efficacy of AS\u00a0+\u00a0SP, with no evidence of adaptive evolution in the he 1990s . While uhe 1990s , it is nP. falciparum for a substantial period of time as Afghanistan moves towards elimination.AS\u00a0+\u00a0SP, the first-line treatment for falciparum malaria in Afghanistan, remains efficacious therapy for uncomplicated falciparum malaria, both in terms of initial parasite clearance and cure. Accompanying molecular data indicate a stable situation with regard to SP resistance, and suggest that AS\u00a0+\u00a0SP is likely to retain a high level of efficacy as treatment for"}
+{"text": "Between 2011 and 2013 the number of recorded malaria cases had more than doubled, and between 2009 and 2013 had increased almost 4-fold in MSF-OCA  programmes in the Democratic Republic of the Congo (DRC). The reasons for this rise are unclear. Incorrect intake of Artemisinin Combination Therapy (ACT) could result in failure to treat the infection and potential recurrence. An adherence study was carried out to assess whether patients were completing the full course of ACT.One hundred and eight malaria patients in Shamwana, Katanga province, DRC were visited in their households the day after ACT was supposed to be completed. They were asked a series of questions about ACT administration and the blister pack was observed (if available).Sixty seven (62.0%) patients were considered probably adherent. This did not take into account the patients that vomited or spat their pills or took them at the incorrect time of day, in which case adherence dropped to 46 (42.6%). The most common reason that patients gave for incomplete/incorrect intake was that they were vomiting or felt unwell (10 patients (24.4%), although the reasons were not recorded for 22 (53.7%) patients). This indicates that there may be poor understanding of the importance of completing the treatment or that the side effects of ACT were significant enough to over-ride the pharmacy instructions.Adherence to ACT was poor in this setting. Health education messages emphasising the need to complete ACT even if patients vomit doses, feel unwell or their health conditions improve should be promoted. MSF-OCA operates health programmes in Mweso and Walikale in North Kivu; Baraka and Kimbi Lulimba in South Kivu; and Shamwana, Katanga (DRC).Between 2011 and 2013, the number of recorded malaria cases more than doubled and between 2009 and 2013, increased almost 4-fold in MSF-OCA programmes in DRC . The numIn Katanga the general trend is less clear for malaria cases. However this is likely due to the variable numbers of projects supported between 2009 and 2013 . FocussiIn 2013 MSF clinic staff in Shamwana began to identify patients returning to the clinic with complicated severe malaria after having already been prescribed the anti-malarial treatment. It was not known if these were treatment failures or re-infections.This was an alarming increase in malaria incidence which cannot be explained by displacement of non-immune populations from non-endemic regions (mountains) to endemic malaria regions as the populations have been relatively stable since 2008.3. These strategies work against both the transmission of the parasite from mosquito vector to humans (and from humans to mosquitoes) and the development of illness and severe disease in humans3. The former is achieved in Shamwana through the distribution of long-lasting insecticide-treated bednet (LLIN) in a targeted manner  and the latter through Artemisinin Combination Therapy (ACT) of confirmed malaria cases  administered in a fixed dose combination (FDC) as a first-line ACT). These strategies do not seem to be having an impact on malaria incidence in MSF settings in DRC.The World Health Organisation (WHO)-recommended strategies for malaria control fall into two major areas: prevention and case managementThe current study explored whether this increase in cases could be explained by patients failing to complete the full three day course and adhering to the time schedule and dosage prescribed for ASAQ. Poor adherence increases drug pressure and thus the risk of developing resistance. Moreover, incorrect or incomplete intake could result in failure to treat the infection and potential recurrence.The study was conducted in Shamwana, Katanga, DRC. Katanga has an average temperature of 24\u00b0C and two seasons, rainy between November and March and dry from April to October. MSF-OCA has been working in the province of Katanga since 2003. MSF-OCA supports Shamwana hospital to provide free secondary health care and supports more than four health centres. The MSF catchment population included Shamwana town and the nearby village of Nsangwa.9. The sample size was therefore based on the conservative estimated adherence of 50% (in order to obtain the maximum sample size). With a precision of 10% and an \u03b1-error of 5%, n=97 patients were required. Assuming a proportion of patients lost to follow-up or withdrawn of 20%, the total sample size needed for the home visits was thus n=117 patients. Similarly, a sample size of 117 was calculated for the exit interviews.For previous ACT adherence assessments, adherence was always approximately 60%To prevent bias due to a change in prescribing and educating behaviour by health staff, the health staff were informed that a study would start but the exact purpose of the study (measuring patient adherence) was at no time revealed to them. The explanation was that this was a mapping exercise of where patients attending the health centre were mainly coming from.At the health centre level, all patients were interviewed using a short \u201ccentre\u201d-questionnaire. This included the patient\u2019s name, age, sex, the number and type of prescriptions, and the patient's address in sufficient detail.Study patients for the \u201chome\u201d-questionnaire were selected through systematic sampling of eligible patients  result that had been prescribed ASAQ) from the \u201ccentre\u201d-questionnaires. Study patients were visited at home on Day 3  and interviewed using the \u201chome\u201d questionnaire to assess patient adherence. Socio-demographic questions about the household were followed by a systematic account of how the pills were taken. A correct treatment schedule instrument was used to guide the questions on adherence. Lastly, some questions about knowledge of malaria cause and prevention were asked. Any sick patients were referred to the health centre.After all \u2018home\u2019-questionnaires were completed, exit interviews were carried out at the health centre using the \u201cexit\u201d-questionnaire to provide further information on patient/caretaker understanding of ACT and pharmacy dispensing practices.A dose was considered correctly taken if according to the patient or caretaker\u2019s verbal account the prescribed amount of pills was taken and it was neither spat out nor vomited.There are no standard definitions for adherence so the following were used .Certain incomplete intake or certain non-adherence: Any patient that showed blister packaging still containing any ASAQ pills was certain to have taken an incomplete treatment. There was absolute certainty that the treatment schedule was not completed.Probable incomplete intake or probable non-adherence: According to the patient/caretaker\u2019s account, all pills were not taken so this patient was probably non-adherent. The blister packaging presented was empty or could not be shown.Probable incorrect intake or probable non-adherence: If according to the patient/caretaker\u2019s account ASAQ was not taken following the prescribed time schedule or dosage, it was assumed the patient took the complete treatment but in an incorrect way. The blister packaging presented was empty or could not be shown.Probable correct intake or probable adherence: Only those patients who stated they had taken the complete treatment schedule, following the treatment protocol exactly were considered \u201cprobably adherent\u201d (this was not \u2018certain\u2019 as each dose had not been observed). The blister packaging presented was empty or could not be shown.10. Data were summarised by number and percentages for categorical variables. Continuous variables were summarised using means, medians and ranges. Adherence status was calculated and expressed as a percentage. Confidence intervals were calculated using binomial and multinomial formulae. To calculate the associations between risk factors and adherence status logistic regressions was used and the results expressed as odds ratios and 95% confidence intervals. Data analysis was conducted using Stata 13.011.Data for each subject were entered into EpiData 3.1 softwareIn total, 274 people completed the \u2018centre\u2019 questionnaires. Among those, 150 (54.7%) were diagnosed with malaria and given a prescription for ACT. Of these malaria patients 17 were aged <1 year so were excluded. Onehundred and seventeen (42.7%) of the 133 (48.5%) patients that satisfied the inclusion criteria were selected for home visits as these were the maximum that could be achieved during daily visits. However, only 108 (39.4%) of these eligible patients completed home questionnaires. The remaining 9 (3.3%) patients were considered lost to follow-up . A totalThe mean age of the patients was 10.5 years and ranged from 1 to 57 years . The mean number of children aged <5 years per household were 2.6 and ranged between 0 and 13  . The maiA total of six (5.6%) patients had one or more pills left at the time of the home visit on Day 3 when all pills should have been taken . They weHowever it should be noted that in order to analyse these data, assumptions had to be made for missing data, primarily an assumption had to be made for 25 patients on Day 0 that ASAQ had been taken only once that day (at the clinic) and had been taken at the same time as the dose on Day 1 and Day 2 .This adherence was calculated based on the number of pills taken, the number of times pills were taken per day and the number of days completed. However the timing of the daily dose and whether the medication was retained (not vomited or spat out) are also important. Both could result in a reduction of anti-malarial activity during the treatment phase and potential treatment failure. Using this stricter definition of adherence, 21 patients that had been defined as probably adherent became probably non-adherent (data not shown). Twelve (57.1%) of these patients did not take doses at the correct time of day, eight (38.1%) patients vomited or spat their pills and one (4.8%) patient vomited or spat their pills and also did not take doses at the correct time of day.Reasons for 1) pills remaining or 2) a description of incorrect intake or 3) a description of correct intake were noted for each patient. However, 22 (62.9%) of the 35 probably non-adherent patients had missing data for reasons for incorrect intake and 19 of the 67 (28.4%) probably adherent patients had missing data for reasons for correct intake.This analysis only considered the individuals classified into the appropriate categories (non-strict adherence). The main reason for pills remaining (incomplete treatment intake) were forgetting to give/take the treatment and patient didn\u2019t feel better/treatment wasn\u2019t working + felt unwell  . UnfortuThe most common reasons for incorrect intake were that the patient forgot to take/caretaker forgot to give the pills, claimed the wrong instructions were given at the clinic or the patient was vomiting  .Taken together the most common reason patients gave for incomplete and/or incorrect intake was that they were vomiting or felt unwell (10 patients (24.4%)).The most common reasons for correct ACT intake was that the correct instructions were given at the clinic  and the patient/caretaker had been given the same medicine before and knew how to take it + Correct instructions were given at the clinic .2 = 8.1, P= 0.017,2 = 8.9, P= 0.031,2 = 8.1, P= 0.004 for non-strict adherence only,Univariate analyses of potential risk factors for non-adherence were explored (using the non-strict adherence classification). These included sex, weight classes (instead of age groups), education level of the patient, knowledge that mosquito bites can cause malaria, knowledge that LLINs prevent malaria and presence of observed LLINs in the household. Significant association with adherence was observed only for sex .2=9.34, P=0.026 for age group and \u03c72=9.16, P=0.05 for weight class).The weight categories showed significantly higher infants and adults in the exit group  knew the name of the disease or could name the correct signs/symptoms of malaria . SimilarHowever, the Outpatient Department (OPD)/pharmacy performance was not consistent. Despite 107 (91.5%) of the patients/caretakers being asked if they had understood the instructions given to them, only 41 (35.0%) had been asked to repeat these instructions and only 90 (76.9%) were given additional information, mainly advised to return to the clinic to get a dose if they vomited or spat up a dose  and informed that ACT can cause fatigue but they must continue the treatment . Also a high but not optimal number of patients  had taken a dose of ACT at the clinic under supervision.This study found that six (5.6%) patients were certainly non-adherent; therefore approximately one out of 18 patients did not complete the course of ACT treatment within the timeframe of 3 days. From the verbal account of the patients or caretakers, another 35 (32.4%) patients took the treatment in a different way to how it was prescribed (probable non-adherence). Therefore, only 67 (62.0%) patients could be considered probably adherent.12.This did not take into account the patients that vomited or spat their pills or took the pills at the incorrect time of day. In that case the adherence dropped to 46 (42.6%). The effect of a decrease in anti-malarial activity that would follow vomiting/spitting or failure to take a timely dose is not clear but it is thought that this risks a period of monotherapy to the partner drug in the ACT combination. Advice about timing of ACT dosages is not clear but it is thought that a 3-hour margin either side of the expected timing is acceptable8.The most common reason patients mentioned for incomplete (67%) and incorrect (20%) intake was that they were vomiting or felt unwell. This appears to indicate that there was a poor understanding of the seriousness of the disease and the importance of finishing the treatment or that the side effects of ASAQ treatment were significant enough to over-ride the pharmacy instructions, unlike previous MSF studies using Coartem\u00ae13.More than a third of the study population claimed not to know the cause of malaria nor how to prevent malaria during home visits, but this was not significantly associated with non-adherence to ACT. Only sex of the patient was associated with adherence . As the side effects were unlikely to affect males more than females, this may demonstrate an indifference to malaria in the male population, a common disease seen regularly in Shamwana. Similarly in Ethiopia, a study of factors associated with non-use of bed nets found that malaria was not perceived by the population to be a problem despite high prevalence of the diseaseWhen assessing the main reason for correct ACT intake, 45 (66.2%) patients claimed they were given the correct instructions at the OPD/pharmacy. When assessing the exit-questionnaires the quality of ACT prescriptions and the related instructions given at the OPD/pharmacy were less clear: 86.3% of the patients in the exit-questionnaires group were able to identify ACT as the anti-malarial treatment given to them amongst other concomitant treatments and 99.2% could correctly repeat the number of days that ACT should be taken. However, only 74.4% could repeat the number of times per day pills should be taken and 65.0% could repeat the number of pills to take.7. Knowledge that mosquito bites cause malaria did not show a statistically significant association with ACT adherence unlike findings of an ACT adherence study in Bo, Sierra Leone8. No association was seen between adherence and age despite some observations in MSF projects that adherence is poor amongst adolescents.Unfortunately the education level of the patients only was assessed, and not that of the caretakers. Therefore, any link between education level and adherence to antimalarials, which have been shown previously to be associated, could not be deduced8. These study settings had similar characteristics to Shamwana, such as a remote rural study area, low education level, treatment intake over 3 days, and treatment already implemented for a certain amount of time. Furthermore, our results are in line with a study carried out in two districts in Western Kenya that found a certain non-adherence of 31.7% and probable non-adherence of 4.2%14 and another study in rural Sri Lanka using post-treatment interviews where 26% of patients self-reported defaulting. The main reasons given for not taking the entire regimen were side effects and disappearance of symptoms15.Three ACT adherence studies carried out in MSF malaria projects had similar findings; certain non-adherence was 22.9% and probable non-adherence 28.8% in a rural area of Sierra Leone, 18% and 29% in a remote area in South Sudan and 21% and 39% respectively in a refugee settlement in Zambia respectively16. In Tanzania, non-adherence was as low as 25%17. These higher adherences might be explained by the fact that adherence assessments were carried out after the successful introduction of a new malaria treatment. The introduction of a new, efficient treatment is always accompanied by special training for health workers, which is linked with greater motivation, and giving a more detailed verbal explanation to patients.However, studies in Uganda and Ghana showed certain non-adherence to be as low as 7% and probable non-adherence to be 3%18. However all had varying definitions of adherence and used different study designs and methods to measure adherence. Standardised methodologies for both self-report and bioassay measurements would improve the evidence base on ACT adherence and effectiveness.A recent systematic review found 37 studies that measured ACT adherenceTwo different methods were used to measure adherence in patients: the pill count (observation of the blister pack) and a systematic questionnaire. Both assessment methods have their limitations: the pill count is a more objective measurement but gives incomplete information; the patient\u2019s account is subjective and not verifiable but provides more information. By classifying patients as either certainly or probably non-adherent, the study accounted for the limitations of both methodologies.19. Combination treatment with ACT when taken correctly reduces, but does not completely eliminate, the chance of resistant strains developing. Poor adherence could potentially expose greater numbers of parasites to the more slowly-eliminated partner drug in ACTs, increasing the risk of resistance to the partner drug20. Once resistance has developed to the partner drug the ACT treatment would effectively become artemisinin monotherapy and hence render artemisinin vulnerable to resistance.This study has shown that ACT adherence in this setting is inadequate and may have contributed to the increase in malaria cases (if recrudescence was a major cause). It should be noted however that there is no recommended target for adherence levels. Artemisinin is the best drug available to treat malaria and currently there are no real alternatives. Artemisinin resistance has likely arisen in Cambodia due to the use of artemisinin monotherapyThe effectiveness of ACT relies on both the efficacy of the drug components and on correct compliance. Adherence to ACT should not be taken for granted. At the community level, health communication campaigns that improve malaria knowledge and help them understand the importance of prevention and correct treatment should be carried out. At the clinic level, first ACT dose should always be observed at the pharmacy/OPD clinic and complete and clear patient instructions, including the importance of completing a course of ACT treatment and what to do if a dose is vomited or spat, should be given. The need to complete three sequential days of malaria treatment even if patients feel unwell or improve should be emphasised at both the community and clinic level.Written informed consent was obtained from the patient/parent/caretaker for the home questionnaires.Verbal informed consent was obtained from the patient/parent/caretaker for the exit questionnaires. The authors should be congratulated for conducting this study in difficult circumstances. Given its very large experience, MSF must be encouraged to contribute more to the fight against malaria, with this type of operational research.I have several comments and suggestions with regards to this study:The authors wanted to investigate the reasons behind the rise in the number of malaria cases seen in their programs in DRC and set out to determine whether it may be caused by poor observance to the treatment.In 2013 the clinic staff in Shamwana identified patients returning to the clinic with severe malaria after having been prescribed a course of ACT (artesunate-amodiaquine). It would be very useful to know more about these patients: where they from all age groups and both genders? What was the median time to the recurrent episode? How far from the clinic did they come from? What was the mean duration of fever before presentation? More generally it is important to have more details of the increasing number of cases see in the clinic between 2009 and 2013: age and gender distributions as well as the number of severe/uncomplicated cases.There are several other potential reasons for the increase in cases: a larger population in the catchment area (or population movements), climatic changes, poor quality medicine, antimalarial resistance. Poor treatment observance is unlikely to be the only reason. The results of this study do not suggest that poor observance is a major problem, specially because the definition of probably non-adherence is too strict: taking the pills at a different time each day will not affect effectiveness. An interesting finding of the exit questionnaire is that few patients were given important additional information (Table 12). This suggests that the providers could probably improve the quality of the prescription. A useful indicator is the time taken by the prescribers to explain to the patients how to take the medicines.Finding out the real contribution of poor observance is not too complicated. A dry blood spot (DBS) collected on all patients presenting with a positive RDT will be sufficient. One needs to know (from the clinic records) if and when the patient was treated last time. The DBS can be used to measure the concentration of amodiaquine in the blood as well as finding parasite DNA. Depending on the type of RDT used, there is a possibility of false positive in the days/weeks after a treatment with ACT because of the persisting HRP2 in the circulation. The DBS can be stored at room temperature and analyzed later. It may identify that in fact the problem of increasing cases is mostly due to the emergence of resistance to amodiaquine. In 2006 Swarthout and colleagues found a failure rate with ASAQ of 6.7%  although in 2008-2009 another study  provided reassurance about the efficacy of ASAQ (cure rate 98.3 (94.1-99.8) by day 42.One suggestion to improve the impact of the intervention (LLIN and ACT) is to ensure that the diagnostic (RDT) and treatment (ACT) is given as soon as possible after the beginning of fever (<48h). This is because in order to prevent transmission, patients must be treated before the emergence of mature gametocytes in the circulation. The addition of the low single dose primaquine (as recommended by WHO) will help reducing transmission.In conclusion, this is a useful paper, but some more epidemiological details about the case-load and some simple investigations would help understanding the factors behind the increasing number of malaria cases seen in MSF clinics. Setting up malaria posts in the villages, where people can go early in the course of their fevers would help maximizing the impact on transmission.I have read this submission. I believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. This is an interesting paper from MSF epidemiologists that aims to determine some of the reasons for significant increases in malaria cases in DRC. It is imperative to prevent the emergence of anti-malarial drug resistance and patient adherence and compliance is critical to this issue. The data presented here provides evidence of what is happening in the field and touches on patients\u2019 attitudes and knowledge in this area. The authors focus on patient compliance and statistically examine socio-demographic and other factors for non-adherence to the prescribed time schedule and dosage. A key finding is the low rate of adherence, which has significant impact on drug-based treatment of malaria. Non-adherence due to vomiting or feeling unwell in a proportion of individuals is an important finding for the drug formulation community; how can this be overcome by changing the formulation or possibly the route,\u00a0 in the future?\u00a0 Timing was also highlighted as an obstacle to correct drug administration; again this is an obstacle that should be tackled both by drug developers, practitioners in the field and those interested in health literacy. The authors should consider minor revisions to their paper to discuss these issues in more depth. For example, based on their data, what are the next critical steps that should be focused on; from both a practice, and more long-term, drug design perspective? How can we succeed in patient education relating to the importance of adherence and designing and implementing a system that results in better patient adherence? Given that the authors found that gender is a significant factor; can the authors suggest how this finding should be used to develop strategies to improve patient compliance? Finally, the authors discuss the problems of trying to compare across studies due to different study designs and methods. From a methodology perspective, therefore, it would be useful if the authors included the full questionnaires used in this study as an appendix, with the objective of replication of the study in other areas.I have read this submission. I believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard."}
+{"text": "In the Democratic Republic of Congo, artesunate-amodiaquine (ASAQ) is the first-line medication recommended for uncomplicated malaria treatment. We conducted a study in Kinshasa to describe the clinical features of the disease and assess the efficacy of ASAQ and its impact on the multiplicity of infection in children with uncomplicated malaria.P. falciparum malaria were treated with ASAQ and followed up passively for 42 days. To distinguish new infections from recrudescent parasites, samples were genotyped using a stepwise strategy with three molecular markers . We then assessed PCR-corrected and -uncorrected day-42 cure rates and multiplicity of infection (MOI).Children aged 12 to 59 months with uncomplicated In total, 2,796 patients were screened and 865 enrolled in the study. Clinical features were characterized by history of fever (100%), coryza (59.9%) and weakness (59.4%). The crude and PCR-corrected efficacies of ASAQ were 55.3% (95%CI: 51.8\u201358.8) and 92.8% (95%CI: 91.0\u201394.6) respectively, as 83.6% (95%CI: 79.1\u201387.2) of the recurrences were new infections. Compared to monoclonal infections, polyclonal infections were more frequent at enrollment (88.1%) and in recurrences . The median MOI at enrollment  decreased to 3 (IQR: 1\u20135) in the recurrent samples (p<0.001). Patients infected with a single haplotype on day 0 had no recrudescence; the risk of recrudescence increased by 28% with each additional haplotype .The PCR-corrected efficacy of ASAQ at day 42 was 92.8%, but crude efficacy was relatively poor due to high reinfection rates. Treatment outcomes were positively correlated with MOI. Continued monitoring of the efficacy of ACTs\u2014ASAQ, in this case\u2014is paramount.NCT01374581ClinicalTrials.gov  The Democratic Republic of Congo (DR Congo) is considered to be one of the countries most severely affected by malaria . MalariaMonitoring the efficacy of ACTs is essential, especially since the reported emergence of resistance to artemisinin derivatives in Southeast Asia ,7,8. TheThis study describes malaria features in Kinshasa and assesses the efficacy of ASAQ after 42 days of treatment. In addition, it presents a cohort in which children who experienced clinical failures became eligible for inclusion in a randomized clinical trial (RCT) to assess the efficacy and safety of three rescue treatments, known as the Quinact study . FurtherThis is a report on the first phase of the Quinact study in DR Congo, a bi-center, randomized, open label, 3-arm trial performed in three phases . In DR CPlasmodium falciparum ; hemoglobin (Hb) value \u22656.0 g/dl; signed  informed consent granted by the parents or guardians and parents\u2019 or guardians\u2019 willingness and ability to comply with the study protocol for the duration of the study. The body weight threshold of 9 kg was selected because the pre-qualified quinine used in the randomized phase was not available for children below this weight.We screened boys and girls aged between 12 and 59 months attending Lisungi Health Center for fever (tympanic temperature \u226538.0\u00b0C) or history of fever. Enrollment in the study was based on the following criteria: body weight \u22659 kg; microscopically confirmed mono-infection or mixed infection containing We excluded patients with severe malaria  or dangeASAQ  was administered once daily over the course of three days. The tablets contained 50 mg of artesunate and 135 mg of amodiaquine for patients weighing 9 to 17.9 kg; and 100 mg of artesunate and 270 mg of amodiaquine for those weighing 18 to 35.9 kg. Treatment was observed directly by a study nurse. Patients were required to remain at the clinic for at least 60 minutes after drug intake. A full or half dose was repeated if vomiting occurred within 30 minutes or between 30 and 60 minutes of drug administration, respectively. If vomiting persisted, the patient was withdrawn from the study and referred to the health facility. The TREND checklist and the study protocol are available; see After enrollment, patients were asked to attend the study site on the two following days for directly observed treatment in order to complete the 3-day course of ASAQ and then every two weeks until day 42. They were encouraged to attend the study site on any unscheduled day if they experienced any health problems. Adverse events (AEs) were documented at each visit. Follow-up was passive. Malaria infection was not screened systematically; this decision was made for logistical reasons, because the study team planned to implement standard active follow-up for the two subsequent phases . Blood sBlood samples were collected on filter paper (Whatman 3MM) on day 0 and on days when blood smears were performed, to enable subsequent parasite genotyping. Serum samples were also collected upon recruitment and frozen for further immunological assessment (to be reported elsewhere).Symptoms and physical examination findings were graded according to their severity  using the severity grading scale developed by the World Health Organization and the United States National Institute of Health, Division of Microbiology and Infectious Diseases.Thick and thin blood films were prepared, dried and stained with Giemsa 10% for 10 minutes. Thin smears were fixed with methanol before staining. Slides were examined using a light microscope at 1000 times magnification. The density of asexual parasites was determined on the basis of the number of parasites per 200 white blood cells (WBC), assuming a total WBC count of 8,000/\u03bcl. If fewer than 10 parasites were read per 200 WBC, the count was extended to 500 WBC. An independent double reading was performed, with the average taken as the final result. A third reader was required in the event of discrepancies. Hb was measured using a portable spectrophotometer . The data management team based at the Infectious Diseases Institute, Makerere University in Uganda, produced a list of random slides to be sent periodically to the Institute of Tropical Medicine (ITM) in Antwerp, Belgium, for external quality control. In addition, two laboratory-specific monitoring visits were conducted by a lab technician from ITM\u2019s malaria unit.Plasmodium clones. Each clone was identified by a particular genotype defined by the alleles present at three loci, encoding GLURP (glutamate-rich protein), MSP (merozoite surface protein) 1 and MSP2, respectively.PCR genotyping was performed at ITM, Antwerp, Belgium, to distinguish recrudescence from reinfection. DNA was extracted from dried blood spots using the QIAGEN QIAamp96 DNA blood kit. Each punch (5 mm) was eluted in 150 \u03bcl of water. The blood spot samples taken from the malaria patients were characterized by the presence of one or more In order to characterize the parasites in each blood spot, alleles from each locus were determined and interpreted as follows:GLURP: alleles were characterized by size, differences in which were caused by varying numbers of a repeated unit.MSP2: two sequence families (3D7 and FC27) were recognized at this locus, both of which varied in size because of different repeated numbers. In order to achieve the highest assay sensitivity, we used fluorochrome-labelled reverse primers that were specific either to the 3D7 or FC27 allelic family. Alleles were distinguished by their fluorescent dye (indicating the family) and by their size, which was determined using an automated sequencer.MSP1: each allele was amplified using a PCR primer set specific to its sequence family .P. falciparum was not identified in the samples collected at D0. All analyses, gel electrophoresis and data entries were double-checked by a second technician. GeneTools software version 4.03.05.0  was used to score fragments of GLURP and MSP1, and GeneMapper version 4.1  was used for MSP2.If only one clone was present in a sample, then only one allele was found at each of the loci described above. If several clones were present, however, the number of alleles found at each locus could range between one and the number of clones. To determine whether it was a new infection or recrudescence that was causing clinical or parasitemic failure, we compared the genetic signatures of the samples at day 0 (D0) and day X . If at least one identical allele was found in these paired samples at each of the three loci, the failure was classified as recrudescent. If only new alleles were observed at one or more loci at DX, the case was classified as a new infection. When the amplification reaction for one locus proved to be negative for either one or both of the paired samples, the outcome was classed as indeterminate. Outcomes were classified as protocol violations when The alleles recovered were binned according to allele size : 50 bp for GLURP; 3 bp for MSP2; and 25 bp for MSP1. The highly divergent alleles found at the MSP2 locus were grouped into dimorphic families known as FC27 and 3D7. Likewise, the MSP1 alleles were grouped into three families: K1, Mad20 and RO33. The stepwise genotyping strategy chosen for this study led to the genotyping of a different number of samples using each marker. In order to avoid bias due to repeated counts of alleles within one patient with multiple infections, the basal level of allelic patterns and genetic diversity was initially assessed using D0 samples only.He = expected heterozygosity, i.e. a measure of the degree of polymorphism in each marker. The infections were classified as baseline samples (D0) or recurrent infections (DX) and then according to the number of P. falciparum-carrying clones: monoclonal  or polyclonal infection . Multiplicity of infection  was estimated using the locus with the highest number of alleles as a proxy [We then calculated and compared the allelic patterns and level of genetic diversity described by each marker between the baseline (D0) and recurrent infections (DX), where Na = number of alleles, Na frequency \u22655% = alleles found with a frequency \u22655% and  a proxy .The sample size was calculated on the basis of the randomized phase of the Quinact trial (reported separately). Recruitment in the first phase was to be continued until the number of failures required for the RCT phase was reached . Data we2 test and average MOIs compared using the Mann-Whitney U test. The allelic patterns\u2014number of alleles and private alleles  and level of genetic diversity \u2014were calculated using GenAlEx v.6.5 [Frequencies, percentages, means and medians were obtained using descriptive statistics. The association between MOI and treatment failure was determined with Kaplan Meier survival analysis and a Cox proportional hazard model adjusted for age, D0 parasite density, Hb concentration and temperature. The proportions of polyclonal infections at D0 and DX were compared using a Pearson \u03c7Ex v.6.5 ,19. The The primary outcome was adequate clinical and parasitological response (ACPR) at day 42, as the passive form of follow-up used in this study phase did not allow us to establish treatment outcomes according to the standard WHO classification . We consThe study protocol was approved by the ethical committees of the University of Antwerp (Reference: UA-A11-02) and the School of Public Health, University of Kinshasa (Reference: ESP/CE/012B/2012). Parents or guardians were asked to sign (or thumb-print if they were illiterate) informed consent forms. The study was externally monitored by the Amsterdam Institute for Global Health and Development in order to guarantee the quality of the data. The study team complied with good clinical and laboratory practice requirements. The protocol was registered under the references NCT01374581 and PACTR201203000351114 in ClinTrials.gov and in the Pan African Clinical Trials Registry, respectively.A total of 2796 patients were screened for eligibility in the first phase of the Quinact trial between August 2012 and January 2014. Of these, 145 patients (5.2%) were excluded on clinical bases. Subsequently, 2651 patients (94.8%) were tested for malaria; more than half tested negative (52.8%). In total, 865 patients fulfilled the selection criteria and were treated with ASAQ. At recruitment, the mean age was 36.2 months (SD: 13.2) and mean Hb was 9.3 g/dl (SD: 1.6). Approximately one third of patients (34.8%) had slept under a mosquito net the night before enrollment . Those wThe parents/guardians of 495 patients (57.2%) reported that their child had been sick in the last two months, but information about the condition was provided in only 106 (21.4%) of these cases. The most frequently reported condition was malaria, including co-morbidities (58.5%). Fever not clearly related to disease was reported by 41 (38.7%) parents/guardians. The parents/guardians of 278 patients were able to describe individual treatment courses prescribed during the month before enrollment. These had either been prescribed at health facilities or represented self-medication. The most common medication class that participants had received in the month before enrollment was antipyretics (44.2%), followed by antimalarial drugs (15.1%) and antibiotics (8.6%).All participants exhibited fever and/or history of fever, and this was the key symptom for screening. The most frequent symptoms were weakness, cough, behavioral changes and coryza. Jaundice and dehydration were rare, but chest examinations revealed abnormal results in 78% of patients . ParasitHe: 0.90\u20130.97). MSP2 was found to be the most polymorphic, exhibiting up to 114 different alleles , while 20 and 19 alleles  were discovered for MSP1 and GLURP, respectively (He) of the three markers did not change among baseline samples (D0) or recurrence after treatment (DX). MSP2 revealed the largest number of private alleles in D0 and DX (26.3% and 17.5%), followed by MSP1 (15.0% and 19.0%) and GLURP (5.3% in both) (The three markers showed high levels of polymorphism (ectively . The levin both) . The proWithin-host parasite diversity, multiplicity of infection (MOI) and risk of recrudescenceThe majority of infections (78.8% of 146) contained a mixture of MSP2 family-specific alleles (3D7 and FC27), while only 13.7% and 7.5% exhibited parasites carrying only alleles 3D7 or FC27, respectively. Similarly, most infections contained a mixture of MSP1 family-specific alleles (79.7% of 64); and infections containing alleles from all three MSP1 allelic families reached 40%. Among infections containing parasites carrying only one MSP1 allelic family, K1 alleles were found to be more frequent (17.2%) than Mad20 and RO33 (1.6% in both cases) . MultiplIn total, 64 and 146 samples  were genotyped using MSP1 and MSP2, respectively. The infections contained alleles belonging either to a single allelic family or to two or three different families. Multiple alleles belonging to the same family were found within one infection. Polyclonal infections were more frequent at D0 and DX  than were monoclonal infections (p = 0.005). The median MOI found in recurrence samples  was lower than the MOI found in D0 samples  (p<0.001). The odds of having a polyclonal infection were 1.8 times higher (95%CI: 1.20\u20132.8) for pre-treatment samples than for recurrences. Among patients with treatment failure, those with new infections were found to be carrying up to 10 haplotypes, while those with recrudescence were harboring up to 14 haplotypes .P. falciparum haplotype found within the D0 infection .No risk of recrudescence was found when the D0 infection was monoclonal , while pP. ovale, at recruitment, but neither of them experienced P. ovale-related recurrence.No clinical failure occurred before day 14. Forty-five percent of patients experienced treatment failure during the 42-day follow-up period . PCR-corPlasmodium infection. In Nigeria, upper respiratory tract infections have been reported in 70% of patients testing positive for malaria in an outpatient department [This study highlights the clinical and molecular features of malaria in Kinshasa, DR Congo, and provides up-to-date data on ASAQ efficacy when treating uncomplicated malaria in children under five years of age. Malaria is clinically characterized by non-specific symptoms which may mimic flu-like syndromes and even lead to misdiagnosis of gastrointestinal infection . Sixty ppartment .et al in Tanzania, for example, has succeeded in revealing causes of fever beyond malaria [Plasmodium infection is confirmed, and co-morbidities should be managed accordingly. Our data suggest that parasite density tends to decrease as age increases  [et al [et al [et al [The proportion of patients with ACPR at day 42 was 92.8% after PCR adjustment. However, the proportion of recurrent malaria was high, at 44.7%. Onyamboko ns 27%) . This di) [et al . The rell [et al . Anotherl [et al . The lev% . This l [et al . It is el [et al . The latP. falciparum strains. Children under five years of age carry the majority of the strains in circulation because of their vulnerability to malaria infection [P. falciparum diversity in the study area and therefore represent baseline knowledge of the diversity and multiplicity of infection in Kinshasa. In addition, most malaria cases identified in this study were polyclonal infections. Some studies have demonstrated a positive association between high P. falciparum diversity and MOI, on the one hand, and high malaria transmission, on the other [In general, the patients screened in our study proved to be infected by a high number of nfection . The reshe other \u201333, thouhe other .P. falciparum diversity is not entirely clear. Yet, even in areas where malaria incidence has decreased by 90%, no decrease in parasite diversity has been documented [et al [The odds of having a polyclonal infection were lower among recurrences than at enrollment. In order to reinforce this trend, which probably reflects the impact of effective medication; joint strategies for malaria prevention  are needed to reduce multiple-strain infections . Howevercumented . This stcumented ,36. Whilcumented . In contd [et al , we founThe passive form of follow-up used in this study is likely to have had an impact on the estimation of the proportion of recurrent infections. If the standard procedures for antimalarial efficacy follow-up were applied in the study\u2014with systematic malaria screening on days 1, 2, 3, 7, 14, 21, 28, 35 and 42 \u2014the studP. falciparum was found to predict a positive treatment outcome. Therefore, reducing the diversity of circulating P. falciparum may be key to slowing down the spread of resistant strains.ASAQ remains a suitable treatment for uncomplicated malaria in Kinshasa, with a PCR-corrected efficacy of 92.8% after 42 days of follow-up. The proportion of crude treatment failures was relatively high in this study, perhaps because of the poor prophylactic effect of amodiaquine, the high level of malaria transmission or the high host susceptibility to malaria among children participating in the study. Infection with a single haplotype of in vitro amodiaquine resistance profiles and the consequences of MOI on treatment efficacy, malaria transmission and immunity development; and entomological studies for assessing malaria transmission levels. In vivo efficacy studies of ACTs are also of paramount importance.This study emphasizes the need for further research on S1 TREND Checklist(PDF)Click here for additional data file.S1 Protocol(PDF)Click here for additional data file.S2 Protocol(PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum and P. vivax malaria. The objective of this study was to evaluate the population pharmacokinetics and pharmacodynamics of piperaquine in patients with P. vivax malaria in Thailand after a standard regimen of dihydroartemisinin-piperaquine to determine whether residual piperaquine prevents or delays the emergence of P. vivax relapse. Sparse blood samples were collected from 116 patients. Piperaquine pharmacokinetics were described well by a three-compartment distribution model. Relapsing P. vivax malaria was accommodated by a constant baseline hazard (8.94 relapses/year) with the addition of a surge function in a fixed 3-week interval and a protective piperaquine effect. The results suggest that a large proportion of the first relapses were suppressed completely by residual piperaquine concentrations and that recurrences resulted mainly from emergence of the second or third relapse or from reinfection. This suggests a significant reduction in P. vivax morbidity when using dihydroartemisinin-piperaquine compared with other antimalarial drugs with shorter terminal postprophylactic effects.Dihydroartemisinin-piperaquine is an effective drug in the treatment of  Plasmodium vivax is the most difficult of the five species of human malaria parasites to eliminate. It is the most geographically widespread human malaria parasite, causing an estimated 130\u2013390 million cases every year.1P. vivax is transmitted within 95 countries in America, Africa, and Asia.3 In total, 2.85 billion people were estimated to be at risk of P. vivax transmission in 2009, especially in Central and Southeast Asia, where more than 90% of the world's P. vivax transmission occurs.3 Acute P. vivax malaria is rarely life threatening, but it causes substantial morbidity in higher transmission areas as a result of multiple relapses from latent liver-stage parasites.4 These repeated infections may result in life-threateningly severe anemia due to destruction of both infected and noninfected red blood cells. P. vivax is an important cause of low birth weight.7P. vivax malaria in most endemic areas. However, P. vivax resistance to chloroquine, which was first observed in Papua New Guinea in 1989,8 is increasing in many parts of the world.9 A combination of two drugs with different mechanisms of action is preferred for the treatment of malaria to reduce the risk of the development of resistant parasite strains. The fixed combination of dihydroartemisinin-piperaquine (DHA-PQ) is recommended by the World Health Organization as the first-line treatment of uncomplicated P. falciparum malaria10 and has replaced chloroquine in the first-line treatment of resistant P. vivax malaria in Indonesia. DHA-PQ is currently available as a fixed-dose combination, administered once daily over 3 days. A meta-analysis pooled from 12 different studies in 6 countries between 2003 and 2006 showed high efficacy against P. falciparum malaria (polymerase chain reaction-corrected cure rates of 98.7% at day 28) and good tolerance  of DHA-PQ.11 DHA-PQ also has excellent efficacy against P. vivax malaria.14Chloroquine, a blood schizonticide, is used in first-line treatment and prophylaxis of t1/2) of ~18\u201328 days). Similar pharmacokinetic properties have been reported in children but these pediatric studies suggest that small children need a higher weight-based dose to achieve exposure comparable to that in older children.15 Most piperaquine pharmacokinetic studies have sampled venous plasma. However, capillary sampling is a desirable alternative for field pharmacokinetic studies, especially in children. The population pharmacokinetics and pharmacodynamics of piperaquine have not been reported previously in patients with P. vivax malaria.Piperaquine is distributed extensively into tissues, resulting in multiphasic distribution and a slow elimination from the systemic circulation  or delayed. The aim of this study was to characterize the population pharmacokinetics and antimalarial pharmacodynamics of piperaquine in the treatment of P. vivax malaria and to use this information to determine whether piperaquine prevents or delays the first P. vivax relapse.In South East Asia and Oceania, P. vivax malaria infections. Full clinical details have been published elsewhere and DHA-PQ was found to be an effective alternative treatment with a lower cumulative risk of recurrent P. vivax malaria at 9 weeks compared with chloroquine (54.9 vs. 79.1%).16 Two-hundred and fifty Karen and Burmese patients with P. vivax malaria infections were enrolled in the DHA-PQ-treated arm of the efficacy study (Table 1). All patients received DHA-PQ  tablets containing 40\u2009mg of dihydroartemisinin and 320\u2009mg of piperaquine phosphate. A standard weight-based regimen (2.2\u2009mg/kg/day of dihydroartemisinin and 17.8\u2009mg/kg/day of piperaquine phosphate) was administered with milk once daily for 3 days. The fixed combination of DHA-PQ was well tolerated with no serious adverse events reported during follow-up. In this population modeling, nine patients were excluded: two patients did not meet the inclusion criteria, five patients had an incomplete course of medication, one patient self-medicated with other drugs, and one patient withdrew consent. Overall, 45% (109/241) of patients presented with recurrent P. vivax malaria during the follow-up period at a median (range) of 56 (14\u201365) days (Table 1).This study was conducted at Shoklo Malaria Research Unit clinics, Mae Sot, Thailand, an area of low and seasonal malaria transmission located at the Thailand\u2013Myanmar border. This pharmacokinetic study was nested into a larger efficacy trial in 500 patients with 17 A proportional venous-capillary transformation factor was, therefore, used to compensate for the difference between sampling matrices, resulting in 41% higher capillary concentrations compared with venous concentrations.A total of 791 plasma concentrations (435 venous and 356 capillary samples) from 116 patients were quantified and used in the pharmacokinetic modeling. Venous and capillary plasma concentrations were transformed into their natural logarithms and modeled simultaneously using nonlinear mixed-effects modeling. Substantial differences in matched venous and capillary blood concentrations have been reported in a previous clinical study.CL/F), intercompartment clearance(s) (Q/F), apparent volume of distribution of the central compartment (VC/F), apparent volume of distribution of the peripheral compartment(s) (VP/F), and absorption rate constant. Several absorption, distribution, and covariate models were fitted to the data to construct the best performing model. Piperaquine pharmacokinetics in patients with P. vivax malaria were best described by a three-compartment distribution model . A three-compartment disposition model resulted in a significant improvement in model fit compared with a two-compartment model  of 108) with no additional benefit of an additional peripheral compartment (\u0394OFV = 0.57). A transit-compartment model with a fixed number of transit compartments  described the absorption phase better than all other absorption models (\u0394OFV > 54.2). A relative bioavailability parameter  was implemented to allow quantification of the interindividual variability in the absorption of piperaquine and resulted in a significant improvement in the model fit (\u0394OFV = 81.8).The initial structural base models were parameterized as elimination clearance (VP2/F (\u0394OFV = 11.5) and age on Q1/F (\u0394OFV = 5.46) were selected in the forward covariate selection but could not be retained in the backward deletion. The estimated interindividual variability was small for the transformation factor, CL/F, Q2/F, and VC/F, and could, therefore, be removed with no significant impact in the final model (\u0394OFV = 2.40). Final pharmacokinetic parameter estimates and secondary parameter estimates are reported in Tables 2 and 3.Body weight, incorporated as a fixed allometric function, improved the model fit significantly (\u0394OFV = 31.0) and resulted in better precision and decreased interindividual variability in clearance and volume parameters. Body temperature on admission on Supplementary Figure S1 online) with a small epsilon-shrinkage of 15.0%. However, a relatively high \u03b7-shrinkage (up to 43.5%) could be seen for certain parameters in the final model because of the sparse data. The numerical and prediction-corrected visual predictive checks resulted in 4.6% (95% CI: 2.7\u20138.1%) and 3.7% (95% CI: 2.5\u20138.2%) of piperaquine observations below and above the simulated 90% prediction interval, respectively .The mean parameter estimates from the simultaneous model with both venous and capillary data were also comparable with estimates when capillary and venous data were modeled separately. The final pharmacokinetic model displayed satisfactory goodness-of-fit diagnostics  protective effect on the risk of recurrent malaria infections with a required in vivo venous piperaquine concentration of 6.92\u2009ng/ml for a 50% relative reduction in baseline hazard (PC50). Sex was selected as a significant covariate (P < 0.05) on baseline hazard in the forward selection step, but it was removed during the backward elimination resulting in a covariate-free pharmacodynamic model. Final pharmacodynamic parameters were well estimated with high precision (Table 2) and simulation-based diagnostics showed good agreement between simulated and observed recurrent malaria infections .Piperaquine inhibits asexual parasite multiplication but has no effect on liver-stage parasites. The influence of piperaquine exposure on the risk of recurrent Table 1) corresponded approximately to a total of 1011 parasites in a typical adult at admission. This assumes that P. vivax does not sequester substantially. DHA-PQ is administered for 3 consecutive days, covering two asexual cycles. Assuming a parasiticidal effect (parasite reduction ratio) of a 10,000-fold parasite reduction per cycle by dihydroartemisinin  results in a 100,000,000-fold reduction in total parasite biomass during the first two cycles.20 At an initial parasite burden of 1011 parasites, this would leave ~1,000 parasites to be eliminated by residual piperaquine concentrations to prevent recrudescent malaria. Piperaquine alone has not been adequately assessed in P. vivax malaria, but if it is similar to chloroquine, it should provide parasite reduction ratios of ~100 to 1,000 per cycle, so most patients should be parasite free 6\u20138 days post-treatment if piperaquine levels are sufficient to sustain a maximum parasiticidal effect for this time. If the parasite burden is not eliminated completely, then these remaining parasites can multiply and cause a recrudescent infection. However, a high cure rate of the asexual infection is likely in this study as concentrations of piperaquine after the second asexual cycle  are highly likely to be able to eliminate ~1,000 parasites. In the first relapse of tropical P. vivax malaria (unaffected by drugs and assuming 1\u201310 schizonts and a subsequent parasite multiplication rate of ~10 per cycle), the hypnozoite-derived hepatic schizonts must liberate 13\u201317 days after the onset of the primary illness to produce a symptomatic infection at day 21. However, if residual piperaquine concentrations are sufficiently high, they could suppress and eliminate the asexual parasites derived from 10,000 to 100,000 released merozoites and thereby prevent the first relapse completely. If residual piperaquine concentrations are not sufficiently high for complete elimination, they could cause an initial suppression, resulting in a delayed symptomatic first relapse. Only 14% (15 out of 109) of recurrent P. vivax infections occurred before day 39, which suggests that DHA-PQ treatment prevents a large proportion of the first relapses . The majority of infections  occurred between day 42 and 56, suggesting that these are mainly delayed second relapses . There will be some contribution from reinfections, but as entomological inoculation rates are probably well below 0.5/person/year for P. vivax in this area, this contribution is likely to be small.The individual patient parasite burden  in Thailand.16 This difference could be a result of the very potent artemisinin derivative which generally has a rapid onset of action against P. vivax malaria,21 but it is likely to result mainly from increasing chloroquine resistance and hence the superiority of piperaquine. In fully sensitive P. vivax malaria, recrudescence rates are close to zero with chloroquine treatment. Previous studies in this region have shown high relapse rates of P. vivax within one month of treatment with a short acting artemisinin derivative.19 Thus, both slowly eliminated drugs prevent the first relapse. Whether the lower cumulative risk of recurrent P. vivax malaria, following DHA-PQ, results from a higher cure rate of the primary infection or a more effective suppression of the first or the second relapse has been unclear. As relapses can arise from parasites which are either genetically similar or genetically distinct to those causing the primary infection, it is not possible to distinguish with certainty between relapse, recrudescence, or reinfection in patients with P. vivax malaria who remain in the endemic area. Recrudescence can occur many weeks after treatment with slowly eliminated drugs, but the pharmacokinetic-pharmacodynamic data modeled in this study suggest that only a negligible proportion (<2%) of the recurrent P. vivax infections could be recrudescent.The fixed combination of DHA-PQ is a highly efficacious antimalarial drug with an excellent safety profile. Antimalarial treatment with DHA-PQ resulted in lower cumulative risk (54.9%) of recurrent P. falciparum malaria.24 This is the first study reporting the population pharmacokinetics of piperaquine in patients with P. vivax malaria.The population pharmacokinetic properties of piperaquine have been described previously in patients with uncomplicated P. falciparum malaria.25 Simplified two-compartment structural models in children and adult patients have also been reported reflecting differences in follow-period and/or sampling strategies.24 A transit-compartment absorption model allowed more flexibility in the absorption phase and provided a clear advantage over other absorption models, although there were not enough data in the absorption phase to estimate both KTR and KA separately. Large interindividual variability was seen in the absorption of piperaquine as in previous studies.25 Data collected here were too sparse to allow an estimation of the variability between dose occasions, which is likely to have inflated the inter-individual variability. Indeed interindividual variability was high in this study with values up to 138%. Venous and capillary plasma concentrations were successfully modeled simultaneously with a transformation factor of 1.41 (relative standard error of 2.11%). This factor was similar to that from a linear regression between observed venous and capillary plasma concentrations (slope = 1.29), which further supports the appropriateness of the simultaneous model. However, the exact mechanism of this matrix-dependent difference cannot be elucidated from the data collected in this study. The advantage of a simultaneous modeling approach is that the model can be used to simulate drug exposures from any sampling technique and therefore enables literature comparisons.The structural three-compartment model described in this study was similar to that reported recently in young children and in pregnant and nonpregnant women with 24 and was not unexpected, considering the strong biological prior of body weight as a covariate on pharmacokinetic disposition and elimination parameters.26 All pharmacokinetic parameters were estimated with good precision (relative standard error < 20%) and the simulation-based diagnostics indicated excellent predictive performance of the final pharmacokinetic model.Body weight was the only significant covariate in this model. This has also been reported in previous studiesCL/F in this P. vivax study was 1.08 l/h/kg, total volume of distribution (VD/F) was 802 l/kg, and t1/2 was 28.8 days, which were comparable with previously published reports in patients with an uncomplicated P. falciparum malaria (mean (range) CL/F of 1.18 (0.90\u20131.4) l/h/kg, VD/F of 789 (574\u2013877) l/kg, and t1/2 of 23.7 (18\u201328) days).Piperaquine P. vivax malaria was expected as DHA-PQ eliminates only blood-stage parasites and does not provide cure of the latent liver-stage parasites. It was expected that piperaquine would delay the time to relapse, as does chloroquine, and this was supported by the pharmacodynamic results. Tropical frequent relapsing strains of P. vivax typically relapse at 3\u20134-week intervals, and the median time to relapse approximates three weeks (21 days) in Thailand after treatment with rapidly eliminated antimalarial drugs .19 In an area of much higher transmission in Papua, Indonesia, there were significantly more early recurrences of P. vivax malaria following artemether-lumefantrine treatment of P. vivax malaria compared with DHA-PQ (cumulative risk of 38 vs. 10% at day 42)13 reflecting the failure of the more rapidly eliminated lumefantrine to suppress the first relapse. However, with longer follow-up in a high transmission area, the cumulative reinfection rates converge as everyone is reinfected eventually. A critical and unanswered question has been whether the slowly eliminated drugs prevented, or simply delayed, the relapse. Explicitly, prevention would mean that the relapse emerged but was eliminated by the residual drug levels, and thus, the first observed recurrence is in fact the second relapse. Delay would mean that the residual drug levels reduced multiplication of the relapse, so it reached patency after a substantial delay. In the former explanation, overall relapse numbers might be reduced as this would exhaust the liver burden of hypnozoites more rapidly. A pharmacometric time-to-event approach explaining relapsing P. vivax malaria, first presented for amodiaquine treatment in pregnant women,29 described the data observed in this study well. An interval-censoring time-to-event model was successfully used since the exact times of relapses were unknown. The risk of relapsing P. vivax malaria in patients was explained by a constant baseline hazard  with a 123% increased risk every third week. The pharmacodynamic model was significantly improved by adding a protective drug effect, indicating that piperaquine cured the primary blood stage P. vivax infection and fully suppressed a large proportion of the first relapses that would have occurred three weeks after the initial infection. The pharmacodynamic model also indicated that the observed high recurrence rate is likely to be explained mainly by delayed second relapses, although it cannot be excluded that some recurrences are reinfections. This can only be mechanistically evaluated by an individual parasite-data-driven pharmacokinetic-pharmacodynamic model. Simulations using the final pharmacokinetic-pharmacodynamic model  demonstrated that minimum day-7 piperaquine venous and capillary concentrations of 27 and 38\u2009ng/ml, respectively, were needed to suppress the risk of relapse for 30 days . This is in close agreement with a previously defined day-7 threshold venous concentration of 30\u2009ng/ml in patients with P. falciparum malaria to minimize the risk of recrudescent infections.30The observed high relapse rate of P. vivax malaria were described successfully by modeling venous and capillary plasma concentrations simultaneously. Body weight was the only significant covariate and resulted in increasing clearance and volumes with increasing body weight. Times to recurrent P. vivax infections were successfully modeled with a time-to-event approach and resulted in a significantly delayed time to recurrent infections during the post-treatment phase of piperaquine treatment. This was explained by prevention of the first P. vivax malaria relapse by residual piperaquine concentrations. Piperaquine is a good candidate for the treatment of P. vivax malaria and the modeling conducted here demonstrated the added benefit of reduced morbidity compared with other more rapidly eliminated antimalarial drugs.In conclusion, the population pharmacokinetic properties of piperaquine in patients with 16Ethics approval was granted by the Faculty of Tropical Medicine Ethics Committee, Mahidol University, Bangkok, Thailand, and the Oxford Tropical Research Ethics Committee, Oxford University, UK. This study was registered in the ISRCTN Register (ISRCTN87827353). Details on the clinical study design and outcomes have been presented in full elsewhere.All patients received a standard regimen of DHA-PQ. Patients who vomited the dose were excluded from the pharmacokinetic study. A full or half replacement dose was administered if vomiting occurred within 30\u2009min of or between 30\u2009min to 1\u2009h after administration, respectively.P. vivax malaria. This random sampling allowed of an adequate coverage of the entire concentration\u2013time profile. An additional 54 patients provided a single plasma sample only at the time of recurrent malaria. Patients were excluded from the study after recurrent malaria. Plasma samples were shipped on dry ice to the Department of Clinical Pharmacology, Mahidol-Oxford Tropical Medicine Research Unit, Thailand, for drug quantification.31Venous blood samples (3\u2009ml) and capillary blood samples (200 \u03bcl) were drawn from each patient for piperaquine plasma measurements. Samples were collected randomly over 69 days (4\u20137 venous samples and 1\u201311 capillary samples) from 62 patients and an additional venous sample (5\u2009ml) was drawn from these patients at the time of recurrent 32 and Xpose version 4.033 in the programming language R version 2.13.2 . Measurements below the lower limit of quantification were less than 0.5% of the total data and therefore omitted. OFV, calculated by NONMEM as minus twice the log-likelihood up to constant, was used for model selection during the model-building process. A difference in the OFV (\u0394OFV) of >3.84 was considered significant (P < 0.05) when comparing two nested models with 1 degree of freedom difference. Full details of the pharmacokinetic and pharmacodynamic modeling methodology can be found in supplementary material.Piperaquine pharmacokinetics and pharmacodynamics were evaluated using nonlinear mixed-effects modeling . Postprocessing and automation were performed using Pearl-Speaks-NONMEM version 3.5.3, Census version 1.2b2,J.T., P.T., N.P.J.D., and N.J.W. wrote the manuscript. A.P.P., K.M.L., E.A.A., N.P.J.D., F.N., and N.J.W. designed the research. J.T., A.P.P., K.M.L., E.A.A., and F.N. performed the research. J.T., P.T., and W.H. analyzed the data.The Wellcome Trust is a UK-based charity that supports medical research and is independent of any drug companies. It has no financial links with the manufacturers of either the diagnostic tests or the drugs used in this study. The authors declared no conflict of interest."}
+{"text": "P. falciparum malaria has emerged in Western Cambodia and now has been observed in Thailand, Vietnam and Myanmar and Lao. In addition, there is a recent report of a Vietnamese patient, apparently infected with P. falciparum in Angola, who was unresponsive to artemisinin treatment. This makes it even more urgent to have baseline information on early parasite response to ACTs in African patients, the first step in establishing surveillance for this important phenotype.Artemisinin resistant Individual patient data from efficacy trials conducted between 1999 and 2012 were shared with the Worldwide Antimalarial Resistance Network (WWARN) and pooled analyses conducted using standardized methodology. Factors affecting early parasitological response to treatment were investigated using logistic regression with study sites fitted as a random effect.P < 0.001) and day 2  compared to treatment with DP. Treatment with ASAQ-L was also associated with a higher risk of persistent parasitemia on day 2  and day 3  compared to treatment with DP.Data from 85 studies  conducted in 27 countries with Artemether-lumefantrine , Artesunate-amodia-quine-fixed dose combination , Artesunate-amodiaquine-co-blister combination , Artesunate-amodiaquine-loose combination , and Di hydro artemisinin-piperaquine  were included. With all drugs tested, high baseline parasite density and fever were both independent predictors of persistent parasitemia on days 1, 2 and 3. Adjusted for these variables, patients from 1 to 5 years and from 5 to 12 years in areas of low/moderate transmission were at a 2-fold and 4-fold risk of persistent parasitemia on day 3 compared to patients of the same age-group in high transmission settings. Treatment with AL was associated with a higher risk of persistent parasitemia on day 1 (AOR = 1.57 [95% CI: 1.38-1.78], These results show no evidence of slow parasite clearance in these African studies. The greatest risk of slow clearance was observed in patients from areas of low/moderate transmission. The threshold for suspected diminished parasite susceptibility to artemisinins on day 3 was estimated to be 5%, much lower than the currently used World Health Organisation threshold of 10%. However, there are clear gaps in surveillance and any sites exceeding the day 3 parasite positivity rate of 5% should be further investigated."}
+{"text": "Artemisinin-based combination therapies (ACTs) are the cornerstone for the treatment of malaria. However, confirmed resistance to artemisinins in South-East Asia, and reports of reduced efficacy of ACTs raise major concerns for malaria treatment and control. Without new drugs to replace artemisinins, it is essential to define dosing strategies that maximize therapeutic efficacy, limit the spread of resistance, and preserve the clinical value of ACTs. It is important to determine the extent to which reduced efficacy of ACTs reflects true resistance versus sub-optimal dosing, and quantify other factors that determine treatment failure. Pooled analyses of individual patient data from multiple clinical trials, by investigators in the Worldwide Antimalarial Resistance Network, have shown high overall efficacy for three widely used ACTs, artemether-lumefantrine, artesunate-amodiaquine, and dihydroartemisinin-piperaquine. Analyses also highlight that suboptimal dosing leads to increased risk of treatment failure, especially among children. In the most recent study, an analysis of clinical trials of artesunate-amodiaquine, widely used among children in Africa, revealed a superior efficacy for fixed-dose combination tablets compared to loose non-fixed dose combinations. This highlights the benefits of fixed-dose combinations as a practical strategy for ensuring optimal antimalarial dosing and maximizing efficacy.http://www.biomedcentral.com/1741-7015/13/66Please see related article:  P. falciparum malaria in all areas in which malaria is endemic [Artemisinin-based combination therapies (ACTs) have made a major contribution to the reduction in global malaria morbidity and mortality since their use became widespread approximately 10\u00a0years ago. ACTs are recommended by the WHO as the first-line treatment of uncomplicated and severe  endemic . Drugs o endemic . Artesunkelch13 gene (gene ID PF3D7_1343700) [Unfortunately, the early signs of artemisinin resistance have emerged in South-East Asia threatening recent gains and milestones in the treatment and control of malaria ,5. Resis1343700) -8, and t1343700) . Emergin1343700) -15. It iA new study from the Worldwide Antimalarial Resistance Network (WWARN) investigated risk factors for treatment failure with AS-AQ therapy . ReducedAS-AQ is available in three different body-weight-adjusted drug formulations: fixed-dose combinations, loose non-fixed dose combinations, and co-blistered non-fixed-dose combinations. All of these combinations aim at delivering 12\u00a0mg/kg of AS over 3\u00a0days, but the total target dose of AQ ranges from 25 to 30\u00a0mg/kg for loose non-fixed dose combinations and is 30\u00a0mg/kg for co-blistered non-fixed-dose combinations and fixed-dose combinations. The WWARN study included 43 trials , of which 39 trials (95% of subjects) were conducted in Africa, as well as 3 in Asia and 1 in South America, and the great majority were children (87.5% were <12\u00a0years old) . The autP. falciparum malaria using fixed-dose AS-AQ formulations should be promoted for national treatment guidelines.Arguably, the most important finding was that fixed-dose combinations were associated with the highest AS-AQ treatment efficacy in all age groups, including in children less than 5\u00a0years of age, independently of high baseline parasitemia and young age. The fixed dose combination of AS-AQ was developed using weight-for-age values from malaria endemic countries, to ensure optimal dosing ,22. LoosP. falciparum resistance to AQ could not be evaluated in this study and further studies incorporating molecular markers of AQ resistance are warranted. Resistance to AQ is associated with mutations in the pfcrt and pfmdr genes [The study also found that the risk of recrudescence after AS-AQ treatment was higher among young children (<12\u00a0years old), those with a high baseline parasitemia, and in Asian studies compared to African. This may be reflective of the overall higher level of AQ resistance in Asia; however, given the small number of subjects from Asian trials, further studies are needed to examine this. The potentially crucial impact of dr genes ,24, whicdr genes . It willThere have been two related studies performed by WWARN investigators on the efficacy of other ACTs, highlighting the importance of optimal dosing. Dihydroartemisinin-piperaquine is another widely used ACT, and risk factors for recrudescence after treatment were examined in a pooled analysis of individual patient data from 26 efficacy studies  ; overallIn an era of emerging artemisinin resistance, and the push for malaria elimination in many regions, studies such as that of the WWARN collaboration provide important data to inform policymakers and clinicians to optimise antimalarial therapies to maximize efficacy and help reduce the evolution of resistance (see Box 1 for a summary of key points). These findings provide further support for international collaborative networks and data sharing arrangements to address major challenges in global health, and the approach used for malaria has strong relevance to antimicrobial therapy and resistance more broadly. Individual patient meta-analyses of therapeutic efficacy studies are considered the strongest form of clinical evidence, and are essential to inform antimalarial policy and clinical treatment guidelines to ensure the rapid and effective treatment of malaria cases.The WHO recommends artemisinin combination therapies (ACT) as the first-line treatment of malaria1Around 390 million courses of ACTs procured annually Emerging resistance to ACTs: Resistance confirmed in five countries in the Greater Mekong sub-regionSuspected artemisinin resistance is defined as failure to clear parasitemia within 3\u00a0days of commencing treatment, or a parasite clearance half-life of \u22655\u00a0hourskelch13 geneResistance to ACTs is associated with mutations in Resistance to other antimalarials is widespread pfcrt and pfmdr1 genesResistance to amodiaquine and chloroquine is associated with mutations in the Ensuring effective dosing is crucial to maximize efficacy and to protect against the further development and spread of resistanceAS-AQ is widely used in Africa, and is the second most widely used ACTAS-AQ Previously shown to have high efficacy when administered correctlyWWARN study performed an individual patient meta-analysis using data from 43 studies (39 studies in Africa) with over 9,000 treatments, predominantly in childrenStudy evaluated the impact of different dosing strategies on therapeutic efficacyFixed-dose and co-blistered non-fixed dose combinations had high efficacy (98%)Use of loose tablets at non-fixed dose had significantly lower efficacy (up to 3.5-fold higher risk of treatment failure)Findings highlight the benefit of fixed dose combinations and support this strategy in the treatment and control of malariaIncreased risk of recrudescence among young children, those with high baseline parasitemia, and among trials conducted in AsiaBased on WHO figures from 2013.See reference"}
+{"text": "Severe malaria is a medical emergency with high mortality in children below 5 years of age especially in sub-Saharan Africa. Recently, quinine has been replaced by artesunate as the first-line drug in the treatment of severe malaria in Cameroon. No local data are yet available on the efficacy of artesunate with respect to the different quinine regimens used in this setting. This study was undertaken at the Ebolowa Regional Hospital (ERH), which is located in a region of perennial transmission of malaria.Plasmodium falciparum confirmed on microscopy after informed parental consent. Patients were randomized into four groups. Group 1 (ARTES) received parenteral artesunate at 2.4\u00a0mg/kg at H0, H12, H24 and then once daily; Group 2 (QLD) received a loading dose of quinine base at 16.6\u00a0mg/kg followed 8 hours later by an eight-hourly maintenance dose of 8.3\u00a0mg/kg quinine base; Group 3 (QNLD3) received 8.3\u00a0mg/kg quinine base every 8 hours; and, Group 4 (QNLD2) received 12.5\u00a0mg/kg quinine base every 12\u00a0h. All patients invariably received a minimum of 24\u00a0h parenteral treatment, then, oral drugs were prescribed. The endpoints were fever clearance time, time to sit unsupported, time to eat, parasite clearance time, and parasitaemia reduction rate at H24. Survival analysis was used to compare the outcomes.This was a randomized, open-label trial in children aged 3 months to 15\u00a0years, admitted in the hospital with severe malaria due to One-hundred and sixteen patients completed the study: 29 in ARTES arm, 28 in QLD arm, 30 in QNLD3 arm, and 29 in QNLD2 arm. There was no major differences in baseline characteristics in the treatment groups. On analysis of endpoints, fever clearance time and parasite clearance time were significantly shorter for artesunate-treated patients than for quinine-treated patients. Parasitaemia reduction rate at H24 was also significantly higher for artesunate. Time to sit unsupported and time to eat were shorter with artesunate, but the difference was not statistically significant.Artesunate is more effective than quinine in the treatment of severe malaria in Cameroonian children. Malaria control is a cornerstone in the reduction of child mortality and constitutes the fourth Millennium Development Goal . To dateIn Cameroon, malaria is a major endemic parasitic disease and the first cause of morbidity and mortality. Children under 5 years old and pregnant women are the most vulnerable population groups. Malaria alone is responsible for 24\u00a0% of total deaths, 40\u201345\u00a0% of medical consultations and 30\u00a0% of hospital admissions in children under 59\u00a0months old [The mortality of untreated severe malaria  approaches 100\u00a0%. With prompt, effective anti-malarial treatment and supportive care, mortality falls to 15\u201320\u00a0% overall, making it a medical emergency . In lineIn 2013, the Cameroon National Malaria Control Programme (NMCP) revised its treatment directives and adopted the above WHO treatment regimens. The programme also recommended a non-loading dose regimen of 8.3\u00a0mg/kg bw of quinine base (QB) eight-hourly . In someDespite the fact that artesunate has been recommended for severe malaria, uptake of parenteral artesunate for the treatment of severe malaria is still slow in Cameroon due to conservative attitudes and possibly due to the lack of country-specific data. Therefore, this study was undertaken to assess the efficacy of artesunate with respect to other quinine regimens currently used in this context for the treatment of severe malaria in children in a hospital located in a malaria-endemic geographical region in south Cameroon.This study was conducted in the paediatric unit of the Ebolowa Regional Hospital (ERH). This hospital is located in Ebolowa, which is the headquarters of the South Region of Cameroon and is the referral hospital in that region. It has a total capacity of 158 beds for an estimated population of 120,000 inhabitants. The paediatric unit itself has a capacity of 28 beds and is headed by a paediatrician assisted by a physician and ten nurses working in teams of two to three. The Ebolowa health district in which the hospital is located is a heavily malaria-infested area characterized by a stable, perennial malaria transmission.Plasmodium falciparum. Other aetiologies of the presenting symptoms were excluded and parents gave written informed consent. Excluded from the study were children who have had prior side-effects to either artesunate or quinine administration, severely malnourished children, and those who had a concomitant infection.This was an open-label, randomized, clinical trial carried out from 1 September, 2013\u201331 March, 2014. Included in the study were all children with malaria aged 3 months to 15\u00a0years irrespective of sex, presenting with one or more signs of severity of malaria according to the 2013 Cameroon NMCP\u2013adopted criteria  and having an initial positive parasitaemia to Eligible patients were randomly assigned to receive either parenteral artesunate or one of the three quinine regimens. Using Kendall and Babington Smith random number table, an assistant not involved in the study performed the randomization in advance in blocks of 20 composed of five of each regimen. Treatment allocations were placed in numbered, opaque, sealed envelopes to which the investigator was blinded until a patient was admitted. On admission, each patient was attributed an envelope corresponding to his unique identification number.0) was done to determine the Plasmodium species and the parasite load, respectively. The thin and thick films were stained in 10\u00a0% Giemsa solution for 10\u201315\u00a0min and read at \u00d7100 after oil immersion. The parasite load was determined by counting the number of parasites in at least 20 oil immersion fields. Each slide was double-checked blindly and was considered to be negative if no parasites were detected in at least 20 oil immersion fields. The following investigations were also done systematically on admission: capillary glycaemia, a urine dipstick for children aged between 3 months and 5 years and a full blood count. A lumbar puncture was systematically done to patients with neurological symptoms to rule out meningitis. Chest X-ray, blood group and rhesus typing, blood urea nitrogen, creatinine, and bilirubin levels were done depending on the clinical presentation. After obtaining samples for investigation, treatment was initiated while awaiting the results of the blood film. If the initial film was negative, the child was excluded from the study and the initiated treatment was continued.Before recruitment of patients for the study, a pre-study training pilot study was carried out from 13 December, 2012\u201331 January, 2013. During the study period, any patient admitted for malaria in the paediatric unit with one or more signs of severity of malaria was immediately re-evaluated and anthropometric and vital parameters taken. An initial thin and thick blood film  and then mixed with 5\u00a0ml saline solution (provided with the drug) before injecting into an indwelling intravenous catheter. For the second group (QLD) the patients received a loading dose of 16.6\u00a0mg/kg bw of QB by intravenous (IV) infusion over 4 hours followed 8 hours after the start of the loading dose, with a maintenance dose of QB at 8.3\u00a0mg/kg bw over 4 hours every 8 hours. Patients in the third group (QNLD3) received 8.3\u00a0mg/kg bw of QB over 4 hours by IV infusion every 8 hours. Those in the fourth group (QNLD2) received 12.5\u00a0mg/kg bw of QB over 4 hours by IV infusion every 12\u00a0h. For the three quinine regimens, quinine was diluted in 10\u00a0ml/kg bw of 10\u00a0% dextrose. Clinical side-effects of the drugs were monitored during the treatment. Patients received a minimum of 24\u00a0h of parenteral treatment and exited from the protocol if they had clinically recovered and parasitaemia was negative. Thereafter, oral anti-malarials (artemether-amodiaquine or artemether-lumefantrine) were prescribed for 3 days to complete the treatment. Resuscitation and supportive management was done in accordance with WHO guidelines.Patients were randomized into four groups. Those of the first group (ARTES) received 2.4\u00a0mg/kg body weight (bw) of parenteral artesunate at HFrom the onset of treatment, the temperature was monitored every 6 hours until normalization; the state of consciousness was evaluated every 2 hours for the first 24\u00a0h then every 6 hours until normalization, using the Glasgow coma scale (GCS) or Blantyre coma scale (BCS) in children under 5 years of age. Parasitaemia was measured every 6 hours until it became negative and the time the child was able to sit unsupported and to eat if this was not possible on admission was also recorded.24).Endpoints of the study were clinical and parasitological outcome. The clinical outcome variables were: fever clearance time , coma recovery time ((CRT) time from the onset of treatment until the patient was fully conscious), time to sit unsupported, time to eat and time to exit from the protocol . The parasitological outcome variables were: parasite clearance time (PCT\u00a0=\u00a0time from the onset of treatment to the time of the first of two successive negative blood smears) and the parasite reduction rate 24\u00a0h after onset of treatment . A two-group comparison  of the outcome variables was done and Student Authorization from the Director of ERH was obtained before starting the study in the paediatric unit of the hospital. Ethical clearance was obtained from the Ethics Committee of the Faculty of Medicine and Biomedical Sciences. The study procedure and justification was explained to the parents and an informed written consent from the parents was obtained before enrolment of the children in the study. For confidentiality, code names were given to all patients. During the study, the work sheets were kept secret by the investigators from people not involved in the study, in order to respect confidentiality of collected data. The information provided was used for this study only and was not shared for any other purposes or projects. Blood films, capillary glycaemia, urine dipstick and artesunate ampoules were done at the expense of the investigators.A total of 281 children admitted for severe malaria at ERH were assessed for eligibility. Following diligent scrutiny of the inclusion and exclusion criteria, 238 patients were enrolled in the study. Fifty-nine were randomized to the ARTES (artesunate) group, 57 to QLD (quinine loading dose) group, 62 to QNLD3  group, and 60 to QNLD2  group. Figure\u00a0Data were analysed primarily by intention-to-treat for the 238 patients enrolled in the study and secondarily per protocol for the remaining sub-set of patients who completed the treatment assigned (n\u00a0=\u00a0116).Clinical and laboratory parameters on admission did not significantly differ between the artesunate and the quinine groups , severe anaemia (37.0\u00a0%) and convulsions 28.0\u00a0%) were the predominant manifestations of severe malaria in this setting  was significantly shorter for artesunate than for the three quinine regimens. Parasite reduction rate 24\u00a0h after onset of treatment was significantly higher for patients in ARTES arm (92.0\u00a0\u00b1\u00a010.4\u00a0%) than for patients in QLD arm , QNLD3 arm , and QNLD2 arm   and an episode of generalized tonico-clonic seizures lasting less than 30\u00a0min, following an eight-day history of fever. The patient had hyperparasitaemia at 285,300 TPf/\u00b5l and the CSF analysis was normal. The patient developed severe respiratory distress and died 4 hours after onset of treatment.The patient in QNLD2 group was a 20\u00a0months old boy who was also admitted with severe anaemia (packed cell volume\u00a0=\u00a011\u00a0%) and two episodes of generalized tonico-clonic seizures lasting less than 30\u00a0min, following a 5-day history of fever. The patient also had hyperparasitaemia at 272,500 TPf/\u00b5l and the CSF analysis was normal. The patient went into coma and died 18\u00a0h after starting treatment.This small sample size study reveals that artesunate is more effective than quinine in the treatment of severe malaria in Cameroonian children. Among artesunate-treated patients, fever clearance time, time to exit from the protocol and PCT were shorter; the parasite reduction rate 24\u00a0h after onset of treatment was higher than for quinine-treated patients. The four groups did not differ in time to sit unsupported, time to eat and CRT.Fever clearance time (FCT) was shorter for patients in ARTES arm, followed by QLD arm, then QNLD2 arm, and finally QNLD3 arm. Considering ARTES as the reference arm, the two-by-two group comparison showed significant differences in FCT between artesunate and the three quinine groups. This is a consistent finding with studies comparing artesunate with the loading dose regimen of quinine \u20139. This The CRT in ARTES arm was longer than in the QNLD3 and QNLD2 arms and shorter than in the QLD arm, but these differences were not statistically significant. The CRTs obtained for the four treatment groups were all higher than that observed by other authors , 11\u201313: Time to sit unsupported and time to eat were shorter for artesunate-treated patients than for quinine-treated patients, but the differences were not significant. This is probably because of the small number of patients followed up for these variables. Dondorp et al.  also obtTime to exit from the protocol was significantly shorter for artesunate-treated patients than for quinine-treated patients. This was likely due to the faster parasite clearance with artesunate, which leads to a faster clinical recovery.PCT was significantly shorter for ARTES group as compared to QLD, QNLD3 and QNLD2 groups. Hatim et al. in Sudan  obtainedParasite reduction rate 24\u00a0h after onset of treatment was significantly higher for patients in the ARTES arm. This information is important because, according to WHO, death from severe malaria often occurs within hours of admission to the hospital, so it is essential that therapeutic concentrations of a highly effective anti-malarial be achieved and maximum parasite clearance be obtained within the first 24\u00a0h .Some weaknesses should be noted in this study. Firstly, the differences between the groups may have been underestimated because of the small sample size. Secondly, patients had to be na\u00efve to the drugs used in the study but patients who took anti-malarials before admission to the hospital were included in the study because of time limitation and excluding them would have considerably reduced the sample size. However, most of the patients (40/54) who were treated with the study drugs to completion received anti-malarial drugs (quinine tablets or syrup or an artemisinin combination therapy (ACT)) at inadequate doses before admission. Thirdly, only short-term monitoring in the hospital during the hospital stay was done.This trial, in a population of Cameroonian children, shows that artesunate is more effective than quinine in the treatment of severe malaria. These results, together with those of the largest trials on artesunate (AQUAMAT and SEAQUAMAT), should reassure practitioners about the beneficial effects of artesunate over quinine and serve as strong motivation to prescribe artesunate, when available, as first-line drug to patients with severe malaria."}
+{"text": "Plasmodium gametocyte carriage. However, the role of artesunate in monotherapy in vivo, the mechanisms involved, and the utility of gametocyte carriage as a potential tool for the surveillance of antimalarial resistance are poorly understood. In 2010\u20132011, we conducted an open-label, prospective efficacy study of artesunate as monotherapy in children 1\u201310\u00a0years of age with uncomplicated falciparum malaria in Bougoula-Hameau, Mali. Standard oral doses of artesunate were administered for 7 days and patients were followed up for 28 days. The data were compared to a similar study conducted in 2002\u20132004. Of 100 children enrolled in the 2010\u20132011 study, 92 were analyzed and compared to 217 children enrolled in the 2002\u20132004 study. The proportion of gametocyte carriers was unchanged at the end of treatment  and did not significantly decline until day 21 of follow-up . The mean gametocyte density at inclusion remained unchanged at the end of treatment . Overall, 46% of the 71 initial non-carriers had gametocytes detected by day 7. Similar results were found in the 2002\u20132004 study. In both studies, although gametocyte carriage significantly decreased by the end of the 28-day follow-up, artesunate did not clear mature gametocytes during treatment and did not prevent the appearance of new stage V gametocytes as assessed by light microscopy. Baseline gametocyte carriage was significantly higher 6\u00a0years after the deployment of artemisinin-based combination therapies in this setting.Artemisinin-based combination therapies decrease  Plasmodium falciparum. Gametocyte formation within the host involves the differentiation of sexually committed merozoites that undergo five distinct developmental stages (stage I\u2013V) over a period of ~10 days \u00a0h . In comparison, the median GCT in 2002\u20132004 was 168\u00a0h  (n\u00a0=\u00a071) had gametocytes detected at some point between day 0 and day 7 of follow-up. Of those, 10 (14.1%), 13 (18.3%), 10 (14.1%), and 6 (8.5%) had become carriers by follow-up days 1, 2, 3, and 7, respectively. Similarly, in the 2002\u20132004 study, among non-gametocyte carriers at inclusion 35.9% (n\u00a0=\u00a064) had gametocytes by day 7 of follow-up. Among those, 7 (10.9%), 10 (15.6%), and 6 (9.4%) became carriers on follow-up days 1, 3, and 7, respectively. Inter-study comparison between 2010\u20132011 and 2002\u20132004 limiting the analysis to patients recruited during the same months of December and January in each period showed no statistically significant differences in the proportion of patients with de novo gametocytemia .Among patients without gametocytemia at inclusion in the 2010\u20132011 study, 46.5%  of our initial non-gametocyte carriers did develop gametocytemia at some point by day 7 of follow-up, much higher than findings of 9% in a similar study in Vietnam . Two preAmong the gametocyte carriers at enrollment, our median gametocyte clearance time (GCT) of 336\u00a0h in December\u2013January of both 2010\u20132011 and 2002\u20132004 is higher than the median GCT of 163\u00a0h in a pooled analysis of nine Thai trials of ACTs . This diWe found a higher proportion of initial gametocyte carriers in the 2010\u20132011 study than in the entire 2002\u20132004 study, which may be due to the fact that the latter study was a year-round study while the former was done at the end of the malaria transmission season in this area. Indeed, the proportion of gametocyte carriers was shown to increase throughout the malaria transmission season . HoweverAn increased duration and density of gametocyte carriage after sulfadoxine-pyrimethamine treatment was proposed as an early indicator of drug resistance . In thisP. falciparum isolates from sub-Saharan Africa [K13 propeller polymorphisms are considered as the best molecular markers of artemisinin resistance . Howevern Africa , 24, 41.n Africa . In lighThe primary limitation of our study is the likelihood of type II bias due to a rather small sample size. Another limitation is our reliance on standard microscopy for gametocyte detection, which is known to consistently underestimate gametocyte carriage . The abiin vitro efficacy of artemisinins against gametocyte populations over time to detect early signs of resistance [Although gametocyte density does not perfectly correlate with infectivity , becausesistance .This study provides baseline gametocyte clearance dynamics when artemisinins are still highly efficacious in sub-Saharan Africa. Monitoring changes in these clearance dynamics may be used as an additional early warning tool in the surveillance of artemisinin resistance in the field."}
+{"text": "Plasmodium falciparum malaria in Malawi, a therapeutic efficacy trial was conducted.Malaria causes significant morbidity in Malawi, with an estimated 5 million cases in 2014. Artemether-lumefantrine (AL) and artesunate-amodiaquine (ASAQ) are the first- and second-line treatments for uncomplicated malaria, respectively, but emerging resistance threatens their efficacy. In order to understand whether AL and ASAQ remain efficacious for the treatment of uncomplicated P. falciparum malaria  were enrolled in a 28-day randomized in vivo efficacy trial at three sites: one each in northern (Karonga), central (Nkhotakota) and southern (Machinga) Malawi. The study was powered to estimate site-specific efficacy for AL and overall efficacy for ASAQ, with 3:1 randomization to AL or ASAQ. Blood was collected for malaria microscopy and molecular testing on days 0\u20133, 7, 14, 21, and 28. Recrudescence and reinfection were differentiated using polymerase chain reaction (PCR) genotyping of merozoite surface protein. The primary outcome was the PCR-corrected day 28 Kaplan\u2013Meier cumulative success rate.During March\u2013July 2014, febrile children aged 6\u201359\u00a0months with microscopy-confirmed uncomplicated A total of 452 children were enrolled; 303/338 (89\u00a0%) and 98/114 (86\u00a0%) reached a study endpoint in AL and ASAQ arms, respectively. All treatment failures occurred after day 3. The day 28 uncorrected cumulative success rate was 97.1\u00a0%  for ASAQ and 76.8\u00a0% (95\u00a0% CI 72.1\u201381.5\u00a0%) for AL, with 82.5\u00a0% (95\u00a0% CI 75.4\u201389.7\u00a0%), 69\u00a0% (95\u00a0% CI 59.9\u201378.1\u00a0%), and 78.2\u00a0% (95\u00a0% CI 70.2\u201386.3\u00a0%) success in the northern, central, and southern regions, respectively. The day 28 PCR-corrected cumulative success rate was 99\u00a0% (95\u00a0% CI 97.2\u2013100\u00a0%) in the ASAQ arm and 99.3\u00a0% (95\u00a0% CI 98.3\u2013100\u00a0%) in the AL arm, with 98\u2013100\u00a0% efficacy in each site.As evidenced by the day 28 PCR-corrected cumulative success rates, both AL and ASAQ remain efficacious treatments for uncomplicated malaria in Malawi. The lower uncorrected efficacy in the AL arm compared to ASAQ may be explained by the shorter half-life of lumefantrine (3\u20136\u00a0days) compared to amodiaquine (9\u201318\u00a0days). The high reinfection rate suggests that there is a continued need to scale-up effective malaria prevention interventions. Plasmodium falciparum [P. falciparum malaria [Malaria is a cause of considerable morbidity and mortality in Malawi, with 5 million cases in 2014. The vast majority of these cases and deaths were caused by  malaria \u20136.P.falciparum malaria in children aged six to 59\u00a0months was conducted in three sites in Malawi.In 2010, the overall efficacy of AL across six sites in Malawi was 93.4\u00a0% , suggestThis study was conducted between March and July of 2014 at three district-level hospitals: Karonga District Hospital in the northern region, Nkhotakota District Hospital in the central region, and Machinga District Hospital in the southern region. Malaria transmission is stable throughout the year with a peak in the rainy season, December to April.The minimum sample size needed to achieve a precision to detect 5\u00a0% failures, at a confidence level of 95\u00a0%, assuming a drug efficacy of 95\u00a0%, was 94 participants per arm. The target sample size was increased to 113 per arm to allow for approximately 15\u00a0% loss to follow up. As AL is the first-line treatment and widely used, the study was powered to provide regional estimates for AL efficacy and a country-wide estimate for ASAQ, resulting in a target sample size of 452 participants, 339 in the AL arm and 113 in the ASAQ arm.\u00ae, Orchid Biomedical Systems, Goa, India). If the RDT was positive, two thick and one thin blood smears were obtained; an initial rapid reading of the thick smear was used to determine whether the patient met the eligibility criteria for parasitaemia, after enrollment the remaining smears were stained more carefully to obtain more accurate parasite estimates and species. Children who were RDT-positive but ineligible for enrollment were treated with AL according to national guidelines. Children with microscopically detected P. falciparum mono-infection with 1000\u2013200,000 asexual parasites/\u03bcl were eligible for enrollment provided that they were able to swallow oral medication, had no danger signs, including severe malnutrition, reported no allergies to the study medications, were not taking any medications that could interfere with the study drugs, and the parents/guardians provided informed consent.The standard in vivo WHO protocol was used . ChildreAfter enrollment, children were randomized 3:1 to receive either AL  or ASAQ . The randomization list was generated for each site using permuted-block randomization in the R (version 3.2.0) blockrand package . Study staff in Malawi did not have access to this list. Cards with the treatment assignment were created based on the list and concealed in opaque envelopes that were opened at the time of first administration. Treatment was directly administered with clean water following manufacturer\u2019s prescribed weight-based dosing, twice daily for AL  and daily for ASAQ  for 3\u00a0days. Participants were observed for 30\u00a0min after treatment to monitor for vomiting or other adverse events; those who vomited were administered a second dose and observed for an additional 30\u00a0min. Patients who vomited both doses were removed from the study and referred to the hospital for parenteral treatment. Study staff directly observed all three ASAQ doses and all three morning AL doses. Parents or guardians were given the evening AL doses with instructions for administering the drug; parents were asked to verify administration at the visit the following day.Patients were seen routinely on day 0 (enrollment), days 1, 2, 3, 7, 14, 21 and 28 , as wellAt enrollment, two slides were obtained. The first slide (thick smear only) was stained rapidly, using 10\u00a0% Giemsa for 10\u201315\u00a0min, and parasitaemia counted to determine eligibility. In patients deemed eligible for enrollment, the second slide (thick and thin smears) was stained with 2.5\u20133\u00a0% Giemsa for 45\u201360\u00a0min and re-read; the results from the second slide constituted the final enrollment parasitaemia. For all subsequent visits, only the slower stain methodology was used.Two on-site microscopists independently read the smears according to WHO protocol, each was blinded to the other\u2019s read . If specPatients were classified according to the WHO definition of therapeutic responses . Early tmsp1) and merozoite surface protein 2 (msp2), using WHO recommended PCR procedures [In order to differentiate between reinfection and recrudescence, DNA samples extracted from paired dried blood spot samples from day 0 and the day of recurrence were subjected to nested PCR for two microsatellite markers, merozoite surface protein 1  were considered significant for all statistical tests. Greenwood\u2019s formula was used to calculate 95\u00a0% CIs for Kaplan\u2013Meier cumulative success rates. All data analysis was done using SAS version 9.3 .Written, informed consent was obtained from the legal guardian of each study participant. This study was approved by the ethical review boards at the Centers for Disease Control and Prevention (CDC) in Atlanta, GA (protocol #6029) and the Malawi College of Medicine in Blantyre, Malawi (protocol #P.11/13/1486).A total of 1561 children were screened; 925 (59.7\u00a0%) were RDT positive and, of those, 452 (48.8) were enrolled  had parasitaemia greater than 200,000 parasites/\u03bcL. These participants were eligible for study inclusion based on the initial screening of the thick smear but were later found to have an enrollment parasitaemia higher than 200,000 parasites/\u03bcL according to a final study review. All analyses were performed both with  measured on day 0 to 10.12\u00a0g/dL (95\u00a0% CI 10.0\u201310.3) on day 14, reaching 10.8\u00a0g/dL (95\u00a0% CI 10.6\u201311.0) by the last visit on day 28  of participants vomited at least one dose; four patients (1.2\u00a0%) vomited the repeat dose and were withdrawn. In the ASAQ arm, 12 (10.5\u00a0%) vomited at least one dose; 1 (0.9\u00a0%) vomited the repeat dose and were withdrawn. No significant differences in vomiting the first or second dose were found between study arms. All patients who vomited second doses of either treatment were given rescue treatment. AEs during drug administration were infrequent in both arms, with a maximum of 5\u00a0% of patients reporting an AE on day 1, 2, or 3. The most commonly reported AEs were cough, fever, and upper respiratory tract infection. There were a total of four SAEs. One participant in the ASAQ arm was treated with parenteral quinine on Day 1 for a worsening clinical picture. Unfortunately, blood was not drawn for a parasitaemia estimate. Three participants (two in AL arm and one in ASAQ arm) were hospitalized for treatment of pneumonia; none of these were thought to be caused by the study drug.Among ASAQ participants, the median day 0 ALT was 42.3\u00a0U/L  and the median AST was 50.6 U/L   compared to amodiaquine (6\u201318\u00a0days) , 16. In While there were differences in uncorrected AL efficacy between sites, these were not statistically significant. However, this study was not powered to detect differences by site, and these differences may have been significant in an appropriately powered study. Reinfections did differ by site, with significantly more reinfections in Nkhotakota compared to the other sites. The higher number of reinfections might be related to lower reported use of ITNs among children sampled from Nkhotakota 64.2\u00a0%) compared to those from Machinga (91\u00a0%) and Karonga (89.1\u00a0%). Alternatively, the higher number of reinfections might reflect a higher transmission intensity in that study site, although malaria prevalence estimates from the 2014 Malaria Indicator Survey did not show large differences between sites  [\u00a0% comparAs expected, haemoglobin levels among children randomized to both treatments improved significantly over the 28-day follow-up. Both AL and ASAQ were well-tolerated and safe, as has been previously reported , 17\u201319. This therapeutic efficacy study had several limitations. First, several participants with a final parasitaemia greater than 200,000 parasites/\u03bcl were included, as these patients were deemed eligible according to the initial microscopy reading. Treating these participants with AL or ASAQ could have increased the risk of delayed parasite clearance and recrudescence as parasite biomass might be too high to be adequately cleared . ReassurP. falciparum malaria in Malawi. There is no evidence at this time that a change in regimens is warranted. However, continued monitoring of drug efficacy, following WHO recommendations, is needed. Finally, the relatively high reinfection rates observed during the study period are evidence of the continued need to scale-up the ownership and use of effective malaria prevention interventions, such as ITNs.The results of this study demonstrate that AL and ASAQ remain efficacious treatments for uncomplicated"}
+{"text": "Plasmodium falciparum malaria in two sentinel high malaria transmission districts in the Eastern Province of Zambia in persons aged six months and above, excluding women aged 12 to 18\u00a0years.Anti-malarial drug resistance continues to be a leading threat to ongoing malaria control efforts and calls for continued monitoring of the efficacy of these drugs in order to inform national anti-malarial drug policy decision-making. This study assessed the therapeutic efficacy and safety of artemether-lumefantrine (AL)(Coartem\u00ae) for the treatment of uncomplicated Plasmodium falciparum mono-infection. A World Health Organization (WHO)-standardized 28-day assessment protocol was used to assess clinical and parasitological responses to directly observed AL treatment of uncomplicated malaria. DNA polymerase chain reaction (PCR) analysis for molecular markers of AL resistance was conducted on positive blood samples and differentiated recrudescence from re-infections of the malaria parasites.This was an observational cohort of 176 symptomatic patients diagnosed with uncomplicated All patients (CI 97.6-100) had adequate clinical and parasitological responses to treatment with AL. At the time of enrolment, mean slide positivity among study participants was 71.8% and 55.2% in Katete and Chipata, respectively. From a mean parasite density of 55,087, 98% of the study participants presented with zero parasitaemia by day 3 of the study. Fever clearance occurred within 24\u00a0hours of treatment with AL. However mean parasite density declines were most dramatic in participants in the older age. No adverse reactions to AL treatment were observed during the study.Plasmodium falciparum malaria in Zambia, endemic for malaria, with some provinces experiencing high transmission intensity. However, the delayed parasite clearance in younger patients calls for further sentinel and periodical monitoring of AL efficacy in different areas of the country.AL remains a safe and efficacious drug for the treatment of uncomplicated  Malaria is a parasitic infection endemic in Zambia. It is reported to be among the 10 top causes of morbidity and mortality in health facilities in the country and thisThe policy change from CQ to AL in 2003 was well accepted by users on account of its proven therapeutic efficacy even against multi-drug resistant parasites, optimal tolerability and safety profile. However, continued use of these anti-malarial medicines including, AL is threatened by the development of parasite resistance. However, currently, there are no alternative anti-malarial drugs available with proven therapeutic efficacy even against multi-drug resistant parasites, optimal tolerability and safety profile as artemisinin-based combinations. In ordeThis study was conducted to provide efficacy and safety data on artemether-lumefantrine following a standard WHO 28-day follow-up therapeutic efficacy and safety protocol to guide the Zambian anti-malarial treatment policy.This was a one-arm prospective study conducted in May 2012. The study assessed clinical and parasitological responses after administration of anti-malarial treatment with AL to eligible patients aged six months (>5kgs weight) and above, excluding women of the age group (12 to 18\u00a0years) suffering from uncomplicated falciparum malaria.The study was conducted in two primary health facilities located in Chipata and Katete districts of Eastern Province in Zambia Figure\u00a0. These sPlasmodium falciparum malaria infection seeking care at the selected study primary health care facilities who were aged six months (>5kgs weight) and above, excluding women of the age group 12 to 18\u00a0years as requesting this age group to take a pregnancy test and initiate contraception is not acceptable in the local context. All enrolled patients were treated with AL on site as directly observed treatment and monitored for 28\u00a0days, as per recommendation for evaluating clinical and parasitological response for drugs such as AL that have a half-life of less than seven days[The population consisted of consenting patients with uncomplicated ven days. Health Assuming a 5% treatment failure rate to AL, a 95% confidence level and a precision around the estimate of 5%, 73 patients were targeted as a minimum for inclusion into the study. With a 20% increase to allow loss to follow-up and withdrawals during the 28-day follow-up period, 87 patients were planned to be included into the study. The study recruited a total of 177 patients overall.P. falciparum detected by microscopy at parasitaemia of 1,000 to 200,000/\u03bcl asexual forms, axilliary temperature \u226537.5\u00b0C, willing to comply with the study protocol for the duration of the study were included. A detailed inclusion criteria is provided elsewhere[Symptomatic patients aged six months (>5kgs weight) and above, excluding women of the age group 12 to 18\u00a0years, self-presenting to health facilities with uncomplicated malaria due to mono-infection of lsewhere.; unable to drink, or breast feed (in case of children), severe vomiting; reported history of convulsion seven days prior to patient contact; presence of lethargy or decreased consciousness; inability to sit or stand, were all excluded. Patients who failed to complete treatment due to persistent vomiting of the treatment or failed to attend scheduled visits during the first three days or withdrew their consent were also excluded. A detailed list of exclusion criteria is provided elsewhere[Patients with general danger signs or signs of severe falciparum malarialsewhere.Parents or guardians of children were instructed to return to the health centre at any time if they had any general danger signs as described under exclusion criteria above. The study team made home visits as follow ups for study participants that were late for their scheduled visits. Patients who failed to return on days 1 and 2 and missed one dose of the treatment or enrolled patients who could not attend scheduled visits were considered lost to follow up (LFU) and excluded from the final analysis.AL was obtained from WHO and administered by a qualified Medical Officer following a treatment regime of two daily doses for three days based on the patient\u2019s weight. The dayOn the basis of parasitological and clinical outcome of treatment with AL, patients were classified according to the WHO definition of therapeutic responses.The development of danger signs for severe malaria on days 1, 2, or 3 in the presence of parasitaemia; parasitaemia on day 2 higher than the day 0 count irrespective of axilliary temperature; parasitaemia on day 3 with axilliary temperature \u226537.5\u00b0C; parasitaemia on day 3\u2009\u2265\u200925% of count on day 0.The development of danger signs for severe malaria after day 3 in the presence of parasitaemia, without previously meeting any of the criteria of ETF; presence of parasitaemia and axilliary temperature \u226537.5\u00b0C or history of fever on any day from day 4 to day 28, without previously meeting any of the criteria of ETF.The presence of parasitaemia on any day from day 7 to day 28 and axilliary temperature <37.5\u00b0C, without previously meeting any of the criteria for ETF or LCF.It is the absence of parasitaemia on day 28 irrespective of axilliary temperature without previously meeting any of the criteria for ETF, LTF, or LPF. The secondary outcomes were fever clearance rate; proportion of patients who had fever cleared on days 1, 2, and 3. Parasite clearance rate: proportion of patients with negative thick blood film smears on days 1, 2, and 3; Gametocyte carriage: proportion of patients with gametocytes during the course of the study.msp1, msp2 and glurp. The technique differentiated \u201crecrudescence\u201d from a newly acquired infection through a comparison of pre- and post-parasite strain genotype profiles. Details of the protocol used in this analysis are described in detail elsewhere[Genotype analysis (nested PCR amplification) was conducted at the National Malaria Control Centre molecular laboratory based on genetic diversity among the malaria parasite genes lsewhere.Data from both clinical and parasitological assessments from the case report for each study participant were entered into the WHO standardized Microsoft excel data collection form. This foApproval to conduct the study was obtained from the Tropical Diseases Research Centre ethics committee based in Ndola, Zambia. Permission to conduct the study was obtained from the Ministry Of Health. Informed consent was sought from all study participants. Guardians provided assent for the participation of the persons under the consenting age. They were asked to read or have read to them, understand and sign/thumbprint an informed consent form. The consent was available in English and explained in vernacular to the patient, parent or guardian. The benefits and potential risks were explained and a signature requested on the consent form. Confidentiality was maintained through ensuring all patients\u2019 information used unique identifiers. All data collection forms were stored in locked files.A total of 142 and 230 eligible patients from Katete and Chipata, respectively, were screened during the study. The slide positivity rate was 71.8% (102/142) and 55.2% (127/230) respectively for Katete and Chipata, which translated into an overall slide positivity of 61.6% (229/372) for both study sites. Following the eligibility requirements, a total of 177 (88 Katete and 89 Chipata) were enrolled into the study  as 89.5% CI 83.9 - 93.6) in patients treated with AL. No patients showed early treatment failures (ETF), while late clinical failure (LCF) was reported in 6 study participants ). Late parasitological failure (LPF) was observed in 12 ) of the evaluated study population ) of the patients had ACPR to the AL treatment Table\u00a0. The KapPlasmodium falciparum parasitaemia were recorded in the under-five age group on days 3 to day 28 as follows: Day3: 2.2% (2/90), Day 7: 3.3% (3/90), Day 14: 1.1% (1/90) , Day 21: 6.7% (6/90) and Day 28: 4.4% (4/90). Likewise, parasitaemia was reported in the age group 5 to 15 on Day 3: 6.4% (4/63) and Day 28: 9.5% (6/63)  reduction of mean parasitaemia was observed in all age groups on Day 1. However, on Day 2 under five children showed 99.7% and the 5 to 15 age groups showed 98% decline. P. falciparum malaria in all ages in areas with high malaria transmission intensity. The study comes nine years after its adoption as first-line anti-malarial in Zambia[This study has demonstrated that fixed dose artemether-lumefantrine is effective for the treatment of uncomplicated n Zambia. The hign Zambia\u201314.in vivo to artemisinins has been reported elsewhere, particularly on the Thai-Cambodian border[WHO recommends a review of treatment policy for uncomplicated malaria at 10% treatment failure, 16. Thin border, 19.Pre-treatment gametocytaemia was cleared by day 7, indicating that AL is still effective with regard to gametocyte clearance, and, suggesting a potential role in transmission reduction. Thus further suggesting a continued role in malaria control for therapeutic interventions, particularly with current calls for malaria elimination in most endemic countries, 21.Zambia is among a few countries in the sub-region which has conducted or maintained regular studies in its sentinel sites in different areas across the country representing the different geographical and epidemiological profiles to monitor therapeutic efficacy of anti-malarial medicines. In the past three to four years efficacy studies faced a challenge on account of reduced malaria cases, which made it difficult and expensive to attain the required sample size. Therefore, to achieve the required sample size in this study, for the first time in Zambia, persons of all ages were enrolled rather than restricting the study to children under the age of five years as was the case previously. Thus inP. falciparum transmission (>15% parasitaemia) area in Zambia.A key limitation of the study was the lack of a corresponding assessment in anaemia in the study participants, which would have provided data on longitudinal effects of malaria parasite infection in the study areas. An important strength of the study is that the study provides vital data collected in a high Artemether-lumefantrine remains a safe and effective drug for the treatment of uncomplicated falciparum malaria in Zambia. AL is well tolerated in all ages when administered. The efficacy of this ACT needs to be carefully monitored periodically since treatment failures can occur due to resistance as well as sub- therapeutic levels due to non-compliance of therapeutic dosage. It is also important to strengthen routine monitoring of anti-malarial efficacy particularly in light of planned elimination efforts which will in part depend on therapeutic interventions."}
+{"text": "Plasmodiumfalciparumk13(pfk13) gene and delayed parasite clearance after artemisinin treatment. In Africa, P. falciparum remains susceptible and combination therapy regimens which include an artemisinin component display good efficacy. Using quantitative real-time PCR (qPCR), sub-microscopic persistence of P. falciparum has previously been reported in one-third of children treated with artemisinin combination therapy (ACT) in western Kenya. In this study, further investigation was made to evaluate whether these sub-microscopic residual parasites also harbour mutations at the propeller region of pfk13 and whether the mutations, if any, affect treatment outcome.Studies in Southeast Asia reported a strong relationship between polymorphisms at the propeller domain of the Kelch 13 (K13) protein encoded by the pfk13 propeller domain was genotyped in DNA samples obtained in 2009 from Kenyan children treated with artemether\u2013lumefantrine (AL) and dihydroartemisinin\u2013piperaquine (DP). Paired samples at pre-treatment (day 0) and day of treatment failure (day 28 or 42) for 32 patients with documented recurrent parasitaemia were available for genotyping. Additional day 3 DNA samples were available for 10 patients.The pfk13-propeller gene in the day 0 sample, but this allele was replaced by the wild-type (A578) form on day 3 and on the day of recurrent parasitaemia. The mutation at amino acid codon 578 showed no association with any phenotype. Polymorphisms in pfk13 were not responsible for parasite persistence and gametocyte carriage in the children treated with ACT.No mutation associated with artemisinin resistance in Southeast Asia was observed. Only one DP-treated patient harboured a non-synonymous mutation at codon 578 (A578S) of k13-propeller mutations in western Kenya with samples collected from a longitudinal study.This study contributes to the ongoing surveillance of suspected artemisinin resistance parasites in Africa by providing baseline prevalence of Clinical Trials Registration NCT00868465. Plasmodium falciparum malaria in endemic areas. Significantly reduced susceptibility to artemisinins, characterized by delayed parasite clearance in vivo and enhanced survival of early stage parasites in vitro, has been reported in Southeast Asia [pfk13 gene of P. falciparum, encoding the kelch-domain protein K13 [pfk13 loci [Artemisinin combination therapy (ACT) is widely used to treat uncomplicated ast Asia , 2. Mostk13 loci . This wok13 loci .pfk13 mutations [pfk13 mutations in 14 sub-Saharan African countries. The study reported the absence of mutations that were associated with artemisinin resistance in Southeast Asia, with the exception of codon 543 [P. falciparum parasites in western Kenyan children on day 3 after ACT treatment was reported and this was associated with subsequent recrudescence and transmission [P. falciparum parasites carrying certain genotypes at pfmdr1, pfcrt, pfubp1, and pfap2mu genes were found to survive more often after ACT treatment at sub-microscopic level [pfk13 and whether the mutations, if any, affect treatment outcome.In Africa, ACT is the first-line anti-malarial treatment and remains efficacious and safe , 6. Howeutations . Recentlodon 543 . In prevsmission . In addiic level . The aimpfk13 propeller domain polymorphism in DNA samples available from the 2009 study at the following time points: for 32 patients with documented recurrent parasitaemia, paired samples at day 0 (pretreatment) and day of treatment failure ; for 10 patients, additional day 3 DNA samples (Table\u00a0DNA samples were previously obtained from filter-paper blood spots from children (age 8\u00a0months\u201310\u00a0years) treated with either artemether\u2013lumefantrine (AL) or dihydroartemisinin\u2013piperaquine (DP) in a clinical trial carried out in 2009 in Mbita, western Kenya and fully described elsewhere , 10. The\u00ae blend master mix  and 5\u00a0\u03bcl of DNA in a total volume of 20\u00a0\u00b5l was run under the following cycling conditions: 95\u00a0\u00b0C for 15\u00a0min then 30 cycles at 95\u00a0\u00b0C for 30\u00a0s, 58\u00a0\u00b0C for 2\u00a0min and extension at 72\u00a0\u00b0C for 2\u00a0min and final extension at 72\u00a0\u00b0C for 10\u00a0min. For the second round of PCR, a total volume of 25\u00a0\u03bcl made of 18.75\u00a0\u03bcl nuclease free water, 250\u00a0nM primer, 1X hot fire pol\u00ae blend master mix  final concentration and 5\u00a0\u03bcl of DNA was run under the following cycling conditions: 95\u00a0\u00b0C for 15\u00a0min then 40 cycles at 95\u00a0\u00b0C for 30\u00a0s, 60\u00a0\u00b0C for 1\u00a0min and extension at 72\u00a0\u00b0C for 1\u00a0min and final extension at 72\u00a0\u00b0C for 10\u00a0min. A positive control (K13_5) with one of the mutations identified in Cambodia (provided by D M\u00e9nard) was included in every experiment.The K13-propeller gene fragment (coordinates 1726169\u20131726997 on chromosome 13 of 3D7 isolate (PF3D7_1343700) was amplified by nested PCR using previously published primers [pfk13 were determined by direct sequencing of amplicons using ABI BigDye Terminator v3.1 cycling sequencing kit and analysis on an ABI 3730 sequencer  as described previously [pfk13-propeller sequence region of 3D7 isolate (PF3D7_134700).Polymorphisms at eviously  and chroPfk13 sequences for 32 samples on day 0, seven samples on day 3 and 30 samples on day of failure were successfully obtained and analysed for sequence variation. Of these 69 sequenced isolates, 68 harboured pfk13 loci encoding propeller domains identical to the reference. A single DP-treated patient (K1368) harboured a non-synonymous mutation at codon 578 of pfk13-propeller gene in the day 0 sample, but this allele was replaced by the wild-type form on day 3 and on the day of recurrent parasitaemia (day of failure) (Table\u00a0pfk13 was retrospectively sequenced in day 1 and day 2 parasites from this individual. Sequence analysis of day 0, day 1 and day 2 samples revealed the presence of mutation at codon A578S, suggesting that the mutant parasite was present 2\u00a0days after DP treatment (Figure\u00a0pfk13 codon P553P on day of failure but that was not observed on day 0 or day 3.pfk13-encoded propeller domain, including the major resistance-associated mutation C580Y, contribute to ACT treatment failure in Mbita, western Kenya. Only one of the 32 patients from this longitudinal study had mutation in the propeller region of pfk13 gene, despite evidence of parasite persistence in over 30\u00a0% of children [msp1 and msp2 genotyping of the day 0 sample showed that the patient was infected with at least three different parasite clones. Blood from this patient was presented to Anopheles gambiae mosquitoes at day 7 by membrane feeding, as reported in previous study, but no infected mosquitoes were generated [No evidence was found that artemisinin resistance-associated mutations in the children . One patchildren , 14. In enerated .pfk13 and its lack of association with parasite clearance half-life in patients treated with ACT [pfcrt, pfmdr1, pfap2mu, and pfubp1 also contribute to treatment outcomes after either artemisinin monotherapy or ACT treatment [pfk13 and other candidate loci into parasite isolates with an African genetic background could clarify whether pfk13 mutations or polymorphisms in other genes are most relevant in determining artemisinin sensitivity in clinical isolates in Africa [The A578S polymorphism has previously been reported in a cross-sectional study in western Kenya, Kisumu, and in four other African countries , but thewith ACT . Althougreatment , 16. Stun Africa , 17.pfk13 variants in the current small study precludes from making meaningful assessment of the role of A578S and other propeller domain mutations in treatment outcomes in Africa. However, it can be confirmed that the sub-microscopic clearance phenotype observed in western Kenya is not directly related to the parasite clearance half-life observed in Southeast Asia. The sub-microscopic residual parasites have been shown previously to be important phenotypes as children harbouring those parasites were significantly more likely to be infectious to mosquitoes and were more likely to have recurrent asexual parasitaemia on day 28 or 42 [The rarity of 28 or 42 . Factorsk13-propeller mutations in western Kenya with samples collected from a longitudinal study, and thus one of the first to relate genotype and phenotype. This information from samples collected in 2009 does not reflect the current status of ACT, nor the prevalence of k13-propeller mutations now. Therefore, continuous monitoring of ACT in western Kenya and the collation of phenotypic and genotypic data to track the emergence and spread of parasites with reduced susceptibility to artemisinins is recommend.This study contributes to the ongoing surveillance of suspected artemisinin resistance parasites in Africa by providing baseline prevalence of"}
+{"text": "Based on the recommendations of the World Health Organization in 2004, Ghana changed her antimalarial drug policy from mono-therapy to Artemisinin-based Combination Therapy (ACTs). The country is currently using three first line drugs artesunate-amodiaquine, artemether-lumefantrine and dihydroartemisinin-piperaquine for the treatment of uncomplicated malaria. Despite this policy, little or no qualitative studies have been conducted to establish the factors influencing adherence to the new treatment for malaria. This study explored factors influencing adherence to the use of ACTs in northern Ghana.This was a qualitative study comprising forty (40) in-depth interviews with patients with malaria who visited selected public and private health facilities and received ACTs. Systematic sampling technique was used to select participants who were given ACTs for the interviews. Nvivo 9 software was used to code the data into themes for further analysis.The study revealed very important differences in knowledge about ACTs. As expected, the less or illiterates could not mention the type of ACT they would prefer to use for treating their malaria. The educated ones had a good knowledge on ACTs and preferred artemether-lumefantrinee in treating their malaria. The reason was that the drug was good and it had minimal or no side effects. Individual attitudes toward the use of medications and the side effects associated with the use of these ACTs were found to be the main factors affecting adherence to the use of ACTs. Perceived cure of illness after the initial dose greatly affected adherence. Other factors such as forgetfulness and lack of information also influenced patient adherence to ACTs use.Individual knowledge, attitudes and behaviors greatly influence patients\u2019 adherence to ACTs use. Since ACTs take a number of days to complete, continuous education by health professionals could improve on adherence to ACTs use by patients with malaria. According to the World Health Organization, malaria remains a major public health problem causing about 207 million cases with 627,000 deaths in 2012 mainly in sub- Saharan Africa , 4.Despite these interventions, malaria remains endemic in Ghana and is the single most important cause of morbidity and mortality especially among young children, pregnant women and the poor . About 3falciparum malaria [In early 2000, the World Health Organization (WHO) recommended to all countries experiencing resistance to mono-therapies to use artemisinin-based combination treatments (ACTs) in treating uncomplicated  malaria . Based o malaria , 8.Countries with high malaria burden need robust information to inform policy decisions about the benefits of wider implementation of new products. Large scale studies to determine the effectiveness of new drugs when delivered through real-life settings are therefore necessary. For instance, data on patients\u2019 adherence and factors affecting the use of new anti-malaria drugs when delivered outside trial conditions are needed to inform policy decisions. The INDEPTH Network Effectiveness and Safety Study (INESS) platform was therefore established to providing effectiveness data to inform policy direction .Adherence refers to the extent to which patients use medications as prescribed by health providers and is an important component of infectious disease control , 11. ForIn 2004, Ghana changed the national anti-malarial drug policy recommending the use of artesunate-amodiaquine combination as the first line drug for treating uncomplicated malaria. However, the implementation process was faced with several challenges related to side effects from the use of these drugs and lack of other treatment options . It therThese medications have since been in use for about a decade now. However, little is known about the factors influencing the use and adherence to these treatments. This study was therefore designed to explore the factors influencing adherence to the use of ACTs in treating uncomplicated malaria in real-life settings following the introduction of the three ACTs as first line treatments for uncomplicated malaria in Ghana.Ethical approval for the study was received from the Navrongo Health Research Centre Institutional Review Board (NHRCIRB 152). The board emphasized on the need to maintain confidentiality of Participants\u2019 information. In line with the approved procedure of obtaining consent for the study, oral consent was obtained from each potential participant prior to being interviewed and this was approved as part of the protocol for this study. Oral consent was solicited and obtained as the majority of the respondents had no formal education and those with formal education also opted for this method of consent.The interview moderators read and translated the consent form into the preferred local language of each participant on the purpose of the study, study procedure, right to withdraw and efforts to ensure confidentiality. The oral consenting process was recorded on a separate voice recorder from the one used for the actual interview. In addition, they were made to recommend a member of the household to witness the consenting process and the demographic data of the witnesses were collected. All children from 10 to 17 years old gave assent while their parents/caretakers gave consent before the interviews were conducted.The study was conducted in the Kassena-Nankana East and West Districts by the Navrongo Health Research Centre. The research centre operates the Navrongo Health and Demographic Surveillance System (NHDSS) in the two districts. The districts cover an area of 1,675 square kilometres of Sahelian savannah with a population of about 153,000 . The maiThe districts have one hospital, eight health centers, two private clinics, which provide curative and preventive health care services to community members. There are 28 Community-based Health Planning and Services (CHPS) compounds located in various communities and providing reproductive health services and treatment of minor health conditions , 21. TheThe two districts have been demarcated into five zones by the NHDSS . Two zonAll patients who visited and received an ACT in the selected health facilities/chemical shops qualified to participate in the study. This was to enable patients share their experience on the use of ACTs that they received. The data collectors spent two days in each of the six study health facilities/chemical shops, systematically selecting every second patient who was given an ACT. The longest duration for taking ACTs is three days. All recruited participants were therefore followed-up on the fourth day when all of them were expected to have completed their doses. The longest period for the interviews after treatment was three days after the last dose was supposed to have been taken. In all, 40 interviews were conducted in the six health facilities/chemical shops. The themes explored included preference of ACTs in the management of malaria, factors responsible for non-adherence to the use of ACTs and suggested ways to address the issue of non-adherence.In-depth interviews were conducted with individuals who were given ACTs at the selected health facilities/chemical shops. Addresses of potential participants were obtained to enable the researchers trace them for the interviews at home after day three when they were expected to have completed the course of treatment. A total of forty (40) in-depth interviews were conducted with patients with malaria who were given ACTs. The interviews were tape recorded with the consent of participants. Notes were taken by the moderator to serve as backup in case the recording was not done properly. Each interview lasted for about 40 minutes. The interviews were conducted in Kassem, Nankani and English depending on the participant\u2019s preference.Two university graduates with experience in conducting qualitative interviews were recruited and trained for one week. They were given an overview of the study and trained on consent procedures, interviewing, probing and transcribing of interviews. Role plays were done during the training session where trainees interviewed each other in both English and the local languages and received feedback from the researchers. A pretest was conducted to evaluate their performance and help finalize the interview guide before the actual data collection.The recorded interviews were transcribed verbatim and entered into a computer using Microsoft word. Guided by the objectives of the study and the themes contained in the guide, a coding list was prepared for data analysis. The data analysis was initiated simultaneously with data collection. This was to ensure that new themes were incorporated into the guide and that thematic saturation was monitored. The data was organized using QSR Nvivo 9 software before analysis. A codebook was developed based on the major themes of the study and transformed into tree nodes and free nodes. Free nodes are generally referred to as open standalone codes usually used at the beginning of the data coding process and are non-hierarchical in nature. Tree nodes are however, hierarchical in nature and they can have relationships with other nodes. They are usually used for secondary coding after open coding from the first round of data coding. The sampling approach was to enable us elicit ethnic differences in the use of ACTs by the two ethnic groups, Kasem and Nankani. However, the analysis did not show any pattern of differences and so this theme was not considered in the presentation of the results. The results of the study were then based on the major and sub-themes that emerged from the study.The age of respondents was grouped into four categories. Most of the respondents, sixteen were between 31\u201340 years while only four of them were between 41 years and above. The level of education of the respondents was also grouped into three, those who had never attended school, primary to junior high level and secondary to tertiary level. From the table, twenty-one respondents had between primary to junior high education while only six (6) of them had secondary to tertiary education .Our findings revealed that there was vast variation of knowledge about ACTs among the educated and uneducated participants. As expected, most of the uneducated participants had little or no knowledge on the type of ACT they were given. They could not mention the names of any of the three ACTs currently being used to treat uncomplicated malaria as captured below by study participants.\u201cWell, I do not know the name because when I went they gave me some medicine that has a mosquito on the pack\u201d.\u201cI wouldn\u2019t be able to tell you the name of the medicine they gave me\u201d.However, the educated participants were able to mention the names of the ACT they were given at the health facility or chemical shop. Others could not mention the name but were able to describe the color of the ACT given. White and yellow is commonly used by community members to describe artesunate\u2014amodiaquine. Most of the educated participants knew artesunate-amodiaquine.\u201cThey gave me the artesunate-amodiaquine\u2026\u201d.\u201cThe doctor gave me artemether-lumefantrine at the health facility\u201d\u201cIt is the yellow and white medicine that was given to me\u201d\u201cI prefer the artemether-lumefantrine because it is good for me. I took the artesunate-amodiaquine some time ago and I nearly died. My feet and hands were all stiff, I could not feel anything, I could not open my eyes and I finally ended in the hospital\u201d.\u201cI was given the artemether-lumefantrine; and it is that drug I prefer using anytime I have malaria. For artesunate-amodiaquine no, no, no, I do not like taking it because when I take it, it is like I have added another illness to the one I already have. For artemether-lumefantrine when I took it I was fine because it did not disturb me at all and it is also good\u201d.Diverse views were expressed by educated participants on the ACT they would mostly prefer using to treat their malaria. However, the results showed that artemether-lumefantrine was reported by participants as the most preferred ACT used to treat uncomplicated malaria. Twenty-six respondents reported preference for artemether-lumefantrine for the treatment of malaria. Though, eight of these patients received artesunate-amodiaquine at their last malaria episode but reported that they would have preferred artemether-lumefantrine. The main reason participants gave for their choice of artemether-lumefantrine was that the drug was good and it had minimal or no side effects as compared to the other ACTs especially the artesunate-amodiaquine. Participants expressed their views this way on the most preferred ACT for treating uncomplicated malaria:Almost all respondents who completed their treatment using artesunate-amodiaquine reported feeling well on the day of the interview. However, participants who did not complete the drug regimen for artesunate-amodiaquine attributed it to the side effects such as body weakness, dizziness, vomiting and loss of appetite. They however, acknowledged that the drug was very good in treating malaria.\u201cThere are two things, though, the artesunate-amodiaquine that I said I got serious side effects when I took it, since then I have never had malaria up to date\u2026. So I think when you force and take artesunate-amodiaquine it will take you long before you get malaria again\u2026.\u201d.\u201cThe one I said is white and yellow (refers to artesunate-amodiaquine), I was given that medicine and after taking them I did not want to hear the scent (smell it) again. After taking the full dose, I felt very weak and could hardly hold an object. After a while, I saw that my situation had improved so the issue is that the drug has side effects but when you persevere and take it to the end (take the full dosage) you will be fine\u201d\u201cI don\u2019t know their names and I take any malaria drug that is given to me to treat my illness\u201d.Some of the uneducated participants on the other hand reported that they had no choice on the type of ACT they would prefer using to treat their malaria. They said that since they did not have knowledge on these ACTs or know the names of these ACTs, they had no problem using any of them that was given at the health facility. A 35 year old female patient had this to say:Sixteen respondents reported having difficulty completing their malaria doses. Perceived cure of malaria after initial dose affected adherence. According to the participants, they felt that their illness or condition was better after they had taken the initial dose of the medication and that made them to stop taking the remaining medicine. Respondents said this was the core reason why they could not complete the course of treatment.\u201cWhen you take and you feel better, you stop taking the drugs. I will use myself as an example because when I took it I realized it was better and I had to stop taking the other tablets\u201d.\u201cWhen I took the malaria drug they gave me at the hospital for the first and second days, I felt better and so I had to stop. I don\u2019t usually like taking drugs and because of that when I manage and take them small and realize that my illness is better I stop taking the rest.The side effects patients experience from using ACTs also greatly affected adherence to malaria treatments especially with respect to artesunate-amodiaquine. Nine out of fifteen patients who were given artesunate-amodiaquine reported that they had to stop using the drug half way during the treatment when they started experiencing side effects such as body weakness, dizziness and loss of appetite. Respondents reported that artesunate-amodiaquine unlike the other ACTs was such that when you take it, your condition would become rather more serious than before because of the side effects and that accounted for their inability to complete the treatment.\u201cIt is because some malaria medicines make people weak when you take them; so after taking the first one and getting weak, you get discouraged. Then you feel you should wait till tomorrow and eventually you stop taking it. That was what I did with the artesunate-amodiaquine they gave me at the hospital. I was weak so I had to stop taking it\u201d\u201cThe reason for this is because of the side effects people get from using these drugs. I was given the yellow and white (artesunate-amodiaquine) drug and when I was taking the medicine, I could not cook and because I do not have somebody to do it for me I had to stop taking it. These are some of the reasons why we are sometimes not able to take the drugs.\u201dQ: Did you take the ACT as prescribed to you?R: No, I could not take all. Laughter by respondent and moderatorQ: Why were you not able to take all of it?R: I did some work and became tired and I forgot and went to bed. I took it the following dayThe problem of recall was also reported in this study as a factor affecting adherence to ACTs use. Participants reported that forgetfulness was a factor responsible for non-adherence to the use of ACTs especially the older people because of the number of times one needed to take the drug in a day and the time they were supposed to take it. A female participant had this to say:\u201cThey said I should take it in the morning and evening but I was always taking it only in the evenings. The reason is that I don\u2019t eat heavy food in the morning because they told me to eat heavy food before taking the drugs and once I cannot eat heavy food in the morning was the reason why I was not taking the drug in the morning. Some people because of the \u201cbitterness\u201d (taste) of some of the drugs, they do not want to take it.\u201d.The requirement for patients to eat before taking medications including malaria drugs affected adherence. The study found that the inability to eat heavy food in the morning before taking the medications also affected adherence to ACT use by patients with malaria. The taste of some ACTs (being bitter) greatly influenced their proper use by some study participants. A female respondent shared her views this way on the issue:\u201cIt is ignorance and sometimes influence from peers. Yeah, some it is due to peer influence because we don\u2019t consider our individual body system when it come to the use of medicine but one may just come and tell you that as for this drug when they prescribed it to me, I took it this and that way, even a drug like paracetamol some may take the whole sachet when you are only asked to take two tablets when you have headache. Some come and tell you that this is how I took it so also take the same especially if the illness is critical. So, I think it is the peer influence\u201d\u201cSometimes when there are so many patients, the health workers do not have patience and because of that they do not take their time to explain how people should take their medications. They are also not able to let them know that if they do not take the medicine according to instructions, it will not work. The health workers usually want the crowd to disperse to enable them go home. So when a patient is told to take twice a day, when he takes it in the morning within a short period, the person will take it again\u201dOther factors mentioned by participants that affected adherence were lack of knowledge as a result of health providers\u2019 inability to educate patients on the proper use of the medication. Twenty-nine respondents mentioned this as one of the factors affecting ACT use. They reported that they were not properly educated by health providers on how they could use the ACT given. Lack of education was mentioned by both educated and non-educated respondents in this study. Peer influence was also reported by study participants. Respondents reported that patients with malaria were not educated by health providers on how to properly use these ACTs. They said that wrong information received from friends and other relations led to wrong use of ACTs. In the exact words of some of the respondents:Some speculative factors affecting adherence were also mentioned by participants. Study participants blamed health providers for patients\u2019 inability to use ACTs according to prescription. They were of the view that advices given by some health providers on the use of ACTs were not accurate and that contributed greatly to non-adherence. They said that was the reason why some people would usually ignore certain prescriptions given by health providers and rather use the drugs their own ways. Participants were of the opinion that people would usually want their illness to heal very fast and as a result they would take over dose of the medication in order to help them in that regard. These factors according to participants were also responsible for malaria patients\u2019 inability to adhere to the use of ACTs.\u201cOthers feel that the advices given at the hospital are not good and that is why they usually ignore what the doctor told them to take the drugs and instead use it the way they want. Though I have not done that but I know of somebody who said that she had to use her own knowledge to take the drug because the way the health worker asked her to take was not correct\u201d\u201cFor some people they have the motive that when they take the drugs more (overdose) it will help to cure the illness faster. So when the doctor says they should take two in the morning and two in the evening they will rather take three, three with the motive that it will cure the illness faster\u2026.\u201d.Despite the fact that there are various factors affecting adherence to the use of ACTs, our study participants proposed various strategies to help address the problem. Thirty-five of the respondents said that there was the need for vigorous continuous education on the use of ACTs at the Out Patient Department (OPD) level, the pharmacy department and at the chemical shops by health providers and chemical shop owners. They were of the view that the education should include the need for people to strictly follow dosage prescriptions and use malaria medications and the likely side effects people could get as a result of wrong use of these ACTs at home.\u201cThe pharmacists and the doctors should do the education; because most of the people here are illiterates they should have time to explain to them how they can take the drugs and why they have to follow prescriptions and take the drugs. We the patients too when they give us the drugs, we should try and take the drugs according to how we have been asked to take\u2026\u201d.\u201cAt the OPD when patients are seated and doctors have not started consulting, a nurse should give a talk to all patients on how they should take their medicines. Sometimes when they write three times on the envelop, someone can take it four times. The talk will make patients understand why they should take medicines according to the way they have been instructed\u201d.Thirteen respondents suggested a single dose treatment for malaria in order to enhance adherence to ACTs use. This suggestion was made because some people do not usually like taking medications because of the taste, the number of times and tablets they have to take in a day and if the single dose treatment had been made, it could help solve these problems. Besides, some people stop using ACTs half way through the treatment course because of the side effects they experience. Therefore, participants were of the view that if the single dose treatment was made for the patient to take it once, the side effects could be tolerated since the medication is not repeated.\u201cWell, if they can reduce the dosage and it can still treat and kill(cure) the malaria or if they can make it one tablet such that you can just take it once and you don\u2019t take again, I think that will be better. If you take it once and that is all, people can manage and take it and that will be better than where you will take it and get these side effects and yet you have to take the remaining tablets, that is why other people do not normally want to continue and finish.\u2026 The other thing is if they can make the injection of it that will also be good because other people do not like taking the tablets because of the taste and the quantity. For me, I prefer taking injection as compared to the tablets\u201dRespondents also suggested that follow-up visits by health workers to the communities to ascertain how people who were given ACTs were using them at home could also improve adherence to malaria treatments.Currently, there are three recommended ACTs for the treatment of uncomplicated malaria in Ghana. The malaria treatment policy requires that artesunate-amodiaquine, artemether-lumefantrine and dihydroartemisinin-piperaquine be used to treat uncomplicated malaria . This stAdherence to treatment regimens is the extent to which patients use medication as prescribed for them by health care providers . It is tThis study found that side effects, forgetfulness, inadequate health information by prescribers affected adherence to the use of medications. Various side effects experienced by patients with malaria from the use of ACTs contributed greatly to non-adherence. The inability of some patients with malaria to contain these side effects has made them to stop using these ACTs half way through the treatment regimen , 12, 14.It is very important for health providers to educate patients on correct use of drugs including ACTs. Based on the views presented by participants in this study, health providers are usually not able to give frequent and enough education to their clients on the need for them to use all medications given even if the condition is better after the patient has taken the initial dosage and the possible side effect they might have experienced. The possible reason for health providers\u2019 inability to provide enough health education to their clients may be as a result of the huge number of clients they usually have to attend to at the Out-Patient Department (OPD) level. Because of the pressure, health providers will usually write on the park on how the patient would use the drug making it difficult for some patients to correctly interpret and use the drug correctly at home especially the uneducated clients.It is reported that health providers\u2019 inability to provide health education to patients on the correct use of ACTs and the possible side effects associated with the use of these ACTs negatively influenced adherence , 14. It Our study participants suggested intensive continuous health education by health providers and chemical shop owners on the side effects people are likely to experience when they use these ACTs wrongly. This will help address wrong use of medications at home by patients. Evidence exist that patients who received health information were more likely to adhere to malaria treatment than those who have not received health information . PatientThough this study provides information required to increase adherence to the use of malaria treatments, some limitations must be taken into consideration. The interviews were mostly conducted in the main local languages and translated into English. Therefore some words could have lost their original meaning. To help mitigate this, two independent people were made to do the translations and transcription and the transcription were verified. However, given the limitation of such a method, in doing the analysis emphasis was placed on the overarching themes and not specific word choices or phrases.Inadequate health information and attitude of some individuals towards the use of medications especially ACTs, side effects and the issue of recall especially among the older people are the main factors influencing adherence to ACTs use in the study area. Despite the fact that some education has been given to patients with malaria at the health facility level on how to use these malaria medications, much efforts still need to be made by health providers in this regard. It is recommended that intensive regular education be carried out at the health facility level on why it is very important for patients to use ACTs correctly. Patients should also be encouraged to use all medications given to them even when there is improvement after the initial dose of the treatment has been taken. There is also the need for health providers to educate their clients on the side effects they are likely to experience from using these ACTs and the need for them to continue and complete the prescribed treatment This will improve on the adherence and individual behavioral factors affecting the use of ACTs and therefore contribute effectively to the reduction in malaria cases."}
+{"text": "Fixed-dose combinations of artemisinin combination therapy are strongly recommended to facilitate drug administration and compliance. New fixed-dose combinations must nevertheless be evaluated in relevant populations in terms of efficacy and pharmacokinetics.Plasmodium falciparum malaria to investigate the efficacy and the pharmacokinetics of mefloquine when combined with artesunate in a fixed-dose combination (400/200 mg of mefloquine base/artesunate). The pharmacokinetic analysis was performed using a population approach.A single-arm, open-label, clinical trial was performed in Indian patients with acute uncomplicated -1 (75%) for the absorption rate constant, 1.13 L/h (30%) for the apparent plasma clearance, 271 L (21%) for the apparent central distribution volume, 344 L (54%) for the apparent peripheral distribution volume, and 1.43 L/h for the apparent distribution clearance. These values were consistent with the pharmacokinetic results described in Thai patients. No significant covariate was found for clearance. Body weight explained the inter-individual variability of the apparent central and peripheral distribution volumes. The PCR-adjusted efficacy of the treatment was 100%.Seventy-seven patients were included in the study. Mefloquine pharmacokinetics obeys a two-compartment model with first-order absorption and elimination. Mean parameter estimates  were as follows: 0.16 hP. falciparum malaria.The lack of significant covariate explaining the inter-individual variability of mefloquine clearance, combined with the excellent efficacy, supports the use of the standard 200/400 mg of artesunate-mefloquine fixed-dose combination in Indian patients with uncomplicated ISRCTN70618692Clinical Trial Registration:  Plasmodium falciparum malaria 1/2) and the calculation of C7 and C28 was made by NONMEM.Bayesian estimates of the individual pharmacokinetic parameters (obtained in NONMEM output) were used to calculate individual MQ elimination half-life, the individual area under the curve from time zero to infinite (AUC), and MQ concentration at D7 (C7) and D28 (C28). Elimination half-life was calculated as follows: ln2/\u03b2 with \u03b2\u2009=\u20090.5 \u00d7 (Q/Vc\u2009+\u2009Q/Vp\u2009+\u2009CL/Vc-[(Q/Vc\u2009+\u2009Q/Vp\u2009+\u2009CL/Vc)The primary efficacy outcome of the study was the PCR-uncorrected cure rate  on D63 and PCR-corrected cure rate on D63 (proportion of patients without recrudescence/ inconclusive/ no results as classified by PCR genotyping).Comparison of MQ C7 and C28 between treatment success and treatment failure was expected to be performed by non-parametric statistical analyses.Seventy-seven patients  were included. Their demographic and biological characteristics are given in Table\u00a0The optimal model was a two-compartment model with first-order absorption and elimination. The estimated parameters were the absorption rate constant (ka), the apparent central distribution volume (Vc/F), the apparent peripheral distribution volume (Vp/F), the apparent elimination clearance (CL/F), and the apparent distribution clearance (Q/F), where F is the bioavailability. Inter-individual variability of Q/F could not be estimated. Residual variability was described by a proportional error model.BW was the only significant covariate and was found to explain the inter-individual variability of Vp/F and Vc/F. All other covariates, including the day of dosing was not found to significantly influence the inter-individual variability of the PK parameters. The parameter estimates are given in Table\u00a0Epsilon shrinkage was 19.8%, eta shrinkage was 21% for CL/F, 34% for Vc/F, 53% for Vp/F, and 20% for ka.The lack of bias of the final model is observed in Figures\u00a0Median (range) mefloquine AUC was 1055 (596 \u2013 1741 mg.h/l). Median (range) C7 was 1.58 (1.08 \u2013 2.90) mg/l, and median (range) C28 was 0.29 (0.11 \u2013 0.57) mg/l. Since no recrudescence and only one new infection was observed over the follow-up of the study (PCR-corrected cure rate of 100%), no correlation between MQ C7 or C28 and treatment efficacy could be investigated.et al.[The present model accurately described the data and was in agreement with previous results seen in adult patients with uncomplicated malaria Table\u00a0, even thet al.. A slighDifferences in study design probably also explain why no modification in MQ pharmacokinetics during the treatment administration period could be detected. This change was previously described and attributed to the rapid clinical improvement seen due to treatment efficacy . There wet al. [et al.[In agreement with previous findings , BW did et al. , who repet al. . However. [et al.. The mecP. falciparum malaria. The lack of relevant biological covariates on MQ CL/F, combined with the excellent efficacy results that were obtained, support the use of the 200/400 mg dose of AS/MQ in this population.The present study provided population pharmacokinetics parameters for MQ, administered as a FDC of AS/MQ, in Indian adult patients with acute uncomplicated ACT: Artemisinin combination therapy; AS: Artesunate; ASAT: Aspartate aminotransferase; ALAT: Alanine aminotransferase; BW: Body weight; DHA: Dihydroartemisinin; MQ: Mefloquine; NPDE: Normalized predictions errors; PRED: Population predictions.The authors have declared that they have no competing interests.VJ pharmacokinetic analysis, writing of the manuscript; BSh study coordination and monitoring; NV study management and execution; BSr technical overview; J-RK study pharmacokinetics protocol, contribution to the manuscript and discussion of the results. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum infections. The recrudescence rates were low, gametocyte carriers lessened, and the curative rate increased remarkably. The combination therapy effectively deferred the emergence of drug resistance in the parasite. The regulation \u201cThe guidelines and regimens for the use of antimalarial drugs in China\u201d was issued to guide rational application and standardize malaria treatment in the country. As the recommended first-line drug to treat falciparum malaria in the world, ACT was adopted in the regulation. In response to the global initiative of malaria eradication proposed by the UN Millennium Development Goals (MDGs), the Chinese government has set a target to eliminate malaria by 2020.This historical review covers antimalarials developed in China, which include artemisinin, artemether, artesunate, and dihydroartemisinin, as well as other synthetic drugs such as piperaquine, pyronaridine, benflumetol (lumefantrine), and naphthoquine. The curative effects of these antimalarials in the treatment of falciparum malaria, including chloroquine-resistant strain, are especially discussed. Following the World Health Organization (WHO) recommended artemisinin-based combination therapy (ACT), different combinations of artemisinin, or its derivative, along with another antimalarial drug were orally used to treat  Please see Additional file In China, malaria was not only widespread but also one of the most threatening diseases to people\u2019s health. Historically, traditional Chinese medicinal herbs were used for thousands of years as a folk remedy to treat malaria. Before the 1950s, the antimalarial quinine was imported into the country in great quantities [Cinchona officinalis L., Rubiaceae, grew wild in South America and were cultivated in Java, Indonesia. During the Second World War, there was no supply of quinine because Java was occupied by Japanese troops. Because of these circumstances, Chinese scientists tried to isolate an active principle against malaria from the root of the plant Dichroa febrifuga L., Saxifragaceae, another traditional herbal medicine, in the early 1940s. The principle was named \u2018\u03b2-dichroine\u2019 or \u2018febrifugine\u2019, and its chemical structure was identified. Two other principles with antimalarial activity were also isolated from the kernel of the plant Brucea javanica (L.) M., Simarubaceae, and were named \u2018bruceine D\u2019 and \u2018bruceine E\u2019, respectively. Their chemical structures were also clarified. However, the investigations did not make further progress into their clinical use [Quinine was first isolated from the cinchona bark by European scientists in the 1920s. The trees of In the early 1940s, chloroquine, an erythrocytic schizonticide of plasmodia, was developed in Western countries. Primaquine and pyrimethamine were synthesized in 1946 and 1951, respectively, and used for radical cure and causal prophylaxis in malaria control.In 1958, the abovementioned chloroquine, primaquine, and pyrimethamine were also manufactured in China for malaria control. The productive techniques of the drugs were improved in order to reduce the costs to meet the needs of large-scale malaria control [Plasmodium falciparum was found and spread in the early 1960s in Southeast Asia [P. berghei and the chloroquine-resistant strain of P. berghei in mice, and P. inui in rhesus monkeys (Macaca mulatta). Later, two more experimental models, P. cynomolgi and P. knowlesi in rhesus monkeys, were established. The new antimalarials were studied and synthesized at a few institutes in the country. Some of the new synthesized compounds were found to have an effect against plasmodia in the blood [Chloroquine-resistant ast Asia . To imprDuring the Vietnamese War in the 1960s, there was an increase of incidence and mortality of chloroquine-resistant falciparum malaria on the border of the Yunnan Province. This situation inspired researchers to actively develop alternative antimalarials that were effective to treat chloroquine-resistant malaria.P. falciparum with chloroquine resistance, including artemisinin isolated from a medicinal herb, which, together with its derivatives, aroused a great interest globally at a later stage.To strengthen the research on antimalarials, an extensive cooperation was organized by the Chinese government in 1967 for drug development with participation of relevant institutions and pharmaceutical companies. Systematic researches from chemical syntheses to pharmacotoxicology to clinical trials were carried out. Among the synthetic compounds and principles extracted, a few were clinically proven to be highly effective to treat In line with the requirement of malaria control, antimalarial drugs should be rationally used for achieving a curative effect, interruption of transmission, and avoidance or deferment of the development of drug resistance. Therefore, the regulation \u201cThe guidelines and regimens for the use of antimalarial drugs in China\u201d was issued and revised in 2009 . ArtemisP. berghei in mice. It was a totally new agent: artemisinin. Its derivatives were then semi-synthesized and proven to be active antimalarials  was recommended for the treatment of falciparum malaria .Artesunate was developed by semi-synthesis from artemisinin  see Fig. It was One hundred and fifty-nine patients with falciparum malaria received a total dosage of 400\u00a0mg of artesunate administered by intravenous or intramuscular injection in the course of three days. The mean fever subsidence and parasite clearance times were 26.1\u00a0h and 28.5\u00a0h, respectively. The recrudescence rate was as high as 52.4% . The totOne hundred and six patients with cerebral malaria received a regimen of 420\u00a0mg of artesunate by injection over six days. Ninety-eight patients were cured and eight died .Consequently, a total dosage increased to 480\u00a0mg by injection over seven days  was recommended in the treatment of falciparum malaria .The semi-synthetic dihydroartemisinin was a middle compound in synthesis . The regimen was given to 64 patients with falciparum malaria. The mean times of fever subsidence and parasite clearance were two and three days, respectively. The recrudescence rate was about 4% followed-up for 30\u00a0days or 35\u00a0days. Gametocytes were still present 14\u00a0days after treatment ,29. The Clinically, the side effects of artemisinins were similar. In general, they were well tolerated. Some patients complained of dizziness, headache, nausea, vomiting, abdominal pain, diarrhea, and palpitation, etc. A few patients had rashes which disappeared after treatment with suitable medicine. Some patients had reticulopenia, leucopenia, and increased SGPT, and urea nitrogen, sinus bradycardia, arrhythmia or premature ventricular beat. Usually the side effects disappeared in one to three days or seven days after treatment.Chemical syntheses have been another important way for developing new antimalarials. Among numerous chemicals, four compounds . The mean fever subsidence time was about two days and the mean parasite clearance time was about three days. The recrudescence rate was 14.9% or 18% ,35, follThe side effects noted were headaches, dizziness, listlessness, somnolence, vomiting, abdominal pain, diarrhea, facial tingling, and reduction of reticulocytes. They were mild and recoverable after treatment. A few cases showed asthma, stuffiness in the chest or dyspnea, palpitations, or I\u00b0 A-V block, which disappeared soon after a proper rest. A few women complained of irregular menstruation which normalized after two months. Piperaquine was contraindicated to the patients with severe acute hepatism, nephropathy, and heart disease, and pregnant women were advised to use with caution.P. falciparum to the drug was 72.9\u201396.45% [In the late 1980s in China, piperaquine was used as a prophylactic and therapeutic agent in malaria control for 10\u00a0years. The resistance rate of 9\u201396.45% , with a 9\u201396.45% .Piperaquine is presently used only for suppressive prophylaxis of vivax malaria in specific populations, i.e. mobile populations such as migrant workers, personnel making crossings at the border, travellers, etc. This drug can accumulate in the liver, and should not be given for longer than four consecutive months .Pyronaridine . No teratogenesis was noted [Pyronaridine could dose-dependently increase the rate of fetal resorption. It possessed embryotoxicity, which was still lower than that of the known positive control drug, dexon . The total dosage by intramuscular injection or intravenous drip was 100\u2013300\u00a0mg divided into two doses given at 4\u20138\u00a0h intervals. The regimens of pyronaridine in the treatment of malaria were efficient and safe. Generally, the mean fever subsidence time of the patients infected by  malaria . The recOver 100 cases of cerebral malaria and other severely complicated malaria were successfully cured with emergency treatment with pyronaridine.More than 10 pregnant patients during their mid or late trimester were also cured with pyronaridine without any adverse effects.In general, pyronaridine was well tolerated by patients. Some patients complained of discomfort in the stomach or epigastrium, dizziness, nausea, headache, abdominal pain, and slight diarrhea. A few patients experienced vomiting, palpitation, and allergic skin rashes which recovered in two days after suitable medicine was taken.P. ovale and P. malariae infections. The patients were all cured with no recrudescence. The side effects were minor and transient [Pyronaridine was used orally to treat falciparum or multidrug-resistant falciparum malaria in Africa, specifically in children, and ransient -50.Benflumetol or lumefantrine  was used to treat falciparum malaria. The mean times of fever subsidence and parasite clearance were three days and two days, respectively, with a recrudescence rate of 11.5%, and most patients carried gametocytes for a long time after medication . A totalP. falciparum to the drug or by a lower dosage of the drug. The side effects of the drug included diarrhea, abdominal pain, vomiting, dizziness, etc., which could disappear after a nap.In Yunnan, 61 cases with falciparum malaria were treated by an oral total dosage of 2.0\u00a0g divided into five doses . The average times of fever subsidence and parasite clearance were within 60\u00a0h. Two days after treatment, four out of 61 cases showed a high fever (39\u00b0\u201340.4\u00b0C) and with the rate of asexual forms being 48\u2013261%. The four cases failed treatment. The recrudescence rate was 9.8% (five out of 51 cases), followed-up for 28\u00a0days. The occurrence rate of gametocytes was 3.9% 28\u00a0days after treatment . The faiP. berghei[Naphthoquine  and piperaquine 375\u00a0mg), in a fixed-ratio of 1:6, was mixed to form one compound tablet of artemisinin and piperaquine, or co-artemisinin. A total oral dosage of the compound tablet was four tablets divided into two doses (one dose daily for two consecutive days) to treat falciparum malaria [75\u00a0mg, inDihydroartemisinin (40\u00a0mg) and piperaquine phosphate (320\u00a0mg), in a fixed-ratio of 1:8, was formulated to obtain one compound tablet of dihydroartemisinin and piperaquine. An oral total dosage was four tablets . Sixty-two cases of falciparum malaria were treated in Yunnan, with average times for fever subsidence and parasite clearance of 34.99\u2009\u00b1\u200916.51\u00a0h and 33.14\u2009\u00b1\u200911.91\u00a0h, respectively. The negative rate of gametocytes was 95.5%. No recrudescence was found 28\u00a0days after follow-up . SimilarOne compound tablet of artemether and benflumetol (lumefantrine), or co-artemether, was formed by combining artemether (20\u00a0mg) with benflumetol (120\u00a0mg), in a fixed-ratio of 1:6. Synergism was shown between the two drugs. An oral dosage was 16 tablets divided into four doses . One hundred and sixty-seven cases with falciparum malaria were treated in 2005. The mean fever subsidence and parasite clearance times were 20.4\u2009\u00b1\u20098.4\u00a0h and 37.9\u2009\u00b1\u20097.9\u00a0h, respectively. The recrudescence rate was 5.9%, followed-up for 28\u00a0days . In prevArtemisinin (125\u00a0mg) and naphthoquine (50\u00a0mg), in a fixed-ratio of 2.5:1, was mixed to form one compound tablet of artemisinin and naphthoquine, or co-naphthoquine.A total oral dosage of eight tablets was given as a single dose to 100 patients with falciparum malaria in Hainan. The average fever subsidence and parasite clearance times were 17.5\u2009\u00b1\u200912.3\u00a0h and 30.0\u2009\u00b1\u20098.8\u00a0h, respectively. The recrudescence rate was 3%. The adverse reactions included headache, nausea, weakness, and loss of appetite. These were recoverable two days after withdrawal . NaphthoP. vivax appeared between day 21 and 28 after treatment. No recrudescence was found in the group taking pyronaridine alone and the combination group, followed-up 28\u00a0days. The gametocyte carriers occupied 16.7% in the dihydroartemisinin group, 60.9% in pyronaridine group, and 20.0% in the combination group, significantly lower than the group of pyronaridine alone (P\u2009<\u20090.01). No abnormality was found in blood, urine, and ECG examinations before or after treatment. A few patients complained of headaches and dizziness, but recovered in one to two days [A combination therapy of dihydroartemisinin (300\u00a0mg) plus pyronaridine (800\u00a0mg) divided into four doses  was orally administered. The two separate drugs were simultaneously taken. Thirty-two patients with multidrug-resistant falciparum malaria were treated. A double blind clinical trial was performed with routine dosage of dihydroartemisinin or pyronaridine alone as control groups in Hainan in 2001. The mean fever subsidence time of the combination therapy group was 35.7\u2009\u00b1\u200924.7\u00a0h, and asexual forms clearance time was 23.8\u2009\u00b1\u200910.1\u00a0h. In the group taking dihydroartemisinin alone, the recrudescence of parasites was found in one case with a temperature of 39.6\u00b0C on day 21, who failed the treatment, and in four cases two days . It was two days .The above regimen of dihydroartemisinin plus pyronaridine was applied in the state of Eritrea, Africa in 2000. Thirteen cases with chloroquine-resistant falciparum malaria received the regimen, and similar results were obtained. Gametocyte carriers were found in 18% of the cases on day seven after treatment .A total oral dosage of artemether or artesunate (300\u00a0mg) or dihydroartemisinin (200\u00a0mg) plus pyronaridine (800\u00a0mg), in a course of two days and given as two separate drugs at the same time, was taken for the treatment of falciparum malaria. No recrudescent rate, and the gametocytes disappeared on day 20 after treatment. Less adverse effects were recorded ,77.The Shin Poong Pharmaceutical Co. and WHO/TDR/PRD, as partners, have developed the pyronaridine-artesunate project for the treatment of uncomplicated malaria .P. falciparum infections. As amodiaquine could lead to agranulocytosis and liver damage [An oral total dosage of artesunate (600\u00a0mg) plus amodiaquine (1.8\u00a0g) was divided into three doses, one each for three consecutive days. The two separate drugs were taken simultaneously for the treatment of r damage , it was r damage .in vitro test revealed that the resistance rate of P. falciparum to amodiaquine was 85.3\u2013100% [Amodiaquine possessed cross-resistance to chloroquine. In 1996, an 5.3\u2013100% , which wP. falciparum in the country, and based on the experience from other countries, ACT and \u201cThe guidelines and regimens for the use of antimalarial drugs in China\u201d  are theFor the treatment of vivax malaria, a predominant species and still sensitive to chloroquine, an oral dosage of 1.2\u00a0g of chloroquine in a three-day course, plus a dosage of 180\u00a0mg of primaquine in an eight-day course is recommended as routine therapy of radical cure. The dosage of 180\u00a0mg of primaquine in an eight-day course is designed as an anti-relapse treatment of vivax malaria.As for prophylaxis in specific populations, i.e. mobile populations such as the ones mentioned previously, a single dose of 600\u00a0mg of piperaquine orally administered once monthly is recommended. A single dose of 300\u00a0mg of chloroquine, once every 7\u201310\u00a0days, is also recommended for prophylaxis.These drugs are evidently effective and affordable for vivax malaria control. There are large numbers of vivax malaria cases in the country and therefore, the drugs for treatment of vivax malaria are considered as the first-line antimalarials in China.Because of the persistent efforts in the past decades to control malaria, the number of indigenous falciparum malaria cases has considerably decreased and accounts for about 10% of the total malaria cases, mostly evident in the border area of Yunnan and in the forest hill area of Hainan. At the end of 2012, the percentage decreased to 6% , showingFor the emergency treatment of cerebral or other complicated, severe malaria cases, artemether, artesunate, or pyronaridine can be the drugs of choice, administered by intramuscular or intravenous injections, or intravenous drip. These drugs are considered as the third-line antimalarials.P. falciparum. Chinese scientists have made great contributions to the world by developing new drugs, including artemisinins and a few others. These progresses have made it possible for artemisinin-based combination therapy (ACT) to not only effectively cure numerous cases but also defer the emergence of resistance of P. falciparum to the antimalarials.Drug development is one of the most important components in malaria control due to the critical situation of drug resistance of P. falciparum.During treatment, the results were sometimes not the same for the same drug or the same drug combination therapy. Besides the different fields, the different results can be explained by the different dosages, courses of treatment, and the numbers of cases which followed-up. Generally, artemisinins are rapidly active against plasmodia, have a long course, and have a high recrudescence rate. After treatment with artemisinins, the gametocyte carriers are less significant than those with piperaquine, benflumetol, and pyronaridine. The actions against malaria parasites are slow when benflumetol and naphthoquine are used alone, and a large dosage of benflumetol is needed. It has been suggested that naphthoquine should be used in combination with a rapid active antimalarial . By usinP. falciparum.With the varying results, the suitable treatment regimens for the drugs and drug combination therapies should be provided with regulations. Therefore, the regulation \u201cThe guidelines and regimens for the use of antimalarial drugs in China\u201d was issued by the health authority. As a recommended first-line drug to treat falciparum malaria in the world, ACT has been adopted in the regulation. It provides the norms for drug selection and standardized treatment of malaria cases. It is also a valid measure to avoid or defer the development of drug resistance of As a response to the initiative of global eradication of malaria proposed by the UN Millennium Development Goals, and on the basis of the great successes achieved in malaria control in the past decades, since 2010, the Chinese government has launched a program to eliminate malaria in most regions by 2015, and in the whole country by 2020 .The author declares that he has no competing interests.Multilingual abstracts in the six official working languages of the United Nations.Click here for file"}
+{"text": "Plasmodium vivax malaria especially from Southeast Asian regions. The present study was conducted to assess the therapeutic efficacy of chloroquine\u2013primaquine (CQ\u2013PQ) combined regimen in a cohort of uncomplicated P. vivax mono-infection.Of late there have been accounts of therapeutic failure and chloroquine resistance in P. vivax malaria as per the World Health Organization\u2019s protocol of in vivo assessment of anti-malarial therapeutic efficacy. Participants were treated with CQ 25\u00a0mg/kg body weight divided over 3\u00a0days and PQ 0.25\u00a0mg/kg body weight daily for 2\u00a0weeks.A tertiary care hospital-based prospective study was conducted among adult cohort with mono-infection P. vivax and Plasmodium falciparum infection by nested polymerase chain reaction test. The majority of subjects  became aparasitaemic on day 2 followed by 35.2% (44/125) on day 3, and 8% (10/125) on day 7, and remained so thereafter. Overall only one therapeutic failure  occurred on day 3 due to persistence of fever and parasitaemia.Of a total of 125 participants recruited, 122 (97.6%) completed day 28 follow up, three (2.4%) participants were lost to follow-up. Eight patients (6.4%) were ascertained to have mixed P. vivax malaria and should be retained as treatment of choice in the study region. One case of treatment failure indicates possible resistance which warrants constant vigilance and periodic surveillance.CQ\u2013PQ combined regimen remains outstandingly effective for uncomplicated  Plasmodium vivax accounts for more than 50% of total malaria burden in India [P. vivax malaria in India [P. vivax cases with normal glucose-6-phosphate dehydrogenase (G6PD) activity [P. vivax have been observed in 23 countries across the globe, including India [P. vivax has been confirmed in only ten countries across the globe, excluding India [in India . Chloroqin India . Additioactivity . TherapeP. vivax remains valid and provides useful indices where regional health policy mandates such [P. vivax mono-infection cohort as per WHO\u2019s protocol [Therapeutic assessment remains the mainstay of monitoring the efficacies of anti-malarial regimens and is recommended to be undertaken once every 2\u00a0years . Surveiltes such . This stprotocol .P. vivax patients aged \u226518\u00a0years attending Kasturba Hospital (KH), Manipal from September 2012 until October 2014. Patients were excluded if they had other febrile illnesses including mixed malaria or P. vivax malaria treated with artesunate combination therapy or prior anti-malarial medication outside KH before presentation or pregnancy or unwillingness to provide a written informed consent.A prospective cohort study was conducted among microscopically proven symptomatic mono-infection Ethical approval (IEC 193/2011) was obtained from the institutional ethics committee of Kasturba Medical College and Kasturba Hospital, Manipal University, Manipal. Participation in study was strictly voluntary and each participant had privilege to retract their participation any time throughout the study. Patients\u2019 information and study data were anonymized.P. vivax mono-infection cases were required [Presuming the proportion of anti-malarial treatment failure (ATF) of CQ\u2013PQ combined regimen to be 5%, for a desired precision of 5% and confidence interval of 95%, a cohort of 73 microscopically proven required . FurtherP. vivax mono-infection and to estimate the parasite index (PI), respectively. DNA was extracted from 200\u00a0\u00b5L of EDTA anticoagulated blood using QIAamp DNA Blood Mini Kit as per the manufacturer\u2019s instructions and stored at \u221220\u00b0C until conduct of nested polymerase chain reaction (nPCR) test for confirmation of the mono-infection with P. vivax malaria.The date of initiation of anti-malarial medication and study enrolment were the same and was considered as day 0. Peripheral blood smear was examined on days 0, 2, 3, 7, 14, 21, and 28. Before initiation of anti-malarial medications, 1\u00a0ml venous blood sample was obtained from each participant in an ethylenediaminetetraacetic acid (EDTA) anticoagulated vacutainer and used for making peripheral smears, total leukocyte count and DNA extraction. On subsequent follow-up days capillary blood sample was obtained by finger prick on cleaned glass slides to prepare both thin and thick smears. Leishman\u2019s stained thin and thick peripheral blood smears were examined to ascertain the \u00ae LH 750 Haematology Analyzer and used for PI calculation. PI was determined as the absolute number of asexual and/or sexual parasites present in 1\u00a0\u00b5L of peripheral blood, as described elsewhere [Axillary temperature was recorded on each respective follow-up day as mentioned above. Participants\u2019 total leukocyte count per \u00b5L was estimated in constituent laboratory of KH, Manipal by Beckman CoulterP. vivax [ATF was considered as the primary outcome. Anti-malarial therapeutic responses were classified as per WHO\u2019s recommendation for P. vivax . HoweverP. vivax and P. falciparum, as described by Snounou et al. [2, 50\u00a0mM KCl, 10\u00a0mM Tris\u2013HCl pH 8.3, 125\u00a0\u00b5M of each dNTPs, 250\u00a0nM of each oligonucleotide primers, 0.5 units of Taq DNA polymerase (Sigma-Aldrich) and 1\u00a0\u00b5L of template DNA. Product of the first amplification reaction was further amplified in second reaction as step 1: 94\u00b0C for 5\u00a0min; step 2: 94\u00b0C for 30\u00a0s; step 3: 55\u00b0C for 30\u00a0s; step 4: 72\u00b0C for 1\u00a0min; steps 2\u20134 for 10 cycles, then step 6: 94\u00b0C for 10\u00a0s; step 7: 60\u00b0C for 10\u00a0s; step 8: 72\u00b0C for 1\u00a0min; steps 6\u20138 for 20 cycles and then final extension at 72\u00b0C for 5\u00a0min followed by termination temperature at 4\u00b0C. Reference samples of mono-infection P. vivax and P. falciparum were procured from the National Institute of Malaria Research (ICMR), Sector 8, Dwarka, Delhi-110077 (India). DNA extracted from the reference samples were used as positive controls in every batch of nPCR tests. While, DNA extracted from a normal healthy volunteer was used as negative control.Small sub-unit ribosomal RNA was amplified using genus and species-specific oligonucleotide primers separately for only u et al. , with moP. vivax malaria, specific anti-malarial medications were prescribed as per the clinicians\u2019 judgements and the national guideline for treatment of malaria [On presentation, patients were treated symptomatically as required. After confirmation of  malaria . Resochi2 test or Fischer\u2019s exact test. All tests of significance were two-tailed with a p value <0.05 indicating statistical significance. Parasite index was summarized as geometric mean with 95% confidence interval of geometric mean. The probability of therapeutic failure was assessed by life table analysis. Data analysis was done using Statistical Package for the Social Sciences version 15.0 .Gaussian distribution of continuous variables was determined by Kolmogorov\u2013Smirnov test. Data having normal distribution were summarized as mean with standard deviation and compared by independent t test. Skewed data were summarized as median with interquartile range and compared by Mann\u2013Whitney U test. Categorical variables were summarized as frequency with proportion and compared by either \u03c7A total of 125 participants sustaining the study selection criteria were enrolled. Pertinent clinical and demographic characteristics of the study cohort have been summarized in Table\u00a0P. vivax was given to all subjects except one participant whose G6PD was low (5.5 U/gm Hb), thus was given a 45\u00a0mg stat PQ dose followed by weekly dose of 45\u00a0mg PQ for next 7\u00a0weeks. Mean G6PD activity in study cohort was determined as 15.2\u00a0\u00b1\u00a04.1 U/gm Hb and ranged from 5.5 to 27.2 U/gm Hb. A complete adherence for the prescribed CQ\u2013PQ regimens was noted among study cohort. Empiric antibiotic (ceftriaxone) was administered to one (1.2%) inpatient.Chloroquine (25\u00a0mg/kg body weight over 3\u00a0days) and primaquine (0.25\u00a0mg/kg body weight daily for 2\u00a0weeks) standard anti-malarial regimen for A total of 122 (97.6%) participants completed day 28 follow-up. Three participants 2.4%) were lost to follow up after day 3. One of them remained parasitaemic on day 3 but had become afebrile within 24\u00a0h of hospitalization. Another two participants had become aparasitaemic by day 3 and their defervescence time was less than 24\u00a0h. More than half of the subjects  became aparasitaemic on day 2 followed by 35.2% (44/125) on day 3; and 8% (10/125) on day 7 and remained so thereafter to day 28  infections among total of eight (6.4%) subjects. The DNA sample of one participant (0.8%) could not be amplified. Notably, the one subject with therapeutic failure had PCR proven P. vivax mono-infection.Nested PCR test confirmed a total of 116 (93.6%) participants to have P. vivax malaria. The KH, Manipal caters for over 200 malaria patients annually from southwestern India comprising Goa, coastal Karnataka and Kerala. Notably, P. vivax infection constitutes more than 55% of total annual malaria incidence in KH, Manipal. Male preponderance observed in the current study is a common finding and has also been reported from the same hospital [P. vivax mono-infection by PCR test and also did not have rigorous 28\u00a0days follow-up, as in the current study. Furthermore, similar therapeutic efficacy until day 28 has also been reported from other parts of India viz. 100% (41/41) from Mangalore [P. vivax recurrences [P. vivax despite having adequate plasma concentration of CQ [Prime index of suspicion of emergence/existence of resistance to anti-malarial drugs surfaces during therapeutic drug trials or surveillances. Present study was undertaken to determine the therapeutic efficacy of CQ\u2013PQ combined regimen in hospital \u20139. Signihospital . Howeverangalore , 98.1% (angalore  and 96.9angalore  for CQ oangalore  for CQ\u2013Purrences \u201316. Noneon of CQ .P. vivax is in Indonesia [P. vivax in India has been documented from north, northeast and western parts, whereas CQ remains sensitive throughout southern India [P. vivax across eastern neighbouring countries of India, including Myanmar [P. vivax strain from those countries along with travellers and/or migrants.The hub of global CQ resistance in ndonesia , 18, 19.rn India . Importa Myanmar , Thailan Myanmar  and Viet Myanmar  is a matP. vivax cases. Statistically, valid sample size, robust analysis and standard methodology render the study outcomes legitimately comparable with parallel series across the globe. Outstanding therapeutic efficacy of CQ\u2013PQ combined regimen for P. vivax malaria observed in the current study is augmenting evidence to support the continuation of the existing national anti-malarial drug policy for P. vivax malaria in India. The probably resistant case of one therapeutic failure could not be ascertained as truly resistant P. vivax strain. Arguably, persistence of fever and increasing parasitaemia from 2,200 parasite/\u00b5L on day 0 to 9,652 parasite/\u00b5L on day 2 and residual 7,012 parasite/\u00b5L on day 3 despite completion of CQ dosage features a true resistant P. vivax strain. However, this argument could possibly have been true only if CQ and desethylchloroquine concentration in participant\u2019s blood [The present study poses considerable strength over studies which did not have PCR-proven mono-infection \u2019s blood , 23 was \u2019s blood , 24, 25 The nPCR test proved an insubstantial proportion of participants to have mixed malaria infection. However those entities of mixed malaria seemed inconsequential clinically as nPCR proven mixed malaria did not affect either the nature of illness or treatment/outcome and conclusions of the study. Notably, this inference of clinical inconsequentiality of merely nPCR-proven mixed malaria must be methodically evaluated further as it is beyond the scope of the current study. Lost to follow-up ensued in only three patients but seemingly they all had ACPR until their day 3 follow-up. However, we cannot rule out the possibility of treatment failure beyond day 3. Remarkably, this study did not assess the efficacy of CQ alone as per the WHO protocol , rather P. vivax malaria remains intact in the study region and thus must be continued as the treatment of choice for uncomplicated P. vivax malaria. Periodic surveillances must be conducted to remain vigilant for taking immediate measures to prevent the emergence and spread of anti-malarial resistance in P. vivax.Therapeutic efficacy of CQ\u2013PQ combined regimen in uncomplicated"}
+{"text": "Plasmodium spp. In the Saharan zone of the country, recent studies have shown that Plasmodium vivax largely predominates over Plasmodium falciparum. Anti-malarial drug response of P. vivax has not been evaluated in Mauritania. The aim of the present study was to evaluate the clinical efficacy and tolerance of chloroquine to treat P. vivax malaria in Mauritanian patients.In 2006, the Mauritanian Ministry of Health adopted a new therapeutic strategy based on the systematic use of artemisinin-based combination therapy (ACT), artesunate-amodiaquine and artemether-lumefantrine, for the first- and second-line treatment of uncomplicated malaria, respectively, regardless of Plasmodium vivax-infected patients aged\u2009>\u20096\u00a0months old were enrolled in Nouakchott and Atar in September\u2013October 2013. Chloroquine was administered at the standard dose of 25\u00a0mg base/kg body weight over three days. Patients were followed until day 28, according to the standard 2009 World Health Organization protocol.A total of 128 patients (67 in Nouakchott and 61 in Atar) were enrolled in the study. Seven patients (5.5%) were either excluded or lost to follow-up. Based on the per protocol analysis, chloroquine efficacy  was 100%. Treatment was well-tolerated. One patient was excluded on day 1 due to urticaria and treated with artesunate-amodiaquine.P. vivax malaria, chloroquine may still have an important role to play in anti-malarial chemotherapy in Mauritania. Further epidemiological studies are required to map the distribution of P. vivax and P. falciparum in the country.Although the current national treatment guideline recommends artesunate-amodiaquine for the first-line treatment of uncomplicated malaria, including  Plasmodium falciparum infections occur in the sahelian south of Mauritania. In contrast, recent epidemiological data indicate that Plasmodium vivax infections predominate in the Saharan zone of the country [Malaria is one of the major public health problems in Mauritania. Every year, between 200,000 and 300,000 malaria cases are officially notified, mostly without laboratory confirmation of clinical diagnosis . Most PlP. vivax malaria is not well known in the neighbouring countries, such as Mali and Senegal. Malaria risk due to P. vivax is limited in Algeria [Plasmodium vivax was eliminated from Morocco in 2010 [P. vivax had been registered in the past in Cape Verde islands, only P. falciparum has been diagnosed during the last 20\u00a0years [P. vivax occurs in S\u00e3o Tom\u00e9 and Pr\u00edncipe, in some East African countries, and in Madagascar [The epidemiology of  Algeria . Plasmod in 2010 . Althoug20\u00a0years . Elsewhedagascar -9.P. falciparum is widespread in West Africa [P. falciparum and P. vivax infections in endemic areas, where mixed infections with these two Plasmodium species occur frequently, the efficacy of different forms of ACT to specifically treat P. vivax infections is not well established.Chloroquine-resistant t Africa . As a coP. vivax is endemic, chloroquine is still the drug of choice to treat this infection [P. vivax, well-tolerated, safe in young children and pregnant women, and much cheaper than ACT. The efficacy of chloroquine to treat P. vivax malaria has not been evaluated in Mauritania. The present single-arm study was performed to evaluate the efficacy and tolerance of chloroquine in P. vivax-infected, symptomatic patients in the Saharan zone in Mauritania where P. vivax has emerged in recent years.In other parts of the world, where nfection . ChloroqP. vivax malaria in the northern districts of Nouakchott [P. vivax malaria, confirmed by rapid diagnostic test for malaria.The clinical studies were conducted simultaneously in two study sites in central and northern Mauritania: Nouakchott , the capital city of Mauritania, and Atar   or history of fever 48\u00a0h preceding medical consultation, and ability to swallow oral medication [P. vivax malaria  [Plasmodium species, severe malnutrition, fever due to concomitant diseases, and history of allergic reaction to chloroquine were excluded.Patients spontaneously consulting at the health centres in September\u2013October 2013 were enrolled after informed written consent if the following criteria were met: age\u2009>\u20096\u00a0months old, dication . Patientulsions) , mixed PFingerprick capillary blood was collected to prepare Giemsa-stained thin and thick smears to be examined under the microscope, according to standard World Health Organization (WHO) procedures . Blood sChloroquine  was administered once daily on day 0 (10\u00a0mg base/kg body weight), day 1 (10\u00a0mg base/kg body weight), and day 2 (5\u00a0mg base/kg body weight) at the standard total dose of 25\u00a0mg base per kg body weight. Each dose administration was supervised by one of the team members, and patients were observed for at least 30\u00a0minutes after drug intake for possible vomiting or adverse effects. In case of vomiting during the observation period, the same dose was given, and the patient was observed for an additional 30\u00a0minutes. In case of repeated vomiting, the patient was withdrawn from the study protocol and rescue treatment was administered . Paracetamol (10\u00a0mg/kg body weight) was administered to all patients three times a day for fever and headache.Patients were followed on days 1, 2, 3, 7, 14, 21, and 28 . FingerpThe primary end point was determined on day 28 and classified into one of the following categories: early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), or adequate clinical and parasitological response (ACPR), as defined by the WHO . TreatmePlasmodium species identity was determined using 18S rRNA as the target molecule [P. vivax recrudescence (i.e. reappearance of the same parasite population as that of pre-treatment infection), reinfection (i.e. appearance of different parasite populations), and relapse (reappearance of the same parasite populations or appearance of different parasite populations from reactivation of hypnozoites) with certainty.Parasite DNA was extracted from filter papers using the Chelex method . Plasmodmolecule . GenotypThe sample size of 50 patients , i.e. a minimum of 60 patients, was calculated on the basis of an expected failure rate of 5%, 10% precision, and a confidence level of 95% .t-test. The significance level was P\u2009<\u20090.05.As recommended in the WHO protocol , per proad hoc Mauritanian national ethics committee and the ethics committee of the WHO . The purpose of the study was explained to the patients  in local dialect. Informed written consent was obtained from adult patients or caretakers of paediatric patients aged less than 12\u00a0years old. For adolescents aged between 12 and 18\u00a0years old, an informed written consent was obtained from both the patients themselves and their parents or legal guardians.The study was reviewed and approved by an P. vivax.A total of 128 patients were enrolled in the study: 67 in Nouakchott and 61 in Atar. Of 128 enrolled patients, 3  were excluded after inclusion for protocol violation on day 1 (n\u2009=\u20091), withdrawal of consent on day 1 (n\u2009=\u20091), and development of urticaria on day 1 (n\u2009=\u20091) (Table\u00a0In both Nouakchott and Atar, per protocol analysis showed 100% chloroquine efficacy Table\u00a0. Due to P. vivax in West Africa. The results indicate that chloroquine is highly efficacious and well-tolerated in the predominantly Moor population residing in Nouakchott and Atar. Fever and parasitaemia were rapidly cleared. Generalized urticaria developed 2\u20133 hours after the first dose of chloroquine in a 7-year old boy. The patient was immediately treated with injectable dexamethasone and oral antihistaminics, and the patient was excluded from the protocol by precaution and treated with artemether-lumefantrine.This is the first evaluation of chloroquine efficacy against P. vivax has been reported from some foci in Asia, Oceania, South America, and more recently, from Madagascar and Ethiopia in eastern Africa [P. vivax parasitaemia after treatment cannot be attributed at present to recrudescence, reinfection, or relapse with certitude due to technical limitations. A co-administration of chloroquine and primaquine may reduce the recurrence rate due to relapse.Although chloroquine was found to be highly effective in the present study, chloroquine-resistant n Africa ,19-23. IP. vivax malaria in Mauritania.A nationwide survey to establish an up-to-date epidemiology of glucose-6-phosphate dehydrogenase deficiency, which presents a potential risk of primaquine-induced haemolysis, and clinical studies on chloroquine co-administered with primaquine are warranted to further assess and determine the role of chloroquine for the treatment of P. vivax and glucose-6-phosphate dehydrogenase deficiency in Mauritania are required before implementing concrete actions to control and eliminate malaria from the country. Based on the results of the present clinical data, the current national treatment guideline that recommends the systematic use of artesunate-amodiaquine for the first-line treatment of acute uncomplicated P. falciparum and P. vivax malaria in Mauritania may need to be revised if further studies confirm chloroquine efficacy in other sites in the country.More precise, up-to-date epidemiological data on the distribution of"}
+{"text": "The Republic of Congo adopted a new anti-malarial treatment policy in 2006, with artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL) as the first- and second-line anti-malarial drugs, respectively. Only three clinical studies had been conducted before the policy change. A randomized study on these two artemisinin-based combinations was conducted, and the effect that sickle cell trait may have on treatment outcomes was evaluated in children under 10\u00a0years old followed during 12\u00a0months in Brazzaville in 2010\u20132011.Plasmodium falciparum episodes were randomly treated with co-formulated ASAQ (Coarsucam\u00ae) or AL (Coartem\u00ae). Patients were followed according to the 2009 World Health Organization protocol for the evaluation of anti-malarial drug efficacy. Plasmodium falciparum recrudescent isolates were compared to pre-treatment isolates by polymerase chain reaction (PCR) to distinguish between re-infection and recrudescence. PCR-uncorrected and PCR-corrected responses to treatment were determined using per protocol analysis. Haemoglobin type  was determined by PCR.A cohort of 330 children under 10\u00a0years of age living in a suburban area in the south of Brazzaville were passively followed for registration of malaria episodes. Uncomplicated Of 282 clinical malaria episodes registered during 1-year follow-up period, 262 children with uncomplicated malaria were treated with ASAQ (129 patients) or AL (133 patients). The PCR-corrected efficacy, expressed as the percentage of adequate clinical and parasitological response, was 97.0\u00a0% for ASAQ and 96.4\u00a0% for AL. Among ASAQ-treated patients, 112 (86.8\u00a0%) carried AA genotype and 17 (13.2\u00a0%) were AS carriers. The PCR-corrected efficacy was 96.4\u00a0% for AA-carriers and 100\u00a0% for AS-carriers treated with ASAQ . Among 133 AL-treated children, 109 (82\u00a0%) carried AA, and 24 (18\u00a0%) AS genotypes. The PCR-corrected efficacy was 96.7\u00a0% among AA-carriers and 95.2\u00a0% among AS-carriers . Nausea, jaundice, headache, dizziness, vomiting, pruritus, abdominal pain, and diarrhoea were registered as adverse events in both groups. ASAQ was associated with significantly more frequent adverse events (P\u00a0<\u00a00.05).This first randomized study in Brazzaville confirmed the excellent efficacy of these co-formulated anti-malarial drugs in children. Sickle cell genotype did not influence the treatment efficacy of artemisinin-based combination therapy. In the 1990s, with the emergence and spread of chloroquine resistance, malaria accounted for 37\u00a0% of hospitalizations and represented the first cause of admissions to paediatric service of the main hospital in Brazzaville, Republic of Congo (RoC) . In healPlasmodium falciparum isolates analysed in earlier studies carried the key pfcrt K76T mutation associated with chloroquine resistance, while quadruple mutations [defined as dihydrofolate reductase (dhfr) mutant alleles Asn51Ile\u00a0+\u00a0Cys59Arg\u00a0+\u00a0Ser108Asn and dihydropteroate syntase (dhps) mutant allele Ala437Gly] were present in more than 50\u00a0% of the isolates obtained from patients with SP treatment failure .P\u00a0=\u00a00.6). There were 17 of 129 (13.2\u00a0%) children and 24 of 133 (18.0\u00a0%) children with AS in the ASAQ and AL groups, respectively. The baseline characteristics of enrolled patients in each group were similar, with the exception of the mean geometric parasite density (P\u00a0<\u00a00.001) (Table\u00a0There were 112 (86.8\u00a0%) and 109 (82.0\u00a0%) AA haemoglobin genotype in ASAQ and AL treatment groups, respectively  children received this drug during their first malaria episode, 51/129 39.5\u00a0%) during their second, third or fourth malaria episodes  received this drug during the first malaria episode, of which 69 (94.5\u00a0%) were successfully treated (ACPR), four (5.5\u00a0%) failures, four new infections, and 10 lost-to-follow-up  still had positive smears on day 2, with a geometric mean of 87 asexual parasites/\u00b5L. On day 3, none of the patients followed (n\u00a0=\u00a0121) had a positive smear.P\u00a0=\u00a00.02). This side effect was followed by vomiting, abdominal pain, diarrhoea, headache, jaundice, and dizziness, with similar frequencies. Pruritus occurred in four ASAQ treated children, while only one AL treated child reported pruritus. Rashes appeared in one ASAQ treated child.Between day 0 and day 7, a total of 166 and 111 adverse events were reported by patients treated with ASAQ and AL, respectively Table\u00a0. AstheniDizziness, nausea, headache and jaundice were reported more frequently (P\u00a0<\u00a00.05) in children aged 5\u201310\u00a0years in both ASAQ and AL treatment groups. Skin manifestations  were mostly observed in under-five children in both treatment groups. The difference in the frequency of fatigue, vomiting, and abdominal pain was not significantly different (P\u00a0>\u00a00.05) between the age groups <5\u00a0years and 5\u201310\u00a0years old.This is the second study on ASAQ and AL efficacy and safety in Brazzaville, the capital city of RoC. Under 10\u00a0years children of the cohort with uncomplicated falciparum malaria were randomly treated with ASAQ or AL. The presence of sickle cells was determined to assess if they influence ACT efficacy in children.All children recruited in the cohort reside in the suburban area in the south of Brazzaville. Despite the impact of urbanization, malaria transmission is still intense in this area where children who are frequently infected with high parasite loads without signs of severe malaria are usually treated in health facilities for uncomplicated malaria. In the present cohort, high parasitaemias  were similarly distributed in under-five children and those between 5 and 10\u00a0years old.Data of this randomized study confirm the high and comparable efficacy of these two forms of ACT in Brazzaville, confirming previous non-randomized trials conducted from 2005 to 2006 , 18. DatTreatment was systematically switched after a previous uncomplicated malaria episode: the patient was treated with AL if he or she had been previously treated with ASAQ, and the patient was treated with ASAQ, if the previous treatment had been AL. Both combinations for the first- and second-line malaria management had a comparable efficacy. In accordance with the Congolese national policy, ASAQ treatment failures were treated with AL. These results suggest that in public health centres either ASAQ or AL can be used as the first-line drug and offer the possibility to use AL to treat patients who are reluctant to take ASAQ due to side effects or past history of allergic reactions. In the early 2000s, sub-Saharan African countries started to implement the new anti-malarial drug policy based on ACT . Among 4In this study the percentage of patients with at least one side effect associated with ASAQ intake was higher than that of patients treated with AL. In particular, asthenia was reported more frequently after ASAQ treatment. Previous studies have reported that patients receiving AQ monotherapy or AQ in combination with another anti-malarial drug complained of fatigue, abdominal pain, and vomiting , 33. In P. falciparum malaria [Sickle cell disease is the most common genetic disease in sub-Saharan African countries. It was estimated that in 2010 the world recorded 305,800 newborns with sickle cell trait, with DR Congo  and Nigeria  being the most affected . In Gabo malaria \u201341. It i malaria . MoreoveThis first randomized study in Brazzaville, conducted 6\u00a0years after the change of policy against malaria and 6\u00a0years after the first ASAQ and AL non-randomized studies, confirms the high efficacy of these combinations, both of which are currently used in co-formulation whereas in 2006, only AL was co-formulated. ACT efficacy in patients carrying AA and AS haemoglobin allows the use of these drugs regardless of sickle cell trait. The recurring problem of adverse events associated with ASAQ warrants further studies on the acceptability of both ACT."}
+{"text": "Artemisinin-based combination treatments (ACTs) or intravenous artesunate are used in over 100 countries for uncomplicated or severe falciparum malaria. Although intravenous artesunate may cause delayed haemolytic anaemia, there is little evaluation of the temporal changes in haematocrit following ACTs.Clinical and parasitological parameters were measured before and following treatment of uncomplicated falciparum malaria in children with artesunate-amodiaquine (AA) or artemether-lumefantrine (AL) over 6-weeks. Changes in haematocrit were characterized in individual patients based on a haematocrit <30\u00a0% or \u226530\u00a0% before and following treatment. Kinetics of the deficit in haematocrit from <30\u00a0% until attainment of \u226530\u00a0% were estimated by a non-compartment model.-1 asexual forms were significantly higher in children with \u22655 units compared to <5 units fall in haematocrit 21 or 28\u00a0days after treatment began. Irrespective of pattern, declines in haematocrit deficit from <30\u00a0% were mono-exponential, with similar half-times for AA- and AL-treated children (1.32 d versus 1.14 d). Anaemia half-time correlated significantly positively with anaemia recovery time in the same patients . Bland-Altman analysis of 9 or 10 multiples of anaemia half-time and anaemia recovery times showed narrow limit of agreement with insignificant biases .In 248 of 1180 children eligible for evaluation, common temporal patterns were: no change or increase in haematocrit from\u2009\u2265\u200930\u00a0% [50\u00a0% of patients], haematocrit >30\u00a0% at presentation declining to <30\u00a0% within 2\u00a0weeks  [19\u00a0% of patients], and haematocrit <30\u00a0% at presentation increasing to\u2009\u2265\u200930\u00a0% [23\u00a0% of patients]. Haematocrit >30\u00a0% at presentation declining to <30\u00a0%, 3\u20135 weeks later  occurred in 7 children (3\u00a0%). Fall in haematocrit \u22655 units following treatment occurred in 57 children [23\u00a0%] between 14 and 28\u00a0days after treatment began. Baseline parasitaemia and proportion with > 100,000\u03bcLIn uncomplicated falciparum malaria, increases or falls in haematocrit are common following ACTs. Falls in haematocrit\u2009\u2265\u20095 units are common and may or may not result in early or late anaemia. In children who recovered from acute falciparum malaria-associated anaemia following ACTs, decline in haematocrit deficit is mono-exponential.PACTR201508001188143, 3 July 2015; PACTR201508001191898, 7 July 2015 http://www.pactr.org.Pan African Clinical Trial Registry The online version of this article (doi:10.1186/s12879-015-1219-y) contains supplementary material, which is available to authorized users. Artemisinin-based combination treatments (ACTs) are the first line treatments of uncomplicated falciparum malaria in over 100 countries  and intrPlasmodium falciparum malaria-associated anaemia, is common in children [Plasmodium falciparum infections to ACTs, namely rapid clearance of asexual parasitaemia and recovery from the symptoms and signs of the infections [children , 13, occchildren , and is children \u201318. Two fections , are oftfections , 20. A rfections .Despite adoption of ACTs as first line treatments in many endemic countries, there is little information on the patterns of change in haematocrit in individual African children following ACTs of uncomplicated infections. Such information may not only assist with the community management of malaria-associated falls in haematocrit to <30\u00a0%, but also with understanding the extent of falls in haematocrit in the different endemic settings, the relationship between falls and time-course of treatment, and the characteristics of the children with different pattern of change in haematocrit following ACTs.In the present study, the temporal patterns of haematocrit after artemisinin-based combination treatments of uncomplicated falciparum malaria were evaluated in a group of children resident in an endemic area of southwestern Nigeria. The main aims were to: (i) characterize the changes in haematocrit with time in the individual patients, (ii) evaluate the factors contributing to moderate fall  in haematocrit following treatment, and (iii) characterize, kinetically, recovery from the fall in haematocrit below 30\u00a0% associated with the different patterns.Plasmodium falciparum malaria-associated anaemia in children before, during and after artemisinin-based combination treatments . The details of the larger study have been reported elsewhere [The study was a prospective study conducted between April 2008 and December 2011 in Ibadan, southwestern Nigeria- an endemic area. It was nested in a larger study of lsewhere , 23. ThePlasmodium falciparum mono-infection \u22652000\u00a0\u03bcL\u22121 of blood, no history of antimalarial drug ingestion in the two weeks prior to enrolment, absence of severe malaria [Briefly, patients were enrolled in the study if they were aged 6\u00a0months\u201315\u00a0years, had symptoms compatible with acute uncomplicated malaria with  malaria  and writEnrolled patients were randomized to receive artemether-lumefantrine or artesunate-amodiaquine (co-formulated). Artemether-lumefantrine  was given according to body weight: patients weighing 5\u201314\u00a0kg received one tablet, those weighing 15\u201324\u00a0kg received two tablets, those weighing 25\u201334\u00a0kg received three tablets, and those weighing >34\u00a0kg received four tablets at presentation (0\u00a0hour), 8\u00a0hours later and at 24, 36, 48 and 60\u00a0hours after the first dose. Each tablet of artemether-lumefantrine contains 20\u00a0mg of artemether and 120\u00a0mg of lumefantrine. Artesunate-amodiaquine  was also given according to body weight: patients weighing \u22654.5 to <9\u00a0kg received one tablet, those weighing \u22659 to <18\u00a0kg received one tablet and those weighing \u226518 to <24\u00a0kg received one tablet of the following formulations: 25\u00a0mg/67.5\u00a0mg, 50\u00a0mg/135\u00a0mg, 100\u00a0mg/270\u00a0mg of fixed dose combination of artesunate/amodiaquine, respectively daily for 3\u00a0days. Children weighing 24\u201336\u00a0kg and >36\u00a0kg received 1.5 and 2 tablets, respectively of 100/270\u00a0mg of fixed dose combination of artesunate/amodiaquine daily for 3\u00a0days.All drugs were given orally. All doses of artesunate-amodiaquine were given under direct observed therapy as were the doses of artemether-lumefantrine given at 0, 8, 24 and 48\u00a0hours. Doses of artemether-lumefantrine at 36 and 60\u00a0hours were given by parents/guardians of the children at home and enquiries were made by telephone calls at the expected times of administration to confirm that the doses were actually given. Artemether-lumefantrine was not given with fatty meals.Thick and thin blood films prepared from finger prick were stained with Giemsa and examined by light microscopy under oil immersion objective lens 1000 x magnification by two assessors who did not know the drug regimen of the patients. A senior member of the study team reviewed the slides if there was any disagreement by the two microscopists. In addition, the slide of every fourth child enrolled in the study was reviewed by the senior member. Thick and thin blood films obtained from each child as soon as they came to the clinic on to blood slides were carefully labeled with the patients\u2019 codes and air-dried before being stained. Follow-up with clinical and parasitological evaluation was done daily on days 1\u20133 and on days 7, 14, 21, 28, 35 and 42.\u22121 of blood. A slide was considered parasite negative if no asexual or sexual parasite was detected after examination of 200 microscope fields. The cure rates on days 21 and 28 were adjusted on the basis of the PCR (polymerase chain reaction) genotyping results of paired samples of patients with recurrent parasitaemia after day 7 of starting treatment as previously described [Parasitaemia, asexual or sexual, in thick films were estimated by counting asexual and sexual parasites relative to 500 leukocytes, or 500 asexual or sexual forms whichever occurred first. From this figure, the parasite density was calculated assuming a leukocyte count of 6000\u00a0\u03bcLCapillary blood collected before treatment and during follow-up was used to measure hematocrit using a microhaematocrit tube and microcentrifuge . Anaemia was defined as a haematocrit <30\u00a0% , 13. AnaHaematocrit \u226530\u00a0% before and following treatment.Haematocrit \u226530\u00a0% before treatment, declining to haematocrit <30\u00a0% within 2\u00a0weeks of treatment, and thereafter increasing to \u226530\u00a0% by 6\u00a0weeks after treatment .Haematocrit \u226530\u00a0% before treatment followed by a decline to haematocrit <30\u00a0%, 3\u20135 weeks following treatment and then increasing to \u226530\u00a0% by 6\u00a0weeks after treatment .Haematocrit \u226530\u00a0% before treatment, 2 consecutive haematocrit <30\u00a0% within 14\u00a0days of treatment, followed by a rise to haematocrit \u226530\u00a0% between 14 and 21\u00a0days and a fall to <30\u00a0% between day 28\u201335 .Haematocrit <30\u00a0% before and following treatment (persistent anaemia).Haematocrit <30\u00a0% before treatment followed by an increase to \u226530\u00a0% after treatment .Haematocrit\u2009<\u200930\u00a0% at presentation and till day 14, followed by a rise to \u226530\u00a0% on days 21\u201328 and a fall to <30\u00a0% on days 35\u201342.Haematocrit <30\u00a0% before treatment, followed by further falls to \u226425\u00a0%, 3\u20135 weeks following treatment.Unclassifiable.Haematocrit <30\u00a0% and \u226530\u00a0% were the reference points in all classified patterns. Temporal changes in haematocrit were classified into the following patterns Fig.\u00a0.Fig. 1Clversus time were obtained, by the trapezoidal rule using the computer program Turbo Ken  as previously described . AUC was also obtained manually by calculating the average haematocrit values between two consecutive time measurements and multiplying it by the time interval between the measurements, and summing up all the values, in a manner similar to that for the numerical estimation of area under a drug concentration-time curve [versus time, and the apparent terminal half-time of anaemia (t1/2(anaemia)) was calculated from ln/(\u03bb).The kinetics of the disposition of deficit in haematocrit from 30\u00a0%, that is, of anaemia, was as previously described , 26. Brime curve . Both meme curve . The appP. falciparum asexual and sexual forms. Proportions were compared by calculating \u03c72 using Yates\u2019 correction, Fisher\u2019s exact or Mantel Haenszel tests. Normally distributed, continuous data were compared by Student\u2019s t test and analysis of variance (ANOVA). Data not conforming to a normal distribution were compared by the Mann-Whitney U tests and the Kruskal Wallis tests. The relationship between two continuous variables was assessed by Spearman\u2019s rank correlation coefficient. Agreement between pharmacodynamic and pharmacokinetic methods of evaluating recovery from anaemia was assessed by Bland-Altman analysis [P-values of <0.05 were taken to indicate significant differences. Data were double entered serially using patients\u2019 codes and were only analyzed at the end of the study.Data were analyzed using version 6 of Epi-Info software  and the analysis . P-value\u22121) was found in 8.9\u00a0% of the patients. In general, PCR-corrected cure rates were similar for both artesunate-amodiaquine and artemether-lumefantrine .Between April 2008 and December 2011, 1180 patients were enrolled in prospective studies of the efficacy of artemisinin-based combination treatments. All patients, at enrolment and during follow-up, had haematocrit measured. However, two hundred and forty eight patients who had haematocrit measured during the entire period of follow-up were analysed. Of these, 177 and 71 were treated with artesunate-amodiaquine and artemether-lumefantrine, respectively. The clinical characteristics of the patients at enrolment are summarized in Table\u00a0post-hoc comparison). Parasitaemia was similar in all three temporal patterns. The features of the other temporal patterns  are summarized in Table\u00a0Tables\u00a0Patterns , 5 and 7\u22121, parasitemia cleared within 1\u00a0day in 6 of 7 children, 3 of 5 children treated with artesunate-amodiaquine had total dose of artesunate >11\u00a0mg/kg, and none of the children required blood transfusion. Also, none of the patients was older than 9\u00a0years. On the whole, virtually all of these children were anaemic for a period of approximately 14\u00a0days . After day 7, overall, 57 of 122 children and 55 of 213 children with \u22655 and <5 units fall in haematocrit, respectively between days 14 and 28 were anaemic . However, the proportion of children who were anaemic on day 14 in the 2 groups, respectively were similar . On day 21, 20 of 48 children and 18 of 75 children in the 2 groups, respectively were anaemic . On day 28, 8 of 22 and 5 of 73 children .Because of very few patients with late fall in haematocrit (n\u2009=\u20097), the features of children with \u22655 units and <5 units fall in haematocrit from baseline following artemisinin-based combination treatments were evaluated. Based on the number of patients evaluated per visit day, on day 7, 16 of 23 children and 62 of 87 children had \u22655 and <5 units fall in haematocrit, respectively, following treatment. The difference between these proportions was not significant . The median doses of the drugs were also similar in the two groups (data not shown). Time to recover from anaemia was significantly shorter in children with early than in those with late monophasic falls in haematocrit . The mean AUC of deficit in haematocrit versus time was significantly lower in children with early monophasic fall than in those with late monophasic fall . Declines of deficit in haematocrit were monoexponential in both patterns .Forty-eight (31 treated with artesunate-amodiaquine and 17 treated with artemether-lumefantrine) and 7 (5 treated with artesunate-amodiaquine and 2 treated with artemether-lumefantrine) children, respectively, had early and late monophasic falls in haematocrit. The children from the two groups, respectively had similar clinical and parasitological parameters at presentation  and it was similar in anaemic children treated with artesunate-amodiaquine and artemether-lumefantrine   of 1.3 d . Estimated mean t\u00bd were similar in children treated with artesunate-amodiaquine and artemether-lumefantrine .The demographic and other characteristics of the 57 children (n\u2009=\u200939 for artesunate-amodiaquine and n\u2009=\u200918 for artemether-lumefantrine) have been summarized in Table\u00a0r\u2009=\u20090.55, P\u2009<\u20090.0001, see Additional file P\u2009=\u20090.19 and 0.63, respectively). However, there was a statistically significant bias at multiples of 7 or 8 anaemia half-times .The relationship between the half-time of decline in haematocrit deficit from 30\u00a0% and anaemia recovery time in the same patients with anaemia at presentation was evaluated in 57 children. The mean half-time of decline in haematocrit deficit from 30\u00a0% was 1.3\u00a0days . The median anaemia recovery time was 14\u00a0days (range 1\u201321). There was a significantly positive correlation between half-time of decline in haematocrit deficit from 30\u00a0% and anaemia recovery time  and anaemic (Pattern 6) children at presentation , 25, 32.Of theoretical interest, for which no child was classified is: no anaemia at presentation, followed by anaemia, recovery from anaemia, and a late fall in haematocrit to anaemia level (Pattern 4). This pattern appears to have combined features of Pattern 2 and Pattern 3 and may be considered a variant of both Patterns 2 and 3. It would appear that the 3 patients included under pattern 7 appeared to have \u2018undulating pattern\u2019. We do not have an explanation for this pattern.It is noteworthy that a late fall in haematocrit to a level below normal occurred in 3\u00a0% of the children  syndrome , 45 consDeclines in haematocrit deficit from 30\u00a0%  were mono-exponential irrespective of whether the deficit was early or late, suggesting that using a non-compartment model, recovery from apparently uncomplicated falciparum malaria-associated anaemia is a first order process. In this and other contexts, early fall in haematocrit to <30\u00a0% differs from late fall to <30\u00a0% by its shorter half-time of anaemia. These two patterns also differ in their anaemia recovery times and AUC of deficit in haematocrit versus time. These differences suggest that, on the whole, the late monophasic anaemia may be more \u2018intense\u2019 than the early monophasic anaemia. A further application of pharmacokinetic principles should permit treatment of the increases in haematocrit associated with artemisinin-based combination treatments of uncomplicated falciparum malaria.In this study, a pharmacodynamic-pharmacokinetic approach was used to quantify the time to resolve uncomplicated malaria-associated anaemia following artemisinin-based combination treatments. The approach showed the following: (i). Anaemia half-time correlated significantly with anaemia recovery time in the same patient. (ii). Greater than 8 multiples of anaemia half-times showed an insignificant bias when a Bland-Altman analysis was used. Thus, confirming that >8 multiples of anaemia half-times can be used interchangeably with anaemia recovery time for the evaluation of recovery from uncomplicated malaria-associated anaemia following artemisinin-based combination treatments. Therefore, the agreement between 9 or 10 multiples of anaemia half-times and anaemia recovery time was not unexpected since it takes 9 or 10 half-times for 99.6 or 99.9\u00a0% completion of an elimination process in a simple one compartment pharmacokinetic model. The ratio of anaemia recovery time to anaemia half-time of approximately 10 in the patients also indirectly support the model.There are limitations of the present study. First, although the clinical and parasitolgical features of children with fall in haematocrit to anaemia level on days 21\u201342 after commencement of treatment were characterised, the nature of the anaemia was not fully characterized (i.e. whether it was haemolytic or not in nature). Second, in the children with late fall in haematocrit to anaemia level, quantification of once-infected and infected red blood cells and the disposition of these red cells during the course of follow-up was not evaluated. Third, The precise contribution of the background causes of anaemia in children from the endemic area  to the time-course of haematocrit was not assessed.Increases in haematocrit are common following artemisinin-based combination treatments of uncomplicated falciparum malaria, but falls in haematocrit from baseline \u22655 units are also relatively common and may or may not result in early or late anaemia. In children who recovered from acute falciparum malaria-associated anaemia following artemisinin-based combination treatments, declines in haematocrit deficit  are mono-exponential."}
+{"text": "Plasmodium falciparum resistance to artemisinin has been reported in South-East Asia. Long half-life drugs are increasingly being used for malaria prevention. The potential spread of parasite resistance to these regimens is real and makes regular efficacy surveillance a priority.versus SP\u2009+\u2009amodiaquine (AQ), and SP alone, was conducted in two villages of Mali. PCR was used to distinguish new infections from recrudescent P. falciparum infections. Patients were followed for 28\u00a0days to assess treatment efficacy.From August to December 2004 and July to December 2005, a randomized open label trial of sulphadoxine-pyrimethamine (SP)\u2009+\u2009artesunate (AS) P. falciparum malaria were recruited. Baseline characteristics were similar in the three treatment arms. Crude ACPRs were 94.9%; 98.6% and 93.5% for SP\u2009+\u2009AS; SP\u2009+\u2009AQ and SP alone arms respectively . After PCR adjustment, cACPRs were 99%; 100% and 97.2% for SP\u2009+\u2009AS; SP\u2009+\u2009AQ and SP alone arms, respectively .Overall 912 children aged between six to 59\u00a0months, with uncomplicated P. falciparum malaria in Mali.Sulphadoxine-pyrimethamine\u2009+\u2009amodiaquine therapy was as efficacious as sulphadoxine-pyrimethamine\u2009+\u2009artesunate, but more efficacious than sulphadoxine-pyrimethamine alone in the treatment of uncomplicated  Plasmodium falciparum resistance to chloroquine (CQ) has challenged malaria control efforts. Previous studies indicated that CQ resistance rates were above 25% in several sites of Mali [The emergence of  of Mali -3. These of Mali . The two of Mali . However of Mali ,8 and ma of Mali ,10.in vivo efficacy trial was designed to evaluate this non-artemisinin-based combination and compare it with SP\u2009+\u2009artesunate (AS), and SP alone.A nationwide effort of implementation of ACT was ongoing by the Malian National Malaria Control Programme with support from several partners. WHO recommendations allowed the use of non-artemisinin combination treatment regimens for the treatment of uncomplicated malaria in settings, where ACT was not available and the component drugs were efficacious and well tolerated . In MaliAnopheles gambiae sensu lato (95.5% of vector population) and Anopheles arabiensis (4.5%). The mean monthly entomologic inoculation rate was 2.8 infectious bites per person, with marked seasonal variations [The trial was conducted in two malaria hyper endemic rural villages, Kolle and Bancoumana, both located 57 to 60 kilometers southwest of Bamako, Mali. Data were collected in Kolle and Bancoumana Heath centers, respectively. Falciparum malaria is endemic and seasonal with parasitaemia prevalence ranging from 40\u201350% in the dry season (October-May) and 70\u201385% in the rainy season (June-September) . The mairiations .The Ethic Committee of the Faculty of Medicine, Pharmacy and Odonto-Stomatology, University of Bamako approved the study protocol. Permission was also obtained from community and local authorities. Informed consent was obtained from parents or infant guardians, according to previously described procedures . Brieflyin vivo criteria [P. falciparum with a parasitaemia of 2,000 \u2013 200,000/\u03bcl; ii) axillary temperature of\u2009\u2265\u200937.5\u00b0C; iii) haemoglobin\u2009\u2265\u20095\u00a0g/dL; iv) absence of febrile illness caused by diseases other than malaria, and v) absence of danger signs , informed consent granted by parents of legal guardians. Exclusion criteria were i) haemoglobin\u2009<\u20095\u00a0g/dL; ii) severe conditions, such as prostration, respiratory distress, renal failure, hypoglycaemia, shock, bleeding, severe vomiting; and iii) a history of allergy or other severe adverse reaction to the study drugs.The study was a randomized open label clinical trial conducted between August and December 2004 and July and December 2005. Patients were enrolled if they were aged between six and 59\u00a0months and fulfilled the following WHO criteria : i) micrBlock-randomization was used to allocate patients to the three treatment arms . Treatment assignments were done through sequentially numbered opaque envelops that concealed the actual treatment to which the patient was randomized. The study statistician generated the randomization code using a computer system. The study microscopists assessing malaria smears were kept blinded throughout the study duration.Enrolled patients were followed-up for 28\u00a0days. They were asked to return for clinical and/or biological follow-up on days 1, 2, 3, 7, 14, 21, 28, or on days of recurrent illness. Patients or guardians were asked about drug consumption since the last clinic visit, and about any adverse event. At each visit, a brief physical examination, including axillary temperature, was performed and blood was taken for thick smears. Patients were asked to return to the clinic if they became ill outside of regular visit times. Cases of treatment failures were treated with quinine. Patients failing to report to the clinic for two consecutive scheduled visits were considered lost to follow up.Arm 1 received SP\u2009+\u2009AS and arm 2 received SP\u2009+\u2009AQ, both in a loose combination and arm 3 received SP alone. Treatment was administered according to body weight at the following doses: AQ , 10\u00a0mg/kg/day on days 0, 1 and 2; AS , 4\u00a0mg/kg/day on days 0, 1 and 2; SP, 25\u00a0mg/kg of sulphadoxine and 1.25\u00a0mg/kg of pyrimethamine  in single dose on day 0. All drugs were administered directly by the study team at the respective health centres. Thereafter, each child was observed for 60\u00a0minutes. If vomiting occurred within 30\u00a0minutes, the dose was repeated; for vomiting after 30\u00a0minutes, a half-dose was administered. In the case of persistent vomiting (3 or more consecutive vomiting incidents), the child received a rescue treatment with intravenous quinine and withdrawn from the study. After the day of enrolment (day 0), patients were assessed on days 1, 2, 3, 7, 14, 21 and 28 using the same procedures described above.Capillary blood was obtained by fingerpick; thick and thin blood films were made and stained with 5% Giemsa for 20\u00a0minutes. Parasite densities were determined from thick blood smears by counting the number of asexual parasites per 300 WBCs assuming a WBC count of 7,500/\u03bcl . The sammsp2) genes was used on samples from patients who failed after day 7. At enrolment and during the follow-up visits or unscheduled visit , blood spots were obtained on filter papers (3MM Whatman) for molecular analysis.To distinguish recrudescent from new infections, molecular genotyping of the parasite merozoite surface protein 2  re-infection, if the alleles of the pre- and post-treatment samples were distinct; (iii) mixed recrudescence and re-infection, if similar alleles were found in the pre- and post-treatment samples, but with additional distinct alleles identified; (iv) indeterminate, if either or both the pre- and post-treatment samples could not be amplified.DNA was extracted using the \u201cmethanol method\u201d . SamplesTreatment outcomes were classified following the WHO anti-malarial drug efficacy guidelines 2003 as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), and adequate clinical and parasitological response (ACPR) . A per pThe required sample size was calculated using the non-inferiority assumption that the three study drugs were equally effective in terms of the primary endpoint, with a two-sided \u03b1 of 0.05 and a power of 85%. The maximum accepted difference in efficacy between the two study treatments was set at 5%. On the basis of an efficacy estimate of 95% on day 28, a total of 900 subjects were necessary (300 in each arm including the 10% lost during follow-up). No interim analysis was planned.U-test for non-normally distributed continuous variables. The proportion of patients failing treatment was compared across groups using the Chi-square test. The paired t-test was used to estimate the increase in haemoglobin content from baseline to different time-points of regular follow-up. Statistical significance was set at p\u2009<\u20090.05.Data were double entered using Microsoft ACCESS and statistical analysis was performed using SPSS version 11.0 . Baseline characteristics of subjects among groups were compared using the Fisher Exact Test and Chi-square tests for categorical variables and the Mann-Whitney The trial profile is summarized in Figure\u00a0versus SP\u2009+\u2009AQ, p\u2009=\u20090.01; SP\u2009+\u2009AS versus SP, p\u2009=\u20090.5; SP\u2009+\u2009AQ versus SP, p\u2009=\u20090.001)  (Table\u00a0There were 1.4% n\u2009=\u2009294) early treatment failures for SP alone but none in the SP\u2009+\u2009AS and SP\u2009+\u2009AQ arms   and SP\u2009+\u2009AQ . No such decrease was found for the SP alone arm   Figure\u00a0b. Using SP\u2009+\u2009AS, SP\u2009+\u2009AQ and SP alone were well tolerated. The number of patients reporting any symptoms or signs within the first week after treatment began was statistically similar between the three treatment arms Figure\u00a0. There wP. falciparum malaria in children less than five years of age. After PCR correction, the highest cACPR was observed with SP\u2009+\u2009AQ although the difference was not significant. Although these data are now over a decade old, they are similar to another study from a different site in Mali [This study found that the non-artemisinin-based combination of SP\u2009+\u2009AQ was as efficacious as an artemisinin-based combination of SP\u2009+\u2009AS for the treatment of uncomplicated  in Mali  and also in Mali . The dat in Mali . In SP\u2009+P. falciparum to SP was found, but assessment of the clinical efficacy of combinations containing SP showed adequate clinical and parasitological response rates of 90.3% [The artemisinin-based combination, SP\u2009+\u2009AS, showed faster parasite clearance than SP\u2009+\u2009AQ, but both regimens resulted in prompt fever reduction. Despite the lower proportion of fever and parasitaemia observed during the first three days after treatment in the ACT arm (SP\u2009+\u2009AS), there were more new infections in this group than in the non-ACT arm (SP\u2009+\u2009AQ). Other studies have made similar observations. For example, in Rwanda , the Uniof 90.3% , 94% [27of 90.3%  in the tof 90.3% . Both coThe Malian Ministry of Health is providing AS\u2009+\u2009AQ and AL free to children less than five years of age. Widespread implementation of ACT in this vulnerable population should produce a significant reduction in malaria mortality and morbidity in Mali. However, it is essential to continue monitoring the efficacy of this regimen, and to consider alternatives should evidence of resistance develop. Intermittent preventive treatment of children with SP\u2009+\u2009AQ, given during the malaria transmission season provided substantial protection against clinical episodes of malaria, malaria infection, and anaemia in children using an Long Lasting Insecticidal Nets (LLIN). As was found in this study, SP\u2009+\u2009AQ was shown to be safe and well tolerated ,32. ThesP. falciparum malaria in Mali. This study further supports the use of SP\u2009+\u2009AQ for SMC and of SP for IPTp in West Africa.Sulphadoxine-pyrimethamine\u2009+\u2009amodiaquine therapy was as efficacious as sulphadoxine-pyrimethamine\u2009+\u2009artesunate, but more efficacious than sulphadoxine-pyrimethamine alone in the treatment of uncomplicated"}
+{"text": "P. falciparum and P. vivax malaria.In a randomized controlled trial Tim Davis and colleagues investigate Artemisinin-naphthoquine versus artemether-lumefantrine for the treatment of Please see later in the article for the Editors' Summary Plasmodium species are transmitted, especially in children, who bear the brunt of infection in endemic areas. In Papua New Guinea (PNG), artemether-lumefantrine is the first-line treatment for uncomplicated malaria, but it has limited efficacy against P. vivax. Artemisinin-naphthoquine should have greater activity in vivax malaria because the elimination of naphthoquine is slower than that of lumefantrine. In this study, the efficacy, tolerability, and safety of these ACTs were assessed in PNG children aged 0.5\u20135 y.Artemisinin combination therapies (ACTs) with broad efficacy are needed where multiple P. falciparum PCR-corrected adequate clinical and parasitologic response (ACPR) and the P. vivax PCR-uncorrected 42-d ACPR. Non-inferiority and superiority designs were used for falciparum and vivax malaria, respectively. Because the artemisinin-naphthoquine regimen involved three doses rather than the manufacturer-specified single dose, the first 188 children underwent detailed safety monitoring. Of 2,542 febrile children screened, 267 were randomized, and 186 with falciparum and 47 with vivax malaria completed the 42-d follow-up. Both ACTs were safe and well tolerated. P. falciparum ACPRs were 97.8% and 100.0% in artemether-lumefantrine and artemisinin-naphthoquine-treated patients, respectively , and P. vivax ACPRs were 30.0% and 100.0%, respectively . Limitations included the exclusion of 11% of randomized patients with sub-threshold parasitemias on confirmatory microscopy and direct observation of only morning artemether-lumefantrine dosing.An open-label, randomized, parallel-group trial of artemether-lumefantrine (six doses over 3 d) and artemisinin-naphthoquine (three daily doses) was conducted between 28 March 2011 and 22 April 2013. Parasitologic outcomes were assessed without knowledge of treatment allocation. Primary endpoints were the 42-d Artemisinin-naphthoquine is non-inferior to artemether-lumefantrine in PNG children with falciparum malaria but has greater efficacy against vivax malaria, findings with implications in similar geo-epidemiologic settings within and beyond Oceania.ACTRN12610000913077 Australian New Zealand Clinical Trials Registry Please see later in the article for the Editors' Summary Plasmodium falciparum causes most of these deaths, but P. vivax is the most common and most widely distributed cause of malaria outside sub-Saharan Africa. Infection with malaria parasites causes recurring flu-like symptoms and must be treated promptly with antimalarial drugs to prevent the development of anemia and potentially fatal damage to the brain and other organs. In the past, malaria was treated with \u201cmonotherapies\u201d such as chloroquine, but the parasites quickly developed resistance to many of these inexpensive drugs. The World Health Organization now recommends artemisinin combination therapy (ACT) for first-line treatment of malaria in all regions where there is drug-resistant malaria. In ACT, artemisinin derivatives  are used in combination with a slower acting, more slowly eliminated partner drug to prevent reemergence of the original infection and to reduce the chances of the malaria parasites becoming resistant to either drug.Malaria is a mosquito-borne parasitic disease that kills more than 600,000 people (mainly young children in sub-Saharan Africa) every year. P. falciparum, for first-line treatment of uncomplicated (mild) malaria even though this ACT is ineffective against the more common P. vivax. In this open-label randomized trial (a study in which participants are randomly assigned to receive different drugs but know which drug they are being given), the researchers ask whether an alternative ACT might be preferable for the treatment of uncomplicated malaria in young children in Papua New Guinea by comparing outcomes after treatment with artemether-lumefantrine versus artemisinin-naphthoquine . Specifically, the researchers test the non-inferiority of artemisinin-naphthoquine compared to artemether-lumefantrine for the treatment of falciparum malaria (whether artemisinin-naphthoquine is not worse than artemether-lumefantrine) and the superiority of artemisinin-naphthoquine compared to artemether-lumefantrine for the treatment of vivax malaria (whether artemisinin-naphthoquine is better than artemether-lumefantrine).Because falciparum and vivax malaria respond differently to antimalarial drugs, wherever there is transmission of both types of malaria but limited facilities for species-specific malaria diagnosis\u2014as in Papua New Guinea\u2014compromises have to be made about which ACT should be used for the treatment of malaria. Thus, Papua New Guinea's national guidelines recommend artemether-lumefantrine, which is effective against the more deadly P. falciparum, 97.8% of those treated with artemether-lumefantrine and 100% of those treated with artemisinin-naphthoquine were clear of their original P. falciparum infection  42 days after treatment. By contrast, among the patients infected with P. vivax, 30% of those treated with artemether-lumefantrine and 100% of those treated with artemisinin-naphthoquine were clear of P. vivax infection 42 days after treatment. Both ACTs were safe and well tolerated.The researchers assigned nearly 250 children (aged 0.5 to 5 years) with falciparum malaria, vivax malaria, or both types of malaria to receive six doses of artemether-lumefantrine over three days or three daily doses of artemisinin-naphthoquine. They then followed the children to see how many children in each treatment group and with each type of malaria were free of malaria 42 days after treatment . Among the patients originally infected with Plasmodium species and that artemisinin-naphthoquine may be better than artemether-lumefantrine for the treatment of uncomplicated malaria in young children in regions where P. vivax predominates.These findings indicate that artemisinin-naphthoquine was non-inferior to artemether-lumefantrine for the treatment of uncomplicated falciparum malaria among young children in Papua New Guinea and had greater efficacy than artemether-lumefantrine against vivax malaria. The accuracy of these findings may be limited by several aspects of the study design. For example, not all the artemether-lumefantrine doses were directly observed, so some children may not have received the full treatment course. Moreover, because all the study participants lived in coastal communities in Papua New Guinea where malaria is highly endemic, treatment responses among children living in areas with lower levels of malaria transmission might be different. Nevertheless, these findings suggest that artemisinin-naphthoquine should be considered alongside other ACTs for the treatment of uncomplicated malaria in regions where there is transmission of multiple http://dx.doi.org/10.1371/journal.pmed.1001773.Please access these websites via the online version of this summary at malaria ; the World Malaria Report 2013 provides details on the current global malaria situation, including information on malaria in Papua New Guinea; the World Health Organization's Guidelines for the Treatment of Malaria is availableInformation is available from the World Health Organization on malaria (in English and Spanish), including personal stories about malariaThe US Centers for Disease Control and Prevention provides information on global control of malaria, including information about malaria in Papua New Guinea, malaria in children, and ACTsInformation is available from the Roll Back Malaria Partnership on the Plasmodium vivax malariaThe Malaria Vaccine Initiative has a fact sheet on malaria (in English and Spanish)MedlinePlus provides links to additional information on trial is availableMore information about this  Plasmodium species, P. vivax remains a major obstacle to eradication because of its complex life cycle and transmission biology P. vivax infections have both been underestimated Malaria control programs incorporating artemisinin combination therapy (ACT) have contributed to a decline in malaria morbidity and mortality worldwide, renewing interest in global eradication P. falciparum and P. vivax can complicate the choice of antimalarial treatment. A recent trial of ACTs involving children from Papua New Guinea (PNG) with uncomplicated malaria showed that the most efficacious were artemether-lumefantrine for P. falciparum and dihydroartemisinin-piperaquine for P. vivax infections Transmission of both P. vivax relapses in patients treated for vivax or falciparum malaria There is a need for assessment of other ACTs that might replicate the high cure rate of artemether-lumefantrine in falciparum malaria while ensuring that vivax malaria is also effectively treated in PNG and similar epidemiologic settings. Of the few available alternatives, artemisinin-naphthoquine has been marketed in a range of countries in Africa, Asia, and Oceania as single-dose treatment based on limited pharmacologic, efficacy, and safety data in adults The aim of the present study was to compare the efficacy of three daily artemisinin-naphthoquine doses to that of a six-dose, 3-d artemether-lumefantrine regimen in PNG children aged 0.5\u20135 y with uncomplicated malaria. We hypothesized that artemisinin-naphthoquine would be well tolerated and safe in this younger vulnerable pediatric population, and that its efficacy would be (i) non-inferior to that of artemether-lumefantrine for falciparum malaria and (ii) superior to that of artemether-lumefantrine for vivax malaria.P. falciparum within 42 d of treatment after correction for reinfections identified by polymerase chain reaction (PCR) genotyping of polymorphic parasite loci, and (ii) appearance of any P. vivax parasitemia within 42 d after treatment for vivax malaria. Secondary endpoints for falciparum malaria, assessed over 42 d, were (i) reappearance of PCR-uncorrected P. falciparum parasitemia, (ii) appearance of P. vivax parasitemia, and (iii) persistence/appearance of P. falciparum gametocytes. For vivax malaria, secondary endpoints were (i) persistence/appearance of P. vivax gametocytes over 42 d and (ii) reappearance of PCR-corrected P. vivax over 42 d  reappearance of P. falciparum day-42 PCR-corrected ACPR for artemether-lumefantrine would be 95.2% (as in our previous study P. vivax ACPR for artemisinin-naphthoquine or artesunate-pyronaridine would be superior (at least double the 30.3% ACPR for artemether-lumefantrine Since artemether-lumefantrine has a high (>95%) day-42 adequate parasitologic and clinical response (ACPR) for falciparum malaria in the PNG pediatric population The original study design involved a comparison of conventional artemether-lumefantrine with both artemisinin-naphthoquine and artesunate-pyronaridine. Artesunate-pyronaridine was not initially available because concerns regarding hepatotoxicity were being addressed The study was approved by the PNG Institute of Medical Research Review Board, the Medical Research Advisory Committee of PNG, and the University of Western Australia Human Research Ethics Committee. Written informed consent was obtained from parents/guardians of all children.P. falciparum  or P. vivax (>250/\u00b5l) were eligible if (i) there were no features of severity Children aged 0.5\u20135 y presenting with an axillary temperature >37.5\u00b0C or a history of fever during the previous 24 h were screened using on-site blood film microscopy. Those with An initial standardized clinical assessment was performed, and blood was drawn for measurement of hemoglobin and blood glucose , as well as hepatic and renal function  and a full blood count . A 12-lead electrocardiograph was taken for measurement of the QT interval. Each QT interval was corrected for heart rate using Bazett's formula , eligible patients were allocated 1\u22361 to artemether-lumefantrine  twice daily for 3 d or to artemisinin-naphthoquine  daily for 3 d. Randomization was independent of Treatments were not blinded, primarily because the endpoints were based on objective clinical and parasitologic criteria. In addition, we sought to simplify drug administration as much as possible to maximize patient adherence and retention, including only once daily dosing and the avoidance of potential gastrointestinal side effects with the unnecessary ingestion of milk in children allocated artemisinin-naphthoquine Standardized assessment, including axillary temperature and blood film microscopy, was repeated on days 1, 2, 3, 7, 14, 28, and 42. Blood was drawn for full blood count and hepatorenal function, and an electrocardiograph was taken, on days 0, 3, and 7. An additional electrocardiograph was performed 4 h after the day-2 dose  and MSP1 based on paired samples collected on day 0 and at reappearance P. vivax recrudescence was determined based on genotyping of MSP1F3 and Microsatellite 16 Efficacy was assessed using WHO definitions Day-7 plasma lumefantrine concentrations were determined using a validated ultra-high-performance liquid chromatography\u2013tandem mass spectrometry assay as previously described Plasmodium species was detected, and those who defaulted from follow-up despite repeated attempts at contact. These excluded patients were retained in prespecified modified intention-to-treat analyses utilizing (i) a worst-case approach  and (ii) a best-case approach  Per-protocol prespecified analyses included children with complete follow-up or a confirmed treatment failure, and excluded those treated for malaria without confirmatory microscopy, those for whom the alternative t test, for non-normally distributed variables by Mann-Whitney U test, and for proportions by Fisher's exact test. Safety and tolerability were assessed from the incidence of symptoms/signs to day 7 using Poisson regression. Kaplan-Meier estimates were computed for each endpoint by Plasmodium species, and treatments compared by log-rank test. Differences in parasitemia, gametocytemia, and hemoglobin concentration by treatment type and time were assessed using generalized linear modeling with Bonferroni correction to adjust for multiple pairwise comparisons, and with adjustment for parasitemia in the case of hemoglobin. Unless otherwise stated, all p-values are two-tailed, with p<0.05 taken as significant.Kaplan-Meier estimates were computed for each endpoint defined by parasite species. Statistical analysis was performed using IBM SPSS Statistics version 20  and STATA/IC 11.2 . Two-sample comparisons for normally distributed variables were by Student's The first patient was recruited on 28 March 2011, and the last follow-up was completed on 22 April 2013. No adverse events occurred in the first 50 patients. A further safety assessment was scheduled when one-third of the total planned sample size of 560 had been recruited. Safety data were available for 91 artemether-lumefantrine-treated and 97 artemisinin-naphthoquine-treated children at that time . In addition, there were no differences between the treatments allocated to the 23 excluded patients who had falciparum malaria or between the treatments allocated to the six with vivax malaria (p>0.67). The 15 children (6.5% of the randomized sample) presenting with mixed P. falciparum/P. vivax infections at densities above each species-specific threshold were included in both species arms P. malariae infection completed all study procedures but was not included in analyses.Of 267 randomized children, 36 (13.5%) were excluded post hoc because of protocol violations  consistent with all six doses having been administered successfully Of the remaining 230 children, the day-42 retention was \u226592% for all species/treatment groups. Ten children  withdrew during follow-up. None of these children had experienced adverse effects; they had all relocated outside the study catchment areas. Two children with p\u22650.21; see p\u200a=\u200a0.60) and no children were considered to have jaundice.There were no significant differences in baseline demographic and anthropometric characteristics , vital signs , hematologic parameters , or blood glucose concentrations between the 91 children allocated to artemether-lumefantrine and the 97 who received artemisinin-naphthoquine in the detailed safety assessment (p\u200a=\u200a0.033). This symptom was mild and short-lived in all cases  . There were no significant changes in QTc in the artemether-lumefantrine group. Twenty-six of 102 children  with an interpretable electrocardiograph on day 2 had QTc>460 msec0.5. The longest QTc at this time point was 505 msec0.5, in an artemisinin-naphthoquine-treated child. No dysrhythmias, dyspnea, syncope, or other transient neurologic events were recorded during follow-up.Despite the technical challenges associated with performing electrocardiography in unwell children in a rural clinic, interpretable traces were obtained from between 68% and 88% of the children in each group at each time point. The QTThe only severe adverse event was considered non-drug related. A 48-mo-old child allocated to artemisinin-naphthoquine was hospitalized with, and treated successfully for, lobar pneumonia diagnosed on day 23 of follow-up. This patient was negative by both microscopy and PCR for malaria at the time of re-presentation.p\u200a=\u200a0.25) and parasitemia  by day 3  and 100%  in the artemether-lumefantrine and artemisinin-naphthoquine groups, respectively, on day 28 (a secondary endpoint), and 97.8%  and 100% , respectively, on day 42  in the artemether-lumefantrine group and 100%  in the artemisinin-naphthoquine group , but artemisinin-naphthoquine-treated patients were more likely to have gametocytemia between days 3 and 14 (see n\u200a=\u200a32 in the artemether-lumefantrine group and n\u200a=\u200a39 in the artemisinin-naphthoquine group), the equivalent odds ratios for gametocyte carriage on days 7 and 14 were 2.6 (95% CI 1.0\u20136.8) and 10.7 (95% CI 1.3\u201388.8).The percentage of children carrying d 14 see . The oddIn patients with vivax malaria, there was prompt gametocyte clearance in both groups by day 4, but, consistent with more frequent treatment failure in the artemether-lumefantrine group, a significant proportion of patients (40.0%) had become positive for gametocytes by day 42, while none of the artemisinin-naphthoquine-treated children were gametocytemic at that time see .p<0.001; see Hemoglobin concentrations in children treated for falciparum malaria were similar in the two treatment groups until day 42 after treatment, when the artemisinin-naphthoquine-treated children had a significantly higher mean concentration  show in vitro evidence of chloroquine resistance with near fixation of the resistance-associated pfcrt allele Despite potential pharmacokinetic/pharmacodynamic shortcomings, initial asexual parasite clearance was as rapid with artemisinin-naphthoquine as with artemether-lumefantrine. There is in vitro evidence of cross-resistance between chloroquine and naphthoquine in c prolongation between baseline and 4 h after the final dose in our artemisinin-naphthoquine-treated children. Since there were no significant between-group differences in pulse rate and fever clearance during initial monitoring, this QTc prolongation is consistent with a naphthoquine-specific effect rather than reflecting a disproportionately larger reduction in sympathetic activity with recovery in artemisinin-naphthoquine-treated children that could have independently promoted an increase in the QTc interval in this group Naphthoquine is a tetra-aminoquinoline drug, and our previous preliminary safety study of artemisinin-naphthoquine in older PNG children did not find evidence of aminoquinoline-specific side effects such as hearing loss, orthostatic hypotension, and hypoglycemia c prolongation. Chloroquine increases the QTc by a mean of up to 30 msec0.5 in healthy volunteers c prolongation similar to that in the present study for children treated with artemisinin-naphthoquine c prolongation observed in our artemisinin-naphthoquine-treated children had resolved by day 7, when plasma naphthoquine concentrations would still have been high Many aminoquinoline drugs cause QTc prolongation has been observed in young children presenting with untreated malaria c prolongation or cardiac arrhythmias , and that they should not be taken with other drugs that prolong the QTc interval This latter observation, amongst other considerations, suggests that the artemisinin-naphthoquine regimen used in the present study was not pro-arrhythmic. Relatively frequent QTPlasmodium species is found. It is possible that treatment responses in children with uncomplicated malaria who have been raised in areas with lower-level transmission may be attenuated because of their reduced or absent malarial immunity. Nevertheless, ACTs such as artemether-lumefantrine are considered efficacious where there is unstable transmission Our study had limitations. Baseline microscopy by study staff under field conditions led to 11% of the 267 randomized patients being included when they had sub-threshold parasitemia based on expert microscopy, while electrocardiography proved difficult in a significant minority of children. We did not directly observe all artemether-lumefantrine doses, but parents/guardians were instructed as to the importance of all doses given at home, adherence was assessed at the next follow-up visit, and day-7 plasma lumefantrine concentrations were consistent with full adherence in each case. The children screened and recruited were drawn from coastal PNG communities that are typical of those in Oceania where hyper- or holo-endemic transmission of multiple Plasmodium species. Artemisinin-naphthoquine may have advantages where P. vivax predominates. Although pharmaco-economic analyses were beyond the scope of the present study, it seems likely that the additional cost of three doses compared to the single dose currently recommended will be more than offset by the substantial reduction in post-treatment vivax malaria.The efficacy, tolerability, and safety of three daily doses of artemisinin-naphthoquine suggest that this regimen should be considered together with other currently available effective ACTs for treatment of uncomplicated malaria in PNG and similar epidemiologic settings with transmission of multiple Text S1Consort checklist for a randomized clinical trial of artemisinin-naphthoquine versus artemether-lumefantrine for uncomplicated malaria in Papua New Guinean children.(DOC)Click here for additional data file.Text S2Protocol for the clinical trial \u201cNovel artemisinin-based combination therapies for Papuan New Guinean children exposed to high transmission of multiple Plasmodium species.\u201d(DOC)Click here for additional data file."}
+{"text": "Plasmodium falciparum to the ACT justify the need for continued search for alternative anti-malarial drugs. The use of antibiotics with anti-malarial properties represents a potentially valuable chemotherapeutic option for the management of drug resistant infections. Thus, the intrinsic anti-malarial activity of the combination of clinical doses of rifampicin with amodiaquine and artemether was evaluated in an animal model using Plasmodium berghei.Artemisinin-based combination therapy (ACT) remains the most effective chemotherapeutic strategy in the management of malaria. However, reports of reduced susceptibility of in vivo was employed. The anti-malarial activity of standard doses of amodiaquine (AQ) with or without artemether (ART) and combined with varying doses of rifampicin (RIF 15\u00a0mg/kg or RIF 30\u00a0mg/kg body weight) was evaluated in 40 mice sub-divided into eight groups and inoculated intraperitoneally with 1\u2009\u00d7\u2009107 red blood cells infected with chloroquine-resistant P. berghei ANKA strain. There were two control groups of animals, one group received amodiaquine alone while the other group received saline. Parasiticidal activity and survival of the animals were assessed over 21\u00a0days.A modification of the suppressive tests Parasitaemia in the control animals peaked at 38% on day 9 and all animals died by day 10. The combination of amodiaquine with rifampicin 15\u00a0mg/kg body weight was the most effective of all the combinations and more efficacious than amodiaquine alone. The order of superiority of anti-malarial efficacy of the combinations was as follows; AQ\u2009+\u2009RIF 15\u2009>\u2009AQ\u2009>\u2009AQ\u2009+\u2009ART\u2009+\u2009RIF 30\u2009>\u2009AQ\u2009+\u2009ART\u2009+\u2009RIF 15\u2009>\u2009AQ\u2009+\u2009RIF 30.The combination of the clinical dose of rifampicin (15\u00a0mg/kg) with amodiaquine represents a potentially valuable treatment option in management of drug resistant malaria. In addition, the role of pharmacokinetic interaction in multiple drug therapy cannot be over-emphasized. Plasmodium falciparum malaria. Unfortunately, Plasmodium resistance to anti-malarial drugs continues to threaten effective control of malaria in endemic areas. Some evidence from in vitro monitoring over time in China and Viet Nam indicated increased IC-50, IC-90 or IC-99 values for artemisinins [in vitro susceptibility was observed from west to east [P. falciparum to artemisinin derivatives in patients in western Cambodia has also been documented [The World Health Organization recommends that artemisinin and its derivatives are combined with other anti-malarial drugs that have different mechanisms of action and longer half-lives in order to maximize the effectiveness of the artemisinins and to protect them from the development of resistance -3. Artemmisinins . Further to east . In a se to east . Reducedcumented . These rin vitro and in vivo[P. falciparum in vitro. The lower activity of the antibiotics has been attributed to a delayed effect of the drugs on the parasites, which becomes progressive with the duration of parasite-drug incubation [The use of antibiotics with anti-malarial activities represents a valuable option in malaria chemotherapeutic strategies. Several antibiotics including tetracycline, azithromycin, fluoroquinolones and rifampicin have been shown to possess anti-malarial activity d in vivo-15. Althd in vivo-19. In acubation ,20.Plasmodium vivax in humans [Plasmodium chabaudi in rodents and chloroquine resistant P. falciparum in vitro[P. falciparum infections [in vivo during malaria infection in an animal model using Plasmodium berghei.Rifampicin is an antitubercular drug with potent anti-malarial activity against n humans , Plasmod in vitro. Combinafections . Rifampifections  while arfections ,26. Indufections . The artfections  and thisRifampicin (RIF) was kindly provided by Bond Pharmaceuticals, Awe, Oyo State, Nigeria, and artemether and amodiaquine were obtained from the Walter Reed Army Institute for Research, USA. Stock solutions of the compounds were prepared in distilled water and stored at -20\u00b0C till required.The animals used in this study were male Swiss albino mice (6 \u2013 8\u00a0weeks old) weighing 18 \u2013 22 grams. The animals were obtained from the animal house of the Malaria Research Laboratories, Institute of Advanced Medical Research and Training (IMRAT), University of Ibadan, Ibadan. The mice were used in accordance with the NIH Guide for the care and use of laboratory animals, NIH publication , revised 1996.7 red blood cells infected with the CQ-resistant P. berghei ANKA strain. The infected animals were randomly divided into eight groups of five mice each and were treated by the oral route with rifampicin alone at 15 or 30\u00a0mg/kg body weight daily for seven days, amodiaquine 10\u00a0mg/kg body weight for three days\u2009+\u2009rifampicin 15 or 30\u00a0mg/kg daily for seven days or amodiaquine 10\u00a0mg/kg body weight for three days\u2009+\u2009artemether 8\u00a0mg/kg daily for three days\u2009+\u2009rifampicin or rifampicin 30\u00a0mg/kg body weight daily for seven days. Two controls groups were used, one treated with amodiaquine 10\u00a0mg/kg body weight given daily for three days while the second group of animals were treated with saline.A modification of the suppressive tests in vivo  was usedThe stock solutions of the drug samples were diluted to the desired final concentration with distilled water so that each animal received 200\u00a0\u03bcl at time of administration of each drug. Parasiticidal activity was assessed daily from day 3 post-infection till day 14, and then on day 21. Blood smears were prepared from the tail, methanol-fixed, stained with Giemsa and microscopically examined by determining parasitaemia in 1,000 erythrocytes. Mortality was monitored daily, until four weeks after infection. Inhibition of parasite growth in drug-treated group was calculated in relation to parasite growth in the non-treated control group. All compounds and combinations were tested in three independent experiments.t test was used to analyse the differences in mean parasitaemia level on days following treatment initiation, and analysis of variance between groups (ANOVA) was used to compare difference in percentage inhibition of parasite growth.Student-Parasitaemia in the untreated control animals ranged from 2.0% on day 3 post-infection to 38.5% on day 9 when it peaked. All animals in the control group died by day 10. Parasitaemia did not clear completely in animals that received AQ or RIF alone. There was however, a significant decrease in parasitaemia (54% to 86%) in the animals treated with AQ alone during the follow up days (days 4-9) compared with the untreated control animals (P\u2009<\u20090.05) , mean parasitaemia was significantly lower (P <0.05) compared with the group of animals that received AQ only Figure\u00a0. In thisParasitaemia in the group of animals treated with AQ\u2009+\u2009ART\u2009+\u2009RIF (15\u00a0mg/kg) or AQ\u2009+\u2009ART\u2009+\u2009RIF 30\u00a0mg/kg) significantly decreased by 75% to 96% or 76% to 94%, respectively, during follow up days 4 to 9 compared to the untreated controls (P\u2009<\u20090.05)  in the group of animals treated with the combination of AQ plus 15\u00a0mg/kg body weight RIF. This is the first report describing beneficial interaction between RIF and AQ. Rifampicin alone given at standard clinical doses (15-30\u00a0mg/kg) in this study exhibited slight anti-malarial activity against P. berghei, as no substantial drop in parasitaemia was observed as reported in previous studies [et al. reported a clear dose-response effect of rifampicin with substantial drop in peripheral parasitaemia of P. chabaudi within 24\u00a0hrs of treatment following the use of 100-200\u00a0mg/kg body weight rifampicin. This dose is six folds higher than the dose employed in the present study. In a study by Schmidt, doses of rifampicin up to 20\u00a0mg/kg failed to effect cures or prevent relapses when combined with chloroquine [This study describes beneficial interaction between amodiaquine and rifampicin in mice infected with chloroquine resistant strains of  studies . Strath oroquine .The beneficial interaction between amodiaquine and rifampicin observed in this study especially at doses of rifampicin used clinically in the management of tuberculosis is a welcome discovery as co-existence between TB and malaria leads to various concerns about their treatment. The suitability of rifampicin rather than tetracycline for the treatment of children might be of particular interest in the potential clinical use of the combination. The combination of amodiaquine with 15\u00a0mg/kg bodyweight of rifampicin in this study was superior to the combination with 30\u00a0mg/kg bodyweight of rifampicin in reducing peripheral parasitaemia and enhancing survival rate. The reason for this disparity is not clear but it may not be unconnected with the auto-inducing effects of rifampicin which may also be dose related. Rifampicin is a great CYP450 inducer and one of the most potent inducers of all drugs used clinically . Thus a et al. who reported unfavourable pharmacokinetic interaction between artemether, lumefantrine and rifampicin in healthy adults [P. berghei in the present study. Similarly, reports of pharmacokinetic interaction between rifampicin and quinine which adversely affects the efficacy of quinine in uncomplicated falciparum malaria have been documented [It would have been expected that the inclusion of artemether, a rapidly acting anti-malarial drug in the combination would produce a more significant effect in terms of enhanced parasite clearance and survival rate. Surprisingly, presence of artemether in the combination of amodiaquine with rifampicin did not produce a more superior anti-malarial effect over AQ\u2009+\u2009RIF 15 or AQ alone against the parasites. Superiority of the combination was as follows AQ\u2009+\u2009RIF 15\u2009>\u2009AQ\u2009>\u2009AQ\u2009+\u2009ART\u2009+\u2009RIF30\u2009>\u2009AQ\u2009+\u2009ART\u2009+\u2009RIF15\u2009>\u2009AQ\u2009+\u2009RIF 30. The potential for drug-drug interaction cannot be overemphasized during combination or multiple drug therapy as it may pose serious challenges to treatment outcome. The lack of the expected enhanced anti-malarial effects when artemether was included in the combination may be as a result of pharmacokinetic interaction. Rifampicin is a potent inducer of hepatic and intestinal CYP450 enzyme activity  and all artemisinin derivative compounds are metabolized by the CYP450 enzymes to the active dihydroartemisinin, also available as a drug itself . The arty adults . In thatcumented . Pharmaccumented ,36.Theoretically it would be expected that rifampicin would alter amodiaquine activity when co-administered because amodiaquine is metabolized by CYP2C8, an enzyme also inducible by rifampicin. However this was not the case, as amodiaquine activity was significantly enhanced in the presence of rifampicin 15\u00a0mg/kg by enhancing the rate of fall in parasitaemia and survival rate of the animals. The higher dose rifampicin (30\u00a0mg/kg) did not appear to induce metabolism of AQ either, as response of infection to AQ\u2009+\u2009RIF30 was similar to that of AQ alone. Rifampicin auto induction, which seems to be dose dependent appears to be a plausible explanation.P. berghei and provides a potentially useful chemotherapeutic alternative in the management of malaria in endemic areas. Detailed pharmacokinetic and toxicological studies on the interaction between amodiaquine and rifampicin are required prior to clinical application of the combination.In conclusion, this study reveals a beneficial effect of rifampicin in combination with amodiaquine against a chloroquine-resistant strain of The authors declare that they have no competing interests.JAB participated in developing research protocol, data collection, data analysis and manuscript preparation, OOA participated in protocol development, data analysis, and manuscript writing, OA participated in data analysis and manuscript preparation, CTH participated in study design, protocol development, and data interpretation, AS participated in study design. GOG was responsible for conceptualization of study, study design, data analysis and interpretation, and manuscript preparation. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum to available non-artemisinin antimalarial drugs, new and novel pharmaceuticals are needed. ARCO\u00ae is a new generation ACT, one of several artemisinin-based combinations developed in China to counter antimalarial drug resistance. ARCO\u00ae is a derivative of two independently developed antimalarials, artemisinin and naphthoquine phosphate, which were combined to form the artemisinin-naphthoquine combination. Both artemisinin and naphthoquine drugs have proven to be efficacious, safe and well tolerated as monotherapies. The artemisinin-naphthoquine combination offers a novel advantage over existing ACTs: it can be administered as a single oral dose (or a 1-day treatment). Several therapeutic studies conducted recently indicate that a single oral dose administration of artemisinin-naphthoquine combination is equally effective and safe as the 3-day treatment with artemether-lumefantrine combination and other existing ACTs. This would make ARCO\u00ae the next generation ACT for the treatment of uncomplicated falciparum malaria. With the rapidly spreading resistance of  Plasmodium falciparum, have been on the rise in the past decade, mainly because this species of malaria parasite have developed multiple strategies to deal with the parasiticidal actions of available non-artemisinin antimalarial drugs . (ii) \u00ae vs. artemether-lumefantrineARCO (AL). Two of these studies were conducted in children populations of Nigeria and Uganda, and one in an adult population with uncomplicated falciparum malaria in Uganda. For the children, the number of ARCO\u00ae tablets administered was based on the bodyweight (25/10 mg/kg artemisinin-naphthoquine combination) and for the adult population ARCO\u00ae tablets were administered as a single dose of eight tablets. The therapeutic responses were monitored for 28 days in the Nigerian study and 42 days in the Ugandan study. There was no significant difference in the efficacy and safety profiles in children in the two studies between the single dose ARCO\u00ae treatment and 6-dose regimen of AL at day 28 and day 42, respectively. Similar observations were made between the two treatments in the adult population in the Ugandan study at day 28 [Source: KPC archived data].(iii) \u00ae vs artemether-lumefantrine vs artesunate-amodiaquineARCO (three-arm study). This study was conducted in Nigeria in mixed population (children + adults) with uncomplicated falciparum malaria. ARCO\u00ae tablets were administered according to the dosing schedule described above. For AL, tablets were administered twice a day for three days based on bodyweight for children and predetermined number of tablets for adults while for AA, once a day treatment for 3 days based on bodyweight for children and predetermined number of combination tablets for adults. The therapeutic responses were monitored for 28 days. The study concluded that ARCO\u00ae and AA treatments were marginally better than artemether-lumefantrine in these settings [Source: KPC archived data]. (iv) \u00ae (2x/day in divided dosage) vs. artemether-lumefantrineARCO (AL). This study was undertaken in Ivory Cost, West Africa, in a mixed population (children and adults) with uncomplicated falciparum malaria. Drugs were administered as described above. The therapeutic responses were monitored for 28 days. There was no significant difference in the efficacy and safety of 1-day (2x/day in divided dosage) treatment with ARCO\u00ae and 3-day with AL (cure rate: ARCO\u00ae 100% vs. 98% AL) [(v)  98% AL) .\u00ae (single dose) vs ARCO\u00ae (2x/day divided doses)ARCO. This study was conducted in Benin  in a child population with uncomplicated falciparum malaria. Children received number of tablets based on bodyweight as a single dose versus same dose divided into two doses give 12 h apart. The therapeutic responses were monitored for 28 days. Therapeutic efficacy and safety were similar for both therapeutic dose regimens .(vi) \u00ae aloneARCO. Two studies were conducted in the adult populations with uncomplicated falciparum malaria: one in Nigeria and one in Myanmar [\u00ae in the respective country settings. The above information is summarized in (vii)  Myanmar , with noBecause the methodology of clinical and/or therapeutic assessment had not been prospectively standardized prior to the conduct of the studies, there existed substantial inter-study differences in defining, assessing, reporting, and classifying efficacy and adverse events. However, all studies were conducted in accordance with the World Health Organization (WHO) guidelines for antimalarial drug efficacy assessment . \u00ae treatment and other artemisinin-based combinations  in the rate of reduction of parasite biomass in the blood, except for the non-artemisinin-based combination [\u00ae, it would seem remarkable for a drug combination with shorter duration of treatment within the family of ACTs. With the exception of few cases, in all the studies fever was cleared within 24 h following treatment  course of ARCO\u00ae. The cure rates however, were similar in all treatments on day-28; extension of observations beyond 28 days did not result in better outcomes nor was there any significant differences in cure rates whether ARCO\u00ae was given as a single dose (8 tablets for adults or on a body-weight basis for children) or in two divided doses over 24 h (1-day treatment regime).All the studies demonstrated that ARCOThe safety data provided in relation to individual patients were primarily clinical. Where laboratory data was available, the laboratory evaluation schedules were not consistent between the studies, making comparative interpretation of safety data difficult. Because the trial methodology included in the pooled analysis had not been prospectively standardized, there existed substantial inter-trial differences in defining, assessing, reporting, and classifying adverse events. Furthermore, reliably distinguishing drug side effects from clinical symptoms of malaria infection is often difficult and much of the reporting is largely dependent upon a subjective assessment performed at the time of the event. The safety data obtained for this analysis were from individual patient data case record forms, which were archived at the Centre for clinical studies at Kunming Pharmaceutical Corporation. A total of 16 different adverse events, with varying frequencies and intensities, were recorded in the pooled analysis of 952 adult patients who received artemisinin-naphthoquine combination for uncomplicated malaria. The five most common adverse events were, in order of frequency, headache, nausea, vomiting, dizziness, and abdominal pain. However, it was difficult to discern which of the adverse events were malaria related and which were due to drug treatment because almost all of these events were reported during the first 24-hrs following the commencement of treatment. No adult or paediatric patients discontinued the treatment or any part of the treatment prematurely due to adverse experiences.\u00ae and extrapolation from this data to different populations and settings should be made with caution. Despite the methodological limitations of this analysis, the overall safety profile of ARCO\u00ae treatment in these series of studies appeared to be benign. The total incidence of drug-related adverse events considered by clinicians and/or principal investigators was estimated to be low (\u22645%). The majority of the adverse events reported has been of gastrointestinal-related in nature and were self-limiting. In general, safety profile of ARCO\u00ae treatment appears to be excellent. However, the data from this analysis do not suggest that there are no additional concerns about ARCO\u00ae use as a therapeutic agent in a wider context but pharmacovigilance is important for its continued deployment. A meaningful comparison of safety profiles between different treatments including artemisinin-based products is beyond the scope of this analysis. It should be noted however, that most of the safety data presented were derived from patients treated with ARCO\u00ae) is a new generation ACT. An analysis of the pooled data from several studies in different countries indicates that this combination is quite safe and equally effective (efficacious) as other ACTs recommended by World Health Organization (WHO) which include artemether-lumefantrine (AL), artesunate-amodiaquine (AS + AQ), artesunate-mefloquine (AS + MQ), artesunate plus sulphadoxine-pyrimethamine (AS + SP) and dihydro-artemisinin-piperaquine (DHA + PPQ) combinations for treatment of uncomplicated falciparum malaria [\u00ae will ever be considered for malaria treatment one day (from a drug resistance point of view) on a wide scale remains to be debated but these series of studies undertaken in different countries provide additional information on the potency and/or efficacy of ARCO\u00ae in different environmental and epidemiological conditions/settings. These and other studies further affirms that ACTs including ARCO\u00ae, represent a promising approach to drug-resistant malaria treatment [\u00ae easily develops resistance if given as single dose. Resistance to the artemisinin\u2019s partner medicines may compromise the efficacy of the ACT. Therefore, the efficacy of ARCO\u00ae needs careful monitoring after its introduction and widescale use, particularly in multidrug resistant areas. At present however, there is seemingly no doubt that a single dose or a 1-day treatment with the combined artemisinin and naphthoquine for uncomplicated falciparum malaria provided high efficacy in adult populations, as well as in the trialed children population. Artemisinin-naphthoquine combination  as a new generation ACT. The findings are consistent with observations made with other ACTs [\u00ae, may be an advantage in that the health workers can administer the drug as directly observed treatment. With the shorter treatment duration, the potential to be deployed for home management of malaria at community levels needs to be evaluated. However, the impact of shorter dosing regimens on morbidity and mortality due to falciparum malaria remains to be ascertained.The pooled data from multi-national therapeutic studies presented here provides scientific evidence for clinical use of ARCOher ACTs ,36,37,38her ACTs ,42,43. T\u00ae versus multiple doses DHA-PPQ on vivax malaria; both treatments provided equivalent cure rates  on 42-day follow-up. Data is limited on the role of ARCO\u00ae in other malaria parasite species including Plasmodium vivax malaria. Only one study in these series compared the efficacy of a single dose ARCO\u00ae) compares favourably with existing ACTs, so may be the next new generation ACT. Review of available data concludes that, from the clinical efficacy and safety point of view, ARCO\u00ae offers no advantages over existing ACTs except for the fact that this combination may be given as a single dose than three daily doses or three twice daily doses. It will be more useful particularly in situations where noncompliance to the currently practiced 3-day course of ACTs is common. Artemisinin-naphthoquine combination  or scaling-down of number of tablets on body-weight basis must be supported and guided by pharmacokinetic data to avoid sub-therapeutic levels. Substantially little pharmacokinetic data is available to support and guide different dosage schedules, so further studies are needed in these areas to optimize therapy for different dosing schedules.ARCO"}
+{"text": "The long half-lives of malaria \u2018partner\u2019 drugs are a potent force selecting for drug resistance. Clinical trials can quantify this effect by estimating a window of selection (WoS), defined as the amount of time post-treatment when drug levels are sufficiently high that resistant parasites can re-establish an infection while preventing drug-sensitive parasites from establishing viable infections.The ability of clinical data to accurately estimate the true WoS was investigated using standard pharmacokinetic\u2013pharmacodynamic models for three widely used malaria drugs: artemether\u2013lumefantrine (AR-LF), artesunate\u2013mefloquine (AS-MQ) and dihydroartemisinin\u2013piperaquine (DHA-PPQ). Estimates of the clinical WoS either (1) ignored all new infections occurring after the 63-day follow-up period, as is currently done in clinical trials, or, (2) recognized that all individuals would eventually be re-infected and arbitrarily assigned them a new infection day.The results suggest current methods of estimating the clinical WoS underestimate the true WoS by as much as 9\u00a0days for AR-LF, 33\u00a0days for AS-MQ and 7\u00a0days for DHA-PPQ. The new method of estimating clinical WoS  was significantly better at estimating the true WoS for AR-LF and AS-MQ.Previous studies, based on clinically observed WoS, have probably underestimated the \u2018true\u2019 WoS and hence the role of drugs with long half-lives in driving resistance. This has important policy implications: high levels of drug use are inevitable in mass drug administration programmes and intermittent preventative treatment programmes and the analysis herein suggests these policies will be far more potent drivers of resistance than previously thought.The online version of this article (doi:10.1186/s12936-015-0810-4) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria and now widely deployed in most endemic countries [Artemisinin combination therapy (ACT) is effective, first-line treatment for uncomplicated ountries . The artountries , 3. Howeountries \u20136. This The putative consequences of long half-life partner drugs driving resistance enters debates on the politics of anti-malarial drug deployment. Mass drug deployment (MDA) policies were widely used in the 1950s and 1960s (reviewed by Von Seidlein and Greenwood ), but fedhfr108 mutation could be observed in patients 15\u00a0days after treatment with sulfadoxine-pyrimethamine (SP), whereas wild-type infections were only observed after 50\u00a0days, thus implying a WoS of 35\u00a0days. Similarly, Sisowath et al. [pfmdr1 D1246Y mutation after lumefantrine treatment. Routine genotyping in clinical trials means such data are readily available and have been used previously to estimate WoS [dhfr108 mutation. This implies that selection for resistance via WoS may be widespread and intense.The genetic process whereby parasites evolve increasing tolerance to the partner drug is usually quantified as a window of selection (WoS). As a specific example, Watkins and Mosobo  noted pamate WoS \u201318. This5 parasites that emerge from the liver are below patency. It has, therefore, been widely assumed that the clinical WoS reflects the true window (op cit). However, it is not clear how well the clinical WoS estimates the true WoS and there are several plausible reasons why they may be poor estimators, discussed further in Fig.\u00a0These WoS estimates, obtained from clinical observations, are widely cited and refered to here as \u2018clinical\u2019 WoS to denote their origin in clinical observations. In fact, the \u2018true\u2019 WoS is the period during which infections bearing the mutation can emerge from the liver (assuming the drugs do not kill the parasites while in the liver stage) and survive to produce a viable infection, while sensitive parasites are killed by residual drug concentrations . UnfortuThe mechanistic PK/PD model of Winter and Hastings  was usedMass drug treatment in human populations was simulated assuming 5,000 simulated patients were each given the same ACT; all patients were treated at day 0 and the simulations run for 365\u00a0days; 63\u00a0days is the recommended period of follow-up in clinical trials but the 365-day \u2018follow-up\u2019 in the simulations reveal how finishing patient observations at 63\u00a0days may affect the results. The focus was solely on increasing resistance to the partner drugs and this was incorporated into simulations by increasing the IC50 of parasites to the partner drug; hence \u2018genotypes\u2019 in this study refers to parasite genotypes with different values of IC50 to the partner drug. Any parasites present on the day of treatment  were ignored on the assumption that the ACT is clinically effective and would eliminate them.The principle underlying the analyses is simple. First, the probability that parasites with different drug-resistant \u2018genotypes\u2019 emerging from the liver could survive residual drug levels on any given day post-treatment was established; this estimated the \u2018true\u2019 WoS  parameters were randomly assigned using the means and distributions in Additional file The question then arises as to how to translate these distributions of the earliest emerging infections into a strict definition of the true WoS. It would be inappropriate to define the start of the WoS as the earliest possible emergence day in the distribution, because this will depend on chance and on sample size. Therefore the true WoS was defined as the difference between the day post-treatment on which 50% of clones could survive residual drugs for any IC50 \u2018genotype\u2019, i.e., the median of the distribution  of the patency data to reduce the element of chance. For example, the data of Sisowath et al.  on Fig.\u00a0Method 2 is analogous to the first and compares centile scores of each allele to estimate the clinical WoS. The one difference is that Method 2 recognizes that each patient would eventually have become re-infected with the allele if followed up for a sufficiently long time and hence that the re-infection data are censored. Patients with no re-infections in the 63-day follow-up period  were arbitrarily scored as having re-infections occurring on day 70. The comparison to obtain WoS relies on centiles (not averages) so it is immaterial which day after 63 is arbitrarily chosen provided the number of censored individuals is not above 50% 1). The t10 and 1012 parasites (chosen from a uniform distribution) from a single clone were present at the time of treatment, the outcome  was simulated and the probability of each IC50 \u2018genotype\u2019 surviving direct treatment determined.The probability of surviving direct treatment (POST) with the partner drug monotherapy was also recorded for the 5,000 patients to provide a baseline probability of successful treatment, against which the survival probabilities of later emerging parasites could be compared. It was assumed that between 1010\u20131012 parasites) than the newly emerging infections on day 0 (105 parasites). Secondly, POST is calculated for the partner drug monotherapy, whereas emergence on day 0 encounters both partner drug and an artemisinin derivative.The \u2018true\u2019 WoS are shown on Fig.\u00a0The presence or absence of a true WoS between the different genotypes can be determined from Fig.\u00a0The pattern of treatment outcome following AR-LF Fig.\u00a0a was broThe clinical WoS were formally defined as the differences between genotypes based on the tenth, 25th and 50th  centile values of patient data and are given on Additional file The key research question addressed here is whether the clinical WoS reliably estimates the \u2018true\u2019 WoS; the estimates are given on Table\u00a0Calculating the clinical WoS using Method 2 generally provided better estimates of the true WoS Table\u00a0 but overIn summary, Table\u00a0These results were determined assuming patients acquire 16 new infections per year . There is currently interest in the improved diagnostics of malaria treatment, primarily to ensure treatment is restricted to confirmed malaria cases, so treatment is appropriate to the underlying clinical condition . A seconThis study was designed to identify the best methods to analyse clinical data that generate results consistent with the \u2018true\u2019 WoS. The resulting recommendations are as follows. First, if possible, a PK/PD analysis similar to that shown in Fig."}
+{"text": "Plasmodium falciparum malaria in infants and children weighing 5 to <35\u00a0kg. However, there are no clinical studies with artemisinin-based combination therapy in infants <5\u00a0kg.Artemether-lumefantrine (AL) dispersible formulation was developed for the treatment of uncomplicated This multicentre, open-label, single-arm study evaluated the efficacy, safety and pharmacokinetics of AL dispersible in infants aged >28\u00a0days and <5\u00a0kg of body weight, who were treated with one AL dispersible tablet (20\u00a0mg artemether/120\u00a0mg lumefantrine) given twice-daily for three days and followed up for six weeks (core follow-up) and at 12\u00a0months of age (long-term follow-up).A total of 20 patients were enrolled and completed the six-week core study follow-up. In the per protocol population, PCR-corrected cure rate at days 28 and 42 was 100% . AL dispersible was well tolerated with reported adverse events of mild to moderate severity. Pharmacokinetic data showed that lumefantrine levels were similar, however, artemether and dihydroartemisinin levels were on average two- to three-fold greater than historical values in infants and children \u22655\u00a0kg.P. falciparum malaria, but artemether and dihydroartemisinin exposures could not be supported by the preclinical safety margins for neurotoxicity. Hence, dosing recommendations cannot be made in infants <5\u00a0kg as implications for toxicity are unknown.A three-day regimen of AL dispersible formulation was efficacious and generally well tolerated in infants weighing <5\u00a0kg with uncomplicated NCT01619878.Clinicaltrials.gov  Plasmodium falciparum malaria in adults and children weighing \u22655\u00a0kg [Artemether-lumefantrine ((AL) Coartem\u00ae/Riamet\u00ae, Novartis Pharma AG, Basel, Switzerland) is the first fixed-dose artemisinin-based combination therapy (ACT) to be prequalified by the World Health Organization for the treatment of uncomplicated ng \u22655\u00a0kg -3. The png \u22655\u00a0kg . The effng \u22655\u00a0kg ,7.P. falciparum malaria. This study aimed to evaluate the efficacy, safety and pharmacokinetics (PK) of AL dispersible following treatment with a three-day regimen in infants <5\u00a0kg with uncomplicated P. falciparum malaria.While much is known about malaria in infants and children weighing \u22655\u00a0kg, there are no clinical trials or national guidelines for treatment of infants <5\u00a0kg. The current treatment for this subgroup is oral quinine, which is associated with a poor safety profile . Other aP. falciparum malaria, including asexual P. falciparum parasitaemia >1,000 and <100,000 parasites/\u03bcL, were included. Patients were recruited from three healthcare facilities in two countries in Africa, one in Benin and two in Burkina Faso. Patients with severe malaria or general danger signs  based on the Integrated Management for Childhood Illnesses (IMCI) criteria for sick infants were excluded from the study. The trial protocol was approved by an Independent Ethics Committee (IEC) and the Institutional Review Board (IRB) at each study centre . Signed informed consent was obtained from parent or legal guardian before any study-related procedure. The study was conducted in accordance with the International Conference on Harmonization Guidelines for Good Clinical Practice, Declaration of Helsinki, and local regulations of each participating country. This trial is registered with with ClinicalTrials.gov number NCT01619878.Male and female infants aged >28 days and weighing <5 kg, with microscopically confirmed acute uncomplicated This multicentre, open-label, single-arm study was planned to be conducted in two sequential cohorts of infants <5\u00a0kg; cohort 1 in infants >28\u00a0days old and cohort 2 in neonates \u226428\u00a0days of age. Based on an interim analysis of cohort 1 results after the six-week core follow-up period, cohort 2 was not initiated as recommended by a data monitoring committee. The study was terminated after all patients from cohort 1 completed their long-term follow-up.Patients were admitted to the hospital during a three-day treatment phase and followed up until day 42 (week 6) after treatment, with regular assessments for efficacy and safety during the six weeks. A long-term follow-up was conducted at 12\u00a0months of age to assess neurodevelopmental status. Patients in cohort 1 received directly observed treatment of AL dispersible tablet (20\u00a0mg artemether/120\u00a0mg lumefantrine) following a regimen of one dispersible tablet twice daily for three consecutive days. The consumption of food or drink (mother\u2019s or formula milk) was recommended after dose to enhance lumefantrine absorption. In the event of vomiting within an hour of dosing, a repeat dose was administered and a maximum of two doses were replaced throughout the entire treatment phase.Patients who developed severe malaria or early treatment failure signs were to receive rescue therapy as per local treatment guidelines. Presence of parasitaemia on day 7, irrespective of clinical state, and vomiting of replacement dose within one hour warranted rescue therapy.Clinical and parasitological examinations were performed on days 0, 1, 2, 3, 7, 28, and 42. Giemsa-stained thick and thin smears were examined locally at each visit using a light binocular microscope fitted with an oil immersion lens for assessment of parasitaemia. Blood smears were also systematically read at a central facility. Blood samples for molecular diagnostics (PCR-based methods) were taken at baseline, days 14, 28 and 42, and at any unscheduled visit when reappearance of parasitaemia was confirmed microscopically.Physical examination was performed on days 2, 3, 4, 7, 14, 28, and 42, or at the time of withdrawal. Vital signs including systolic and diastolic blood pressure , temperature, and pulse rate were measured. Blood samples were collected and analysed locally for haematology and blood chemistry tests.Safety monitoring included recording of all adverse events (AEs) and serious AEs (SAEs). Neurodevelopmental status was assessed using an age-appropriate scale  at 12\u00a0months of age .Blood samples (250\u00a0\u03bcL each) were drawn at predefined timepoints: one and two hours after the first AL dose for measurement of artemether and its active metabolite dihydroartemisinin (DHA) in plasma; six hours after the fifth and sixth doses, 24\u00a0hours after the sixth dose, and on day 7 for lumefantrine measurement. Artemether and DHA were analysed using reversed-phase HPLC with MS/MS detection and the limit of quantification (LOQ) was 5\u00a0ng/mL. Lumefantrine was determined using reversed-phase HPLC with MS/MS detection and the LOQ was 50\u00a0ng/L.It was aimed to enroll 40 patients to obtain a sample size of at least 30 patients evaluable for the primary endpoint . Based on an assumed PCR-corrected day 28 cure rate of 95% and a sample size of 30 evaluable patients, the study had 80% power to result in an estimate of cure rate associated with a lower exact 95% confidence limit >77%. For secondary efficacy variables, Exact Pearson-Clopper two-sided 95% confidence limits were constructed for dichotomous outcome variables . Incidence of gametocyte carriage over time was summarized by baseline gametocyte carriage status.Descriptive summary statistics were presented for values and change from baseline to each follow-up. For PK exposure to artemether, DHA and lumefantrine, descriptive statistics of concentrations at discrete timepoints were presented. However, no formal PK analysis was performed due to limited number of sample points.The infant neurodevelopmental scale included the total scores for motor milestones (section A), coordination (section B), tone (section C), behaviours (section D), and the sum of all sections together. All patients who received at least one dose of the study drug and had confirmed falciparum malaria at baseline were included in the full analysis set (FAS). The evaluable patient set (EPS) comprised patients from the FAS who had a known 28-day cure status, i.e., those who completed the study at least until day 28, or were considered failures before day 28 for reasons other than premature withdrawal. Patients from the EPS weighing <5\u00a0kg at baseline, who took at least five of six doses of the study drug, had baseline asexual parasite counts >1,000 and <100,000 parasites/\u03bcL based on central microscopy reading, and did not take any other medication with anti-malarial effect (excluding rescue medication) up to day 28 were included in the per protocol set (PPS).PCR-corrected parasitological cure rate at day 28 was the primary efficacy outcome defined as the proportion of patients clearing asexual parasites within seven days of initiating the study treatment without recrudescence before or at day 28, corrected for re-infection using PCR. The secondary efficacy outcomes included PCR-corrected parasitological cure rates at days 14 and 42 and uncorrected cure rates at days 3, 7, 14, 28, and 42.Artemether, DHA and lumefantrine concentrations were determined at discrete timepoints during and after treatment with AL dispersible. Safety parameters included incidence of AEs, SAEs and routine safety laboratory assessments.A total of 20 infants were recruited into the study and completed the six-week core follow-up. One patient discontinued study treatment because of vomiting but remained in the study. No patients received rescue therapy. A total of 17 patients completed the long-term follow-up visit at 12\u00a0months of age  was 82 37-214) days and median body weight (range) was 4.8 (2.6-4.9) kg. There was an equal number of male and female patients, and all patients were of African origin. The patients\u2019 demographic and baseline characteristics are shown in Table\u00a04 days anPlasmodium ovale in addition to P. falciparum at baseline, which persisted up to 72\u00a0hours and cleared. AL dispersible rapidly cleared fever with a mean time to fever clearance of 4.0\u2009\u00b1\u20096.4 and 4.1\u2009\u00b1\u20097.0\u00a0hours in the FAS and EPS/PPS populations, respectively. Complete (100%) asexual parasite clearance was achieved by 48\u00a0hours : 79.4-100), whereas it was 80% for the FAS population (95% CI: 56.3-94.3). Four patients did not have a microscopy reading after day 7 and were hence conservatively classified as treatment failures. The uncorrected cure rate on day 14 was also 100%  in the EPS and PPS populations. However, the uncorrected cure rates were lower on days 28 and 42 in all populations  during the core follow-up period. The most frequently reported AEs were new episodes of uncomplicated malaria  reported in 55% of patients during the follow-up period, followed by anaemia  and bronchitis (30%). Pyrexia, a manifestation of malaria, was also reported as an AE. Vomiting was reported as a drug-related AE in four (20%) patients  was 27.1\u2009\u00b1\u20094.5, coordination (Section B) 24.8\u2009\u00b1\u20094.6, tone (Section C) 16.6\u2009\u00b1\u20092.6, and behaviour (Section D) 14.4\u2009\u00b1\u20091.2. The overall total score mean\u2009\u00b1\u2009SD  was 82.8\u2009\u00b1\u20099.4. The scores were within the normal range in all four domains of the scale.The mean artemether and DHA concentrations at one and two hours post-first dose were two- to three-fold greater compared with historical data in infants/children \u22655\u00a0kg Table\u00a0. Mean luIn the current study that evaluated the efficacy, safety and PK of AL dispersible in infants aged >28\u00a0days and weighing <5\u00a0kg, AL dispersible was efficacious for the treatment of uncomplicated falciparum malaria. The PCR-corrected cure rate (100%) on days 28 and 42 for the EPS and PPS populations was slightly higher than earlier reports in infants and children \u22655\u00a0kg . The uncAL dispersible was well tolerated with no new AEs reported. Except for one patient who developed cerebral malaria and anaemia in the long-term follow-up, all AEs were mild to moderate in intensity and the most commonly reported AEs were indicative of signs or symptoms of malaria, in line with previous reports in older infants and children ,12. As eArtemether and DHA concentrations, on average, were found to be two- to three-fold greater in infants <5\u00a0kg as compared with historical levels  in olderArtemether exposure was consistently high in infants <5\u00a0kg and aged less than three months, and infants with the lowest body weight and age had the highest exposure. Artemether undergoes a high first pass metabolism and is predominantly metabolized by the CYP3A4 enzyme . CYP3A4 DHA concentrations also increased, though to a lesser extent than artemether. This could result from the different metabolic pathways and the relatively lower first pass effect of DHA. DHA undergoes metabolism by glucuronidation, predominantly by UGT1A9 and UGT2B7, which are mostly hepatic. UGTs have decreased activity in newborns and young children as compared with adolescents and adults . In neonGiven the known neurotoxic effects of artemisinins in animal studies -20, the max and AUC) and greater systemic toxicity with decreasing age of rat pups after artemether oral administration were reported [Higher exposures  systemic exposures were two- to three-fold greater for patients in this study as compared to infants and children \u22655\u00a0kg. Although no neurotoxicity has been reported after oral doses, implications for potential toxicity in infants and neonates are unknown. Therefore, based on the results of the current study, clear dosing recommendations cannot be made in infants <5\u00a0kg. Furthermore, the findings from this study underscore the need for detailed characterization of the pharmacokinetic/pharmacodynamic and safety profile of ACT and other anti-malarial drugs for use in this population to determine optimal doses rather than relying on empirical doses."}
+{"text": "Dihydroartemisinin-piperaquine (DHA-PQ) is one of five WHO recommended artemisinin combination therapy (ACT) for the treatment of uncomplicated malaria. However, little was known on its post-registration safety and effectiveness in sub-Saharan Africa. DHA-PQ provides a long post-treatment prophylactic effect against re-infection; however, new infections have been reported within a few weeks of treatment, especially in children. This paper reports the clinical outcomes following administration of DHQ-PQ in real-life conditions in public health facilities in Burkina Faso, Ghana, Mozambique, and Tanzania for the treatment of confirmed uncomplicated malaria.An observational, non-comparative, longitudinal study was conducted on 10,591 patients with confirmed uncomplicated malaria visiting public health facilities within seven health and demographic surveillance system sites in four African countries  between September 2013 and April 2014. Patients were treated with DHA-PQ based on body weight and followed up for 28\u00a0days to assess the clinical outcome. A nested cohort of 1002 was intensely followed up. Clinical outcome was assessed using the proportion of patients who reported signs and symptoms of malaria after completing 3\u00a0days of treatment.A total of 11,097 patients were screened with 11,017 enrolled, 94 were lost to follow-up, 332 withdrew and 10,591 (96.1\u00a0%) patients aged 6\u00a0months\u201385\u00a0years met protocol requirements for analysis. Females were 52.8 and 48.5\u00a0% were\u00a0<5\u00a0years of age. Malaria was diagnosed by microscopy and rapid diagnostic test in 69.8\u00a0% and 29.9\u00a0%, respectively. At day 28, the unadjusted risk of recurrent symptomatic parasitaemia was 0.5\u00a0% . Most of the recurrent symptomatic malaria patients (76\u00a0%) were children\u00a0<5\u00a0years. The mean haemoglobin level decreased from 10.6\u00a0g/dl on day 1 to 10.2\u00a0g/dl on day 7. There was no significant renal impairment in the nested cohort during the first 7\u00a0days of follow-up with minimal non-clinically significant changes noted in the liver enzymes.DHA-PQ was effective and well tolerated in the treatment of uncomplicated malaria and provides an excellent alternative first-line ACT in sub-Saharan Africa. Plasmodium falciparum is endemic as at 2013 . The exclusion criteria included: known allergy to artemisinin or to PQ; history of taking a DHA-PQ dose in the previous 4\u00a0weeks, known pregnancy, lactating women, complicated malaria, history of taking medicinal products that are known to prolong the corrected QT (QTc) interval on echocardiogram (ECG) , and family history of sudden unexplained death or personal/family history of predisposing cardiac conditions for arrhythmia/QT prolongation. Patients or their guardians were encouraged to comply with all scheduled follow-up visits.P. falciparum, 92.9\u201395.5\u00a0% for non-P. falciparum and specificity of 97.5\u201399.5\u00a0%.Patients were recruited into two groups, the main and nested cohort with no special criteria apart from the stated inclusion and exclusion criteria. Sites determined the cohort criteria based on their follow-up plan. Each eligible patient had a detailed clinical assessment including past medical and drug history after a confirmed parasitological (microscopy and RDT) diagnosis. Capillary blood sample was used in preparing thick blood smear and parasite density calculated per 200 leucocytes. The sites used different HRP2-based RDTs with sensitivity ranging from 90.5\u201399.7\u00a0% for A detailed laboratory assessment beside a prerequisite confirmed microscopic parasitaemia at baseline was carried out for the nested cohort. Investigations carried out were haematology , biochemistry , and plasma concentration of PQ as well as triplicate baseline ECG before day 1 drug administration. On each visit, 5\u00a0ml of venous blood was used for the investigations. Different types of Sysmex  and ABX micros B were used for haematology at the various sites while Selectra Junior, Humalyzer primus with Integra 400 plus and Vitalab flexor E were used for biochemistry analysis of patient samples.\u00ae (DHA-PQ) was administered once daily based on body weight for 3\u00a0days. Two concentrations were used: 20/160 and 40/320\u00a0mg of DHA and PQ, respectively, as shown in Table\u00a0EurartesimPatients were followed up for 28\u00a0days with active follow-up on day 5 (\u00b12\u00a0days) and day 28 by telephone or physical contact to document recovery status and AEs. Compliance to treatment was assessed in the main group by asking if patient adhered to the study prescription on day 5 (\u00b12\u00a0days). The nested cohort was additionally followed on days 2, 3 and 7. Day 2 visit was at home, for DOT of the second dose of the study medication, monitor clinical status and record any AEs. Patients were seen at the health facilities on day 3 for treatment completion, ECG recording (single pre and 3- to 4-hour triplicate post dosage) and then repeat haematology, biochemistry and plasma PQ concentration. These investigations were repeated on day 7 with a single ECG recording. Each facility visit included detailed clinical workup, screening for AEs and appropriate case report forms were completed. Patients were also encouraged to report any AEs experienced from day 1 to day 28 by telephone or visit to the health facility (unscheduled visit). Patients with signs and symptoms of malaria on all unscheduled visits were re-tested for malaria and managed appropriately as per national guidelines.The primary outcome of interest was adequate clinical response at day 28, defined as absence of clinical signs and symptoms for malaria at day 28. Treatment failure was defined as unresolved symptoms and signs and parasitaemia within 2\u00a0weeks of full treatment or recurrence of fever and parasitaemia after 2\u00a0weeks of full treatment . The def\u00ae (version 11.2). Descriptive analysis of all data recorded at study entry was carried out to characterize the population studied. The main outcome of interest, frequencies and respective percentages were calculated. Proportions, means and standard deviations were used in estimating the secondary outcomes. Means were estimated for other continuous independent variables. The parasite densities were grouped as\u00a0<500 parasites/\u00b5L, 500 to\u00a0<5000 parasites/\u00b5L and 5000 parasites/\u00b5L and above.Data were double-entered using Open Clinica software from all sites. Statistical analyses were performed using the software package STATAThe Institutional Ethics Committee of all seven HDSS sites involved in the trial and the Independent National Ethics Review Committees in these four countries approved the protocol. The protocol and case report forms are available and annexed as supporting documents. The study was conducted in accordance with Good Clinical Practice and the Helsinki declaration for biomedical research on human subjects. The Safety Committee, Central Data Monitoring Team and Independent Monitors provided oversight of study conduct. The study was registered with Clinicaltrials.gov; Trial registration number NCT02199951.A total of 10,591 participants successfully completed the study and were included in the analysis Fig.\u00a0.Fig.\u00a01StThe mean weight and height for the main group were 26.0\u00a0kg and 117.2\u00a0cm, and that for the nested cohort were 28.7\u00a0kg and 125.5\u00a0cm, respectively. Weight and height measurements were taken on day 1. The mean temperature for both cohorts on day 1 was 37.4\u00a0\u00b0C and declined to 36.6\u00a0\u00b0C on day 7 for the nested cohort. Out of the 1002 patients in the nested cohort, 441 had temperature\u00a0\u226537.5\u00a0\u00b0C on day 1. This decreased to 35 and 39 patients on day 3 and day 7, respectively. Forty-three per cent (4104) of the main group had fever on day 1. With the exception of blood pressure, all other vital signs decreased with age and no significant change was recorded among the nested cohort within the first 7\u00a0days of follow-up.P. falciparum cases. There were 94 cases of mixed infections from the microscopic findings: 90 of P. falciparum and P. malariae and four of P. falciparum and P. ovale with no case of P. vivax reported. Gametocyte carriage was 2.7\u00a0% (199/7260) and 11.1\u00a0% (13/117) for P. falciparum and P. malariae cases, respectively, by microscopy. Majority of patients had parasitaemia\u00a0<5000/\u00b5L  and 29.9\u00a0% (3170) patients, respectively, with 30 patients diagnosed clinically. The RDTs were 91 and 29The mean Hb of the nested cohort decreased from 10.6\u00a0g/dl on day 1 to 10.2\u00a0g/dl on day 7, however, it increased by 0.2\u00a0g/dl from day 3 to day 7  from day 4 to day 28. Fever, cough, headache, abdominal pains, and anorexia were the common symptoms presented on these visits . Of these, 47\u00a0% (24/51) were seen within 2\u00a0weeks of treatment (4\u201314\u00a0days) (median 7.7\u00a0days) and 53\u00a0% (27/51) after 2\u00a0weeks of treatment (15\u201328\u00a0days) (median 21\u00a0days). Most patients 76\u00a0% (39/51) with recurrent symptomatic parasitaemia were\u00a0<5\u00a0years. Navrongo, Nouna and Rufiji sites recorded 80\u00a0%  of these cases as shown in  administration in \u2018real-life clinical situations\u2019 at public health facilities for the treatment of confirmed uncomplicated malaria, WHO acceptable (>95\u00a0%) adequate clinical response with very low unadjusted rate of recurrent symptomatic parasitaemia of 0.5\u00a0% by day 28 was observed. This rate is lower than previously observed in more controlled studies in Southeast Asia  and sub-This study found that the majority of the recurrent symptomatic parasitaemia occurred almost in equal proportions within 14\u00a0days and after 14\u00a0days\u2019 post-treatment. This is in contrast to the review of 39 trials on ACT in the WHO 2010 treatment guidelines, which had enrolled 6124 participants. No treatment failure was recorded within two weeks in 32 of the trials. In the remaining seven trials, failure rates at day 14 ranged from 1\u20137\u00a0%, highlighting unusual rate of treatment failure within 2\u00a0weeks of treatment \u20137 and reThe majority of treatment failure cases were found in the\u00a0<5 age group corroborating earlier reports . In younOf the 51 patients who tested positive during the unscheduled visit, 22\u00a0% (11/51) were not treated with an anti-malarial contrary to the WHO malaria treatment guideline  leading The mean Hb outcome on day 7 was lower than the baseline level. This confirms similar results by Tjitra et al.  and OlliThis is the largest phase IV study in sub-Saharan Africa on DHA-PQ for the treatment of uncomplicated malaria, showing that DHA-PQ is very effective and tolerable. DHA-PQ should be an appropriate alternative to other first-line ACT in sub-Saharan Africa and its deployment will reduce drug pressure on the two widely used ACT (AS/AQ and AL)."}
+{"text": "Plasmodium vivax malaria is an important public health issue in the Amazon region, and it accounts for approximately 84\u00a0% of cases of the disease. Migration across the border between Brazil and French Guiana contributes to the maintenance of the disease. The aim of this study was to evaluate the therapeutic and parasitological responses of patients with P. vivax malaria treated with chloroquine and primaquine in the socio-environmental context of cross-border interactions between Brazil and French Guiana. The factors controlled were diagnostic agreement, adherence, adjustment of primaquine doses for patient weight, and quality of the drugs used.P. vivax treated with chloroquine  and primaquine for 7\u00a0days, who were followed for 28\u00a0days. The primaquine doses were adjusted for the patients\u2019 weight. A number of factors were determined: epidemiological characteristics, origin of patients, signs and symptoms, initial parasitaemia and parasitaemia clearance time, blood concentrations of chloroquine and primaquine, quality of anti-malarial drugs and diagnostic agreement.A prospective study was conducted in 2011 with 103 individuals aged 10\u201360\u00a0years with a positive diagnosis of Ninety-five patients were followed for 28\u00a0days. There was a 100\u00a0% agreement in microscopic diagnosis between field laboratory and reference centre. The adhesion to the treatment was 100\u00a0%. Of these patients, 32.6\u00a0% received a weight-adjusted dose of primaquine. The chloroquine and primaquine tablets were consistent with the optimal quality limits for human consumption. The investigated patients achieved optimal blood exposure to anti-malarial drugs. The parasitological and therapeutic response was adequate in 99.0\u00a0% of cases.P. vivax malaria using chloroquine combined with primaquine remains effective, when external factors are controlled, such as the quality of anti-malarial drugs, the adhesion to the treatment prescribed, the correct diagnostic and the adjustment of primaquine dose for patient body weight.In the municipality of Oiapoque, the therapeutic regime used for the treatment of  Plasmodium vivax malaria is an important public health issue in the Amazon region, and it accounts for approximately 84\u00a0% of malaria cases .The study was conducted in the municipality of Oiapoque-AP in the Amazon region of far northern Brazil. Oiapoque shares an international border with French Guiana. Oiapoque is situed at sea level and has an area of 22.625 KmIndividuals of both genders aged 16\u201360\u00a0years who spontaneously searched for public health care services in Oiapoque with signs and symptoms suggestive of malaria, from August to December 2011, and with a positive thick blood smear for vivax malaria were included in the study. Individuals with chronic and infectious concomitant disease, patients with mixed, severe or complicated malaria, pregnant women, indigenous people, patients with reports of prior adverse reactions to the drugs and those who disagreed or did not sign the consent form were excluded.Coordenadoria de Assist\u00eancia Farmac\u00eautica do Amap\u00e1). Prior to use, the validity, concentration and lots of all drugs were carefully checked. Moreover, physicochemical analyses were performed prior to use, which consisted of determining the weight of the tablets according to the general methods described in the Brazilian Pharmacopoeia [The drugs used were supplied by the Coordination of Pharmaceutical Assistance of Amap\u00e1  and after one, three, seven, 14, 21, and 28\u00a0days. The typical signs and symptoms of malaria as well as non-specific symptoms were classified as \u2018present\u2019 or \u2018absent\u2019 , 4. PatiThe first-line regimen used to treat vivax malaria consisted of chloroquine diphosphate  at doses of 10\u00a0mg base/kg on the first day followed by 7.5\u00a0mg/kg on the second and third days, corresponding to a total dose of 25\u00a0mg/kg, combined with primaquine diphosphate  at a dose of 0.50\u00a0kg base/kg for 7\u00a0days, corresponding to a total dose of 3.0\u20133.5\u00a0mg/kg. The dose of primaquine was adjusted for weight for patients weighing 70\u00a0kg or more as shown in Table\u00a0The adherence by patients to treatment was evaluated during follow-up clinical interviews, as well as by pill counting performed daily at home of patients. Subsequently, these values were compared with blood anti-malarial drug concentrations , 21, 22.P. vivax were not found. Three levels of parasitaemia were defined: low, intermediate, and high, which corresponded to up to 15,000, 15,001\u201360,000, and equal to or greater than 60,001 parasites per mm3 of blood, respectively. The parasitaemia for inclusion in the study ranged from 200 to 100,000 parasite per mm3 of blood. The parasitaemia clearance time (PCT) of the patients was checked every 24\u00a0h using thick-smear slides on days 1, 2, 3, 4 5, and 7. Values are expressed as the means and standard deviation [The thick smear was stained with Giemsa following Walker\u2019s method and examined under a light microscope. Parasitaemia was determined by counting the parasites in 100 microscopic fields (750 times), and this value was multiplied by five, with the result expressed as parasites per cubic millimetre of blood. Smears were considered negative  when, after examining at least 200 microscopic fields, asexual forms of eviation , 24.Laborat\u00f3rio Central de Sa\u00fade P\u00fablica do Amap\u00e1\u2014LACEN-Amap\u00e1) reference laboratory.All positive slides were reviewed in the Central Laboratory of Public Health of Amap\u00e1 , and dried at room temperature for 4\u00a0h. Then, the filter paper was placed in a paper envelope.Blood samples (approximate volume of 20 \u03bcL) were collected during clinical assessments using finger-prick lancets. Samples were then mounted on glass slides to obtain thick blood smears . For theChloroquine and primaquine were measured in order to assess the adherence. A reverse-phase high-performance liquid chromatography system was used for the measurement of both drugs. The chromatographic conditions for determination of primaquine were as follows: UV detection: 254\u00a0nm; flow rate: 1\u00a0mL/min; column: RP-8.15\u00a0cm, 5\u00a0\u03bcm and 4.6\u00a0mm internal diameter ; mobile phase: acetonitrile:methanol:water 30:26:95, v/v); pH 5; injection volume: 50\u00a0mL. The validated analytical procedure for the determination of primaquine concentrations showed that the method was linear from 10 to 900\u00a0ng/mL. The mean intra- and inter-day coefficients of variation were 8.3 and 12.1\u00a0%, respectively. The limit of detection was 5\u00a0ng/mL and the limit of quantification was 10\u00a0ng/mL. The mean retention time of primaquine was 8.8\u00a0min and of quinidine  was 7.0\u00a0min. The mean recovery of primaquine was 90.3\u00a0%. The stability of samples supplemented with primaquine was 120\u00a0days under the conditions described above [:95, v/v;For chloroquine, the chromatographic conditions were as follows: column: C-8, Xterra 4.6\u00a0\u00d7\u00a0150\u00a0mm in size, 5-\u03bcm particles, reverse phase RP18; pre-column: Xterra, RP18 3.9\u00a0\u00d7\u00a020\u00a0mm, 5-\u03bcm particles; mobile phase: 40\u00a0% acetonitrile, 60\u00a0% (1\u00a0% triethylamine in water); pH 10.5, flow rate: 1.2\u00a0mL/min; UV detection: 333\u00a0nm; excitation: 330\u00a0nm; temperature: 25\u00a0\u00b0C; injection volume: 50 \u03bcL. The mean retention time for chloroquine was 20.0\u00a0min and for quinine  was 6.0\u00a0min. The limit of detection was 25\u00a0ng/mL and the limit of quantification was 38\u00a0ng/mL. The method was linear from 38 to 250\u00a0ng/mL. The mean recovery was 87\u00a0%. The intra- and inter-day coefficients of variation were 10.5 and 13\u00a0%, respectively. The stability of samples supplemented with chloroquine was 90\u00a0days .The World Health Organization classification criteria for parasitological response in areas with intense transmission was adopted, with sensitive (S) or resistant types I, II and III . ParasitThe sample size was based on the World Health Organization protocol for studies of the efficacy of anti-malarial drugs in the Americas, specifically of the efficacy of chloroquine in the treatment of vivax malaria. A minimum of 81 patients was required, but the study included 103 patients, with 95 who were fully monitored .Samples are expressed as the means (SD) and as median and range when required. The Fisher\u2019s exact, Chi square and G tests were used to compare the variables: gender, age, origin of cases, and signs and symptoms of the disease. The correlation between parasitaemia and the presence of signs and symptoms of infection was determined by the Spearman\u2019s correlation coefficient (s). The Mann\u2013Whitney test was used to compare parasitaemia clearance times. The accepted level of significance was 5\u00a0%.The study was approved by the Research Ethics Committee of the SEAMA School under protocol number CEP/SEAMA No. 079/08 of 11/27/2008 as part of the main project. Candidate participants were only included in the study after receiving clarification and providing written agreement to participate in the study.Plasmodium falciparum, which was diagnosed during the first week of follow-up.A total of 103 patients with laboratory diagnosis of vivax malaria were initially included in the study. Of these, 95 completed the follow-up. Six participants were excluded due to loss to follow-up, because they went away of Oiapoque. Another two patients were excluded due to concurrent infections with p\u00a0=\u00a00.0002). Of cases originating in French Guiana, the illegal gold mining site of \u2018Sikini\u2019 was the most commonly cited with 21.9\u00a0% of cases (p\u00a0<\u00a00.0001).The majority of patients were males (67\u00a0%). The mean age was 30 (\u00b112) years, ranging from 16 to 60\u00a0years. The age group most affected by the disease was 20\u201329\u00a0years followed by 30\u201339\u00a0years for both genders. A total of 19\u00a0% of cases corresponded to primary infection. Reports of previous infection were divided into one, two and more than four, corresponding to 2, 15 and 64\u00a0% of cases, respectively. Autochthonous cases in Oiapoque represented 69\u00a0%  in 45\u00a0min, which is above the upper limit of 70\u00a0%. The primaquine diphosphate tablets exhibited a release rate of 85\u00a0% (\u00b13.2) in 45\u00a0min, which was also above the minimal tolerance value of 80\u00a0%. The mean chloroquine diphosphate content in the tablets was 98.31 (2.11)\u00a0%, ranging from 96.33 to 103.45\u00a0% of the labelled value. The mean primaquine diphosphate content in the tablets was 99.87 (2.28)\u00a0%, ranging from 98.12 to 101.13\u00a0%, i.e., within the acceptable range of 93.0\u2013107.0\u00a0% of the labelled value.p\u00a0<\u00a00.0001).Several clinical symptoms were reported, including fever (88\u00a0%), chills and headache (86\u00a0%), sweating (83\u00a0%), myalgia (43\u00a0%), and nausea (56\u00a0%). Despite fever being the most frequent clinical manifestation, it was not the initial complaint in 12\u00a0% of patients. Other symptoms, such as headache, myalgia and nausea were initially mentioned. The classic triad of malarial symptoms  was reported in 46.7\u00a0% of cases. There was a significant correlation between parasitaemia and the presence of signs and symptoms of infection . It is noteworthy that the relapsed patient did not require adjustment, as the patient weighed 55\u00a0kg.The dose of primaquine was adjusted in 32.6\u00a0% of patients weighing 70\u00a0kg or more by adding two 15-mg tablets on each day, which is equivalent to 0.30\u00a0mg/day. In this group, the total dose of primaquine ranged from 240 to 304\u00a0mg. The duration of administration was increased from 8 to 10\u00a0days to reach the total dose of primaquine. The adjustment of doses did not alter blood concentrations of primaquine . There were no cases of high parasitaemia.The geometric mean of parasitaemia was 1168 \u00b13.32) parasites/mm parasitep\u00a0=\u00a00.0284).The parasitaemia clearance time ranged from 48 to 120\u00a0h. All patients were negative for parasitaemia during the first week of treatment. Of these, 80\u00a0% were negative on D3 and the rest during the first week (D0\u2013D7). Patients with an intermediate parasite density showed a longer parasitaemia clearance time compared to those with low parasite density  with complete disappearance of symptoms and clearance of parasites, which was classified as sensitive (S). One participant (1.0\u00a0%) had late treatment failure, with a parasitological response classified as resistance type RI /MS in Oiapoque indicated that 14.2 and 13.5\u00a0% of cure verification slides were positive in 2009 and 2010, respectively. However, one should consider that the examination in this study was conducted on day 28 after beginning treatment, as opposed to the 60\u00a0days recommended by the Ministry of Health after treatment for P. vivax.The adequate parasitological response at the end of clinical-laboratory follow-up showed the therapeutic efficacy of the standard treatment in 99\u00a0% of cases, confirming previous findings in patients treated with the same therapeutic regimen in Bel\u00e9m, in the Brazilian state of Para . Howeverreatment , 36. In P. vivax , 35, 36.P. vivax , 15, 16.P. vivax , 15, 16.The control of factors responsible by treatment failures in vivax malaria evaluated in the current study, such as the adjustment of the dose of primaquine for patient weight, the intrinsic quality of the drugs, and the adherence of all patients to the prescribed treatment assessed through interviews, pill counts and confirmation of drug exposure by the concentrations of anti-malarial drugs in blood may have contributed to the high effectiveness of the standard treatment. In addition, the efficiency of disease control policies in Brazil, based on early and correct diagnosis, as well as in the timely and appropriate treatment of cases, may have also contributed to these findings, which was corroborated by the agreement of thick blood smear diagnoses between different treatment centres, as well as by the absence of anti-malarial drugs in pre-dose samples .P. vivax at concentrations above the minimum inhibitory concentration for chloroquine-sensitive strains. The treatment failure could be explained by the fact that there are different parasite genotypes circulating in the region [Late treatment failure in one patient could be classified as resistance type RI. The plasma concentration of chloroquine at D28 was greater than 100\u00a0ng/mL, indicating adequate drug exposure as well as the presence of blood strains of e region , 25, 37.P. vivax isolates have no significant resistance profile to the currently recommended treatment. The control over external factors, such as the dose adjustment of primaquine based on the weight of the patient, the quality of anti-malarial drugs and adherence to treatment, contributed to the high effectiveness of the standard treatment found in the study.In the municipality of Oiapoque the treatment regimen of chloroquine combined with primaquine used for the treatment of vivax malaria remains effective. In addition, circulating"}
+{"text": "Plasmodium vivax in Bolivia, where 95% of the cases of malaria are attributed to this species. The aim of this study was to evaluate the therapeutic efficacy of CQ in this setting.Chloroquine (CQ) over three days plus primaquine (PQ) for seven days is the treatment of choice of infections by P. vivax mono-infection received 25\u00a0mg/Kg body weight of CQ over three days. The concentrations of CQ\u00a0+\u00a0desethylchloroquine (DCQ) in blood were determined at days 7 and 28 of follow up; at follow-up and on the day of treatment failure was administered PQ.Patients in the Amazon region of northern Bolivia, were included in the study from May to November 2011 and the therapeutic efficacy of CQ was evaluated over a 28-day follow-up period. Patients with One hundred patients fulfilled the inclusion criteria, two were lost to follow up and another two were later excluded for protocol violation. Of the 96 patients who completed the follow up 10 showed TF; one presented continued parasitaemia until day 7 of follow up, three on day 21 and six on day 28 of follow up. The geometric mean of CQ\u00a0+\u00a0DCQ on day 7 was 321.7\u00a0ng/ml (range 197\u2013535\u00a0ng/ml). In six patients with TF the CQ\u00a0+\u00a0DCQ concentrations in blood on the day of TF were >100\u00a0ng/ml. The rate of resistance was 6.5%.P. vivax in the Amazon region of Bolivia. New clinical trials are needed to establish alternative treatments against these parasites in this region of South America.The present study demonstrates the presence of resistance to CQ in the treatment of malaria by  Plasmodium vivax while, in Bolivia, 93% correspond to this parasite and the remaining 7% to Plasmodium falciparum. In Bolivia, the first-line therapeutic schedule for infections by P. vivax includes 25\u00a0mg/Kg body weight of chloroquine (CQ) for three days plus 3.5\u00a0mg of primaquine (PQ) for 7\u00a0days. The first is aimed at reducing the parasitic load in young forms of the parasite and eliminating immature gametocytes while the second drug is active against the gametocytes and hypnozoites of this species [In South America, 60%  of the cP. vivax was in Papua New Guinea in 1989 [P. vivax were reported in Guyana in 1996, with three patients maintaining parasitaemia in the presence of adequate serum levels of CQ [The first description of resistance to CQ in infections by  in 1989 , with th in 1989 , 5. In Lls of CQ  and in BIn 2003, an in vivo evaluation of resistance to CQ was carried out in the municipality of Riberalta in Bolivia, reporting 15% 9/59 cases) of TF over a 28-day period. However, none of the cases showed serum levels of the drug /59 cases. The preP. vivax are reported in Bolivia [The study was developed in the department of Beni, in the municipality of Riberalta, in the north of the Amazonia of Bolivia where 18% of the diagnoses of malaria and 21% of the cases of malaria by  Bolivia .P. vivax, a parasite density of asexual forms of 250\u2013100,000 parasites/\u03bcl of blood, a temperature >37.5\u00b0C or in its absence a history of a rise in temperature within the last 48\u00a0h. Microscopic diagnosis was performed according to the practical guidelines of diagnosis of the National Malaria Control Programme [The screening methods for surveillance of anti-malarial drug efficacy (MSADE) of the WHO  were folrogramme , (thick The exclusion criteria were: pregnant women confirmed by rapid pregnancy diagnostic tests (detection of human chorionic gonadotropin in urine), patients with signs and symptoms of severe malaria, a history of allergy to anti-malarial drugs, concomitant presence of severe or chronic diseases such as tuberculosis or HIV/AIDS and a previous history of having received anti-malarial drugs.The treatment consisted of the use of chloroquine phosphate  25\u00a0mg/Kg body weight administered orally over 3\u00a0days: on days 0 and 1 of the study 10\u00a0mg/Kg body weight were given and on day 2, 5\u00a0mg/Kg body weight were administered under strict supervision of the investigative team. Patients vomiting within half an hour after the administration of the drug were given another treatment cycle with the same dose and patients with more than two episodes of vomiting were excluded from the study.P. falciparum, plus PQ 0.5\u00a0mg/Kg over 7\u00a0days.The patients who accepted to participate in the study were followed on days 2, 3, 7, 14, 21 and 28 after the treatment. The day of diagnosis was denominated day 0. After the 28\u00a0days of follow-up all the patients were treated with PQ 0.5\u00a0mg/Kg body weight/day administered orally during seven days. Patients with TF or those in whom mixed malaria was detected were excluded from the study. These patients were then administered artemisinin-based combination therapy (ACT) used in Bolivia for the treatment of Two milliliters of blood were obtained by venopuncture from all the patients in the study, with or without parasitaemia, on days 7 and 28 of the follow up. The concentrations of CQ and DCQ in blood were also determined by high performance liquid chromatography (HPLC) according to the standardized technique of the Center for Disease Control in Atlanta, USA in patients with TF. The samples obtained were kept at 4\u00b0C until analysis.n-hexane, t-butyl methyl ether and diethylamine (1:1:0.003), flow gradient: 1.0\u00a0ml/min; injection volume 7\u00a0\u00b5l and column temperature: 30\u00b0C. A concentration of CQ\u00a0+\u00a0DCQ of 70\u201399\u00a0ng/ml was considered effective to eliminate all the asexual and sexual forms of P. vivax in blood [The HPLC was performed using the Shimadzu LC-10ATVP with RF-10 AXL fluorescence detector under the following chromatographic conditions: chromatographic column: Agilent Zorbax SIL, silane packing (corresponding to L3 according to USP). Detector: fluorescence, emission 380\u00a0nm, excitation 320\u00a0nm, mobile phase A: methanol and diethylamine (100:0.3); mobile phase B: in blood  and a CQin blood \u201317.\u00ae 7  software was used to analyse the data of frequency and to measure the data of CQ\u00a0+\u00a0DCQ. The statistical package SPSS\u00ae v 20  14 was used to compare the means and medians of the results of CQ\u00a0+\u00a0DCQ and associated variables, with parametric and non-parametric models of analysis of variance. Kaplan-Meier analysis of survival and confidence intervals was analysed by the WHO programme for study the in vivo therapeutic efficacy [The Tableauefficacy . All theThe study was performed following the recommendations of the National Committee of Bioethics of Bolivia and the Ethical Committee of PAHO (PAHOERC). All patients provided informed consent to participate in the study, and in those under legal age informed consent was obtained from their legal guardians.P. vivax were reported out of a total of 5,290 cases evaluated in the urban area of Riberalta. One hundred of these patients (51 in CM and 49 in LU) were included in the study. Of the total number of cases evaluated 64% were males and 36% females, with a median age of 20\u00a0years (range 5\u201369\u00a0years), and 25% of whom were under the age of 15\u00a0years. Thirty-one percent (n\u00a0=\u00a031) had an axillary temperature >37\u00b0C, the geometric mean of PD on day 0 was 3,837.43\u00a0parasites/\u00b5l , The geometric mean of CQ administered over the three days of treatment was 1,332.19\u00a0mg . The mean haemoglobin values on day 0 in the females was 10\u00a0g/dl (range 7\u201314\u00a0g/dl) and 12\u00a0g/dl (range 8\u201316\u00a0g/dl) in males, and 75% (n\u00a0=\u00a075) had anaemia before initiating the treatment  eliminated the parasitaemia on day 2; 91% (n\u00a0=\u00a087) on day 3 and 99% (n\u00a0=\u00a095) on day 7 of follow up. Parasitaemia or clinical deterioration were not observed until the end of follow up in 89.6% (n\u00a0=\u00a086). Treatment failure was observed on days 7, 21 and 28 follow-up, with 1.04% (n\u00a0=\u00a01) on day 7, 3.1% (n\u00a0=\u00a03) on day 21 and 6.3% (n\u00a0=\u00a06) on day 28. The median age of the patients cured was 21\u00a0years (range 5\u201361\u00a0years) and that of patients with TF was 14\u00a0years (range 5\u201332\u00a0years), with the differences being statistically significant (P\u00a0=\u00a0<0.05). With the Kaplan-Meier analysis the cumulative incidence of treatment failure was 0.104 (95% CI 0.057\u20130.185).Of the 100 patients included in the study two were lost to follow up and another two were later excluded; one for presenting co-infection by The geometric mean CQ\u00a0+\u00a0DCQ value in blood of all the patients followed with and without TF (n\u00a0=\u00a096) on day 7 was 319.46\u00a0ng/ml (range 50\u2013743\u00a0ng/ml), being 109.24\u00a0ng/ml (range 34\u2013602\u00a0ng/ml) on day 28 of follow up (n\u00a0=\u00a092). In the patient with continued presence of parasites in blood until day 7 inclusive the CQ\u00a0+\u00a0DCQ values on day 7 were 535\u00a0\u00b5g/ml and the patient was excluded from the study. In the 10 patients presenting TF the geometric mean of CQ\u00a0+\u00a0DCQ was 321.7\u00a0ng/ml (range 197\u2013535\u00a0ng/ml) on day 7, being 113.46\u00a0ng/ml (range 75\u2013223\u00a0ng/ml) on the day of TF. In six patients the CQ\u00a0+\u00a0DCQ level on the day of TF was above the minimum elimination concentration (MEC) (100\u00a0ng/ml)  demonstrating the CQ presented adequate absorption through the intestinal tract . In 6 ofP. vivax with the levels of the drug in blood, describing resistance in two out of 177 patients in the Amazon region of Peru [Ruebsh et al. confirmed resistance to CQ in infections by  of Peru . In addi of Peru .P. falciparum because of their demonstrated rapid effectiveness in eliminating the parasitic load and achieving complete cure of the disease [P. vivax. Several clinical trials have demonstrated the efficacy of the ACT in the treatment of malaria P. vivax, with dihydroartemisinin-piperaquine being the ACT most frequently studied [According to the recommendations of the WHO, a change in the treatment schedule should be considered in cases of proven resistance >10% . The rat disease . This is studied . Taking  studied , the cho studied , 24, witP. vivax in the Amazon region of Bolivia. Considering the percentage of resistance found and the rate of TF with MEC of the drug in blood on the day of TF this resistance could be greater. New treatment schedules should be evaluated to guarantee the control of malaria in this region.In conclusion, 6.5% of resistance to CQ was observed in infections by"}
+{"text": "Plasmodium falciparum or Plasmodium vivax compared with 2.2% and 30.0%, respectively, in AM-LM-treated children during 42\u00a0days of follow-up. To determine whether, consistent with the long elimination half-life of NQ, this difference in efficacy would be more durable, clinical episodes of malaria were assessed in a subset of trial patients followed for six months post-treatment.In a recent trial of artemisinin-naphthoquine (artemisinin-NQ) and artemether-lumefantrine (AM-LM) therapy in young children from Papua New Guinea (PNG), there were no treatment failures in artemisinin-NQ-treated children with For children completing trial procedures and who were assessable at six months, all within-trial and subsequent clinical malaria episodes were ascertained, the latter by clinic attendances and/or review of hand-held health records. Presentations with non-malarial illness were also recorded. Differences between allocated treatments for pre-specified endpoints were determined using Kaplan-Meier survival analysis.P\u2009=\u20090.033, log rank test). The median (interquartile range) time to first episode of clinical malaria was 64 (50\u2013146) vs 116 (77\u2013130) days, respectively (P\u2009=\u20090.20). There were no between-group differences in the incidence of first presentation with non-malarial illness (P\u2009=\u20090.31).Of 247 children who were followed to Day 42, 176 (71.3%) were included in the present sub-study, 87 allocated to AM-LM and 89 to artemisinin-NQ. Twenty children in the AM-LM group (32.8%) had a first episode of clinical malaria within six months compared with 10 (16.4%) in the artemisinin-NQ group (The greater effectiveness of artemisinin-NQ over conventional AM-LM extends to at least six months post-treatment for clinical malaria but not non-malarial illness.ACTRN12610000913077.Australian New Zealand Clinical Trials Registry  Plasmodium vivax is transmitted, late post-treatment recurrences can result from relapses from dormant liver stages. Durations of follow-up of up to 63\u00a0days have been employed in this situation . Two-sample comparisons for normally-distributed variables were by Student\u2019s Of the 267 children randomized in the parent trial, 247 (92.5%) were followed to Day 42. Of this latter number, 176 (71.3%) were included in the six month analysis, comprising 87 allocated to AM-LM and 89 to artemisinin-NQ  within six months of treatment  children had a first episode of clinical malaria (due to either P\u2009=\u20090.008).If only those children who had an ACPR at Day 42 in the parent trial were considered, 12 of the 37 in the AM-LM group (32.4%) and 10 of 89 in the artemisinin-NQ group (11.2%) had an episode of clinical malaria between six weeks and six months after treatment  definite P. falciparum (slide or RDT positive for this species), ii) definite P. vivax (slide or RDT positive for this species), iii) definite mixed P. falciparum/P.vivax (slide positive for these species), iv) P. falciparum with or without P. vivax , and v) unknown . There was no difference between the proportions of children in each category by allocated treatment (see Table\u00a0P\u2009=\u20090.12 by Fisher\u2019s exact test). In 19 out of the 30 cases (63.3%), there was definite slide or RDT evidence of P. falciparum infection.From the available data relating to species of P\u2009=\u20090.36; see Table\u00a0P\u2009=\u20090.31 by log rank test). There was also no significant between-group difference in the time to the first episode of a non-malarial illness .Twenty-one 24.1%) and 16 (18.0%) of children in the AM-LM and artemisinin-NQ groups, respectively, had a first episode of non-malarial illness during the six-month follow-up period , there  EIR 562 . In the m EIR 37 ), the poin vivo, in vitro and/or genetic monitoring of parasite resistance should be an integral part of this process.Antimalarial drugs such as NQ with long elimination half-lives are more likely to promote the development of parasite resistance as a result of prolonged sub-therapeutic plasma concentrations ,27. In tPlasmodium species nor genotyping to differentiate recrudescences from reinfections was performed, but the primary endpoint  was a pragmatic one designed to allow the long-term real-world effectiveness of the two treatment regimens to be assessed. For logistic and financial reasons, not all participants in the parent trial at each site were followed for the full six months, but most children were included and the two treatment groups were similar in number and baseline characteristics. Indeed, the significantly greater number of vivax cases in the artemisinin-NQ group and the potentially high rate of associated late relapses may have led to bias against this ACT, but it nonetheless proved more effective than AM-LM.The present study had limitations. Neither accurate differentiation between The present study is one of the few published in the literature that have employed long-term follow-up of patients with malaria treated with artemisinin combination therapy. The data suggest strongly that the long terminal elimination half-life of three daily doses of NQ confers benefit of artemisinin-NQ over AM-LM to at least six months post-treatment. This observation should favourably influence cost-effectiveness analyses. In addition, the present long-term follow-up data also add to the reassuring results of detailed safety monitoring over 42\u00a0days in the parent trial , in that"}
+{"text": "Plasmodium vivax, but the local efficacy has not been assessed. The impact of cases imported from India on the genetic make-up of the local vivax populations is currently unknown.Bhutan has made substantial progress in reducing malaria incidence. The national guidelines recommend chloroquine (CQ) and primaquine (PQ) for radical cure of uncomplicated P. vivax mono-infection were enrolled into a clinical efficacy study and molecular survey. Study participants received a standard dose of CQ (25\u00a0mg/kg over 3\u00a0days) followed by weekly review until day 28. On day 28 a 14-day regimen of PQ (0.25\u00a0mg/kg/day) was commenced under direct observation. After day 42, patients were followed up monthly for a year. The primary and secondary endpoints were risk of treatment failure at day 28 and at 1\u00a0year. Parasite genotyping was undertaken at nine tandem repeat markers, and standard population genetic metrics were applied to examine population diversity and structure in infections thought to be acquired inside or outside of Bhutan.Patients over 4\u00a0years of age with uncomplicated HE 0.87 and 0.90) in both populations. There was no notable differentiation between the autochthonous and imported populations.A total of 24 patients were enrolled in the clinical study between April 2013 and October 2015. Eight patients (33.3\u00a0%) were lost to follow-up in the first 6\u00a0months and another eight patients lost between 6 and 12\u00a0months. No (0/24) treatment failures occurred by day 28 and no (0/8) parasitaemia was detected following PQ treatment. Some 95.8\u00a0% (23/24) of patients were aparasitaemic by day 2. There were no haemolytic or serious events. Genotyping was undertaken on parasites from 12 autochthonous cases and 16 suspected imported cases. Diversity was high  contains supplementary material, which is available to authorized users. Plasmodium vivax infection worldwide [Plasmodium falciparum decreases in many areas of Asia, P. vivax has emerged as the more prominent species. P. vivax exhibits important biological differences from P. falciparum, including the development of dormant liver stages and the emergence of gametocytes before the onset of clinical symptoms [P. falciparum, posing a challenge to control efforts and rendering P. vivax highly prone to resurgence. Furthermore P. vivax also exhibits greater genetic diversity than P. falciparum [P. vivax strains with adaptive genetic backgrounds to emerge in response to environmental challenges, such as anti-malarial drugs or host immune pressure. The large burden of low density P. vivax infections presents a diagnostic challenge, further impeding containment efforts [More than 2.8 billion people are reported to be at risk of orldwide , 2, the orldwide . As the symptoms . These plciparum \u20138, enhan efforts .P. vivax, which accounts for over 60\u00a0% of cases [P. vivax isolates to lie dormant in the liver for weeks or months before relapsing greatly enhances the parasite\u2019s \u2018mobility\u2019, compounding the risks of importation of this species. Diligent surveillance of imported infections and ongoing local parasite transmission dynamics are critical to the success of malaria elimination efforts. As demonstrated in other studies, parasite genotyping can provide useful insights in monitoring both local and imported infection during the pre-elimination phase [Bhutan has made remarkable progress in reducing the malaria burden in the country, with a significant decline in clinical cases from nearly 40,000 in 1994 to 436 in 2010 [of cases . The vecof cases . Currentof cases , which con phase .P. vivax in Bhutan. However, given the accumulating reports of chloroquine-resistant P. vivax in other parts of the world [P. vivax in Bhutan.To date, there has been no clinical-parasitological evidence of chloroquine-resistant he world , there iP. vivax population.Maintaining momentum in the late stages of malaria elimination becomes increasingly challenging as the numbers of clinical cases falls and the public health priority for malaria control activities declines. However, during this phase, informed decision-making on the optimal cost-effective strategies is critical, and yet the clinical and molecular characterization of the parasite population is rarely addressed. The current study was conducted to address these issues in a very low-endemic pre-elimination setting, assessing for the first time the efficacy of CQ plus primaquine (PQ) against uncomplicated vivax malaria in Bhutan, and gauging the transmission dynamics of autochthonous and suspected imported cases in the local 2, and sharing borders with China in the north and India in the south, west and east. The population size is estimated over 772,000 with a floating population (expatriates) of ~37,000 persons [P. falciparum and P. vivax. Reported mosquito vectors for these species include Anopheles pseudowillmori and Anopheles culicifacies. Anopheles minimus, Anopheles fluviatilis and Anopheles dirus were present in the past, but have not been recorded in recent years. The incidence of malaria has declined sharply in Bhutan from 194 cases : 0.4/1000 risk population) in 2011 to 48 cases (API: 0.1/1000 risk population) in 2013. Among the districts that reported malaria cases in 2013, the majority (60\u00a0%) was reported in Sarpang district (API: 10/1000 risk population). The remaining cases were reported in S/jongkhar, Dagana and Samtse districts, with API of 3, 2 and 1.5 per 1000 risk population, respectively.Bhutan is located in the eastern Himalaya region, encompassing an area of approximately 38,000\u00a0km persons . About 7The study was designed as a 1-year trial at 12 sentinel sites: eight health centres in Sarpang district and four other sites in Samtse, Dagana and Samdrup Jonghkar districts. These sites were chosen based on the number of reported cases in 2011 and 2012. Due to low recruitment, the study period was extended for a further 16\u00a0months and the number of sites expanded to include 35 health centres across an additional four high-risk districts   and on days 1, 2, 3, 7, 14 and 28. A complete medical history and demographic information were recorded at baseline and a capillary or venous blood sample taken for microscopy, Hb and molecular analyses. At each follow-up visit, any adverse events were documented, the axillary temperature was measured and a finger-prick blood sample collected for microscopy. Hb was measured using Hemocue\u2122  on days 7, 14, 28, 35, 41 and on the last visit.Thick and thin blood films were examined by two qualified microscopists who read the slides independently, and the parasitaemia was calculated by averaging the two counts. Blood smears with discordant results (differences between the two microscopists in parasite species or the density of parasitaemia >50\u00a0%) were re-examined by a third, independent microscopist from the VDCP, and parasite density was calculated by averaging the two closest counts. Slides were declared negative when no parasites were seen in 200 high power fields.Plasmodium species were confirmed by PCR using the protocol described by Padley et al. [Pv3.27, msp1F3, MS1, MS5, MS8, MS10, MS12, MS16 and MS20 [Pv3.27, MS16 and msp1F3 loci were amplified using methods described previously [P. vivax genotyping studies, minor alleles were only called if they had a minimum 33\u00a0% height of the predominant allele [DNA was extracted from 200\u00a0\u00b5l of whole blood using the QIAamp DNA Blood mini kit (Qiagen) according to the manufacturer\u2019s instructions. y et al.  with theand MS20 , 20. Theand MS20 . In addiand MS20 . The Pv3eviously . The proeviously , 24. Thet allele .P. vivax detected by microscopy during the follow-up period. The primary efficacy endpoint was the risk of recurrent parasitaemia by day 28 and the secondary efficacy endpoints were the cumulative risk of recurrent parasitaemia by 12\u00a0months and the proportion of patients with detectable parasitaemia on days 1 and 2. Safety outcomes were the type and number of adverse events and serious adverse events occurring during the entire follow-up and the Hb profile.Parasite recurrence was defined as any parasitaemia with t test for parametric comparisons. For categorical variables percentages and corresponding 95\u00a0% confidence intervals (95\u00a0% CI) were calculated using Wilson\u2019s method. Proportions were examined using \u03c72 with Yates\u2019 correction or Fisher\u2019s exact test. The cumulative risk of failure was assessed by survival analysis using the Kaplan\u2013Meier method. Patients lost to follow-up were censored on the last day of follow-up.Clinical data were entered into an access database. All statistical analyses were undertaken using STATA 13 . The Mann\u2013Whitney U test or Kruskal\u2013Wallis method were used for non-parametric comparisons, and Student\u2019s An infection was defined as polyclonal if more than one allele was observed at one or more loci. The multiplicity of infection (MOI) for a given sample was defined as the maximum number of alleles observed at any of the loci investigated. With the exception of measures of polyclonality and MOI, only the predominant allele at each locus in each isolate was used for analysis .HE) was measured as an index of population diversity using the formula n is the number of isolates analysed and pi is the frequency of the ith allele in the population. The probability (P) that two unrelated parasites exhibited homologous genotypes (expected homozygosity) was calculated for each marker using the formula P\u00a0=\u00a0\u03a3pi2, where pi is the frequency of the ith allele in the population [Pi) of two unrelated parasites exhibiting homologous genotypes at multiple loci were calculated by multiplying P for the individual loci in question.The expected heterozygosity (pulation . CumulatIAS) using the web-based LIAN 3.5 software [IASestimates was assessed using 10,000 random permutations of the data. LD was assessed in (1) all samples, and (2) with repeated MLGs represented once.Multi-locus genotypes (MLGs) were reconstructed from the predominant allele at each locus in isolates with no missing data. Multi-locus linkage disequilibrium (LD) was measured by the standardized index of association (software . The sigps). Using (1-ps) as a measure of genetic distance [r-test with 10,000 permutations using the ade4 package in R [The genetic relatedness between sample pairs was assessed by measuring the proportion of alleles shared between MLG pairs , and the Human Research Ethics Committee of the Northern Territory Department of Health and Menzies School of Health Research, Australia (HREC 2012-1871). The study was registered at ClinicalTrial.gov with the registration number NCT01716260.Between May 2013 and October 2015, a total of 28 patients were screened for enrolment, four of whom declined consent. Of the remaining 24 patients, adherence to follow-up was achieved in 19 (79.2\u00a0%) patients at day 28, 16 (66.6\u00a0%) at 6\u00a0months and 8 (33.3\u00a0%) at 12\u00a0months  with no patients receiving less than 20\u00a0mg/kg. The median total dose of PQ was 3.7\u00a0mg/kg (IQR 3.3\u20134.8) with a range between 2.8 and 5.7\u00a0mg/kg.There was no treatment failure by day 28 and no peripheral parasitaemia was detected during the remaining follow-up after PQ treatment. On day 1, 45.2\u00a0% (11/24) patients remained parasitaemic by microscopy, but on day 2 this had fallen to 4.4\u00a0% (1/23). All patients had cleared asexual and sexual forms of parasites by day 3 (0/22).The mean Hb at enrolment was 11.4\u00a0g/dl (95\u00a0% CI 10.8\u201311.9) with no significant difference between baseline and day 7. By day 28, before start of PQ, the Hb concentration had risen to 12.0\u00a0g/dl (95\u00a0% CI 11.1\u201312.9) and this did not change significantly after 14\u00a0days of PQ (mean 12.1\u00a0g/dl (95\u00a0% CI 11.3\u201312.8). The Hb did not fall by more than 25\u00a0%\u00a0from baseline in any of the patients and none required a blood transfusion. There was no difference between Bhutanese and non-Bhutanese\u00a0patients regarding their Hb concentrations before or after PQ treatment. Two patients reported stomach pain on day 1, and another two reported nausea on the same day. Five patients complained of headache on days 1, 2, 3 and 7. No serious adverse events occurred.P. vivax infection could not participate in the clinical survey but their specimens were used for parasite genotyping. Forty-six percent (13/28) of the samples were autochthonous and 54\u00a0% (15/28) were classified as imported. Genotyping was successfully undertaken in all of these samples, with only two isolates failing at a single marker (MS20 locus in both cases). All markers, including the five \u2018balanced\u2019 markers had high diversity (HE range 0.85\u20130.97)  of patients enrolled in the clinical survey. An additional five patients with microscopy and PCR-positive 3/24 of pHE 0.91). Diversity levels were high in both the autochthonous (mean HE 0.87) and the imported infections (mean HE 0.90). Diversity remained high (mean HE 0.89) for autochthonous (mean HE 0.85) and imported (mean HE 0.89) cases when analysis was restricted to the five balanced markers.Six 21.4\u00a0%) infections demonstrated evidence of polyclonality, including five (38\u00a0%) autochthonous and one (7\u00a0%) imported case (Table\u00a01.4\u00a0% infp) that two unrelated parasites would exhibit homologous genotypes at all nine loci was 2\u00a0\u00d7\u00a010\u221210. Aside from the four isolates with identical/highly related MLGs, there was no evidence of significant correlation between the distance in sampling date and the proportion of alleles shared between infections . LD was high in the autochthonous population  but dropped more than two-fold after adjustment for the repeated MLGs in the identical strains  .There was no notable separation between the Bhutanese vs non-Bhutanese isolates in the neighbour-joining tree Fig.\u00a0. Four inP. vivax population, likely sustained by infections imported from India.This study presents the first description of the genetic diversity and structure of the parasite population and the sustained efficacy of CQ and PQ against uncomplicated vivax malaria in Bhutan. The clinical and molecular features of the parasite population were gathered as the country enters the final stages of malaria elimination. In this very low endemic setting, over 60\u00a0% of the infections sampled originated from non-Bhutanese patients travelling across the border from northern India. The results reveal extensive genetic diversity in the P. vivax appears to be effective with rapid early parasite clearance and no recurrences observed by day 28 [P. vivax remains sensitive to CQ [The Bhutanese treatment policy for y day 28 . These fve to CQ .No relapses were recorded over the subsequent year following administration of directly observed PQ. The one-year follow-up in the current study should have been adequate to have captured even late recurrences, as described in parts of northern India , 33. AlmComparison of baseline data between Bhutanese nationals and non-Bhutanese nationals indicated a trend towards more symptomatic disease among the Bhutanese patient population with significantly higher mean temperatures at enrolment and higher parasite densities, which may reflect differences in prior exposure and acquired immunity. Interestingly, no children presented at any of the participating health centres and the youngest patient enrolled in the study was 23\u00a0years old. This is in line with reports from India where the highest reported number of cases is among young adults, most likely attributable to young males being at greater risk of malaria from employment-seeking activities.In many countries PQ roll-out is hampered by concerns over drug-induced haemolyses in G6PD-deficient patients . The curP. vivax infections (HE 0.87\u201390) was high, comparable to high transmission regions of Southeast Asia and the Pacific [P. falciparum populations, where expected heterozygosity generally exhibits a positive correlation with endemicity [P. vivax population as local transmission levels declined [P. vivax infections to spread undetected as dormant hypnozoites, it is highly probable that infections introduced from India sustain the high P. vivax diversity observed in the local Bhutanese population. The autochthonous Bhutanese infections exhibited nearly as high diversity (HE 0.87) as those from imported infections (HE 0.90), and there was no notable genetic differentiation between these two groups. India currently has the highest burden of P. vivax infection in the world, thus presenting a potentially highly diverse reservoir of infection [Even though the study was carried out in a very low transmission area, the genetic diversity in both autochthonous and imported ns HE 0.8\u201390 was h Pacific . This pademicity . A recendeclined , leadingdeclined , and thenfection .p\u00a0=\u00a02\u00a0\u00d7\u00a010\u221210). These results highlight the capacity of the local vector population to sustain transmission within Bhutan despite best efforts in vector control, and thus underline the importance of maintaining diligent surveillance, especially at border areas, in the final stages of malaria elimination to prevent outbreaks.The genetic analyses also revealed evidence of local transmission events, reflected by four genetically identical autochthonous strains sourced from patients residing in the same district (Sarpang) and presenting at the local health facility between 1 and 46\u00a0days of one another. Based on the diversity observed at each of the loci, the probability that two unrelated infections would present with the same genetic profile across all nine loci by chance alone was extremely low  in the imported P. vivax population is consistent with these infections having been sourced largely from a high transmission setting within which frequent recombination between genetically distinct parasites breaks down LD. After adjusting for the identical strains, LD in the autochthonous population declined more than two-fold but remained significant . Given the low reported incidence of P. vivax infection in Bhutan and low complexity of infections, it is likely that the local levels of LD reflect a complex combination of both local transmission, with a degree of inbreeding, and importation of a diverse array of infections. However, high rates of asymptomatic and sub-patent infections will also sustain local transmission and parasite diversity; further studies are required to assess the depth of this hidden reservoir of infection.The absence of LD  of all reported cases in Bhutan during the study period.The small sample size in Bhutan highlights the challenges for surveillance of the declining symptomatic illness as a country moves towards malaria elimination within its borders. At the end stages of elimination, selective drug pressure increases often resulting in the residual parasite population being the most drug resistant . In thiseview in ), but thP. vivax infection and preventing relapse in uncomplicated patients in Bhutan. Over half of the cases presenting at the local health facilities were visiting patients from India and, in conjunction with the genetic analyses, highlight that local transmission is being sustained primarily by importation of parasites from neighbouring countries. The risk that these imported infections could introduce drug-resistant parasites demands continued surveillance of the Bhutanese P. vivax and P. falciparum populations.In summary, this study demonstrates that supervised treatment with CQ plus 14\u00a0days of PQ remains highly effective at treating blood stage"}
+{"text": "Given increasing rates of resistance to existing therapy, new options for treatment and prophylaxis of malaria are needed.Plasmodium falciparum were randomised in both studies to either azithromycin (AZ) 1,000\u00a0mg plus chloroquine (CQ) 600-mg base  once daily for three days or mefloquine hydrochloride (MQ) 1,250\u00a0mg (split dose). In the first study, an additional regimen of AZ 500\u00a0mg plus CQ 600-mg base (AZCQ 500\u00a0mg) once daily for three days was included. All study participants were hospitalised until three consecutive daily blood smears were negative for asexual P. falciparum parasitaemia. Study participants were evaluated weekly for 42\u00a0days, with Day 28 polymerase chain reaction (PCR)-corrected parasitological clearance rate as primary endpoint.Two randomised, comparative, non-inferiority studies were conducted in Africa, one double-blinded and one open-label. Adults with fever, a positive peripheral blood smear, and a positive rapid diagnostic test for vs MQ 111/112 . Among the participating countries, in vitro CQ resistance based on pfcrt mutation frequency in the baseline isolates across both studies ranged from 20.8% (Zambia) to 96.1% (Uganda). Serious adverse events  were observed more frequently with MQ compared with AZCQ , though discontinuations for AEs were similar . Common AEs in the AZ-containing arms included pruritus, vomiting, dizziness, and headache.A total of 467 subjects were randomised in the two studies. At 28\u00a0days\u2019 follow-up, PCR-corrected parasitological clearance rates in the per protocol population in the first study were 101/103 (98%) with AZCQ 1,000\u00a0mg compared with 102/103 (99%) with MQ . The AZCQ 500-mg regimen was stopped during an interim study review . In the second study, clearance rates were similar: AZCQ 1,000\u00a0mg 107/107 (100%) Among adults with symptomatic uncomplicated falciparum malaria in Africa, the combination of AZ 1,000\u00a0mg and CQ 600-mg base once daily for three days resulted in Day 28 PCR-corrected parasitological clearance rates of \u226598% and was non-inferior to treatment with MQ. AZCQ was well tolerated.NCT00082576 and NCT00367653ClinicalTrials.gov identifiers The online version of this article (doi:10.1186/1475-2875-13-458) contains supplementary material, which is available to authorized users. Plasmodium species led to increased morbidity and mortality  once daily for three days), whereas in study 1155 medication was provided unblinded with the same two treatment regimens as in study 1134. As a consequence, the methods described below relate to study conduct and analysis for both trials with exceptions specifically noted where relevant.P. falciparum asexual parasitaemia between 1,000 and 100,000 parasites/\u03bcL and fever, documented or by history, within the prior 24\u00a0hours. Subjects also were required to have a serum glucose \u226560\u00a0mg/dL and a rapid diagnostic test positive for P. falciparum. Women of childbearing potential were required to have a negative urine human chorionic gonadotropin test before study entry, and enrolled subjects were to use adequate contraception during the entire study. Subjects were excluded for any of the following reasons: clinical or laboratory evidence of severe or complicated malaria; the presence of non-falciparum species on microscopy; pregnancy or breast-feeding; history of allergy or hypersensitivity to AZ, CQ, MQ, or related compounds; history of epilepsy or psoriasis; treatment with any anti-malarial drug or with any antibacterial with known anti-malarial activity within two weeks before enrolment into the study; laboratory evidence of abnormal renal or liver function; any major psychiatric disorders; inability to swallow oral medication; treatment with other investigational drugs 30\u00a0days prior to enrolment; alcohol and/or any other drug abuse; requirement to use medication during the study that might have interfered with the evaluation of the study drug; other medical conditions that would have interfered with the evaluation of the therapeutic response or safety of the study drug; and inability to comprehend and/or unwillingness to follow the study protocol; or, prior participation in this study. Finally, subjects were excluded if they had not lived continuously in a malaria-endemic area for at least the previous year.Eligible subjects were \u226518\u00a0years of age with symptomatic uncomplicated malaria as demonstrated by blood smears positive for Subjects were to be withdrawn from the study and placed on alternative therapy for any of the following reasons: impaired consciousness, respiratory distress, seizures, hypoglycaemia, gross haematuria, increase in parasitaemia to >100,000 parasites/\u03bcL 48\u00a0hours after the first treatment dose, failure of parasitaemia to decrease to 25% of the baseline value on Day 2 and investigator opinion that other anti-malarial therapy was indicated or treatment failure.Study drug was administered as blinded therapy with matching placebo (in study 1134) or unblinded (in study 1155). In study 1134, eligible subjects were initially randomised to oral treatment for three days in one of three active treatment arms: AZ 1,000\u00a0mg plus a CQ 600-mg base  once daily for three days, AZ 500\u00a0mg plus a CQ 600-mg base (AZCQ 500\u00a0mg) once daily for three days, or MQ  1,250\u00a0mg . In study 1155, subjects were randomised to either AZCQ 1,000\u00a0mg once daily for three days or MQ  1,250\u00a0mg . MQ was used as the comparator based on the recommendation of the FDA, as this was the only FDA-approved oral anti-malaria agent at the time. MQ was expected to have high efficacy rates in all of the chosen study locations at the time of the study conduct.P. falciparum. Giemsa-stained blood smears were read and interpreted by experienced microscopists who were blinded to all clinical information, including treatment allocation and the readings by the other microscopists. Three slides were prepared for each subject; two of these were read locally, whereas the third slide was maintained for possible third party review in case of any discordance \u226550% between the first two readers. Microscopy results at the site guided subject management; any subject with persistent or recurrent parasitaemia during the follow-up period was treated with anti-malarial drugs according to local treatment guidelines and withdrawn from the study after documentation of clearance of parasitaemia.A blood dipstick-based test  was used for rapid diagnostic testing of For subjects who developed asexual parasitaemia after a period of clearance, paired blood blots from baseline and the time of recurrence were analysed to distinguish recrudescence from re-infection. Genotyping was performed by nested polymerase chain reaction (PCR) at two different laboratories, the Naval Medical Research Unit 2 in Jakarta, Indonesia (study 1134 only) and the Malaria Research and Training Centre in Bamako, Mali (both studies). In both studies, merozoite surface protein (MSP)-1, MSP-2, and the microsatellite CA1 gene loci (Su) were amplified at the Malaria Research and Training Centre. Recrudescence was defined as the reappearance of asexual blood-stage parasites of the same genotype as Day 0 parasites, whereas re-infection was defined as infection by a different genotype. Subjects were censored at the time of re-infection.P. falciparum CQ resistance transporter gene (pfcrt). For study 1134, molecular detection of the K76T mutation associated with CQ resistance was detected by means of a standardised real-time PCR-based diagnostic assay [pfcrt gene mutations. Based on differential melting-curve analysis, isolates were identified as CQ-sensitive isolates with a CVMNK haplotype or CQ-resistant isolates carrying a CVIET, SVMNT, CVMNT, or CVMET haplotype. For study 1155, nested PCR methods were performed for pfcrt (K76T) and P. falciparum multi-drug resistance protein-1  with the specific primer pairs followed by restriction fragment length polymorphism [Paired blood specimens, collected before treatment and at the time of recurrent asexual parasitaemia (treatment failure), were evaluated for mutations in the ic assay , 10. In P. falciparum parasitaemia and the investigator deemed discharge from the hospital appropriate. Peripheral blood smears for parasite counts were obtained at eight-hour intervals until clearance was demonstrated, then on Day 7 and weekly thereafter through Day 42 to monitor recrudescence. Vital signs, clinical signs and symptoms, adverse events (AEs), and concomitant medications were assessed for all subjects on each day of treatment  and at each post-therapy visit . Haematology and serum chemistry laboratory tests were performed at Baseline and Day 3 and at subsequent visits if clinically indicated. Haematology tests included red blood count, white blood count with differential, haemoglobin, haematocrit, and platelets. Serum chemistry tests included electrolytes, urea nitrogen, creatinine, aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase.All subjects were hospitalised and monitored until three consecutive blood smears were negative for asexual P. falciparum parasite clearance rate, adjusted for molecular testing (PCR-corrected) to differentiate recrudescence (true failures) from re-infection. Efficacy was assessed at Days 28 (primary endpoint) and 42 in the per protocol (PP) population. Treated subjects were excluded from the PP population if they did not meet the disease definition, did not receive all three days of study medication , or did not have a blood smear at the specified time point (unless due to recurrent parasitaemia before that time point). Subjects were assigned a response of eradicated if parasitaemia cleared within seven days after initiation of treatment and did not recur through the time point of interest. Failure was defined as not achieving clearance of asexual P. falciparum parasitaemia within seven days or, if after achieving clearance, PCR-corrected parasitaemia recurred.The primary endpoint was the asexual Secondary efficacy analyses included time required for asexual parasite clearance, fever clearance time, and assessment of the percentage of subjects with early and late treatment failures as defined by the World Health Organization .All subjects who received at least one dose of study medication were evaluated for safety. Subjects were monitored closely for clinical evidence of illness progression. All subjects with persistent or recurrent parasitaemia received therapy consistent with the local standard of care and were monitored until parasite clearance was documented. Other safety evaluations included AE and vital sign monitoring throughout the study, haematology and serum chemistry laboratory evaluations, and physical examinations.P. falciparum parasite clearance rates in the AZCQ 1,000\u00a0mg and MQ treatment arms at Day 28 in the PP population. A two-sided confidence interval (CI), using the appropriate confidence level, was constructed for the difference between treatment groups in asexual parasite clearance rates using normal approximation to the binomial, and also for within group rates. A CI was computed for the clearance rate within a treatment group using exact methods when no failures were observed. The confidence level in the final analysis for study 1134 required a small adjustment upward from 95 to 95.04% to prevent the overall false-positive rate from exceeding 0.05, due to the planned interim analysis of the primary efficacy outcome. There was no adjustment for centres in the analyses. Non-inferiority was to be concluded if the lower boundary of the CI for the difference in parasite clearance rates  was -10% or more. A non-inferiority margin of 10% was based on regulatory guidance available at the time of study conception. Clearance time comparisons up to Day 7 were generated using Kaplan-Meier estimates. Parasite clearance time was defined as the time in days from Baseline to the first of the three consecutive zero parasite counts. Fever clearance time was defined as the time in days from Baseline to the first of the two consecutive time points without evidence of elevated temperature.The primary efficacy analysis compared the PCR-corrected asexual Total sample size was determined based on the following assumptions: 80% power to show non-inferiority of the AZCQ 1,000-mg arm relative to the MQ arm, 85% of randomised subjects satisfying criteria for inclusion in the PP analysis at the Day 28 visit, and expected parasite clearance rates of 95% in the AZCQ 1,000-mg arm and 95% in the MQ arm.These studies were conducted in compliance with the Declaration of Helsinki, institutional review boards, informed consent regulation, and the International Conference on Harmonisation Good Clinical Practice guidelines. All local regulatory requirements were followed. The clinical protocol was conducted in accordance with FDA regulations. Pfizer Inc conducted the clinical monitoring of the study.A total of 467 subjects were randomised to study medication Figure\u00a0. The demThe AZCQ 500-mg regimen was stopped early in study 1134 on the recommendation of the data safety monitoring board after a review of data from studies conducted in South America and India demonstrated a dose response with lower efficacy rates for this regimen compared with AZCQ 1,000\u00a0mg. At that point, a total of nine subjects had been randomised to the AZCQ 500-mg regimen, of which six of seven subjects in the PP population had responded to therapy through Day 28  compared with 99.0% parasite clearance  in those who received MQ based on PCR-corrected results . Median time to resolution of fever was observed to be <0.5\u00a0days after starting therapy for subjects randomised to AZCQ 1,000\u00a0mg and 0.5\u00a0days for those given MQ.The median time to clearance of asexual Similar findings were observed in study 1155, whereas subjects in the PP population randomised to AZCQ 1,000\u00a0mg had 100% parasite clearance  compared with 99.1%  in those randomised to MQ  at Day 28. For corresponding PCR-uncorrected results, subjects in both the AZCQ 1,000-mg  and MQ  groups had a 99.1% parasite clearance. At Day 42, PCR-corrected parasite clearance remained at 100% for the AZCQ group  and 99.1% for the MQ group .The median time to clearance of parasitaemia was 44\u00a0hours for subjects given AZCQ 1,000\u00a0mg and 40\u00a0hours for those given MQ. Median time to resolution of fever was observed to be 1.5\u00a0days after starting therapy for subjects randomised to AZCQ 1,000\u00a0mg and 1\u00a0day for those given MQ, although the difference was not statistically significant.Two late parasitological failures (LPFs) were observed in study 1134 for subjects in the AZCQ 1,000-mg treatment group Table\u00a0, whereaspfcrt haplotype  of subjects randomised to AZCQ 1,000\u00a0mg and in 61.7% 71/115) of those randomised to MQ in study 1134, and in 70.8% (80/113) and 62.1% (72/116) of subjects, respectively, in study 1155 . A severe AE of vomiting was seen in one (0.4%) subjects given AZCQ 1,000\u00a0mg, whereas six severe AEs were observed in five (2.2%) subjects given MQ . The majority of all other AEs with AZCQ 1,000\u00a0mg and MQ were mild in severity.P. falciparum. In addition to similar clinical and parasitological outcomes, the AZCQ fixed-dose combination also achieved an overall efficacy rate of 99%, which is consistent with other treatment modalities recommended in treatment guidelines for this subject population [The fixed-dose combination of AZ 1,000\u00a0mg and a CQ 600-mg base was found to be non-inferior to MQ in two randomised studies conducted in Africa in adults with symptomatic uncomplicated malaria due to pulation . The meaA pre-specified non-inferiority margin of 10% was based on regulatory guidance at the time of study conception. Although this margin has been used by other phase III trials as recent as 2011, many trials of late have switched to a more conservative margin of 5% . Given tIn Zambia, the rate of clinical resistance to CQ was >70% in 2003 at the time of switching from CQ to artemether/lumefantrine. Based on this observation, it is clear that AZCQ displayed efficacy in areas with a high prevalence of CQ-resistant falciparum malaria.vs the other countries included in this study.Molecular marker resistance to CQ across these seven African countries between 2004 and 2007 ranged from approximately 21 to 96%. There was a trend to lower rates of CQ resistance in the clinical trial sites in Ghana and Zambia over this time period, which may be related to anti-malarial policies in these countries pfcrt CQ-resistant haplotype most frequently described in Africa has been CVIET, which is thought to have possibly been introduced from southeast Asia through India [The gh India . As prevgh India , 16.in vitro CQ resistance would be expected to be significantly lower than those observed on this combination therapy [While no CQ monotherapy control arm was included in this trial, the historical rates of clinical response to CQ in areas with significant  therapy . WhetherMoreover, anti-malarial immunity in malaria-endemic areas also can assist in parasite clearance and may play a role in the enhanced efficacy of the combination therapy, although innate immunity is equally likely to be effective in those subjects who received either AZ or CQ alone .Discontinuations due to AEs were infrequent and numbers were similar in each regimen. Pruritus was seen more frequently with the AZCQ combination, a well-described effect associated with the use of CQ in subjects with melanoderma . CentralFor a regimen to be considered appropriate as a therapy for malaria, it first must demonstrate that it is safe and effective in the target subject population. Other considerations, however, should be addressed with regard to the AZCQ combination. CQ is no longer recommended for the treatment of malaria in Africa due to the generally high rates of background CQ resistance and subsequent poor clinical response to therapy throughout the continent. The position of the World Health Organization is that all therapy for the treatment of malaria should be combination therapy, primarily combinations that include an artemisinin derivative . The comA limitation of this study was the recruitment of adults with symptomatic uncomplicated malaria only and thus, other target populations, including pregnant women, should be studied, as it is known that pregnant women are more susceptible to malaria compared with non-pregnant women within the same age group . The proA phase III trial  that evaluated a blended tablet of AZ and CQ for intermittent preventive treatment of malaria in pregnancy was stopped early, as an interim analysis indicated it was unlikely to meet the primary endpoint  defined as superiority of AZCQ over sulfadoxine/pyrimethamine in the proportion of patients meeting a sub-optimal pregnancy outcome . As the primary endpoint was analysed using the intent-to-treat population, lack of tolerability of the combination treatment may have resulted in a higher dropout rate leading to the inclusion of missing data as a failure in the interim analysis.New therapy for the prevention and treatment of malaria in the context of a broader campaign of public health interventions is, therefore, needed to help limit the morbidity and mortality associated with this disease.The results of this study indicate that among adults with symptomatic uncomplicated falciparum malaria in Africa, a fixed-dose combination of AZ 1,000\u00a0mg and a CQ 600-mg base once daily for three days resulted in Day 28 and Day 42 PCR-corrected parasitological clearance rates of \u226598%. AZCQ was non-inferior to treatment with MQ and was well tolerated.Additional file 1: Table S1: Median change from Baseline to last observation in laboratory values in study 1134. Description: The data in the table provides details of laboratory values from study 1134. (DOCX 38 KB)Additional file 2: Table S2: Median change from Baseline to last observation in laboratory values in study 1155. Description: The data in the table provides details of laboratory values from study 1155. (DOCX 35 KB)"}
+{"text": "Plasmodium falciparum may be threatened by parasites with reduced responsiveness to artemisinins. Among 298 ACT-treated children from Mbita, Kenya, sub-microscopic persistence of P. falciparum on day 3 post-treatment was associated with subsequent microscopically detected parasitaemia at days 28 or 42.The efficacy of ACT for treating malaria patients infected with pfcrt, pfmdr1, pfubp1 and pfap2mu were determined in the Mbita cohort before treatment, on days 2 and 3 after initiation of treatment and on the day of treatment failure.Parasite density post-treatment was measured by duplex probe qPCR at days 1, 2 and 3. DNA sequences of resistance-associated parasite loci P. falciparum on day 3 post-treatment was associated with subsequent microscopically detected parasitaemia at days 28 or 42. Parasites surviving ACT on day 2 or day 3 post-treatment were significantly more likely than the baseline population to carry the wildtype haplotypes of pfcrt  and pfmdr1 . In contrast, variant alleles of the novel candidate resistance genes pfap2mu  and pfubp-1  were significantly more prevalent post-treatment. No genetic similarities were found to parasites with reduced sensitivity to short-course artemisinin monotherapy, recently described in Cambodia.Among 298 ACT-treated children from Mbita, Kenya, sub-microscopic persistence of P. falciparum genotypes are more likely to survive ACT at sub-microscopic level, and contribute to onward transmission and subsequent patent recrudescence. These surviving parasites may have an important public health impact that has been overlooked. The possible role of both innate and acquired immune mechanisms in this phenomenon will be discussed.We conclude that, among treated children in western Kenya, certain multi-locus"}
+{"text": "Plasmodium falciparum infection. Some earlier studies point to a direct impact of Plasmodium falciparum infection on the electric conduction system of the heart. The aim of this study was to analyse infection- and drug-induced effects on the electric conduction system.While several anti-malarials are known to affect the electric conduction system of the heart, less is known on the direct effects of Children aged 12\u00a0months to 108\u00a0months with severe malaria were included in Kumasi, Ghana. In addition to basic demographic, clinical, biochemical and parasitological, biochemical data were measured data upon hospitalization (day 0) and 12-lead electrocardiograms were recorded before (day 0) and after (day 1) initiation of quinine therapy as well as after 42 (\u00b13) days.A total of 180 children were included. Most children were tachycardic on day 0 but heart rate declined on day 1 and during follow up. The corrected QT intervals were longest on day 1 and shortest on day 0. Comparison of QT intervals with day 42  after stratification for age demonstrated that in the youngest (<24\u00a0months) this was mainly due to a QT shortage on day 0 while a QT prolongation on day 1 was most pronounced in the oldest (\u226548\u00a0months). Nearly one third of the participating children had measurable 4-aminoquinoline levels upon admission, but no direct effect on the corrected QT intervals could be shown.P. falciparum infection itself can provoke changes in the electrophysiology of the heart, independent of anti-malarial therapy. Especially in young - thus non immune - children the effect of acute disease associated pre-treatment QT-shortage is more pronounced than quinine associated QT-prolongation after therapy. Nevertheless, neither malaria nor anti-malarial induced effects on the electrophysiology of the heart were associated with clinically relevant arrhythmias in the present study population.Severe  Plasmodium falciparum infection on the electric conduction system of the heart is incomplete. Like other febrile diseases, malaria increases the sympathetic tone in patients, leading to an acceleration of the electric conduction and repolarization of the heart, which can be shown as shortening of the QT intervals in electrocardiographic recordings [P. falciparum infection on the electric conduction system of the heart, which belongs to one of the most severely parasitized organs in humans [et al., for example, found a correlation between parasitaemia and corrected QT prolongation in Gambian children with uncomplicated falciparum malaria [P. falciparum malaria, found abnormal ECG findings in 14.3 per cent of all patients, including T-segment or T-wave alterations in 15 patients and delayed conduction in eight patients [The systematic assessment of the effect of severe cordings . Howevern humans . Von Sei malaria . Anotherpatients .P. falciparum malaria, such as quinine and halofantrine, are associated with an alteration of the electrophysiology of the heart, in particular provoking QT prolongation [In addition to potential malaria-related effects, several drugs used for the treatment of ongation ,9. This This study was designed to measure intra-individual ECG changes at three major points: Day 0 representing malaria without drug effects, day 1 representing malaria and drug effects as well as day 42 representing the healthy status without malaria and anti-malarial drugs. In addition, potential influence of pre-hospitalization medications, age as well as signs and symptoms defining severe malaria on cardiac electrophysiology were assessed.The study was conducted at the Paediatric Emergency Unit of the Komfo Anokye Teaching Hosptial (KATH) in Kumasi, Ghana, between March 2009 and October 2009. After written consent was obtained by parents or legal guardians children aged 12\u00a0months to 108\u00a0months with signs of severe malaria and a positive rapid diagnostic test (OptiMAL\u00ae) were screened. Severe malaria was defined as presence of asexual blood stage parasites, symptoms and conditions according to WHO criteria [On admission (day 0) the patients were clinically examined and parameters, such as blood pressure, heart rate, respiratory rate as well as body temperature were recorded and signs of severe malaria were ascertained. Patients were treated with parenteral quinine according to national guidelines (20\u00a0mg/kg loading dose followed by 10\u00a0mg/kg twice daily) for seven days or until they were able to receive orally administrated artemether-lumefantrine. Patients were followed-up on days 1 and 42 (\u00b13).et al. was chosen as it more effectively removes the rate dependence of the adjusted QT in children [A standard 12-lead electrocardiogram (ECG) was conducted on day 0 before the start of quinine therapy, on day 1 after treatment initiation and on day 42. ECGs were recorded at a chart rate of 50\u00a0mm/sec. All ECG results were printed, scanned and reviewed using the software GraphicConverter X (Version 6.7.3). An electronic ruler was used to measure RR, PQ, QRS, and QT. To minimize variability due to lead selection, QT-intervals in lead II and V5 were measured and the longer interval was selected for further analysis ,12. U-wachildren . For comBlood samples were taken on day 0 and on day 42. Plasmodium falciparum malaria was ascertained from Giemsa-stained thick and thin films. Blood chemistry parameters were measured by the I-STAT\u00ae analysing system including but not limited to creatinine, glucose, pH, lactate and base excess. Concentrations of 4-aminoquinoline drugs  were determined from urine samples obtained on day 0 using high-performance liquid chromatography (HPLC) with ultra-violet (UV) detection. The method has already been published with some minor deviations . In brieThe study was approved by the ethics committee of the Kwane Nkrumah University of Science and Technology (KNUST), Kumasi. Statistics software SPSS 18.0 was used to analyse data. Statistical significance was defined a p\u2009<\u20090.05. Dependent t-test  was used for comparing findings between day 0, day 1 and day 42. Independent t-test was used to analyse differences in ECG parameters in subgroups.Severe malaria was ascertained and complete day 0-data sets were available for 180 (82%) of 220 initially screened children, 103 (57%) were males. The median age was 36\u00a0months (range 12 to 108\u00a0months), 125 of the 180 (70%) children were available for follow up ECG assessment on day 42. Criteria for severe malaria were prostration (158 cases), multiple convulsions (94 cases), hyperlactataemia (63 cases), jaundice (55 cases), metabolic acidosis (55 cases), hyperparasitaemia (53 cases), coma (50 cases), severe anaemia (41 cases), haemoglubinuria (36 cases), respiratory distress (22), circular collapse (14), hypoglycaemia (10 cases), renal impairment (3 cases) and abnormal bleeding (1 case). Five children (3%) died. Demographic, clinical and biochemical characteristics are shown in Table\u00a0Most children had moderate to severe tachycardia on day 0 but heart rates (HR) declined on day 1 and during follow up Table\u00a0. The corOf the 153 cases in which urine concentrations of 4-aminoquinoline drugs were measured on the day of admission 50 (33%) were positive for one or more than one of these drugs. (N-desethyl)-chloroquine was detectable in 28 of the 50 cases (56%), (N-desethyl-) amiodiaquine was detectable in 37 of the 50 cases (74%), thus 15 cases (30%) were positive for (N-desethyl-) chloroquine and (N-desethyl-) amiodiaquine.HR was significantly slower in children who had taken 4-aminoquinolines prior to admission compared to those who had not . There was no significant difference in the mean of hQT between children with or without 4-aminoquinoline levels .The effect of the pre-hospitalization intake of 4-aminoquinolines was further stratified on hQT according to age Figure\u00a0C and D. P. falciparum infection on the conduction system of the heart, the impact of severe malaria defining conditions on heart rate and hQT intervals upon hospitalization was analysed  as well as cardiac arrhythmias have the potential to influences cardiocirculatory function and, therefore, to gain clinical relevance. Children with severe malaria were tachycardic on the day of admission but the heart rate declined with the onset of anti-malarial therapy to a level considered as normal on day 42 reflecting the state of acute febrile illness and subsequent recovery.et al., who also saw a relative shortening of pre-treatment QT intervals in children with uncomplicated malaria [NA) and thereby widening QRS complex, an effect earlier described in other studies [Corrected QT intervals (hQT) were shorter before therapy than during therapy and also shorter before therapy compared to after cure. The hQT prolongation on day 1 was probably at least partially related to quinine [ malaria . The QRS studies ,19 and aet al. observed that QRS prolongation after the onset of quinine therapy was most pronounced in children younger than 24\u00a0months [Stratification revealed some age-specific effects on both heart rate and repolarization. HR was highest in the youngest and lowest in the oldest children on day 0, 1 and 42, respectively, resembling the physiological decline of heart rates with age. The age stratified hQT intervals on day 42, reflecting the healthy status, were comparable in all age groups confirming the validity of the applied correcting formula for the QT interval. Nevertheless, there are differences in the characteristics of the hQT intervals between the different age groups during acute disease: In the groups 24 to <48\u00a0months of age and >48\u00a0months of age there was a significant prolongation of hQT on day 1 after the start of quinine therapy in comparison to day 0 as well as to day 42. In the group of children younger than 24\u00a0months of age, on the other hand, there was no significant difference in hQT intervals between day 1 and day 42  on parasite density in Ghanaian children with severe malaria with the highest NO levels in the youngest children . NO is kA relatively high percentage of children had taken 4-aminoquinoline drugs prior to admission. Despite the change of national policies in 2009 an unexpected high portion was still treated with chloroquine. Chloroquine is considered to potentially prolong QT interval ,24, but Of the symptoms and characteristics of severe malaria, some conditions seem to influence the electrophysiology of the heart prior to quinine therapy while others do not. Metabolic acidosis - present in one-third of all children \u2013 as well as respiratory distress and hyperlactataemia were associated with a shortening of hQT intervals. Respiratory distress as a clinical sign of metabolic acidosis was present in most of the acidotic children and both respiratory distress and acidosis are considered as independent predictors of fatal outcome in falciparum malaria ,26. How The study had a number of limitations: a problem involved the determination of the corrected QT intervals: Many formulas have been proposed to correct QT intervals for the heart rate ,15,30,31In conclusion, the findings suggest that severe malaria itself is associated with changes in the electrophysiology of the heart, independent of anti-malarial therapy. Especially in young - thus non-immune \u2013 children acute malaria-associated QT-shortage seemed to be more prominent than anti-malarial-associated QT-prolongation. Nevertheless, it remains unclear whether these ECG-changes are of high clinical relevance as cardiac arrhythmias and subsequent circulatory dysfunction does not seem to be a major contributor to unfavourable outcomes in severe malaria.cQT: Bazett corrected QT interval; ECG: Electrocardiogram; hQT: Wernicke corrected QT interval; HR: Heart rate; KATH: Komfo Anokye Teaching Hosptial; KNUST: Kwame Nkrumah University of Science and Technology; Min: Minute; ms: Milliseconds; NO: Nitric oxide; SD: Standard deviation.The authors declare that they have no competing interests.DP and SH participated in patient recruitment and data collection. LR and DP performed the statistical analyses and drafted the manuscript. JNH participated in statistical analysis. JPC and SBN conceived of the study and participated in its design and coordination and helped to draft the manuscript. JB performed the urine sample analyses. TF, DA, JS, JFH, PF, CT and GDB significantly contributed to the protocol development and/or study implementation. All authors read and approved the final manuscript."}
+{"text": "Regular efficacy monitoring of these two combinations is conducted every 2 or 3\u00a0years. This paper reports the latest efficacy assessment results and the investigation of mutations in the Plasmodium falciparum, were treated with either AL  or ASAQ . The WHO standard protocol for anti-malarial treatment evaluation was used. The primary end-point was 28-day adequate clinical and parasitological response (ACPR), corrected to exclude reinfection using polymerase-chain reaction (PCR) genotyping.The study was conducted in 2012\u20132013 on three sentinel sites of Togo . Children aged 6\u201359\u00a0months, who were symptomatically infected with k13 propeller domain, only 9 (1.8\u00a0%) mutations were reported, three in each site. All mutant parasites were cleared before day 3. All day 3 positive patients were infected with k13 wild type parasites.A total of 523 children were included in the study. PCR-corrected ACPR was 96.3\u2013100\u00a0% for ASAQ and 97\u2013100\u00a0% for AL across the three study sites. Adverse events were negligible: 0\u20134.8\u00a0% across all sites, for both artemisinin-based combinations. Upon investigation of mutations in the The efficacy of AL and ASAQ remains high in Togo, and both drugs are well tolerated. ASAQ and AL would be recommended for the treatment of uncomplicated malaria in Togo. Plasmodium falciparum malaria transmission is stable, but with seasonal outbreaks which are equatorial-type in the south and tropical-type in the north of the country. Since 2005, the National Malaria Control Programme (NMCP) has recommended two different forms of artemisinin-based combination therapy (ACT) for the treatment of uncomplicated malaria: artemether\u2013lumefantrine (AL) and artesunate\u2013amodiaquine (ASAQ) [Despite several decades of malaria control efforts, malaria-related mortality and morbidity remains a public health problem in sub-Saharan countries , 2. In Te (ASAQ) . Severe e (ASAQ) . On preve (ASAQ) .P. falciparum kelch (k13) protein have recently been linked with artemisinin resistance [At a global level, increases in the availability and use of ACT, together with the increased use of insecticide-treated bed nets, have substantially reduced malaria burden. Between 2000 and 2015, malaria incidence rates decreased by 37\u00a0% globally, and by 42\u00a0% in Africa. During this same period, malaria mortality rates fell by 60\u00a0% globally and by 66\u00a0% in the African Region \u201312. Poinm kelch k protein k13 propeller domain.This article reports the results of the latest ACT efficacy evaluation carried out in 2012\u20132013 on three sentinel sites, and results from the investigation of mutations in the This evaluation was conducted in Togo on three of the five sentinel sites designated for the surveillance of anti-malarial treatments Fig.\u00a0: (1) CacP. falciparum malaria, conducted according to the World Health Organization (WHO) standard protocol [This was a prospective study of the clinical and parasitological efficacy of ASAQ and AL for the treatment of uncomplicated protocol .The sample size calculation was based on an assumed efficacy of 95\u00a0% for both, artemisinin-based combinations a 10\u00a0% significance level and a power of 95\u00a0%, with 20\u00a0% in addition, to account for patients who are likely to be either lost during follow-up, withdraw or be excluded after detection of reinfection with PCR correction , 16.P. falciparum mono-infection . Children with severe malaria symptoms according to the WHO case definition, and symptoms of severe malnutrition and chronic diseases, or with mixed Plasmodium infection were excluded.Eligible patients were children aged between six and 59\u00a0months, presenting to the participating health facilities with fever or a history of fever  in the last 24\u00a0h, who had microscopically-confirmed \u00ae, Novartis Pharma, Switzerland) was administered as 20\u00a0mg artemether/120\u00a0mg lumefantrine fixed-dose combination tablets twice daily during 3\u00a0days according to body weight: one tablet for 5\u201314\u00a0kg and two tablets for 15\u201324\u00a0kg. ASAQ , artesunate (25 or 50\u00a0mg) and amodiaquine (67.5 or 135\u00a0mg), were administered once daily during 3\u00a0days according to body weight: 25\u00a0mg artesunate/67.5\u00a0mg amodiaquine tablets for 5\u20138\u00a0kg and 50\u00a0mg artesunate/135\u00a0mg amodiaquine tablets for 9\u201317\u00a0kg. As antipyretic, 60\u00a0mg/kg of Paracetamol per day during the first 3\u00a0days (day 0 to day 2) was given systematically to every child included in the study.Patients were randomly assigned to receive AL or ASAQ, with all doses administered under medical supervision. AL . Asexual parasitaemia was determined from Giemsa-stained thick blood smears against the number of parasites per 200 white blood cells on day 0 and 1000 at follow-up assessments, based on a putative count of 6000 white blood cells per microlitre of blood. Gametocytes were similarly enumerated [Children enrolled in the cohort received drug treatment on days 0, 1 and 2, with follow up on days 3, 7, 14, 21 and 28. Adverse events were screened and recorded at each visit. Clinical examination, including fever assessment, was performed on days 1, 2, 3, 7, 14, 21 and 28. Capillary blood was taken for parasite counts and assessment of haemoglobin levels were performed at screening (day 0) and at days 2, 3, 7, 14, 21 and 28. Haemoglobin level was measured using a point-of-care testing: the HemoCuemsp-1), merozoite surface protein-2 (msp-2), and glutamate-rich protein (glurp) polymorphism, as per WHO methods [Dried blood spots were obtained for PCR at enrolment (day 0) and on follow-up days 1, 2, 3, 7, 14, 21 and 28. In the case of parasitaemia detected after day 7, to distinguish between recrudescence and reinfection, PCR genotyping was performed on paired dried blood spots to determine polymorphisms in merozoite surface protein-1  protocol followed by Sanger sequencing using primers specific to P. falciparum. The amplicon used for sequencing covered 740\u00a0bp, which included the k13 propeller domain [To assess the treatment effect on parasites mutations that modulate treatment response, mutations in stigated , 13. TheTreatment outcomes were classified based on parasitological and clinical outcomes assessment as recommended by the WHO . TherapeThe primary outcome measure was ACPR, corrected for reinfection using PCR genotyping at day 28. Patients were excluded from the analysis for the following reasons: lost to follow-up, protocol violations, treatment failure due to reinfection, or type of treatment failure could not be determined using PCR. Severe malaria cases developed during follow-up were treated with parenteral quinine according to the NCMP recommendations.\u00ae Excel-based applications [Data management and analysis were completed with Epi Info 6.0.4 adapted to the WHO\u2019s Microsoftications  that ena3, 2 (0.2\u00a0%) for parasitaemia\u00a0>250,000 parasites/mm3, 3 (0.2\u00a0%) for signs of severe malaria, 11 (0.9\u00a0%) for mixed P. falciparum and Plasmodium malariae or Plasmodium ovale infection, 6 (0.5\u00a0%) for parents refusal to consent, and 7 (0.6\u00a0%) for children refusal to take medicines.Among 1239 children with fever screened, 523 were included in the study  and 7 (2.7\u00a0%) cases of mild adverse events, respectively in the ASAQ and AL cohorts Table\u00a0.Table\u00a03Ak13 propeller domain. Overall, 491 (98.2\u00a0%) were wild type or presented with synonymous mutations. Only 9 (1.8\u00a0%) mutations were reported, three in each site: S522M, A578S and C532S in Lom\u00e9; S522C, A578S in Sodok\u00e9; and S522C and A578S (n\u00a0=\u00a02) in Niamtougou. All the patients carrying mutant parasites cleared their parasites before day 3. In addition, all day 3 positive patients were infected with k13 wild type parasites.Among the 523 patients, 500 isolates were successfully sequenced for the Following the WHO guidelines, the NMCP of Togo is recommending AL and ASAQ for the treatment of uncomplicated malaria, since 2005. A monitoring of these two artemisinin-based combinations is conducted every 2 or 3\u00a0years to provide timely information on trends of their efficacy and safety to enhance evidence-based decision making by the NMCP .The results showed that the two combinations induced fever clearance as quickly as day 1 following the treatment for more than 50\u00a0% of participants across all study sites. Complete parasite clearance was observed by day 3 for all participants on two sentinel sites (Sokod\u00e9 and Lom\u00e9). Moreover, the gametocytocidal effect was observed by the rapid reduction in gametocytes, as early as day 1 for AL and from day 3 for ASAQ. With ASAQ, the initial increase in the mean gametocytaemia before its rapid decrease from day 3 suggests the appearance of new gametocytes following initiation of the treatment. Nevertheless, complete clearance was achieved by day 28 for both combinations. Parasites and gametocytes clearances are consistent with the significant improvement of haemoglobin levels following treatment.Beside fever, parasite and gametocytes clearance, and improvement of haemoglobin level, efficacy results obtained in our study were also very satisfying. The PCR-corrected therapeutic efficacy of the two forms of artemisinin-based combinations recommended by the NMCP for uncomplicated malaria cases management in Togo remains above the 95\u00a0% efficacy threshold recommended by the WHO . In addiStudies conducted in the same settings in Togo between 2005 and 2009 showed the same significant improvement of haemoglobin levels following treatment. Moreover, the gametocytocidal effect of ACT was observed by the initial, rapid reduction in gametocytes. They also showed a high therapeutic efficacy: adequate clinical and parasitological response (ACPR), corrected for reinfection by PCR genotyping was about 95\u00a0% on all sentinel sites. In fact, the PCR-corrected 28-day cure rates using the per-protocol analysis were between 94\u2013100\u00a0% for ASAQ and 96\u2013100\u00a0% for AL .The efficacy assessment of ACT has also shown a high efficacy level of the combinations in other West Africa countries. In Ghana, a bordering country of Togo, Abuaku et al.  in a stuIn Benin, another bordering country, a 42-day therapeutic efficacy study of AL conducted by Ogouy\u00e8mi-Hounto et al.  in 2014 Results of this study are close to those of other similar studies conducted in the sub-Saharan African region during the last 5\u00a0years. These studies also showed high efficacy of ACT adopted in the area to treat uncomplicated malaria. Mekonnen et al.  in a stuAlthough such high levels of ACT efficacy are found in sub-Saharan Africa more than 10\u00a0years after its adoption in the region, there is a concern of emergence of artemisinin resistance in Africa. So far, artemisinin resistance is confined and has been detected in five countries in the Greater Mekong Subregion  , 6. In tP. falciparum malaria for children in Togo. No evidence of the emergence of artemisinin resistance in Togo was found upon investigation of mutations in the k13 propeller domain. The use of these two combinations (ASAQ and AL) adopted by health authorities, could be continued for the treatment of uncomplicated malaria in Togo.Both artesunate\u2013amodiaquine (ASAQ) and artemether\u2013lumefantrine (AL) have been shown to be safe and highly effective in the treatment of uncomplicated"}
+{"text": "Plasmodium falciparum malaria, but the effects of repeated, long-term use are not well documented. We conducted a 2-year randomized, open-label, longitudinal, phase IV clinical trial comparing the efficacy and safety of fixed-dose ASAQ and AL for repeated treatment of uncomplicated malaria in children under 5 years at Nagongera Health Centre, Uganda. Participants were randomized to ASAQ or AL and all subsequent malaria episodes were treated with the same regimen. 413 children were enrolled and experienced a total of 6027 malaria episodes . For the first malaria episode, the PCR-corrected-cure rate for ASAQ (97.5%) was non-inferior to that for AL . PCR-corrected cure rates for subsequent malaria episodes that had over 100 cases (episodes 2\u201318), ranged from 88.1% to 98.9% per episode, with no clear difference between the treatment arms. Parasites were completely cleared by day 3 for all malaria episodes and gametocyte carriage was less than 1% by day 21. Fever clearance was faster in the ASAQ group for the first episode. Treatment compliance for subsequent episodes (only first dose administration observed) was close to 100%. Adverse events though common were similar between treatment arms and mostly related to the disease. Serious adverse events were uncommon, comparable between treatment arms and resolved spontaneously. Anemia and neutropenia occurred in <0.5% of cases per episode, abnormal liver function tests occurred in 0.3% to 1.4% of cases. Both regimens were safe and effective for repeated treatment of malaria.The safety and efficacy of the two most widely used fixed-dose artemisinin-based combination therapies (ACT), artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL) are well established for single episodes of uncomplicated NCT00699920Current Controlled Trials  Since 2001, artemisinin-based combination therapy (ACT) is recommended by the World Health Organization (WHO) for the first-line treatment of uncomplicated malaria Until 2007, the combination of artemether and lumefantrine (AL) was the only fixed-dose ACT widely available in malaria-endemic countries. A fixed-dose combination of artesunate and amodiaquine (ASAQ Winthrop [ASAQ]), developed by a partnership between the Drugs for Neglected Diseases initiative (DNDi) and Sanofi, was prequalified by the WHO in 2008 and included in the List of Essential Medicines in 2011 P. falciparum malaria in children less than 5 years old. The study was conducted between 2nd June 2008 and 2nd June 2010 at Nagongera health centre, in Tororo district in Eastern Uganda, an area of intense perennial malaria transmission  We conducted a phase IV study to assess the efficacy and safety of repeated use of ASAQ and AL for the treatment of recurrent uncomplicated This was a randomized, open-label, longitudinal, phase IV clinical trial comparing the efficacy and safety of ASAQ and AL for the repeated treatment of uncomplicated malaria during a 2 year study period, in children less than 5 years old. The study was approved by the Faculty of Medicine Makerere University Research and Ethics Committee, the Uganda National Council of Science and Technology, and the Uganda National Drug Authority. The study was carried out in accordance with International Conference on Harmonization Guidelines for Good Clinical Practice and the Declaration of Helsinki. Caregivers of children included in the trial were explained the study objectives and procedures, and written informed consent was obtained from them. The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Plasmodium falciparum, while Anopheles gambiae s.s., and to a lesser extent Anopheles fenestus, are the major vectors The study was conducted between June 2008 and June 2010 at Nagongera Health Centre, Nagongera sub-county, Tororo district, Uganda. Nagongera is a rural sub-county located in Tororo district in eastern Uganda with a population of 37,715 people in 2009\u20132010 P. falciparum mono infection with parasitemia \u22652000/\u00b5L 3) had fever (axillary temperature \u226537.5\u00b0C) or history of fever within the previous 24 hours, 4) body weight \u22655 kg, and 5) hemoglobin (Hb) concentration \u22655 g/dL. Children were excluded from the study if they had a known allergy to one of the study medications, if they had a history of hepatic or hematological impairment during treatment with amodiaquine, a history of cardiac disease, or had a concomitant febrile illness. Children were excluded if they had severe malaria or danger signs of severe malaria (a recent history of convulsions (1 to 2 within 24 h), unconsciousness, lethargy, inability to drink or breast feed, vomiting, prostration); severe concomitant disease; a known disturbance of electrolyte balance ; were receiving a medication metabolized by or that inhibits cytochrome CYP 2D6; were receiving a medication known to prolong the QTc interval; or had received artesunate-amodiaquine or artemether-lumefantrine combination treatment within the previous 2 weeks. Children who experienced a new malaria attack at least 14 days after the previous one were included for subsequent follow up if they had a) a repeat infection with Plasmodium falciparum, b) fever (axillary temperature \u226537.5\u00b0C) or history of fever within the previous 24 hours, c) body weight \u22655 kg, and d) hemoglobin (Hb) concentration \u22655 g/dL, e) no severe malaria or danger signs. Participants were treated with the same drug and followed up for 42 days.Children were included in the study if they were 11) aged 6 to 59 months 2) had uncomplicated malaria with asexual P. falciparum malaria, children were referred to a study nurse responsible for the treatment group assignment and allocation of study medications but who was not responsible for patient assessment. A randomization list was computer generated by a statistician in blocks of 4 and provided to the nurse who randomized the children 1\u22361 to either the ASAQ or AL treatment group. The randomization codes were provided in an opaque list corresponding to the patient study number. During randomization, the study nurse scraped the opaque case corresponding to the study number in order to determine the allocated treatment. Only the study nurse had access to the sealed treatment randomization list. However, given the variation in appearance, taste and dosing, the other research staff may have known to which arm children were assigned. Supervised treatment allocation and administration of medications was performed by the study nurse who administered all medications orally as follows: ASAQ  was administered once daily for 3 days as 1 tablet of 25 mg artesunate/67.5 mg amodiaquine for body weights \u22655 to <9 kg, 1 tablet of 50 mg artesunate/135 mg amodiaquine for weights of \u22659 to <18 kg, or 1 tablet of 100 mg artesunate/270 mg amodiaquine for weights of \u226518 to <36 kg. AL  was administered twice daily for 3 days as 1 tablet of 20 mg artemether/120 mg lumefantrine for body weights <15 kg, 2 tablets for weights \u226515 to <25 kg, or 3 tablets for weights of \u226525 to <35 kg. If the child vomited or rejected the medication within the monitoring period, the same dose was re-administered. The administration of all doses for the first episode was directly observed. Patients were kept for 30 minutes after treatment and the dose was re-administered if vomiting occurred. All patients were provided with a 3-day supply of paracetamol for the treatment of febrile symptoms. Patients with hemoglobin <10\u20220 g/dL were treated with ferrous sulphate for 14 days and given mebendazole if they were over one year of age and had not been treated in the previous 6 months.Following the initial diagnosis of uncomplicated Participants included for the second and subsequent follow ups received the same treatment as for the first malaria episode, but treatment was unsupervised after the first dose. If vomiting occurred within 30 min, the child's caregivers were asked to return the child to the study clinic to receive the same treatment. For all episodes, if the child vomited or rejected the medication a second time, a replacement treatment  was administered. All patients who developed severe/complicated malaria during active follow-up were treated with parenteral quinine as per the national guidelines.At enrolment, children's parents or guardians were asked about use of other medications and presence of common symptoms. Axillary temperature and weight were recorded, and a physical examination performed. At follow up visits scheduled for Days 1, 2, 3, 7, 14, 21, 28, and 42, a standardized history was collected and physical examination performed. At each visit, including unscheduled ones, blood samples were collected by venipuncture or finger prick sampling on days 0, 1, 2, 3, 7, 14, 21, 28, and 42 for the first episode of malaria and on days 0, 3, 7, 14, 21, 28, and 42 for subsequent episodes. Aliquots (\u223c25 \u00b5L) were collected onto Whatman Gr3 filter paper, air dried, and stored in sealed sample bags at ambient temperature with a desiccant. The Hb concentration was assessed in blood samples collected on days 0, 3, 7, and 28. If Hb concentration was abnormal (<10 g/dL) on day 7, it was assessed again on day 14. If it was abnormal on day 28, it was assessed again on day 42. Blood hematology and biochemistry was performed on days 0, 7, and 28. Hematology and biochemistry measurements were repeated on day 14 and 42 if results were abnormal on day 7 and day 28 respectively. Parasitemia was assessed in all blood samples by thick and thin blood smears, and malaria parasite genotype was assessed for all patients with outcomes classified as late clinical failure or late parasitological failure. Spleen palpation was performed at each visit for each malaria episode and was scored according to Hackett's classification. Patients' parents were encouraged to attend the clinic at any time if the child was ill. Patients not attending the clinic at the scheduled visits were actively followed up at home. Each patient was followed up actively after recruitment for 42 days (first follow up) and then passively for 2 years. If within the 2-year period the patient presented with a new malaria attack, he/she was enrolled again in the study and treated with the same drugs.Thick and thin blood smears were stained with 2% Giemsa for 30 minutes. Parasite density was determined by reading the thick blood smear and counting the number of asexual parasites per 200 white blood cells (WBCs), assuming a WBC count of 8,000/\u00b5l. Slides were considered negative if no parasite was detected after reading 100 high-powered fields. Presence of gametocytes was also recorded. Thin blood smears were reviewed for non-falciparum infections. Two microscopists independently read all the slides and parasite densities were calculated by averaging the two counts. Readings with discordant results  were re-examined by a third microscopist; parasite density was calculated by averaging the two closest densities while the final parasite species was determined by two concordant reads. Hemoglobin measurements were made using a portable spectrophotometer Hemocue Hb 201 or using a Humacount 5 hematology counter .MSP1 and MSP2) and the glutamate rich protein (GLURP) were amplified using nested PCR as described previously Molecular genotyping was carried out at the Institut de recherche biom\u00e9dicale des arm\u00e9es-IRBA, Ex-IMTSSA, Marseille, France on samples collected from patients with late treatment failure to discriminate between a recrudescent and a new infection. Parasite deoxyribonucleic acid (DNA) genotype was determined for samples collected on day 0 and on the day of any recurrent parasitemia. For first malaria episodes, parasite DNA was prepared from filter papers by Chelex extraction , and for subsequent episodes, DNA was extracted using a MagMAX DNA Multi-Sample Kit . Selected regions of the merozoite surface proteins 1 and 2  and compared using Genemapper 4.0 software (Applied Biosystems). Genotyping patterns were compared using GelCompar II software .MSP1, MSP2, and GLURP). Children were considered to have a new infection if the length polymorphism was different between the sample collected at day 0 and the sample collected on the day of recurrent parasitemia for at least 1 marker. Results were considered indeterminate if DNA amplification was unsuccessful for DNA samples collected on either day 0 or the day of recurrent parasitemia.As described previously The primary objective of the study was to demonstrate the non-inferiority of PCR-adjusted adequate clinical and parasitological response at D28 of ASAQ versus AL during the first malaria episode. The secondary objectives of the study were: 1) to compare the two treatment regiments in terms of D42 efficacy, parasitological and fever clearance, clinical and biological tolerability and evolution of gametocyte carriage during the first malaria episode (observed treatment administration) 2) to compare the two groups of treatment in terms of fever and parasite clearance at D3 and 3) to compare the two groups of treatment in terms of D28 and D42 clinical and parasitological effectiveness, clinical and biological tolerability, fever and parasite clearance at D3, evolution of gametocyte carriage and treatment compliance during the second and following malaria episodes (non observed administration). During the total follow up of the cohort, the study aimed to compare the two groups of treatment in terms of treatment incidence density, clinical and biological tolerability, impact on anemia and Hackett score.Treatment outcomes were classified according to the WHO guidelines for areas of intense transmission as adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF), and late parasitological failure (LPF) The primary efficacy endpoint was the clinical and parasitological outcome at day 28 adjusted by genotyping for the first malaria episode. Secondary efficacy endpoints included clinical and parasitological outcome at day 28 adjusted by genotyping for subsequent episodes, and clinical and parasitological outcome at day 28 unadjusted by genotyping, clinical and parasitological outcome at day 42, unadjusted and adjusted by genotyping, fever and parasite clearance, gametocytaemia  at day 7, 14, 21, 28 and 42, hemoglobin changes between day 0 and day 28, and incidence of adverse events for all episodes. The treatment incidence density during the total duration of the study was compared.3; blood alanine amino transferase concentration>8\u00d7 upper limit of the normal range (0\u201345 IU/L); and blood alanine amino transferase concentration>3\u00d7 upper limit of the normal range associated with a total bilirubin concentration>2\u00d7 upper limit of the normal range (0.2\u20131.0 mg/dL). Serious adverse events (SAEs) were defined as any untoward medical occurrences that resulted in death, were life-threatening, required inpatient hospitalization or prolongation of existing hospitalization, resulted in persistent or significant disability or incapacity, were congenital anomalies or birth defects, or were considered medically important. Hospitalization during the first 3 days of the first malaria episode for social reasons  was not considered an AE or SAE.Adverse events (AEs) were defined as any untoward medical occurrence  or clinical investigation in a treated subject, irrespective a causal relationship with the treatment . Treatment-emergent AEs (TEAEs) were defined as AEs that developed or worsened within the study period (42 days) for each episode. All events were graded by the investigator by severity  using the WHO toxicity grading scale for determining the severity of adverse events and National Institute of Health paediatric toxicity tables, May 2001 guidelines, and the relationship to study treatment was recorded. Physical or clinical symptoms at each scheduled and unscheduled follow-up visit were recorded by the study investigator as an AE if they appeared or worsened during the study. Laboratory and vital sign abnormalities were recorded as AEs only if they were symptomatic, required corrective treatment, led to discontinuation, fulfilled a severity criterion, or were defined as an adverse event of special interest (AESI). AESIs included neutropenia as defined by a neutrophil count <400/mmTreatment of the first episode was supervised. For subsequent episodes, for which only the first treatment dose was supervised, blister packages were returned at the end of the third day of treatment by the parent or guardian. Tablets remaining in packages were counted by the study nurse. Treatment compliance was calculated as the percentage of tablets actually taken vs. tablets that should have been taken.Because of the high number of malaria infections observed during this study, the study stakeholders, in consultation with the ethical review board, decided in July 2009 to provide each child's family with an ITN . Parents were instructed by a study nurse to place an ITN over the child's bed.In May 2010, children's families were surveyed about their use of the provided ITNs by a study investigator and, when needed for translation, a community healthcare worker. Briefly, parents or legal guardians were asked about the use of ITNs and the child's sleeping habits. Also, as part of the survey, correct placement of the ITNs in the home over the bed of the child was checked.The sample size was calculated using NQuery version 4.0  based on the non\u2013inferiority testing of ASAQ versus AL for the first attack (supervised intakes). The acceptable non-inferiority margin (\u0394) in proportion of success between ASAQ and AL was chosen at 5%. The test of non inferiority between ASAQ and AL was based on the unilateral confidence interval method with \u03b1\u200a=\u200a5%, and \u03b2\u200a=\u200a20%. According to previous studies in East Africa, Statistical analysis was performed using SAS version 8.2 . All statistical tests were performed at a 5% significance level and were two tailed except for non-inferiority tests. Continuous variables were compared using Student's t-test for normal distributions or a non-parametric Wilcoxon test for distributions that were not normal. Categorical variables were compared using the chi-square test or Fisher's exact test. The intent-to-treat population was defined as all patients who received at least one dose of study medication and who did not have two rejections or vomiting; the per-protocol population comprised all ITT population children who completed the study without major protocol violation and the safety population comprised all patients who received at least one dose of study treatment.Plasmodium falciparum infections detected on the basis of genotyping were also censored.Parasite densities were normalized using logarithmic transformation. Risks of treatment failure were estimated using the Kaplan-Meier product limit formula. Data were censored for patients who did not complete follow-up or were reinfected with non-falciparum species. For the PCR-adjusted ACPR, new The primary objective, non-inferiority of ASAQ vs. AL in the day-28 PCR-corrected ACPR rate during the first malaria episode, was assessed in the per-protocol and intent-to-treat populations. In addition, in the secondary analyses, PCR-corrected rates for subsequent episodes were compared in the intent-to-treat population. ASAQ was considered non-inferior to AL if the lower limit of the one-sided 95% confidence interval for the difference in rates was greater than \u22125%. Parasite density, gametocyte carriage, and Hb concentrations were examined in the intent-to-treat population. Safety was assessed in all the patients who received at least one dose of the study treatment.Of 422 patients who underwent screening, 416 were randomized to treatment , and 413 were enrolled in the study . Three cThe 413 enrolled children experienced a total of 6027 malaria episodes over the 2 years of the study. The children experienced on average (\u00b1 standard deviation) 15\u00b15 malaria episodes  . ChildreA peak in malaria episodes was observed from October 2008 to January 2009, April to May 2009 and from December 2009 to April 2010 . After dFull treatment compliance was observed for most malaria episodes with the exception of episodes 2, 6, 10, and 20 where one child in the AL group did not take all the study medications. Overall treatment compliance was 100% for ASAQ and greater than 99% for AL.The PCR-corrected ACPR rate during the first malaria episode was 97.5% for participants treated with ASAQ versus 97.0% for AL in the per-protocol population  confirming non - inferiority . The PCRThe D28 PCR-corrected ACPR rate in subsequent malaria episodes with more than 100 cases (episodes 2 to 18), varied from 88.8% to 98.9% for ASAQ and from 88.1% to 95.6% for AL in the PP population. In the ITT population the D28 PCR-corrected ACPR rate for episodes 2 to 18 varied from 88.3% to 98.9% for ASAQ and from 88.1% to 95.6% for AL in the PP population, . We had Fever clearance rate was significantly higher in participants treated with ASAQ compared to those treated with AL on day 1 of the first malaria episode (p<0.001), however there was no significant difference on days 2 and 3, with fever clearance rates>96% . The proThe mean time to parasite clearance was 1.8 days for both treatment regimens . ParasitDuring the first malaria episode, mean Hb level was low (<10 g/dl) between days 0 to 7 in both treatment groups however levels became normal by day 14 and continued to increase up to day 42 . In contSplenomegaly was detected on day 0 of the first episode in 8.6% (17/198) of children in the ASAQ group and 11.5% (23/201) of those in the AL group (p\u200a=\u200a0.525). All cases were grade 1 or 2. For subsequent episodes, splenomegaly was present in at most 1.2% of the children in each group, and no differences were detected between the ASAQ and AL groups (data not shown).Similar numbers of children in the ASAQ and AL groups reported TEAEs  and SAEs  as shown in AESIs were reported in 14 children in the ASAQ group (6.7%) and 14 (6.8%) in the AL group . The mosth malaria episode. Blood tests indicated that she had recrudescence of her infection. These events and her death were not considered by the investigator to be treatment-related.SAEs were reported in 16 children in the ASAQ group and in 9 in the AL group . The mosITNs were distributed to study participants in July 2009 because of the high number of malaria episodes experienced. A survey performed in May 2010 showed that ITNs were distributed to all homes and were correctly placed over the bed of the study participants in 100% of the homes . The surIn malaria-endemic areas, patients suffer from repeated malaria infections and therefore repeatedly use antimalarial drugs. However, most clinical studies have assessed the efficacy and safety of ACTs for only a single malaria episode. This longitudinal phase IV study was conducted in an area of perennial high malaria transmission to compare the safety and efficacy of ASAQ and AL when administered repeatedly over a long period. The study was performed in an area of Eastern Uganda known to have high malaria transmission intensity, with an annual entomological inoculation rate of 125 infectious bites per person per year The 413 children treated in this study experienced a total of 6027 malaria episodes (range 1 to 26) and on average (\u00b1 standard deviation) 15\u00b15) episodes over the course of the 2-year study. This is to our best knowledge the highest number of episodes ever reported in studies that have evaluated the repeated use of ASAQ and AL. Patients experienced on average 2 to 3 episodes per year in a 2-year study in Mali This study showed that ASAQ and AL have high and comparable efficacy profiles for the repeated treatment of malaria in children. The PCR-corrected ACPR for ASAQ was 97.5% and was non-inferior to the rate for AL (97.0%). The PCR-corrected cure rates for ASAQ are comparable to findings reported previously in West and Central Africa (94% to 98%) The number of malaria episodes was slightly, higher for participants treated with ASAQ compared to AL in this study, whereas it was lower or comparable in studies conducted in West Africa The median number of malaria episodes was slightly but significantly higher in the ASAQ group than in the AL group, and the average time between episodes significantly longer in the AL group, with a mean time from D0 of one attack to the D0 of the previous one significantly greater in the AL group for the 3rd, 4th, 7th, 10th and 19th attack. This finding differs from that of a similar study conducted in Mali which showed that patients treated with ASAQ had significantly less risk of recurrent malaria compared to those treated with AL Following an episode of malaria, even people who are asymptomatic can carry gametocytes and therefore remain reservoirs for infection The current study showed that both treatments rapidly reduced gametocyte carriage during the first episode and that this reduction in gametocyte carriage persisted during the subsequent episodes. Therefore, repeated treatment with ACTs is not only effective for treating individuals with malaria but might also reduce malaria transmission within the community. In such settings a post treatment dose with an ACT one month after a malaria episode could reduce recurrent episodes and improve the health of the population.Fever clearance rate was significantly higher in participants treated with ASAQ compared to those treated with AL on day 1 of the first malaria episode (p<0.001), however there was no significant difference on days 2 and 3, where fever clearance rates were>96% on both treatment arms. The exact reason for this is not clear but could be related to the antipyretic properties of amodiaquine, however the supply of antipyretics to patients on the first 3 days, could have led to an over estimation of the fever clearance in both treatment arms.Repeated administration of ASAQ and AL over a 2-year period in this study did not lead to unexpected safety issues. Safety profiles for both ACTs were good and comparable, and there was no evidence of emerging toxicity due to repeated use. AEs and SAEs were mainly reported during the first malaria episode and were consistent with features of malaria infection or concomitant infection or injuries. Anemia and neutropenia were observed in less than 0.5% of the children per episode, and only 0.3% to 1.4% of the participants had an abnormal liver function test. Increased alanine aminotransferase levels were rarely associated with increased total bilirubinemia. Furthermore all episodes of neutropenia spontaneously returned to normal and did not recur during subsequent treatment administrations.There was a large reduction in the incidence of the AEs after the first malaria episode. This was probably due to a larger proportion of cases with fever (18.6%) during the first episode compared to less than 5.5% in the subsequent episodes. In the subsequent malaria episodes, children were included on the basis of positive parasitemia without regard for fever or history of fever. Furthermore there could have been more AEs in the first episode due to the closer follow-up and administration of all the treatment doses by the study staff. In this study, repeated treatments did not appear to be associated with hepatic or hematologic toxicity. Similar safety profiles were reported in two other studies that assessed the repeated use of ASAQ and AL, although there were fewer episodes than in the current study Although one of the studies reported significantly more vomiting with the artesunate-amodiaquine co-blister than with AL  Administration of treatments was unsupervised for all but the first episode and first intake of subsequent treatments, but full compliance was obtained in all cases except for a few isolated episodes in children treated with AL. Excellent compliance for repeated treatment with ASAQ has been reported previously Anopheles numbers due to increasing insecticide resistance, as found in a recent longitudinal study on the effect of ITNs in a Senegalese community Anopheles biting behavior, as reported in 3-year study in Benin One year into the study, the unexpectedly high number of malaria episodes raised ethical concerns that led to a decision to distribute ITNs to the children's families. Although study staff members confirmed that the ITNs were properly placed and used, the ITNs did not appear to have a lasting impact on the incidence of malaria episodes. Although study participants had ITNs, coverage of ITN use in the study area was low. The proportion of children <5 who slept under a mosquito net in the area increased from 33.85 in 2006 to 47.8 in 2009 while the proportion of children under 5 who slept under an ITN increased from 12.8% in 2006 to 41.8% in 2010 Recent reports of delayed parasite clearance after artemisinin derivative treatment in Southeast Asia have raised concerns of emerging resistance to artemisinin This study was conducted in a high transmission area and our findings may be general sable to other settings in Africa with similar malaria transmission intensity.This study is remarkable because of the number of malaria episodes studied, and it clearly showed that ASAQ is safe and effective for repeated use, reduces gametocyte carriage and is as well tolerated and effective as AL.Approval S1Makerere Institutional Review Board/Ethics Committee approval document.(PDF)Click here for additional data file.Approval S2Uganda National Council of Science and Technology approval document.(PDF)Click here for additional data file.Checklist S1Completed CONSORT checklist.(DOC)Click here for additional data file.Protocol S1Study protocol.(DOC)Click here for additional data file."}
+{"text": "Both drugs were found to be well tolerated by the patients. This study demonstrates the effectiveness and tolerability of ASAQ and AL supporting their continuous use for the treatment of uncomplicated P. falciparum malaria infection in C\u00f4te d'Ivoire.Two years after the introduction of free Artesunate-Amodiaquine (ASAQ) and Artemether-Lumefantrine (AL) for the treatment of uncomplicated malaria in public health facilities in C\u00f4te d'Ivoire, we carried out this study to compare their efficacy and tolerability in three surveillance sites. It was a multicentre open randomised clinical trial of 3-day ASAQ treatment against AL for the treatment of 2 parallel groups of patients aged 2 years and above. The endpoints were (1) Adequate Clinical and Parasitological Response (ACPR) at day 28 and (2) the clinical and biological tolerability. Of the 300 patients who were enrolled 289, with 143 (49.5%) and 146 (50.5%) in the ASAQ and AL groups, respectively, correctly followed the WHO 2003 protocol we used. The PCR-corrected ACPR was 99.3% for each group. More than 94% of patients no longer showed signs of fever, 48 hours after treatment. Approximately 78% of the people in the ASAQ group had a parasite clearance time of 48 hours or less compared to 81% in the AL group ( P. falciparum to various antimalarial drugs available, the WHO introduced and recommended the use of artemisinin-based combination therapy (ACT) for the treatment of malaria [Malaria remains a serious health concern in sub-Saharan Africa . In C\u00f4te malaria , 4. P. falciparum resistant strains as a result of drug pressure. Furthermore, self-medication, poor adherence to treatment, counterfeit drugs, and human and Plasmodium genetic makeup may influence the efficacy and safety profiles of ACTs in C\u00f4te d'Ivoire.This recommendation was adopted by C\u00f4te d'Ivoire in 2007 . Since 2 P. falciparum to artemisinin derivatives has been recently documented in the Cambodia-Thailand border region [Indeed, resistance ofr region , 7. Thisr region , 9. It iThe aim of this study was to compare the efficacy and tolerability of Artesunate-Amodiaquine and Artemether-Lumefantrine for the treatment of uncomplicated falciparum malaria two years after their large-scale use in C\u00f4te d'Ivoire as first-line and second-line treatment.The study was carried out between June and September 2012 in three surveillance sites for antimalarial drug efficacy in C\u00f4te d'Ivoire: Abengourou (forest zone), San Pedro , and Yamoussoukro (forest transition zone). In each site, two popular and well known health centres were chosen for the survey. Throughout the country, there are four distinct seasons divided into two raining seasons (December\u2013July and October-November) with high malaria transmission and two dry seasons (December\u2013March and August-September).A controlled randomized multicentre and open therapeutic trial with a 28-day follow-up period comparing the efficacy, safety, and tolerability of two Fixed-Dose Combinations (FDC) which are ASAQ and AL in patients from 2 years old and above was used. Throughout the survey, the standard WHO 2003 efficacy assessment protocol was followed .Based on previous studies \u201313, the  P. falciparum monoinfection with parasitaemia from 2,000 to 200,000/\u03bcL of blood. Patients with signs or evidence of severe malaria/malnutrition, repeated vomiting, intercurrent infectious disease, history of previous serious side effects to the drugs used during the trial, and past cardiac, hepatic, or renal history or those who were pregnant (positive test) or breast-feeding were excluded. Criteria to stop the treatment and/or withdrawal of a patient from the study included the following: (1) occurrence of serious adverse effects; (2) unsatisfactory therapeutic response; (3) violation of the protocol; (4) withdrawal of consent; and (5) being lost to follow-up. Before inclusion, written informed consent was obtained from the patient or the patient's legal guardian. Approval was obtained from the national ethics committee before study onset.The study population consisted of outpatients who came to the health facilities with uncomplicated malaria-like symptoms. Patients were referred to the study team for recruitment. Inclusion criteria for the study were as follows: (1) being at least two years old; (2) fever with axillary temperature \u226537.5\u00b0C; (3)For each patient involved in the study, the protocol was read and explained to him/her or to the legal guardian (as regards children). On acceptance, patient or legal guardian had to sign the informed consent forms to take part in the study. For all the recruited patients, baseline examinations and laboratory investigations were conducted immediately and free of charge. Those among the patients who met inclusion criteria at baseline were randomly assigned to one of the two treatment groups following a randomization list. In each study site, computer generated randomization codes were prepared by an independent individual. These codes were enclosed in sequentially numbered opaque sealed envelopes, each of which contained the treatment allocation. The envelopes were assigned in sequential order to participants after inclusion.Each patient was allocated to one of the two treatment groups. Artesunate- (AS) Amodiaquine (AQ) (ASAQ)  was administered as a single daily dose for three days. Each tablet of ASAQ contained either 50\u2009mg of AS and 135\u2009mg of AQ or 100\u2009mg of AS and 270\u2009mg of AQ. ASAQ treatments varied according to body weight: 9\u201317\u2009kg, one tablet (50\u2009mg/135\u2009mg) per dose; 18\u201336\u2009kg, one tablet (100\u2009mg/270\u2009mg) per dose; and over 36\u2009kg, two tablets (100\u2009mg/270\u2009mg) per dose. Artemether-Lumefantrine (AL) tablets  were administered at 0 and 8 hours on day 1 and then twice daily for two subsequent days according to body weight: 5\u201314\u2009kg, one tablet per dose; 15\u201324\u2009kg, two tablets per dose; 25\u201334\u2009kg, three tablets per dose; 35\u2009kg and over, four tablets per dose. All treatments were given under direct supervision of a member of the study team. If the patient vomited within 30 minutes after taking the drug, the whole dose was readministered. However, if the vomiting persisted, the patient was removed from the study and referred to the health centre for an in-depth investigation and treatment according to the current national policy. The dose could not be administered again if vomiting occurred more than 60 minutes after administration.Concomitant treatment refers to the treatment of diseases other than malaria. Antipyretic and antiallergic were provided when needed during the follow-up. Plasmodium which could interact or lead to false evaluation of the drug efficacy evaluation.Antibiotics such as sulphonamide, tetracycline, quinolone, and macrolide were contraindicated during the study because of their possible activity against P. falciparum molecular biology analysis at baseline and then after day 7 in case of parasitaemia. The response to treatment was measured and defined according to WHO guidelines [After inclusion, patients were scheduled for follow-up examinations on days 1, 2, 3, 4, 7, 14, 21, and 28 using WHO in vivo tests with a follow-up period of 28 days , 14. Theidelines . P. falciparum in the peripheral blood was determined by counting the number of asexual parasites in 200 white blood cells (WBC). All of the thick and thin blood films were reread to double check. A slide was considered negative after reading 200 microscopic fields. The presence of gametocytes was also noted. In case of discrepancy, a third reading was made by a third microscopist. An external quality control was carried out on 10% of the slides. Venous blood was collected on days 1 and 4 for conducting haematological (full blood count) and biochemical  investigations.At each visit, thick and thin blood films were performed. The density ofmsp1) and merozoite surface protein-2 (msp2) using nested PCR as described by Soulama et al. [In order to distinguish recrudescence from new infection, filter-paper blood spots  were collected from finger pricks on day 1 and on the day of recurrent parasitaemia (after day 7) and used for molecular genotyping. Parasite DNA was extracted from filter-paper blood spots using the Chelex methods  and anala et al. .Primary efficacy parameter was the cure rate at day 28. It is the proportion of patients for whom removal parasitaemia is obtained within 7 days of the study without recrudescence within 28 days after the start of study treatment. The recurrence is defined as a new clinical manifestation of the infection after initial removal of parasites in the peripheral blood. However, in case of reinfection (verified by PCR), parasitological recurrence is not considered as a treatment failure of malaria drug received.Secondary efficacy endpoints were as follows: (a) Cure rate at 14 days: proportion of patients for whom removal parasitaemia is obtained within 7 days of the study without recrudescence within 14 days after the start of study treatment. (b) Parasite clearance time: time elapsed between the first administration and the first total and continued disappearance of parasite asexual forms and persisting for at least another 24 hours. (c) Thermal clearance time: time elapsed between the first dose and the first lowering of the temperature below 37.5\u00b0C for at least another 24 hours. (d) Gametocyte carriage evolution. (e) Improvement in haemoglobin rate compared to the start of the study.Consider the following:It consisted of monitoring and registration of any adverse event , biological monitoring , and the assessment of the clinical status of subject  during follow-up. Any clinical or biological sign not present in inclusion and which appeared during follow-up or any sign present at day 1 and worsening thereafter was considered as adverse event.t-test. The cases of protocol violation and withdrawn consent were censored at the time they left the study. The distributions of fever and parasite clearance were compared using Pearson's Chi-square test. Differences of haemoglobin and biochemical parameters values within individuals between day 1 and day 4 were computed. Changes in haemoglobin concentrations and in biochemical parameters were compared using the paired t-test. The level of significance for statistical tests was set at 0.05. Data were done in Per Protocol (PP) analysis.All data were recorded and checked using Epi data version 3.1 and analysed with SPSS for windows (version 16.0). The characteristics of patients in the two groups at inclusion were compared using Pearson's Chi-square test and independent samples A total of 300 patients were included in the study. 151 patients were randomized to ASAQ and 149 to AL. In the ASAQ group, six patients were lost to follow-up and two patients have withdrawn their consent. In the AL group, there were two patients lost to follow-up and one case of consent withdrawal. Finally, 143 (49.5%) patients and 146 (50.5%) patients were successfully followed up, respectively, in ASAQ and AL groups .Baseline characteristics of patients receiving either ASAQ or AL are summarized in  P. falciparum infections and preventing parasite recurrences compared to AL. Only nine treatment failures were observed: 1 (0.7%) in the ASAQ group and 8 (5.5%) in the AL group. Most of the therapeutic failures were classified as LCF. And most of LCF cases were found among children between five and fifteen (6.8%). Any case of Late Parasitological Failure (LPF) was observed. After PCR correction, both treatments had the same Adequate Clinical and Parasitological Response (ACPR) (99.3%).The results of the treatment efficacy are presented by treatment group, day of follow-up, unadjusted and adjusted by genotyping, and age of study patients . Cure raAt day 14, cure rate was 100% with ASAQ and 99.3% with AL. Indeed, one case of Early Treatment Failure (ETF) was observed in AL group in an under-five child.p = 0.496)  .p = 0.00086).In both treatment groups, the majority of subjects had fever clearance \u2264 24 hours. Beyond 72 hours, all the patients treated by ASAQ were nonfebrile. Seven patients in the AL group were febrile after 96 hours . The difAt inclusion, there were only two gametocyte carriers in the ASAQ group and five in the AL group. In both therapeutic groups, the number of gametocyte carriers decreased throughout the follow-up and was zero at day 28 .Among the 289 patients followed up, in 175 (121 ASAQ and 54 AL) some side effects were observed (60.5%). Generally, adverse events were regular in the ASAQ group and included pruritus, asthenia, and drowsiness and vomiting. The AL seemed clinically better tolerated than ASAQ. However, side effects were not severe in both treatments as to interrupting the treatment .p < 0.001). Between day 1 and day 4 in both treatment groups we observed that the mean of ALT decreased but was not significantly different, the level of creatinine varied but not significantly, and finally the amount of bilirubin decreased significantly in both treatment arms (p < 0.001)  than ASAQ group (\u22120.86\u2009g/dL). The decrease was significant within both groups (< 0.001) . P. falciparum malaria treatment among patients more than two years old.This randomized trial enabled an evaluation of the efficacy and tolerability of ASAQ versus AL in three sentinel sites in C\u00f4te d'Ivoire as part of the uncomplicatedASAQ appeared to be a better treatment option on the basis of non-PCR-corrected responses, based on the lower percentage of recurrent parasitaemia observed. However, the PCR-corrected cure rates which indicated the true efficacy were the same for both treatments. The results showed a high cure rate for both regimens after a standard 28-day follow-up with an ACPR rate adjusted by 99.3%. This is consistent with efficacy results reported from several sub-Saharan African countries , 17\u201323.The fact that there was more reinfection than recrudescence shows that malaria transmission is high in all three sentinel sites . Indeed,Fever clearance was fast in both treatment groups confirming the previous data . HoweverThe decrease of gametocyte rate during the treatment with ACTs has been demonstrated. This action decreases transmission and therefore leads to a significant reduction in the spread of resistance , 31. In The clinical tolerance was good with minor adverse events in both treatment groups confirming previous studies . Regardi falciparum malaria. Therefore, this work supports the continued use of these ACTs in the management of malaria with added advantage provided in public health facilities in slowing the spread of malaria drug resistance and of global reduction or elimination of malaria in C\u00f4te d'Ivoire.During our study, we found that ASAQ was as effective and well-tolerated as was AL in the treatment of uncomplicated"}
+{"text": "The antimalarial mefloquine was used to directly measure the PK/PD in both models, which were compared to previously published trial data for malaria patients. The clinical part was a single-center, controlled study using a blood-stage Plasmodium falciparum challenge inoculum in volunteers to characterize the effectiveness of mefloquine against early malaria. The study was conducted in three cohorts (n = 8 each) using different doses of mefloquine. The characteristic delay in onset of action of about 24 h was seen in both NSG and IBSM systems. In vivo 50% inhibitory concentrations (IC50s) were estimated at 2.0 \u03bcg/ml and 1.8 \u03bcg/ml in the NSG and IBSM models, respectively, aligning with 1.8 \u03bcg/ml reported previously for patients. In the IBSM model, the parasite reduction ratios were 157 and 195 for the 10- and 15-mg/kg doses, within the range of previously reported clinical data for patients but significantly lower than observed in the mouse model. Linking mouse and human challenge models to clinical trial data can accelerate the accrual of critical data on antimalarial drug activity. Such data can guide large clinical trials required for development of urgently needed novel antimalarial combinations. Effective progression of candidate antimalarials is dependent on optimal dosing in clinical studies, which is determined by a sound understanding of pharmacokinetics and pharmacodynamics (PK/PD). Recently, two important translational models for antimalarials have been developed: the NOD/SCID/IL2R\u03b3 The cumulative success rate from candidate selection to registration for the development of anti-infectives is below 5%  immunodeficient rodent model of malaria  and their overall response to malaria . One appTo investigate the potential of the combined use of these two models, we evaluated a well-established and metabolically stable malaria drug, mefloquine , in bothIn vivo therapeutic efficacy against P. falciparum was assessed in a 4-day parasite-normalized standard assay (PNSA) previously described (\u2212/\u2212 (NSG) mice  were injected daily with 1 ml of human erythrocytes (50% hematocrit)  obtained from P. falciparum-infected donor mice. The Plasmodium strain had been generated at GlaxoSmithKline (GSK), Tres Cantos, Spain (escribed . NOD SCIatocrit)  and apprs, Spain . Parasitin vivo studies were performed in the P. falciparum humanized mouse model: (i) per os (p.o.) multidose administration once a day for 4 days at 0.2, 1, 3, 10, and 30 mg/kg of body weight (n = 3 mice per dose level); (ii) multidose administration p.o. once a day for 4 days at 1, 2.5, 5, 10, 20, 30, 40, 50, 60, and 70 mg/kg (n = 1 mouse per dose level); (iii) bolus i.v. single administration at 1 mg/kg in both infected  and noninfected humanized mice (n = 3 mice per group); and (iv) single administration p.o. at 20 mg/kg of infected  and noninfected humanized mice (n = 3 mice per group). Data from the three experiments were used to model exposure kinetics.Four in vivo studies. The samples were taken at 0.25, 0.5, 1, 2, 3, 6, 8, and 23 h after the first dose and diluted with 25 \u03bcl of water containing 0.1% saponin for erythrocyte lysis, flash-frozen on dry ice, and stored at \u221280\u00b0C until analysis. The lower limit of quantification of mefloquine in blood was 2.5 ng/ml.Mefloquine levels in whole blood were measured in serial samples of peripheral blood (25 \u03bcl) from every mouse in the Parasitemia was measured in experiments i and ii by flow cytometry as described previously , with a P. falciparum-infected human erythrocytes administered i.v. (Plasmodium 18S rRNA genes). If clinical or parasitological evidence of malaria  occurred, treatment began at this time. If clinically well at the end of the 48 h confinement period, participants were discharged and monitored on an outpatient basis for safety, blood mefloquine levels, and the presence of malaria parasites by qPCR. The effect of mefloquine on parasitemia was observed for up to 7 days before compulsory commencement of treatment with artemether-lumefantrine  administered as four tablets as a single dose every 12 h for 60 h. As an additional safety measure, parasitemia was monitored until the end-of-study visit, scheduled approximately 28 days after the inoculation. Dose escalation between cohorts occurred only after review by the Safety Review Team of the observed mefloquine safety and pharmacodynamic outcome up to day 14 postinoculation for the previous cohort. Details of parasitemia and plasma mefloquine determinations are described in the supplemental material.A single-center, controlled study was undertaken by following methods previously described . The stured i.v. , 22. Parred i.v.  results Parasitemia was quantified by qPCR as described previously  at the fThe PK of mefloquine was assessed in terms of the plasma concentration-time profiles in blood samples collected predose and at the following times after dosing: 0.25, 0.5, 1, 2, 3, 4, 6, 8, 12, 24, 36, 48, 72, 96, 120, and 144 h. Plasma was separated and frozen at \u2264\u221270\u00b0C within 30 min. It was found that mefloquine standards in plasma could be stored for 10 days at 4\u00b0C with no change in measured concentration (data not shown). Mefloquine was assayed in 200-\u03bcl plasma samples following acetonitrile extraction as described previously . A one-compartment model had been used previously to model mefloquine exposure . A two-cConcentration-versus-time data were analyzed by noncompartmental analysis methods using Phoenix software . Additional statistical analysis and modeling were performed with GraphPad Prism  and NONMEM software .10 transformed, and the mean of the log10-transformed parasitemia was implemented as a summary parasitemia value per time point per subject. For the historical data sets for which aggregated parasitemia data were available, PRR estimates were calculated using the mean parasitemia for each study and dose of interest over time (hours since treatment).Parasite reduction ratio (PRR) estimates were calculated as previously described . In brieThe parasitemia data were modeled as part of the simultaneous PK and PD (PK/PD) modeling . SuccessGk is a first-order parasite growth constant, Dk is a first-order parasite clearance constant, C is the drug concentration in the effect compartment, IC50 is the drug concentration corresponding to when parasite clearance is half maximal, and H is the Hill coefficient, determining the steepness of the drug concentration-parasite clearance relationship.The PK/PD model was compiled in NONMEM using differential equations based on the initial PK and baseline modeling. Our earlier PK/PD model , based oThe clinical protocol and associated documents were reviewed and approved by the Queensland Institute of Medical Research Berghofer Human Research Ethics Committee. The trial was registered at anzctr.org.au (registration number ACTRN12612000323820). All the animal experiments performed at GSK were approved by the DDW Ethical Committee on Animal Research, performed at the DDW Laboratory Animal Science facilities accredited by AAALAC, and conducted in accordance with European Directive 86/609/EEC and the GSK policy on the care, welfare, and treatment of animals . The hum10 growth rate of 0.013 h\u22121 . In mefloquine-treated animals (10 parasite reduction ratio (log10 PRRMax) was calculated as 3.3 . The average age of volunteers was 25 (\u00b14) years, with a weight of 72 (\u00b111) kg, a height of 175 (\u00b111) cm, and body mass index (BMI) of 23 (\u00b12). The only serious adverse event (recurrent tonsillitis) was considered unrelated to the clinical trial treatment. The mild and moderate adverse events were either malaria related or mefloquine related . These events were comparable to those seen earlier in the IBSM studies or reported following the use of mefloquine. Among the 52 adverse events attributed to malaria, all were judged to be of mild to moderate severity, and they were generally of a transient nature. The most common adverse events attributable to malaria were fever symptoms (n = 6) and mild leukopenia (n = 7). Of the adverse events attributable to mefloquine, all except two (lucid dreams and nausea) occurred in the highest-dose cohort (15 mg/kg). Of note, three of the five subjects who experienced dizziness reported a significant duration of symptoms . Four subjects reported nausea and/or vomiting following receipt of the highest dose of mefloquine, and two reported insomnia, with one of these two reporting that this lasted for 6 days.Twenty-two volunteers  were enrolled in a clinical trial to evaluate the PK/PD relationship of mefloquine administered in three doses  and 2.20  for the 10-mg cohort and 195  and 2.29  for the 15-mg cohort. For the 10-mg/kg administration, the MIC of mefloquine was reached between 96 and 108 h. The geometric mean MIC was 784 ng/ml (CV = 18%). This is equivalent to an IC50 of 667 ng/ml, or 1.8 \u03bcM, comparable to the value we found in mice .Rates of clearance of parasitemia, as measured by the dose-specific PRRWe identified two earlier clinical trial studies in which serial quantitative parasite count data were reported following administration of mefloquine monotherapy. Using the published data , we calc50 derived in NSG mice infected with P. falciparum, 768 ng/ml, closely matches that estimated by modeling in infected humans  binding are roughly equivalent, measured at 95 to 96% in humans and \u223c97% in rodents . The est65 ng/ml ).We found that mefloquine-induced parasite clearance kinetics observed in the IBSM matched those reported in earlier clinical studies , 28, desin vivo IC50 measured in the mouse and human challenge models and in clinical patients. However, PRRs differ substantially between the models and also between published clinical studies, suggesting that the IC50s may be more useful in predicting optimal dosing of new drugs. Moreover, other factors, including parasite biology and immunity , are inin vivo system. It remains to be seen if the correlations that we have found between the in vivo model and in volunteers also hold for drugs with a different mechanism of action, with exposure characteristics different from those of mefloquine, and that produce significantly different exposures in rodents and humans. But with antimalarials typically targeting either the parasite directly or components in the human infected erythrocyte compartment, it would seem that the two models that we have evaluated in this study have all the critical elements in place for confidently guiding clinical development of molecules currently progressing through the antimalarial pipeline.While falciparum malaria models involving both NSG mice and human volunteers have been developed and refined recently, a gap to date has been a formal assessment of these models' validity in a coherent pathway for drug discovery using a clinically well-established antimalarial as a benchmark. In this study, we have evaluated mefloquine both in infected NSG mice and in volunteers, fitted the key parameters in a PD model, and found an excellent correlation between the variables that describe each"}
+{"text": "Plasmodium falciparum threatens the health of millions of people and poses a major challenge to the control of malaria. Monitoring drug efficacy in 2-year intervals is an important tool for establishing rational anti-malarial drug policies. This study addresses the therapeutic efficacy of artemether-lumefantrine (AL) for the treatment of Plasmodium falciparum in southwestern Ethiopia.The development and spread of chloroquine-resistant P. falciparum. All 93 eligible patients were treated with AL and followed for 28\u00a0days. For each patient, recurrence of parasitaemia, the clinical condition, and the presence of gametoytes were assessed on each visit during the follow-up period. PCR was conducted to differentiate re-infection from recrudescence.A 28-day in vivo therapeutic efficacy study was conducted from September to December, 2011, in southwestern Ethiopia. Participants were selected for the study if they were older than 6\u00a0months, weighed more than 5\u00a0kg, symptomatic, and had microscopically confirmed, uncomplicated P. falciparum and one (1.1\u00a0%) was due to P. vivax. From 89 study subjects, 12 (13.5\u00a0%) carried P. falciparum gametocytes at day 0, whereas the 28-day gametocyte carriage rate was 2 (2.2\u00a0%).Seventy-four (83.1\u00a0%) of the study subjects cleared fever by day 1, but five (5.6\u00a0%) had fever at day 2. All study subjects cleared fever by day 3. Seventy-nine (88.8\u00a0%) of the study subjects cleared the parasite by day 1, seven (7.9\u00a0%) were blood-smear positive by day 1, and three (3.4\u00a0%) were positive by day 2. In five patients (5.6\u00a0%), parasitaemia reappeared during the 28-day follow-up period. From these five, one (1.1\u00a0%) was a late clinical failure, and four (4.5\u00a0%) were a late parasitological failure. On the day of recurrent parasitaemia, the level of chloroquine/desethylchloroquine (CQ-DCQ) was above the minimum effective concentration (>100\u00a0ng/ml) in one patient. There were 84 (94.4\u00a0%) adequate clinical and parasitological responses. The 28-day, PCR-uncorrected (unadjusted by genotyping) cure rate was 84 (94.4\u00a0%), whereas the 28-day, PCR-corrected cure rate was 87 (97.8\u00a0%). Of the three re-infections, two (2.2\u00a0%) were due to P. falciparum malaria and reducing gametocyte carriage in southwestern Ethiopia.Years after the introduction of AL in Ethiopia, the finding of this study is that AL has been highly effective in the treatment of uncomplicated  Although there have been encouraging reports of declining of morbidity and mortality from malaria in most endemic countries , it remaPlasmodium falciparum and P. vivax are the two dominant parasite species, with relative frequencies of about 60 and 40\u00a0%, respectively . Still, Plasmodium falciparum is the dominant parasite species, causing severe and complicated manifestations and is responsible for most malarial deaths [In Ethiopia 75\u00a0% of the area is malarious, and approximately 52 million people live in high-risk areas, mainly at altitudes below 2,000 metres . Plasmod. Plasmodium falciparum has an extraordinary ability to do this, creating a major challenge for the control efforts and increasing the number of deaths in sub-Saharan Africa [Early diagnosis and effective treatment are essential elements for the control of malaria. However, one of the obstacles to controlling malaria is the capability of the parasites to evolve resistance to various anti-malarial drugsn Africa , 6.P. falciparum to a standard triple-dose of chloroquine (25\u00a0mg base/kg) [P. falciparum and P. vivax, brought a policy change, and sulfadoxine-pyrimetamine (SP) has been used as an affordable alternative treatment of uncomplicated malaria cases since 1998 [P. falciparum [P. falciparum malaria for all countries experiencing resistance to monotherapies in 2001 [In Ethiopia, resistance of base/kg) , and a rlciparum . As a re in 2001 . Accordi in 2001 . Replaci in 2001 .ACT was designed to attack malaria parasites with different mechanisms of action simultaneously. It reduces the emergence of drug resistance, and it gives a faster relief from clinical symptoms and parasite clearance . ArtemetP. falciparum malaria after treatment with ACTs emerged from observational data collected in Cambodia [To insure timely changes to treatment policy, WHO recommended therapeutic efficacy studies of the drug every 2\u00a0years . AccordiCambodia . MoreoveCambodia . Delay iCambodia .P. falciparum malaria in Southwestern Ethiopia. The secondary objectives of the study were to determine the prevalence of post-treatment gametocyte carriage in Southwestern Ethiopia. This information will inform policy makers with respect to appropriate antimalarial strategies.The development of resistance to ACT could have a major impact on malaria control efforts at time when there are no other drugs in the development pipeline. Therefore, the primary aim of this study was to assess the therapeutic efficacy of AL for the treatment of uncomplicated P. falciparum malaria with artemether\u2013lumefantrine , 2001, at Omo Nada health center in southwestern Ethiopia. Following WHO protocols , we evalOmo Nada  is one of the woredas (the name for districts in Ethiopia) in Jimma Zone, Oromia Region of Ethiopia. Nada is the administrative center of the woreda; other towns in Omo Nada include Asendabo. The altitude of this woreda ranges from 1,000 to 3,340\u00a0meters above sea level, and its total population is 2,48,173. As in most other places, malaria transmission in Omo Nada follows rainy seasons, with transmission peaking in the months between September and December and between April and May. The main malaria control strategy in the woreda includes use of Long Lasting Insecticidal Nets (LLINs), malaria case management with ACTs, and intermittent preventive treatment during pregnancy (IPTp) .2 P (1\u00a0\u2212\u00a0P), where P stands for the anticipated prevalence, d for the margin of error, and Z for the Z statistic at a given level of confidence. For the level of confidence of 95\u00a0%, the Z value is 1.96. In this study a 5\u00a0% failure rate was expected, the precision level was set a 4.5\u00a0%, and a loss-to-follow-up rate was anticipated to be 20\u00a0% over 28-days. Thus the calculated sample size was 93, which was considered sufficient to address the study objectives.For therapeuthic efficacy studies, sample sizes (n) were calculcated based on recommendations by the Technical Expert Group on Malaria Chemotherapy , using tP. falciparum-positive patients visiting health centres who met the WHO inclusion guidelines for the assessment and monitoring of anti-malarial drug efficacy for the treatment of uncomplicated malaria [The study subjects were recruited among  malaria . The detOver 6\u00a0months of ageLiving in areas of low-to-moderate transmissionP. falciparum detected by microscopyMono-infection with Asexual parasite count of 1,000\u2013100,000/\u00b5l in areas of low-to-moderate transmissionAxillary temperature\u00a0\u226537.5\u00a0\u00b0C or history of fever during the 24\u00a0h before recruitmentAbility to swallow oral medicationAbility and willingness to comply with the protocol for the duration of the study and to comply with the study visit scheduleInformed consent from the patient The inclusion criteria were as follows:P. falciparum malaria, according to the definitions of WHOGeneral danger signs in children under 5\u00a0years (coughing or difficulty breathing) or signs of severe Severe malnutrition according to WHO growth standards .Febrile condition due to diseases other than malaria  or other known underlying chronic or severe diseases Regular medication which might interfere with anti-malarial pharmacokinetics History of hypersensitivity reactions to any medicine tested or used as an alternative treatment or contraindication to ALPregnant or breastfeeding womenThe exclusion criteria were as follows:Thick and thin blood smears were prepared and stained with 10\u00a0% Giemsa for 10\u00a0min. The stained blood films were examined on days 0, 1, 2, 3, 7, 14, 21, and 28, as well as on any unexpected visits with the patient due to the patient suffering a malaria attack before the day of the next appointment. Two laboratory personnel read the blood film slide independently. Blood smears with discordant results  were re-examined by a third, independent microscopist, and parasite density was calculated by averaging the two closest counts. A blood film was considered negative when no parasite was found after examining 100 fields. On day 0, hemoglobin was determined for those study subjects who were willing to participate on the study. Parasite count was based on the number of asexual parasites observed against 200 leukocytes. This number was then multiplied by 40 to gain an approximate count per microlitre.Parasite densities for all participants were calculated using an assumed WBC of 8.0\u00a0\u00d7\u00a010(9)/L of blood, which has been set by WHO to be used for convenience in facilities which lack the tools to determine patients\u2019 absolute full blood cell count (FBC) values. In addition, the whole Giemsa-stained thick film was examined for gametocyte carriage before and after artemether\u2013lumefantrine treatment on all follow up days.As a routine procedure within the health system of Ethiopia, study subjects were treated with AL (Artefan) , Mumbai 400 067, India) twice daily on days 0, 1, and 2. Study medication was administered based on weight; the first dose were administered under the supervision of a qualified member of the study staff. Study subjects were followed for 30\u00a0min post treatment as an additional procedure from the routine ones. If vomiting occurred, a second full dose was administered. If repeated vomiting occurred, patients were withdrawn from the study and offered rescue therapy. Patients who failed to respond to AL treatment during the 28\u00a0days of follow-up were treated with oral quinine (8\u00a0mg/kg per day for 7\u00a0days), which is a second line of drugs and a regular treatment regime for the treatment of malaria in Ethiopia.Patients were asked to return to the health centre on days 1, 2, 3, 7, 14, 21, 28 or if suffering from a malaria attack before the day of the next appointment. On each follow-up day, blood was examined for the presence or absence of parasites, and temperature was measured. Those patients with recurrence of parasites during the follow-up days were treated with quinine. Patients were considered lost to follow-up when they did not come to the clinic as scheduled and became unreachable.DNA was isolated using QIAgen DNA Mini Kit for blood and tissue , based on the manufacturer\u2019s instructions, and stored at \u221220\u00a0\u00b0C until use. A nested PCR assay was then carried out as previously described elsewhere .msp1, msp2 and glurp\u2014were used in this here; these markers have been shown to have adequate discriminatory power for recrudescence versus re-infection testing [P. falciparum can be divided into 17 distinct blocks. Exept block 2 region, the non-conserved sequences can be grouped into major families represented by the MAD20 [The most widely used genetic markers for malaria genotyping\u2014the antigen genes  testing . Merozoihe MAD20 . Alleliche MAD20 . The msphe MAD20 , 27. Allhe MAD20 .Table\u00a01STreatment outcomes for AL were determined based on the WHO classification of treatment outcomes as follows: (1) early treatment failure (ETF), (2) late clinical failure (LCF), (3) late parasitological failure (LPF), and (4) adequate clinical and parasitological response (ACPR) .Ethical clearances were obtained from Aklilu Lemma Institute of Pathobiology (ALIPB), Armauer Hansen Research Institute (AHRI/ALERT) and Jimma University before commencing the projects. The Investigator explained the study to each potential subject verbally, provided all the information , and gave time for any queries. Then the potential subject was provided with a written consent form and afforded sufficient time to read it. Once an individual had all his/her questions answered and agreed to participate in the study, they or their family or guardian signed an informed consent prior to inclusion in the study. Confidentiality of information and freedom to withdraw from the study anytime were guaranteed. In addition, study subjects were compensated for transportation costs during the 28-day follow-up period.P. falciparum-positive patients visiting Omo Nada health centers, 93 fulfilled the inclusion criteria and were included on the AL efficacy study. Of these, two vomited twice within 30\u00a0min, and two were lost to follow-up; thus these were excluded from the study.Among 150 The majority of the study subjects (59.8\u00a0%) were male. The mean age and temperature of the study subjects were 17.3\u00a0years and 37.7\u00a0\u00b0C, respectively. The minimum haemoglobin level was 8.0\u00a0g/ml, while the maximum haemoglobin level was 14.6\u00a0g/ml. The mean parasite load counted from the thick blood film was 8,404  (Table\u00a0Seventy-four 83.1\u00a0%) of the study subjects cleared fever by day 1, ten (11.2\u00a0%) still had fever on day 1, and five (5.6\u00a0%) had fever on day 2. All study subjects cleared fever by day 3. In addition, 79 (88.8\u00a0%) of the study subjects cleared the parasite by day 1, seven (7.9\u00a0%) were blood-smear positive by day 1, and three (3.4\u00a0%) were positive by day 2  was due to P. vivax  were classified as treatment failures. One (1.1\u00a0%) was a late clinical failure, and four 4.5\u00a0%) were late parasitological failures. From the four late parasitological failures, parasites were identified from three study subjects on day 14 and from one study subjects on day 21. There were 84 (94.4\u00a0%) adequate clinical and parasitological responses. PCR uncorrected cure rate (unadjusted by genotyping) was 94.4\u00a0% (95\u00a0% CI 88.0\u201397.9\u00a0%) and the PCR corrected cure rate was 97.8\u00a0% (95\u00a0% CI 92.8\u201399.6\u00a0%). Of the three re-infections, two (2.2\u00a0%) were due to .5\u00a0% wereThere were 33 study subjects \u22645\u00a0years of age and the rest (56) were above 5\u00a0years of age. In\u00a0\u22645\u00a0years of age the PCR-uncorrected cure rate was 97.0\u00a0% (95\u00a0% CI 86.0\u2013100.0\u00a0%) while PCR-corrected cure rate was 100.0\u00a0% (95\u00a0% CI 91.3\u20130.0\u00a0%). On the other hand, PCR-uncorrected and corrected cure rates in study subjects above 5\u00a0years of age were 92.9\u00a0% (95\u00a0% CI 83.7\u201397.7\u00a0%) and 96.4\u00a0% (95\u00a0% CI 88.7\u201399.4\u00a0%) respectively   carried .5\u00a0% carrP. falciparum to evolve resistance to a number of anti-malarial drugs has recently increased the number of deaths from malaria worldwide [P. falciparum [P. falciparum malaria since 2004 in southwestern Ethiopia. All 93 P. falciparum mono-infections in our study received AL, and 74 of them (83.1\u00a0%) resolved their fever by day 3. AL is known to clear fevers in a very short period of time (less than 3\u00a0days). The parasite clearance in this study was faster than the result reported from Thai\u2013Cambodian border where the parasite resistance was characterized in this efficacy study [The remarkable ability of orldwide , 28. Chllciparum  and highlciparum , 29 havecy study .falciparum malaria in Ethiopia. In the current study there were 84 (94.4\u00a0%) adequate clinical and parasitological responses. That means PCR uncorrected cure rate (unadjusted by genotyping) was 94.4\u00a0% (95\u00a0% CI 88.0\u201397.9\u00a0%) and The PCR corrected cure rate was 97.8\u00a0% (95\u00a0% CI 92.8\u201399.6\u00a0%). It is concluded that the efficacy of AL for the treatment of uncomplicated falciparum malaria was high in this study.Unfortunately, the description of artemisinin resistance in Southeast Asia  and the P. falciparum and the activity of AL on gametocytes. Modeling studies have suggested that the spread of anti-malarial drug resistance is primarily driven by a \u201cwindow of selection\u201d after therapy, and the duration of this window is increased for drugs with longer elimination half-lives. The artemisinin drugs have short elimination half-lives and are thus less likely to select for resistance [Likewise, the high efficacy of AL has been reported in earlier studies from Ethiopia \u201335. Anotsistance , 40. Morsistance . In Cambsistance .P. falciparum malaria in southwestern Nigeria: fter treatment with AL, 8, 1, 1, 3, 3, and 3 children were gametocyte carriers on days 0, 3, 7, 14, 21, and 28, respectively [P. falciparum malaria during the next 2\u00a0years [Plasmodium species and kills gametocytes, the sexual stage of the malaria parasite responsible for infection of the mosquito. Artemisinins also kill immature and developing gametocytes, the sexual stages that are essential for transmission, thereby reducing transmission and contributing to malaria-control programs [In the current study the whole thick blood films were examined for the presence of the gametocyte stage, and there was a signifiant reduction of gametocyte carriage at day 28. Gametocyte carriage reduction after treatment was also seen in children with uncomplicated ectively . In addi 2\u00a0years . Furtherprograms .One of the limitations of this study was AL was given without fatty food. As well known, food enhances the oral absorption of artemether and lumefantrine. In healthy volunteers, the relative bioavailability of artemether increased by two- to threefold and that of lumefantrine by 16-fold when administered after a high-fat meal, as opposed to under fasting conditions .P. falciparum malaria and reducing gametocyte carriage in southwest Ethiopia. The result of this study supports the continuation of AL as a first line treatment for uncomplicated malaria, and there is presently no threat of artemisinin resistance developing in Southwest Ethiopia. This result can perhaps be generalized to areas with similar setting in East Africa.Eight years after the introduction of AL, this efficacy study revealed that it has been highly effective in the treatment of uncomplicated"}
+{"text": "P. vivax. To compare efficacy and safety of primaquine regimens currently used to prevent relapses by P. vivax of primaquine regimen 0.5 mg/kg/ day for 7 or 14 days compared to standard regimen of 0.25 mg/kg/day for 14 days. Efficacy of primaquine according to cumulative incidence of recurrences after 28 days was determined. The overall relative risk with fixed-effects meta-analysis was estimated.  A systematic review was carried out to identify clinical trials evaluating efficacy and safety to prevent malaria recurrences by  For the regimen 0.5 mg/kg/day/7 days were identified 7 studies, which showed an incidence of recurrence between 0% and 20% with follow-up 60-210 days; only 4 studies comparing with the standard regimen 0.25 mg/kg/day/14 days and no difference in recurrences between both regimens  were found. 3 clinical trials using regimen 0.5 mg/kg/day/14 days with an incidence of recurrences between 1.8% and 18.0% during 330-365 days were identified; only one study comparing with the standard regimen . High risk of bias and differences in handling of included studies were found. P. vivax. Clinical trials are required to guide changes in treatment regimen of malaria vivax. Available evidence is insufficient to determine whether currently PQ regimens used as alternative rather than standard treatment have better efficacy and safety in preventing relapse of  Plasmodium vivax is a major public health problem; about 20 million cases of malaria worldwide occur by this species, mainly in Asia and The Americas P. vivax, but two are prevailing 1) The pattern of tropical regions having a first relapse early and later relapses in short time intervals, usually every month; this pattern prevails in South America, Southeast Asia and Oceania Malaria caused by P. vivax, the combination of two antimalarials is used: A blood schizontocidal that eliminates blood forms and tissue schizontocide that eliminates liver forms P. vivax, such as chloroquine, amodiaquine, artesunate, artemether, lumefantrine, dihydroartemisinin, piperaquine, chloroquine remains the most widely used as first-line treatment for this species P. vivax For the treatment of P. vivax, these must be administered and supervised so patients must be monitored during the first post-treatment month in order to ensure the cure of the primary attack and then monitor relapses In order to evaluate the therapeutic efficacy of treatment regimens for P. vivax strain, the origin of the strain and the diagnostic method for evaluating recurrences The World Health Organization (WHO) recommends the use of PQ under the regimen 0.25 mg/kg/day for 14 days , for which a better efficacy has been demonstrated in six months compared with regimens shorter in 3 or 5 days of the same daily dose  P. vivax malaria, some countries are currently using alternative regimens such as applying twice the standard daily dose (0.5 mg/kg/day) during 7 days  or 14 days  Although most endemic countries follow the WHO recommendation to use PQ in the first-line treatment of P. vivax malaria. The secondary objective was comparing the PQ regimen of 0.5 mg/kg/day for 14 days with the standard regimen.The primary objective of this systematic review was to compare the efficacy and safety of the PQ regimen of 0.5 mg/kg/day for 7 days with the standard regimen of 0.25 mg/kg/day for 14 days in patients infected with uncomplicated A protocol (unpublished) was designed following the recommendations of the Cochrane collaboration for systematic reviews and the PRISMA guide  for the reporting of the results was followed P. vivax malaria and follow-up longer than 28 days. The main outcome was the cumulative incidence of recurrences after a 28-day follow-up since the first dose, defined as a P. vivax infection in patients who previously had an adequate clinical response to the treatment. As a secondary outcome, the treatment adverse events  were assessed. Clinical trials with and without random allocation evaluating the regimen of 0.5 mg/kg/day for 7 or 14 days in participants diagnosed with uncomplicated Plasmodium vivax) AND (recurrence OR recurren* OR relapse) AND primaquine\". For searching other databases, the keywords \" OR vivax AND primaquine\" were used. The references of the selected articles from the electronic search and the systematic and narrative reviews published previously were reviewed. The European database Open Grey was revised, in search of unpublished studies using the term \"primaquine\". Finally, the available reports of recent events by the ASTMH Annual Meeting, of the Latin American Parasitology Convention and of the Convention of the Colombian Association of Parasitology and Tropical Medicine were revised; for searching in these reports, the term \"primaquine\" was used, or \"primaquina\" when the information was only available in Spanish.Searches were performed on the following electronic data bases: MEDLINE, EMBASE, LILACS, SCIELO, the Central Register of Controlled Clinical Trials from the Cochrane Group and Clinical Trials. The search was done until August 20, 2015; unrestricted by language or publication date. The searching strategy in MEDLINE and EMBASE was: \" OR , the number of arms, the intervention and the comparison group, the adjustment of the PQ dose according to body weight, the diagnosis test used for detect recurrence, if the follow-up of participants took place in an endemic area, the population type, if the parasitaemia was or was not an inclusion criterion, the performance of tests for glucose-6-phosphate dehydrogenase (G6PD), the simultaneous or non-simultaneous administration of the PQ and the blood schizonticide, the PQ daily dose, the treatment time, the total dose, the treatment supervision, the basal characteristics (age and gender), the number of participants, the length of follow-up, the absolute frequency of recurrences, the cumulative incidence of recurrence and the adverse events associated with the PQ treatment.In order to assess the quality of the studies, the approach proposed by the GRADE  group was followed p< 0.10 value indicating heterogeneity, and the I2 statistic assuming significant heterogeneity if it was higher than 20%. The cumulative incidence of recurrences  was used as a summary measure for individual studies; besides, the relative risks ratio (RR) and their respective confidence intervals (95% CI) were calculated for studies that made the comparison with the standard regimen. For the primary objective, comparing the efficacy and safety of PQ 0.5 mg/kg/day for 7 days with the standard regimen, a meta-analysis of fixed effects was performed, with which the overall RR and its 95% CI was estimated. For the secondary objective, comparing the PQ regimen of 0.5 mg/kg/day for 14 days with the standard regimen, a meta-analysis could not be performed because only one study was found. The assessment of the heterogeneity of the results between studies was performed with the Chi Square test with a A total of 626 articles were identified in the electronic databases, 213 in MEDLINE, 344 in EMBASE, 49 in LILACS, 28 in SCIELO, 103 in CENTRAL of Cochrane and 33 in Clinical Trials; and, additionally, an article was identified in other sources. 376 were duplicated and 251 articles were screened, out of which 229 were excluded. Twenty-three articles were fully read for review in the second phase, out of which 9 clinical trials met with the eligibility criteria; 7 studies for the qualitative systematic review of the primary objective Table 1 shows the main characteristics of the studies. Three studies were conducted in Brazil P. vivax malaria. Only two studies considered a parasitemia greater than 1,000 parasites /\u00b5L Clinical trials included a total of 1,996 participants, 1,486 received PQ 0.5 mg/kg/day for 7 days and 510 participants received PQ 0.5 mg/kg/day for 14 days. The participants were recruited from the civilian population in eight studies et al., where a maximum of 45 mg per day were administered to participants whose body weight was over 70 Kg Seven clinical trials used PQ 0.5 mg/Kg/day for 7 days, in combination with a single dose chloroquine (CQ) of 10 mg/Kg The regimens used to compare the PQ 0.5 mg/Kg/day for 7 or 14 days were heterogeneous; three studies used an arm without PQ for comparison P. vivax malaria recurrences was the primary outcome in all clinical trials included. Eight studies reported the assessment of adverse events to treatment The percentage of cumulative incidence of Figure 2a shows the judgment of the authors regarding the items assessed about methodological quality for each of the included studies. Table 2 shows the results for each study. A cumulative recurrence incidence mean of 8.41% (range = 0%-22.80%) was found for PQ 0.5 mg/Kg/day for 7 days For the primary objective, a meta-analysis was performed, which showed that the rate of PQ 0.5 mg/kg/day for 7 days does not have an efficiency lower than that of the standard regimen (RR= 0.977 (95% CI= 0.670-1.423) For the secondary objective, only one study compared PQ 0.5 mg/Kg/day for 14 days with the standard regimen P. vivax relapses P. vivax malaria with other regimens. Including studies until 2012, they have shown that the total doses lower than the standard regimen have lower efficacy A priority for the control and elimination of malaria in the world is having an effective treatment to prevent We only included trials with a follow-up longer than 28 days, we identified four clinical trials that compared the standard PQ regimen with the PQ regimen of 0.5 mg/Kg/day for 7 days, and only one study comparing with the PQ regimen of 0.5 mg/Kg/day for 14 days. Additionally, we identified 5 clinical trials where one of these two alternative regimens was assessed, but without the comparison with the standard regimen. This systematic review could omit other studies, because here we only consider some of the search sources different from electronic databases  but the search for abstracts in other malaria-related conferences and contact with investigators to try to identify other unpublished or ongoing studies was not included.P. vivax malaria use at least one long half-life blood schizontocidal such as chloroquine; and generally the therapeutic response to them is assessed during the first post-treatment month Considering that recurrences can occur in less than 28 days after the primary episode 2 and I2 statistical test did not show such heterogeneity possibly due to the lack of power, important differences were found regarding the length of follow-up, the time when the study was conducted, the blood schizontocides used, in addition to the deficiencies in the conduction of studies that cast doubt on the validity of the comparisons. We observed a mean of 8.38% of recurrence incidence for the regime which doubles the total dose of PQ, i.e. 0.5/mg/Kg/day for 14 days; this value was similar to the mean for the standard regimen as reported in a previous systematic revision (9.9%) P. vivax strains according to the relapse pattern, there is some evidence of the simultaneous presence of several patterns in the same region Another source of heterogeneity in the results could be the origin of the participants, since there are different patterns of relapse by geographic region P. vivax relapses. Most studies had methodological limitations in the allocation concealment, in the blinding of participants and staff evaluating the outcome, and in the management of incomplete data; these limitations could bias results favoring efficacy (lower recurrence incidence) of the PQ regimen of 0.5 mg/Kg/day for 7 or 14 days in relation to the standard regimen.The evidence found in this systematic review is considered to have low quality due to the high bias risk identified in the clinical trials included or due to the lack of clarity for the items considered in the methodological quality assessment Eight out of the nine clinical trials included in this review assessed adverse effects to treatment, these considered mainly abdominal pain, nausea, vomiting, itching, dizziness, epigastric pain, hemolysis and methamoglobinemia, which are the major side effects for the PQ During the conduction of this systematic revision we identified 4 prospective observational studies evaluating PQ regimens of our concern P. vivax in the studies intending to assess recurrences, since it could contribute to a better approach to the therapeutic efficacy of PQ. Only two studies looked at the genetic characterization of recurrences to be classified as relapses or reinfections P. vivax relapses, though here it was found that apparently the 7-day regimen is not inferior than the standard one, this systematic revision suggests that given the small number of clinical trials reported and methodological limitations thereof, there is insufficient evidence to determine which PQ regimen used as first-line treatment for P. vivax in the world has the best efficacy and safety in preventing relapses. It is necessary to conduct clinical trials with a high methodological quality to compare these regimens and thus have therapeutic alternatives to prevent relapses, which may guide changes in treatment protocols for P. vivax malaria.The PQ regimen of 0.5 mg/Kg/day for 7 or 14 days is currently used in some countries instead of the standard treatment to prevent"}
+{"text": "Adherence to multidosing is challenging worldwide. This study assessed the extent of adherence to multidosing artemether-lumefantrine (ALu) in a rural community in Tanzania, six years after switching from single dose policy of sulphadoxine-pyrimethamine.This study was a prospective observational, open label, non-randomized study involving 151 patients with uncomplicated malaria recruited at Fukayosi dispensary in Bagamoyo district in Tanzania. Patients treated with ALu were visited at home on day 3 for interview on drug intake, capillary blood sample collection for microscopy and ALu tablets count. Venous blood samples (2\u00a0ml) for determination of blood lumefantrine concentrations and blood slides for microscopy were collected on day-7. Kappa\u2019s coefficient was used to assess agreement between pill count and self-report. Adherence was categorized depending on the tablets remaining and what the patient reported. Only those with empty blister pack available but no tablet remaining and reported taking all six doses of ALu at a correct dose and correct time were regarded as definite adherent. The rest were either probable adherent or probable non-adherent.Only 14.9% of the patients were definite adherent the rest took the drug at incorrect time or did not finish the tablets. Out of 90 patients with analysed plasma samples for lumefantrine blood concentrations, 13/90 (14.4.0%) had lumefantrine concentrations <175\u00a0ng/ml. There was no difference in mean lumefantrine concentration in the patients who stated to have taken all doses as required (561.61\u00a0ng/ml 95% CI\u2009=\u2009419.81-703.41) compared to those who stated to have not adhered well to drug intake . None of the patients had detectable parasites by microscopy on day-3 and day-7 regardless of adherence status and the level of day-7 blood lumefantrine. There was strong agreement between the self-reported responses on drug intake and pill-counts (kappa coefficient\u2009=\u20090.955). Age, sex, education and place where first dose was taken were associated with adherence.The overall adherence six years after the change of malaria treatment policy was low. It is, therefore, important to continuously monitor the level of adherence to treatment in order to get the current situation and institute corrective measures on time. Adherence to multi-dosing regimen is a global challenge facing both developed and developing countries ,2. AdherSince the introduction of ALu, a number of studies which assessed the extent of adherence have been published -12. AsseSeveral studies have demonstrated the usefulness of drug plasma concentrations in the assessment of adherence to drug intake ,5,10,14.et al..Baseline parasitaemia is shown in Table\u00a0et al. and Simba et al. reports [This study has demonstrated a relatively low adherence to ALu treatment regimen in a rural community in Tanzania, more than six years after ALu had been introduced as first line drug of choice for treatment of non-severe falciparum malaria. The study has shown low adherence to ALu, since only 10/143 7.0%) of the participants took all six doses at correct dosage and time. These findings disagree with what Kabanywani .0% of th reports .Despite the differences in adherence rates obtained by the use of different methods, no detectable parasite was observed by microscopy and fever subsided in all patients Table\u00a0. There wThis study applied patient/catetaker self report as measures for adherence Table\u00a0 and was To further ascertain the adherence obtained by self report and pill count, day-7 lumefantrine plasma concentration was also used as supplementary marker for assessing adherence to ALu intake Table\u00a0. LumefanIn this study the majority of patients (68.5%) preferred to take the first dose at home. The main reason given was the lack of water and/or food required for ALu intake at the dispensary and therefore majority initiated the dosing at home. It can be seen that about 40% of patients missed the correct timing of the second dose and a worse scenario is observed in dose 3 and the last dose .However it is important to note that, those who were definitely non-adherent were only 20.3% implying that, it is the component of those who were categorized as probably non-adherent (72.7%) that affected the overall adherence rate in this study. The low adherence rate hereby reported is mainly attributed by taking the tablets at an incorrect time interval (Figure\u00a0The present study has shown that education level had no significant influence on the extent of adherence in the rural population. There was no statistically significant difference between those who had no formal education and patients with at least primary education (The limitation of this study is a recall bias by some patients on the actual time for taking each dose in line with the dosage regimen. Lack of genetic profiles from the study population with regard to variation in drug metabolism among individuals is also a limitation to this study.status quo. This is important in instituting corrective measures without delay. It is important to continuously provide adequate adherence counseling to patients on the importance of taking the doses at the recommended time interval.Adherence to treatment with artemether-lumefantrine drug combination six years after the change of malaria treatment policy declined compared to what was reported at the point of policy introduction. The mainly reason that contributed to low adherence rate was due to poor timing of intake of the subsequent doses. Continuous monitoring of the extent of adherence to treatment is essential in ensuring the The authors declare that they have no competing interests.OMSM and SP conceived the study, and participated in its design, coordination, data analysis and writing of the manuscript. BN participated in the data analysis as well as manuscript preparation. SM participated in study design, data collection and analysis and contributed in the preparation and writting of the manuscript. All authors read and approved the final manuscript."}
+{"text": "The objective of this study is to show the pharmacokinetic (PK) profile of intravenous artesunate (AS), which was manufactured under good manufacturing practice (GMP) conditions, in adults with uncomplicated Plasmodium falciparum malaria. Plasma concentrations of AS and dihydroartemisinin (DHA) were measured using a validated liquid chromatography\u2013mass spectrometry (LC-MS/MS) methodology. Pharmacokinetic data were analysed with a compartmental analysis for AS and DHA.The PK parameters of intravenous AS manufactured under current cGMP were evaluated after a single dose of drug at 2.4 mg/kg infused over 2 min in 28 adults with uncomplicated max) of AS was shown to be 28,558 ng/mL while the Cmax of DHA was determined to be 2,932 ng/mL. Significant variability was noted in the PK profiles of the 28 patients tested. For example, Cmax values of AS were calculated to range from 3,362 to 55,873 ng/mL, and the Cmax value of DHA was noted to vary from 1,493 to 5,569 ng/mL. The mean area under the curve (AUC) of AS was shown to be approximately half that of DHA . The DHA/AS ratio observed was 1.94 during the one-day single treatment, and the AUC and half- life measured for DHA were significantly larger and longer than for AS.The results suggest there were no drug-related adverse events in any of the patients. After intravenous infusion, the concentration of the parent drug rapidly declined, and the AS was converted to DHA. AS and DHA showed mean elimination half-lives of 0.17 hours and 1.30 hours, respectively. The high mean peak concentration (CIntravenous AS can provide much higher peak concentrations of AS when compared to concentrations achieved with oral therapy; this may be crucial for the rapid elimination of parasites in patients with severe malaria. Given the much longer half-life of DHA compared to the short half-life of AS, DHA also plays a significant role in treatment of severe malaria. In vitro bioassay tests have shown that the activity of DHA is similar to AS[falciparum malaria and for treatment of severe malaria in Vietnam, Thailand, China, and Myanmar, however, limited studies have been carried out in Africa[Intravenous (IV) artesunate (AS) has been shown to completely inhibit parasite growth in infected human patients within two to four hours after dosing, and its active metabolite, dihydroartemisinin (DHA), is the only artemisinin derivative with activity against all asexual blood stage parasites. AS treaar to AS and threar to AS,7. AS han Africa. Currentn Africa. The Chin Africa.falciparum malaria infections[Following recent clinical studies with severe malaria patients, IV AS was shown to have the highest success for treatment of malaria with a low incidence of adverse events-12. The fections,12.falciparum malaria. The PK estimates will be also used to compare with the PK profiles observed in populations of healthy adults administered AS in another manuscript. The overall goal in the present study was to provide a well-validated, efficacious, and safe product for the treatment of severe malaria licensed by the US Food and Drug Administration (FDA) available for use in the USA and by US soldiers who encounter severe malaria infections.Alternatives to treatment for severe malaria are needed in the USA and Europe for treatment of severe malaria. While nations in the Third World can choose to use AS preparations such as the Guilin AS product for treatment of disease, there are issues with the use of this AS product in First World countries where the Guilin AS formulation cannot be sold legally. The objective of this study is to show the pharmacokinetic (PK) profile of IV AS in Kenyans infected with uncomplicated The bulk artesunic acid (4-(10\u2019dihydro-artemisinin-oxymethyl) succinate) substance was purchased from Knoll AG (Switzerland). BASF Pharmaceuticals rebottled it from the original company under GMP conditions. The clinical trial AS (Batch #: 14462\u201316) was tested for sterility and short-term stability. The formulation was contained in sterilized bottles with 110 mg artesunic acid per bottle. The injection buffer for AS was manufactured as a GMP phosphate salt with 0.3 M PBS (pH 8.0). The product was prepared for administration by reconstitution in sodium phosphate buffer prior to infusion, which took place within 1 hr after solution was prepared due to a precipitation in 3\u20134 hours at room temperature.Plasmodium falciparum malaria and body weights of 42\u201385 kg. The subjects of this study resided in a malaria-endemic region of Kenya and in districts surrounding the Province of Nyanza. The majority of the population in these areas is considered semi-immune, however, malaria attack rates in the city are low, and many adults in this area may well have waning immunity. The prevalence of malaria in Nyanza Province has been shown to range from 25 to 45%.All subjects gave written informed consent prior to the commencement of any study procedure . Overall, 30 adult subjects were enrolled, 20 males and ten non-pregnant females, aged 18\u201365 years, with uncomplicated Adult subjects were recruited with signs and symptoms of malaria who were seen at the Nyanza Medical Centre, Kisumu or one of its neighbouring health clinics , Nyanza Province. Local clinicians made a provisional clinical diagnosis of malaria, and these patients were subsequently referred to the hospital laboratory for microscopic examination of blood smears or rapid malaria antigen tests. Those subjects from the hospital laboratory who had not initiated therapy after a positive test were recruited for this study.P. falciparum malaria with at least 200 parasites per \u03bcL confirmed by malaria blood smear.The screening process for subject participation consisted of questionnaires and clinical examinations including medical history, physical examination, and laboratory analyses. Phlebotomy was performed to obtain samples for laboratory testing and malaria blood films. A clean-catch urine specimen was also collected for urinalysis and pregnancy testing, as applicable. Baseline safety testing included both clinical chemistry and haematology testing. Analyses included blood urea nitrogen (BUN), creatinine, aspartate aminotransferase (AST), alanine aminotransferase (ALT), total and direct bilirubin, reticulocyte, and a complete blood cell (CBC) count with differential assessment. Ultimately, enrolment determinations consisted of compliance with inclusion and exclusion criteria, including a diagnosis of uncomplicated This was a Phase II, open-label, non-randomized PK clinical trial of IV AS. Subjects completed an outpatient visit at the study centre for confirmation of study eligibility and baseline electrocardiogram monitoring. All subjects received an identical dose of the test article: injectable IV AS daily for two days consisting of 2.4 mg/kg delivered over 2 min. Safety, PK and efficacy assessments occurred intermittently with follow-on standard therapy with a three-day course of Malarone\u00ae to ensure cure. After completion of each injection, the study nurse examined how well AS injection was tolerated by performing inspection of the injection site and subject response. Normal activities, excluding strenuous exercise, were permitted four hours after dosing. The clinical trial identifier in ClinicalTrials.gov is NCT00298610. The humPK sampling was performed using a flush saline technique from an IV line or phlebotomy if the IV line was compromised. Samples were collected on day 1 approximately 10 min before dosing (0 hours), and at the following nominal time points after dosing: 5, 10, 20, and 40 min, and 1, 2, 4, and 6 hours. The blood (6 ml) will be collected from each subject via a cannula or by repeated venipuncture (opposite arm from infusion) into lithium heparin tubes. Actual time points were obtained within\u2009\u00b1\u200920% of the stated time points, and the plasma samples were immediately stored at approximately \u221270\u00b0C after collection, and shipped to the Armed Forces Research Institute of Medical Sciences (AFRIMS) Bangkok, Thailand on dry ice, for analysis. Finger-stick sampling was used for any malaria blood films collected at time points not requiring phlebotomy.Subjects were treated and monitored as outpatients or inpatients as deemed necessary by the subject, study staff and the principal investigator. Subjects managed as outpatients were permitted to leave the study site after the first dose of study drug (day 0) and sampling (approximately eight hours). Subjects were required to return the following day for scheduled follow-up.An LC-MS method for the quantitation of AS and DHA (100 \u03bcL) in human plasma was validated in a range from 1.4-1,153 ng/mL for AS and from 1.1-853 ng/mL for DHA. The analytes were extracted from human plasma after protein precipitation using two volumes of ice-cold acetonitrile. After centrifugation at 10,000 rpm, the clear supernatant extracts were analysed on a single quadrupole Mass Spectrometer  in the positive ion electrospray ionization (+ESI) mode. The AS and DHA compounds were monitored using single ion recording. The LC-MS with a reversed phase column  and a pre-column of the same material (2.1\u2009\u00d7\u200910 mm) was mounted on a liquid chromatography system . The method was performed a gradient elution with the following mobile phases: A) 6.25 mM ammonium acetate in water (pH 4.5) and B) 100% acetonitrile gradient from 20 to 40% in ammonium acetate buffer in 9 min at the flow rate of 0.4 mL/min. The ammonium-adducts of DHA were detected at a mass to charge ratio (m/z) of 302 and eluted in 5 min, while adducts of AS were detected at a mass to charge ratio of 402 and eluted in 6 min. The total analysis time for both compounds was 12 min.In these methods, the intra-assay and inter-assay accuracy and precision (n\u2009=\u20096) had an inaccuracy rate of\u2009\u00b1\u20098.4% and coefficient of variations (CVs) of \u22647.9% at 19.2/14.2 (50), 192/142 (500), 961/711  and 1,153/853  ng/mL (nM) for AS/DHA. The intra-assay and inter-assay inaccuracy and precision were within\u2009\u00b1\u20096.0 and \u226417.6%, respectively[In order to compare only the clinically relevant AS concentrations, a compartmental PK analysis was based on a short-term (2 min) IV infusion on individual patient level in accordance with a previous PK evaluation,16. The Cmax) were calculated from the corresponding model equations  at the respective time points. The AUCinf was estimated by direct integration of equation with CA modeling from zero to infinity. The total clearance (CL) is calculated with dose divided by total AUC. The Cmax and the time to maximum drug concentration (Tmax) were obtained from the PK modeling. The predicted AS concentration at the end of the infusion (2 min), however, was calculated by compartmental modelling. The values of the following PK parameters were derived by a compartmental method. The elimination half-life (t\u00bd), mean residence time (MRT), the AUC from time zero to the last sampling time (AUCt), the AUC from time zero extrapolated to infinity (AUCinf), total systemic clearance (CL), and the volume of distribution during steady state (Vss) were obtained by PK modeling.The drug plasma concentrations at the end of IV infusion , and the volume of distribution during steady state (Vss/F). The AUCinf ratios of AS to its metabolite (DHA) were calculated to determine exposure to any metabolite compared with the parent drug.Previous PK analysis and PK eIV AS manufactured to be cGMP compliant was administrated to 30 adult patients. PK samples were not collected for two subjects (Two cases have been excluded due to the insufficient data). Efficacy and safety evaluations will be discussed in detail in a separate manuscript.P. falciparum malaria were treated with an intravenous infusion of AS (2.4 mg/kg given over 2 min) daily for two days. For each subject, adverse events (AEs) were recorded throughout the post-dosing period. Multiple doses of this formulation of AS administered intravenously at a dose of 2.4 mg/kg daily for two consecutive days were well tolerated in all patients. No dose related toxicity was found for AS at the doses studied. There were no subject dropouts for AEs or other treatment-related issues. The dose-related decrease in reticulocyte count was noted that reached its lowest point four days after dosing and returned to normal by study day 7 in most cases. There were no other clinically significant laboratory abnormalities detected. No deleterious haemodynamic or electrocardiography (ECG) effects were seen. A transient, reversible sensation of altered or unusual taste, which lasted less than 30 min in all cases, was associated with intravenous dosing of AS. All remaining side effects were generally mild and all were reversible.In this study, 30 patients with uncomplicated max of AS was shown to be 28,558 ng/mL at the ending time of the infusion for the study population. The comparison results support the suggestion that PK analysis of AS may be performed only with compartment modelling[Vss were calculated to be 1,728 mL/h/kg and 139 mL/kg, respectively  of AS following IV infusion ranged from 3,362 to 55,873 ng/mL, and the AUC of AS ranged from 477 to 6,434 ng\u2009\u00b7\u2009h/mL, suggesting a large inter-individual variability in these PK parameters. Similarly, the Cmax of DHA ranged from 1,493 to 5,569 ng/mL, and the AUC values calculated ranged from 2,024 to 5,424 ng\u2009\u00b7\u2009h/mL.In the 28 malaria patients evaluated in this study, the peak concentrations  and non-compartment (model-independent) PK analytical techniques. PK evalanalysis,16. SimiDHA/AS ratio of 1.94, the peak concentration of AS was shown to be much higher than DHA with a CmaxAS/Cmax DHA ratio of 10.24.In this study and as previously reported, artesunate is rapidly converted to DHA. DHA wasCmax achieved after drug administration, and further studies have shown that AS is superior to other artemisinin analogues in terms of the very high Cmax observed following oral or IV administration[In this study, DHA was shown to have 7.65-fold longer half-life than that of AS. Therefore, the efficacy of AS has been attributed both to its own intrinsic activity and to the activity of its principle metabolite, DHA-4. The hstration. Other Pstration,20. A nustration.Cmax of AS ranging from 130\u201316,149 ng/mL in adult patients, demonstrating the significant variability  following intravenous AS administration  of 47.4-99.8% Table\u00a0, which iThe dose of intravenous AS currently used for clinical use was designed in accordance with prior experience of physicians over the last 25 years who were concerned with neurotoxicity and dose-dependent haemotoxicity and thus, the dose was limited to 2\u20132.4 mg/kg to prevent a submaximal anti-malarial effect, and to sustain a minimum curative rate >90%. The lowIn theory, patients with severe malaria require a peak concentration of AS ranging from 7,500-30,000 ng/mL to efficaciously exterminate malaria parasites where parasitaemia exceeds 5%,16. HoweThe study presented here underlines the need for appropriate PK analysis of AS and DHA following intravenous infusion. Injectable AS is a superior anti-malarial agent yielding very high peak plasma concentrations over a very short exposure time. The higher peak level serves to eliminate parasites rapidly, and the short exposure time aids in avoiding fatal neurotoxicity. PreviouThe authors have declared that they have no competing interests.SR and MP conceived the study. SR, BO and WO conducted the clinical trial study. PT, VM and BS conducted LC/MS/MS study. PW, SM and MH reviewed and edited this manuscript. QL analysed PK data and wrote the manuscript. All authors read and approved the final manuscript."}
+{"text": "Chlorproguanil-dapsone (CD) has been linked to hemolysis in symptomatic glucose-6-phosphate dehydrogenase deficient (G6PDd) children. Few studies have explored the effects of G6PD status on hemolysis in children treated with Intermittent Preventive Treatment in infants (IPTi) antimalarial regimens. We sought to examine the joint effects of G6PD status and IPTi antimalarial treatment on incidence of hemolysis in asymptomatic children treated with CD, sulfadoxine-pyrimethamine (SP), and mefloquine (MQ).A secondary analysis of data from a double-blind, placebo-controlled trial of IPTi was conducted. Hemoglobin (Hb) measurements were made at IPTi doses, regular follow-up and emergency visits. G6PD genotype was determined at 9 months looking for SNPs for the A- genotype at coding position 202. Multivariable linear and logistic regression models were used to examine hemolysis among children with valid G6PD genotyping results. Hemolysis was defined as the absolute change in Hb or as any post-dose Hb <8 g/dL. These outcomes were assessed using either a single follow-up Hb on day 7 after an IPTi dose or Hb obtained 1 to 14 or 28 days after each IPTi dose.Relative to placebo, CD reduced Hb by approximately 0.5 g/dL at day 7 and within 14 days of an IPTi dose, and by 0.2 g/dL within 28 days. Adjusted declines in the CD group were larger than in the MQ and SP groups. At day 7, homo-/hemizygous genotype was associated with higher odds of Hb <8 g/dL  and greater absolute reductions in Hb . There was no evidence to suggest increased reductions in Hb among homo-/hemizygous children treated with CD compared to placebo, SP or MQ.While treatment with CD demonstrated greater reductions in Hb at 7 and 14 days after an IPTi dose compared to both SP and MQ, there was no evidence that G6PD deficiency exacerbated the adverse effects of CD, despite evidence for higher hemolysis risk among G6PDd infants. Substantial progress has been made in malaria control over the last decade, with many malaria endemic countries now planning for malaria elimination . The patG6PD deficiency is the most common X-linked enzyme deficiency in humans, affecting more than 400 million people worldwide . Due to Pneumocystis carinii [While variants of G6PD deficiency appear to provide partial protection against malaria  it can a carinii , especia carinii .Plasmodium falciparum  malaria in response to concerns of growing resistance to chloroquine and sulfadoxine-pyrimethamine (SP) in Africa [et al. undertook a study to examine the protective efficacy and safety of three antimalarials\u2013SP, mefloquine (MQ) and CD in an area of high SP resistance in northeast Tanzania in search of alternative regimens to SP for Intermittent Preventive Treatment for malaria in infants (IPTi) [Pfdhfr) and dihydropteroate synthetase (Pfdhps) mutations, offers an explanation for this lack of efficacy in episodes of clinical malaria [In the late 1990s, chlorproguanil-dapsone (CD) was developed by a public-private partnership as a low-cost treatment for uncomplicated n Africa , 18. CD n Africa . Prior ts (IPTi) . At the  malaria . Regardi malaria . DespiteTo address this gap, using existing data from the Kilimanjaro IPTi Drug Options Trial, we explored differences in susceptibility to hemolysis and other adverse events among asymptomatic G6PDd infants treated with three antimalarial drugs in sub-Saharan Africa. Here, we examine the joint effects of G6PD status and IPTi antimalarial treatment on incidence of hemolysis. We hypothesized that incidence of hemolysis is increased among homo-/hemizygous infants treated with CD compared to placebo and other antimalarials, especially within 7 days following treatment. First, we contrast absolute changes in hemoglobin levels up to 7, 14 and 28 days after an IPTi dose by treatment. Then we estimate the impact of G6PD status on hemolysis among all infants who received active antimalarial treatment and assess for evidence of modification of the effects of IPTi treatment on hemolysis by G6PD status. Finally, we assess G6PD genotype effects on incidence of other adverse event episodes up to two years of age following IPTi.S1 Protocol). Briefly, the trial tested the protective efficacy and safety of three antimalarial regimens for IPTi. The trial was a randomized, double-blind placebo-controlled trial of SP, CD, and MQ, compared to placebo, conducted at neighboring moderate- and low-transmission sites in northeast Tanzania. Children aged 8\u201316 weeks who attended clinics for WHO\u2019s Extended Program on Immunization (EPI) were eligible for inclusion. Enrolled children who met the inclusion criteria were randomly assigned to receive full treatment doses of SP, CD, MQ, or placebo, given alongside routine immunizations at approximately 2, 3, and 9 months of age. Blood samples were collected before children were given their first and third course of IPTi. Blood was also collected on filter paper bloodspots (Whatman 3MM) using the buffy coat. Hemoglobin measurements were made at IPTi doses, regular follow-up and emergency visits. A total of 800 infants had their hemoglobin concentration systematically measured on day 7 after IPTi drug administration, including the first 200 infants to receive the first course of IPTi and the second 200 infants to receive the third course at each site. Every child was followed up at home on days 2 and 3 following treatment. During these visits, health workers assessed for possible adverse events and checked for adherence to drug regimens. In addition, all children were followed up at 10, 18, and 24 months of age. Additional follow-up visits at 11 and 12 months of age were done on random samples .This is a secondary analysis of data from a clinical trial that took place between 2004 and 2008. The drug trial has been described in detail elsewhere . The proet al. [G6PD screening was conducted using samples collected at routine visits at 9 months, focusing on the G6PD deficiency allele 202A G6PD A-, the most common in sub-Saharan Africa. Whole blood samples were collected and centrifuged on the day of collection. Plasma, buffy coat and red cells were stored at -20\u00b0C locally and transferred to the central laboratory for G6PD testing at a later date. Filter paper bloodspots (Whatman 3MM) for each patient were prepared using the collected buffy coat. DNA extraction from the impregnated filter papers was done using the Chelex method as previously described . The extet al.  to detecClinicalTrials.gov (identifier: NCT00158574).The clinical protocol of the original study was approved by the National Medical Research Coordination Committee of the National Institute for Medical Research of Tanzania (NIMR-MRCC) and by the London School of Hygiene and Tropical Medicine ethics committee and registered with For this analysis, only children whose G6PD status could successfully be determined were included.Homozygous females and hemizygous males were considered G6PDd and analyzed together, because of small numbers. Heterozygous G6PD females were analyzed as a separate category.The primary safety endpoint considered for this analysis was incidence of hemolysis. We used hemoglobin as a surrogate for hemolysis using two measures. First, hemolysis was defined as the absolute change in hemoglobin, from the day of the most recent IPTi dose. We also defined hemolysis as any post-dose hemoglobin measurement <8 g/dL (a measure of moderate anemia).For each outcome definition, we evaluated hemolysis using two approaches. The first restricted the analysis to children who had their hemoglobin concentration systematically measured on day 7 after an IPTi dose. The second considered any follow-up hemoglobin measurements obtained from 1 to 14 or 28 days after each IPTi dose administration, provided it was obtained before the next IPTi dose. The time frames for follow-up hemoglobin measurements were chosen to reflect what is known about the duration of drug-induced hemolytic episodes. Typically, within 24\u201372 hours of drug dosing, clinically detectable hemolysis and jaundice become apparent, worsening until days 7\u20138 . StudiesWe also assessed treatment and genotype effects on incidence of adverse events  throughout the study period.Directed acyclic graphs were used to identify potential confounders of G6PD status. In contrast to many conventional epidemiologic risk factors, genotype is not affected by most risk factors for hemolysis or malaria. However, G6PD deficiency does vary by sex, a potential risk factor for study outcomes. Moreover, genotype varies by distance above sea level (and accordingly site), a risk factor for malaria if not hemolysis, probably due to the selective pressure of malaria endemicity on the source population. Accordingly, genotype effect estimates were adjusted for these potential confounders.All analyses were performed using STATA 13.1 .Linear and logistic models were used for continuous and binary hemolysis outcomes measured only once for each infant. For repeated continuous and binary outcomes, mixed effects linear and logistic models with random intercepts were used. Analyses exploring G6PD genotype effects on hemolysis were restricted to children in the active arms, motivated by the hypothesis that adverse effects of G6PD deficiency would only be observed in infants receiving active treatment. Analyses were adjusted for sex and elevation, either directly or using inverse weighting by way of propensity scores for rare binary and failure time outcomes. Propensity scores, an alternative approach to standard adjustment for covariates, are useful when a binary or categorical exposure is common, but the binary or failure time outcome is rare, and when there are a large number of potential confounders that must be accounted for . In analPoisson models were used to estimate genotype effects on incidence of hemolysis and secondary adverse event outcomes, adjusting for treatment, gender, site, and elevation, with follow-up censored at 24 months of age or on the date of exit for children lost to follow-up due to migration, refusal, or exclusion. As in the main trial analysis, children were considered not at risk for malaria for 21 days after receipt of treatment with an antimalarial drug, not at risk for hemolysis for 28 days after a hemolytic episode [Fig 1).A total of 2419 children were enrolled in the original trial, of whom 1842 (76%) were screened for G6PD deficiency. Among those screened, 1557/1842 (85%) were successfully genotyped and included in this analysis. G6PD results were not obtained for the remaining 285 patients (15%) either because samples were not received or DNA extraction was not successful. Among the genotyped children, 1129 (73%) returned for at least one follow-up hemoglobin measurement within 28 days of an IPTi dose. In addition, a randomly selected 447 (28%) returned for a day 7 follow-up hemoglobin measurement (Table 1). Sex differences (p<0.001) are explained by the fact that the heterozygous genotype only exists in females. Children from the high transmission site (Korogwe) (p = 0.003) and those who lived at lower median elevations from sea level (p = 0.002) were more likely to be homo-/hemizygous. Homo-/hemizygous children also displayed lower median hemoglobin levels (p = 0.004) and slightly higher median weight values at enrollment (p = 0.005). Compared to children with successful G6PD genotyping results, included in this analysis, excluded children  were more likely to be male (p = 0.004), lived closer to the nearest clinic (p = 0.002), and displayed higher bednet coverage (p = 0.02) (S1 Table).Demographic and clinical characteristics at enrollment among children with valid genotyping results were generally similar across genotypes, with a few exceptions (Table 2). Adjusted declines in the CD group were also generally larger than in the MQ and SP groups, which did not differ from placebo.In analyses adjusting for genotype, sex, dose number, site, and elevation, treatment with CD reduced hemoglobin levels by approximately 0.5 g/dL 7 days after an IPTi dose, by a similar amount within 14 days, and by 0.2 g/dL within 28 days, compared to placebo (Table 3). This was an uncommon outcome, with only 6 events (2%) in the normal group (n = 281), 4 events (17%) in the homo-/hemizygous group (n = 24), and no events in the heterozygous group (n = 29) (p = 0.01). On day 7, there was also borderline evidence of a greater average decline in hemoglobin among homo-/hemizygous children compared to normal children . In contrast, we found little or no evidence for a genotype effect on either outcome within 14 or 28 days of an IPTi dose.In analyses examining the impact of G6PD status on hemolysis among 329 children in the active treatment arms, adjusting for sex and elevation using inverse weighting, we found a strong association of homo-/hemizygous genotype with hemoglobin <8 g/dL 7 days after IPTi treatment  (Table 4), but there was no persuasive evidence for treatment-genotype interaction in any of these analyses . Due to few events, we were unable to explore effect modification by G6PD status on hemogobin <8 g/dL at day 7 or within 14 or 28 days of an IPTi dose.In analyses stratified by genotype and adjusting for dose number, sex, site, weight and elevation, treatment with CD reduced hemoglobin by nominally larger amounts in the homo-/hemizygous group (range: 0.49\u20131.31 g/dL) than among normal genotype children (range: 0.18\u20130.48 g/dL) (S2 Table).During a median follow-up time of 21.8 months, 314 children experienced at least one episode of clinical malaria, for a total of 707 episodes, including 598 in the normal genotype group, 67 among heterozygous children, and 42 among the homo-/hemizygous group. There were also a total of 954 hospitalizations: 362 participants had one hospitalization, 155 had two, and 85 were hospitalized three or more times. There was no evidence for differences in malaria incidence or hospitalizations by G6PD genotype. Thirty-four children received blood transfusions, including three who required more than one transfusion, for a total of 38 events. Seven children died, including two in the homo-/hemizygous group (one in the CD arm); the remaining deaths were among G6PD normal children. These outcomes were too uncommon for meaningful statistical analysis , although mutations other than G6PD A- could have affected hemolysis risk and explain part of the observed results. However, this is unlikely to be important, because G6PD A- accounts for 90% of G6PD deficiency in Africa .P. vivax and P. falciparum. Both drugs also cause oxidant hemolysis in individuals with G6PD deficiency, and as malaria control programs begin to consider the use of primaquine or tafenoquine (if approved and licensed), identifying subgroups for which antimalarial treatment may be harmful or likely to have the largest benefit will become increasingly important. While this may prove challenging given the rarity of G6PD deficiency, even if fixed by design, pooled analyses with larger samples of G6PDd individuals could offer new insights. Interaction analyses can enable us to substantiate clinically important differences in hematological effects for oxidative antimalarial treatments across G6PD genotypes, and approaches to examine such trends in this important patient population are needed.These limitations render it challenging to find true subgroup effects, partially explaining why there is currently limited data concerning the hemolytic risk associated with diverse antimalarial drug regimens across G6PD genotypes in most populations. This analysis was underpowered to detect interaction; nonetheless, the direction of trends is consistent with expectations that IPTi with CD may produce greater reductions in hemoglobin among homo-/hemizygous children compared to other antimalarials. CD was withdrawn from the market in 2008 due to post-licensure hemolytic toxicity in patients with G6PD deficiency; however, dapsone is still used in the prevention and treatment of a variety of diseases \u201317, 36 aThis re-analysis of data from the Kilimanjaro IPTi Drug Options Trial among nearly 1600 children from northeastern Tanzania showed that treatment with CD as well as G6PD deficiency were associated with declines in hemoglobin and increased risk of moderate anemia. However, despite a fairly large sample, the study was underpowered to determine whether the adverse effects of CD are exacerbated among infants with G6PD deficiency, primarily because these deficient G6PD genotypes were uncommon. Combining data from a series of similar, well-conducted primary studies could offer a way to answer this question and optimize the targeting of treatment, which is increasing in importance as systematic treatment with drugs with hemolytic potential, such as the 8-aminoquinolines, primaquine and tafenoquine, becomes more widespread.S1 Fig(TIFF)Click here for additional data file.S1 Protocol(DOC)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file."}
+{"text": "Plasmodium falciparum in India has historically travelled from northeast India along the Myanmar border. The treatment policy for P. falciparum in the region was, therefore, changed from chloroquine to artesunate (AS) plus sulphadoxine-pyrimethamine (SP) in selected areas in 2005 and in 2008 it became the first-line treatment. Recognizing that resistance to the partner drug can limit the useful life of this combination therapy, routine in vivo and molecular monitoring of anti-malarial drug efficacy through sentinel sites was initiated in 2009.Anti-malarial drug resistance in Between May and October 2012, 190 subjects with acute uncomplicated falciparum malaria were enrolled in therapeutic efficacy studies in the states of Arunachal Pradesh, Tripura, and Mizoram. Clinical and parasitological assessments were conducted over 42\u00a0days of follow-up. Multivariate analysis was used to determine risk factors associated with treatment failure. Genotyping was done to distinguish re-infection from recrudescence as well as to determine the prevalence of molecular markers of antifolate resistance among isolates.P. falciparum cleared parasitaemia within 24\u00a0hours of treatment, but eight, including three patients who failed treatment, remained parasitaemic on day 3. Risk factors associated with treatment failure included age\u2009<\u2009five years, fever at the time of enrolment and AS under dosing. No adverse events were reported. Presence of dhfr plus dhps quintuple mutation was observed predominantly in treatment failure samples.A total of 169 patients completed 42\u00a0days of follow-up at three sites. The crude and PCR-corrected Kaplan-Meier survival estimates of AS\u2009+\u2009SP were 60.8% (95% CI: 48.0-71.4) and 76.6% (95% CI: 64.1-85.2) in Gomati, Tripura; 74.6% (95% CI: 62.0-83.6) and 81.7% (95% CI: 69.4-89.5) in Lunglei, Mizoram; and, 59.5% (95% CI: 42.0-73.2) and 82.3% (95% CI: 64.6-91.6) in Changlang, Arunachal Pradesh. Most patients with AS\u2009+\u2009SP treatment failure was widespread in northeast India and exceeded the threshold for changing drug policy. Based on these results, in January 2013 the expert committee of the National Vector Borne Disease Control Programme formulated the first subnational drug policy for India and selected artemether plus lumefantrine as the new first-line treatment in the northeast. Continued monitoring of anti-malarial drug efficacy is essential for effective malaria control. Plasmodium falciparum was a major contributor to global resurgence of malaria in the 20th Century. Southeast Asia (SEA) has been the focus of drug resistance for all anti-malarials with loss of valuable drugs from time to time. Chloroquine resistance in P. falciparum malaria was first reported in 1957 in SEA region, later followed by sulphadoxine-pyrimethamine resistance after ten years [P. falciparum[Plasmodium vivax, another parasite that contributes equally to human malaria in the country.Anti-malarial drug resistance, particularly in en years . India sen years . HoweverP. falciparum malaria in the country is based on artemisinin combination therapy (ACT) as per the recommendation of World Health Organization (WHO) [dihydropteroate synthase (dhps) and dihydrofolate reductase (dhfr) genes, both coding for essential enzymes in the folate biosynthesis pathway, led to resistance to antifolate drugs [Since 2005, treatment of uncomplicated on (WHO) . The ACTon (WHO) . Howeveron (WHO) , which tte drugs .Recent reports on artemisinin resistance in the SEA region  also reqdhfr plus dhps) are potential early warning signs; the same were also monitored for the study sites which prompted more focus in these study areas.The Indian northeastern region has varied ecological diversity, inaccessible areas where vector control measures as well early treatment and diagnosis facilities are difficult to assess. Malaria control remains a challenge in these areas. ACT was introduced in the study areas in 2007 and since 2009 continuous monitoring of recommended anti-malarials has been done through nationwide sentinel site system. Also, partner drug resistance markers ; from August to September 2012 in Tripura ); from September to October 2012 in Arunachal Pradesh . The study sites  cases.The state contributes 1.3% to the country\u2019s malaria burden . The peaPlasmodium falciparum is a major malaria parasite in this region, causing 70 to 90% of malaria infections. The hot and humid climate in the region is ideal for survival and multiplication of malaria vectors [Tripura is one of the states in the northeastern region which shares a long international border with Bangladesh. The hilly and undulating terrain and the movement of people across the border have led to persistence of malaria in villages near the border. The regions adjacent to the Indo-Bangladesh border are mostly covered with thick forests and have poor communication and health infrastructure.  vectors .P. falciparum has increased from 56 to 64% in the last five years.Changlang district in the state of Arunachal Pradesh is bounded by Assam and Arunachal Pradesh in the north and Myanmar in the southeast. The state contributes 1.1% of the country\u2019s malaria burden . The capin vivo monitoring was followed [This was an open-label, one-arm prospective study of clinical and parasitological responses after administration of ACT treatment and WHO protocol for followed .Informed, written consent was obtained from enrolled adults and from a legal guardian of each child enrolled. The study protocol was approved by the Institutional Ethics Committee of the National Institute of Malaria Research (NIMR) in New Delhi. These studies were part of monitoring and surveillance studies jointly carried out by NIMR and National Vector Borne Disease Control Programme (NVBDCP) with added quality assurance, monitoring and molecular markers.P. falciparum with parasitaemia between 500 and 100,000 asexual parasites/\u03bcl blood, absence of other febrile conditions and informed consent. Patients having severe malnutrition as per WHO guidelines, severe malaria or danger signs and inability to come for follow-up visits were excluded from the study.Adults and children over 6\u00a0months presenting with fever (axillary temperature \u2265\u200937.5\u00b0C) at visit or a history of fever for the preceding 24\u00a0hours were included in the study. Other criteria for inclusion were mono-infection with P. falciparum received artesunate (AS)\u2009+\u2009SP (AS 4\u00a0mg/kg for three days plus SP 25/1.25\u00a0mg/kg single dose on day 0) and primaquine (PQ) (0.75\u00a0mg/kg) on last day of treatment. The treatment was directly observed on all three days and used quality assured drugs through state government supply which consisted of manufacturers (site name in brackets): Medicamen Biotech Ltd, India (Lunglei), Medico Remedies Pvt Ltd, India (Gomati), ZEST Pharma, India (Changlang). These drugs are supplied in the trade name of \u2018Antimalarial combi blister pack\u2019. All drugs were used within their expiry period and batch number and expiry dates were recorded in each case record form (CRF). The dosing was based on age with five categories of combi blister packs with age category\u2009<\u2009one year, one to four years, five to eight years, nine to 14\u00a0years and >15\u00a0years (adult pack). The study followed the standard WHO protocol for assessment of therapeutic efficacy of anti-malarial drugs for uncomplicated falciparum malaria for moderate transmission [msp1, msp2 (merozoite surface protein) and glurp (glutamate rich protein) gene loci of pre- and post-treatment sample pairs. The outcome of treatment with PCR correction was based on the number of true recrudescence excluding cases of novel infections. In addition, pfdhfr and pfdhps mutations for partner drug resistance were used as molecular markers to analyse different samples.Patients with uncomplicated smission . On enroThick and thin blood films were collected and stained with Giemsa. Slides were examined on day 0 by experienced microscopist(s) for species identification and quantification of parasites. Slides were also prepared on days 1, 2, 3, 7, 14, 21, 28, 35, and 42 to determine asexual and sexual parasite density by counting the number of parasites against 200 white blood cells (WBCs), and were expressed assuming WBC count to be 8,000/microlitre. A slide was considered negative when counting 1,000 WBC in thick smear did not show asexual parasites. All the slides were cross-checked at NIMR, Delhi.msp2 or glurp. The third marker analysed was msp1 to differentiate recrudescence from new infections [pfdhfr gene and pfdhps gene at codon 436, 437, 540, 581, 613 were also analysed in the day 0 samples.Paired blood samples of patients collected on day 0 and the day of recurrent parasitaemia (14\u201342 days) were analysed sequentially starting with the highest discriminatory marker, fections . A new ifections . In addimsp1 , two allelic families of msp2 (Fc 27 and Ic) and glurp. Pfdhfr and pfdhps gene products were PCR amplified using earlier reported methods [dhfr and dhps gene. Applied Biosystem thermocycler was used for all PCR amplification reactions. Digested PCR product (5\u20138 microlitre) was analysed on 1.5% agarose gel containing ethidium bromide (0.5\u00a0\u03bcg/ml) and 0.5X TBE running buffer (pH\u00a08.0). PCR products were visualized under UV transilluminator (280\u00a0nm) and digitally captured with the help of gel documentation system (Alpha Imager EP). Molecular sizes of PCR fragments were calculated using gene tool .Genomic DNA was isolated from blood spots using QIAamp DNA minikit, Germany. Genotyping PCR assays were carried out following the protocols reported earlier . Separat methods  and thenThe case record forms were completed for each patient and all the clinical and parasitological data from days 0 to 42 were recorded. The data were entered in WHO software  and bothAll the patients with ETF or LTF were treated with oral quinine in standard dose of 10\u00a0mg/kg body weight, three times for seven days. CRFs were used to capture adverse events, if any.P. falciparum were categorized as being \u2265\u200987.5%, <\u200987.5 to 75% and <\u200975% of the recommended dose per kg. The parasite clearance time (PCT) was recorded in CRF till day 2 of treatment, although slides were made on day 3 as well. However the delayed PCT (PCT \u2265\u200972\u00a0hours) was calculated on day 3 based on the mean PCT up to day 2\u2009\u00b1\u2009SD. Kaplan-Meier survival analyses of treatment failure and parasite clearance were conducted with and without the results of the PCR-based identification of the parasitaemia that resulted from post-treatment re-infection. Log-risk models were used to evaluate the multivariate associations observed between risk factors and endpoints. Multivariate analysis was based on full models that included age, sex, fever at enrolment, level of parasitaemia at enrolment, infection with a parasite that harboured any of the investigated mutations in dhfr or dhps, and AS dose, as well as the interactions between age and AS dose, age and level of parasitaemia on enrolment, and between presence of mutations and level of parasitaemia on enrolment. A complete case (per protocol) analysis was performed. A strategy of backward elimination was followed in which p-value of <\u20090.10 and <\u20090.15, respectively, were used as the cut-offs in eliminating individual factors and interaction terms as statistically significant risk factors for treatment failures. All data analysis was performed using version 7.2 of a software package developed by WHO\u2019s Global Malaria Programme for evaluating therapeutic efficacy or SPSS version 14.Patient and demographic variables that could be associated with treatment failure were investigated. Using the body weights recorded on day 0, the doses of AS given to patients with P. falciparum receiving AS\u2009+\u2009SP treatment on first three days and PQ on last day of treatment were enrolled from Lunglei district in Mizoram (71), Gomati district in Tripura (77) and Changlang district in Arunachal Pradesh (42), the three far- flung regions of northeast India. Fifteen patients were withdrawn after cross-checking due to mix infection, presence of other species, or out of range parasitaemia at the time of enrolment. Thus, 175 patients were found to be eligible, 169 (96.6%) of whom completed the 42\u00a0days of follow-up  of the patients were confirmed as treatment failure, 24 were found to have P. falciparum re-infection and one had P. vivax infection on day 21. These 32 patients who were confirmed to have failed AS\u2009+\u2009SP treatment came from Lunglei (11), Gomati (15) and Changlang (six)  in <\u200924\u00a0hours, 35.5% (60 of 169) in intervals of 24 to <48\u00a0hours, 8.8% (15 of 169) in 48 to <\u200972\u00a0hours and 4.7% (eight of 169) of the patients in \u2265\u200972\u00a0hours, respectively  relative to adults 15\u00a0years or older and low administered dose of AS  relative to 3.5\u00a0mg/kg or more were associated with higher risk. The risk of failure increased as the dose of AS decreased. However, optimal or higher dose of SP was observed in 90.6% patients. Fever at enrolment was positively and significantly associated with treatment failures. Also, relative risk was highest with the presence of triple or quadruples mutation  and triple mutations (n\u2009=\u200969) were common. The most frequent haplotype was double mutant 108/59 followed by triple mutant 108/59/51. Only five isolates were single mutant while three were found to have quadruple mutation (5.2%).Out of the 190 isolates, 155 could be successfully genotyped for dhps, triple (n\u2009=\u200975) and quadruple mutations (n\u2009=\u200935) were common. Mutations in codon 437 and 540 were most frequent and the most frequent haplotype was triple mutant 540/437/436 followed by quadruple mutant 581/540/437/436  recommended for This study provides the first report on the emerging treatment failure of AS\u2009+\u2009SP in remote areas of the country, including areas directly across the border from Bangladesh and Myanmar where artemisinin resistance has been confirmed recently. Treatment failure to this ACT is now evident at all the three study sites of northeast India. The failure of AS\u2009+\u2009SP was likely as the molecular marker of partner drug resistance showed increasing trend since 2009 in the nationwide sentinel site monitoring where 20% random samples were assayed to monitor drug resistance at molecular level. The trend showed increase from single to double mutation in majority of the samples. These studies were part of nationwide sentinel site monitoring system, where 15 sentinel sites are being monitored across the country to ascertain the efficacy of recommended anti-malarials against the predominant species of Plasmodium.Based on the results of this study, the expert committee of the NVBDCP created the first subnational drug policy for India and selected artemether plus lumefantrine as the new first-line treatment in the northeast .ACT treatment failure >\u200910% is the cost-effective threshold and requires change of treatment policy for malaria along with continuous monitoring in adjacent areas . In the dhps remained wild in majority of the samples, however, increasing trend in dhps point mutations were observed later, particularly in northeastern region. These findings prompted more focus in northeastern region. Thus, three sites at different ecological conditions were selected and efficacy of AS\u2009+\u2009SP in P. falciparum was studied.Until recently, the efficacy and safety of recommended ACT (AS\u2009+\u2009SP) was high at multiple sites in the country . The natdhfr and dhps gene was observed from all the three study sites with higher frequency in dhps gene. Sulphadoxine resistance in P. falciparum is associated with mutations at five pfdhps codons; 436ala/phe, 437gly, 540glu, 581gly, and 613ser [pfdhps mutation at codon A436 causes alteration in binding of sulphadoxine followed by sequential mutations at G437, E540, G581, and T/S613 which may cause increase in sulphadoxine resistance [Data on molecular studies for SP resistance are conclusive with increased frequency of quintuple mutation over time. Increased frequency of quadruple mutation in d 613ser . The pfdsistance . Mutatiosistance .pfdhfr codons (108asn\u2009+\u200951ile\u2009+ 59arg) and two pfdhps codons (437gly\u2009+\u2009540glu) [A strong indicator for SP treatment failure is the quintuple mutations in three \u2009540glu) . Quintup\u2009540glu) .in vitro and in vivo has been proposed [Following these findings, further new studies on tracking resistance to artemisinin are being undertaken at two sites in this region. Recently association of K13-propeller polymorphism with artemisinin resistance proposed . FurtherVarious factors including host immunity and pharmacokinetics of the drug play an important role in achieving complete cure or treatment failure in patients. In the present study, younger age category (<five years) relative to adults 15\u00a0years or older, fever at enrolment and low administered dose of AS  relative to 3.5\u00a0mg/kg or more were associated with higher risk of treatment failure. Negative correlation was observed between the risk of failure with the dose of AS (in mg/kg body weight). Although, the recommended daily dose of AS is 4\u00a0mg/kg, 15.6% of treatment failure patients received 3.0 to <\u20093.5\u00a0mg of AS per kg, and 9.4% received <\u20093.0\u00a0mg/kg. Due to variation in body weight within age categories, dosing of drugs based on age category presents the risk of under-dosing or over-dosing. Thus, the criteria to use age rather than body weight for calculating the dose of anti-malarial needed for a patient is the probable cause of suboptimal dosing. However, optimal or higher dose of SP was observed in majority of the patients with only 9.4% patients receiving under-dose of SP. Fever at enrolment was positively and significantly associated with treatment failures. Earlier results of nation-wide sentinel system also observed younger age, fever and low AS dose as the potential risk factors for treatment failure . Partnerin vitro monitoring in the northeast for early warning of this ACT is required.Recently increasing failure rates of artesunate mefloquine in Thailand and dihydroartemisinin piperaquine in Cambodia have been reported ,33. WithThe efficacy of AS\u2009+\u2009SP was declining in northeast India and exceeded the threshold for changing drug policy. Based on the results of the study, the expert committee of the National Vector Borne Disease Control Programme formulated the first subnational drug policy for India and selected artemether plus lumefantrine as the new first-line treatment in the northeast. Continued monitoring of anti-malarial drug efficacy in northeastern region is essential for effective malaria control.The nation-wide sentinel site monitoring system created in 2009 accelerated the assessment of emerging ACT treatment failure in areas with international borders and is vigilant to detect delayed parasite clearance, if any. Few of the earlier reported limitations of nationwide sentinel system were overcome during 2012. Firstly, follow-up was extended up to 42\u00a0days, delayed parasitaemia was monitored by looking at parasite clearance interval of \u2265\u200972\u00a0hours; partner drug resistance markers were analysed in all the isolates. The other limitations include 24\u00a0hours sampling for parasite monitoring, as the patients were from distant places. The sample size was relatively low in Changlang district, Arunachal Pradesh. To ascertain the role of partner drug resistance, drug levels of SP could not be done as the samples for drug concentration could not be collected.The authors declare that they have no competing interests.NM, NV and GSS were responsible for the design of the project proposal and monitored progress. NM, KK, BS, JPN and RSB were involved in quality check of data and molecular studies. NM, VD and SP monitored the studies at the site. NM, RR and NKS compiled and performed statistical analysis. NM and NKS wrote the first draft and ACD, GSS, ARA and NV corrected the draft and all authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum resistance to artemisinin combination therapy (ACT) is a worrying development. It calls for close surveillance to monitor the efficacy of the drugs. The objectives of this study were to determine the performance of SD Bioline malaria AgPf(HRP-2/pLDH) 3 band Rapid Diagnostic Test (RDT) against Giemsa-stained blood smear and evaluate the suitability of this test in assessing the therapeutic efficacy of ACT in pediatric malaria patients in the Democratic Republic of the Congo (DRC).The emergence of P. falciparum in Kinshasa, DRC. Of the 166 patients who tested positive for P. falciparum at recruitment (day 0), 103 consented to participate in this study and were followed up and retested for P. falciparum on day 3, day 7, day 14, day 21 and day 28.Five hundred and one patients with malaria symptoms were screened for pLDH band. Some patients turned positive during follow up but pLDH readings remained consistent with blood smear readings.Sensitivity and specificity of the test were significantly high on day 0 and so were their positive and negative predictive values. Higher proportions of false positive cases were observed on the HRP-2 band irrespective of patient parasite densities during the follow up but these were barely seen on the Pf(HRP-2/pLDH) RDT demonstrated high performance in DRC. Thus, the test can be employed to assess the efficacy of ACT in pediatric malaria patients and prioritize areas that require the deployment of advanced testing like polymerase chain reaction (PCR).SD Bioline malaria Ag Plasmodium falciparum antigen histidine-rich protein 2 (HRP-2), (ii) RDTs that detect lactate dehydrogenase (pLDH) from P. falciparum and (iii) RDTs that detect P. vivax aldolase in human whole blood [Commercially available rapid diagnostic tests (RDTs) are numerous in the market. They all fall into three categories: (i) RDTs that detect the le blood .P. falciparum (Pf) HRP-2 remains in the bloodstream for an extended time following a successful clearance of the parasite, thus, contributing to false positive results and limiting specificity of the test category that targets PfHRP-2 [pLDH is known to be rapidly cleared from the bloodstream and becomes undetectable at about the same time as blood smears become negative after a successful anti-malarial therapy [Generally, it is known that  therapy , 4.pLDH-based tests seem to be more useful since they become negative soon after parasites are cleared from the blood [pLDH-based tests better discriminate false positive results from treatment failure than HRP-2 tests. SD Bioline malaria AgPf(HRP-2/pLDH) RDT has been reportedly proven to be effective in reducing the rate of false positives caused by the persistence of HRP-2 after a successful drug therapy and guiding accurate prescription and monitoring efficacy of drug therapy in countries like Burkina Faso [In order to monitor the success of an anti-malarial drug therapy, particularly in the presence of fever, he blood . Thus, pina Faso . HoweverPf(HRP-2/pLDH) 3 band RDT in symptomatic pediatric malaria patients against microscopy of Giemsa-stained blood films and to evaluate the effectiveness of SD Bioline malaria AgPf(HRP-2/pLDH) in monitoring the efficacy of ACT (artesunate amodiaquine) among pediatric patients in Kinshasa, DRC where P. falciparum transmission is characterized as high [The objective of this study was to determine the performance of SD Bioline malaria Ag as high .Pf(HRP-2/pLDH) RDT were collected from the patients on the day of recruitment (day 0). Blood slides were stained with Giemsa 3% for 30 minutes. The first slide readings were performed at the study site while the second readings were conducted at the National Biomedical Research Institute in Kinshasa. Those with discrepant results were read by a third reader under the supervision of the principal investigator, a parasitologist, until when the final agreement was reached. Technicians reading blood slides were blinded to the results of the SD Bioline malaria AgPf(HRP-2/pLDH) RDT. Likewise, those reading the RDT were blinded to the results of the blood slides.This study was conducted at Tembo Health Center in Kinshasa in DRC from October to December 2012. The health facility was chosen because of its relatively high number of pediatric patients in the 3 months preceding this study. This high number of pediatric patients was used as indicative of the availability of sufficient study participants. All pediatric patients who presented with symptoms of malaria during the study were invited to participate. After obtaining consent from patients\u2019 parents or guardians, demographic and clinical data were collected including previous anti-malarial drug intake. Blood samples (finger prick or venous blood) for thick and thin blood films and for the SD Bioline malaria AgPf(HRP-2/pLDH) RDT was performed according to the manufacturer's instructions and read after 15 minutes by the first laboratory technician and then after 30 minutes by the second laboratory technician who was blinded to readings from the first technician. In case of a discrepancy between the readings, the test was repeated. Results were recorded as follows: (i) when the 3 bands of the test kit were all colored, the test was positive for both PfHRP-2 and PfpLDH, (ii) when bands C and T1 were colored, the results were positive for Pf HRP-2 and not for PfpLDH, (iii) when bands C and T2 were colored, the results were positive for PfpLDH and not for PfHRP-2 and (vi) when only band C was colored, the results were negative. Parasite density was computed by counting the number of parasites/200 white blood cells (WBC) and assuming a WBC count of 8,000/\u00b5l [SD Bioline malaria Ag8,000/\u00b5l .pLDH in the blood, respondents were followed for 28 days from day 0. Thus, besides day 0, blood samples for thick and thin blood films as well as for the SD Bioline malaria Pf (HRP-2/pLHD) RDT were collected on day 3, day 7, day 14, day 21 and day 28. The tests were performed following the same procedure as described above.Positive cases of malaria were managed as recommended by the DRC National Malaria Control Program and World Health Organisation (WHO) . To moniSensitivity of the RDT assay is defined herein as its ability to correctly detect blood specimens containing malaria antigen. It is the percentage of true positive (TP) malaria specimens identified by the RDT as positive divided by the number of specimens identified by the gold standard assay (Giemsa-stained blood smear) as positive (TP)+false negative (FN).Specificity of the RDT assay is its ability to correctly discriminate specimens that do not contain malaria antigen. It is the percentage of true negative (TN) specimens identified by the RDT as negative divided by the number of specimens identified by the gold standard assay as negative (TN)+false positive (FP).Positive predictive value (PV+) is the probability that when the test is reactive, the specimen actually contains malaria antigen. PV+ is calculated as TP/(TP+FP). Negative predictive value (PV-) is the probability that when a test is negative, a specimen does not have malaria antigen. PV- is calculated as TN/(FN+TN).Pf(HRP-2/pLDH) RDT was assessed by measuring its sensitivity, specificity, positive and negative predictive values against blood smear microscopy as the gold standard [Data were entered into MS Excel  independently by two people. The two datasets were then compared and discrepancies were corrected to create the final version that was analyzed using IBM SPSS Statistics version 20 . The performance of SD Bioline malaria Agstandard .PfHRP-2/pLDH antigen present in <1,000 parasites/\u00b5l, 1001 - 5,000 parasites/\u00b5l, 5,001 - 50,000 parasites/\u00b5l and in > 50,000 parasites/\u00b5l on day 3, 7, 14, 21 and 28. In other words, this represented the percentage of participants who tested positive for AgPf(HRP-2/pLDH) RDT while having a negative blood smear test for a given parasite density level and day of follow up. We compared groups using Fisher's exact test. The level of significance was set up at p<0.05.The proportion of false positives among the children were estimated as a function of This study was first approved by the DRC National Biomedical Research Institute's Research Committee. Written informed consent was sought from parents or guardians of the children who participated in this study. The parents or guardians were assured that the data would be kept confidential and used only for research purposes. Participants who tested positive for malaria were given standard malaria management as stipulated by the DRC National Malaria Control Program and WHO.P. falciparum in this study. Their mean (standard deviation) age was 26.9 (5.8) months. The youngest was 3.4 months old while the oldest was 53.2 months old. Two hundred and thirty six (47.1%) participants who were screened were females and 265 (52.9%) were males. Three hundred and thirty five (66.9%) participants tested negative for malaria while 166 (33.1%) tested positive for malaria infection. Parents/guardians of 103 children (out of the 166) gave consent to participate in the follow up part of the study, giving a response rate of 62.0%.In total, 501 children were screened for Pf(HRP-2/pLDH) RDT performance against blood smear microscopy as the gold standard are presented in Pf(HRP-2/pLDH) RDT exhibited high sensitivity, specificity, positive and negative predictive values. Pf(HRP-2/pLDH) RDT by level of P. falciparum parasitaemia. The sensitivity of SD Bioline malaria AgPf(HRP-2/pLDH) ranged from 85.1% to 100%. The RDT exhibited its lowest sensitivity when the parasitaemia was less than 10,000/\u00b5l.The frequency distribution of the study participants by parasite density level is presented in P. falciparum on day 0 and their parents or guardians consented to participate in this study were followed up till day 28. They were 48 females and 55 males. Their mean age (standard deviation) was 31.4 (7.4) months. The youngest was 6 months old and the oldest was 50 months. After treatment, on day 3 and day 7, all patients tested negative by blood smear microscopy. On day 14, two patients became positive by blood smear microscopy and two were lost to follow up. On day 21, four more patients became positive and six were lost to follow up. On day 28, eleven other patients became positive making a total of 17 patients who became positive. These cases represented either treatment failure or new infections and six more were lost to follow up making a total of 14 children lost to follow up. The performance of SD Bioline malaria AgPf(HRP-2/pLDH) against blood smear microscopy by day of follow up is presented inOne hundred and three patients who were positive for pLDH on day 3, 7, 14, 21 and 28. Band HRP-2 had higher proportions of false positives than pLDH. Though the proportion of false positives dropped with days of follow up until day 28 it remained significantly higher in band HRP-2. The proportions of false positives in band pLDH were significantly (p<0.05) lower than those in band HRP-2. In the pLDH band, except for patients having parasite densities of 5,001-50,000/\u00b5l where a few cases of false positives were observed till day 14, no single case of false positives was observed among patients with parasite densities \u2264 1,000/\u00b5l on day 3, 7, 14, 21 and 28. There were no false positives among children with parasite density \u2264 50,000/\u00b5l on 7, 14, 21 and 28 in band pLDH.Pf(HRP-2/pLDH) RDT among pediatric patients with symptomatic malaria in Kinshasa, DRC was high. The test attributes for testing and monitoring the efficacy of ACT treatment among pediatric patients in the population studied were reasonably high. The sensitivity and specificity of the test were found to be high. The corresponding positive and negative predictive values were also high indicating the high performance of this test.The main finding of this study is that the performance of SD Bioline malaria AgPf(HRP-2/pLDH) RDT in this population, the probability of testing positive when one truly has malaria is more-or-less 100% which indicates high performance of this test in Kinshasa, DRC.Although the sensitivity of the test went a bit down to 85.1% among patients with parasite densities \u2264 1,000/\u00b5l, this study showed sensitivity of about 100% for parasite densities > 1,000/\u00b5l. Similar results are found in the literature , 10. ThePf(HRP-2/pLDH) for monitoring the efficacy of ACT treatment showed higher proportions of false positives with respect to the HRP-2 band irrespective of patient parasites density and day of follow up. This means that readings from the test band HRP-2 are not suitable for monitoring the efficacy of ACT treatment. The fact that false positive cases were barely seen on the pLDH band makes this band the most suitable for monitoring the efficacy of ACT treatment. Though few cases of false positives were observed among patients with parasite densities of 5001- 50,000/\u00b5l up to day 14, no single case of false positive was observed among patients with other parasite densities up to the end of the follow up. The concomitant capability of the pLDH band to turn negative when blood smear microscopy is negative or positive when blood smear microscopy is positive as demonstrated herein is an attribute that makes SD Bioline malaria AgPf(HRP-2/pLDH) RDT a suitable test for monitoring the efficacy of ACT treatment in this area of highP. falciparum transmission.The evaluation of the performance of SD Bioline malaria AgP. falciparum transmission in Kinshasa, the major limitation of this test is its inability to differentiate between true treatment failure and reinfection cases as in total 17 cases (16.5%) became positive before the end of the study after initial successful treatment. Whether these were true cases of treatment failure or cases of new infection, only a polymerase chain reaction (PCR) procedure can differentiate. PCR is not easily accessible in this resource-poor country. Thus, SD Bioline malaria AgPf(HRP-2/pLDH) RDT can serve as a guide and a prioritization tool for monitoring the efficacy of ACT treatment in communities were the study was conducted. Once decreased ACT efficacy is suspected in a community, PCR should then be employed to discriminate between treatment failure and reinfection.Considering the high level of Pf(HRP-2/pLDH) RDT has been shown to have high sensitivity, specificity, positive and negative predictive values among pediatric patients with symptomatic malaria in Kinshasa, DRC. The RDT's relatively good attributes make it suitable for use as a guide in the monitoring of the efficacy of ACT treatment among pediatric patients in resource-poor areas like Kinshasa, DRC.SD Bioline malaria Ag"}
+{"text": "Studying the parasite reservoir is a tool for monitoring the effectiveness of control strategies. The gametocyte carriage is more common in patients with asexual forms, and the occurrence of recrudescence or reinfection in patients could also be favored and contribute to the maintenance of a large reservoir of parasites. The aim of this study was to determine the prevalence of submicroscopic gametocytaemia in patients treated for uncomplicated malaria.Gametocytes carriage and density were estimated by Pfs25mRNA amplification using QT-NASBA in samples obtained at enrolment and during the follow-up (day 21 to day 42 post-treatment) of children treated with either artesunate-amodiaquine (ASAQ) or artemether-lumefantrine (AL). Data were analyzed according to the study visit, the presence of asexual parasites, the type of treatment and the treatment response.P = 0.6). During the post-treatment visits, it was 58% and 44% in the ASAQ and in the AL groups respectively (P = 0.4). When paired samples of 23 children were analyzed, the gametocytaemia was positively correlated with the asexual form density the day of treatment failure (rho = 0.4 in the ASAQ group and rho = 0.5 in the AL). Logistic regression analysis showed that recrudescent infection (aOR: 12.9 [1.1-14.9]) were independent risk factors for SMG carriage whereas no association was found with the type of treatment, age and number of episodes.Samples from 48 children were analyzed; 23 were treated with ASAQ and 25 with AL. They had 147 visits, all corresponding to treatment failure with either ASAQ or AL. None of the patients had a microscopic gametocytaemia. Overall, the frequency of SMG carriage was 51%, comparable at day 0 between the ASAQ (53%) and the AL (56%) patients (The frequency of SMG carriage is high after ACT treatment whatever the combination used. A strong association between the presence of gametocytes and a recurrent infection is also observed."}
+{"text": "Artesunate\u00a0+\u00a0sulfadoxine-pyrimethamine (AS\u00a0+\u00a0SP) has been the first-line treatment and artemether-lumefantrine (AL) the second-linetreatment for uncomplicated falciparum malaria in Yemen since 2005. This paper reports the results of studies conducted to monitor therapeutic efficacy of these two drugs in sentinel sites in Yemen.Eight therapeutic efficacy studies were conducted in six sentinel sites during the period 2009\u20132013 in Yemen. Five studies were for the evaluation of AS\u00a0+\u00a0SP  and three studies  for the evaluation of AL. The studies were done according to standard WHO protocol 2009 with 28-day follow-up.In the evaluation of AS\u00a0+\u00a0SP, the PCR-corrected cure rate was 98\u00a0% (95\u00a0% CI 92.2\u201399.5\u00a0%) in one site and 100\u00a0% in all of the other four sites. In the sites where AL was evaluated, the PCR-corrected cure rate was 100\u00a0% in all the sites. All patients were negative for asexual parasitaemia on day 3 in both the AS\u00a0+\u00a0SP and the AL groups. There was a higher rate of clearance of gametocytaemia in the AL-treated group when compared with the AS\u00a0+\u00a0SP groups from day 7 onwards.AS\u00a0+\u00a0SP remains the effective drug for uncomplicated falciparum malaria in Yemen. AL is also highly effective and can be an appropriate alternative to AS\u00a0+\u00a0SP for the treatment of falciparum malaria. AL demonstrated a higher efficacy in clearing microscopic gametocytaemia than AS\u00a0+\u00a0SP.Trial registration: Trial registration number ACTRN12610000696099 Plasmodium falciparum and only 1\u00a0% due to Plasmodium vivax. No indigenous malaria has been detected in Socotra since 2006 [There are four major epidemiological strata of malaria in the Republic of Yemen. These are coastal plain, foothills, mountains, and the island of Socotra. Malaria-free zones include the mountain plateau and arid slopes from the highlands to the desert. In 2013, 25\u00a0%  of a total population of 24,400,000 lived in areas of high transmission (>1 case/1000), 53\u00a0%  in areas of low transmission (0\u20131 case/1000) and 22\u00a0%  in malaria-free areas [P. falciparum based on the earlier versions of standard WHO protocol [Plasmodium falciparum to anti-malarial drugs, which cited nine therapeutic efficacy studies in Yemen with median treatment failure rate of 42.4\u00a0% (range 9\u201357\u00a0%) for CQ and one trial for SP with no treatment failure [Most of the early studies on anti-malarial drug efficacy that were carried out in Yemen in the 1980s and the 1990s were done in the southern parts of the country. These were mainly in vivo studies based on the standard WHO seven-day test to assess response of falciparum malaria to chloroquine (CQ). The studies, which were conducted by WHO consultants for malaria control, reported no significant levels of CQ resistance at the time . In 2002protocol . Since tprotocol , 7 basedprotocol . By 2004The emergence and spread of drug-resistant malaria has been a major factor in the global resurgence of falciparum malaria in the 20th Century. Drug resistance has been implicated in the spread of malaria to new areas and re-emergence of malaria in areas where the disease had been eliminated. Drug resistance has also played a significant role in the occurrence and severity of epidemics in some parts of the world . In YemeIn 2005, Yemen switched to artemisinin-based combination therapy (ACT) for the treatment of uncomplicated falciparum malaria as per the recommendation of WHO . The selAS\u00a0+\u00a0SP has been evaluated extensively in adults and children with uncomplicated malaria in other parts of the world and was found to be sufficiently efficacious in areas where 28-day cure rates with SP alone exceeded 80\u00a0% , 14. AL P. falciparum with parasite density of 500\u2013200,000 asexual parasites/\u03bcl. A patient was excluded if she/he had a febrile illness other than malaria or severe and complicated falciparum malaria.The studies were conducted in six sentinel sites representing the different malaria-endemic provinces of the country see Fig.\u00a0. Each stPregnant women and females of reproductive age who could not be tested for pregnancy were\u00a0excluded. Pregnancy testing of unmarried women and female minors aged 12\u201317\u00a0years in Yemen is not acceptable according to the local customs and culture.The methods of treatment, follow-up and analysis of outcomes was based on WHO guidelines . After o\u00ae, Novartis Pharmaceutical Corporation) according to body weight bands. Patients weighing 5\u201314\u00a0kg received one tablet (20\u00a0mg artemether plus 120\u00a0mg lumefantrine) per dose, those weighing 15\u201324\u00a0kg received two, those weighing 25\u201334\u00a0kg three tablets, and those weighing\u00a0\u226535\u00a0kg received four tablets. In total, six doses were administered at hours 0, 8, 24, 36, 48, and 60. Study medications were provided by WHO. The intake of all doses of treatment was directly observed. All drugs were within their expiry period, and batch number and expiry date were recorded in each case record form (CRF). If a patient vomited within 30\u00a0min of treatment, a full dose was re-administered. All patients were followed up on days 1, 2, 3, 7, 14, 21, and 28. On each of these follow-up visits clinical and parasitological assessments were repeated. Patients were also asked to report to the clinic at any time if new or recurrent symptoms occurred.All patients received a standard regimen of the drug being tested . In the All patients who failed to be cured with AS\u00a0+\u00a0SP were treated with AL. Those who failed treatment with AL were treated with quinine orally at 10\u00a0mg salt/kg three times daily for 7\u00a0days.Thick and thin blood films for parasite counts were obtained from each patient and examined at screening and on days 2, 3, 7, 14, 21, 28 or on any other day if the patient returned spontaneously and parasitological reassessment was required. Blood smears were stained with 2.5\u00a0% Giemsa for 45\u00a0min and examined at a magnification of 1000\u00d7\u00a0to identify the parasite species and to determine the parasite density.The thick blood smear was used to calculate the parasite density, by counting the number of asexual parasites against 200 white blood cells (WBC) with a hand tally counter. The count was terminated when either 500 parasites or 200 WBC were reached, whichever came first. A blood smear was declared negative after examination of 1000 WBC revealed no asexual parasites. Parasite density, expressed as the number of asexual parasites per \u00b5l of blood, was calculated by dividing the number of asexual parasites by the number of WBC counted and then multiplying by an assumed WBC density of 8000 per \u00b5l.For quality assurance, all blood smears were re-read by a second microscopist. Blood smears with non-concordant results (differences in species or differences in parasite density of\u00a0>50\u00a0%) were re-read by a third microscopist and the average parasite density of the two most concordant counts was used.msp1, msp2 (merozoite surface proteins) and glurp (glutamate-rich protein). Subsequently, gel analysis was performed using Bionumerics V.5.10 in order to determine the size of the amplified gene targets. Identification of recrudescence and new infections was performed according to WHO guidelines [Two to three drops of blood were collected through finger pricks on filter paper (Whatmann No 3) during enrolment and each time blood smears were required, according to the protocol, on and after day 7. Specimens were labelled anonymously , kept in individual plastic bags with desiccant pouches and protected from light, humidity and extreme temperature until analysed. The specimens were genotyped to distinguish between recrudescence and new infections, according to methods recommended by WHO . Blood sidelines .Treatment outcomes were classified on the basis of an assessment of the parasitological and clinical outcome of anti-malarial treatment according to WHO guidelines . A patiePatients were assessed clinically for drug tolerability. Both adverse events and serious adverse events were monitored at enrolment and on each of the follow-up visit. An adverse event was defined as any untoward medical occurrence irrespective of its suspected relationship to the study medications. Serious adverse event included untoward medical occurrence requiring hospitalization or resulting in death.The treatment failure rate of the two drugs in the study areas was estimated to be 5\u00a0%. At a confidence level of 95\u00a0% and a precision estimate of 5\u00a0%, a minimum of 73 patients had been planned to be enrolled in each site. With a 20\u00a0% increase to allow loss to follow-up and withdrawals during the 28-day follow-up period, 87 patients were targeted to be included in the study per site per drug. Data were double entered and validated using a programme developed by WHO . Per pro\u00ae to generate descriptive statistics and analyse data . A 2\u00a0\u00d7\u00a02 Chi square table was used to analyse associations between proportions using Epi-Info 7 .Statistical analysis was performed using SPSS 22Permission to conduct the studies was obtained from the General Doctorate for Research and Studies, Ministry of Health, Yemen, which is the national body with oversight to ethically review research proposals involving human subjects, and WHO Ethical Research Committee. Local authorities (community leaders) of the study areas were informed of the study objectives, procedures and duration and their permission was sought. Individual informed consent was obtained from adults and parents/guardians of children. Those who were illiterate selected a witness not related to the research team to sign on their behalf.Eight therapeutic efficacy studies were conducted in six sentinel sites in Yemen to assess the therapeutic efficacy of AS\u00a0+\u00a0SP (five studies) and AL (three studies) during the period November 2009 to March 2013. Table\u00a0Table\u00a0P. falciparum asexual stages/\u00b5l. The second case (LCF) was a 5\u00a0year old female who presented on day 28 with a recent history of fever and her blood film was positive with 45,454 P. falciparum asexual stages/\u00b5l. Both cases had treatment doses within the therapeutic range.In the individual studies and after PCR-correction Kaplan\u2013Meier analysis, the cure rate with AS\u00a0+\u00a0SP ranged between 98 and 100\u00a0% in the different sites. The two cases of treatment failure were in Bajil 2013. The first case (LCF) was a 2\u00a0year old male who presented on day 28 with a history of fever, his blood film showed 880 In the AL studies, a total of 268 patients with falciparum malaria satisfied the inclusion criteria Table\u00a0. The stuFigure\u00a0The 28-day parasitological cure observed after first-line AS\u00a0+\u00a0SP therapy in the present study exceeded 95\u00a0% in all the sites after PCR-correction and also in the uncorrected analyses, meeting the WHO recommendation that cure rates for falciparum malaria should be at least 90\u00a0% and preferably\u00a0>95\u00a0%. Detection of asexual parasites on day 3 is an early warning sign of slow clearance of parasites by artemisinin , 19. In P. falciparum isolates from 100 malaria patients in Tihamah, Yemen [P. falciparum in the study area.The findings of high efficacy of AS\u00a0+\u00a0SP against falciparum malaria are supported by the fact that at present there is no evidence that SP has lost its efficacy against falciparum malaria in Yemen. In 2005, in vivo and in vitro tests were conducted by Al-Kabsi et al. to determine the SP efficacy against h, Yemen . In the dhfr Arg-59 mutation in 99 amplified samples, while the dhps Glu-540 was not detected in 119 amplified samples [dhps [pfcrt, pfmdr1, dhfr, and dhps) were genotyped in 108 P. falciparum isolates collected in three sites in Yemen: Dhamar, Hodeidah and Taiz. The investigators concluded that the absence of the triple mutant dhfr genotype (IRN) and dhps mutations supports the use of AS\u00a0+\u00a0SP as first-line therapy. They suggested that the previous report on the presence of C59 mutation could possibly be explained by inclusion of expatriates in the sample of patients studied. In another study, isolates from 90 patients with microscopically confirmed P. falciparum infection from Al-Hodaida were analysed for the molecular dhfr 108\u00a0N by Abdul-Ghani et al. [P. falciparum isolates in its pure and mixed-type forms. They suggested that the high frequency of dhfr 108\u00a0N among parasite isolates should be cause for concern about the efficacy of SP as partner with AS. However, dhfr 108 mutation is a first step and is followed by other mutations before significant resistance occurs [Published studies with SP resistance-associated molecular markers in Yemen are few and have not provided consistent results about the prevalence of these markers. A molecular marker study in Meseimeer found a 5\u00a0% prevalence of  samples , 7. Thises [dhps . In a moes [dhps , four dri et al. . The mute occurs . The hige occurs  and othee occurs  and Somae occurs , calls fIn the present study, the 28-day, PCR-uncorrected cure rate after treatment with AL is 100\u00a0% in Bajil (95\u00a0% CI 95.1\u2013100\u00a0%), in Jabal-Al-Sharq it is 97.8\u00a0% (95\u00a0% CI 92.2\u201399.7\u00a0%) and in Tur Bani Qa\u2019is it is 94.3\u00a0% (95\u00a0% CI 87.2\u201398.1\u00a0%). The PCR-corrected, 28-day cure rate after AL in each of the three sites is 100\u00a0%. This is higher than the findings of a recently pooled analysis of a 28-day, PCR-corrected parasitological cure rate of 97.1\u00a0% in adults and 97.3\u00a0% in children . The twoThe effect of AL in the present study on microscopic gametocytaemia is consistent with findings from other endemic areas, which show a significant impact of AL on gametocytaemia . The datAS\u00a0+\u00a0SP remains a safe and effective first-line drug for the treatment of uncomplicated falciparum malaria in Yemen. However, monitoring the efficacy of this ACT should be continued since there is a high risk of failure of the SP component due to sub-therapeutic levels resulting from inadequate use of anti-malarial drugs in the country. Surveillance for SP resistance-associated molecular markers should also be monitored as a supportive tool to in vivo efficacy. AL is highly efficacious in Yemen and remains the appropriate option as a second-line treatment for uncomplicated falciparum malaria. AL shows higher efficacy than AS\u00a0+\u00a0SP in the clearance of microscopic gametocytaemia."}
+{"text": "This study aimed to synthesize the existing evidence on the efficacy and safety of a single dose artemisinin\u2013naphthoquine (ASNQ) for treatment of uncomplicated malaria in endemic countries.A meta-analysis of randomized, controlled trials (RCT), assessing efficacy and safety of single dose ASNQ was carried out. Comparator drugs included artemether\u2013lumefentrine (AL), chloroquine plus sulfadoxine-pyrimethamine (CQSP) and dihydroartemisinin\u2013piperaquine (DHP). The efficacy and safety profile of non-comparator, single-arm studies on the single dose ASNQ was also assessed. The primary endpoint was efficacy defined as an absence of PCR-confirmed parasitaemia. The methodological quality of the included studies was assessed using the six domains for the risk of bias.Five RCTs and three single-arm studies were included in this review. As RCT studies did not compare the same anti-malarial drugs, it was difficult to do a pooled analysis. At day 28, a pooled analysis of two RCTs (n\u00a0=\u00a0271) showed a comparable efficacy on PCR-confirmed parasitaemia between ASNQ and AL. Another RCT, which compared ASNQ and CQSP or ASNQ and DHP, also showed comparable efficacy. At day 42, one RCT comparing ASNQ and DHP and another RCT comparing ASNQ and AL reported comparable levels of efficacy. The proportion of parasite clearance was faster in the ASNQ groups than the comparators at day 1, and almost all parasites were cleared by day 3 in the ASNQ groups.The present review provides some evidence to support that there is similar efficacy and safety of the single dose ASNQ compared to other anti-malarial drugs in treating uncomplicated malaria. Larger, adequately powered, well-designed studies are recommended to substantiate the efficacy and safety in different populations and in different epidemiological settings. As the potential evolution of drug resistance is a great concern and this cannot be addressed in a short-term study, the use of single dose ASNQ needs further evaluation.The online version of this article (doi:10.1186/s12936-015-0919-5) contains supplementary material, which is available to authorized users. The target set under Millennium Development Goal 6 will be reached by 55 countries that are on track to reduce their malaria burden by 75\u00a0% . DespitePlasmodium falciparum malaria to ensure efficacy and reduce the emergence of drug-resistant parasites [The development of resistance to conventionally used anti-malarial drugs, such as chloroquine (CQ) and sulfadoxine-pyrimethamine (SP) has been documented . WHO recarasites , 4, 5. Sarasites . The conarasites , 7 to raarasites .P. falciparum resistance to artemisinin  in five countries of the Greater Mekong Sub-region, such as Cambodia, Laos, Myanmar, Thailand, and Vietnam [Resistance of malaria parasites to currently used ACT has emerged and is following a similar pattern of resistance previously observed with other anti-malarial drugs. Thus far, studies have documented evidence of  Vietnam  and this Vietnam . Thus, iA recent development of an oral single dose ACT therapy is a coformulated combination of artemisinin and naphthoquine phosphate (NQ) , 7, 9. NIndividual studies assessing efficacy and safety of the single dose ASNQ are available. As transmission of malaria varies even over small distances , informaThe present study adhered to the preferred reporting items for systematic reviews and meta-analyses (PRISMA) . The staPlasmodium OR falciparum OR vivax).Studies on the assessment of efficacy of single dose ASNQ in treating uncomplicated malaria were searched in electronic databases such as MEDLINE, EMBASE, CINHAL and the Cochrane Library. For ongoing and unpublished trials, the websites of WHO, the clinical trials database, and the drug manufacturer were checked. The reference sections of the selected studies and relevant reviews were also checked for the possibility of any additional papers. The search was limited to human studies, published in English, French and Chinese languages until May 2015. Medical subject headings (MeSH) terms and text words were (artemisinin\u2013naphthoquine OR ARCO) AND (treatment success OR treatment failure OR efficacy OR tolerability OR safety) AND  Participants residing in endemic countries and having uncomplicated malaria, regardless of age, gender, pregnancy status, and species of malaria parasite were considered. Diagnosis of malaria was based on microscopy of Giemsa-stained peripheral blood films or a rapid-onsite diagnostic test. Subsequent PCR-based analysis for species confirmation was an additional merit.Intervention (I) Studies using fixed combination single dose ASNQ by participants in one arm, regardless of route of administration and brand name were considered.Comparison (C) Studies which compared the efficacy of single dose ASNQ to alternative anti-malarial drug(s) or placebo were included.Outcomes (O) For simplicity sake, the primary and secondary outcomes were efficacy and safety, respectively. Efficacy was defined as the proportion of absence of (1) PCR-confirmed parasitaemia in patients at day 28 and at day 42, (2) PCR-unconfirmed parasitaemia in patients at day 28 and at day 42, or, (3) parasitological and fever clearance time. The safety outcomes were incidence of adverse events (AEs) and serious adverse events (SAEs). Studies were included if the effect estimates of each study such as relative risk (RR), odds ratio (OR) or hazards ratio (HR) and its 95\u00a0% confidence interval (CI) were provided or made available for computation.Study design (S) Randomized controlled trials (RCTs) which assessed efficacy of fixed-combination single dose ASNQ in treating uncomplicated malaria were included. Information from single-arm trials were also considered separately for the therapeutic efficacy of ASNQ, although they were not included in the main meta-analysis. Abstracts and conference reports were included, if they provided adequate data on the comparable efficacy between single dose ASNQ and other anti-malarial drug(s).Two authors independently screened the title and abstract yielded from the electronic search. Any discrepancies between the two authors were resolved by consensus. The two authors individually collected information from each included study using the piloted data extraction form. Information collected included characteristics of participants, study design, characteristics of the experimental drug , confirmation of malaria infection, specific speciation, duration of follow-up and the reported clinical outcomes. For studies with overlapping of study population, the one that provided the most comprehensive data was used.The methodological quality of the included studies was assessed using the risk of bias tool applied for the Cochrane systematic reviews . The sixP. falciparum, Plasmodium vivax, mixed malaria infection), transmission intensity, brand and dosing of ASNQ. Due to an insufficient number of studies and paucity of data, stratification was not possible.RR and its 95\u00a0% CI for dichotomous data and/or mean difference (MD) and standard deviation (SD) for continuous data from each study were recorded. Meta-analysis was performed, if two or more studies with direct head-to-head comparisons were included. This was only possible for the comparison between ASNQ and artemether\u2013lumefantrine (AL) due to limited number of studies in the similar manner. Initially, it was planned to stratify analyses by the targeted malaria parasite  overlapped with a study included in this review , (2) ass overlappOnly five RCTs and three single-arm studies were identified for the current review. The characteristics of the included RCTs are provided in Additional file Overall, most of the RCTs  included in the current review had \u2018low risk of bias\u2019 as they met five of the six domains assessed , 21\u201323. I-square value: 0\u00a0%) [As the RCTs included in the present review did not compare ASNQ with the same anti-malarial drugs, it was difficult to do a pooled analysis. At day 28, a pooled analysis of two RCTs (n\u00a0=\u00a0271) showed a similar efficacy between ASNQ single dose  and AL   , 23. Of ue: 0\u00a0%)  on PCR-cue: 0\u00a0%)  P. falciparum malaria.At day 42, there was a similar efficacy between ASNQ  and DHP  on PCR-confirmed parasitaemia   and betwThe mean parasite clearance time (PCT) was comparable between ASNQ (28\u00a0\u00b1\u00a011.7) and DHP (25.5\u00a0\u00b1\u00a012.2)  . The proFever clearance time (FCT) was reported in two RCTs; a comparable FCT was shown between ASNQ and DHP   or ASNQ p\u00a0>\u00a00.05) [Only one RCT reported this data. The gametocytaemia by day 3 was reduced from 67.3 to 18.6\u00a0% in ASNQ and from 70.3 to 17\u00a0% in DHP (\u00a0>\u00a00.05) . A singl\u00a0>\u00a00.05) .vs comparator group). Table\u00a0In all five RCTs, AEs were recorded and compared between treatment groups  [p: 0.56) on PCR-confirmed parasitaemia [Per protocol analysis, all patients who were not available  were removed from the denominator, and this was done in two RCTs , 23. At sitaemia . Due to sitaemia .In order to facilitate the development of treatment policies for the deployment of effective anti-malarial drugs, a systematic monitoring of anti-malarial drug efficacy and drug resistance is needed , 34. TheOverall, the findings could provide some evidence that there was a comparable efficacy between ASNQ single dose and comparators such as CQSP, DHP and AL. Moreover, the efficacy estimates of ASNQ were consistent at day 28 and at day 42. Using both day 28 and day 42 in this analysis was relevant because a 28-day follow-up captured the majority of treatment failures with drugs inclusive of artemisinin derivatives, while the longer follow-up for 42\u00a0days was optimal for these drugs . RCTs, aP. falciparum clearance continued to be achieved in sub-Saharan African patients treated with ACT [A pooled analysis of studies assessing artesunate\u2013amodiaquine including a single dose regimen in treating uncomplicated malaria documented that rapid with ACT .The current analysis showed that ASNQ single dose had a property of rapid reduction in parasite biomass within the initial 24\u00a0h. In fact, the rate at which treatment clears parasites within the first few days is the most useful practical test for ACT. This is because early response to treatment relies predominantly on the parasite response to artemisinin, independent of whether parasites are later cleared permanently through the combination of longer-lived companion drug and the host\u2019s immune response .Although ACT reduced the density of gametocytaemia and the proportion of infected mosquitoes, sub-microscopic levels of gametocytes (which were present in a significant number of patients after treatment) appeared to be sufficient to drive post-treatment transmission . The detOne of the key elements in any drug development and evaluation is the issue of safety for the population for whom the drug is intended . The incPlasmodium malariae. In each cycle, artemisinin and its derivatives would reduce parasite biomass by a factor of ~10,000 [A study assessing a dose comparison of ASNQ in children with malaria showed similar efficacy and tolerability between single dose with water/with milk and twice daily dose regimens, indicating single dosing can be expected to improve better compliance . A short ~10,000 , 40.P. vivax infections [A dose range study in PNG showed that a lower single ASNQ dose was associated with relatively frequent recurrence of fections . The recfections , 33.Any anti-malarial regimen must have robust evidence of both optimal efficacy for patient survival and an ability to reduce the potential for drug resistance . The finThere are limitations to the present study. A small number of studies with small samples on the assessment of the ASNQ single dose were identified for the present review. In order to demonstrate the non-inferiority (by a margin of 5\u00a0%) of an alternative treatment (ASNQ in this case) to a current treatment known to be 95\u00a0% effective, at least 299 patients would be necessary in each study arm, with a one-sided test that has a statistical power of 80\u00a0% and a significance level of 2.5\u00a0% . Using tP. falciparum and P. vivax co-exist. For instance, the efficacy of ASNQ and CQSP were comparable in the PNG study where P. falciparum and P. vivax are equally important [P. vivax was likely to be missed in PCR-uncorrected parasitaemia. This could be more pronounced in the CQSP group as parasite resistance to this combination drug is already confirmed in PNG [The PCR-confirmation of parasites was done in only two RCTs in the current review , 22, henmportant . Thus, Pd in PNG . The effThree RCTs in this review were open-label trials , 22, 23.Plasmodium species. All single-arm studies were conducted on patients having P. falciparum [P. falciparum.Due to limited data, the current review is unable to provide evidence for different age groups or different lciparum , 10, 11.lciparum  needs caBased on available data, the current analysis shows comparable efficacy and safety profiles of the single dose ASNQ. Single dose ASNQ can improve treatment compliance and is simple and practicable. This is because this single dose can be delivered as directly observed treatment, immediately after confirmation of malaria . Should P. falciparum and P. vivax for a longer period than piperaquine and, in particular lumefantrine. This is because the peak plasma concentration of benflumetol  is attained slowly (8\u00a0h plus a 2-h absorption lag time), and the elimination half-life is estimated to be 4.5\u00a0days [The relatively long terminal half-life and wide therapeutic index of NQ  could co4.5\u00a0days . Althoug4.5\u00a0days . There iThe present review provide some evidence to support the comparable efficacy and safety of the single dose ASNQ compared to other comparator anti-malarial drugs in treating uncomplicated malaria. Larger, adequately powered, well-designed studies are recommended to substantiate the efficacy and safety in different populations and in different epidemiological settings. As the potential evolution of drug resistance is a great concern and this cannot be addressed in a short-term study, the use of single dose ASNQ needs further evaluation."}
+{"text": "Parenteral artesunate is recommended as first-line therapy for severe and complicated malaria. Although its efficacy has been proven, long-term safety profile is still under evaluation. Several cases of delayed haemolytic anaemia occurred after initial clinical improvement and resolution of parasitaemia in non-immune travellers and children living in endemic areas. Reports have generated concern that this phenomenon might be related to the treatment itself, either by direct toxicity or immune-related mechanism. This is a report of the first case of autoimmune haemolytic anaemia following treatment of severe malaria initially managed with parenteral artesunate with strong indication for drug-immune related mechanism.Plasmodium falciparum infection with a parasitaemia of 0.8%. Severe malaria was diagnosed according to the WHO criteria. The patient was initially managed with artemether/lumefantrine combination and then parenteral artesunate for 48\u00a0hours. Empiric antibiotic course was also initiated with ceftriaxone, metronidazole, gentamycin, and then piperacillin and ciprofloxacin. At day 14, haemoglobin dropped to 4.6\u00a0g/dL with biologic features indicative of haemolysis . At that time, parasitaemia was negative and other infections or hereditary disorders were excluded, while Coombs\u2019 direct antiglobulin test was positive for IgG and C3d. Antinuclear antibodies were absent. Further investigations evidenced drug-induced antibodies related to artesunate. It was concluded a drug-mediated autoimmune haemolytic anaemia. A corticosteroids regimen was initiated at 1\u00a0mg/kg/day. Outcome was favourable and corticosteroids were progressively tapered during two months. At present the patient\u2019s condition remains stable without recurrence of haemolytic anaemia.A 17-year old Ivoirian female travelling in France presented with fever, headache and abdominal pain seven days after her arrival. Physical examination was indicative of septic shock while blood analysis showed normal haemoglobin level, but profound thrombocytopaenia and hyperlactataemia. Blood smear analysis showed This is the first case of delayed haemolytic anaemia related to artesunate with a strong indication for drug-immune related mechanism. Further research is warranted to better characterize this plausible cause of post-treatment haemolysis following parenteral artesunate administration in severe malaria patients.The online version of this article (doi:10.1186/1475-2875-13-398) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria remains a major risk for northern countries\u2019 travellers returning from malaria-endemic areas. According to WHO guidelines and recommendations of the European Society for Clinical Microbiology and Infectious Diseases, intravenous (iv) artesunate should be considered as first-line treatment for severe malaria, instead of quinine [Infection with  quinine . While s quinine  and Afri quinine , little  quinine \u20138. Most 3 , a haemoglobin level of 12.6\u00a0g/dL (12\u201316) with abnormalities indicative of haemolysis: rise in lactate dehydrogenase (LDH) at 500 U/L (5\u2013248) and total bilirubin at 105\u00a0\u03bcmol/L (3\u201318) with a low haptoglobin of 0.15\u00a0g/L (0.3-2). She was diagnosed with uncomplicated malaria as peripheral thin blood film showed P. falciparum trophozoites (0.8% of parasitized erythrocytes). Abdominal ultrasonography ruled out biliary tract or gall bladder infection. A treatment with oral artemether/lumefantrine combination (Riamet\u00a9) was initiated with respectively, 80 and 480\u00a0mg trice within the first 24\u00a0hours of hospitalization . On day 2, her clinical condition deteriorated, her blood pressure dropped to 80/40\u00a0mmHg together with a pulse rate of 130\u00a0bpm. Laboratory tests indicated a fall of thrombocyte count at 6,000/mm3 and elevation of blood lactate up to 7.4\u00a0mmol/L (N\u2009<\u20092). The patient was now classified as complicated malaria and admitted to intensive care unit. Treatment was switched to intravenous artesunate , started at three doses of 120\u00a0mg (2.4\u00a0mg/kg body weight) with 12-hour interval. Concurrently she was managed with supportive care, that was administration of 2\u00a0l normal saline solution and two units of packed thrombocytes. The use of norepinephrine up to 0.5\u00a0\u03bcg/kg/min was required during 24\u00a0hours to restore a normal blood pressure. The septic shock condition was managed by a course of empiric antibiotic treatment, i.e., ceftriaxone 2\u00a0g/day, gentamycin 3\u00a0mg/kg/day and metronidazole 1.5\u00a0g/day. Evolution was marked by rapid improvement as blood pressure maintained at normal values, and fever disappeared. Lactate level returned within normal range and parasitaemia declined under 0.1% of red blood cells (RBCs). Twelve hours after the last dose of artesunate , the patient was transferred to a regular ward of tropical medicine (day 4). Anti-malarial treatment was continued orally with two daily doses of 80\u00a0mg artemether and 480\u00a0mg lumefantrine until day 6 . Antibiotics were changed to piperacillin/tazobactam (12\u00a0g daily) and ciprofloxacin 500 mgx2/day. Despite initial favourable evolution and total clearance of parasites, fever reappeared on day 8. Of note, a drop in haemoglobin up to 6.3\u00a0g/dL was evidenced while thrombocytosis was associated: 489\u00a0G/L. The Coombs\u2019 test performed at that time was negative. The patient left hospital against medical advice but was re-admitted on day 14 because of persistent fever and marked asthaenia.A 17-year old Ivorian female without remarkable medical history was admitted for fever, chills, headache, and abdominal pain in a French University Hospital Centre (day 1). She had left Ivory Coast seven days earlier to live in France for studying purpose and symptoms began two days before her admission. Initial physical examination showed a temperature of 39\u00b0C and pain when palpating right hypocondrium. Blood tests demonstrated a normal leucocyte count, a thrombocyte count of 11,000/mm3, LDH flare up to 658 U/L and undetectable haptoglobin level. Repeated blood films for malaria as well as for schistocytes  were negative. Glucose-6-phosphate dehydrogenase (G6PD) deficiency was promptly ruled out and haemoglobin electrophoresis did not reveal any abnormality. On the other hand, direct Coombs\u2019 test revealed positivity for both IgG and complement factor C3d while this test was negative five days earlier. The irregular antibody testing was negative. It was concluded AIHA and complementary aetiological inquiry was performed. Antinuclear antibodies were absent and the investigation was negative for numerous pathogens such as bacteria , viruses  or parasites .In the context of unusual immunohaematological haemolysis, no transfusion was attempted and corticosteroids were introduced. Methylprednisolone pulses of 60\u00a0mg per day during seven days were performed. The haemoglobin level gradually improved . This testing was conducted according to a home-made method developed by the French Blood Institute according to standardized reports [Concurrently, she was additionally tested for drug-dependent antibodies by  reports \u201311 and dAmong all tested drugs, the test was positive only with artesunate in the papain-pretreatment condition Table\u00a0. Thus, nThe patient\u2019s samples (15\u00a0mL of ethylene diamine tetra-acetate blood and 15 mL of serum) were referred to the French Blood Institute of Bordeaux. Sera of healthy individuals of AB blood group that willingly give their blood to the French Blood Institute served as negative controls and were referred as healthy donors. The initial antibody identification was performed using an IAT (anti-IgG and anti-C3d) tube method, with native and papain-treated RBCs . Papain is an enzyme that potentiates the agglutination reaction by reducing the negative charges on the surface of RBCs, thus allowing greater accessibility of some epitopes. Papain treatment of the tested RBCs was performed according to the manufacturer\u2019s recommendations : one volume of papain solution was added to one volume of washed RBCs. After an incubation of 15 min at 37\u00b0C, RBCs were washed three times. Serum samples of the patient were incubated during one hour at 37\u00b0C with a panel of erythrocyte and various concentrations of the drugs mentioned above: pure, diluted to 1/10, and estimated therapeutic concentration. Then, agglutination of RBCs was tested with indirect antiglobulin test using anti-IgG or anti-C3d as well as papain pretreated erythrocytes. As negative controls, saline solution and the complement source  were both tested with and without the drug solution added to reagent RBCs. The positive control referred to erythrocytes incubated with a serum-containing antibody directed against a cell surface protein of RBCs, namely Fya. The result of the test was estimated by the degree of RBCs agglutination with a macroscopic evaluation. If RBCs were not sensitized by antibodies, they were not stuck together and were found at the bottom of the tube (reaction scored \u20180\u2019). If RBCs were sensitized by antibodies, they would remain clustered at the surface column and this reaction was graded from 1 to 4+, 4+ being the strongest positive reaction.To date, up to 19 cases of putative artesunate-related, late-onset haemolysis have been described in the literature among patients with severe malaria who returned from malaria-endemic regions \u20138. All pThe fact that mainly hyperparasitaemic patients developed this condition has been related by some to a mechanism called \u2018pitting\u2019. After extraction of blood stage parasites during splenic passage, these once-infected erythrocytes have a reduced lifespan compared to na\u00efve erythrocytes with a mean lifespan of around 180 hours and with a total removal of pitted erythrocytes after 28 days. A hypothesis is that haemolytic activity may increase two weeks after acute malaria due to synchronized destruction of pitted erythrocytes , 14. Of Mycoplasma pneumoniae or Epstein Barr Virus-related-mononucleosis. It seems that malaria itself has never been described as a potential cause of AIHA. Although most cases of AIHA are idiopathic, the context of multiple drugs intake and recovery within a few weeks were in favour of a drug-induced immune haemolytic anaemia (DIIHA). Indeed, when a drug responsible for DIIHA is discontinued, the haemolytic anaemia resolves soon afterwards, while during idiopathic AIHA evolution is often chronic or recurrent. In the present case, all suspected treatments were stopped approximately at the same time, what does not permit to charge one in particular. Of note, the patient did not receive artemisin-derivative for previous malaria fever.Haemolytic anaemia is a challenging situation for physicians as multiple causes may be involved , 16. TheImmune-mediated haemolysis may be another mechanism responsible of artesunate-related delayed haemolytic anaemia. In the cases of DIIHA, the direct Coombs\u2019 test (DAT) is usually positive, thus being a prerequisite for hypothesize DIIHA . Thus, aDIIHA is a rare condition, as the estimated incidence is of one per million population per year. Three mechanisms are described: drug absorption (hapten-induced), immune complex formation with the drug on RBC surface or auto-antibody production resulting in IgG and/or IgM (C3d complement) positivity in direct antiglobulin test . Numerouin vitro induction of haemolysis with several of the employed drugs. Usually these tests can show immune-mediated haemolysis dependent of the presence of the medication, indicative of drug-dependent antibodies, but they are not performed routinely by most of the laboratories. The results of these tests were indicative of an artesunate-mediated auto-immune haemolysis. Of note, the positive result was evidenced using papain pretreated RBCs, which is considered to enhance the sensitivity for detection of antibody response challenges, as described elsewhere [Concerning lumefantrine, some cases of haemolytic anaemia are reported . Howeverlsewhere . Such reWhether this suspected immune-mediated haemolysis is a consequence of artesunate itself or of its excipients is questionable. Indeed, no case of artesunate-associated haemolysis were reported in USA, where the medication is produced by the Army Medical Material Development Activity, while in other countries the manufacturing process is different .Finally, outcome was slowly favourable while using glucocorticoids, although this may be coincidental. Management of DIIHA requires discontinuing the suspected medication: it is often the only treatment. Actually, efficacy of steroids is uncertain because data are limited to case reports and the discontinuation of the drug at the same time is a confounding factor.Delayed haemolysis is a frequent and relevant complication in patients treated with parenteral artesunate for severe malaria. The aetiology of this phenomenon is still unknown but the drug may act as substrate for autoimmune mechanism. Patients undergoing this drug should be closely monitored with prolonged follow-up including, in patients with delayed haemolysis, a specific immunological investigation.Written informed consent was obtained from the patient for publication of this Case Report and any accompanying images. A copy of the written consent is available for review by the Editor-in-Chief of this journal.Cases reported in the literature of delayed haemolytic anaemia with positive Coombs\u2019 test after intravenous artesunate therapy for severe malaria.(DOCX 14 KB)Additional file 1:"}
+{"text": "In resource-poor settings, treatment adherence is a major determinant of response to anti-malarial drugs as most are taken at home without medical supervision. Evidence on adherence to artemisinin-based combination therapy (ACT) is limited. The study aimed to measure adherence and identify reasons for non-adherence to a 3-day, fixed-dose combination (FDC) of artesunate\u2013amodiaquine (ASAQ), the first-line treatment for uncomplicated malaria in the M\u00e9decins Sans Fronti\u00e8res project in the Shabunda Health Zone, South Kivu, Democratic Republic of Congo, a highly malarious and conflict-affected area.The study took place in the health centres/outpatient departments of the Shabunda general hospital, the quarter Mbangayo, and participant households. Patients prescribed FDC ASAQ were visited at home on the day after their regimen finished and asked to complete an adherence questionnaire. Patients/caretakers were also interviewed when exiting the outpatient department to understand their attitude towards FDC ASAQ and assess the quality of the prescribing process.148 patients/caretakers completed the adherence questionnaire: 11.5\u00a0%  had \u22651 tablet left at the time of the home visit and were defined as certainly non-adherent; 13.5\u00a0%  were probably non-adherent; thus total non-adherence was 25.0\u00a0% . 75\u00a0%  were defined as probably adherent. In exit interviews, 87.5\u00a0% (105/120) knew they had malaria or could name the correct signs/symptoms. 89\u00a0% (107/120) could identify FDC ASAQ as anti-malarials among all tablets given and correctly repeat the intake instructions given at the outpatient department.This is the first study to assess adherence to an FDC of ACT under real treatment conditions in a context of high instability. High quality prescribing of anti-malarials at health centre level and patient adherence to the correct intake of ACT were possible in this setting. Adherence to treatment regimen requires careful and constant monitoring which might be better guaranteed at health centre rather than community level. It could, nevertheless, be a precondition to the successful introduction of home- or community based management of malaria. Plasmodium falciparum is endemic [Malaria, despite being treatable, remains a major killer in many low-income countries, with around half the world\u2019s population at risk from the disease. Approximately 627,000 deaths occur every year, 90\u00a0% in sub-Saharan Africa and 85\u00a0% in children under 5\u00a0years of age , 2. In a endemic . Incorre endemic . WorryinThe correct use of a treatment involves correct diagnosis, prescription, dosage, and adherence. Factors that influence patient adherence include frequency of dosing, number of pills, duration of treatment, side-effects, cost of treatment, household income, concomitant treatment intake, education level, sufficient explanation of the importance of full-course adherence by the treatment prescriber, attitudes towards the sharing and saving of medications, and caregivers\u2019 perception of the severity of the illness \u201314. OnlyAdherence to treatment is a major determinant of the response to anti-malarial drugs, especially for remote and unstable settings where home-based or community-based programmes for managing malaria might be one of the few strategies able to overcome barriers to accessing health care , 16. AdhFixed-dose combinations (FDC) might improve adherence compared with co-blistered drugs \u201321. With\u00ae) or microscopy at hospital level when indicated. Malaria is holoendemic  in Shabunda Health Zone, with the highest number of cases occurring between September and December. In January\u2013October 2013, malaria cases in MSF-supported health facilities in Shabunda Health Zone accounted for 31.4\u00a0% of consultations in the outpatient department (OPD) ; 48.4\u00a0%  were in children aged under 5\u00a0years. Uncomplicated malaria was the principal cause of morbidity and the main cause for admission to a reference hospital  .In accordance with the national protocol of the Programme Nationale de Lutte contre le Paludisme  of the Democratic Republic of the Congo (DRC), FDC ASAQ is the first-line malaria treatment used in the MSF project in the Shabunda Health Zone, South Kivu, DRC. All malaria cases are confirmed with a rapid diagnostic test  with a catchment area of 172,854 habitants, located in the South Kivu Province of DRC, bordering with Rwanda and Burundi to the east . The area has long been affected by conflict. MSF supports a general hospital run by the Ministry of Health in Shabunda town, a hospital centre in the village of Matili (around 30\u00a0km south of Shabunda) both providing secondary-level health care, and seven health centres (OPDs) providing primary-level health care to the whole catchment area.The study took place in 2013, in the OPD of the general hospital of Shabunda and the OPD Mbangayo  and in the households of the study population. Malaria activities are completely integrated in the hospitals and OPD activities. The study was conducted during November to coincide with the highest period of malaria incidence and therefore to reflect the level of adherence when the duration and quality of malaria care might vary because of the high caseload.\u00ae) of uncomplicated falciparum malaria, with or without additional diagnoses; receipt of a course of FDC ASAQ during normal consultation hours in the MSF-supported OPDs; no previous participation of a household member in the study; and signed informed consent (adherence study) or verbal consent (exit interview). Patients were excluded from the adherence study if they were resident \u226530\u00a0min from the OPDs by foot or lived in Kitete, a \u201cquartier\u201d (quarter) of Shabunda town considered inaccessible due to security constraints. Kitete borders the river Ulindi, and rebel forces have their base on the other bank. Moreover, the airstrip, which was under fire 3\u00a0days before the start of the study, must be crossed to reach Kitete.Patient inclusion criteria were: age \u22651\u00a0year; a confirmed diagnosis via RDT  or in French. The study was supervised by the principal investigator and a field study supervisor who was fluent in all spoken languages. Prior to initiating the study, 2-days of training were given to all interviewers and data collectors. Extra attention was paid by the supervisors to discuss and agree on the same wording for all interviewers and data collectors while carrying out the interviews in the different local languages. With the help of role plays, all interviewers and data collectors had sufficient time to familiarize themselves with the questionnaires.The usual procedures of consultation, treatment prescription and dispensing in the MSF-supported OPDs were followed: patients received a health card and a malaria RDT if they were suspected of having malaria, defined as fever or history of fever in the previous 48\u00a0h; were prescribed FDC ASAQ in the case of a positive RDT and further drugs if necessary; took the first dose of FDC ASAQ under supervision at the pharmacy; and were given the second and third daily doses to take at home with intake instructions.Before leaving the OPD, a short screening questionnaire was administered by three data collectors: a full description of all drugs prescribed was copied from each patient\u2019s health card and their address recorded to enable subsequent location of their household. All patients, irrespective of diagnosis, received the screening questionnaire to avoid raising suspicion among health staff and patients/caretakers of patients about the specific purpose of the study. Health staff were informed that a study was taking place, but the exact purpose of the study was not revealed to avoid influencing routine prescribing habits.Patients prescribed FDC ASAQ were visited at home on the day after the last dose should have been consumed. If the visit could not be completed, the interviewer tried again the following day. No interviews were carried out at a later date. Patients needed to be at home to answer the questionnaire themselves. For children or patients not able to take FDC ASAQ on their own, the caretakers who gave them their treatment had to be present to answer the adherence questionnaire. Patients or caretakers who were not able to be located were classified as lost to follow-up and excluded from the study.The semi-structured adherence questionnaire was developed in French. It included initial general questions about the patient and household: sex, age, education level of patient or caretaker, household composition, and occupation of head of household. A systematic account of how the tablets had been taken was recorded. Patients or caretakers were also asked to show the original co-formulated blister package of the FDC ASAQ; if it was still available any remaining tablets were counted. Finally, two questions on general malaria knowledge were asked. Five interviewers administered the adherence questionnaires at the home of the patients.After all the screening questionnaires had been completed, exit interviews were carried out at the OPDs to help understand patient/caretaker attitudes towards FDC ASAQ prescription and to assess prescription quality (the information and instructions provided to the patient/caretaker and supervision of the first dose). All patients who received FDC ASAQ during normal consultation hours were eligible. They or their respective caretakers were interviewed upon exiting the OPD consecutively until the required number was achieved. Patients/caretakers were asked to describe how they were planning to give/take the FDC ASAQ, followed by a series of systematic questions for each treatment dose. Three interviewers  conducted the exit interviews at each of the two OPDs.Since there is no standard definition for adherence we used definitions shown to be reliable in similar settings \u201329. PatiOn the basis of malaria adherence studies in similar areas, adherence was conservatively estimated at 50\u00a0% \u201328. Withp value and relative risks (with 95\u00a0% CI) were calculated where appropriate. The adherence and exit study populations were compared by comparison of proportions using the Chi squared test.Data were entered into EpiData 3.0 software . Data were cleaned by checking inconsistencies in data entry and responses. Data analysis was conducted using SPSS 10.0 for Windows . Baseline characteristics of patients/caretakers were described . The proportions of probable/certain (non-) adherence and complete/incomplete/correct treatment intake were expressed with 95\u00a0% confidence intervals (95\u00a0% CIs). Risk factors for non-adherence such as age, education level and knowledge of malaria were analysed. Both The Ethics Review Board of MSF and the Ethics Committee of DRC granted ethical approval. Authorities at all health levels and heads of the different quarters in Shabunda town were informed via personal visits before the study started. Written consent was sought from patients or responsible caretakers of patients before the adherence questionnaire was administered. Verbal consent was sought from patients, or responsible caretakers participating in the exit interviews.The adherence and exit questionnaires were anonymous. All data remained, throughout data entry and analysis, anonymous at the patient level. All participants received an explanation of the study purpose in the language they were most comfortable with\u2014Swahili, Kirega, or French. Participation was voluntary and all participants were offered the opportunity to refuse participation at any time without penalty.Data were collected between November 13 and 21, 2013. In total, 856 patients were screened at the two OPDs. Of these, 432 received an FDC ASAQ prescription (50.5\u00a0%), 262 (60.6\u00a0%) of whom were eligible for inclusion in the adherence study; 148 patients/caretakers were finally included . Seventeen  of the interviews at home were carried out 2\u00a0days after the last dose should have been consumed. No difference in adherence could be seen between the interviews carried out 1 or 2\u00a0days after the last dose should have been consumed.Among the 148 patients interviewed, 11.5\u00a0%  had one or more tablets left at the time of the home visit and were defined as certainly non-adherent; 13.5\u00a0%  were defined as probably non-adherent, giving a total of 25.0\u00a0%  non-adherence. Seventy-five percent  were defined as probably adherent  and no food/sugar available in the household for FDC ASAQ intake  Table\u00a0; 70.6\u00a0% None of the risk factors such as age, education level, and knowledge of malaria were associated with increased risk of non-adherence (n\u00a0=\u00a037). When adherence study patients or their respective caretakers were asked about the cause of malaria, 60.8\u00a0%  mentioned mosquito bites and 39.2\u00a0%  claimed not to know the cause of malaria or mentioned a wrong cause. In total, 79.1\u00a0%  mentioned that sleeping under an insecticide-treated mosquito net is a way of preventing malaria and 20.9\u00a0%  did not know any means of prevention or mentioned an incorrect means of prevention.p\u00a0>\u00a00.05).Exit interviews were carried out with 120 patients. The general characteristics of this group are shown in Table\u00a0Of the 120 patients in the exit interview group, 28 (23.3\u00a0%) had come without and 92 (76.7\u00a0%) with a caretaker to the OPDs. Of this group, 87.5\u00a0% (105/120) knew that they had malaria or could name the correct signs/symptoms of malaria. Eighty-nine percent (107/120) were able to identify the FDC ASAQ tablets (in combination with paracetamol) as anti-malarials among all the tablets given Table\u00a0. Eighty-According to patients/caretakers, 72.5\u00a0% (87/120) of the health staff in the OPDs had asked them if they had understood the FDC ASAQ intake instructions. Only 32.5\u00a0% (39/120) were asked to repeat these instructions and 35.8\u00a0% (43/120) were given additional information by health staff related to FDC ASAQ. In most cases  this advice was to take sugar water or eat before the FDC ASAQ intake.The results of this study show that good adherence to the correct intake of FDC ASAQ is possible even in this remote, conflict-affected region. The high reported rate (75\u00a0%) of probable adherence in this rural area is encouraging and could help prolong the effectiveness of first-line malaria treatment regimens. However, there was a (non-significant) finding that patients younger than 5\u00a0years were less likely to be completely adherent (62\u00a0%), which is concerning since children are more likely to develop severe disease, increasing the risk of drug resistance.When assessing the main reason for correct FDC ASAQ intake, almost all patients claimed they understood the instructions given to them at the OPDs. When assessing the exit interview group, an equally positive statement can be made for the quality of FDC ASAQ prescriptions and the instructions given at the OPDs: almost all patients were able to identify FDC ASAQ as the anti-malarial treatment amongst other concomitant treatments and could correctly repeat the instructions given to them in the OPDs. This shows much better quality of anti-malarial prescriptions than in a similar study conducted in The Gambia where only a third of caretakers were able to name the disease for which they had visited the health facility , 31.ACT should ideally be prescribed in combination with paracetamol. Very positively, almost 96\u00a0% of patients had received a prescription for paracetamol. According to the home interviews, adherence to paracetamol was high, with more than 90\u00a0% of patients with a paracetamol prescription taking it with the FDC ASAQ.Only 11.5\u00a0% of patients were certainly non-adherent with another 13.5\u00a0% meeting the criteria for probable non-adherence. Of the patients that were non-adherent (certain and probable), in two-thirds this was related to incomplete intake and in one-third to incorrect intake. The main reasons patients gave for incomplete intake was that the treatment made them feel sick or there was neither food nor sugar water at home. Two-thirds of the patients with incomplete intake did not take their third and last dose of FDC ASAQ. This result might be linked to the finding in the exit interviews that only a third of patients or caretakers were given any additional information related to FDC ASAQ. Of those who received additional information, none mentioned being told that the treatment may cause sickness and fatigue. The health staff placed emphasis on taking the treatment together with food or sugar water, which might have led to patients deciding to miss a dose rather than take it on an empty stomach. Around two-thirds of patients who took the drug incorrectly did so because they vomited immediately after intake. The remaining few patients with incorrect intake mostly took the second dose in the evening of the first day of treatment intake. These results matched the results of the exit interview group: only 10\u00a0% could not correctly repeat the instructions for FDC ASAQ intake, the most common error was to take the second dose in the evening of the first day.None of the risk factors such as age, education level, and knowledge of malaria were associated with an increased risk for non-adherence. Nevertheless the study population seems to have a relatively high level of understanding of the disease, shown previously to be significantly linked to good adherence . This coTwo different methods, considered to be complementary, were used to measure adherence in patients: the pill count and a systematic questionnaire. Biological assays may offer more precise adherence measurements \u201336. TheyOne of the most difficult things to avoid in adherence studies is influencing health staff such that they change prescribing habits. It is also important not to alert patients that there might be a later check-up regarding the treatment in order to avoid good-will bias , 17, 37.Initially the authors\u2019 wanted to include the hospital centre in the village of Matili, located roughly an hour\u2019s car journey from Shabunda town, as a study site. However, due to the deteriorating security situation in the region, only Shabunda town health facilities were able to be included. As very remote areas are usually linked to lower adherence, lower adherence levels might have been found if the most remote areas of Shabunda Health Zone had been included.Although more patients were eligible to participate in the study, the deteriorating security situation in the region meant that data collection needed to be completed as quickly as possible. Participant recruitment was therefore stopped once the calculated sample size had been reached.In 2005, the first systematic review on malaria adherence concluded that there was insufficient information on this subject and research would benefit from of standardization of methodologies . In 2014In the seven studies carried out on adherence to the ACT combination ASAQ in Africa between 2008 and 2012, adherence varied between 48 and 97\u00a0%. In three studies in which adherence was >90\u00a0% the purpose of the study was known both by health staff and the study population \u201341. In tThe adherence result of 75\u00a0% in this study is higher than in studies with a related design , methodology (self-reports from patients/caregivers in combination with pill counts), similar study areas , and preconditions . In Ethiopia, adherence was 39\u00a0%, in Zambia 40\u00a0%, in Sierra Leone 48\u00a0%, in South Sudan 53\u00a0%, in the Central African Republic 61\u00a0% and in Kenya 64\u00a0% , 43, 44.In the exit interview group, almost half the patients or their caretakers claimed not to have taken the first treatment dose under supervision in the OPDs. According to adherence interviews, the main reason was that the patients had not eaten prior to the OPD visit and therefore were told to take the first dose at home together with food. Given the high overall adherence in the study, the intake of the first dose under supervision does not seem to have a big influence on adherence and contradicts the results of previous studies where supervised intake of the first anti-malarial treatment was directly correlated with higher adherence , 42, 44.Access to prompt and effective treatment is a cornerstone of the current malaria control strategy. A delay in starting appropriate treatment is a major contributor to malaria mortality. Many patients and especially children with suspected malaria, where medical services are not easily accessible, start treatment too late or do not receive it at all and die at home. Therefore, since 2004, WHO has recommended home- or community-based management of malaria by trained community health workers as one of the strategies for improving access to prompt and effective malaria case management . HoweverTo the authors\u2019 knowledge, this is the first study to assess adherence to an FDC of ACT under real treatment conditions in a context of high instability. High quality prescribing of anti-malarials at health centre level and patient adherence to the correct intake of ACT were possible in this setting. Adherence to treatment regimens requires careful and constant monitoring, which might be better guaranteed at health centre rather than community level. It could, nevertheless, be a precondition to the successful introduction of home- or community based management of malaria."}
+{"text": "Plasmodium falciparum genes associated with resistance to AS  and SP ; and (c) to assess the adequacy of this ACT to clear gametocytes.In Yemen, artesunate plus sulfadoxine-pyrimethamine (AS\u00a0+\u00a0SP) has been used as first-line treatment for uncomplicated falciparum malaria, which accounts for about 99\u00a0% of malaria cases. There is evidence that resistance to SP is increasing, with potential negative impact on efficacy, and in particular on curbing transmission. This study aims: (a) to evaluate the therapeutic efficacy of AS\u00a0+\u00a0SP treatment for uncomplicated falciparum malaria in Yemen; (b) to investigate the frequency of mutations in pfdhfr, pfdhps, and pfK13 genes were obtained by sequencing following amplification.A 28-day in vivo evaluation of the clinical and parasitological response to three-day course of AS\u00a0+\u00a0SP was carried out in two areas of high endemicity  in Yemen according to standard WHO protocol 2009. Clinical and parasitological indices were monitored over a 28-day follow-up, and the outcome was PCR-corrected. The frequencies of mutations in the pfk13 propeller domain wild-type allele, and mutations associated with SP failure were observed only for pfdhfr with the double mutation (S108N\u00a0+\u00a0N51I) found in 65.4\u00a0% of the isolates sequenced.Eighty-six patients completed the study, with a cure rate of 96.5\u00a0% (94.2\u00a0% PCR-uncorrected). Whereas four (4.7\u00a0%) patients still showed parasitaemia on day 2 post-treatment, all were found negative for asexual malaria stages on days 3 and 7. The efficacy of gametocyte clearance was poor , with gametocytes persisting throughout the study in some patients. All the isolates sequenced had the pfk13 or pfdhps, though double mutations were observed for pfdhfr. The observed persistent gametocytaemia re-enforces calls to add a single dose primaquine to this ACT in order to minimizes the potential for transmission and enhance regional efforts to eliminate malaria.In Yemen, AS\u00a0+\u00a0SP therapy remains effective for the treatment of uncomplicated falciparum malaria. Mutations were not detected in The online version of this article (doi:10.1186/s12936-016-1344-0) contains supplementary material, which is available to authorized users. Plasmodium falciparum resistance to anti-malarial drugs led to the development of new strategies to treat malaria and extend the life of novel drugs. Over the last decade, artemisinin-based combination therapy (ACT) has become the cornerstone of malaria treatment policies [P. falciparum lines with reduced susceptibility to artemisinin [pfk13 propeller domain have been reported from African P. falciparum, but none was associated with artemisinin resistance [The worldwide spread of policies , 2. Thisemisinin \u20135 raisesemisinin . To dateemisinin \u20139, with emisinin , 10\u201312. sistance .P. falciparum (99\u00a0%), with Anopheles arabiensis, Anopheles sargentii and Anopheles azaniae as the major insect vectors [P. falciparum infections, with a combination of artemether with lumefantrine (AL) as a second-line treatment [P. falciparum populations in Yemen that whereas no mutations associated with drug resistance occur for pfdhps, the prevalence of such mutations in pfdhfr is increasing [In Yemen, malaria is still a major life-threatening health problem for more than 60\u00a0% of the population of nearly 25 million people, and in particular in the southwestern corner of country [ vectors . The cou vectors , with tw vectors , 20. The vectors \u201324 prompreatment , 26, thecreasing \u201330. Finacreasing .pfk13 gene propeller domain, pfdhfr and pfdhps genes of the P. falciparum parasites circulating in this area. Furthermore, the impact of the AS\u00a0+\u00a0SP treatment on microscopic gametocytaemia was also evaluated throughout the follow-up period.The present in vivo trial aimed to assess the clinical and parasitological efficacy of AS\u00a0+\u00a0SP treatment in an area of high endemicity close to the Saudi Arabian border, and to investigate the polymorphisms in the An active case study was carried out among febrile individuals, suspected to have malaria infection, was conducted from March to May 2014 in some malaria-endemic areas in two provinces, Hodeidah and Al-Mahwit located in Tehama region, northwest Yemen , Yemen. These included AdDahi and Bajil Districts in Hodeidah Province, and Khamis Bani-Saad District in Al-Mahwit Province. According to the NMCP records for 2013 . The filter papers were left to dry in air away from dust and kept in an aluminium foil pouch with desiccant silicon bags at room temperature (25\u201327\u00a0\u00b0C) until use. This sample obtained prior to the administration of treatment  was considered to be the day 0 sample.A finger-prick blood sample was collected from each participant for rapid diagnostic test (RDT) , for preparing thick and thin blood smears, and to obtain blood spots on 3MM WhatmanThe blood smears were Giemsa-stained prior to microscopic examination for the identification of parasite species and stages. The parasitaemia was determined using a modified procedure for counting the asexual stages. Parasites were enumerated against 300 white blood cells (WBC) with a hand tally counter and the number was then multiplied by 25 to obtain the number of parasites per \u00b5L of blood (assuming an average of 7500\u00a0WBC/\u00b5L blood). The parasitaemia was graded as low (<1000 parasites/\u00b5L blood), moderate (1000\u20139999\u00a0parasites/\u00b5L blood), or severe . The presence of gametocytes was examined against 500 WBC; though gametocytaemia was treated as qualitative variable in data analysis. Each microscopy-positive slide and a selection of negative slides were examined by two senior specialists and the average parasite count was considered.P. falciparum mono-infection, and ability to swallow oral medication were invited to participate in the in vivo trial. Details concerning the study, its benefits and any possible potential risks were fully explained. Only those who willingly agreed to participate and gave informed written and signed consents as well as declared their willingness to comply with the study protocol for the duration of the study and to comply with the study visit schedule were enrolled in the study. Patients who had signs of severe or complicated falciparum malaria or had mixed infection with another Plasmodium species (P. vivax) or had febrile conditions due to diseases other than malaria or had a history of adverse effects due to any anti-malarial drug or any other medicines were excluded from the in vivo study. Moreover, pregnant women and females with positive pregnancy test or females who were unable to or unwilling to do a pregnancy test were also excluded.All the patients who fulfilled the inclusion criteria, including age of 6\u00a0months and above, axillary temperature of\u00a0\u226537.5\u00a0\u00b0C or history of fever within the last 72\u00a0h, microscopically confirmed as uncomplicated Treatment efficacy was evaluated by monitoring the clinical and parasitological parameters over a 28-day follow-up period from the day the first drug dose was administered (day 0). The methods of treatment, schedule of follow-up and analysis of outcomes were as per the WHO guidelines . The druACT in the form of AS\u00a0+\u00a0SP is the first-line treatment in Yemen for uncomplicated falciparum malaria. AS\u00a0+\u00a0SP used in this study was provided by WHO through NMCP: 50\u00a0mg artesunate tablets in a strip of 12 tablets  and sulfadoxine 500\u00a0mg/pyrimethamine 25\u00a0mg tablets in a pack of 1000 tablets . The drug was administered at a daily dose of artesunate 4\u00a0mg per kg of body weight (BW) for three successive days  plus a single dose of 25/1.25\u00a0mg per kg BW of SP, respectively, on the first day (day 0). Additionally, NMCP provided the study with the second-line treatment AL should AS\u00a0+\u00a0SP therapeutic failure occur, and parenteral quinine therapy for those who might vomit the treatment twice and/or could not take the medication orally, for pregnant women during first trimester, for patients with severe or complicated malaria and in case of AL therapeutic failure.\u00ae Blood & Tissue Kit, Cat. no. 69506, Germany) according to manufacturer\u2019s instruction. DNA was eluted using 100 \u00b5L of the elution buffer AE  and kept at \u221220\u00a0\u00b0C until use.Two to three discs (6-mm diameter) of the blood spot dried on the filter paper were cut using a flamed-sterile puncher, and were then used for DNA extraction using a Qiagen blood and tissue kit , merozoite surface protein-2 (msp2) and glutamate rich protein (glurp) genes.Recurrent cases were distinguished from new infections (PCR-correction) according to the protocols described previously , 36 for \u00ae safe DNA gel stain  using UV documenting system . The allelic variants observed were then grouped into different size bins of 25\u00a0bp for msp1 and msp2, and of 50\u00a0bp for glurp. For those cases where parasites were observed during the follow-up period, msp1 and msp2 allelic variants were compared between the samples obtained before and after treatment  by capillary electrophoresis in order to increase sensitivity and resolution. Forward oligonucleotide primers of secondary PCR for msp1  and for msp2 (M2-FCF and IC-F) were labelled with 6-FAM fluorescent dye. Amplicons were analysed by the automated ABI 3730XL Genetic Analyzer and then interpreted using GeneMapper\u00ae analysis software version 4.0 . Details of primers sequences and the PCR conditions used to amplify msp1, msp2 and glurp are available in Additional file Amplification of the three markers was achieved using nested PCR . AmplicoP. falciparum isolates, and the mutations identified following direct sequencing or through restriction fragment length polymorphism (RFLP) analysis.Selected domains of the parasite genes associated with drug resistance were amplified by conventional or nested PCR using genomic DNA purified from the pfk13 propeller domain A single run PCR using forward and reverse primers K13-F (5\u2032-GTTGGTGGAGCTATTTTTGAAACATCTAG-3\u2032) and K13-R (5\u2032-GCCAAGCTGCCATTCATTTGTATC-3\u2032) that were designed based on P. falciparum 3D7 kelch gene , yielding 1062\u00a0bp amplicon that corresponds to nucleotides 1094\u20132127 (codons 364\u2013709). The PCR products were then visualized by 1\u00a0% agarose gel stained with Sybr\u00ae safe DNA gel stain under UV, then gel-purified, sequenced in both directions, and the resulting sequences aligned using BioEdit Sequence Alignment Editor Software (version 7.1.9) and compared to P. falciparum 3D7 Kelch 13 gene (Gene ID: 814205). All positions were evaluated for the presence of mutations. Details of the primer sequences and PCR conditions used are available in Additional file pfdhfr and pfdhps.pfdhfr was analysed for mutations at six codons  and pfdhps at seven codons . Amplification of pfdhfr was achieved by a modified nested PCR protocol [pfdhps amplification was slightly modified: the upstream primers for both primary and secondary PCR were changed so as to yield a longer amplicon (1005\u00a0bp). PCR amplification was performed using PS1-F (5\u2032-GAATTTTTATCCATTCCTCATG-3\u2032)\u00a0+\u00a0O2 (5\u2032-TTCCTCATGTAATTCATCTGA-3\u2032) and PSA-F (5\u2032-GTATACAACACACAGATATAG-3\u2032)\u00a0+\u00a0O2 primers pairs for the primary and secondary amplification reactions, respectively [P. falciparum.  for data analysis. For descriptive analysis, frequency and proportion were used to present the distribution categorical variables. All quantitative variables were examined for normality by Shapiro\u2013Wilk test before analysis. Statistical associations between point mutations and explanatory variables, including age, gender, sites, and parasitaemia, were assessed using the Chi square test or Fisher\u2019s Exact test where applicable. Per-protocol and Kaplan\u2013Meier survival analysis was used to evaluate the treatment outcome and to plot the decline in gametocytaemic cases after treatment among individuals who were gametocyte-positive at enrolment. Patients were censored\u00a0from the per-protocol analysis of treatment outcome if they were lost to follow-up, identified with re-infection (after PCR-correction) or decided to withdraw from the study, but they were considered in the Kaplan\u2013Meier survival analysis until the day of withdrawal. A The study protocol was approved by the Medical Ethics Committee of the University of Malaya Medical Centre, Kuala Lumpur (Ref. 974.19), and by the Ministry of Health and the NMCP in Yemen. At the villages, the residents were informed about the aims and procedures of the study. Moreover, the participants were also informed that they could withdraw from the study at any point of time without citing reasons for doing so. Afterwards, written and signed or thumb-printed informed consents were taken from adult participant or from the parents on behalf of their children before enrolment, as approved by the mentioned Ethics Committees. The consent request form was translated into Arabic language and was read entirely to the patients or their parents/guardians before they were asked to sign the document. Furthermore, permission for pregnancy testing was sought from married female participants aged 18\u00a0years and above. RDT-positive individuals were treated according to the national malaria treatment policy, Ministry of Health and Population, Yemen (as stated in Methods). Any patient who decided not to participate or to continue in the in vivo study was also treated and followed-up until total clearance of parasitaemia by day 28.Of the 622 individuals screened, 188 (30.2\u00a0%) and 189 (30.4\u00a0%) individuals were found positive for malaria by using the CareStart\u2122 HRP2-RDT and microscopy, respectively. However, only 89 met the inclusion criteria and consented to be enrolled in the study. All were given AS\u00a0+\u00a0SP and followed up for 28\u00a0days. Two patients were lost to follow-up from day 14 onwards and one patient refused to continue from day 2, leaving 86 patients  who were successfully followed up until day 28. Admission asexual parasite levels varied from 561 to 55,555 and with mean parasitaemia of 8199\u00a0parasites/\u00b5L blood. The general characteristics of the 86 participants are shown in Table\u00a0Overall, asexual parasite clearance was rapid as no patient was found parasitaemic on days 3 and 7. By day 1, 51 patients (59.3\u00a0%) were cleared of parasites and only four (4.7\u00a0%) patients still had microscopically detectable parasitaemia on day 2 which subsequently cleared on day 3. Asexual parasites with fever reappeared (recrudescent/re-infection cases) on day 14 in two patients, on day 21 in another, and finally on day 28 in two further patients . Moreover, almost similar mean baseline parasitaemia was reported between the LCF and the ACPR cases .After Kaplan\u2013Meier analysis, therapeutic outcome of AS\u00a0+\u00a0SP was 94.2\u00a0% (before PCR-correction) with 81 cases showing adequate clinical and parasitological response Fig.\u00a0. Thus, tPfk13-propeller sequences were obtained from all 86 (100\u00a0%) isolates collected in this study. None of the study isolates analysed for mutations of the pfK13 propeller domain (including the three recrudescent isolates) carried any mutations at the 39 codons previously published to be associated with artemisinin resistance [m all 86 0\u00a0% isolaPfdhfr and pfdhps. For pfdhfr, sequences were obtained from 81 (94.2\u00a0%) of the study isolates. Double mutations at S108N and N51I were observed in 53 isolates (65.4\u00a0%), and all the other codons  were found to be wild type. Thus, the ACICNI pfdhfr haplotype was predominant (65.4\u00a0%) over the wild-type ACNCSI haplotype. For pfdhps, sequences were obtained from 76 isolates (88.4\u00a0%), and no mutations were observed for codons 436, 437, 540, 581, and 613, though mutations were in two other codons: 640 (n\u00a0=\u00a06) and 645 (n\u00a0=\u00a04). The distribution of the pfdhfr double-mutated haplotype frequency was found significantly associated with the districts and sites from which the participants were recruited: in 87.5 and 80.4\u00a0% of those from Ad-Dahee and Khamis Bani-Saad, respectively, while patients from Bajil districts harboured parasites that were mostly wild type (5.9\u00a0% with mutated haplotypes). Parasites obtained from patients from the villages of Al-Humarah (AdDahee) and Shat Hajal (Khamis Bani-Saad) all carried the double mutation. No association was reported between the frequency of pfdhfr mutations and other variables such as sex and age, admission parasite density, or gametocytaemia.P. falciparum during the initial survey, and a similar proportion of the patients recruited (35/86) were also gametocytaemic at the initiation of AS\u00a0+\u00a0SP treatment. Most of these patients still harboured gametocytes by day 7 post-treatment, and 14\u00a0% were still gametocytaemic at the end of the trail follow-up period . High levels of resistance to SP have generally precluded the selection of SP as the ACT companion drug, and the AS\u00a0+\u00a0SP combination has been uniquely adopted in countries of WHO\u2019s Eastern Mediterranean Region (except for Djibouti) and India where it is supplemented by primaquine, but it is administered alone in Azerbaijan, Somalia, Sudan, and Yemen. Increased frequencies of mutations associated with SP failure have been recorded in Sudan , and in pfdhfr double mutation. Given high cure rate and the absence of resistance-associated mutations in the pfdhps gene and the rarity of triple mutations in the pfdhfr gene in Yemen [The study presented here indicates that the efficacy of AS\u00a0+\u00a0SP treatment is still high in Yemen (>95\u00a0%), which concords with the only other in vivo efficacy trial that was recently conducted in the same country . In bothin Yemen , a recomin Yemen , 43.pfdhfr and pfdhps mutations has been reported in the Jazan region in southwest Saudi Arabia that shares borders with this study area [pfdhfr double mutation  predominates [pfdhfr\u00a0and\u00a0pfdhps genes reported in Pakistan, the\u00a0triple pfdhfr\u00a0mutant was not observed, though 52\u00a0% (88/170) of the isolates studied harboured the pfdhps A437G mutation with the double mutated (C59R\u00a0+\u00a0S108N) pfdhfr [pfdhfr\u2009\u00a0+\u00a0\u2009pfdhps triple mutations (pfdhfr C59R\u00a0+\u00a0S108N and pfdhps A437G) during 2008\u20132010 following the adoption of AS\u00a0+\u00a0SP in 2007 [In neighbouring and other Eastern Mediterranean countries, a similar profile of udy area , and in ominates . Despite) pfdhfr . Studies in 2007 , but a s in 2007 . Ultimatpfk13 propeller domain in P. falciparum from Yemen indicated that the parasites are still free of the mutations associated with artemisinin resistance in Asia. This might be due to the relatively short exposure of the parasites to artemisinin as well as the use of SP as the partner drug. Indeed, although ACT was introduced in 2009, its distribution at district level was gradual and its widespread use is slowed by continuing availability and prescription of chloroquine in both urban and rural areas [pfk13 should be regularly monitored in Yemen.This first investigation of the al areas , 42, 43.In the context of sustainable malaria control, the use of SP for treatment suffers from a major disadvantage, namely a poor efficacy in eliminating the sexual stages. Thus, when SP was assessed in in vivo trials alone or in combination with artemisinins, the rapid clearance of asexual parasites was not observed for the gametocytes that often persisted for many days, sometimes throughout the follow-up period, in the circulation and remained infectious to mosquitoes. It is likely that the prevalence of post-treatment gametocytaemia was underestimated when microscopy alone was used to detect the gametocytes. Indeed, numerous studies showed that molecularly detectable sub-microscopic gametocytes are common following treatment and could be sufficient to sustain transmission \u201354. In YThe data presented here reinforce the suggestion that the fate of the sexual stages should be considered as an additional factor to measure in standardized protocols of in vivo anti-malarial efficacy studies when the objectives of the malaria control programme also aim to reduce transmission . Furtherpfdhfr\u00a0genotype was found in 65.4\u00a0% of the isolates while no mutation was reported in the pfdhps as well as pfk13 genes. Moreover, persistent gametocytaemia after AS\u00a0+\u00a0SP treatment suggests the addition of primaquine in order to reduce malaria transmission sustainably. This is all the more important in Yemen, where disruption of the health infrastructure and population displacements due to the current political situation are likely to lead to an exacerbation of malaria and to threaten the surrounding countries in the Arabian Peninsula.The data from Yemen indicates that AS\u00a0+\u00a0SP is still efficacious for treatment of uncomplicated falciparum malaria. Double mutant"}
+{"text": "Plasmodiumfalciparum has posed an obstacle to effective treatment and challenges many malaria control programmes in endemic areas. In Angola, until 2003, chloroquine (CQ) was used as first-line therapy for uncomplicated malaria. It was replaced initially by amodiaquine and, in 2006, by artemisinin-based combination therapy (ACT) with artemether-lumefantrine . Efficacy study of ACT, conducted in Angola between 2004 and 2005, showed a baseline efficacy of \u224899\u00a0%.Drug resistance in pfmsp2 were genotyped by nested-PCR, to distinguish parasite recrudescence from new infections; the polymorphisms at codons 86 and 1246 of pfmdr1 gene, and 769 of pfatp6 gene were assessed by PCR\u2013RFLP and sequencing for pfk13-propeller genotype.103 malaria patients were enrolled according to WHO proceedings. Patients were followed up with clinical and parasitological evaluations for 28\u00a0days, parasite density and identification was evaluated by microscopy, the pfmdr1 N86Y position and only one sample carried the mutant allele (Y) for pfmdr1 1246; 14\u00a0% of these samples showed increased pfmdr1 copy number; 100\u00a0% (n\u00a0=\u00a021) had wild-type allele of k13 gene in all the studied positions.The cure rate was 91.3\u00a0%. The obtained results showed that from 103 patients, 12.6\u00a0% (n\u00a0=\u00a013) still had parasitaemia 1\u00a0day after the treatment was finished. On day 0, of the 94 evaluated samples, wild-type alleles were identified in 73.4\u00a0% (n\u00a0=\u00a069) for These results showed changes in parasite profile susceptibility to AL in comparison to the baseline data from 2002 to 2004 and on the genotyping characteristics; the clinical outcome after treatment with AL did not link a particular genotype with treatment failure; observed changes do not provide sufficient evidence for a treatment policy change, but they suggest that a carefully monitoring is needed in this area. Malaria is the leading cause of morbidity in countries where it is endemic; an estimated 3.3 billion people are at risk of being infected with malaria and developing disease worldwide , 2 thougMalaria remains a public health challenge in Angola with changes in mortality over the last 3\u00a0years. According to an unpublished report 2014) of National Malaria Control Programme (NMCP), between 2010 and 2012 deaths attributed to malaria decreased from 8114 to 5736, but in 2013, 1,999,868 cases and 7300 deaths due to malaria were reported [014 of NaAn evaluation of anti-malarial efficacy of chloroquine (CQ), amodiaquine (AQ) and sulfadoxine-pyrimethamine (SP) carried out by the Angolan NMCP, in the period between 2000 and 2005, revealed treatment failure of 57.7, 3.8 and 15.4\u00a0%, respectively. Studies of artemether-lumefantrine (AL) and artesunate\u00a0+\u00a0amodiaquine (ASAQ), efficacy, conducted in 2004 and 2005, revealed efficacy of \u224899\u00a0% , 7. BaseDespite the WHO recommendation of ACT, there are worrying signs associated with loss of efficacy. Existence of isolates from different areas with markedly reduced susceptibility to ACT was reported. A loss of efficacy of ACT in western Cambodia, with slower parasitological responses to anti-malarial drugs, principally an unusually high failure rate of artesunate\u00a0+\u00a0mefloquine, suggesting the emergence of artemisinin resistance, were published in 2009. In addition, prolongation of parasite clearance time (PCT) has been reported on the Thai-Myanmar border, indicating that artemisinin resistance could be spreading outside Cambodia , 10, 11.Plasmodium falciparum multidrug resistant 1 (pfmdr1) gene copy number, or specific point mutations in its codons 86, 184, 1034, 1042 and 1246 have been associated with the susceptibility to this class of anti-malarials [pfmdr1 copy number and the pfmdr186N allele [pfmdr1 gene has been linked with lumefantrine and mefloquine resistance and to the risks of treatment failure with artesunate\u2013mefloquine and AL in Southeast Asia, while mutations at the codon 86 of the pfmdr1 gene can modulate lumefantrine efficacy [pfatp6 can also mediate changes in artemisinin susceptibility; specially the mutation S769N [k13) propeller domain were found to be associated with delayed parasite clearance; this may open new possibilities for tracking resistance to artemisinins [The genetic basis of artemisinin resistance is not yet known; several polymorphism as N allele , 14. Mulefficacy . Polymoron S769N , 16, 17.misinins .\u00ae for the treatment of uncomplicated P.falciparum infection, in Luanda, Angola, and to analyse the polymorphisms in pfmdr1, pfatp6, and K13 genes previously associated to susceptibility to AL, 6\u00a0years after its introduction on this area.The present study aims to evaluate the therapeutic efficacy of CoartemThis was an interventional prospective single cohort study conducted in Luanda, Angola, considered as malaria mesoendemic stable area. Blood sample collection was carried out in 2011, 2012 and 2013 in the period of high malaria transmission (between April and June). Patients were recruited and followed at three health centres: a Catholic Church health care unit  and two military health units (Cl\u00ednica do Ex\u00e9rcito and Instituto Superior T\u00e9cnico Militar). Subjects were selected among children and adults with confirmed malaria infection, based on the WHO 2009 guidelines for drug efficacy study .P. falciparum confirmed by microscopy. Further, other criteria were included: (a) to have non-complicated malaria ; (b) to have an initial parasite density between 1000 and 100,000 asexual parasites/\u03bcL; (c) exclusion of the concomitant diseases, after a clinical interview and examination; (d) voluntary participation in the study with written informed consent; (e) commitment to attend the follow-up examination on indicated days.Patients were eligible to participate, if older than 6\u00a0months of age, axillary temperature of at least 37.5\u00a0\u00b0C or history of fever within the last 24\u00a0h, with symptoms compatible with malaria, and infection with Patients with evidence of severe malaria or any other disease, presence of severe malnutrition or with appearance or history of adverse effects due to any anti-malarial drug, to other medicines, or to other causes were excluded from the study. Patients who did not meet these basic enrolment criteria were treated in accordance with routine practice of each institution. Patient\u2019s selection was conducted by a skilled team, including physicians, nurses, microscopy\u2019s technicians and one epidemiologist.Ethical approval was obtained from the Ethic Committee of National Public Health Institute . All participants received information about study objectives and protocols and informed written consent was obtained from all eligible participants/parents or guardians.Sample size should be calculated based on the expected proportion of treatment failures in the studied population . As this\u00ae, Novartis, Basel Switzerland). For adults, four tablets were administered twice a day; doses were adjusted according to weight for children under 35\u00a0kg [\u00ae was given to the patient to be taken at home. Patients were recommended to take the tablets with fatty food or milk to improve their absorption [Patients were treated with six doses of AL  and on each follow-up day . Thick blood films stained with Giemsa were prepared and examined , on eachOutcomes were classified according to WHO anti-malarials drug efficacy test guidelines 2009  as: \u2018EarPlasmodium falciparum nested-PCR was performed according to the protocol described by Snounou et al. [Plasmodium species and for merozoite surface protein2 (pfmsp2\u2014FC and IC families) genotyping of on D0 and to positive samples from post-treatment (after D7) in order to distinguish parasite recrudescence from reinfection [Extraction of DNA from dried blood spot on filter paper was carried out using phenol\u2013chloroform method previously tested , 26. Plau et al. , 28, fornfection , 30. Recpfmdr1, and 749 of pfatp6 genes were performed by conventional RFLP-PCR [pfmdr1 86N, EcoRV (New England Biolabs) assessed to the 1246Y alleles and enzyme RsaI (New England Biolabs) for pfatp6 769S allele [\u00ae LEAgarose, ref. 50004) gel stained with ethidium bromide (Sigma-Aldrich E1510) and visualized under UV [The analysis of polymorphisms of the parasite, based on mutations at positions 86 and 1246 of RFLP-PCR . ApoI , as previously described [\u00ae Green Supermix , 300\u00a0nM primers and 2\u00a0\u00b5L de DNA was added to the mixture. All samples were run in triplicate and the DNA of P. falciparum strains, 3D7 and Dd2 were used as single and multiple copy calibrators, respectively. pfmdr1 copy number was calculated using a comparative threshold method (2\u2212\u0394\u0394Ct method) [pfmdr1 copy number when the sample had pfmdr1 folds >1.5 by RT-PCR.The variation of escribed , 33, 34.K13-propeller domain was amplified according to previously described methods [The  methods , for ide methods , 20.Data were entered and analysed using Epi-info 7 version. Mixed infections thus contribute to the prevalence of both alleles were excluded of all statistical analyses.During the 3\u00a0years of study, a total of 3628 patients (adult and children) were screened for malaria parasites in the selected health care units. Of these, 972 (26.8\u00a0%) had positive microscopic malaria diagnosis; this result shows the need to consider the diverse causes of febrile syndrome in malaria endemic regions. Initially, 122 patients were selected and from these, nineteen were excluded: two vomited persistently, nine were lost during the follow-up period and 8 had a negative PCR at day 0. A total of 103 patients met the required criteria and were enrolled in the study; and from these 21 (20.4\u00a0%) were children under 5\u00a0years of age.Treatment was well tolerated, no patients presented relevant complaints or treatment adverse effects.In this the study, in 2011, 37 patients were enrolled, 33 in 2012 and 33 in 2013, giving a total sample of 103 patients, aged between 6\u00a0months to 56\u00a0years of age; 716 blood samples were collected during the follow up days for molecular analysis. The group of children under 5\u00a0years of age represented 16\u00a0% of sampling (114 samples from 21 children). Distribution of observed patients and microscopy evaluation by days of follow-up is shown on Table\u00a0\u00ae treatment), 13 of the 103 patients (12.6\u00a0%) had parasites detectable by microscopy, six of those patients  still had parasites detectable by microscopy on D7. Using the PCR method, 48 patients (46.6\u00a0%) on D3 and 35 (34\u00a0%) on D7 gave positive results for presence of P. falciparum. Samples with positive PCR had correspondence with follow-up days with positive microscopic results for asexual forms and/or gametocytes. The high sensitivity of the PCR method to detect the parasites at sub-microscopic densities, gametocytes presence and errors in microscopic diagnostic are factors that can generally explain the difference on results obtained by microscopic and PCR methods [On D3  of the patients on D0. Seven patients (6.8\u00a0%) still had gametocytes detectable on D7 and were still found in three patients on D14, in one on D21 and one on D28, after the asexual parasites had been cleared. The presence of gametocytes on D0 can suggest the usual existence in the endemic areas of long-lasting infections, initially asymptomatic with low parasitaemia, which can turn manifest when there is debility of host defense or new additional infection. This condition promotes maintenance of disease transmission in the community .The average fever clearance time was \u224824\u00a0h after the initial dose of treatment, 97\u00a0% of the patients had no fever at the second day and 99\u00a0% were not febrile on the third day of medication (D2). One patient was still febrile and with parasitaemia on D2, being classified as ETF. By D3 the totality of patients was cleared of fever. During the follow-up period 13 and 6 patients still had parasite positive blood smear on days 3 and 7 respectively, not presenting any clinical signs.Of the 103 patients, in the study, 90.3\u00a0% (n\u00a0=\u00a093) were classified as ACPR. Nine patients exhibited parasitaemia after treatment with AL. The global failure rate before PCR correction was 9.7\u00a0% (n\u00a0=\u00a010). One patient, under 5\u00a0years of age, had persistent fever and parasitaemia until day 2, which was classified as ETF, according to WHO protocol, eight patients were classified as LPF, being six asymptomatic patients with parasite positive blood smear on D7 and two asymptomatic with blood parasitaemia on D14 and D28 respectively. One febrile patient presented parasites on D28 and was classified as LCF. After PCR correction, the overall cure rate was 91.2\u00a0% (n\u00a0=\u00a094); recrudescence was found in eight patients and reinfection in one. Among the 21 children \u22645\u00a0years of age included in the study, five (23.8\u00a0%) presented treatment failure and three of these had anti-malarial medication under direct observed regime, at Divina Provid\u00eancia Hospital.According to the age group, the frequency of ACPR was 76.2\u00a0% in patients \u22645 (n\u00a0=\u00a016/21) years and 95.1\u00a0% (n\u00a0=\u00a078/82) in those aged >5\u00a0years; there was no statistically significant association between treatment outcomes and age groups (p\u00a0=\u00a00.334). The treatment outcome is shown in Table\u00a0P. falciparum sensitivity to artemisinin and its derivatives, defining suspected resistance to artemisinin as an increase in parasite clearance time, as evidenced by 10\u00a0% or more cases with parasites detectable on day 3 of treatment with ACT. Changing of anti-malarial treatment policy is recommended when the treatment failure rate exceeds 10\u00a0% [The WHO considers the proportion of patients with parasitaemia on day 3 of treatment as routine indicator in monitoring of the eds 10\u00a0% , 10, 11.In this study, 12.6\u00a0% n\u00a0=\u00a013) of the 103 followed patients had still microscopic parasites detectable on D3 and failure rate of 8.8\u00a0% was met. Considering WHO parameters, the results met in this study might suggest an elongated parasite clearance time , 11. The of the 1pfmsp2 genotyping was performed for all samples on day 0 and for those with microscopic positive result from D14 to differentiate between new infection and recrudescence. The pfmsp2 genotyping was successfully assessed in 96 samples from (D0). Of these, 32 samples (33.3\u00a0%) were collected in each year, respectively 2011, 2012 and 2013. From the pfmsp2 genotyping analysis, 72\u00a0% (n\u00a0=\u00a069) of the 96 samples had just one of the studied alleles (FC27 or IC1), therefore termed as monoclonal infections and the IC1 family was predominant with 62.3\u00a0% (n\u00a0=\u00a043) of monoclonal samples.pfmsp2 in the 3 microscopic positive samples from day 14 demonstrated that 2 had the matching allelic bands of day 0, indicating recrudescence. One positive sample on day 28 did not have the matching allelic bands of day 0, indicating reinfection. Seven of eight amplified samples from patients with treatment failure carried IC1 family alleles. The statistic tests did not evidence significant association between msp2 families identified and treatment responses (P\u00a0=\u00a00.115). In this study, the reduced number of samples with parasites detectable from day 14 constitutes a limitation for relevant conclusions on recrudescent and new infection rates.The remaining mixed infections, with FC and IC families, were found in 28\u00a0% (n\u00a0=\u00a027) of the analysed samples. Polyclonal infections were found in 37.5\u00a0% of cases (n\u00a0=\u00a036/96) on day 0. A progressive elimination of mixed variants post-treatment was noticed. After day 3, no more samples with mixed families were detected. The genotyping of pfmdr1 86 SNP, out of 94 analysed samples, 28 (29.8\u00a0%) were collected in 2011, 33 (35.1\u00a0%) isolates collected in 2012 and 33 (35.1\u00a0%) in 2013. The global ranges of frequency were 73.4\u00a0% for the wild allele (N86), 18.1\u00a0% for the mutant (86Y) and 8.5\u00a0% for the mixed alleles (N\u00a0+\u00a0Y). The ranges of frequencies by years for pfmdr1 N86 allele were 68\u00a0% (19/28) in 2011, 69.7\u00a0% (23/33) in 2012 and 82\u00a0% (27/33) in 2013. In the respective years, the frequencies of the mutant 86Y were 21, 18.2 and 15\u00a0%; and the mixed infections was found in 11, 12 and 3\u00a0% in 2011, 2012 and 2013, respectively.Gene fragments containing the SNP 86 (N/Y) and 1246 (D/Y) were successfully amplified on D0 in 94 and 96 samples, respectively. For the analysis of pfmdr1 86 position, there was no significant association between the presence of pfmdr1 86 polymorphism and the year o collection (p\u00a0=\u00a00.869). The pfmdr1 D1246 was the predominant allele in the isolates for all time points, and one isolate only, collected in 2012, presented the mutant allele 1246Y, this isolate was obtained from a patient classified as ACPR and which carried a mixed 86\u00a0N\u00a0+\u00a0Y allele.These results show a predominance of the wild-type allele for the The results of this study are in agreement with those reported by Fan\u00e7ony et al. in northern U\u00edge province of Angola in 2009, where rates of 68.1, 17.4 and 14.5\u00a0% for N86, 86Y and 86\u00a0N\u00a0+\u00a0Y, respectively, were encountered ; howeverThe analysis of pooled data from 31 studies (1995\u20132010) conducted by Venkatesan et al. in 2014 reported rates of 37\u00a0% (N86), 44\u00a0% (86Y) and 19\u00a0% (86\u00a0N\u00a0+\u00a0Y) for Est Africa, 51\u00a0% (N86), 14\u00a0% (86Y) and 34\u00a0% (86\u00a0N\u00a0+\u00a0Y) for West Africa and 71\u00a0% (N86), 29\u00a0% (86Y9) and 0\u00a0% (86Y\u00a0+\u00a0Y) in Asia/Oceania .pfmdr1 genotype profile trend. Recently, several field studies have suggested that AL appeared to select the N86 wild-type allele of pfmdr1 gene associated to reduced susceptibility to lumefantrine. Also, the predominance of this wild-type allele can be related to the elimination of CQ and AQ monotherapy [Variations in drug exposure seem to affect the otherapy , 45. Thepfmdr1 1246 polymorphism, the wild-type (D1246) was present as the major allele in the isolates. Just one isolate collected in 2012 presented the mutant allele 1246Y, this isolate was from a patient classified as ACPR and which carried a mixed 86\u00a0N\u00a0+\u00a0Y alleles. The results of this study are in accordance with those found in a majority of studies in the African region where usually the mutant 1246Y is not frequent [pfmdr1 1286Y and 76.7\u00a0% for D1246 alleles [For the frequent , but dif alleles .pfmdr1 1246Y in isolates from West Africa (Guinea Bissau), in DRC  an in this study (Angola) reveals a emergence of this genotype previously considered rare in Africa but common in South America. The presence of these genotypes on this area may due to the increased migration phenomenon and globalization.Detection of the pfatp6 gene were assessed in all samples (103), none of the isolates displaying the 769\u00a0N mutation; it should be noted that even the recrudescent samples exhibited the wild-type S769 haplotype. This result coincides with Menegon et al., where wild-type allele S769 was found in all isolates collected in 2004. Similar results were reported from studies conducted in the Africa Region [SNP at 769 (S769N) position of the a Region , 15.pfK13-propeller gene, all samples (n\u00a0=\u00a021) processed by sequencing exhibited the wild type allele in all analysed positions, including in SNPs considered associated to artemisinin resistance . The results of this study are in agreement with those reported by Plucinski and by Escobar et al., where wild-type alleles were found at the rendered positions of pfk13 gene, in isolates from Angola and Mozambique [Regarding zambique . The frepfmdr1 copy number on D0. The results demonstrated that 13 isolates (14\u00a0%) showed increased copy number, ranging from 2 to 3 copies. The majority of analysed isolates had a single copy of the gene pfmdr1 , 12 isolates (13\u00a0%) had two copies (1.6\u00a0\u2264\u00a02.5 folds) and one sample (1\u00a0%) had three copies (3.44 folds). The proportions of samples with amplified pfmdr1 by year of collection were: 12.5\u00a0% (n\u00a0=\u00a04/32) in 2011, 13.8\u00a0% (n\u00a0=\u00a04/29) in 2012 and 15.6 (n\u00a0=\u00a05/32) in 2013.The Table pfmdr1 increased copy number, 92.3\u00a0% (n\u00a0=\u00a012/13) were from patients classified as ACPR. The nine patients classified as treatment failure exhibited a single copy of pfmdr1 whenever tested, whilst the patient initially classified as LCF was finally confirmed as a new infection had presented two copies of the gene on D0 and a single copy for D28.According to AL treatment outcomes, of the 13 samples with pfmdr1 gene has been associated with clinical failures and within vitro resistance to aryl-amino-alcohols, particularly mefloquine (MQ), quinine (QN), and halofantrine, but also to lumefantrine [pfmdr1 and therapeutic response to Coartem\u00ae. However, the frequency of isolates with increase copy number of pfmdr1 gene found in this study seems higher in comparison to average reported in various studies from African region [An increase in the copy number of fantrine , 47, 49;n region .pfmdr1 copy amplification has been rarely found in the African region [pfmdr1 copy number from Angolan patients; however from other countries, and from a total of 131 isolates collected in 2005\u20132009 from West Africa (n\u00a0=\u00a099) and Central Africa (n\u00a0=\u00a032), only 4 had pfmdr1 copy number amplification. Menard et al. in Cameron in 2009 did not identify samples with pfmdr1 gene amplification, and similar results were reported by Vaughan-Williams et al. in Kwazulo Natal, South Africa, in 2012 [According to other studies the n region , 50. The in 2012 .\u00ae treatment response, the failure rate of 8.8\u00a0% is higher in comparison to that of 1.2\u00a0% found in the 2004 study for AL efficacy in Angola [Relatively to Coartemn Angola , and then Angola .pfmdr1 86 and 1246 positions found in this study is in agreement with those reported in various studies conducted in the African region [86Y allele previously selected by CQ. Detection of the pfmdr1 1246Y in this study as well as in others studies of isolates from West Africa (Guinea Bissau) and DRC  seems to indicate a emergence of this genotype previously considered rare in Africa, may be due to globalization and migration. The frequency of isolates with increase copy number of pfmdr1 gene found in this study seems higher in comparison to others reported in various studies from the African region where copy amplification of this gene has been rarely found. The analysis of the clinical outcomes after treatment with Coartem\u00ae did not link a particular genotype with treatment failure met in this study.The predominance of the wild-type allele for the n region . Recentlpfmdr1 genotyping in comparison to many studies previously realized in the area. However, the low number of patients in the study and with treatment failure could be a limitation for a consistent conclusion.Globally, the result of this study suggests a change of the profile of the parasite population circulating in Angola, reflected by a decreased susceptibility to AL and a relative alteration in"}
+{"text": "Day 7 plasma concentrations of lumefantrine (LF) can serve as a marker to predict malaria treatment outcome in different study populations. Two main cut-off points (175 and 280\u00a0ng/ml) are used to indicate plasma concentrations of LF, below which treatment failure is anticipated. However, there is limited data on the cumulative risk of recurrent parasitaemia (RP) in relation to day 7 LF plasma concentrations in pregnant women. This study describes the prevalence, severity, factors influencing treatment outcome of malaria in pregnancy and day 7 LF plasma concentration therapeutic cut-off points that predicts treatment outcome in pregnant women.Plasmodium falciparum malaria receiving artemether-lumefantrine (ALu) participated in pharmacokinetics and pharmacodynamics study. Blood samples were collected on days 0, 2, 7, 14, 21 and 28 for malaria parasite quantification. LF plasma concentrations were determined on day 7. The primary outcome measure was an adequate clinical and parasitological response (ACPR) after treatment with ALu.This was a one-arm prospective cohort study whereby 89 pregnant women with uncomplicated The prevalence of malaria in pregnant women was 8.1\u00a0% (95\u00a0% CI 6.85\u20139.35) of whom 3.4\u00a0% (95\u00a0% CI 1.49\u20138.51) had severe malaria. The overall PCR-uncorrected treatment failure rate was 11.7\u00a0% (95\u00a0% CI 0.54\u201313.46\u00a0%). Low baseline hemoglobin (<10\u00a0g/dl) and day 7 LF concentration\u00a0<600\u00a0ng/ml were significant predictors of RP. The median day 7 LF concentration was significantly lower in pregnant women with RP (270\u00a0ng/ml) than those with ACPR (705\u00a0ng/ml) (p\u00a0=\u00a00.016). The relative risk of RP was 4.8 folds higher (p\u00a0=\u00a00.034) when cut-off of\u00a0<280\u00a0ng/ml was compared to\u00a0\u2265280\u00a0ng/ml and 7.8-folds higher (p\u00a0=\u00a00.022) when cut-off of\u00a0<600\u00a0ng/ml was compared to\u00a0\u2265600\u00a0ng/ml. The cut-off value of 175\u00a0ng/ml was not associated with the risk of RP (p\u00a0=\u00a00.399).Pregnant women with day 7 LF concentration\u00a0<600\u00a0ng/ml are at high risk of RP than those with\u00a0\u2265600\u00a0ng/ml. To achieve effective therapeutic outcome, higher day 7 venous plasma LF concentration\u00a0\u2265600\u00a0ng/ml is required for pregnant patients than the previously suggested cut-off value of 175 or 280\u00a0ng/ml for non-pregnant adult patients. Plasmodium falciparum infection in pregnancy are most pronounced for women in their first pregnancy whereas in low transmission settings malaria affects all pregnant women, regardless of the number of times they have been pregnant [Over 93\u00a0% of the Tanzania mainland population lives in areas where malaria is endemic. In Tanzania, there is great variation in the risk of malaria transmission and prevalence ranging from 1\u201333\u00a0%, with an average of about 10\u00a0% . Despitepregnant . It is epregnant .P. falciparum malaria in the second and third trimester of pregnancy [P. falciparum parasites and prevents recrudescence, thereby ensuring malaria cure [Artemisinin-based combination therapy is recommended by WHO as first-line treatment for uncomplicated regnancy . The fixregnancy . Artemetregnancy . Lumefanria cure .Plasma LF concentrations at day 7 reflects the degree to which the residual parasites are exposed, and is generally considered as a useful pharmacokinetic marker to predict malaria treatment outcome , 11. DifMalaria during pregnancy is associated with high maternal and perinatal mortality . PregnanIt is well recognized that plasma LF concentration on day 7 LF levels is a surrogate marker to predict treatment outcome, but the therapeutic cut of point may vary between different patient populations and transmission areas (low versus high). The therapeutic day 7 LF concentration threshold below which RP is anticipated needs to be defined better, particularly for high risk group living in malaria endemic countries , includiP. falciparum is the predominant species [This was a one-arm prospective cohort study that included all pregnant women who gave consent to be screened for malaria when attending antenatal clinics  at Kisar species .The study received ethics approval from the institutional review board of Muhimbili University of Health and Allied Sciences (MUHAS). Participants were informed about the aim of the study and gave written consent before participating in the study. Confidentiality was ensured to all individuals who participated in the study, whereby, all collected samples and the filled confidential report forms (CRF) have a coded identification number.The study aimed to obtain a sample size of 50, which is the minimum recommended by the WHO regardless of rates of failure anticipated, in order to be representative . This isP. falciparum infection and haemoglobin level of\u00a0>8\u00a0g/dl. Full medical history, including current illnesses and medication used, were recorded. Clinical examination on the day of enrollment and during follow-up visits on days 2, 7, 14 and 28 were done including axillary temperature measurement and malaria-related symptoms evaluation.Between May 2014 to April 2015, 1835 pregnant women attending the antenatal cares (ANCs) were screened by using malaria rapid diagnostic test (MRDT). Pregnant women with MRDT positive results were enrolled in the study. Inclusion criteria included women aged 18\u00a0years and above, resident of Mkuranga or Kisarawe districts, in the second or third trimester, with uncomplicated \u00ae MRDT . The blood samples were collected by finger prick. About 5\u00a0\u03bcl of whole blood were added into the \u2018sample well\u2019 of respective test devices using a micropipette supplied with the test kit. Four drops of assay diluent were added into the \u2018sample diluent well\u2019. All the test results were recorded within 30\u00a0min. The sample was positive when there was appearance of a control line and a test line on the result window but it was negative when there was only a control line.Malaria was tested by using SD BIOLINE Malaria Ag P.f/PanP. falciparum within 28\u00a0days of follow-up were treated with either artesunate or artemether or quinine injection as per the national malaria treatment guidelines [Enrolled study participants received four tablets of ALu  (20\u00a0mg artemether and 120\u00a0mg lumefantrine) over the course of 3\u00a0days at 0, 8, 24, 36, 48, and 60\u00a0h. In order to enhance absorption of ALu and standardize dose administration, patients were supplied with packets of milk containing 4.5\u00a0g of fat  and were\u00ae machine  following manufacturers\u2019 instructions.To estimate the parasite density, capillary blood from a finger prick was taken at days 0, 2, 7, 14, 21 and 28. Samples were collected on slides, Giemsa-stained thick smears were examined by two different experienced microscopists using light microscope. Plasmodium parasites were counted against 200 white blood cells (WBC) on the thick film. Five hundred WBCs were counted where less than ten parasites were observed. The parasite count was then multiplied by a factor of 40 or 16 depending on the counted WBCs. Haemoglobin was measured by HemoCue Hb 201\u00a0+\u00a0g for 5\u00a0min, and the plasma samples were stored in cryotubes. Samples were stored at \u221280\u00a0\u00b0C at MUHAS before analysis. Plasma LF concentration was analysed using a validated high performance liquid chromatography (HPLC) with ultraviolet detection at Sida/MUHAS bioanalytical laboratory in Dar-es-Salaam, Tanzania as described previously [In total, three millilitres of venous blood were drawn from the patients at random times on days 0 and 7 to determine plasma LF concentrations. Day 0 blood sample was collected before starting the medication as a baseline in order to determine the presence of LF in the patient\u2019s plasma prior to treatment , 35. Bloeviously . The coeTreatment outcome classification was based on WHO recommendations on the methods for surveillance of anti-malarial drug efficacy as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), and Adequate clinical and parasitological response (ACPR) . An ACPRt test . The cumulative risk of RP was estimated using the Kaplan\u2013Meier product limit formula and data were censored. A P value\u00a0<0.05 was considered statistically significant.The data were analysed using the statistical package for social sciences  software, version 20. The intention-to-treat approach was used to analyse the anti-malarial treatment response. All day 7 LF plasma concentrations were first transformed into log 10 values before applying parametric tests. The relation of the outcome variable (treatment outcome) and explanatory variables were tested using independent Baseline characteristics of all enrolled patients are presented in Table\u00a0P. falciparum infection. Severe malaria was prominent in primigravidae (four women) than multigravid pregnant women (one woman). MRDT positive samples were further analysed using microscope whereby, 111 out of 148 (75\u00a0%) samples were confirmed to have P. falciparum infection.One thousand eight hundred and thirty five pregnant women were screened using MRDT during the study period and out of these 148 were positive for malaria. Prevalence of malaria in pregnancy during the study period was 8.1\u00a0% (95\u00a0% CI 6.85\u20139.35). Out of these malaria positive pregnant women, 5  had severe malaria and the rest (143 pregnant women) had uncomplicated P. falciparum malaria consented and were enrolled in this study. Flowchart for study enrolment is presented in Fig.\u00a0Eighty-nine patients with uncomplicated There were a total of seven  therapeutic failures in this study among pregnant women. No ETF or LCF was observed in this study; nonetheless, seven pregnant women had LPF. Hence, the overall ACPR after 28\u00a0days of follow up was 88.3\u00a0% (95\u00a0% CI 40.37\u201365.63\u00a0%).Day 7 LF concentration was significantly lower among pregnant women with RP than those with ACPR (p\u00a0=\u00a00.016). The median plasma LF concentration among pregnant women with ACPR was 705\u00a0ng/ml (140\u20133059\u00a0ng/ml), whereas for women with LPF, it was 270\u00a0ng/ml (123.6\u2013602\u00a0ng/ml)  for pregnant women. There was marked inter-individual variability in day 7 plasma LF concentrations . There was no association between day 7 LF plasma concentrations with age (p\u00a0=\u00a00.784), gravida (p\u00a0=\u00a00.314), trimester (p\u00a0=\u00a00.496) or baseline parasitaemia (p\u00a0=\u00a00.644). Amongst of the studied population 6.7\u00a0% of the patients had day 7 concentrations below the therapeutic cut-off of 175\u00a0ng/ml, 21.7\u00a0% below 280\u00a0ng/ml, and 43.3\u00a0% below 600\u00a0ng/ml. More than half of the pregnant women (57\u00a0%) had day 7 concentration of\u00a0>600\u00a0ng/ml which is required for maximal efficacy. Equally, 15\u00a0% had 175\u2013280\u00a0ng/ml, and 21.7\u00a0% had 281\u2013600\u00a0ng/ml . The relative risk of RP was 2.3 (when comparing LF concentrations\u00a0<175 to\u00a0>175\u00a0ng/ml), 4.8 (when comparing LF concentrations\u00a0<280 to\u00a0>280\u00a0ng/ml) and 7.8-folds higher when comparing LF concentrations\u00a0<\u00a0600 to\u00a0>600\u00a0ng/ml. The relative risk of RP in patients at the cut-off point LF\u00a0<175\u00a0ng/ml was not significantly higher than in those with LF concentration of\u00a0>175\u00a0ng/ml (p\u00a0=\u00a00.399). On the other hand, relative risk of RP was statistically significant using the LF cut-off point of\u00a0<280\u00a0ng/ml compared to\u00a0>280\u00a0ng/ml (p\u00a0=\u00a00.034) and\u00a0<600\u00a0ng/ml compared to\u00a0>600\u00a0ng/ml (p\u00a0=\u00a00.022) . Using the cut-off value of 600\u00a0ng/ml, the cumulative risk of RP was increasing in patients with\u00a0<600\u00a0ng/ml from day 7 onwards, whereas, there was a slight increase on the cumulative risk of RP on day 14 in patients with LF concentration\u00a0>600\u00a0ng/ml (log-rank p\u00a0<\u00a00.017)  pregnant women were MRDT positive of whom 111 (6.0\u00a0%) had parasitologically-proven s 12.2\u00a0% . The pres 12.2\u00a0%  and Souts 12.2\u00a0% ), but los 12.2\u00a0% , 42 and s 12.2\u00a0% ). Markeds 12.2\u00a0% . BaselinThe study is well controlled, by excluding data from study participants in whom plasma LF concentration was detected at baseline (day 0). Residual anti-malarial concentrations before treatment in patients with malaria may interfere with outcome of the treatment under investigation and creates bias in drug safety and efficacy assessment , 35. A pPharmacokinetics of drugs in pregnancy is altered by several factors, including physiological changes that lower drug absorption, speed up drug clearance, and increase body fluid volume of distribution , 29, 45.As many as 43.3\u00a0% pregnant women did have day 7 LF concentrations of\u00a0<600\u00a0ng/ml which is slightly higher than 31\u00a0% which was previously reported in a study conducted by Mosha et al. in Rufiji, Coast region in Tanzania . MoreoveThe cure rate in this study was 88.3\u00a0%, which is similar to previous report from Rufiji, Tanzania . The medThe relative risk of RP increased proportionately when day 7 LF concentration cut-off values increased from\u00a0<175 to\u00a0<600\u00a0ng/ml. The host immune responses are reduced during pregnancy and this potentially means that a different LF day 7 concentration threshold is required in this special population . The relIn areas of high or moderate malaria transmission, response to malaria treatment mainly depends on the host\u2019s immunity, genetic and the amount of drugs available in human plasma to clear the parasites , 50. TheOther baseline characteristics, such as age of the patient, gravida, trimester and baseline parasitaemia were not important factors determining the day 7 LF concentration and subsequent therapeutic response in study participants. This is contrary to what has been reported in a previous study involving pregnant and non-pregnant patients in which patients with higher baseline parasitaemia were more likely to fail treatment .The findings of this study support that day 7 LF concentrations can be used as a reliable predictor of treatment outcome of malaria in pregnant women. The relative and cumulative risk of RP is higher when using the cut-off point of\u00a0<600\u00a0ng/ml as compared to the previous cut-off points of\u00a0<175 and\u00a0<280\u00a0ng/ml indicating that even pregnant women with day 7 LF concentrations\u00a0>280\u00a0ng/ml but less than 600\u00a0ng/ml are still at high risk of RP. In order to achieve effective therapeutic outcome, higher day 7 venous plasma LF concentration\u00a0\u2265600\u00a0ng/ml is required in pregnant patients for maximal efficacy than a cut-off value of 175 or 280\u00a0ng/ml, which has been suggested for non-pregnant adult patients. More studies in different areas should be conducted because malaria treatment outcome after ALu administration can be influenced by a number of other factors besides day 7 LF concentration."}
+{"text": "Current day malaria cases and deaths are indicative of a lack of access to both methods of prevention, diagnosis, and treatment; an important determinant of treatment efficacy is adherence. This study is a follow up to the baseline study of adherence to artemether-lumefantrine (AL) carried out in Garissa District in 2010. The study presented evaluates any changes in adherence levels which may have occurred in the area during this period and after nearly three years of sustained use of ACT across the public health sector.The study was carried out in Garissa County in the North Eastern Province of Kenya and included patients fitting the suspected malaria case definition and having been prescribed AL, regardless of confirmatory diagnosis. A questionnaire assessed the intake of AL via both self-reporting by the participant and observation of blister packs by the interviewer. On separate occasions exit interviews with patients and observations of prescribers were also carried out.Of the 218 participants enrolled, 195 were successfully followed up. 60% of participants were found to be adherent to the three-day AL regimen, this is 4.7% lower than the proportion of participants adherent in 2010; the result of a two-sided z-test was not significant (p\u2009=\u20090.23). The odds of the patient being adherent to AL increased by 65% with each additional correct statement regarding how to take AL that a patient could recall (between zero and four statements), this was the only variable significantly associated with patient adherence (p\u2009=\u20090.01).Sustaining the ACT adherence rates at the 2010 levels, through 2.5\u00a0years of insecurity in the study area is an achievement and suggests that if security can be improved barriers to improving health service quality and patient adherence to AL would be removed. This study, by looking specifically at anti-malarial adherence over a prolonged period and in a setting of severe conflict, provides a valuable and rare insight in to the challenges and barriers to ACT adherence in such settings. Plasmodium falciparum [. It was as part of the search for new and safer anti-malarials that artemisinin-based drugs emerged from China, becoming widely available outside of China for the first time in the 1990s. With the hope of preserving the efficacy of this anti-malarial it is recommended only in combined use with other anti-malarial, as artemisinin combination therapy (ACT) [There were 198 million range 124\u2013283) cases of malaria and 584,000  deaths from malaria in 2013. These large figures are a considerable decrease on the 2000 levels since when malaria cases have dropped by 26% worldwide, and the number of deaths due to malaria has fallen by 47% worldwide [4\u2013283 casACT has a more complicated dosing regimen than monotherapies, in the case of artemether-lumefantrine (AL) this ranges from one to four tablets (determined by a patient\u2019s weight) taken twice a day for three day. The efficacy of ACT is very high, however, this is assuming a patient is adherent to the complete course of treatment -7. The dP. falciparum accounts for almost 100% of malaria infections [Anopheles gambiae, Anopheles arabiensis, and Anopheles funestus [. Extrapolating on the 2012 Kenyan population estimate of 43.18 million with the yearly growth rate of 2.7% [In Kenya, funestus . Extrapo of 2.7% , the est of 2.7% . In 2013 of 2.7% .The baseline study carried out by The MENTOR Initiative in 2010 in Garissa County, North Eastern Province, Kenya, found that of a sample of 272 patients, 176 (64.7%) were \u201cprobably adherent\u201d, defined as patients claiming to have completed all doses whether or not they were able to show the empty blister packet. The strongest predictor of adherence was found to be patient knowledge of the ACT regimen, if a patient was able to cite at least one correct instruction as to how to take AL they had 1.76 , the odds of being fully adherent compared to a respondent that could not cite any instructions [Since the baseline study in 2010 The MENTOR Initiative has supported the Ministry of Health (MoH) to reinforce malaria case management in Garissa, through the introduction of rapid diagnostic testing (RDT) for patients with suspected malaria, prior to treatment. Health workers received regular technical coaching to encourage them to explain the AL treatment regime to patients and there was a comprehensive package of malaria prevention activities. In addition, and in line with the 2011 study recommendations , InformaThe purpose of this study is to measure current day patient adherence to the three day AL treatment regimen, and determine any changes in adherence since the 2010 study. Whilst the 2010 study was conducted in the areas of Bunyala, in Western Kenya, and Garissa, this 2013 study included only the area of Garissa; for this reason all comparative analysis between the two studies shall only take in to account the 2010 data collected in Garissa.2). It borders Wajir County to the North, Tana River County to the West, Lamu County to the South, and Isiolo to the North West and shares an international boundary with Somalia to the East  or history of fever in the last 48\u00a0hours with other obvious causes of fever excluded , patSix health facilities in Garissa were involved in the study, three of which had participated in 2010. Health workers at the health facilities informed their patients to keep the empty AL blister packs after treatment had been completed. Patients were not told why they were being asked to keep the blister packs in order to maintain the validity of the results. At the end of each day, the registers of the health facilities were consulted to identify patients who met inclusion criteria, these patients or their caretakers were visited at home by the study team and community health workers on the fourth day after receiving treatment.At the beginning of the household visit, written consent was requested from each adult patient, and the parent or guardian of every child patient included in the study. The patient\u2019s axillary temperature was taken. A structured questionnaire \u2013 available in English and translated into Somali and field-tested \u2013 was given to the participants. The questionnaire assessed the intake of AL via both self-reporting by the participant and observation of blister packs by the interviewer. Socio-demographic information on the patient was collected and the patient or caretaker\u2019s language, age and education level, the number of people who spent the previous night at the house, and the number of children cared for were all recorded.The questionnaire was concluded by counting any remaining tablets. In cases where tablets remained, patients/caretakers were asked to specify why not all tablets were taken by the patient/ given to the child. Questions were asked in such a way that honest answers were encouraged, without placing blame on interviewees. The team of interviewers were supervised by MENTOR Initiative clinical staff.Patients were categorized as definitely non-adherent; probably non-adherent; and probably adherent based on the following classifications. Those who presented blister packs with AL doses still remaining were definitely non-adherent; patients or caregivers who claimed doses not completed but who did not show a blister pack were classified as probably non-adherent; those who claimed to have completed all doses and could show an empty blister pack were classified as probably adherent as were patients who could not show an empty blister pack but claimed to have taken all of the tablets. Non-adherent patients were advised to return to the health clinic for a new assessment to avoid negative consequences from an incomplete course of treatment.In order to identify possible factors influencing adherence, two additional investigations were carried out. The week after the completion of collection of adherence data, patients or their caretakers were systematically interviewed after leaving the pharmacy (out of sight of the pharmacy in order to reduce bias) to assess how well they had understood the AL prescription. A short questionnaire, asking for a spontaneous account of how the treatment was to be taken/ given to the child, was carried out followed by systematic questioning for each treatment dose.e.g. timing of drug intake, importance of completing the treatment. The observers stood next to the patient during the explanation so the pharmacist/person dispensing the medication knew a study was being conducted, but he/she was blinded as to the purpose of the observation.On separate occasions the pharmacists or other persons dispensing medication in each health facility were systematically observed while explaining the AL prescription to patients or caretakers. A check-list was used to verify what information was given: Open-ended questions were reviewed and grouped into thematic categories that were coded for data entry. Results were double entered into Epi Info 3.5.1  and all data were analysed in STATA 13.0 . Data were summarized using proportions and means and medians, where appropriate. Comparisons of proportions between categorical variables were made using two-tailed Z-tests. Univariate and multivariate logistic regression was used to test for significance between potential explanatory variables and the dichotomous outcome variable of the patient being adherent or not. Significance tests were determined at the 10% level.The study was not an intervention study and was not designed to influence the diagnosis and treatment of the participants. Permission to carry out the study was gained from the University of Nairobi, The Ministry of Health, Kenya, and Kenya National Hospital ethics review committee.At the beginning of the visit written consent was requested from every adult and parent or guardian of every child considered for inclusion in the study.A total of 218 respondents were enrolled in the study. They came from six health facilities from within Garissa district . Of these enrolled participants 23 (10.5%) were either lost to follow-up (n\u2009=\u200919) or didn\u2019t respond to later questioning (n\u2009=\u20094), bringing the total number of participants successfully followed-up down to 195. The majority of participants were male (81.0%) and over 15\u00a0years of age (99.0%), half of participants owned their own businesses (53.8%), when asked about education 32.3% reported never having attended a school or college, just over half of all households contained 0\u20135 people (55.4%) with the remaining containing six or more people. Characteristics of the study population are shown in Table\u00a0Of the 195 participants successfully followed up, 60.0% were found to be probably adherent to the three day ACT regimen. Of the 78 participants classified as non-adherent, 31 (39.7%) were definitely non-adherent and 47 (60.3%) were probably non-adherent. This proportion of patients found to be probably adherent in 2013 is 4.7% lower than the proportion of participants probably adherent in 2010. The result of a two-sided z-test between the proportion adherent in 2010 and 2013 was not significant at the p\u2009\u2264\u20090.05 level  and 112 males (44.2%), in 2013 there were only 37 females (19.0%) and 158 males (81.0%). In 2013 the proportion adherent differed slightly between male and female, 59.5% probably adherent and 62.2% probably adherent, respectively. The 2010 study population also contained many more participants under the age of 15\u00a0years. In 2013 only two study participants (1%) were children under 15\u00a0years of age compared to the 136 participants (50%) in the 2010 study population in Garissa. Of the 2013 study population the majority tested RDT positive for malaria (80.5%) with 37 participants testing negative (18.9%), one participant did not know their RDT result (0.5%).Univariate logistic regression compared 17 variables thought to be potential predictors of patient adherence. Of these 17 variables, 11 were not fitted in to a multivariate logistic regression model as their crude (bivariate) odds ratios were not significant (p\u2009\u2265\u20090.10); one variable- whether or not the patient had difficulty in understanding the prescription instructions- was excluded as collinearity with adherence was shown. The remaining five variables with p-values \u22640.10 were fitted in to the multivariate logistic regression model Table\u00a0. The onlThe follow up questionnaire/survey of patients four days after their visit to the health facility and their being prescribed ACT collected information on their knowledge and perceptions regarding malaria and its treatment. Table\u00a0Data collected via both observations of prescriptions and exit interviews Table\u00a0 showed tThe ACT adherence study carried out in 2010 in Garissa and Bunyala was the first of its kind in Kenya and was important in informing the Ministry of Health nationally, and regionally, on the success of the introduction of ACT since 2006, and the factors associated with either high or low levels of adherence . The folSimilar studies in to the adherence of patients to different treatment regimens for uncomplicated malaria show varying results. Using self-reporting, pill counts, and blood drug levels to investigate the adherence of patients to a six dose regimen of AL in Uganda found 90% of patients to be probably adherent . In the The 4.7% decrease in proportion adherent in Garissa from the 2010 level of 64.7% is not statistically significant and so should be interpreted as no significant change in adherence levels between the years, rather than a true decrease. This result is disappointing, however, the lack of apparent improvement does not mean the data is of no interest, nor does it reflect the improvements seen in the uptake and usage of RDTs and ACT by health workers across the region . PossiblDifferences in the study populations between 2010 and 2013 in Garissa may have impacted the adherence levels seen. The sample size of patients in 2013 (n\u2009=\u2009195) was much smaller than the sample size in the 2010 study (n\u2009=\u2009918) . This drThe methods used to measure adherence and the categorisation of patients as adherent or not differ between adherence studies. The majority of methods can be summarised with five overarching approaches; \u201cCompleted treatment\u201d identifying individuals who say they completed treatment, \u201cverified completed treatment\u201d corroboration of reported completed treatment with a pill count, \u201ctimely completion\u201d refers to patients reporting timely completion, \u201cverified timely completion\u201d refers to those reporting timely completion and with a pill count to confirm no tablet left. The last method is \u201cbiological assay\u201d which uses the detection of sufficient levels of the drug in biological samples to confirm adherence . The metWhile policy is to give ACT for free at health facilities, the study has shown that this is often not the case. 37 patients (19%) reported paying for their treatment; the largest proportion of these paid 100 Kenyan Shillings (KSH) , equivalent to one United States Dollar (USD), five people paid less, and nine paid more. The highest price reported was 800 KSH. Of those reportedly paying for treatment, 83.8% (n\u2009=\u200931) received treatment from a clinical pharmacist, 13.5% (n\u2009=\u20095) received treatment from a doctor, or nurse, and 2.7% (n\u2009=\u20091) received treatment from a private pharmacy. The proportion of Kenyans living below the international poverty line of 1.25 USD per day (roughly 110 KSH) is 43.4% . Having Interventions aiming to improve patient adherence to a treatment regime have been numerous and varied. From improving adherence to short-term treatments, such as anti-malarials, to treatments for chronic conditions, such as hypertension, various methods have been investigated including changing dosing schedules , reminder pill packaging, individual counselling, home visits by research assistants, and reward schemes for high adherence ,37. A stIn the study area of Garissa the ethnic Somali population is a marginalized one anecdotally sceptical of outside interventions and IEC campaigns, making the challenge of real behaviour change even greater. The number of correct statements reported by the patient was the most significant variable in predicting adherence in this survey and the 2010 survey , this reLocal contextual factors are also likely to have had an effect on adherence levels. Security in Garissa has historically been fragile. From 2010 onwards conflict inside Somalia intensified resulting in significant influxes of new refugees into the camps in Garissa Town. In October 2011, Kenya sent 10,000 troops into southern Somalia, largely through Garissa, to attack and clear Al Shabaab terrorist groups that controlled areas neighbouring Garissa ,41. The This paper has laid out the findings of an ACT adherence study in Garissa County, Kenya, nearly three years after a base-line study and efforts to improve ACT prescription practices and adherence. Results showed no significant change in adherence levels over the period. The ability of patients to recall prescription practise statements was significantly associated with adherence, a finding important in informing future interventions aimed at improving adherence. Changes in the study populations, successes and failures of prescribers to provide accurate and free health care, and the challenging security context of the area are all possible explanations for the lack of an increase in adherence levels. Given all of these factors, sustaining the ACT adherence rates at the 2010 levels, through three years of insecurity is an achievement and suggests that if security can be improved barriers to improving health service quality and patient adherence to ACT could be removed."}
+{"text": "Plasmodium falciparum extends across Southeast Asia where it is associated with worsening partner drug resistance and a decline in the efficacy of frontline artemisinin-based combination therapy. Dihydroartemisinin-piperaquine (DP) is an essential component of preventive and curative treatment in the region, but its therapeutic efficacy has fallen in Cambodia.Artemisinin resistance in A prospective clinical and parasitological evaluation of DP was conducted at two sites in Upper Myanmar between August 2013 and December 2014, enrolling 116 patients with acute uncomplicated falciparum malaria. Patients received DP orally for 3\u00a0days together with primaquine 0.25\u00a0mg/kg on admission. Parasite clearance half-lives based on 6 hourly blood smears, and day 42 therapeutic responses were assessed as well as parasite K13 genotypes.k13 gene. The k13 F446I mutation was found in 25.4\u00a0% of infections and was associated with a median clearance half-life of 4.7\u00a0h compared with 2.7\u00a0h for infections without k13 mutations (p\u00a0<\u00a00.001). There were no failures after 42\u00a0days of follow-up, although 18\u00a0% of patients had persistent parasitaemia on day 3.Median parasite clearance half-life was prolonged, and clearance half-life was greater than 5\u00a0h in 21\u00a0% of patients. Delayed parasite clearance was significantly associated with mutations in the propeller region of the parasite k13 mutation observed in Upper Myanmar, F446I, appears to be associated with an intermediate rate of parasite clearance compared to other common mutations described elsewhere in the Greater Mekong Subregion. Discerning this phenotype requires relatively detailed clearance measurements, highlighting the importance of methodology in assessing artemisinin resistance.The dominant The online version of this article (doi:10.1186/s12936-016-1240-7) contains supplementary material, which is available to authorized users. Plasmodium falciparum is now prevalent across much of mainland Southeast Asia   remained parasitaemic 3\u00a0days after treatment [30.2\u00a0% of patients in Myitkyina and 9.9\u00a0% in Thabeikkyin (p\u00a0=\u00a00.01)]. Fever resolution was rapid   until 42\u00a0days  mutations was 28.9\u00a0% (33/114) (Table\u00a0k13 mutations compared with 12 of 71 (16.9\u00a0%) in Thabeikkyin. All k13 mutations observed had been reported in previous studies. No mutations were found in the non-propeller region of k13 (amino acids 210\u2013440).The k13 and 4.7\u00a0h for F446I mutation (p\u00a0<\u00a00.001)  compared to those with wild type alleles  (p\u00a0=\u00a00.02).Median parasite clearance half-life was 2.7\u00a0h for wild type k13 propeller mutation had an odds ratio of 5.0  while recruitment in Myitkyina had an odds ratio of 4.0 . Using a cutoff of 5\u00a0h, these ratios were 2.3  and 3.5 , respectively  Table\u00a0; in thisk13 propeller mutations at the time of enrollment (p\u00a0=\u00a00.42) or after treatment (p\u00a0=\u00a01.0), suggesting that artemisinin resistance had no effect on the proportion of patients with gametocytes at or after admission.Ten patients (8.8\u00a0%) had patent gametocytaemia at the time of enrollment, with nine of these recruited in Myitkyina Table\u00a0. The geoThe median (IQR) durations of gametocyte carriage in Myitkyina and Thabeikkyin were 57 (6\u201381) and 16.5 (12\u201324) h respectively. Overall gametocyte carriage rate was 26 PGW per 1000\u00a0weeks of follow up. There was no association between gametocyte carriage duration and geographical location (p\u00a0=\u00a00.31), or parasite clearance half-life (p\u00a0=\u00a00.25). All patients with gametocytaemia cleared gametocytes within 96\u00a0h from the time gametocytes were first seen.One third of participants 35\u00a0%) were anaemic on enrolment. The nadir in mean haematocrit occurred on day 3, after which haematocrit values gradually increased  received by patients for dihydroartemisinin, piperaquine and primaquine were 2.2 (2.1\u20132.3), 17.5 (16.6\u201318.5) and 0.26 (0.24\u20130.28) mg/kg respectively. Study medications were generally well tolerated although six patients vomited the day 0 medication, and one vomited on day 2; all were retreated according to protocol. One serious adverse event was reported at the Myitkyina site; one patient who had already missed follow-up visits died in a road traffic accident approximately 2\u00a0weeks after the last visit.P. falciparum  where primaquine was not used [52 h] whek13 mutation observed, k13 F446I, was associated with a clearance half-life of slightly under 5\u00a0h, suggesting an \u201cintermediate resistance\u201d phenotype. The study also emphasizes the clearance half-life remains the gold-standard in research studies of artemisinin resistance and shows the value of frequent in vivo sampling of parasitaemia in locations where susceptibility to artemisinins is previously unknown [In summary, this study shows that while treatment efficacy remains high with DHA-piperaquine, artemisinin resistance has emerged and is established in Upper Myanmar. However, the main  unknown . The sam unknown , but per unknown ."}
+{"text": "Controlled human malaria infection (CHMI) studies which recapitulate mosquito-borne infection are a critical tool to identify protective vaccine and drug candidates for advancement to field trials. In partnership with the Walter Reed Army Institute of Research, the CHMI model was established at the Seattle Biomedical Research Institute's Malaria Clinical Trials Center (MCTC). Activities and reagents at both centers were aligned to ensure comparability and continued safety of the model. To demonstrate successful implementation, CHMI was performed in six healthy malaria-na\u00efve volunteers.Plasmodium falciparum by the bite of five infected Anopheles stephensi mosquitoes under controlled conditions and were monitored for signs and symptoms of malaria and for parasitemia by peripheral blood smear. Subjects were treated upon diagnosis with chloroquine by directly observed therapy. Immunological (T cell and antibody) and molecular diagnostic  assessments were also performed.All volunteers received NF54 strain All six volunteers developed patent parasitemia and clinical malaria. No serious adverse events occurred during the study period or for six months post-infection. The mean prepatent period was 11.2 days (range 9\u201314 days), and geometric mean parasitemia upon diagnosis was 10.8 parasites/\u00b5L (range 2\u201369) by microscopy. qRT-PCR detected parasites an average of 3.7 days (range 2\u20134 days) earlier than blood smears. All volunteers developed antibodies to the blood-stage antigen merozoite surface protein 1 (MSP-1), which persisted up to six months. Humoral and cellular responses to pre-erythrocytic antigens circumsporozoite protein (CSP) and liver-stage antigen 1 (LSA-1) were limited.The CHMI model was safe, well tolerated and characterized by consistent prepatent periods, pre-symptomatic diagnosis in 3/6 subjects and adverse event profiles as reported at established centers. The MCTC can now evaluate candidates in the increasingly diverse vaccine and drug pipeline using the CHMI model.NCT01058226ClinicalTrials.gov  Plasmodium sporozoites under controlled conditions. To date, CHMI has mostly been performed using Plasmodium falciparum-infected mosquitoes Plasmodium vivax studies have recently taken place as well In the absence of defined immune correlates of protection and consistently predictive animal models, controlled human malaria infections (CHMI) have become the most effective means of assessing early-stage efficacy of candidate pre-erythrocytic and erythrocytic vaccines and anti-malarial drugs. Under this model, malaria parasite-infected mosquitoes are allowed to bite human volunteers to inoculate them with P. falciparumCHMI studies have been conducted in the United States of America (USA) and elsewhere for decades. Reviews of the published literature indicate the model is safe, reproducible and well-tolerated by subjects participating in clinical challenge trials with Despite an expanding pipeline of candidate malaria interventions in development, the number of clinical research centers capable of conducting CHMI is limited The primary objective of the study was to demonstrate the safety, tolerability and infectivity of CHMI in a newly-established facility. The secondary objectives were to assess the malaria-specific immune response following CHMI in healthy malaria-na\u00efve adults. Additionally, we sought to evaluate the detection and quantification of prepatent parasitemia by real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR).The study was conducted at the MCTC at Seattle BioMed in Seattle, WA. The institute also houses the Center for Mosquito Production and Malaria Infection Research (CeMPMIR), an entomology facility that was upgraded for the production of malaria sporozoites and malaria-infected mosquitoes under phase-appropriate current Good Manufacturing Practices (cGMPs) to support MCTC trials. The Clinical and Translational Research Laboratory (CTRL) was simultaneously established to support specimen processing and to perform standardized assays according to Good Clinical Laboratory Practices (GCLPs) for clinical trials.Seattle BioMed and WRAIR established a Cooperative Research and Development Agreement to institute the CHMI model at the MCTC. WRAIR research and clinical staff from the US Military Malaria Vaccine Program served in an advisory capacity throughout preparation and conduct of the trial to ensure alignment with WRAIR practices and participant safety.P. falciparum sporozoites by the bites of five infected Anopheles stephensi mosquitoes under controlled conditions. Volunteers were closely monitored in the post-challenge period and treated with standard oral doses of chloroquine phosphate upon diagnosis of malaria parasitemia by positive thick blood films. Periodic clinical assessments, physical examinations and laboratory monitoring were performed for evaluation of protocol-defined safety, infectivity and immunology endpoints.This was a prospective, open-label, single intervention study of six healthy malaria-na\u00efve adult volunteers undergoing CHMI. All subjects were enrolled as a single cohort and followed the same study schedule Figure 1http://clinicaltrials.gov/show/NCT01058226). The protocol for this trial and supporting CONSORT checklist are available as supporting information; see Checklist S1 and Protocol S1.This study was conducted in accordance with the International Conference of Harmonization (ICH) Good Clinical Practices (GCP) and applicable Food and Drug Administration (FDA) regulations. Based on evolving regulatory standards for human challenge studies, the P. falciparum sporozoite was considered an investigational product and for the first time, the CHMI procedures and study protocol were conducted under an investigational new drug application (IND) filed with the FDA. Prior to study initiation, the protocol and study-related documents were approved by the Western Institutional Review Board. The study was registered on ClinicalTrials.gov .Subjects were recruited using IRB-approved messaging from the greater Seattle area. Subjects provided written acknowledgment of informed consent and were required to pass an assessment of understanding quiz covering key study concepts and risks related to participation. Adult males and non-pregnant females aged 18\u201350 years were eligible to participate in the study if they were in good general health as demonstrated by detailed medical history review, physical examination and laboratory assessment performed within 56 days of enrollment. Subjects were required to have a low risk of coronary heart disease based on the NHANES 1 screening criteria P. falciparum-infected A. stephensi mosquitoes for use in CHMI were reared in compliance with phase-appropriate cGMPs in the secure CeMPMIR entomology facility according to standard procedures adapted from WRAIR. The A. stephensi mosquitoes were a laboratory-reared strain originally received from WRAIR. Mosquitoes were infected with P. falciparum by standard membrane feeding methods on parasite cultures derived from the WRAIR master cell bank (MCB) containing a sufficient proportion of mature gametocytes. Prior to the CHMI, sensitivity testing of the parasite MCB to the anti-malarial drugs chloroquine, doxycycline, atovaquone and quinine was performed. The results confirmed the sensitivity of the NF-54 parasite MCB to these drugs at published values and were virtually identical to the sensitive type strain, P. falciparum 3D7, a clone of NF-54. On the day of challenge, five infected mosquitoes were placed in a screen-covered carton and allowed to feed on the subject's forearm for five minutes. Following the feed, mosquitoes were evaluated to confirm both an adequate blood meal  and the presence of adequate numbers of sporozoites in their salivary glands . The sporozoite load was rated microscopically according to a semi-quantitative scale: 0 (no sporozoites observed), +1 (1\u201310), +2 (11\u2013100), +3 (101\u20131000) and +4 (>1000) Wild-type NF54 strain Subjects were closely monitored for 30\u201345 minutes post-challenge for acute reactogenicity and were issued a symptom diary and thermometer to record symptoms and oral temperatures daily for five days. Subjects were evaluated in clinic on Day 1 post-challenge, contacted by phone daily on Days 2-4 and evaluated again in clinic daily on Days 5\u20138 post-challenge. From Day 9 post-challenge, subjects were housed in a local hotel with study staff for close observation during the time they were expected to develop patent parasitemia and symptoms of clinical malaria infection. Clinical assessments conducted at each visit included symptom review, vital signs and physical examination. Peripheral blood smears were examined for detection of malaria parasites at least daily beginning on Day 5 post-infection and continuing through diagnosis and treatment. Blood samples for safety endpoint assays were collected prior to challenge, on the day of the first positive blood smear and on Days 35 and 56 post-challenge. Blood samples for immunology endpoint assays were collected at baseline, on the day of the first positive blood smear, on Days 1, 5, 35 and 56 post-challenge.Malaria treatment decisions were based on diagnosis of patent (microscopic) parasitemia by a positive peripheral blood smear. Subjects were treated upon detection of patent parasitemia with a standard oral regimen of oral chloroquine phosphate  under direct observation. Subjects continued their overnight hotel stays until completion of treatment and confirmation of three consecutive daily negative blood smears, after which they were followed weekly for eight weeks. Subjects were invited to return for optional long-term immunology assessments at three and six months post-infection.Protocol S1). AEs were coded using the Medical Dictionary for Regulatory Activities (MedDRA) and reported using System Organ Class and Preferred Term.Primary safety endpoints were assessed by clinical and laboratory evaluations following the challenge and clinically significant changes from baseline status were reported as adverse events (AEs). Solicited AEs were captured on a diary card and/or by direct questioning during the 28 days post-challenge. A solicited AE was defined as a predetermined event that may reflect safety concerns related to the investigational product or events that could be reasonably expected to occur as part of the intervention. These included local and systemic signs and symptoms related to the challenge and/or clinical malaria. Table 1.Development of patent parasitemia was monitored by microscopic evaluation of peripheral blood smears. All subjects undergoing CHMI were expected to develop patent parasitemia. Blood smears were assessed by microscopy daily from Day 5 post-infection through diagnosis and after treatment until three consecutive daily blood smears were negative. Additional blood smears were prepared at other times as needed for diagnosis in symptomatic subjects. The prepatent period was defined as the time from CHMI to microscopic diagnosis, whereas the incubation period was the time from CHMI to the onset of malaria-related symptoms.Table S1).Malaria microscopy was conducted according to standard challenge microscopy procedures adapted from WRAIR. Giemsa-stained thick blood smears were prepared in duplicate from 10 \u00b5L of blood sample spread over a 1\u00d72 cm rectangle and examined under a high power oil immersion objective (100X) by microscopists certified to read thick blood smears for CHMI studies. Smears were considered positive if at least two unambiguous parasites per slide were identified by a study microscopist and confirmed by the lead microscopist. For asymptomatic or treated subjects, up to five passes were read for an area equivalent to \u223c290\u2013320 total high-power fields (hpf), allowing for detection of a parasite density of approximately three parasites/\u00b5L before declaring a blood smear negative. For symptomatic volunteers, up to 15 passes (\u223c870\u2013960 hpf) were scanned before declaring a smear to be negative. Quantification of microscopic parasite densities on slides from the day of diagnosis was performed by two independent microscopists who examined a minimum total of 1 \u00b5L of blood for each subject. Average parasitemia density was calculated from the number of parasites observed divided by the volume of blood examined microscopically (using conversion of volume per hpf)  and at regular intervals after challenge including Day 1 and Day 5 (corresponding to the liver stage of parasite development), the day of the first positive blood smear (corresponding to the blood stage) and post-treatment at Days 35 and 56 following challenge. In addition, subjects had the option to provide separate written consent to participate in a long-term immunology follow-up assessment with collection of serum samples on at three and six months post-challenge.42) were evaluated by enzyme-linked immunosorbent assay (ELISA) as described Levels of antibodies against liver- and blood-stage antigens including circumsporozoite protein (CSP), apical membrane antigen 1 (AMA-1) and the 42 kDa fragment of merozoite surface protein 1 (MSP-16 PBMCs and \u22654-fold above the mean of the negative control wells as described Cell-mediated immune responses to CSP and liver-stage antigen 1 (LSA-1) were evaluated by interferon-gamma (IFN\u03b3) ELISpot assay using peripheral blood mononuclear cells (PBMCs) as previously described The study was an open-label single intervention trial to demonstrate the reproducibility of CHMI at a new center. Primary and secondary analyses involved summaries and descriptive statistics for safety, efficacy (infectivity) and immunology endpoints. Comparative analyses of the primary (microscopy) and exploratory (qRT-PCR) infectivity endpoints was undertaken for the safety population and is presented as the difference in the cumulative distribution of the number of days between challenge and the first positive test (blood smear minus qRT-PCR) using a Kaplan-Meier estimator. Analyses of immunology endpoints (by ELISA and ELISpot) were performed for all subjects for whom data was available at a specific time point. Qualitative assay data analysis was performed by tabulating the frequency of positive responses for each assay at each timepoint an assessment was performed.A total of 18 subjects were screened from January \u2013 March 2010 for eligibility to enroll in the study Figure 1Figure S1).Six subjects successfully completed the experimental infection, receiving a minimum of five invective bites. One subject inadvertently received a total of six infective bites, rather than the five defined in the protocol. This deviation was reviewed by the Medical Monitor and Safety Monitoring Committee. After reviewing safety data, both agreed that the additional infective bite did not put the subject at increased risk. As noted in No SAEs were reported for any subject through Day 56 of the trial or during six months following the CHMI. The incidence of solicited and unsolicited AEs reported as related to the malaria challenge or infection is summarized in During the acute reactogenicity period , the most common AE was pruritus localized to the site of inoculation, occurring in 3 (50%) subjects. No other AE occurred in >1 subject during this period. All subjects experienced at least one systemic AE during the 28 days post-challenge. The most frequently reported systemic AEs related to malaria were headache and myalgia each occurring in 5 (83.3%) subjects, followed by fever, malaise, nausea and arthralgia each occurring in 4 (66.7%) subjects Table 1.All subjects reported at least one unsolicited AE. The majority of unsolicited AEs were mild, and most were considered unrelated to challenge procedures or malaria infection. Related events included cough, decreased appetite, dizziness, insomnia and exertional dyspnea Table 1.Four of six subjects had no laboratory abnormalities throughout the trial. One subject had an asymptomatic mild-moderate increase in liver transaminases (ALT 160 U/L [2.5 xULN] and AST 105 U/L [2.6 x ULN)] occurring on the same day as diagnosis of patent parasitemia . The other subject experienced a mild, asymptomatic decrease in Hb from baseline (1.8 g/dL) occurring on Day 10 post-challenge that remained within the normal range. Both abnormalities resolved by Day 35.All subjects undergoing CHMI developed microscopic parasitemia with a mean prepatent period of 11.2 days  Table 2.Table S1). Similar to reports from other centers, there was no apparent correlation between incubation or prepatent periods and density of parasitemia at diagnosis.The geometric mean parasite density (by microscopy) at the time of microscopic diagnosis was 10.8 parasites/\u00b5L across all slides examined  (10 parasites/mL (Table S2). The parasite density by qRT-PCR on the day of corresponding microscopic diagnosis ranged from 3,730 to 120,700 parasites/mL, which was in agreement with reports from other CHMI centers We previously reported the performance characteristics of our qRT-PCR assay, and its comparability to microscopic diagnosis in this trial Subjects were treated with chloroquine upon microscopic diagnosis and were released from the hotel after three consecutive negative daily blood smears. All subjects were blood smear-negative within two days of initiating treatment Table 3.P. falciparum antigens were analyzed for all six subjects through Day 56 and, for five of the six subjects who consented to the extension portion of the protocol, at three and six months post-challenge. Humoral responses to P. falciparum were measured by ELISA for pre-erythrocytic (CSP) and blood-stage (AMA-1 and MSP-142) antigens  observed in one subject were likewise consistent with uncomplicated malaria and resolved within 24 hours of initiating antimalarial treatment.Consistent with the literature In this study, prepatent parasitemia was detected by qRT-PCR an average of 3.7 days before blood smear diagnosis, and modeling of the qRT-PCR data suggested that 28\u201360 (range 8\u2013111) hepatocytes were infected per subject as previously reported In this study, mosquito infectivity was approximately 50% as assessed by the post-challenge dissection and sporozoite rating system As expected from the limited antigenic exposure with only five to six infected mosquito bites, limited humoral and cellular immune responses to pre-erythrocytic and erythrocytic antigens were observed. Although two subjects had transient measurable antibody levels to CSP, neither maintained a detectable response at six months post-challenge. A third subject had pre-existing humoral and cellular responses to CSP that persisted at all timepoints, did not markedly increase after infection and remains unexplained; anomalous cross-reactive responses to CSP have also been reported by others P. falciparum in our study was not sufficient to induce significant antibody response to AMA-1 as detected by our standardized assay. The peak cellular response was expected at Day 35+/\u22123 days, as previously reported in experimental malaria infection models Likewise, a single challenge with P. falciparum sporozoites, and all subjects developed patent parasitemia and were successfully treated and cured with standard doses of chloroquine. The CHMI was safe and well tolerated. The AE profile, prepatent and incubation periods were consistent with the expected clinical course and response to CHMI reported in previous malaria studies conducted at other centers worldwide. All volunteers developed antibody responses to the blood-stage antigen MSP-1, which was detectable up to six months post-challenge. Both humoral and cellular responses to pre-erythrocytic antigens CSP and LSA-1 were limited. Results from a diagnostic qRT-PCR assay correlated with blood smear density results and identified prepatent infection nearly four days before peripheral blood smears. Seattle BioMed plans to support additional CHMI studies to accelerate development of effective malaria drugs and vaccines.In summary, this study demonstrates the successful implementation of the CHMI mosquito bite model at Seattle BioMed, thereby expanding the global capacity for conducting these important studies. Subjects were successfully infected with Figure S1Cumulative mosquito sporozoite ratings for all mosquitoes used to challenge each subject. The sporozoite load was rated microscopically according to a semi-quantitative scale: 0 (no sporozoites observed), +1 (1\u201310), +2 (11\u2013100), +3 (101\u20131000) and +4 (>1000) (TIF)Click here for additional data file.Table S1Microscopy-based parasite density assessment.1Volume per 1 cm pass based on the field number for each microscope, corresponding to 290\u2013320 hpf. Microscope/equipment numbers: Nikon Eclipse 50i, (FN 22), Nikon microscope E200, (FN20)(DOCX)Click here for additional data file.Table S2Comparison of microscopy and qRT-PCR-based parasite density assessments for first smear-positive sample.1qRT-PCR measurements as reported in 2Quantitative agreement denoted if microscopic and qRT-PCR parasite density measurements were within 0.5 log10 parasites/mL of each other.(DOCX)Click here for additional data file.Checklist S1CONSORT checklist for MC-001.(DOC)Click here for additional data file.Protocol S1IRB-approved study MC-001 protocol.(PDF)Click here for additional data file."}
+{"text": "Malnutrition and malaria frequently coexist in sub-Saharan African countries. Studies on efficacy of antimalarial treatments usually follow the WHO standardized protocol in which severely malnourished children are systematically excluded.Few studies have assessed the efficacy of chloroquine, sulfadoxine-pyrimethamine and quinine in severe acute malnourished children. Overall, efficacy of these treatments appeared to be reduced, attributed to lower immunity and for some antimalarials altered pharmacokinetic profiles and lower drug concentrations. However, similar research on the efficacy and pharmacokinetic profiles of artemisinin-combination therapies (ACTs) and especially artemether-lumefantrine in malnourished children is currently lacking.The main objective of this study is to assess whether artemether-lumefantrine is less efficacious in children suffering from severe acute malnutrition (SAM) compared to non-SAM children, and if so, to what extent this can be attributed to a sub-optimal pharmacokinetic profile.P. falciparum malaria aged between 6 and 59 months will be enrolled. Two non-SAM children will be enrolled after the enrolment of each SAM case. Children with severe manifestations of malaria or complications of acute malnutrition needing intensive treatment will be excluded.In two sites, Ouelessebougou, Mali and Maradi, Niger, children with uncomplicated microscopically-confirmed Treatment intakes will be supervised and children will be followed-up for 42 days, according to WHO guidance for surveillance of antimalarial drug efficacy. Polymerase Chain Reaction genotyping will be used to distinguish recrudescence from re-infection. SAM children will also benefit from the national nutritional rehabilitation program.Outcomes will be compared between the SAM and non-SAM populations. The primary outcome will be adequate clinical and parasitological response at day 28 after PCR correction, estimated by Kaplan-Meier analysis. To assess the pharmacokinetic profile of lumefantrine, a sparse sampling approach will be used with randomized allocation of sampling times (5 per child). A total of 180 SAM children and 360 non-SAM children will be recruited during the 2013 and 2014 malaria seasons.This study will provide important information that is currently lacking on the effect of SAM on therapeutic efficacy and pharmacokinetic profile of artemether-lumefantrine. If it shows lower therapeutic efficacy and decreased lumefantrine concentrations, it would inform dose optimization studies in SAM children.NCT01958905 ClinicalTrials.gov:  Plasmodium falciparum malaria are two major public health problems in sub-Saharan Africa. In Sahel countries, both diseases have a marked seasonality, with concomitant peaks in the second half of the year. The rainy season, roughly from July to October, corresponds with the high transmission season of malaria, and to the hunger gap period. In these countries, children under 5 years of age are the highest-risk population for both diseases, and it has been estimated that more than 50\u00a0% of child deaths were attributable to malnutrition potentiating effects on infections [Malnutrition and fections , 2.Child malnutrition is a diverse spectrum: stunting i.e. chronic malnutrition reflected by a low height-for-age, and wasting i.e. acute malnutrition reflected by a low weight-for-height. The latter allows detecting acute or sub-acute episodes, and is a good predictive marker of short-term mortality. It is an indication to refer children into nutritional rehabilitation programs , togetheChild malnutrition is associated with a higher risk of infections and infectious episodes contribute to deteriorate the nutritional status , resultiGenerally, studies on efficacy of antimalarial treatments follow the WHO standard protocol  in whichResearch on the efficacy of artemisinin-combination therapies (ACTs) in malnourished children is greatly lacking. One case of treatment failure has been reported in a child with SAM treated with artemether-lumefantrine (AL) in India, without evidence of genotype resistance . The autIn the population of malnourished children, specific PK properties might alter the efficacy of antimalarials. Malnutrition is associated with increased total body water leading to increased volume of distribution of drugs . MoreoveFurther studies evaluating the effect of malnutrition on ACTs efficacy are warranted. These studies should also assess PK of ACTs in severely malnourished children. Unless drug concentrations are measured, it is impossible to distinguish clinical treatment failure resulting from an inadequate drug exposure from failure due to drug-resistant parasites.The MAL-NUT study aims to address the lack in knowledge regarding the efficacy and PK of AL in children with SAM. AL is becoming even more widely used as first line treatment recommended in Sahel countries, with the implementation of malaria seasonal chemoprophylaxis with SP plus Amodiaquine precluding the use of artesunate-amodiaquine and artesunate-SP in these areas.The research question is: Is the current recommended dose of AL for the treatment of uncomplicated malaria less efficacious in the children with severe acute malnutrition (SAM children) compared to the non-SAM children and is PK in cause? We hypothesize that AL efficacy might be impaired in SAM children, due to altered lumefantrine PK profile in this population.The study aims to assess whether the current treatment dose is adequate for SAM children. It is expected that results will inform further recommendations for malaria treatment in this important target population.The primary objective of the study is to compare the rates of adequate clinical and parasitological response (ACPR), after PCR correction, between SAM and non-SAM children, during 28 days of follow-up.To compare the day 7 lumefantrine concentration and PK profile of lumefantrine between SAM and non-SAM children (The PK of artemether will not be assessed due to feasibility concerns)To assess the effect of malnutrition on i) therapeutic efficacy, ii) PK profile of lumefantrine, with adjustment for other cofactorsTo compare the time to parasite clearance between SAM and non-SAM childrenTo compare the rates of different types of failure (early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF)) between SAM and non-SAM children;To evaluate the relationship between treatment failure and the concentration of lumefantrine in the blood;To compare the rates of reinfection and recrudescence during a follow-up period of 42 days between SAM and non-SAM children; andTo compare the nature and frequency of adverse events between SAM and non-SAM children.The secondary objectives are:As an exploratory objective, we also propose to evaluate the effect of malnutrition on the acquisition of specific antimalarial immunity. Malnutrition is known to cause relative immunosuppression, but very few studies documented its effect on specific antimalarial antibodies. We will compare the level of anti-AMA-1 antibodies at baseline between SAM and non-SAM children, after adjustment for age.MAL-NUT is a non-randomized non-blinded comparative trial designed to compare the efficacy of AL and the lumefantrine pharmacokinetic profile in SAM versus non-SAM children. The two groups to be compared will not be randomly allocated since they include different populations of children. Similarly, blinding is not applicable.Enrolled children will receive AL orally at the WHO-recommended weight-based dose over 3 days under directly observed treatment, and will be followed prospectively for 42 days according to the standardized WHO methods for surveillance of antimalarial drug efficacy in high transmission settings .Follow-up will allow detecting early and late clinical or parasitological failure, with PCR genotyping to distinguish recrudescence from reinfection. The lumefantrine PK profile will be assessed using a population-based approach to restrict the number of samples required per child.Sample size calculations were designed to detect a minimum crude difference in the proportion of adequate clinical and parasitological response (ACPR) between SAM and non-SAM children, with a ratio of 1/2 between the two groups (to ease the recruitment of SAM children). A total number of 540 children (180 SAM and 360 non-SAM) would allow to detect a minimum difference of 8\u00a0% in ACPR proportion (87\u00a0% in SAM versus 95\u00a0% in non-SAM children), with a statistical power of at least 80\u00a0% and a 2-sided significance level of 5\u00a0%, and 15\u00a0% expected missing endpoints. Two thirds of the children will be enrolled in Mali during the 2013 and 2014 malaria seasons, and one third will be enrolled in Niger during the 2014 malaria season.The MAL-NUT study will be conducted in two sites, Oulessebougou in the region of Koulikoro and Maradi in the district of Maradi, located in in the south of Mali and Niger respectively. In both countries, malaria transmission is seasonal with an annual peak of high transmission during and shortly after the rainy season, usually between July and December. AL is one of the recommended first-line malaria treatments. Since 2012, seasonal malaria chemoprevention (SMC) with SP and amodiaquine has been adopted by both countries and is being implemented in several areas , but not in the areas were enrolments will be conducted.Regarding malnutrition, despite progress its prevalence remains at the emergency level in Mali, and Niger remains the country the most affected worldwide . In the In Mali, the study will be conducted in the district hospital of Ouelessebougou, located in the Koulikoro region, about an hour\u2019s drive from Bamako. This hospital hosts the Malaria Research and Training Centre (MRTC) for clinical research activities, and a nutritional rehabilitation program conducted by the Medical Alliance Against Malaria.In Niger, children will be recruited in the primary care health center of Andoum\u00e8 in Maradi city, where malnutrition activities are supported by UNICEF. In Maradi, Epicentre has a permanent research center that has been collaborating with the Ministry of Health on malnutrition and malaria research for many years including clinical trials.Plasmodium falciparum malaria (confirmed by thick and thin blood films), and whose parents or guardians have given their free and informed written consent to participate in the study.The study population will consist of children aged 6\u201359 months with uncomplicated Age between 6 and 59 monthsWeight\u2009\u2265\u20095 kgAxillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C or history of fever during the previous 24 h as reported by the parent/guardianP. falciparum monoinfection confirmed on blood filmParasitic density between 1,000 and 200,000 asexual forms/\u03bcL of blood.High probability of compliance with follow-up visits (no near-term travel plans)Written consent of a parent or guardian who is at least 18 years of ageAccording to the group: in SAM children, weight-for-height z-score\u2009<\u2009-3 SD or MUAC <115 mm and/or bilateral edema, and in non-SAM children, weight-for-height z-score\u2009\u2265\u2009- 3 SD, and MUAC\u2009\u2265\u2009115 mm and absence of edema.A child will be eligible to be enrolled in the study if he/she meets all the inclusion criteria listed below:General danger signs or signs of severe malaria as defined by the WHO;Plasmodium species detected by microscopy;Mixed or mono-infection with another Severe anemia (hemoglobin <5 g / dL);Known underlying chronic or severe disease ;Presence of febrile conditions due to diseases other than malaria which could alter the outcome of the study;Known history of hypersensitivity or contra-indication to any of the study medications: artemether, lumefantrine (first-line medications), or artesunate, amodiaquine (rescue medications)History of a full treatment course with AL in the past 14 daysHeight-for-age\u2009<\u2009-3 Z scores Severe complications of malnutrition requiring hospitalization in intensive care or stabilizationA child will not be eligible if at least one of the exclusion criteria listed below is present:The definition of severe acute malnutrition is a weight-for- height\u2009<\u2009-3 Z score according to the WHO standards (2006) and / or MUAC less than 115 mm and/or presence of bilateral edema . ChildreTo recruit SAM children, children presenting at the nutritional center, meeting the following criteria and interested to participate in the study will be referred to the study staff: Aged 6 to 59 months, SAM according to WHO definition , fever oRegarding the enrolment of non-SAM children, malaria cases will be sought at the routine pediatric consultation. Children with fever or a history of fever and positive RDT who are interested to participate into the study will be referred to the study staff and after informed consent is obtained, a capillary blood sample will be taken from a finger prick for thick blood film and measurement of hemoglobin concentration. Enrolments will be limited to the first two eligible non-SAM children received after the enrolment of a SAM child , to allow carrying out inclusions in parallel for the two study groups, with benefit in terms of representativeness of the study population throughout the malaria season.Screening blood films will be read on-site. If the parasite density is between 1000 and 200\u2009000 parasites per microliter, the child will be referred to the clinical investigator who will complete the eligibility screen. Eligible patients will be enrolled in the study and assigned an identification number after written informed consent has been obtained.The treatment under evaluation is artemether 20 mg - lumefantrine 120mg: Coartem\u00ae Novartis  will be administered six times, twice daily for 3 days .The interval between the first and second administration is 8 +/-1 h, then subsequent intakes will be administered in the morning and evening, 12 h +/- 2 h apart, i.e. 24, 36, 48 and 60 h after the first dose. To improve absorption, treatment intake will be associated with fat intake. In non-SAM children, the tablets should be administered with a glass of milk, or in the middle of a breastfeeding. In SAM children, treatment should be administered with ready-to-use therapeutic food (RUTF) or therapeutic milk.All AL doses will be directly observed by the study nurse. Patients will then remain under observation for 30 min. If vomiting occurs, a second dose should be administered. If the child vomits after the second dose, he will be removed from the study and rescue treatment will be administered.All children will receive an insecticide-treated bed net at enrolment.One tablet of 5 mg of folic acid will be administered to each child at Day 0. A systematic deworming by albendazole will be administered at Day 14, according to national guidelines. If the axillary temperature exceeds 38.5\u00a0\u00b0C and external cooling measures are insufficient, a dose of 15 mg/kg of paracetamol can be administered and repeated every 6 h. Iron supplementation will be administered in case of anemia for children not requiring nutritional supplements .Additionally in SAM children, feeding medications and associated therapies will be provided according to the national nutrition programs of Mali and Niger: RUTF (Plumpynut\u00ae) will be provided, corresponding to an intake of approximately 170 kcal/kg/day. The child will be closely monitored, if his nutritional status does not improve or if he encounters a medical complication, he will be transferred to the intensive care unit. A systematic treatment with oral amoxicillin will be administered for 7 days, twice a day at the dosage of 50 to 100 mg/Kg/day. A dose of vitamin A will be administered at Day 28, except for children who have already received a dose in the previous 4 months. Finally, SAM children without prior vaccination will be vaccinated against measles at Day 28.Other drugs may be administered for concomitant diseases. However, antibiotics with antimalarial activity will be prohibited for the duration of follow-up, unless no alternative treatment is available.P. falciparum.Antimalarial rescue treatment will be administered in case of treatment failure, repeated vomiting or malaria infection with species other than In case of treatment failure without signs of severe malaria, children will receive Artesunate-Amodiaquine fixed combination  orally, which is the second-line treatment in Mali and Niger, administered at fixed hours for 3 days .In case of treatment failure with signs of severe malaria or oral administration impossible (repeated vomiting), artesunate will be administered IV at a dose of 2.4 mg / kg at 0, 12 and 24 h, and then every 24 h until the patient can tolerate oral treatment. An alternative is the use of artemether IM at a dose of 3.6 mg/kg on day 1 followed by 1.6 mg/kg the following days. After that, a full 3-day course of ASAQ should be administered.After rescue treatment is administered, the weekly clinical follow-up will continue until D 42 to allow the research team to verify its efficacy, and to monitor for any adverse events.At baseline, socio-demographic and anthropometric characteristics, medical history, clinical data, and biological data will be collected: in addition to the blood film and hemoglobin concentration performed at screening, capillary blood will be collected from a finger prick, and a few drops will be placed on filter paper to perform PCR genotyping  and to measure Anti-AMA1 IgG .Children will be kept under observation in the hospital for the 72 first hours to ensure supervised administration of the treatment. On Day 1, Day 2, Day 3, a clinical examination will be performed, axillary temperature measured, and thick and thin blood films collected from capillary blood. In addition, the determination of parasite clearance time will involve extra blood films at h6, h12, h36.The child will return home after the study team has noted their contact information. An insecticide-treated mosquito net will be provided and the appointment schedule for weekly follow-up visits will be clearly explained to the parent/caretaker.On day 7, 14, 21, 28, 35 and 42, the child will undergo a physical examination with temperature measurement, and thick and thin blood films from capillary blood. Adverse events and concomitant medications will be reported. The measurement of hemoglobin concentration will also be performed on day 28.If the child attends the study center for an unscheduled visit, a physical examination will be performed. If symptoms are suggestive of malaria, blood films will be performed.In case of treatment failure, capillary blood will be collected on filter paper for PCR genotyping and lumefantrine concentration assay and the rescue treatment will be administered.Children not attending their appointment will be actively traced by home visitors the next morning.For children with SAM, each weekly visit in the study will be combined with a visit to the nutritional rehabilitation center for monitoring and treatment of malnutrition.The schedule of follow-up activities is summarized in Fig.\u00a0Lumefantrine concentrations will be analysed using a population-based approach to restrict the number of samples required per child . To achiFor ethical and practical reasons, wherever possible, we will combine the PK samplings with other samplings performed for assessing parasitological efficacy. PK modelers have the following population sampling scheme: The first collection will be performed at h6, h12, h24, h36 or h48 , the second at h60 (the evening of day 2), the third at h72 (just before the discharge), the fourth at day 7, and the fifth at day 14 or day 21 . An additional dosage will be made on the day of failure, if parasitological failure occurs.The random allocation sequence of PK sampling times will be computer-generated by Excel before the study starts. The randomization list with be retained in each study site by the investigators, who will directly implement the allocation sequence (no blinding is required).The PK of artemether and its active metabolite, dihydroartemisinin, will not be performed due to the heavy burden in this vulnerable population as well as logistical concerns as three venous blood samplings between h0 and h8 then plasma storage and shipment at -80\u00a0\u00b0C would be required. As an alternative, the parasite clearance will be determined in all patients, as it is considered the best indicator of impaired efficacy of the artemisinin component of an ACT . A proloDuring this study, only capillary blood from finger pricks will be collected. The total amount of blood required for each patient included will approximate 7 ml for the total duration of follow-up.) will be used.For malaria screening, the SD Bioline\u00ae HRP2 RDT  on day 0 and day 28.glurp, msp-2 and msp-1 loci are a sign of treatment failure, while distinct genetic profiles indicate reinfection [PCR genotyping of malaria parasites will be performed to differentiate between a recrudescence and a re-infection. For this purpose, samples will be collected at enrolment and at treatment failure (if any) on filter paper and stored individually in re-sealable, zippered pouches containing a desiccant and sheltered from moisture, excessive heat and light. PCR genotyping will be carried out at the MRTC laboratory in Bamako. PCR adjustment consists of comparing primary infection parasites with those responsible for secondary infections. Identical genotypes at the nfection . The paiFor lumefantrine concentration assessment, capillary blood will be collected in a microcapillary tube coated with lithium heparinate, then immediately transferred to a filter paper previously treated by the laboratory of Cape Town (50 microliter of whole blood per spot). Once dry, the filter papers will be stored away from heat and excessive light and regularly sent to the Division of Clinical Pharmacology, University of Cape Town for analysis. Quality controls spiked by the analytical laboratory, specifically designed to control for the environmental conditions at the site and during storage, will be stored and sent under the same conditions. PK assays will be performed using liquid chromatography and tandem mass spectrometry (LC MS/MS).At h0, for children enrolled in Mali, 3 drops of capillary blood will be placed on filter paper for the dosage of anti-AMA1 specific antimalarial antibodies by Elisa method. Analysis will be performed by MRTC, Bamako.The primary outcome of this study is the proportion of patients having an adequate clinical and parasitological response (ACPR) as defined by the WHO  on day 2\u25cb Corrected ACPR proportion on day 42\u25cb Failure proportion by type  by day 28 and 42, as defined by WHO \u25cb Proportion of reinfection and recrudescence\u25cb Time to parasite clearance and parasite clearance half-lifeRegarding treatment efficacy\u25cb Area under curve (AUC) and other PK parameters \u25cb Concentration on day 7\u25cb Concentration on day of failure (in case of failure)Regarding PK of lumefantrine\u25cb Type and frequency of adverse eventsRegarding safetySecondary outcomes are:P. falciparum, death, severe adverse event, loss to-follow-up\u2026). In all cases, the date and reasons for the interruption will be collected in the report form.Children will be followed until day 42 whenever possible. However, some reasons may lead to the interruption of monitoring , seriousness, relationship with AL, action(s) taken, date of resolution, outcome.A physician investigator will be available on site 24h per day for the management of adverse events. In case of a SAE, as defined by ICH , a specific SAE declaration form will be filled in by the site principal investigator within 48 h and sent to the investigator coordinator in Epicentre, the Ethics Committees and the DSMB. They will recommend any action to be taken (including early termination of the study). In addition, ethics committees who reviewed and approved the protocol will also receive periodical reports of aggregated data on AEs, at regular intervals throughout the study.Study data will be double entered using REDCap electronic data capture tools hosted at Epicentre  then datP. falciparum malaria, number of patients infected by other Plasmodium species, number of patients excluded (and reasons why they were not eligible), number of patients enrolled and analyzable in each group of SAM and non-SAM children.A flowchart will present the following information: number of patients screened, number of patients infected with For all enrolled children, baseline characteristics will be presented for each group (SAM versus non-SAM).Clinical and parasitological efficacy outcomes will be presented for SAM and non-SAM children: number of ETF, LCF, LPF, ACPR, re-infection, recrudescence, mixed infection, by day 28 and day 42.The Kaplan-Meier survival analysis will be used to calculate estimates of survival without subsequent infection in both intent-to-treat and per-protocol populations. This analysis will allow accounting for censored data , 34.Logistic regression analysis will be used to compare at day 28 (and then day 42) the proportion of ACPR in patients for which the endpoint has been determined, excluding those lost to follow-up or with missing endpoint.Two methods will be used to estimate the therapeutic efficacy in each group:After univariate analyses, multivariate analyses using survival models (time to recurrence) and logistic regression (proportion of ACPR) will be performed where the therapeutic success will be explained by the degree of malnutrition and other cofactors, including lumefantrine PK profile.PK modeling will estimate the lumefantrine area under the curve, Cmax, Tmax, Volume of Distribution and Clearance in SAM and non-SAM children using nonlinear mixed effect models (NONNEM).The values obtained will be compared between groups, and related to parasitological efficacy.Regression models of the log-transformed parasite counts will be fitted in order to estimate parasite clearance using the Parasite Clearance Estimator Tool (PCE) developed by the WorldWide Antimalarial Resistance Network (WWARN) . ParasitA French version of the protocol and informed consent form has been approved by the Ethics Committee of the Faculty of Medicine and Odonto-Stomatologie and the Faculty of Pharmacy in Bamako, Mali and Niger National Ethics Committee of the Ministry of Health. An English version has been approved by MSF Ethical Review Board.Any amendment to the protocol that may affect the rights, safety and / or welfare of the participants or the conduct of research will be submitted again to Ethics Committees before being implemented.Individual data will not be disclosed to anyone outside the research team, unless it is necessary for the proper medical management of children . The database will not contain any personally identifiable information; patients are only identified by their inclusion number.The study will comply with the recommendations of Good Clinical Practice and with standards of care established by the national health programs and the Declaration of Helsinki. An external Good Clinical Practice monitor will conduct regular visits to the sites, as detailed in a dedicated monitoring plan. Data report forms will be checked on site in relation to source documents to ensure they are complete and clear. The monitor will also review compliance with written standardized operating procedures.For this study, a DSMB will be instituted including members with the following expertise: pharmacometrician, epidemiologist, malaria specialist, and pediatrician. As described in a dedicated DSMB charter, it will be responsible for regularly review the results regarding the enrolment of patients, and the efficacy and safety of AL. The DSMB will advise the Scientific Committee on the continuation or, if needed the amendment or discontinuation of the trial.Inclusion in the study is voluntary and will only be possible if the adult parent or guardian gives written informed consent.Participants will receive a free malaria treatment and will benefit from active monitoring in the health facility for 42 days. In case of treatment failure, they will receive a rescue treatment. Concomitant diseases will also be actively sought and promptly managed. Children with SAM will benefit appropriate nutritional care. All the care provided meets the highest quality standards, and is in accordance with national guidelines.Participants will be informed of the possible occurrence of side effects related to the study treatments, and these will be systematically recorded and managed.The constraints for the child (and caregiver) will be to stay in the health center for the 3 days of treatment, and then return to the study site every week for 6 weeks. Reimbursements will be given in compensation for the time spent for visits and travel costs, according to the recommendations of the community leaders. During follow-up, the child will have several capillary blood collections that are not performed routinely, although the total blood volume sampled will be limited to under 10 mL so is not associated with a particular risk. The risks are fear and pain at the puncture site.This study will be conducted during 2013 and 2014 malaria seasons in Mali, and 2014 malaria season in Niger. Enrolments are expected to end in early 2015 and results should be available in second semester 2015.The study report will be discussed and circulated to the stakeholders, including the local officials. A poster summarizing the main results will be posted at the two study hospitals to inform the participants. Several oral feedback sessions will also be held on the study sites and with scientific partners and policy makers. The results will be published in scientific journals and reported at international conferences.In the study areas, a better understanding of the efficacy of antimalarial drugs (in SAM and non-SAM children) will contribute to updating the protocols for case management of malaria, and thus contribute to improving the health of the population.More generally, this study will provide important, lacking information on the efficacy of AL in SAM children and could inform further recommendations for the management of malaria in this specific population. It will therefore benefit all malnourished children living in malaria-endemic areas (the entire Sahel Africa is concerned). If our results indicate a lack of efficacy of AL at usual doses, we will recommend evaluating the use of increased doses through dose optimization studies.Trial status: active recruitment"}
+{"text": "P = 0.006) and the proportion of children with parasitemia 1 day after treatment began was significantly lower (P = 0.016) in artesunate-amodiaquine\u2014compared with artemether-lumefantrine-treated children. Parasite clearance times were similar with both treatments. Overall efficacy was 96.3% , and was similar for both regimens. Polymerase chain reaction-corrected parasitologic cure rates on Day 28 were 96.9% (95% CI 93.9\u201398.2%) and 98.3% (95% CI 96.1\u201399.3%) for artemether-lumefantrine and artesunate-amodiaquine, respectively. Gametocyte carriage post treatment was significantly lower than pretreatment (P < 0.0001). In anemic children, mean time to recovery from anemia was 10 days (95% CI 9.04\u201310.9) and was similar for both regimens. Both treatments were well tolerated and are safe and efficacious treatments of uncomplicated falciparum malaria in young Nigerian children.The efficacy of 3-day regimens of artemether-lumefantrine and artesunate-amodiaquine were evaluated in 747 children < 5 years of age with uncomplicated malaria from six geographical areas of Nigeria. Fever clearance was significantly faster ( Plasmodium falciparum in some parts of Southeast Asia where ACTs have been deployed for over a decade as first-line treatments,P. falciparum has not yet been reported. Historically, resistance to antimalarial drugs originates from Southeast Asia and South America, and then spreads to East and Central Africa, and to West Africa.9Artemisinin-based combination treatments (ACTs), the recommended first-line treatments of uncomplicated falciparum malaria globally,P. falciparum to artemether-lumefantrine and artesunate-amodiaquine, we evaluated the therapeutic efficacy of artemether-lumefantrine and artesunate-amodiaquine in Nigerian children < five years of age. The primary aims were to compare the efficacy of these ACTs in all malaria-endemic settings in Nigeria and to assess changes in P. falciparum susceptibility to ACTs.In Nigeria, artemether-lumefantrine and artesunate-amodiaquine were adopted as first-line treatments in 2005.This study was an open-label study trial carried out between October 2009 and November 2010 at the following locations: Agbani, Ikot Ansa, Barkin Ladi, and Damboa, in Enugu, Cross River, Plateau, and Borno states, respectively (the eastern flank of the study sites), and in Ijede, Sabo quarters of Ibadan and Makarfi in Lagos, Oyo, and Kaduna states, respectively (the western flank) . In virtP. falciparum parasitemia between 1,000 and 200,000 asexual forms per \u03bcL, a body (axillary) temperature > 37.4\u00b0C, or history of fever in the 24 to 48 h preceding presentation, absence of other concomitant illness, no history of antimalarial use in the 2 weeks preceding presentation, and written informed consent given by parents or guardians. Patients with severe malaria,Patients were eligible to join the study if they were < 5 years of age, had symptoms compatible with acute uncomplicated falciparum malaria such as fever, anorexia, vomiting, or abdominal discomfort with or without diarrhea, with Enrolled patients were randomized to receive artemether-lumefantrine or artesunate-amodiaquine (co-formulated) given according to body weight. Artemether-lumefantrine  was given as follows: patients weighing 5\u201314 kg received one tablet, and those weighing 15\u201324 kg received two tablets at presentation (0 hour), 8 hours later, and at 24, 36, 48, and 60 hours after the first dose. Each tablet of artemether-lumefantrine contains 20 mg of artemether and 120 mg of lumefantrine. Artesunate-amodiaquine  was also given as follows: patients weighing \u2265 4.5 to < 9 kg received one tablet, those weighing \u2265 9 to < 18 kg received one tablet, and those weighing \u2265 18 to < 24 kg received one tablet of the following formulations: 25 mg/67.5 mg, 50 mg/135 mg, 100 mg/270 mg of fixed dose combination of artesunate/amodiaquine, respectively, daily for 3 days.All drugs were given orally. In patients treated with artesunate-amodiaquine, the drug was given as single day or single daily doses in the clinic by the physician. In patients treated with artemether-lumefantrine, the 0-, 24-, and 48-hour doses were given in the clinic by the physician, and the 8-, 36-, and 60-hour doses were given by parents or guardians of the children at home. Parents or guardians were questioned at follow-up on the time and events after drug administration. After drug administration in the clinic, all patients waited for at least 30 minutes to ensure the drug was not vomited. If it was, the dose was repeated. If the repeat dose was vomited, the patient was excluded from the study. If necessary, patients were provided with antipyretics (paracetamol tablets 10\u201315 mg/kg every 8 hours for 24 hours). The randomization was computer generated and treatment codes were sealed in individual envelopes. Patient evaluation and follow-up after administration was performed by another physician blinded to the drug treatment. Thick and thin blood films were obtained from each child as soon as they came to the clinic and the slides were carefully labeled with the patients' codes and air-dried before being stained.Follow-up with clinical and parasitological evaluation was done daily on Days 1\u20133 and on Days 7, 14, 21, and 28. This consists of enquiry about the patients well being, presence or absence of initial presenting symptoms, presence of additional symptoms, measurement of body temperature, heart and respiratory rates, and a blood smear for quantification of parasitemia.Side effects were defined as symptoms and signs that first occurred or became worse after treatment was started and were checked for at every visit. Laboratory tests to further elicit adverse events were not performed routinely at every visit. Any new events occurring during treatment were also considered as side effects.Thick and thin blood films prepared from a finger prick were stained with Giemsa and examined by light microscopy under an oil-immersion objective at 1,000\u00d7 magnification by two independent assessors who did not know the drug regimen of the patients. A senior member of the study team reviewed the slides if there was any disagreement by the two microscopists. In addition, the slides of every fourth child enrolled in the study were reviewed by the senior member. Parasitemia, asexual or sexual, in thick films were estimated by counting asexual and sexual parasites relative to 500 leukocytes, or 500 asexual or sexual forms, whichever occurred first. From this figure, the parasite density was calculated assuming a leukocyte count of 6,000/\u03bcL of blood. A slide was considered parasite negative if no asexual or sexual parasite was detected after examination of 200 microscope fields.Capillary blood, collected before and during follow-up, was used to measure packed cell volume (PCV). The PCVs were measured using a microhematocrit tube and microcentrifuge .Blood was spotted on filter paper on Days 0, 3, 7, 14, 21, and 28, and at the time of treatment failures for parasite genotyping. Paired primary and post treatment samples were analyzed using parasites' polymorphic genes to distinguish recrudescence from new infections. Briefly, block 2 of merozoite surface protein-1 (MSP-1) and the block 3 of merozoite surface protein-2 (MSP-2) and region II of glutamine-rich protein (GLURP) were amplified by two rounds of polymerase chain reaction (PCR) using specific primers as previously described.The banding pattern of post-treatment parasite was compared with matched primary parasites in each of the patients who had parasitemia after treatment with either artemether-lumefantrine or artesunate-amodiaquine. Post treatment and primary infection parasites showing identical bands were considered as recrudescence, whereas non-identity in banding patterns was considered as newly acquired infections. To confirm the absence of recurrent parasitemia, samples obtained in one in every four patients with microscopically negative blood films were also subjected to PCR analysis.in vivo clinical classification criteria.Response to drug treatment was assessed using a modified version of the World Health Organization (WHO) 2003 Cure rates were defined as the percentages of patients whose asexual parasitemia cleared from peripheral blood and who were free of patent asexual parasitemia on Days 14, 21, and 28 of follow-up. The cure rates on Days 21 and 28 were adjusted on the basis of the PCR genotyping results of paired samples of patients with recurrent parasitemia after Day 7 of starting treatment.All patients failing treatment, defined as reappearance of parasitemia after its initial complete clearance or failure of complete clearance within 28 days, were retreated whether they became symptomatic or not and after samples had been obtained for microscopy and PCR analysis. Patients failing treatment with artemether-lumefantrine were retreated with artesunate-amodiaquine and vice versa, and were followed up for another 28 days.P. falciparum asexual and sexual forms. Proportions were compared by calculating \u03c72 using Yates' correction, Fisher's exact or Mantel Haenszel tests. Normally distributed, continuous data were compared by Student's t test and analysis of variance. Data not conforming to a normal distribution were compared by the Mann-Whitney U tests and the Kruskal Wallis tests (or by Wilcoxon ranked sum test). The risk for parasite reappearance was calculated by survival using the Kaplan-Meier method. In vivo parasitological measures of artemisinin susceptibility were calculated as previously described.P values of \u2264 0.05 were taken to indicate significant differences. Data were double entered serially using patients' codes and were only analyzed at the end of the study.Sample size was calculated so that the study will be able to detect a 5% absolute difference in parasitologic failure rate between artemether-lumefantrine and artesunate-amodiaquine treatment groups with 95% power and at a 5% significance level. The expected treatment success rates were 100% for artemether-lumefantrine and 95% for artesunate-amodiaquine on Day 28. Except at Ikot Ansa, where a total of 63 children was enrolled in the two treatment arms, a minimum of 50 patients per treatment arm were enrolled at each site. Data were analyzed using version 6 of Epi-Info softwareBetween October 2009 and November 2010, 747 patients were recruited: 360 in artemether-lumefantrine and 387 in artesunate-amodiaquine groups . Response to both treatment regimens was not related to age: 13 of 123 and 18 of 236 of \u2264 2 and > 2 years of age, respectively, treated with artemether-lumefantrine failed treatment by Day 28 (P = 0.46). Similarly, 10 of 114 and 18 of 271 \u2264 2 and > 2 years of age, respectively, treated with artesunate-amodiaquine failed treatment by Day 28 (P = 0.60).Overall, ACPR in both treatment groups on Days 14, 21, and 28 was seen in 713 of 719 children , 681 of 707 children , and 658 of 678 children , respectively. In children treated with artemether-lumefantrine, ACPR on Days 14, 21, and 28 was seen in 344 of 348 children , 329 of 339 children , and 316 of /327 children , respectively. In children treated with artesunate-amodiaquine, ACPR on Days 14, 21, and 28, was seen in 369 of 371 children , 352 of 368 children , and 342 of 351 children , respectively. There were no significant differences in these parameters at all times in both treatment groups . In artemether-lumefantrine-treated children, PCR corrected parasitologic cure rates on Days 14, 21, and 28 were 344 of 348, 98.9%, 95% CI 96.9\u201399.6%; 333 of 339, 98.2%, 95% CI 96.0\u201399.3%; and 316 of 327, 96.6%, 95% CI 93.9\u201398.2%, respectively. In artesunate-amodiaquine-treated children, PCR corrected parasitologic cure rates on Days 14, 21, and 28 were 369 of 371, 99.5%, 95% CI 97.9\u201399.9%; 356 of 368, 96.7%, 95% CI 94.2\u201398.2%; and 345 of 351, 98.3%, 95% CI 96.1\u201399.3%, respectively. There were no significant differences in these parameters at all times in both treatment groups .Of the 72 recurrent infections during the 28-day follow-up period, 21 were new infections and 34 were recrudescent infections of P = 0.48). Overall, the probability of reappearance of asexual parasitemia after treatment with both combinations were similar . Similarly, recrudescent infections were not related to age: 1 and 37 infections were in the < 1 and = 1 year olds, respectively .The median times to recrudescence and new infections (reinfections) were similar . Recurrent infections were similar in both treatment groups: artemether-lumefantrine, 24 of 349; artesunate-amodiaquine, 21 of 375, N = 314) versus 60 . The median dose of artemether was also significantly higher in children without recrudescence 12.6  versus 10.0 . In children treated with artesunate-amodiaquine, the median dose of amodiaquine was similar in children without recrudescence and in those with recrudescence: 31.2  versus 31.2 . The median dose of artesunate was also similar in children without and with recrudescence 11.5 , versus 11.5 . The findings on the median dose in the two treatment groups were seen at all study sites (data not shown).In children treated with artemether-lumefantrine, the median dose of lumefantrine was significantly higher in children without recrudescence than in those with recrudescence: 75.8 . However, parasite positivity rate on Day 2 were similar in children with recrudesce and those without recrudescence . In children treated with artemether-lumefantrine, parasite positivity rate on Day 1 were similar in children with recrudesce and those without recrudescence . Parasite positivity rate on Day 2 were also similar in children with recrudesce and those without recrudescence . In children treated with artesunate-amodiaquine, parasite positivity rate on Day 1 were similar in children with recrudesce and those without recrudescence . Parasite positivity rate on Day 2 were also similar in children with recrudesce and those without recrudescence . Parasite positivity rate on Day 3 was also similar in children with recrudescence and in those without recrudescence .Overall, parasite positivity rate on Day 1 was significantly higher in children with recrudesce compared with those without recrudescence . However, treatment outcome was more likely to be different in Agbani  and Barkin Ladi  for both drugs if adequate clinical and parasitological response in Damboa (Borno State) or other sites was used as the reference. Recrudescent infections were not related to enrollment parasitemia: 4 of 71 and 31 of 675 children with a parasitemia \u2265 100,000/\u03bcL and < 100,000/\u03bcL, respectively, had a PCR-confirmed recrudescence (P = 0.52); geometric mean parasitemia : 14 of 35 versus 296 of 274, P = 0.77.The overall rate of recrudescent infections was 4.8% (34 of 711 children) and was significantly higher at sites on the eastern than on the western flanks 26 of 311 versus 8 of 319,  had a parasitemia < 100,000/\u03bcL: 322 and 353 children in artemether-lumefantrine and artesunate-amodiaquine treatment arms, respectively. In these 675 children, parasitemia on Day 3 was seen in 5 children: 2 in artemether-lumefantrine and 3 in artesunate-amodiaquine-treated children\u2014an overall parasite positivity rate on Day 3 of 0.7%. The five patients with parasitemia on Day 3, were scattered among five different study sites. Patient enrollment at each of these study sites was \u223c50\u201360 patients per treatment arm. Thus, there was no evidence of any cluster of cases with slow parasite clearance after treatment.19N = 724): 349 children treated with artemether-lumefantrine and 375 children treated with artesunate-amodiaquine). By Day 1, parasitemia cleared in 46% of the children (N = 724), including 41% (N = 349) of the children treated with artemether-lumefantrine and in 50% (N = 375) of the children treated with artesunate-amodiaquine. The difference between these two proportions was statistically significant (P = 0.013). By Day 2, parasitemia was still present in 6% of children treated with artemether-lumefantrine and in 4.5% of children treated with artesunate-amodiaquine. The difference between these two proportions was not statistically significant (P = 0.47). Overall, parasite clearance times were similar with both artemether-lumefantrine and artesunate-amodiaquine treatments . Parasite clearance times were similar in the two treatment arms at all study sites (data not shown). In one 34-month-old child, treated with artesunate-amodiaquine, parasitemia persisted until Day 7 of follow-up. The child was hyperparasitemic at enrollment . No patient received rescue medication.Clearance of parasitemia was evaluated in all children who completed a minimum follow-up of 7 days . However, fever clearance was similar in children with and without recrudescence . The mean weights in children with and without recrudescence were similar .Parasite clearance was significantly longer in children with recrudescent infections compared with those without recrudescence . By Day 2, fever had cleared in 28 children treated with artemether-lumefantrine and in 19 children treated with artesunate-amodiaquine. The difference between these proportions was also not statistically significant (P = 0.09). Overall, fever clearance was significantly faster with artesunate-amodiaquine compared with artemether-lumefantrine-treated children . At all sites fever clearance times were similar in artemether-lumefantrine-treated children (data not shown). Similarly, at all sites fever clearance times were similar in artesunate-amodiaquine-treated children (data not shown).Five hundred and forty children were febrile at enrollment: 258 of 349 in the artemether-lumefantrine and 282 of 375 in the artesunate-amodiaquine groups. By Day 1, fever had cleared in 227 children (88%) treated with artemether-lumefantrine and in 260 children (92%) treated with artesunate-amodiaquine. The difference between these proportions was not statistically significant (P = 0.36). Overall, post-treatment gametocyte carriage was significantly lower than pretreatment gametocyte carriage in both treatment groups (P < 0.0001). Overall, baseline gametocyte carriage was significantly higher in children with recrudescent infections than in those without recrudescence . On Day 14, gametocytes were not seen in peripheral blood of all children with recrudescent infections (N = 34) but were detectable in the peripheral blood in 1.3% of those without recrudescence (N = 553). In children treated with artemether lumefantrine, 50% (N = 14) of those with recrudescence had gametocytes at baseline, but gametocytes were not detectable in the peripheral blood of these children on Day 14 (N = 13).Gametocytes were detected in the peripheral blood of 159 of 623 children (25.5%) from the two treatment groups . The detP = 0.87). After treatment, the prevalence of anemia was similar in both treatment groups (P = 0.71). Post-treatment prevalence of anemia was significantly lower than pretreatment in both treatment groups (P < 0.0001). Only 2% of children (12 of 646) who were anemic at enrollment were still anemic by Day 28. Changes in hematocrit before, during, and after treatment with both treatment regimens were similar (N = 272), and was similar in artemether-lumefantrine- and artesunate-amodiaquine-treated children . Anemia recovery time was not related to age: in \u2264 2 and > 2 years of age, mean times were 11.3 days, range 1\u201328, 95% CI 9.6\u201312.9, N = 92, and 9.3 days, range 1\u201328 days, 95% CI 8.2\u201310.4, N = 179, respectively, P = 0.53). Overall, anemia recovery time was significantly longer in children with enrollment parasitemia > 50,000/\u03bcL than in children with enrollment parasitemia \u2264 50,000/\u03bcL . In children treated with artesunate-amodiaquine, anemia recovery time was also significantly longer in children with enrollment parasitemia > 50,000/\u03bcL than in children with enrollment parasitemia \u2264 50,000/\u03bcL . In children treated with artemether-lumefantrine, anemia recovery times were similar in the children with enrollment parasitemia > 50,000/\u03bcL and those with enrollment parasitemia \u2264 50,000/\u03bcL .Hematocrit data were available in 672 children at enrollment and in 646 children 28 days after enrollment . Anemia P < 0.0001). In 142 anemic children treated with artemether-lumefantrine, hematocrit was also significantly higher after treatment than before treatment . Similarly, in 147 anemic children treated with artesunate-amodiaquine, hematocrit was also significantly higher after treatment than before treatment .In 289 anemic children (from both treatment arms) in whom hematocrit data were available at enrollment and 28 days after enrollment, hematocrit was significantly higher after treatment than before treatment . Seven children (3 in artemether-lumefantrine and 4 in artesunate-amodiaquine) reported more than two adverse events. Many of the children reporting adverse events were > 2 years of age. Adverse events were carefully documented at two sites: Barkin Ladi and Sabo Quarters of Ibadan. Overall 78 of 223 children (32 of 104 in artemether-lumefantrine and 46 of 119 in artesunate-amodiaquine) reported at least one adverse event within the first week of starting treatment. There was no significant difference in the proportions of patients reporting adverse events in both treatment groups  haplotypePfmdr 1 and Pfcrt (SVMNT),In this relatively large study conducted at sites representing most geographical areas of Nigeria, and at \u223c5 years after adoption of artemether-lumefantrine and artesunate-amodiaquine as first-line treatments,As anticipated, fever clearance was significantly faster in children treated with artesunate-amodiaquine compared with those treated with artemether-lumefantrine. This may be caused by the antipyretic effect of the amodiaquine component of artesunate-amodiaquine.31in vivo resistance of P. falciparum to lumefantrine in children who received the lowest dose of lumefantrine, because no previous studies in Nigeria have shown that such lower doses of lumefantrine resulted in clearance of parasitemia in the region, that would indicate a change in susceptibility to antimalarial drugs.32The significantly reduced gametocyte carriage post treatment by the two ACTs is in keeping with previous reports.in vivo sensitivity to artemisinin in the different regions of Nigeria 5 years after adoption of artemisinin-based combination treatments. The evidences to support the latter are: overall efficacy of 96.5%Based on the 3% threshold of parasite positivity on Day 3 defined by Stepniewaska and others,P. falciparum to antimalarial drugs in Nigeria. For example, chloroquine or sulfadoxine-pyrimethamine resistance was more frequently reported in the eastern than in the western region of the country.P. falciparum strains with reduced susceptibility to artemisinin will emerge or spread to Nigeria, a positive relationship should be found between parasite density and recrudescence of infections in endemic areas of Nigeria.Recrudescent infections occurred in 5% of the children and the rates were significantly higher at sites located on the eastern than on the western flank . The reaP. falciparum has emerged to artemisinin in Southeast Asia, particularly in Cambodia, Thai-Cambodian border and the western border of Thailand where artemisinin combination drugs have been deployed,in vitro test is not reliable for artemisinin compounds, and new tests are being developed,P. falciparum isolates in vitro to artemisinin drugs in NigeriaP. falciparum isolates in vivo, and in vitro, and by molecular markers in endemic areas of Nigeria. The recent identification of molecular markers will help in monitoring the distribution and spread of resistance and to understanding the evolution and mechanism(s) of resistance in P. falciparum to artemisinin drugs.Recent reports indicate that resistance in P. falciparum-infected red blood cells, particularly when parasitemias are high.Compared with non-artemisinin or non- artemisinin-based combination treatments, artemisinin drugs and artemisinin-based combination treatments may retard the elimination, by the spleen, of Reported adverse events to both treatment regimens were similar, did not necessitate discontinuation of treatment in any child, and were, in general, indistinguishable from the symptoms and signs of acute malaria infections. It is noteworthy that no child reported pruritus, an adverse event frequently encountered in Nigerian children treated with 4-aminoquinolines (chloroquine or amodiaquine), and to a lesser extent, halofantrine and mefloquine."}
+{"text": "A regular evaluation of therapeutic efficacy in sentinel sites and a system of surveillance are required to establish treatment guidelines and adapt national anti-malarial drug policy to the rapidly changing epidemiology of drug-resistant malaria. The current anti-malarial treatment guideline in Mauritania, officially recommended since 2006, is based on artemisinin-based combination therapy. The aim of the present study was to evaluate clinical efficacy and tolerance of artesunate-amodiaquine, the first-line treatment for acute uncomplicated malaria, in Mauritanian paediatric and adult patients to validate its continued use in the country.Plasmodium falciparum-infected symptomatic patients aged\u2009>\u2009six months were enrolled in Kobeni and Timbedra in southern Mauritania in September to October 2013. Co-formulated artesunate-amodiaquine was administered at the recommended dose over three days. Patients were followed until day 28. Parasitological and clinical response was classified according to the standard 2009 World Health Organization protocol.A total of 130 patients (65 in Kobeni and 65 in Timbedra) were enrolled in the study. Seventeen patients (13.1%) were either excluded (before PCR correction) or lost to follow-up. Based on the per protocol analysis, artesunate-amodiaquine efficacy  was 96.6% in Kobeni and 98.2% in Timbedra before PCR correction. Late clinical failure was observed in two patients in Kobeni and one patient in Timbedra. After PCR correction, the efficacy rate in the two study sites was 98.2%. On day 3, all patients were afebrile and had negative smears. Treatment was well tolerated.P. falciparum malaria. In the majority of patients, fever and parasitaemia were rapidly cleared before day 3. The results support the national anti-malarial drug guideline for a continued use of artesunate-amodiaquine as a first-line drug for uncomplicated malaria in southern Mauritania.Artesunate-amodiaquine is well tolerated and highly efficacious for the treatment of uncomplicated  Plasmodium falciparum infections occur in the sahelian south of the country, where transmission is seasonal , and malaria represents the first cause of outpatient consultation (25%), hospitalization (35.5%) and mortality (39%).Malaria is one of the major public health problems in Mauritania. Every year, between 200,000 and 300,000 malaria cases are officially notified, mostly without laboratory confirmation of clinical diagnosis . Most PlResistance to chloroquine and sulphadoxine-pyrimethamine is widespread in West Africa . Since ain vitro was reported in thre of five P. falciparum isolates obtained from travellers returning from Mauritania to France [P. falciparum was confirmed in another clinical study conducted in 1998 in Aioun and Kobeni (southern Mauritania) in which it was reported that 33 of 85 (38.8%) patients treated with chloroquine and followed for 14\u00a0days failed to respond to the treatment [P. falciparum chloroquine resistance transporter (pfcrt) found in that study supports clinical resistance to chloroquine in southern Mauritania. By contrast, analysis of key codons in the molecular markers for sulphadoxine-pyrimethamine resistance (dihydropteroate synthase (dhps) and dihydrofolate reductase (dhfr), respectively) in the same samples suggested low prevalence (<22%) of antifolate resistance [In Mauritania, clinical data on anti-malarial drug efficacy are scarce. In one of the first published papers on a regional survey of clinical resistance to chloroquine between 1987 and 1990 in francophone West Africa, it was reported that Mauritania was still free of chloroquine resistance . Howevero France . Chloroqreatment . The higP. falciparum-infected, symptomatic paediatric and adult patients in southern Mauritania where P. falciparum is known to be endemic.Since 2006, in line with the anti-malarial drug policies in neighbouring countries, notably Senegal and Mali, Mauritanian Ministry of Health recommends the systematic use of ASAQ and artemether-lumefantrine for the first- and second-line treatment of acute uncomplicated malaria, respectively. This new treatment guideline is not based on data obtained in the country as ACT has never been evaluated before in Mauritanian patients using a standardized protocol. The present single-arm study was performed to evaluate the efficacy and tolerance of ASAQ in moughataa) of Kobeni in 2013. Only one health centre attends to the health needs of the population residing in Kobeni.The clinical studies were conducted in two study areas in southern Mauritania: Kobeni (Hodh El Gharbi province) and Timbedra (Hodh Echargui province) Figure\u00a0. The towla route de l\u2019Espoir, about 1,100\u00a0km from Nouakchott. There were 76,932 inhabitants in the department (moughataa) of Timbedra in 2013. The agricultural and pastoral lifestyle in these two cities is similar. During the rainy season (July to October or November), an average of 30 febrile cases per day are treated in the health centres in Kobeni and Timbedra.The city of Timbedra  is situated in the southeastern region of the country (Hodh Ech Chargui region) and lies along the major road, P. falciparum mono-infection with parasitaemia between 250 and 100,000 asexual parasites/\u03bcL, presence of fever (axillary temperature \u226537.5\u00b0C) or history of fever 24\u00a0hours preceding medical consultation, and ability to swallow oral medication [Plasmodium species, severe malnutrition, fever due to concomitant diseases, and history of allergic reaction to amodiaquine or artemisinin derivatives were excluded. Due to the relative contra-indication of artemisinin derivatives during the first trimester of pregnancy and during breastfeeding, married women with a positive pregnancy test and breastfeeding women were excluded from the present study. In addition, due to the ethical problem in performing pregnancy test in adolescent girls aged between 12 and 18\u00a0years old and single adult women >18\u00a0years old, these two categories of patients were excluded from the study protocol.Patients spontaneously consulting at the health centres in September and October 2013 were enrolled after informed written consent if the following criteria were met: age\u2009>\u2009six months old (without upper limit), dication . Patientdication , mixed P\u00ae Hb 301 system .Finger-prick capillary blood was collected to prepare and examine Giemsa-stained thin and thick smears under the microscope, according to standard WHO procedures . Blood sASAQ  was administered on days 0, 1 and 2 at the doses recommended by the manufacturer on mg base per weight basis. Each dose administration was supervised by one of the team members, and patients were observed for at least 30\u00a0minutes after drug intake for possible vomiting or adverse effects. In case of vomiting during the observation period, the same dose was given, and the patient was observed for an additional 30\u00a0minutes. In case of repeated vomiting, the patient was withdrawn from the study protocol and rescue treatment was administered . Paracetamol (10\u00a0mg/kg body weight) was administered to all patients three times a day for fever and headache.Patients were followed on days 1, 2, 3, 7, 14, 21, and 28 [The primary endpoint was determined on day 28 and classified into one of the following categories: early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), or adequate clinical and parasitological response (ACPR), as defined by the WHO [Plasmodium species was determined using 18S rRNA as the target molecule [msa1), merozoite surface antigen-2 (msa2), and glutamine-rich protein (glurp), as recommended by the WHO [Parasite DNA was extracted from filter papers using the Chelex method . Plasmodmolecule . Genotyp the WHO .The sample size of 50 patients , i.e., a minimum of 60 patients, was calculated on the basis of an expected failure rate of 5% extrapolated from data obtained in Senegal, 10% precision and a confidence level of 95% .t-test. Proportions were arranged in a 2\u2009\u00d7\u20092 contingency table and compared using Fisher\u2019s exact test. All statistical tests were two-sided, and the level of statistical significance was fixed at P <0.05. SigmaStat 3.5  software was used for these analyses.Haemoglobin values on day 0 and day 28 were compared using the paired t-test. The significance level was fixed at P <0.05.As recommended in the WHO protocol , per proad hoc Mauritanian national ethics committee and the ethics committee of the WHO . The purpose of the study was explained to the patients  in local dialect. Informed written consent was obtained from adult patients or caretakers of paediatric patients aged under 12\u00a0years. For adolescents aged between 12 and 18\u00a0years old, an informed written consent was obtained from both the patients themselves and their parents or legal guardians.The study was reviewed and approved by an P >0.05) in the mean age and body weight of the two patient populations in Kobeni and Timbedra. However, the mean body temperature and parasitaemia were significantly higher (P <0.05) in Kobeni than in Timbedra. This observation was probably related to the fact that the majority of patients seen in Kobeni health centre  were febrile at the time of consultation, whereas in Timbedra most patients  presented with a recent history of fever but were afebrile at the time of consultation and enrollment.A total of 130 patients (65 patients in each study site) were enrolled in the study , or withdrawal of consent (n\u2009=\u20092), and eight (6.2%) were lost to follow-up  were excluded after inclusion for protocol violation (n\u2009=\u20093), repeated vomiting (n\u2009=\u20093), mixed sent n\u2009=\u2009, and eigIn Kobeni, per protocol analysis showed ASAQ efficacy of 96.6% before PCR correction. There were two cases of LCF, one occurring on day 5 and the other on day 27. The patient responding with LCF on day 5  was a 20-month-old boy presenting with a high-grade fever (39.8\u00b0C), a parasitaemia of 17,890 asexual parasites/\u03bcL, and haemoglobin of 7.8\u00a0g/dL on day 0. He was afebrile from day 1 to day 3 and had a positive smear on day 2  and a negative smear on day 3, but was seen on day 5 with a high-grade fever of 39.6\u00b0C and a positive smear . There were no signs and symptoms of clinical aggravation. The patient was treated with artemether-lumefantrine. The other patient responding with LCF was an eight-year-old boy presenting with a high-grade fever of 40.2\u00b0C and 17,140 asexual parasites/\u03bcL on day 0. He initially responded well to ASAQ treatment. He was afebrile from day 1 to day 21 and had a parasitaemia of 460 asexual parasites/\u03bcL on day 2 and negative smears from day 3 to day 21. On day 27, the patient was febrile (37.6\u00b0C) and had a positive smear . PCR analysis showed that the recurrence of fever and positive smear on day 27 was due to re-infection. After PCR adjustment, ASAQ efficacy was 98.2% in Kobeni.In Timbedra, the PCR-unadjusted outcome also showed high efficacy (98.2% ACPR) in the per protocol population, with only one LCF occurring on day 21 in a seven-year-old boy. This patient presented with fever (38.2\u00b0C) and 1,782 asexual parasites/\u03bcL on day 0. He was afebrile from day 1 to day 14 and aparasitaemic from day 2 to day 14. On day 21, the patient was seen with a low-grade fever (37.7\u00b0C) and 432 asexual parasites/\u03bcL. PCR analysis showed recrudescence.The overall efficacy of ASAQ was 98.2% after PCR correction (56 of 57 patients with ACPR (one patient was censored due to re-infection) in Kobeni and 54 of 55 patients with ACPR in Timbedra) Figures\u00a0. In bothASAQ was well-tolerated. Thirty-three of 130 patients (25.4%) reported the following mild and transient adverse effects : nausea or vomiting , headache , dizziness , anorexia , pruritus , and palpitation . Three patients with LCF (two in Kobeni and one in Timbedra) did not report vomiting or any other adverse effects. Although it is difficult to distinguish between malaria-associated symptoms and drug-induced adverse effects, the adverse events reported by the patients resolved spontaneously within 24\u00a0hours following administration of the first treatment dose. Pruritus that occurred on day 3 in one patient disappeared on day 4 without any specific treatment. No serious adverse effect was observed.P <0.05) increase in the mean (\u00b1 SD) haemoglobin value from 9.94\u2009\u00b1\u20091.66\u00a0g/dl on day 0 to the mean (\u00b1 SD) haemoglobin value of 11.2\u2009\u00b1\u20091.5\u00a0g/dl on day 28. The improvement in haemoglobin values is another benefit of a rapid and effective anti-malarial treatment for patients, in particular in young African children who tend to suffer from malaria-associated anaemia more than adults.Paired haemoglobin values were available from all 56 patients followed until day 28 in Kobeni. There was a statistically significant (The rapid and high efficacy of ASAQ observed in the present study is comparable to that found in the neighbouring countries, Senegal and Mali \u201316. The"}
+{"text": "Plasmodium vivax mono-infections were enrolled in a prospective, open-label, randomized trial to receive either chloroquine or dihydroartemisinin\u2013piperaquine (DHA-PPQ). The proportions of patients with adequate clinical and parasitological responses were 47% in the chloroquine arm (31 of 65 patients) and 66% in the DHA-PPQ arm (42 of 63 patients) in the Kaplan\u2013Meier intention-to-treat analysis , thus establishing non-inferiority of DHA-PPQ. Fever clearance time , parasite clearance time , and parasite clearance half-life  were all significantly shorter in the DHA-PPQ arm. All cases of recurrent parasitemia in the chloroquine arm occurred from day 33 onward, with corresponding whole blood chloroquine concentration lower than 100 ng/mL in all patients. Chloroquine thus remains efficacious for the treatment of P. vivax malaria in southern Vietnam, but DHA-PPQ provides more rapid symptomatic and parasitological recovery.A total of 128 Vietnamese patients with symptomatic  Plasmodium vivax is the second most prevalent malaria in the world. It is estimated that 2.85 billion people live at risk of infection.Plasmodium vivax is endemic throughout the tropics except for much of sub-Saharan Africa. The global burden of P. vivax malaria is estimated to range from \u223c70 to 80 million cases annually, with about 80\u201390% of cases occurring in the Middle East, Asia, and the Western Pacific.2P. vivax has been documented in Columbia, Peru, Bolivia, Brazil, Guyana, Turkey, Ethiopia, Madagascar, India, Myanmar, Thailand, Vietnam, Indonesia, Papua New Guinea, and South Korea.Plasmodium falciparum and P. vivax are the most widespread malaria species in Vietnam, representing 61% and 34% of all cases, respectively.P. vivax malaria in all countries except Indonesia, Papua New Guinea, the Solomon Islands, Vanuatu, and Cambodia where artemisinin combination therapies (ACTs) have been adopted since 2009.P. vivax on or before day 28, or prophylactic failure, has been observed in Asia, Africa, and Latin America.P. vivax in Vietnam.P. vivax was detected.P. vivax in other provinces in Vietnam such as Ninh Thuan and the Central, is of concern.14Chloroquine-resistant P. falciparum malaria in southeast AsiaP. vivax malaria. Dihydroartemisinin\u2013piperaquine (DHA-PPQ) is the first-line treatment of P. falciparum malaria in Vietnam. Clinical studies have shown that DHA-PPQ clears P. vivax infections more quickly than chloroquine.P. vivax infections and to evaluate the efficacy of DHA-PPQ, we conducted a randomized controlled trial in patients with P. vivax mono-infections in Vietnam.Recent worsening of antimalarial drug resistance of The study took place at Bu Gia Map and Dak O communes of Bu Gia Map District, Binh Phuoc Province, an area of high malaria transmission in Vietnam. Patients \u2265 3 years old with uncomplicated vivax malaria were eligible for enrollment if they had mono-infection with parasitemia \u2265 250/\u03bcL asexual forms, had axillary or tympanic temperature \u2265 37.5\u00b0C (or history of fever during the past 24 hours) and if they or their guardians gave fully informed consent. Patients were excluded if they had febrile conditions due to diseases other than malaria or other known underlying chronic or were severely ill, were on regular medications that may interfere with antimalarial pharmacokinetics, had received antimalarial drug in the previous 48 hours, had a history of hypersensitivity or contraindications to study drugs, had splenectomy, or were in the first trimester of pregnancy.P. vivax malaria.This study was a prospective open-label randomized comparison of the efficacy of DHA-PPQ versus chloroquine in Patients were randomized to receive either chloroquine (25 mg base/kg for 3 days) or DHA-PPQ (dihydroartemisinin 40 mg + piperaquine phosphate 320 mg per tablet) for 3 days; doses depend on body weight according to National Guidelines.DHA-PPQ was sourced from OPC pharmaceutical company  (under the brand name CV Artecan) in Vietnam and chloroquine was provided by the Institute of Malaria-Parasitology-Entomology as part of the National Malaria Program.r2 > 0.99. The limit of detection was 4 ng/mL and limit of quantification was 10 ng/mL in both plasma and blood for both chloroquine and MCQ. The intra-, inter- and total assay precisions were less than 10% for chloroquine and MCQ in plasma and whole blood samples. In plasma, the accuracies varied between 101% and 103% for chloroquine and MCQ; whereas in whole blood, the accuracies ranged from 97.0\u2013102% for chloroquine and MCQ. The mean recoveries of chloroquine and MCQ were 89.58\u201391.24% and 84.22\u201391.32% for plasma and 77.74\u201382.06% and 75.88\u201379.76% for blood, respectively. Furthermore, the recovery of quinine  in all validation batches was 90.19% (relative standard deviation = 2.46%) for plasma and 91.05% (relative standard deviation = 4.17%) for whole blood.Chloroquine and monodesethylchloroquine (MCQ) were measured in both whole blood and plasma of patients on the day of enrollment, days 7, 28 and at the time of recurrent parasitemia. Drug levels were determined by high-performance liquid chromatography (HPLC). In brief, the liquid chromatography system was a LaChrom Elite  controlled by EZChrom Elite version 3.18 HPLC System Manager Software (Merck\u2013Hitachi). Solid phase extraction (SPE) on Isolute-96-CBA  was used to process 100 \u03bcL of plasma/whole blood samples. After the SPE procedure, 50 \u03bcL of reconstituted solution was injected into HPLC system. Chloroquine, MCQ, and quinine  were separated using a mobile phase consisting of phosphate buffer 25 mM (pH 2.60) and acetonitrile  with 2 mM sodium perchlorate on a ZORBAX SB-CN 150 \u00d7 4.6-mm, 5-\u03bcm column equipped with 5-\u03bcm guard cartridges ZORBAX SB-CN 12.5 \u00d7 4.6 mm  at a flow rate of 1.2 mL/minute at ambient temperature in 10 minutes. The retention time of chloroquine was 4.62 minutes, MCQ was 5.95 minutes, and that of quinine  was 7.50 minutes. The diode array detector was set at a wavelength of 343 nm. The method was linear over the range of 10\u20135,000 ng/mL for both chloroquine and MCQ in plasma and whole blood with \u00a9  screening test, was also performed on day 0.Parasitemia and hematocrit were determined every 6 hours until parasite clearance, and then on days of follow-up. Malaria blood films were stained with Giemsa solution, and parasites counted against 500 white blood cells or against 1,000 red blood cells. Two qualified microscopists read all the slides independently, and parasite densities were calculated by averaging the two counts. A glucose-6-phosphate dehydrogenase (G6PD) deficiency rapid diagnostic test, the CareStartP. vivax. Follow-up activities included clinical assessment, measurement of temperature, hematocrit, parasitological assessment, and alternative treatment in case of treatment failure.Patients were admitted to the communal health stations for at least 48 hours or longer until two consecutive blood smears were negative for parasites. A clinical, microscopy and hematological assessment was performed on day 3 (at 72 hours), day 7, and then once a week until day 63 for follow-up of P. falciparum malaria.21The primary endpoint of this study was the proportion of patients classified as having an adequate clinical and parasitological response during a follow-up period of 63 days. In addition, the proportions of early treatment failures, late clinical failures, and late parasitological failures, as well as the time to treatment failure, were also reported as secondary endpoints. Adequate clinical and parasitological response, early treatment failure, late clinical failure, and late parasitological failure were defined using the same criteria as for the assessment of T1/2, that is, the time for parasitemia to fall by half during the log-linear phase of parasite clearance as defined by the Parasite Clearance Estimator developed by the Worldwide Antimalarial Resistance NetworkP. falciparum only and hence usage for P. vivax is exploratory); 2) parasite clearance time, defined as the time in hours from the first treatment dose to the first of two consecutive parasitemia counts below the detection limit of 20 parasites/\u03bcL (patients without documented parasitemia clearance were censored at the time of the last measured positive parasite count); 3) proportion of patients with a parasite clearance time > 48 hours after starting treatment; and 4) fever clearance time, defined as the time in hours from the first treatment dose to the start of the first sustained period of 24 hours without fever .Other secondary outcomes included 1) parasite clearance half-life The trial was powered to demonstrate the non-inferiority of DHA-PPQ as compared with the standard of care of chloroquine treatment with respect to the primary endpoint, the proportion of patients with an adequate clinical and parasitological response on day 63. Assuming an identical adequate clinical and parasitological response probability of 90% on day 63 in both treatment groups, a non-inferiority margin of 10%, a one-sided significance level of 2.5%, and an 80% power, a minimum of 142 patients per study arm were required. To accommodate losses to follow-up and protocol violations, the sample size was increased by 16%, giving a target sample size of 330 patients .P values for testing superiority of DHA-PPQ were also reported. The parasite clearance half-life was compared between the two arms using the t test. Parasite and fever clearance times were summarized with Kaplan\u2013Meier estimates of median and interquartile range (IQR), and comparisons between groups were based on the score test of a Cox-regression analysis with the treatment arm as the only covariate. All endpoints were reported and compared on the intention-to-treat (ITT) population containing all randomized patients. The primary endpoint was additionally reported for the per-protocol population, which excluded all patients who were withdrawn or lost to follow-up at any time point during the 63-day follow-up. The reported standard estimates, two-sided 95% CIs, and tests do not account for the fact that the trial was stopped early. All analyses were performed with the statistical software R3.2.0 .Following World Health Organization guidelines,`The study was approved by the Ethical Committee of the Ho Chi Minh Institute of Malaria-Parasitology-Entomology and the Oxford University Tropical Research Ethics Board. Clinicaltrial.gov registration number: NCT01887821.P. vivax malaria.When the study started in early 2013 radical treatment was not routinely provided, as primaquine was not routinely available. Availability improved substantially during the trial and new national guidelines on malaria treatment were introduced. As a result the trial steering committee decided to terminate the trial and provide radical treatment to all patients with The trial started in February 2013 and stopped in November 2014 after 128 of 330 planned patients had been enrolled. The proportion of patients with positive smears among those presenting to the health stations during the above period was 374/2,144 (17.4%). There were 65 patients enrolled in the chloroquine arm and 63 in the DHA-PPQ arm. Follow-up to day 63 or day of failure  was achieved in 49/65 (75.4%) patients treated with chloroquine and in 52/63 (82.5%) patients given DHA-PPQ . No early treatment failures were observed in either treatment arm. Late clinical failures and late parasitological failures occurred in 10 and 18 patients in the chloroquine arm compared with five and 14 in the DHA-PPQ arm. The earliest failure events were one late parasitological failure in the DHA-PPQ arm on day 28 and one late clinical failure on day 33 in the chloroquine arm. The rate of recurrent infections was significantly lower in the DHA-PPQ group  . The median (IQR) parasite clearance time were 36  hours in the chloroquine arm and 18  hours in the DHA-PPQ arm (P < 0.001). The proportions of patients with parasite clearance time > 48 hours were 25% in the chloroquine arm and 3% in the DHA-PPQ arm (P < 0.001). Median (IQR) fever clearance time were 24  hours in the chloroquine arm and 12  hours in the DHA-PPQ arm. Fever clearance was significantly faster in the DHA-PPQ arm .The mean parasite clearance half-lives were 3.98 hours in the chloroquine arm and 1.80 hours in the DHA-PPQ arm  patients had detectable chloroquine and MCQ at low concentrations in whole blood . By day 28, only 4/54 patients had whole blood chloroquine and MCQ concentrations \u2265 100 ng/mL , the putative minimal effective concentration for chloroquine-sensitive The treatments were well tolerated, and no serious adverse events were recorded in either arm.P. vivax malaria for over 50 years. Few studies have been conducted to assess the efficacy of chloroquine against P. vivax. Data from 1995 in southern, central Vietnam showed that of the evaluable P. vivax patients, 23/23 (100%) had sensitive infections,P. vivax cases, also conducted in central Vietnam (2009), there were 9.4% recurrences.13In Vietnam, chloroquine has been the drug of choice for s, 23/23 0% had seP. vivax malaria. DHA-PPQ was definitely not inferior to chloroquine and in some therapeutic aspects was superior. Fever and parasite clearance were more rapid, and overall recurrence rates were lower.17Our 63-day study at two sentinel sites in an area of moderately high malaria transmission in southern Vietnam confirmed that both chloroquine and DHA-PPQ are well-tolerated and efficacious treatments for P. falciparum in Vietnam over many years showed that this is a safe and well-tolerated regimen. If DHA-PPQ is accepted for the treatment of both P. falciparum and P. vivax, the procurement and distribution of antimalarials of the national malaria control/elimination program will be simplified.Experience from the use of DHA-PPQ in the treatment of There were 47 \u201clate treatment failures\u201d in this study which, except for one recurrent parasitemia on day 28 in the DHA-PPQ arm, happened after day 35 when blood concentrations of both drugs had declined. Primaquine was not given in this study, and so it is difficult to determine the nature of these recurrences . According to Baird, recurrent parasitemia within 35 days of chloroquine therapy supports a provisional diagnosis of resistance.P. vivax malaria are often lower as G6PD deficiency provides some protection. All the patients in our study received 14-day primaquine on day 63 or day of recurrent parasitemia (except the one G6PD deficiency case mentioned above), and there was no hemoglobinuria or other adverse effects.Regarding G6PD status among enrolled patients, only one case of deficiency was recorded in a male of S'Ti\u00eang ethnic minority. Confirmation of G6PD status using the standard enzyme-linked immunosorbent assay method in our study population is in progress and results will be reported later. Previous studies have shown that G6PD deficiency determined by the methylene blue reduction test and by G-6-PDH kit  was 8.7% (23/266) in Kinh ethnic people and 14% 36/258) in S'Ti\u00eang people, respectively,8 in S'TiP. vivax malaria but that DHA-PPQ offers clinical and operational advantages.Taken together, these findings suggest that chloroquine is still an effective treatment of P. falciparum and P. vivax malaria may be a concern for policy makers, but in Vietnam the cost difference would not be large because DHA-PPQ is locally produced. In any case, the clinical and operational benefits of a single simple effective treatment would outweigh the disadvantages.In recent years, benefits and disadvantages of using ACTs, especially combinations with partner drugs that have long half-lives , have been reviewed.P. vivax malaria in southern Vietnam. DHA-PPQ, not inferior to chloroquine in efficacy, is an alternative treatment with many advantages over chloroquine: shorter parasite clearance half-life, parasite clearance time, and fever clearance time and lower proportion of parasite clearance time > 48 hours.To conclude, chloroquine remains efficacious for the treatment of"}
+{"text": "Plasmodium falciparum has been confirmed in the Greater Mekong subregion (GMS). Dihydroartemisinin-piperaquine (DAPQ) is the most commonly used ACT in China. To understand the DAPQ sensitivity of P. falciparum, DAPQ resistance was monitored in vivo along the China-Myanmar border from 2007 to 2013.Artemisinin-based combination therapy (ACT) is the recommended first-line treatment of falciparum malaria in all endemic countries. Artemisinin resistance in P. falciparum were recruited to this study after obtaining full informed consent. DAPQ tablets for different categories of kg body weight ranges were given once a day for three days. Patients were followed up for 42\u00a0days. Polymerase chain reaction (PCR) was conducted to distinguish between re-infection and recrudescence, to confirm the Plasmodium species. The data were entered and analysed by the Kaplan-Meier method. Treatment outcome was assessed according to the WHO recommended standards.Eligible patients with mono-infections of 2\u2009=\u20092.81, P\u2009=\u20090.7288).243 patients were completed valid follow-up. The fever clearance time (FCT) and asexual parasite clearance times (APCT) were, respectively, 36.5\u2009\u00b1\u200910.9 and 43.5\u2009\u00b1\u200911.8\u00a0hours, and there was an increasing trend of both FCT  and APCT  from 2007 to 2013. Eight  patients present parasitaemia on day three after medication; however they were spontaneous cure on day four. 241  of the patients were adequate clinical and parasitological response (ACPR) and the proportions of ACPR had not changed significantly from 2007 to 2013 (XP. falciparum had not significantly changed along the China-Myanmar border of Yunnan Province, China. However more attentions should be given to becoming slower fever and parasite clearance.In terms of efficacy, DAPQ is still an effective treatment for falciparum malaria. DAPQ sensitivity in  Plasmodium falciparum has been identified and confirmed in Cambodia, Thailand, Myanmar, and Vietnam [P. falciparum is one of important components to prevent the emergence of new foci of resistance, as well as to limit the spread of resistance [Malaria remains one of the major global public health problems . Intensi Vietnam -10. Othe Vietnam . The art Vietnam  and no osistance .P. falciparum, were reported along the border in China, and the annual parasite incidence was only 0.9  per ten thousand. Monitoring artemisinin resistance in Yunnan Province would contribute both to malaria elimination in China and in containing global artemisinin resistance. From 2007 to 2013, DAPQ resistance was monitored in vivo to determine the dynamics of P. falciparum sensitivity to DAPQ on the China- Myanmar border.China has declared a national policy for malaria elimination by 2020 ,13. ACT P. falciparum among slides with parasites was 38.1%  in the five Special Regions of Myanmar. In eliminating settings, malaria predominantly occurs in border areas and imported cases tend to represent a majority of recorded cases [China and Myanmar share 2,185 kilometre border. There are 19 counties of China and fives special regions of Myanmar on the border. Populations includes 4,687, 896 residents of 19 counties of Yunnan Province of China and 586,000 residents of the five Special Regions of Myanmar. Due to the complex emergency situation in the five Special Regions of Myanmar, where fell outside the coverage of national malaria control efforts supported by Myanmar\u2019s Ministry of Health, effective access of malaria interventions could only be achieved from China. In March, 2008, the baseline survey of evaluation indicator for the sixth grant to China of the Global Fund to fight AIDS, Tuberculosis and Malaria showed that the parasite prevalence rate was 13.6% , and the proportion of P. falciparum were recruited to this study, after obtaining full informed consent. Only patients older than one year, body weight\u2009\u2265\u20095\u00a0kg, presenting with parasite density 500\u2013100,000 parasites per \u03bcl were enrolled into the study. Imported malaria was identified as patients who had travelled from endemic areas of Myanmar within one month and were diagnosed as malaria in China [Patients whose axillary temperature was\u2009\u2265\u200937.5\u00b0C or with a history of fever during the previous 24\u00a0hours were diagnosed by using microscopy of thick and thin blood smears. Patients with mono-infections of in China . Patientin China ,21.DAPQ was manufactured by Zhejiang Holley Nanhu Pharmaceutical Co. Ltd, and provided by the West Pacific Office of WHO (WPRO/WHO). The drug quality was controlled by WPRO/WHO too. The batch numbers of DAPQ were 600807 (manufactured on 30 Aug 2007), 460909 (23 Sep 2009) and 400911 (20 Sep 2011). Each tablet contains 40\u00a0mg base dihydroartemisinin and 320\u00a0mg piperaquine phosphate. DAPQ was given once a day for three days and the dosing were based on the recommendation of WPRO/WHO. For convenient administration, the doses were calculated into tablets for different categories of kg body weight ranges , late clinical failure (LCF), late parasitological failure (LPF), and adequate clinical and parasitological response (ACPR). The ETF definition was to conform to any one of the criteria: (1) danger signs or severe malaria on day 1, 2 or 3, in the presence of parasitaemia; (2) parasitaemia on day 2 higher than on day 0, irrespective of axillary temperature; (3) parasitaemia on day 3 with axillary temperature\u2009\u2265\u200937.5\u00b0C; and, (4) parasitaemia on day 3\u2009\u2265\u200925% of count on day 0. The LCF definition was to satisfy any one of the criteria: (1) danger signs or severe malaria in the presence of parasitaemia on any day between day 4 and day 42 in patients who did not previously meet any of the criteria of ETF; and, (2) presence of parasitaemia on any day between day 4 and day 42 with axillary temperature\u2009\u2265\u200937.5\u00b0C in patients who did not previously meet any of the criteria of ETF. LPF definition is to satisfy presence of parasitaemia on any day between day 7 and day 42 with axillary temperature\u2009<\u200937.5\u00b0C in patients who did not previously meet any criteria of ETF or LCF. ACPR definition was to satisfy absence of parasitaemia on day 42, irrespective of axillary temperature, in patients who did not previously meet any criteria of ETF, LCF or LPF ,21.The data were by double independent data entry and analysed by the Kaplan-Meier method ,26. The According to the Helsinki Declaration, ethical approval for the study was granted by the Ethics Committee of Yunnan Institute of Parasitic Diseases, China. The purpose of the study was explained and then approval was sought from patients and their caretakers. Informed written consent was obtained from patient or carers of Child patients. All results were kept confidential and were unlinked to any identifying information.2\u2009=\u20092.81, P\u2009=\u20090.7288) , 22(7.6%) withdrew, 25(8.6%) were lost at follow-up, and 243 (83.8%) completed valid follow-up, of which 178 (72.3%) were Burmese .orldwide ,29. The orldwide ,28-30. Torldwide . The marP. falciparum had been monitored on the China-Myanmar border. Sun et al. reported that DAPQ was efficacious for falciparum malaria treatment in 2006 [et al. reported a 97.0% of cumulative success rate (CSR) and reduced DAPQ sensitivity in P. falciparum of a trial in which the only eight tablets of DAPQ (a dosage for >40\u00a0kg body weight) were used for uncomplicated falciparum malaria treatment [et al., the technical limitation was to not use PCR to distinguish between re-infection and recrudescence, and they conduct their study in Wa State of Myanmar, where P falciparum prevalence was high during 2007\u20132008. The technical limitation and the different regimen might result in the declined CSR, and geographic differences might be one of explanations for reported differences too. Huang et al. reported a 95.9% (47/49) of artesunate CSR and a high frequency of mutations in pfcrt, pfdhfr and pfdhps associated with chloroquine and sulphadoxine\u2013pyrimethamine resistance and no pfatp6 mutation in P. falciparum [et al. reported low levels of point mutations in pfmdr1 and pfatp6 prevalence and no pfmdr1 gene amplification detected [P. falciparum along the China-Myanmar border.DAPQ sensitivity of  in 2006 , howeverreatment . In the lciparum . Wang etdetected . The resPF3D7_1343700 kelch propeller domain (K13-propeller) are important determinants of artemisinin resistance [P. falciparum along the China-Myanmar border.Several limitations should be considered while using the results of study. Firstly, both total and yearly sample size were limited by the difficulty in recruiting patients because of low malaria incidence in eliminating settings, only total 1011 falciparum malaria cases were reported during 2010\u20132013. Secondly, the surveillance activity was interrupted in 2008 because of shortage of funding. Thirdly, the surveillance sites had to be changed in order to recruit falciparum malaria patients, in Yingjiang in 2007, Menglian in 2009, in both Tengchong and Yingjiang from 2010 to 2013, however all the three sites are on the China-Myanmar border. Fourthly, the study did not use any pharmacokinetics (PK) measurement to ascertain drug absorption and to characterize the concentration\u2013time profile of drugs and the relevant covariates , so it did not exclude confounding effect of some variables that could influence APCT . Fifthlysistance ; the stusistance ,39. The P. falciparum was still sensitive to artemisinins on the China-Myanmar border. The genetic diversity of malaria parasites and multiclonal infections are correlated with transmission intensity as well as the spread of anti-malarial resistance. Despite the intensified control efforts and the decline of malaria prevalence along China-Myanmar border, the parasite population size and transmission intensity remained high enough to allow effective genetic recombination of the parasites and continued maintenance of genetic diversity in China-Myanmar border area [P. falciparum should be maintained and containment activities should be further strengthened to monitor the spread of resistance, define therapeutic and operational strategies to understand its molecular basis and counter its impact [While artemisinin resistance was found on the western border of Cambodia-Thailand and the western border of Thailand-Myanmar , P. falcder area . Therefos impact ,42. In ts impact , so it mP. falciparum treatment had been reported in Hainan Province of China in 2004 [P. falciparum has been eliminated by using the strategy of early diagnosis and treatment (EDT) with ACT [P. falciparum, the strategy of EDT with ACT has reduced malaria in the migrant population living on the Thai-Myanmar border [P. falciparum. Lower doses of the gametocytocide would be safer, might still be very effective for blocking transmission.Detection of drug efficacy is only the first step to producing accessible and useful information for decision makers. The translation of increased access to data on health outcomes into usable evidence for rational policy and planning requires a global coordination and communication effort . In term in 2004  and then in 2004 , P. falcwith ACT . The emer border . The addr border . DespiteP. falciparum had not significantly changed along the China-Myanmar border of Yunnan Province, China. However more attentions should be given to becoming slower fever and parasite clearance, surveillance for artemisinin resistance in P. falciparum should be maintained and containment measures are urgently needed.In terms of efficacy, DAPQ is still effective for falciparum malaria treatment. DAPQ sensitivity in"}
+{"text": "Plasmodium falciparum malaria had biochemical evidence of thiamin deficiency, which was associated with a higher incidence of adverse events. Thiamin supplementation might, therefore, reduce adverse events in this population.In a recent study one third of Lao patients presenting with uncomplicated An exploratory, double-blind, parallel group, placebo-controlled, superiority trial of thiamin supplementation in patients of all ages with uncomplicated and severe falciparum malaria was conducted in Xepon District, Savannakhet Province, southern Laos. Patients were randomly assigned to either oral thiamin 10\u00a0mg/day for 7\u00a0days immediately after standard anti-malarial treatment then 5\u00a0mg daily until day 42, or identical oral placebo.After interim analyses when 630 patients (314 in thiamin and 316 in placebo groups) had been recruited, the trial was discontinued on the grounds of futility. On admission biochemical thiamin deficiency  was present in 27% of patients and 9% had severe deficiency . After 42\u00a0days of treatment, the frequency of thiamin deficiency was lower in the thiamin  compared to the placebo  groups (p\u2009<\u20090.001 and p\u2009=\u20090.05), respectively. Except for diarrhoea, 7% in the placebo compared to 3% in the thiamin group (p\u2009=\u20090.04), and dizziness on day 1 , all adverse events were not significantly different between the groups (p\u2009>\u20090.05). Clinical, haematological, and parasitological responses to treatment did not differ significantly between the two groups.Thiamin supplementation reduced biochemical thiamin deficiency among Lao malaria patients following anti-malarial drug treatment, but it did not reduce the frequency of adverse events after anti-malarial therapy or have any detected clinical or parasitological impact.ISRCTN 85411059 Plasmodium falciparum malaria from chloroquine (CQ) and sulphadoxine-pyrimethamine (SP) to artemisinin-based combination therapy (ACT) (artemether-lumefantrine), which has been highly efficacious , 314 and 316 received thiamin and identical placebo, respectively  with High-performance liquid chromatography (HPLC), using the method of the United States Pharmacopeia , of threPatient admission demographic, clinical and laboratory details differed little between the thiamin and placebo groups Table\u00a0. All patt-test p\u2009<\u20090.001 for both groups, Table\u00a0Fever and parasite clearance times, post-treatment haematocrit at all time points and gametocyte rates were not significantly different between patients who received thiamin and placebo Table\u00a0. BaselinP. falciparum reappearance, PCR genotyping indicated that 17/50 (34%) had recrudescent infections . One patient in placebo group with P. falciparum reappearance had an indeterminate PCR result and, considering this patient as having a recrudescent infection, the overall 42-day cure rates per protocol, excluding patients who were lost to follow up, withdrew consent, had persistent vomiting, or re-infection, were 97% (599/617)  . Of 18 patients (6 in thiamin and 12 in placebo groups) considered as having recrudescent infection, 16 (6 in thiamin and 10 in placebo groups) had parasite recurrence at or before day 28. The 28-day cure rates by ITT analysis were 308/314 (98%) for thiamin and 306/316 (97%) for placebo groups. The median (range) interval to recrudescent falciparum infections was 21 (14\u201342) days. By ITT and per protocol analyses, the 42-day PCR uncorrected cure rates for thiamin and placebo groups were 294/314 (94%) 0 . The mean (95%CI) day 42 alpha values were significantly lower in the thiamin [12 (11\u201313)%] compared to the placebo [17 (16\u201317)%] groups (p\u2009<\u20090.001). The overall proportion of patients with thiamin deficiency at day 42 was 6% and with severe thiamin deficiency was 2% and was significantly lower in the thiamin  compared to the placebo  groups (p\u2009<\u20090.001 and p\u2009=\u20090.05), respectively.t-test p\u2009<\u20090.001); -7.8 % in the thiamin group and -3.6 % in the placebo group (paired t-test p\u2009<\u20090.001 for both). The mean (95%CI) changes in basal and activated ETK (micromoles/min/gHb) values between the thiamin versus placebo groups were  and , respectively.Following anti-malarial and thiamin/placebo treatments, there was a significant decrement in alpha. For 614 patients with paired admission and day 42 samples, the overall mean  percentage change in alpha was -5.7 % (paired vs 66/313 (21%), p\u2009=\u20090.001] and , respectively. Mean alpha values and the proportion of the patients with thiamin deficiency on admission and at day 42 were not statistically different between pregnant and non-pregnant women (p\u2009>\u20090.05). The median (range) basal and activated ETK on admission and at day 42 were significantly higher in severe than in uncomplicated malaria patients (p\u2009<\u20090.01). The proportion of patients with thiamin deficiency on admission  was significantly lower in severe [3/34 (9%)] compared to uncomplicated [166/596 (28%)] malaria (p\u2009=\u20090.01) but were similar for severe [1/30 (3%)] and uncomplicated [37/584 (6%)] groups at 42-day follow-up (p\u2009=\u20091.00).Higher admission and day 42 basal and activated ETK but lower admission and day 42 alpha values were observed among children compared to adults . The proportion of patients with thiamin deficiency  was also significantly higher in the adults [89/216 (41%)] compared to children [80/414 (19%)] on admission (p\u2009<\u20090001) and [24/209 (11%)] vs [14/405 (3%)] at day 42 (p\u2009<\u20090.001). The proportion of patients with thiamin deficiency  on admission and at day 42 was significantly higher in males than in females . The mean glucose at day 42 was not different between the groups but the mean (95%CI) lactate (mmol/L) at day 42 was slightly lower in the thiamin [2.5 (2.4 \u2013 2.6)] than in the placebo [2.7 (2.6 \u2013 2.8)] groups, (p\u2009=\u20090.048).The mean (95%CI) whole blood glucose and lactate on admission was similar between thiamin and placebo groups Table\u00a0. Followiversus day 42 mean (95%CI) glucose (mmol/L) and lactate (mmol/L) among this group of patients was  and , respectively.Among patients with severe malaria (n\u2009=\u200934), the mean glucose and lactate also significantly decreased following anti-malarial and thiamin/placebo treatments with the mean (95%CI) difference between admission and day 42 of 1.2 (0.6 \u2013 1.9) mmol/L for glucose and 2.4 (1.8 \u2013 3.0) mmol/L for lactate. The admission No consistent relationships were observed between lactate and basal and activated ETK or alpha at admission and day 42.No patient had obvious symptoms or signs suggesting thiamin deficiency on admission or during or after treatment. Adverse event symptoms were recorded for only those 461 (73%) patients aged >4\u00a0years old and able to answer questions Table\u00a0. The medFor both groups combined, the admission and day 42 median (range) basal and activated ETK were significantly higher in patients without any subsequent adverse events compared to those with at least one subsequent adverse effect . Admission mean (95%CI) alpha was significantly lower in patients without subsequent adverse events [18% (17-19%) than in the patients with least one adverse event [22% (21-24%)] (p\u2009<\u20090.001). The corresponding values at day 42 were 13% (12-14%) for those without adverse events and 16% (15-17%) for those with at least one adverse event (p\u2009<\u20090.001). The proportion of patients with thiamin deficiency  was significantly higher in those with at least one adverse event compared to those without adverse events  on admission and  at day 42, respectively. The proportion of the patients with at least one possible adverse event and the frequency of various side effects among the patients with severe malaria (n\u2009=\u200934) were not different between the thiamin and placebo groups (p\u2009>\u20090.05). Adverse events only in the first and last week were similar between thiamin and placebo groups .et al.[A large randomized, double-blind, parallel group, placebo-controlled trial to compare the incidence of adverse events between malaria patients who were supplemented with and without thiamin in addition to standard ACT treatment was conducted. No major differences were identified between the two groups so the trial was discontinued, following the interim analyses of data from 630 recruited patients. The incidence of adverse effects, including dizziness, headache and nightmare, and the proportion of the patients with one or more subsequent adverse effects was not statistically different between the thiamin and placebo groups, suggesting that oral thiamin supplementation does not reduce adverse effects during and after anti-malarial treatment. In contrast with the post-hoc analysis of data published by Mayxay Limitations of the study include a delay of 7\u201317 months between sample collection and ETK assays, the lack of a parenteral thiamin versus placebo arm for immediate treatment of those with severe malaria and the relative rarity of patients with severe malaria recruited to the study.et al. [et al. [In the study of Mayxay et al. , it was et al. . Earlieret al. . Indeed, [et al.  at 30% a [et al. . Severe Lao adults have multiple risk factors for thiamin deficiency including hard physical labour of rice farming, alcohol intake, consumption of polished rice and thiaminase-containing food, such as \u201cpaa dek\u201d (fermented fish paste), and thiamin antagonists such as betel nut. The significantly higher prevalence of biochemical thiamin deficiency on admission and at 42-day follow-up among adults (\u226515\u00a0years) compared to children aged <15\u00a0years old suggests that hard physical labour of the rural rice-farming adults may play an important role in biochemical thiamin deficiency .Thiamin supplementation did not reduce the frequency of adverse events after anti-malarial therapy among patients with falciparum malaria in southern Laos. Thiamin supplementation could help to reduce biochemical thiamin deficiency among malaria patients following anti-malarial drug treatment, but this did not have a measurable clinical or parasitological impact.The authors declare that they have no competing interests.MM designed the study, recruited and followed up the patients, analysed data and drafted paper. MK and OS designed the study, recruited and followed up the patients, and revised the paper. MI designed the study, performed the molecular genetic studies, and revised the paper. TP, BH, VV, SP and NJW designed the study and revised the paper. LC oversaw the measurement of ETK activity and haemoglobin in the lysed erythrocytes. PNN designed the study, analysed data, drafted and revised the paper. All authors read and approved the final manuscript."}
+{"text": "Plasmodium falciparum infections adversely affect pregnancy. Anti-malarial treatment failure is common. The objective of this study was to examine the duration of persistent parasite carriage following anti-malarial treatment in pregnancy.P. falciparum malaria cases. Women with microscopy confirmed P. falciparum malaria had a PCR blood spot from a finger-prick sample collected. Parasite DNA was extracted from the blood-spot samples using saponin lysis/Chelex extraction method and genotyped using polymorphic segments of MSP1, MSP2 and GLURP. Recurrent infections were classified by genotyping as novel, recrudescent or indeterminate. Factors associated with time to microscopy-detected recrudescence were analysed using multivariable regression techniques.The data presented here are a collation from previous studies carried out since 1994 in the Shoklo Malaria Research Unit (SMRU) on the Thailand-Myanmar border and performed using the same unique methodology detailed in the Materials and Methods section. Screening for malaria by microscopy is a routine part of weekly antenatal care (ANC) visits and therapeutic responses to anti-malarials were assessed in P. falciparum and there were 909 recurrent episodes (481 novel and 428 recrudescent) confirmed by PCR genotyping. Most of the recurrences, 85\u00a0% (770/909), occurred after treatment with quinine monotherapy, artesunate monotherapy or artesunate-clindamycin. The geometric mean number of days to recurrence was significantly shorter in women with recrudescent infection, 24.5 (95\u00a0%: 23.4-25.8), compared to re-infection, 49.7 (95\u00a0%: 46.9-52.7), P <0.001. The proportion of recrudescent P. falciparum infections that occurred after days 28, 42 and 63 from the start of treatment was 29.1\u00a0% (124/428), 13.3\u00a0% (57/428) and 5.6\u00a0% (24/428). Recrudescent infections \u2265100\u00a0days after treatment occurred with quinine and mefloquine monotherapy, and quinine\u2009+\u2009clindamycin and artesunate\u2009+\u2009atovaquone-proguanil combination therapy. Treatments containing an artemisinin derivative or an intercalated Plasmodium vivax infection increased the geometric mean interval to recrudescence by 1.28-fold (95\u00a0% CI: 1.09-1.51) and 2.19-fold (1.77-2.72), respectively. Intervals to recrudescence were decreased 0.83-fold (0.73-0.95) if treatment was not fully supervised (suggesting incomplete adherence) and 0.98-fold (0.96-0.99) for each doubling in baseline parasitaemia.From December 1994 to November 2009, 700 women were treated for P. vivax infections treated with chloroquine. Accurate determination of drug efficacy in pregnancy requires longer duration of follow-up, preferably until delivery or day 63, whichever occurs last.Prolonged time to recrudescence may occur in pregnancy, regardless of anti-malarial treatment. Long intervals to recrudescence are more likely with the use of artemisinin-containing treatments and also observed with intercalated  Plasmodium falciparum are prevalent, frequent screening and treatment of all positive malaria episodes is required. This has been a focus area of the antenatal clinics (ANCs) of Shoklo Malaria Research Unit (SMRU) on the Thailand-Myanmar border [P. falciparum parasite recurrence was introduced to distinguish novel or new infections from recrudescent infections [P. falciparum at SMRU have been published within the context of treatment efficacy trials in pregnant women [P. falciparum anti-malarial drug trials is limited to 63\u00a0days [In order to reduce malaria-related maternal mortality in a low-transmission area where multidrug-resistant (MDR) strains of r border , 2. In 1fections . This isfections , 5. PCR nt women \u201311 and int women , 13. The 63\u00a0days . In preg 63\u00a0days , 187\u00a0day 63\u00a0days , 85\u00a0days 63\u00a0days  and 121\u00a0P. falciparum infections (South Carolina strain) in neurosyphilis patients. The average duration of infection was 222\u00a0days with three of the infections persisting for more than one year, and the longest 480\u00a0days. In a similar report with the Panama strain of P. falciparum there were 23 incompletely treated infections which persisted for an average of 280\u2009\u00b1\u200920 (range 114 to 503) days, with four cases persisting for more than one year [P. falciparum is well established [P. falciparum in an individual is 13\u00a0years [P. falciparum genotyping was used to examine the duration of persistent P. falciparum carriage in pregnant women and examine the factors associated with time to recrudescence.In the era of malaria therapy from the 1920s to the 1950s, and the volunteer studies conducted to assess new anti-malarial drugs conducted in Australia and the USA, natural infections in previously non-immune subjects were observed to last for many months. Transfusion malaria infections acquired from donors who had left the endemic area years previously were also documented . In the one year . These rablished . The lon13\u00a0years . Here P.Pregnant women in this series attended the weekly ANC of SMRU on the northwestern border of Thailand. In this hilly, forested area malaria transmission is low and seasonal  in the whole population, including pregnant women . AcquireThis study spans 15\u00a0years during which treatment of falciparum malaria has changed from monotherapy to artemisinin-based combination therapy (ACT)  for seven days; artesunate (outside the first trimester) once daily  for seven days ; artemether-lumefantrine : four tablets twice daily for three days, mefloquine 25\u00a0mg/kg stat dose or as a split dose (15\u00a0mg/kg and 10\u00a0mg/kg on consecutive days); dihydroartemisinin-piperaquine once daily for three days ; artesunate (4\u00a0mg/kg/day) combined to atovaquone-proguanil (atovaquone 20\u00a0mg/kg/day and 8\u00a0mg/kg/d) for three days . Since 2007, clindamycin was combined with artesunate or quinine at a dose of 5\u00a0mg/kg three times daily . Chloroquine  was given once a day for three days, with a dose of 10\u00a0mg base/kg/day for two days and 5\u00a0mg base/kg/day on the third day, for Contrary to the changes in treatment over time there has been no change in the unique methodology applied to pregnant women in this area including the screening by microscopy for malaria offered at every antenatal visit during pregnancy, in the basic data collected at the time of each malaria episode and in the way the sample is collected. At the time of each malaria episode the date, body temperature, history of fever in the previous 48\u00a0h, parasitaemia, haematocrit, gestational age of the pregnancy, and malaria history were recorded. After microscopy diagnosis and before treatment, fresh blood from a finger-prick sample was used to make three blood spots (approximately 30\u00a0\u03bcl each) on a strip of 3\u00a0M filter paper. This was dried overnight before DNA extraction the next day.Symptomatic malaria was defined as slide-confirmed parasitaemia with a history of fever in the previous 48\u00a0h or a measured axillary or aural temperature \u226537.5\u00a0\u00b0C . AsymptoFor recrudescence risk factor based analysis anti-malarial therapies were grouped into artemisinin based: including ACT and artesunate monotherapy and non-artemisinin based including combinations such as quinine and clindamycin or monotherapy such as mefloquine.Plasmodium falciparum infection from consecutive malaria episodes during pregnancy was analysed by using the three loci genotypes: MSP1, MSP2 and GLURP. Nested PCR was the amplification strategy used to genotype P. falciparum and this is explained in detail elsewhere [lsewhere , 27, 28.P. falciparum infection was positively skewed  transformed. Linear regression models were fitted to the loge transformed outcome, and either geometric means or ratios of geometric means were derived. Separate sub-group analyses were performed for women with recrudescent infection(s) and women with novel infection(s). In the multivariable linear regression analyses, a risk factor was deemed to be associated with the loge transformed outcome if the p-value was below the nominal significance level of 0.1; multi-collinearity between each of the covariates  was assessed using variance inflation factors ; and all estimates derived from the regression analyses were adjusted for age (years), weight (kg), and study period categorized into four three-year intervals . As resistance appears with increasing years of drug use, e.g., artemisinin, and as the cohort was conducted over many years with different numbers of women available in different years, four blocks were chosen for a reasonably even distribution of women in each block, so time could be controlled for in regression analysis. To account for the correlation between the number of days to recurrence of multiple P. falciparum infections during a single pregnancy, robust standard errors were calculated using the Huber-White sandwich estimator. Tests for linear trends were performed by fitting a linear model to the outcome variable with the categorical covariate treated as pseudo continuous. Proportions were compared using Pearson\u2019s Chi-squared test. Data were analysed in Stata/IC version 11.1 .Continuous variables were described using median (range) and categorical variables using frequency (percentage). The distribution of the number of days to recurrence of wed Fig.\u00a0 and therThe data presented here represent a collation of data from women enrolled in treatment or in one prevention trial in pregnancy all previously published and approved by the Faculty of Tropical Medicine Ethics Committee in Bangkok and in later years the Oxford Tropical Research Ethics Committee \u201311, 29.P. falciparum of which 19 were classified as indeterminate by PCR; seven failed to amplify; no PCR spot was found for two episodes and pregnancies with episodes of non-sequential pairs were removed, leaving 1,609 episodes for analysis in 700 women  had two and 54 (6.0\u00a0%) experienced three or more recurrences within the same pregnancy. The maximum number of consecutive P. falciparum episodes within a single pregnancy was five and all four recurrent infections could be characterized by PCR in two women. The demographic characteristics of the pregnant women at the time of the primary infection are reported in Table\u00a0From December 1994 to November 2009, there were 700 women with 1,647 episodes of Most of the recurrent infections, 85\u00a0% (770/909), occurred following treatment with one of the three most commonly used regimens: quinine monotherapy, artesunate monotherapy or artesunate-clindamycin, which are all rapidly eliminated seven-day treatments Table\u00a0. Not allP <0.001  was 24 (95\u00a0% CI: 23\u201326) days; first to the second recrudescence (N\u2009=\u200955) was 25 (95\u00a0% CI: 22\u201329) days; and, from the second to the third or more recrudescence (N\u2009=\u200920) was 28 (95\u00a0% CI: 22\u201335) days, P\u2009=\u20090.44. For recurrent episodes that were novel infections, the observed times were 50 (95\u00a0% CI: 46\u201353) days for primary to first (N\u2009=\u2009339); 51 (95\u00a0% CI: 46\u201358) days first to second (N\u2009=\u2009108); and, 45 (95\u00a0% CI: 38\u201353) days for second to third or more infection (N\u2009=\u200934), P\u2009=\u20090.38.The geometric mean number of days to recurrence was significantly shorter in women with recrudescent infection, 24.5 (95\u00a0%: 23.4-25.8) days, compared to re-infection, 49.7 (95\u00a0%: 46.9-52.7) days, P. falciparum recurrences, 19.3\u00a0% (176) had an intercalated Plasmodium vivax infection between the P. falciparum primary and recurrent episode. There were 145 recurrent P. falciparum episodes with one intercalated P. vivax episode; and 23, seven and one with recurrent P. falciparum episodes with two, three and five intercalated P. vivax episodes, respectively. The proportion of recrudescent P. falciparum infections following an intercalated P. vivax infection was significantly lower than those without an intercalated P. vivax episode: 19.9\u00a0% (35/176) compared with 53.6\u00a0% (393/733), P <0.001. In both novel and recrudescent infection, intercalated P. vivax infection resulted in a longer geometric mean number of days to recurrence compared to those without intercalated P. vivax: 71 (95\u00a0% CI: 65\u201378) versus 43 (95\u00a0% CI: 40\u201346) days, and 49 (95\u00a0% CI: 41\u201359) versus 23 (95\u00a0% CI: 22\u201324) (P <0.001 for both), respectively. The proportion of intercalated P. vivax infections was lower with medium- and long-acting anti-malarial treatments (Table\u00a0Of the 909 N\u2009=\u2009381), medium- (N\u2009=\u200922) and long- (N\u2009=\u20097) acting anti-malarials was 24 (95\u00a0% CI: 23\u201326), 28 (95\u00a0% CI: 25\u201333) and 33 (95\u00a0% CI: 26\u201343) days (linear trend: P\u2009=\u20090.02), when mefloquine monotherapy was excluded from the analysis . When mefloquine monotherapy was included, the geometric mean number of days to recrudescent infection for long-acting anti-malarials fell to 25 (95\u00a0% CI: 20\u201332) days and the linear trend was no longer significant (linear trend: P\u2009=\u20090.85). There was no evidence of a linear trend in number of days to novel infection for short-, medium- and long-acting anti-malarials.The geometric mean number of days to recrudescent infection(s) for short- (Nearly one third 29\u00a0% (124/428) of recrudescent infection occurred after day 28 with 13.3\u00a0% (57/428) and 5.6\u00a0% (24/428) occurring after day 42 and 63, respectively. The proportion of quinine treatments that were true recrudescent infections that would be wrongly classified as \u2018treatment success\u2019, if follow-up ceased at day 28, day 42 and day 63 were 26.1\u00a0% (81/310), 11.6\u00a0% (36/310) and 5.8\u00a0% (18/310), respectively .For four anti-malarial treatments , women were observed to have recrudescent infections 100 or more days from the start of drug treatment Table\u00a0. There wP. falciparum infection. Of the eight risk factors, anti-malarial therapy, therapy supervision, intercalated P. vivax infection, and initial parasitaemia were found to be associated with number of days to recrudescent infection P <0.1, in linear regression analyses, including a single risk factor and adjusting for age, weight and study period  was observed for women whose treatment was incompletely supervised compared to completely supervised; and 0.98-fold (95\u00a0% CI: 0.96-0.99) for each doubling in the baseline parasitaemia.Estimated ratios of geometric means are presented in Table\u00a0P. falciparum malaria parasitaemia in pregnancy.This study reports the largest, single-site, longitudinal population study of PCR-genotyped P. falciparum. Supervising the treatment was associated with an increased interval to late recrudescence. Several factors contributed to the high rates of recrudescence. While all women were treated with standard adult dosing of anti-malarials, pregnancy alters the pharmacokinetic properties of many anti-malarials lowering their plasma concentrations [The interval from treatment of falciparum malaria in pregnancy to PCR-confirmed recrudescent infection can be prolonged. The maximum observed time in this large dataset was 126\u00a0days. Pregnant women in this study generally received supervised rather than unsupervised treatment of trations , 18. It trations , 31. Dortrations , 33. Howtrations .P. vivax infection resulted in a significantly lower proportion of recrudescent P. falciparum episodes than in women who did not have intercalated P. vivax. Intercalated P. vivax prolonged the time to recurrence of both novel and recrudescent infections and was the most significant risk factor for prolonged time to recrudescence. Although the chloroquine inhibitory concentration of 50\u00a0% (IC50) for P. falciparum isolates in this area are amongst the highest reported in the world [P. falciparum in a double-blind, randomized, placebo-controlled, prophylaxis trial of 1,000 pregnant women in this setting [P. vivax may activate host-defences which affect P. falciparum [P. vivax has previously been reported in this area: mixed  infection was associated with a four-fold reduction in the risk of developing severe malaria [Intercalated he world , 36 some setting . In addilciparum . The two malaria .P. falciparum parasitaemia in pregnancy are harmful to the pregnant woman and the foetus and prompt and efficacious treatment is required [The proportion of recrudescent infections with quinine and mefloquine monotherapy was high and these poorly efficacious regimens should no longer be used in this area , 7. Paterequired . Frequenrequired . Treatmerequired , which cThis study demonstrates that a significant proportion of pregnant women, 29.0\u00a0% by day 28, 13.3\u00a0% by day 42 and 5.6\u00a0% by day 63 of follow-up, have recrudescent infections beyond the traditional boundaries used for follow-up in anti-malarial efficacy trials. These proportions are large enough to underestimate significantly true failure rates with shorter periods of follow-up. Accurate assessment of anti-malarial efficacy in pregnancy requires follow-up to delivery, failing this, day 63. This is contrary to modelling data from trials in non-pregnant patients where 42\u201363 days follow-up captures nearly all anti-malarial treatment failures [P. falciparum no trial that includes both pregnant and non-pregnant women has been conducted. Attempts to pool data on recrudescent P. falciparum infections for age-matched women of reproductive age treated with the same anti-malarial and same year of treatment within the population, i.e., artemether-lumefantrine and dihydroartemisinin-piperaquine, were unsuccessful.There are limitations to this analysis: only three drug treatments  accounted for 85\u00a0% of the data. In this setting, with MDR-per se many predispose to a longer duration to recrudescence this study is unable to prove it. Only settings with data including prolonged duration of follow-up; the same regimen for diagnosis and treatment of P. falciparum infections in pregnant and non-pregnant women which is most often not the case; and preferably with women matched for age and gravidity would have the potential to elucidate this suspicion.While there are reasons to suspect that pregnancy P. falciparum are very low it is difficult to justify the use of IPT as the risk benefit ratio may not be in favour of providing this drug to all pregnant women. The current annual incidence of P. falciparum in pregnancy in the area is now less than 0.5 infections per woman per year [No effect of anti-malarial drugs was observed on the post-treatment prophylactic effect, i.e., the time to novel recurrence however without a group who receives no anti-malarials this cannot be proven. There was limited power to detect a post-treatment prophylactic effect and there were few patients who received long-acting anti-malarials and re-infection rates were very low, so any effect would have been small. Regarding IPT in this area a study conducted in men followed for nine months in a randomized, placebo controlled IPT trial required dihydroartemisinin-piperaquine to be provided monthly at full treatment doses (two tablets twice per day for three days) to be effective . When reper year .P. vivax occurs and receives treatment, time to recrudescence of P. falciparum can be prolonged. In this area of low, seasonal malaria transmission, recrudescence occurred after day 42 in approximately 15\u00a0% and after day 63 in 5\u00a0%, of pregnant women. Accurate characterization of drug efficacy in pregnancy requires follow-up to delivery or day 63, whichever occurs last.The interval to recrudescence of falciparum malaria in pregnancy can be prolonged, regardless of the anti-malarial used for treatment. In areas where intercalated"}
+{"text": "Plasmodium vivax malaria is a major cause of morbidity and recognised as an important contributor to mortality in some endemic areas. The current recommended treatment regimen for the radical cure of P. vivax includes a schizontocidal antimalarial, usually chloroquine, combined with a 14\u00a0day regimen of primaquine. The long treatment course frequently results in poor adherence and effectiveness. Shorter courses of higher daily doses of primaquine have the potential to improve adherence and thus effectiveness without compromising safety. The proposed multicentre randomised clinical trial aims to provide evidence across a variety of endemic settings on the safety and efficacy of high dose short course primaquine in glucose-6-phosphate-dehydrogenase (G6PD) normal patients.P. vivax parasitaemia over 12\u00a0months of follow-up, for all individuals, controlling for site, comparing the 7 versus 14-day primaquine treatment arms. Secondary endpoints are other efficacy measures such as incidence risk at different time points. Further endpoints are risks of haemolysis and severe adverse events.This study is designed as a placebo controlled, double blinded, randomized trial in four countries: Indonesia, Vietnam, Afghanistan and Ethiopia. G6PD normal patients diagnosed with vivax malaria are randomized to receive either 7 or 14\u00a0days high dose primaquine or placebo. G6PD deficient (G6PDd) patients are allocated to weekly primaquine doses for 8\u00a0weeks. All treatment is directly observed and recurrent episodes are treated with the same treatment than allocated at the enrolment episode. Patients are followed daily until completion of treatment, weekly until 8\u00a0weeks and then monthly until 1\u00a0year after initiation of the treatment. The primary endpoint is the incidence rate (per person year) of symptomatic recurrent P. vivax malaria treatment policy through peer-reviewed publications and academic presentations. Findings will contribute to a better understanding of the risks and benefits of primaquine which is crucial in persuading policy makers as well as clinicians of the importance of radical cure of vivax malaria, contributing to decreased transmission and a reduce parasite reservoir.This study has been approved by relevant institutional ethics committees in the UK and Australia, and all participating countries. Results will be disseminated to inform NCT01814683. Registered March 18, 2013ClinicalTrials.gov Identifier: The online version of this article (doi:10.1186/s12879-015-1276-2) contains supplementary material, which is available to authorized users. Plasmodium vivax malaria is a major cause of morbidity and recognised as an important contributor to mortality in some endemic areas. Unlike P. falciparum malaria, P. vivax infections form dormant liver stages (hypnozoites) which cause relapses of the infection weeks to months after the initial attack. In South-East Asia relapse rates commonly exceed 50\u00a0%, making relapse the main cause of vivax illness. Recurrent episodes of febrile illness and haemolysis inflict a significant public health burden particularly in vulnerable groups such as pregnant women and young children. The first line treatment of vivax malaria is a combination of chloroquine , and primaquine . Primaquine, an 8 aminoquinoline, is currently the only licensed drug with activity against hypnozoites. An important constraint on the global deployment of primaquine is its potential to cause haemolysis in patients with glucose-6-phosphate dehydrogenase deficiency (G6PDd), which typically occurs in 2\u201315\u00a0% of patients in endemic regions [P. vivax infection results in healthcare providers rarely prescribing primaquine even when recommended in policy. The lack of a safe and reliable radical cure of P. vivax is a major threat to current malaria control and elimination efforts. regions . In pracThe main determinant of primaquine efficacy is the total dose of primaquine administered, rather than the dosing schedule . The 14\u00a0P. vivax so larger daily doses may substantially augment chloroquine\u2019s blood stage activity in areas of low level chloroquine resistance. In Thailand directly observed primaquine (1\u00a0mg/kg/day) administered over 7\u00a0days was well tolerated and reduced relapses by day 28 to 4\u00a0% [Shorter courses of higher daily doses of primaquine have the potential to improve adherence and thus effectiveness without compromising efficacy . Primaqu8 to 4\u00a0% . This isP. vivax in patients with known G6PDd is challenging. Current WHO guidelines recommend a weekly dose of 0.75\u00a0mg/kg for 8\u00a0weeks which mitigates primaquine-induced haemolysis [The radical cure of emolysis .P. vivax. A better understanding of the risks and benefits of primaquine is crucial in persuading policy makers and clinicians of the importance of the radical cure of vivax malaria that will reduce the parasite reservoir and decrease transmission. The strength and limitations of this study are listed in Table\u00a0Due to the long duration of standard primaquine treatment regimens, courses are difficult to supervise, are poorly adhered to and lack effectiveness. The proposed multicentre randomised clinical trial will provide evidence across a variety of endemic settings on the safety and efficacy of high dose-short course primaquine in G6PD normal patients. In a parallel single arm study data on the safety of weekly primaquine in patients with G6PDd will be obtained. The study aims to generate evidence that will directly inform global public health policy for the radical cure of P. vivax relapse in G6PD normal patients. Secondary objectives are to assess the absolute risks and benefits of radical treatment regimens in different endemic settings, to provide data on the safety of a weekly dose of primaquine (0.75\u00a0mg base/kg) in patients with G6PD deficiency and to identify the most cost-effective strategies for the management of P. vivax with respect to the use of G6PD tests, the dosing schedule and the epidemiological context.The primary objective of this study is therefore to determine whether a 7-day primaquine regimen is safe and not inferior to the standard 14-day regimen  in preventing \u25cb Intervention 1: Standard blood schizontocidal therapy plus 14\u00a0days of supervised primaquine  administered once per day (0.5\u00a0mg/kg).\u25cb Intervention 2: Standard blood schizontocidal therapy plus 7\u00a0days of supervised primaquine  administered once per day (1.0\u00a0mg/kg OD) followed by 7\u00a0days of placebo.\u25cb Control arm: Standard blood schizontocidal therapy plus 14\u00a0days placebo.This is a randomized, double-blind, placebo-controlled, non-inferiority trial in G6PD normal patients with uncomplicated vivax malaria in seven participating study sites in Indonesia (two sites), Vietnam, Ethiopia (2 sites) and Afghanistan (two sites). Patients presenting to a participating treatment centre with uncomplicated vivax malaria and fulfilling the enrolment criteria will be randomly assigned to one of three treatment arms:Patients tested initially and found to be G6PD deficient will be excluded from the randomised study but offered enrolment in a single arm non-randomised observational study to receive standard schizontocidal therapy plus primaquine 0.75\u00a0mg/kg/week for 8 doses .All patients will receive standard medical care for the management of uncomplicated malaria, with blood schizontocidal treatment administered as either chloroquine  or an artemisinin combination therapy depending on local recommendations and known chloroquine efficacy.Recurrences of any species within 28\u00a0days will be considered treatment failures and treated with local second line alternatives (such as ACT or 7\u00a0days quinine) according to national guidelines. After 28\u00a0days, treatment failure is less likely and so patients will be treated with the same treatment regimen as that allocated at enrolment. Primaquine/placebo will be administered with food (crackers or a biscuit), which has been shown to reduce gastrointestinal side effects. All doses of study drugs will be supervised. If participants cannot visit the study centre, or fail to attend during the 14\u00a0days of supervised therapy, team members will visit them in their homes or places of school or work to ensure complete dosing.P. vivax will be confirmed by microscopy and recorded. Intercurrent P. falciparum parasitaemia will be treated with the locally recommended ACT .Treatment efficacy and patient safety will be ensured by close monitoring over a 12\u00a0months follow up period following a schedule of visits and corresponding clinical and laboratory examinations. Each recurrent Male and female patients over 6\u00a0months of age presenting to a participating treatment centre with uncomplicated vivax malaria will be enrolled into the study if they fulfil the following inclusion and exclusion criteria. All patients will be screened for G6PD status using the NADPH spot test and those found to be deficient will be excluded from the main trial but encouraged to enrol in a parallel G6PDd arm. Participants in this non-randomised, observational study will receive the standard blood schizontocidal therapy plus a single supervised weekly dose of primaquine 0.75\u00a0mg base/kg weekly for 8\u00a0weeks , with a similar follow up regimen as those patients in the primary study.\u25cb Participant (or parent/guardian of children below age of consent) is willing and able to give written informed consent to participate in the trial; verbal consent in the presence of a literate witness is required for illiterate patients. In addition, written assent  from children 12 to 17\u00a0years as per local practice.P. vivax of any parasitaemia in countries which use CQ as blood schizontocidal therapy. Mixed infections with P. vivax and P. falciparum can be enrolled in countries which use an artemisinin combination therapy.\u25cb Monoinfection with \u25cb Diagnosis of malaria can be based on either malaria rapid diagnostic test or slide microscopy, as per site preference.\u25cb Over 6\u00a0months of age\u25cb Weight 5\u00a0kg or greater\u25cb Fever (axillary temperature \u226537.5\u00a0\u00b0C) or history of fever in the last 48\u00a0h.\u25cb Able, in the investigator\u2019s opinion, and willing to comply with the study requirements and follow-up.The participant may enter the study if ALL of the following apply:\u25cb Female participant who is pregnant or lactating\u25cb Inability to tolerate oral treatment\u25cb Previous episode of haemolysis or severe haemoglobinuria following primaquine\u25cb Signs/symptoms indicative of severe/complicated malaria or warning signs requiring parenteral treatment\u25cb Haemoglobin concentration less than 9\u00a0g/dL\u25cb Known hypersensitivity or allergy to the study drugs\u25cb Blood transfusion in last 90\u00a0days, since this can mask G6PD deficient status\u25cb Presence of any condition which in the judgment of the investigator would place the participant at undue risk or interfere with the results of the study \u25cb Coadministration of other medication known to cause haemolysis\u25cb Currently taking medication known to interfere significantly with the pharmacokinetics of primaquine and the schizontocidal study drugs\u25cb Previously been a study participant in IMPROV (i.e. the same patient cannot be enrolled twice)The participant may not enter the study if ANY of the following apply:P. vivax. Some sites will also routinely use rapid diagnostic tests to complement microscopy. After the diagnosis is confirmed microscopically or by Rapid Diagnostic Test (RDT), the patient will be approached for informed consent. In some centres, the following tests are considered part of clinical practice: malaria blood film, RDT, G6PD test and haematocrit or haemoglobin concentration. If any one of these tests is not routine, then consent and assent will be obtained before these tests are performed.When malaria is suspected, a thick and thin blood smear will be obtained for microscopic diagnosis of The participant (or parent/guardian of children below age of consent) must personally sign and date the latest approved version of the informed consent form before any study specific procedures are performed. Alternatively, verbal consent in the presence of a literate witness will be obtained from illiterate patients. In addition to the Informed Consent, Assent will be obtained from children 12 to 17\u00a0years of age if locally required.Demographics: The patient\u2019s date of birth  and gender will be recorded. If the patient is a female of childbearing age, she will be asked if she is currently pregnant, lactating, planning to get pregnant and the date of the first day of her last menstrual period. These questions will be culturally adapted in countries with strict prohibitions and restrictions on marriage and pregnancy.Pre-treatment temperature: The patient (or parent/guardian of children) will be asked if he/she had fever during the past 48\u00a0h and his/her current temperature (axillary) will be measured. This will be done on screening and each subsequent visit.Medical history and physical examination: The details of any disease/surgical conditions, and drug allergies will be recorded. The patient\u2019s pulse rate, respiratory rate, weight, mid upper arm circumference (MUAC) in children\u2009<\u20095\u00a0years, and the results of a baseline physical examination will be recorded. This will be done on screening and each subsequent visit.Prior medication in the last 28\u00a0days: All over-the-counter or prescription medication, vitamins, and/or herbal supplements will be recorded. This will include the medicine name , starting and ending dates, total dose, route of administration and the indication for use. This will be done on screening and each subsequent visit.Capillary blood sample (up to 400\u00a0\u03bcl) will be obtained at enrolment for the following tests :Field Blood Haemoglobin. Samples will be obtained at the initial visit, Day 3 and each weekly or monthly follow up visit .\u25cb Rapid Diagnostic test (RDT) for malaria. A multi-species  RDT for the diagnosis of falciparum and vivax malaria will be undertaken on initial screening. The decision whether to enrol a patients is based on the RDT result.\u25cb Parasite microscopy. A thick and thin malaria smear will be made at each scheduled visit  and at any unscheduled visit if malaria is suspected. Microscopy will be used to confirm the presence of parasitaemia and to estimate the parasite density.\u25cb G6PD Testing. Blood will be collected in an EDTA tube or heparinised haematocrit capillary tube on initial screening for glucose-6-phosphate-dehydrogenase semi-quantitative fluorescent spot test for initial screening in the field. A reagent solution containing Glucose-6-P\u2009+\u2009NADP+ is mixed with whole blood or a dried blood spot. Samples obtained from normal or slightly reduced G6PD activity will show strong fluorescence. Failure to fluoresce after 10-min of incubation suggests a total or marked deficiency of G6PD. This test may fluoresce falsely if the study participant has had a blood transfusion within the last 90\u00a0days. Definition of G6PD status for the purpose of enrolment will be decided based on the NAPD Spot test. If the result is deficient or borderline, the patient will be enrolled into the G6PD deficient arm\u25cb Other tests on remaining capillary blood include the following:P. vivax infection.\u25aa Parasite genetic studies . These will be used to explore parasite factors associated with recurrent \u25aa Host genotyping for G6PD and other red cell and cytochrome P450 drug metabolism polymorphisms (such as 2D6). These will be used to explore whether host factors influence primaquine blood concentrations which may affect treatment failure.\u25aa Drug concentrations .\u25aa Quantitative analysis of G6PD status will be carried out at a suitable reference centre.\u25aa Repeat G6PD testing with novel G6PD tests. To assess the potential of rapid point of care diagnostic tests.\u25aa Serology, to assess the immune status of the patient and whether this affects symptomatic disease.\u25cb A venous blood sample will be obtained on enrolment, days 7 and 13/14 , the first day of each subsequent recurrence, and at a mid-term review at 6\u00a0months for the following tests:Complete Blood Count (CBC). Automated CBCs will be analysed using the Sysmex pocH-100i\u2122, or equivalent, machine, if the machine is available.\u25cb Pharmacokinetic Analysis (PK). Blood samples will be collected on day 7 and day 13/14 for HPLC analysis of blood concentrations including CQ, PP, primaquine and carboxyprimaquine.\u25cb Other tests on remaining venous blood include the following:P. vivax infection.\u25aa Parasite genetic studies . These will be used to explore parasite factors associated with recurrent \u25aa Host genotyping for G6PD and other red cell and cytochrome P450 drug metabolism polymorphisms (such as 2D6). These will be used to explore whether host factors influence primaquine blood concentrations which may affect treatment failure.\u25aa Flow cytometry \u2013 to detect the G6PD activity in heterozygous women.\u25aa Drug concentrations \u25aa G6PD quantitative assay in sites where laboratory facilities permit timely sample processing (if not done on capillary blood). This will be used to explore whether the degree of G6PD activity influences the side effect profile.\u25aa Serology, to assess the immune status of the patient and whether this affects symptomatic disease.\u25cb The total amount of blood to be collected during the study period of 12\u00a0months will vary depending on the number of recurrent episodes of malaria. If there are no recurrences, the minimum blood volumes will be approximately 31\u00a0mL (patient aged\u2009>\u20095y), 19\u00a0mL (age 12\u201360\u00a0m) and 13\u00a0ml (age 6\u2013<\u200912\u00a0m). If there are two recurrences, these volumes become approximately 41, 23 and 14\u00a0mL. These volumes anticipated over a 12\u00a0month follow up are well within the acceptable limits for patients aged 6\u00a0months to 14\u00a0years.Blood for protocol mandated tests that cannot be done straight away, will be stored for future analysis. This may result in unused blood remaining. Permission will be sought for the long term storage of this unused blood for future tests relevant to the study outcomes. The length of storage will follow local ethics committee guidelines. Any new tests will need ethical approval. Any transfer of specimens will follow all relevant national and IATA regulations.Urine \u03b2-HCG pregnancy test will be performed in all women of childbearing age (unless menstruating) eligible for enrolment. The decision to perform the test in unmarried women will be culturally adapted and follow local practice in countries with strict prohibitions and restrictions on marriage and pregnancy. Bioline HCG test strips or an equivalent test will be used. After the initial screening, the test will be performed during scheduled and unscheduled visits if recurrent P. vivax or P. falciparum is diagnosed.In participating centres, where the G6PD result is available straight away, regimen allocation and administration of the study agent will be on Day 0. However, in participating centres where the G6PD result is not available on the day of enrolment, regimen allocation and administration of the study drugs will be on Day 1. Participating centres can decide whether they prefer to give PQ on Day 0 immediately following schizontocidal therapy or on Day 1 following the start of the schizontocidal therapy. It is important that once a decision is made for each site there will be consistent adherence to the chosen starting day within that site.Once G6PD results are available, those testing normal will be randomized in the 3-arm main study whereas, G6PD deficient participants will be enrolled into an observational (non-randomised) open-label arm.A randomization list for participants will be prepared centrally at the Mahidol-Oxford Tropical Medicine Research Unit (MORU), Bangkok, Thailand. Randomisation will be in blocks of 20 for each dosing band. Individual patients drug kits containing primaquine/placebo drug cards will be labelled by weight bands: A \u2013 5\u201334\u00a0kg, B - 35\u201345\u00a0kg, and C-\u2009\u2265\u200946\u00a0kg. There will be 20 individual patient drug kits in a box.The study staff responsible for generating the randomization list and for selecting code letters for the study agents will not be involved in any other way in the conduct of the trial and will not be present at the study site.Randomisation to determine the regimen allocation will be carried out as soon as a participant is enrolled and will be based on his/her weight. The first drug kit in the relevant box will be given to the first patient; the second drug kit will be given to the next patient in that weight category and so on. The number on the drug kit will be the unique drug randomisation number. It will be recorded onto the CRF as well as the subject number.Primaquine and primaquine placebos have been manufactured specifically for the study. In order to conceal the allocation of primaquine versus placebo and dose regimen of primaquine, the study will use 7.5\u00a0mg and 15\u00a0mg tablets primaquine and a placebo with similar appearance. Primaquine will be administered as a single daily dose: 0.5\u00a0mg/kg/day \u00d7 14\u00a0days or 1.0\u00a0mg/kg/day \u00d7 7\u00a0days.All study participants will receive blood schizontocidal treatment administered as either chloroquine  or an artemisinin based combination treatment (ACT), depending on local recommendations and known chloroquine efficacy. Chloroquine (each tablet containing 155\u00a0mg of base) will be given at initially (4 tablets or 10\u00a0mg/kg for children) and then at 6\u20138, 24 and 48\u00a0h (2 tablets or 5\u00a0mg/kg for children). Artekin\u2122 (each tablet containing 40\u00a0mg dihydroartemisinin and 320\u00a0mg piperaquine) will be given at 0, 24 and 48\u00a0h. Artemether-lumefantrine (CoArtem\u00ae) is another ACT in use in some countries. One tablet contains 20\u00a0mg of artemether and 120\u00a0mg of lumefantrine. Dosing will be by weight administered twice daily for three days.P. vivax. It is essentially a pro-drug that is extensively and rapidly metabolized to carboxyprimaquine with only a small fraction of the parent drug excreted unchanged. However, little is known of the pharmacokinetics of the metabolites that are responsible for both its antimalarial activity and toxicity. Its principal metabolite, carboxyprimaquine, is formed as a result of oxidative deamination, which is thought to involve both the cytochrome-P450 enzyme complex and monoamine oxidases (MAO). Primaquine is rapidly and almost completely absorbed following oral administration, with peak plasma concentrations (Cmax) reached within 3\u00a0h. It is cleared by hepatic biotransformation, with an elimination half-life of 8\u00a0h. The pharmacokinetics of primaquine does not seem to be time-dependent, showing similar kinetics after repeated dosages. The most important adverse effect is acute haemolytic anaemia in patients with G6PD deficiency. Primaquine may also cause abdominal pain if administered on an empty stomach. Other side effects include nausea and vomiting. Uncommon effects include mild anaemia and leucocytosis. Primaquine causes methaemoglobinaemia , but this seldom causes symptoms in healthy individuals and is self-limiting.Primaquine is an 8-aminoquinoline with cidal activity against the gametocytes and hypnozoites of \u25cb Primaquine regimen 1: 14\u00a0days of supervised primaquine  administered once per day (0.5\u00a0mg/kg)\u25cb Primaquine regimen 2: 7\u00a0days of supervised primaquine  administered once per day (1.0\u00a0mg/kg OD) followed by 7\u00a0days of supervised placebo\u25cb Control Arm: 14\u00a0days of supervised placeboEligible G6PD-normal patients will be randomized to two primaquine regimens and the control arms in a ratio of 2:2:1.Eligible G6PD deficient participants will be enrolled into an observational (non-randomised) open-label arm. Aside from standard blood schizontocidal therapy, these participants will receive a single supervised weekly dose of primaquine 0.75\u00a0mg base/kg weekly for 7\u00a0weeks i.e. 8 doses . During an 8-week treatment regimen haematological screening and clinical assessments will be made prior to each weekly dose of primaquine. Haemoglobin concentration will be assessed using HemoCue\u2122  done on Days 3, 7, 10, 14, 17, 21 etc. in the G6PDd arm. The proportion of subjects completing all 8 doses will be classified as \u201csafely treated\u201d. Subjects deemed at risk with further dosing will not be treated further with primaquine. Providing that consent is not withdrawn, all patients, irrespective of completion of therapy, will continue to be followed up to document safety and efficacy parameters at 12\u00a0months.All patients in either group will be observed for one hour after treatment. If vomiting occurs within one hour, the investigator has the option of redosing with both drugs or omitting the primaquine/placebo or open label primaquine. If the repeat dose of ACT / CQ is also vomited within one hour, the patient will be stopped oral treatment and will be given parenteral rescue treatment.When the patient is able to eat and drink normally, they will continue with their study drugs to complete their courses of both the schizonticidal treatment and the primaquine/placebo or open primaquine. Drugs which have been vomited will be counted as administered treatment on that day.Patients may develop persistent vomiting, even after the acute phase of illness. In this case the primaquine/placebo will be stopped and restarted once the vomiting has resolved.Patients will be seen daily until PQ treatment is completed, weekly until week 8 and then monthly until 1\u00a0year after enrolment. For the weekly visits a window of 3\u00a0days either side will be acceptable for the scheduled follow up. For the monthly visits, 7\u00a0days either way will be acceptable. Some subjects may be unable to attend the clinic or be away from home. If such patients own a mobile phone, they can be contacted and interviewed by phone to complete a symptom questionnaire to assess safety outcomes. The most important adverse effect of primaquine is haemolytic anaemia in patients with G6PD deficiency. Haemoglobin concentration  will therefore be checked regularly.Methaemoglobinemia will be assessed using a non-invasive oximeter (Masimo) at baseline, daily for the first three days then D7 and 13/14. MetHb will be measured in a subgroup of patients at two selected sites in the first instance; thereafter, the Masimo machine will be moved to another site. To reduce the risk of unblinding of patient allocation, metHb concentrations will be measured by a research team member who will not be involved in other patient assessments. These data will be recorded in a register and not on the case report form to reduce the risk of unblinding the treatment regimen.Participants will attend the study centre according to the scheduled times and will also be encouraged to report to the study centre in the event of any illness. The blood film will only be read immediately if the patient is symptomatic (i.e. has a fever or a history of fever in the preceding 24\u00a0h); this mirrors the real world scenario of passive case detection when patients only present to a clinic if they are sick.If the patient is well at a routine follow up the blood film will be stored and processed at a later date in the reference laboratory. This will provide data on asymptomatic parasitaemia and how often it resolves spontaneously; this has important implications for understanding the transmission of malaria back to the community. Asymptomatic patients subsequently found to have peripheral parasitaemia will not be sought and treated.\u25cb A recurrent vivax parasitaemia within 28\u00a0days is potentially a recrudescent infection. These patients will be treated according to national guidelines, usually an ACT or quinine. They will remain in the study, have their parasitaemia checked on Day 7 and will then resume routine follow up.\u25cb After 28\u00a0days, a recurrent vivax parasitaemia is more likely to represent a relapse. These patients will be treated with a repeat supervised course of the same regimen that was administered on Day 0  with additional follow-up on days 7 and 14, and thereafter at their scheduled monthly follow up visit until the end of the study.\u25cb Those in the G6PDd group will also be treated with the same schizonticidal drug administered on Day 0 and will either complete their 8 doses of primaquine, if this has not been completed when the recurrence occurred or will be given a new course of 8 doses, if not on primaquine at the time of the recurrence.P. falciparum parasitaemia , they will be treated according to national guidelines . Some countries also recommend a single dose of primaquine 0.45\u00a0mg/kg for transmission blocking. Patients who develop severe malaria will be admitted to hospital and treated with the recommended parenteral drug. Some CQ treated patients may develop falciparum parasitaemia while still taking CQ, if this happens, they will be treated with an ACT.\u25cb If patients develop symptomatic malaria with Treatment will only be offered to patients who are symptomatic patients.P. vivax or P.falciparum, will have their treatment supervised and clinical and parasitological recovery documented and continue in the study.Patients with recurrent malaria either due to P. vivax infection. Patients may experience multiple episodes of P. vivax infection. Each episode will be documented and the patient retreated with the study drug. If a patient has more than four symptomatic recurrences of P. vivax, outside the time defined for a recrudescent infection, then they will not be prescribed any further study drug. Patients on their fifth or subsequent symptomatic P. vivax recurrence will be treated with supervised open primaquine at a dose of 0.5\u00a0mg/kg \u00d7 14\u00a0days. They will remain in the study and be followed up to Day 365.Patients may experience symptomatic recurrent vivax parasitaemia during follow up after they have received treatment for their initial P. vivax recurrences within the first six months of the study. Such patients will then be treated with be treated with supervised open primaquine at a dose of 0.5\u00a0mg/kg \u00d7 14\u00a0days. They will remain in the study and be followed up to Day 365.In Vietnam, retreatment with study drug will only be allowed for three symptomatic P. vivax recurrences will be treated with supervised open primaquine at a dose of 0.5\u00a0mg/kg \u00d7 14\u00a0days, if this has not already been given. Those patients who did not have any recurrent vivax parasitaemias are at low risk of further relapse, and will not be offered supervised open primaquine.At the end of the study, patients who have had 1 or more Throughout the study investigators may prescribe other concomitant medications or treatments deemed necessary to provide adequate supportive care, e.g. paracetamol for fever. Any medication, other than the study medication taken during the study will be recorded in the CRF. Drugs conferring a risk of hemolysis are listed in table\u00a0A total of two household cost questionnaires will be completed on enrolment and on day 13/14. The second one will capture all costs following the attendance on day 0, including the duration of the episode and associated productivity losses, further expenditure for consultation, medication, investigations or admissions, travel costs (other than those for research purposes).The end of trial is the date of the last visit of the last participant, which would be a maximum of 365\u00a0days after recruitment of the last participant. No participant will be followed up for longer than 365\u00a0days from enrolment regardless of repeated episode of parasitaemia.Patients withdrawn from the study will not have any further protocol mandated study investigations or procedures at and after the point of withdrawal. Each participant has the right to withdraw from the study at any time .\u25cb Ineligibility (either arising during the study or retrospective having been overlooked at screening)\u25cb Significant protocol violation e.g. given wrong dose of primaquine/placebo\u25cb Significant non-compliance with treatment regimen or study requirements\u25cb An adverse or serious adverse event which results in inability to continue to comply with study procedures\u25cb Disease progression which results in an inability to continue to comply with study procedures\u25cb Lost to follow up (patients fail to return for follow up right up to study day 365)Other reasons for study withdrawal include:Summary information on withdrawn patients will be recorded in the final status CRF. If the participant is withdrawn from the study because of an adverse event, the investigator will arrange for follow-up visits or telephone calls until the adverse event has resolved or stabilised.\u25cb Persistent vomiting of study drug/s\u25cb A serious adverse event which requires stopping study drug/s\u25cb Disease progression which requires discontinuation of the study medication An emergency event requiring unblinding of the drug regimen code allocated to the participantThere will be instances when patients may have adverse events that result in temporary interruption of their study drugs or a break in their follow up. When these have resolved, patients will still remain in the study and be followed up according to the study schedule. Examples include:Adverse Event (AE): An AE or adverse experience is any untoward medical occurrence in a patient or clinical investigation participants administered a medicinal product, which does not necessarily have to have a causal relationship with this treatment (the study medication). An AE can therefore be any unfavourable and unintended sign , symptom or disease temporally associated with the use of the study medication, whether or not considered related to the study medication.Adverse Reaction (AR): Adverse reactions are defined as untoward and unintended responses to a medicinal product related to any dose. The phrase \u201cresponses to a medicinal product\u201d infers that a causal relationship between a study medication and an AE is at least a reasonable possibility, i.e., the relationship cannot be ruled out. All cases judged by either the reporting medically qualified professional or the sponsor as having a reasonable suspected causal relationship to the study medication qualify as adverse reactions.Serious Adverse Event (SAE): A serious adverse event is any untoward medical occurrence that results in death, is life-threatening, requires inpatient hospitalisation or prolongation of existing hospitalisation, results in persistent or significant disability/incapacity, or is a congenital anomaly/birth defect.Other events that may not result in death are not life threatening, or do not require hospitalisation, may be considered a serious adverse event when, based upon appropriate medical judgement, the event may jeopardise the patient and may require medical or surgical intervention to prevent one of the outcomes listed above.Serious Adverse Reaction (SAR): An adverse event (expected or unexpected) that is both serious and, in the opinion of the reporting investigator, believed with reasonable probability to be due to one of the study treatments, based on the information provided.Suspected Unexpected Serious Adverse Reaction (SUSAR): A serious adverse reaction, the nature or severity of which is not consistent with the applicable product information .Causality and Expectedness: The relationship of an adverse event to study medication is to be assessed according to the following definitions:\u25cb Definitely Related: Strong evidence exists that the study drug caused the adverse event. There is a temporal relationship between the event onset and administration of the study drug. There is strong therapeutic and pharmacologic evidence that the event was caused by the study drug. The subject\u2019s clinical state or concomitant therapies have been ruled out as a cause. In the case of cessation or reduction of the dose, the event abates or resolves, and reappears upon re-challenge.\u25cb Probably Related: A temporal relationship exists between the event onset and the administration of study drug, and appears with some degree of certainty to be related based on the known therapeutic and pharmacologic actions of the study drug. It cannot be readily explained by the subject\u2019s clinical state or concomitant therapies. In the case of cessation or reduction of the dose, the event abates or resolves.\u25cb Possibly Related: A temporal relationship exists between the event onset and the administration of study drug. Although the adverse event may appear unlikely to be related to the study drug, it cannot be ruled out with certainty; and/or the event cannot be readily explained by the subject\u2019s clinical state or concomitant therapies.\u25cb Not Related: Evidence exists that the adverse event definitely has aetiology other than the study drug  and does not meet any criteria listed above.Recording AEs: All grades 3 and 4 adverse events should be recorded for up to 28\u00a0days after the last day of administration of primaquine in G6PD normal patients (i.e. Day 42) and up to 14\u00a0days after the last dose in G6PD deficient patients. All related AEs that result in a participant\u2019s withdrawal from the study or are present at the end of the study, should be followed up until a satisfactory resolution occurs. It will be left to the Investigator\u2019s clinical judgment whether or not an AE is of sufficient severity to require stopping the participant\u2019s treatment. A participant may also voluntarily withdraw from treatment due to what he or she perceives as an intolerable AE. If either of these occurs, the participant must undergo an end of study assessment and be given appropriate care under medical supervision until symptoms cease or the condition becomes stable.The severity of events will be assessed on the following scale: 1\u2009=\u2009mild, 2\u2009=\u2009moderate, 3\u2009=\u2009severe, 4\u2009=\u2009potentially life-threatening. Any pregnancy occurring during the clinical study and the outcome of the pregnancy should be recorded and followed up for congenital abnormality or birth defect.Reporting Procedures for Serious Adverse Events: All SAEs will be reported by the site investigator to the MORU study coordinator and pharmacovigilance officer within one working day of awareness, who will in turn notify the DSMB and PI within 48\u00a0h of their being notified of the event. The site PI will be responsible for notifying the local Ethical Committees as appropriate. All information received for a case will be detailed on a full SAE report form.DMSB: A Data Safety Monitoring Board (DSMB) will be appointed to review the study progress, safety data, critical efficacy outcomes, at regular intervals and recommend to the sponsor whether to continue, modify, or stop the trial.Annual Safety reports: In addition to the expedited reporting above, the PI shall submit an annual safety report throughout the clinical trial, and any additional reports on request.Malaria can cause a temporary fall in haemoglobin that can be exacerbated by primaquine  AND a fractional fall in haemoglobin\u2009\u2265\u200925\u00a0% from baselinePatients may report symptoms that are suggestive of anaemia and / or acute intravascular haemolysis or may have decline in the HemoCue measured Hb that may be concerning for the attending clinician. All patients in whom the attending clinician is concerned about will be assessed, including a history, recent non study drug intake, a clinical examination, urine inspection for colour, measure the Hb and retest for G6PDd (daily arm only) with the fluorescent spot test. If any of the following are present then primaquine/placebo study drug will be stopped:Patients reaching any of these criteria will have their primaquine/placebo stopped. They will be unblinded from the study but will continue to be followed for the remainder of their 1\u00a0year follow up. The need for admission to hospital for observation or to receive treatment including a blood transfusion is a serious adverse event (SAE) by definition.A fractional Hb drop\u2009\u2265\u200925\u00a0% from baseline ORUrine colour\u2009\u2265\u20095 (macroscopic haemoglobinuria) OROther significant clinical concern of increasing anaemia or severe haemolysisIf the patient has any of the following features primaquine will be withheld, pending close monitoring:An adverse event form must be filled for those patients with clinical concern who had their primaquine/placebo trial drug therapy withheld but later recommenced.Following a severe haemolytic reaction, patients are at significant risk of further haemolysis if dosed with primaquine again. In some settings local clinicians regard the risk of multiple recurrent episodes warrant careful rechallenge with primaquine. However the decision to redose with open label primaquine will be made by the attending local clinician. It is not a protocol requirement. The physician will assess the risks and benefits and discuss them with the patient. In patients who have had severe haemolysis, the risk of further drug induced haemolysis will be weighed against the risk of recurrent malaria causing haemolysis and anaemia.If primaquine is prescribed then this should be at the weekly dose of 0.75\u00a0mg /kg to complete a total dose of 6\u00a0mg/kg. Redosing should be limited to those patients with no signs of ongoing haemolysis, a Hb above 9\u00a0g/dL, no anaemia related symptoms and no signs evidence of clinical compromise.The primary aim of this trial is to demonstrate non-inferiority of a 7\u00a0day primaquine regimen to the standard 14\u00a0day primaquine regimen with respect to incidence rate of vivax symptomatic parasitaemia over 12\u00a0months. This will reflect mainly relapse in these relatively low transmission settings. The sample size calculation is based on an assumed incidence rate of 0.2 infections per person-year in both arms, a non-inferiority margin of 0.07 infections per person-year and a one-sided significance level of 2.5\u00a0%. Based on these assumptions, a total sample size of 1200 evaluable patients, randomly allocated to receive a primaquine regimen (600 patients in each treatment arm), followed for one year will provide a power of 80\u00a0% to show non-inferiority or, equivalently, that the two-sided 95\u00a0% confidence interval for the difference in incidence rate of malaria between the two arms excludes an excess rate of 0.07 infections per person-year or more in favour of the 14\u00a0day regimen.A further 300 patients in the control arm will also be followed for one year. With 300 patients in the control arm and 600 patients in each treatment arm (i.e. 75 controls and 150 patients in each treatment arm at each of the five study sites), the study has 95 power and 95\u00a0% confidence to detect a difference (i.e. a superiority comparison of either regimen) at each of the study sites assuming an incidence rate of 0.2 infections per person-year in each of the treatment arms and 0.6 infections per person-year in the control arm (ranging from 0.2 in the Indian subcontinent and 1.0 in Vietnam and Indonesia). The combined proportion of losses to follow-up and major protocol violations is expected to be no more than 20\u00a0%, so to account for this, a total of 1875 G6PD normal patients  will be randomized in this trial. In addition, up to 380 patients (approximately 20\u00a0% of the screened participants) with G6PD deficiency will be enrolled for the non-randomised observational study.With the addition of two extra sites, Ethiopian and northern Sumatra, we expect to exceed the original sample size calculation and, thus, have increased power.A detailed statistical analyses plan has been developed and agreed on among the investigators  is obtained from the following review boards: Oxford Tropical Research Ethics Committee OxTREC (Ref number 1014\u201313) and the Human Research Ethics Committee of the Northern Territory Department of Health, Australia HREC (Ref Number 13\u20131991). In addition local approvals were obtained from the National Bioethics Committee (NBC) in Pakistan (this site was dropped before start of enrolment), the Institutional Review Board, Ministry of Public Health, Afghanistan, the Health Research Ethics Committee, Faculty of Medicine University of Indonesia, Cipto Mangunkusumo Hospital, Jakarta, Indonesia, the Ministry of Health Evaluation Committee on Ethics in Biomedical Research Vietnam, the Institutional Scientific & Ethical Review Committee of the Ethiopian Public Health Institute, the National Research Ethics Review Committee, Ethiopia and the Institutional Review Board of the Columbia University Medical Centre, US.Indemnity for the trial is provided by the University of Oxford and Menzies School of Health Research. SAEs will be reported to the DSMB, ethics committees and the sponsor\u2019s research office. Good Clinical Practice (GCP) training is provided to all staff/investigators prior to commencing the studies. Other ethical consideration can be found in Additional file All Investigators will be involved in reviewing drafts of the manuscripts, abstracts, press releases and any other publications arising from the study. Authorship will be determined in accordance with the ICMJE guidelines and other contributors will be acknowledged."}
+{"text": "Plasmodium vivax infections. Although malaria transmission has declined, current treatment strategies must be evaluated to advance towards malaria elimination.In Mexico, combined chloroquine (CQ) and primaquine (PQ) treatment has been used since the late 1950s to treat P. vivax with the 14-day (T14) treatment or intermittent single dose (ISD) regimen was evaluated in southern Mexico between February 2008 and September 2010. Patients over 12\u00a0months old with P. vivax mono-infection and asexual parasitaemia\u00a0\u2265500 parasites/\u00b5l were treated under supervision. After diagnosis (day 0), treatment began immediately. T14 patients received CQ for 3\u00a0days  and PQ daily for 14\u00a0days (0.25\u00a0mg/kg), while ISD patients received a single dose of CQ (10\u00a0mg/kg) and PQ (0.75\u00a0mg/kg) on days 0, 30, 60, 180, 210, and 240. Follow-up was done by observing clinical and laboratory  outcome, considering two endpoints: primary blood infection clearance and clinical response at\u00a0~28\u00a0days, and the incidence of recurrent blood infection during 12\u00a0months. Parasite genotypes of primary/recurrent blood infections were analysed.The clinical and parasitological outcome of treating symptomatic During the first 28\u00a0days, no differences in parasite clearance or clinical outcome were observed between T14 (86 patients) and ISD (67 patients). On day 3, 95\u00a0% of patients in both groups showed no blood parasites, and no recurrences were detected on days 7\u201328. Contrarily, the therapeutic effectiveness (absence of recurrent parasitaemia) was distinct for T14 versus ISD at 12\u00a0months: 83.7 versus 50\u00a0%, respectively (p\u00a0=\u00a00.000). Symptomatic and asymptomatic infections were recorded on days 31\u2013352. Some parasite recurrences were detected by PCR and/or serological testing.P. vivax transmission. Alternatives for meeting the challenge of T14 supervision are discussed.T14 was effective for opportune elimination of the primary blood infection and preventing relapse episodes. The first single dose of CQ-PQ eliminated primary blood infection as efficiently as the initial three-dose scheme of T14, but the ISD regimen should be abandoned. A single combined dose administered to symptomatic patients in remote areas while awaiting parasitological diagnosis may contribute to halting Trial registration: NIH-USA, ClinicalTrial.gov Identifier: NCT02394197The online version of this article (doi:10.1186/s12936-015-0938-2) contains supplementary material, which is available to authorized users. Plasmodium falciparum was completely eliminated in 2000, the same year in which 7259 Plasmodium vivax cases were reported. The number of cases of the latter Plasmodium species was reduced to 2702 in 2009. The country is currently considered to be in the pre-elimination phase [According to malaria records, which have existed in Mexico since 1949, the last epidemic occurred in the 1980s. At that time, more than 120,000 cases were reported each year and malaria transmission was present in over 50\u00a0% of the country. Due to the intensification of control activities, transmission has now been reduced to a few residual foci. In fact, local transmission of P. vivax blood and liver infections [In most affected areas around the world, a combination treatment of chloroquine (CQ) and primaquine (PQ) was introduced in the 1950s to treat fections . In regifections , 8. In fP. vivax malaria to date. However, PQ treatment is frequently not completed due to its undesirable side effects. Although haemolysis due to glucose 6-phosphate dehydrogenase deficiency (G6PDd) in PQ-treated patients is a concern in malaria-affected regions [Like most of the Americas, Mexico has been using the combination of CQ and PQ as the standard treatment for  regions , the Mex regions . In this regions \u201313.In locations of difficult access, the intervals between blood sampling, parasitological diagnosis and patient treatment used to be as long as 3\u20134\u00a0weeks. Hence, to reduce the risk of transmission, febrile patients were blood sampled and immediately treated with a single dose of 10\u00a0mg of CQ and 0.75\u00a0mg of PQ per kg bw , 15. TheP. vivax infections during a 12-month period.The first single combined CQ-PQ dose is expected to eliminate blood parasites, since PQ may destroy parasites surviving from the effect of CQ , 18, 19.The protocol of this study was approved by the Ethics in Research Review Committee of the National Institute of Public Health (Mexico). Informed consent was obtained from all patients or the guardians of minors.An open-labelled, non-randomized, clinical trial was conducted with a group of malaria patients receiving a combined dose of CQ and PQ with either an ISD schedule or the standard 14-day treatment (T14). In both cases, the first dose was given on day 0, the day of diagnosis. The treatment dosing was adjusted according to the age group of a patient, as recommended by the Mexican MCP for ISD and T14 . The stuThe study was carried out in the Tapachula municipality and surrounding communities in southern Chiapas, Mexico, a region on the Guatemala border with an altitude ranging from\u00a0~50\u20131200 meters above sea level. This is a hypo-endemic region showing variable malaria incidence, with an annual parasite incidence (API) of 0.579\u20131.523 cases/1000 inhabitants when considering only the affected villages for the period 2003\u20132007 . Patient recruitment was carried out in the diagnosis facility at the Regional Centre for Public Health Research-INSP (CRISP) in Tapachula City, where febrile individuals seeking malaria diagnosis and treatment can obtain free service.P. vivax mono-infection, confirmed by microscopy; (3) parasite densities of 500-50,000/\u03bcl of blood; and, (4) elevated axillary temperature (\u226537.5\u00a0\u00b0C) or a history of a fever episode within the previous 48\u00a0h. Patients were excluded if they: (1) lived in communities that could not be reached by travelling for 1\u00a0h by motor vehicle from the CRISP centre; (2) presented signs of severe malnutrition or anaemia; (3) had taken an anti-malaria treatment or had had a malaria infection within the previous 12\u00a0months (to reduce the probability of incorporating patients with a relapse) [Patients were recruited from February 2008 to October 2009 and the follow-up ended in September 2010. Inclusion criteria were: (1) patients over 12\u00a0months old; (2) rature \u22653.5\u00a0\u00b0C or For practical purposes, patients were not randomized to form the two treatment groups (ISD and T14). Patients living in Tapachula City and the nearby area were preferred to administer the T14 treatment in order to facilitate daily supervision. On the day of diagnosis (day 0), patients were examined for axillary temperature, body weight, spleen swelling, and clinical symptoms such as fever, headache, myalgia, arthralgia, erythema, and jaundice. Capillary blood, collected by finger pricking, was used to prepare thin and thick smears and to impregnate filter paper (Whatman #2) for parasitological and serological (DNA and antibody) analysis, respectively.For the ISD regimen, the standard single dose consisted of 10\u00a0mg/kg of CQ and 0.75\u00a0mg/kg of PQ, which the patients were scheduled to take on days 0, 30, 60, 180, 210, and 240. For the T14 treatment, the optimal dose was 25\u00a0mg/kg of CQ administered over a three-day period (10\u00a0mg/kg on days 0 and 1 and 5\u00a0mg/kg on day 2) and PQ at 0.25\u00a0mg/kg/day given during 14\u00a0days [The CQ and PQ tablets were provided by the local malaria control programme  at any time during the follow-up period, patients were encouraged to visit the facility at CRISP or to contact the field team by mobile phone (most patients or their parents had a mobile phone). The same recommendation was given in the event that patients had any question about the study. Whether during the treatment or follow-up period, participants unable to reach the facility were visited at their homes for treatment, clinical revision and blood sampling. All treated patients were scheduled for examination on days 2, 3, 7, 14, 21, and 28 . AfterwaP. vivax episode was confirmed, they were immediately treated with T14 [Two thick and thin blood films and a sample of blood impregnated in filter paper (Whatman #2) were prepared at every scheduled or extra visit. Patients with axillary temperatures\u00a0\u226537.5\u00a0\u00b0C were symptomatically treated with paracetamol (acetaminophen) (10\u00a0mg/kg). A rapid diagnostic test (RDT) OptiMAL\u00a9  was applwith T14 .Thick and thin blood smears were stained with 10\u00a0% Giemsa (pH 7.2) for 5\u00a0min. The two thick smears were independently examined under a light microscope by two trained laboratory technicians by using oil immersion 100\u00d7. Asexual and sexual parasite densities were determined by counting the number of parasites found per 200 white blood cells (WBC) assuming 7000 WBC/\u03bcl of blood , 25. In P. vivax IgG antibody titre , were subjected to molecular diagnosis due to suspicion of recent malaria infection. Six 5-mm punches of filter paper impregnated with dried blood were used to extract DNA, using the QIAamp\u00ae DNA Blood Mini Kit  and following the manufacturer\u2019s instructions. The DNA obtained was suspended in 50\u00a0\u00b5l of water and stored at \u221220\u00a0\u00b0C until used. The presence of the P. vivax 18S small sub-unit ribosomal RNA gene was assessed as previously reported [P. vivax: 500, 2000 and 10,000 p/\u00b5l) and uninfected blood samples were included as controls.Molecular diagnosis has proven to be very sensitive in different regions affected by malaria , 27. Allreported . InfecteP. vivax blood stage can last for months or years in exposed patients. However, antibody titres fade out more rapidly in treated patients [P. vivax re-exposure, with and without clinical symptoms and/or low parasitaemia in recurrent blood infections, retrospective research was conducted at the end of the 12-month follow-up period to look for native P. vivax blood-stage proteins in preserved blood samples by using ELISA, as previously reported [\u00ae model EL312) at 405\u00a0nm. Cut-off values were previously determined using unexposed individuals as the mean and 2SD as 0.25 OD values (95\u00a0% confidence). The ELISA-OD values were plotted according to time point per patient.Antibodies against the patients \u201331. To rreported . BrieflyP. vivax infected blood was analysed for cspr and msp3\u03b1 by polymerase chain reaction and restriction fragment length polymorphism (PCR\u2013RFLP) [P. vivax genotypes among groups was examined by Fisher\u2019s exact test (\u03b1\u00a0=\u00a00.05). In addition to cspr and msp3\u03b1, parasite genotypes were analysed using msp3\u03b2 after Alu I digestion to compare parasites producing primary and secondary episodes in each patient [To determine the frequency of parasite genotypes, all CR\u2013RFLP) . The hom patient .For each patient that completed the supervised treatments and the follow-up (for 28\u00a0days and 12\u00a0months post-diagnosis), parasitological and clinical data were analysed using STATA v12. To discard the possibility that a sub-dose caused delay in parasite clearance, the body weight recorded on day 0 was used to determine the accuracy of doses of CQ and PQ given to patients using age group dosing.The first endpoint, parasitological and clinical cure of the primary blood infection, was evaluated by determining the early clinical and parasitological outcome at 2\u20133\u00a0days post-diagnosis, and the possible clinical and/or parasitological asexual parasite persistence or reappearance within seven to 28\u00a0days post-diagnosis. Therapeutic failure (TF) was defined as the presence of parasitaemia, while an adequate clinical and parasitological response (ACPR) was considered with the absence of parasites on days 7\u201328: for these analyses, both microscopic and PCR results were included.P. vivax genotype on day 2 or 3. The Wilcoxon rank sum test was used for non-parametric comparisons: age, asexual and sexual parasitaemia, drug doses, number of days of symptoms, and axial temperature. All tests of significance were two-tailed and P values\u00a0<\u00a00.05 were considered statistically significant.Fisher\u2019s exact test was used to compare two or more independent proportions between treatment groups, considering gender, PCR positivity, parasite persistence, the presence of symptoms, and the distribution of the P. vivax blood infection (first recurrent case) detected at each month of sampling, divided by the total number of patients followed up. The Z-test was used to determine differences in the proportion of patients or samples with a recurrent infection. To plot the cumulative incidence of patients with the first recurrence, the Kaplan\u2013Meier failure estimate was used.The second endpoint evaluated was by determining whether or not recurrent blood infections existed during the follow-up ending 12\u00a0months after day 0. The effectiveness was calculated as the per cent of the accumulated number of patients without P. vivax by microscopy. Of the 153 self-reporting malaria-symptomatic patients that satisfied the inclusion criteria, 86 received the T14 treatment and 67 the ISD regimen .Most patients recruited for the study were living in an area of 18\u00a0\u00d7\u00a010 sq km and they were comprised of mostly indigenous and Mexican mestizo populations. Although a higher proportion of T14 patients lived in the Tapachula City and its surrounding areas (zone 1), and a higher proportion of ISD patients lived in the foothills (zone 2)  and 23 (34.3\u00a0%) completed the programmed follow-up visits and the flexible-schedule visits, respectively. Patients missing three continuous scheduled visits were excluded from the follow-up. Thirteen patients were not accessible for one visit during the first 14\u00a0days, and another four missed two follow-up visits. There were nine patients who withdrew, including four on day 2, two on day 3, one on day 7, and two on day 14. Six patients were not willing to be subjected to finger pricking, two moved to inaccessible communities, and another took an extra medication of CQ-PQ on day 3.Out of 86 patients with the T14 treatment, 49 (56.9\u00a0%) completed the 14 supervised doses. Of these, 44 completed all follow-up visits . Three patients missed one visit, on day 21 or 28. There were two withdrawals of patients afraid of finger pricking on day 21.P. vivax according to both methods). There were 32 samples (4.1\u00a0%) only PCR-positive, indicating that parasite density was at the sub-microscopic level. Eight samples (1.22\u00a0%) were microscopy-positive but PCR negative. These discordant samples had parasite densities below 100/\u03bcl . Three samples (with parasite densities below 100/\u03bcl) were positive in only one of the two thick smears.A total of 778 samples obtained from days 2 to 28, examined by both microscopy and PCR (including three samples with only gametocytes), were used to compare the performance of the diagnostic methods. Of these, 90.3\u00a0% had concordant results . The number of PCR-positive samples on days 2 and 3 was higher than that determined by microscopy. On day 2, 41.3\u00a0% (1.87-fold higher than microscopy) and 34.1\u00a0% (2.3-fold higher than microscopy) of samples were PCR-positive for the ISD and T14 treatments, respectively. Persistent parasitaemia, detected by microscopy and/or PCR on days 2 and 3, was inversely correlated to the parasite count on day 0. That is, a higher parasite density  was found on day 0 in patients that earlier cleared blood parasites, compared to the density observed  in patients that on days 2 and 3 still harboured blood parasites (p\u00a0=\u00a00.0375). On the other hand, parasite persistence was not associated with parasite genotype, lower CQ doses, or patient gender or age.No difference in parasitaemia clearance was observed between treatments. Few patients had asexual blood parasites detected by microscopy on days 2 or 3 . The presence of these symptoms was not associated with parasitaemia levels on day 0, parasite persistence (observed on days 2 and 3), gender, or PQ dosage.Light jaundice in T14-treated patients was associated with the number of days with malaria symptoms before recruitment. The group with light jaundice had a higher proportion of patients with more symptomatic days previous to recruitment , compared to patients with no jaundice   (Table\u00a0The comparison between the number and timing of recurrent blood infections in the T14 0) Table\u00a0. Eightee0) Table\u00a0. Upon coP. vivax genotype could be obtained, the primary blood infections matched their respective recurrent parasites , and three patients with genotype cspvk210-msp3\u03b1 B/msp3 \u03b2 III also relapsed (with homologous parasites) after distinct intervals . With all recurrent infections treated with supervised T14, the patients had no further detectable recurrent infections, with one exception .For 16 of the 20 samples from ISD patients in which the es Table\u00a0. No assoP. vivax blood stage were detected by ELISA in 151 of 153 samples, with OD values from 0.3 to\u00a0>3.0 (mean 1.14\u00a0\u00b1\u00a00.73) on day 7 after the initial treatment. Antibody OD values decreased post-treatment, causing nearly 50\u00a0% of the patients to be ELISA-negative within 3\u00a0months. The antibody response in two patients of the ISD group\u00a0remained positive, with low OD values, until the end\u00a0of the study. However, there were no clinical symptoms of a recurrent blood infection or parasitaemia , it was not possible to demonstrate malaria blood infection by microscopy or PCR. No correlation was documented between the low PQ dosage in T14 or ISD and the presence of recurrent blood infections .In the T14 group, recurrent T14 Fig.\u00a0a. AntiboT14 Fig.\u00a0, but in ns Table\u00a0. InteresP. vivax parasitaemia, in a similar way as the first three doses of T14. The effectiveness of a single dose in clearing primary blood infections, reported earlier in El Salvador [The current findings demonstrate a lower performance in preventing relapses exerted by the ISD regimen compared to the T14 standard treatment. Nonetheless, the first ISD single dose (CQ 10\u00a0mg and PQ 0.75\u00a0mg/kg bw) effectively cleared the primary Salvador , NicaragP. vivax infection has been proven in many different geographic sites, including Korea [P. vivax resistance to CQ has been evidenced. In this region, a combined regimen of CQ and PQ to treat malaria was introduced in the 1950s and is still effective today, probably because PQ can effectively eliminate parasites with low sensitivity to CQ [The efficacy of a standard 25\u00a0mg/kg bw CQ dose to cure ng Korea , Colombing Korea , Peru [3ng Korea , India [ng Korea , Pakistang Korea , China [ng Korea , Thailanng Korea , Afghaning Korea , Vanuatung Korea , Iran [4ng Korea , Ethioping Korea , and Maung Korea . A similng Korea . No dataty to CQ .P. falciparum gametocytes [Primaquine is the only licensed 8-aminoquinolone available to malaria control programmes for the elimination of liver-dwelling hypnozoites. If left untreated, hypnozoites can produce relapses upon reactivation at variable periods of time , 49. Howetocytes \u201313, and In this study, it is likely that most cases of light jaundice, detected in about 12\u00a0% of the patients recruited, were due to infected erythrocyte lysis, and others resulted from PQ administration. The condition of light jaundice occurred mostly in patients under T14 treatment. Indeed, as this treatment progressed there was a greater frequency of jaundice. Although no severe AHA was observed and light jaundice did not worsen in the participants, the current findings warrant further studies to determine G6PDd in the Mexican population affected by malaria infection, as well as its implications for treatment of the same. G6PDd screening would be facilitated by the development of diagnostic tests applicable in field conditions .Mild pruritus was observed in 10\u00a0% of the patients in both treatment groups after the administration of CQ\u2013PQ, a condition that disappeared within 1\u00a0week. CQ-related pruritus has been reported in a high proportion of Africans, in 1.9\u00a0% of Asians , and in P. vivax blood stage as well as the PCR diagnosis were powerful for discovering recurrent parasite exposure and/or infections, albeit retrospectively. The serological test was very sensitive to antibody titres, which increased after re-exposure to parasites (due to recurrent parasitaemia), as confirmed by PCR. Accordingly, the serological follow up confirmed parasite clearance and the absence of new blood infection. It is possible that there were some recurrent infections cleared without treatment, as evidenced by the antibody response. This situation was reported in Peru [Although many recurrent episodes were detected by microscopy, the serological analysis of the IgG antibody response to the  in Peru .P. vivax transmission [P. vivax occur within 1\u00a0year of the primary infection. The low transmission rate in southern Mexico [Relapse episodes represent the most important difficulty that impedes the elimination of smission . Early rsmission , 59\u201361. smission , 62\u201364. smission  and Afghsmission , are alsn Mexico  makes ren Mexico . AccordiP. vivax strains produced higher numbers of long-term relapse episodes. Worldwide, the T14 treatment has proven to be the most effective treatment for eliminating P. vivax relapses [The lack of effectiveness of the ISD regimen was evidenced by the fact that relatively few of the recurrent blood infections (33\u00a0%) were suppressed by the programmed single doses. Moreover, as these patients developed parasitaemia they might have been involved as sources of infection before the ISD was administered. The time interval from the primary to the first short- and long-term recurrent infections, as well as the overall pattern of re-infection, partially resembles the outcome reported in Central America, where relapses . Since trelapses . Similarrelapses  and Colorelapses , in Mexirelapses . In the relapses  and 85\u00a0%relapses  after trrelapses , 71.Although the efficacy of ISD for eliminating primary infections is herein documented, relapses occurred unpredictably at any time within 12\u00a0months after the first dose. This treatment scheme has been abandoned by the Mexican MCP. The present pre-elimination status of Mexico requires new strategies for the most thorough detection and treatment of cases . AlthougP. vivax blood infections, in a similar way as the initial three doses of the T14 scheme. A single combined dose administered to symptomatic patients in remote areas while awaiting parasitological diagnosis may contribute to halting P. vivax transmission. The supervised T14 treatment was much more effective for preventing recurrent infections. Moreover, the semi-supervised T14 (with family supervision) was also effective for patients that did not strictly comply with the supervised scheme. On the other hand, it is clear that at the individual level, ISD scored very low compared to T14 in preventing relapses and should therefore be abandoned. Further research should be conducted to determine the ideal PQ dosing.The first dose of CQ-PQ in the ISD regimen effectively cleared primary clinical and parasitological"}
+{"text": "Case management based on prompt diagnosis and adequate treatment using artemisinin-based combination therapy (ACT) remains the main focus of malaria control in Ghana. As part of routine surveillance on the therapeutic efficacy of ACT in Ghana, the efficacy of amodiaquine-artesunate (AS-AQ) and artemether-lumefantrine (AL) were studied in six sentinel sites representing the forest and savannah zones of the country.Three sites representing the two ecological zones studied AS-AQ whilst the other three sites studied AL. In each site, the study was a one-arm prospective evaluation of the clinical, parasitological, and haematological responses to directly observed therapy for uncomplicated malaria with either AS-AQ or AL among children aged 6\u00a0months and 9\u00a0years. The WHO 2009 protocol for monitoring anti-malarial drug efficacy was used for the study between July 2013 and March 2014.Per-protocol analyses on day 28 showed an overall PCR-corrected cure rate of 100\u00a0% for AS-AQ and 97.6\u00a0%  for AL: 97.2\u00a0%  in the forest zone and 100\u00a0% in the savannah zone. Kaplan\u2013Meier survival analysis showed similar outcomes. Prevalence of fever decreased by about 75\u00a0% after the first day of treatment with each ACT in the two ecological zones. No child studied was parasitaemic on day 3, and gametocytaemia was generally maintained at low levels (<5\u00a0%). Post-treatment mean haemoglobin concentrations significantly increased in the two ecological zones.Therapeutic efficacy of AS-AQ and AL remains over 90\u00a0% in the forest and savannah zones of Ghana. Additionally, post-treatment parasitaemia on day 3 is rare suggesting that artemisinin is still efficacious in Ghana. Malaria remains a major public health problem in the world after decades of deployment of different interventions. Globally, an estimated 3.3 billion people are at risk of malaria infection and disease. It is estimated that in 2013 there were approximately 198 million cases of malaria worldwide, representing a decline of 30\u00a0% since year 2000. In the same year, malaria accounted for approximately 584,000 deaths worldwide, representing a decline of 47\u00a0% since year 2000. Africa remains the continent with the heaviest burden of malaria, accounting for about 90\u00a0% of all malaria deaths in the world. It is also estimated that about 78\u00a0% of all malaria deaths occur among children aged under 5\u00a0years [Case management based on prompt diagnosis and adequate treatment remains a major focus of malaria control in Ghana. Other interventions include intermittent preventive treatment among pregnant women (IPTp), long-lasting insecticide-treated nets (LLINs), indoor residual spraying (IRS), and recently seasonal malaria chemoprevention (SMC) . EvidencSince 2005 the Noguchi Memorial Institute for Medical Research (NMIMR), in collaboration with the National Malaria Control Programme (NMCP), has been monitoring the therapeutic efficacy of first-line anti-malarial drugs from sentinel sites across Ghana with the view of providing continuous data to inform malaria treatment policy in the country. This paper covers the therapeutic efficacy of AS-AQ and AL across two ecological zones in Ghana during the period July 2013 to March 2014 using the 2009 WHO protocol for surveillance of anti-malarial drug efficacy .The study was a one-arm prospective evaluation of the clinical, parasitological, and haematological responses to directly observed therapy for uncomplicated malaria with either AS-AQ or AL among children aged 6\u00a0months to 9\u00a0years in sentinel sites representing the savannah and forest zones of Ghana. The primary objective of the study was to assess cure rates in the two ecological zones whilst secondary objectives were to assess patterns of fever and parasite clearance as well as changes in haemoglobin levels and gametocyte carriage rates after treatment with AS-AQ and AL.Therapeutic efficacy of AS-AQ was studied in the Yendi Municipal and Wa Regional Hospitals (representing the savannah zone) and Hohoe Municipal Hospital (representing the forest zone). Therapeutic efficacy of AL was studied in Navrongo War Memorial Hospital (representing the savannah zone) and Bekwai Municipal and Begoro District Hospitals (representing the forest zone).The Yendi Municipal is located in the Northern region of Ghana with an estimated municipal population of 117,780 and an annual average rainfall of 1125\u00a0mm. Wa regional hospital is located within Wa municipal in the Upper West region of Ghana with an estimated municipal population of 107,214 and an annual average rainfall of between 840\u00a0mm and 1400\u00a0mm. The Navrongo War Memorial Hospital is located in the Kassena Nakana East district in the Upper East region of Ghana with an estimated district population of 109,944 and an annual average rainfall of 950\u00a0mm. Malaria transmission in the savannah zone of Ghana is generally perennial with marked seasonal variation and an annual entomological rate of about 418 infective bites per person per year \u201315.Hohoe is located in the Volta region of Ghana with an estimated municipal population of 262,046 and an annual average rainfall of 1300\u00a0mm. The rainfall pattern is bimodal with two distinct rainy seasons: April to July as major and September to November as minor. Malaria transmission in the municipality is intense and peaks with the two rainy seasons. Bekwai municipal is located in the Ashanti region of Ghana with an estimated municipal population of 118,024. Annual rainfall in the municipality is 1600\u20131800\u00a0mm with double maxima rainfall in May and October each year. Malaria transmission in the municipality is intense and perennial. Begoro is located in the Fanteakwa district in the Eastern region of Ghana. The Fanteakwa district has an estimated population of 108,614. Annual rainfall in the district is 1500\u20132000\u00a0mm with double maxima rainfall in June and October each year. Malaria transmission in the district is intense and perennial. Annual entomological inoculation rate in the forest zone can be as high as 866 infective bites per person per year , 16\u201318.Plasmodium falciparum; parasite count ranging between 1000 and 250,000 per \u00b5l; haemoglobin level\u00a0>5\u00a0g/dl; and absence of signs/symptoms of severe malaria.Children aged between 6\u00a0months and 9\u00a0years with a history of fever suggestive of malaria were screened for the study. Blood samples from a finger prick were used to prepare thick and thin smears for malaria microscopy and haemoglobin level determination. Children who met the inclusion criteria were recruited into the study after parental consent had been obtained. Inclusion criteria included axillary temperature\u00a0\u226537.5\u00a0\u00b0C or history of fever during the past 24\u00a0h; mono-infection with \u00ae  at 0, 8, 24, 36, 48, and 60\u00a0h. Children weighing 5 to\u00a0<15\u00a0kg were given one tablet per hour of treatment; those weighing 15 to\u00a0<25\u00a0kg were given two tablets per hour of treatment; and those weighing 25 to\u00a0<35\u00a0kg were given three tablets per hour of treatment. All tablets were dissolved in water and given under direct observation by a study nurse and children were observed for 30\u00a0min to ascertain retention of anti-malarial. Children who vomited during the observation period were re-treated with the same dose of anti-malarial and observed for an additional 30\u00a0min. Children with repeated vomiting were given parenteral therapy with quinine as per national standard treatment guidelines and excluded from the study. All children were allowed use of antipyretics. Children who showed signs/symptoms of severe malaria were withdrawn from the study.All anti-malarials used in the study were fixed-dose combinations, and were supplied by the WHO, Geneva. For sites studying AS-AQ, children enrolled received single daily weight-based products from Sanofi Aventis. The products were 25/67.5\u00a0mg  for children weighing 4.5 to\u00a0<9\u00a0kg; 50/135\u00a0mg  for children weighing 9 to\u00a0<18\u00a0kg; and 100/270\u00a0mg  for children weighing 18 to\u00a0<36\u00a0kg. For sites studying AL, children enrolled received weight-based 20/120\u00a0mg of Coartemmsp1, msp2), and glutamate-rich protein (glurp) were used to distinguish between re-infection and recrudescence [Children enrolled into the study were followed-up for 28\u00a0days. Children were seen at the outpatient department on days 1, 2, 3, 7, 14, 21, and 28 (day of commencement of treatment was counted as day 0). At each follow-up visit, the children were clinically examined by a study Physician, who recorded findings on a Case Record Form (CRF). Parasitaemia levels  were assessed on days 2, 3, 7, 14, 21, 28, and any day within the 28-day follow-up period that a child is brought to the clinic with fever. Thick and thin smears were stained with 3\u00a0% Giemsa for 30\u201345\u00a0min for quantification of asexual parasites against 200 white blood cells using a hand tally counter. Sexual parasite counts were done per 1000 white blood cells. Parasitaemia levels were expressed per \u00b5l blood assuming white blood cell count of 8000 per \u00b5l blood. A smear was declared negative when examination of 100 thick-film fields did not show presence of asexual parasites. For quality control purposes, all blood slides were read by two qualified independent microscopists, and discordant readings were re-examined by a third qualified independent microscopist. Discordance was defined as differences between the first and second microscopists regarding presence/absence of asexual or sexual parasites; species diagnosis; and day 0 counts meeting the inclusion criterion of 1000\u2013250,000 per \u00b5l blood. The first or second reading was taken as final depending on whichever agrees with the third reading. Filter paper blots were obtained on day 0 and at recurrence of parasitaemia for PCR genotyping. Merozoite surface proteins 1 and 2 , Late Parasitological Failure (LPF), Late Clinical Failure (LCF), and Adequate Clinical and Parasitological Response (ACPR) [The Institutional Review Board of the Noguchi Memorial Institute for Medical Research, University of Ghana, reviewed and approved the study. Written informed consent was obtained from each parent/guardian at the start of the study. Each parent/guardian was informed of the objectives, methods, anticipated benefits and potential hazards of the study. They were also informed that they were at liberty to withdraw their children from the study at any time without penalty.A cumulative total of 310 children participated in the study in the two ecological zones: 140 received AS-AQ and 170 received AL. Of the children who received AS-AQ, 81 (57.9\u00a0%) were within the savannah zone. Of the children who received AL, 124 (72.9\u00a0%) were in the forest zone. For both anti-malarials, there were no significant differences between the two ecological zones, in terms of male/female ratio as well as baseline mean haemoglobin levels  in the savannah zone and remained 1.000 in the forest zone (p\u00a0=\u00a00.895). Incidence of treatment success with AS-AQ remained 1.000 and 0.987 in the forest and savannah zones, respectively until day 21, when both zones experienced a decline to 0.981  and 0.974 , respectively (p\u00a0=\u00a00.721), and remained same between day 21 and day 28. For AL, PCR-uncorrected treatment success remained 1.000 in the two ecological zones until day 14 when cumulative incidence of treatment success declined to 0.984  in the forest zone and 0.929  in the savannah zone (p\u00a0=\u00a00.208). Cumulative incidence of treatment success further declined on day 21 to 0.942  in the forest zone and 0.667  in the savannah zone (p\u00a0<\u00a00.001). Cumulative incidence of treatment success remained 0.942 in the forest zone until day 26, when it declined to 0.934 , and further declined to 0.851  on day 28 compared with 0.601  on the same day in the savannah zone (p\u00a0=\u00a00.004)   on days 2 and 3 after treatment with AS-AQ, and remained absent from day 7 to day 28. For AL, gametocytaemia in the forest zone declined from 3.3\u00a0%  on day zero to 0.9\u00a0%  on day 28 (p\u00a0=\u00a00.413), and was present only on day 7  in the savannah zone.Mean haemoglobin concentration in the forest zone significantly increased from 8.7 (\u00b11.0) at baseline to 9.2 (\u00b10.8) on day 28 after treatment with AS-AQ (p\u00a0=\u00a00.006). Similarly, mean haemoglobin concentration in the same zone significantly increased from 10.2 (\u00b11.6) at baseline to 10.7 (\u00b11.3) on day 28 after treatment with AL (p\u00a0=\u00a00.011). Within the savannah zone, mean haemoglobin concentration significantly increased from 8.7 (\u00b11.9) at baseline to 10.3 (\u00b12.1) after treatment with AS-AQ (p\u00a0<\u00a00.001) Table\u00a0.Table\u00a04CThe therapeutic efficacy of two of Ghana\u2019s first-line anti-malarial drugs for the treatment of uncomplicated malaria (AS-AQ and AL) were studied in six sentinel sites representing the forest and savannah zones during the 2013/2014 surveillance period, with the view of providing information to guide malaria treatment policy in the country.The study showed that PCR-uncorrected cure rates for AS-AQ were 98\u00a0%  in the forest zone and 97.3\u00a0%  in the savannah zone whilst AL showed PCR-uncorrected cure rates of 85\u00a0%  in the forest zone and 56.3\u00a0%  in the savannah zone. Although the study was not a comparative study, the findings compare well with the observation of higher PCR-uncorrected cure rate for AS-AQ, compared with AL, in Mozambique . This obBoth AS-AQ and AL achieved effective fever and parasite clearance in the two ecological zones. Prevalence of fever in both zones declined by 75\u00a0% after first day of treatment with each ACT; no child was parasitaemic on day 3; and prevalence of gametocytaemia generally declined from 3.3 to 0.9\u00a0% within the 28-day follow-up period. These findings suggest that ACT still has a rapid effect on fever and parasite clearance as well as gametocyte carriage in Ghana , 27, 28.Post-treatment mean haemoglobin concentrations assessed on day 28, compared with pre-treatment concentrations, showed significant increase with both AS-AQ and AL treatment in the forest zone, and AS-AQ in the savannah zone (there was no available post-treatment data for AL treatment in the savannah zone). These findings suggest that ACT treatment for uncomplicated malaria has a positive effect on post-treatment haemoglobin levels. Though some studies have not shown significantly improved post-treatment haemoglobin levels after ACT treatment , 28, 30,Therapeutic efficacy of AS-AQ and AL remains over 90\u00a0% in the forest and savannah zones of Ghana. The two drugs were effective in clearing fever and parasites as well as maintaining gametocytaemia at low rates (less than 5\u00a0%), and improving post-treatment mean haemoglobin levels. Artemisinin is still efficacious in Ghana by virtue of the absence of parasitaemia on day 3 post-treatment."}
+{"text": "Hemolysis occurred in a low proportion of patients and did not increase transfusion requirements. Artesunate is the most effective treatment for severe malaria. However, delayed-onset hemolytic anemia has been observed in \u224820% of travelers who receive artesunate, \u224860% of whom require transfusion. This finding could discourage physicians from using artesunate. We prospectively evaluated a cohort of 123 patients in France who had severe imported malaria that was treated with artesunate; our evaluation focused on outcome, adverse events, and postartesunate delayed-onset hemolysis (PADH). Of the 123 patients, 6 (5%) died. Overall, 97 adverse events occurred. Among the 78 patients who received follow-up for >8 days after treatment initiation, 76 (97%) had anemia, and 21 (27%) of the 78 cases were recorded as PADH. The median drop in hemoglobin levels was 1.3 g/dL; 15% of patients with PADH had hemoglobin levels of <7 g/dL, and 1 required transfusion. Despite the high incidence of PADH, the resulting anemia remained mild in 85% of cases. This reassuring result confirms the safety and therapeutic benefit of artesunate. Intravenous (IV) artesunate has been the recommended first-line treatment for severe malaria worldwide since 2010 . Data were collected and analyzed anonymously. After diagnosis, all patients received IV artesunate (2.4 mg/kg) at 0, 12, and 24 hours and daily thereafter until oral antimalarial treatment could be administered to complete treatment as recommended in France  became available in France through the Agence Nationale de S\u00e9curit\u00e9 du M\u00e9dicament, the French national drug agency. The product, manufactured by Guilin Laboratories in China, was imported to Europe by ACE Pharmaceuticals. Within the framework of a temporary use authorization program, data were prospectively collected during May 2011\u2013May 2013 from medical charts and by using Agence Nationale de S\u00e9curit\u00e9 du M\u00e9dicament forms that were completed by attending physicians at the beginning and end of treatment. A dedicated team at the National Reference Center for Malaria (NRCM) retrieved the data. Additional data were obtained from the national pharmacovigilance system and an NRCM database, as described . Hemolysis was defined as a plasma haptoglobin level of <0.1 g/L , plasma lactic dehydrogenase (LDH) level of >390 IU/L , or both. We defined 3 patterns of anemia as previously described , 7 (\u00b12), 14 (\u00b13), 21 (\u00b13), and 28 (\u00b13) after treatment initiation .The death rate at day 28 was the main clinical endpoint. Parasitological cure was defined as a Travel characteristics and demographic, clinical, and laboratory variables were evaluated. Quantitative variables were expressed as medians (quartiles 1\u20133 [Q1\u20133]) or, when appropriate, as means (SEMs). Qualitative variables were expressed as percentages. Differences between groups  were analyzed by using the Fisher exact test for categorical variables and Mann-Whitney test for continuous variables. Statistical analyses were performed by using IBM SPSS Statistics version 20 . All reported p values are 2-tailed.A study flowchart is provided in Among the 123 patients with severe malaria treated with artesunate, 117 fully or partially recovered and 6 died from malaria . All deaths were related to severe multiorgan failure and occurred within 3 days of receiving artesunate. = 0.098), and severe anemia at day 0 were not significantly associated with death. The median time between symptom onset and initiation of artesunate treatment was 1.5 days (Q1\u20133: 1\u20135) in the 6 patients who died versus 4.0 days (Q1\u20133: 2\u20135) in those who survived (p = 0.13). Artesunate was used as second-line treatment after quinine in 2 of 6 patients who died versus 49 of 117 patients who survived (p = 1). All survivors had complete parasite clearance before treatment day 7. Only 1 relapse was observed; it occurred 26 days after a 3-day course of IV artesunate that was not followed by the recommended oral course of antimalarial drug therapy.The following characteristics were seen more frequently at admission in patients who died versus those who survived: lower median Glasgow Coma Scale (median score 10 [Q1\u20133: 3\u201313]) vs. 14 [Q1\u20133: 14\u201315], p = 0.001; reference score 15); respiratory distress ; higher median parasitemia level ; higher median lactate level ; higher total bilirubin concentration ; lower glucose level ; and renal insufficiency . Age, sex, immunocompromised state, place of malaria acquisition, cardiocirculatory impairment  with liver toxicity as a potential side effect . All these episodes occurred before day 8. Vision loss occurred in 1 patient and was considered by the attending ophthalmologist to be associated with hypertensive retinopathy. Two cases of acute cerebellar syndrome occurred; both were thought to be associated with a postmalaria neurologic syndrome. Tinnitus was reported in 1 patient who received quinine just before artesunate. One patient experienced continuous tremors that resolved spontaneously. Liver enzyme levels increased in 8/117 patients who survived, including 6 who concurrently received QTc lengthening  and transient bradycardia were recorded for 1 and 2 patients, respectively. One transient bradycardia (54 bpm) episode occurred between 2 artesunate injections and resolved spontaneously. In the patient with QTc lengthening , artesunate treatment  was changed to artemether/lumefantrine  and then to atovaquone/proguanil  because of persistent QTc lengthening (560 ms) accompanied by low potassium levels . These 2 patients were 13 and 15 years of age and weighed 62 kg and 40 kg, respectively. Another patient experienced severe disseminated intravascular coagulation that led to arterial ischemia of extremities and central nervous system ischemia . Amputation of fingers and legs was necessary. All sequelae in this patient were considered related to severe malaria. One patient with myasthenia gravis received artesunate and experienced no worsening of the disease   , 3. The = 0.002) ; these c= 0.002) . During = 0.002) . Maximum= 0.002) ; Table 3Of the 78 patients, 15  received a total of 20 blood transfusions, 15 (75%) of which were performed before day 8, during the acute phase of the disease . Among tAccording to NRCM data, each year in France, \u2248250 patients are treated for severe imported malaria. In this cohort of 123 patients treated for severe malaria in high-care settings, IV artesunate was effective and generally safe. The death rate was 5%, and blood transfusion was necessary for <20% of all patients and for <5% of patients with PADH anemia. Compared with retrospective case series, our prospective approach reduced bias toward severe anemia cases and provided a robust evaluation of artesunate safety, particularly as concerns PADH anemia.P. falciparum malaria cases treated with quinine (10% death rate), the major factors associated with death were low Glasgow Coma Scale score, respiratory failure, severe renal impairment, hyperlactemia, or hypoglycemia , reticulocytopenia (0.6%), and elevated liver enzymes (1.1%) . Nevertheless, it is unlikely that severe cases of delayed hemolysis or other severe side effects would be overlooked in a temporary use authorization program implemented at facilities with high levels of care, and, as mentioned, this study captured a fairly high proportion of mild to moderate cases of delayed hemolysis. Furthermore, the study was not randomized, but in the setting of severe imported malaria, it is considered unethical to repeat artesunate versus quinine trials already performed in malaria-endemic countries (Our prospective analysis joins other reports (Baseline data and complementary qualitative features of anemia for patients with severe imported malaria treated with artesunate, France, 2011\u20132013."}
+{"text": "Plasmodium falciparum malaria were randomly allocated to each study arm. Children were hospitalized for 3 days, given supervised treatment, and followed up weekly for 42 days. All regimens were well tolerated and rapidly effective. The median parasitemia clearance half-life was 2.2 h, and half-lives were similar between arms (P = 0.19). The PCR-uncorrected cure rates by day 42 were 73.0% for amodiaquine-artesunate, 70.2% for artemether-lumefantrine, and 86.3% for dihydroartemisinin-piperaquine (P = 0.001). Early treatment failure occurred in three patients (0.5%), one in each arm. The PCR-corrected cure rates were 93.4% for amodiaquine-artesunate, 92.7% for artemether-lumefantrine, and 94.3% for dihydroartemisinin-piperaquine (P = 0.78). The last provided a longer posttreatment prophylactic effect than did the other two treatments. The day 7 plasma concentration of piperaquine was below 30 ng/ml in 47% of the children treated with dihydroartemisinin-piperaquine, and the day 7 lumefantrine concentration was below 280 ng/ml in 37.0% of children who received artemether-lumefantrine. Thus, although cure rates were all satisfactory, they could be improved by increasing the dose. An open-label, randomized controlled trial was carried out in 2011\u20132012 in the Democratic Republic of the Congo to test the efficacy, safety, and tolerability of the artemisinin-based combination treatments dihydroartemisinin-piperaquine, amodiaquine-artesunate, and artemether-lumefantrine. Six hundred eighty-four children aged 3 to 59 months with uncomplicated  Plasmodium falciparum malaria consists of amodiaquine-artesunate (AA) or artemether-lumefantrine (AL), although artemether-lumefantrine, which was introduced in 2010, has very limited availability in the public sector. Amodiaquine-artesunate remains the most widely distributed antimalarial therapy in DRC. It was introduced in 2006, replacing sulfadoxine-pyrimethamine, which is now used only as an intermittent preventive treatment in pregnancy. The distribution and access of antimalarials in the rural areas of the country are organized through the public sector, whereas in the urban setting the private sector is predominant. Due to the civil unrest that has affected the country for many years, there is a paucity of data concerning the efficacy of antimalarial drugs in DRC. Available studies show substantial geographic variation in therapeutic efficacy, with similar variation in the prevalence of polymorphic alleles in P. falciparum genes associated with parasitological failure (2The Democratic Republic of the Congo (DRC) is one of the five countries with the greatest malaria burden in the world . The curfailure 24. This rDihydroartemisinin-piperaquine (DP) is an artemisinin-based combination therapy (ACT) with a good safety and tolerability profile which is as effective as other ACTs in areas of endemicity in Asia and Africa . PiperaqP. falciparum malaria in children in Kinshasa, DRC, 5 years after its introduction as a first-line treatment, and to compare this with the efficacies of potential alternatives, dihydroartemisinin-piperaquine and artemether-lumefantrine, the latter recently added to the first-line treatment policy. The study was registered with the International Standard Randomized Controlled Trial Number Register (www.isrctn.org) under registration no. ISRCTN20984426.The aim of this trial was to assess the efficacy of amodiaquine-artesunate for the treatment of uncomplicated The study was carried out in a research center located in an urban district of Kinshasa (DRC). Malaria transmission in the area is intense and perennial, with two annual peaks corresponding to the rainy seasons.P. falciparum, parasitemia density between 2,000 and 200,000 asexual parasites/\u03bcl, axillary body temperature of \u226537.5\u00b0C or history of fever in the preceding 24 h, hemoglobin level of \u22655.0 g/dl, and ability to take oral medication. Patients with severe malaria , artemether-lumefantrine (AL) and amodiaquine-artesunate (AA).Patients were randomly allocated to receive one of the three study treatments.AA  was administered once a day for 3 days according to body weight at a mean dosage of 3.8 mg/kg of body weight/day of artesunate and 10.2 mg/kg/day of amodiaquine. Three types of fixed-dose tablets were used, containing artesunate at 25 mg, 50 mg, or 100 mg plus amodiaquine at 67.5 mg, 135 mg, or 270 mg. Tablets were administered according to the manufacturer's instructions: 4.5 to 8.9 kg of weight, 1 tablet of 25 mg artesunate/67.5 mg amodiaquine; 9 to 17.9 kg, 1 tablet of 50 mg artesunate/135 mg amodiaquine; and 18 to 35.9 kg, 2 tablets of 100 mg artesunate/270 mg amodiaquine.DP  was administered once a day for 3 days according to body weight with the following scheme: 5 to 7.9 kg, 0.5 tablet; 8 to 9.9 kg, 0.75 tablet; 10 to 14.9 kg, 1 tablet; 15 to 20.9 kg, 1.5 tablets; and 21 to 29.9 kg, 2.0 tablets. This dosage scheme was different from the one recommended by the manufacturer . We used 5 intervals of weight instead of 3 to improve the therapeutic dose of dihydroartemisinin at the upper limit of the range for each interval. The mean dosage was 3.3 mg/kg/day of dihydroartemisinin and 26.6 mg/kg/day of piperaquine.AL  was administered in 6 doses over 3 days . Each tablet contained 20 mg artemether and 120 mg lumefantrine, and the mean dose was 2.0 mg/kg of artemether and 12.7 mg/kg of lumefantrine for each dose. Tablets were administered according to the manufacturer's instructions: 5 to 14.9 kg, 1 tablet; 15 to 24.9 kg, 2 tablets; 25 to 34.9 kg, 3 tablets; and >35 kg, 4 tablets.Tablets were administered under medical supervision and with 100 ml of milk (the fat in the milk improves drug absorption). Patients were observed for 1 h after drug ingestion. The full dose was repeated if the patient vomited within 30 min, and a half-dose was given if the patient vomited within 1 h.Patients who failed treatment were treated with intravenous (i.v.) quinine if severe  or oral quinine if uncomplicated (10 mg/kg three times daily for 7 days) according to national policy. Parenteral artesunate was not available.All children were hospitalized for 3 days and followed up actively once a week for 42 days after treatment. Caretakers were invited to come back to the center or to contact the study nurse in the case that the child was unwell. If the patient did not report for the scheduled visits, every effort was made to locate him or her at the home address. At each visit, the medical history, clinical signs and symptoms, body temperature, and a blood sample for parasitemia were collected.n = 684).For the calculation of the sample size, we assumed a cure rate of 95% with AL, 99% with DP, and 85% with AA. A sample size of 621 patients would have been adequate to detect a 10% difference between the standard treatment (AA) and AL or DP at the 5% level and with 90% power. The sample size was increased by 10% to allow for loss to follow-up , Bangkok, Thailand. Patients were enrolled by the study physician and assigned to treatment by the study nurse who opened the next consecutively numbered envelope. Once an envelope was opened, the patient was considered included in the study.The primary outcome measure was the PCR-corrected cure rate by day 42. Secondary outcome measures were parasite and fever clearance and occurrence of adverse events (AE). Treatment outcome was established according to the standard WHO classification . Early tSafety reporting was performed according to the ICH Harmonized Tripartite Guideline for Good Clinical Practice .Asexual and sexual malaria parasites were identified and counted on Giemsa-stained thick films and reported per 200 leukocytes (WBC), assuming a total WBC count of 8,000/\u03bcl . Slides 50) and (ii) as parasite clearance rate derived from the log-linear section of the log parasitemia-time curve and expressed as the parasite clearance half-life .Blood films were prepared at baseline and 6 and 12 h and then repeated every 12 h until 2 consecutive negative blood films were observed. Parasite clearance was assessed (i) as the time for the parasite count to decrease to 50% of its initial value (PC1/2 measured in this study with the more recent data collected in 2013 during the Tracking Resistance to Artemisinin Collaboration (TRAC) project, all slides were read a second time after the study was terminated by the same microscopist team using a different counting technique: if more than 20 parasites were seen on the thick smear after 10 fields, parasitemia per 1,000 erythrocytes (RBC) was counted on the thin smear. Below that threshold, parasites were counted on the thick smear per 500 WBC.To compare the PCtHemoglobin was measured on admission using a portable photometer . Thereafter, the hematocrit was measured at baseline, daily during the hospitalization, and at days 7 and 14 of the follow-up by microhematocrit centrifugation .Total and differential WBC counts were assessed daily during the hospitalization and at day 7 and 14 of the follow-up .Liver function tests were performed, and aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatinine levels were measured from plasma at the hospital laboratories (SEAC-Screenmaster) at baseline and 48 h.A dried blood spot (DBS) was prepared at admission, daily during the hospitalization, and at each follow-up visit for further molecular analysis.A random sample of tablets of DP was analyzed for content and quality at the Department of Pharmacology of MORU, and 2 ml of venous blood was taken at day 7 from 246 consecutive patients to measure plasma concentrations of lumefantrine and piperaquine.Paired filter paper samples from enrollment and the follow-up day on which parasites were detected by microscopy were analyzed at the Shoklo Malaria Research Unit (SMRU) to distinguish between recrudescence and reinfection. Parasite DNA was purified , and the three polymorphic markers MSP-1, MSP-2, and GLURP were genotyped. A recrudescent infection was defined as one that matched in size at least one allele of each marker between the first and second samples. If any pair of alleles of a polyclonal primary infection was detected during a second episode, this was considered a recrudescence.The study was approved by the Oxford University Research Ethic Committee (OXTREC), the Institutional Review Board of Kinshasa School of Public Health (KSPH), and the Ministry of Public Health of DRC. A verbal consent was obtained from caretakers before screening children for malaria and anemia. A written consent form was obtained from caretakers whose children fulfilled all inclusion criteria before enrolling the patient in the study.The study was monitored regularly by a qualified internal monitor  for adherence to Good Clinical Practice (GCP) regulations. All Investigators and the Research Ethic Committee (REC) of KSPH were notified of serious adverse events (SAEs).2 was used to compare proportions. Analysis of variance (ANOVA) was used for normally distributed continuous data, and the nonparametric Kruskal-Wallis test was used to analyze continuous data with a nonnormal distribution.Data were double entered in Microsoft Access 2007 and validated using Epi Info 6.4b . Statistical analyses were performed using STATA v.11 . Descriptive statistics were used to summarize demographic data and baseline values. For the per-protocol analysis, \u03c7For the intention-to-treat (ITT) analysis, the log rank test was used to test the equality of the survivor function across groups and Cox regression was used to estimate hazard ratio of infections posttreatment.The overall fractional reduction in hematocrit was defined as the difference between the patient's lowest level of hematocrit and that at baseline  divided by the hematocrit at baseline. The percentages of patients whose hematocrit fell >20% or 25% were compared between groups. No interim analyses for efficacy or futility were done.Between September 2011 and November 2012, 684 patients were included in the study, 228 in each treatment group. Forty-two patients (6.1%) discontinued the study: 5 children were withdrawn during the hospitalization because the families changed their minds and 37 were lost to follow-up between days 7 and 42 . One patient, in the AA group, died at day 29 from causes unrelated to malaria or the study drug. These cases were not included in the per-protocol analysis, and they were censored on the last day that the patients were visited by the doctor and tested for malaria in the intention-to-treat (ITT) analysis. The flow of patients through the study is outlined in the patient flow diagram .At enrollment, patients had similar demographic, clinical, and parasitological characteristics . The tabP = 0.001) , 92.7% for AL , and 94.3% for DP  (P = 0.76).The cure rates by day 42 (primary outcome), PCR uncorrected, were similar in patients treated with AA (73.5%) and those treated with AL (70.6%), whereas the cure rate was significantly higher in patients treated with DP (86.8%) (Early treatment failure occurred in three patients (0.5%), one in each arm. Data are reported also using day 28 cure rates as the endpoint .2 = 18.83, P = 0.0001, and PCR corrected, \u03c72 = 1.00, P = 0.61) (P > 0.0001). The results were not affected by age or initial parasitemia.The ITT analysis showed similar results  . The risP = 0.003; On admission, 26.8% of patients had fever (axillary temperature of \u226537.5\u00b0C). For the other patients, the parent or guardian reported a history of fever in the preceding 24 h as the main reason for seeking a doctor at the health center. After 24 h, 97% of children were afebrile and there was a significantly higher proportion of children with fever in the AL group at day 2  than for AA  and DP  (P < 0.001)  with no significant differences between arms, indicating similar efficacies of the three different artemisinin derivatives (P = 0.08) (All treatments were associated with a rapid clearance of parasitemia. The parasite positivity rate (proportion of children with a positive slide at day 2) was significantly higher in the AL arm (< 0.001) . The med = 0.08) .n = 195/684) of patients were gametocytemic with no significant differences between groups. Treatment with AL resulted in lower gametocyte carriage rates than did the other two treatments in the follow-up period, days 7 to 21 (P < 0.001)  . In 7 chP = 0.65) (P < 0.001).The mean packed cell volume (PCV) at admission was 30.1% . In the first week, there was an overall mean fractional reduction in the PCV of 10%  with no differences between arms ( = 0.65) . HyperpaP < 0.001).A reduction of >25% of the initial PCV value was observed in 14.9% of hyperparasitemic children and 2.3% of nonhyperparasitemic children (P = 0.62). The risk of receiving a blood transfusion was 6.5 times higher in the hyperparasitemic children . By day 14, the levels were comparable to those at admission in all patients.Ten patients developed decompensated anemia within 4 days of recruitment and required a blood transfusion: 3 in the AA group, 4 in the AL group, and 3 in the DP group . Fourteen of these patients developed severe neutropenia (<500 neutrophils/\u03bcl) .The median WBC counts were similar between the treatment groups on recruitment and at each day of follow-up with an increase from day 0 to day 7 to normal values . NeutropMean serum levels of aspartate aminotransferase (U/liter), alanine aminotransferase (U/liter), and creatinine (mg/dl) were similar at baseline among groups. With minor fluctuations (a trend toward a reduction for AST and ALT), the mean levels at day 2 remained similar to those of day 1 with no statistical differences between groups (data not shown).n = 21; 9.2%) than did children treated with AA  or AL  (P = 0.03). The second dose of AL\u2014administered after 8 h\u2014was vomited in seven cases, and taking this into account, the overall difference between treatments during the first day was not significant (P = 0.06). These children were all given a second dose of the drug. On the second day, there was no difference in vomiting postdose between children treated with AA  and those treated with DP , whereas in the AL arm only 2 (0.9%) cases vomited after the first dose and none after the second dose (P = 0.02).During the first day, children treated with DP vomited within 1 h of the first dose significantly more often . There were otherwise no differences in the numbers of AEs between treatment groups. In 17 cases, the adverse event was graded as severe or life-threatening. All cases were classified as unlikely to be related to the study treatment. Ten patients developed decompensated anemia during the hospitalization and required a blood transfusion (described above). There was one case of severe skin eruption 18 days posttreatment (DP), one case of chickenpox 12 days posttreatment (AL), 1 case of abscess 11 days posttreatment (DP), 1 case of leukocytosis 2 weeks posttreatment (AL), and 3 cases of asthenia (2 AL and 1 AA). One child in the AA group died in a different hospital at day 29. The cause of death was unknown, but the death was considered unlikely to have been caused by either malaria or the drug treatment.At least one adverse event was reported in 37.4% of patients during the posttreatment period that was not present on admission or increased in intensity and was classified as possibly or probably related to the study drug. Most AEs were graded as of minor or moderate intensity. The most frequent AEs were weakness, anorexia, and gastrointestinal disorders . However, these symptoms overlap known malaria symptomatology. Anorexia and weakness were reported more frequently in children treated with AA than in those treated with DP and AL during the second and third days of treatment . The plasma level was significantly lower in children weighing <15 kg  than in those weighing \u226515 kg . Accordingly, 43.7% of children weighing <15 kg had a plasma level of \u2264280 ng/ml, considered the cutoff for therapeutic efficacy (P = 0.018). The 7 children in this subsample with a PCR-confirmed recrudescence had a median level of 429 ng/ml , not significantly different from those who successfully cleared the infection .One hundred twenty-one samples of venous blood were collected at day 7 from patients who received AL . The medefficacy , comparer = 0.22; test for trend, P = 0.04). In 47.2% (59/125) of patients, the plasma level was below 30 ng/ml, the previously published threshold associated with therapeutic efficacy , which has been extensively used in DRC since its introduction in 2006.The PCR-corrected day 28 cure rate for AL in the present study was 96.8%, which is comparable to the 97.9% day 28 cure rate observed in the same area 3 years previously .In the subsample analyzed, the drug level of piperaquine at day 7, reflecting the concentration of drug to which residual parasites are exposed and thus predictive of outcome, was suboptimal in 47% of patients. This confirms previous results , 14 showIn this clinical trial, we measured efficacy of treatments administered under supervision, with a glass of milk, and retreatment was given if the first dose was vomited. Drug exposure, in a nontrial setting , is expected to be lower . Dose opAlthough the efficacies in terms of ACPR rates of the three ACTs were comparable, children treated with DP were at lower risk of having a second episode of malaria during the follow-up period because of the longer posttreatment prophylactic effect of DP related to the longer plasma half-life of piperaquine. This chemoprophylactic effect is important in areas of endemicity such as the study area and makes this drug a good candidate for replacing sulfadoxine pyrimethamine for intermittent preventive treatment in pregnant women and children , 19.This study population was characterized by hyperparasitemia, and the initial high levels of parasitemia affected, as expected, the recovery of hematocrit after the initial episode of malaria, but not the treatment efficacy. The initial level of gametocytemia was also high (30%), and AL was significantly more effective in clearing the sexual stages than were the other ACTs. Data in literature on the gametocytocidal properties of the different ACT are conflicting, and the effect, if any, on malaria transmission is unclear .1/2 was 2.2 h  and comparable to the results observed in 2013 during the TRAC project, 2.2 h  (50s with the three therapies (but not with the PCt1/2) could be attributed to the relatively slow conversion of artemether to dihydroartemisinin  compared to artesunate  and dihydroartemisinin in the acute phase of malaria  . The dif malaria , resultiThe three combinations were well tolerated, and there were no significant differences in the types and numbers of adverse events between arms.P. falciparum malaria. Dihydroartemisinin-piperaquine had the longest-lasting chemoprophylactic effect which prevented repeated clinical attacks in the treated children. The recommended dosage of DP provides suboptimal piperaquine plasma concentrations, particularly in small children.The three combinations tested were equally efficacious and well tolerated for the treatment of children with acute uncomplicated"}
+{"text": "Plasmodium falciparum accounts for approximately 60% of malaria cases in Ethiopia and artemether\u2013lumefantrine has been used as a first-line treatment for uncomplicated P. falciparum malaria since 2004. The aim of this study was to assess the therapeutic efficacy of artemether\u2013lumefantrine (AL) for the treatment of uncomplicated P. falciparum malaria in north-western Ethiopia.P. falciparum malaria was conducted in Enfranze Health Centre in accordance with the 2009 WHO efficacy study guidelines. Patients were treated with a 3-day course of AL and clinical and parasitological parameters were monitored over a 28-day follow-up. All data from recruited patients were imported into an electronic data base and Kaplan\u2013Meier survival analysis was used for analysing primary  and secondary  outcomes.A 28-day one-arm, prospective evaluation of the clinical and parasitological response to the first-line treatment for uncomplicated Eighty patients were enrolled and all of them completed the 28-day follow-up period. The PCR-corrected cure rate was 95.0% (95% CI 87.0\u201398.4%) and there were two ETF, one LCF and three LPF. Two of the LPF were classified as re infections by PCR. Seventy three point seven five percent, 91.25 and 95% of patients had cleared their parasitaemia by days 1, 2, and 3, respectively, and 75, 91.25 and 96.25% of patients had cleared their fever by days 1, 2, and 3. All patients completely cleared their gametocytes by day 7.The relatively high cure rate, low proportion of patients still positive on day 3 as well as parasite clearance times in this study would indicate no imminent threat of artemisinin resistance development in the region. However, the threat of spreading or de novo development of artemisinin resistance warrants regular monitoring of drug efficacy throughout the region. Plasmodium and transmitted by female Anopheles mosquitoes. Plasmodium falciparum is by far the most important specie, responsible for nearly all severe malaria cases [Malaria is a disease caused by protozoan parasites of the genus ia cases , 2. Abouia cases .P. falciparum and Plasmodium vivax are the main species accounting for roughly 60 and 40% of malaria cases, respectively [Plasmodium falciparum causes severe malaria with a case fatality rate of about 10% in hospitalized adults and up to 33% in children less than 12\u00a0years old in Ethiopia [Malaria is the leading communicable disease in Ethiopia and an estimated 68% of the population of Ethiopia lives in areas at risk of malaria . In the ectively , 6. PlasEthiopia .P. falciparum malaria, but has also been identified in P. vivax [Early diagnosis and prompt treatment is one of the main strategies in malaria prevention and control and it is also the key to reducing morbidity and preventing mortality . HoweverP. vivax , 11.P. falciparum malaria [In Ethiopia, high level resistance to chloroquine (CQ) in 1998 necessitated a change to sulfadoxine\u2013pyrimethamine (SP) as first-line anti-malarial drug. However, high treatment failure rates with SP of up to 72% were reported in some areas which have led to increasing acceptance of using a combination of two or more drugs in an attempt to reduce malaria transmission and resistance development. Consequently, artemether\u2013lumefantrine (AL) was adopted in 2004, which currently is being used as the first-line drug for the treatment of uncomplicated  malaria , 13.P. falciparum malaria in all endemic countries [P. falciparum to artemisinin has been confirmed in western Cambodia and Thailand [P. falciparum malaria in north-western Ethiopia.The World Health Organization (WHO) recommends artemisinin-based combinations, such as AL, as first-line treatment forum complicated ountries . HoweverThailand . This reThailand \u201319. ThisThailand  and drugThailand . The maiThe study was conducted at Enfranze Health Centre. Enfranze is a sub-district, located in North Gondar administrative zone, Amhara Region, 675\u00a0km north of Addis Ababa and 60\u00a0km from Gondar town and at an elevation of 1,500\u00a0m above sea level. This area is malaria-endemic with a total population of about 45,686  and the majority of the population depends on subsistence farming.P. falciparum malaria conducted between January and May 2013.The design was a one-arm, prospective evaluation of the clinical and parasitological response to directly observed treatment for uncomplicated P. falciparum malaria [The study subjects were recruited among febrile patients attending Enfranze Health Centre using inclusion criteria as defined in the WHO guidelines for assessing the therapeutic efficacy of anti-malarial drugs against  malaria .P. falciparum, above 6\u00a0months of age, a parasitaemia of 1,000\u2014100,000/\u03bcl, weight >5\u00a0kg, presence of axillary temperature (\u226537.5\u00b0C) and no use of anti-malarial drugs 2\u00a0weeks prior to enrollment into the study. Patients with danger signs of severe and complicated malaria according to WHO criteria (including severe anaemia defined as haemoglobin <5\u00a0g/dl), history of allergic reactions to the study drug AL, mixed infection with another Plasmodium species, concomitant presence of febrile conditions with the potential to confound study outcome , severe malnutrition , as well as pregnant and lactating women, were not included in the study.The following inclusion criteria were used for the study: mono-infection with The sample size was determined using a single population proportion formula according to the WHO guidelines: assuming a maximum of 25% clinical failures, 10% precision, and a confidence level of 95% with up to 10% losses to follow up a sample size of 80 was calculated .A rapid screening procedure was used in an outpatient setting to identify patients who meet enrolment criteria. The typical screening data set included age, sex, temperature, body weight, pregnancy test, initial blood slide examination and haemoglobin. All patients meeting the basic enrolment criteria during the screening procedure were evaluated in greater depth by a member of the study team. Physical examination was performed at baseline (day 0 before dosing) and on days 1, 2, 3, 7, 14, 21 and 28. Body weight was determined on day 0 using a weight scale. The screening weight was used to calculate the dose (number of tablets) to be administered. Axillary temperature was measured at baseline (day 0 before dosing) and on days 1, 2, 3, 7, 14, 21 and 28. Female patients of child-bearing age (12\u201349\u00a0years) were asked to provide a urine sample for pregnancy testing before enrolment in the study and if sexually active were provided with condoms for the duration of the study.\u00ae filter paper on day 0 during enrollment and in case of recurrent parasitaemia. The filter paper was air dried and stored in a self-sealing plastic bag with desiccators for further molecular analysis.Finger-prick blood samples were collected from consenting patients for malaria parasite identification and haemoglobin level measurement. Patients that satisfied the criteria were enrolled into the study and followed up on days 1, 2, 3, 7, 14, 21, and 28 where finger-prick samples were taken for microscopic glass slides. Another drop of blood was collected on Whatman 903Thick and thin blood smears were prepared and stained with 10% Giemsa (pH 7.4) for 10\u00a0min and read by two senior microscopists. Blood films were taken at least eight times for each patient during the study period  and during any unscheduled visit. A blood film was considered negative when no parasites were seen after examining 100 high power fields on the thick film. Parasites were counted on thick films relative to 200 leukocytes by two microscopists blinded to each other\u2019s results. Blood smears with discordant results  were re-examined by a third, independent microscopist, and parasite density was calculated by averaging the two closest counts.P. falciparum monoinfections were genotyped. Gene loci\u2014glurp, msp1 and msp2\u2014of these samples were compared by PCR as described previously [In order to differentiate a recrudescence from a newly acquired infection, blood spots were collected from all patients at day 0 (before drug intake) and in case of LPF on Whatman filter paper and sent to Medical University of Vienna for genotyping of merozoite surface protein 1 (MSP1), merozoite surface protein 2 (MSP2) and glutamate-rich protein (GLURP). To exclude mixed infections or infections with other human malaria parasites the samples were analysed with nested PCR for species classification as reported previously , 23. Afteviously .Finger-prick blood samples were used to measure haemoglobin. Due to limited resources the actual haemoglobin concentration should be measured by hemocue. However, this study assessed the haematocrit value only. In healthy persons, the haematocrit (expressed as a percentage) is roughly three times the haemoglobin concentration (expressed in grams per decilitre). This ratio is maintained in normocytic anaemia, but in most of the tropical forms of chronic anaemia the ratio is 3.3:1.\u00ae)  twice daily on days 0, 1, and 2. Study participants were advised to take the study drug with milk to improve absorption. Study medication was administered based on weight; the first and each morning dose were directly observed by the study staff [P. falciparum parasitaemia were treated with quinine.All eligible patients were treated with AL (Coartemdy staff . The evePatients were classified as early treatment failure (ETF), late clinical failure (LCF), and late parasitological failure  as per WHO definition .After checking for completeness all data were imported into Excel and Kaplan\u2013Meier survival analysis was used for analysing primary  and secondary  outcomes, Cox regression was used to identify predictor variables of secondary outcomes. P values <0.05 were considered statistically significant.The study protocol was reviewed and approved by the Ethical Review Committee of the School of Biomedical and Laboratory Sciences, College of Medicine and Health Sciences, University of Gondar. Written informed consent was obtained from all study participants or their legal representatives after being translated and read in the vernacular language.A total of 80 patients  were enrolled. Baseline demographics are presented in Table\u00a0The PCR-corrected cure rate by Kaplan\u2013Meier analysis was 95.0% (95% CI 87.0\u201398.4%). Two participants developed ETF on days 2 and 3, respectively. One participant had a LCF on day 14 and three participants developed LPF on days 21 and 28, respectively, out of which two were classified as re-infections based on molecular analysis of paired samples. The remaining 74 patients completed the follow-up without recurrence of parasitaemia  and gametocyte (P\u00a0=\u00a00.798) clearance times. There was statistically significant association between parasite density at baseline and mean parasite clearance time (P\u00a0=\u00a00.002) Table\u00a0.Table\u00a04PBased on axillary temperatures at baseline participants had similar mean parasite (P\u00a0=\u00a00.760), fever (P\u00a0=\u00a00.329) and mean gametocyte (P\u00a0=\u00a00.498) clearance time Table\u00a0.Table\u00a05TP. falciparum malaria in the region. These results are consistent with studies reported from neighboring Kenya, 96% [This study suggests that with a cure rate of around 95% AL remains an efficacious treatment for uncomplicated nya, 96%  and Burknya, 96%  or Togo,nya, 96%  and slignya, 96% , Senegalnya, 96% , Congo, nya, 96% , Tanzaninya, 96% , southernya, 96%  and soutnya, 96% . Most ofUnlike previous reports from south-western Ethiopia , particiUnlike most previous studies, in this study there were two ETF \u201334. The All except seven participants had cleared parasitaemia by day 2. However, four patients  were still parasitaemic on day 3 (72\u00a0h) after initiation of treatment. All of these patients had initial parasite counts above average and one of them  later developed a LCF. This is considered to be well below the threshold indicating potentially emerging resistance and is comparable to previous findings in southern Ethiopia  and BurkClinical improvement was swift and fever clearance was similarly rapid in most of the participants with only three participants  remaining febrile up to day 3. Fever clearance largely depends on inclusion criteria (e.g. febrile vs. only history of fever) and is, therefore, difficult to compare across study sites but seemed similar to fever clearance reported in previous studies across Africa , 36, 37.The relatively high cure rate, low proportion of patients still positive on day 3 as well as parasite clearance times in this study would indicate no imminent threat of artemisinin resistance development in the region. Artemether\u2013lumefantrine remains highly efficacious in the treatment of uncomplicated falciparum malaria in small as well as older children and adults. However, the threat of spreading or de novo development of artemisinin resistance warrants regular monitoring of drug efficacy throughout the region."}
+{"text": "Asymptomatic and symptomatic malaria during pregnancy has consequences for both mother and her offspring. Unfortunately, there is insufficient information on the safety and efficacy of most antimalarials in pregnancy. Indeed, clinical trials assessing antimalarial treatments systematically exclude pregnancy for fear of teratogenicity and embryotoxicity. The little available information originates from South East Asia while in sub-Saharan Africa such information is still limited and needs to be provided.A Phase 3, non-inferiority, multicentre, randomized, open-label clinical trial on safety and efficacy of 4 ACT when administered during pregnancy was carried out in 4 African countries: Burkina Faso, Ghana, Malawi and Zambia. This is a four arm trial using a balanced incomplete block design. Pregnant women diagnosed with malaria are randomised to receive either amodiaquine-artesunate (AQ-AS), dihydroartemisinin-piperaquine (DHA-PQ), artemether-lumefantrine (AL), or mefloquine-artesunate (MQAS). They are actively followed up until day 63 post-treatment and then monthly until 4\u20136 weeks post-delivery. The offspring is visited at the time of the first birthday. The primary endpoint is treatment failure (PCR adjusted) at day 63 and safety profiles. Secondary endpoints included PCR unadjusted treatment failure up to day 63, gametocyte carriage, Hb changes, placenta malaria, mean birth weight and low birth weight. The primary statistical analysis will use the combined data from all 4 centres, with adjustment for any centre effects, using an additive model for the response rates. This will allow the assessment of all 6 possible pair-wise treatment comparisons using all available data.The strength of this trial is the involvement of several African countries, increasing the generalisability of the results. In addition, it assesses most ACTs currently available, determining their relative \u2018-value-\u2019 compared to others. The balanced incomplete block design was chosen because using all 4-arms in each site would have increased complexity in terms of implementation. Excluding HIV-positive pregnant women on antiretroviral drugs may be seen as a limitation because of the possible interactions between antiretroviral and antimalarial treatments. Nevertheless, the results of this trial will provide the evidence base for the formulation of malaria treatment policy for pregnant women in sub-Saharan Africa.NCT00852423 The risk of malaria is higher in pregnant women than in the general population. There is insufficient information on the safety and efficacy of most antimalarial drugs in pregnancy  as they We propose to assess the efficacy and safety of the most important ACTs currently available, namely artemether-lumefantrine (AL), amodiaquine-artesunate (AQ-AS), mefloquine-artesunate (MQ-AS) and dihydroartemisinin-piperaquine (DHA-PQ). The choice of ACTs is based on several criteria, including known treatment efficacy in children, safety in pivotal phase-3 trials in children and adults, practicality of the dosing regimen , fixed dose combinations, drug tolerance, current availability in the population and affordable cost. The rationale for contemporaneously testing several drug regimens is to shorten the time of data collection and determine the relative \u2018value\u2019 of each treatment, providing the basis for an informed choice by malaria control programmes and policy makers.in vitro drug sensitivity) that may help in interpreting the observed results.Reliable data on the safety and efficacy of ACTs in pregnant women can be rapidly collected only within the context of a randomized-controlled trial. The production of such a large dataset will advance considerably the knowledge on the treatment of malaria in pregnancy in a relatively short period of time compared to pregnancy registers. One of the major issues for this trial is the altered pharmacokinetic of antimalarial drugs during pregnancy and the influence on the outcome of treatment. There has been a recent increase in trials that measure the pharmacokinetics of antimalarial drugs in pregnant women . Recent ACTs have been shown to be extremely efficacious in children . EfficacP. falciparum infection during the second and the third trimester, and collect explanatory variables for therapeutic response. The primary hypothesis is the clinical equivalence (pair-wise non-inferiority) of the 4 treatment regimens, with clinical equivalence defined as difference in treatment failure rates (PCR corrected) of 5% or less.The main objective of the trial is to determine the safety and efficacy of 4 ACTs when administered to pregnant women with Specific objectives are:P. falciparum malaria;to compare the efficacy of AL, AQ-AS, MQ-AS and DHA-PQ in terms of treatment failure by 63\u00a0days after start of treatment with or without genotyping; Parasite clearance time; Haematological recovery by 14, 28, 42 and 63\u00a0days post-treatment and at delivery; Birth weight measured within 72\u00a0hours of delivery; and prevalence of placenta to describe the safety profile of AL, AQ-AS, MQ-AS and DHA-PQ in terms of tolerability; incidence of adverse events until one year post-partum;to determine the relation between drug pharmacokinetics (Day 7 levels of the partner drug) and response to treatment;in vitro susceptibility of P. falciparum isolates collected before treatment to several drugs, including the partner drug tested, and to correlate their IC50 to treatment response.to assess the This is a non-inferiority, multicentre, randomized, open label study on 4 antimalarial treatments, namely DHA-PQ, MQAS, AQAS and AL, assessed in each site using a \u201cbalanced incomplete block design\u201d \u2013 with 3 out of 4 arms used in each site. There are 7 sites in the study distributed in the four countries, i.e. Burkina Faso (Nanoro and Nazoanga), Ghana , Malawi (Chikwawa) and Zambia (Nchelenge).The treatments tested are distributed in a way to allow a head-to-head comparison and the establishment of the treatment\u2019s relative value according to a series of outcomes Table\u00a0. This apThe primary analysis is the assessment of therapeutic equivalence of the 4 treatments  with respect to therapeutic success at day 63 and their safety throughout the follow up, i.e. up to one year after delivery.The randomization list is generated prior to the beginning of the study.The interpretation of the PCR reading is blinded or masked with regard to the treatment allocation of the patients.An independent Data Safety and Monitoring Board (DSMB) reviews all safety data.The following procedures are used to ensure an unbiased assignment of treatment safety and efficacy:Treatment Failure (TF) (PCR adjusted) at day 63 defined according to the WHO criteria [criteria  as the sSafety profiles including significant changes in relevant laboratory values. Subjects are monitored for 63\u00a0days for possible development of adverse events. All adverse events are recorded on the specific form in the electronic CRF. Vital signs, blood chemistry and haematology are monitored and changes in relevant laboratory parameters are assessed.There are two primary end points:PCR unadjusted treatment failure up to day 63 (TF63U); Time to treatment failure (PCR adjusted and unadjusted) during 63\u00a0days of active follow-up after treatment; Asexual parasite clearance time (PCT): Asexual parasite clearance time is defined as the time (in days) from time of randomization to 2 consecutive negative blood slides (collected at different days) - the time to the event is taken as the time to the first negative slide; Gametocytaemia  at day 7, 14, 21, 28 and 63 after treatment; Hb changes day 14, 28, 42 and 63; Acute, chronic or past infection of the placenta ; Mean birth weight and prevalence of low birth weight.\u00ae, Angelholm, Sweden, according to manufacturer\u2019s instructions.Maternal haemoglobin is measured using Hb301 HemocueBlood samples are collected by finger prick at specified time points during the trial for blood slides (thick blood film) and blood spots on filter paper. Thick blood smears are stained with 3% Giemsa for 30\u00a0minutes and read by trained microscopists at each site. Parasite densities are calculated by counting the number of asexual parasites per 200 leukocytes , assuming a leukocyte count of 8,000/\u03bcl. A blood smear is considered negative when the examination of 100 high power fields does not reveal asexual parasites. Each slide is read separately by two experienced microscopists and discrepancies resolved by a third reader.Plasmodium falciparum genome. For the three genes, each PCR-amplification product of a different size is considered to originate from a different clone of Plasmodium falciparum, reflecting a different genotype. For the samples collected from the same patient at day 0 and day of recurrent parasitaemia, the length polymorphism of MSP1, MSP2 and GLURP are determined, i.e. the number of bands in each PCR reaction and their respective size. Results are interpreted as follows:Blood spots on filter paper are used for genotyping recurrent malaria infections during follow up and compared them with pre-treatment samples. This is done by characterizing MSP1, MSP2 and GLURP, single-copy genes in the Recrudescence: For each marker , at least one identical length polymorphism is found in the sample collected at day 0 and day of recurrent parasitaemia.New infection: For at least one marker, length polymorphism is different between the sample collected at day 0 and that at day of recurrent parasitaemia.Indeterminate: Samples that fail to produce a result due to an inability to amplify DNA at day 0 and/or day of recurrent parasitaemia.3 biopsy specimen is obtained from the maternal-facing side of the placenta as soon as possible after delivery. Biopsy specimens are preserved in 10% neutral buffered formalin which are processed and embedded in paraffin wax by standard techniques. Pending histological evaluation, all biopsies are kept at 4\u00b0C. Paraffin sections 4\u00a0mm thick are stained with haematoxylin-eosin stain.A 1\u00a0cmAcute infection Chronic infection Past infection (no parasite but pigment present)No infection Placental biopsies are classified according to the following definitions :Acute inIndividual pharmacokinetic studies are often underpowered for identifying the factors that influence antimalarial pharmacokinetic parameters, which may have a major influence on the observed therapeutic response. Complementing efficacy data with some information on the pharmacokinetic properties of the treatment used allows a better interpretation of the observed recurrent infections or true recrudescences/new infections, as these may be the results of inadequate drug levels because of altered distribution, poor absorption or metabolism. The day 7 drug concentration has been shown to be the most important single measure, in terms of correlation with the area under the concentration time curve and association with treatment response, for lumefantrine, piperaquine and mefloquine . Therefoin vitro tests. Venous blood samples (5\u00a0ml) are collected at day 0 before treatment from women with a parasite density of at least 4,000/\u03bcL. The HRP2 ELISA [P. falciparum in the presence of lumefantrine, monodesethylamodiaquine (active metabolite of amodiaquine), mefloquine, piperaquine and dihydroartemisinin (active metabolite of artemisinin derivatives).This component is carried out at the sites in Burkina Faso only. The sensitivity of the parasites to the drugs used is determined by carrying out P2 ELISA , 18 is uThe sample size for this study was determined by simulation with the following assumptions and requirements: (1) study conclusions are determined by two-sided 95% confidence intervals for difference in response rates (% of therapeutic success), with decision rule as described below, (2) all 4 treatments have identical true response rates of 95%, (3) 95% power for each of the 6 pair-wise comparisons and 80% power for the combined hypothesis that all treatments are therapeutically equivalent is required. With these assumptions, approximately 700 patients/treatment arm are needed. If the true response rate for one of the treatments is lower than 95%, then power is reduced to 80% (for a true response rate of 94%) or 50% (for a true response rate of 93%). Allowing for a 20% loss to follow-up, a total of 870 patients are recruited to each treatment; this is equivalent to 290 patients in each treatment group in each centre  \u2013 and hence a total study sample size of 3480 patients. Inclusion of HIV-infected women is not expected to have a major influence on the sample size calculation. The percentage of therapeutic success may be slightly but not dramatically lower in this subgroup of patients.For safety, when combined, the trial is able to detect with 90% power major adverse events occurring at the frequency of at least 2-3%.Baseline comparability: Patients in each treatment group in each site are described separately with respect to baseline characteristics. The clinical importance of any imbalance will be noted, though statistical tests of significance are not undertaken.Primary analyses: The primary analysis of the study is the assessment of therapeutic equivalence of the 4 treatments  with respect to therapeutic success at day 63 and their safety throughout the follow-up, i.e. up to one year after delivery.A detailed analysis plan is drawn up prior to the analysis.Therapeutic equivalence is assessed using the pair-wise difference in response rates (percentage of women with therapeutic success). Assessment of the difference in true response rates is performed by calculating the two-sided 95% confidence interval for the difference in response rates from the observed data, using the following decision rule:if the two-sided 95% confidence interval for the difference in response rates lies entirely between \u22125% and +5%, then therapeutic equivalence of the two treatments is concluded;if the 95% confidence interval for the difference in response rates includes \u22125% or +5%, then therapeutic equivalence cannot be established;if the 95% confidence interval for the difference in response rates lies entirely below \u22125% or entirely above 5%, then one treatment is clinically inferior to the other.The primary analysis uses the combined data from all 4 centres together, with adjustment for any centre effects, using an additive model for the response rates . This allows the assessment of all 6 possible pair-wise treatment comparisons using all available data. Equivalence will be established using two-sided confidence intervals. Though 6 treatment comparisons will be performed, no adjustment for multiplicity is needed as the focus of the study is on the individual pair-wise treatment comparisons. In addition, combined hypotheses of interest  require each of the individual hypotheses to be accepted. Consequently, there is no inflation of the type I error rate due to multiple testing. However, the power for combined hypotheses is lower than for the individual pair-wise comparisons. Thus, the power calculation of this study required a high (95%) power for individual pair-wise treatment comparisons, resulting in an acceptable (80%) power for the combined hypothesis. For the efficacy analysis, both an intention-to-treat and a per-protocol approach are adopted, with the per-protocol analysis being the primary approach, as recommended for equivalence studies.For safety analysis, all non-serious and serious adverse events (SAE) are grouped according to a pre-specified side-effect coding system and tabulated. The number (and percentage) of patients experiencing any adverse event, any SAE, and any drug-related SAE are compared between treatment groups using Fisher's exact test. Safety is analyzed using the all-patients-treated approach.P. falciparum monoinfection of any density, regardless of symptoms, and a Hb concentration of at least 7\u00a0g/dL. Pregnant women with a negative blood slide are not included in the study and go through the routine antenatal clinic procedures according to national policy and receive a dose of sulfadoxine-pyrimethamine (SP) as intermittent preventive treatment (IPT). Exclusion criteria include history of allergic reactions to the study drugs, of known pregnancy complications or bad obstetric history such as repeated stillbirths or eclampsia, of presence or history of major illnesses likely to influence pregnancy outcome, e.g. diabetes mellitus, severe renal or heart disease, or active tuberculosis, current cotrimoxazole prophylaxis or ARV treatment. Table\u00a0All pregnant women in the second and third trimester (<37\u00a0weeks) and attending the antenatal clinic of the study health facilities are systematically screened for malaria infection with a rapid diagnostic test; if positive they are further assessed for eligibility. They are included if they are at least 15\u00a0years old, with a pregnancy of at least 16\u00a0weeks, a For the informed consent, all interviews are conducted in the native language of the patients by a qualified person identified by the Investigator. Written information and consent forms in the local language are provided to the women or Legally Authorized Representatives (LAR) for their review. After the interview, the patients and, in case they are of minor age/not emancipated, the parents or guardians are asked to confirm their willingness to participate in the study by signing (or thumb-printing if illiterate) the consent form.Each eligible pregnant woman who agrees to give informed consent is assigned a unique study number and enrolled. Besides malaria infection and parasite density, Hb, total white blood cell count, differential count, total bilirubin, ALAT and creatinine are measured. In addition, a blood sample is collected on filter paper for later genotyping..Randomisation is carried out according to a pre-established list comprising blocks of varying size and stratified according to the number of recruitment points in each site. Allocation of treatment according to the randomization list is in sealed envelopes labelled with the patient\u2019s unique code, guaranteeing concealment until recruitmentThe study treatment is administered during the first 3 study days (days 0\u20132) by the study doctor or nurse and the patient kept for one hour in the clinic. If vomiting occurs within 30\u00a0minutes, a full treatment is re-administered, half a dose if after 30\u00a0minutes. In case of persisting vomiting, an alternative treatment, e.g. quinine, is provided.Scheduled visits are at day 3, 7 and then every week until day 63 post-treatment. However, women are encouraged to attend the antenatal clinic if they felt ill between scheduled visits. At the end of the active follow-up period, women are asked to attend the antenatal clinic monthly and at any time they feel unhealthy until delivery. At each visit, scheduled and unscheduled, the medical history since the last visit (including any treatment taken), current signs and symptoms (if any) are collected. A blood sample for thick and thin blood film and later genotyping to determine the rate of re-infection is collected and the body temperature checked. Haematology  is measured at day 7, 14, 28 and 63; biochemistry  at day 7 and 14.If pregnant women recruited during the third trimester deliver before the end of the 63-day active follow, scheduled visits continue after delivery until the day 63 is completed. The outcome of pregnancy, including any congenital abnormality, the birth weight and maternal Hb are collected as soon as possible after delivery. In addition, a placenta impression smear and a placenta biopsy for later histopathological analysis are collected. Both the mother and the new-born are reassessed twice after delivery for any adverse event: between 4 and 6\u00a0weeks and then after one year  and their active follow up stopped. Nevertheless, they are still followed up (safety data) until one year post-delivery.Safety is closely monitored during the course of the study in compliance with ICH/GCP guidelines. At each visit, the investigator ascertains the occurrence of any adverse events since the previous visit; including those involving laboratory values which are out of normal range and are of clinical importance to be considered as AEs, and the proper AE reporting procedure is followed. The severity of a clinical adverse event is scored according to the following scale: mild, moderate, severe and life-threatening.Serious Adverse Events (SAEs) are defined as any untoward medical occurrence that, at any dose of the medication given, fulfilled the following criteria; death, life-threatening, requiring hospitalization or prolongation of hospitalization, resulting in persistent or significant disability or incapacity, congenital anomaly/ birth defect, or other situations such as important medical events that may not be immediately life-threatening or result in death or hospitalization but may jeopardize the subject or may require medical or surgical intervention to prevent one of the other outcomes listed in the above definition. The reporting investigator assesses the relationship between investigational product and the occurrence of each AE/SAE. The relationship of an adverse event to study drug is assessed according to the following definitions: \u2018Definitely unrelated\u2019, \u2018Unlikely\u2019, \u2018Possible\u2019, \u2018Probable\u2019 and \u2018Definitely related\u2019. The outcome of each AE is assessed according to the classification as follows \u2013 \u2018Completely recovered\u2019, \u2018Not yet completely recovered\u2019, \u2018Deterioration\u2019, \u2018Permanent damage\u2019, \u2018Death\u2019, \u2018Ongoing\u2019 and \u2018Unknown\u2019.Each SAE has to be reported to the sponsor and to the concerned ethical bodies in the study countries within 24 working hours since the time the study staff becomes aware of it, and any reporting delay has to be explained. All the SAE forms are further sent by the sponsor to the concerned ethical bodies in Belgium and to the independent DSMB. Each SAE is followed up until resolution.Each site is visited at least 3 times during the conduct of the trial plus a study initiation visit at the start of clinical activities and a close-out visit after the last patient has completed the follow up. The monitor will perform the tasks as described in International Conference on Harmonisation (ICH)-Good Clinical Practice (GCP) E6, section 5.18 and will carry out at least 10% source data verification (SDV). For all sites, the SDV percentage will be increased by the monitor if the quality of data entry is found not to be satisfactory.Each patient has her own source document file, according to a common source document template provided by the Sponsor, with all the original documents, e.g. laboratory results. This data is captured into an electronic case report form (e-CRF) developed in the GCP-compliant software MACRO  for clinical trials. The e-CRF has in-built consistency checks; data can be entered either online or offline and then uploaded to be sent to the central server. The final database is obtained after the resolution of all queries and then locked for later statistical analysis done according to a pre-established statistical data analysis plan.The Consortium Secretariat (CS) acts as a steering committee. It comprises one investigator from each site and will assess the progress of the trial. The members of the CS will address policy and operational issues related to the protocol. The CS has responsibility for protecting the scientific conduct and integrity of the trial. Its functions include review of the protocol before ethical approval; and formulation of recommendation for any change in the design and operations of the trial during the course of the trial, when needed.The independent Data Safety and Monitoring Board (DSMB) is composed by four independent scientists, i.e. a paediatrician, a gynaecologist, a statistician and a malariologist. They meet every three months during recruitment or can be called together if the necessity arises.Several amendments Table\u00a0 have beeAnother amendment was done on the basis of the modification of blood piperaquine concentrations by food intake which could result in a QTc prolongation, a risk factor for serious cardiac arrhythmia. Therefore, the manufacturer advised to administer DHAPQ tablets with water only and at least 3\u00a0hours apart from meal, mainly for the second and third administration, and advice the woman not to eat for the next 3\u00a0hours.Pregnant women are one of the high risk groups affected by the malaria burden and few antimalarials are available to treat them. This study assesses the efficacy and safety of four ACTs for the treatment of uncomplicated malaria in pregnant women in Africa. This is the largest clinical trial of its kind and will provide the evidence base for the formulation of treatment guidelines for malaria in pregnancy.Evidence of the interaction between malaria and HIV-1 has already been reported. HIV-1 infected pregnant women have a higher prevalence of peripheral parasitaemia and placental malaria , 20 and Considering that most pregnant women recruited in the study will have an infection with a relatively low parasite density and that they will be treated with an ACT, it is expected that the number of treatment failures in this specific sub-group would be extremely low. They will have to be treated in any case as the consequences of the malaria infection on the woman\u2019s health and that of her offspring are well known.In the second and third trimester of pregnancy, WHO recommends the use of ACT known to be effective in the country/region. Despite this recommendation, it should be recognised that the available information on the treatments to be used in this trial is limited. ACT should not be used in the first trimester of pregnancy, the time of greatest concern for potential teratogenicity, and particular care will be taken in excluding women of this gestational age. DHA-PQ is the least used of the 4 ACTs studied in this trial and it is not among the WHO recommended ACTs during pregnancy because of insufficient information . HoweverData collection completed."}
+{"text": "Plasmodium falciparum malaria among six to 59 months old Malawian children.The resistance of malaria parasites to sulphadoxine-pyrimethamine (SP) in 2007 led to the Malawi Ministry of Health changing to artemether-lumefantrine (AL) as first-line for uncomplicated malaria treatment. This study determined the efficacy and safety of AL for the treatment of uncomplicated This was a prospective study of children six to 59 months old treated with AL after presenting with uncomplicated malaria in the six health facilities in Malawi. The children were followed up on days 1, 2, 3, 7, 14, 21 and 28 days post-treatment and assessed for clinical and parasitological responses. The Kaplan Meier survival estimate was used to measure the efficacy of AL by calculating the cumulative risk of failure at day 28.A total of 322 children were recruited into the study across the six sites. The overall intention-to-treat (ITT) polymerase chain reaction (PCR)-corrected cure rate was 93.4%. Per protocol overall PCR-corrected cure rates for the study sites were; Karonga 98.0%, Kawale 97.4%, Machinga 90.2%, Mangochi 95.4% and Rumphi 91.3%. Nkhotakota study site had the lowest cure rate of 78.0%.There is evidence of good efficacy of AL in Malawi notwithstanding geographical contrasts and this supports the continued use of AL as the first-line treatment for uncomplicated malaria. However there may be need to further investigate the comparatively low efficacy rate found in Nkhotakota district in order to identify possible determinants of treatment failure. Malaria is still one of the leading causes of mortality among children under five years of age in sub-Saharan Africa despite efforts to control it ,2. GlobaPlasmodium [Plasmodium falciparum malaria among Malawian children aged between six to 59 months.The World Health Organization (WHO) recommended the use of artemisinin-based combination therapy (ACT) for treatment of uncomplicated malaria . The curasmodium ,10, inclasmodium . Evidencasmodium ,13. Howeasmodium -16. Delaasmodium . Passiveasmodium . Sentineasmodium . The stuP. falciparum malaria in Malawian children. Malawi is in south-eastern Africa. The climate is tropical-continental with some maritime influences and a rainy season varying between November and April. Malaria is endemic in Malawi, with a prevalence of 28% among children under five years of age [P. falciparum malaria at the health centre in the selected study sites were recruited in the study.This was a prospective study conducted between the months of March to June 2010 aimed at monitoring the efficacy and safety of AL for the treatment of uncomplicated s of age . The stuP. falciparum detected by microscopy with parasite density of 1,000 - 200,000/\u03bcl asexual forms, axillary temperature \u226537.5 \u00b0C or history of fever for the previous 24 hours, guardians willing to return for weekly routine and unscheduled visits, planning to remain in the study area during the study and provided informed consent. The exclusion criteria for the study were: presence of general danger signs or severe P. falciparum malaria in the children as per WHO classification, mixed or mono-infection with other Plasmodium species, severe malnutrition , other diseases like measles, acute lower respiratory tract infection, severe diarrhoea with dehydration, cardiac, renal and hepatic diseases. Further exclusion included regular medication, which might have interfered with anti-malarial drugs pharmacokinetics and history of contraindications or hypersensitivity reactions to AL. Assuming that treatment failure rate of AL in Malawi will be about 5%. At a confidence level of 95% and a precision around the estimate of 5%, a minimum of 73 patients were to be included. With a design effect of 2.0, a number of 146 participants were reached. Allowing for a 20% loss to follow-up and withdrawals, a minimum 176 participants was to be included in the study.Study participants were screened as they presented to the health facility. The inclusion criteria for the study were: children aged from six to 59 months, presence of mono-infection with Children enrolled after meeting the inclusion criteria were given a personal identification number after guardian signed an informed consent. All children were treated with AL. Regular tablets of AL  were administered orally according to weight bands using a standard dose of 2mg/kg artemether with 10mg/kg lumefantrine (co-formulated tablets) twice daily on Days 0, 1 and 2 . All sixThe study was conducted for a period of 28 days. Day 0 was designed as the day patients were enrolled and received the first dose of AL. Follow up was from Day 1, 2, 3, 7, 14, 21 and 28. During the follow up visits, children had clinical and laboratory assessments done. Haemoglobin was determined using a Hemocue\u00ae. Thick and thin blood smears were prepared for determination of malaria parasitaemia. Smears were double read and if any discrepancies between the two readers, the slide was read by a third person independently. PCR analysis was conducted at the Malawi Liverpool Wellcome Trust laboratories, which provided a PCR-corrected outcome.any untoward medical occurrence in a patient administered a medicinal product and which does not necessarily have to have a causal relationship with this treatment\u201d [Guardians of participants were advised to return on any day during the follow-up period if symptoms recurred or became severe, or any occurrence of adverse events. International Conference on Harmonization (ICH) Tripartite Guideline for 2003 defines an adverse event as \u201ceatment\u201d .The primary outcomes for the study were overall treatment success and overall treatment failure. Treatment outcomes were classified according to WHO guidelines as; early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), loss to follow-up, withdrawal from the study and adequate clinical and parasitological response (ACPR) . The secData was collected on paper using a Case Report Form (CRF). These were checked on a daily basis before data entry. Then the data was independently double entered into an Excel database specifically designed by the WHO for anti-malarial efficacy studies. Data was then exported and analysed in STATA. Both intention to treat analysis and per protocol analysis methods were used when analysing the data. Intention to treat analysis estimated the effect of the assigned treatment (AL). Per protocol analysis estimated the effect of adhering to the assigned treatment protocol. A description of baseline characteristics of all participants involved in the study was presented. The Kaplan Meier method was used to measure the efficacy of AL by calculating the cumulative risk of failure at day 28. There was comparison among the study sites Rumphi, Mangochi, Nkhotakota, Machinga, Kawale, and Karonga on: the proportion of lost to follow up patients and those patients withdrawn from the study, proportion of ETF, LCF, LPF, ACPR on day 28, success and failure cumulative incidence rates on day 28 with 95% confidence interval and p value of less than 0.05 was considered significant. The differences in efficacy between sites were assessed using Cox proportional hazards model.This study was approved by the National Health Sciences Research Committee  in Malawi.An overall number of 322 participants were recruited into the study. Intention to treat analysis included loss to follow up and withdrawals in the denominator while per protocol analysis excluded participants withdrawn 16.2% (52/322) and lost to follow up 2.5% (8/322) from the study. Table\u00a0All 322 children presented with fever during recruitment into the study with the mean temperature of 38.5\u00b0C. All 322 children reported with parasitaemia at recruitment on Day 0. Day 1 reported 68.1% (211/310) children who had not cleared parasites, while Day 2 had 9.1% (28/305) children with parasitaemia. Day 3 had 0.9% (3/305) children reporting with parasitaemia. Figure\u00a0All participants who presented with malaria during the follow-up period had their filter paper sample undergo PCR. The PCR-corrected cure rates were measured. Using intention to treat analysis, ETF was reported among 1.6% 5/320) study participants. LPF was reported among 0.9% (3/320) study participants while LCF was reported among 4.1% (13/320) study participants during the 28 days follow up period. This makes a total of 21 children with treatment failure and total time at risk of 7,926 days. The incident rate (hazard function) was estimated as 2.65 per 1,000 persons per day with the assumption that the incident rate remains constant. Intention-to-treat Kaplan-Meier survival estimate curve is displayed in Figure\u00a00 study pThe treatment outcomes by study site were as follows; Nkhotakota study site had the highest LCF of 6, while Machinga study site had 3 LCF and Karonga had 0 LCF. Karonga, Machinga and Nkhotakota study sites showed one child with ETF in each study site while Kawale and Mangochi had no children observed to ETF. There were three children with LPF, one in Mangochi and two in Nkhotakota which were confirmed by PCR Table\u00a0. Table\u00a03Gametocytes were found in peripheral blood of seven children at recruitment into the study. One child each was found with gametocytes on Day 7 and 21. No gametocyte carriage was reported on any other follow-up day.A number of adverse events occurred during the 28 day follow-up period however none of the events were related to the effect of AL Figure\u00a0. The mosThe study showed that the overall efficacy of AL after PCR correction was high in Malawi at 93.4%. This corresponds with other studies done in other countries of Africa on the efficacy of AL ,13,21,22There was a dramatic reduction in parasitaemia in the study participants after only one day\u2019s dose of AL. This is consistent with other studies that have shown that the artemisinin derivative act more rapidly than other types of anti-malarial, both in killing parasites and in inhibiting their major metabolic processes, such as glycolysis, nucleic acid and protein synthesis . They alThe recommendations for the WHO malaria treatment guidelines in the year 2010 emphasises on switching to another anti-malarial drug when the first-line has more than a 10% overall failure rate . Even thP. falciparum malaria in Malawi since the study has demonstrated that AL is safe and has not exceeded the more than 10% cut-off failure rate.At policy level, AL should remain to be used as first-line treatment for the treatment of uncomplicated This study aimed at determining whether AL was efficacious and safe, and investigated the difference in efficacy between the study sites. The findings of the study showed that there was high efficacy of AL in Malawi. Therefore, it is important to monitor efficacy in order to detect and prevent resistance of AL. Surveillance of the established sentinel sites should continue. However there may be need to further investigate the comparatively low efficacy rate found in Nkhotakota district in order to identify possible determinants of treatment failure. Therefore, AL should remain the first-line treatment for uncomplicated malaria among children in Malawi."}
+{"text": "Plasmodium falciparum has emerged in the Greater Mekong sub-region and poses a major global public health threat. Slow parasite clearance is a key clinical manifestation of reduced susceptibility to artemisinin. This study was designed to establish the baseline values for clearance in patients from Sub-Saharan African countries with uncomplicated malaria treated with artemisinin-based combination therapies (ACTs).Artemisinin-resistant a priori statistical analytical plan. Factors affecting early parasitological response were investigated using logistic regression with study sites fitted as a random effect. The risk of bias in included studies was evaluated based on study design, methodology and missing data.A literature review in PubMed was conducted in March 2013 to identify all prospective clinical trials , including ACTs conducted in Sub-Saharan Africa, between 1960 and 2012. Individual patient data from these studies were shared with the WorldWide Antimalarial Resistance Network (WWARN) and pooled using an P <0.001); fever (>37.5\u00a0\u00b0C) (AOR\u2009=\u20091.50 (95\u00a0% CI: 1.06\u20132.13), P\u2009=\u20090.022); severe anaemia (AOR\u2009=\u20092.04 (95\u00a0% CI: 1.21\u20133.44), P\u2009=\u20090.008); areas of low/moderate transmission setting (AOR\u2009=\u20092.71 (95\u00a0% CI: 1.38\u20135.36), P\u2009=\u20090.004); and treatment with the loose formulation of artesunate-amodiaquine (AOR\u2009=\u20092.27 (95\u00a0% CI: 1.14\u20134.51), P\u2009=\u20090.020, compared to dihydroartemisinin-piperaquine).In total, 29,493 patients from 84 clinical trials were included in the analysis, treated with artemether-lumefantrine , artesunate-amodiaquine  and dihydroartemisinin-piperaquine . The overall parasite clearance rate was rapid. The parasite positivity rate (PPR) decreased from 59.7\u00a0% (95\u00a0% CI: 54.5\u201364.9) on day 1 to 6.7\u00a0% (95\u00a0% CI: 4.8\u20138.7) on day 2 and 0.9\u00a0% (95\u00a0% CI: 0.5\u20131.2) on day 3. The 95th percentile of observed day 3 PPR was 5.3\u00a0%. Independent risk factors predictive of day 3 positivity were: high baseline parasitaemia (adjusted odds ratio (AOR)\u2009=\u20091.16 (95\u00a0% CI: 1.08\u20131.25); per 2-fold increase in parasite density, The three ACTs assessed in this analysis continue to achieve rapid early parasitological clearance across the sites assessed in Sub-Saharan Africa. A threshold of 5\u00a0%\u00a0day 3 parasite positivity from a minimum sample size of 50 patients provides a more sensitive benchmark in Sub-Saharan Africa compared to the current recommended threshold of 10\u00a0% to trigger further investigation of artemisinin susceptibility.The online version of this article (doi:10.1186/s12916-015-0445-x) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria in the Greater Mekong region [The increasing availability of artemisinin-based combination therapies (ACTs) and long-lasting insecticidal nets (LLINs) over the last decade has contributed to a substantial reduction in malaria morbidity and mortality in Sub-Saharan Africa (SSA) , 2. Howeg region \u20139 threatDelayed parasite clearance is the hallmark of artemisinin resistance , 10, 11.It is known that the speed of parasite clearance is influenced by a number of host, parasite and drug factors , 11, 21,P. falciparum malaria enrolled in ACT clinical efficacy trials conducted between 1999 and 2012. The aim was to provide a baseline of early parasitological response profiles so that sites at high risk (hot spots) for artemisinin resistance can be identified going forward, to inform malaria control and containment efforts.Therefore, to assess the dynamics of early parasitological response after artemisinin combination therapy observed in SSA, parasite clearance data were compiled from patients with uncomplicated World malaria report 2014 [A literature review was conducted in PubMed in March 2013 and updated in 2014 to identify all published clinical trials of antimalarials since 1960. All antimalarial clinical trials published since 1960 were identified by the application of the key terms  AND ) through the PubMed library. All references containing any mention of antimalarial drugs were tabulated and manually checked to confirm prospective clinical trials. Studies on prevention or prophylaxis, reviews, animal studies or studies of patients with severe malaria or in pregnant women were excluded. When pdfs were available further details of the publications were reviewed, and basic details on the study methodology, treatment arms assessed and the study locations were documented. These are provided in the WorldWide Antimalarial Resistance Network (WWARN) publication library . SpecifiP. falciparum ; ii) clinical trials conducted in SSA with one of the following three ACTs: artemether-lumefantrine (AL) (six-dose), dihydroartemisinin-piperaquine (DP) and one of the three formulations of artesunate-amodiaquine (AS-AQ): fixed dose combination (ASAQ-FDC), non-fixed dose combination in a loose formulation (ASAQ-loose NFDC) or non-fixed dose combination in a co-blister formulation (ASAQ-coblistered NFDC); and iii) parasitaemia was sampled at least on days 2 (48\u00a0hours) and 3 (72\u00a0hours) following treatment. Individual study protocols were available for all trials included, either from the publication or as a metafile submitted with the raw data. All data were uploaded to the WWARN repository and standardized using a methodology described in the clinical module data management and statistical analysis plan [All research groups in the systematic review were contacted to share the entire dataset of their study with WWARN. Those who had contributed studies previously to the WWARN data repository were also invited to participate and asked whether they were aware of any unpublished or ongoing clinical trials involving ACTs, and these additional unpublished studies were also requested. Studies were included in the meta-analysis provided that they were: i) prospective clinical efficacy studies of uncomplicated sis plan .Anaemia was defined according to WHO guidelines  , based on the fact that all studies contributed to WWARN must have already obtained all necessary ethical approvals and informed consent.All statistical analyses were carried out based on an a priori statistical plan . The prim) was determined based on the fraction of missing information (\u03b3) assuming 5\u00a0% loss in efficiency (\u03b7) using m\u2009\u2265\u2009\u03b3*(\u03b7/1\u2013\u03b7) [Univariable and multivariable analyses of risk factors associated with parasite positivity status on days 1, 2 and 3 were conducted using generalized linear mixed model (logit link), in a one-stage analysis by combining all of the individual patient data. In order to account for within study clustering, study sites were fitted as random effects; the statistical significance of which was assessed using a likelihood ratio test. Statistical heterogeneity was quantified as the variance of the random effects using maximum likelihood method and the proportion of total variance contributed by the site-level variance component (\u03c1) was reported. Missing covariates were dealt with using multiple imputation methods. The number of imputations (*(\u03b7/1\u2013\u03b7) . Known cThe robustness of the coefficients in the final multivariable model was examined using bootstrap sampling. Sensitivity analysis was performed by excluding one study site at a time and the coefficient of variation around the parameter estimates was calculated. The final model was used to simulate outcome for each patient and the observed PPRs were plotted against the simulated PPRs to assess model adequacy.U test or Kruskal\u2013Wallis test. All statistical analyses were carried out using R  using lme4 package.Continuous variables were compared between groups using generalized linear regression with study sites fitted as random effects. Data that were not normally distributed were compared with Mann\u2013Whitney In accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, the risk of bias within studies was assessed based on: 1) study design ; 2) microscopy methodologies for parasite quantification; and 3) the proportion of patients with (a) missing outcomes (missing outcome on days 2 and 3) and (b) missing baseline covariates .meta package. Publication bias was assessed through the use of a funnel plot of the log-transformed odds ratio, the asymmetry of which was tested using Egger\u2019s method.To assess whether the non-availability of some individual participant data could have biased the results, we extracted data on PPRs from studies not providing individual patient data and performed a two-stage meta-analysis of proportions using logit transformation; a continuity correction of 0.5 was applied to studies with zero cell count using The systematic literature review identified 140 published clinical studies of ACT efficacy that were potentially relevant to this analysis. Researchers agreed to share individual patient data from 71 trials (50.7\u00a0%) including 25,731 patients (59.9\u00a0% of the targeted population). Additional data were available for 3,762 patients from 13 unpublished trials. In total, individual records were available from 29,493 patients enrolled in 27 different countries between 1999 and 2012 . The median baseline parasitaemia was 20,200 parasites/\u03bcl  with slight differences between treatment groups (Table\u00a0The baseline characteristics of the included patients are given in Table\u00a0P <0.001) and day 2 , but there was no age-related difference on day 3. Patients with an initial parasite density >100,000 parasites/\u03bcl had a PPR of 82.7\u00a0%  on day 1, 14.3\u00a0%  on day 2 and 1.3\u00a0%  on day 3. The corresponding proportions for patients with parasitaemia less than 100,000 parasites/\u03bcl were 57.0\u00a0% , 5.9\u00a0%  and 0.8\u00a0% , respectively for days 1, 2 and 3 . There were no regional differences or temporal trend in the PPRs on any days during the time period studied, that is, 1999\u20132012. A detailed summary of the PPRs for each of the treatment regimens stratified by country and calendar year is presented in Additional file The presence and density of parasites on day 1 could only be assessed in 55\u00a0% 16,196/29,493) of patients (52 studies). The overall parasite clearance rate for all studies was rapid. The PPR decreased from 59.7\u00a0% (95\u00a0% CI: 54.5\u201364.9) on day 1  to 6.7\u00a0% (95\u00a0% CI: 4.8\u20138.7) on day 2  and 0.9\u00a0% (95\u00a0% CI: 0.5\u20131.2) on day 3 . The PPRs on days 1, 2 and 3 were similar for AL, DP and ASAQ-FDC, but higher for the non-fixed formulations of AS-AQ on days 2 and 3  compared to those treated with DP or those treated with ASAQ-FDC (AOR\u2009=\u20091.33 (95\u00a0% CI: 1.08\u20131.63), P\u2009=\u20090.005). Similarly, patients treated with ASAQ-loose NFDC had an increased risk of remaining parasitaemic on day 2 compared to DP (AOR\u2009=\u20091.46 (95\u00a0% CI: 1.05\u20132.01), P\u2009=\u20090.022) and compared to ASAQ-FDC (AOR\u2009=\u20091.61 (95\u00a0% CI: 1.14\u20132.29), P\u2009=\u20090.007). In the same multivariable model, patients from low/moderate transmission sites were also at greater risk of remaining parasitaemic on day 2 compared to those from high transmission sites (AOR\u2009=\u20091.88 (95\u00a0% CI: 1.09\u20133.24), P\u2009=\u20090.024) , fever (AOR\u2009=\u20091.50 (95\u00a0% CI: 1.06\u20132.13), P\u2009=\u20090.022), severe anaemia (Hb\u2009<\u20097\u00a0g/dl) (AOR\u2009=\u20092.04 (95\u00a0% CI: 1.21\u20133.44), P\u2009=\u20090.008) and being from areas of low/moderate transmission ; see Table\u00a0P\u2009=\u20090.020) increased risk of being parasitaemic on day 3 compared to patients treated with DP and 3.36-fold ((95\u00a0% CI: 1.61\u20136.98), P\u2009=\u20090.001) higher risk compared to patients treated with ASAQ-FDC. Similarly, patients treated with ASAQ-coblistered NFDC were at 4.18-fold ((95\u00a0% CI: 1.28\u201313.68), P\u2009=\u20090.017) greater risk compared to those treated with ASAQ-FDC , for every 2-fold increase in parasite density, P\u2009=\u20090.003)  was available in 72\u00a0%  of the patients. Adjusted for the baseline confounders, the mg/kg dose of artemisinin component was not associated with the risk of parasite positivity on any day for patients treated with DP or AS-AQ (either for the fixed or the loose combinations). However, in patients treated with AL, an increased mg/kg dose of artemether was associated with a lower risk of patent parasitaemia only on day 1. Every unit increase in daily mg/kg artemether dose reduced the risk of parasite positivity by 5\u00a0% ((95\u00a0% CI: 1\u20137\u00a0%), The overall day 3 PPR was 0.58\u00a0% (95\u00a0% CI: 0.34\u20130.82) for AL, 0.54\u00a0% (95\u00a0% CI: 0.14\u20130.94) for ASAQ-FDC and 0.77\u00a0% (95\u00a0% CI: 0.11\u20131.42) for DP. In studies with a sample size greater than 50 patients, the observed PPR was unlikely to exceed 5\u00a0% positivity on day 3 Fig.\u00a0, Fig.\u00a04.Attrition biases of the included studies are presented in Additional file This large pooled analysis of nearly 30,000 patients from trials conducted before 2012 highlights that parasite clearance after treatment with an ACT is still extremely rapid in Sub-Saharan Africa. More than 90\u00a0% of the patients were aparasitaemic by day 2 and 99\u00a0% by day 3, consistent with previous reports demonstrating rapid parasite clearance after treatment with ACTs in high transmission settings .In areas of intense transmission, immunity develops at a relatively young age , 39 and After adjusting for these parasite and host factors, the risks of persistent parasitaemia on days 1 and 2 were higher in patients treated with AL compared to those treated with DP and ASAQ-FDC, but this difference was no longer apparent by day 3. Artemether is a lipophilic compound and is more slowly absorbed than artesunate or dihydroartemisinin, and this difference may explain the slower action of AL , 45. Morin vitro testing [The study period encompasses 1999 to 2012, covering the period during the introduction of the large scale deployment of ACTs across Africa. Overall, there were no differences in the early parasitological response post-ACT treatment in different sub-regions of SSA and there was no evidence of decreased susceptibility to artemisinin in Africa over this time period. Nevertheless, there were 22 sites where PPR on day 3 exceeded 3\u00a0% (the threshold below which artemisinin resistance in unlikely), with two sites exceeding day 3 PPR of 10\u00a0% . In Miandrivazo (Madagascar), the reported PPR was 10.3\u00a0% in 2006  but less testing  and mole testing  to rule Our analysis has a number of limitations. First, the literature search was limited to prospective clinical trials indexed in PubMed and some relevant studies may have been overlooked. However, we actively looked for relevant trials (unpublished) and the research groups contacted represent the majority of the malaria community, which is relatively small and highly interactive. It is highly unlikely that any studies were missed. The assessment of publication bias (PB) showed that effect sizes were symmetrical suggesting low risk of bias in studies included. Of the 140 trials identified, individual patient data were available for inclusion for 71 of the published studies (50.7\u00a0%). To address this potential bias, included studies were compared with the published studies that were not available. There were no apparent differences in patient population and/or outcomes between the studies included and those where individual patient data were not available. Reassuringly, the results from two-stage meta-analyses, which combined studies with and without individual patient data, were also similar to the results obtained from studies where only individual patient data were available, suggesting that systematic attrition bias was unlikely. A second issue is that, although the days of follow-up were recorded in the studies, the actual time of blood collection was not. Daily samples were taken over a range of times and the interval between days is likely to have varied significantly from the desired 24-, 48- or 72-hour timelines. Third, the data used rely on quantitative microscopy and quality control on microscopy procedures were reported in only 60\u00a0% of the studies. Accurate recording of the time of sampling, harmonizing microscopy procedures and appropriate quality control procedures could greatly improve the precision of the parasite clearance time . To faciThis large dataset provided a unique opportunity to identify a threshold for day 3 parasite positivity based upon African studies, below which artemisinin resistance is highly unlikely. The upper limit of the 95\u00a0% CI for day 3 PPR, indicative of the worst-case scenario, defines maximum PPR which could be observed reliably in a clinical trial. This threshold was vulnerable to the initial parasitaemia and study sample size. For example, in studies with 50 or less patients, the confidence interval around any threshold value was wide, hence its predictive utility under those circumstances is limited. Our results demonstrate that the 95th percentile of the observed day 3 PPR in Africa was 5.3\u00a0%, substantially lower than the currently recommended threshold of 10\u00a0% for suspected partial artemisinin resistance. These findings strongly suggest that a \u2018one size fits all\u2019 threshold of 10\u00a0% should be used with caution. A simple sensitive parameter indicative of potential artemisinin resistance would be an extremely useful surveillance tool. Our analysis suggests that although the widely proposed 10\u00a0% threshold would be specific, it lacks sensitivity in detecting an early stage changes of delayed parasite clearance. Moreover, a previous WWARN meta-analysis of published literature showed that the PPR on day 3 over the same period (1999\u20132012) was much lower in Africa (1\u00a0%) compared to Asia (3.8\u00a0%) . A thresIn conclusion, this pooled analysis provides critical baseline information regarding early parasitological response post-treatment with ACTs in SSA. The assessment of the host, parasite and drug determinants which influence the early parasitological response can provide evidence-based guidance for monitoring the early signs of artemisinin resistance and effective case management that will be critical in optimizing malaria control and containment efforts."}
+{"text": "This randomized, open-label study was conducted to establish the non-inferiority of a combination of azithromycin (AZ) and chloroquine (CQ) to artemether-lumefantrine (AL) for treatment of uncomplicated malaria in children from six sites in sub-Saharan Africa.Plasmodium falciparum malaria between six and 59\u00a0months of age were randomized 1:1 to either AZCQ (30\u00a0mg/kg AZ\u2009+\u200910\u00a0mg/kg CQ base) or AL per prescribing information for three days . Each site could enrol in the study population once the treatment of uncomplicated malaria in five children five to 12\u00a0years of age was deemed to be effective and well tolerated. The primary efficacy evaluation was the proportion of subjects in both the modified intent-to-treat (MITT) and per-protocol (PP) populations with an adequate clinical and parasitological response (PCR corrected) at Day 28. Non-inferiority was concluded if the lower bound of the 95% confidence interval comparing the two groups was 10 percentage points or greater.Children with uncomplicated versus 98% (AL) for MITT, a difference of -9.10  and 93% (AZCQ) versus 99% (AL) for PP, a difference of -6.08 . Early and late treatment failures were more common in subjects receiving AZCQ. Adverse events were more common in subjects treated with AZCQ. Drug concentrations obtained at specified time points following AZCQ administration had a large coefficient of variation partially due to sparse sampling with sample collection time window.A total of 255 children were enrolled in the efficacy analysis . The PCR corrected clearance rates were 89% (AZCQ) In this study, non-inferiority of AZCQ to AL was not demonstrated.NCT00677833.ClinicalTrials.gov The online version of this article (doi:10.1186/s12936-015-0620-8) contains supplementary material, which is available to authorized users. Plasmodium, continues to be a global health problem [Plasmodium falciparum accounted for 91% of total cases. The vast majority of the estimated 627,000 malaria deaths occurred in 2012, 90% in Africa, and children under five years accounted for 77% of these deaths [P. falciparum [.Malaria, a disease caused by five different species of  problem . Throughin vitro and in vivo against CQ-resistant strains of P. falciparum [P. falciparum malaria in adults were recently demonstrated in two multicentre phase 3 clinical studies in Africa [P. falciparum malaria in African children.The combination of azithromycin (AZ) and chloroquine (CQ) has been shown to have synergistic activity lciparum -5. AZ ann Africa ,7 and inn Africa -10. In an Africa . The objThis was an open-label, randomized, phase 2/3 study at six sites  in sub-Saharan Africa that compared the efficacy ) of AZCQ with that of AL in treatment of children with symptomatic uncomplicated falciparum malaria. An independent external data monitoring committee (EDMC), comprising international and regional malaria researchers, had oversight of the study. The study was conducted between June 2008 and September 2010.P. falciparum, mono-infection with parasite counts between 1,000 to 100,000 parasites/\u03bcL, and documented fever , or history of fever within the previous 24\u00a0hours. Subjects also had to have a blood glucose \u226560\u00a0mg/dL and haemoglobin \u22656\u00a0g/dL or haematocrit \u226518% without signs of anaemia-induced congestive heart failure, and a negative urine pregnancy test for females\u2009\u2265\u2009ten years of age. Informed consent for the study and permission from subjects\u2019 legal guardian for the three-day inpatient stay was also needed. Concomitant use of medications  was permitted. Use of medications with known drug interactions with macrolides was closely monitored. Medications metabolized by the CYP2D6 isoenzyme were not permitted (contra-indicated with AL). In addition, agents that prolong the QT interval and concomitant administration of other anti-malarial drugs were avoided because of limited safety data with AL.To be eligible for participation in the study, subjects were required to be able to receive treatment on an outpatient basis. Subjects were included if age ranged from five to 12\u00a0years of age (screening cohort) or between six and 59\u00a0months of age (primary study cohort) and had uncomplicated, symptomatic malaria. The presence of malaria was defined as positive blood smears for Plasmodium species, any history of allergy/hypersensitivity to or contraindication for use of any of the study drugs or history of treatment with anti-malarial drugs within two weeks prior to enrollment, body weight <5\u00a0kg or severe malnutrition, and known or suspected cardiovascular, hepatic, or renal abnormality, specific systemic diseases, or other medical conditions that would interfere with the evaluation of therapeutic response or safety of the study drug, other severe acute or chronic medical or psychiatric condition or laboratory abnormality, or other common febrile conditions, such as tonsillitis, measles, etc.Exclusion criteria included subjects with severe or complicated malaria or a peripheral blood smear indicating a mix of In Burkina Faso, the National Ethical Committee for Health Research reviewed and approved the final protocol, informed consent, and any amendments. At all other study centres, this review and approval was done by the institutional review board at each study centre. Oversight of safety data evaluation was provided through the activities of the EDMC. This study was conducted in compliance with the ethical principles originating in or derived from the Declaration of Helsinki and in compliance with all International Conference on Harmonization Good Clinical Practice Guidelines. The study protocol was submitted to the US Food and Drug Administration under an Investigational New Drug application. All local regulatory requirements were also followed.P. falciparum before proceeding with the primary study population cohort (Cohort 2) of younger children (six to 59\u00a0months of age). The decision to proceed to Cohort 2 for each site was based on the safety and efficacy of five evaluable AZCQ subjects observed in Cohort 1 at that site. The site could begin enrolling in Cohort 2 if all five evaluable subjects in the AZCQ group of Cohort 1 demonstrated a clinical and parasitological response within the first seven days of treatment initiation, continued clearance of parasitaemia through Day 28, and no clinically significant safety and tolerability issues were noted. If four of five subjects in the AZCQ arm demonstrated parasitological clearance and a clinical response within the first seven days of treatment initiation, the centre continued to enrol approximately ten additional subjects with five evaluable subjects receiving AZCQ. Continuing onto Cohort 2 required that eight or more of ten AZCQ-treated subjects demonstrated a clinical response and parasitologic clearance up to Day 28, and no significant safety and tolerability issues were noted. If fewer than four of five subjects in the AZCQ arm met the above criteria, the EDMC evaluated data from all sites and made recommendations on when to proceed with Cohort 2 enrolment. In all other cases, the decision to move from Cohort 1 to Cohort 2 for each centre was made by the investigator, the sponsor (Pfizer), and occasionally the EDMC.Subject enrolment was carried out in two stages. Each site was required to initially enrol ten evaluable subjects in a screening cohort (Cohort 1) of older children (five to 12\u00a0years of age) who were assumed to have some degree of immunity to In each cohort, subjects were randomized 1:1 to either receive an AZCQ fixed-dose combination tablet (30\u00a0mg/kg AZ\u2009+\u200910\u00a0mg /kg CQ base) orally once daily for three days  or AL orally per prescribing information for three days . A computer-generated randomization list was provided to investigators and randomization numbers were assigned sequentially as subjects were deemed eligible to participate in the study. Random assignment to either the AZCQ or AL treatment groups was determined by the assigned number from the randomization list. Two strengths of AZCQ were available for this study: AZ 300\u00a0mg/CQ 100\u00a0mg and AZ 150\u00a0mg/CQ 50\u00a0mg. Tablets were scored to allow for dosing by body weight. Subjects received AZCQ 300/100-mg tablets for \u226520\u00a0kg body weight: and AZCQ 150/50-mg tablets for 5 to <20\u00a0kg body weight. When possible, the study drugs were administered with or immediately after food consumption. Any dose of AZCQ that was vomited within 30\u00a0minutes of administration and any dose of AL that was vomited within 60\u00a0minutes of administration was repeated once. If vomiting recurred, the subject was discontinued from the study treatment.P. falciparum was identified using a blood dipstick-based test (Binax NOW immunochromatographic test (ICT)) and confirmed by microscopy on a Giemsa-stained peripheral blood smear. All eligible subjects were then hospitalized for\u2009\u2265\u2009three days for study drug administration and monitoring of the study key parameters. During this time, blood smears (thick and thin) were obtained every eight hours until two consecutive smears showed absence of parasitaemia. Clinical assessments, including adverse events (AEs), thick and thin peripheral smears and vital signs were evaluated on study Days 0, 3, 7, 14, 21, 28, 35, and 42. Electrocardiograms were performed in a limited number of subjects on study Days 0 and 2. At each site, clinical presentation and smear results from each study day were used to determine subject management. Discharge from the hospital occurred following clearance of asexual parasitaemia and investigator approval. All subjects were given insecticide-treated bed nets for use in the hospital and for use at home once discharged.At enrolment, the presence of Sparse pharmacokinetic (PK) blood samples were collected only in the AZCQ treatment group to determine serum AZ, plasma CQ, and desethyl-CQ concentrations. Whole blood samples were collected on Day 0 (prior to the first dose) at 0\u00a0hours (window: -1 to 0\u00a0hour); on Day 2 (the day of the third dose) at 0 , three (window: 2\u20134 hours), and eight hours (window: 6\u201310 hours), and randomly on Day 7. Samples were either centrifuged at 3,500\u00a0rpm for 10\u00a0min at 4\u00b0C or the centrifuge bucket and rotor were chilled on ice or in a refrigerator for two hours prior to use. Samples were stored within one hour of collection at -20\u00b0C until shipped for assay. AZ, CQ and desethyl-CQ concentrations were analysed using validated, sensitive and specific high-performance liquid chromatography tandem mass spectrometric methods in compliance with Pfizer standard operating procedures. AZ assays were performed by Bioanalytical Systems, Ltd (BASi)  and CQ/desethyl-CQ assays were performed by Cetero Research  .P. falciparum parasitologic clearance at Day 28 irrespective of axillary, oral, rectal, or tympanic temperature without previously meeting the criteria of early treatment failure or polymerase chain reaction (PCR)-corrected late treatment failure. PCR-corrected refers to the use of molecular testing to differentiate recrudescence from re-infection when evaluating efficacy  completed the Day 28 visit. Interim data were reviewed by the EDMC, and a determination to continue the study was made based on a predefined futility criterion (ACPR). ACPR was defined as asexual P. falciparum parasites at baseline and when parasitaemia occurred after initial clearance. Recurrence  was categorized as either recrudescence , or re-infection (infection by a different genotypic parasite as documented by molecular testing).Peripheral blood smear examinations were conducted to evaluate eligibility for inclusion by assessing parasite counts and identifying parasite species, parasite susceptibility to CQ and response to treatment. Molecular testing was done to genotype the Cohort 1 and Cohort 2 were analysed independently. Because Cohort 1 was only a small screening cohort, although data were analysed, no formal statistical inference was made. Three study populations were defined: 1) all treated subjects ; 2) modified intent-to-treat (MITT), defined as a subset of subjects in the all-treated subjects population who met the specified disease criteria at baseline; and, 3) the per-protocol (PP) population, defined as a subset of subjects in the MITT population who received all three days of assigned study medication.External quality checks were conducted for on-site microscopists. At Ivory Coast, all three microscopists failed the parasite-count evaluation; therefore, data (nine subjects) from Ivory Coast were excluded from all ACPR analyses for both the MITT and PP populations, but included in sensitivity analysis for the MITT population.Subjects who withdrew from the study due to any reason other than treatment failure were censored on the day of the last available blood smear measurement.Subjects who received an anti-malarial concomitant medication for treatment of a reinfection  were censored at the visit date if the anti-malarial was given after collection of the blood smear on that day. Otherwise, the subject was censored on the day of the last available blood smear measurement taken prior to the visit. Note that in a PCR-uncorrected analysis, this type of censoring only occurred when a subject was treated at a visit based on symptoms but later found not to have parasitaemia.The primary efficacy endpoint was the proportion of subjects with PCR-corrected ACPR at the Day 28 evaluation. Two datasets were analysed for efficacy: the MITT and PP populations (excluding data from Ivory Coast). Cohort 2 was the focus of the efficacy analysis. The primary analysis variable was the time (days) to the first occurrence of treatment failure analysed using the Kaplan-Meier method , 35, 42) was estimated from the Kaplan-Meier curve. A two-sided 95% confidence interval (CI) for the difference in ACPR proportions (AZCQ \u2013 AL) was constructed. Non-inferiority was concluded if the lower boundary of the CI was greater than or equal to -10 percentage points for both the MITT and PP populations at Day 28 (PCR-corrected).A sensitivity analysis was conducted to support the primary efficacy analysis for both the MITT and PP populations. This sensitivity analysis considered subjects who prematurely discontinued from the study due to any reason, without having an event, as treatment failures at the time of withdrawal.P. falciparum gametocytaemia  without subsequent recurrence through the day of consideration. With the exception of ACPR, all other secondary endpoints were analysed using the MITT study population. Descriptive statistics of concentration data for serum AZ, plasma CQ and plasma desethyl-CQ were provided by treatment, cohort, study day, and nominal time post-dose.Secondary study endpoints included ACPR (PCR-corrected and -uncorrected) at Days 7, 14, 21, 28 (uncorrected), 35, and 42 as well as early and late treatment failures (Table\u00a0A target sample size of approximately 244 subjects in Cohort 2 (approximately 122 subjects per arm) was determined based on the following assumptions: 1) an evaluability rate (for the purpose of sample size determination) of 85% corresponding to the Day 28 visit ; 2) an expected ACPR (PCR-corrected) rate of 95% in the AZCQ group and 95% in the AL group; 3) a power of 80% to show non-inferiority of the AZCQ group relative to the AL group; and, 4) accounting for any loss in power due to the efficacy futility interim analysis.P. falciparum malaria following blood smear microscopy with mean baseline parasite counts that ranged from 1,000/\u03bcL to 110,240/\u03bcL. All study centres met the criteria for enrolling subjects into Cohort 2.The screening cohort included 106 subjects. After excluding subjects from the Ivory Coast, the MITT population included 85 subjects. Four subjects or their parent/legal guardian in the AZCQ treatment group were no longer willing to participate and discontinued from the study. There were no treatment discontinuations in the AL group. The mean age was 7.4\u00a0years for AZCQ-treated subjects and 7.9\u00a0years for AL-treated subjects. The AL group had a higher proportion of males. All subjects had confirmed The primary study population included 255 subjects who were screened and randomized Figure\u00a0. After eDemographic characteristics of treatment groups were similar, except for gender ratio; there was a higher proportion of males in the AZCQ group Table\u00a0. The meaAt baseline, all subjects had either fever or a history of fever within 24\u00a0hours of consent to participate in the study. The mean duration between subjects\u2019 first malaria symptom and initiation of consent process for this study was 1.9\u00a0days, and\u2009\u2264\u2009three hours elapsed between consent and drug administration in all but 12 subjects.versus 98%  in the AL group achieved the primary endpoint  in the AL group achieved the primary endpoint (Table\u00a0. The between-group differences in the PP population (AZCQ-AL) was -6% , which also did not meet the non-inferiority criterion. Although not a pre-specified comparison in the study protocol, since the CI for both the MITT and PP populations also excluded zero, AZCQ was inferior to AL at the Day 28 time point.Efficacy data were used to determine if AZCQ was non-inferior to AL; based on the primary endpoint of ACPR at Day 28 (PCR-corrected), non-inferiority of AZCQ to AL was not demonstrated. In the MITT population, 89% 95% CI: 83%, 96%) of subjects in the AZCQ group %, 96% ofversus 73% in the AL group at Day 28, with similar findings in the PP population.The proportion of subjects with ACPR (PCR-corrected) was numerically higher in the AL group than in the AZCQ group at all visits for both analysis populations Table\u00a0. A signiMost  of the treatment failures (up to Day 28 including early treatment failures) were from a single centre . Estimates of ACPR at Day 28 (MITT) were essentially unchanged when accounting for each subject\u2019s baseline parasite count . Early treatment failures were more common in the AZCQ group (5.83% (MITT) and 1.75% (PP)) than in the AL group (0.79% (MITT) and 0% (PP)). Also, higher proportions of late parasitological failures were observed in the AZCQ group (4.17% (MITT) and 4.39% (PP)) than in the AL group (0.79% (MITT) and 0.81% (PP)). No late clinical failures were observed in either treatment group (MITT or PP).Plasmodium falciparum gametocyte clearance rates were higher at all time points for AL-treated subjects (>90%) compared with that for AZCQ-treated subjects (>80%).The time to fever clearance between groups was approximately 24\u00a0hours in both treatment groups with no statistically significant difference between groups. One subject in each treatment group had fever that persisted for more than 72\u00a0hours despite being aparasitaemic from study Day 1 (AL) or study Day 2 (AZCQ). Serum concentrations of AZ and plasma concentrations of CQ and desethyl-CQ in each cohort and for the combination of cohorts are reported in Table\u00a0versus one (0.8%) in the AL group. No meaningful differences in laboratory parameters were noted between AZCQ and AL treatment groups  were reported. Three SAEs occurred in Cohort 1 , sepsis and hepatitis B ), and one SAE  in Cohort 2. None of these SAEs were considered related to study drug by investigators. Seven Cohort 2 subjects (5.6%) in the AZCQ group withdrew from treatment due to an AE, The most frequently reported AEs in Cohort 2 overall were asymptomatic parasitaemia (coded as infection parasitic), vomiting, malaria, and pyrexia Table\u00a0. The mosBased on the predefined primary endpoint in this study, non-inferiority of AZCQ relative to AL was not demonstrated. Although both treatment groups had a high proportion of subjects achieving ACPR at Day 28 (PCR-corrected), the proportion was lower (i.e. inferior) in the AZCQ group compared with the AL group.In the PP population, the efficacy was 93% in AZCQ-treated subjects compared with 99% in the AL-treated subjects at Day 28. Similarly, in the MITT, efficacy was 89% in AZCQ-treated subjects compared with 98% in AL-treated subjects. The conclusions from the sensitivity analyses performed for all ACPR endpoints, including subjects from the Ivory Coast and classifying study discontinuations as failures, were no different from the main study conclusion. The evaluability criteria may explain the small difference in AZCQ efficacy observed between the PP and MITT populations, since subjects in the PP population were required to receive all three days of study medication. This study used the Kaplan-Meier curve/product limit estimator to evaluate the primary endpoint. Use of this limit estimator is recommended by WHO  and has P. falciparum malaria, treatment with AZ 1,000\u00a0mg and CQ 600\u00a0mg resulted in Day 28 PCR-corrected parasitologic clearance rates of 98% [P. falciparum malaria in Bangladesh [P. falciparum malaria in children six to 59\u00a0months of age in Tanzania [versus 30\u00a0mg/kg). In addition, CQ, alone and in combination with artesunate, AZ, or atovaquone-proguanil, has been used to treat uncomplicated P. falciparum malaria in similar-aged children in Malawi [P. falciparum, and the incidence of malaria was similar across the treatment groups.Absence of non-inferiority of AZCQ observed in this study was an unexpected outcome because there is evidence of effectiveness of AZ in combination with CQ and other agents for treatment of malaria. In two randomized, comparative studies in African adults with s of 98% . In one ngladesh . AlthougTanzania . Treatmen Malawi . In thisIn the present study, the serum AZ and plasma CQ concentrations were generally comparable between Cohorts 1 and 2. The large coefficient of variation may limit the ability to draw conclusions from these data. The mean values of AZ, CQ, and desethyl-CQ concentrations at 0\u00a0hours on Day 2 (prior to the third dose) and on Day 7 (after the third dose) that were observed in this paediatric study were similar to the concentrations at same PK sampling time points observed in African adults treated with 1,000\u00a0mg AZ plus 600\u00a0mg CQ once daily for three days. In the African study, the combination of AZ and CQ resulted in an overall efficacy rate of 100% at Day 28 [The overall proportion of subjects with AEs was similar across treatment groups within each cohort, although treatment-related AEs were reported more frequently in the AZCQ group in both cohorts. Posthoc analysis of the frequency of AEs in cohort 2 showed that, for most, there was no statistically significant difference between treatment groups, however, vomiting was more frequent in the AZCQ group and abdominal pain was more frequent in the AL group. Study discontinuation due to AEs was higher in the AZCQ group in Cohort 2. No unexpected safety signals were identified beyond the known AE profile of individually dosed AZ or CQ.In this study, efficacy of AZCQ was inferior to AL. Therefore, consideration of future use of AZCQ for IPT or seasonal malaria chemoprophylaxis may require additional study to determine whether a combination of AZ with other malaria drug(s) or use of an alternative dosing regimen with higher AZ dosing improves response to treatment with AZCQ in children aged six to 59\u00a0months."}
+{"text": "Plasmodium falciparum malaria, respectively. However, recent studies suggest that delayed anaemia is associated with these treatments in non-immune travellers. This paper aimed to assess the risk factors associated with delayed anaemia after falciparum malaria treatment with artemisinin-containing drugs in malaria-endemic populations.In sub-Saharan Africa, artemisinin-based combination therapy (ACT) and injectable artesunate are the first-line treatments for uncomplicated and severe Pooled, individual malaria patient data were extracted from 13 clinical trials performed from 2002 to 2011 in various settings of Mali. Treatment regimens were artemether-lumefantrine, artesunate plus amodiaquine, artesunate plus sulphadoxine-pyrimethamine, artesunate plus sulphamethoxypyrazine-pyrimethamine, artesunate plus mefloquine, artesunate-pyronaridine, artesunate monotherapy, chloroquine, sulphadoxine-pyrimethamine, amodiaquine and sulphadoxine-pyrimethamine plus amodiaquine. Univariate and multivariate analyses were performed using the generalized linear and latent mixed model procedures to assess risk factors associated with haemoglobin concentration evolution and anaemia during the treatment follow-up.A total of 5,990 participants were recruited and followed from day 0 to day 28. The participants\u2019 median age was five years, ranging from three months to 70\u00a0years. There was a decrease in haemoglobin level on day 7 in all treatment arms, but the magnitude varied across treatments. There was a significant risk of haemoglobin level decrease on day 7 in the artemisinin-based therapy compared to the non-artemisinin treatments. The risk of haemoglobin concentration drop was associated with age group\u2009<\u2009five years old (0.61\u00a0g/dL 95% CI (0.71 to 0.51), p\u2009<\u20090.001), baseline high parasite density (0.43\u00a0g/dL 95% CI (0.51 to 0.35), p\u2009<\u20090.001) and treatment failure (0.40\u00a0g/dL 95% CI (0.59 to 0.20), p\u2009=\u20090.018), while high haemoglobin level at baseline was a protective factor (0.53 to 0.59) p\u2009<\u20090.001). No association was found between artemisinin-based therapy and severe delayed anaemia.P. falciparum malaria are associated with a transient and clinically moderate haemoglobin decrease by day 7 but not associated with a delayed severe anaemia.Oral artemisinin derivative treatments for uncomplicated The online version of this article (doi:10.1186/1475-2875-13-358) contains supplementary material, which is available to authorized users. Plasmodium falciparum to monotherapy, and to improve treatment outcome, WHO recommends the use of artemisinin-based combination therapy (ACT) for the treatment of uncomplicated P. falciparum malaria [According to the World Health Organization (WHO) 2013 report, 207 million cases of malaria were estimated to have occurred in 2012 with 627,000 deaths [ malaria . However malaria \u20139. WHO r malaria , 11. Theth day of follow-up, but did not provide the specific day of the occurrence of anaemia during the follow-up period. As precise knowledge of the specific occurrence day of haemoglobin change or anaemia occurrence is of clinical relevance, this paper aimed to perform a pooled analysis of data from 13 clinical trials conducted in Mali. The goal of this pooled data analysis was to describe the dynamics of haemoglobin levels, delayed anaemia and the associated risk or protective factors during each follow-up day, up to 28\u00a0days.There is limited large-scale data in the literature from malaria-endemic areas assessing haematological parameters, particularly anaemia among patients treated with the artemisinin-containing malaria treatments. Recently a publication from a large-scale, pooled, multi-country study data from malaria-endemic areas assessed haematologic parameter changes in patients treated with ACT  and other therapy  . That stet al. Host immunological factors involved in clearance of drug resistant falciparum malaria, unpublished observation; Maiga H et al. Dhfr-dhps quadruple mutant predicts sulfadoxine-pyrimethamine resistance in Mali, an emerging sulfadoxine-pyrimethamine resistance setting, unpublished observation).A pooled, malaria-treatment, clinical trial analysis was performed, using individual data extracted from clinical trial databases of 13 trials conducted by researchers from Malaria Research and Training Centre (MRTC), Bamako, a research institution within the University of Sciences, Techniques and Technologies of Bamako (USTTB), Mali. The different studies were performed from 2002 to 2011 in various malaria-transmission settings of Mali   ACT: artemether-lumefantrine ((AL) Coartem\u00ae from Novartis); artesunate plus amodiaquine ((AS-AQ), loose or fixed-dose combination, Arsucam/Coarsucam\u00ae, both from Sanofi-Aventis); artesunate plus sulphadoxine-pyrimethamine ((AS-SP), loose combination; AS, Arsumax\u00ae from Sanofi-Aventis and SP, Fansidar\u00ae from Roche); artesunate plus sulphamethoxypyrazine-pyrimethamine ((AS-SMP), loose combination, Coarinate\u00ae from Dafra Pharma); artesunate plus mefloquine ((AS-MEF), loose combination, Artequin\u00ae from Mepha Pharma); and, artesunate-pyronaridine ((AS-PYR), fixed-dose combination, Pyramax\u00ae from Shin Poong); (ii) artesunate oral monotherapy ((AS), Arsumax\u00ae or Asunate Denk\u00ae respectively from Sanofi-Aventis and Denk Pharma); (iii) non ACT: chloroquine (CQ); amodiaquine (AQ); sulphadoxine-pyrimethamine (SP) and sulphadoxine-pyrimethamine plus amodiaquine ((SP-AQ), loose combination).et al. Host immunological factors involved in clearance of drug resistant falciparum malaria, unpublished observation; Maiga H et al.: Dhfr-dhps quadruple mutant predicts sulfadoxine-pyrimethamine resistance in Mali, an emerging sulfadoxine-pyrimethamine resistance setting, unpublished observation) and also in a Additional file Treatment courses durations were three days for all except for: a) SP given as a single dose; and, b) AS given up to five days in one study  and seveData were systematically analysed according to three outcomes: dynamic of haemoglobin level, haemoglobin drop and anaemia occurrence.Haemoglobin levels were analysed as a continuous variable (haemoglobin changes outcome) and as categorical variables (haemoglobin drop and anaemia occurrence). Drop of haemoglobin value was defined from day 0 to the day of the different follow-up visit  with the following thresholds: \u22651\u00a0g/dL or \u22652\u00a0g/dL. Anaemia was defined as a haemoglobin value <10\u00a0g/dL and severe anaemia as haemoglobin value <8\u00a0g/dL as described elsewhere . DelayedFor these outcomes, multivariate analyses assessed the different associated risk factors during each visit after treatment  in order to determine any significant changes during the follow-up. These analyses included all participants for whom haemoglobin data were available on day 0 and at least one follow-up day (day 7 or 14 or 28). Specifically for the delayed severe anaemia occurrence, associated risk factors were assessed with different time points: (i) from day 7 to day 28; (ii) from day 14 to day 28; and, (iii) on day 28. These analyses used a subset of participants for which the haemoglobin data were available for all follow-up visits . Different late or delayed severe anaemia periods were assessed: (i) an absence of severe anaemia on day 0 followed by its occurrence during any subsequent days for follow-up (7 or 14 or 28); (ii) an absence of severe anaemia on day 0 and day 7 followed by its occurrence during any subsequent days for follow-up ; and, (iii) an absence of severe anaemia on day 0 and day 7 and day 14 followed by its occurrence on day 28.Descriptive statistics provided the total number of participants, means, standard deviations, medians, minimums, and maximums for quantitative variables and proportions for qualitative variables.Univariate analyses assessed the haemoglobin changes between day 0 and the day of post-treatment evaluation  using paired Student test in each treatment arm.For both continuous variable of haemoglobin changes and categorized variable of haemoglobin drop or anaemia outcomes, multivariate analyses were performed using generalized linear and latent mixed model (GLLAMM), with study site as a latent factor and as random effect. Models were systematically adjusted for covariates such as age, parasite density at baseline (log transformed), malaria treatment, and treatment failure during the follow-up. For the haemoglobin changes outcome, the gaussian family  was used to estimate the coefficients of covariate effects. For the categorized haemoglobin drop and anaemia outcomes, the binomial family  was used to estimate odds ratios associated with covariates.The confidence intervals (CIs) were estimated at 95% and p value <0.05 was considered significant. Data were analysed with GLLAMM procedure using Stata Software version 12.1 (Stata Corp).Each study had a prior approval from the institutional ethical committee at the Faculty of Medicine, Pharmacy and Odonto-Stomatogy (FMPOS)/USTTB, Bamako, Mali. Written consent was obtained in each study from each participant or parent/legal guardian.This pooled data of 13 trials in eight Malian sites  in all treatments except for AQ Table\u00a0. The freMultivariate analyses showed a higher risk for haemoglobin decrease in the ACT (AS or the combined AS-AQ/AS-SP/AS-PYR and AL) compared to the non ACT (CQ/AQ/SP). The risk of haemoglobin decrease was significant for the AS, mean decrease of 0.69\u00a0g/dL 95% CI 0.95-0.44), p\u2009<\u20090.001; and the AS-containing combinations: AS-AQ/AS-SP/AS-PYR, 0.34\u00a0g/dL, 95% CI (0.57-0.11), p\u2009=\u20090.003  frequency on day 0, 7, 14, and 28 was 35.5% , 45.7% , 35.3% , and 23.9% , respectively.The severe anaemia (haemoglobin <8\u00a0g/dL) frequency on day 0, 7, 14, and 28 was, respectively, 9.0% , 10.1% , 6.2% , and 4.0% .Among a total of 1,053 eligible patients , 135 patients (12.8%) experienced severe anaemia from day 7 to day 28. Multivariate analysis showed a risk of severe anaemia associated with the AS arm from day 7 to day 28 (OR\u2009=\u20091.85 (1.05-3.27); p\u2009=\u20090.03) compared to the non-artemisinin-contained treatments   compared to the non-artemisinin-contained treatments  from day 7 to day 28 , 38 patients (4%) experienced severe anaemia from day 14 to day 28. Multivariate analyses , 23 patients (2.4%) experienced severe anemia on day 28. Multivariate analyses  conducted in 2002\u20132004, which enrolled 948 patients in CQ arm.The risk of blood transfusion was assessed: a total of five cases of blood transfusion were documented, all in children\u2009\u2264\u2009three years old. Transfusion was done based both on low haemoglobin and clinical anaemia symptoms as judged by the physician. There were three in AS-AQ arm , one in AS arm (on day 7) and one in SP arm (unknown day). Data of these patients were excluded from this analysis once they received the blood transfusion. The transfusion information was not available for one study  were associated with the risk of haemoglobin drop. As reported in this study, haemoglobin increase or anaemia recovery, which is common by day 28, was also documented in several studies , 24, 25.The frequency of severe anaemia, which was about 9% on day 0 and 10% on day 7, decreased gradually to 4% by day 28. Multivariate analyses indicated that there was a risk of severe anaemia associated with the AS arm from day 7 to day 28, compared to the non-ACT (CQ/AQ/SP/SP\u2009+\u2009AQ). But this was not evident from day 14 to 28 or on day 28. These observations could be explained by the relatively important decrease of haemoglobin shown in this study on day 7 particularly with AS arm Table\u00a0. ArtemisFollowing the previous study reporting anaemia without haemoglobin evolution assessment , this stThis study could not investigate the mechanisms involved in the haemoglobin evolutions observed. Indeed, the decrease on day 7 needs more study to improve the understanding of anaemia mechanisms, such as the reticulocytes evaluation. There is also a need for malaria treatment data from various countries, injectable AS treatment, vulnerable groups .P. falciparum malaria are associated with a transient and clinically moderate haemoglobin decrease by day 7 but not associated with a delayed severe anaemia.No association was found between ACT and severe delayed anaemia. Oral artemisinin derivative treatments for uncomplicated Study treatment doses.(PDF 107 KB)Additional file 1:"}
+{"text": "This post hoc analysis assessed the safety, tolerability and effectiveness of long-term treatment with aripiprazole adjunctive to either bupropion or selective serotonin reuptake inhibitors (SSRIs)/serotonin\u2013norepinephrine reuptake inhibitors (SNRIs) in patients with major depressive disorder (MDD).Data from de novo patients (did not participate in 2 previous studies) in a 52-week, open-label safety study of adjunctive aripiprazole after documented inadequate response to 1\u20134 antidepressant treatments  were analyzed post hoc. Assessments included safety and tolerability, sexual functioning  and Clinical Global Impressions\u2013Severity (CGI-S).Forty-seven patients received bupropion plus aripiprazole and 245 received an SSRI/SNRI plus aripiprazole; 19 (40.4%) and 78 (31.8%), respectively, completed 52\u00a0weeks of treatment, and 46 and 242, respectively, received \u22651 dose of study medication (safety sample). Median time to discontinuation (any reason) was 184.0\u00a0days. Overall, 97.8% of patients in the bupropion group and 93.8% in the SSRI/SNRI group experienced \u22651 adverse event. The most common treatment-emergent adverse events were fatigue (26.1%) and somnolence (21.7%) with bupropion and fatigue (23.6%) and akathisia (23.6%) with an SSRI/SNRI. Mean change in body weight at week 52 (observed cases) was +3.1\u00a0kg for bupropion and +2.4\u00a0kg for an SSRI/SNRI. Treatment-emergent, potentially clinically relevant abnormalities in fasting glucose occurred in 8.3% of patients with bupropion and 17.4% with an SSRI/SNRI; for abnormalities in fasting total cholesterol, the incidence was 25.0% and 34.7%, respectively. Mean (SE) change from baseline in fasting glucose was 1.4 (1.9) mg/dL with bupropion and 2.7 (1.5) mg/dL with an SSRI/SNRI. Baseline MGH-SFI item scores indicated less severe impairment with bupropion versus an SSRI/SNRI; in both groups most MGH-SFI items exhibited improvement at week 52. Mean CGI-S improvement at week 52 (last observation carried forward) was -1.4 with bupropion and -1.5 with an SSRI/SNRI (efficacy sample).There were no unexpected AEs with long-term adjunctive aripiprazole therapy when added to either bupropion or SSRIs/SNRIs, and symptom improvement was similar between ADT groups. Sexual functioning in patients with MDD on antidepressants was also modestly improved after adding aripiprazole.NCT00095745 .ClinicalTrials.gov: Furtheceptors. In thisIn the previously conducted, short-term, placebo-controlled trials-8, aripiBeyond issues pertaining to safety, there are other reasons to examine the combination of bupropion and adjunctive aripiprazole. For example, whereas SSRIs and SNRIs can have adverse effects on sexual function, bupropiHypodopaminergic activity may be associated with diminished drive, amotivation, reduced cognitive function and lack of positive mood. MoreoveAripiprazole as adjunctive treatment to bupropion was permitted in patients directly enrolling in the 52-week, open-label trial of aripiprazole augmentation; data frde novo subgroup were aged \u226518\u00a0years with Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, Text Revision criteria for a major depressive episode and a duration of current depressive episode of at least 8\u00a0weeks. All patients entered a 7- to 28-day pre-treatment screening phase consisting of a screening and baseline visit to assess study eligibility criteria and allow washout of prohibited concomitant pharmacotherapy. Full details of inclusion criteria have been presented previously[de novo enrollment. Patients or their legal representatives provided written informed consent before participation. The study protocol was approved by the institutional review board at each site either through a local IRB or one of the following central IRBs: the Institutional Review Board, Inc., Laguna Hills, California; the Schulman Associates IRB, Inc., Cincinnati, Ohio; the IRB of the Office of Scientific Affairs, Philadelphia, Pennsylvania; and the Western Institutional Review Board, Olympia, Washington. The study was conducted in accordance with the ethical principles set forth in the Declaration of Helsinki.This was a post hoc analysis of data from a subgroup of patients directly enrolled  in a multicenter, 52-week, open-label study [Patients were eligible to enter 52\u00a0weeks of open-label treatment if they had an inadequate response  to at least one but no more than four ADT trials (each of at least 6\u00a0weeks duration at an adequate dose); a MontgAll patients received ADT in accordance with current product labelling, with dose adjustments permitted within the recommended dose range. Adjunctive aripiprazole was initiated at 5\u00a0mg/day, and dosed in the range of 2\u201330\u00a0mg/day for patients receiving venlafaxine XR, escitalopram, mirtazapine or sertraline and 2\u201315\u00a0mg/day for patients on fluoxetine, paroxetine, duloxetine or bupropion .Safety was evaluated by monitoring adverse events (AEs) and vital signs (at baseline and each study visit), body weight  and 12-lead electrocardiogram (ECG) . Laboratory tests, including fasting metabolic parameters, were conducted at open-label treatment baseline and Weeks 8, 26, 38 and 52. Effectiveness was assessed throughout the open-label treatment period using the Clinical Global Impressions\u2013Severity (CGI-S) of illness (decrease in score signifies improvement in severity of illness). In addiFor this post hoc analysis, data were pooled into the two ADT subgroups of bupropion plus aripiprazole and SSRI/SNRI plus aripiprazole. Safety analyses included all patients who received at least one dose of aripiprazole as adjunctive therapy to either bupropion or an SSRI/SNRI ADT, and analyses of effectiveness included all patients in the safety sample who had at least one CGI-S assessment during open-label treatment (efficacy sample). Summary statistics for change in effectiveness and safety rating scale scores are presented for patients treated with bupropion and for all other SSRI/SNRI ADTs combined. Data for patients receiving bupropion XL or bupropion SR were pooled into one group. Data for aripiprazole plus bupropion were compared only to the subset of patients receiving aripiprazole plus an SSRI/SNRI because all patients in the registered trials received aripiprazole as an adjunct to an SSRI/SNRI. This exploratory, post hoc subgroup analysis is limited by reduced power, increased variance, and an increased influence of chance; thus, nde novo patients  of whom 296 began the open-label treatment phase; the remaining 74 patients provided informed consent but did not meet eligibility criteria and were excluded from the open-label treatment protocol. Patient disposition of those treated with bupropion plus aripiprazole or an SSRI/SNRI plus aripiprazole is shown in Figure\u00a0Disposition, demographics, efficacy, safety, and tolerability in the overall study population were previously reported. This stAt entry into open-label treatment, the distribution of ADTs was as follows: bupropion XL/SR, n\u2009=\u200946 (15.8%); duloxetine, n\u2009=\u20097 (2.4%); escitalopram, n\u2009=\u200966 (22.7%); fluoxetine, n\u2009=\u200942 (14.4%); paroxetine, n\u2009=\u200928 (9.6%); paroxetine CR, n\u2009=\u200910 (3.4%); sertraline, n\u2009=\u200939 (13.4%); and venlafaxine XR, n\u2009=\u200950 (17.2%). The mean doses of ADT at study endpoint (the last day of each ADT dosing) were as follows: bupropion XL/SR 340\u00a0mg/day (n\u2009=\u200946); duloxetine 60\u00a0mg/day (n\u2009=\u20097); escitalopram 16.8\u00a0mg/day (n\u2009=\u200965); fluoxetine 41.2\u00a0mg/day (n\u2009=\u200942); paroxetine 34.4\u00a0mg/day (n\u2009=\u200927); paroxetine CR 29.3\u00a0mg/day (n\u2009=\u200910); sertraline 125\u00a0mg/day (n\u2009=\u200939); and venlafaxine XR 167\u00a0mg/day (n\u2009=\u200950). The mean dose of aripiprazole at study endpoint was 9.6\u00a0mg/day in patients receiving bupropion plus aripiprazole and 9.3\u00a0mg/day in patients receiving an SSRI/SNRI plus aripiprazole .Forty-six patients treated with bupropion plus aripiprazole and 242 treated with an SSRI/SNRI plus aripiprazole received at least one dose of study medication and were included in the safety sample. The baseline demographic characteristics of patients are presented in Table\u00a0During open-label treatment, 272 (94.4%) patients experienced at least one AE, and the rate of AEs was similar between bupropion plus aripiprazole  and an SSRI/SNRI plus aripiprazole . Treatment-emergent AEs, by ADT, that occurred at an incidence \u226510% in either ADT group are shown in Table\u00a0With respect to less common AEs, no patient treated with bupropion plus aripiprazole experienced seizures. Two SSRI/SNRI plus aripiprazole-treated patients reported seizures; a 53-year-old man with a remote history of a grand mal seizure reported a convulsion while receiving paroxetine 60\u00a0mg/day and aripiprazole 5\u00a0mg/day, and a 22-year-old female patient receiving escitalopram who had discontinued the study on Day 118 reported a convulsion on Day 155. Neither event was considered related to study medication.Both ADT groups experienced a mean increase in weight over the course of treatment. Mean weight change at Week 52 (OC) was +3.1\u00a0kg for bupropion plus aripiprazole, and +2.4\u00a0kg for an SSRI/SNRI plus aripiprazole. Clinically significant weight gain (increase of \u22657% from baseline) occurred in 20.6% of bupropion plus aripiprazole and 26.2% of SSRI/SNRI plus aripiprazole patients at Week 52 (LOCF). Clinically significant weight loss (decrease of \u22657% from baseline) occurred in 5.9% and 7.1% of bupropion plus aripiprazole and SSRI/SNRI plus aripiprazole patients, respectively.The metabolic findings for patients treated with bupropion plus aripiprazole and an SSRI/SNRI plus aripiprazole are shown in Table\u00a0Mean MGH-SFI overall improvement scores over the course of treatment for patients treated with bupropion plus aripiprazole and an SSRI/SNRI plus aripiprazole, by gender, are shown in Figure\u00a0Mean change in MGH-SFI item scores from baseline to Week 52, by gender, for patients treated with bupropion plus aripiprazole and an SSRI/SNRI plus aripiprazole are shown in Table\u00a0C (Bazett) prolongation in patients treated with an SSRI/SNRI plus aripiprazole (8.8%), with 1.8% of patients experiencing the abnormality at more than one time point; all other abnormalities occurred in <3% of patients. Only one ECG abnormality was reported in bupropion plus aripiprazole-treated patients: one case of QTC (Bazett) prolongation (2.4%). The incidence of potentially clinically relevant serum chemistry, hematology and electrolyte measurements was also low in both ADT groups. Exceptions to this were the incidence of potentially clinically relevant prolactin abnormalities (14.6% for bupropion plus aripiprazole and 16.0% for an SSRI/SNRI plus aripiprazole), hemoglobin A1C (7.3% for bupropion plus aripiprazole and 10.2% for an SSRI/SNRI plus aripiprazole) and uric acid levels in females (11.1% for bupropion plus aripiprazole and 2.0% for an SSRI/SNRI plus aripiprazole).The incidence of potentially clinically relevant electrocardiographic abnormalities was low with both bupropion plus aripiprazole and an SSRI/SNRI plus aripiprazole. The most common ECG abnormality was QTImprovement in symptom severity, as measured by mean change in CGI-S from baseline to Week 52 for bupropion plus aripiprazole and an SSRI/SNRI plus aripiprazole patients are shown in Figure\u00a0Aripiprazole is an atypical antipsychotic with a unique pharmacological profile, and while the exact mechanism of action of atypical antipsychotic augmentation is not well understood, it is possible that the pharmacological profile of aripiprazole may be particularly effective as an augmentation agent in depression. In this study, improvement in depressive symptoms occurred when aripiprazole was added to either bupropion or an SSRI/SNRI and symptom improvement was maintained over a 52-week treatment period at a similar level with both ADT groups. Thus, combining two agents with effects on dopaminergic activity does not appear to inhibit the benefits of aripiprazole as an augmentation agent. However, the pharmacodynamic effects of concomitant use of aripiprazole and bupropion on dopamine neurotransmission warrant further investigation.With respect to safety, this is to our knowledge the largest prospective study of the use of aripiprazole adjunctive to bupropion conducted to date. Aripiprazole augmentation of bupropion was not associated with any unexpected AEs and had a long-term safety profile that was comparable to that of aripiprazole augmentation of SSRIs/SNRIs in patients with MDD who had experienced an inadequate response to ADT monotherapy. Importantly, there was no evidence of an adverse synergistic or additive effect on dopamine associated with co-administration of aripiprazole and bupropion, compared with other aripiprazole/ADT combinations, and reasons for discontinuation were similar between adjunctive ADT groups. Notably, rates of akathisia were not higher with bupropion plus aripiprazole than with aripiprazole adjunctive to SSRI/SNRI ADTs. Seizures are one of the neurologic side effects reported with bupropion,11. WhilBoth bupropion and SSRIs/SNRIs adjunctive with aripiprazole were associated with a mean increase in weight over the 52\u00a0weeks of treatment, with no apparent association between type of antidepressant and the extent of weight gain. Although it is difficult to establish the relative contribution of aripiprazole to the weight gain reported here given that long-term administration of antidepressant medications is also associated with medically relevant weight gain, all patMetabolic disturbances associated with medication are particularly important during long-term treatment. The incidence of treatment-emergent, potentially clinically relevant abnormalities in fasting glucose levels was lower in patients receiving bupropion plus aripiprazole than an SSRI/SNRI plus aripiprazole and mean changes in glucose levels tended to favor the group treated with bupropion plus aripiprazole. The incidence of treatment-emergent potentially clinically relevant abnormalities in fasting total cholesterol was also lower for patients receiving bupropion plus aripiprazole than an SSRI/SNRI plus aripiprazole, although there was no difference in mean change from baseline between the treatment groups. Findings for other metabolic parameters are harder to interpret. However, the relatively favorable metabolic profile of aripiprazole among atypical antipsychotics support the possibility that changes in metabolic parameters stem from SSRI/SNRI treatment in this patient group or theirSexual dysfunction is common in patients with MDD and is an important concern during treatment, especially as some antidepressant therapies can contribute to the emergence or exacerbation of sexual dysfunction. Although variability exists within the therapeutic classes, SSRIs and SNRIs generally worsen sexual function, while nThis analysis explored the effects of aripiprazole as adjunctive therapy to bupropion. One of the strengths of this analysis is that the large patient population included in the original multicenter study consisted of a relatively large subgroup of patients treated with bupropion plus aripiprazole, permitting further analysis of this data set; previously published research on the use of aripiprazole adjunctive to bupropion is limited to four case reports. HoweverThe addition of aripiprazole augmentation to bupropion was not associated with any unexpected AEs and the overall tolerability of adjunctive aripiprazole was similar between bupropion and SSRI/SNRI. The addition of aripiprazole augmentation to antidepressant therapy results in improvements in depression symptoms and sexual function in patients with MDD who had not responded to treatment with one or more ADTs and the benefits did not vary when added to the norepinephrine\u2013dopamine reuptake inhibitor bupropion or an SSRI/SNRI, the latter of which is now a well-established combination.ADT: Antidepressant treatment; AE: Adverse event; CGI-S: Clinical Global Impressions\u2013Severity score; CR: Controlled release; ECG: Electrocardiogram; LOCF: Last observation carried forward; MADRS: Montgomery-\u00c5sberg Depression Rating Scale; MDD: Major depressive disorder; MGH-SFI: Massachusetts General Hospital Sexual Functioning Inventory; OC: Observed case; SNRI: Serotonin\u2013norepinephrine reuptake inhibitor; SR: Sustained release; SSRI: Selective serotonin reuptake inhibitor; XL: Extended release (bupropion); XR: Extended release (venlafaxine).During the past 5\u00a0years, Anita H. Clayton has received research grants from BioSante Pharmaceuticals, Inc., Boehringer-Ingelheim, Bristol-Myers Squibb, Eli Lilly and Company, Forest Research Institute, Inc., Novartis, Palatin Technologies, Pfizer, Inc., Repligen Corporation, sanofi-aventis, Takeda, and Trimel Pharmaceuticals; has been an advisor/consultant to Apricus Biosciences, Inc., Astra Zeneca, Boehringer-Ingelheim, Bristol-Myers Squibb, Concert Pharmaceuticals, Eli Lilly and Company, Euthymics, Forest Research Institute, Inc., Labopharm, Inc, Lundbeck, New England Research Institutes, Inc., Novartis Pharmaceuticals, Palatin Technologies, Pfizer, Inc., PGxHealth, sanofi-aventis, S1 Biopharmaceuticals, Inc., Sprout Pharmaceuticals, Takeda Global Research & Development, TransTech Pharma, Inc., Trimel Pharmaceuticals, and Wyeth; has participated in a disease-state speakers bureau for Boehringer-Ingelheim and Eli Lilly and Company; has received royalties/copyright fees from Ballantine Books/Random House, Changes in Sexual Functioning Questionnaire, Guilford Publications, and Healthcare Technology Systems, Inc.; has held shares/restricted stock units in Euthymics and S1 Biopharmaceuticals, Inc.; and has had a US provisional patent application .During the past 3\u00a0years, Michael E. Thase has been an advisor/consultant for Aldolar, Alkermes, AstraZeneca, Bristol-Myers Squibb., Eli Lilly & Co., Forest Pharmaceuticals , Johnson & Johnson (Janssen Pharmaceutica), Lundbeck, MedAvante, Inc., Merck, Neuronetics, Inc., Otsuka, Pfizer Inc., PharmaNeuroBoost, Rexahn, Roche, Shire US, Inc., and Takeda. Dr Thase has received honoraria for talks from AstraZeneca, Bristol-Myers Squibb, Eli Lilly & Co., Merck, and Pfizer. Dr Thase has received research grants from the Agency for Health Care Research and Quality, Alkermes, the National Institute of Mental Health, Eli Lilly & Co., Forest Pharmaceuticals, GlaxoSmithKline, Otsuka, PharmaNeuroBoost, and Roche. He has equity holdings in MedAvante, Inc. and receives income from royalties from American Psychiatric Publishing, Inc., Guilford Publications, and Herald House. Dr Thase\u2019s wife is an employee of Embryon, Inc.Robert M. Berman, Zachary J. Cain, Ronald Marcus, and John Sheehan are employees of Bristol-Myers Squibb; Sabrina Vogel Marler was an employee of Bristol-Myers Squibb when these post hoc analyses were conducted. John Sheehan is a former employee of Janssen Pharmaceuticals, Inc.Ross A. Baker is an employee of Otsuka Pharmaceutical Development & Commercialization; Robert A. Forbes was an employee of Otsuka Pharmaceutical Development & Commercialization when these post hoc analyses were conducted.AHC, RAB, JJS, SVM and MET contributed to the conception and design of the analysis. MET was a study investigator and participated in data collection. RAB and JJS managed the analyses. SVM undertook the statistical analysis. All authors contributed to the analysis and interpretation of data, critically reviewed all drafts to provide important intellectual content, and made the decision to submit the manuscript. All authors have approved the final manuscript.John J. Sheehan, Sabrina Vogel Marler, Ronald Marcus, and Robert M. Berman were employees of Bristol-Myers Squibb when this study was performed.Treatment-emergent AEs occurring in \u226510% of patients in any ADT group (safety sample).Click here for file"}
+{"text": "Plasmodium falciparum transmission-blocking due to concerns over drug-related haemolysis risk, especially among glucose-6-phosphate dehydrogenase-deficient (G6PDd) people, without evidence demonstrating that it can be safely deployed in their settings. Pharmacovigilance methods provide a systematic way of collecting safety data and supporting the rollout of SLD PQ.Countries remain reluctant to adopt the 2012 World Health Organization recommendation for single low-dose (0.25\u00a0mg/kg) primaquine (SLD PQ) for The Primaquine Roll Out Monitoring Pharmacovigilance Tool (PROMPT), comprising: (1) a standardized form to support the surveillance of possible adverse events following SLD PQ treatment; (2) a patient information card to enhance awareness of known adverse drug reactions of SLD PQ use; and (3) a database compiling recorded information, was developed and piloted. Data on patient characteristics, malaria diagnosis and treatment are collected. Blood samples are taken to measure haemoglobin (Hb) and test for G6PD deficiency. Active follow-up includes a repeat Hb measurement and adverse event monitoring on or near day 7. A 13-month prospective pilot study in two hospital facilities in Swaziland alongside the introduction of SLD PQ generated preliminary evidence on the feasibility and acceptability of PROMPT.PROMPT was well received by nurses as a simple, pragmatic approach to active surveillance of SLD PQ safety data. Of the 102 patients enrolled and administered SLD PQ, none were G6PDd. 93 (91.2\u00a0%) returned on or near day 7 for follow-up. Four (4.6\u00a0%) patients had falls in Hb \u226525\u00a0% from baseline, none of whom presented with signs or symptoms of anaemia. No patient\u2019s Hb fell below 7\u00a0g/dL and none required a blood transfusion. Of the 11 (11\u00a0%) patients who reported an adverse event over the study period, three were considered serious and included two deaths and one hospitalization; none were causally related to SLD PQ. Four non-serious adverse events were considered definitely, probably, or possibly related to SLD PQ.Improved pharmacovigilance to monitor and promote the safety of the WHO recommendation is needed. The successful application of PROMPT demonstrates its potential as an important tool to rapidly generate locally acquired safety data and support pharmacovigilance in resource-limited settings.The online version of this article (doi:10.1186/s12936-016-1410-7) contains supplementary material, which is available to authorized users. Plasmodium falciparum gametocytes (stages I to III) and asexual blood stages of human malaria species [P. falciparum gametocytes. Primaquine (PQ) is currently the only licensed anti-malarial in this class of drugs.As malaria programmes move towards elimination, stopping onward transmission from all detected cases becomes increasingly important. The most commonly used anti-malarials are artemisinin derivatives, which destroy early developing  species . However species \u20134. The 8Plasmodium vivax and Plasmodium ovale species [P. falciparum [Since its approval by the FDA in 1952, PQ has been used as 14-day anti-relapse treatment against  species . In addilciparum . Howeverlciparum . The extlciparum \u201311.P. falciparum gametocytes. However, low uptake of this recommendation and concerns over the potential for haemolytic toxicity in G6PD-deficient (G6PDd) individuals prompted the issue of a new WHO directive, recommending the use of a lower 0.25\u00a0mg/kg dose of PQ in areas threatened by artemisinin resistance and in settings targeting malaria elimination. This 0.25\u00a0mg/kg dose should be given to all patients with parasitologically-confirmed P. falciparum malaria together with ACT, except for pregnant women and infants. Prior G6PD testing is not required [Previously, the World Health Organization (WHO) recommended the addition of a single 0.75\u00a0mg/kg dose of PQ to ACT for the clearance of required . Despiterequired . There rThe use of standardized, prospective pharmacovigilance methods for PQ provides an opportunity to generate data that can be used to confirm or refute safety concerns. To date, pharmacovigilance on PQ is weak and relies predominately on passive reporting. In most countries in sub-Saharan Africa, passive reporting systems, which lack an appropriate denominator of persons exposed to treatment, are limited or non-existent . From a P. falciparum malaria.To support the safe rollout of SLD PQ, the Primaquine Roll Out Monitoring Pharmacovigilance Tool (PROMPT) was developed in collaboration with the National Malaria Control Programme (NMCP) in Swaziland. The objective of this study was to assess the feasibility and perceived acceptability of using a simple pharmacovigilance tool to collect standardized safety data in those prescribed SLD PQ therapy for the treatment of This project involved two phases: (1) a tool development phase during which the components of PROMPT were developed and (2) a pilot phase, when intended end-users in Swaziland participated in testing PROMPT alongside the introduction of SLD PQ.Through an interactive process, involving a literature review and discussions with national and international experts, a tool was developed comprising three parts: (1) a standardized form (on either a paper or electronic platform) collecting a minimum set of essential data elements to support the surveillance of possible adverse events following SLD PQ treatment; (2) a patient information card to enhance awareness of known adverse drug reactions of SLD PQ use; and (3) a database compiling recorded information.Data collection begins once the prescriber has decided to treat confirmed, uncomplicated malaria cases with PQ (day 0), in addition to the standard blood schizonticide recommended by that particular country. The prescriber collects basic socio-demographic data , contact information, patient and family history data , malaria diagnosis information, and drug dosage and frequency data.For those involved with active surveillance, a blood sample is collected to measure haemoglobin or haematocrit and to screen for G6PD deficiency. Patients are asked to return for a scheduled follow-up visit on or near day 7 because drug-induced hemolysis and jaundice are clinically detectable within the first 7\u00a0days . At thisAt the time of prescribing PQ (day 0), patients are given a patient information card with oral and written instructions on how to identify the signs and symptoms of known adverse drug reactions of PQ  and to monitor the colour of their urine . A 25\u00a0% or greater reduction was regarded as the minimum clinically important threshold that would signify caution for implementing the addition of SLD PQ to ACT for the treatment of Plasmodium falciparum is the predominant parasite species accounting for over 99\u00a0% of malaria cases. One-third of Swaziland\u2019s population is at risk of infection, with residents in the Lubombo region near the eastern borders with Mozambique and South Africa at greatest risk.In response to a request from the Ministry of Health (MoH), PROMPT was first piloted in the Lubombo and Manzini regions of the Kingdom of Swaziland alongside the introduction of SLD PQ. Good Shepherd Mission Hospital, a rural, regional referral hospital in Lubombo, and Raleigh Fitkin Memorial Hospital, a hospital in Manzini run by the Catholic mission and government were both purposively selected by the NMCP to represent facilities with the highest burden of malaria cases. Lubombo and Manzini reported a total of 266 and 164 malaria cases, respectively, this past year (March 2014\u2013April 2015) . MalariaThe pilot was tested following discussions with key stakeholders, including the MoH, an experienced database developer, infectious disease specialists with pharmacovigilance expertise, and local health workers  over 6\u00a0months. Consultations with the database developer allowed us to build a user-friendly, electronic data collection tool configured to country capacity. Infectious disease and pharmacovigilance specialists offered guidance on how to standardize and define the data elements (and minimum reporting criteria) collected. Health workers reviewed the steps necessary to use PROMPT and increased awareness around its use. Observations were made at each facility and discussions were conducted with clinic staff to best integrate PROMPT into the existing clinical workflow and capture confirmed malaria cases.Nurses from both facilities received a two-day training workshop followed by 7\u00a0days of pre-pilot on-site training. Staff from the NMCP, health facility doctors and nurse matrons were also invited to the group training to enable additional support to trained health workers during study implementation. Half way through study implementation (August 2014) trained health workers received a 1-day refresher training. Both trainings focused on illustrating routine study procedures. The health workers were trained in data reporting procedures and electronic data collection. Each facility received job aids  to offer cues and direction on key procedures. Instruction on identifying the signs and symptoms of acute haemolytic anaemia, the main adverse effect of SLD PQ use, was also given. Training objectives also included opportunities for the study team to complete tool procedures in a pre-testing environment and to solicit feedback from the team on components of PROMPT needing further refinement.P. falciparum malaria were eligible to receive SLD PQ. Patients were not deemed suitable for SLD PQ treatment if they had any of the following: evidence of severe malaria [All patients over 1\u00a0year of age and prescribed standard doses of first-line artemether-lumefantrine (AL) for confirmed, uncomplicated  malaria ; history\u00ae, Micro Labs Ltd Hosur, Bangalore, India) using 7.5\u00a0mg PQ tablets, according to four weight bands established following discussions with country doctors and senior officials from the NMCP  and haemoglobin was measured using HaemoCue 201+ photometers . Patients were then treated with the WHO recommended single dose of PQ (0.25\u00a0mg/kg)  3G  tablets running Android OS to enable automated data collection in the health facilities  and reactions to and ease or challenges of study procedures were collected, using a semi-structured questionnaire, to investigate perceptions related to acceptability among trained nurses. Discussions  took about 60\u00a0min to complete. A moderator collected handwritten or electronic notes.To assess the safety of introducing SLD PQ, the haematologic response and occurrence of adverse events in those receiving SLD PQ therapy were investigated. Haemoglobin concentrations (defined in g/dL) were expressed as the absolute and relative change from baseline values (day 0). Anaemia was classified according to WHO guidelines  . Qualitative data from interview notes were summarized after familiarization of the data using an iterative approach. Emergent themes from the data were identified based on frequency of appearance and summarized.The study was approved by review committees at the Swaziland MoH (reference MH/599C/FWA 000 15267) and the University of California, San Francisco (IRB no. 13-11626), and granted non-engaged status by the United States Centers for Disease Control and Prevention. Participants or a parent or guardian for children less than 7\u00a0years of age gave written, informed consent; children between the ages of 7 and 17\u00a0years gave assent following national guidelines.Over a 13-month pilot period (March 2014\u2013April 2015), 166 confirmed, uncomplicated malaria cases from two hospital facilities in Lubombo (77) and Manzini (89) regions were reported through Swaziland\u2019s Health Management Information System and Immediate Disease Notification System. Of those reported, 112/166 (67.5\u00a0%) patients were seen by four nurses trained in the use of PROMPT and screened for inclusion. Ten patients were excluded: one tested positive for pregnancy, seven had haemoglobin concentrations less than 8\u00a0g/dL, and two declined to participate. A total of 102 patients (59 from Lubombo and 43 from Manzini) were enrolled and administered SLD PQ and 98/102 (96.0.\u00a0%) were followed up with 93/102 (91.2\u00a0%) successfully completing all follow-up procedures. This included obtaining a repeat haemoglobin measurement and collecting data on all adverse events experienced since taking SLD PQ  returning promptly on day 7 and 87/102  returning within 10\u00a0days (range: 4\u201310) post-treatment for a follow-up visit.All four trained nurses participated in informal feedback discussions to offer initial perceptions of PROMPT\u2019s acceptability and to suggest ways to improve upon the existing tool. The standardized data collection form using handheld electronic devices was popular. Nurses noted several benefits of using an electronic data entry system over paper for monitoring the safety of SLD PQ. All nurses agreed that this included the time saved on data entry and half of nurses felt that the electronic devices kept patient records confidential and accessible only by trained nurses and staff. There was also general agreement amongst the nurses that the average staff time required for screening and enrolment , and follow-up visits  per patient was acceptable. This period of time excludes the time taken for retrieving missing or additional information.These discussions also revealed their views on counselling sessions with patients on G6PD deficiency and known adverse drug reactions associated with SLD PQ use. All nurses agreed that the patient information cards were a useful component of PROMPT that facilitated the performance of study procedures and conveyed key messages. However, nurses agreed that the language used to communicate G6PD deficiency and adverse effects of SLD PQ use to patients was challenging to develop. In addition, nurses highlighted challenges with the adverse event reporting process, including challenges with identifying cases as relevant to report. Nurses more frequently reported on events that were either severe or life-threatening in nature or expected signs and symptoms of SLD PQ use .The main challenge described when using an electronic platform was server connectivity; while nurses were still able to enter information and store data while the server was offline, data could not be synchronized and uploaded to the country-level database. This limited capacity to monitor and analyse data in real-time and resulted at times in delayed or duplicated SMS text messages being sent to patients.P. falciparum malaria patients treated with SLD PQ who completed follow-up (n\u00a0=\u00a093), the mean age was 25\u00a0years, ranging from 13\u00a0months to 68\u00a0years  experiencing declines in haemoglobin concentration post-treatment. The mean absolute reduction in haemoglobin concentration within 10\u00a0days, relative to day 0 was 0.6\u00a0g/dL   and microscopy. He was admitted to the hospital as a precaution to monitor his diarrhoea. The following day he was enrolled and treated with AL plus SLD PQ, and panadeine. The diarrhoea continued throughout the day. He was confused. Diclofenac was added to his treatment. Two days after symptom onset, the patient died. Another patient presenting with complaints of weakness, jaundice, and coughing, was prescribed AL based on signs and symptoms, despite testing negative for malaria by RDT and microscopy. When the patient returned to the clinic to pick up his AL treatment (two\u00a0days after event onset), the patient was incorrectly enrolled into the study and treated with AL plus SLD PQ. Despite anti-malarial therapy, the patient\u2019s condition continued to deteriorate. Upon active follow-up by telephone, the study nurses learned of the patient\u2019s death. A verbal autopsy suggested that the patient\u2019s health had deteriorated to the point where the patient was unable to pass urine. He died within 10\u00a0days of initial presentation. One patient, with complaints of diarrhoea, vomiting, headache, and dizziness, was admitted and hospitalized one day after treatment with SLD PQ but recovered within 48\u00a0h of onset.In general, SLD PQ was well tolerated by patients. Out of all the patients who received SLD PQ and returned for follow-up, 89\u00a0% (87/98) of patients did not report any adverse event since receiving treatment. Of the 11 (11\u00a0%) patients who reported an event, three were considered serious using ICH guidelines and included two deaths and one hospitalization; none were classified as related to SLD PQ Table\u00a0. In all None of the patients vomited in the 30\u00a0min following drug administration. The most frequently reported events in all 11 patients were dark urine (36\u00a0%), vomiting (27\u00a0%), fever (18\u00a0%), and diarrhoea (18\u00a0%). Four non-serious adverse events were considered definitely, probably or possibly related to SLD PQ. Of the non-serious adverse events, all recovered except for one whose outcome remains unknown despite multiple telephone attempts at follow-up. This patient, following a diagnosis of vivax malaria confirmed by microscopy was referred to one of the study facilities and received the recommended higher dose of PQ (30\u00a0mg). Following treatment, the patient reported malaise and worsening of symptoms.Key findings from the safety assessment were disseminated and recommendations were made Fig.\u00a0 to key sP. falciparum malaria. Results suggest that PROMPT achieved its objective and successfully yielded standardized safety data on the range and frequency of adverse events, including haematologic changes, experienced after SLD PQ therapy. The data in turn contributed to a limited but growing local evidence base and offered the program insight prior to countrywide adoption of WHO policy. These findings are supported by the following observations.This study is the first to demonstrate the feasibility and acceptability of performing active safety monitoring using a simple pharmacovigilance tool in those prescribed SLD PQ therapy for the treatment of P. falciparum patients who presented with an adverse event recovered. Moreover, repeat haematologic testing ensured that clinically important drops in haemoglobin  were detected. No patient\u2019s haemoglobin fell below 7\u00a0g/dL and none required a blood transfusion.Firstly, PROMPT promoted and facilitated the capture of treatment emergent adverse events after initiation of SLD PQ therapy with a known denominator of the number of people exposed to the drug (n\u00a0=\u00a0102). Calculating adverse drug reaction frequencies or incidence rates to examine risk for particular treatments is limited with current systems that rely primarily on passive, spontaneous reporting . ImportaSecond, this study showed high adherence (>90\u00a0%) to visits on or near day 7 and demonstrated the acceptability of this approach among trained nurses. The use of simple approaches\u2014real-time electronic data collection, SMS text reminders, follow-up phone calls, and patient information cards\u2014encouraged patients to return to clinics and helped ensure the timely reporting of safety data to a central repository. This is in line with prior research illustrating how sensitization and training of health workers and consumers can be strong incentives for reporting and adherence to scheduled visits . FurtherThis study did, however, face challenges that are inherent to establishing pharmacovigilance systems with reliable reporting processes . UnderreThere are limitations to the study also worth noting. Sampling was restricted to two hospital facilities in a low transmission African setting in relatively healthy participants with haemoglobin concentrations of 8\u00a0g/dL or greater, and no enrolled patients were G6PDd, which may limit the generalizability of our findings. Furthermore, by not enrolling any G6PDd individuals, the targeted G6PDd sample size was not achieved. This could be due to a low prevalence of G6PD deficiency in this malaria-infected population or due to a multitude of factors that affect the performance of G6PD tests. Recent haemolytic events can leave patients with a large proportion of young red blood cells in circulation. These cells have high G6PD enzyme function and can obscure correct diagnosis and produce false normal results. In addition, borderline colourimetric results can produce subjective and variable readings . For theThis study also relied on the views of four trained nurses to produce opinions on tool acceptability thus restricting the breadth of opinions and experiences captured. Nonetheless, perceptions did demonstrate promise that PROMPT would be well received on a larger scale. Additional interviews among end users would allow us to capture a broader range of views in the future.Expanding PROMPT\u2019s use beyond Swaziland presents an opportunity to address these limitations and improve the systematic collection of safety data where data are needed. Data that include G6PDd participants are being collected from four additional countries across Asia and Africa  with differing levels of risk associated with PQ therapy. As data from these sites increases in size, it will be possible to pool individual patient data, directly compare, and determine the risks of PQ therapy with ever-greater accuracy. These new insights could serve to inform the highest safe dose of SLD PQ in G6PDd individuals that is critical to defining the therapeutic dose range and guiding appropriate dosing regimens .P. vivax or P. ovale. Women of childbearing age inadvertently exposed to PQ in very early, unsuspected pregnancy could also be identified at around 3\u00a0months and prospectively followed at regular intervals  to assess pregnancy and birth outcomes. Data on the use of PQ in pregnancy remains limited and further study on the safety of PQ in pregnancy are needed. Finally, these tools could be expanded beyond PQ to other drugs for a variety of other diseases.PROMPT could also extend to support pharmacovigilance activities under diverse contexts and in select populations. For example, subjects could be followed up at additional time points  to assess outcomes after PQ treatment of longer duration for P. falciparum treatment guidelines, albeit in different doses and schedules. And demand for its use is growing rapidly in parts of Africa with Botswana, Ethiopia, Namibia, South Africa, Swaziland, Zanzibar, and Zimbabwe who have written SLD PQ into national strategic plans or treatment guidelines for falciparum malaria [As access to SLD PQ increases, a larger evidence-base to support the safety of the latest WHO recommendation will be needed. Several countries in the Asia Pacific, including Bhutan, Indonesia, Malaysia, Myanmar, Thailand, the Philippines, and Vietnam include SLD PQ in their  malaria . Althoug malaria , no medi"}
+{"text": "Gametocytes are responsible for transmission of malaria from human to mosquito. Artemisinin combination therapy (ACT) reduces post-treatment gametocyte carriage, dependent upon host, parasite and pharmacodynamic factors. The gametocytocidal properties of antimalarial drugs are important for malaria elimination efforts. An individual patient clinical data meta-analysis was undertaken to identify the determinants of gametocyte carriage and the comparative effects of four ACTs: artemether-lumefantrine (AL), artesunate/amodiaquine (AS-AQ), artesunate/mefloquine (AS-MQ), and dihydroartemisinin-piperaquine (DP).Factors associated with gametocytaemia prior to, and following, ACT treatment were identified in multivariable logistic or Cox regression analysis with random effects. All relevant studies were identified through a systematic review of PubMed. Risk of bias was evaluated based on study design, methodology, and missing data.P\u2009=\u20090.005 compared to AL) and AS-AQ fixed dose combination (FDC) . Among individuals who had gametocytaemia before treatment, gametocytaemia clearance was significantly faster with AS-MQ  and slower with DP  compared to AL. Both recrudescent (adjusted odds ratio (AOR), 9.05; 95 % CI, 3.74\u201321.90; P\u2009<\u20090.001) and new  infections with asexual-stage parasites were strongly associated with development of gametocytaemia after day 7.The systematic review identified 169 published and 9 unpublished studies, 126 of which were shared with the WorldWide Antimalarial Resistance Network (WWARN) and 121 trials including 48,840 patients were included in the analysis. Prevalence of gametocytaemia by microscopy at enrolment was 12.1 % , and increased with decreasing age, decreasing asexual parasite density and decreasing haemoglobin concentration, and was higher in patients without fever at presentation. After ACT treatment, gametocytaemia appeared in 1.9 %  of patients. The appearance of gametocytaemia was lowest after AS-MQ and AL and significantly higher after DP (adjusted hazard ratio (AHR), 2.03; 95 % CI, 1.24\u20133.12; AS-MQ and AL are more effective than DP and AS-AQ FDC in preventing gametocytaemia shortly after treatment, suggesting that the non-artemisinin partner drug or the timing of artemisinin dosing are important determinants of post-treatment gametocyte dynamics.The online version of this article (doi:10.1186/s12916-016-0621-7) contains supplementary material, which is available to authorized users. Plasmodium falciparum gametocytaemia has been associated with asexual parasite densities, the duration of malaria symptoms, anaemia and immunity [Malaria remains a leading cause of morbidity and mortality in endemic countries, with an estimated 584,000 deaths and 198 million clinical cases of malaria globally in 2013 [immunity . As a coimmunity . Upon inimmunity  and hostACT is now recommended universally for the treatment of uncomplicated falciparum malaria. Artemisinins are highly effective against the pathogenic asexual parasite stages  and immaBecause gametocytes are only detected in a fraction of patients by microscopy, individual trials are often insufficiently powered to compare gametocytocidal properties between ACTs or disentangle host and parasite factors that influence gametocyte dynamics. To address this, a pooled analysis of individual-level patient data was undertaken in patients before and after treatment with artemether-lumefantrine (AL), artesunate-amodiaquine (AS-AQ), artesunate-mefloquine (AS-MQ), and dihydroartemisinin-piperaquine (DP).P. falciparum malaria ; (2) asexual parasite quantification at enrolment; (3) gametocyte quantification or prevalence at enrolment; (4) well described methodology for quantifying asexual parasites and gametocytes; and (5) haemoglobin (or haematocrit) estimation at enrolment.A search was conducted in PubMed in September 2014 to identify all antimalarial clinical trials published between 1990 and 2014, in which gametocytes were recorded using the search strategy described in the legend of Additional file Individual study protocols were available for all trials included, either from the publication or as a metafile submitted with the raw data. Individual patient data from eligible studies were shared, collated and standardised using a previously described methodology , 20. StuStatistical analyses were carried out using STATA (Version 13.1) according to an a priori Statistical Analysis Plan . BrieflyGametocyte carriage at any time after treatment in patients with no recurrent parasitaemia, patients with recrudescent infections and patients with reinfections were compared using multilevel logistic regression models with random effects for study site and subject.Methods to detect gametocytes by microscopy differed between trials. The sensitivity of microscopy methods was included in the analyses, by classifying studies into one of four categories, as follows: (1) studies in which slides were specifically read for gametocytes, reviewing at least 100 microscopic high power fields or against\u2009\u2265\u20091000 white blood cells (WBC) (4 studies); (2) microscopists specifically instructed to record gametocytes but slides were primarily read for asexual parasites; \u2265 100 microscopic high power fields per\u2009\u2265\u20091000 WBC were read (26 studies); (3) microscopists were specifically instructed to record gametocytes; 50\u201399 microscopic high power fields per 500\u2013999 WBC were read (33 studies); (4) microscopists were not specifically instructed to record gametocytes or the number of examined high power fields was\u2009<\u200950 or the number of WBC was\u2009<\u2009500 (40 studies). For 18 studies, the information on the sensitivity of the microscopy was not available.Risk of bias within studies was assessed based on (1) study design ; (2) methodology for gametocyte detection; and (3) the number and proportion of patients with (a) missing outcomes and (b) missing baseline covariates . For the final models, two sets of sensitivity analyses were performed. Firstly, a model was refitted with each study\u2019s data excluded, one at a time, and a coefficient of variation around the parameter estimates calculated. This would identify any influential studies, that is, studies with unusual results  that affect the overall pooled analysis findings. Secondly, for the outcome measure time to gametocytaemia, the impact of incomplete gametocyte carriage data was investigated by refitting the final multivariable model in a subset of patients with complete weekly data for 28 days.All data included in this analysis were obtained in accordance with the laws and ethical approvals applicable to the countries in which the studies were conducted, and were obtained with the knowledge and consent of the individual to which they relate. Data were fully anonymised either before or during the process of uploading to the WWARN repository. Ethical approval to conduct individual participant data pooled analyses was granted to WWARN by the Oxford Tropical Research Ethics Committee (OXTREC-48-09).P. falciparum gametocytes at enrolment or during follow-up. Investigators of 117 clinical trials  agreed to contribute their data. In addition, nine unpublished studies  were shared, one of which was published subsequently. After exclusion of duplicate studies, studies in returning travellers, multiple infection episodes and participants with protocol violations, 48,840 study participants from 121 individual clinical trials were retained  or Asia 13,546; 27.7 %) with a minority coming from South America  , and was not significantly influenced by the slide reading method. In Africa, fractional polynomial analysis indicated a gradual decline in the proportion of gametocyte-positive smears with increasing age Fig.\u00a0; Fig.\u00a02.In Asia, there was a gradual decline in prevalence of gametocytaemia with increasing asexual parasite density across the entire range of asexual parasite densities that were observed Fig.\u00a0. In AfriP\u2009<\u20090.001) and Asian  patients. Male gender was a predictor of prevalence of gametocytaemia at enrolment in studies in Asia  and South America  but not Africa (Table\u00a0P\u2009<\u20090.001) and in Asia  but this was not significant in the multivariable analysis  were less likely to present with gametocytaemia and this remained significant after adjusting for covariates in both African (adjusted odds ratio (AOR), 0.63; 95 % CI, 0.58\u20130.69; P\u2009=\u20090.005 compared to AL) and AS-AQ   but not after treatment with any of the other ACTs  Fig.\u00a0. This prAL) Fig.\u00a0, Table\u00a04P\u2009<\u20090.001 for log increase in gametocyte density) and the type of ACT given  and slower with DP  , and non-significantly slower compared to AL . The overall observed proportion of patients who cleared gametocytes by day 7 was 64.4 % for AL, 61.7 % for AS-MQ, 52.3 % for DP, and 47.8 % for AS-AQ, while by day 14 gametocytes were cleared by 85.7 %, 90.2 %, 70.3 %, and 72.1 % of patients, respectively.A total of 2433 patients treated with an ACT were gametocytaemic at enrolment and had no recurrent infection. Overall, 57.4 %  of these patients cleared gametocytaemia by day 7, 78.4 %  by day 14 and 88.2 %  by day 21. The only independent determinants of gametocyte clearance were initial gametocyte density  Fig.\u00a0. For theP\u2009=\u20090.025) and develop gametocytaemia after day 7  compared to patients with no recorded recurrence and at least 28 days follow-up. Similarly, the increased risk of gametocytaemia on any day during follow-up  and of developing gametocytaemia after day 7  was observed in individuals with reinfection . Gametocyte densities commonly fluctuate around the microscopic threshold for detection and the use of molecular gametocyte detection tools would have uncovered a higher proportion of gametocyte carriers [Whilst our analysis focuses on peripheral gametocytaemia, it is important to acknowledge that this is a surrogate marker of malaria transmission potential. The infectivity of persisting or appearing gametocytes may be affected by the type of antimalarial treatment . Antimalcarriers . The addcarriers  and prevcarriers , 47 but carriers , 48, thicarriers , 7.Our analysis was purposefully restricted to microscopic findings on gametocytaemia, for which most data are available. Although this approach will have missed some gametocyte carriers, this would not affect the comparison of treatment arms. Studies where microscopy, molecular gametocyte data and infectivity results are available indicate that these methods lead to the same conclusions on the comparative effects of antimalarials on post-treatment gametocyte dynamics and infectivity , 18.In conclusion, we identified independent risk factors for the prevalence of gametocytaemia in patients with uncomplicated falciparum malaria in studies conducted on three continents. AS-MQ and AL are superior ACT options in preventing gametocytes shortly after treatment compared to DP or AS-AQ. We hypothesize that this difference is due to the non-artemisinin partner drug defining post-treatment gametocyte dynamics."}
+{"text": "Sickle cell disease (SCD) is a genetic disorder common in malaria endemic areas. In endemic areas, malaria is a major cause of morbidity and mortality among SCD patients. This suggests the need for prompt initiation of efficacious anti-malarial therapy in SCD patients with acute malaria. However, there is no information to date, on the efficacy or safety of artemisinin combination therapy when used for malaria treatment in SCD patients.Children with SCD and acute uncomplicated malaria (n\u2009=\u200960) were randomized to treatment with artesunate-amodiaquine (AA), or artemether-lumefantrine (AL). A comparison group of non-SCD children  with uncomplicated malaria were also randomized to treatment with AA or AL. Recruited children were followed up and selected investigations were done on days 1, 2, 3, 7, 14, 28, 35, and 42. Selected clinical and laboratory parameters of the SCD patients were also compared with a group of malaria-negative SCD children (n\u2009=\u200982) in steady state.The parasite densities on admission were significantly lower in the SCD group, compared with the non-SCD group (p\u2009=\u20090.0006). The parasite reduction ratio (PRR) was lower, clearance was slower (p\u2009<\u20090.0001), and time for initial parasitaemia to decline by 50 and 90% were longer for the SCD group. Adequate clinical and parasitological response (ACPR) on day 28 was 98.3% (58/59) in the SCD group and 100% (57/57) in the non-SCD group. Corresponding ACPR rates on day 42 were 96.5% (55/57) in the SCD group and 96.4% (53/55) in the non-SCD group. The fractional changes in haemoglobin, platelets and white blood cell counts between baseline (day 0) and endpoint (day 42) were 16.9, 40.6 and 92.3%, respectively, for the SCD group, and, 12.3, 48.8 and 7.5%, respectively, for the non-SCD group. There were no differences in these indices between AA- and AL-treated subjects.The parasite clearance of SCD children with uncomplicated malaria was slower compared with non-SCD children. AA and AL showed similar clinical and parasitological effects in the SCD and non-SCD groups. The alterations in WBC and platelet counts may have implications for SCD severity.ISRCTN96891086.Current controlled trials  S globin gene (SS) or in individuals heterozygous for the \u03b2S allele and other abnormal \u03b2 globin gene alleles such as \u03b2C (SC), S \u03b20 thalassemia, or S\u03b2+ thalassemia. SCD is associated with significant morbidity and mortality and is characterized clinically among others, by chronic haemolysis, vaso-occlusive events and increased susceptibility to infections. The highest frequencies of homozygous SCD are found in sub-Saharan Africa, where intense malaria transmission occurs.Sickle cell disease (SCD) is the most common monogenic disease worldwide. SCD is an autosomal recessive condition that results from substitution of glutamic acid with valine in the sixth position of the beta-globin chain of haemoglobin. SCD occurs in individuals homozygous for the \u03b2In endemic areas, malaria is considered a major cause of the morbidity and mortality that SCD subjects experience. A linkage exists between the presence of sickle haemoglobin (HbS) and protection from malaria in the heterozygous state \u20133; howevin vitro[It is possible that response to anti-malarial therapy may differ between SCD and non-SCD subjects undergoing anti-malarial treatment. Indeed, plasma chloroquine concentrations have been shown to be 2-3 fold lower in subjects with SCD (HbSS) compared to those with HbAA or HbAS, and chloroquine accumulation has been shown to differ between SCD and non-SCD erythrocytes in vitro. Platelein vitro, suggestin vitro\u201319, coulIn the present study, the efficacy of artesunate-amodiaquine (AA) and artemether-lumefantrine (AL), the two most commonly adopted ACT regimens across sub-Saharan Africa, were evaluated among SCD children with parasitologically confirmed acute uncomplicated malaria in Accra, Ghana. Selected haematological parameters in treated SCD children were compared with those of non-SCD children (HbAA genotype), as well as with comparable haematological parameters of a group of apparently well, microscopically and malaria rapid diagnostic test (RDT)-negative SCD children in steady state.The study was conducted at the Outpatients Department (OPD) and Paediatric Sickle Cell Clinic of the Department of Child Health (DCH), Korle Bu Teaching Hospital (KBTH), between January 2010 and December 2011. The KBTH is a 1,600-bed tertiary referral hospital in Accra, Ghana. Malaria transmission occurs all year in Ghana, peaking in July-August immediately after the major rainfall season. The prevalence of malaria parasitaemia in Accra ranges between 6 and 22% (average 14.8%) depending on the community  and an SThe DCH is a referral centre within KBTH for children aged less than 13\u00a0years with acute or chronic medical and surgical conditions. The OPD is the first point of call for referred cases to the DCH. The OPD has an average yearly attendance of 36,000, and is divided into an acute care outpatients and specialty clinics. The Paediatric Sickle Cell Clinic is one of the specialty clinics at the DCH and has about 5,000 SCD patients below 13\u00a0years of age registered. The Paediatric Sickle Cell Clinic provides routine and follow-up care for children with SCD. The Clinic is held once a week and has an average weekly attendance of about 60-70. Aside from the Clinic, SCD children with acute illness are seen at the Paediatric Emergency Department 24\u00a0hours a day throughout the week.Children of known SCD status registered at the Paediatric Sickle Cell Clinic, who presented to the Paediatric Emergency Department, or the Paediatric Sickle Cell Clinic with an acute febrile illness were referred to the study team. A study physician examined the patient and recruited subjects if study criteria were fulfilled. At the adjoining primary care facility (Korle Bu Polyclinic) children not known to have SCD presenting with an acute febrile illness were also referred to the study team and recruited if study criteria were fulfilled. A group of known SCD children visiting the Paediatric Sickle Cell Clinic for routine scheduled follow-up visits, who were determined to be in steady state on the day of visit (n\u2009=\u200982), were also recruited for the purpose of comparing selected haematological indices between malaria-positive and malaria-negative SCD children.Plasmodium falciparum infection of parasite density <200,000/\u03bcL; and, willingness of the accompanying parent/guardian to comply with the study procedures and follow-up schedule. Exclusion criteria were: symptoms or signs of severe malaria; known intolerance or allergy to study medications; and, reported treatment with any of the study drugs one month preceding enrolment. The inclusion and exclusion criteria for the non-SCD (HbAA) children were the same as for the SCD children apart from haemoglobin genotype. The SCD children in steady state were children with confirmed SCD status registered at the Paediatric Sickle Cell Clinic who were asymptomatic and were visiting the Clinic for routine scheduled evaluations. These children had a negative blood film and negative malaria RDT result on the day of recruitment. A standardized evaluation was done for assessing and recording presenting symptoms and clinical signs and for recording initial laboratory investigations.Inclusion criteria for the study were: child of confirmed SCD status aged six months to 12\u00a0years with an acute febrile illness ; A computer-generated simple randomization scheme was prepared in advance. Allocated treatments were kept in sealed, opaque envelopes which were opened after completion of all formal enrolment procedures and were then administered.\u00ae, Sanofi Aventis, France; 25/50/100\u00a0mg artesunate and 67.5/135/270\u00a0mg amodiaquine), single daily dose, was administered for three days according to body weight: \u22654.5-\u2009<\u20099\u00a0kg (25\u00a0mg/67.5\u00a0mg), one tablet/dose; \u22659-\u2009<\u200918\u00a0kg (50\u00a0mg/135\u00a0mg), one tablet/dose; \u226518-\u2009<\u200936\u00a0kg (100\u00a0mg/270\u00a0mg), one tablet/dose; \u226536\u00a0kg (100\u00a0mg/270\u00a0mg), two tablets/dose. Artemether-lumefantrine  was administered at zero and eight hours on the first day, and then twice daily for two subsequent days according to body weight: 5-14\u00a0kg, one tablet/dose; 15-24\u00a0kg, two tablets/dose; 25-34\u00a0kg, three tablets/dose; 35\u00a0kg and over, four tablets/dose. All drug doses were administered by study nurses at the clinic. At the clinic, children were observed for one hour after each drug administration. Treatments were re-administered if the child vomited within the observation period. Children who vomited the re-administered dose were withdrawn.Artesunate-amodiaquine , physical examination, and laboratory investigations, described below.\u00ae, Premier Medical Corporation Ltd, India) and parasite lactic dehydrogenase  were done according to the manufacturers\u2019 instructions. The total WBC and differential counts were measured by means of an automated haematology analyzer . Clinical chemistry parameters were measured, using a chemistry analyzer . A nested PCR method was used to analyse polymorphisms in the genes coding for the merozoite surface protein (MSP) 1 and 2 to distinguish between recrudescent and new infections [Venous blood was collected into EDTA and heparinized sample tubes on days 0, 3, 7, 14, 28, and 42. On follow-up days 1 and 2, only a finger prick sample for a blood smear for malaria microscopy was done. Malaria parasitaemia (expressed as parasites per microlitre of whole blood) was determined in Giemsa-stained thick blood films. Parasite density was determined by counting the number of asexual stage parasites relative to 200 white blood cells (WBC), and multiplied by the measured WBC count. Each slide was read independently by two microscopists and the average of two readings recorded. Approximately 10% of all slides were read by a third expert microscopist who was independent of the study. Any discrepancies in the readings were checked with all three microscopists present and a consensus reached. RDT for the histidine-rich protein 2 , and by 90% (PC90).The primary endpoint was the parasite clearance rate on days 1, 2 and 3 of treatment, determined by the parasite reduction ratios (PRR), and the time for initial parasitaemia to decline by 50%  overall cure rates on day 28 or 42) after treatment with either AA or AL \u201328. Thes or 42 afOther outcome measures included parasitaemia recurrence over the follow-up period after initial clearance and changes in selected haematological profiles in the respective groups during follow-up. In the context of SCD, the extent of changes in parameters such as haemoglobin, leucocytes, neutrophils, and platelets could have important ramifications for disease-related morbidity. Other secondary outcome measures included day 28 and 42 cure rates, classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), or adequate clinical and parasitological response (ACPR), according to the WHO 2005 guidelines .Adverse events were defined as any untoward medical occurrence, irrespective of its suspected relationship to study medications according to International Conference on Harmonization (ICH) guidelines.50; PC90) between the SCD and non-SCD groups. A sample size of 27 per group was sufficient to detect, at 90% power and 95% confidence, a 30% difference in parasite clearance, or at least a two-fold change in PRR between the SCD and non-SCD groups on day 2. The PRR was defined as the ratio of the parasite count before treatment to the parasite count on days 1, 2 or 3. Time to 50 and 90% parasite clearance were obtained from the intercept on a plot of parasitaemia versus time. A random-effect generalized least square regression method was used to compare parasite densities between the SCD and non-SCD groups on successive days. The parasite clearance rates on the specific days were also compared, using survival analysis. Categorical variables were compared, using the Chi square test with Yates\u2019s correction, or the Fisher\u2019s exact test as appropriate. Continuous variables were compared, using the student\u2019s t-test, or one-way analysis of variance, or the Kruskall-Wallis test, as appropriate. For the survival analysis, subjects who did not complete 42\u00a0days of follow-up were censored on their last follow-up date. Probability values <0.05 were considered significant. Data were analysed using Stata\u2122 (Version 10).The study was designed to detect a difference in parasite clearance and fractional reduction in parasite count , or AL (n\u2009=\u200932). The SCD genotype distributions among the recruited subjects were: SS, n\u2009=\u200943 (71.66%); SC, n\u2009=\u200911 (18.33%); SS/SD, n\u2009=\u20095 (8.3%); and, S\u03b2thal, n\u2009=\u20091 (1.6%). A total of 243 non-SCD (HbAA) children presenting with an acute febrile illness were screened at the Korle Bu Polyclinic for inclusion, out of which 59 with confirmed P. falciparum uncomplicated malaria were randomized to treatment with AA (n\u2009=\u200931) or AL (n\u2009=\u200928). The flow of subjects through the study is shown  were not statistically significant. Selected characteristics of the SCD-steady state subjects were significantly different from that of the malaria-positive SCD patients  groups Table\u00a0. Within The predominant presenting symptom for both groups was a history of fever. However, the proportion of SCD subjects presenting with elevated temperature at admission was comparatively low. The remaining presenting symptoms among the SCD subjects were mostly vaso-occlusive complex-related Table\u00a0. The repThe parasite clearance of the SCD subjects appeared slower, compared with the non-SCD group Figure\u00a0. A randoWithin the SCD group, the parasite clearance rate per day was 0.58  for the AA-treated group, compared with 0.50,  for the AL-treated group . Similar results were obtained, using survival analysis  in the SCD group, and 100% (57/57) in the non-SCD group, respectively. The corresponding ACPR rates on day 42 were 96.5% (55/57) in the SCD group and 96.4% (53/55) in the HbAA group. The PCR-corrected ACPR rates on day 42 were 98.2% (56/57) in the SCD group and 98.2% (54/55) in the non-SCD group  was 96.4% (27/28) on day 28 and 92.6% (25/27) on day 42. The corresponding ACPR rates in the non-SCD group were 100% (30/30) on day 28 and 100% (29/29) day 42. The proportion of AL-treated subjects with ACPR in the SCD group was 100% (31/31) on day 28 and 100% (30/30) on days 28 and 42, respectively. The corresponding ACPR rates in the non-SCD group were 100% (27/27) on day 28 and 92.3% (24/26) on day 42, respectively. Selected characteristics of children with recurrent parasitaemia during follow-up are shown -malaria-positive subjects on admission and all scheduled follow-up days Figure\u00a0. Among tversus 7.5%, (non-SCD); platelets  versus 48.8%, (non-SCD); and, haemoglobin , respectively.The fractional change differences in the selected haematological indices  between admission  and endpoint  in the SCD and non-SCD groups were WBC  The individual plasma DEAQ concentrations in the two AA-treated subjects who experienced recurrent parasitaemia were 20\u00a0ng/ml and 21\u00a0ng/ml, respectively.Adverse events emerging during and after treatment overlapped substantially with known malaria or SCD symptomatology, could not be attributed to administered treatments, and were in most cases adjudged as unrelated to study medications.This study reports treatment response of paediatric SCD patients with acute uncomplicated malaria to recommended first-line ACT regimens. In particular, the study reports for the first time systematically collected data on parasite clearance and on selected haematological parameters  in SCD children undergoing ACT treatment.The data showed significantly lower parasite densities on admission, slower clearance dynamics, lower PRR values, and longer time to 50 and 90% parasite clearance in the SCD, compared with the non-SCD children. The significantly lower parasite densities among SCD patients compared with non-SCD patients has been reported in previous studies , 30, 31.versus non-SCD group data on one hand, and the comparable clearance rates between the two groups from the Cox regression analysis may reflect a convergence effect due to the significant initial parasitaemia differences between the two groups. The several-fold difference in admission parasite density between the two groups makes direct comparison problematic. However, the lower PRR and clearance rate in the SCD group in spite of a lower initial parasitaemia suggests a true between-group difference. The cure rates on day 28 or 42 were high for both the SCD and non-SCD groups. This is consistent with the high efficacy rates for AA and AL that have been reported across sub-Saharan Africa [The lower PRR and longer time to 50 and 90% parasite clearance among the SCD-malaria patients compared with non-SCD malaria patients has not been previously reported. This is partly because there have been no previous studies reporting parasite clearance changes among SCD patients undergoing anti-malarial treatment. The apparent disparity between the slower clearance, lower PRR\u2019s and time to 50 and 90% clearance of the SCD n Africa \u201328, 40. n Africa \u201343. It iin vivo day 3 DEAQ concentration of 135\u00a0ng/ml is the minimum level required for adequate treatment efficacy [The individual plasma drug concentrations of DEAQ  in the two AA-treated subjects with recurrent parasitaemia were lower than the mean plasma concentrations  , 45 thatThe lower fever incidence and higher WBC counts in the SCD group at presentation may present specific difficulties, especially with respect to diagnosis of diseases of bacterial origin. However, a raised WBC in SCD is also a recognized risk factor for complications ranging from acute chest syndrome, silent cerebral infarction, clinically overt stroke, and early death \u201352. In tThe majority of SCD subjects presented on admission with symptoms and signs consistent with vaso-occlusive complex (VOC) -related phenomena. Vaso-occlusion crisis is a hallmark of SCD but the mechanism(s) responsible for triggering VOC in SCD is likely a result of complex interplay of precipitating factors at multiple stages. However, it has been shown that SCD subjects in VOC have higher levels of pro-inflammatory cytokines compared with those in steady state . Since mIt is possible that the observed PRR differences between the SCD and non-SCD patients reflect intrinsic differences in peripheral blood components and specific variations in cell types between SCD and non-SCD subjects. It is usually assumed that malaria parasites and leucocytes are both distributed evenly on a thick smear; however, it has been shown that in cases of scanty parasitaemia, parasites tend to aggregate in a specific area of a blood smear . This suThe observed profiles for the selected haematological indices for the non-SCD subjects , are consistent with those reported for African children with uncomplicated malaria , 57. TheIn SCD subjects, elevated platelet counts have been ascribed among others, to absent splenic pooling or functional asplenia . As a reTreatment of children with uncomplicated malaria showed lower PRR and slower parasite clearance in the SCD subjects compared with non-SCD subjects. However, the comparably high day 28 and 42 cure rates in both SCD and non-SCD groups suggest an unlikely effect of this finding on acute malaria management in SCD patients. There were marked variations in leucocyte and platelet counts between acute illness and recovery stages among the SCD patients with malaria. The overall implications of these changes with respect to malaria-related morbidity among SCD patients are uncertain."}
+{"text": "Plasmodium vivax malaria in southern Ethiopia.Chloroquine (CQ) is the first-line treatment for vivax malaria in Ethiopia, but there is evidence for its declining efficacy. Defining the extent and regional distribution of CQ resistance is critical to ensure optimal treatment guidelines. This study aimed to provide data on the therapeutic efficacy of CQ against P. vivax mono-infection aged between 8\u00a0months and 65\u00a0years were enrolled in a clinical efficacy trial. The study was conducted at four sites in southern Ethiopia. Study participants were treated with a supervised course of CQ (25\u00a0mg/kg over three consecutive days), followed by weekly blood film examination and clinical assessment for 28\u00a0days. CQ blood concentrations were not assessed. The primary endpoint was the risk of failure at 28\u00a0days by survival analysis.Patients with Plasmodiumfalciparum parasitaemia and 29 lost to follow up). Two (0.7\u00a0%) patients experienced early treatment failure and 23 (8\u00a0%) late treatment failure. The overall risk of recurrence by day 28 was 9.4\u00a0% (95\u00a0% CI 6.4\u201313.6\u00a0%) with site-specific estimates of 3.8\u00a0% (95\u00a0% CI 1.2\u201311.3) for Shele, 21.9\u00a0% (95\u00a0% CI 12.2\u201336.1) for Guba, 5.9\u00a0% (95\u00a0% CI 1.9\u201317.3) for Batu and 9.2\u00a0% (95\u00a0% CI 4.5\u201317.6) for Shone.Between May 2010 and December 2013, 288 patients were enrolled in the study . Baseline characteristics varied significantly between sites. In total 34 (11.8\u00a0%) patients were censored during follow up (five with There is evidence of reduced CQ efficacy across three of the four study sites, with the degree of resistance severe enough in Guba to suggest that review of treatment policy may be warranted. Plasmodium vivax infection worldwide , and this remained significant after controlling for baseline parasitaemia. There was no difference in the total dose per kilogram administered in patients who failed treatment and those who were treated successfully. There were a total of 18 (6.3\u00a0%) patients who vomited any dose of CQ. From those, only one (5.5\u00a0%) had a late treatment failure compared to 8.1\u00a0% (22/270) in those who did not vomit (p\u00a0=\u00a00.69).Although the time to recurrence was not correlated significantly with baseline parasitaemia, patients failing treatment had a significantly higher baseline parasitaemia (geometric mean 5434\u00a0\u00b5lThe mean haemoglobin concentration at baseline was 12.5\u00a0g/dL (95\u00a0% CI 12.3\u201312.78) and 12.5\u00a0g/dL (95\u00a0% CI 12.3\u201312.8) at day 28, the risk of anaemia (Hb\u00a0<\u00a010\u00a0g/dL) being 18.5\u00a0% (51/175) and 16.4\u00a0% (35/214) at baseline and at day 28 respectively. Haemoglobin was significantly correlated with baseline parasite density , and also with age . The risk of anaemia varied significantly between sites being greatest in Shele (49.4\u00a0%) ; although the greater the degree of resistance the earlier the recurrence is likely to occur. Whereas genotyping of polymorphic loci is undertaken routinely in P. falciparum drug efficacy trials to distinguish new infections from true parasite recrudescence [P. vivax the interpretation of molecular typing is less informative. Heterologous paired isolates from pre- and post-treatment help to rule out true recrudescence, however homologous infection can be due to either recrudescence or relapse [Recurrent descence , in P. v relapse . ImportaP. falciparum clinical trials [P. vivax have demonstrated a similar relationship [There was noted heterogeneity in the risk of recurrence between and within sites. Treatment failure was higher in patients who were younger, and had a higher parasitaemia at enrollment. Both of these factors are well recognized as important risk factors for failure in l trials , 25. Higtionship  suggestitionship . In the tionship .Overall this study highlights evidence of clinical treatment failure after CQ most likely due to CQ resistance emerging in three out of four sites in southern Ethiopia. The data supports calls for increased drug resistance monitoring and re-evaluation of treatment guidelines."}
+{"text": "Plasmodium infections were common at baseline, with 41\u00b71% of children positive for Plasmodium falciparum/Plasmodium malariae, 9\u00b72% for P. falciparum/ Plasmodium ovale spp. and 8\u00b70% for all three species. Moreover, on day 17, 39\u00b79% of children infected with falciparum malaria at baseline were again positive for the same species, and 9\u00b72% of those infected with P. malariae at baseline were positive for P. malariae. Here, chronic multi-species malaria infections persisted in children after AL treatment(s). Better point-of-care diagnostics for non-falciparum infections are needed, as well as further investigation of AL performance in asymptomatic individuals.During a longitudinal study investigating the dynamics of malaria in Ugandan lakeshore communities, a consistently high malaria prevalence was observed in young children despite regular treatment. To explore the short-term performance of artemether-lumefantrine (AL), a pilot investigation into parasite carriage after treatment(s) was conducted in Bukoba village. A total of 163 children (aged 2\u20137 years) with a positive blood film and rapid antigen test were treated with AL; only 8\u00b77% of these had elevated axillary temperatures. On day 7 and then on day 17, 40 children (26\u00b73%) and 33 (22\u00b73%) were positive by microscopy, respectively. Real-time PCR analysis demonstrated that multi-species  Challenges to effective malaria control in this country include an extremely high transmission intensity with entomological inoculation rates greater than 100 per year in many areas, a weak health system with a fragile supply chain for front-line antimalarial treatments, inadequate case management with poor diagnostic tools in rural health centres and ineffective monitoring and evaluation of control interventions  and lactate dehydrogenase , display marked differences in their sensitivity, stability and dynamics of clearance from the blood after antimalarial treatment  promotes a \u2018test-and-treat\u2019 strategy for malaria involving parasitological diagnosis rather than presumptive treatment based on clinical indicators WHO, a. The goP. falciparum remains the major cause of morbidity and mortality in sub-Saharan Africa, non-falciparum infections, such as Plasmodium vivax, Plasmodium malariae and Plasmodium ovale spp., are also present in humans and their clinical significance should not be overlooked   or First Response RDTs , OptiMAL RDTs  and microscopy on Giemsa-stained blood films as described  community-based longitudinal study was conducted in six villages on the shores of Lake Albert and Lake Victoria in Uganda, commencing in May 2009  within the SIMI cohort (N\u00a0=\u00a0188) were recruited to the study on the basis of a positive First Response RDT result and microscopy-confirmed malaria. Children were subsequently followed up 7 and 17 days after day 0 (day of initiation of treatment). Due to financial and logistical constraints, only individuals who were RDT-positive on day 17 (see below), who had been given oral quinine sulphate as an alternative exit treatment, were followed up on day 24. A blood smear archive was made on days 0, 7 and 17, and confirmatory microscopy was carried out 2 or 3 days after each sample was taken . In addition, haemoglobin (Hb) levels were recorded for each child using a HemoCue spectrometer  on days 0, 7 and 17.Bukoba village, Mayuge District, showed the highest prevalence of falciparum and non-falciparum malaria in young children see  and so wP. falciparum infections  and albendazole (400\u00a0mg) according to WHO guidelines  and glutamate-rich protein (glurp) was carried out according to published methods  and R v2.10.1 . Anaemia was categorized based on haemoglobin levels as follows: mild 10\u201311\u00a0g\u00a0dLL\u22121 WHO, . To deteL\u22121 WHO, .Ethical approval was provided by The London School of Hygiene and Tropical Medicine and the Ugandan National Council of Science and Technology. Informed consent was given by mothers on behalf of their children and documented in writing or by thumbprint (in cases of illiteracy).S. mansoni infection was very low, 9\u00b70% of children had egg-patent hookworm infections.\u03bcL\u22121 blood. On day 7, 40 children (26\u00b73%) remained malaria positive by microscopy and on day 17, 33 children (21\u00b73%) had microscopically detectable malaria infections. After genotyping using msp1, msp2 and glurp loci, 33 out of 40 and 17 out of 33 children were shown to have recrudescent P. falciparum infections on days 7 and 17, respectively. By real-time PCR 96\u00b79% of children were positive for P. falciparum on day 0 and multi-species Plasmodium infections were common, with 41\u00b71% positive for P. falciparum/P. malariae, 9\u00b72% positive for P. falciparum/P. ovale spp. and 8\u00b70% for all three species . Thus strong evidence for clustering of malaria infections after AL treatment was not found, although this may reflect a lack of power due to the small numbers of infected individuals on days 7 and 17.To determine whether there were differing risks of infection after treatment for children living in different parts of the village, their households and infection status on days 7 and 17 were mapped . SpatialP. falciparum parasitaemia 7 days after treatment and that 22\u00b73% (11\u00b77% recrudescent) had parasites in their blood after 17 days, despite the fact that 33% of these positive children had already been retreated with AL at day 7. A substantial number of additional recurrent infections were identified by real-time PCR, including P. malariae and P. ovale spp. infections. Although none of these infections could be considered \u2018complicated\u2019 clinical malaria, their significance needs to be better quantified.In light of the challenges in diagnosis of falciparum and non-falciparum malaria and the chronically high prevalence of infections in Bukoba village, Uganda, we document the short-term effect of AL treatment on multiple malaria species in a cohort of young children living in this area. The study was designed to gain insights into treatment responses when AL was distributed to parasitaemic children in a community-based setting (unsupervised treatment), rather than as a carefully controlled standard efficacy trial. It is outside the remit of this paper to explore how well the results of standard efficacy trials predict the effectiveness of home-based management strategies in the same population. However, we found by microscopy that 26\u00b73% of treated children (21\u00b79% recrudescent by PCR genotyping) showed et al.et al.et al.et al.et al.et al.et al.et al.et al.et al.et al. reported a small decline in parasitological response rates to ACTs (including AL) in coastal Kenya  in recurrent infections soon after AL treatment , there was negligible risk of parasitaemia before day 21 and that risks of treatment failure by day 28 ranged from 1\u20139%  and those with lower parasitaemia (data not shown). Similar responses to AL were also observed when children with fever or moderate/severe anaemia on day 0 were compared with those with no fever or anaemia. However, the number of symptomatic individuals in our study may be too small to detect subtle differences in responses to AL treatment between symptomatic and asymptomatic individuals.This study was based on assessment of parasitaemia rather than clinical illness and most children had asymptomatic infections on day 0. The 21-day efficacy of the ACT dihydroartemisinin piperiquine (DP) for treatment of et al.et al.et al.et al.et al.et al. in Kenya, who showed that residual sub-microscopic parasitaemia on day 3 after commencement of AL or DP treatment was associated with longer gametocyte carriage, increased transmission to mosquitoes and recurrence of microscopically detectable malaria infections on day 24 or day 48 (Beshir et al.There has been little work assessing the dynamics of parasite clearance after AL treatment using PCR, although a median clearance time of 2 days has been reported (Aydin-Schmidt P. malariae and P. ovale infections detected by real-time PCR at baseline were mixed infections with P. falciparum, which were difficult to detect on thick films despite cross-checking. Plasmodium malariae was not fully sensitive to AL in this study as around 9% of infected individuals were still positive on day 17, and two children were P. malariae positive at day 24 after three antimalarial treatments. The fact that 50% of study participants were infected with P. malariae and 18% with P. ovale indicates that the number of mixed infections receiving ACTs is substantially larger than previously thought. Mombo-Ngoma et al. reported, in 38 Gabonese patients, a 28-day cure rate by microscopy of 100% for AL in treatment of P. malariae, P. ovale and mixed infections in Gabon, although post hoc PCR revealed that only 19 of the patients in the study had been correctly diagnosed with non-falciparum malaria (Mombo-Ngoma et al.et al. detected persistent P. malariae and P. ovale spp. infections in Ghanaian schoolchildren 21 days after ACT treatment using qPCR as the endpoint measure (Dinko et al.P. malariae and P. ovale spp. would be of great assistance in these endeavours.The vast majority of et al.It must be borne in mind that our study did not formally determine how well the participants complied with the approved antimalarial treatment regime, i.e. two doses a day for 3 days after consumption of fatty food WHO, a. Measurin vivo study protocols, with sensitive molecular endpoints, must now be developed that are specifically designed to measure the efficacy of antimalarial regimens against asymptomatic and sub-patent Plasmodium spp. infections.The burden of malaria remains high in Uganda. Whilst the Ugandan malaria policy promotes AL as the first-line antimalarial treatment and is doubtless saving many lives, other challenges in its implementation remain. Our results suggest that even though ACTs remain highly efficacious in treatment of clinical disease in sub-Saharan Africa, asymptomatic and submicroscopic falciparum and non-falciparum infections often persist after treatment. These infections may be significant in terms of later recurrence of clinical disease and parasite transmission. Improved diagnostics are needed for field-based detection of non-falciparum malaria infections, gametocytes and drug-resistance markers. Increased community-based surveillance using a range of diagnostic tests is required to provide further insights into the extent of persistent parasitaemia after ACT treatment and for early detection of ACT-tolerant parasites in sub-Saharan Africa. Finally, new"}
+{"text": "Widespread parasite resistance to first-line treatment for uncomplicated malaria leads to introduction of new drug interventions. Introducing such interventions is complex and sensitive because of stakeholder interests and public resistance. To enhance take up of such interventions, health policy communication strategies need to deliver accurate and accessible information to empower communities with necessary information and address problems of cultural acceptance of new interventions.To explore community understanding of policy changes in first-line treatment for uncomplicated malaria in Kenya; to evaluate the potential role of policy communication in influencing responses to changes in first-line treatment policy.Data collection involved qualitative strategies in a remote district in the Kenyan Coast: in-depth interviews (n\u2009=\u200929), focus group discussions (n\u2009=\u200914), informal conversations (n\u2009=\u200911) and patient narratives (n\u2009=\u20098). Constant comparative method was used in the analysis. Being malaria-prone and remotely located, the district offered an ideal area to investigate whether or not and how policy communication about a matter as critical as change of treatment policy reaches vulnerable populations.Three years after initial implementation (2009), there was limited knowledge or understanding regarding change of first-line treatment from sulphadoxine-pyrimethamine (SP) to artemether-lumefantrine (AL) for treatment of uncomplicated malaria in the study district. The print and electronic media used to create awareness about the drug change appeared to have had little impact. Although respondents were aware of the existence of AL, the drug was known neither by name nor as the official first-line treatment. Depending on individuals or groups, AL was largely viewed negatively. The weaknesses in communication strategy surrounding the change to AL included poor choice of communication tools, confusing advertisements of other drugs and conflicts between patients and providers.Effective health policy communication is important for the uptake of new drug interventions and adherence to treatment regimens. Besides, prompt access to effective treatment may not be achieved if beneficiaries are not adequately informed about treatment policy changes. Future changes in treatment policy should ensure that the communication strategy is designed to pass sustained, accurate and effective messages that account for local contexts. Changes in first-line treatment for uncomplicated malaria have occurred from chloroquine to antifolate combination drugs and to the current artemisinin-based combination therapy (ACT). However, there is no uniformity across countries in the stages of change from one drug to another because there are differences in geographical distribution of resistance to any single antimalarial. Chloroquine (CQ) was the most widely used anti-malarial drug since it was first synthesized in 1934 and for many decades was the drug of choice for treatment of non-severe or uncomplicated malaria, until drug resistance greatly reduced its usefulness [Changing treatment policy is a standard scientific procedure but such a change needs to be understood by the entire citizenry who are the targeted beneficiaries. To reach everyone with adequate and accurate information about the change in treatment policy, there is a need for concerted health communication. As noted by  poor heaSulphadoxine-pyrimethamine (SP) was introduced in Kenya in 1998 but by 2001 the treatment failure rate was over 25%, a threshold for changing first-line treatment for uncomplicated malaria. The Kenya Drug Policy Technical Working Group met in 2003 and proposed artemether-lumefantrine (AL) (Coartem\u00ae) to replace SP because it was the only co-formulated ACT recommended by the WHO as having passed rigorous international regulatory scrutiny. Artemether-lumefantrine was finally rolled out country-wide in 2006 after much debate about sustainable financing and procurement regulations .To implement the change, the MOH conducted training for health workers at the national, provincial and district levels. The training recommended parasitic diagnosis before administering Coartem\u00ae even though the government of Kenya  indicateThe study was conducted in a remote rural district on the Kenyan coast. The district was chosen because of the need to explore how policies get communicated and implemented in remote areas. More specifically, was the need to understand how policy changes in first-line treatment of uncomplicated malaria are implemented and how the same are understood by populations in hard-to-reach locations. The focus on a hard-to-reach population in an economically marginalized district was important to the study as it sought to understand how health interventions respond to the needs, characteristics and local circumstances of populations in remote locations. As stated by  vulnerabA cross-sectional qualitative study was carried out in three purposively sampled administrative zones out of the four that make up the district. Two of the divisions were farthest from the urban Kinango district headquarters and the other was the location of the headquarters. This sampling approach was intended to capture any differences in experience between people living closest to the district headquarters and those from the farthest part of the district. Two villages per division were randomly selected to participate in the study.Data were collected between January and February 2009 using document reviews, focus group discussions (FGDs) (n\u2009=\u200914), in-depth interviews (n\u2009=\u200929), informal conversations (n\u2009=\u200911) and patient narratives (n\u2009=\u20098). The number of interviews was not predetermined and interviews were stopped on saturation of information. Kiswahili was the main language of communication as it was the language of the respondents. Participants for FGDs were selected on the basis of age and gender. They included parents with young children aged below ten years because young children and pregnant women are most vulnerable to malaria infection. The range of topics explored in the FGDs included local information networks and sources of health information and perceptions about anti-malarials.In-depth interviews were conducted with district health managers, community leaders and managers (in-charges) of six dispensaries. They provided information on the policy communication processes and their effectiveness, implementation challenges and uptake of AL. Table\u00a0Informal conversations were based on chance and dwelt on communication about health issues and accounting for gender differences in the sources of health information as well as people\u2019s perceptions about changes in malaria drugs. Patient or caretaker narratives were conducted with purposively selected patients or caretakers who were willing to talk to the researcher at the health facilities. Those selected were asked to tell their story about their illness or illness of the child; how it started, diagnosis, the range of treatment options undertaken and their effectiveness as well as their interaction with health workers. All interviews were audio-recorded except for informal conversations.All interviews were transcribed and translated into English. Coding and analysis were performed manually by developing a matrix for emerging categories and themes. Data from various sources were first analysed separately and emerging themes and categories compared later. The themes and categories centred on phrases, incidents and behaviour during interviews, discussions, and observations. In accordance with recommendations of ,11, dataThe criterion for selecting data to present was based on relevance to the stated objectives. Data presented here were based on the level of importance as judged by emphasis and consistency in raising specific issues during interviews by study participants.Ethical approval was sought from the Kenya Medical Research Institute (KEMRI) Ethical Review Committee and the University of Cape Town Ethical Review Board, Faculty of Health Sciences. All participants gave free and informed, written and/or verbal consent for the study.Based on relevance to the objectives, the results were summarized and presented in relation to three sub-topics that represented the study participants\u2019 main lines of understanding of AL and the changes in first-line treatment policy for uncomplicated malaria. The first part looks at the study population\u2019s perceptions about general changes in first-line treatment policy for uncomplicated malaria over time, because such perceptions have a bearing on take-up of new drug interventions. The second part explores perceptions specific to the change from SP to AL including views of the study participants about the importance of AL as a first-line drug. The last part addresses barriers to health policy communication that most likely influenced poor uptake of AL and identifies key variables that could be useful for future interventions.It was not clear from the literature or from the study participants how they got communication about earlier drug changes from chloroquine to SP. However, the majority of participants indicated that they learnt about new drugs mainly through media advertisements and at health facilities. Given the profit motivation of drug companies advertising through the media, most study participants strongly believed that any change in treatment policy was mainly driven by the desire for profit maximisation. This view was shared by all FGD participants, 17 key informant interviews, two district health managers, and four primary health facility in-charges. A community leader said of the changes:\u201cI think chloroquine is no longer available because the company associated with it had made a lot of money so the government thought it was time to give another company chance to make money. That explains why every time we have different types of drugs for malaria.\u201dOther study participants believed that drugs change because the government keeps testing different drugs to establish the most effectiveness one against malaria. Asked whether the change in malaria drugs was good for the community, most participants in 12 FGDs and nine informal interviews reported that it did not contribute to better treatment for malaria. They believed that any changes were confusing, as stated by female FGD participant:\u201cThe change of drugs is very bad because when we are already used to one type of drug then one day we are given a new one whose name we do not know. This makes treatment difficult because we do not know which drug to use anymore\u2026.\u201dHowever, six key informants out of 20 at the village level understood the scientific basis for change of drugs and explained that drugs change due to treatment failure.Nearly all study participants were aware of the existence of AL but knew it only as \u201cthe drug with many tablets\u201d. They had no idea that it replaced SP as the official first-line treatment for uncomplicated malaria. Only six people (four women and two men) among all study participants at the village level knew AL by name and recognized it as the official first-line anti-malarial. These individuals had formal jobs and better education than their peers had, and were residents of a village near the urban section of the study district.There were mixed responses at the village level when asked about the history of changes in first-line treatment. Most respondents explained that drugs have changed from CQ to amodiaquine (AQ) or from SP to a number of painkillers as advertised through the radio. The respondents further explained that the variety of drugs mentioned reflected people\u2019s preferences since there was no single drug that suited every malaria patient.In the narratives, three malaria patients thought AL was a painkiller and most men in the more remote villages believed that malaria drugs were seasonal and preferred CQ to AL. A 51\u00a0year old man said in an FGD:\u201cWe do not know that drugs have changed, but we know there are seasons for malaria drugs and it is now the season for malaratab (amodiaquine) and that other one with many tablets. When chloroquine season returns, we will have no problems over which drug to use to treat malaria\u2026\u201dWhenever people have negative attitudes toward a health intervention, uptake is likely to be affected. A policy communication strategy that accompanies change of health interventions should pre-empt or change such negative attitudes. However, in the study area, a number of respondents regarded AL as harmful. This was identified in 14 in-depth interviews, nine FGDs and seven informal interviews. Older individuals (50\u00a0years and above) were particularly concerned that AL was more harmful and less effective than CQ. One male key informant remarked:\u201cWhen taking the new drug (Coartem), we ask ourselves many questions: \u2018Where are they from and why can\u2019t we be given the ones that we know? What harm can they cause to us?\u2019 This makes us afraid\u2026.. We use this drug but inwardly we ask ourselves why it is different from the ones we used before and whether they are any better in treating malaria\u2026.\u201dThe perceived harm in AL was further emphasized in the fact that it was not recommended for expectant mothers. A young man said:\u201cRecently, I asked my wife why she was given fewer drugs than those given to everyone who goes to the dispensary and she told me that the other drug with many tablets cannot be given to pregnant women. That confirms the new drug can be harmful to anyone else.\u201dstrong and weak malaria because using AL to treat \u2018weak\u2019 malaria when it is a strong drug could render it ineffective against \u2018strong\u2019 malaria and this could be fatal. Those who perceived AL as weak cited its numerous tablets (24 tablets for adult dose while SP had three tablets). A female FGD participant said of AL tablets:Study participants classified malaria into two categories: \u2018weak\u2019 and \u2018strong\u2019. Weak malaria meant mild or non-serious illness in which the patient is not incapacitated in any way. Strong malaria on the hand meant that the patient is perceived to be seriously ill with malaria and requires urgent medical attention. A number of study participants complained that one type of drug cannot be used to treat both \u201cWe heard that malaria drugs that we are used to are no longer good and hoped for a new and better drug, but that is not the case because a full dose of the new drug has 24 tablets instead of just a few. Why are the tablets so many if it is a good drug?\u201dAccording to health workers, some of those who thought that AL was weak were cases of recurrent malaria, which sometimes led to the judgement that AL was weak. A health worker confirmed that a few people who experienced recurrent malaria after taking AL were reluctant to use the drug again or seek any formal treatment at all. Those seeking formal treatment were keen to use a different health facility in the hope that they will not be given the same drug (AL). Should they be given the same drug, they were unlikely to take it, as confirmed by a female FGD participant:\u201cIf we suffer from malaria soon after taking all those tablets, we demand for a different drug. That means we would go to a different dispensary but if that dispensary also issues the same drug, we bring those drugs home to keep, then turn to herbs or go to private clinics for injections....\u201dA number of factors emerged as clearly influencing communication around the change of first-line drugs for uncomplicated malaria.The findings indicate that the choice of policy communication strategies used by the government during the change to AL were not the most preferred in the study area. Figure\u00a0Among the information, education and communication (IEC) tools used to facilitate the change from SP to AL, only radio and posters seemed to have reached the study district. However, the impact of these tools on the primary caretakers, mostly women, was negligible because most radios were owned and controlled by men. Although most people at health facilities were women and potentially had access to posters advertising the change to AL, widespread illiteracy made it impossible for them to read the posters. A young woman said of the posters:\u201c\u2026all we see are graphics and pictures, which we think are decorations on the dispensary walls.\u201dStudy participants emphasized that they preferred interpersonal communication to radios and posters and the best sources would be public health officers (PHOs), local authorities and teachers, and complemented by radio communication. The interpersonal communication allowed for exchange of views and prompt response to community concerns about the newly introduced drugs. For interpersonal communication however, the PHOs were not part of the change process for lack of resources to train them but a health manager acknowledged the mistake in leaving out the PHOs:\u201cIt was a mistake to leave out the PHOs because their involvement would have ensured a more successful implementation of the change to AL.\u201dThe health manager explained that emphasis on interpersonal communication would have best clarified that AL was the most suitable drug to treat malaria and could also have eliminated confusion over which drugs to use. A typical confusion over drugs was captured in the following statement:\u201c\u2026the dispensary gave my wife what it said were the latest malaria drugs but the shopkeeper also had the latest drugs different from the dispensary. This is confusing.\u201dConflicts were manifest in three areas: provider-patient communication, perceived health worker incompetence and professional integrity. Study participants complained of poor interpersonal relationships with health workers. In the narratives, two patients complained about rude health workers who refused to answer their queries about AL. A mother said:\u201cI wanted to know why I was given malaria drugs different from the ones I have known all along but the nurse asked me whether I came to be treated or to ask questions\u2026.\u201dHealth workers admitted they rarely engaged patients about the change to AL because of the limited time to attend to large numbers of patients. At least one in-charge mentioned that it was not their responsibility to discuss issues of adherence with patients and saw no need to inform them about any changes in treatment:\u201cThe patients are usually very many and we cannot engage each of them in lengthy conversations about adherence. Besides, I have never considered it important to engage the community on such issues as change of drugs....\u201dThe perceived inability of health workers to give proper treatment affected use and adherence to AL. In six FGDs, some study participants regarded presumptive treatment as poor quality care and preferred diagnostic tests. A young mother said:\u201cThe health worker just looks at the baby and prescribes treatment. How does he know what he is suffering from without tests?\u201dSome health workers were also alleged to engage in unethical practices including running private businesses within public health facilities and allegations of sale of facility drugs. A young man said:\u201c\u2026the health worker shows you two types of drugs\u2026. He would say, \u2018This drug belongs to the government, and these other one is mine. Mine is the best because it is stronger; so you choose which one you want.\u2019 His drugs are indeed better but more expensive.\u201dA range of anti-malarials and other drugs sold over the counter are often the subject of sustained marketing advertisement through the print and electronic media. The continuous advertisement of such drugs by respective companies overshadowed government advertisement of AL the same media. Consequently, community members could not differentiate between the drugs advertised by the private companies and AL as the recommended first-line drug. Concerns about the role of drug companies in limiting uptake of AL were expressed by health workers:\u201cThere is a lot of information coming through radios and posters regarding different types of malaria drugs and without anyone to differentiate for the ordinary people and show them that Coartem is the best and recommended drug\u2026 they get confused by the adverts....\u201dhoma\u2019 can be translated to mean fever, common cold and flu, but it also refers to febrile conditions such as malaria. It was not always clear from the adverts what type of \u2018homa\u2019 the advertised drugs treated and so patients often used painkillers to treat malaria with the understanding that they were drugs for \u2018homa\u2019. A health worker commented:In a few but important cases, the language used in the advertisements by drug companies lacked clarity; for example, among the study community and the wider Coastal region, the term \u2018\u201cIn this community, the word \u2018homa\u2019 means different illnesses including common cold, stomach upsets and malaria. Unless these are clarified in the advertisement for drugs, the locals will continue using painkillers in treating malaria....\u201dThere was very limited knowledge within the study district about what prompts changes in treatment policy. Lack of such knowledge led to mostly negative interpretations about why there had been changes in first-line treatment for uncomplicated malaria. Specific to the treatment policy change from SP to AL, the communication strategy employed to facilitate take up of AL had little impact in the study district and instead seemed to perpetuate negative perceptions about the change from SP to AL. Although this study was conducted in 2009, the issues it raises remain important for consideration in future changes in treatment policies and accompanying communication, particularly in remote settings.One of the things that changing treatment policy and the accompanying communication have to contend with is the entrenched health seeking and treatment habits of the target population. A number of authors such as -16 have To a large extent, our findings indicate that weaknesses in policy communication during the change from SP to AL contributed to the poor uptake. Part of the reason for the communication problems was that a number of contextual factors relevant to the study population were apparently not considered in the strategy applied. Contextual factors, according to  influencThe government\u2019s overreliance on the print (written) and electronic media meant that communication about the change to AL remained largely inaccessible to most of the study population because of illiteracy and lack of control of such resources as radio and the financial means to maintain them. According to , poor coRecommendations by ,19 also Central to interpersonal communication during health policy communication is patient-provider engagement. Community members understood health workers as experts in their field, which explained their strong preference for public health officers as their preferred main sources of health information. Health information communicated by experts, as perceived by the community, is better received compared to communication from ordinary persons . FurtherFinally, negative perceptions about change of treatment policy for uncomplicated malaria and about AL in particular, could also have emanated from lack of trust in overall political governance given wider media and other reports that described massive high level corruption, for example . Being aThe study highlights the need for effective policy communication using locally available channels during the change of first-line treatment for uncomplicated malaria, and perhaps for other drug regimen changes. In addition, communication of the treatment policy change should be supported by other actions such as training health workers to open dialogue and effectively communicate with patients to inspire confidence in new drugs as well as a prolonged public education campaign that allows the public to adjust to the newly introduced drug. Such an approach should be able to increase the amount and quality of information received by beneficiaries, which is important for take-up of new health interventions. Although this study was undertaken only in one district, its detailed investigation of experience suggests that prompt access to effective treatment may not be achieved in any other setting if beneficiaries are not adequately prepared for changes in drug regimens such as first-line treatment for uncomplicated malaria. The study suggests that future changes in first-line drugs should ensure that treatment policy communication is well designed to inspire trust and confidence in the public, particularly among people in remote settings, and to facilitate acceptance and take-up of new health interventions. The choice of a communication channel is as important as the intervention itself.The authors declare that they have no competing interests.VO designed the study, collected and analysed data and wrote the first and final drafts. LG supported the study design and supervised the entire study. Both authors read and commented on all drafts."}
+{"text": "Pneumonia is the leading cause of infectious death amongst children globally, with the highest burden in Africa. Early identification of children at risk of treatment failure in the community and prompt referral could lower mortality. A number of clinical markers have been independently associated with oral antibiotic failure in childhood pneumonia. This study aimed to develop a prognostic model for fast-breathing pneumonia treatment failure in sub-Saharan Africa.We prospectively followed a cohort of children (2\u201359 months), diagnosed by community health workers with fast-breathing pneumonia using World Health Organisation (WHO) integrated community case management guidelines. Cases were followed at days 5 and 14 by study data collectors, who assessed a range of pre-determined clinical features for treatment outcome. We built the prognostic model using eight pre-defined parameters, using multivariable logistic regression, validated through bootstrapping.We assessed 1,542 cases of which 769 were included . The treatment failure rate was 15% at day 5 and relapse was 4% at day 14. Concurrent malaria diagnosis  and moderate malnutrition  were associated with treatment failure. The model demonstrated poor calibration and discrimination (c-statistic: 0.56).This study suggests that it may be difficult to create a pragmatic community-level prognostic child pneumonia tool based solely on clinical markers and pulse oximetry in an HIV and malaria endemic setting. Further work is needed to identify more accurate and reliable referral algorithms that remain feasible for use by community health workers. Haemophilus influenzae B (HiB) and pneumococcal conjugate vaccines (PCV) [Globally, pneumonia is the leading infectious cause of death amongst children under five, with an estimated 0.9 million deaths annually, and the highest burden found in Africa , 2. Whiles (PCV) , 4, effeThe World Health Organisation (WHO) integrated community case management (iCCM) guidelines clinically stratify pneumonia into either non-severe pneumonia (fast breathing only) or severe pneumonia requiring referral (presence of chest indrawing or danger signs) . Non-sev2) <90%) and poor antibiotic adherence [Treatment failure is the persistence of symptoms or deterioration following antibiotic initiation . Common dherence \u201316. To ddherence .A referral algorithm that could be implemented at the community-level  presents an opportunity for further reduction in pneumonia morbidity and mortality, through more rapid referral of higher risk children when they are more amenable to alternative treatments. The aim of this study was to develop a pragmatic prognostic algorithm for treatment failure in children diagnosed with fast-breathing pneumonia by community health workers (CHWs), treated with oral co-trimoxazole (the first line treatment during the study period) in the community.We conducted a prospective cohort study in Malawi from September 2013 to June 2014, nested within a larger parent study assessing the impact of PCV introduction in Malawi. This study was conducted in sub-populations of two districts in the central region of Malawi (Mchinji and Lilongwe). The populations covered were all rural, with mobile phone ownership of 35% and the populations were predominantly subsistence farmers .The primary health care system in rural Malawi consists of government employed CHWs, called Health Surveillance Assistants locally. CHWs receive 12 weeks basic training delivered by the government (including iCCM), and they run designated village clinics one or two times per week as well as conducting house visits and outreach campaigns. These CHWs cover catchment areas of approximately 1,200 people, although this varies according to region, and are supervised by senior CHWs based at health facilities.Patients were recruited at weekly village clinics run by 34 CHWs, 18 in Mchinji and 16 in Lilongwe rural, with a catchment area of approximately 50,000 people and 8,500 children under five. CHWs were responsible for the initial clinical assessment, following slightly modified iCCM guidelines with concurrent diagnoses and treatments allowed; in Malawi HIV and malaria rapid diagnostic testing (RDTs) are not fully implemented at the community-level and it is common for acute fevers to be presumptively treated as malaria . This coIf diagnosed with pneumonia, information from the clinical assessment (including pulse oximetry with Lifebox\u00ae) was recorded on case report forms (CRFs) by the CHW, along with their referral or treatment decision. To measure PCV effectiveness, the parent PCV study required a consistent clinical pneumonia definition applied across the community, health centre, and hospital health system levels. Therefore, the CHW CRFs included additional respiratory signs associated with more severe disease and routinely collected at the facility level, but not previously included at the community level, including nasal flaring, head nodding, and grunting. Pulse oximetry and the CRFs were introduced among these CHWs in late 2011 as part of the parent PCV study and were also used for this sub-study, both are not currently standard care at this level in Malawi [Although CHWs are recommended to follow up children after pneumonia treatment, this is not done routinely in actual practice, and this study required routine, timely, and rigorous outcome assessments. Community-level outcome assessments were not included in the parent PCV study. For each village clinic, we therefore employed local village level data collectors (VDCs) to recruit and conduct routine, timely, and comprehensive follow up interviews. VDCs received training on conducting a clinical assessment for pneumonia signs and symptoms, using a pulse oximeter, the study protocol and questionnaires. Clinical assessment training included the use of videos, demonstrations, practical sessions and supervised assessments with real cases at local health centres. All CHWs also attended the training. A one month field pilot (August 2013) was conducted, followed by a half day refresher training for all VDCs and CHWs prior to study recruitment.Patient recruitment was done by the VDCs at the village clinics (on \u2018Day 0\u2019). Patients were eligible for recruitment if the CHW had diagnosed them with fast-breathing pneumonia and prescribed oral antibiotics for home treatment. VDCs followed up patients at their homes on day 5 and day 14. If the patient was not located on day 5, the VDCs re-visited the home on day 7. At the follow up interviews, VDCs conducted a clinical assessment of the child (including pulse oximetry), asked about additional care seeking and antibiotic adherence. We used smart phones for follow up data collection, with data downloaded onto field supervisor\u2019s laptops every 2 weeks and cleaning checks run each month. Data was linked with the CHW filled CRFs through the unique barcode, which the VDCs scanned to register a case at the village clinic.data quality assurance exercises were carried out to ensure reliable and consistent data, including: 1) an initial vital signs standardization audit conducted by a paediatric pulmonologist, which all VDCs passed. 2) Random GPS spot checks by field supervisors of follow up locations. 3) All VDCs had at least one interview supervised by a pediatrician to ensure standardized clinical assessments including oximetry, in which all performed satisfactorily. 4) Monthly data review and feedback meetings, to highlight and address data quality issues. 5) Regular shadowing of interviews by field supervisors, with concurrent recording of clinical signs. 6) Remote clinical validation for the presence of lower chest indrawing from a random sample of VDC collected video clips.Several all children aged 2\u201359 months with CHW-diagnosed WHO fast-breathing pneumonia who were adherent to home oral co-trimoxazole treatment. First line treatment for fast-breathing pneumonia in Malawi is a 5-day BD (twice a day) course of co-trimoxazole .2 <90%, and grunting, nasal flaring and head nodding as signs of severe respiratory distress. All children who were severely malnourished (mid-upper arm circumference (MUAC) <11.5cm) were referred.defined according to the 2013 iCCM guidelines, as the defined as the presence of any of the following on day 5 of follow up : fast breathing for age, axillary temperature >37.5\u00b0C, lower chest indrawing, any danger sign (as defined above), change of antibiotic, hospital admission or death . Changedefined as completing at least 80% of the prescribed antibiotics and confirmed by visual examination of remaining antibiotics by the VDC. We restricted the model to \u2018adherent\u2019 children only, as non-adherence has been shown to be associated with treatment failure in previous studies; the aim of this study was to make a referral tool based on clinical presentation only.a priori for inclusion in the prognostic model based on a literature and expert panel review and practicality for use at the community level: oxygen saturation; respiratory rate; fever (axillary temperature >38\u00b0C); malnutrition, as determined by MUAC measurements; age; PCV-13 and pentavalent vaccination (including HiB). We also decided to include concurrent clinical malaria diagnosis. The continuous variables were categorized before inclusion into the model, based on substantive knowledge as follows: oxygen saturation of SpO2 90\u201394% and \u226595%; MUAC as moderate malnutrition (11.5\u201313.5cm) and normal nutrition (\u226513.5cm); age as 2\u20135, 6\u201311, and 12\u201359 months; respiratory rate as fast  and very fast breathing .We described the cohort, including levels of missing data. The following variables were determined 2; the Hosmer-Lemeshow goodness of fit test ; c-statistic ; slope shrinkage in the internal validation . Additional information is found in Multivariable logistic regression was used to create the prognostic model, retaining all pre-defined risk variables. Missing data were imputed using multiple imputation with chained equations , assuminThe sample size calculation was based on the requirement of 10 treatment failure cases per degree of freedom in the model, therefore with the eight candidate predictors evaluated using 11 degrees of freedom, we required a sample of at least 110 treatment failures.th percentile, median, 75th percentile and maximum values by CHW. CHWs were flagged if for any variable, the 25th and 75th percentiles were equal and the model was re-fit without the data from these CHWs. All descriptive analysis was done using Stata SE11 and statistical analysis was done using R version 3.1.1 [We performed sensitivity analyses to compare: 1) the baseline characteristics between those lost to follow up and those included in the final analysis; 2) check for outlying CHWs to determine whether data collection inaccuracies could give rise to spurious prediction results. For baseline clinical observations we calculated the mean, standard deviation, minimum, 25on 3.1.1 .Informed consent for study recruitment and prospective follow up was sought from the child\u2019s accompanying care-giver at the initial clinical assessment and was confirmed at each subsequent follow up with the care-giver present. The study information sheet was read aloud by the VDC before starting the interview. Consent was given verbally and recorded in the electronic form on the smart phone. Verbal consent was sought as literacy is low in this population. Ethical approval for this informed consent procedure and the overall study was granted by the National Health Sciences Research Committee in Malawi (reference: 941).The VDCs assessed 1,542 children diagnosed with fast-breathing pneumonia by the CHWs for enrollment over a 9 month period, of which 1,055 were eligible. Reasons for ineligibility were: presence of lower chest indrawing and/or danger signs (60%), no fast breathing (20%), children were not given antibiotics (16%) or children were too old or too young (4%). A total of 197 children (19%) were lost to follow-up; 2 value). The model equation is:Odds ratios, following imputation for missing values, for the eight pre-defined predictors of treatment failure are reported in th percentile. The observed risks of treatment failure for each quintile of predicted risk were plotted at the mid-point of the quintile. Children in the highest-risk quartile were 3.19 times more likely to fail oral co-trimoxazole than children in the lowest-risk quartile, a measure of discrimination. The c-statistic  demonstrated that the model had a low ability to detect subtle differences or discriminate. We found evidence of adequate calibration as the observed risks agreed approximately with the predicted risks for each quintile. However, the Hosmer-Lemeshow goodness of fit test showed some evidence of lack of fit of the model (P = 0.04).The predicted risk of treatment failure was plotted with a risk predictiveness curve . The preth percentile), 33.3%  of the treatment failure cases would be referred; this is the sensitivity. The same decision rule would result in a specificity of 82.4% , a positive predictive value of 24.8%  and negative predictive value of 87.7% . While the sensitivity and specificity are properties of the prediction model and the decision rule for referral, the predictive values depend on the incidence of treatment failure in a given population and setting. If CHWs referred those children in the top risk quintile  validation, we found some evidence of over-fitting the logistic model to the data because the slope-shrinkage statistic was 0.6. 2 of 5.1%. We found the best predictors for failure were a concurrent clinical malaria diagnosis and moderate malnutrition, while predictors such as young age and respiratory rate did not show any significant associations. Possible explanations of our findings include: lack of key variables ; challenges in community diagnosis of pneumonia; and quality of antibiotics, adherence and additional care seeking.We found a treatment failure rate of 14.8% at day 5 of follow up, for fast-breathing pneumonia treated with co-trimoxazole. This is consistent with rates found in previous studies of fast-breathing pneumonia treatment failure with co-trimoxazole, with rates ranging from 11\u201321%, albeit from studies conducted in South Asia at the facility level, and not with CHWs , 24. TheIn the context of this stud y setting, we were unable to reliably or ethically collect HIV status of the children included, as the study was implemented within an existing larger parent PCV13 effectiveness study. The cohort of CHWs that were originally selected to participate in the wider study were not trained or qualified to provide HIV care or use RDTs for malaria testing, something which is common in community settings. Prior studies have demonstrated that being HIV-infected or -exposed increases the odds of treatment failure in pneumonia  , 26. MalWe found concurrent clinical malaria diagnosis to be a predictor of antibiotic treatment failure. A study from Mozambique found that malaria diagnosis was the strongest risk factor associated with having an acute respiratory infection (ARI) diagnosis in infants . Prior sWe excluded infants who were non-adherent from the analysis, based on information given by the care-giver and where possible, visibly counting remaining tablets. However, in this setting maternal education and literacy are low (35% completely illiterate ) and it This study had many strengths, in that it was a prospective community-based cohort implemented in a quasi-programmatic setting, generating evidence not only on outcomes but also the practicality of developing a reliable pragmatic prognostic without laboratory support. It also included the use of pulse oximetry at the CHW level to support diagnosis and referral decision-making. However, our key limitations were the lack of HIV and malaria testing and relatively high loss to follow-up, despite dedicated VDCs. Cases were recruited into the cohort during village clinics, where the VDCs would arrange to visit the patient\u2019s house for follow up. Despite VDCs being recruited from the local area, there were issues in locating households, and families being unavailable. The challenges with the active follow-up, included: lack of consistent address information; people using multiple names; follow ups taking place over the rainy season making travel challenging, and during planting and harvesting time when care-givers were busy farming. We have learnt several lessons in attempting to prospectively evaluate risks of treatment failure in clinically diagnosed pneumonia at the community level. Firstly, the reliability of CHW clinical diagnosis and treatment/referral decisions is a challenge and, despite active mentorship, misclassification and therefore incorrect treatment was common. Malawi guidelines do allow for CHWs to provide HIV testing services and recommend the use of RDTs but this is only realistically implemented with private support. Nevertheless, to address these challenges future studies should consider recruiting cohorts from higher levels of the healthcare system with access to malaria testing, HIV status and more qualified healthcare staff, although this may limit generalizability to the community setting. Secondly, our choice of VDCs was based on their local knowledge of the communities and not on clinical experience; in spite of this, we still had high loss to follow-up. Future studies could consider incentivizing care-givers to return to the healthcare facility for follow-up assessments as active community-based follow-up poses complex challenges, despite significant efforts. As co-trimoxazole is no longer the WHO recommendation for outpatient pneumonia treatment and newly revised guidelines allow for home treatment of children with lower chest indrawing, further research on the performance of a prognostic algorithm is needed.While we have hypothesized several reasons for our findings, there is a lack of evidence around rates, causes and predictors of treatment failure in fast-breathing pneumonia at the community level in sub-Saharan Africa, with which to compare this study . The difS1 Text(DOCX)Click here for additional data file."}
+{"text": "Plasmodium falciparum but its effect on post-treatment gametocyte circulation and infectiousness to mosquitoes has not been quantified.A single low dose (0.25\u00a0mg/kg) of primaquine is recommended as a gametocytocide in combination with artemisinin-based combination therapies for In this randomised, double-blind, placebo-controlled trial, 360 asymptomatic parasitaemic children aged 2-15 years were enrolled and assigned to receive: artemether-lumefantrine (AL) and a dose of placebo; AL and a 0.25\u00a0mg/kg primaquine dose; or AL and a 0.40\u00a0mg/kg primaquine dose. On days 0, 2, 3, 7, 10 and 14, gametocytes were detected and quantified by microscopy, Pfs25 mRNA quantitative nucleic acid sequence based amplification (QT-NASBA), and quantitative reverse-transcriptase PCR (qRT-PCR). For a subset of participants, pre- and post-treatment infectiousness was assessed by mosquito feeding assays on days -1, 3, 7, 10 and 14.Both primaquine arms had lower gametocyte prevalences after day 3 compared to the placebo arm, regardless of gametocyte detection method. The mean  number of days to gametocyte clearance in children with patent gametocytes on day 0 (N\u2009=\u2009150) was 19.7 (14.6 \u2013 24.8), 7.7 (6.3 \u2013 9.1) and 8.2 (6.7 \u2013 9.6) for the AL-placebo, the 0.25\u00a0mg/kg primaquine dose and the 0.40\u00a0mg/kg primaquine dose arms, respectively. While 38.0\u00a0% (30/79) of selected gametocytaemic individuals were infectious before treatment, only 1/251 participant, from the AL-placebo group, infected mosquitoes after treatment.We observed similar gametocyte clearance rates after 0.25 and 0.40\u00a0mg/kg primaquine doses. Infectivity to mosquitoes after AL was very low and absent in primaquine arms.NCT01935882The online version of this article (doi:10.1186/s12916-016-0581-y) contains supplementary material, which is available to authorized users. Infectiousness to malaria vectors depends on the presence of sexual stage parasites, gametocytes, in the peripheral blood and is essential to sustain transmission in endemic areas . ShortenPlasmodium falciparum malaria in elimination and artemisinin resistance containment scenarios ) were used. After the first 143 children were recruited, mosquito feeding experiments were amended: gametocytaemic children were randomised to have feeding assays on days -1 (one day before treatment initiation) and 7 or 3 and 7.Treatment concealment was achieved by the addition of a syrup, which masked the colour and taste of primaquine and placebo. Participants, investigators, and staff who were not involved in study drug administration were blinded to study arm allocation.AL  was administered as half a tablet (20\u00a0mg of artemether and 120\u00a0mg of lumefantrine) per 5\u00a0kg of body weight in a six-dose regimen over three days. Primaquine doses were prepared as previously described  and giveParticipants were asked to return to the study clinic on days 0, 1, 2, 3, 7, 10 and 14. On day 0 (day of enrolment), AL administration was initiated. On each follow-up day, study subjects were examined and, except on day 1, had a blood slide prepared to detect malaria parasites and their haemoglobin levels quantified by Hemocue photometer . Smears were screened for asexual stage parasites and gametocytes, and double-read. Biochemistry and full blood count assessments were performed on days 0, 3 and 7 on venous samples.Finger-prick-blood samples (50\u00a0\u03bcL) were collected on all visits, except on day 1, stored in 250\u00a0\u03bcL of RNAprotect\u00ae cell reagent (Qiagen), and used for gametocyte detection and quantification: quantitative nucleic acid sequence based amplification (QT-NASBA)  was perfAnopheles gambiae mosquitoes. Fully fed mosquitoes were selected, kept on glucose at 27\u201329\u00a0\u00b0C and dissected seven days later. Midguts were examined for the presence of oocysts.A subset of recruited children was invited to participate in membrane feeding assays  at two tThe primary efficacy endpoint was gametocyte clearance time (i.e. number of days to undetectable gametocyte levels) in the 0.25\u00a0mg/kg primaquine arm compared to the 0.40\u00a0mg/kg primaquine arm. The primary safety endpoint was maximal fall in haemoglobin levels during follow-up. Secondary efficacy endpoints were: gametocyte prevalence on days 3, 7, 10 and 14, and proportion of mosquitoes developing infection and their oocyst counts in feeding assays performed after treatment administration. Secondary safety endpoints included: number of participants requiring blood transfusion, maximal percentage decrease in haemoglobin concentration, proportion of participants with haemoglobin levels below 5\u00a0g/dL and number of serious adverse events.Sample size calculations were based on non-inferiority of 0.25\u00a0mg/kg versus 0.40\u00a0mg/kg primaquine dose. For the gametocyte clearance endpoint, the non-inferiority margin was 2.5\u00a0days . If mean2 or Fisher\u2019s exact test. Non-parametric tests  were used to compare parasite levels among different study groups. To quantify the effect of primaquine on transmission, post-treatment mosquito infection rates were compared between treatment arms. The maximal fall in haemoglobin levels measured by Hemocue photometer was presented as mean (95\u00a0% CI) per study arm; Student\u2019s t-test was used for pair-wise comparisons.Stata 12.0  and SAS 9.3  were used for statistical analysis. The mean time to gametocyte clearance was estimated for each treatment arm using a non-linear model   and enrolled 360 children  compared to the AL-placebo and the 0.40\u00a0mg/kg primaquine arms . Both primaquine arms had lower microscopically-detectable gametocyte prevalence on days 7, 10 and 14, but not on days 2 and 3, compared to the AL-placebo group , 7 (N\u2009=\u200952), 10 (N\u2009=\u200954) and 14 (N\u2009=\u200952); during this Study Phase, no mosquitoes developed infection , occurring in three, one and four individuals assigned to the AL-placebo, the 0.25\u00a0mg/kg primaquine and the 0.40\u00a0mg/kg primaquine arms, respectively, were considered to be possibly related to study participation  membrane feeding assays underestimate infectivity compared to direct skin feeding assays and consequently low level post-treatment infectivity might be misclassified as non-infectiousness when using this artificial system; (2) the number of mosquitoes used in each assay might not represent the number of mosquito bites an infectious individual would receive in natural settings , and binary outcomes (e.g. \u201cinfectious\u201d versus \u201cnot-infectious\u201d) might be less relevant than within-individual changes in mosquito infection rates; (3) baseline infectivity will depend on the inclusion criteria and although recruiting individuals with high gametocyte densities would maximize the power to observe changes in infectiousness with treatment, these individuals might not be representative of an \u201caverage\u201d infected person. As duration and intensity of infectiousness are likely to depend on host age and transmission setting, individuals from different ages and study sites, representing a wide spectrum of endemicities, would need to be recruited.Primaquine-induced haemolysis in individuals with G6PD deficiency is a concern for control programs and one major reason why this drug is not widely used in Africa. Here, a rapid diagnostic test was used to exclude individuals with this condition. No cases of severe haemolysis were observed, corroborating previous observations made in symptomatic children with normal G6PD activity receiving a single 0.40\u00a0mg/kg primaquine dose . HoweverThe ultimate goal of using primaquine in falciparum malaria in conjunction with ACT is to minimise post-treatment malaria transmission, reduce the transmission of artemisinin-resistant parasites and accelerate attempts to eliminate malaria . Althoug"}
+{"text": "P. falciparum malaria infections. It was recommended that split dosing should be incorporated into all artemisinin combination regimen designs. To explain why parasite clearance rates were not faster with split dose regimens it was concluded that splenic malaria parasite clearance capacity was readily exceeded, resulting in the accumulation of dead parasites in the circulation, that parasite clearance was therefore an unreliable measure of drug efficacy, and instead that human immunity is the primary determinant of clearance rates. To test these various hypotheses we performed a logistic meta-regression analysis of cure rates from all falciparum malaria treatment trials (n\u2009=\u200940) with monotherapy arms containing artemisinin or a derivative (76 arms). There was no evidence that split dosing enhanced cure rates.It has been suggested recently, based on pharmacokinetic-pharmacodynamic modelling exercises, that twice daily dosing of artemisinins increases malaria parasite killing and so could \u201cdramatically enhance and restore drug effectiveness\u201d in artemisinin resistant  The artemisinins are eliminated rapidly (t1/2~1\u2009hour) but giving them twice or even three times in one day did not appear to provide additional benefit over once daily administration, so this became the norm14. There were a few exceptions. In uncomplicated malaria artemether-lumefantrine required twice daily administration because of the readily saturated oral absorption of lumefantrine. In severe malaria there was concern that in a highly synchronous infection in which mature schizonts predominated, sub-maximal effects might result from the first parenteral administration so a second dose was given at 12\u2009hours as an \u201cinsurance policy\u201d.When artemisinin and its derivatives were first evaluated in the treatment of malaria a variety of doses and dosing schedules were assessed. Following single or multiple doses, rates of parasite clearance in falciparum and vivax malaria were faster than observed previously with other classes of antimalarial drug, but satisfactory cure rates with artemisinins alone in falciparum malaria required dosing for more than five days17 and one study went as far as to claim that it could \u201cdramatically enhance and restore drug effectiveness\u201d in artemisinin resistant infections18. The authors further recommended \u201cthat twice-daily dosing should be incorporated into all artemisinin combination treatment (ACT) regimen design considerations as a simple and effective way of ensuring the continued long-term effectiveness of ACTs\u201d. The Liverpool group\u2019s strong recommendation for a major change in dosing, which would have a profound effect on current and future practices if followed, was based on PK-PD modelling and was not supported by clinical trial data19. The modelling predicted that splitting current once daily ACT doses into twice per day administration would increase parasite killing enormously (by a factor of 108)18. So have treatment recommendations been wrong all these years - or is there something wrong with the modelling?Recently, based on pharmacokinetic-pharmacodynamic (PK-PD) modelling studies, it has been suggested that twice daily dosing of artemisinins could increase parasite killing4 then a dose 6\u20138\u2009hours later would also reduce numbers by 104 fold resulting in a 108 total reduction. However, similar rates of parasitaemia decline (over 48\u2009hours) are observed whether artemisinins are given once, twice or even three times in a day19. Furthermore parasite clearance is actually slower with protracted exposures to the antimalarial peroxides following slowly absorbed intramuscular artemether and slowly eliminated artefenomel than it is with parenteral artesunate which is eliminated very rapidly21. This may be considered an example of \u2018all models are wrong, but some are useful\u2019. The standard PK-PD models are useful for predicting PRR for once daily administration of artemisinin but they fail to predict the observed PRR when more than one dose is given per day. Recent modelling claims to have \u2018solved\u2019 this model misfit by hypothesising that\u00a0the splenic clearance capacity for\u00a0 infected erythrocytes\u00a0is exceeded by parasite killing by artemisinin antimalarials17. It was conjectured that this \" saturation\" of clearance capacity resulted in the accumulation of dead parasites in the circulation thereby dissociating parasite killing from parasite clearance. Based on this conjecture, it was argued that parasite clearance was an unreliable measure of drug efficacy (as the proportions of live and dead parasites could not be distinguished). Furthermore it was concluded unreservedly that \u201chuman immunity is the primary determinant of clearance rates, unless or until artemisinin killing has fallen to near-ineffective levels\u201d22 and more recently that \u201c the impact of human immunity in clearing erythrocytes containing dead or dying parasites makes parasite clearance rates highly insensitive and non-specific diagnostics of resistance\u201d23. If indeed these conjectures are all true they would deal a serious blow to current epidemiological assessments of artemisinin resistance, which rely heavily on this metric for phenotyping and for validating the parasite genotyping used in molecular surveillance28. This study examines whether evidence from previous clinical studies of the efficacy of artemisinin and its derivatives supports this hypothesized model structure and the derived therapeutic recommendations.Standard PK-PD models of antimalarial drug concentration-effect relationships generally parameterise the \u2018PD\u2019 component (parasite killing) as a sigmoid \u2013Emax relationship driven by plasma concentrations in which parasite numbers decline as a first-order process for a given drug concentration. From observed 48-hour cycle parasite reduction ratios (PRR) and measured plasma concentration profiles in artemisinin treated patients, these models imply that parasite killing rates are very high for a few hours following drug administration, and then decline rapidly as concentrations of artesunate (or artemether) and their main metabolite dihydroartemisinin all fall. It follows logically from this model construct that the effect of each dose is equivalent (in terms of PRR) when suitably spaced out in time, i.e. if the first dose reduced the number of parasites by a factor of 1028. In studies where the effects of immunity, or age as a surrogate of cumulative exposure, on parasite clearance rates have been quantitated, the effects are much smaller than the effects of artemisinin resistance30. In assessing treatment responses in artemisinin resistant falciparum malaria the hypothesis that human immunity is the primary determinant of parasite clearance rate17 does not fit the facts.The radical suggestions of the Liverpool group are not supported by clinical observations. If immunity is the primary determinant of parasite clearance rates it cannot explain why parasite clearance rates are twice as slow in patients of similar age and geographic origin, who have K13 mutant artemisinin-resistant compared with K13 wild type artemisinin-sensitive parasites18, then irrespective of effects on parasite clearance there should be substantial differences in cure rates with twice daily versus once daily administration.If parasite killing substantially exceeds splenic clearance capacity, and giving artemisinins twice daily is so much better than once daily as claimedR (version 3.1.1) using the function glmer from the lme4 package. The database of extracted study meta-data can be found in the supplementary materials.To test this hypothesis we performed a logistic meta-regression analysis of all trials with monotherapy arms containing artemisinin or a derivative. The dependent variable was cure rate, and the independent variables were duration of treatment (in days) and number of artemisinin doses. If the Liverpool group\u2019s hypothesis was correct then the number of doses given should have been a significant covariate. A further meta-regression model was run on all studies of oral artesunate with the mg/kg dose as an independent variable. The logistic meta-regressions were run with a random effect for each study (random intercept term) and fixed effects for the number of doses and duration of treatment (and dose). We fitted the model in 31. Studies were eligible if once daily or more frequent administration of artemisinin or a derivative was used as monotherapy to treat uncomplicated falciparum malaria and if cure rates at 28 days were reported for each treatment group (PCR adjusted or unadjusted). For two studies the numbers of failures by day 28 were not reported and were requested from the corresponding author.We searched the WorldWide Antimalarial Resistance Network (WWARN) Clinical Trials Publication library for eligible studies. This online resource contains all antimalarial clinical efficacy trials conducted and published since 1960In early studies where patients were kept in hospital for 28 days to assess cure rate (i.e. reinfection was not possible) the result was combined with later community based studies with PCR adjusted estimates.The search identified 40 studies, comprising 76 distinct treatment arms, which met the inclusion criteria (37 from the WWARN database and 3 from secondary searches of references in those articles). The earliest study was reported in 1984, and the most recent in 2016. The majority of studies were performed in Asia (n\u2009=\u200933) with seven reported from Sub-Saharan Africa. Twenty-five studies included participants aged 15 years or older, thirteen enrolled children and adults and for two studies this information was not available. The treatment arms were artesunate (n\u2009=\u200948) artemether (10), artemisinin (10), dihydroartemisinin (7) or beta-cyclodextrin-artemisinin complex (1) all given in a variety of doses, dosing intervals, routes and treatment course durations  and that the number of doses explained none of the residual variance . Thus for regimens of artemisinins that were sub-optimal , having more than one dose per day provided no benefit over once daily administration. To check the assumption that each dose administered in these various studies produces the maximum parasiticidal effect a logistic meta-regression was performed using only studies with the most frequently evaluated drug, oral artesunate, (n\u2009=\u200939). This found that the dose in mg/kg was not a significant predictor of efficacy (lowest doses were 1.6\u2009mg/kg). This confirms that ~2\u2009mg/kg gives almost maximum antiparasitic effects in artemisinin-sensitive 32. In studies of artemisinin monotherapies where twice or thrice daily administration has been evaluated, and in contemporary or sequential comparisons with once daily administration there is no evidence that cure rates are substantially higher with more than once daily dosing38.For antimalarial treatments with artemisinin or its derivatives there is no evidence that split dosing either accelerates parasite clearance or augments cure rates significantly. In those studies where cure rates were sub-maximal, and the conjectured \u201cdramatic enhancement of drug effectiveness\u201d should have been evident, none was found. An effect of dose frequency on treatment efficacy was observed in a small study of 43 patients in which once daily artemether-lumefantrine was compared with standard twice daily dosing. In that study PCR adjusted cure rates were 85.1% and 94.4% respectively but this difference was explained entirely by 30% lower lumefantrine levels in the former group since lumefantrine exposure is the principal determinant of cure following treatment with this ACT39. Taken together there is no clinical support for these PK-PD modelling predictions, which appear to be wrong. The fundamental problem is probably the modelling of parasite killing and clearance in falciparum malaria as a simple first order process39. Whilst the decline in parasite densities is log linear, and can therefore be described as a first order process, it seems that once daily exposures to artemisinin or its derivatives produce maximum effects in the majority of patients. The kinetics of malaria parasite killing and parasite recovery are more complex than currently modelled.Furthermore there is no evidence that\u00a0splenic clearance functions are as low as\u00a0 the values needed to sustain this hypothesisThe data are uploaded as a supplementary file.Dataset 1"}
+{"text": "Artemisinin-based combination therapy (ACT), together with other control measures, have reduced the burden of falciparum malaria in sub-Saharan countries, including Sudan. Sudan adopted ACT in 2004 with a remarkable reduction in mortality due to falciparum malaria. However, emergence of resistance to the first-line treatment artesunate and sulfadoxine/pyrimethamine (AS/SP) has created new challenges to the control of malaria in Sudan. A search for an alternative drug of choice for treating uncomplicated malaria has become inevitable. The objective of this study was to evaluate the therapeutic efficacies of dihydroartemisinin/piperaquine (DHA\u2013PPQ) and AS/SP in an area of unstable transmission in Blue Nile State, Sudan in 2015\u201316.A total of 148 patients with uncomplicated malaria were recruited in the study from November 2015 to end of January 2016. Seventy-five patients received DHA\u2013PPQ while 73 received AS/SP. Patients were monitored for clinical and parasitological outcomes following the standard WHO protocol for a period of 42\u00a0days for DHA\u2013PPQ and 28\u00a0days for AS/SP; nested PCR (nPCR) was performed to confirm parasite re-appearance from day 7 onwards.Fifty-five patients completed the DHA\u2013PPQ arm protocol with success cure rate of 98.2% (95% CI 90.3\u2013100%) and one late clinical failure 1.8% (95% CI 0.0\u20139.7%). The AS/SP showed adequate clinical and parasitological response (ACPR) of 83.6% (95% CI 71.9\u201391.8%), early treatment failure was 1.6% (95% CI 0.0\u20138.8%) and late parasitological failure (LPF) was 14.8% (95% CI 7\u201326.2%). The respective PCR uncorrected LPF was 20%.DHA\u2013PPQ is an efficacious ACT and candidate for replacement of first-line treatment in Sudan while AS/SP showed high treatment failure rate and must be replaced.The online version of this article (doi:10.1186/s12936-017-1817-9) contains supplementary material, which is available to authorized users. Malaria is endemic in Sudan and creates a burden to the health system. It has been reported that 7.5 million cases and 35,000 deaths per year were recorded before 2005 . HoweverSudan adopted artemisinin-based combination\u00a0therapy (ACT) in 2004 with artesunate and sulfadoxine/pyrimethamine (AS/SP) as the first-line of treatment, and artemether/lumefantrine (AL) as second-line \u20137. HowevPlasmodium falciparum together with a gametocidal effect. However, it has a short half-life of 50\u201360\u00a0min [Dihydroartemisinin/piperaquine (DHA\u2013PPQ) combination has been used in Southeast Asia and other countries in Africa with profound results. DHA is very effective in removal of the asexual stages of 0\u201360\u00a0min , 11. Its0\u201360\u00a0min , 12. ThiWith reports of failure and reduced efficacy of AS/SP, especially in eastern Sudan , Sudan iThis was an open-label, clinical trial according to WHO guidelines adopted by the National Federal Ministry of Health, Sudan. It is a non-randomized in vivo efficacy clinical trial conducted in two health centres in Dazamin. There was no intention of direct comparison of the patients and treatment outcomes of the two health centres.This study was conducted in two health centres  in Damazin, the capital town of the Blue Nile State in southeastern Sudan. Its population is 281,403, while the population of the entire State is over one million. Rainfall is seasonal from June to October  and malaria transmission follows this season. However Damazin has a second transmission season in December to February, similar to that reported from another area in eastern Sudan .Seventy-five patients were studied for DHA\u2013PPQ and 73 were studied for AS/SP combination therapy according to WHO guidelines  during tPatients were followed up in the above-stated days, when temperature and adverse events of treatment were recorded If parasites were not cleared or symptoms existed, a rescue treatment with AL combination of artemether and lumefantrine was administered.The data from patient sheets were entered in a predesigned Excel sheet including gender, age, temperature, parasite counts until day 42 for DHA\u2013PPQ and day 28 for AS/SP. Treatment outcomes were based on parasitological and clinical responses according the WHO guidelines  and were\u00ae Whatman paper) on day 0 and the follow-up days, dried and stored in sealed plastic bags. Samples with positive asexual parasitaemia on/or after the seventh day were subjected for molecular analysis. Briefly, DNA was extracted from day 0 and the respective days of re-appearance of parasites using the QIAamp\u00ae DNA Mini Kit  for dried blood according to manufacturer protocol. Nested PCR (nPCR) was performed for identification of P. falciparum MSP1 and MSP2 allele variants, as described previously [Blood from finger pricks was collected on filter paper  in the DHA\u2013PPQ centre and seven (10%) in the AS/SP centre. During follow-up no gametocytes were recorded in DHA\u2013PPQ group while four (5.7%) patients from the AS/SP centre had gametocytes.Table\u00a0This efficacy study has shown that DHA\u2013PPQ is effective in the treatment of uncomplicated falciparum malaria in an area of unstable transmission in Sudan. The single late parasitological failure on day 35 proved to be a recrudescence after verification by nPCR. The response to the treatment was dramatic and all slides were negative from day 1 and continued to be so to the end of follow-up. There were no complaints of adverse complications due to the treatment. There were no severe adverse events recorded during the follow up. As this is the first time the drug has been tried in Sudan there are no previous reports for comparison.Other research in neighbouring countries has shown similar effectiveness , 20. In Anopheles mosquitoes and thus raise a question of potency of artesunate as a gametocidal therapy. The success of malaria control in Sudan largely depends on the use of AS/SP and AL in the treatment of cases beside other control measures. Failure of any of the control measures, especially treatment, will jeopardize that success. The health authority has acknowledged the failure of AS/SP and the need for change with DHA\u2013PPQ.On the other hand, AS/SP has lost its effectiveness against falciparum malaria in Blue Nile region. Besides the single case ETF, 20% of the patients showed late parasitological failure and four patients had gametocytes on days 21 and 28 of the follow-up (Table\u00a0Sudan is on its way to changing its first-line malaria treatment and the proposed DHA\u2013PPQ is an ideal alternative. The policy makers at the Federal Ministry of Health have decided to choose AL as first-line malaria treatment and DHA\u2013PPQ as a second-line malaria treatment. A decree has been issued to this effect by the Federal Minister of Health, Sudan on March 9th, 2017. This study has shown that DHA\u2013PPQ would suitable as a first-line treatment in an area of unstable transmission in Sudan, but the Ministry of Health has chosen it as second-line. AS/SP has lost its place as first-line therapy for uncomplicated falciparum malaria and the results of this study justify its replacement.Additional file 1. DHA-PPQ treatment group.Additional file 2. AS-SP treatment group."}
+{"text": "Plasmodium falciparum malaria and assesses the limitations of the current practices. Alternative approaches are explored and their implementation is discussed using example data from a large multi-site study.The Kaplan\u2013Meier (K\u2013M) method is currently the preferred approach to derive an efficacy estimate from anti-malarial trial data. In this approach event times are assumed to be continuous and estimates are generated on the assumption that there is only one cause of failure. In reality, failures are captured at pre-scheduled time points and patients can fail treatment due to a variety of causes other than the primary endpoint, commonly termed competing risk events. Ignoring these underlying assumptions can potentially distort the derived efficacy estimates and result in misleading conclusions. This review details the evolution of statistical methods used to derive anti-malarial efficacy for uncomplicated The online version of this article (doi:10.1186/s12936-017-2074-7) contains supplementary material, which is available to authorized users. Despite a rich pharmacopeia of anti-malarial agents, the emergence and spread of anti-malarial drug resistance has been relentless, with resistance now documented to all recommended treatment regimens in widespread use. Efficacy estimates derived from in vivo clinical trials, including post marketing and surveillance studies, form the basis for monitoring the status of anti-malarial drug resistance, with in vitro drug susceptibility testing and molecular analyses providing important complementary and confirmatory information. The main measure of anti-malarial clinical efficacy is the risk of recrudescent infection, which is defined as recurrent parasitaemia with parasites that were present prior to the initiation of treatment. Recrudescence needs to be differentiated from reinfections arising from inoculation with a new parasite strain during the follow-up period. New infections can be either from the same parasite species or a different one Fig.\u00a0 1].Fig..Fig.1].FRecrudescent parasitaemia arises from a variety of factors which can be broadly categorized into those related to the host, the parasite or the drug Fig.\u00a0. PharmacAnti-malarial efficacy needs to be monitored routinely in endemic areas, so that early indications of drug resistance are recognized and malaria control activities can be revised accordingly . DefininIn this review, the evolution of the methods for defining anti-malarial drug efficacy since the 1960s and the key statistical approaches currently available are documented. Challenges associated with these statistical methods and how they apply to stand-alone efficacy trials and comparative drug studies are discussed.Plasmodium falciparum malaria in 1965, primarily to monitor chloroquine resistance which had been identified a few years earlier [The WHO released its first standardized protocol for assessing in vivo efficacy against  earlier , 16. Thi earlier . These e earlier . The lat earlier . The rec earlier . Whilst Two approaches have been used in deriving efficacy estimates in anti-malarial efficacy studies: (i) the calculation of the proportion of patients cured within a specified period of follow-up (this proportion is often referred to as the \u201ccure rate\u201d) and (ii) survival analysis, which provides a cumulative probability of cure. The term \u201ccure rate\u201d statistically speaking is misleading since this is not a rate, but a point estimate of risk at a predefined time point. The proportion cured is usually estimated using a per-protocol (PP) and intention to treat (ITT) approaches. In PP approach, the proportion cured is derived from all patients followed until treatment failure or a set period of time, excluding those with protocol deviations, those who develop new infection or who are lost to follow-up. Whilst relatively easy to calculate, this approach ignores valuable information provided by the patients who experience protocol deviations or are lost to follow-up. Patients failing treatment are more likely to become symptomatic and seek retreatment and thus be detected passively. Whereas patients who are cured are more likely to tire from active detection and be lost to follow up\u2014these attrition biases result in an underestimation of treatment efficacy . In a moSurvival analysis using Kaplan\u2013Meier (K\u2013M) method provides an alternative strategy. This approach maximizes the information available from each patient, thereby increasing the precision of the estimates , 13. TheSeveral reports have compared the use of PP approach and K\u2013M survival analysis in deriving anti-malarial efficacy. Guthmann et al. pooled datasets from 13 trials (n\u00a0=\u00a02576) to examine the discrepancies in derived estimates when PP and K\u2013M approach were used . OverallSince the K\u2013M approach was being increasingly recommended for deriving efficacy estimates, Price et al. provided a classification table for different possible outcomes . However, patients can experience new infections with P. falciparum or other species such as Plasmodium vivax during the ensuing weeks  and the time point of interest The K\u2013M method makes a fundamental assumption of independent (non-informative) censoring, i.e. patients who are censored have the same risk of observing the outcome as those who are still being followed-up. When a patient experiences new infections, censoring is no longer non-informative (as they will be retreated) and in such situations the use of K\u2013M leads to an upwards biased estimate of treatment failure , 27, 28.mentclass2pt{minimcmprsk package in R software  and present an overall K\u2013M estimate without considering the multi-centric nature of the data, an approach recommended by Srinivasan and Zhou provided the data from several sites (studies) are independent . Howeversoftware .The meta-analytic approach proposed by Comberscure et al. has been illustrated with the example dataset on AS\u00a0+\u00a0AQ arm . The K\u2013MTo investigate suitable alternative treatment regimens, a comparative randomized clinical trial is required and these comparative trials raise further difficulties in the analyses and interpretation of the data.Anti-malarial drugs with different elimination half-lives may vary considerably in the period of time during which peripheral parasite growth is suppressed. Since post treatment prophylaxis reduces the risk of new infection and delays the timing of recurrent parasitaemia, for drugs with similar efficacy the comparative results may be biased against the drug with the shorter half-life. Conversely, a new drug which has a long elimination half-life may result in the false impression of good efficacy. Consider dihydroartemisinin\u2013piperaquine (DP) and artemether\u2013lumefantrine (AL). Piperaquine is a slowly eliminated regimen with a terminal elimination half-life of 13.5\u201328\u00a0days and will suppress parasitic growth for a far longer period compared to lumefantrine, which has an elimination half-life ranging from 1.4 to 11.5\u00a0days .Fig.\u00a05Cu8 parasites) in 7 parasite life cycles, which is approximately 14\u00a0days assuming an efficient multiplication of tenfold per cycle and a parasitic developmental cycle of 48\u00a0h. In this scenario recrudescences begin to appear by day 33. Under the same assumption, a drug which provides more prolonged prophylaxis  will result in recrudescent parasites reaching the limit of detection on day 45. Hence, comparing these two drugs on day 42 will result in a biased conclusion. The comparative efficacy should account, therefore, for differences in the pharmacokinetic profile of the drugs. This is further confounded by transmission setting, which determines the risk of new infection (a competing risk event). Characterization of the duration of follow-up required to appropriately capture the treatment failures would provide a basis for comparison and this is currently being investigated [Ideally, the comparison overall efficacy of anti-malarial regimens should be carried out at a time, when anti-malarial drug concentrations cease to suppress parasite growth  and after allowing the time for the parasites to reach the limit of detection. If the drug concentrations fall below MIC on day 18, and assuming 10 parasites are circulating in the blood, these parasites will reach the threshold for detection  it appears to be more relevant to focus on the overall proportion of failures observed during the follow-up time. Consequently, an alternative approach test is needed which allows comparison of the cumulative Kaplan\u2013Meier estimates at a specific time point. Such a test can be constructed from the difference of complementary log\u2013log transformed K\u2013M estimates at a specified time point and the appropriately estimating the standard error for this difference .X2 test statistic for the comparing the difference in two estimates is given by [Var(\u03b4) denotes variance estimate of \u03b4   can be tested using the K\u2013M estimates. Suppose \u0394 is the margin for non-inferiority. The 95% CI for the difference of two K\u2013M estimates can be constructed by adding individual variance of the two K\u2013M estimates . For antt. Let \u03b3 be the corresponding relative risk of failure for new treatment compared to the standard regimen (estimated as hazards ratio from Cox\u2019s model) and let Under the proportional hazards assumption, the non-inferiority between regimens can be tested based on the relative risk measure (e.g. hazards ratio). Let  hazards ,\\documenThe upper limit for non-inferiority based on relative risk then can be derived based entirely on the efficacy of the reference (standard) arm as:t. Logarithm of the hazard ratio corresponds to the difference between K\u2013M survival estimates on the complementary log\u2013log scale. The 95% confidence interval  for this difference can be calculated using expression in Additional file Non-inferiority is demonstrated if the 95% CI for the estimated hazards ratio remains below the non-inferiority limit on the relative risk scale . FurtherSignificant resources have been spent to overcome recognized difficulties in estimating anti-malarial efficacy by standardizing methodological procedures and these have facilitated the monitoring of anti-malarial drug resistance over time and geographical location. However, even a robust design and analysis requires that the derived point estimates of efficacy are interpreted with caution and the confidence interval around such point estimates should be given equal importance when interpreting the result of a study. The study design, transmission setting, laboratory and genotyping procedures, patient demographics and adherence to protocols all need to be considered. Uncertainty associated with PCR genotyping can be ameliorated to a certain extent if allelic distribution and clonality of infection in the study population is known. This allows for adjustment of pre and post treatment alleles matching purely by chance; various modelling approaches have been proposed for estimating the haplotype frequencies and for adjusting drug efficacy estimates and novel genotyping approach has been recently suggested , 52\u201356."}
+{"text": "Pivmecillinam is also one of the first-line drugs recommended in the international guidelines for LUTIs . The international recommended duration is based on evidence saying that a 7-day regimen is better than a 3-day regimen. However, no data says that a 5-day regimen is superior to a 3-day regimen. With this study we aim to identify and to compare the efficacy of pivmecillinam 400\u00a0mg\u00a0t.i.d in a 3-day respectively 5-day regimen, against community acquired uncomplicated LUTI, i.e., in women at the age of 18\u201370 year old.Uncomplicated lower urinary tract infections (LUTI) are very common, and presumably around 200,000 female patients are treated for this annually in Denmark. The current Danish national clinical practice guidelines recommend pivmecillinam as a first-line drug  will be given to the patient. We aim for 161 evaluable patients in each arm.The general practitioner will at consultation give a suitable patient the opportunity to participate in the study. If the patient will give her consent, a double-blinded kit (i.e.Pivmecillinam is an excellent choice against urinary tract infections and we believe this study will fill in the gaps and strengthen the evidence on the treatment against one of the most common infections in our society. Thus, aiming to provide a more rational and ecological beneficial antimicrobial therapy.2014-001321-32. EudraCTno.:  Escherichia coli  / \u2265104 bacteria/ml [other uropathogenic bacteria]) the isolates will also be examined for resistance mechanisms and patterns, minimal inhibitory concentration (MIC) to mecillinam  and determining the bacterial specimen . All urine samples will processed according to laboratory routine and susceptibility tested according to EUCAST guidelines [One pre-treatment urine sample (day 0) with optional urine-dipstick, and two control urine samples  will be sampled from the patient. The three urine samples will be sent to the DCM, Hvidovre University Hospital, Denmark, for examination of the presence and count of uropathogenic bacteria. In presence of significant bacteriuria  to the included patients to follow-up on the questionnaires and control urine-samples, as well as answering any questions the patients might have. The phone calls or text-messages are app. made on day 3\u20135, day 9\u201311 and day 28\u201330, respectively. However, not at any time will we try to increase the adherence to the treatment, since the study ought to be mirroring a realistic patient situation.If a patient during the study would develop signs of upper UTI (i.e. pyelonephritis), treatment failure, serious adverse effects or allergic reactions to the medicine, or decision of withdrawal, she will have to terminate from the study and possibly get a new evaluation by a physician.Patients will be told at recruitment to visit the GP again if she develops worsening of symptoms or not experiencing any symptom relief during the study period. In that case the GP is able to choose a new treatment strategy for the patient and will have the potential microbiological diagnostic answers (i.e. from pre-treatment urine sample) to take in consideration in this approach.Co-primary endpoint no. 1 \u2013clinical efficacy .Co-primary endpoint no. 2 \u2013 bacteriological efficacy .We have two co-primary endpoints:Clinical signs and symptoms in uncomplicated LUTIs in women.Frequency and timing of relapse of symptoms and/or bacteriuria.Bacteriological findings (i.e. ESBL producing bacteria).Complications (i.e. urosepsis and/or pyelonephritis) within 28\u00a0days.Clinically cured is defined as cumulative symptom score of <2.Clinical relapse is defined as initial clinically cured followed by cumulative clinical score of \u22652 at day 28.2 fold decrease in uropathogenic bacteria) or potentially full eradication of uropathogenic bacteria (i.e. sterile culture or asymptomatic contaminations of non-uropathogenic bacteria).Bacteriologically cured is defined as significant reduction  analysis will be consisting of all included patients (i.e. patients that present with symptoms of uncomplicated LUTI and are included in the study). The per-protocol (PP) analysis will be consisting of all the patients that follow-up with clinical and bacteriological data (i.e. excluding the dropouts). The need-to-treat (NTT) analysis will be stratified from the PP analysis to only include patient with true uncomplicated LUTI .p\u2009<\u20090.05, and a statistical power of 0.8, we will need 113 patients in each group for the NTT analysis. With an estimated fall-out rate of app. 30% between each arm; 161 patients per group will need to be included for the PP analysis, and 230 in each group for the ITT analysis. There is a possibility for an interim analysis after 300 included participants, since we cannot reject a possibility for a higher (i.e. 20%) absolute efficacy.We hypothesize that there is a 15% absolute differences in efficacy between the groups, measured in clinical efficacy. We believe there to be an even bigger difference in bacteriological efficacy. Assuming an efficacy of 90 and 75% for 5\u00a0days and 3\u00a0days therapy, respectively, using a double-sided test with The source data is the GP\u2019s and patient\u2019s questionnaires and the diagnostics answers from the microbiological lab at the DCM. All source data will all be stored by the investigator and sponsor, in originals and scanned copies, at the DCM. The collected source data are transferred, logged and processed in the computers of the DCM. Analysis will be made in ITT, PP, and NTT form, respectively. All data will be analyzed coded, and only de-coded after the analysis is done and the analytic results are written in the relevant sections in the future article. The statistical software packages SAS  will be used for all analyses. Categorical data will be analyzed using a Fisher\u2019s Exact test.Results  will be presented at scientific meetings, and in scientific publications in accordance to CONSORT 2010 statement .The study will comply with all the requirements to secure patients safety and to be carried out according to the Declaration of Helsinki, national laws and regulations. Ethical approval has been obtained by the Danish committee on biomedical research ethics for the capitol region of Denmark (No. H-4-2014-072). Approvals have also been obtained from the Danish National Health and Medicines Authority and the Danish Data Protection Agency. The study will be conducted according to the principle of Good Clinical Practice (GCP) and monitored by the GCP Unit of Bispebjerg Copenhagen University Hospital, Denmark. All patients will be included in concordance with rights of a research subject in clinical research, according to the national committee of health and research ethics document, which each subject will be provided the option to read before deciding to participate. The informed consent is obtained at the GP office, where the participant also will be informed about potential risks, benefits and procedures etc. based on an information sheet that the central investigators have prepared in concordance with above mentioned committees\u2019 standards. The patient can withdraw their consent at any time without disadvantages.The study will comply with the Danish regulation about management of personal information by the act on processing of personal data (Act No. 429 of 31 May 2000). All patient data will be carefully processed and stored at the Department of Clinical Microbiology, Hvidovre University Hospital, Denmark. All data will be anonymous in any possible future publication.resistant to mecillinam  and/orPseudomonas spp., Acinetobacter spp., or Enterococcus spp. and other Gram positive uropathogenic bacteria etc.a specimen to which pivmecillinam is not recommended for various reasons  - i.e. The only time during the study in which we will intervene with antimicrobial therapy is if we during microbiological diagnostics find out that the patient is receiving a possibly ineffective therapy. This will for example be the case if we find causing uropathogenic bacteria to be:We will then make contact and recommend another more suitable antimicrobial therapy.Pivmecillinam has been proven in RCTs to be safe and effective in the treatment of UTIs , 8, 20. Pivmecillinam is an excellent choice against urinary tract infections, and has shown to be both safe and effective against UTI , 12, 22,Ferry et al.  have demt.i.d., since that is the dose we recommend in Denmark; and we believe 200\u00a0mg\u2009t.i.d to be an insufficient dose (i.e. supported by pharmacokinetic mouse models [. [We have chosen the 400\u00a0mg dose e models  and the odels [. ). With tWe designed this protocol similarly to previous similar clinical studies on uncomplicated LUTI , 24, 25,We want the inclusion procedures to be as realistic as possible. Therefore, the consulting with the GPs will be changed as little as possible and restricted to the inclusion day. However, patients will be clearly instructed to return to the GP as usual at sign of treatment failure, or if they wish to consult for any other reason.The primary outcome we want to focus on is the clinical effect, since that is of most importance to the patient. However, it is important to know that the bacteria disappear and not cause clinical relapses in the future; therefore, we included two follow-up urine sample at day-4 and day-28, respectively.This study will give us the knowledge of what drug therapy of pivmecillinam that is most effective and sufficient. If the 5-day therapy will show to be significantly superior to the 3-day regimen, the current guidelines will have to be changed in best interests of the patients. Also through the perspective of the patients, it will be beneficial if the 3-day regimen is proven to be non-inferior to the 5-day regimen , and we can keep using the 3-day regimen in concordance with the principles of evidence-based medicine.We believe this study will fill the scientific gaps and provide sufficient evidence for the most efficient dose duration of pivmecillinam against uncomplicated lower urinary tract infections. Thus, strengthening the guidelines for one of the most common community acquired infections in our society."}
+{"text": "Malaria is a major cause of morbidity and mortality in many African countries and parts of Asia and South America. Novel approaches to combating the disease have emerged in recent years and several drug candidates are now being tested clinically. However, it is long before these novel drugs can hit the market, especially due to a scarcity of safety and efficacy data.To reduce the malaria burden, the Medicines for Malaria Venture (MMV) was established in 1999 to develop novel medicines through industry and academic partners\u2019 collaboration. However, no reviews were focused following various preclinical and clinical studies published since the MMV initiation (2000) to till date.We identify promising approaches in the global portfolio of antimalarial medicines, and highlight challenges and patient specific concerns of these novel molecules. We discuss different clinical studies focusing on the evaluation of novel drugs against malaria in different human trials over the past five years.The drugs KAE609 and DDD107498 are still being evaluated in Phase I trials and preclinical developmental studies. Both the safety and efficacy of novel compounds such as KAF156 and DSM265 need to be assessed further, especially for use in pregnant women. Synthetic non-artemisinin ozonides such as OZ277 raised concerns in terms of its insufficient efficacy against high parasitic loads. Aminoquinoline-based scaffolds such as ferroquine are promising but should be combined with good partner drugs for enhanced efficacy. AQ-13 induced electrocardiac events, which led to prolonged QTc intervals. Tafenoquine, the only new anti-relapse scaffold for patients with a glucose-6-phosphate dehydrogenase deficiency, has raised significant concerns due to its hemolytic activity. Other compounds, including methylene blue  and fosmidomycin (DXP reductoisomerase inhibitor), are available but cannot be used in children.At this stage, we are unable to identify a single magic bullet against malaria. Future studies should focus on effective single-dose molecules that can act against all stages of malaria in order to prevent transmission. Newer medicines have also raised concerns in terms of efficacy and safety. Overall, more evidence is needed to effectively reduce the current malaria burden. Treatment strategies that target the blood stage with transmission-blocking properties are needed to prevent future drug resistance.The online version of this article (doi:10.1186/s40249-016-0196-8) contains supplementary material, which is available to authorized users. Please see Additional file Plasmodium, transmitted through the bite of the female Anopheles mosquito. It is a major public health problem in many endemic countries including Sub-Saharan Africa (SSA); in 2015, an estimated 438 000 malaria deaths were reported globally ) and P. vivax (median EC50\u2009=\u20090.51 nM [range 0.25\u20131.39 nM]) ) . DDD1074P. falciparum and P. vivax strains in the liver, asexual erythrocytic, and transmission stages. One recently published paper reported a KAF156 Phase II proof-of-concept trial [KAF156 , a promising chemoprevention molecule, is a cyclic amine resistance locus inhibitor (PfCARL) developed by the Novartis research consortium . In vitrpt trial  conducteP. falciparum and P. vivax parasites is being assessed in adult patients using a single dose treatment (400\u00a0mg) [DSM265, a dihydroorotate dehydrogenase (DHODH) inhibitor acting against the liver (schizont formation) stage, is proving to be promising as a one-dose (400\u00a0mg) malaria cure in a Phase I trial in healthy volunteers, with an encouraging safety profile. DSM265 is currently in the clinical developmental stage (Phase II) in Peru NCT02123290). Its activity against uncomplicated 290. Its In most countries in SSA, malaria in pregnancy contributes to significant maternal and perinatal mortality. It is not recommended to use ACTs during the first trimester due to side effects observed in preclinical models . CurrentPlasmodium species at different stages of infection. Another similar phenotypic molecule known as MMV390048, developed by researchers at the University of Cape Town in South Africa, targets lipid phosphatidyl inositol 4-kinase (PfPI4K) [pfcrt is associated with chloroquine resistance. Additional polymorphisms (dhfr and dhps) for sulfadoxine-pyrimethamine and polymorphism of P. falciparum multidrug resistance protein 1 (pfmdr1) are associated with resistance to chloroquine, mefloquine, quinine, and artemisinin [P. falciparum ap2-mu (Pfap2-mu) homologue [pfmdr1, and sarco-endoplasmic reticulum calcium ATPase6 (PfSERCA) [pfmdr 1, pfcrt, and pf3d7-1343700 Kelch propeller (K13-propeller) mutations are potential markers indicating that P. falciparum is developing resistance to artemisinin and its derivatives [Several molecules are currently being tested in preclinical models. Examples include SJ557733, developed in collaboration between St. Jude Children\u2019s Research Hospital, TN, USA and Rutgers University, NJ, USA , and PA2(PfPI4K) . The MMVemisinin . Novel lomologue , gene muPfSERCA)  may be aivatives , 33.TablP. falciparum malaria was shown to be not as effective as artemisinin. This was indicated by the reduced parasitic clearance on day 28 after seven days (60\u201370\u00a0%) compared to artesunate dose\u2013response (95\u00a0%) [P. falciparum malaria in India and received mark approval from the Drug Controller General of India. It was subsequently put on the market in seven African countries [Plasmodium [Quinine, first used in Europe in the 17th century, chloroquine , and 4-ae (95\u00a0%) . Thus, iountries . Due to ountries . The resasmodium .P. falciparum and P. vivax malaria. Due to the reduced elimination half-life of 46\u201362 h, a single dose of OZ439 alone or in combination with piperaquine can eliminate 98.0\u00a0% of P. falciparum and 99.6\u00a0% of P. vivax within 36\u00a0h. Artefenomel has demonstrated a higher parasitic clearance within the first 24\u00a0h in P. vivax patients as compared to P. falciparum patients (30\u201336 h). However, gametocyte clearance was 100\u00a0% in patients who were administered 1 200\u00a0mg of artefenomel within 48\u00a0h [Different doses of artefenomel (200\u20131 200\u00a0mg) were tested in a Phase IIA exploratory, open-label trial and revealed promising safety and efficacy profiles among Southeast Asian adults with uncomplicated P. falciparum and P. vivax in the first trimester, with no risk of spontaneous abortions or major congenital malformations [One of the major concerns about the use of OZ compounds is that they have a similar endoperoxide structure to artemisinin, indicating possible treatment failures. Previous data suggest that artemisinin derivatives are associated with the risk of spontaneous abortions in early pregnancies , but recrmations . Similarrmations . No clinP. falciparum and multidrug resistant strains [P. falciparum malaria planned for Western Cambodia (NCT00936767) has been withdrawn for unknown reasons. Some studies reported neurological and auditory side effects such as ataxia and slurred speech [50s correlation with pfmdr1 Y184F mutations, which potentially reduces sensitivity to artemisinin-resistant strains and contributes to emerging ACT resistance [P.falciparum phosphatidylinositol-3 kinase (PfPI3K) [Two interesting endoperoxides from artesunate derivatives including artemisone BAY 44\u20139585) and tetraxoane (TDD E209), are examples of other synthetic candidates currently under development. Artemisone is a semi-synthetic second-generation artemisinin derivative developed in collaboration between Bayer HealthCare Pharmaceuticals in Germany and the Hong Kong University of Science and Technology. Results of preclinical studies are highly promising as compared to other novel artemisinins. Artemisone is more effective than artesunate against  strains , 41. Dosd speech , 43 due \u20139585 and(PfPI3K) .Plasmodium strains when compared to artesunate. Several preclinical studies have shown its benefits, particularly for treating patients infected with chloroquine-, amodiaquine-, and mefloquine-resistant malaria strains [P. falciparum and P. vivax malaria at the multicenter level. One study has been completed (NCT00988507) but no results are yet available. Most recently, a ferroquine-artesunate dose-ranging Phase II trial on P. falciparum-infected adults and children in eight African hospitals was conducted [CI: 90\u2013100) after treatment with 2\u00a0mg/kg ferroquine combined with 4\u00a0mg/kg artesunate. However, the cure rate was reduced  when ferroquine monotherapy 4\u00a0mg/kg/day for 3-days regimen was used. Furthermore, exacerbated malaria symptoms were observed in 14\u00a0% of the individuals in the treatment cohort.Ferroquine is an ameliorated blood-schizonticidal 4-aminoquinoline developed by Sanofi-Aventis. Along with OZ439, it is a more effective parasite-killing compound against  strains \u201348. The onducted . The resAnother 4-aminoquinoline derivative called AQ-13 (Ro47-0543), a similarly structured chloroquine with a modified propyl side chain from the aminoquine panel, was developed in collaboration between Tulane University and Louisiana State University both located in LA, USA. Preclinical studies have indicated increased efficacy of AQ-13 when compared to other derivatives . Phase IP. falciparum, while fewer studies evaluate treatments against P. vivax and P. ovale malaria. A deficiency of glucose-6-phosphate dehydrogenase (G6PD) is a hereditary enzyme defect condition that causes episodic hemolysis. Patients with a G6PD deficiency are common in malaria-endemic countries and are at high risk of hemolysis due to treatment with antimalarial drugs . These patients are generally not included in trials due genotypic variations. For these individuals, tafenoquine (WR 238605) is a good alternative drug. It is an 8-aminoquinoline derivative and has a similar mode of action to primaquine against hypnozoites, gametocytes, and liver stages [P. vivax malaria. Tafenoquine with chloroquine was tested in studies against P. vivax malaria. In a Phase IIB dose-ranging trial, different doses of tafenoquine alone  or in combination with 15\u00a0mg primaquine for 14\u00a0days were tested, with a fixed dose of chloroquine for three days. A single dose of tafenoquine (300\u00a0mg) co-administered with chloroquine was shown to prevent relapse in 89.2\u00a0% (95\u00a0% CI: 77\u201395) of people as compared to chloroquine alone  during the first six months of follow-up [P. vivax patients for radical cure showed that single-dose tafenoquine (300\u00a0mg) combined to chloroquine is more efficacious in preventing relapses as compared to chloroquine alone, with a similar safety profile. Based on these observations, GSK and MMV announced two new Phase III studies: 1) a DETECTIVE study (TAF112582) to evaluate the efficacy, safety, and tolerability of tafenoquine co-administered with chloroquine as a radical cure for P. vivax malaria ; and 2) a GATHER study (TAF 116546) to assess the incidence of hemolysis and the efficacy and safety of tafenoquine over primaquine [The majority of clinical trials focus on malaria caused by r stages . Tafenoqollow-up . Recent y TAF 1166 to asseP. falciparum isolates [P. falciparum malaria. Although results were promising, adverse effects were reported including vomiting, as well as the discoloration of urine, mucous surfaces, and teeth [P. falciparum malaria. This combination showed poor efficacy (71\u00a0%) when compared to the control group  [P. falciparum transmission. A Phase I trial testing the combination of methylene blue with primaquine is currently registered in the clinicaltrials.gov database (NCT01668433), but results are not yet available.A century ago, the German scientist Paul Ehrlich discovered the antiplasmodial activity of methylene blue . The cheisolates . Methylend teeth . Drug rend teeth . In 2006nd teeth . In 2011e; 85\u00a0%) . HoweverStreptomyces lavendulae bacterial isolates. This compound inhibits the non-mevalonate pathway , essential for the synthesis of parasite isoprenoids [P. falciparum malaria. On day 28, recrudescence was observed in seven out of nine patients, indicating monotherapy failure [P. falciparum malaria. Poor efficacy was observed due to poor immunity in children aged between one an>d two years [P. falciparum malaria, aged between one and 60\u00a0years and with a body weight between 5 and 90\u00a0kg, is currently registered in the clinicaltrials.gov database (NCT02198807). Overall, studies indicated that fosmidomycin is only effective for short-term treatment. Studies on finding a potential partner drug to prove the efficiency of fosmidomycin urgently need to be conducted.Isoprenoids are derived from the mevalonate pathway in humans, an essential metabolic pathway for parasite synthesis. Jomaa Pharma GmbH developed a synthetic antibiotic agent called fosmidomycin derived from prenoids . Fosmido failure . Fosmidowo years . Two addwo years . HoweverP.falciparum and protein inhibition in blood- and liver-stage parasites could be ideal for future drug development.In this review, we summarized the different approaches tested over the years to control the malaria pandemic, and possibly reduce global malaria incidence and mortality by 90\u00a0% before 2030. Novel chemotherapeutic approaches have emerged over the past five years, with promising results. Nevertheless, the efficacy and safety of these drugs need to be studied further. These novel antimalarial approaches are multifaceted, thus there is an urgent need for effective single-dose molecules to act during the liver and blood stages of malaria. Effective compounds should be developed before global emergence of resistance to artemisinin derivatives and 4-aminoquinoline. There is currently no low-dose primaquine regimen for pediatric use. Novel blood-stage compounds such as DDD107498 and tafenoquine should focus on blocking parasite transmission in children and adolescents, and pregnant women. Molecules such as ferroquine should be combined with a potential partner drug to enhance efficacy. Additional challenges in preventing the relapse of malaria episodes include hemolysis in patients with a G6PD deficiency, treatment for drug-resistant strains, pediatric dosing, serious drug-drug interactions, transmission blocking, radical cure, and relapse prevention. Potentially targeting mitochondrial electron-transport chain of"}
+{"text": "Leishmania parasite resides and replicates within host macrophages during visceral leishmaniasis (VL). This study aimed to evaluate neopterin, a marker of macrophage activation, as possible pharmacodynamic biomarker to monitor VL treatment response and to predict long-term clinical relapse of VL. Following informed consent, 497 plasma samples were collected from East-African VL patients receiving a 28-day miltefosine monotherapy (48 patients) or 11-day combination therapy of miltefosine and liposomal amphotericin B . Neopterin was quantified with ELISA. Values are reported as median (inter-quartile range). Baseline neopterin concentrations were elevated in all VL patients at 98.8 (63.9\u2013135) nmol/L compared to reported levels for healthy controls (<10 nmol/L). During the first treatment week, concentrations remained stable in monotherapy patients (p = 0.807), but decreased two-fold compared to baseline in the combination therapy patients (p < 0.01). In the combination therapy arm, neopterin concentrations increased significantly 1 day after L-AMB infusion compared to baseline for cured patients , but not for relapsing patients . The neopterin parameter with the highest predictive power for VL relapse was a higher than 8% neopterin concentration increase between end of treatment and day 60 follow-up (ROC AUC 0.84), with a 93% sensitivity and 65% specificity. In conclusion, the identified neopterin parameter could be a potentially useful surrogate endpoint to identify patients in clinical trials at risk of relapse earlier during follow-up, possibly in a panel of biomarkers to increase its specificity.The  Leishmania parasite. VL is affecting mostly the poorest of the poor and remains a devastating neglected tropical disease with high morbidity and mortality, with over 200,000 new cases and over 20,000 deaths annually  is a systemic disease caused by the Parasite recrudescence and clinical relapse occur in a relatively large proportion of VL patients  to produce free radicals that kill the intracellular Leishmania donovani parasites. L. donovani infection causes an increase in monocyte load in the infected organs  and 5.34 \u00b1 2.7 nmol/L (n = 359) for children (<18 years) and adults, respectively  are 6.78 \u00b1 3.6 nmol/L  and miltefosine.Neopterin concentrations were determined as part of a randomized multicentre trial (registered as NCT01067443) assessing the safety and efficacy of different VL treatments in Eastern Africa. The clinical results and pharmacokinetic analysis have been published elsewhere  and Sudan (Institute of Endemic Diseases) prior to the start of the trial in each country , spleen aspirates , or bone marrow samples . Samples originated from patients receiving either a 28-day 2.5 mg/kg/day miltefosine monotherapy (48 patients), or a combination treatment of one dose 10 mg/kg L-AMB on day 1 of treatment, followed by a 10-day 2.5 mg/kg/day miltefosine treatment (48 patients).Patients that required rescue treatment during treatment or patients who had a fatal outcome before the end of treatment were indicated as \u201cinitial treatment failure.\u201d Final cure was determined at 6 months after end of treatment (day 210). Patients indicated as \u201crelapse\u201d were cured at the end of treatment, but received rescue treatment within 6 months after treatment due to reappearance of VL clinical signs and symptoms and parasite recrudescence confirmed by microscopy.To decrease the invasiveness of sampling for patients, neopterin concentrations were quantified in the same samples collected for miltefosine pharmacokinetic analysis , following the manufacturer's instructions. Two calibration curves  were included in every analysis together with two quality control samples in duplicate. Samples above the upper limit of quantitation were reanalyzed in a 10x dilution with a dilution buffer provided by the manufacturer. The optical density (OD) was measured at 450 nm by an Infinite\u00ae M200 Microplate Reader . The OD values were converted to neopterin concentrations from the standard curve using a 4 parameter non-linear logistic regression model in Prism .Incurred sample reanalysis was performed for 4% of all samples. The acceptance criterion was adapted from FDA guidelines for bioanalytical method validation, and stated that at least two-thirds of the analyzed concentrations should be within 20% deviation of the initially analyzed concentration . As incurred sample reanalysis was performed >1.5 years after initial analysis for a proportion of samples, these results were used to assess the influence of long-term storage on neopterin quantification.Data cleaning and interpretation was performed with R (version 3.1.2) and packages \u201cggplot2,\u201d \u201cHmisc,\u201d and \u201cplyr.\u201d All values are reported as the median . In the display of results, nominal time points are depicted instead of actual time points.t-test on log-transformed data was used when comparing groups, unless indicated otherwise.Absolute neopterin concentrations and relative concentration changes over time\u2014during and after treatment\u2014were evaluated for their ability to reliably discriminate between cured and relapsed patients. In statistical comparisons, absolute and log-transformed data were checked for normality and equal variances. The two-sided Subsequently, a logistic regression was performed in R to evaluate the significance of the evaluated neopterin parameter as a predictor of clinical outcome. Finally, receiver-operating characteristic (ROC) curves were generated with the R package \u201cpROC.\u201d The interplay between sensitivity and specificity of neopterin as biomarker in isolation was interpreted and the optimal cut-off value was determined with the same package.n = 4).A total of 497 plasma samples were available from 96 patients; 48 patients in combination therapy and 48 patients in monotherapy. In both treatment arms, two patients experienced initial treatment failure. Six patients in the combination therapy arm and nine patients in the monotherapy arm relapsed within 6 months after treatment. Of patients experiencing treatment failure, samples were only included up to the day they received rescue treatment; subsequent samples were omitted (n = 15) received rescue treatment at day 112 , approximately 3 months after treatment.Patient characteristics are depicted in Table During treatment, all actual sample collection time points were within \u00b115% of the nominal time points. During follow-up, the variability in actual sample collection time points was larger, with day 60 samples collected between day 54\u2013157 and day 210 samples between day 185\u2013345. Nonetheless, >85% of samples were collected within \u00b115% of the nominal time point during follow-up.n = 12) of samples analyzed >1.5 years after initial analysis (11 out of 12 within \u00b120% deviation). This indicates adequate stability of neopterin in plasma for at least 1.5 years when stored at \u221220\u00b0C.For all runs, quality control samples were within the acceptable range according to ELISA kit specifications. Incurred sample reanalysis was found to be acceptable . Incurred sample reanalysis was also acceptable for the subset (n = 46) at 98.8 nmol/L (IQR 63.9\u2013135)  trend toward higher neopterin baseline levels in monotherapy patients cured at the end of treatment  compared to patients requiring rescue therapy during or within 6 months after treatment . There were no significant differences in baseline neopterin levels between age categories (adult/child), country and gender .Baseline neopterin concentrations were elevated in all monotherapy VL patients , while neopterin levels remained unchanged in patients receiving monotherapy with a concentration of 91.3 nmol/L . Interestingly, for both treatment arms, day 210 neopterin concentrations were still elevated  compared to the reported healthy control levels of <10 nmol/L.Neopterin concentrations regressed during treatment to comparable end of treatment values of 33.6 nmol/L  and 21.9 nmol/L . There was, however, a difference in the rate of neopterin decline between the two treatment arms . This was not observed for combination therapy patients who eventually failed treatment or relapsed . Despite the significantly higher day 2 neopterin concentrations in cured patients, this parameter is not a significant predictor of final cure (p = 0.0853). The ROC AUC , but not for the monotherapy arm alone. ROC curves of these parameters are depicted in Figure The day 60 neopterin concentration was a significant predictor of relapse in both arms combined and in the combination therapy arm (n = 14) experienced a significantly higher neopterin concentration increase during the first month of follow-up  compared to patients that remained cured  . For patients that relapsed, there was no correlation between the D60/EoT ratio and the day they received rescue treatment (linear regression R2 = \u22120.009).An increase in neopterin concentrations was observed for relapsing patients between end of treatment and day 60 Table , but notp < 0.001), the monotherapy (p < 0.01), and combination therapy (p < 0.05) separately. ROC curves are depicted in Figure The D60/EoT neopterin concentration ratio was a significant predictor of relapse for both arms combined , which might have impeded statistical power to detect predictors for relapse.As a subset of cured patients also demonstrated an increase in neopterin concentration after end of treatment, specificity was relatively low. No clinical explanation could be identified for the neopterin concentration increase on day 60 in these cured patients: there were no consistent trends in fever, hematological or clinical chemistry parameters, nor were there more co-infections or concomittant medications reported in these patients. An additional limitation of the study was the relatively small group of relapses  (Skogmar et al., Currently there are no biomarkers either to identify treated VL patients at risk of relapse, or to establish final cure during the follow-up in clinical trials. This lack of early markers or test of cure is impeding the development of new antileishmanial treatment regimens. This study is the first evaluation of neopterin as a predictor of relapse in VL patients.In conclusion, the identified 1.08 D60/EoT ratio cut-off\u2014an >8% neopterin concentration increase between end of treatment and day 60\u2014could serve as a surrogate endpoint identifying the patients in clinical trials who have an increased risk of relapse. Identified at-risk patients could be more intensively followed up in clinical trials, possibly using qPCR to quantify parasite loads in the blood and/or tissue to enable early detection of parasite recrudescence. The use of this neopterin parameter as a predictive biomarker for relapse in VL should be formally evaluated in a prospective trial, possibly in a panel of biomarkers to increase specificity.MW, AM, EK, FA, and TD were responsible for the clinical study conception and design. MW, AM, and EK were responsible for acquisition of the clinical data and samples. JB and JS provided the laboratory setting to perform the analysis. AK performed the laboratory analysis. AK and TD performed the data analysis and interpretation of the results. AK took the lead in writing the manuscript. All authors provided critical feedback and helped shape the research, analysis, and manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Plasmodium falciparum transmission in areas threatened by artemisinin resistance. To date, no randomized controlled trials have measured primaquine\u2019s effect on infectiousness to Anopheline mosquitoes in Southeast Asia.Single low dose primaquine  is recommended in combination with artemisinin-based combination therapy (ACT) as a gametocytocide to prevent Anopheles dirus on days 0, 3, 7, and 14 of blood-stage therapy. Gametocytemia was evaluated by microscopy and reverse-transcriptase PCR.Cambodian adults with uncomplicated falciparum malaria were randomized to receive a single 45mg dose of primaquine  or no primaquine after the third dose of dihydroartemisin-piperaquine (DHP) therapy. A membrane-feeding assay measured infectiousness to Prior to trial halt for poor DHP treatment efficacy, 101 participants were randomized and 50 received primaquine. Overall microscopic gametocyte prevalence was low (9%), but gametocytemic subjects given primaquine were gametocyte-free by day 14, and significantly less likely to harbor gametocytes by day 7 compared to those treated with DHP-alone, who remained gametocytemic for a median of two weeks. Only one infectious subject was randomized to the primaquine group, precluding assessment of transmission-blocking efficacy. However, he showed a two-fold reduction in oocyst density of infected mosquitoes less than 24 hours after primaquine dosing. In the DHP-alone group, four subjects remained infectious through day 14, infecting roughly the same number of mosquitoes pre and post-treatment. Overall, microscopic gametocytemia was an excellent predictor of infectiousness, and performed better than submicroscopic gametocytemia post-treatment, with none of 474 mosquitoes infected post-treatment arising from submicroscopic gametocytes.In a setting of established ACT resistance, a single dose of 45mg primaquine added to DHP rapidly and significantly reduced gametocytemia, while DHP-alone failed to reduce gametocytemia and prevent malaria transmission to mosquitoes. Continued efforts to make single dose primaquine widely available are needed to help achieve malaria elimination. Plasmodium falciparum cases in the region, but have failed to prevent the emergence of slow-clearing parasites throughout Southeast Asia [Malaria containment efforts launched in western Cambodia over the last 7\u20138 years to curb artemisinin-resistant malaria have contributed to a marked decline in ast Asia \u20133. Camboast Asia \u20136. As feP. falciparum to mosquitoes in areas threatened by artemisinin resistance [Anopheles gambiae mosquitoes in West Africa [In 2010, the WHO recommended that single dose primaquine  be used as a gametocytocide in combination with ACT to prevent transmission of sistance \u201311. Howesistance . Earliert Africa .Anopheles dirus mosquitoes, the major malaria vector in the region. Unfortunately, DHP failure in the cohort was unexpectedly high indicating rapid progression of clinical resistance, and the trial had to be halted early, as previously reported [In 2012\u20132014, we similarly undertook an open label randomized trial to determine the transmission-blocking efficacy of primaquine added to DHP in western Cambodia. We used membrane feeding assays to measure infectiousness to reported . Here weP. falciparum or mixed P. falciparum/P. vivax (Pf/Pv) infection diagnosed with microscopy and confirmed with real-time polymerase chain reaction (PCR). Testing for glucose-6-phosphate dehydrogenase (G6PD) deficiency was carried out using both the fluorescent spot test  and quantitative testing of enzyme activity . Participants with severe deficiency (WHO Class I or II) defined as 10% or less of the lower limit of normal activity were excluded, while those with mild or moderate G6PD deficiency were included in the trial. More details of the study design have been previously presented [This study was carried out between 10 December 2012\u201324 February 2014 in Oddar Meanchey Province, northwestern Cambodia. Malaria transmission is low, heterogeneous, and seasonal with entomological inoculation rates generally below one/person/year. The majority of clinical cases occur during the rainy season between May and December [resented . ParticiAll participants received three doses of dihydroartemisinin-piperaquine (40mg and 320mg per tablet) as directly observed therapy over 3 days . After the third dose, subjects were randomly allocated 1:1 to receive 45mg of primaquine or no primaquine. This dose typically occurred 52 hours from baseline (on day 2). Participants were released to outpatient follow-up on day 3 (72 hours) or once afebrile with two consecutive negative smears, returning for weekly follow-up visits until day 42.Giemsa-stained thick and thin blood smears collected at baseline, every 8 hours during treatment, and at weekly follow-up visits were examined by two microscopists blinded to each other\u2019s results, with gametocytes counted per 2000 white blood cells . MoleculAnopheles dirus mosquitoes, with the goal of yielding approximately 200 engorged (fed) mosquitoes. Further details on the membrane feeding assay have been previously published [P. falciparum infection in mosquitoes fed on subjects with mixed Pf/Pv infection.Mosquito infectivity was measured in all study participants, regardless of gametocyte status, via membrane feeding experiments at baseline (day 0); and days 3 (72 hours from baseline), 7, and 14 post-treatment. For each experiment, 2mL of fresh venous blood was fed to ~300 colony-reared female ublished . Nine daublished . In partWe assessed the transmission-blocking efficacy of a 3-day dose of DHP with or without a single oral 45 mg dose of primaquine by comparing the proportion of individuals infecting at least one mosquito out of fifty, at one and two weeks post-treatment in the two arms. We also assessed the effect of the two treatment regimens on the risk of gametocyte carriage as measured by microscopy and RT-PCR. This was done by comparing gametocyte prevalence at weekly timepoints post-treatment and the time to gametocyte clearance in the two arms. We also analyzed the number of infected mosquitos per treatment arm, the relationship of gametocytemia to mosquito infectivity, and within-person changes in hemoglobin four days post-primaquine treatment among volunteers with G6PD-deficiency.The study was not powered for transmission-blocking efficacy, but for the main primary objective of measuring the therapeutic efficacy of DHP with and without primaquine . A samplOut of 107 subjects who were enrolled and given dihydroartemisinin-piperaquine (DHP) therapy, six withdrew from the study before being randomized, leaving 101 who were randomized to primaquine (PQ) or no primaquine on Day 2; 50 received DHP+PQ and 51 DHP-only [A total of 387 membrane feeding assays were successfully conducted in the 101 subjects pre and post-treatment . Feeding assays were performed a median of 1.0 hours (interquartile range 28 min to 2.2 hours) after venous blood draw. In every experiment, 50 mosquito midguts were dissected 9 days after initial membrane feeding and examined for oocysts. Positive mosquito infection (at least 1/50 with midgut oocysts) was confirmed in all cases by real-time PCR of mosquitoes also saved at day 9 post-feeding.Only 7/101 (6.9%) participants were infectious to mosquitoes pre-treatment\u20146 in the DHP-only arm and 1 in the DHP+PQ arm . Of the Of note, the one gametocytemic and infectious subject in the primaquine group (SN-060) remained infectious at the day 3 membrane feeding, which occurred just ~20hrs after primaquine dosing, but displayed a two-fold reduction in oocyst density in the infected mosquitoes , Table 3At baseline, 9/101 (9%) participants were gametocyte positive by microscopy, while 46/101 (46%) were gametocyte positive by Pfs25 RT-PCR. Just prior to primaquine dosing on day 2, the proportion of participants with microscopic gametocytemia increased to 12/101 (12%). By one week post follow-up, 5 days after primaquine dosing, those given primaquine had a lower rate of gametocyte carriage, with 2.1% of subjects in the DHP + PQ group displaying patent gametocytes vs. 15% of those in the DHP-only arm  , Fig 2. Accordingly, gametocyte clearance was faster in the primaquine group. Among the 12 participants who were gametocytemic by microscopy at day 2, the median time to clearance was 1 day in the primaquine group vs. 12 days in the non-primaquine group (hazard ratio 7.3 (95% CI 1.3\u201342), logrank p = 0.01) . The detWe previously showed a close relationship between microscopic gametocytemia and infectiousness to mosquitoes at baseline pre-treatment in this cohort . Using aG6PD screening by both qualitative and quantitative tests led to the exclusion of 5 persons with severe G6PD deficiency  prior to enrollment. Of 101 participants enrolled in the study, 8 had mild or moderate G6PD deficiency, with activity ranging from 0.6 to 3.2 U/Hgb (median 1.2 U/Hgb). Among these 8 subjects, a greater drop in hemoglobin at day 7 was seen among those treated with primaquine but was not statistically significant: median fractional reduction in hemoglobin was 24.8% (range 0.9% to 29.8%) in the six subjects given primaquine vs. 7.1% (-4.4% and 18.7%) in the two subjects not given primaquine (p = 0.14). The largest drop was to 9.9 g/dL from a baseline Hgb of 14.1; all Hgb values were >11.0 g/dL by day 14 . In the Anopheles dirus, one of the predominant outdoor biting malaria vectors residing in the forest and forest fringes throughout Southeast Asia [Our study is one of now three randomized control trials using a mosquito infection endpoint to provide evidence for the WHO\u2019s 2010 recommendation to add single dose primaquine to ACT as a transmission-blocking intervention in areas nearing elimination or threatened by artemisinin resistance ,19. It iThis study used the single 45mg dose (0.75 mg/kg) of primaquine recommended by WHO at the time it was conducted. The WHO revised their recommendation to use a lower dose of 0.25mg/kg or 15mg in 2012 . The swiFrom an efficacy standpoint, dose-finding transmission-blocking studies recently completed in Mali and Burkina Faso now support adequate transmission-blocking efficacy of the lower 0.25mg/kg dose recommended by the WHO ,19,27 inAs we have previously noted, microscopic gametocytemia was an excellent predictor of infectiousness in our cohort. Of 21 episodes of human to mosquito transmission in the cohort (7 pre-treatment and 14 post-treatment), all but two arose from subjects with microscopic gametocytemia. The two that arose from submicroscopic gametocytes occurred pre-treatment and resulted in low-level mosquito infection . Our molPerhaps the most unsettling finding here was how much post-treatment gametocytemia still remained in the ACT-alone \u201ccontrol\u201d arm in this setting of prevalent artemisinin and piperaquine resistance. In areas where ACTs have not been compromised, artemisinin-based therapies rapidly clear asexual parasite stages and are thought be effective against immature gametocytes as well, resulting in low rates of post-treatment gametocytemia compared to non-artemisinin drugs ,11,29. INot all ACTs are equal in their ability to reduce gametocytemia. While DHP offers advantages of once daily dosing and a long elimination half-life, its Achilles heel may be more post-treatment gametocyte carriage compared to other ACTs. Two large studies both found that gametocyte carriage rates following DHP treatment were 2\u20133 fold higher than those following artesunate-mefloquine (ASMQ), the other ACT frequently used in Southeast Asia ,37, a fiThe weakness of DHP as an anti-gametocyte ACT is overcome when given with single dose primaquine 8,14,24). Our study shows that this is true even in the setting of high-grade DHP failure, at least with the higher 0.75mg/kg dosing ,14,41\u201343,24. Our P. falciparum transmission. They highlight the importance of adding primaquine as a transmission-blocking intervention in areas where ACTs are failing, as we found that Cambodian patients treated with dihydroartemisinin-piperaquine cleared gametocytes slowly and still contributed substantially to the infectious reservoir after treatment. Fortunately, most infectious subjects were readily identified by the presence of patent gametocytes. Despite adoption of primaquine into the official drug policy of many countries, a large gap remains between policy and real-world availability [Overall, our findings, though limited by small numbers and a higher dose of primaquine than currently recommended, lend support to the WHO recommendation to give single dose primaquine to reduce lability . OvercomS1 FigThis figure is similar to (TIFF)Click here for additional data file.S2 FigHemoglobin values during follow-up for the 8 participants with G6PD deficiency Class III (>10% enzyme activity) (A) and the calculated fractional change compared to day 0 pre-treatment in those same subjects (B). Trend lines for the subjects in the primaquine group are red (for those with >25% drop in Hgb at day 7) and green . Trend lines for the 2 subjects in the DHP-alone group are black.(TIFF)Click here for additional data file.S1 TableThe tables include membrane-feeding assays conducted on subjects pre and post-treatment. In the upper table, PCR positivity at both Day 9 (oocyst stage) and Day 16 (sporozoite stage) was required for an overall positive determination. In the lower table, only PCR positivity at day 9 was used. In 4 of the 9 discordant results, PCR was positive in 1/5 pools of 10 mosquitoes. In the remaining 5 time points, 2/5 pools were PCR-positive. Subjects with mixed Pf/Pv infections are excluded from this table because of different mosquito processing procedures.(PDF)Click here for additional data file.S1 File(PDF)Click here for additional data file.S2 File(DOCX)Click here for additional data file.S3 File(XLS)Click here for additional data file."}
+{"text": "Laboratory analyses included haematology and clinical chemistry. The main objective of the ECG assessment in this study was to evaluate the effect of administration of DHA/PPQ on QTc intervals and the association of QTc intervals with changes in blood biochemistry, full and differential blood count over time after the DHA/PPQ administration. A total of 1315 patients gave consent and were enrolled of which 1147 (87%) had complete information for analyses. Of the enrolled patients 488 (42%), 323 (28%), 213 (19%) and 123 (11%) were from Ghana, Burkina Faso, Tanzania and Mozambique, respectively. Median (lower\u2014upper quartile) age was 8 (5\u201314) years and a quarter of the patients were children under five years of age (n = 287). Changes in blood biochemistry, full and differential blood count were temporal which remained within clinical thresholds and did not require any intervention. The mean QTcF values were significantly higher than on day 1 when measured on day 3 before and after administration of the treatment as well as on day 7, four days after completion of treatment . In all age groups the values of QT, QTcF and QTcB were highest on day 3 after drug intake. The mean extreme QTcF prolongation from baseline was lowest on day 3 before drug intake  and highest on day 3 after the last dose . There were 79 (7%) events of extreme mean QTcF prolongation which were not clinically significant. Nearly a half of them (n = 37) were grade 3 and mainly among males (33/37). Patients in Burkina Faso, Mozambique and Tanzania had significantly lower mean QTcF than patients in Ghana by an average of 3, 4 and 11 ms, respectively. We found no evidence that Eurartesim\u00ae administered in therapeutic doses in patients with uncomplicated malaria and no predisposing cardiac conditions in Africa was associated with adverse clinically significant QTc prolongation.The antimalarial drug piperaquine is associated with delayed ventricular depolarization, causing prolonged QT interval (time taken for ventricular de-polarisation and re-polarisation). There is a lack of safety data regarding dihydroartemisinin/piperaquine (DHA/PPQ) for the treatment of uncomplicated malaria, which has limited its use. We created a platform where electrocardiograms (ECG) were performed in public hospitals for the safety assessment of DHA/PPQ, at baseline before the use of dihydroartemisinin/piperaquine (Eurartesim Dihydroartemisinin/piperaquine (DHA/PPQ) is one of the artemisinin based combination therapies (ACTs) effective against uncomplicated malaria. Its use has been limited due reported cardiotoxic effects . Currenttorsades de pointes (TdP). In clinical pharmacology prolonged QT interval is a heart rhythm\u2019s toxicity indicator recorded upon confirmation of Fridericia\u2019s correction > 450 milliseconds  as compared to counterpart adult females. In children, boys aged between 6 months and 5 years had significantly 5  higher mean QTcF compared to girls of the same age .Mean extreme QTcF prolongation from baseline was highest (60 ms) on day 3 after the last dose and lowest on the same day (33 ms) before last dose. On day 7 after treatment there was a downturn in the mean QTcF as compared to baseline which was recorded as 47 ms. In total we observed 79 (7%) extreme mean QTcF prolongation events from baseline which were also not clinically significant. The distribution of the extreme QTcF prolongations was 15 (19%), 58 (73%) and 6 (8%) recorded on day 3 before drug intake, day 3 post drug intake and on day 7 after treatment respectively. Almost a half of all extreme QTcF prolongation events that occurred were grade 3, (37/79) whereas male accounted the majority (33/37) and children aged 5-<12 years were more than a half of all grade 3 extreme prolongation cases (n = 19). Male patients had majority of QTcF prolongation events than female throughout the treatment period .\u00ae) evaluated by the incorporation of ECG and laboratory measurements of changes in patients treated with Eurartesim\u00ae. DHA/PPQ is one of the antimalarials that has shown a good efficacy against P.falciparum infection [P.falciparum malaria fever, especially the severe form is also associated with tachyarrhythmia a manifestation form of shortening of QT interval [\u00ae. There were prolongations in the mean QTcF during the course of treatment with Eurartesim\u00ae that was provided on Day 1 through Day 3. These changes were recorded in 79 cases of extreme QTcF prolongation that reverted to initial baseline QTc values in most patients on Day 7. In this study, QTc maximum prolongation was observed on Day 3 corresponding to what would be considered onset of peak plasma concentration of piperaquine following the last dose administration [torsades de pointes. In our study QTcF prolongation that we observed did not worsen to torsades de pointes most probably due to the therapeutic dose that we used being within safety margin of therapeutic thresholds. Reversible ECG changes from baseline on day 1 as demonstrated through the recession of mean QTc interval values recorded on day 7 suggests the safety of the optimal therapeutic dose of Eurartesim\u00ae among African patients with uncomplicated malaria is warranted. One would suggest that perhaps we did not see untoward extreme maximum QTc prolongation in the study population because all patient with the history of cardiac problems were excluded at baseline. This position has some bearing and cannot go without comment. Our approach was to exercise utmost safety precautions and this might have enabled us to avoid any unnecessary torsadogenic events that might resulted, had we included-pro-arrhythmic patients. These findings also reflect the classic overall presentation of plasma concentration levels that are expected to decline around day 7 corresponding QTc recovery [Whereas use of ECG for cardiotoxicity evaluation of other ACTs has been well detailed in literature, this study gives additional evidence of safety of DHA/PPQ Click here for additional data file.S1 Protocol(PDF)Click here for additional data file."}
+{"text": "Artemisinin-based combination therapy (ACT) and novel drug combinations are available and used in African countries to treat uncomplicated malaria. Network meta-analysis methods are rarely and poorly applied for the comparison of their efficacies. This method was applied on a set of randomized controlled trials to illustrate its usefulness.A literature review available in Pubmed was conducted in July 2016. Eligible studies, conducted in sub-Saharan Africa, published between 2002 and 2016, focused on randomized controlled trials of at least two artemisinin-based combinations to treat uncomplicated malaria in children and adults. Agglomerate data were: the number of PCR-corrected adequate clinical and parasitological response (ACPR) on day 28, used as the primary endpoint in all interventions, the number of participants and the list of treatments. A Bayesian random effect meta-analysis using a binary outcome was the method to compare the efficacy. Ranking measure was used to obtain a hierarchy of the competing interventions.In total, 76 articles were included; 13 treatment regimens were involved and tested in 36,001 patients. Using artemether\u2013lumefantrine (AL) as the common comparator for the entire network, 12 relative treatment effects were estimated and indirect comparisons were obtained. Dihydroartemisinin\u2013piperaquine (DHAP) was shown to be more effective than AL , ASAQ , and amodiaquine\u2013sulfadoxine\u2013pyrimethamine (AQSP): OR\u00a0=\u00a02.20; 95% CI 1.21\u20133.96; 8863 patients. Artesunate\u2013amodiaquine (ASAQ) was comparable to AL . No significant difference was found between artesunate and mefloquine (ASMQ) and AL . According to treatment ranking, among the WHO-recommended ACT medicines, DHAP was shown to be the most efficacious.Based on the available evidence, this study demonstrated the superiority of DHAP among currently recommended artemisinin-based combinations. The application of the methods described here may be helpful to gain better understanding of treatment efficacy and improve future decisions. However, more data are needed to allow robust conclusions about the results in comparison with novel drugs. Further surveillance of the efficacy of anti-malarial drugs and clinical trials are needed to closely follow the evolution of the epidemiology of drug-resistant malaria in Africa.The online version of this article (doi:10.1186/s12936-017-1963-0) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria in Africa and elsewhere in the world. Since 2006, four ACT medicines have been recommended by the World Health Organization (WHO): artemether\u2013lumefantrine (AL), artesunate\u2013amodiaquine (ASAQ), artesunate\u2013mefloquine (ASMQ) and artesunate\u2013sulfadoxine\u2013pyrimethamine (ASSP) : \u201c2016/06/30\u201d [PDAT]) AND \u201chumans\u201d. In addition, reference lists of reviews were also screened to include potential articles. Studies were eligible if they involved at least 2 ACT medicines and reported clinical efficacy as ACPR corrected by polymerase chain reaction (PCR). Cure, i.e. the term \u201cadequate clinical and parasitological response\u201d, was defined as undetectable parasitaemia with or without fever, without previously meeting the criteria of treatment failure on the last day of follow-up .K\u00a0=\u00a013 treatments were involved. The quinine arm was removed from a 3-arm study ) ) , 44, 45.Table\u00a0Indirect estimates were calculated from each model assuming the consistency equation Table\u00a0. The resModel 2 was selected for calculating the rank probabilities. Table\u00a0To provide an answer to the best treatment among WHO-recommended ACT medicines and AQSP, a second analysis was carried out using model 2, with AL as the overall control group. Treatments were numbered as follows: AL\u00a0=\u00a01, AQSP\u00a0=\u00a02, ASAQ\u00a0=\u00a03, ASMQ\u00a0=\u00a04, ASSP\u00a0=\u00a05, DHAP\u00a0=\u00a06, 7\u00a0=\u00a0ASAQCPH, 8\u00a0=\u00a0ASATPG, 9\u00a0=\u00a0ASCD, 10\u00a0=\u00a0ASNAPH, 11\u00a0=\u00a0ASPY, 12\u00a0=\u00a0ASSMP, and 13\u00a0=\u00a0DHAPT, allowing the WHO-recommended ACT medicines to be the first 6 treatments. Ranking was only up to 6. Table\u00a0NMA of randomized clinical trials is becoming a promising tool to analyse grouped data with multiple interventions. The use of this methodological approach was assessed for the evaluation of malaria treatment efficacy and the selection of the most efficacious drug. Several clinical trials were identified and assessed using the proportions of PCR-adjusted ACPR. The rationale of choosing 2002\u20132003 as the starting point was that it is considered as the early years of the introduction of ACT in some African countries and the year when the WHO protocol requiring a 28-day follow-up to guide clinical trials was updated . When usThe results become more robust when a Bayesian estimation method is used. One of the strengths of this method is that, at a point of time when the efficacy of a drug is not established, the contribution of other trials could suggest its relative efficacy. In addition, pooling trial increases the sample size and allows the selection of the best treatment. This is an advantage of NMA approach that may facilitate the selection of the best treatment through rank probabilities and is, therefore, one feature of the novel method to guide decision making. Two models were examined in the present study, and both provided similar results.P. falciparum malaria in Africa [P. falciparum has become a global research goal. Since their adoption in African countries, few data on the selective impact of ACT in the circulating parasite population are available. A recent study assessed the selective impact of the treatment with ASAQ and AL on Pfcrt and Pfmdr1 alleles and found no association between the presence of Pfcrt and Pfmdr1 alleles before treatment and at the time of treatment failure [kelch 13 [The results did not show any difference in the efficacies of ASAQ and AL. AL was the first co-formulated fixed-combination ACT to become available, followed by ASAQ. The six-dose regimen of AL is highly effective and represents a challenging comparator for any new drug combination although the non-inferiority of ASAQ versus AL had been shown in an earlier multi-centric study . Both ACn Africa , 48. Hown Africa , 50. Aftn Africa . Since c failure . Hence, kelch 13 .The results highlighted that DHAP is superior to AL, contrary to some published reports in which the overall efficacy of DHAP was found to be similar to that of AL in multi-centric studies in Africa , 53 and Studies in Africa also demonstrated the efficacy and tolerability of ASMQ , 59. Howdhfr) and dihydropteroate synthase (dhps) genes and where SP is employed for intermittent preventive treatment in pregnancy and infancy and AQSP combination is employed for seasonal malaria chemoprevention [Regarding ASSP, it has been efficacious and well tolerated as ASAQ and, has been used in some African countries as the first-line treatment . Recent evention .In addition to five ACT medicines currently recommended by the WHO , severalP. falciparum gametocytes in an effort to reduce transmission in areas where malaria elimination programme is being implemented [ASPY is a newly introduced form of ACT which may possibly be deployed together with primaquine to kill mature lemented , 71. A mlemented . In the So far, only a few studies have presented results based on the suggested methodological changes. However, Donegan et al.  exploredAmong non-ACT drug combinations, AQSP had been evaluated and compared to ACT in some African countries during the pre-ACT period , 74. In Based on this novel methodological approach, among the 13 combination therapies included in the present analysis, ASATPG emerged at the top rank as the best treatment followed by ASAQCPH and ASNAPH in spite of the limited number of trials. These findings show that NMA tends to be too sensitive when treatments are tested once or twice. The method allowed these treatments to be at first rank. However, to counter this methodological limitation, further randomized clinical trials with AL, ASAQ, AQNAPH and ASATPG would be necessary to confirm these initial findings based on a limited number of trials.The second analysis with the most widely tested combinations in Africa showed that DHAP was more effective than AL and found to be the best treatment, followed by ASMQ, and AL at the last rank. The combination AL has been found in most studies to be the most efficacious treatment. In the present analysis, the finding that AL is not the most efficacious ACT could be due to the fact that it was fixed as the overall control drug in the network. However, this result does not bring into question its efficacy because in the present analysis it was ranked among the best treatments. As an additional option, ASMQ, which has not been extensively employed in Africa, could be used as an alternative for the first-line anti-malarial therapy.The present study provides the first overview of NMA methods for anti-malarial drug efficacy. It follows the work initiated by Donegan et al. and offers further research opportunities . HoweverNMA technique may have a role to play in the evaluation of different public health policies and interventions for malaria control. DHAP was found the best treatment overall but this observation should be treated with caution in decision-making since the present analysis was based on the outcome on day 28 and did not take into consideration the outcome on day 42 and 63 for drugs with long elimination half-life due to inadequate available data. More comparative studies are needed with novel drug combinations to assess their efficacy as compared to the currently recommended artemisinin-based combinations in Africa.Additional file 1. Description of the trials characteristics. The file contains the description within each article in terms of trial characteristics, including the number of treatment arms (a), the list of treatment (k), the enrolled population (n) and the number of observed PCR-corrected ACPR (r). The file also contains the number of studies for each pairwise comparison.Additional file 2. WinBUGS codes for the random effect models. The file contains two parts. Part 1 is the code to run the random effect model under the hypothesis of consistency; part 2 is the code for the random effect model under the inconsistency assumption. These codes are to be run using the free WinBUGS 1.4 software downloadable at http://www.mrc-bsu.cam.ac.uk/bugs/winbugs/contents.shtml.Additional file 3. Forest plots and other direct and indirect comparisons. Direct and indirect comparisons are extracted from the model with inconsistency."}
+{"text": "Adding low-dose primaquine to malaria treatment reduced gametocyte carriage by 73%. Patients who received primaquine had more frequent hemoglobinuria and there was a greater reduction in haemoglobin concentration in G6PD-deficient patients. One patient who received primaquine developed moderately severe anemia. More information is needed about the safety of low-dose primaquine in populations where G6PD deficiency is common.Plasmodium falciparum malaria were randomized to receive 1 of 3 artemisinin combination therapies (ACTs) with or without primaquine (0.25 mg/kg). Glucose-6-phosphate dehydrogenase (G6PD) status was determined using a rapid test. Patients were followed for 28 days to record hemoglobin concentration, adverse events, and gametocyte carriage. The primary end point was the change in Hb at day 7.Adults with P = .01). One G6PD normal patient who received primaquine developed moderately severe anaemia (Hb < 8 g/dL). Dark urine was more frequent in patients who received primaquine. Primaquine was associated with a 73% (95% CI 24\u201390) reduction in gametocyte carriage (P = .013).In sum, 274 patients were randomized, 139 received an ACT alone, and 135 received an ACT + primaquine. The mean reduction in Hb at day 7 was similar in each group, a difference in the ACT + PQ versus the ACT alone group of \u22120.04 g/dL , but the effect of primaquine differed according to G6PD status. In G6PD-deficient patients the drop in Hb was 0.63 g/dL  greater in those who received primaquine than in those who received an ACT alone. In G6PD-normal patients, the reduction in Hb was 0.22 g/dL  less in those who received primaquine PACTR201411000937373 ( Plasmodium falciparum malaria as a component of pre-elimination or elimination programs [Plasmodium vivax and as a P. falciparum gametocytocide [P.vivax hypnozoites [Progress in the fight against malaria  has led programs . Primaqucytocide . Primaqucytocide . Howevercytocide . Primaqucytocide , but thecytocide . The A- cytocide , 10 but cytocide . The G6Pcytocide . In neigcytocide . Althougcytocide . This cacytocide . Hemolyscytocide , 17 thannozoites .In studies in Mali and Burkina Faso, administration of a single dose of primaquine (0.25 mg/kg or 0.4 mg/kg) reduced gametocyte carriage and infectiousness without clinically significant haemolysis, but these studies did not include G6PD-deficient participants , 20. In P. falciparum malaria.In Senegal, elimination of malaria is planned in the central and northern parts of the country where incidence is low . IntroduP. falciparum malaria were enrolled if they had monospecific infection with parasite density 1000\u2013100000 trophozoites/\u03bcL and gave signed consent, and were randomized individually to receive AL, DHA-PQ, or ASAQ either alone or with primaquine. Patients were excluded if they were pregnant (confirmed by urine testing), breastfeeding, had a history of hypersensitivity to any of the study drugs, had severe malaria, moderately severe anemia (hemoglobin <8 g/dL), or had a chronic illness.An open-label randomized trial was undertaken in adult patients presenting with malaria at Deggo health post, Pikine, Dakar, Senegal. Patients >18 years with Treatment assignments, prepared in permuted blocks of 18 using Stata 12 , were placed in numbered, sealed, opaque envelopes, which were opened in sequence by the study pharmacist at the time of treatment. The pharmacist was not involved in patient screening or assessment of outcomes. The study was not blinded, but laboratory technicians responsible for hemoglobin measurement and microscopy were unaware of treatment allocations. Fixed ACT combinations were used. For AL, tablets containing 20 mg artemether and 120 mg lumefantrine (Novartis) were given twice daily at the clinic; for DHA-PQ, patients received 3 tablets  per day for 3 days, whereas for ASAQ, 2 tablets of 100 mg artesunate plus 270 mg amodiaquine (Sanofi-Aventis) were given per day for 3 days. One tablet of primaquine  was given to patients who were randomized to the ACT + primaquine groups. Primaquine was given at day 0 in addition to the first dose of ACT under the direct observation of the research team. After each treatment dose, participants were observed for 30 minutes and treatment readministered if the patient vomited. Patients who vomited a second time were withdrawn from the study and treated with quinine. Primaquine and AL were given with biscuits (to provide fat as recommended by the manufacturer). Clinical assessment was done on days 0, 1, and 2; subsequent visits were scheduled on days 3, 7, 14, 21, and 28. At each of these visits, hemoglobin concentration (Hb) was measured and adverse events were recorded. Adverse events, graded using a severity scale adapted from the WHO toxicity grading scale for determining the severity of adverse events, were classified as minor, moderate, or severe . Urine sCareStart G6PD deficiency Rapid Diagnostic Test (RDT) is a qualitative enzyme chromatographic test based on the reduction of colorless nitro blue tetrazolium dye to dark-colored formazan. The sensitivity and specificity for this test range from 90% to 98% and from 87% to 96%, respectively [Hb was measured using a HemoCue machine . A qualitative test of G6PD activity  was done on day 0 using a finger-prick blood sample. The ectively , 26. ThiP-values were obtained for testing interaction between G6PD status and treatment group at each time point, and the effect of treatment in G6PD normal and deficient patients, with a 95% confidence interval, at each time point was obtained from the model. Logistic regression was used to compare the proportion of G6PD normal and deficient patients who had a drop in Hb of 2 g/dL or more by day 7. Data were double-entered into an Access database. Analyses were conducted using Stata .The primary endpoint was the change in Hb from day 0 to day 7. A reduction in Hb of 0.3 g/dL or less due to primaquine was considered to be small enough to be of no clinical concern. An interim analysis conducted to assess safety showed that the standard deviation of the change in Hb was 2.54 g/dL and on this basis a total sample size of 350 patients would be needed to have a 95% confidence interval that excluded a difference of 0.3 g/dL or more between groups. Secondary endpoints included change in Hb by day 3, day 14, day 21, and day 28; anemia (Hb < 11 g/dL) at any time up to day 28; clinical adverse events up to day 28, and the prevalence and density of gametocyte carriage during follow-up. The effect of primaquine on Hb was estimated using a mixed model, with time, treatment group, interaction between time and treatment, ACT regimen and gender as fixed effects, baseline Hb as a covariate, and patient as a random effect. The treatment difference at each time point, with a 95% confidence interval, was obtained from this model. A planned subgroup analysis estimated the effect of primaquine in G6PD normal and G6PD deficient patients, by fitting a model that included G6PD status, the interaction between time and G6PD status, and the interaction between treatment group and G6PD status at each time point, as fixed effects, baseline Hb as a covariate, and patient as a random effect. Wald The protocol was approved by the Conseil National de Recherche en Sant\u00e9 in Senegal.During 2 malaria transmission seasons, 402 individuals with a history of fever and a positive malaria RDT were screened. The most frequent reason for exclusion was Hb < 8 g/dL at the time of screening (72 patients). In sum, 274 patients were randomized; 262 (96%) completed 28 days of follow-up . RecruitP-value for the interaction of ACT type with primaquine was not significant . Similar results were obtained at other time points. The estimates of differences at each time point obtained from the mixed model, adjusted for effects of ACT type, sex, and baseline Hb, are shown in The mean Hb on day 7 was similar in patients who received ACT alone and those who received ACT + primaquine. The difference  adjusted for baseline was \u22120.23 g/dL  for AL, \u22120.02  for ASAQ and +0.54  for DHA-PQ. As the  0.0728, , for subP-value of .01. There was no evidence of interaction on other days (Fifty-four patients (20%) were G6PD-deficient; in this subgroup, the mean Hb on day 7 was 0.63 g/dL  lower in those who received primaquine than in those who received ACT alone . In G6PDher days . The meaAmong G6PD-normal patients, the proportion with a drop in Hb of 2 g/dL or more at day 7 was 29% in the ACT group versus 24% in the ACT plus primaquine group, a risk ratio (RR) of 0.85  (Table S3). Among G6PD-deficient patients, 26% had a drop in Hb of 2 g/dL or more after treatment with ACT by day 7 compared with 31% after treatment with ACT plus primaquine .The proportion of patients with moderate anaemia (Hb < 11 g/dL) during the 28 day follow-up period was similar in those who received primaquine and those who received ACT alone, and there was no evidence of an interaction with G6PD status (Table S4). One patient presented with moderately severe anemia (Hb < 8 g/dL), a man in the AL + primaquine group, whose baseline Hb was 12 g/dL and who was G6PD-normal. He received AL and one tablet of primaquine (0.22 mg/kg). His Hb was 8.4 g/dL on day 3 and 7.3 g/dL on day 7 (a 39% reduction from baseline) but with no clinical features of anemia. After supplementation with iron and folate, his Hb recovered to 11.2 g/dL by day 28.P < .001). The estimates on day 2 were 25% and 12% , and 4.5% and 1.5%  on day 3. Those who had dark urine (grade 3 or above) on day 3 had a lower Hb on day 3 than those who did not .The incidence of adverse events was similar in all treatment groups (Table S5) except for the occurrence of dark-coloured urine, which was more common in patients who received primaquine and occurred during the first 3 days after the start of treatment; 79/135 (59%) of patients who received primaquine had dark urine (grade 3 or above) on day 1 after treatment versus 46/139 (33%) in those who received ACT alone . The area-under-the-curve of gametocyte density over time was 106.7 gametocyte-days in the ACT group and 29.5 gametocyte-days in the ACT + primaquine group, a reduction of 72% (95% CI 16%, 91%; P = .024, The prevalence and density of gametocyte carriage were substantially lower in patients who received primaquine than in those who did not . The aveIn 2012, the WHO revised the previously recommended primaquine dose of 0.75 mg/kg to one of 0.25 mg/kg when used together with an ACT as a pre-elimination strategy . HoweverWe are aware of only one other trial has investigated safety of low-dose primaquine in G6PD deficient patients in Africa . In thatP. falciparum gametocytes with a high degree of efficacy, and it can do this in a single dose [In our study, addition of low-dose primaquine to ACT treatment in adult patients resulted in a substantial reduction in gametocyte carriage. Reduced gametocyte carriage was also observed in studies in G6PD-normal patients in Mali and Burkina Faso , 20. Thugle dose , 31.In our trial, patients with Hb < 8 g/dL were not enrolled. Under routine conditions, and especially during mass drug treatment studies, it may not be feasible to assess Hb prior to treatment for all individuals, and it would be difficult to predict how safe the drug would be in patients with lower Hb. In this trial, direct measurement of hemolysis was not possible. Measuring bilirubin, haptoglobin, methemoglobin, and reticulocyte count would be useful to determine whether the hemoglobin reduction is due to hemolysis or other factors.Primaquine has become a key tool for malaria elimination . This stClinical Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at r_tine_supplementary_information_pm_revClick here for additional data file."}
+{"text": "Between 2009 and 2012, malaria cases diagnosed in a M\u00e9decins sans Fronti\u00e8res programme have increased fivefold in Baraka, South Kivu, Democratic Republic of the Congo (DRC). The cause of this increase is not known. An in vivo drug efficacy trial was conducted to determine whether increased treatment failure rates may have contributed to the apparent increase in malaria diagnoses.In an open-randomized non-inferiority trial, the efficacy of artesunate\u2013amodiaquine (ASAQ) was compared to artemether\u2013lumefantrine (AL) for the treatment of uncomplicated falciparum malaria in 288 children aged 6\u201359\u00a0months. Included children had directly supervised treatment and were then followed for 42\u00a0days with weekly clinical and parasitological evaluations. The blood samples of children found to have recurring parasitaemia within 42\u00a0days were checked by PCR to confirm whether or not this was due to reinfection or recrudescence (i.e. treatment failure).Out of 873 children screened, 585 (67\u00a0%) were excluded and 288 children were randomized to either ASAQ or AL. At day 42 of follow up, the treatment efficacy of ASAQ was 78\u00a0% before and 95\u00a0% after PCR correction for re-infections. In the AL-arm, treatment efficacy was 84\u00a0% before and 99.0\u00a0% after PCR correction. Treatment efficacy after PCR correction was within the margin of non-inferiority as set for this study. Fewer children in the AL arm reported adverse reactions.ASAQ is still effective as a treatment for uncomplicated malaria in Baraka, South Kivu, DRC. In this region, AL may have higher efficacy but additional trials are required to draw this conclusion with confidence. The high re-infection rate in South-Kivu indicates intense malaria transmission.Trial registration NCT02741024The online version of this article (doi:10.1186/s12936-016-1444-x) contains supplementary material, which is available to authorized users. The Democratic Republic of Congo (DRC) is one of the five countries with the highest malaria burden in the world . The easM\u00e9decins sans Fronti\u00e8res\u2013Operational Centre Amsterdam (MSF-OCA or MSF) has been working in the provinces of North Kivu and South Kivu since the early 1990s and in Katanga since 2003  has also observed an increase in malaria cases. In the Baraka programme, without any significant changes to the programme activities, the number of parasitologically confirmed uncomplicated malaria cases has risen from 7457 in 2009 to 44,317 cases in 2012  protocols for surveillance of anti-malarial drug efficacy . The stuThe study was approved by the Ethical Review Board (ERB) of MSF on June 17th 2013 and by the Zone Chief Medical Officer (M\u00e9decin Chef de Zone) and the Provincial Medical Inspector  of South Kivu (N251/969/B.MIP/SK/2013). It was registered at ClinicalTrials.gov under number NCT02741024. Eligible patients were included in the study after an adult parent/caretaker gave written informed consent. If the parent/caretaker was illiterate, a literate witness was asked to sign next to the parent/caretaker\u2019s fingerprint. Free health care for malaria and other illnesses is provided by MSF to the general population and was, therefore, also provided throughout the study follow-up period to all patients included in the study.P. falciparum with an asexual parasite density between 2000 and 200,000/\u00b5l of blood, (3) an ability to swallow (crushed or dissolved) oral medication, (4) a high probability of respecting follow up visits, and (5) a signed informed consent by their adult (\u226518\u00a0years old) parent/caretaker. Children were excluded if they presented with general danger signs according to the WHO protocol \u201cMethods for surveillance of antimalarial drug efficacy\u201d [Children aged between 6 and 59\u00a0months, presenting with fever (axillary temperature\u00a0\u226537.5\u00a0\u00b0C) or reported history of fever in the last 24\u00a0h, and with a positive rapid diagnostic test (RDT) were enrolled in the study if they had: (1) a body weight\u00a0\u22655\u00a0kg, (2) a slide confirmation of mono-infection with Sample size calculations were based on the data that was available before October 2012. At that moment, studies determined the 42-day risk of recurrent parasitaemia due to recrudescence (treatment failure) in children to range from 0.9 to 6\u00a0% with AL and ASAQ . In orde\u00ae, 20\u00a0mg artemether/120\u00a0mg lumefantrine, Novartis) given with milk twice daily over three days  over 3\u00a0days. Drugs were given according to the manufacturer\u2019s instructions and dosage. Study medication was imported and stored under strict regulations, ensuring control within the provided storage temperature and humidity ranges. The first dose of the study drug at day 0 (D0) was dispensed at the health clinic, and was taken by the child while under supervision of a study team member. For children who could not swallow tablets, AL tablets were dissolved, and ASAQ tablets were crushed in minimal amounts of water and dispensed with a spoon. After intake, the child was observed for 30\u00a0min. If the child vomited or spat out the medication within the monitoring period, a resting period of 15\u00a0min was observed before re-administering a repeat dose. If the repeat dose was also vomited within 30 min, the child was administered a rescue treatment  and was excluded from the study. Patients returned to the clinic on days 1 and 2 for observed administration of the study drug and a clinical assessment. In order to observe the intake of the evening doses of AL, patients randomized to receive AL were visited at home at around 8\u00a0h, 32\u00a0h and 56\u00a0h after the first dose. A follow up was scheduled for all patients on day 3, 7, 14, 21, 28, 35 and 42 for clinical assessment and blood smear evaluation.All patients who were eligible for the study had a medical examination, and a MUAC and WHZ screening. A thick and a thin blood smear were performed and capillary blood was collected on a fast technology for analysis (FTA) card . If the patient was eligible, informed consent was requested from the parent/caretaker after the explanation of the study by one of the study team members. Once the informed consent was obtained, patients were randomized to one of two treatment regimens according to which treatment had been randomly pre-allocated to that unique patient number. Treatment regimens consisted of: (1) ASAQ fixed dose (Winthrop Sanofi Aventis), given as 1 tab/day over three days , or (2) AL fixed combination  were not assigned an efficacy treatment outcome.Study endpoints were classified according to the WHO guidelines  as adequInitial testing for malaria infection was done on finger-prick blood using an RDT, the SD Bioline 05FK50 Malaria Ag P.f . This RDT relies on detection of the Histidine Rich Protein 2 (HRP2), and is amongst the most sensitive according to the WHO product testing programme .P. falciparum asexual parasitaemia on the thick smear was performed according to the WHO protocol \u201cMethods for surveillance of antimalarial drug efficacy\u201d [P. falciparum gametocytes was recorded.Thick and thin blood smears were stained with 10\u00a0% Giemsa for 15\u00a0min. Smears were read to 100 fields before they could be declared negative. Species were confirmed on the thin smear. Quantification of fficacy\u201d . PresencEach slide was read, independently, by two qualified microscopists in the Baraka hospital laboratory, Parasite densities were calculated by taking a mean of the two counts. Blood smears with discordant results  were re-examined by a third microscopist who was blinded to the results of the first two, and parasite density was calculated by taking a mean of the two closest counts. In addition, external quality control of the slides was performed by blinded re-checking of 50 randomly selected malaria slides by an expert microscopist at the Epicentre Mbarare Research Base in Uganda.\u00ae Hb 301 System , according to manufacturer\u2019s instructions. PCR genotyping analysis was performed in order to distinguish true recrudescence (same parasite strain) from a newly acquired infection (different parasite strain) on capillary blood samples stored on FTA cards. The genotyping was performed at the Department of Medical Microbiology, at the Academic Medical Centre  and post-treatment (failure) samples was performed according to the WHO protocol [Haemoglobin levels on D0 or during follow-up were determined using the HemoCueprotocol . Pre- anprotocol . Speciesprotocol .Plasmodium species other than the P. falciparum .All data were double entered in the WHO global database for anti-malarial drug efficacy by two different epidemiologists, as well as in the worldwide anti-malarial resistance network (WWARN) database provided online . Statistt test was used for continuous data, as appropriate. The log rank test was used to compare Kaplan\u2013Meier survival curves. p\u00a0<\u00a00.05 were considered statistically significant.Chi square tests or Fisher\u2019s exact tests were used to compare categorical data and Student\u2019s Data are available on request in accordance with MSF\u2019s data sharing policy .rd party (n\u00a0=\u00a09), or having been re-infected with a malaria species other than P. falciparum (n\u00a0=\u00a02).A description of the study outline is shown in Fig.\u00a0The baseline characteristics of the patients enrolled in the study are presented in Table\u00a0Five children were classified as early treatment failures as per study protocol. All of these children had a parasitological improvement, but had either anaemia or jaundice that warranted admission to the hospital. They received intravenous artesunate according to the study protocol and the WHO guidelines for the treatment of malaria  and wereP. ovale), as confirmed in weekly parasitological examinations; this was an involuntary protocol violation and these two children were censored in the final analysis. In total, 192 (80\u00a0%) children had an adequate clinical and parasitological response (92/119 for ASAQ and 100/119 for AL) at day 42. The proportion of recurring infection (including the two reinfections with other species) during the 42\u00a0days follow-up was higher in the ASAQ group  compared to the AL group , but this difference was not statistically significant. Similarly, cumulative failure on day 28 was not statistically significantly different between arms.Treatment efficacy estimates are presented in Table\u00a046 paired FTA filter paper samples were PCR genotyped, including all late clinical and parasitological failure samples. Following the PCR genotyping, 21 of the 27 recurrent parasitaemic episodes in the ASAQ arm (77.8\u00a0%) were classified as re-infections and 6 (22.2\u00a0%) as recrudescence. In the AL arm, 15 out of 19 samples (78.9\u00a0%) were classified as re-infections and 1 as (5.3\u00a0%) as recrudescence. There were no indications that treatment dosing was lower for the children experiencing treatment failure.For three samples with recurring parasitaemia in the AL arm outcome could not be classified by PCR; these samples were censored at the last date of follow-up for which the child was still malaria negative. The proportions of children reaching each a study endpoint, before and after PCR adjustment, are presented in Table\u00a0The cumulative probability of recurrence-free survival over the 42\u00a0days follow-up was determined by Kaplan\u2013Meier survival analysis. The PCR un-adjusted curves shown in Fig.\u00a0A relatively large proportion  of children still had parasites on day 2 of follow-up. This proportion fell to below 5\u00a0% in both arms on day 3. There was no significant difference in the proportion of parasites at D2 and D3 between the two treatment arms . The adverse event most reported was asthenia  for both arms. Children receiving ASAQ reported significantly more asthenia and anorexia (loss of appetite) than children receiving AL Table\u00a0.Table\u00a04AP. falciparum malaria in children aged six to 59\u00a0months in the study setting. In 2004, a similar study in DRC (Pool Region) showed comparable results (98.5\u00a0% efficacy of ASAQ and 100\u00a0% efficacy of AL at 28\u00a0days) [The results of this study indicate that both study drugs retain adequate efficacy (PCR corrected cure rate at day 42 of 94.7\u00a0% for ASAQ and 99.0\u00a0% for AL) in the treatment of uncomplicated 28\u00a0days) . This wa28\u00a0days)  in Owand28\u00a0days) . In the 28\u00a0days) , 25.The high efficacy of both ASAQ and AL in this region is reassuring. WHO only recommends a switch in the first line treatment if the efficacy falls below 90\u00a0% . AlthougWhilst re-infections do not indicate failing anti-malarial drugs, they are of public health relevance and pose a considerable burden to the population and health systems. The results of this study seem to support recent findings that the period over which lumefantrine and amodiaquine provide post-treatment protection is 10\u201314\u00a0days \u201328. HoweIn this study\u2019s setting children will normally receive ASAQ each time they have a malaria infection. Reassuringly, Yeka et al.  showed tWith regards to tolerability, there were no serious adverse events registered during the trial. Asthenia and anorexia were the most commonly recorded minor adverse events with a marked difference between the two drugs. It was shown by the results of this study that AL was better tolerated. During the study it was observed that the parents/caretakers would prefer that their child be randomized to AL rather than ASAQ as the former was perceived as more tolerable by the children. In fact, the chance to be randomized to receive AL seemed to be a major driving force to participate in the study. Although asthenia and anorexia may be classified as minor adverse effects, it can have a negative effect on adherence, and thus prevent a complete cure.There was a relatively high proportion (67\u00a0%) of children excluded from the study at the time of screening. There were two major reasons for exclusion: the high number of children with co-morbidities, and the high number of children who had taken ACT in the previous 28\u00a0days (either from a clinic or from a private vendor). In the children with co-morbidities, it is not clear what relation malaria had to their clinical illness; the children could be presenting with clinical illness due to malaria, or have symptoms primarily from their co-morbidity but happen to have malaria parasites. This is a dilemma that is faced often in the field, and may lead to an overestimation of the burden of malaria. Furthermore, an RDT for malaria is often performed as a first means of triage for fever, and when the test is positive, there is the risk that the child may not be adequately examined for other illnesses. In this study, 49 of the 873 (5.6\u00a0%) children with a positive RDT (based on HRP2 detection) had a negative thick smear. This could be the result of circulating HRP2 from a previous, well treated malaria infection. This is expected considering that Grandesso et al.  have shown that over 50\u00a0% of HRP2 RDTs remain positive more than 6\u00a0weeks after adequate treatment of a malaria infection.ASAQ is still effective as a treatment for uncomplicated malaria in Baraka, South Kivu, DRC, 9\u00a0years after its introduction as a first-line treatment. There is no evidence that ASAQ is inferior to AL in this setting with the non-inferiority margin of 7\u00a0% that was set for the current study. AL may have higher efficacy but additional trials are required to draw this conclusion with confidence. There is a high rate of re-infection in South-Kivu within six weeks of adequate treatment of either ASAQ or AL, indicating intense malaria transmission. AL was better tolerated by patients than ASAQ. Although earlier estimates of anti-malarial drug efficacy are unavailable for the current population, the high efficacy of ASAQ makes it unlikely that declining drug efficacy is a reason for the apparent increase in malaria cases observed in the region."}
+{"text": "Plasmodium falciparum to chloroquine was conducted from January 2013 to March 2015 in six localities in the south of Haiti.Malaria is considered a public health priority in Haiti, with a goal to eliminate by year 2020. Chloroquine is the first-line treatment recommended by the Ministry of Public Health and Population. In order to verify the suitability of chloroquine for uncomplicated malaria treatment, an in vivo study of susceptibility of P. falciparum malaria were included in the study and followed until day 28 after having taken 25\u00a0mg/kg of chloroquine orally over 3 days. The sample included 28 children under the age of 10, 9 adolescents aged 10\u201319\u00a0years, and 24 adults aged 20\u00a0years and over. Among them, 30 were monitored on day 3 (49%) and 33 on day 28 (59%). Clinical and parasitological monitoring was carried out on day 7 on 28 subjects, on day 14 on 13 subjects and on day 21 on 18 subjects. Residual parasitaemia with presence of trophozoites was found in 7 of 30 subjects on day 3 (23%), and in 6 of 28 subjects on day 7 (21%) who had a temperature less than 37.5\u00a0\u00b0C. These patients can be considered as late parasitological failures. All monitoring performed on day 28 was negative. Gametocytes were found in 3 patients (9%) despite the use of primaquine. The continuing low parasitaemia on day 3 and 7 in more than one fifth of cases raises the question of the efficacy of chloroquine in southern Haiti.Sixty-one patients who presented with confirmed P. falciparum malaria cases in southern Haiti. Consequently, there is a need to strengthen malaria treatment surveillance and to study the effectiveness of chloroquine in Haiti by monitoring patients after treatment.Results suggest a decrease of chloroquine susceptibility for treatment of  Plasmodium falciparum, and strains are believed to be susceptible to chloroquine, which is still often used in therapy, and remains the first-line treatment recommended by the Minist\u00e8re de la Sant\u00e9 Publique et de la Population . Surveillance information of patients treated for malaria in Haiti is very limited. Recent studies have found P. falciparum isolates carrying chloroquine resistant genes [P. falciparum was conducted from January 2013 to March 2015 in three departments in Haiti .Malaria is considered a public health priority in Haiti \u20134, even nt genes , 7. Addint genes . In ordeThe study was carried out in six coastal sites in three geographic departments in Haiti Fig.\u00a0. In thesP. falciparum mono-infection with a parasitaemia \u2265to 1000 asexual forms per microlitre of blood , axillary temperature \u226537.5\u00a0\u00b0C on the day of the survey or fever and chills within the prior 24\u00a0h, no signs of severe or complicated malaria, and no anti-malarial treatment within the previous 2 weeks. Study objectives were explained in Haitian Creole and patients were included after having given their written consent, or that of a parent or legal guardian for minors.Study participation included febrile patients and villagers with an axillary temperature greater or equal to 37.5\u00a0\u00b0C at baseline, and those who had a fever with chills within the prior 24\u00a0h. Inclusion criteria were: P. falciparum and non-P. falciparum malaria  was performed. Questioning and a clinical examination looking for signs of malaria were conducted by the same physician (CPR) among the positive patients included in the study. For each subject, a capillary blood sample was taken from the fingertip to make a thick and thin blood smear. Research, identification and counting of sexual and asexual forms of P. falciparum were conducted at 1000\u00d7 magnification. Subjects with less than 1000 asexual forms per microlitre were excluded from the study. Blood was collected on Whatman 903\u00ae filter paper for genotyping of the parasite strain (results not yet published). Patients were treated with chloroquine and primaquine according to dosage recommended by the Ministry of Public Health and Population, namely, 600\u00a0mg of chloroquine and 60\u00a0mg of primaquine at baseline (D0) and 300\u00a0mg of chloroquine on day 1 and 2 for adults. Children received 10\u00a0mg/kg at baseline and 5\u00a0mg/kg on day 1 and 2. After administration of treatment, patients were kept under observation for 30\u00a0min to verify absence of vomiting. In case of vomiting before 30\u00a0min passed, a new dose was administered.A rapid diagnostic test (RDT) for Whenever possible, subjects were monitored on day 1, 2, 3, 7, 14, 21 and 28, axillary temperature was monitored, a clinical exam and a thick smear were performed. The criteria for assessing treatment efficacy were those defined by the World Health Organization (WHO)  to distiP. falciparum was performed compared to 200 leukocytes and evaluated by microlitre of blood while considering a number of 8000 leukocytes per microlitre of blood.Thick and thin blood smears were stained using the Giemsa method and examined under a microscope. The count of asexual forms of P. falciparum detected by a fever greater than 37.5\u00a0\u00b0C and/or a positive RDT confirmed by microscopy of the parasite on the thick smear were enrolled. Only 61 patients met the inclusion criteria, nine having too low of parasitaemia levels (between 261 and 933 trophozoites/\u00b5L), and one subject was lost after day 1. The 61 retained subjects came from six sites across three geographic regions (departments) in the south of Haiti . The distribution of patients according to age and parasitic density is given in Table\u00a0P. falciparum trophozoites in the thick smear, however, in a smaller quantity than on day 0 and without clinical signs (temperature <37.5\u00a0\u00b0C). Six out of 28 still had P. falciparum throphozoites on day 7, but with an axillary temperature <37.5\u00a0\u00b0C.Of the 61 patients included, 30 were monitored on day 3 (49% of the sample). Seven (23.3%) still showed By monitoring at a distance, 13 patients were reviewed at day 14, 18 patients at day 21, and 33 patients at day 28 at the end of the study. Two 2-year-old children showed persistence of trophozoites in the thick smear until day 21, but with an axillary temperature <37.5\u00a0\u00b0C. At day 28, all 33 controlled subjects had a negative thick smear without trophozoites and an axillary temperature <37.5\u00a0\u00b0C. Only 3 of them were still gametocyte carriers. During clinical follow-up, we observed the appearance on day 28 of a level 1 spleen according to the Hackett classification in a 6-year-old girl and a 72-year-old woman.P. falciparum malaria cases in southern Haiti. Six patients failed therapy. As chloroquine is inexpensive and commonly used in Haiti this study has important implications. As stated by the World Health Organization [The results of this study showed a decrease of chloroquine susceptibility for treatment of nization , high panization , 5, it sClose monitoring of patients during treatment resulted in a rapid decrease of parasitic density in four out of five cases. Two 2-year-old children in Anse-\u00e0-B\u0153uf retained trophozoite parasitaemia on day 14 and 21, but not on day 28 when only gametocytes were found. Throughout the entire follow-up period, their axillary temperature was <37.5\u00a0\u00b0C. For one patient, parasitaemia decreased dramatically, going from 3579 trophozoites/\u00b5L at baseline to 204/\u00b5L at day 14\u201344/\u00b5L at day 21. For the other, there were 1280 trophozoites at baseline and parasitaemia remained at this same level on day 7, 14, and 21. These two cases can thus be considered as two late parasitological failures.Among all patients, the majority or most of the clinical symptoms noted on day 0 disappeared as early as the day 2 or 3 monitoring, except for the pallor of the mucous membranes, attesting to the frequent anemia among children in these villages. The fever decline was rapid: while nearly half of the patients had an axillary temperature between 37.5\u00a0\u00b0C and 40.8\u00a0\u00b0C at baseline, only two of 31 patients had, respectively, an axillary temperature of 37.9\u00a0\u00b0C and 38.1\u00a0\u00b0C on day 1, only one out of 25 on day 2 (37.5\u00a0\u00b0C), and one out of 30 on day 3 (38.1\u00a0\u00b0C). Afterward, all patients that were monitored weekly until day 28 had an axillary temperature of less than 37.5\u00a0\u00b0C.P. falciparum isolates remain susceptible, in vivo, to chloroquine in Haiti, as was observed some 30 years ago [Through questioning and examination, the improvement of clinical status was observed among all patients during monitoring, indicating that chloroquine remains effective in Haiti. This study shows that the majority of ears ago , 12 and ears ago .P. falciparum haplotypes carrying chloroquine resistant genes in Artibonite [However, in a recent study , the pertibonite .Two major difficulties were encountered during this study. The first was the recruitment of cases that met the pre-established inclusion criteria. It is currently difficult to recruit malaria cases in Haiti, except in a few specific areas such as Anse-\u00e0-B\u0153uf in the Sud-Est department (N\u00b06 on the map in Fig.\u00a0P. falciparum parasites.The second limitation concerns the routine monitoring of included subjects. Despite their consent to participate in the study, some subjects were lost and did not return to their follow-up appointments, which resulted in variation of the days where follow-up monitoring was performed. This loss of patients was experienced less in the fishing villages of Anse-\u00e0-B\u0153uf and Bariadelle, where it was easier to find the subjects in their home, in case they missed monitoring appointments. However, despite this limitation, the study still indicates the decrease of chloroquine susceptibility by P. falciparum throughout the country in order to identify the regions affected by these failures in chloroquine treatment. This information is critical in determining appropriate therapeutic and control measures in Haiti.The implications of this study highlight the need to strengthen surveillance of decreasing in vivo susceptibility of"}
+{"text": "ABCB1 c.3435C>T, CYP3A4*1B (g.-392A>G), CYP3A5*3 (g.6986A>G) SNPs and artemether\u00a0+\u00a0lumefantrine treatment outcome in 103 uncomplicated malaria patients from Angola. No significant associations with the CYP3A4*1B and CYP3A5*3 were observed, while a significant predominance of the ABCB1 c.3435CC genotype was found among the recurrent infection-free patients (p\u00a0<\u00a00.01), suggesting a role for this transporter in AL inter-individual performance.Malaria treatment performance is potentially influenced by pharmacogenetic factors. This study reports an association study between the  Artemisinin combination therapy (ACT) has contributed to the remarkable decline by 48% in the malaria mortality rate between 2000 and 2015 [Artemether\u2013lumefantrine (AL) is the most adopted antimalarial by national malaria control programs worldwide. In Angola, it represents the first-line treatment of choice for uncomplicated malaria.Upon AL oral administration, artemether shows an elimination half-life of 1\u20133\u00a0h, CYP3A4 being the main enzyme involved in its conversion towards the  dihydroartemisinin (DHA) metabolite . Only\u00a0<1CYP3A4 and ABCB1 pharmacogenetic characteristics influencing AL performance.Both lumefantrine and DHA are essentially eliminated through the bile. In the apical biliocanalicular membrane of the hepatocyte, the ABCB1 (MDR1/Pgp) ATP-binding cassette (ABC) transporter is a major biliary efflux pump, particularly for lipophilic substrates, as lumefantrine , 8. SignCYP3A4 and ABCB1 tag SNPs, particularly the promoter located g.-392A>G in the former (CYP3A4*1B) and the synonymous c.3435C>T in the latter, did not yield significant associations [ABCB1 c.3435C>T with altered LUM exposure among HIV positive subjects under Efavirenz based therapy.A previous attempt to correlate lumefantrine pharmacokinetic (PK) parameters with In the present work, we hypothesized that small pharmacokinetic differences might have observable pharmacodynamic consequences, the parasite reaction being a more sensitive parameter of the individual pharmacogenetic influence. Parasite clearance on day 3 post-treatment, recurrent infection prevalence and the 28-day cure rate endpoint of adequate clinical and parasitologic response (ACPR) were herein used as parameters to assess the effect of the patient pharmacogenetic status on AL in vivo anti-parasite performance. To test this hypothesis, a previously performed AL efficacy trial was analysed.Plasmodium falciparum malaria involved in an AL  efficacy trial in the Luanda region, Angola, conducted during the 2011\u20132013 period [2, D3, D7, D14, D21 and D28. At each time-point, thick blood films were examined for the presence of parasites and a capillary blood sample obtained for PCR analysis.One-hundred and three unrelated patients with microscopy confirmed  uncomplicated 3 period . ClinicaEthical approval was obtained from the Angolan National Public Health Institute/Ministry of Health Ethics Committee. All procedures followed the latest version of the Declaration of Helsinki.\u00ae Classic Card, Whatman). DNA extraction was done by phenol\u2013chloroform methods. The ABCB1 c.3435C>T, CYP3A4\u00a0g.-392C>G and CYP3A5\u00a0g.6986A>G SNPs were analysed by established PCR\u2013RFLP protocols [pfmsp2 PCR amplification of all samples at all time spots [pfmsp2 positive PCR by D3, informative concerning the artemisinin partner performance, in accordance with WHO guidelines; and, (b) pfmsp2 positive PCR during the 28-day follow up (lumefantrine prophylactic effect). The number of clinical failure events (PCR-corrected recrudescence) was too small to warrant their specific analysis. All recrudescences were included in the general recurrence group.Capillary blood sample were collected on filter paper  test and Z statistics were used to determine significant differences between proportions . Five individual associations were herein tested: recurrence vs CYP3A4*1B allele status, recurrence vs ABCB1 c3435C>T SNP status, recurrence vs CYP3A5*3 SNP status, D3 positivity vs CYP3A4*1B SNP status, and D3 positivity vs ABCB1 c.3435C>T SNP status). Accordingly, a Bonferroni-corrected significance threshold at p\u00a0<\u00a00.01 was considered for these tests. Multivariate Log-linear analysis was performed in order to specifically investigate associations of key three categorical variables: (a) D28 follow-up outcome (recurrence), (b) ABCB1/MDR1 c.3435C>T genotype, and (c) pfmdr1 N86Y status of the initial infection. The multiple testing was Benjamini\u2013Hochberg corrected, assuming a 10% false discovery rate (q).Data on SNPs were analysed using IBM SPSS version 23. Chi square  had positive PCR. During the 28-day follow up, 29/103 patients experienced recurrent parasitaemia, as detected through PCR. Ninety-eight patients were successfully analysed for the CYP3A4 -392A>G SNP. The genotype frequency in this Angolan population was 0.112  for the wild type (g.-392AA), 0.541  minor allele homozygous (g.-392GG) and 0.347  for the heterozygous (g.-392AG). The population was found in Hardy\u2013Weinberg equilibrium for this locus (p\u00a0>\u00a00.05).The 28-day PCR-corrected cure rate was 91.3%. On DCYP3A4*1B genotype was not found to be significantly associated with either D3 parasite positivity    for the wild type (c.3435CC), 0.079  for the minor allele homozygous (c.3435TT) and 0.158,  for the heterozygous (c.3435CT). The population was found in Hardy\u2013Weinberg equilibrium for this locus (p\u00a0>\u00a00.05).Concerning the c.3435C>T SNP was also not significantly associated with the D3 parasite PCR positivity   (Table\u00a0The CYP3A5*3 (c.6986A>G) in 84 samples, as this is the allele is the most investigated as having a robust deleterious effect in the expression of the gene [c.6986AA), 0.048,  for the heterozygous (c.6986AG) and 0.202  for the minor allele homozygous (c.6986GG). The sample population was found in Hardy\u2013Weinberg equilibrium for this locus (p\u00a0>\u00a00.05).During the completion of the present work, a new report has suggested CYP3A5 as a contributor to lumefantrine metabolism . Followithe gene . GenotypCYP3A4*1B, no significant association was observed between the patient status for carrying a CYP3A5*3 alleles and the parasitological outcome during the 28\u00a0day follow up , namely the 2\u00a0=\u00a010.84, df\u00a0=\u00a04, p\u00a0=\u00a00.0284 vs p (corrected)\u00a0=\u00a00.0286), and the association between recurrence during follow-up and the ABCB1 c3435C>T status \u00a0=\u00a00.0143), after filtering the effect of the presence/absence of the pfmdr1 N86 allele  of 10% for Benjamini\u2013Hochberg multiple test correction, only two associations stood out as near the threshold of significance: the overall interaction between the three analysed variables (GCYP3A and ABCB1 pharmacogenetics and AL pharmacodynamics in vivo endpoints. D3 positivity was not significantly associated with the patient CYP3A4*1B status (Table\u00a0P. falciparum parasites. Pharmacogenetic driven variable rates of artemether to DHA bioconversion are likely not to be readily visible in terms of the artemisinin effect on the infection. As for the ABCB1 c.3435C>T, the negative observations possibly result from the specific capacity of this transporter to efflux more lypophilic compounds then the final phase II glucuronidated DHA extracted from the liver.The present work focused on finding links between patient CYP3A4*1B and CYP3A4*3 status were not seen to influence the risk of malaria recurrence. The AL post-treatment protective effect is essentially related with the action of lumefantrine, the long half-life partner. In regular conditions, it is expected that the large majority of lumefantrine is eliminated unchanged [CYP3A4*1B and CYP3A5*3 SNPs in modulating AL post-treatment prophylactic action.The nchanged , a resulnchanged . This menchanged . Also, inchanged -and as sABCB1 c.3435TT genotype was found among patients suffering recurrent infections during the 28-day follow up, suggesting a role of the encoded P-glycoprotein. The synonymous c.3435C>T polymorphism has been proposed to be linked with altered rates of protein synthesis, leading to proteins that albeit having the same primary sequence, emerge from the process of translation with different tertiary conformations [ABCB1 c.3435CC genotype among the recurrence-free patients, signalling an increased lumefantrine exposure associated with this genotype, which better shielded the recovering patients from new infections.A significant increase in the frequency of the rmations . The funABCB1 c.3435TT genotype, was suggested to be associated with a significantly decreased D7 lumefantrine levels among patients undergoing malaria treatment with AL. Such an effect in drug exposure can explain the increase risk of these patients towards recurrent infection.A shortcoming of the present study is the unavailability of pharmacokinetic data, namely D7 LUM levels, in order to have a complete pharmacokinetic/pharmacogenetic picture. Nevertheless, it is interesting to note that the present results are in agreement with recent data from Maganda et al. , where tThese data suggest lumefantrine as part of the group of ABCB1 substrates where this genotype is associated with increased drug exposure, probably due to a less efficient efflux. Other examples include tacrolimus , 23, silABCB1 c.3435C>T SNP in the performance of artemether\u2013lumefantrine. The present observations join other recent reports pointing for the importance drug transporter pharmacogenetics in ACT pharmacokinetics and pharmacodynamics [By exploring potential pharmacodynamics/pharmacogenetic associations in anti-malarial therapy, this report shows a non-negligible influence of the host dynamics , 27."}
+{"text": "Plasmodium falciparum malaria in Ethiopia. This was done by performing a meta-analysis of recent studies conducted in the country on this topic.As Ethiopia is one of the sub-Saharan countries with a great burden of malaria the effectiveness of first line anti-malarial drugs is the major concern. The aim of this study was to synthesize the available evidence on the efficacy of artemether-lumefantrine in the treatment of uncomplicated P. falciparum malaria in Ethiopian patients were searched for using the PubMed and Google Scholar databases. Ten prospective single-arm cohort studies that followed patients for 28\u201342\u00a0days were included in this analysis. All of the included studies were deemed to be of high quality.Studies published between January 2010 and January 2017 that reported on the efficacy of artemether-lumefantrine in the treatment of Ten studies involving 1179 patients that were eligible for meta-analysis were identified. At recruitment, the average parasite count per patient was 1 2981/\u03bcl of blood. On the third day of treatment, 96.7% and 98.5% of the study subjects become fever-free and parasite-free, respectively. Based on the per protocol analysis, the cure rate after use of artemether-lumefantrine was 98.2% (polymerase chain reaction corrected) and 97.01% (polymerase chain reaction uncorrected) after 28\u00a0days of follow-up. The reinfection rate within 28\u00a0days was 1.1% and the recrudescence rate was 1.9%.P. falciparum malaria using artemether-lumefantrine in Ethiopia is still high enough to recommend the drug as a first-line agent. There should be careful periodic monitoring of the efficacy of this drug, as treatment failure may occur due to resistance, sub-therapeutic levels that may occur due to non-adherence, or inadequate absorption.This review found that the cure rate for uncomplicated The online version of this article (10.1186/s40249-017-0372-5) contains supplementary material, which is available to authorized users. Please see Additional\u00a0file\u00a0Malaria is one of the major public health problems in the world. Its burden is very high in Sub-Saharan Africa, where about 90% of all malaria deaths occur . Each miPlasmodium falciparum and P. vivax; roughly 60% of malaria cases in the country are due to the former [P. falciparum in Ethiopia is about 33% in children below 12\u00a0years of age and 10% in hospitalized adults [Malaria is an important communicable disease affecting large numbers of people in Ethiopia. About 68% of the Ethiopian population lives in high-risk malaria areas . The come former , 5. An eP. falciparum strain was the main threat to malaria prevention and control in Ethiopia [P. falciparum malaria in 2004 [Early diagnosis and timely treatment of malaria with an effective drug is an important strategy to control the disease . HoweverEthiopia . A failuEthiopia , 10. HowEthiopia . After tEthiopia  and the Ethiopia , 13, the in 2004 . At that in 2004 .The commThe World Health Organization (WHO) recommends above 90% parasitological and clinical curative rates for an antimalarial drug to be approved as a first-line agent . There aP. falciparum malaria in Ethiopia.The WHO recommends that the efficacy of first-line antimalarial drugs should be checked at least every two years . As EthiP. falciparum malaria in Ethiopia was searched for using the online databases PubMed (https://www.ncbi.nlm.nih.gov/pubmed/) and Google Scholar (https://scholar.google.com/).Literature published between January 2010 and January 2017 that assessed the efficacy of AL in the treatment of uncomplicated Plasmodium falciparum malaria\u2019, \u2018antimalarial drug\u2019, and \u2018Ethiopia\u2019. Selected reference lists of retrieved articles were also searched manually using Google.The search was conducted by combining the following words: \u2018efficacy\u2019, \u2018therapeutic efficacy\u2019, \u2018artemether-lumefantrine\u2019, \u2018Coartem\u2019,\u2018cohort\u2019, \u2018in vivo\u2019, \u2018P. falciparum mono-infected Ethiopian patients and which were published between January 2010 and January 2017 were included in this review. Studies done on non-human subjects and articles of which the full text could not be obtained were excluded.The author reviewed all of the identified articles in order to assess eligibility for inclusion based on predefined criteria. Studies done to assess the efficacy of AL on Validity and methodological quality of all included studies were assessed according to the National Institutes of Health (NIHs) quality assessment tool for observational cohort studies.  The toolRelevant information such as study design, study setting, follow-up period, sample size, baseline characteristics of study subjects, fever clearance, parasite clearance, treatment failure, and cure rates were extracted from each article using the well-prepared and piloted data extraction format in the form of table.Meta-analysis was performed using Comprehensive Meta-Analysis software version 2.2.064 . ContinuA total of 235 articles were initially retrieved. After excluding duplicates, the titles of 212 articles were assessed and 197 were found to be irrelevant. Abstracts of the remaining 15 articles were checked to determine if they fulfilled the inclusion criteria. Of those, five were rejected: two were done on non-human subjects, one was published before 2010, and the full text was not available for the other two. In the end, 10 articles were found to be suitable to be included in this study. Figure\u00a0P. falciparum malaria in Ethiopia.All 10 studies included in this review were prospective single-arm cohort studies that determined the efficacy of AL in the treatment of uncomplicated The studies were conducted in different malarious parts of the country . The WHO guide for surveillance of antimalarial drug efficacy was used in all of the studies to select subjects and to conduct the study.P. falciparum mono-infection were included. Most  of the studies followed patients who were older than six months. One study excluded patients who were below one year of age and another excluded children under five years of age. Details of each study are summarized in Table\u00a0In the majority 8/10, 80%) of the studies, subjects were followed for 28\u00a0days, while in the remaining  studies, the follow-up period was 42\u00a0days. In each study, anywhere from 66 to 384 patients with 0, 80% ofEach article was evaluated against 14 criteria using the NIHs quality assessment tool for observational cohort studies . FortunaThe 10 studies included a total of 1179 participants from 14 study sites. The mean age of study participants was 15.8\u00a0years. At recruitment, the average parasite count per patient was 12,981/\u03bcl of blood and gametocytes were found in 7.7% of patients. Table\u00a0Fever and parasite clearance was rapid. On the third day of treatment, 96.7% and 98.5% of study subjects become fever-free and parasite-free, respectively. There was also a significant decrease in gametocyte carriage from 7.7% at baseline to 0.4% on the 28th day of treatment. Table\u00a0Of the 1179 subjects included in the 10 studies reviewed, only 27 (2.29%) showed treatment failure. As shown in Table\u00a02\u00a0=\u00a038.8). As shown in the forest plots provided in Figs.\u00a0Heterogeneity between the studies was minimal I^\u00a0=\u00a038.8. P. falciparum cases have been reported in the country [P. falciparum malaria in Ethiopia.AL has been the first-line drug used in Ethiopia for the treatment of uncomplicated falciparum malaria since 2004 after the development of resistance for older antimalarials such as chloroquine and sulphadoxine-pyrimethamine . Since i country , 32, 38.P. falciparum malaria reduces the number of patients who require paracetamol during the follow-up period, which means that the number of drugs taken by patients and thus the risks associated with them is reduced. Other studies also confirm the rapid fever clearance capacity of AL [The mean temperature of patients on the day of enrolment was 38.2\u00a0\u00b1\u00a00.35\u00a0\u00b0C. Closer mean temperature at day zero was reported in similar studies conducted in Sudan (38.5\u00a0\u00b1\u00a00.6\u00a0\u00b0C)  and Zambty of AL \u201343. ThisFrom the 1179 patients included in the 10 studies, 1161 (98.5%) had parasitic clearance within the first three days of treatment with AL. This rate of parasite clearance was consistent with other reports \u201347. In aIn contrast to the finding of the current review, a slower parasite clearance rate and increased third day parasitemia was reported in studies conducted on the Thai-Cambodian border , 24.The P. falciparum malaria. Similar findings were reported by other studies conducted in different parts of the world [The PCR corrected cure rate of AL in the current review was found to be 98.2%. This reveals that AL is highly effective in the treatment of uncomplicated he world , 53\u201355. he world \u201359. A mehe world . A commuhe world .P. falciparum malaria. Therefore, in Ethiopia AL can still be used as a first-line agent in the treatment of uncomplicated P. falciparum malaria. This will not, however, guarantee the avoidance of subsequent timely monitoring for its efficacy. The WHO has said that the efficacy of first-line medicines must be cheeked at least every two years [The WHO recommends that for an antimalarial drug to be selected as a first-line treatment agent, it should have a clinical and parasitological cure rate of 90% or higher . The reswo years .The follow-up period in most of the studies reviewed was 28\u00a0days. Two of the studies followed patients for 42\u00a0days and one of them reported that the rate of recrudescence was higher at day 42 than at day 28  . This inIn all of the 10 studies reviewed, drug levels were not tested, therefore it is difficult to attribute treatment failure to the drug\u2019s ineffectiveness or development of resistance, as it may be due to insufficient drug levels. It is thus better if future studies conducted on AL efficacy measure the serum drug levels and correlate these with the outcome. Sensitivity of PCR genotyping may have also affected the results of this review, as similar strains, defined as recrudescence, could be new infections, especially in low to moderate transmission areas with a limited diversity of strains .P. falciparum population.Even though the morning doses of AL were taken in front of investigators in most of the studies, the night time intake of the drug was not directly controlled. Therefore, it was difficult to determine whether some of the treatment failure cases reported were due to low adherence. There are also many other factors that may influence the treatment outcome. Further studies that identify risk factors for treatment failure should be conducted. Studies that describe in detail the characteristics of patients with recrudescence are also very important. Molecular surveillance may also play an important role in detecting genetic markers associated with AL resistance in the local P. falciparum malaria even after a number of years of its widespread use in the country. There should, however, be careful periodic monitoring of the efficacy of this drug, as treatment failure may occur due to resistance, sub-therapeutic levels that may occur due to non-adherence, or inadequate absorption.AL is highly efficacious in Ethiopia for the treatment of uncomplicated Additional file 1:Multilingual abstracts in the five official working languages of the United Nations. (PDF 799 kb)Additional file 2:Treatment outcome from individual study. (DOCX 18 kb)"}
+{"text": "Plasmodium falciparum malaria with either pyronaridine\u2013artesunate (PA) or artemether\u2013lumefantrine (AL).Artemisinin-based combinations differ in their impact on gametocyte prevalence and density. This study assessed female and male gametocyte dynamics after treating children with uncomplicated Plasmodium falciparum malaria were included and randomly assigned to PA or AL treatment. Filter paper blood samples were collected as a source of RNA for quantitative reverse-transcription PCR (qRT-PCR) and nucleic acid sequence based amplification (QT-NASBA) to detect female gametocytes (targeting Pfs25 mRNA). Male gametocytes were detected by qRT-PCR (targeting PfMGET mRNA). Duration of gametocyte carriage, the female and male gametocyte response and the agreement between qRT-PCR and QT-NASBA were determined.Kenyan children with uncomplicated Pfs25 qRT-PCR . qRT-PCR based female gametocyte prevalence decreased from 100% (75/75) at baseline to 6.06% (4/66) at day 14 in the AL group and from 97.7% (83/85) to 13.9% (11/79) in the PA group. Male gametocyte prevalence decreased from 41.3% (31/75) at baseline to 19.7% (13/66) at day 14 in the AL group and from 35.3% (30/85) to 22.8% (18/79) in the PA group. There was good agreement between Pfs25 qRT-PCR and QT-NASBA female gametocyte prevalence .The mean duration of female gametocyte carriage was significantly longer for PA (4.9\u00a0days) than for AL (3.8\u00a0days) as estimated by QT-NASBA (P\u2009=\u20090.036), but this difference was less clear when determined by This study indicates that female gametocyte clearance may be slightly faster after AL compared to PA. Male gametocytes showed similar post-treatment clearance between study arms. Future studies should further address potential differences between the post-treatment transmission potential after PA compared to AL.Trial registration This study is registered at clinicaltrials.gov under NCT02411994. Registration date: 8 April 2015. https://clinicaltrials.gov/ct2/show/NCT02411994?term=pyronaridine-artesunate&cond=Malaria&cntry=KE&rank=1The online version of this article (10.1186/s12936-018-2373-7) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria, it considerably contributed to the decline of the disease burden [P. falciparum malaria and the blood stage of Plasmodium vivax malaria [Since artemisinin-based combination therapy (ACT) became widely adopted as first-line treatment for uncomplicated e burden \u20134. Howeve burden , 6. New  malaria \u201312. Mild malaria .P. falciparum (gametocytes), has not been extensively studied in the clinical setting. In vitro data are contradicting: a strong gametocytocidal effect of pyronaridine against stage II\u2013IV gametocytes has been found [So far, the effect of PA on the transmission stages of en found , but wasen found . Delves en found , 16. Witen found , 17\u201319. en found . Among pPfs25 [Pfs25) and male (PfMGET) gametocytes [To accurately evaluate the gametocyte response after ACT treatment, molecular tools are informative since post-treatment gametocyte densities are often below the detection threshold of microscopy . QuantitPfs25 , 23. Recetocytes . This dietocytes . Thus, fPfs25 qRT-PCR and QT-NASBA for the detection of female gametocytes was determined.In this study, the QT-NASBA and qRT-PCR based female specific gametocyte response after PA-treatment was compared to that after AL. Furthermore, qRT-PCR was used to evaluate and compare male and female gametocyte dynamics. Finally, the agreement between P. falciparum malaria [P. falciparum mono-infection with a parasitaemia between 1000 and 200,000/\u00b5l. Exclusion criteria were signs and symptoms of complicated malaria, non-P. falciparum or mixed Plasmodium infection, a history of hepatic and/or renal impairment, a haemoglobin (Hb) concentration\u2009<\u20096\u00a0g/dL, severe malnutrition (defined as having a weight-for-age or height-for-age z-score of\u2009<\u2009\u2212\u20093) [This observational study was part of a phase III randomized clinical trial investigating the efficacy and safety of PA compared to AL in Kenyan children with uncomplicated  malaria . The stuf\u2009<\u2009\u2212\u20093) , having Study participants were randomized to receive a 3-day course of either artemether\u2013lumefantrine  or pyronaridine\u2013artesunate . All study staff except the pharmacists responsible for drug administration were blinded to treatment allocation. Dosing was body-weight dependent and drugs were administered according to manufacturer\u2019s instructions . Giemsa-stained thick smears were used for determination and counting of asexual parasites and gametocytes, according to WHO procedures [Pfs25 QT-NASBA and sex-specific Pfs25 and PfMGET qRT-PCR were used for gametocyte detection on day 0, 3, 7 and 14. To perform these assays, 2\u2009\u00d7\u200950\u00a0\u00b5l finger-prick blood was collected on Whatman 903 protein saver cards , dried at room temperature for 24\u00a0h, packed individually with silica and stored at \u2212\u200920\u00a0\u00b0C until shipment to the Netherlands. Nucleic acid extraction was done by Nuclisens EasyMag  and DNA/RNA was stored at \u2212\u200970\u00a0\u00b0C. QT-NASBA was performed as previously described [3 to 10\u22121 gametocytes/\u00b5l, which were produced from in vitro cultures as reported [3 to 10\u22122 gametocytes/\u00b5l) for quantification [Female specific escribed , with mireported . qRT-PCRfication . Samplesfication .\u22121 days), and gametocyte prevalence and density on day 3, 7 and 14 as determined by QT-NASBA and Pfs25/PfMGET qRT-PCR. Finally, the agreement between QT-NASBA and Pfs25 qRT-PCR for the detection but not quantification of female gametocytes was determined.The primary outcome was the mean duration of female gametocyte carriage in the PA arm compared to the AL arm, based on QT-NASBA. Secondary outcomes were: the qRT-PCR based mean duration of female gametocyte carriage, the QT-NASBA and qRT-PCR based female gametocyte circulation time, the QT-NASBA based area under the curve (AUC) of female gametocyte density over time  based on variance components, taking repeated measures into account [Stata software version 14.0  and SAS version 9.4  were used for statistical analyses. A deterministic compartmental model, as previously published, was fitted to determine the duration of female gametocyte carriage and gametocyte circulation times . The AUC account , 34.A consecutive subset of 160 children from the main clinical trial participated in the present study. Of these 160 participants, 85 received PA and 75 received AL .Baseline characteristics were similar between intervention groups Table\u00a0. QT-NASBPfs25 qRT-PCR  in the PA group and 31.0% 22/71) in the AL group (P\u2009=\u20090.433). At day 7, prevalence decreased to 21.7% (18/83) in the PA group and 16.7% (12/72) in the AL group (P\u2009=\u20090.430). Prevalence at day 14 was 15.2% (12/79) in the PA group and 7.58% (5/66) in the AL group (P\u2009=\u20090.156). Female gametocyte prevalence estimates were highly similar when assessed by 2/71 in tWhile prevalence estimates were comparable between QT-NASBA and qRT-PCR, some differences in density measurements were observed. As can be seen in Fig.\u00a0The mean duration of female gametocyte carriage as estimated by QT-NASBA was significantly longer in the PA group , compared to the AL group  (P\u2009=\u20090.036). By qRT-PCR, the mean duration of female gametocyte carriage was also longer in the PA group , compared to the AL group , but this difference was not significant (P\u2009=\u20090.166). Similarly, the QT-NASBA based mean gametocyte circulation time was longer for PA  compared to AL  (P\u2009=\u20090.003). This was also the case for the qRT-PCR based mean gametocyte circulation time   of participants in the PA group and 100% (75/75) of the participants in the AL group P\u2009=\u20090.499). This prevalence decreased to 13.9% (11/79) on day 14 in the PA group and 6.06% (4/66) in the AL group P\u2009=\u20090.122)  gametocytes/\u00b5l at baseline to 0.58 (IQR 0.30\u20131.66) on day 3 in the PA group. In the AL group, female baseline density was 1.94 gametocytes/\u00b5l (IQR 0.77\u20135.35), which decreased to 0.30 (IQR 0.14\u20130.62) on day 3. By day 14, median female gametocyte density was 0.33 (IQR 0.14\u20136.42) in the PA group and 0.12 (IQR 0.08\u20130.90) in the AL group (P\u2009=\u20090.322) . Out of 560, 12 samples were positive by QT-NASBA but not by qRT-PCR. Similarly, 10 samples were positive by qRT-PCR but negative by QT-NASBA. The two tests were in agreement for the remaining 538 samples (209 positive and 329 negative). The CCC was 0.85 (95% CI 0.82\u20130.87), indicating good agreement between Pfs25 qRT-PCR and QT-NASBA for the detection of female gametocytes.The agreement between binary female prevalence outcomes of This is the first paper describing kinetics of submicroscopic gametocytes after PA treatment using molecular detection methods in comparison with the most widely used first-line treatment for malaria in Africa, AL. The duration of female gametocyte carriage and gametocyte circulation time appeared to be slightly longer for PA compared to AL. There were no indications that PA or AL preferentially cleared male gametocytes.The failure of conventional anti-malarials, including ACT, to clear circulating mature gametocytes may allow persisting malaria transmission in the week(s) following treatment . GametocDifferent effects of ACT on the gametocyte response have previously been reported and in a recent meta-analysis AL was shown to be better in preventing the microscopic occurrence of gametocytes shortly after treatment compared to DP or AS-AQ . A pointBaseline prevalence of female gametocytes (estimated by qRT-PCR) was 98.8% (158/160), while male baseline prevalence was only 38.1% (61/160). This is in contrast to the data presented by Stone et al. , from thNo evidence of faster male compared to female gametocyte clearance was found. In fact, the present data suggest that even though the proportion of participants with male gametocytes at baseline was lower than that with female gametocytes, males may actually be cleared slower. Previous studies that examined gametocyte sex ratio after DP or SP-AQ alone or with primaquine observed that during the course of follow-up gametocyte sex ratios became more female-biased while primaquine initially resulted in a male-biased sex ratio , 36. In The apparent increase in male density  could not be explained by an absolute increase within individuals, but rather reflects a difference between the population positives on day 0 and that on day 14. Stone et al. performed a similar analysis and reported a small decrease of male density after DP treatment (from 3.8/\u00b5l at baseline to 0.9/\u00b5l at day 7) . Both stPfs25 qRT-PCR female gametocyte prevalence was observed. This confirms data from a previous study where both assays were shown to be suitable to detect and quantify submicroscopic levels of gametocytes, although the reproducibility of qRT-PCR was found to be better than that of QT-NASBA [A good level of agreement between QT-NASBA and QT-NASBA .Microsporidia species, which has been shown to inhibit the survival of Plasmodium in mosquitoes [A limitation of the present study is that gametocyte infectiousness to mosquitoes could not be established. This was due to an infection of the established mosquito colony with squitoes . Since oP. falciparum malaria. These data may contribute to estimates of impact differences between artemisinin-based combinations, for example based on a model that demonstrated a higher reduction of clinical episodes using long-acting combinations in high transmission settings, while combinations with shorter half-lifes but more pronounced gametocytocidal effects were shown to be more suitable for low-transmission settings [This study provides important data on the submicroscopic gametocyte response after PA compared to AL treatment of uncomplicated settings .Additional file 1. Weight-based dosing of pyronaridine\u2013artesunate and artemether\u2013lumefantrine."}
+{"text": "Leishmania (Viannia) panamensis predominated among the strains isolated . Noncompartmental analysis demonstrated that plasma and intracellular miltefosine concentrations were, overall, lower in children than in adults. Exposure to miltefosine, estimated by area under the concentration-time curve and maximum concentration, was significantly lower in children in both the central and intracellular compartments (P < 0.01). Leishmania persistence was detected in 43% of study participants at the end of treatment and in 27% at 90 days after initiation of treatment. Clinical response was not dependent on parasite elimination. In vitro miltefosine susceptibility was similar for Leishmania strains from adults and children. Our results document PK differences for miltefosine in children and adults with cutaneous leishmaniasis that affect drug exposure and could influence the outcome of treatment, and they provide bases for optimizing therapeutic regimens for CL in pediatric populations. An open-label pharmacokinetics (PK) clinical trial was conducted to comparatively assess the PK and explore the pharmacodynamics (PD) of miltefosine in children and adults with cutaneous leishmaniasis (CL) in Colombia. Sixty patients, 30 children aged 2 to 12 years and 30 adults aged 18 to 60 years, were enrolled. Participants received miltefosine (Impavido) at a nominal dose of 2.5 mg/kg/day for 28 days. Miltefosine concentrations were measured in plasma and peripheral blood mononuclear cells by liquid chromatography-tandem mass spectrometry of samples obtained during treatment and up to 6 months following completion of treatment, when therapeutic outcome was determined. Fifty-two patients were cured, 5 pediatric patients failed treatment, and 3 participants were lost to follow-up.  Leishmania infection in children (Children with cutaneous leishmaniasis (CL) present several diagnostic and therapeutic challenges compared to adults with CL. These include greater pathogenicity of children , propenschildren , and subchildren . There ichildren . LimitedLeishmania species causing CL has been suggested by in vitro evaluations (Leishmania (Viannia) guyanensis and Leishmania (Viannia) braziliensis are endemic in Brazil  and CL \u201310, alth\u2013luations , 12. Milluations , and itsn Brazil , 9. Neven Brazil , 15. PK Leishmania parasites are intracellular pathogens, the drug concentration within host cells is critical to the direct antimicrobial effect. PK are generally determined in plasma under the assumption that systemic drug exposure is proportional to and predictive of exposure in target tissues/cells  species were isolated from 83% of adults and 70% of children. L. (V.) panamensis was the most prevalent species, at 95% (20/21) in children and 88% (22/25) in adults. L. (V.) braziliensis was isolated from one child and two adults.Sixty-three patients were assessed for eligibility; two did not meet inclusion criteria based on clinical laboratory analysis, and one declared unavailability for follow-up. Among the 60 enrolled participants, two adults and one child were lost to follow-up . In bothMiltefosine concentrations in plasma were detectable for up to 6 months following completion of treatment . IntraceCmax) than in plasma (50) for strains from a previously reported patient cohort  were lower (P < 0.01) in children than in adults was precluded by unavailability of \u22653 data points from the elimination phase. Although no significant differences in time to Cmax (Tmax) or elimination half-life were observed among groups, miltefosine plasma concentrations declined >40% between days 15 and 29 for 6 adults and one child, and this was reflected in the paired intracellular concentrations.Intracellular miltefosine concentration-time profiles followed those in plasma. Intracellular accumulation of miltefosine was evident, resulting in a 2-fold higher maximum concentration , which 90% (53/59) of participants attended . Although the sampling window established for visits was \u00b17 days, the median window of attendance was 2 to 3 days for any visit. Adherence to treatment based on diary and pill count was similar for children and adults . Adherence by two participants who did not return their diary (one adult and one child) could be assessed only by pill count, indicating \u226596.4% drug use. Additionally, one adult presented a discrepancy between reported doses and pill count; the adherence based on pill count was 76%. The previously mentioned decline in miltefosine concentration for six adult patients and one pediatric patient is discordant with the corresponding diary and pill count and could indicate nonadherence that was not perceived by self-reporting. Notably, these seven patients were cured.Leishmania in lesions before treatment was demonstrated in all patients. Parasite detection declined to 44% (26/59 patients) at end of treatment (EoT) based on aspirate or swab samples obtained from active or apparently healed lesions, and it diminished to 27% (16/59) at day 90  and measurable parasite loads at the lesion site. Likewise no relationship between parasite clearance and clinical outcome was evident.The presence and viability of t day 90 . Among pt day 90 . Neitherme point  and B. Nin vitro, 20% (9/45) were tolerant, and 15.6% (7/45) were classified as indeterminate. No differences were observed between the susceptibility profiles of strains isolated from children and adults . There was no apparent relationship between parasite drug susceptibility and clinical outcome.Among strains isolated from study participants, 64.4% (29/45) were susceptible to miltefosine d adults . LeishmaP = 0.052). Notably, treatment failure occurred only among children \u22647 years of age. No significant differences in time to cure were found between study groups  and 82.7% of children (24/29). Despite the higher frequency of therapeutic failure in children, this difference was not statistically significant (y groups . The logP = 0.29). Eighty-nine percent of clinical adverse events were mild (grade 1); no reported events were classified as serious. In no case did intolerance require interruption of treatment. The most frequent AE were nausea, vomiting, abdominal pain, and mildly increased creatinine levels (At least one adverse event (AE) was observed in 56% of participants; no differences in the frequency of AEs were detected between study groups (e levels . AEs werL. (V.) panamensis and Leishmania (Viannia) guyanensis . Future population-based compartmental modeling methods may allow more precise predictions of total exposure in plasma and peripheral blood mononuclear cells (PBMCs) in these patients and reveal an exposure-response relationship. We have previously proposed allometric dosing of miltefosine for VL patients, aiming to overcome differences in exposure observed by modeling of PK in children and adults . The lowIn addition to PK differences, other patient characteristics influence the outcome of treatment in CL patients \u201328. In o\u201328\u2013Cmax values of miltefosine in our study population, both children and adults, were similar to those reported in prior clinical studies from which PK were modeled  for clinical strains of Leishmania (Viannia)  .In vitro drug susceptibility did not differ among strains from patients who responded to or failed treatment. Similarly, miltefosine susceptibility did not differ for promastigotes of L. donovani strains from VL patients who failed or responded to treatment  and the Colombian National Institute for Food and Drug Safety (INVIMA) and followed international guidelines. All individuals participated voluntarily, providing written informed consent. Legal guardians of pediatric patients provided written informed consent, and children aged \u22657 years provided written informed assent .An open-label trial was conducted to determine the pharmacokinetics of miltefosine in children and adults with CL. Sixty patients, 30 children and 30 adults, were enrolled from January 2012 through October 2013 at CIDEIM outpatient clinics in Cali and Tumaco, Colombia. Timing intervals of blood samples to determine the PK of miltefosine were based on published PK models generated with data from adult patients with Old World CL . ClinicaEligible participants were adults aged 18 to 60 years and children aged 2 to 12 years  with parasitologically confirmed CL and availability for 6 months follow-up after treatment. Exclusion criteria were mucocutaneous disease, use of any antileishmanial drug during 6 months prior to diagnosis, medical history of cardiac, renal, or hepatic disease, menarche , pregnancy, and baseline values for hemoglobin, aspartate aminotransferase, alanine aminotransferase, creatinine, or serum urea nitrogen outside the normal range. In cases presenting borderline values, inclusion/exclusion was supported by clinical assessment. Contraception (Depo-Provera) was administered to women of reproductive age during treatment and throughout follow-up.Patients received a nominal dose of 2.5 mg/kg/day miltefosine rounded to the nearest 10- or 50-mg capsule, with a maximum dose in adults of 150 mg/day, during 28 days. Doses received were registered in an individual patient diary, and the number of capsules and empty blisters were counted during follow-up visits to assess adherence. Peripheral blood samples (10 ml for adults and 3 ml for children) were collected for isolation of plasma and peripheral blood mononuclear cells (PBMCs) for quantification of miltefosine. Blood samples were collected at eight intervals over 7 months: pretreatment, after the first day of treatment (day 2), and days 15, 29, 60, 90, 120, and 210. A sampling window of \u00b17 days was accepted for all visits except pretreatment, day 2, and end of treatment (day 29) visits. Clinical laboratory tests performed at baseline were repeated at the end of treatment to monitor potential drug-related toxicity.Adverse events (AEs), defined by clinical and laboratory criteria, were evaluated and recorded at every visit throughout treatment and follow-up. Severity of AEs was graded according to Common Toxicity Criteria of the National Cancer Institute, V.4 , and cauClinical response was evaluated at end of treatment and days 90 and 210. Cure was defined as complete reepithelialization and absence of inflammatory signs for all lesions. Definitive cure was established at day 210. Clinical failure was defined as incomplete reepithelialization and/or presence of induration, raised borders, or other evidence of inflammation of any lesion, reactivation of the original lesion(s), or appearance of new lesions during the follow-up period.https://cran.r-project.org/web/packages/ncappc/index.html).Miltefosine concentrations in plasma were determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) . Each anLesion/scar aspirates and swab samples were obtained at diagnosis and days 29 and 90. Duplicate samples were obtained from the border of the most recent lesion for independent RNA and DNA extraction. DNA was extracted using the DNA blood and tissue kit (Qiagen), and RNA was detected using TRIzol followed by RNA cleanup with the RNeasy extraction kit (Qiagen). Purified RNA was treated with DNase I and eluted in a total volume of 35 \u03bcl. cDNA was synthesized from 10 \u03bcl total RNA using the high-capacity cDNA reverse transcription kit (Applied Biosystems). The quantity and quality of nucleic acids were evaluated using a NanoDrop 2000 spectrophotometer.Leishmania kinetoplast DNA (kDNA) was amplified from samples by PCR using LV-B1 primers, followed by Southern blot hybridization (Leishmania 7SLRNA (dization . Amplifidization . Live paa 7SLRNA , calculaa 7SLRNA . QualitaLeishmania was isolated by culture of tissue fluid obtained by needle aspiration of cutaneous lesions. Parasites were identified using subgenus- and species-discriminating monoclonal antibodies (tibodies . Drug sutibodies .t tests. Differences in proportions were determined using \u03c72 or Fisher exact tests. Relationships between therapeutic outcome and Leishmania species, parasite burden, drug susceptibility of isolated strains, Cmax, AUCd0\u201329 in plasma and PBMCs, patient age, and size and number of lesions were explored using logistic regression. Analyses were performed using Stata-14.Data were verified by double entry prior to analysis. The Shapiro-Wilk test was used to assess the distribution of continuous data. Differences in variance of quantitative data were estimated using Mann-Whitney and"}
+{"text": "Plasmodium vivax malaria, but this is rarely implemented because of concerns over potential hemolysis in patients who have G6PD deficiency. Between August 2009 and February 2014, we conducted an open-label, randomized controlled trial of chloroquine (CQ) alone versus chloroquine plus primaquine (0.25 mg base/kg/day for 14 days) (CQ+PQ) in patients aged 6 months and older with microscopy confirmed P. vivax infection. In the CQ+PQ group, G6PD deficiency was excluded by fluorescent spot testing. The primary outcome was P. vivax recurrence assessed by survival analysis over one year follow-up. Of 593 patients enrolled, 570 attended at or after 14 days of follow-up. Plasmodium vivax recurrences occurred in 37 (13.1%) of 282 patients in the CQ+PQ arm versus 86 (29.9%) of 288 in the CQ arm  . Protection against recurrence was greater in the first 6 months of follow-up  than later . Five of seven patients requiring hospital admission were considered possible cases of PQ-related hemolysis, and PQ was stopped in a further six; however, in none of these cases did hemoglobin fall by \u2265 2 g/dL or to below 7 g/dL, and genotyping did not detect any cases of Mediterranean variant G6PD deficiency. PQ 0.25 mg/kg/day for 14 days prevents relapse of P. vivax in Afghanistan. Patient visits during the first week may improve adherence. Implementation will require deployment of point-of-care phenotypic tests for G6PD deficiency.Afghanistan\u2019s national guidelines recommend primaquine (PQ) for radical treatment of  Plasmodium vivax.2 Limited resources and security challenges hamper control efforts.2 Control of vivax malaria is particularly challenging because the dormant liver stages cause multiple relapses providing a source for new transmission.3 Trials of primaquine (PQ), the only widely available radical therapy, in Afghan refugees living in Pakistan documented good efficacy for PQ 0.25 or 0.5 mg/kg/day given over 14 days5 or 0.75 mg/kg/week for 8 weeks,5 compared with a 5-day PQ regimen.6 Although adherence to the 14-day treatment course is a concern, efficacy was preserved with an unsupervised regimen accompanied by appropriate instructions.4Malaria remains a significant problem in Afghanistan where cases are caused by 9 G6PD phenotypic tests are generally unavailable in Afghanistan as in many other tropical regions. Hence, administration of PQ according to the national guideline is rarely undertaken.The national treatment policy for vivax malaria in Afghanistan is chloroquine (CQ) plus 0.25 mg/kg/day PQ for 14 days. Unfortunately, PQ causes hemolysis in individuals with G6PD deficiency, a common red cell enzyme deficiency in malaria endemic regions.To assess the effectiveness of unsupervised PQ in preventing relapse of vivax malaria in Afghanistan and thereby support local evidence-based policy development, we compared the relative therapeutic effectiveness of CQ and chloroquine plus primaquine (CQ+PQ) in the treatment of vivax malaria at two provincial sites in Afghanistan.11 Malaria is seasonal with cases commencing in May, peaking in July and August, and rare by November.In Afghanistan, malaria is confined to the northern plains, the Jalalabad basin, and valleys fringing the central mountains to the west and south.The study was conducted from August 2009 to February 2014 at two provincial malaria control centers. Jalalabad is a referral center for the whole eastern region. Asadabad is the capital city of Kunar province, a largely mountainous area. Both areas border Pakistan with which there is uncontrolled movement.This prospective, open label, randomized controlled trial was conducted in patients aged 6 months and older with uncomplicated microscopy confirmed symptomatic vivax malaria. Clinical and demographic findings and vital signs were documented, including axillary temperature. Capillary blood was collected for hemoglobin measurement , Giemsa-staining of thick and thin blood films for parasite speciation and counting (40 \u00d7 number of parasites per 200 white blood cells on the thick film) and G6PD fluorescent spot test (see below). Parasitemia was considered negative after examination of 30 thick film high-power fields.P. vivax, axillary temperature \u2265 37.5\u00b0C or oral/rectal temperature \u2265 38\u00b0C or history of fever in preceding 24 hours, a negative urine pregnancy test in women at risk of pregnancy, ability to swallow oral medication and comply with study requirements, and written informed consent by the patient or attending parent/guardian. Exclusion criteria were any clinical or laboratory feature of severe malaria,12 hemoglobin concentration \u2264 8 g/dL, known G6PD deficiency, significant comorbidity, known hypersensitivity to any study drugs, mixed species Plasmodium infection and pregnancy or lactation.Inclusion criteria were monoinfection with asexual stages of The two treatment arms were CQ and CQ+PQ. Patients were allocated using a pregenerated randomization list in blocks of 20 held independently of the field teams by a statistician. The individual allocations were kept in sealed envelopes and opened only after enrolment. The microscopists but not the patients and clinical field workers were blinded to treatment arm.13Patients randomized to CQ+PQ were tested for G6PD deficiency before receiving antimalarial medication. Samples were tested using the G-6-PD OSMMR 2000  NADPH fluorescent spot test kit. When fluorescence was absent (indicating severe deficiency of G6PD) patients were withdrawn and given CQ alone with routine follow-up. Patients with normal fluorescence were treated with CQ+PQ (see below). All patients were advised to return as soon as possible if new symptoms developed (such as dark urine) or their condition worsened significantly. As far as possible, baseline dried blood spots were retained for later testing for the prevalent G6PD Mediterranean variant (563C>T) by polymerase chain reaction (PCR)-restriction fragment length polymorphism.Patients received quality-controlled CQ  aiming for a target total dose of 25 mg base/kg in divided doses over 3 days, with or without PQ (Bangkok GPO) 0.25 mg base/kg/day for 14 days. According to body weight, individual doses were rounded to the nearest quarter tablet or tablets were crushed, mixed with 5 mL water and the appropriate volume given. CQ and the first three PQ doses were observed at the malaria treatment center and patients remained for 60 minutes after drug administration. Patients vomiting within this time received the same dose again. Patients who vomited the medication twice were withdrawn from the study. Aside from the day 7 PQ dose (observed at the center), the remainder of the PQ doses were taken at home.PQ was stopped if there were significant symptoms suggesting hemolysis, hemoglobin concentration fell \u2265 2 g/dL, or absolute hemoglobin was < 7 g/dL at any time. Clinicians were free to stop PQ for clinical reasons outside these criteria. If necessary patients were admitted to hospital (a serious adverse event).15 Patients were also requested to attend if they felt unwell at other times during follow-up. At each visit a standard symptom questionnaire was completed. Thick and thin blood slides were examined and hemoglobin measured. Other tests were undertaken as indicated.Patients were seen daily for the first 3 days, weekly to 28 days and monthly for 12 months.P. vivax infections had capillary blood collected on filter paper for confirmatory PCR analysis and were treated with the same treatment as before (CQ or CQ+PQ); patients in the CQ+PQ arm who had failed to complete the initial PQ course and then experienced recurrent P. vivax were treated with CQ only.Patients with recurrent P. falciparum infections were treated with artesunate plus sulphadoxine-pyrimethamine16 and retained in the study. Patients failing to attend follow-up were visited at home and asked to reattend the center. Remuneration for transportation was provided for follow-up visits. All patients were given one long-lasting insecticide treated bed net.Patients were censored from analysis if there was stated withdrawal of consent, severe malaria or persistent vomiting during the acute phase  or development of concomitant disease interfering with classification of treatment outcome. Patients who developed P. vivax recurrence (detected by microscopy) assessed by survival analysis. Secondary outcomes were the total number of recurrences, and the safety and tolerability of the treatments. Patients who completed study treatment entered the per protocol (PP) analysis. Those in the CQ+PQ arm who failed to complete PQ continued to be followed up and were included in the intention-to-treat (ITT) analysis. Patients who did not attend at or after 14 days were excluded from both efficacy analyses.The primary outcome was P. vivax within a year after treatment from 25% to 15%. Using Z-test with continuity correction, one-sided significance level (\u03b1) = 0.025 and power = 80%, calculated sample size was 270 per arm; 600 patients were planned to allow for 10% loss to follow up.It was hypothesized that PQ would reduce the proportion of patients with at least one recurrent attack of t test, Mann-Whitney U and \u03c72 (or Fisher\u2019s exact) tests were used for comparison of baseline variables, as appropriate. Parasitological failure rates were assessed by recurrence-free survival analysis (Kaplan-Meier). Cox regression was performed to identify possible independent predictors of recurrence, using a stepwise elimination method.Data were entered onto record forms, transferred electronically to an Open Clinica database and downloaded via Excel into STATA (v10) . Student\u2019s P. vivax and P. falciparum.17DNA was extracted from filter-paper blood spots from recurrent cases and nested PCR performed to detect The study was approved by the Ethics Committee of the Faculty of Tropical Medicine, Mahidol University, Thailand, the Oxford Tropical Research Ethics Committee, Oxford University, United Kingdom and the Institutional Review Board of the Afghan Public Health Institute, Ministry of Public Health, Afghanistan. The trial was registered under the identifier NCT01178021.P. vivax were enrolled in the study and randomized to the CQ (N = 295) and CQ+PQ (N = 298) arms  arms .Six patients  randomized to CQ+PQ were judged G6PD deficient on fluorescent spot testing, treated with CQ and withdrawn from the study. Later G6PD molecular testing using retained baseline dried blood spots showed that three males were hemizygous for the Mediterranean deficient variant, while the female was wild-type .Baseline characteristics were similar between the two treatment groups .P. falciparum monoinfection during follow-up, were treated with artesunate plus sulphadoxine-pyrimethamine, and retained in the study.Of the 570 individuals seen at or after 14 days, 23 (4.0%) were lost to follow-up before 6 months. A total of 59 (10.4%) were lost to follow-up (without experiencing the primary endpoint) before 1 year. Security issues in the Kunar region in 2010 prevented the latter stages of follow-up in a set of 27 consecutive patients recruited at the Kunar site. Two patients had P. vivax recurrence occurred in 86 (29.9%) of 288 patients in the CQ arm and 37 (13.1%) of 282 in the CQ+PQ arm . The PP analysis (excluding six patients not completing PQ) gave similar results: CQ+PQ HR 0.35 (0.24\u20130.52).Protection from recurrence afforded by PQ was largely confined to the first 6 months of follow-up, with 45 and four patients with recurrence in the CQ and CQ+PQ arms respectively. The Cox proportional estimate for protective effect of PQ in the first 6 months was 0.082 (95% CI 0.029\u20130.23) by ITT analysis. In months 7\u201312 protection was no longer significant .Univariate analysis showed that age, gender, weight, baseline hemoglobin and parasitemia were not significantly associated with recurrence. Baseline gametocytemia on microscopy was associated with recurrence , and remained significant in a multivariable analysis with treatment arm. However, in contrast to the effect of PQ, the association was not significant in the first 6 months  and was only significant for months 6\u201312 .P. vivax before day 42 with six in the CQ arm occurring at day 28 (or 1 day before). The median parasitemia at recurrence in these six cases was 1,980/\u03bcL .At day 1 after treatment, 84.3% of patients in the CQ arm and 90.3% in the CQ+PQ arm had cleared parasites on the peripheral blood film; by day 2, this proportion was 98.6% in both arms. Eight patients (1.4%: 7 in the CQ arm) had recurrent P. vivax recurrences were speciated by PCR. Of 88 available samples, 79 were found to be P. vivax, four Plasmodium falciparum and two mixed infections (three cases had no amplification product). Of the P. falciparum monoinfections (misidentified as P. vivax), three were in the CQ arm and one in the CQ+PQ arm; one of these CQ cases went on to have a true P. vivax recurrence (confirmed by PCR). Inaccurately speciated cases are hence unlikely to have significantly impacted the main findings.Dried blood spots from reported P. vivax was associated with a fall and rise in hemoglobin of very similar magnitude   increase compared with baseline of 0.44 g/dL (0.36\u20130.53) . Recurre26\u20130.58) .Both treatments were generally well tolerated. Repeated vomiting of the study medication was not reported. Seven patients deteriorated, requiring hospital admission, during the treatment phase and were withdrawn from the study ; five weThere were seven other cases where signs or symptoms considered compatible with hemolysis occurred but the patient was not hospitalized or withdrawn. Six of these were in the CQ+PQ arm, and PQ administration was stopped . Some paP. vivax in Afghanistan. Measuring the efficacy of radical treatment involves consideration of several methodological issues. Molecular tests cannot distinguish relapse from reinfection or recrudescence , requiring a control arm of CQ, efficacious only against the blood-stage infection. The follow-up period needs to take into account the natural history of relapses in the given location.18 In Southeast Asia and Oceania, where relapses are frequent and early, differences in relapse rate can be discerned with a relatively short period of follow-up. In South Asia, relapse rates are lower, and in some areas, long latency temperate strains are prevalent,19 so a longer period of observation with high rates of follow-up is required to obtain sufficient power.The trial\u2019s primary aim was to measure the efficacy of unsupervised PQ (14 days at 0.25 mg/kg/day) as radical treatment of 20 but still the lack of early infections in the CQ+PQ arm suggests that these were relapses rather than recrudescences, indicating ongoing efficacy of CQ against blood stages.21 For the first 6 months, the recurrence-free survival curve for CQ indicated a constant recurrence frequency of about 2.5% per month; again, these are likely to have been relapses because recurrences remained rare in the CQ+PQ arm. After 6 months, the two survival curves became more similar, suggesting that reinfections predominated, although there was a possible excess of recurrences in the CQ group after 8 months that may represent late relapses of long latency infections as seen previously19 . The overall recurrence rates in the two arms are broadly similar to those previously found in Afghan refugees in Pakistan.2Given its high rate of follow-up, our study provided a clear view of the natural history of vivax relapse in Afghanistan. The survival curves in the two study arms appeared to diverge from the outset, with seven of eight early recurrences (before day 42) in the CQ only arm. PQ has significant blood-stage activity,18 and has been shown to provide useful antirelapse activity (of varying magnitude) in other randomized controlled studies undertaken in Pakistan,4 India,23 Thailand,25 and Ethiopia26 as well as the multi-center DETECTIVE study27; these studies have also been summarized in relevant meta-analyses.29The overall efficacy of 0.25 mg/kg/day PQ for 14 days over a year was approximately 65%. The efficacy during the first 6 months, in which relapses are likely to have predominated, was around 90%. This study hence provides clear evidence that the current PQ regimen is efficacious in radical cure. The total dose of 3.5 mg/kg prescribed is categorized as \u201clow dose\u201d according to the stratification proposed by John et al.30Vivax recurrence was associated with a fall of approximately 0.5 g/dL hemoglobin. Although unlikely to be clinically detrimental in isolation, this could carry a cumulative effect with repeated recurrences (particularly if diagnosed late and/or associated with other forms of anemia).31 and consistent with previous studies.21 In addition, microscopic parasite clearance rate was fast in both CQ and CQ+PQ arms, with more than 98% of cases clear of parasites by day two, another marker of CQ sensitivity.31CQ has been used as first-line schizonticidal treatment of vivax malaria in Afghanistan for many decades. The data obtained in this study suggest that it remains an efficacious schizonticide in this region. Six cases in the CQ arm experienced relapse at 28 days (or just before). Without drug levels at the time of recurrence it is not possible to exclude the possibility that these cases represent CQ-resistant parasites, but the overall proportion of such cases  is indicative of a sensitive population4The study had certain weaknesses which might impact on its generalizability. Most PQ doses were unsupervised, but PQ was observed on days zero, one, two, and seven, with further reinforcement of adherence by ongoing follow-up, so it was partly supervised, potentially overestimating effectiveness. However, previous work suggests good levels of adherence in this community.32The work was undertaken in a resource-poor setting with challenging logistic and security issues; at one stage, the Kunar site closed because of civil unrest. The malaria treatment centers lack facilities for detailed assessment of possible hemolysis, leading to the possibility of subjective decision making by staff. Linked to this, the study was nonblinded, with clinical staff aware of treatment allocation. None of the hospitalized patients judged to have a syndrome compatible with PQ-induced hemolysis, or in whom PQ was stopped, met laboratory criteria for hemolysis, suggesting that these were not hemolytic episodes; furthermore, retrospective G6PD genotyping (limited to the Mediterranean locus) indicated only wild-type alleles. Notably, some of these patients were anemic at baseline, suggesting that study doctors may have been influenced by disease rather than treatment effects. Use of placebo would have offset this problem.30 and also contribute to the infectious reservoir in low transmission settings.34 Molecular validation by PCR of dried blood spots indicated that a small proportion (around 5%) of recurrent infections thought to be P. vivax were actually P. falciparum, an unsurprising result given previously identified challenges in accuracy of malaria diagnosis in Afghanistan.35 The relatively low number of possibly misclassified cases appears unlikely to have compromised the study\u2019s overall conclusions.For pragmatic reasons, detection and speciation of recurrent malaria, clearly central to assessment of the primary endpoint, were undertaken by microscopy, thereby focusing on the efficacy of treatment in terms of symptomatic cases. Detection of recurrence via the more sensitive technique of PCR (on dried blood spots) would have allowed assessment of the impact of PQ on all recurrences, including submicroscopic infections which can increase the risk of anemiaP. vivax patients in Afghanistan. Because of ongoing safety concerns, methods for exclusion of G6PD deficiency that can be applied a range of field settings38 are needed if PQ is to be widely deployed.PQ administered in a largely unsupervised fashion at 0.25 mg/kg/day for 14 days provides efficacious radical cure in"}
+{"text": "Plasmodium falciparum malaria in Tanzania.The World Health Organization (WHO) recently recommended the addition of a single low-dose of the gametocytocidal drug primaquine (PQ) to artemisinin-based combination therapy (ACT) in low transmission settings as a component of pre-elimination or elimination programmes. However, it is unclear whether that influences the ACT cure rate. The study assessed treatment outcome of artemether-lumefantrine (AL) plus a single PQ dose (0.25\u00a0mg/kg) versus standard AL regimen for treatment of acute uncomplicated P. falciparum malaria patients aged \u22651\u00a0year, with the exception of pregnant and lactating women, were enrolled and treated with AL plus a single PQ dose (0.25\u00a0mg/kg) or AL alone under supervision. PQ was administered together with the first AL dose. Clinical and laboratory assessments were performed at 0, 8, 24, 36, 48, 60, and 72\u00a0h and on days 7, 14, 21, and 28. The primary end-point was a polymerase chain reaction (PCR)-adjusted adequate clinical and parasitological response (ACPR) on day 28. Secondary outcomes included: fever and asexual parasitaemia clearance, proportion of patients with PCR-determined parasitaemia on day 3, and proportion of patients with Pfmdr1 N86Y and Pfcrt K76T on days 0, 3 and day of recurrent infection.A randomized, single-blinded, clinical trial was conducted in Yombo, Bagamoyo district, Tanzania. Acute uncomplicated p\u00a0=\u00a00.79). Day 28 PCR-adjusted ACPR and re-infection rate was 105/105 (100\u00a0%) and 101/102 (99\u00a0%) (p\u00a0=\u00a00.31), and 5/107 (4.7\u00a0%) and 5/8 (4.8\u00a0%) (p\u00a0=\u00a00.95), in AL\u00a0+\u00a0PQ and AL arm, respectively. There was neither any statistically significant difference in the proportion of Pfmdr1 N86Y or Pfcrt K76T between treatment arms on days 0, 3 and day of recurrent infection, nor within treatment arms between days 0 and 3 or day 0 and day of recurrent infection.Overall 220 patients were enrolled, 110 were allocated AL\u00a0+\u00a0PQ and AL, respectively. Parasite clearance by microscopy was fast, but PCR detectable parasitaemia on day 3 was 31/109 (28.4\u00a0%) and 29/108 (26.9\u00a0%) in patients treated with AL\u00a0+\u00a0PQ and AL, respectively  contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria globally .The distribution of P. falciparum malaria in Tanzania, with similar PCR-ACPR as previously has been reported from the same study area of AL alone [This study provides much needed data on treatment outcome of the new WHO recommendation of adding a single low-dose (0.25\u00a0mg/kg) of PQ to AL compared with standard AL treatment alone. Both regimens provided high cure rates for the treatment of acute uncomplicated AL alone \u201321, and AL alone , 22. SimPfmdr1 N86 and Pfcrt K76 among recurrent infections during follow-up after AL treatment [Pfmdr1 N86 and Pfcrt K76 in the study area ever since AL was introduced as first-line treatment of uncomplicated malaria in 2006 [Pfmdr1 N86 and Pfcrt K76, neither during the early treatment phase nor among recurrent infections during follow-up after treatment. Thus, the findings suggest that addition of this single low-dose PQ (0.25\u00a0mg/kg) does not apparently spur the selection potential of drug resistance markers previously associated with lumefantrine tolerance/resistance.Previous studies have reported selection of reatment \u201325, and reatment . A simil in 2006 , which hThe study has some potential limitations including that it involved patients of all ages with all levels of parasitaemia despite the WHO recommendation to conduct efficacy studies in children under the age of 5\u00a0years with parasitaemia level of 2000\u2013200,000/\u00b5l of blood in highly malaria-endemic countries such as Tanzania, to minimize the influence of immunity on treatment outcome in older individuals. However, this study was done to assess whether adding PQ to AL would potentially compromise the efficacy of latter drug probably due to drug\u2013drug interaction, that can be done at all ages and levels of parasitaemia.P. falciparum malaria in Tanzania. The findings support the use of a single low-dose of PQ together with AL as a potential component of ongoing P. falciparum malaria elimination efforts.The new WHO recommendation of adding a single low-dose (0.25\u00a0mg/kg) of PQ to AL did not compromise treatment outcome of uncomplicated"}
+{"text": "Plasmodium vivax are also treated with 14\u00a0days primaquine  (PQ). The aim of this study was to assess the efficacy of the national policy.First-line schizontocidal treatment for uncomplicated malaria in the Republic of the Sudan is artesunate  plus Sulphadoxine/pyrimethamine (25/1.25\u00a0mg/kg) (AS/SP). Patients with Plasmodium falciparum and/or P. vivax malaria were treated with AS/SP. Patients with P. falciparum were randomized to no primaquine (Pf-noPQ) or a single 0.25\u00a0mg/kg dose of PQ (Pf-PQ1). Patients with P. vivax received 14\u00a0days unsupervised 3.5\u00a0mg/kg PQ (Pv-PQ14) on day 2 or at the end of follow up (Pv-noPQ). Primary endpoint was the risk of recurrent parasitaemia at day 42. G6PD activity was measured by spectrophotometry and the Accessbio Biosensor\u2122.Patients above 1\u00a0year, with microscopy-confirmed, P. falciparum (74.8%), 77 (24.9%) with P. vivax and 1 (0.3%) patient with mixed infection were enrolled. The PCR corrected cumulative risk of recurrent parasitaemia on day 42 was 3.8% (95% CI 1.2\u201311.2%) in the Pf-noPQ arm compared to 0.9% (95% CI 0.1\u20136.0%) in the Pf-PQ1 arm; . The corresponding risks of recurrence were 13.4% (95% CI 5.2\u201331.9%) in the Pv-noPQ arm and 5.3% (95% CI 1.3\u201319.4%) in the Pv-PQ14 arm . Two (0.9%) patients had G6PD enzyme activity below 10%, 19 (8.9%) patients below 60% of the adjusted male median. Correlation between spectrophotometry and Biosensor\u2122 was low .231 patients with P. falciparum and P. vivax. The addition of PQ reduced the risk of recurrent P. falciparum and P. vivax by day 42, although this did not reach statistical significance. The version of the Biosensor\u2122 assessed is not suitable for routine use.AS/SP remains effective for the treatment of Trial registrationhttps://clinicaltrials.gov/ct2/show/NCT02592408The online version of this article (10.1186/s12936-018-2266-9) contains supplementary material, which is available to authorized users. There has been a decline in the incidence of malaria in many sub-Saharan countries in the last decade \u20133, whichPlasmodium vivax , the risk being greater in patients treated with Pf-noPQ (3.4% [95% CI 1.1\u201310.3%]) compared to 0.9% (95% CI 0.1\u20136.0%) in those treated with Pf-PQ1; Hazard Ratio HR\u2009=\u20090.26 [95% CI 0.03\u20132.47], p\u2009=\u20090.203.In the P. falciparum had gametocytes detected on blood film examination during follow up.By day 42 the risk of parasite recurrence was 2.5% [95% CI 2.06\u20136.01%]; 4.8% (95% CI 1.8\u201312.4%) in the Pf-noPQ and 0.9% (95% CI 0.1\u20136.0%) in the Pf-PQ14 ; Fig.\u00a0P. vivax arms, a total of 6 patients had recurrent parasitaemia at days 14 (n\u2009=\u20091), 21 (n\u2009=\u20091), 28 (n\u2009=\u20091), 35 (n\u2009=\u20093). In patients not receiving 14\u00a0days primaquine the risk of recurrent P. vivax at day 28 was 9.9% [95% CI 3.3\u201327.8%] compared to 0% in those treated with AS/SP plus 14\u00a0days primaquine; p\u2009=\u20090.046. At day 42 the respective risks had risen to 13.4% [95% CI 5.2\u201331.9%] in the Pv-noPQ arm and 5.3% [95% CI 1.3\u201319.4%] in the Pv-PQ14 arm; HR 0.36, [95% CI 0.1\u20132.0], p\u2009=\u20090.212  patients with P. vivax  of patients receiving a sub-optimal dose of any of the study drugs were excluded, leaving 127 (54.9%) of patients with 09 of patNo patient received sub-optimal AS Table\u00a0. For patP. falciparum and 11.0\u00a0g/dl in patients with P. vivax , compared to patients infected with P. vivax whose Hb did not change  over the same period; p\u2009=\u20090.015 .At baseline the median Hb concentration was 11.7\u00a0g/dl in patients with on Table\u00a0. On day P. falciparum the Hb concentration at day 7 had risen 6.0% (IQR \u2212\u00a00.8 to 14.8) from baseline following treatment with Pf-noPQ compared to 13.2% (IQR 0.0 to 23.9) after treatment with Pf-PQ1; p\u2009=\u20090.007. In patients with P. vivax, there was no change in Hb (IQR 0.0 to 10.0) between day 0 and day 7 following treatment with Pv-noPQ compared to 9.1% (IQR 0.0 to 15.4) in vivax patients treated with primaquine (Pv-PQ14); p\u2009=\u20090.106. None of the patients included required a blood transfusion.The median Hb levels increased in all treatment arms between days 0 and 7 Fig.\u00a0, Table\u00a04A total of 551 adverse events (AE) were reported, of which 497 (90.2%) were categorized as mild and 51 (9.8%) as moderate. Three (0.5%) mild events were recorded as possibly related to AS/SP. The most frequent AEs were gastrointestinal symptoms including poor appetite (40.3%), nausea (25.6%) and abdominal pain (20.7%)  patients had G6PD activity between 30 and 60%  patients. The adjusted male median by spectrophotometry was 9.7 U/gHb, compared to 7.9\u00a0U/gHb derived by the Biosensor\u2122 (p\u2009<\u20090.001). By spectrophotometry 2 (0.9%) patients with 60% Fig.\u00a0.Fig.\u00a04G6s\u2009=\u20090.330 (p\u2009<\u20090.001), with a mean difference of 2.1 U/gHb and the 95% limit of agreement (LoA) ranged from \u2212\u00a05.1 U/gHb to 9.2 U/gHb  with activities between 10 and 30% activity, and 27 patients with G6PD activity between 30 and 60% , although these benefits did not reach statistical significance. Results on gametocyte carriage are limited by the small denominator applied for microscopy (200WBC).This efficacy study demonstrates that when the target dose of AS/SP was achieved the risk of recurrent parasitaemia was low for patients with either 25\u00a0mg/kg . While tP. falciparum [P. falciparum by day 28 generally exceeds 95%, except for one site in Gedaref, in the East of the country, where the efficacy fell from 91% in 2012 to 87% in 2015 [In 2006, a study in the east of the country reported a day 28 PCR corrected efficacy of 93.5% for AS/SP for lciparum . Other c in 2015 .pfdhfr and pfdhps genes and a single case with a triple mutation in the pfdhfr gene, all associated with antifolate resistance [One study from Kassala, Eastern Sudan reported a PCR corrected efficacy of 93.7% by day 28 , while asistance .P. falciparum [In Somalia, the efficacy of AS/SP has fallen to 88% and artemether\u2013lumefantrine has replaced AS/SP as the first-line treatment for lciparum . In MarcP. falciparum and P. vivax AS/SP has been used as a universal schizontocidal treatment for both species particularly in locations where diagnostic facilities are unable to distinguish reliably between species. To date 3 published trials have assessed the efficacy of AS/SP against P. vivax. Tjitra et al. found AS/SP to have almost 90% efficacy against P. vivax in Papua, Indonesia [P. vivax by day 42 after AS/SP rose to 25% in Afghanistan [P. vivax recurrence at day 42 was 13.4% in patients treated with AS/SP alone, less than half that expected in our a priori power calculation. The risk of recurrence fell to 5.3% in those treated with AS/SP plus 14\u00a0days primaquine although this did not reach statistical significance (p\u2009=\u20090.212); Table\u00a0In areas co-endemic for ndonesia . Howeverhanistan , and 67%hanistan . In the P. vivax can be achieved with a 14-day regimen of primaquine, although the long course of treatment is associated with poor adherence and compromised effectiveness [WHO guidelines recommend that the radical cure of tiveness , 40\u201344. tiveness , and indP. falciparum infection there was no further fall in Hb observed between day 2 and day 7, with or without a single dose of primaquine. These observations are reassuring and concur with previous reports from Tanzania [P. vivax infection had G6PD activity below 30%, although 2/22 (9.1%) had intermediate activity between 30 and 60%, reassuringly none of the patients had a significant fall in Hb after day 2.PQ is known to cause haemolysis in G6PD deficient individuals with the nadir usually observed 2\u20137\u00a0days after initial exposure . The greTanzania  supportiTanzania . None ofHowes et al. had earlier documented a country wide G6PD deficiency prevalence of 15.3% with higher frequencies towards the West of the country and lower frequencies in the East . These pThe study also evaluated a novel point of care Biosenor\u2122  which was compared to the reference method of spectrophotometry. The results highlight that the current Biosensor\u2122 underdiagnosed severe G6PD deficiency, a finding consistent with a recent similar study conducted in Bangladesh . Out of P. falciparum and a 14-day regimen in patients with P. vivax were well tolerated, and associated with a modest reduction in the number of recurrences by day 42, however reduction was not significant for P. falciparum. The Biosensor\u2122 achieved suitable operational characteristics, but test performance is insufficient for routine diagnosis. A second generation with integrated haemoglobin measurement will be introduced to the market and may provide a more suitable solution.AS/SP appears to retain adequate efficacy in uncomplicated malaria in New Halfa and Gezira Slanj. Both a single dose of primaquine in patients with Additional file 1. Scatter plot body weight vs. total dose SP received. red line\u2009=\u2009target dose recommended by the WHO.Additional file 2. Database."}
+{"text": "Uncommon and rare adverse events (AEs), with delayed onset may not be detected before new drugs are licensed and deployed. The present study examined the post licensure safety of dihydroartemisinin-piperaquine (DHP) as an additional treatment for malaria in Ghana. The relationship between the incidence of AEs, treatment completion rate, participant characteristics and concomitant medications are reported.\u00ae (20/160mg dihydroartemisinin and 40/320mg piperaquine by Sigma-Tau Incorporated) was given, according to the body weight of patients. First treatment doses were under observation but the second and third doses were taken at home except in a sub-study involving a nested cohort. Patients were contacted at Day 5 (\u00b1 2 days) either on telephone or by a home visit to document any AEs experienced. Patients were asked to report to the study team any other AEs that occurred within 28 days post-treatment. All patients in the nested cohort had electrocardiogram (ECG).A study conducted from September 2013 to June 2014 in Navrongo, Kintampo and Dodowa health research centres in Ghana is presented. Participants had confirmed malaria and no known allergy to study drug. Patients provided informed consent and had their symptoms and results of their clinical examinations documented. Treatment with EurartesimA total of 4563 patients, 52.1% females and 48.2% <6 years completed the study. A total of 444 patients were enrolled into the nested cohort. About 33% had temperature \u2265 37.5\u00b0C at enrolment. Approximately 3.4% reported taking prior antimalarials, 19.4% other medications and 86% took at least one concomitant medication. Incidence of AEs was 7.6% including infections (4.6%), gastrointestinal disorders (1.0%) and local reactions at the site of venesection (0.5%). Others were respiratory disorders (0.4%) and nervous system disorders (0.3%). There were nine adverse events of special interest (AESI); itching/pruritus (7), dizziness (1), and skin lesions (1). Patients who took medications prior to enrolment had higher incidence of AEs compared with those without . Statistically significant associations were found between the reported AEs and age of patients (P<0.001), their body mass index (BMI) (P< 0.001) and parasite densities (P< 0.001).Dihydroartemisinin-Piperaquine was well tolerated with no serious safety concerns identified. Obesity and prior enrolment medication were among significant factors associated with increased AEs reporting. Systematic clinical studies in human volunteers are often carried out in order to discover the effects of new medicinal products , 2. Somefalciparum malaria. Currently, dihydroartemisinin plus piperaquine (DHP) has been made an option for the first-line treatment of uncomplicated malaria [Artemisinin-based combination therapies (ACTs) are presently recommended for the treatment of uncomplicated  malaria . The cho malaria . The milTo contribute to post-licensure safety information on antimalarials, the INDEPTH-Network in 2009 established an effectiveness and safety platform for post licensure evaluation of newly registered antimalarials across Africa . The PhaIn Ghana, three first-line artemisinin-based combination drugs are available for treatment of uncomplicated malaria\u2014Artesunate-amodiaquine (ASAQ) and Artemether-lumefantrine (ALU) have been in the public health system for some time but Dihydroartemisinin-piperaquine (DHP) has recently been introduced and has limited pharmacovigilance safety data.This study used the INDEPTH-Network safety-monitoring platform and data from a prospective phase IV study that evaluated clinical safety of fixed-dose dihydroartemisinin-piperaquine .This study assessed the safety of DHP as an additional first-line combination treatment for acute uncomplicated malaria across three different ecological zones with health and demographic surveillance systems in Ghana.The dihydroartemisinin (DHA) component of DHP is very rapidly absorbed with Tmax being about 1\u20132 hours after single and multiple dosing. The Piperaquine (PQ) component is highly lipophilic, slowly absorbed and has a Tmax of about 4\u20135 hours after single and repeated dose . PiperaqIn view of the concerns about cardiotoxicity of DHP \u201316, this2 and characterized by coastal savannah vegetation [2 surface area and a combined resident population of about 143 000 [2, population of about 156 000 and Guinea Savannah vegetation [The study was conducted across Ghana in three health research centres where there are on-going Health and Demographic Surveillance Systems (HDSS) getation . The Kin 143 000 . The Navgetation .Anopheles gambiae s.l and Plasmodium falciparum being the most predominant malaria vector and parasite respectively. Recent parasite prevalence estimates ranged from 5% along the coast to about 40% in northern Ghana [Malaria is endemic in all the three study sites with rn Ghana . The Gharn Ghana .\u00ae on blood biochemistry, full blood count, white cell count differential count and QTc intervals. In this subset, the presence of Plasmodia of any species was confirmed microscopically before treatment and was intensively followed-up. Blood was taken for haemoglobin, full blood count and biochemistry at Day 1 before drug administration, Day 3 between 3\u20134 hours after the last dose of treatment, and Day 7. If the results were abnormal and clinically relevant, the blood examination was repeated weekly until normalization. They also had ECGs undertaken on Day 1 before drug administration, twice on Day 3 (i.e. before and 3\u20134 hours after the last drug administration) as well as on Day 7.This was a prospective, observational, open-label, non-comparative, multi-site study conducted from September 2013 to June 2014 across the three sites [All patients visiting selected health facilities in the study sites during the study period were screened for inclusion. The description and analysis presented in this report covers mainly detailed data from the three study sites in Ghana: Navrongo, Kintampo and Dodowa health and demographic surveillance sites.Study participation included persons of both sexes with presence or history of fever within the previous 48 hours, chills, headache, general malaise and/or loss of appetite. All eligible patients were invited to participate in the study after the necessary individual and/or parental informed consent and/or assent had been obtained. Each patient had detailed symptoms enquiry, clinical examinations and all significant conditions documented. Each patient was allowed to enter the study once and had to satisfy the selection criteria.The inclusion criteria were as follows; uncomplicated malaria diagnosed as per WHO recommendations, age \u22656 months and weight \u22655 kg. Others were the patient\u2019s ability to take oral medication, willingness to participate based on a witnessed signed informed consent, and access to a health facility. The patient had to agree to comply with all scheduled follow-up visits. The exclusion criteria included a known allergy to artemisinin or to piperaquine, known pregnancy, lactating women and patients with complicated malaria. All patients who were on drugs that are known to prolong the QTc interval or have antimicrobial effects or are non-sedating antihistamines or have taken a DHP in the previous four weeks were also excluded. Others included a family history of sudden unexplained death, or personal or family history of predisposing cardiac conditions for arrhythmia/QT prolongation.\u00ae . The first dose was administered under observation to all patients in the clinic. Patients were instructed to remain in the clinic for observation for at least one hour. The second and third daily doses were self-administered at home on consecutive days and at the same time as the first dose. Those in the small nested cohort took all three treatment doses under direct observation. Patients who vomited within 30 minutes of administration of the first dose were re-treated with the same dosage, and if vomiting occurred within 30 to 60 minutes, half a dose was re-administered. Re-dosing was not attempted more than once.Drug treatment was based on weight bands using the two available dosage strengths of EurartesimPatients were contacted on Day 5 (\u00b1 2 days) either by telephone or by a home visit by trained field supervisors for assessment of their recovery status; all adverse events experienced after treatment were documented during this visit or phone call. For all instances where adverse events information collected during the telephone contact was considered incomplete, a study staff was asked to visit the patient to complete the interview. In addition, patients were asked to report to the study team any other adverse events that occurred within 28 days after drug administration. Patients whose symptoms worsened between intake of the first dose and within 28 days of study enrolment were asked to visit the nearest health facility or contact study staff using the telephone numbers on their consent form. In cases of serious and / or severe adverse events classified as of special interest, a field supervisor was detailed to bring the participant for further assessment by the study clinicians. If any patient reported a cardiac event, an ECG was performed and the trace was inspected for other abnormalities. Any patient with events deemed to be serious were cared for in line with the national standard practices.The AESI were predefined in the study and in all situations patients were directed to the health facility for evaluation and recording of all relevant information. AESI suggestive of possible cardio-toxicity and prolonged QT were palpitations, seizures, pounding or pain in the chest area and fainting or syncope. AESI suggestive of neurotoxicity were seizures, dizziness, pins and needles sensations, visual disturbance, difficulties in coordination and tinnitus. Those suggestive of possible cutaneous reactions and phototoxicity were urticaria, angioedema, skin lesions, itching pruritus, discoloration and dermatitis.Patients in the nested cohort had a baseline 12-leads digitalized ECG done in triplicate. Print outs of the ECGs with at least five complexes for each lead were read by the study physicians before the first dose was administered. For each test, automatic calculation for QTcF was verified and participants with average recordings of \u2265 450ms excluded and prescribed other antimalarials. Electronic copies of all the ECGs were sent out to a central cardiac laboratory (Cardiabase) in France for further evaluation. Quality control checks included the assessment of the intra-reader and inter-reader variability. Pre-dose ECG was performed before day 3 dose was administered and patients were observed for 3 to 4 hours before the day 3 post-treatment ECGs in triplicate were performed. Participants whose day 3 pre-treatment QTc interval were \u2265 500 ms were observed for 6 hours till the QTc interval was less than 480ms before the third dose was administered. Alternate antimalarial medicines were administered if the QTc interval persistently remained \u2265 480ms. On day 7, ECGs were repeated for each participant.The initial diagnosis of malaria was confirmed with malaria rapid diagnostic test as per the World Health Organization (WHO) recommendation . In addi\u00ae (version 11.2) software package. Descriptive analyses were conducted for all data. In addition, results from bivariate analysis, logistic regression, Chi-square, odds ratios are presented. Body Mass Index (BMI) calculated as weight in kilograms divided by the square of height in meters was estimated. The BMI is an indicator of the level of body fatness and is therefore used for the assessment of overweight and obesity. For adults a BMI value of 30 or more is regarded as obesity. We estimated BMI for those aged \u226518 years. Adverse events were coded and described using the Medical Dictionary for Regulatory Activities  terminology. All events reported were grouped by MedDRA\u00ae system organ class classification. The estimates of the incidence of AEs were based on crude rates with no causality assessment of individual cases. A two-sided 95% confidence interval (CI) was constructed for the estimates. The QTc intervals (ms) were evaluated after correcting for the heart rate with Fredericia\u2019s formula (QTcF = QT/RR1/3). Descriptive analysis of the mean QTcF was done. The proportion and mean difference between the baseline and day 3 pre-dose, day 3 post-dose and day 7 were computed. The pre- and post-dose mean and median values of all the laboratory parameters were described.Data were double-entered into an on-line system using OpenClinica software, which is good clinical practice compliant. Statistical analysis were performed using STATAThe justification for the use of human participants for this study was to evaluate the safety of DHP when used in patients with signs and symptoms of uncomplicated malaria in a real life situation. The protocols received approvals from the Ghana health service ethics review committee and the Navrongo, Kintampo and Dodowa health research centre institutional review boards respectively. Approval was also obtained from Ghana\u2019s foods and drugs authority. The approvals were followed by community engagement to seek the support of the communities and their leaders. The study was conducted in compliance with good clinical and laboratory practices. Written informed consent was obtained from all study participants before any study-related activity was undertaken. Parental consent was obtained for children and assent for older children (12\u201317 years). The study was registered with Clinicaltrials.gov (NCT02199951).A total of 4838 patients with acute febrile illness were screened, 4777 enrolled and 4563 completed the study. A total of 444 patients were enrolled into the nested cohort . KintampAbout 33% (1498/4563) of patients had axillary temperature \u2265 37.5\u00b0C at enrolment and this varied across the sites; Kintampo (40%), Navrongo (29.2%) and Dodowa (25.1%). Age specific fever prevalence were; < 6 years (43.2%), 6\u201312 years (31.0%), 13\u201318 (21.7%) and >18years (13.1%). The frequency of baseline symptoms were fever (26%), anorexia (11%), headache (11%), vomiting (10%), cough (9%), weakness (9%), abdominal pain (7%), muscular pain (5%), diarrhea (4%) and nausea (4%). About 12% of all patients had parasite density less than 500/\u03bcL and 24% > 50,000 /\u03bcL. Geometric mean parasite density (95%CI) by site was 4752/\u03bcL  in Navrongo, 25503/\u03bcL  in Kintampo and 7995 /\u03bcL  in Dodowa. About half of the patients from Kintampo (48.5%) had parasite density of >50,000 /L compared to 15% in Navrongo and 33.3% in Dodowa. Age specific parasite density (95% CI) was 23802/\u03bcL  in < 6year olds, 9897/\u03bcL  in 6\u201312 year olds, 2434 /\u03bcL  in 13\u201318 year olds and 1540/\u03bcL  those >18 years .Approximately 3.4% (156/4563) of patients reported taking antimalarials within the last four weeks prior to enrolment. The prevalence was highest in Navrongo 4.3% (82/1901), followed by Kintampo 3.5% (62/1778) and Dodowa 1.4% (12/884). About 70% (106/156) of all antimalarials taken prior to enrolment were in children < 6years of age. The age specific prevalence was <6years (4.8%), 6-12years (1.9%), 13\u201318 years (1.1%) and >18years (3.0%). The prevalence in females and in males were similar . Approximately 19.4%  of patients reported taking other medications prior to enrolment. The site-specific prevalence was Navrongo (15.9%), Kintampo (17.8%) and Dodowa (30%). About 20% were in children < 6 years, 19.0% in females and 20% in males.2 = 18.7; P< 0.0001). Children aged <6 years had the highest (88.2%) compared to those aged 6\u201312 years (83.4%), 13\u201318 years (87.8%) and above 18 years (84.6%). Approximately 86% (3938/4565) of the participants took at least one concomitant medication during the study. The proportions of patients who took concomitant medications per site were Navrongo (90.2%), Kintampo (96.5%) and Dodowa (57.5%). 2 = 18.7; P <0.0001), ranging from 9.2%  in Navrongo, 8.4%  in Dodowa to 5.5%  in Kintampo. Infection and infestations, and gastrointestinal disorders were the leading incidence of AEs reported in all sites 2 = 2.9; P> 0.05) and by concomitant medication . Patients who took other medications prior to enrolment had higher incidence of AEs compared with those who did not . Statistically significant heterogeneity of AEs was found between the study treatment and different age groups , body mass index       and mala 0.0001) . There w 0.0001)  but para 0.0001) . LogistiChange in QTcF values in each individual patient were assessed by comparing the number of patients who had increases and decreases in the parameters. In all, findings from 378 patients on day 3 were found to have increased, 90 were decreased and 9 had no change from the baseline findings. The mean QTcF at baseline (day 1) was 394.8ms . There was no significant difference in the QTcF between males (393.8ms) and females (395.6 ms) at baseline . The mean QTcF (95% CI) on day 3 pre-dose, day 3 post-dose and day 7 were 409.3ms , 422.9ms  and 399.2ms  respectively. Compared to the baseline value there were significant changes in mean QTcF in day- 3 pre-dose -14.5ms , day-3 post-dose -28.2ms  and day-7 post-enrolment -4.5ms . For three patients who had QTcF above the 500ms cutoff value, one patient had QTcF of 509ms on day-3 post-dose and the same patient had QTcF of 516ms on day-7 post-enrolment. These were however not clinically significant and resolved after follow-up. The number of patients with QTcF prolongation > 60ms from their baseline were seven on day-3 pre-dose, 38 on day-3 post-dose and only one on day-7 post-enrolment and none of them was clinically significant.6/\u03bcL  and total white cells x 106/\u03bcL . For the clinical chemistry, the respective mean baseline (day 0) values compared to day 3 estimates were total bilirubin, \u03bcmol/L , alanine transaminases, U/L , aspartate transaminases, U/L . Others were blood urea nitrogen, \u03bcmol/L , chloride, \u03bcmol/L  and potassium, \u03bcmol/L . The changes in the biochemical parameters were not clinically significant.Patients who were enrolled into the nested arm of the study had safety evaluation done for their laboratory parameters. Of the 444 patients who provided blood samples 61.7% (274), 34.9% (155) and 3.4% (15) were recruited from the Navrongo, Dodowa and Kintampo sites respectively. Approximately 46% (95% CI 41.2 50.7) were males. Change in parameter values in individual patient, before and after treatment was assessed by comparing increases and decreases in the parameter. The findings were as follows; haemoglobin , total RBCs , total WBCs , total bilirubin  and total Chloride (63.4% vs. 34.2%: p<0.0001). Others were total alanine transaminase , total aspartate transaminase  total potassium  and total creatinine . In addition, the means across the entire subgroup- before and after treatment were assessed as follows; total mean haemoglobin g/dL ; total red cells x 10falciparum malaria in Ghana and the capacity of the three sites to undertake post-licensure surveillance of new medicinal products [The study determined additional safety data on DHP as a first line treatment for uncomplicated products .Approximately 95% of all the enrolled patients completed the study as planned. High treatment completion rate in a clinical trial suggests treatment acceptability given that participants are more likely to drop out of a study where adverse events are many and intolerable. The treatment completion rate in this study is similar to previous reports that showed the effectiveness of DHP for malaria in endemic countries , 21, 22.Patients taking antimalarials prior to being enrolled were mostly children with similar characteristics across sites and this did not influence the incidence of AEs. Four out of every five participants took at least one additional medication during the study participation.Taking additional medication during study participation was not associated with higher incidence of AEs compared to those who took medications prior to enrolment. Patients who had taken medications prior to enrolment were two times more likely to report AEs. Though self-medication increases access to medication and relief for acutely sick persons, it can also lead to serious drug interactions and AEs . Self-meHigher incidence of AEs was observed in the rural settings compared to the semi-urbanized settings. This was expected, as rural residents are more likely to have lower incomes, less access to health care and more likely to self-medicate , 24. TheTherapeutic drugs often demonstrate negative side effects on concomitant treatment hence the need to collect data on such drugs over time. Concomitant medications can either increase or decrease the metabolism of an investigational drug. When treatment metabolism is reduced as a result of concomitant medication, the resultant effects will be symptoms of over dosage even if the patient has taken the treatment as prescribed. Conversely, if there is increased metabolism, it may lead to faster excretion of the active metabolite of the investigational drug and it may not be able to reach its peak and potency at the prescribed dosage , 26.Statistically significant heterogeneity was found between AEs and body mass index with a positive correlation between them. This suggests a potential for higher plasma concentrations of DHP in obese persons and therefore its administration to patients based on weight should be done with circumspection. Piperaquine as a lipophilic drug with slow elimination and long half-life , 26 can Safety evaluation of DHP by electrocardiogram showed that, compared to the baseline value, there were significant changes in mean QTcF post-enrolment. For patients who had QTcF above the 500ms cutoff value however, only one person had higher QTcF on both day-3 postdose and day-7 post-enrolment. Only one patient with QTcF prolongation > 60ms had such on day-7 post-enrolment and none of them was clinically significant. These few and clinically insignificant QTcF changes are consistent with earlier reports ,15, whicIt should be noted that while this was a large study, the sample size may still not have been enough to detect rare serious adverse events. Secondly, some of the findings as for example some of the AEs may not have been properly assessed particularly in very young children. Further, the site- specific variation in AE incidence could reflect ascertainment bias and the fact that some patients who had higher parasite densities could be more symptomatic of their malaria.This study has added additional safety data to show that DHP is safe even in challenged and post licensure settings. No significant serious AEs or ASEI were documented. The findings indicate that in rural settings, obese patients and pre-enrolment medication were the significant factors associated with AEs. This study demonstrated the capacity of the three research sites to undertake phase four studies to detect rare AEs of new drugs over a much larger patient population. Further technical support and collaboration will enable the sites to conduct similar studies for other drugs and vaccines across Ghana and beyond.S1 Trend Checklist(PDF)Click here for additional data file.S1 Protocol(PDF)Click here for additional data file."}
+{"text": "Plasmodium vivax malaria differs from that of Plasmodium falciparum malaria in fundamentally important ways. This article reviews the guiding principles, practices, and evidence underpinning the diagnosis and treatment of P. vivax malaria.The diagnosis and treatment of  Plasmodium vivax infection can be broadly categorized into three purposes: identification of clinical cases (passive case detection [PCD]), surveillance (active case detection [ACD]), and clinical trials. Each scenario brings distinct requirements, tools, and pitfalls for diagnosis of the infection.The diagnosis of P. vivax antigen (rapid diagnostic test [RDT]). Clinical signs and symptoms alone, though frequently used, can neither differentiate malaria infection from other causes of febrile illness, nor distinguish between Plasmodium falciparum and P. vivax or malaria caused by another plasmodia. Competent microscopy is typically more sensitive, specific, and informative (with respect to parasite count and stages present) than RDT. However, the sustainability of microscopy services challenges most health-care systems where endemic malaria occurs.2The accurate diagnosis of vivax malaria in an acutely ill patient seeking routine care requires microscopy examination of a Giemsa-stained blood smear (microscopy), or use of an immunochromatographic cassette containing monoclonal antibodies to a Standards for malaria microscopy training, certification, and practice are available from World Health Organization (WHO).The density of parasitemia in patients with acute vivax malaria depends upon many factors, including naive versus a state of semi-immunity, age, delay in seeking treatment, self-treatment behavior before presentation, and likely a variety of host and parasite factors.7P. vivax malaria is typically an order of magnitude lower than P. falciparum in most clinical settings where both these species occur, thus increasing the risk of false negative microscopy diagnosis with acute vivax malaria. Repeated blood film examinations, or increasing the number of microscopy fields to 300 or more in patients suspected of having malaria should be carried out before confidently reporting the patient as negative for malaria parasites.The parasite density in P. falciparum is falling, there may be particularly low sensitivity for P. falciparum.The primary diagnostic threat in the clinical setting is poor sensitivity. Training of clinical microscopists should be aimed at maximizing parasite detection, even if that is at the cost of a lower level of specificity. In some settings where vivax malaria predominates and transmission of In addition to relatively intensive training and certification for the microscopist, competent microscopy requires a clean and well-functioning light microscope, clean glass slides, immersion oil with appropriate optical properties, and fresh filtered reagents for Giemsa staining. In many settings of endemic malaria, these essentials represent substantial challenges that cannot be reliably sustained. Where the quality of microscopy services cannot be assured, the use of RDT is recommended.8P. falciparum infection than that with P. vivax . Reaction to both indicates presence of P. falciparum, either alone or mixed with any other species, whereas reaction to pLDH alone indicates absence of P. falciparum and presence of any other species, but does not distinguish among these. Other RDTs do offer a P. vivax-specific diagnosis employing aldolase antigen capture with satisfactory performance.11The principle advantage of RDTs is their ease of use and sustainability in resource-challenged settings. RDTs are available from many commercial sources at relatively low cost . Most of these kits are stable at ambient temperature storage for many months. WHO offers standards for training and certification in the use of RDT. RDT see  . Some tACD is usually conducted through mass blood surveys with the intent of detecting asymptomatic carriers of infection for the purposes of either malaria control or measurement of prevalence of parasitemia in at-risk populations. Asymptomatic carriers typically have much lower parasitemias compared with those seeking medical attention for illness, and RDTs detect only a minority of these infected individuals. The majority of mass blood surveys  are performed using microscopy. The growing realization of the importance of detecting low-level parasitemias has spurred development of alternative technologies such as loop-mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR).15The process of microscopy for the purpose of ACD is similar to that described for PCD. A microscopist has some flexibility in the clinical setting regarding the degree of effort committed to an examination . However, in ACD, only one blood film is usually collected, and thus care must be taken to ensure that the same degree of diligence is applied for all samples collected.The most widely applied and validated molecular diagnostic is a nested PCR amplification of small subunit ribosomal RNA gene sequences.P. falciparum and pan-genus-specific reagents) showed the technique to be comparable to standard nested PCR technique and superior to expert microscopy.P. vivax in large field surveys in China.19LAMP of parasite-specific DNA is a more recent technology more suited to ACD in endemic settings. The technique does not require expensive thermocyclers or gel electrophoresis, the readout being a visual color change in a small test tube. Recent evaluation of a commercially available kit . PQ can cause serious hemolysis in patients with G6PDd, an inherited X-linked highly diverse disorder affecting about 400 million people. G6PDd occurs at a prevalence varying from < 1% to 30% among residents of endemic zones.The diagnosis of glucose-6-phosphate dehydrogenase deficiency (G6PDd) is an important diagnostic procedure before initiating the radical cure of Because the G6PDd is X-linked, it is either wholly absent or present (hemizygous) in males. In females, in contrast, it may be absent, homozygous, or heterozygous. Homozygosity in females is relatively rare , but heterozygosity is common. This is an important clinical and diagnostic problem due to the phenomenon of lyonization of X-linked genetic traits in women. This process results in two distinct populations of red blood cells, that is, those expressing defective or normal G6PD. The relative proportion of cells expressing abnormal enzyme averages 50% but ranges between 0% and 100%. In other words, heterozygous females may be G6PD normal or fully G6PD deficient; however, most have red blood cell populations presenting a mosaic of the two phenotypes. The clinical significance of this in the context of PQ toxicity is unclear, but challenges the widespread deployment of PQ to vulnerable populations.Ascertainment of G6PDd status in a clinical setting can involve the quantitative or qualitative measurement of G6PD activity in hemolysate, cytochemical microscopic examination of whole cells, or inference from genotyping extracted DNA using PCR technology.Standardized commercially available kits may be used for the spectrophotometric determination of G6PD activity in enzyme units (U) per gram of hemoglobin (gHb). Normal values range approximately from 7 to 10 U/gHb (at 30\u00b0C).In the quantitative assay, as in the qualitative assay, diagnosis of G6PDd in patients having recently suffered acute hemolytic anemia is problematic, since some patients may exhibit a normal phenotype as a consequence of the most vulnerable red blood cells having been removed and their replacement by young erythrocytes that have inherently higher G6PD activity levels. The effects of acute malaria on G6PD tests have not been evaluated.Most G6PD tests require a separate Hb measurement, and this imposes additional complexity and costs. The reason of doing so, however, is the impact of Hb level on the qualitative measurement, for example, below about 8 g/dL Hb, G6PD activity measurements trend sharply higher, probably falsely so.Standardized commercially available kits allow a visual determination of G6PD phenotype. These all involve the enzymatic conversion of NADP+ to NADPH by G6PD, which may be visualized directly (using fluorescent lighting) or indirectly . The most commonly used and widely validated qualitative assays are listed in 26Cytological techniques can be used to measure the degree of X-inactivation by lyonization in heterozygous females. These methods differentially stain individual red blood cells and permit estimation of the proportion of cells expressing defective G6PD enzyme. A variety of techniques have been described,28The principal advantage of G6PD genotyping is a diagnosis un-confounded by the physiological variables affecting G6PD activity such as patients suffering acute hemolytic anemia or lyonization in heterozygous females. The primary disadvantage, however, is the uncertainty of a \u201cnormal\u201d diagnosis when applying primers to selected known genotypes using standard PCR/restriction fragment length polymorphism (RFLP) techniques. The patient may be deficient but with a genotype not represented in the genetic analysis\u2014a significant hazard with G6PD, which is a complex gene with over 200 known variants documented. New variants of G6PD are routinely found wherever sufficiently detailed investigations can be carried out.Most patients with vivax malaria receive care at home or in the community outside of a clinic. Even when patients attend a clinic or hospital with a laboratory facility, there is often no capacity for G6PDd diagnosis. Similarly, diagnosis of G6PDd for survey purposes also occurs in rural settings and often cannot accommodate relatively sophisticated laboratory techniques.There is currently no validated point-of-care (POC) diagnostic device for G6PDd that is practical to apply where most malaria patients live. The obstacles to such a test include cost, complexity, heat sensitivity, and the need for a cold chain to transport and store the kit. A commercial kit in development, however, shows promise in overcoming those issues: the CareStart G6PD\u2122 , which is affordable, easy to use, and is not sensitive to ambient temperature fluctuations. However, although it is capable of reliably detecting G6PD deficiency below 30% of normal activity,33Surveys to assess the prevalence of G6PDd are often undertaken where laboratory capacity is limited, the test readout being subjective and visually categorized into G6PD deficient, intermediate, or normal. A newer quantitative technique\u2014WST8/1-methoxy-PMS\u2014has been successfully used to survey populations with a quantitative enzyme activity readout. Dried blood blots collected in the field and kept refrigerated are returned to the laboratory for G6PD activity determination in a 96-well spectrophotometric assay format.Evaluation of the safety and efficacy of therapies for acute vivax malaria, which necessarily includes both blood schizontocidal and hypnozoitocidal therapies, employs diagnostics for both the infection and G6PDd. Clinical trials of experimental therapies often demand higher standards of diagnosis than may be applied in routine clinical care, to provide greater assurance of subject safety and the precision of efficacy estimates.Trials of blood schizontocidal therapies typically only enroll patients with parasitemias above a certain threshold, to increase the likelihood that the associated fever is due to the infection. When parasitemias exceed 500/\u03bcL, almost any diagnostic technique will suffice. However, microscopy remains the method of choice for staging the parasite and documenting the parasite clearance.Screening for G6PDd is required for PQ therapy in trials and experimental blood schizontocide, which do not have a demonstrated safety profile in G6PD-deficient patients.The methodology of anti-relapse efficacy trials is challenging, but broadening enrollment criteria may be warranted to include those with subpatent infections. In this context, the use of highly sensitive diagnostic techniques such as LAMP may help to discriminate patient risk groups at enrollment. It is important to identify patients with G6PD variants known to be at high risk of severe hemolysis. Historically, the NADPH spot test  has been used for this purpose, including a series of trials of PQ as primary prophylaxis in the past decade where subjects received large cumulative doses of this drug over prolonged periods.Clinical efficacy is defined by the clearance and recurrence of parasitemia. In most patients these occur at relatively low levels of patency, and thus expert microscopy is crucial in the assessment of blood films. In contrast to routine clinical microscopy, the microscopist serving clinical trials must undergo rigorous certification to minimize false-positive outcomes, which have potential to significantly underestimate clinical efficacy.P. vivax are to reduce the immediate risk to the host, eradicate peripheral asexual parasitemia, prevent the recurrent infection, and interrupt the cycle of transmission.P. vivax to form dormant liver stages (hypnozoites) capable of causing relapsing infections weeks to months after the initial blood-stage infection, provides a major challenge to the complete eradication of parasites from the body. Since no single drug achieves all of these aims, a combination of antimalarials is required targeting a variety of specific key elements of the parasite life cycle.1The goals of antimalarial treatment in P. vivax is known to be chloroquine (CQ) sensitive, the WHO recommends 3 days of CQ or an artemisinin combination treatment plus 2 weeks of PQ .P. falciparum and P. vivax.In areas where P. vivax, the exception being the antifolates, which act slowly,P. vivax in clinical trials.Most commonly used antimalarial drugs are also active against the asexual stages of P. vivax.P. vivax malaria, but it is not yet recommended by the WHO. Artemisinin in combination with effective partner drug have shown excellent cure rates in P. vivax infection.Artemisinin combination therapies (ACTs) are the treatment of choice for CQR P. falciparum and P. vivax infections, offering a pragmatic approach with operational efficiencies.P. vivax has led to a number of countries adopting ACTs as first-line treatment for P. vivax. These include DHA\u2013piperaquine in Indonesia and Cambodia, and artemether-lumefantrine in Papua New Guinea, Solomon Islands, Sudan, Namibia, South Africa, and Vanuatu.P. falciparum, where P. vivax is sympatric (see Drug Development for P. vivax section).The deployment of an ACT-based strategy permits a unified policy for treating both Plasmodium vivax is very sensitive to artemisinin and its derivatives. In the absence of comparative drug trials, physicians have tended to adopt a similar treatment approach for severe vivax malaria as for severe falciparum malaria,Severe and fatal vivax malaria has been reported from Indonesia,P. vivax malaria.Large-scale multicentered trials in patients in Asia and Africa have demonstrated clear superiority of intravenous artesunate over quinine in reducing case fatality rate in severe falciparum malaria.\u2022Artesunate: 2.4 mg/kg body weight, intravenously or intramuscularly given on admission (time = 0), then at 12 and 24 hours, and then once a day. This is the treatment of choice.\u2022Artemether: 3.2 mg/kg body weight, intramuscularly given on admission, then 1.6 mg/kg body weight per day.\u2022Quinine: 20 mg quinine salt/kg body weight on admission  followed by maintenance dose of 10 mg/kg body weight 8 hourly .Specific antimalarial treatment recommended in severe vivax malaria includes the following in order of preference:Parenteral antimalarials should be administered for at least 24 hours. Once the patient can accept oral therapy, full course of oral ACT should be given to the patients. Full details are available in the latest Malaria Treatment Guidelines.1P. vivax were published in 1989 from Australian travelers to Papua New Guinea,P. falciparum. Although intrinsic differences in the transmission dynamics between these two species may account for much of the time lag, it is likely that this also reflects the inherent complexity of defining antimalarial treatment efficacy of P. vivax.P. vivax has been documented on the island of New Guinea, where patients treated with CQ have been observed to have early clinical deterioration requiring hospitalization, delayed parasite clearance, and early recurrent parasitaemia.P. vivax, albeit to a lesser degree, has been reported from across the vivax-endemic world  and 21 treatment arms assessing the clinical efficacy of ACTs . CQ efficacy has been quantified at 97 geographical locations, of which 47 (48%) revealed reduced potential susceptibility . Numerous studies demonstrate the great therapeutic benefit of primaquine therapy at low  was 25% (range: 0\u201390%) at 4\u20136 months, compared with 6.7% (range: 0\u201359%) in the 82 studies of low-dose PQ . High-dose PQ regimens  were assessed in 28 treatment arms, and were associated with a median recurrence rate of 0% (range: 0\u201315%) at 1 month. However, comparisons between dosing regimens need to be interpreted with caution, since they are confounded by the marked heterogeneity in study design, dose regimens, and idiosyncrasies of the endemic setting notably variable rates of reinfection.A recent review of the published literature identified 87 clinical trials presenting data on 59,735 patients enrolled in 156 treatment armsP < 0.001). Two studies enrolling 171 patients demonstrated high effectiveness of high-dose PQ compared with a control arm . Two recent studies from Indonesia documented good efficacy (> 95%) against relapse in P. vivax among soldiers followed for a year (without risk of reinfection) after directly observed high-dose PQ therapy either following108The frequency and the timing of relapses are determined by sporozoite inoculum and parasite relapse phenotype, and this results in huge regional variation in the absolute risk of relapse independent of PQ efficacy.Current guidelines recommend a 14-day course, but such prolonged courses of treatment can result in significant problems with adherence.P. vivax. This may reflect the absence of the phenomenon or the extraordinary difficulty of gathering unambiguous evidence of it. Ingram and othersP. vivax experimental challenge subjects.114There is no evidence of resistance to therapeutic doses of PQ by hypnozoites of The most common serious adverse effect (SAE) following PQ is intravascular hemolysis associated with the passage of dark or black urine and mild jaundice. There are two potentially lethal consequences; life-threatening anemia and acute hemoglobinuric renal failure. From 69 studies and additional case reports (excluding data from mass treatment campaigns) that evaluated adverse events to PQ, no SAEs were reported in G6PD-normal individuals, with the possible exception of one psychotic reaction in an individual with undetermined G6PD status.P. vivax and highly prevalent Mediterranean or Mediterranean-like severe G6PDd variantsIn total 15 deaths associated with PQ have been reported over the past six decades, of which 13 were from severe haemolysis.P. vivax episode is associated with malaria-related hemolysis and dyserythropoiesis and a cumulative risk of anemia. This contributes to impaired development, school and work performance, and in high-transmission areas to anemia-related deaths. Without prolonged monitoring over repeated episodes of malaria, the benefits of radical cure may not be appreciated.Primaquine is underused. Several studies highlight that even in areas where PQ is recommended, in practice it is prescribed rarely.If there is no available G6PD POC test, it is difficult to generalize on the correct approach to patient management. The risk versus benefit weighing depends on the prevalence and severity of G6PDd in the area, the degree of anemia, and the availability of blood transfusion  as well as the probability of relapse and the likely dose regimen required (the benefit). In some circumstances the assessment may favor withholding PQ, and in others it may favor starting the radical curative regimen after educating the patient about the possible risks and informing the patient that they should stop the drug if they fail to recover within 3 days, become ill, or their urine becomes red or black.P. falciparum malaria.P. vivax malaria given the 0.75 mg/kg PQ dose weekly for 8 weeks.The evidence of PQ safety in known G6PD-deficient subjects or patients is limited to just four of the many dozens of known G6PD variants  and less than a couple dozen subjects. In general, variants with > 5% residual G6PD activity showed relatively mild and self-limiting hemolysis, whereas those with < 5% activity showed severe hemolysis. There is insufficient evidence on which to guide PQ therapeutic decisions based on known or suspected degree of G6PD enzyme impairment. Any diagnosis of G6PDd in a male hemizygote should be considered indicative of risk of serious harm and prompt withholding standard daily PQ therapy against relapse.Females present a more complex and difficult therapeutic problem. Homozygous females carry two mutant X alleles and phenotypically mirror hemizygous males in having 100% defective red blood cell populations. Female heterozygotes possess both wild type and mutant G6PD phenotypes, expressed as mosaicism in their red blood cell populations. Random inactivation of one or the other X chromosome during embryonic development results in variable proportions of wild type and mutant phenotypes among their red blood cells .There are relatively few data on the safety of PQ in young children, but there is no indication that toxicity in children differs from that in adults. Thus PQ is recommended in young children above 6 months of age, and not in infants .Primaquine is contraindicated in pregnancy because of the unknown G6PD status of the fetus and thus the risk of inducing intrauterine hemolysis. Excretion of PQ in breast milk is not known so caution should be exercised and preferably PQ should not be given. If PQ is prescribed to a lactating mother, the baby needs to be observed closely.Pneumocystis jirovecii infections without apparent complications.There is no specific information on the safety of PQ in this important patient group. Primaquine has been used as a second-line treatment (with clindamycin) of 450 (predominantly 2D6) to reactive intermediates that mediate both the antimalarial effects and hemolytic toxicity. Theoretically, inhibitors such as quinidine, ketoconazole, paroxetine, and fluoxetine may reduce toxicity, but these predictions require further study. Historic examples of likely drug\u2013drug interactions have occurred between pamaquine (a PQ precursor) and mepacrine  impacting safety, and between quinine and CQ impacting efficacy against relapse.126Primaquine is metabolized by monoamine oxidase to the biologically inert, but slowly eliminated, carboxyprimaquine, and via CYPP. falciparum and P. vivax commonly occur together; mixed infection is very common, and many patients with falciparum malaria harbor P. vivax hypnozoites that commonly cause relapse 3\u20138 weeks after the acute illness. The emergence of CQ resistance in many endemic regions, along with > 50% prevalence of hypnozoites of P. vivax in patients diagnosed and treated for P. falciparum, provides a strong rationale for a unified policy for therapy of uncomplicated malaria of any species.P. falciparum are also efficacious against P. vivax.130Outside of Africa, Tafenoquine , is an 8-aminoquinoline with in vivo anti-hypnozoite activity. The mechanism of action of TQ, as for all the 8-aminoquinoline class of drugs, is unknown. TQ has a long half-life enabling radical cure treatment as a single dose.P. vivax malaria patients with G6PDd. The objective of the phase I dose escalation study was to determine the safety  and tolerability of TQ in G6PD-deficient healthy subjects without the influence of disease-related confounding factors. For the initial dose escalation phase, G6PD-deficient heterozygous female healthy volunteers with enzyme activity range between 40% and 60% of the site median normal value were recruited.Phase I and IIb studies have been conducted and phase III efficacy and safety studies are ongoing.Current options for radical cure are dependent upon the 8-aminoquinolines. Compounds that are protective against the oxidative damage from PQ or TQ have potential for safe deployment and accessibility to radical cure. Novel assays exist to screen for such products, for instance an in vivo immunodeficient murine model transfused with human G6PD-deficient blood.During the development of PQ in the late 1940s, one volunteer experienced deep hemolysis after a few 60 mg daily doses of the 8-aminoquinoline isopentaquine. Months later, however, when the same dose of isopentaquine was administered with an oral 500 mg dose of methylene blue , the subject completed the 14-day regimen without signs of hemolysis.134\u00ae, Martindale Pharma, Buckinghamshire, United Kingdom) as a solution for injection, 5 mg/mL for the following indication: treatment of medicinal and chemical products\u2013induced methemoglobinemia. Methylthioninium chloride is indicated in adults and children above the age of 3 months. However, it is currently contraindicated in patients with G6PDd due to the risk of hemolytic anemia.MB is registered . This compound is under investigation as an 8-aminoquinoline-induced hemolysis-blocking agent.Bulaquine (BQ) is a prodrug of PQ and is believed to hydrolyze in the stomach to PQ. Clinical trials of BQ have failed to show that this agents is superior to PQ in terms of safety or anti-relapse efficacy for relapse and this role in clinical practice remains limited.137Plasmodium cynomolgi/macaque relapsing malaria model, tinidazole cured one of six macaques studied with an apparent mild delay to relapse in the other five monkeys. The only study in humans was conducted in healthy G6PD-normal Thai adults with P. vivax infection. Subjects were randomized to treatment with either 2 g of oral tinidazole daily for 5 days or standard therapy with PQ (30 mg base per day for 14 days); all patients received CQ (at 25 mg/kg/day for 5 days). Six of the first seven subjects treated with tinidazole relapsed before day 63. The authors of this study concluded that tinidazole was ineffective in preventing relapse of P. vivax given at 2 g a day for 5 days concurrently with CQ. The macaque relapsing model appeared to predict correctly the outcome in humans.138Tinidazole is a 5-nitroimidazole used for the treatment of amebiasis and giardiasis. In the P. vivax liver-stage assay currently available to drive drug discovery; hence, the current strategy to identify novel compounds relies largely on a pragmatic approach and the application of surrogate assays. The current approach consists of screening a very large number of compounds in a high-throughput P. falciparum blood-stage assay, and selection of the most likely candidates to be tested in a secondary lower throughput assays such as the Plasmodium yoelii liver-stage assay.P. cynomolgi liver-stage assayP. cynomolgi sporozoite infections of primary rhesus hepatocytes to generate, reproducibly, small parasite forms. The latter have been shown to reactivate in vitro, suggesting that they may be hypnozoites.P. yoelii assay are then tested further to find series with a better therapeutic ratio than PQ.There is no reliable P. vivax liver-stage in vitro assay is under development to replace the current model. The aim is to develop assays with a throughput that is sufficient to screen large libraries directly. The development of this assay as well as other tools to aid in the development of anti-relapse molecule have been reviewed elsewhere (see the article by Olliaro and others).A P. yoelii assay.http://www.mmv.org/research-development/rd-portfolio).Analysis of over 5 million compounds from approximately 20 compound collections from the pharmaceutical and biotechnology companies, academic diversity collections, and diversity providers, generated around 25,000 compounds with activity against the blood stage of the parasite IC50s ranging from 1 to 3 \u03bcM. The liver-stage activity of a selection of those hits was then assessed using the P. yoelii liver-stage assay are currently being evaluated in the P. cynomolgi S assay for their potential effect on hypnozoites.Work has also been undertaken to evaluate the liver-stage activity of the current portfolio of schizonticidal compounds, with more than 50 anti-infectives currently in use or under development. Several of these antimalarials with schizontocidal activity in a Progress in trying to identify potential new candidates as anti-relapse agents has been made; it is however important to note that beyond those mentioned here, there are no new compounds in the global malaria portfolio in preclinical development specifically for their anti-relapse activity. The first generation of compounds that have been specifically optimized based on their potential for anti-relapse activity is expected to enter preclinical development in 2014.P. vivax has changed little over the last 60 years although recent technological advances promise to deliver better, more sensitive and field-adapted diagnostics for parasitemia and G6PDd, and to identify novel antimalarial compounds. Plasmodium falciparum is resistant to CQ in most endemic regions. CQR P. vivax, although slower to appear than P. falciparum, is spreading throughout the endemic world. A unified treatment policy for malaria caused by any species of Plasmodium may offer significant individual and public health advantages over the current strategy that relies on CQ plus PQ. ACTs are highly effective against P. vivax, and the more slowly slowly-eliminated ACTs suppress the early relapses, but whether this reduces transmission in the long term remains to be proven. The most important potential chemotherapeutic means of interrupting transmission of vivax malaria will be radical cure and thus prevention of all future relapses by using a safe and well-tolerated hypnozoitocidal medication. Primaquine is the only licensed hypnozoitocidal drug, but studies are underway to find more practical ways of deploying it. Renewed efforts are underway to develop alternative treatments for safer and more effective radical cure.The treatment of P. falciparum and P. vivax.A unified treatment policy for malaria of any parasitological cause will confer significant individual, public health, and operational benefits in regions co-endemic for P. vivax, which in the short term reduces the risk of anemia and the transmissibility of the parasite.Slowly eliminated ACTs offer posttreatment prophylaxis against the first relapse of tropical strains of P. vivax malaria, and may be achieved by adopting short-course, high-dose regimens, although the safety of these regimens has not been established.Ensuring adherence to a complete course of PQ is the greatest challenge in the control of A robust bedside test for G6PD deficiency is urgently needed to define the toxicity profile of PQ and facilitate the safe deployment of anti-relapse therapy.Supplemental Annexes."}
+{"text": "Plasmodium falciparum gametocyte exported protein-5 (PfGEXP5) transcript analysis as an important tool for evaluating the earliest (ring) stage sexual gametocytes in the blood of infected individuals. We show that gametocyte rings are detected in the peripheral blood immediately following establishment of asexual infections\u2014without the need for triggers such as high-density asexual parasitemia or drug treatment. Committed gametocytes are refractory to the commonly used drug piperaquine, and mature gametocytes reappear in the bloodstream 10 days after the initial appearance of gametocyte rings. A further wave of commitment is observed following recrudescent asexual parasitemia, and these gametocytes are again refractory to piperaquine treatment. This work has implications for monitoring gametocyte and transmission dynamics and responses to drug treatment.The recent focus on the elimination of malaria has led to an increased interest in the role of sexual stages in its transmission. We introduce  Plasmodium falciparum causes >200 million cases of malaria each year and kills approximately 400000 children [children . Treatmechildren . Resistachildren .P. falciparum gametocyte transitions through 5 distinct stages over a period of about 10 days. Early ring stage gametocytes are morphologically indistinguishable from asexual rings. Later stage gametocytes (stages II\u2013IV) of P. falciparum gradually elongate to adopt a characteristic crescent or falciform shape [As the World Health Organization (WHO) shifts its focus from disease control to elimination, it is critically important to understand the drivers and dynamics of carriage of both asexual parasites, which cause disease, and sexual-stage gametocytes, which are responsible for disease transmission. Gametocytogenesis is initiated when a sexually committed merozoite invades a red blood cell (RBC) . The sexrm shape . Stage Irm shape . The onlGiven the lack of specific markers for ring-stage gametocytes, it has been difficult to answer a number of fundamental questions, such as whether these early stage gametocytes are sequestered or are freely circulating , 10. IndP. falciparum chloroquine resistance transporter (PfCRT) [Piperaquine is a bisquinoline antimalarial that was developed in the 1960s. It was first used as a monotherapy in China, leading to the development of resistance in that country . It comp (PfCRT) ; indeed  (PfCRT) , 22. PipP. falciparum infections with piperaquine rapidly clears asexual parasitemia but is followed some time later by the appearance of mature gametocytes [The experimentally induced blood-stage malaria (IBSM) infection model  has provetocytes . HoweverPlasmodium falciparum gametocyte exported protein-5 (PfGEXP5) has been recently identified as a specific marker of early stage sexually differentiated parasites [gexp5 transcript levels provides a convenient and reliable method for enumerating ring-stage gametocytes both in vitro and in vivo. Using quantitative reverse transcriptase assays specific for gexp5 (gametocyte rings), skeleton-binding protein-1 , and P. falciparum sexual-stage antigen-25 , we have characterized the production of gametocytes in experimental human malaria infections before and after treatment with piperaquine monotherapy.arasites . In thisVery tightly synchronized schizont stages  of a high gametocyte-producing 3D7 parasite line  were obtThe clinical study component (ANZCTR no. 12613000565741) was conducted at the contract research organization, Q-Pharm . Blood samples were collected from subjects in the third cohort of a previously reported study whose objective was to investigate the antimalarial activity of piperaquine . The PCRgexp5, sbp1, pfs25, and 18S ribosomal RNA, using assays described in detail in the Supplementary Materials. The assays were designed to exclude amplification of any contaminating genomic DNA targets. Sample mRNA and controls were reverse-transcribed to cDNA, and qRT-PCR was performed as described . Positive, negative, and \u201cno template\u201d controls were included, whereas standard curve material (7 \u00d7 10-fold dilutions) was comprised of linearized and purified plasmid DNA target standards. These standard curves were used to report the absolute number of mRNA transcripts in samples by interpolation using a linear regression model of the measured concentration (transcripts per microliter) of the standard curve material .Parasitemia in the clinical trial was monitored using a real-time, quantitative PCR using primers and a hydrolysis probe that targets the 18S rDNA gene . For quagexp5, tightly synchronized (approximately 2-hour window) schizonts (approximately 3% parasitemia) were allowed to reinvade, forming a high parasitemia culture (approximately 13%) containing a proportion of committed gametocytes. Two hours after reinvasion, any remaining schizonts were removed, and samples of the culture were harvested at 6, 12, and 24 hpi  can be detected in the ring stage of parasites that are committed to sexual-stage development from 14 hours post invasion (hpi) but is absent in asexual rings . To deted 24 hpi , top rowgexp5 is transcribed at only very low levels in schizont-stage parasites (pfs25 (PF10_0303), are low in schizonts and during the early stages of commitment, with residual signal likely representing low-level contamination with late-stage gametocytes , sbp1 transcript levels were readily detectable, consistent with the presence of circulating ring-stage parasites  and well before antimalarial treatment with piperaquine was administered. A trough in gexp5 transcript levels on days 5\u20136 is consistent with disappearance of the first round of ring-stage gametocytes due to sequestration. A marked increase in gexp5 transcript levels was observed on day 7 . A rebound in sbp1 transcript level, accompanied by an increase in gexp5 transcript, was observed in the 3 subjects who experienced recrudescent infection on days 11, 23, and 27 after infection  gametocytes appear in the circulation [pfs25 mRNA transcripts remained low both before and after piperaquine treatment . In this context, it should be noted that the limit of detection with Giemsa thick film is approximately 10000 parasites per milliliter [In this study, we followed asexual and sexual blood-stage development in subjects, using lliliter .gexp5 transcript increased dramatically as the parasitemia rose. The paucity of quantitative data prevents us from concluding whether this is more consistent with density-dependent enhancement of commitment or constitutive commitment that rises with increasing parasitemia. Nevertheless these data indicate that gametocytogenesis occurs at very low densities and in clinical malaria is very likely already at a high level by the time patients develop symptoms. These presymptomatic individuals will potentially contribute to malaria transmission.Interestingly we found that the level of There has been significant interest in determining the compartment in which commitment takes place. Stage II\u2013IV gametocytes are not observed in the peripheral blood due to sequestration in the spleen and bone marrow . Gametocpfs25-positive) gametocytes appeared in the circulation 9\u201310 days after the appearance of the initial surge of ring-stage (gexp5-positive) gametocytes and continued to circulate even after the follow-up treatment with a higher dose of piperaquine. This indicates that sequestered gametocytes are not killed by the piperaquine exposure achieved in this study. This is consistent with the relatively poor activity of piperaquine in vitro against midstage gametocytes [We followed changes in circulating asexual and sexual ring stages after treatment with piperaquine. The number of asexual and sexual parasites continued to increase following treatment, consistent with a previous study showing that piperaquine has little activity against ring-stage parasites . After 2etocytes , 40.The parasite line used in this study, 3D7, is piperaquine sensitive; however, there is increasing evidence for the emergence of resistance to this drug in the Greater Mekong subregion. Moreover, dose selection for piperaquine is complicated by variable levels of absorption , by diffThis study reveals that gametocytogenesis is initiated very early in human infections, almost certainly prior to the point at which patients seek treatment. Coupled with the lack of activity of piperaquine against both asexual and sexual ring-stage parasites and its inadequate activity against mid- and late-stage gametocytes, this raises significant concerns about using combinations that include piperaquine, particularly in the face of emerging resistance. The observation that DHA-piperaquine is associated with higher levels of gametocytemia after treatment, compared with other artemisinin combinations , merits gexp5 transcript-based assay with sufficient sensitivity to detect ring-stage gametocytes when the parasite density is <100 parasites per milliliter provides a very important tool to address key epidemiological questions relating to the dynamics of parasite transmission. These tools are critical to current efforts to progress toward malaria elimination.Efforts to eradicate malaria from particular regions will be highly dependent on effective methods of targeting gametocytes. The IBSM model offers a very useful tool to determine whether individual components and particular drug combinations are likely to carry the liability of permitting gametocyte production. Moreover, the availability of a Supplementary DataClick here for additional data file."}
+{"text": "Plasmodium vivax parasites have a unique dormant stage that can cause relapses weeks or months after the initial infection. These dormant parasites are among the main challenges of vivax malaria control as they constitute a reservoir that is difficult to eliminate. Since field studies are confounded by reinfections and possible recrudescence of drug-resistant parasites, most analyses of P.\u00a0vivax relapses have focused on travelers returning from regions of malaria endemicity. However, it is not clear whether these individuals accurately recapitulate the relapse patterns of repeatedly infected individuals residing in areas of endemicity. Here, we present analyses of vivax malaria patients enrolled in a tightly controlled field study in Cambodia. After antimalarial drug treatment was administered, we relocated 20 individuals to a nontransmission area and followed them for 60\u00a0days, with blood collection performed every second day. Our analyses reveal that 60% of the patients relapsed during the monitoring period. Using whole-genome sequencing and high-throughput genotyping, we showed that relapses in Cambodia are often polyclonal and that the relapsing parasites harbor various degrees of relatedness to the parasites present in the initial infection. Our analyses also showed that clone populations differed dynamically, with new clones emerging during the course of the relapsing infections. Overall, our study data show that it is possible to investigate the patterns, dynamics, and diversity of P.\u00a0vivax relapses of individuals living in a region of malaria endemicity and reveal that P.\u00a0vivax relapses are much more pervasive and complex than previously considered.  P.\u00a0vivax parasites can remain dormant in the liver and relapse weeks or months after the initial infection, greatly complicating malaria control and elimination efforts. The few investigations of this dormant stage have relied on travelers and military personnel returning from areas of malaria endemicity. However, it is not clear whether these individuals, exposed to a limited number of infections, accurately represent the patterns of relapses of individuals living in areas of endemicity, who are repeatedly infected by P.\u00a0vivax parasites. Our study combined tightly controlled fieldwork with comprehensive genomic analyses, and our report provides a first opportunity to investigate the patterns, dynamics, and diversity of P.\u00a0vivax relapses directly with individuals living in areas of endemicity. Plasmodium falciparum malaria, the results for P.\u00a0vivax, the main cause of human malaria outside Africa, are much less encouraging  . We then relocated these patients to an area with no transmission of malaria and monitored them for at least 60\u00a0days .We recruited 20 P.\u00a0vivax parasites at 3 and 6\u00a0days after CQ treatment as measured, respectively, by microscopy and PCR . However, among the 20 patients, 12 (60%) displayed recurrences of P.\u00a0vivax parasites within 60\u00a0days . In all five cases, the recurring parasites displayed thousands of such new alleles  genome sequencing data from the  alleles , indicat10.1128/mBio.01888-17.5TABLE\u00a0S1\u00a0P.\u00a0vivax reference genome sequence (and the percentage mapped) are indicated, as well as the mean genome coverage (Mean Cov.) and number of nucleotides sequenced at more than 50\u00d7 coverage (Cov>50\u00d7). Download TABLE\u00a0S1, PDF file, 0.2 MB.The table indicates the results of the whole-genome sequencing for each patient isolate, collected at enrollment (D0) or during relapse (DX). The number of total read pairs generated and number of read pairs mapped to the Copyright \u00a9 2018 Popovici et al.2018Popovici et al.Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the To confirm the genome sequencing results and to test whether the recurring clones that had not been detected in the initial infection might have been present at low abundance, we generated genotyping data with very high coverage  and whole-genome sequencing (y\u00a0axis). The graph shows the data for the parasites present in BL02 at day 0. Download FIG\u00a0S1, TIF file, 0.04 MB.Comparison of the allele frequencies determined by genotyping and whole-genome sequencing. Each dot represents one SNP and is displayed according to the allele frequency determined by genotyping  and will need to be confirmed with a larger sample size.Previous studies have shown that t clones \u201318. Cons clones \u2013. Interes10.1128/mBio.01888-17.2FIG\u00a0S2\u00a0FIG\u00a0S2, TIF file, 0.1 MB.Example of a polyclonal relapse. The figure shows the reference allele frequency plot for the parasites present in the relapse of BL12. Note the two peaks at ~50% and 5%, indicating that at least three parasites  were present at the time of relapse. Download Copyright \u00a9 2018 Popovici et al.2018Popovici et al.Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the P.\u00a0vivax genome, of new alleles observed in relapsing parasites but not detected in the initial infection. In one patient (BL10) who displayed both monoclonal initial and relapsing infections, these new alleles were distributed evenly throughout the genome, indicating that these two clones were unrelated  or by the introduction of a new clone (bottom row), starting from an infection with either two clones (left) and three clones (right). Note that a change in the proportion of the clone populations typically leads to asymmetrical changes in allele frequencies (red arrow), while the introduction of new parasites is more clearly shown by the appearance of new alleles (green circles). Download Copyright \u00a9 2018 Popovici et al.2018Popovici et al.Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the 10.1128/mBio.01888-17.4FIG\u00a0S4\u00a0x\u00a0axis and the y\u00a0axis, respectively). For example, the graphs show that a single clone was present in BL18 at day 56 and day 57 (left panel) but that a minor clone appeared at day 58 . Similarly, two clones were present in BL10 and in similar proportions at day 53 and day 55 , but the minor clone was lost by day 56 (right panel). See also FIG\u00a0S4, TIF file, 0.4 MB.Changes in clone populations during the course of the relapses. The figure shows the allele frequencies determined for each genotyped SNP (blue dots) at two consecutive time points , the number of SNPs successfully genotyped , and the number of clones estimated to be present based on the genotyping data . Download TABLE\u00a0S2, PDF file, 0.2 MB.The table summarizes the results of the monitoring of relapsing infections. For each patient and each day following detection of Copyright \u00a9 2018 Popovici et al.2018Popovici et al.Creative Commons Attribution 4.0 International license.This content is distributed under the terms of the For one of the patients, BL02, who remained asymptomatic for 30\u00a0days during the monitoring period, we were able to complement the genotype data with whole-genome sequencing of the parasites at days 41, 48, and 55 . These gPlasmodium vivax. While many factors contribute to the parasite\u2019s resilience, the existence of a dormant stage in P.\u00a0vivax plays a critical role, since it provides a reservoir of parasites that cannot easily be targeted using current approaches  to exclude possible reinfections. Second, we ruled out the possibility that recurring infections were caused by recrudescence of drug-resistant parasites by (i) testing for P.\u00a0vivax DNA in patient blood every second day after treatment using a highly sensitive detection method, (ii) comparing the genotypes of the parasites in the initial infections and recurrences, (iii) verifying that drugs in patients\u2019 blood on the day of recurrence were below therapeutic levels, and (iv) successfully treating symptomatic recurrences with another full course of chloroquine .Successful elimination of malaria worldwide is notably challenged by the difficulties encountered in our attempts to efficiently control proaches . Unfortuproaches \u201313, 21. P.\u00a0vivax relapses were pervasive among patients living in a low-transmission area of Cambodia; among the 20 patients enrolled in this study, 12 (60%) relapsed within 60\u00a0days . While only five of these patients developed symptoms of malaria from these recurring infections, our results emphasize the importance of relapses for malaria control; without radical cure, these patients may present opportunities for further transmission of the disease despite the effective treatment against blood stage parasites . One hypothesis for explaining this observation is that patients with polyclonal infections had higher levels of previous exposure to P.\u00a0vivax and, therefore, carried a greater load of hypnozoites in their liver . This hypothesis is also consistent with the observation that some of these relapsing clones were not detected at day 0 and were not related to those initially present, indicating that they likely originated from infections predating our study . Overall, these findings indicate that many hypnozoites were present in the liver of these infected patients and emphasize that successful malaria eradication will require specific elimination of this important reservoir.Using this rigorously controlled setting, we observed that arasites . Indeed,arasites , 11, 13 P.\u00a0vivax DNA was able to be detected before) . These observations could suggest that reactivation of P.\u00a0vivax hypnozoites occurs continuously but that sustained parasitemia can be observed only once chloroquine has been cleared. While our findings will need to be validated and to be tested in other settings with different transmission levels, the results from this study indicate that the numbers of hypnozoites in infected individuals might be much greater than previously thought.Beyond enabling rigorous assessment of the prevalence of relapses, our report also provides a unique opportunity to preliminarily assess the biological mechanisms underlying this phenomenon. The observation that many relapses were polyclonal, at the earliest time of genotyping, could suggest that an external stimulus triggers the reactivation of several different hypnozoites at once . Fever aP.\u00a0vivax relapses further but also to understand the pathophysiology of malaria better and to identify host and parasite factors that influence the transition from asymptomatic infection to the development of clinical malaria. Overall, our study data illustrate the promise of modern fieldwork, combined with extensive genomic analyses, for better understanding the biology of P.\u00a0vivax parasites and, hopefully, better controlling vivax malaria.Finally, while we observed pervasive relapses, the fate of the relapsing clones varied considerably among patients, as did the clinical consequences of the infection; the immune system controlled and eliminated the relapsing parasites in some patients in a few days, one patient remained infected and asymptomatic for several weeks, and five patients developed symptoms of malaria and had to be treated. Such variations illustrate the potential of this study design to provide a rigorous framework not only to study P.\u00a0vivax . After written informed consent was obtained, all patients received a supervised standard 3-day course of CQ (250 mg). This study was approved by the National Ethic Committee of the Cambodian Ministry of Health (no. 038NECHR) and registered on ClinicalTrials.gov . Details about the recruitment criteria and patient characteristics are reported elsewhere . For the relapse study presented here, we analyzed only patients relocated to a malaria-free area (city of Banlung) for the entire monitoring period (n = 20).We conducted a prospective clinical drug efficacy study in Ratanakiri Province in Northeastern Cambodia to assess the efficacy of a standard 3-day course of chloroquine (CQ) against vivax malaria. Briefly, febrile patients (or patients with a history of fever in the last 48\u00a0h) seeking antimalarial treatment after non-falciparum rapid diagnostic test (RDT)-positive diagnoses  were offered an opportunity to participate in the study. Exclusion criteria were pregnant or lactating women, signs of severe malaria, known other illnesses, inability to provide informed consent, and age of less than 15\u00a0years. We recruited 40 male and female febrile patients from villages within 50\u00a0km of the Province capital Banlung and infected solely with P.\u00a0vivax DNA by reverse transcription-PCR (RT-PCR), the patient was retreated by chloroquine  and the monitoring continued for a minimum of 14\u00a0days.Before treatment, we collected 4\u00a0ml of blood from each patient. After clearance of the initial infection, which occurred 3 to 5\u00a0days after CQ administration , we recorded axillary temperature and collected 500\u00a0\u00b5l of capillary blood from each patient every second day until the end of the monitoring period at day 60. If one patient developed symptoms of malaria during the monitoring period and was confirmed to be positive for Plasmodium parasites by RT-PCR . We extracted parasite DNA from each blood sample by the use of a QIAamp DNA Blood Minikit and determined the presence and species of y RT-PCR . We alsochrome B ) to humachrome B ) determiP.\u00a0vivax strains  and during all subsequent follow-ups (every second day) using the sequencing-based assay described in reference\u00a016. Briefly, we extracted DNA from each sample and used multiplex PCRs to amplify 100 to 300\u00a0bp of DNA sequence surrounding SNPs among Cambodian  strains . A total strains  using Bo strains  and usedP.\u00a0vivax reference genome  all patients before CQ treatment and (ii) the patients who, during the follow-up period, developed symptoms of malaria or sustained asymptomatic parasitemia. After leucocyte depletion performed using cellulose columns , we isole genome  using Boe genome  and analTo determine if a given infection was polyclonal, we looked for nucleotide positions with multiple alleles  andPRJNA420510.The sequence data are freely available in NCBI SRA under BioProject accession no."}
+{"text": "Plasmodium falciparum, but P. vivax has been more refractory. Its ability to form dormant liver stages confounds control and elimination efforts. To compare the efficacy and safety of primaquine regimens for radical cure, we undertook a randomized controlled trial in Ethiopia.Recent efforts in malaria control have resulted in great gains in reducing the burden of P. vivax mono-infection were enrolled and randomly assigned to receive either chloroquine (CQ) or artemether-lumefantrine (AL), alone or in combination with 14 d of semi-supervised primaquine (PQ) . A total of 398 patients  were followed for 1 y, and recurrent episodes were treated with the same treatment allocated at enrolment. The primary endpoints were the risk of P. vivax recurrence at day 28 and at day 42.Patients with normal glucose-6-phosphate dehydrogenase status with symptomatic P. vivax infection at day 28 was 4.0% (95% CI 1.5%\u201310.4%) after CQ treatment and 0% (95% CI 0%\u20134.0%) after CQ+PQ. The corresponding risks were 12.0% (95% CI 6.8%\u201320.6%) following AL alone and 2.3% (95% CI 0.6%\u20139.0%) following AL+PQ. On day 42, the risk was 18.7% (95% CI 12.2%\u201328.0%) after CQ, 1.2% (95% CI 0.2%\u20138.0%) after CQ+PQ, 29.9% (95% CI 21.6%\u201340.5%) after AL, and 5.9% (95% CI 2.4%\u201313.5%) after AL+PQ . In those not prescribed PQ, the risk of recurrence by day 42 appeared greater following AL treatment than CQ treatment . At the end of follow-up, the incidence rate of P. vivax was 2.2 episodes/person-year for patients treated with CQ compared to 0.4 for patients treated with CQ+PQ  and 2.3 episodes/person-year for AL compared to 0.5 for AL+PQ . There was no difference in the occurrence of adverse events between treatment arms.The risk of recurrent The main limitations of the study were the early termination of the trial and the omission of haemoglobin measurement after day 42, resulting in an inability to estimate the cumulative risk of anaemia.P. vivax, the risk of recurrence in this study was greater following treatment with AL unless it was combined with a supervised course of PQ. PQ combined with either CQ or AL was well tolerated and reduced recurrence of vivax malaria by 5-fold at 1 y.Despite evidence of CQ-resistant NCT01680406ClinicalTrials.gov  Kamala Thriemer and colleagues report on a randomized controlled trial of treatments for Plasmodium vivax malaria. Plasmodium vivax is endemic, recurrent parasitaemia arises from chloroquine (CQ) resistance and the reactivation of dormant hypnozoites. Recurrent parasitaemia is associated with significant morbidity and mortality. Providing safe and effective radical cure of blood and liver stages of the parasite is critical for the control and elimination of P. vivax malaria.In areas where P. vivax clinical trials highlighted a declining efficacy for CQ in many P. vivax endemic areas and marked heterogeneity in study design for determining primaquine (PQ) efficacy. The conclusions of these reviews were that clinical trials to determine the efficacy of radical cure should follow up patients for a prolonged period to capture late relapses and should quantify the incidence rate of multiple recurrences, not just the time to the first recurrence.Two systematic reviews of P. vivax provides a good rationale for artemisinin combination therapy for P. vivax malaria; however, the efficacy of artemisinin combination therapies in combination with PQ radical cure is poorly documented.The rise in CQ-resistant P. vivax malaria in Ethiopia), the second group received CQ plus PQ, the third group received artemether-lumefantrine (AL) alone, and the fourth group received AL plus PQ. All patients were followed up for a year and were treated with the same treatment for every P. vivax malaria episode.We randomized patients to four treatment arms: one group received CQ only (the current recommended treatment for P. vivax infections at day 28 after treatment and also over 12 months. The risk of recurrence by day 28 and 42 was greater following AL than CQ. The addition of PQ to either CQ or AL reduced the risk of recurrence 3-fold by day 42, and 2- to 3-fold over one year.We quantified the risk of P. vivax malaria episodes per year, whereas patients not treated with PQ had on average two episodes per year. The efficacy of PQ treatment for recurrences, which was unsupervised, was 3- to 4-fold lower than that of the initial treatment, which was semi-supervised.Patients treated with PQ had on average only 0.5 In Ethiopia there is evidence of CQ resistance; nevertheless, in this study CQ monotherapy had greater efficacy than AL therapy at day 42.The addition of PQ radical cure to either CQ or AL provided major benefits in reducing subsequent recurrent infection.PQ radical cure should be included in the treatment schedule in Ethiopia and other areas with high relapse risk to reduce relapsing infection and transmission, but further work is needed to improve adherence to the current 14-day regimen to ensure maximum public health impact. Plasmodium vivax infection  < 80 g/l) or history of haemolysis, known hypersensitivity to any of the study drugs, other significant comorbidities, or regular medication that could interfere with their antimalarial treatment. All participants or their guardian/caregiver agreed to finger prick sampling and provided written informed consent.The study was designed as an open-label randomized controlled trial with four arms, conducted at the Bishoftu Malaria Control Center and the Batu Health Center in Oromia Region, in central Ethiopia . Bishoftr survey . PatientPatients were randomized in a two-stage process: (i) at enrolment, randomization to either CQ or AL schizonticidal treatment and (ii) on day 2, after G6PD testing, randomization into the PQ or no-PQ arms. Patients with severe or intermediate G6PD deficiency were excluded from the second randomization to PQ. The randomization sequence was computer-generated by one of the investigators and kept in sealed opaque envelopes. Randomization was done in blocks of eight for each site separately. Clinicians enrolled the patients, and nurses assigned the patients sequentially according to the sealed envelopes.CQ  was prescribed according to national guidelines at 25 mg base/kg over 3 d . AL  was administered twice daily for 3 d according to the manufacturer\u2019s recommendations. The CQ and AL tablets used in the study were certified and provided by the World Health Organization Global Malaria Programme. Initial schizonticidal treatment with CQ was fully supervised, whereas for AL only the morning dose was supervised, and the second dose taken unsupervised at home. PQ  was prescribed at 0.25 mg/kg daily dose over 14 d starting on day 2, and administration was semi-supervised during the initial episode, with directly observed treatment (DOT) on days 2, 3, 7, 10, and 14; doses in between were self-administered. For all subsequent episodes of malaria, schizonticidal as well as PQ treatment was unsupervised.All patients were observed for 60 min after treatment for adverse reactions or vomiting when administered under supervision. Patients vomiting their medication within the first 30 min received a repeat full dose; patients vomiting within 30\u201360 min received half the original dose. Patients failing treatment in any arm within 14 d were treated with quinine 10 mg/kg every 8 h for 7 d as per the national guideline.During screening, a G6PD test was carried out on all patients, and a pregnancy test on all adult females. At enrolment, patients had a medical examination, a questionnaire was completed, and a capillary blood sample was collected for blood film examination and Hb measurement. Three 50-\u03bcl capillary blood spots were stored on filter papers . In consenting patients, venous blood samples were collected at enrolment and, if malaria recurred, on the day of recurrence.P. vivax malaria occurring more than 14 d following treatment received the same treatment as that allocated at enrolment, with subsequent follow-up according to the initial schedule.Patients were asked to return for routine assessment daily for the first 3 d, then weekly until day 42, and thereafter monthly for an additional 10 mo. Patients were also asked to return to the clinic if they had signs or symptoms consistent with malaria, and those with recurrent clinical A follow-up medical exam and blood film examination was undertaken on days 1, 2, 3, 7, 14, 21, 28, 35, and 42, and monthly thereafter for 10 mo. Adverse drug reactions and concomitant medications were recorded at every visit, with repeat Hb concentration measured on days 3, 7, 14, 21, 28, 35, and 42.G6PD status was determined by NADPH fluorescent spot test . Hb concentration was measured using a HemoCue Hb 301 instrument . Blood film examination was undertaken by two certified microscopists, and all slides read independently. Blood smears with discordant results  were re-examined by a third, independent microscopist, and parasite density was calculated by averaging the two closest counts.P. falciparum duplex PET-PCR assay . Th. ThP. viThe study was conducted between 8 November 2012 and 31 December 2014, during which a total of 1,177 patients were screened, and 398 (33.8%) enrolled in the study . In viewThe majority of patients  were enrolled at the Bishoftu Malaria Control Center. There were no important differences in baseline characteristics between the four treatment arms  or in thP. vivax infections documented within 28 d of follow-up, and a further 34 between day 28 and 42   for patients treated with CQ alone compared to 0% (95% CI 0%\u20134.0%) for those treated with CQ+PQ (p < 0.001). The corresponding risks were 12.0% (95% CI 6.8%\u201320.6%) following AL alone and 2.3% (95% CI 0.6%\u20139.0%) following AL+PQ  (There were 17 recurrent +PQ arm) . An addip = 0.005). The corresponding risk for patients in the AL arm was 29.9% (95% CI 21.6%\u201340.5%) compared to 5.9% (95% CI 2.4%\u201313.5%) in the AL+PQ arm  (p = 0.059)  in the CQ arm and 1.2% (95% CI 0.2%\u20138.0%) in the CQ+PQ arm  .P. vivax isolates that could be assessed were homologous (Genotyping was feasible in 90.9% (362/398) of parasite isolates on day 0 and in 90.2% (46/51) of paired isolates from patients with recurrence prior to day 42. In total, 47.8% (22/46) of the paired mologous .Blood chloroquine concentrations on the day of recurrence could be measured in two of the four patients with recurrent infection before day 28 in the CQ arm, both of whom had CQ blood concentrations greater than 100 nM.p < 0.001). By day 2, the prevalence of parasitaemia was 6.8% in the CQ arms and 2.6% in the AL arms. By day 3, only two patients in the CQ arms remained parasitaemic  of those treated with CQ were still parasitaemic compared to 31.8% (61/192) of those treated with AL regimens   were afebrile within 24 h, with 98.8% (84/85) in the AL arms compared to 93.8% (75/80) in the CQ arms (= 0.109) .P. vivax determined by microscopy , and a further eight had had P. falciparum infections (three following CQ and five after CQ+PQ) (P. vivax was 61.7% (95% CI 51.9%\u201371.7%) following CQ alone compared to 72.4% (95% CI 62.5%\u201381.6%) following AL alone (p = 0.127). Compared to CQ or AL alone, the risk of recurrence was significantly lower when treatment was combined with PQ: 20.5% (95% CI 13.0%\u201331.5%) following CQ+PQ  and 22.0% (95% CI 14.2%\u201333.1%) following AL+PQ . There was no difference in the risk of recurrence at the end of the study between patients treated with CQ+PQ and AL+PQ  . The ris 95% CI 1.0%\u201331.5%P. vivax infections occurred later in the PQ arms than the monotherapy arms. The median time of the first recurrence was 82.5 d (interquartile range [IQR] 61.0\u2013186.5) for CQ+PQ compared to 51 d (IQR 42.0\u201386.0) for CQ alone (p = 0.006) and 87 d (IQR 35.0\u2013117.0) for AL+PQ compared to 49.5 d (IQR 35.0\u201398.0) for AL alone (p = 0.040). In the no-PQ arms, 90.8% (108/119) of the recurrent parasitaemias occurred within the first 6 mo.Recurrent P. vivax recurrent infections were detected by microscopy during follow-up   . The incr CQ+PQ) .p = 0.036 and p = 0.059, respectively.The initial treatment with PQ was supervised on scheduled clinic days, and adherence assessed by tablet review at the subsequent visit. The total PQ dose administered per unit body weight was 3.4 mg/kg (IQR 2.8\u20134.0) in the AL+PQ arm and 3.5 mg/kg (IQR 2.6\u20134.0) in the CQ+PQ arm. The histogram of PQ dose distribution shows distinct populations that correspond to patients who received a total PQ dose below 2.6 mg/kg and the rest . There wp = 0.5) or the AL arm . However, patients were at significantly greater risk of recurrent P. vivax after the unsupervised PQ retreatment than after the partially supervised initial PQ treatment: 86.8% (95% CI 77.3%\u201392.5%) versus 63.2% (95% CI 31.7%\u201393.3%) in patients in the CQ+PQ arm  and 84.0% (95% CI 74.0%\u201390.4%) versus 51.9% (95% CI 24.4%\u201373.7%) in patients in the AL+PQ arm   .A total of 358 AEs were reported, but no serious AEs. There was no difference in the occurrence of any of the categories of AEs between treatment arms or by total PQ dosage received. The most frequent AEs were headache (25.5%), fever (18.8%), chills (9.2%), cough (7.0%), vomiting (6.7%), body pain (5.0%), abdominal pain (4.8%), weakness (4.2%) and diarrhoea (4.0%) .p = 0.577). On day 3, 3.6% (7/195) of patients treated with a PQ regimen had anaemia (Hb < 100 g/l) compared to 4.2% (8/191) of those treated with a no-PQ regimen (p = 0.08). By day 28, haematological recovery occurred in 211 (70.8%) patients. Hb concentration did not fall below 80 g/l in any of the patients during the study period, and no patient required blood transfusion.The nadir of Hb concentration occurred on day 3 irrespective of treatment. The mean percentage fall from baseline by day 3 was 6.3% (95% CI 5.0%\u20137.7%) in the CQ arm, 5.7% (95% CI 4.3%\u20137.1%) in the AL arm, 5.9% (95% CI 4.4%\u20137.4%) in the CQ+PQ arm, and 7.6% (95% CI 5.1%\u201310.0%) in the AL+PQ arm  was well tolerated, without significant adverse effects. Recent clinical trials have highlighted the declining efficacy of CQ against P. vivax in Ethiopia [p = 0.059). In equatorial regions endemic for P. vivax, including Ethiopia, the risk of P. vivax relapse is generally high, with the first recurrence occurring about 21 d following the initial treatment [This study highlights that the combination of PQ with either CQ or AL reduced the risk of early recurrence within 42 d) of Ethiopia \u201320, withEthiopia  to 22% [2 d of Etreatment . Lumefanreatment . By day P. vivax and incidence of recurrence over a 12-mo period. The incidence of infection reflects the likely impact that would result from corresponding policy change. Patients receiving CQ or AL alone had four or five recurrences, with an overall incidence rate of approximately two episodes per PYO. The addition of a supervised 14-d regimen of PQ resulted in a 4-fold reduction in the incidence rate, from 2 to 0.5 episodes per PYO. Radical cure with PQ is a function of the total mg/kg dose administered [p < 0.05).The prolonged duration of the study and repeated administration of the same treatment regimen for each episode of malaria allowed quantification of the cumulative risk of nistered ,26, and There was also a 3- to 4-fold greater risk of recurrence following treatment of patients\u2019 first recurrent parasitaemia (which was unsupervised) compared to that following treatment of their initial parasitaemia (which was semi-supervised). Although this non-randomized comparison is vulnerable to bias, there was no difference in the risk of recurrence following the first and second treatments with CQ or AL monotherapy, suggesting that the difference was likely due to poor adherence to an unsupervised 14-d regimen of PQ. These findings are consistent with previous studies that have shown PQ adherence falling to less than 30% when unsupervised \u201329. SignP. vivax endemic countries have implemented this yet. Further work to understand the barriers to routine G6PD testing is currently underway. In our study, PQ was administered to individuals confirmed to be G6PD normal at a total dose of 3.5 mg/kg  [P. vivax recurrences, each of which were associated with repeated bouts of haemolysis and a cumulative risk of anaemia [There is reluctance of some healthcare providers to prescribe PQ, mainly due to concerns over its safety and potential for causing severe haemolysis . To ensuregimen) . This re anaemia .P. vivax infections irrespective of whether they were homologous or heterologous. Third, the early termination of our trial reduced the final sample size. However, loss to follow-up was lower than assumed in the prior power calculation, and the final sample size enrolled into the study still achieved a power of greater than 80% for the primary outcomes of the study.There were a number of limitations of the study. First, the omission of routine Hb measurements after day 42, except in patients with recurrent malaria, prevented quantification of the long-term effects of recurrent episodes of malaria on the cumulative risk of anaemia. Further studies are needed to define the risk and benefits of alternative PQ dosing strategies in different endemic settings. Second, whilst all of the early recurrent parasitaemias were genotyped and compared with the pre-treatment parasitaemia, this comparison was not undertaken for recurrent infections after day 42. Heterologous recurrences occurring within the first 42 d can be due to either reinfection or relapse events but not true recrudescences. Parasite genotyping allows these events to be censored, enabling a better estimate of early efficacy. However, genotyping of late recurrences cannot distinguish between relapsing infections, which can be either homologous or heterologous, and reinfections. For this reason, we used a conservative approach to quantify and compare the cumulative risk and incidence of all recurrent P. vivax in our study, the risk of early recurrence appeared greater following AL, likely due to relapsing infections emerging after the post-treatment prophylaxis of AL had waned. When administered as monotherapy, there was little benefit of changing the first-line therapy for vivax malaria from CQ to AL in this region of Ethiopia. However, PQ treatment should be included in future revision of the national treatment guidelines to decrease relapse rates, with likely benefits in reducing both malarial anaemia and transmission potential. Better methods of ensuring adequate adherence will be needed if the public health impact is to be maximised.In conclusion, although there was evidence of low-grade CQ resistance of S1 Fig(DOCX)Click here for additional data file.S2 Fig(DOCX)Click here for additional data file.S1 PRISMA Checklist(DOC)Click here for additional data file.S1 Text(PDF)Click here for additional data file.S2 Text(PDF)Click here for additional data file.S3 Text(PDF)Click here for additional data file."}
+{"text": "Plasmodium falciparum infections with mutations in the \u2018K13\u2019 gene, with reduced ring-stage susceptibility to artemisinins, and slow parasite clearance in patients treated with ACTs, are now widespread in Southeast Asia. We review clinical efficacy data from the region (2000\u20132015) that provides strong evidence that the loss of first-line ACTs in western Cambodia, first artesunate-mefloquine and then DHA-piperaquine, can be attributed primarily to K13 mutated parasites. The ring-stage activity of artemisinins is therefore critical for the sustained efficacy of ACTs; once it is lost, rapid selection of partner drug resistance and ACT failure are inevitable consequences. Consensus methods for monitoring artemisinin resistance are now available. Despite increased investment in regional control activities, ACTs are failing across an expanding area of the Greater Mekong subregion. Although multiple K13 mutations have arisen independently, successful multidrug-resistant parasite genotypes are taking over and threaten to spread to India and Africa. Stronger containment efforts and new approaches to sustaining long-term efficacy of antimalarial regimens are needed to prevent a global malaria emergency.Artemisinins are the most rapidly acting of currently available antimalarial drugs. Artesunate has become the treatment of choice for severe malaria, and artemisinin-based combination therapies (ACTs) are the foundation of modern falciparum malaria treatment globally. Their safety and tolerability profile is excellent. Unfortunately,  Plasmodium falciparum malaria is causing failure of artemisinin-based combination therapies across an expanding area of Southeast Asia, undermining control and elimination efforts. The potential global consequences can only be avoided by new approaches that ensure sustained efficacy for antimalarial regimens in malaria affected populations.Artemisinin resistance in   Plasmodium falciparum malaria parasites resistant to multiple drugs  were prevalent in Southeast Asia. The benefits of combinations were well established in infectious disease and cancer treatments, and had been used in the treatment of malaria since the discovery of plasmoquine (pamaquine) in the 1920s; chloroquine, amodiaquine, quinine and mefloquine had individually been combined with sulphadoxine-pyrimethamine. However, all had failed rapidly, unsurprisingly given pre-existing resistance to the components. Artemisinin-based combination therapies (ACTs) were introduced in direct response to this deteriorating situation. ACTs provided rapidly and reliably effective, well-tolerated antimalarial regimens with sustained efficacy. By 2006, ACTs were recommended as standard treatment for falciparum malaria worldwide. Their increasing deployment has been one of the main factors behind the reduction in malaria transmission in Asia . Rapid scientific advances in understanding of this problem have taken place within the last five years . Oral artesunate and artemether are active themselves but are also converted by blood esterases and hepatic cytochrome P450 enzymes respectively to DHA, which provides the majority of their activity. Artemisinins are rapidly absorbed and have a short elimination half-life of approximately 1 h, but this is sufficient time for them to exert maximal effects against both asexual stages and immature gametocytes.et\u00a0al.et\u00a0al.et\u00a0al.et\u00a0al.et\u00a0al.et\u00a0al.et\u00a0al.in vitro susceptibility of recrudescent isolates argue against this resulting from inadequate killing of all parasites.Artemisinins have two distinctive pharmacodynamic properties that determine how they are best used. First, they kill both young rings and more mature trophozoites rapidly, an action important for life-saving efficacy in severe disease as well as cure  the day 28 failure rate for artesunate plus mefloquine was 14%, and in a subsequent 2004 study when follow-up was extended to 42 days the failure rate was 21%  medicines. Evidence that et\u00a0al.A subsequent study in southern Cambodia with a 42-day follow-up period also documented unsatisfactory cure rates with artesunate plus mefloquine  , artemisinin resistance may also enhance the natural propensity for parasites to persist via quiescence (dormancy) after artemisinin exposure, allowing subsequent recovery days or weeks later. In the artemisinin-pressured line in which the K13 mutation was first described, as well as culture-adapted field isolates, a higher proportion of ring stages enter developmental arrest and subsequently exit quiescence in response to artemisinins, as well as other drugs  is a specific target of artemisinins and that its levels are increased in parasites with K13 mutations . Importantly, this method accounts for differences in parasitaemias and parasite stage distributions   and requires measurement of parasitaemia every 6 to 8 h in the early phase  drugs, artemisinin monotherapies and failure to administer or complete full treatment courses correctly are issues well known in the region  appears necessary for maximal effect. This dose is associated with faster parasite clearance (by 8.1%) compared to 2 mg\u00a0kg\u22121 in artemisinin-sensitive populations (WWARN Parasite Clearance Study Group \u22121 artemether) provided slightly slower clearance than artesunate in the TRAC study .Even if correctly taken, current ACT regimens may not contain ideal doses, particularly for vulnerable groups such as children and those with hyperparasitaemia   in et\u00a0al.How can resistance be contained in a region where vector control measures are less effective than in other malaria endemic regions and drugs are an essential pillar of malaria control? Controlling malaria by treatment of symptomatic cases alone could distil malaria parasites down to the least drug sensitive\u2014the \u2018last man standing is the most resistant\u2019  who diagnose and treat malaria should be present in every village in malaria endemic areas. These workers are the central pillar of effective malaria control, but coverage with effective and well-supported VHWs is still patchy throughout the region. Once VHWs are in place mass treatment with three monthly rounds of DHA-piperaquine and single low dose primaquine, which appears safe even in G6PD-deficient individuals (Bancone Supplementary DataFEMSRE online.Supplementary data are available at Click here for additional data file."}
+{"text": "P.vivax that could have brought change of treatment regimen is yet to be established in Ethiopia, continuous and regular monitoring of drug\u2019s efficacy is critical for establishing rational anti-malarial drug policies. This study therefore, assessed the therapeutic efficacy of Chloroquine (CQ) for the treatment of Plasmodium vivax infections in Northwestern Ethiopia.Resistance to anti-malarials is a major challenge for effective malaria control in sub-Saharan Africa. This triggered a need for routine monitoring of the efficacy of the antimalarial drugs every two years in all malaria endemic countries. Chloroquine remained the drug of choice for the treatment of vivax malaria in Ethiopia. Though, a strong scientific evidence of chloroquine resistance to An observational, 28- day therapeutic clinical efficacy study was conducted from August to December, 2014, in Northwest Ethiopia. Patients confirmed to have monoinfection of vivax malaria, aged above 6 months were included. All subjects were treated with standard chloroquine dose of 25 mg/kg for three (3) days. Parasitological and clinical outcomes of treated patients were then evaluated on days 1, 2, 3, 7, 14, 21, and 28 during the entire 28-day follow-up period. A portable spectrophotometer  was used to estimate hemoglobin concentration.A total of 69 subjects had completed follow up. Some 57/69 (82.6%) had fever at enrolment and the rest 12 patients 48 hours before enrollment. Out of total, 65/69 (94.2%) and 66/69 (95.6%) of the study subjects were free of fever by day 1 and day 2 respectively but fever was cleared in all subjects by day 3. At base line the mean asexual parasitemia was 3540 parasites/\u03bcL of blood. Parasite carriage on day 3 was 3%. The overall cure rate  was very high (97%) [(95% CI = 93.1\u201399.4)]. The time to parasite, fever and gametocyte clearance as expressed in mean (SD) was 35 (3), 25 (4.6), 28 (3.2) hours respectively. Mean hemoglobin was significantly increased (P<0.001) from 12.2 (7\u201315) g/dl at day 0 to 13.3 (10\u201316) g/dl on day 28.In view of our findings, CQ remains efficacious for the treatment of vivax malaria in the study area. However, there is a need to monitor CQR regularly using molecular and or biochemical tools for better evaluation of treatment outcomes. Plasmodium vivax is the most widely distributed species responsible for 25\u201340% of malaria cases worldwide  and abdominal discomfort [4/69 (7.2%)] occurred most frequently. Nausea, dizziness, vomiting and diarrhea was reported in 4/69 (5.8%), 3 (4.3%), 2 (2.9%) and 2 (2.9%) of patients respectively. No adverse effect/s other than those registered in FDA and or national malaria treatments were reported in this study. Adverse effects were shown to occur more frequently among children aged between 1 and 4 years (p = 0.022).P.vivax mono-infection in Ethiopia, despite the growing concerns of resistance pattern [One of the strategies to control malaria is timely provision of effective anti-malarials to infected individuals. Therefore, monitoring the dynamics of anti-malarial drug resistance could help detect emerging resistant strains early. This prospective, observational follow up study assessed the therapeutic efficacy of CQ, which remained a drug of choice for the treatment of  pattern ,34, 41. P.vivax. Given, the emergence of chloroquine-resistant vivax malaria can occur at any time in different geographical locations depending on the type of parasite genotypes circulating in the population, it is recommended to conduct similar therapeutic efficacy studies to adequately address the problem of drug resistance.Few studies conducted in Ethiopia so far indicated an alarming levels of CQR . HoweverP.vivax mono infected subjects were higher compared to other findings in different parts of Ethiopia [In this study, the proportion of Ethiopia , 43. MorEthiopia ,43,44. TP.vivax as it is in line with WHO recommendation [Of 69 participants who completed a 28 day follow up in current CQ efficacy study, 2 (2.9%) were labeled as risks of treatment failure and 97.1% ACPR. From the two treatment failures one was a LPF which had occurred on day 28 of follow up period and the remaining one was a LCF that occurred on day 21. Both treatment failures were seen in patients less than 15 years old. Similar scenarios have been reported elsewhere ,41,45. Gendation and Debreendation  respectiP.vivax genetic polymorphisms.Similarly a 96.4% cure rate was reported in Serbo (Sothwest Ethiopia) by 2009 . But a sThe current study also showed a fast clearance rate of both parasitemia and fever following CQ mono therapy. Some 98.5% showed no asexual parasitemia on day 3, and only one patient was parasitemic on day 21, and one on day 28 though the density of parasite was minimal. In other studies conducted in Ethiopia ,34, the P.falciparum infections [P.vivax with CQ [In the present study, both mild and moderate types of anemia were relatively prevalent in the pre-treatment phase. However, none of the study subjects with ACPR showed moderate anemia at the end of follow up and a significant reduction in mild anemia was also recorded following therapy. Thus the proportion of patients with anemia (both mild and moderate) was reduced from 46.5% to 14.5% following CQ therapy. Improvement in hemoglobin levels after treatment with anti malarial drugs and successful parasite clearance were reported following treatment of uncomplicated fections ,44 with  with CQ ,42. It i with CQ . Indeed  with CQ .In the current study, a three-dose regimen of CQ was safe and well tolerated anti- malarial drug. Some (20%) of subjects showed common adverse effects of CQ and more than one adverse effect per individuals were rarely reported. Weakness and abdominal discomfort were the adverse effects that occurred frequently. The observed common adverse effects occurred more frequently in less than five years old children. The adverse effects that occurred among the study participants were not different from those acknowledged and registered under FDA and national malaria treatment guide of Ethiopia.This study has got its own strengths and limitations. To the knowledge of authors, it is among few studies of its kind in Ethiopia that would provide useful information on the efficacy of CQ for policy makers. Indeed, abandonment of a given drug is recommended when 10% of infections are not responding to treatment. Moreover, it is relatively cost effectiveness study design particularly in resource constraint settings to monitor the risk of treatment failure regularly. In this follow up study, patients were given directly observed treatment by investigators under supervision and none of them had recurrent vomiting. These conditions are likely to reduce the risk of treatment failure attributable to poor dosing. However, to mention the limitations, we did not determine drug resistance phenotype; neither undertook molecular characterization to determine gene polymorphism, and measured plasma drug concentrations.In view of our findings, the risk of treatment failure to three-dose regimen of CQ therapy for vivax malaria is low. A significant improvement in clinical and parasitologic parameters was as well as minimal adverse events were recorded. However, given conflicting reports indicating alarming levels of treatment failures from other sites, there remains a need to monitor the emergence of chloroquine-resistant vivax malaria across the nation to obtain an adequate representation in different ecological and epidemiological settings.S1 File(DOC)Click here for additional data file.S2 File(DOC)Click here for additional data file.S3 File(DOC)Click here for additional data file."}
+{"text": "Plasmodium are more likely to occur in humans infected in these areas. In many mixed infections, parasite densities of the cryptic species may be low and often not recognized in clinical practice.Since several malaria parasite species are usually present in a particular area, co-infections with more than one species of Plasmodium falciparum and Plasmodium malariae co-infection for both children at admission. They were both treated with atovaquone-proguanil combination for 3\u00a0days. At day 7, both thin blood smears examination remained negative but at day 28, thin blood smear was positive for P. malariae trophozoites and for Plasmodium ovale for the girl and her brother, respectively. Samples collected at day 1 and day 28 were submitted to real-time PCR showing the presence of the three parasite species  in admission blood samples from the two children and only P. ovale at day 28.Two children (3 and 6\u00a0years old) adopted recently from Central African Republic were admitted to hospital because of intermittent fever. Thin blood smears stained with Giemsa showed P. falciparum tending to dominate other species. These observations provide more evidence that recommendations for treatment of imported malaria should take into account the risk of concurrent or cryptic infection with Plasmodium species. Clinicians and biologists should be aware of the underestimated frequency of mixed infections with cryptic species and of the importance of patient follow-up at day 28. Future guidelines should shed more light on the treatment of mixed infection and on the interest of using artemisinin-based combinations for falciparum and non-falciparum species.Twenty-eight days follow-up after treatment led to detection of a third parasite species in the blood of these two patients suggesting covert co-infection and a delayed appearance of one cryptic species following treatment. Concurrently infecting malaria species could be mutually suppressive, with  Plasmodium. However, diagnostic of these mixed-infections is rare in clinical practice, and probably underestimated by microscopy. Expert microscopists are uncommon in clinical laboratory, and distinguishing the species from young trophozoite forms of Plasmodium parasites is sometimes difficult [P. falciparum is obviously taking most of the attention considering the potential consequences of a wrong diagnosis, and other species are sometimes scanned with less intensity. In case of mixed infection, the concurrent species is mostly below the threshold of microscopy. For example, the ratio of Plasmodium malariae/P. falciparum was described as extremely low in a study conducted in Ivory Coast and the protective effect of lengthy subclinical P. malariae infections was suspected [Plasmodium vivax and Plasmodium ovale, which are responsible for relapses, are not detectable [P. falciparum and P. vivax malaria [Mixed-species malaria infections have received sporadic attention , and theifficult . The lifuspected . Moreovetectable . The del malaria .P. malariae when inoculated with P. vivax in an attempt to treat neurosyphilis [It has been known from decades that antagonism between species may lead to a rapid decrease of circulating parasites, such as syphilis .P. falciparum and P. malariae, followed by P. ovale delayed infection) in two adopted children from Central African Republic are presented here to document the risk of late reappearance and the need for clinical and parasitological follow-up. These cases focused more light on the risk of treating each malaria species by drugs with different efficacy.Triple species infection are known, but remain rare in the literature. Two parallel cases of triple infections (Two children (3 and 6\u00a0years old) were admitted to hospital because of intermittent fever and suspected malaria. They have been adopted recently and came back from Central African Republic a week previously.The 6-year-old boy presented with a temperature of 37.9\u00a0\u00b0C, a systolic murmur in mitral focus, and a hepatosplenomegaly. The haemoglobin level was 6.7\u00a0g/dL, the mean corpuscular volume was 65 fL, and the platelet count was 216 G/L. Biochemical tests showed a normal liver function, increased serum levels of lactate dehydrogenase  and C-reactive protein . A coagulation test result for the prothrombin time\u2013international normalized ratio was slightly prolonged . An ultrasonographic abdominal examination showed hepatomegaly , and splenomegaly , and lack of deep lymph nodes. An ultrasonographic heart examination reported no abnormalities.The 3-year-old girl, presented with a temperature of 37\u00a0\u00b0C, bilateral crackles predominant right, and a hepatosplenomegaly. The haemoglobin level was 6.3\u00a0g/dL, the mean corpuscular volume was 71.6 fL, and the platelet count was 138 g/L. Biochemical tests showed a normal liver function, increased serum levels of lactate dehydrogenase (379 U/L) and C-reactive protein (48\u00a0mg/mL). A coagulation test result for the prothrombin time\u2013international normalized ratio was slightly normal. Ultrasonographic abdominal examination reported a splenomegaly . A rapid diagnostic test  showed three positive bands for the control, Plasmodium spp. (pLDH), and P. falciparum (pHRPII) markers.Thin blood smears stained with Giemsa showed Both children received 1 adult co-formulated tablet of Malarone, containing 250\u00a0mg atovaquone and 100\u00a0mg proguanil hydrochloride, per day for 3\u00a0days. The tablets were administered during the meal without addition of fat. The boy weighed 19\u00a0kg, the girl weighed 14.5\u00a0kg and the administered weight-based dose of atovaquone/proguanil was 10/5 and 17/7\u00a0mg/kg/day for the boy and girl respectively. They did not vomit and no other adverse events were noted during treatment. Both children received iron syrup supplementation and paracetamol.P. falciparum\u00a0<0.01%\u00a0+\u00a0P. malariae\u00a0<0.01% and P. falciparum 0%\u00a0+\u00a0P. malariae 0.02%, for girl and boy respectively). These figures are not uncommon because atovaquone-proguanil is known to act relatively slowly. Apyrexia was obtained at day 2 and patients were discharged on day 5 and did not return until scheduled appointments for control of parasitaemia on day 7.Almost complete parasitaemia clearance was obtained at day 3  was detected from her brother\u2019s blood. To confirm the species identification, the samples collected at day 1 and day 28 were submitted to real-time PCR as previously described [P. falciparum, P. malariae and P. ovale) in D0 samples from both children, and only P. ovale at day 28.At day 7, both thin blood smears examination remained negative, while few picnotic parasites were detected on thick blood smears. Surprisingly, at day 28, thin blood smear from the girl was positive and revealed\u00a0<0.01% escribed . MoleculP. ovale, patients were given a total dose of 25\u00a0mg/kg of chloroquine over 3\u00a0days. Both children were not glucose-6-phosphate dehydrogenase-deficient, but primaquine was not used to prevent relapses. Clinical and parasitological follow-up was performed and total parasite clearance was assessed by thin and thick blood smears at day 42, showing the absence of recurrence. Both children had non-haemolytic , microcytic anaemia at admission to hospital and were found to have iron deficiency, which was successfully treated after 3\u00a0months of iron supplementation. Echocardiography examinations revealed that boy had an inorganic heart murmur. Gastrointestinal helminths were not found after 3 stool examinations for ova and parasites. Cysts of Entamoeba coli and Chilomastix mesnili were found in the boy\u2019s stools, who was treated with secnidazole. Patients were then lost to follow-up.Both children were still apyretic and didn\u2019t show any symptoms of infection. Considering the suspected parasitological failure of the atovaquone-proguanil treatment against P. falciparum, P. vivax, P. malariae, P. ovale wallikeri, P. ovale curtisi and P. knowlesi [P. ovale wallikeri is not distinguishable from P. ovale curtisi, and P. knowlesi is impossible to distinguish from P. malariae. Most of the rapid diagnostic tests available on the market showed lower performance for P. vivax (70.7% panel detection score) than for P. falciparum (85% panel detection score) in the round 6 of WHO [Plasmodium ovale and P. malariae RDTs have not been evaluated so far. Molecular tests are the only methods allowing species identification with a good accuracy.Six different malaria species are known to infect humans: knowlesi . RoutineAnopheles species, are able to carry more than one species, including the four main species for Anopheles gambiae [Plasmodium vivax is supposed to reduce the severity of P. falciparum infection; P. falciparum reduces the availability of red blood cells for P. vivax infection, and a theory of species-transcendenting density-dependant (STDD) regulation of multiple malaria infection has been proposed [Mixed, or concurrent infections, are frequently reported in epidemiological studies conducted in endemic areas and probably underdiagnosed at an individual level. It is known that several  gambiae . There iproposed , 11.falciparum malaria, and chloroquine to treat non-falciparum malaria, under-diagnosis of mixed infection may lead to inappropriate treatment of patients. In countries with chloroquine-resistant P. vivax, ACT is the treatment policy for non-falciparum infections. ACT is also recommended treatment of mixed infections [P. falciparum and P. malariae co-infection was successfully treated with atovaquone-proguanil, while asymptomatic P. ovale was detected at day 28. Unfortunately, it was not possible to know if the recurrence of P. ovale was due to drug resistance or relapse of liver stages, and cytochrome b gene mutation was not investigated due to low amount of samples. A plausible cause of this late therapeutic failure is the relatively insufficient dosage due to increased oral clearance of atovaquone in pediatric patients. However, failure to atovaquone-proguanil prophylaxis has been reported in travelers presenting P. ovale infection [There is no evidence that drugs used to treat the dominant specie may be ineffective against the cryptic species. Since ACTs are mainly recommended to treat fections . In the nfection .Plasmodium species presenting different sensitivity to drugs. Considering that information on drug resistance of non-falciparum species is scarce, clinician and biologists should take particular attention to patient follow-up at day 28.These observations provide more evidence that recommendations for treatment of imported malaria may take into account the risk of concurrent or cryptic infection with"}
+{"text": "Recent anti-malarial resistance monitoring in Angola has shown efficacy of artemether\u2013lumefantrine (AL) in certain sites approaching the key 90% lower limit of efficacy recommended for artemisinin-based combination therapy. In addition, a controversial case of malaria unresponsive to artemisinins was reported in a patient infected in Lunda Sul Province in 2013.Plasmodium falciparum infection treated with AL, artesunate\u2013amodiaquine (ASAQ), or dihydroartemisinin\u2013piperaquine (DP). The study comprised two treatment arms in each of three provinces: Benguela , Zaire , and Lunda Sul . Samples from treatment failures were analysed for molecular markers of resistance for artemisinin (K13) and lumefantrine (pfmdr1).During January\u2013June 2015, investigators monitored the clinical and parasitological response of children with uncomplicated pfmdr1 haplotypes associated with decreased lumefantrine susceptibility.A total of 467 children reached a study endpoint. Fifty-four treatment failures were observed: four early treatment failures, 40 re-infections and ten recrudescences. Excluding re-infections, the 28-day microsatellite-corrected efficacy was 96.3% (95% CI 91\u2013100) for AL in Benguela, 99.9% (95\u2013100) for ASAQ in Benguela, 88.1% (81\u201395) for AL in Zaire, and 100% for ASAQ in Lunda Sul. For DP, the 42-day corrected efficacy was 98.8% (96\u2013100) in Zaire and 100% in Lunda Sul. All treatment failures were wild type for K13, but all AL treatment failures had No evidence was found to corroborate the specific allegation of artemisinin resistance in Lunda Sul. The efficacy below 90% of AL in Zaire matches findings from 2013 from the same site. Further monitoring, particularly including measurement of lumefantrine blood levels, is recommended.The online version of this article (doi:10.1186/s12936-017-1712-4) contains supplementary material, which is available to authorized users. Ensuring that current first- and second-line anti-malarials remain efficacious is a key component of the international effort to reduce malaria morbidity and mortality. Following World Health Organization (WHO) guidelines , AngolanIn recent years, strong evidence has accumulated that malaria parasites resistant to artemisinins have emerged and are spreading in certain parts of Southeast Asia , 3. MoniCurrently there is no evidence that artemisinin-resistant parasites have appeared in sub-Saharan Africa, where the majority of global malaria morbidity and mortality is concentrated , 6. No ppfmdr1 genotypes previously associated with decreased lumefantrine susceptibility were reported in both studies.Angola, malaria endemic in its entirety, has occupied a prominent place in recent anti-malarial resistance monitoring in Africa . Its larHere, the results of the 2015 round of biennial therapeutic efficacy monitoring in Angola are presented from three provinces selected based on past efficacy results: Benguela, Zaire and Lunda Sul.Data were collected as part of a six-arm clinical outcomes trial following the standard WHO in vivo therapeutic efficacy protocol . In eachPlasmodium falciparum infection presenting to the sentinel health facilities and meeting standard inclusion criteria [Children with symptomatic, uncomplicated criteria  were invThe target sample enrolment was 100 children per arm, providing enough power to estimate efficacy with 95% confidence limits of\u00a0\u00b15% assuming an expected efficacy of 95% and a maximum loss to follow-up and exclusion rate of 27%.\u00ae; Novartis, Basel, Switzerland), ASAQ , and DP . Study participants in the AL arm in Benguela received the regular formulation of Coartem, whereas those in the AL arm in Zaire received the dispersible formulation. AL was procured by The President\u2019s Malaria Initiative and provided by the Angolan National Malaria Control Programme, and the ASAQ and DP were procured and provided by WHO. All medications passed lot-quality testing, performed in laboratories in the USA (AL), the Netherlands (DP) and South Africa (ASAQ), prior to the study. The participants were treated according to manufacturers\u2019 weight-based dosing recommendations. All three doses of ASAQ and DP were directly observed in the study health facilities. For AL, the morning doses were directly observed at the study health facilities, while the evening doses were given by participants\u2019 guardians. To ensure appropriate administration of evening doses, guardians were counselled at the health facility and reminded in the evening by telephone. Guardians were also instructed to bring back empty blister packs the following day. Study participants not correctly completing the medication were withdrawn from the study.Study participants were administered one of three anti-malarials: AL  or 42\u00a0days (DP arms). At each follow-up visit, clinical response to treatment was monitored through a standardized history and physical examination and parasitological response was assessed through preparation and examination of thick blood films (with the exception of day 1). Guardians were asked about any adverse events experienced by the study participants. Blood slides were prepared and read by two dedicated study laboratory technicians following standard procedures , and a sDried blood spots were transported to CDC laboratories in Atlanta. DNA was extracted using a QIAamp Blood DNA kit  from day 0 and day-of-failure samples for all treatment failures and day-0 samples from 50 randomly selected participants with adequate clinical and parasitological response (ACPR).For all treatment failure samples and the randomly selected treatment successes, the lengths of six microsatellites  were meaP. falciparum K13, P. falciparum multidrug-resistance gene 1 (pfmdr1), and P. falciparum cytochrome b (pfcytb) genes were amplified and sequenced for day 0 and day of failure for all treatment failures. The P. falciparum K13 propeller domain was amplified using a P. falciparum-specific protocol described previously [pfmdr1, covering codons 86\u2013184 and 1034\u20131246, were amplified using a previously described protocol [pfcytb gene was amplified using the following primers: forward primer, 5\u2032 CTATTAATTTAGTTAAAGCACAC 3\u2032 and reverse primer, 5\u2032 ACAGAATAATCTCTAGCACCA 3\u2032. 1 \u00b5L of genomic DNA was amplified using 0.5\u00a0\u00b5M of each primer, 0.2\u00a0mM dNTP, 2\u00a0mM MgCl2 and 1 U High Fidelity Taq . For the primary reaction, the following cycling parameters were used: 2\u00a0min at 98\u00a0\u00b0C, 35 cycles of 98\u00a0\u00b0C for 10\u00a0s, 46\u00a0\u00b0C for 30\u00a0s, 68\u00a0\u00b0C for 2:30\u00a0min, and final extension for 10\u00a0min at 68\u00a0\u00b0C. Amplified PCR products were visualized on a 2% agarose gel after electrophoresis. Sequencing of PCR products was performed using an ABI 3730 sequencer .Regions in the eviously . Two regprotocol . The pfcSanger sequences were analysed using Geneious R8 software  to identify specific single-nucleotide polymorphism combinations. Single-nucleotide polymorphisms were only called if both the forward and reverse strands had the mutation.Study participants not meeting any exclusion criteria were classified as one of three possible study endpoints. Study participants showing signs of severe disease while parasitaemic in the first 3 days following treatment, having a parasite density on day 2 that exceeded day 0 parasite density, clearing less than 75% of day 0 parasites on day 3, or still febrile (\u226537.5\u00a0\u00b0C axillary) and parasitaemic on day 3 were classified as early treatment failures (ETFs). Study participants that were febrile and parasitaemic after day 3 or parasitaemic regardless of body temperature after day 6 were classified as late treatment failures (LTFs). Participants not meeting these criteria with a negative slide on the last day of follow-up were classified as having an ACPR.Kaplan\u2013Meier estimates of the survival curve for each arm were calculated and plotted. Uncorrected efficacy was calculated as the proportion ACPR (per-protocol method), as well as the Kaplan\u2013Meier estimate of the survival function at day 28 and/or 42. Microsatellite data for LTFs were analysed using a previously published statistical algorithm which assigns each LTF a posterior probability of recrudescence . LTFs wiParents or guardians of all study participants provided written informed consent prior to enrolment and were reimbursed for transport costs for clinic visits. The activity was classified as non-research by human subjects research boards at CDC and the Angolan Ministry of Health and was approved by the WHO Ethical Research Committee.Plasmodium vivax infection during follow-up, and one participant in the DP arm in Lunda Sul that developed severe malaria and died fewer than 24\u00a0h after enrolment. The minimal sample size of 73 study participants reaching a study endpoint was achieved in all arms except for the ASAQ arm in Lunda Sul, where only 56 participants finished 28-day follow-up. Participant characteristics at baseline reflected the different inclusion criteria in each site  were enrolled  of participants taking ASAQ, 7.3% (15/205) of participants taking AL, and 8.4% (17/202) of participants taking DP. One participant in the DP arm in Zaire experienced seizures on day 12 of follow-up, was negative for malaria by microscopy, and was excluded. No other adverse events were reported.Clearance of parasites in all six arms was rapid, with clearance rates in each arm ranging from 94 to 100% on day 2, increasing to 99 to 100% on day 3  LTFs having a probability of recrudescence either\u00a0>0.9 or\u00a0<0.1, signifying high confidence  and NFD  were the most common haplotypes, found in 61 (59%) and 33 (32%) of tested samples, respectively. In the AL arms, the recrudescent and incident re-infections were all either NYD  or NFD .All treatment failures were wild type for K13 and tb Table\u00a0. The pfmThese therapeutic efficacy monitoring results from the three provinces demonstrate that the artemisinin components of the three ACT in use in Angola are still highly effective against falciparum malaria, while there is some variation between the efficacies of the partner drugs by site. Although the three sentinel sites are dispersed throughout the country Fig.\u00a0, the effNo K13 mutations were detected in any of the treatment failures. While there is no guarantee that artemisinin resistance in sub-Saharan Africa will necessarily be mediated by K13 mutations , the absAlthough the ASAQ arm in Lunda Sul did not meet its target sample size, the high efficacy of both DP and ASAQ in Lunda Sul, the absence of K13 mutants, and high clearance rates provide no evidence that a focus of artemisinin resistance has been established in Lunda Sul.In general, all three of the partner drugs to the artemisinin derivatives were efficacious in sufficiently suppressing the initial malaria infection. No recrudescences were observed in either the Benguela or the Lunda Sul ASAQ arm. A total of 15 LTFs were observed in the DP arms, with all of these occurring in the Zaire arm. The 42-day uncorrected efficacy of DP in Zaire was estimated to be 82.5%, which is lower than expected, as 42-day uncorrected efficacies of DP are usually reported to be closer to 90% , a resulThe AL arms recorded a total of 38 LTFs, despite only a 28-day follow-up duration, resulting in 28-day uncorrected efficacies below 90% in both Benguela and Zaire. This was likely partly due to lumefantrine having a significantly shorter half-life than piperaquine . The lonAmongst the 38 LTFs observed in the AL arms, nine were classified as recrudescent infections. While only one of the LTFs in Benguela was a recrudescence of the original infection, there were eight recrudescences noted in the AL arm in Zaire. Combined with three ETFs, this resulted in a corrected efficacy, using the Kaplan\u2013Meier estimator, of 88.1% in Zaire, close to the 89.6% estimate reported in 2013 . It is apfmdr1 NYD and NFD haplotypes in AL recrudescent and incident infections during follow-up closely matches data from the 2013 therapeutic efficacy monitoring in Zaire and U\u00edge Provinces, where 91% of AL treatment failures had NYD or NFD haplotypes on day of failure. This is consistent with evidence that NYD and NFD haplotypes are associated with decreased susceptibility to lumefantrine and are under selection following AL treatment [The preponderance of reatment , 28, 29.The fact that lumefantrine drug levels were not measured and that the evening AL doses were not directly observed precludes the exclusion of underdosing or malabsorption as explanations for the decreased efficacy of AL in Zaire Province. Future therapeutic efficacy monitoring should incorporate direct observation of all doses of AL, including strict adherence to food intake during administration of AL, and measurement of day 7 lumefantrine levels to definitively rule out the alternative hypothesis of underdosing or malabsorption.The sequential, non-randomized enrolment of study participants meant that participants in the different arms were exposed to different risk of re-infection as the follow-up period in the second arm in each province took place later in the peak transmission season. As a result, direct comparison of drug efficacies is difficult across study arms.P. falciparum parasites to artemisinin derivatives, points to high efficacy of amodiaquine and piperaquine as partner drugs, and reaffirms a low efficacy of lumefantrine in Zaire Province in northern Angola. Further routine efficacy monitoring, including direct observation of all doses and blood drug level measurement, is recommended.Overall, the 2015 round of efficacy monitoring in Angola confirms the continued sensitivity of"}
+{"text": "Malaria has declined dramatically along the Thai\u2013Myanmar border in recent years due to malaria control and elimination programmes. However, at the same time, artemisinin resistance has spread, raising concerns about the efficacy of parenteral artesunate for the treatment of severe malaria.In November 2015 and April 2017, two patients were treated for severe malaria with parenteral artesunate. Quinine was added within 24\u00a0h due to an initial poor response to treatment. The first patient died within 24\u00a0h of starting treatment and the second did not clear his peripheral parasitaemia until 11\u00a0days later. Genotyping revealed artemisinin resistance Kelch-13 markers.Reliable efficacy of artesunate for the treatment of severe malaria may no longer be assured in areas where artemisinin resistance has emerged. Empirical addition of parenteral quinine to artesunate for treatment is recommended as a precautionary measure.The online version of this article (10.1186/s12936-018-2182-z) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria   was 91.8 (2.5\u20137.1\u00a0mmol/L), creatinine 2.5 (0.67\u20131.17\u00a0mg/dL), potassium 4.47 (3.5\u20135.1\u00a0mmol/L), sodium 135.8 (136\u2013145\u00a0mmol/L), bicarbonate 4.9 (23\u201329\u00a0mmol/L), blood glucose was 74\u00a0mg/dL and Hct dropped to 22%, so 350\u00a0mL whole blood was transfused. His Glasgow Coma Score (GCS) was 15/15; he was eating and walking unsupported, had a respiratory rate of 40/min, clear lungs and a temperature of 36.9\u00a0\u00b0C, blood pressure of 90/40\u00a0mmHg. His asexual parasitaemia was now 27% of iRBC or 746,064\u00a0\u00b5L (>\u00a080% of early ring stage), gametocytaemia 1824\u00a0\u00b5L, schizontaemia 32\u00a0\u00b5L. A loading dose (20\u00a0mg/kg) of quinine 4-hourly infusion was added. Four hours after the start of the loading dose quinine infusion (quinine in 500\u00a0mL of dextrose 10%) the patient\u2019s condition was unchanged and his asexual parasitaemia was now 26% iRBC or 718,432\u00a0\u00b5L (>\u00a080% early ring stage), gametocytaemia 1920\u00a0\u00b5L, schizontaemia 16\u00a0\u00b5L . HIV rapid test was positive, which was confirmed subsequently with a baseline CD4 count of 39\u00a0cells/mL. The patient\u2019s GCS was 8/15, tympanic temperature 39.0\u00a0\u00b0C, blood glucose 51\u00a0mg/dL, and blood pressure 120/80\u00a0mmHg. His liver was palpable, 3\u00a0cm below the costal margin and his spleen was not palpable, although confirmed to be present on abdominal ultrasonography. Intravenous artesunate (2.4\u00a0mg/kg) was given on admission, after 12\u00a0h and again after 24\u00a0h as per protocol [On 3 April, 2017, an 18\u00a0years old male was admitted with cerebral malaria: After 72\u00a0h, the patient recovered consciousness and started eating. However, the parasitaemia had increased again Fig.\u00a0. AtovaquThe anti-malarials used were artemether-lumefantrine  and artesunate injection (Atlantic pharma: LA170333) provided by the Global Fund malaria programme. Quinine injection (A.N.B laboratories Co. Ltd: 555009) and atovaquone\u2013proguanil  were purchased from the supplier. All the drugs were in date.All blood slides were double-checked by an expert microscopist of Shoklo Malaria Research Unit.Leftover blood from the patients was sent for parasite Kelch-13 genotyping with patient or relative written consent. Both had kelch mutant parasites: patient 1 with the P441L mutation and patient 2 the C580Y mutation. Microsatellite genotyping of flanking regions of C580Y haplotype of the Kelch-13 gene revealed a different C580Y lineage from those described recently in Western Cambodia/Northeastern Thailand/Southern Laos/Myanmar, suggesting a different parental origin .In Case 1, artesunate or DHA were not detected which was not unexpected since the blood sampling time was\u00a0~\u00a024\u00a0h after the second (last) dose of artemether-lumefantrine. However, lumefantrine was also undetectable, which could be due to poor absorption of drug since the tablets were taken with water  or non-aIn Case 2, the plasma concentration of artesunate after\u00a0~\u00a02\u00a0h of intravenous injection was 105\u00a0ng/mL and DHA was 1530\u00a0ng/mL, which were well above the plasma levels providing maximum parasiticidal effect .Parasite isolates taken from patient 2, after two doses of intravenous artesunate injection (~\u00a02\u00a0h of second dose) grew normally in drug free plate  was performed on the parasites obtained from these two patients . This advantage is severely compromised by artemisinin resistance with implications for the treatment of severe malaria. A previous study of 69 patients conducted in 1998  showed nIn the absence of conclusive evidence, to minimize the risk to individual patients, it is recommended that all patients treated for severe malaria acquired in areas where artemisinin resistance is established should receive immediate empirical parenteral artesunate and quinine with frequent monitoring of parasite density.Additional file 1. Parasite isolates collected after two doses of parenteral artesunate growing in drug-free plate.Additional file 2. Methods and other table."}
+{"text": "Malaria endemic countries need to assess efficacy of anti-malarial treatments on a regular basis. Moreover, resistance to artemisinin that is established across mainland South-East Asia represents today a major threat to global health. Monitoring the efficacy of artemisinin-based combination therapies is of paramount importance to detect as early as possible the emergence of resistance in African countries that toll the highest burden of malaria morbidity and mortality.\u00ae), dihydroartemisinin\u2013piperaquine  and artemether\u2013lumefantrine  taken under supervision and respecting pharmaceutical recommendations. The study enrolled for each treatment arm 212 children aged 6\u201359\u00a0months living in Maradi (Niger) and suffering with uncomplicated falciparum malaria. The Kaplan\u2013Meier 42-day PCR-adjusted cure rate was the primary outcome. A standardized parasite clearance estimator was used to assess delayed parasite clearance as surrogate maker of suspected artemisinin resistance.A WHO standard protocol was used to assess efficacy of the combinations artesunate\u2013amodiaquine  was higher in the AM\u2013LM arm (32.4%) than in the AS\u2013AQ (13.8%) and the DHA\u2013PPQ arm (24.9%). The parasite clearance rate constant was 0.27, 0.26 and 0.25 per hour for AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM, respectively.All the three treatments evaluated largely meet WHO criteria (at least 95% efficacy). AS\u2013AQ and AL\u2013LM may continue to be used and DHA\u2013PPQ may be also recommended as first-line treatment for uncomplicated falciparum malaria in Maradi. The parasite clearance rate were consistent with reference values indicating no suspected artemisinin resistance. Nevertheless, the monitoring of anti-malarial drug efficacy should continue.Trial registration details Registry number at ClinicalTrial.gov: NCT01755559 Malaria is a leading cause of morbidity and mortality in Niger, with 5.2 million cases and 10 thousands deaths being estimated in 2015 , and 94.Plasmodium falciparum kelch protein gene (Pfk13) [Pfk13, although none of them were among the mutations associated with artemisinin resistance in South-East Asia [Resistance to artemisinin emerged in Thailand\u2013Cambodia border in the early 2000 \u20137. Resis (Pfk13) , 11. A rast Asia .In the effort to monitor the efficacy of anti-malarial treatments in use, a three-arm efficacy study was carried out in Maradi, in the south of Niger. AM\u2013LM and AS\u2013AQ were evaluated, as well as dihydroartemisinin\u2013piperaquine (DHA\u2013PPQ). This latter treatment has proven to be highly efficacious in previous studies \u201315 and rP. falciparum mono-infection confirmed with microscopy. Eligibility for inclusion and clinical and parasitological evaluations were according to WHO standardized protocol for monitoring anti-malarial drug efficacy [This was an in vivo study in children 6\u201359\u00a0months with a efficacy . In addiefficacy  to monitWith an expected efficacy of 95%, an accuracy of 3%, the sample size was estimated to be 184 children per arm. This sample size allowed to test the hypothesis that the expected efficacy of 95% was not statistically equal or inferior to 90%, with an alpha error of 0.05 and a power of 80%. Sample size was increased of 20% to take into account the occurrence of re-infections, indeterminate PCR results and loss to follow-up. The final sample size was 221 children per arm, and therefore 663 children in total.\u00ae Sanofi Aventis), dihydroartemisinin\u2013piperaquine  or artemether\u2013lumefantrine . All treatments were given under supervision of a nurse and respecting pharmaceutical recommendations . The allocation of patients to one of the three treatments was randomized. This procedure allowed carrying out inclusions in parallel, and provided a better representativeness of the study population throughout the study period for each of the study treatment.Children meeting inclusion criteria were enrolled and treated on-site with a 3-day regimen of either artesunate\u2013amodiaquine (AS\u2013AQ WinthropAfter admission (day 0), children were followed-up at pre-determined intervals  and at any time the care giver judged the child did not feel well. At each visit, both thick and thin blood smears were taken from enrolled patients. Microscopic examination was done under 100\u00d7 oil immersion magnification. A physician examined the child\u2019s clinical condition during the entire follow-up period.Patients were classified as (1) early treatment failure (ETF), (2) late clinical failure (LCF), (3) late parasitological failure (LPF), or (4) adequate clinical and parasitological response (ACPR), as per WHO guidelines . ChildreSamples for PCR analysis were collected on FTA cards, air-dried and stored in separated sealed bags in dry and dark conditions at room temperature. Samples were analysed at the Malaria Research and Training Centre in Bamako  using a previously described method .Paired samples (pre-treatment and recurrent parasites) were compared and possible outcomes were: (1) recrudescence if similar alleles were found in the pre- and post-treatment samples for all the markers, (2) re-infection, if the alleles of the pre- and post-treatment samples were distinct; (3) mixed recrudescence and re-infection, if similar alleles were found in the pre- and post-treatment samples for all the markers as mentioned above, but with additional distinct alleles identified; (4) indeterminate, if at least one marker in either the pre- or the post-treatment sample did not allow a definitive conclusion; or (5) no DNA isolated, if one or both the pre- and post-treatment samples could not be amplified .Children who experienced therapeutic failure received a rescue treatment consisting of oral quinine as monotherapy at the dose of 10\u00a0mg/kg every 8\u00a0h for 7\u00a0days. In cases of severe malaria, the rescue treatment was quinine administered intravenously . With the administration of the rescue treatment, the normal study follow-up was terminated. However, patients who received rescue treatment were invited to return to the research centre at least once, to allow the research team to verify the efficacy of the treatment.Two analysis methods were used to estimate the therapeutic efficacy.The Kaplan\u2013Meier survival analysis was used to calculate PCR-adjusted estimates at day 28 and day 42 , 20. PatThe second analysis used the per-protocol method and was carried out to allow a comparison with previous studies. In this method, the efficacy was estimated as proportion of ACPR and was based only on the patients for which a therapeutic endpoint  was reached. All other patients, including reinfections, were excluded from this latter analysis.Percentages and rates were expressed with 95% confidence intervals. Quantitative variables were described by mean and standard deviation if normally distributed, and by median and interquartile 25 and 75 percentiles range (IQR), if not normally distributed. Comparison of continuous numeric variables was tested with the one-way analysis of variance (ANOVA) if normally distributed, and with Kruskal\u2013Wallis test if not normally distributed.A blood smear was collected every 8\u00a0h (with a tolerance window of 2\u00a0h) starting from the first treatment dose intake. Sampling for parasite clearance was stopped when two consecutive blood smears were negative. As ETF endpoint requires a blood smear sample at day 3, this latter was always performed even if most of patient\u2019s blood smears performed for the parasite clearance were negative by day 1 or day 2 . The cleThe safety and tolerability of the three treatments were previously assessed and treatments were recommended for routine clinical use , 22, 23.Written informed consent was obtained from the parent or the guardian of each child enrolled.The MSF Ethical Review Board and the National Ethics Committee of the Ministry of Health of Niger approved the study protocol. The study was registered at ClinicalTrials.gov (NCT01755559).Between 17 June 2013 and 22 September 2014, of 1141 children aged 6\u201359\u00a0months who attended the study clinic , 663 children (221 in each study arm) satisfied the inclusion criteria, and were enrolled in the study.Of the 221 children included in each treatment arm, 4, 3 and 3 children were excluded from the Kaplan\u2013Meier analysis in the AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM arms respectively, while, in the same order, 41, 63 and 86 children were excluded from the per-protocol analysis. The study flowchart, with the reasons for excluding from the analyses, is presented in Fig.\u00a0The median age was 29, 29 and 30\u00a0months and the male/female ratio was 1.1, 1.2 and 1.1 for AS\u2013AQ, DHA\u2013PPQ and AL\u2013LM respectively. No significant differences on baseline characteristics were found between treatment arms , while the time to clearance 50 and 99% of parasitaemia was longer with AM\u2013LM than with the other two treatments (p\u00a0=\u00a00.001)  and the parasite clearance rate constant  were similar among the three treatments (1) Table\u00a0. No patip\u00a0=\u00a00.930). Only one child in the AS\u2013AQ arm was positive at day 14.At admission gametocytes were detected in 5.1, 2.3 and 4.3% of children in the AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM arms, respectively. In the same order 2.4, 1.9 and 2.0% gametocytes were detected at day 3 (p\u00a0<\u00a00.001). In children taking AM\u2013LM, the risk of reinfection was significantly higher for children aged 24\u201359\u00a0months (37.9%) compared to children aged 6\u201323\u00a0months (19.8%) (p\u00a0=\u00a00.007).During the 42-day follow-up, 28, 51 and 64 reinfections were reported in the AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM treatment arms, respectively. These events, expressed in terms of Kaplan\u2013Meier failures, represent a risk of 13.8, 24.7 and 31.9% for AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM respectively , 132 (59.7%), and 140 (63.4%) for AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM, respectively.The most common adverse events in decreasing order were fever, cough, rhinorrhoea, diarrhoea, conjunctivitis, pyoderma, vomiting and anorexia Table\u00a0. No chilThe number of events whose relationship to study treatment was classified as possible was 7, 8 and 4 for AS\u2013AQ, DHQ\u2013PPQ and AM\u2013LM respectively. The number of events whose relationship to treatment was classified as probable was 1 for AS\u2013AQ and DHA\u2013PPQ and none for AM\u2013LM. A total of 4 serious adverse events were reported; 1, 2 and 1 event with AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM, respectively.In the AS\u2013AQ arm, a child had decompensated anaemia (haemoglobin 5.3\u00a0g/dL) on the last day of treatment. At admission, parasitaemia was 2512\u00a0parasites/\u03bcL, haemoglobin level was 8.8\u00a0g/dL and the child had the spleen palpable under the costal rim. The relationship of the event to the study treatment was judged unlikely. The event was resolved in 5\u00a0days with a blood transfusion.The first serious adverse event with DHA\u2013PPQ was a child who experienced uncontrollable agitation requiring administration of sedatives on the second day of treatment. The relationship of the event with the study treatment was judged to be possible. The event was resolved.The second serious adverse event with DHA\u2013PPQ was a 14-month-old child who had a weight loss  19\u00a0days after the last treatment. At admission the weight was 6.6\u00a0kg. Weight loss was associated with a family event that resulted in a severe weaning of the child. The relationship of the event with the study treatment was judged unrelated. The child was treated at a nutritional centre and the event was resolved in 2\u00a0months .In the AM\u2013LM arm, a child had had a respiratory infection requiring hospitalization and administration of antibiotics 20\u00a0days after the last treatment. The relationship of the event with the study treatment was judged unrelated. The event was resolved.All the three anti-malarial treatments evaluated in this study showed to be very effective in treating uncomplicated falciparum malaria. The primary efficacy endpoint, the Kaplan\u2013Meier efficiency estimate, but also other efficacy criteria, the per-protocol analysis and the parasite clearance indicators, yielded very reassuring results and were in line with similar studies in neighbouring countries \u201328.No early therapeutic failure occurred and the estimated efficacies of all the three treatments were well above 95%, which is the WHO minimum threshold for recommending new anti-malarial treatments into the public health guidelines . The thrThe three treatments showed that they are well tolerated, as only two children did not complete the treatment due to vomiting and only one serious adverse event, uncontrollable agitation requiring sedation in a child treated with DHA\u2013PPQ, was thought to be associated with treatment. No adverse events were considered to have a highly probable relationship with the treatment. The number of other adverse events for which a relationship with treatment was considered to be possible or probable was quite low and their typology was similar to events reported in other studies , 29, 30.The lengthening of parasite clearance time is retained as an early indicator of the emergence of artemisinin resistance . The resThe only significant difference among the three study treatments between the three treatments was the number of reinfections that occurred during the follow-up period. While the high number of reinfections was expected, as most children were admitted to the study during the seasonal peak in the region, the number of reinfections after treatment with AM\u2013LM  was significantly higher than with the other two treatments. This might be accounted for the shorter half-life of lumefantrine  comparedThe results of this study indicate that the three treatments evaluated, AS\u2013AQ, DHA\u2013PPQ and AM\u2013LM, largely meet WHO criteria. AS\u2013AQ and AL\u2013LM, the current first-line options, may continue to be used and DHA\u2013PPQ may be recommended for the treatment of uncomplicated falciparum malaria in Maradi. Nevertheless, in the light of the emergence of artemisinin resistance in South-East Asia, their efficacy, as well as the molecular markers associated to artemisinin resistance, must continue to be monitored."}
+{"text": "Plasmodium vivax on prosperity and longevity have been highlighted in several recent clinical case reports. The first line of vivax treatment drugs has seen no radical innovation for more than 60\u00a0years. This study introduces a subtle incremental innovation to vivax treatment: a chloroquine and primaquine co-blister. The co-blister includes a new chloroquine formulation incorporating coated tablets to mask the drug\u2019s bitter taste and user-friendly packaging containing tablets of each drug, which may improve patient adherence and facilitate the appropriate use of the drugs. This new formulation will replace the non-coated chloroquine distributed in Brazil.Malaria remains a major public health problem, with half the world population at risk of contracting malaria. The effects of Patients were orally treated with 150\u00a0mg coated chloroquine tablets for 3\u00a0days: an initial 450\u00a0mg dose, followed by two 300\u00a0mg doses. The patients were treated concomitantly with two 15\u00a0mg primaquine tablets for 7\u20139\u00a0days, according to their weight. The primary objective of this study was to prove parasitological and clinical cure rates above 90\u00a0% by day 28.0\u2013t values of 374.44 (0.35) and 3700.43 (0.36)\u00a0ng/mL, respectively.This single-arm open-label non-comparative trial was conducted according to the WHO recommended methodology for the surveillance of anti-malarial drug efficacy in the Brazilian Amazon. On day 28, the parasitological and clinical response was adequate in 98.8\u00a0% of patients (CI 95\u00a0% 93.4\u2013100\u00a0%). The success rate on day 3 was 100\u00a0%, and the cumulative success rate by day 28 was 98.8\u00a0% (CI 95\u00a0% 91.7\u201399.8\u00a0%). There were no serious adverse events, with most adverse events classified as mild. The pharmacokinetic parameters of chloroquine analysed in whole blood dry spot samples showed mean (coefficient of variation) Cmax and AUCThis study reports an appropriate safety and efficacy profile of a new formulation of coated chloroquine tablets for vivax malaria treatment in the Brazilian Amazon. The cure rates meet the WHO efficacy criteria, supporting current Brazilian guidelines and the use of the formulation for vivax malaria treatment. Nevertheless, further studies should be conducted to address adherence and the effectiveness of the formulation.Trial registration RBR-77q7t3-UTN: U1111-1121-2982. Registered 10th May 2011The online version of this article (doi:10.1186/s12936-016-1530-0) contains supplementary material, which is available to authorized users. Malaria remains a major public health problem. Despite many advances in the past 15\u00a0years, 3.2 billion people are at risk of contracting malaria, which represents half of the population of the world. According the World Health Organization (WHO), in its 2015 World Malaria Report, the number of malaria cases decreased from an estimated 262 million in 2000 (range 205\u2013316 million) to 214 million in 2015 (range 149\u2013303 million), a decline of 18\u00a0% [Plasmodium falciparum has been the primary target of malaria drug development for the past 15\u00a0years. The high mortality, the emergence of drug resistance and the record high number of cases in Africa justify this approach. Although falciparum malaria is still a major public health problem, during the past decade, key achievements have dramatically changed its epidemiological profile, such as new rapid diagnosis tests (RDT) and new treatments comprising fixed dose combinations of artemisinin derivatives and other anti-malarial drugs. In Brazil, the incidence ratio of falciparum/vivax has considerably changed since artemisinin-based combination therapy (ACT) has been recommended as the first-line treatment. In 1988, Plasmodium vivax and P. falciparum had incidence rates of 50\u00a0% each. In 1990, P. falciparum corresponded to 44.3\u00a0% of cases; in 2009, 3\u00a0years after ACT implementation, P. vivax represented 84\u00a0% of the recorded cases, whereas P. falciparum represented only 16\u00a0% [P. vivax infections accounted for 84\u00a0% of cases in 2014. The total number of cases declined by 19\u00a0% compared to 2013. The overall number of cases is currently the lowest it has been for the past 35\u00a0years in Brazil [P. vivax has been highlighted based on its effects on prosperity and longevity [P. vivax was the most frequent cause of hospitalization (63\u00a0%) and death (11/23) due to malaria in the Brazilian Amazon region [The importance of ongevity , 7, and ongevity \u201313. In 2n region . In addin region , among wn region .In addition, various barriers are known to impair transmission control as the early appearance of gametocytes and relapses from hypnozoites. In 2009, Guerra et al.  estimateP. vivax treatment was developed in this study: a chloroquine and primaquine co-blister. This approach is a formulation and packaging improvement of the current vivax treatment recommended by the Brazilian Ministry of Health. The co-blister of chloroquine and primaquine was developed assuming that a user-friendly primary package containing both tablets of concomitant use may improve adherence [The first line drug treatment for vivax malaria has been used for more than 60\u00a0years with no radical innovation. To address the lack of research progress on this most neglected disease , an incrdherence . The co-dherence  especialdherence . In BrazThe new user-friendly primary polyvinylidene chloride (PVDC) package of coated chloroquine and primaquine may replace the non-coated chloroquine packed in paper strips that have been distributed in Brazil. The co-blister includes a new chloroquine formulation of coated tablets. Coating the chloroquine masks the bitter taste of the drug and may improve treatment adherence. Although it was not studied in this population, bitter taste is known as a factor that may impair adherence . In thisThe clinical study protocol and informed consent were reviewed and approved by the Ethics Committee at the Tropical Medicine Research Center of Rond\u00f4nia (N\u00ba 01/11 CEP/CEPEM e N\u00ba 0001.0.046.000-11 CAAE-SISNEP). The Brazilian National Council on Ethics in Research (CONEP), Ministry of Health, accredits this committee. The clinical study was conducted in accordance with the Helsinki Declaration, Good Clinical Practice  and the P. vivax infection confirmed by microscopy, asexual parasite count >250/\u03bcL; either axillar temperature \u226537.5\u00a0\u00b0C or a history of fever during the past 48\u00a0h; the ability to swallow tablets; and the availability to attend the study follow-up visits. Patients who met one of the following criteria were not eligible to participate: prior malaria treatment in the past 63\u00a0days; signs of severe malaria; either other febrile conditions or chronic disease ; the use of any medication that interferes with anti-malarial pharmacokinetics; an intolerance to chloroquine or primaquine; a known glucose-6-phosphate deficiency; pregnancy confirmed by human chorionic gonadotropin (hCG) in their urine or breastfeeding; or among the indigenous population.Patients with vivax malaria mono-infection willing to participate were included in the study after signing the informed consent form if they met the following inclusion criteria: aged between 18 and 70\u00a0years; weight between 50 and 90\u00a0kg, This single arm open-label non-comparative trial was conducted according the WHO recommendations  at the TEligible patients were treated in accordance with the Brazilian Ministry of Health recommendation . All patChloroquine and primaquine administrations were supervised during the first 3\u00a0days. The first dose was taken after diagnosis, and the following doses were administered between 8 and 10 a.m. If the patient vomited within 30\u00a0min after a dose, the same dose was administered again. Treatment adherence to primaquine between days 4 and 7 was assessed by inquiring with the patients at the day 7 follow-up visit. The patients were also asked about the use of concomitant therapies at every visit and recorded.Patients were evaluated on the day of enrolment and on days 1, 2, 3, 7, 14, 21 and 28 after the study period. The scheduled study procedures included a complete clinical anamnesis, a physical exam and a urinalysis at enrolment and an assessment of the most frequent clinical signs of malaria and adverse events (AEs) at all the follow-up visits. Additionally, blood samples were collected for parasite counts , haemoglobin and haematocrit evaluations , genotyping  and chloroquine pharmacokinetic analyses .The primary efficacy endpoint was the proportion of the population with an adequate clinical and parasitological response; otherwise, it was classified either as early treatment failure or late treatment failure.Both per protocol (PP) and intention-to-treat (ITT) analyses were conducted, for which the PP was the primary analysis, and the ITT analysis was performed to confirm the results. The PP population excluded any participant associated with a protocol violation or who was lost by the follow-up visit at the primary endpoint on day 28, whereas the ITT population included any violation or loss by the follow-up as parasitological and clinical failure. The cumulative success rate by day 28, corresponding to the probability of remaining parasite-free at day 28, was calculated using a Kaplan Meyer survival curve. The absolute number of patients who presented symptoms and parasitaemia 72\u00a0h after treatment were defined as early treatment failures. Demographic data were presented for both populations. All the data were summarized as frequencies and percentages for categorical variables and as the means\u00a0\u00b1\u00a0standard deviations (SD) for quantitative variables.The safety analysis was conducted in the ITT population describing the frequency of AEs and stratifying the AEs among the follow categories: serious AEs, AEs leading to treatment suspension, and AEs corresponding to causalities described as doubtful, unlikely, possible or probable. The mean haemoglobin and haematocrit results at baseline and at the follow-up visits are also presented.The patients were encouraged to seek unscheduled assessments if any AE was suspected. All clinical or laboratory abnormalities were categorized as grade I to IV according to the common terminology criteria for adverse events (CTCAE) of the National Cancer Institute . Any susThe parasitological densities were estimated using Giemsa-stained blood slides at a magnification of 1000\u00d7 as the WHO recommended methodology for the surveillance of anti-malarial drug efficacy . Two tramax, Tmax, AUC0\u2013t and AUC0\u2013inf were calculated using the software WinNonlinTM version 6.3 Pharsight and Microsoft Excel version 14.4.0. The presence of parasites with chloroquine blood levels above the 100\u00a0ng/mL threshold was considered to characterize resistance according the protocol and the WHO.To access chloroquine pharmacokinetics, samples of the peripheral blood were collected either at domiciliary visits or at the day of a clinic visit and transferred to Whatman (USA) ET 31 CHR E 3MM filter papers. The whole blood concentrations of chloroquine were measured using validated high performance liquid chromatography tandem to a mass spectrometry (HPLC\u2013MS/MS) method. This method was in accordance with Brazilian  and inteP. falciparum infection (3.8\u00a0%), were less than 18\u00a0years of age (2.5\u00a0%), contracted either other febrile condition or chronic disease (1\u00a0%), declined to participate (1\u00a0%), were over 70\u00a0years of age (0.4\u00a0%), or another justifiable reason (1.6\u00a0%). A flow chart is provided in Additional file A total of 921 patients were screened at the Tropical Medicine Research Centre of Rond\u00f4nia, Porto Velho, Brazil from April 2013 to April 2015. Eighty-eight patients who met all inclusion criteria were enrolled in the study. Patients were excluded from the study if they were unavailable to attend the study follow-up visits, resided in a remote area (41.7\u00a0%), had been administered malaria treatment within the past 63\u00a0days (21.4\u00a0%), weighted >90\u00a0kg (9.2\u00a0%), were already participating in another clinical trial (7.1\u00a0%), had a P. falciparum infected patient (1.1\u00a0%) and one malaria mixed infection patient (1.1\u00a0%). Three patients were lost by the follow-up visit (3.5\u00a0%). All 88 patients were included the ITT efficacy analysis and safety analysis. Qualitative and quantitative demographic population characteristics and the clinical baseline of the PP and ITT populations are presented in Tables\u00a0Among the 88 enrolled patients, 82 (93.2\u00a0%) finished the follow-up and were included in the PP analysis. One patient withdrew the informed consent 1.1\u00a0%). Protocol deviations included one .1\u00a0%. ProThe most frequent symptoms at enrolment were fever (98.7\u00a0%), myalgia (90.9\u00a0%), chills (89.8\u00a0%) and headache (86.4\u00a0%). The concomitant treatments most reported prior to enrolment were administration of antipyretics/analgesics (46.6\u00a0%), antihypertensive agents (3.4\u00a0%) and antihistamines (2.3\u00a0%). After study initiation, 73.3\u00a0% of the population reported the administration of concomitant medications. Most of these patients took a symptomatic treatment two or three times. Non-steroidal anti-inflammatory drugs (NSAIDs) were the drugs most frequently reported concomitant medication (50.3\u00a0%). NSAIDs and antihistamines were often taken within the first 3\u00a0days after anti-malarial treatment initiation, which may be due to either to malaria symptoms or to chloroquine-related pruritus.All treatment doses with the new chloroquine formulation and primaquine were supervised during the initial 3\u00a0days. Patients were asked about their adherence to the primaquine doses at the day 7 follow-up visit. Two patients (2.3\u00a0%) reported disruption of primaquine treatment either on day 4 or 5 but neither were classified as a treatment failure. Therefore, these patients were included in the PP analysis.Among the 82 patients in the PP population, parasitological and clinical responses were adequate in 98.8\u00a0% (CI 95\u00a0% 93.4\u2013100). There was one failure at day 29; however, because the protocol allows a 1-day margin for this follow-up visit, this patient was considered a late clinical and parasitological failure.Secondary endpoints included the cure rate at day 3  and the cumulative success rate at day 28 based on the survival Kaplan\u2013Meyer method. Early parasitological clearance 72\u00a0h after treatment initiation was evaluated in 84 patients. The success rate at day 3 was 100\u00a0%. The cumulative success rate at day 28 or the probability of remaining parasite-free at day 28 was 98.8\u00a0% (CI 95\u00a0% 91.7\u201399.8).There were no deaths in this study, nor any other serious adverse event, as defined by GCP. No AEs led to treatment suspension. A safety analysis was conducted for the ITT population, as all patients received at least three days of chloroquine and primaquine.A total of 380 AEs were reported during the study. The principal investigator described 304 (80\u00a0%) AEs as grade I, 74 AEs (19.7\u00a0%) as grade II, and only one (0.3\u00a0%) AE as grade III. There was no report of a grade IV AE. The number of AEs and their distribution according to the type of casualty are given in Table\u00a0Among the 214 (56.3\u00a0%) AEs classified as possibly related to the test drugs, 186 (48.9\u00a0%) were classified as grade I and 28 (7.4\u00a0%) as grade II. None of the grade III or grade IV AEs were classified as possibly related to the test drugs. There were 82 (21.6\u00a0%) events likely related to the test drugs, of which 61 (16.0\u00a0%) were classified as grade I, and 21 (5.5\u00a0%) as grade II; none were reported as grade III or grade IV. Among the 296 events possibly or likely related to the test drugs, 247 (83.4\u00a0%) were classified as grade I and, therefore, did not require any treatment.The most frequent AEs were pruritus (56\u00a0%), headache (38.6\u00a0%), abdominal pain (35.2\u00a0%), sleep disorder (31.8\u00a0%), nauseas (28.4\u00a0%), diarrhoea (20.5\u00a0%). Vomiting, lack of appetite, cutaneous rash, muscular pain, compartmental disorder, and dizziness were reported in more than 5\u00a0% of the population. Thirty-nine patients (44.3\u00a0%) reported AEs that were classified as doubtfully related to the study drugs. AEs possibly related to the drugs were observed in 64 (72.7\u00a0%) patients, and AEs likely related to the medication were described in 49 (55.7\u00a0%) patients. A total of 74 (84\u00a0%) patients reported either an AE possibly related to or an AE likely related to the study drugs.The distribution of AEs according the main body system disorders and the follow-up visit interval is presented in Fig.\u00a0A similar pattern was observed for the time to disappearance of symptoms. Fevers were recorded in 87 (98.9\u00a0%) patients at the time of enrolment and were present in 4 (4.5\u00a0%) patients at day 3. At the time of enrolment, 48 and 40 patients reported nausea and abdominal pain, respectively. At day 1, there was a 92.4\u00a0% decrease in reported nausea and a 68.7\u00a0% decrease in reported abdominal pain. As these symptoms are related to both the disease and treatment, they were recorded as AEs if they were not present prior to administration or they were recurrent. This overlap in symptoms may have led to an over estimation of AEs; however, it provides a conservative safety profile.The mean haematocrit and haemoglobin on days 0, 14 and 28 are presented in the Table\u00a0The pharmacokinetic parameters of chloroquine in whole blood dry spot samples (n\u00a0=\u00a086 patients) are presented with the mean, standard deviation, coefficient of variation, and minimum and maximum values in Table\u00a0max was 122.20\u00a0ng/mL. The lowest value did not correspond to the patient who exhibited a clinical and parasitological failure at day 29. This patient maintained chloroquine blood levels above the minimum effective blood concentration until day 14. However, the sample collected on the failure day contained a lower concentration than 100\u00a0ng/mL and, therefore, does not characterize a resistant parasite according the protocol and the WHO. The coefficients of variation of the main pharmacokinetic parameters were high (\u22650.35), which might be due to the restricted (6) sample of collection points.All patients achieved whole blood levels above 100\u00a0ng/mL. The minimum value at CDespite the many achievements in malaria control over the past 15\u00a0years, malaria remains a major public health problem . In thisThis study presents the safety and efficacy results of a new coated chloroquine formulation; nevertheless, improving adherence, acceptability and suitability to local needs and behaviours remain clear limitations of malaria treatment, and the effectiveness for which have not yet been evaluated and remain to be proven. Further studies to are currently planned to provide this lack of data and to increase the body of evidence supporting innovative and simple approaches to improve public health. Additional pharmaceutical developments must also be pursued to address the need of children who cannot swallow tablets.P. vivax more than 1\u00a0year prior to this study, and he reported adherence to the primaquine treatment. The pharmacokinetic analysis revealed the chloroquine exposure in this patient that was similar to the other patients. The drug levels on the day of failure did not characterize the parasite as resistant. Due to the one single failure, the known high frequency of heterologous hypnozoite activation during a vivax relapse, and the lack of access to deep sequencing at the end of this study, genotyping to differentiate relapse from possible re-infection was not conducted.The efficacy results reported here are comparable to other recent publications in Brazil . AlthougIn an endemic area, the probability of remaining parasite-free as long as possible is a relevant measure that contributes to patient quality of life. At day 28, the probability of the patients being parasite-free was 98.8\u00a0% in this trial. The treatments were directly observed during the first 3\u00a0days, and primaquine doses were continually adjusted per kilogram. A limitation of this study is highlighted by the fact that the initial 3\u00a0days of treatment were directly observed in this clinical trial setting, which is unlike routine health assistance in this area and may compromise the external validity of this study. On the other hand, primaquine administration was not supervised after day 3, and therefore, the actual efficacy of the results as to whether they represent the complete recommended treatment may be challenged.Many AEs were reported in the safety results, highlighting the awareness of the study team regarding the importance of the patients\u2019 safety profiles in a study associated with regulatory processes. Most of the AEs were mild and did not require any symptomatic treatment. The safety profile concerned treatment adherence, although it is similar to that described in literature. A review of primaquine clinical trials including safety evaluations from 11,466 patients reported a rate of serious AEs of 8.7 per 10,000 and no deaths . AlthougP. vivax resistance to the treatment in the study population. These results support the use of a new formulation of coated chloroquine tablets in Brazil and may be the first step towards a user-friendly co-blister of chloroquine and primaquine to treat malaria vivax.This study shows the safety and efficacy of a new formulation of coated chloroquine for vivax malaria treatment in Brazil. The cure rate was 98.8\u00a0% (CI 95\u00a0% 93.4\u2013100\u00a0%) and met the WHO criteria. This study provides satisfactory evidence of the efficacy of the formulation, supporting the National Malaria Control Programme treatment guidelines. The treatment is also safe in the dose range and population studied. There is no evidence of"}
+{"text": "Malaria remains one of the most serious infections for travellers to tropical countries. Due to the lack of harmonized guidelines a large variety of treatment regimens is used in Europe to treat severe malaria.Plasmodium falciparum malaria according to the 2006 WHO criteria.The European Network for Tropical Medicine and Travel Health (TropNet) conducted an 8-year, multicentre, observational study to analyse epidemiology, treatment practices and outcomes of severe malaria in its member sites across Europe. Physicians at participating TropNet centres were asked to report pseudonymized retrospective data from all patients treated at their centre for microscopically confirmed severe From 2006 to 2014 a total of 185 patients with severe malaria treated in 12 European countries were included. Three patients died, resulting in a 28-day survival rate of 98.4%. The majority of infections were acquired in West Africa . The proportion of patients treated with intravenous artesunate increased from 27% in 2006 to 60% in 2013. Altogether, 56 different combinations of intravenous and oral drugs were used across 28 study centres. The risk of acute renal failure (36 vs 17% p\u00a0=\u00a00.04) or cerebral malaria  was significantly higher in patients\u00a0\u226560\u00a0years than in younger patients. Respiratory distress with the need for mechanical ventilation was significantly associated with the risk of death in the study population . Post-artemisinin delayed haemolysis was reported in 19/70 (27%) patients treated with intravenous artesunate.The majority of patients with severe malaria in this study were tourists or migrants acquiring the infection in West Africa. Intravenous artesunate is increasingly used for treatment of severe malaria in many European treatment centres and can be given safely to European patients with severe malaria. Patients treated with intravenous artesunate should be followed up to detect and manage late haemolytic events. Around 5200 cases of malaria are imported to EU countries per year, of which up to 10% progress to severe malaria  , 9, 10. Quinine is still widely used in Europe, but the rate of patients receiving intravenous artesunate almost doubled in the 8-year study period. In the final year 2014, every second patient received intravenous artesunate. Current surveillance data from national reference centres indicate that the proportion of patients treated with artesunate is further increasing, particularly in countries where participating centres still reported frequent treatment with quinine during this study . AlthougFollowing the initial description of an episode of severe prolonged haemolysis after treatment of a patient with severe malaria with intravenous artesunate in Japan in 2002 , late haMortality in this patient population was very low, reflecting the high standard of intensive care in Europe. Previous single-centre and national studies reported mortality rates between 4 and 15% , 22, 34.Only 10% of patients with severe malaria had taken anti-malarial chemoprophylaxis and very few of them had been fully compliant. These data suggest that correct anti-malarial prophylaxis can effectively prevent severe malaria in European travellers. Counselling of travellers on malaria prevention should be improved and coverage increased, particularly for travellers going to West Africa, where 60% of infections in this study were acquired. Little is known about the proportion of European travellers who take prophylaxis. In a recent study only 60% of travellers from the UK to endemic areas used anti-malarial chemoprophylaxis . VFR traSupportive treatments such as exchange transfusions and erythrocyte apheresis are a matter of controversy and their use is guided by national or local practices. European single-centre studies recently failed to demonstrate improved parasite clearance through whole blood or erythrocyte exchange, compared to patients treated with quinine or artesunate alone , 38. OnlThis observational study has inherent limitations. As patient information was collected retrospectively in TropNet centres and not all patients treated may have been reported equally, selective under-reporting, e.g., for patients who died, may have occurred. Bias in reporting data on PADH must be assumed as the condition was not known at the beginning of the study and no universally accepted clinical definition exists to date. All TropNet centres are referral centres for tropical medicine and patient composition as well as treatment data may not fully reflect treatment practices and outcomes in non-referral hospitals. Moreover, data reported by the participating treatment centres may not always fully reflect treatment practices in the respective countries. For patients with elevated creatinine levels at the end of follow-up, it cannot be ruled out that unknown or undocumented elevation of creatinine had existed before the episode of severe malaria.The data show that the majority of patients with severe malaria in Europe are tourists or migrants acquiring infection in West Africa. Intravenous artesunate is increasingly used for treatment of severe malaria; it is the most effective drug and can be safely given to European patients with severe malaria. There is need for harmonization of guidelines for the treatment of severe malaria in Europe. Patients treated with intravenous artesunate should be followed up to detect late haemolytic events."}
+{"text": "Severe malaria is a medical emergency associated with high mortality. Adequate treatment requires initial parenteral therapy for fast parasite clearance followed by longer acting oral antimalarial drugs for cure and prevention of recrudescence.In a randomized controlled clinical trial, we evaluated the 42-day parasitological outcomes of severe malaria treatment with intravenous artesunate (AS) or intravenous quinine (QNN) followed by oral artemisinin based combination therapy (ACT) in children living in a high malaria transmission setting in Eastern Uganda.P\u2009<\u20090.001). Overall, 63.3% (178/281) of the participants had unadjusted parasitological treatment failure over the 42-day follow-up period. Molecular genotyping to distinguish re-infection from recrudescence was performed in a sample of 127 of the 178 participants, of whom majority 93 (73.2%) had re-infection and 34 (26.8%) had recrudescence. The 42\u00a0day risk of recrudescence did not differ with ACT administered. Adverse events were of mild to moderate severity and consistent with malaria symptoms.We enrolled 300 participants and all were included in the intention to treat analysis. Baseline characteristics were similar across treatment arms. The median and interquartile range for number of days from baseline to parasite clearance was significantly lower among participants who received intravenous AS (2 (1\u20132) vs 3 (2\u20133), In this high transmission setting, we observed adequate initial treatment outcomes followed by very high rates of malaria re-infection post severe malaria treatment. The impact of recurrent antimalarial treatment on the long term efficacy of antimalarial regimens needs to be investigated and surveillance mechanisms for resistance markers established since recurrent malaria infections are likely to be exposed to sub-therapeutic drug concentrations. More strategies for prevention of recurrent malaria infections in the most at risk populations are needed.PACTR201110000321348).The study was registered with the Pan African Clinical Trial Registry ( Plasmodium falciparum [Malaria is a major cause of morbidity and mortality worldwide. In 2015, the WHO estimates that there were 214 million cases of malaria and 438,000 deaths worldwide, with 90% of deaths occurring in Africa [P. falciparum malaria worldwide. Uganda adopted the policy to use intravenous AS in 2013 with intravenous QNN recommended as an alternative. To date, intravenous QNN is still widely prescribed due to inconsistent availability and frequent stock-outs of intravenous AS in health facilities [Two previous multicentre randomized trials demonstrated superiority of intravenous artesunate (AS) over intravenous quinine (QNN) for treatment of severe malaria in Africa and Asia \u20137. Intracilities .The Ugandan malaria treatment policy recommends artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) as the first and second line ACTs for follow-on therapy. Previous studies demonstrated excellent efficacy and effectiveness of both AL and DP for treatment of uncomplicated malaria \u201311 with The study was a randomized single blind clinical trial conducted in Tororo District Hospital situated in Eastern Uganda, an area with perennial malaria transmission and an annual entomological inoculation rate estimated to be 310 infective bites per person per year .P.falciparum malaria with at least one of the following symptoms and signs; repeated convulsions (more than 2 in 24\u00a0h), impaired consciousness (Glasgow Coma Score\u2009<\u200911 for adults and Blantyre Coma Score\u2009<\u20093 for children), coma, hyperpyrexia (axillary temperature >/= 40\u00a0\u00b0C), respiratory distress (acidotic breathing), inability to tolerate oral therapy and vomiting all oral intake, circulatory collapse or shock (systolic Blood Pressure\u2009<\u200980\u00a0mmHg in adults and <50\u00a0mmHg in children), spontaneous bleeding, haemoglobinuria (tea colored urine), jaundice, prostration , severe normocytic anemia (hemoglobin <5\u00a0g/dL), hypoglycemia (blood sugar <2.2\u00a0mmol/l or 40\u00a0mg/dL), renal impairment (serum creatinine >265\u00a0\u03bcmol/l), and hyperparasitaemia (>10%). Patients were excluded if they had obvious concomitant febrile illness, history of allergy to any of the study drugs, were resident more than 20\u00a0km from the hospital and therefore could not return for follow-up visits or if they had history of receipt of an effective antimalarial drug within 72 hours before presenting to hospital. The last exclusion criterion was modified to exclude patients with history of receipt of an effective antimalarial drug within 24 hours before presenting to hospital due to large numbers of patients reporting self medication with antimalarial drugs prior to presenting to the hospital, thus affecting recruitment.Consecutive patients presenting to the hospital with symptoms and signs of severe malaria and a microscopy positive thick blood film for malaria parasites were referred to the study physicians for further assessment for study eligibility. Patients were enrolled if they fulfilled the following inclusion criteria: age 6\u00a0months and above, with history of fever in the last 24\u00a0h or axillary temperature\u2009>\u200937.5\u00a0\u00b0C, a diagnosis of severe malaria and whose parent or guardian provided written informed consent. Severe malaria was defined as presence or history of fever plus a positive blood film for A 2-stage block randomization procedure was performed by an independent statistician using the R computer software in blocks of 20. Participants were first randomized to intravenous AS or intravenous QNN then to AL or DP. Computer-generated randomization codes were enclosed in sequentially numbered opaque sealed envelopes containing treatment allocation. After meeting study eligibility criteria, the study nurse assigned the next envelope to the participant, opened the envelope and assigned treatment allocation. Treatment was immediately initiated.Intravenous AS  was administered as 2.4\u00a0mg/kg at start of treatment, repeated at 12 and 24\u00a0h and every 24\u00a0h till the switch to oral therapy. Each 60\u00a0mg vial of artesunic acid was dissolved in 1\u00a0mL of 5% sodium bicarbonate (provided in the pack) and mixed with 5\u00a0mL of 5% dextrose, then injected as a slow bolus into an indwelling intravenous cannula. Intravenous QNN dihydrochloride  was administered as 10\u00a0mg/kg body weight in 5% dextrose (10\u00a0ml/kg) through an indwelling catheter over 4\u00a0h and repeated 8 hourly till the switch to oral therapy.Intravenous antimalarial therapy was administered for a minimum of 24\u00a0h and when participants could tolerate oral therapy, they were given a full course of the oral ACT assigned. Oral AL  was administered according to body weight as; one (5\u201314\u00a0kg), two (15\u201324\u00a0kg), three (25\u201334\u00a0kg) and four (>35\u00a0kg) tablets given twice daily for 3 days. Parents and care takers were given instructions to administer the second dose of AL 8 hours after the first then every morning and evening to complete a 3\u00a0day course and all doses to be administered after food or a cup of milk. Oral DP  40\u00a0mg\u2009+\u2009piperaquine (PQP) 320\u00a0mg tablets) was administered targeting a total of 6.4 and 51.2\u00a0mg/kg of dihydroartemisinin and piperaquine, respectively, given in three equally divided doses to the nearest quarter tablet. We used a pill cutter to ensure that the tablet fractions were as close to the nearest quarter tablet as possible.All participants received paracetamol in a dose of 15\u00a0mg/kg at 8 hourly intervals. Adjunctive and supportive treatment such as diazepam for convulsions and dextrose for hypoglycemia was given in accordance with the Uganda Ministry of Health guidelines. All caretakers were given counseling on adherence to drugs, follow-up visits and potential drug side effects by the study nurse. They were instructed to observe the participants for 30\u00a0min after drug administration and if vomiting occurred they were to administer another dose. If vomiting occurred repeatedly (> 3 times) they were to bring back the participant to the study clinic for evaluation and treatment plus collection of additional doses of drugs.Serial blood smears were performed at 0, 1, 2, 4, 6, 8, 10, 12, 16, 20, 24\u00a0h post start of intravenous drugs followed by every 6\u00a0h until 6\u00a0h after parasite clearance. Participants were discharged from hospital when the blood smear was negative for malaria parasites and were followed up for 42\u00a0days to ascertain parasitological outcomes and monitor for adverse events. They were brought back to the clinic for follow up assessment on days 1, 3, 7, 14, 21, 28, 35, 42 post the commencement of ACT administration, and any unscheduled day if the participant felt unwell. Follow-up assessment on each of these days consisted of medical history, physical examination and a finger prick to collect blood on slides for thick blood film for malaria diagnosis and on filter paper for genotyping. Participants with positive malaria films were reassessed for severity and treated accordingly. Participants who returned with severe malaria were re-admitted and treated with intravenous AS plus AL. Those who returned with uncomplicated malaria were evaluated for treatment failure and treated according to national guidelines with DP if the blood smear was positive before day 14 and AL if it was positive after day 14 post commencement of ACT. Participants were discontinued from study follow up if they could not continue with the prescribed medication, missed a scheduled follow-up visit and could not be located at home or if they administered additional antimalarial drugs outside the study protocol.Thin smears were performed to determine the type of malaria parasite species and thick smears to determine the parasite density and for detection of parasitemia during follow-up. Thick blood smears were stained using 3% Giemsa for 30\u00a0min and read by experienced laboratory technologists. All slides were read by two independent microscopists and any discrepant results were reviewed by a tie breaker. Microscopists were blinded to participants\u2019 treatment assignment. In order to blind the microscopists, blood samples were drawn by the study nurse and study numbers were used to label all samples. The study numbers were not known by the microscopists and they did not have access to study data including study databases, participants\u2019 assessments and clinical notes. The study doctors and nurses were aware that laboratory technicians were blinded to treatment assignment and were trained not to reveal the treatment assignments to the microscopists.Parasite density was calculated by counting the number of asexual parasites (ring stages) per 200 white blood cells (WBCs) or per 500 if the count was less than ten parasites per 200 WBCs, assuming a WBC count of 8000/uL of blood. A smear was considered negative if no parasites were seen after review of 100 high-power fields. Complete blood count and hemoglobin estimation were performed using the Coulter counter, Beckman coulter, Life Science, United States of America.Filter paper blood samples were collected and subsequently molecular genotyping was conducted for participants with parasitological treatment failure to distinguish re-infection from recrudescence. We used Whatman 3MM Filter paper from Sigma. Molecular genotyping was performed at the Makerere University-University of California San Francisco Molecular Biology laboratory in Mulago, Kampala. Parasite DNA was extracted from filter paper blood samples collected on the day of enrollment and the day of parasitological treatment failure using Chelex 100 Resin extraction  as previously described . Paired To evaluate the 42\u00a0day parasitological treatment outcomes following treatment of severe malaria with intravenous AS or intravenous QNN plus ACT (AL or DP); we selected the primary study outcome as parasitological treatment failure unadjusted by genotyping classified as parasitemia detected by thick blood smear during follow-up. This primary outcome was selected because it best represents the treatment outcome measure used in routine clinical care. The secondary outcomes were parasitological treatment failure adjusted by genotyping classified as positive PCR on any follow-up day categorised as reinfection or recrudescence. Adverse events were defined as any medical occurrence post study drug administration. They were graded as mild, moderate, severe and life threatening and their relationship to the study drug was classified as unrelated, possibly, probably or definitely related to study drug.We estimated the proportion of parasitological failure of 23.6% for intravenous QNN followed by ACT. Assuming a 50% difference in the proportion of patients with parasites at 24\u00a0h following start of IV AS, using an alpha of 5%, power of 80%, and adjusting for possible loss to follow up of 10%, a total of 330 patients were needed.Data were entered and verified using MS ACCESS and analyzed using STATA version 13.1 . Descriptive statistics were used to compare demographic and clinical characteristics among the four study arms. Continuous variables were compared using Wilcoxon test for non-normally distributed data. Categorical variables were compared using Chi-square test. Parasite density was normalized using logarithmic transformation. Intention-to-Treat analysis was used, which included all enrolled participants. Unadjusted treatment failure was classified as a positive blood smear on any of the follow-up days. Adjusted treatment failure was classified as either re-infection or recrudescence based on genotyping. The risk of treatment failure at 28, 35 and 42\u00a0days of follow up (unadjusted and adjusted by genotyping) were estimated using the Kaplan-Meier survival method and compared using the Log Rank test. Time at risk was calculated from day one of ACT allocation to date of treatment failure among participants who failed, last day of follow-up for those who did not complete follow-up, or day 42 for the patients who completed 42\u00a0days of follow-up. In the analysis for adjusted parasitological outcomes, only recrudescence was considered as true parasitological treatment failure. Safety data from all participants were analyzed. In a sensitivity analysis, we imputed missing genotype data among the treatment failures using multiple imputation with chained equations. Variables included in the imputation model were age, sex, weight, parasite density at baseline visit and treatment arm. Five rounds of imputations were performed. Statistical tests were performed at 5% level of significance and 95% confidence intervals.We enrolled and followed up 300 participants between January 2012 and March 2013 . During the 42-day follow-up period, 63.3% (178/281) of the participants were classified as having unadjusted parasitological treatment failure; occurring in 41/71  of participants who received AS\u2009+\u2009AL, 53/79  of those who received AS +DP, 39/79  of those who received QNN\u2009+\u2009AL and 45/71  of those who received QNN\u2009+\u2009DP. The 42\u00a0day risk of unadjusted parasitological treatment failure was 40% lower for participants who received QNN\u2009+\u2009AL  but not different for those who received QNN\u2009+\u2009DP  or AS\u2009+\u2009AL  compared to those who received AS\u2009+\u2009DP. The risk of getting unadjusted treatment failure was not different across the four study arms (p\u2009=\u20090.044)  for number of days to parasite clearance was significantly lower among participants who received intravenous AS compared with participants who received intravenous QNN (2 (1\u20132) vs 3 (2\u20133), Molecular genotyping was performed for the first 127 (71.3%) of the 178 participants with unadjusted parasitological failures, of whom; 30/127 (23.6%) received AS\u2009+\u2009AL, 37/127 (29.1%) received AS\u2009+\u2009DP, 29/127 (22.8%) received QNN\u2009+\u2009AL, and 31/127 (24.4%) received QNN\u2009+\u2009DP. Majority  had re-infection while 34 (26.7%) had recrudescence as the cause of treatment failure.p\u2009=\u20090.155).Reinfection occurred in 24/71  of participants who received AS\u2009+\u2009AL, 30/79  of those who received AS +DP, 21/79  of those who received QNN\u2009+\u2009AL and 18/71  of those who received QNN\u2009+\u2009DP. The risk of getting reinfection was not different across the four study arms (p\u2009=\u20090.154), followed by QNN\u2009+\u2009AL  and AS\u2009+\u2009AL  compared to those who received AS\u2009+\u2009DP. The 42\u00a0day risk of recrudescence was not different among the four ACT study arms (p\u2009=\u20090.055)  of participants who received AS\u2009+\u2009AL, 7/79  of those who received AS +DP, 8/79  of those who received QNN\u2009+\u2009AL and 13/71  of those who received QNN\u2009+\u2009DP. The probability of having recrudescence was highest among patients who received QNN\u2009+\u2009DP . Recrudescence occurred in 6/71  of participants who received AS\u2009+\u2009AL, 12/79  of those who received AS +DP, 13/79  of those who received QNN\u2009+\u2009AL and 23/71  of those who received QNN\u2009+\u2009DP. The risk of getting recrudescence was not different across the four study arms (p\u2009=\u20090.055).In the sensitivity analysis, 41/178 (23.0%) received AS\u2009+\u2009AL, 53/178 (29.8%) received AS\u2009+\u2009DP, 39/178 (21.9%) received QNN\u2009+\u2009AL, and 45/178 (25.3%) received QNN\u2009+\u2009DP. Majority  had re-infection while 54 (30.3%) had recrudescence as the cause of treatment failure. Reinfection occurred in 35/71  of participants who received AS\u2009+\u2009AL, 41/79  of those who received AS +DP, 26/79  of those who received QNN\u2009+\u2009AL and 22/71  of those who received QNN\u2009+\u2009DP. The risk of getting reinfection was not different across the four study arms  experienced severe adverse events, all classified as unrelated to study drugs. Severe adverse events were commonest among participants who received QNN\u2009+\u2009AL  followed by AS\u2009+\u2009DP , QNN\u2009+\u2009DP  and least among those who received AS\u2009+\u2009AL . Pneumonia was the commonest severe adverse event occurring in 12/22 (55%) followed by gastro enteritis in 6 (27%) and severe anemia in 4 (18%). All the adverse events were treated and resolved completely except for one participant who experienced repeated convulsions, after receiving intravenous AS and died within 2\u00a0h post drug administration.We evaluated the 42-day parasitological treatment outcomes and safety following treatment of children with severe malaria using intravenous AS or intravenous QNN plus ACT (AL or DP) in Tororo District Hospital in Eastern Uganda. Participants who received intravenous AS cleared malaria parasites significantly faster than those who received intravenous QNN, which is consistent with data from previous large trials , 5.There was low mortality among our study participants. The risk of death for patients with severe malaria is high and increases with delayed initiation of treatment and in presence of multiple complications. Risk of death is also dependent on the patient\u2019s age, background immunity, pre-morbid and concomitant diseases. We did not study time intervals between start of symptoms and presentation to hospital or receipt of antimalarial drug. Our previous study demonstrated longer duration of illness before receiving antimalarial medication among children presenting to hospital with severe malaria compared to those who had uncomplicated malaria . It is pBoth AL and DP were effective at clearing residual parasites during the 42-day follow-up period. Majority of our study participants classified as treatment failure actually suffered re-infection with malaria parasites during the 42-day follow-up period, with higher reinfection rates among participants who received AS\u2009+\u2009ACT compared to QNN\u2009+\u2009ACT regimens. Our findings are consistent with previous data from similar high malaria transmission settings which demonstrated high rates of re-infection with malaria parasites after initial treatment , 21. OurThe high rate of malaria re-infection in our study population, calls for strengthening of national efforts in malaria prevention, with particular emphasis on high malaria transmission settings. Interventions such as long lasting insecticide treated bed nets, indoor residual spraying with insecticides, malaria chemoprophylaxis for high risk groups, are highly effective in reducing malaria morbidity  and haveIt is important to understand why some children living in malaria endemic areas are at higher risk for development of severe malaria and recurrent malaria attacks than others living in the same areas. Several hypothesis for host susceptibility to severe malaria and re-infection have been suggested and some are under study , 25. TheWe demonstrated low levels of recrudescence among our study participants with no difference in the 42-day risk of recrudescence among the four ACT study arms. Among the few participants in whom recrudescence was demonstrated; the highest probability of recrudescence was among participants who received the combination of QNN\u2009+\u2009DP followed by QNN\u2009+\u2009AL, although this finding was not statistically significant possibly due to the limitation of the small sample size for this comparison. Our study was also limited by the genotyping method used which is known to have low resolution. The long acting drug should ideally mop up all residual parasites to prevent recrudescence while also preventing re-infection. It is possible that the QNN\u2009+\u2009ACT regimens predisposed participants to higher rates of recrudescence due to slower initial clearance of parasite biomass by intravenous QNN compared to intravenous AS. This finding supports the current recommendation to use intravenous AS in preference to intravenous QNN for initial treatment for severe malaria and calls for improved availability and accessibility to intravenous AS especially in the high transmission areas.The study participants\u2019 clinical characteristics did not differ among study groups with the commonest forms of severe malaria documented to be; inability to feed and prostration. Our study participants did not present with very severe forms of malaria, a finding that is consistent with data from previous studies which demonstrated lower risk of very severe forms of malaria among individuals living in areas with high malaria transmission most likely due to development of partial immunity from recurrent malaria infections early in life . IndividWe observed adequate initial treatment outcomes followed by very high rates of malaria re-infection post severe malaria treatment with intravenous AS or intravenous QNN followed by ACT (AL or DP) in a high malaria transmission setting. The impact of recurrent antimalarial treatment on the long term efficacy of antimalarial regimens needs to be investigated and surveillance mechanisms for resistance markers established since recurrent malaria infections are likely to be exposed to sub-therapeutic drug concentrations. More strategies for prevention of recurrent malaria infections in the most at risk populations are needed."}
+{"text": "Plasmodium vivax recurrence. With a favorable safety profile, this antimalarial treatment proved to be a good first-line alternative. Chloroquine resistance is probably underestimated in the area.In the Brazilian Amazon, the artesunate\u2013amodiaquine combination was more effective in preventing  Plasmodium vivax chloroquine (CQ) resistance, there have been no trials comparing its efficacy with that of artemisinin-based combination therapies (ACTs) in Latin America.Despite increasing evidence of the development of \u00ae (ASAQ) versus CQ for treatment of uncomplicated P. vivax infection in Manaus, Brazil. Patients were followed for 42 days. Primary endpoints were adequate clinical and parasitological responses (ACPR) rates at day 28. Genotype-adjustment was performed.This randomized controlled trial compared the antischizontocidal efficacy and safety of a 3-day supervised treatment of the fixed-dose combination artesunate-amodiaquine WinthropFrom 2012 to 2013, 380 patients were enrolled. In the per-protocol (PP) analysis, adjusted-ACPR was achieved in 100% (165/165) and 93.6% (161/172) of patients in the ASAQ and CQ arm  at day 28 and in 97.4% (151/155) and 77.7% (129/166), respectively , at day 42. Apart from ITT D28 assessment, superiority of ASAQ on ACPR was demonstrated. ASAQ presented faster clearance of parasitaemia and fever. Based on CQ blood level measurements, CQ resistance prevalence was estimated at 11.5% (95% CI: 7.5-17.3) up to day 42. At least one emergent adverse event (AE) was recorded for 79/190 (41x6%) in the ASAQ group and for 85/190 (44x7%) in the CQ group. Both treatments had similar safety profiles.ASAQ exhibited high efficacy against CQ resistant P. vivax and is an adequate alternative in the study area. Studies with an efficacious comparator, longer follow-up and genotype-adjustment can improve CQR characterization.Clinical Trials Registration. NCT01378286. Plasmodium vivax is the malaria-causing parasite species with the widest geographical distribution, resulting in 2.85 billion people at risk  vs 2 [1.1%]). At day 28, patients who received CQ presented with a higher increase from baseline . Overall AE rates were similar (There was a relative decrease of hemoglobin (Hb) between inclusion and day 7 for all patients , with greater reduction observed for ASAQ . Interpretation of failures occurring after day 28 is complicated by the possibility of recurrences being relapses or reinfections , 25. DesP. vivax due to the post-treatment prophylactic effect [In the absence of PQ administration, the drug\u2019s elimination half-life is an important property to be considered against c effect . Its infc effect  was not c effect . This rac effect . Thus, oP. vivax efficacy studies with interesting results [P. falciparum, there are no well-defined criteria to differentiate recrudescence from reinfection and relapse [The genotype-adjusted ACPR rates were measured in an attempt to gain a more rigorous case definition of CQR. Genotyping has been applied in  results , 36. How relapse . PCR cor relapse .ASAQ was more effective in clearing patent gametocytemia. CQ may take up to 4 days for gametocyte clearance . StudiesHb reduction at day 7 was higher in patients who received ASAQ. Possible explanations include higher inflammatory response and/or oxidative stress associated with treatment, suppression of the blood marrow by artesunate, and pitting , 51. TheP. falciparum malaria\u201d [The safety and tolerability of ASAQ FDC were consistent with what was found in previous studies that used this combination in the approved indication \u201ctreatment of uncomplicated malaria\u201d . Sinus bA limitation of our study was the small number of children included. This population is at higher risk of treatment failure , 54, forP. vivax efficacy restrict follow-up to 28 days in order to minimize the contribution of relapses, considering CQ elimination half-life. We believe this can result in underestimation of resistance, which, in our study, could be as high as 30% based on the risk difference against an efficacious comparator. This difference highlights the need to develop better designs and strategies for measuring antimalarial efficacy against P. vivax.By not coadministering PQ, we were able to estimate CQR, which at 11.5%, considering concurrent blood drug levels criterion, is very high. This is important, as patients receiving CQ are at higher risk of recurring episodes from resistant parasites that arise from either recrudescence or relapse. These findings also suggest that the usual methods for measuring CQ efficacy are prone to underestimate the true CQR rates. Most studies of P. vivax in Latin America. We demonstrated that CQR is underestimated in the region and that a change in treatment policy should be considered. Recently Plasmodium vivax has received increased attention, requiring development of new strategies and tools [P. vivax infections is complex and should be based on different factors, including the prevalence of CQ-resistant parasites; the current treatment adopted for P. falciparum; efficacy of the partner drug; interaction of the ACT with PQ; and the potential impact of reducing transmission [P. falciparum and P. vivax, it is necessary to provide a more effective option and reduce the risk of treating P. falciparum with CQ due to misdiagnosis [This is the first trial to compare an ACT with CQ as blood schizontocidal against nd tools . The chosmission . In ordeiagnosis . We beliClinical Infectious Diseases online. Consisting of data provided by the author to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the author, so questions or comments should be addressed to the author.Supplementary materials are available at Funding. This study was funded by Sanofi. The study sponsor participated in the development the protocol, interpreted the data, helped write the report, and performed monitoring.Acknowledgements. We acknowledge Director-President of FMT-HVD Professor Maria Alecrim for the institutional support, all the personnel from the Malaria Laboratory for their contribution in running the trial, and the study monitor from Sanofi. We thank the patients and their families for taking part in the study. C. T. D. R. and M. V. G. L. are recipients of a grant from the Conselho Nacional de Desenvolvimento Cient\u00edfico e Tecnol\u00f3gico as Bolsistas de Produtividade and A. M. S. receives a short-term training grant from WHO\u2019s Special Programme for Research and Training in Tropical Diseases.Author contributions. A. M. S., V. L., C. D. T. R., and M. V. G. L. designed and coordinated the study. A. M. S., A. C. A., B. L. M., K. M., M. M. M., and M. C. M. participated in data collection. G. C. M., A. K., M. M. M., I. F., and J. L. F. V. participated in the laboratory analyses. A. M. S., V. L., and M. V. G. L. conducted the statistical analyses. A. M. S. wrote the first draft of the manuscript. All authors participated in data interpretation and approved the final version and take responsibility for accuracy and completeness of data reporting and for the decision to submit for publication.Potential conflicts of interest. V. L. is employed by Sanofi. All other authors report no conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed."}
+{"text": "Plasmodium falciparum malaria with prolonged activity would substantially advance malaria control. DSM265 is an experimental antimalarial that selectively inhibits the parasite dihydroorotate dehydrogenase. DSM265 shows in vitro activity against liver and blood stages of P falciparum. We assessed the prophylactic activity of DSM265 against controlled human malaria infection (CHMI).A drug for causal  prophylaxis of P falciparum sporozoites . An additional group received daily atovaquone-proguanil (250-100 mg) for 9 days, starting 1 day before CHMI (cohort 1B). Allocation to DSM265, atovaquone-proguanil, or placebo was randomised by an interactive web response system. Allocation to cohort 1A and 1B was open-label, within cohorts 1A and 2, allocation to DSM265 and placebo was double-blinded. All treatments were given orally. Volunteers were treated with an antimalarial on day 28, or when parasitaemic, as detected by thick blood smear (TBS) microscopy. The primary efficacy endpoint was time-to-parasitaemia, assessed by TBS. All participants receiving at least one dose of chemoprophylaxis or placebo were considered for safety, those receiving PfSPZ Challenge for efficacy analyses. Log-rank test was used to compare time-to-parasitemia between interventions. The trial was registered with ClinicalTrials.gov, number NCT02450578.At the Institute of Tropical Medicine, Eberhard Karls University , healthy, malaria-naive adults were allocated to receive 400 mg DSM265 or placebo either 1 day (cohort 1A) or 7 days (cohort 2) before CHMI by direct venous inoculation (DVI) of 3200 aseptic, purified, cryopreserved 22 participants were enrolled between Oct 23, 2015, and Jan 18, 2016. Five participants received 400 mg DSM265 and two participants received placebo 1 day before CHMI (cohort 1A), six participants received daily atovaquone-proguanil 1 day before CHMI (cohort 1B), and six participants received 400 mg DSM265 and two participants received placebo 7 days before CHMI (cohort 2). Five of five participants receiving DSM265 1 day before CHMI and six of six in the atovaquone-proguanil cohort were protected, whereas placebo recipients (two of two) developed malaria on days 11 and 14. When given 7 days before CHMI, three of six volunteers receiving DSM265 became TBS positive on days 11, 13, and 24. The remaining three DSM265-treated, TBS-negative participants of cohort 2 developed transient submicroscopic parasitaemia. Both participants receiving placebo 7 days before CHMI became TBS positive on day 11. The only possible DSM265-related adverse event was a moderate transient elevation in serum bilirubin in one participant.A single dose of 400 mg DSM265 was well tolerated and had causal prophylactic activity when given 1 day before CHMI. Future trials are needed to investigate further the use of DSM265 for the prophylaxis of malaria.Global Health Innovative Technology Fund, Wellcome Trust, Bill & Melinda Gates Foundation through Medicines for Malaria Venture, and the German Center for Infection Research. Despite all the success achieved in reducing the burden of malaria, in 2015 nearly half a million people died from the disease.More than 100 million travellers from non-tropical regions visit malaria-endemic countries every year, of which an estimated 30 million travellers are at risk of developing malaria, and about 30\u2008000 acquire the disease.Evidence before this studyClinicalTrials.gov database on the same day with the same term and found six clinical trial registrations: two in Australia , two in the USA, one in Peru, and the reported trial in Germany. None of these studies have yet reported results.DSM265 is a selective inhibitor of the plasmodial dihydroorotate dehydrogenase. On Jan 30, 2017, we did an unrestricted PubMed database search with the term \u201cDSM265\u201d and found nine publications: four reviews and five original articles, none of which were a clinical trial. One study reported the preclinical evaluation of DSM265. We searched the Added value of this studyPlasmodium falciparum sporozoites .To our knowledge, this study is the first to report use of DSM265 for prophylaxis of malaria in human beings. Besides safety, tolerability, and pharmacokinetics of a single dose of 400 mg DSM265, protective efficacy was assessed using controlled human malaria infection (CHMI). To our knowledge, it is also the first time that a chemoprotective regimen was assessed using the highly standardised CHMI model with purified, cryopreserved Implications of all the available evidenceP falciparum malaria when given 1 day before direct venous inoculation of PfSPZ Challenge. Efficacy was lower when DSM265 was given 7 days before direct venous inoculation of PfSPZ Challenge but partial activity against the asexual blood stage was maintained. Future trials are needed to optimise DSM265 formulation, dose, and schedule, and assess its activity against naturally acquired P falciparum malaria.Our study shows that a single dose of 400 mg DSM265 is a well tolerated and efficacious antimalarial for causal prophylaxis of The target candidate and product profiles for antimalarials were recently updated with particular emphasis on elimination and eradication scenarios,Plasmodium falciparum sporozoites  facilitates studies with the required level of standardisation and flexibility for exploratory early phase clinical trials.The development of new clinical study designs that allow the evaluation of chemoprophylactic interventions, including efficacy early in clinical development are of paramount importance. Applying such an improved design, the selection of the most promising drug candidate for further development in large, resource intensive licensure programmes would be more stringent and fast. The recent development of controlled human malaria infection (CHMI) models using aseptic, purified, cryopreserved P falciparum parasites. It is a novel, orally active plasmodium-selective dihydroorotate dehydrogenase (DHODH) inhibitorP falciparum, unlike most human tissues, lacks salvage pathways that could serve as an alternative source of pyrimidines.P falciparum life-cycle stage, and against asexual blood stages.P falciparum and Plasmodium vivax malaria .DSM265 is the most advanced of a new generation of antimalarial drugs with the potential to prevent infection with P falciparum sporozoites .The aim of our study was to assess the safety, tolerability, and pharmacokinetics of DSM265 as well as the chemoprophylactic efficacy of DSM265 by CHMI, using direct venous inoculation (DVI) of 3200 P falciparum sporozoites by PfSPZ Challenge. The day of DVI for CHMI was defined as day 0 for all study procedures .This single centre, double-blind, randomised, placebo-controlled phase 1 clinical trial recruited two sequential cohorts : cohort Recruitment for cohort 2 depended on at least partial efficacy of DSM265 in cohort 1A , and on a positive safety review by a safety review team. The last scheduled follow-up visit was on day 60 following DVI for CHMI (day 60).2 and 30 kg/m2 were included. All participants were required to practice effective contraception, and female participants were to provide a negative pregnancy test. Further inclusion criteria were: no clinically significant findings in history, physical, and laboratory examination, being reachable at all times by mobile phone during the whole study, agreement to share medical information on the volunteer with his or her general practitioner, and understanding of study procedures and risks, assessed by a multiple choice test. Additionally, willingness to undergo CHMI with PfSPZ Challenge, to take a curative regimen if necessary, and being able to comply with all study requirements (in the investigator's opinion) were also required.The study population was selected to represent healthy, malaria-naive adults. Volunteers aged 18\u201345 years with a body-mass index (BMI) between 18 kg/mExclusion criteria were: a history of malaria or plans to travel to endemic regions during the study, participation in another clinical trial within 30 days before enrolment or during the study, previous participation in a malaria vaccine trial, history of serious psychiatric conditions, convulsions, or severe head trauma, any malignancy, and diabetes mellitus. Moreover, arrhythmias, prolonged QTc interval (>450 ms), or any other clinically significant abnormalities in the electrocardiogram, breast feeding, or intention to become pregnant, HIV, hepatitis B or C virus infection, any suspected immunodeficient state, history of splenectomy, and haemoglobinopathies also prevented participation. Further exclusion criteria were smoking more than ten cigarettes or equivalent per day, alcohol or injected drug abuse, moderate risk or higher categories for cardiovascular events within 5 years of the study commencement date, use of any prescription drugs (except contraception), herbal supplements, or over-the-counter medication within 2 weeks before initial dosing, and intake of grapefruit, grapefruit juice, Seville orange, or other products containing these ingredients within 7 days of the first drug administration of DSM265 or placebo. Any symptoms, physical signs, and laboratory values suggestive of systemic disorders, or of conditions that could interfere with the interpretation of the study results, or compromise the health of the volunteers excluded participation. The full study protocol with further details is available from the corresponding author upon request. Eligibility criteria were assessed after written informed consent was given.The study was done at the clinical trial platform of the Institute for Tropical Medicine in T\u00fcbingen, Germany, and received approval from the ethics committee of the Eberhard Karls University and the University Clinics T\u00fcbingen. The study was compliant with the International Council for Harmonisation Good Clinical Practice guidelines and the German Medicines Law.Randomisation for cohort 1 and cohort 2 was done by an interactive web response system service provided by an external contract research organisation, which attributed a volunteer identification code to each eligible volunteer using a computer-generated  allocation sequence without restrictions. In cohort 1, allocation to cohort 1A (DSM265 and placebo) or 1B (atovaquone-proguanil) was open label. Nested in cohort 1A, allocation to DSM265 or placebo was double-blinded as it was in cohort 2. Clinical team, funder, and volunteers remained blinded until database lock. Only the study pharmacist received the randomisation list in a sealed envelope on the day of DSM265 administration and had no other role in the trial. The allocation ratio for DSM265 to placebo was 3:1. A higher proportion of DSM265 against placebo recipients was chosen to increase information on efficacy, safety, and tolerability of DSM265. The atovaquone\u2013proguanil group served as an additional control.Placebo was taste-blended, and visually and haptically indistinguishable from DSM265. Following preparation by the study pharmacist, the product was transferred to the clinical department by a blinded courier. Volunteers were required to ingest the DSM265 or placebo suspension under observation of at least two investigators. All containers used were identical, and were marked with the volunteer identification code. First unblinding was done following an interim database lock after day 28 of cohort 1A to allow assessment of efficacy by an independent statistician. The sponsor and investigators remained blinded until completion of follow-up (day 60). Cohort 2 was unblinded after final database lock.DSM265  was supplied as a 25% (250 mg/g) spray-dried dispersion. The powder was reconstituted in 240 mL of vehicle  on day of dosing. The placebo  was supplied as hydroxypropylmethylcellulose acetate succinate powder in a bottle and reconstituted exactly as DSM265. DSM265 and placebo were given orally as liquid emulsion.Volunteers from cohort 1A and cohort 2 were dosed after fasting for at least 4 h, followed by a standardised breakfast after drug intake. Meal records were taken until day 28 of CHMI. In cohort 1B, atovaquone-proguanil  was given daily for 9 days together with a fat-containing snack or drink, as recommended on the package insert. Blood for pharmacokinetic analyses was sampled at baseline, 15 min, 30 min, 60 min, and 2 h, 4 h, 6 h, 8 h, 14 h, 24 h, 48 h, 72 h, 168 h, 264 h, and 480 h after dosing in cohort 1A and cohort 2 for pharmacokinetic analyses. DSM265 and DSM450 (the main metabolite) concentrations in blood were measured from plasma and dried blood spots by liquid chromatography-tandem mass spectrometry at Swiss BioQuant , according to standard operating procedures (details available upon request from the corresponding author).P falciparum sporozoites, strain NF54  as previously described.CHMI was initiated by DVI of 3200 purified, cryopreserved Adverse events and concomitant medications were captured during all on-site visits and evaluated by the investigators. From day 6 following DVI onwards, blood for parasitological investigations was sampled daily until day 28. In volunteers who became TBS positive, daily sampling was continued until two consecutive TBS were negative. Additionally, volunteers were contacted by telephone at prespecified intervals. The clinical team was available at all times for the duration of the trial. Biochemistry, haematology, and urine analysis were scheduled regularly , and a 12-lead electrocardiogram (ECG) was done before, at 8 h and 24 h after dosing, before malaria treatment, and on day 28 and day 60 of CHMI.The primary efficacy endpoint was the prepatent period in days defined as time from DVI of PfSPZ Challenge (day 0) to parasitaemia, detected by TBS. Secondary outcome measures were number and severity of adverse events, the pharmacokinetic , and pharmacodynamics profile (parasitaemia over time).P falciparum sporozoites of PfSPZ Challenge) in malaria-naive volunteers is 100% with 56 of 56 exposed volunteers enrolled in independent studies at the time of writing the protocol (exact one-sided 95% CI 0\u00b795\u20131).max) and time (tmax) at peak levels were read from the concentration-over-time plot and area under the curve (AUC) was calculated using the linear trapezoidal rule. The safety review team reviewed safety, pharmacokinetic, and pharmacodynamics data on completion of the CHMI in each cohort (day 28) and decided on trial continuation or discontinuation, serious adverse events, or any other crucial findings. The Wilcoxon-Mann-Whitney test was used for comparisons of non-parametric variables. Log-rank test was used to compare time-to-parasitaemia between placebo and DSM265 treatments. All analyses were coded with R,ClinicalTrials.gov, number NCT02450578.The sample size of this study reflects the exploratory nature of the trial. Sample size was selected to demonstrate at least 61% protective efficacy with 95% probability when full protection (six of six) is achieved (exact one-sided 95% CI 0\u00b761\u20131). Infectivity of the inoculum  of 40 screened volunteers were eligible. Volunteers were allocated to receive placebo, atovaquone-proguanil, or DSM265, in cohort 1, and placebo or DSM265 in cohort 2 . One parIn cohort 1 all volunteers treated with DSM265 and atovaquone-proguanil were fully protected; no parasitaemia in TBS and qPCR occurred during follow-up. All volunteers (n=4) allocated to placebo in cohort 1A and cohort 2 developed parasitaemia with a geometric mean time-to-parasitaemia by TBS microscopy (prepatent period) of 11\u00b77 days . The geometric mean of time-to-parasitaemia by qPCR was 7\u00b78 days .Cohort 2 volunteers were partly protected : three oAll TBS-positive volunteers received a full therapeutic course of atovaquone-proguanil (1000-400 mg), once daily for 3 days, starting on the day of TBS positivity. Treatment was successful in all cases, and parasitaemic volunteers recovered without sequelae.All seven volunteers in cohort 1A experienced adverse events: there were 41 grade 1, three grade 2, one grade 3, and two grade 4 events. The only possible DSM265-related adverse event was an episode of transient increase in total, direct, and indirect serum bilirubin 2\u20138 days following dosing. A similar transient elevation was observed as a baseline (pre-treatment) finding in the same participant. During follow-up, one serious adverse event occurred in cohort 1A: a bilateral pulmonary embolism occurred in a non-smoking woman under oral contraceptive treatment 34 days after DSM265 administration while she was on a transatlantic flight. The volunteer never became parasitaemic and recovered fully, except for a grade 1 sinus tachycardia, which still persisted at the last follow-up visit.In cohort 1B, 20 grade 1 and one grade 2 adverse events occurred in five of six volunteers. Five adverse events were attributed to atovaquone-proguanil; all grade 1 in severity . No adverse events were attributed to malaria.In cohort 2, 108 adverse events were reported in eight volunteers: 81 grade 1, 19 grade 2, six grade 3, and two grade 4 events. The most frequent adverse events were fatigue  and headache (15 episodes in six volunteers: ten grade 1 and five grade 2), followed by fever . 60 adverse events were attributed to malaria. The only two severe adverse events (grade 3 and 4) not attributed to malaria were laboratory abnormalities  without clinical signs, most likely due to sampling error. DSM265-related adverse events were not reported in this cohort.Altogether, 58 adverse events  occurred in the placebo recipients in both cohorts. 29 of 58 events were associated with malaria or with antimalarial treatment. The most frequent adverse events were headache (11 episodes in four volunteers: eight grade 1 and three grade 2) followed by fatigue , and fever (six episodes in two volunteers: three grade 1 and three grade 2). Drug-related adverse events were observed after placebo administration in three of four participants: insomnia (one event in one volunteer), abdominal discomfort (two events in two volunteers), palpitations (three events in one volunteer), dizziness (two events in one volunteer), and headache (one event in one volunteer).QTc prolongation was not observed in the DSM265-treated participants. Treatment-emergent adverse events are listed in the In pharmacokinetics analysis, the concentration of DSM265 and DSM450 sampled from plasma and dried blood spots were in agreement, although the concentration of both molecules was systematically lower in dried blood spots over the whole concentration range, probably because of the paper matrix . Therefomax was 12\u00b71 \u03bcg/mL (IQR 9\u00b76\u201313\u00b78), median tmax was 3 h (2\u201312). The median plasma elimination half-life for DSM265 was 137\u00b75 h (IQR 95\u00b71\u2013147\u00b75). AUC0\u2013\u221e values in plasma ranged from 1080 \u03bcg\u00b7h/mL to 2460 \u03bcg\u00b7h/mL (median 1745 \u03bcg\u00b7h/mL [IQR 1442\u20132060]). Median AUC0\u2013168 was 1031 \u03bcg\u00b7h/mL (IQR 867\u20131165). The pharmacokinetics of DSM265 and DSM450 showed a similar profile in cohort 1A and cohort 2  in TBS-negative  compared with TBS-positive infections . The concentration of DSM265 was particularly low in two participants with similar parasite kinetics to placebo .P falciparum malaria when given 1 day before PfSPZ Challenge inoculation. DSM265 is currently being developed as a drug for antimalarial treatment and chemoprophylaxis.Our study shows that a single dose of 400 mg DSM265 was well tolerated and efficacious for causal chemoprophylaxis against A limitation of such studies is the low sample size that misses small differences and rare safety issues. When unreplaceable dropouts occur, as in cohort 1A of this study, power further decreases. Other limitations of CHMI studies are related to their model character and the study population. CHMI is optimised to infect all inoculated volunteers, which is more stringent than in naturally acquired infection, in which only a fraction of bites of infected mosquitoes will lead to malaria.DSM265 was safe and well tolerated in this study's setting, with no clinically significant changes in vital signs, ECGs, or laboratory tests following oral dosing of 400 mg DSM265. The only serious adverse event, a bilateral pulmonary embolism, was not considered to be related to the investigational product, or any other study procedures such as PfSPZ Challenge or antimalarial rescue treatment. The long intercontinental flight and concomitant intake of oral contraceptives, two multiplicative risk factors for thromboembolic events, were the likely triggers for this event.21A preliminary good safety profile makes DSM265 a highly promising candidate for further development as a chemoprophylactic antimalarial. Given 1 day before CHMI, all participants were fully protected. This action contrasts with chemoprophylaxis with several approved drugs that only act against the asexual blood stage . Here, parasites must egress from the liver before they become susceptible to the drug. Other drugs such as the artemisinins have extremely short half-lives and offer no protection beyond a few hours.When given 7 days before CHMI (cohort 2), DSM265 efficacy was reduced. Prevention of malaria depended on the half-life of DSM265 and the concentration at the moment of CHMI, whereas the pre-erythrocytic development of the parasite was not sufficiently suppressed; all volunteers developed parasitaemia. Nevertheless, in three of six participants asexual blood stage growth was suppressed. Furthermore, we observed substantial variation in the time to peak plasma concentration. Despite fasting before and standardised food intake following DSM265 administration, a food effect cannot be excluded since drinking was not restricted following DSM265 intake.Despite the small sample size, this study provides important information on the potential of DSM265 both in chemoprotection and in malaria treatment. Our study suggests that gains in formulation, dosing, and schedule that improve consistency and duration of exposure might translate to longer protection. Since no evidence for drug-related toxicity has been observed, a further dose increase can be envisaged, which we predict could prolong protective efficacy.P falciparum malaria that is correlated with plasma exposure of the compound.To our knowledge, this is the first time that standardised CHMI using DVI of PfSPZ has been used to assess chemoprotective properties of a new chemical entity in early development. We show that DSM265 is well tolerated and has causal prophylactic activity against This online publication has been corrected. The corrected version first appeared at thelancet.com/infection on May 23, 2017."}
+{"text": "Plasmodium falciparum gametocytes that might be present at the time of treatment.Substantial gains have been made in reducing the global burden of malaria, much of which can be attributed to greater access to prompt diagnosis and highly effective treatment. However, as endemic countries commit to eliminating malaria, more aggressive interventions are needed to target the large number of apparently healthy individuals who harbour transmissible malaria parasites. Although most national antimalarial guidelines recommend artemisinin combination therapy for the management of uncomplicated falciparum malaria, chemopreventive strategies have generally adopted non-artemisinin combination therapy regimens such as sulfadoxine-pyrimethamine and amodiaquine. Artemisinin and its derivatives reduce carriage of sexual stages of the malaria parasites (gametocytes) that are infectious to the mosquito vector, but neither artemisinin combination therapies nor sulfadoxine-pyrimethamine and amodiaquine prevent transmission from fully mature P falciparum gametocytes. Although primaquine can induce haemolysis in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency, a single low dose of 0\u00b725 mg/kg is safe and well tolerated, even in G6PD-deficient individuals, and highly effective in reducing transmissibility.Primaquine has potent activity against mature The Lancet Infectious Diseases, Alassane Dicko and colleaguesP falciparum transmissibility. 40 G6PD-normal asymptomatic Malian participants were randomly assigned to receive either sulfadoxine-pyrimethamine and amodiaquine , with or without primaquine . Another 40 participants were randomly assigned to receive dihydroartemisinin-piperaquine with or without methylene blue (15 mg/kg per day for 3 days). Transmissibility was assessed by molecular quantification of sexual stage-specific mRNAs and by membrane feeding blood to mosquitos and counting the oocytes that formed. Both primaquine and methylene blue were highly effective in reducing gametocytaemia and preventing transmissibility within 2 days of starting treatment.In P falciparum gametocytocidal drug in vivo, as suggested by earlier studies. Although the study population was limited to male participants who were G6PD-normal, previous studies have shown that a single low dose of primaquine (0\u00b725 mg/kg) is safe in people with moderate severity G6PD deficiency (G6PD-Mahidol), and that a 3-day regimen of methylene blue was also safe in the generally less severe G6PD A-variant prevalent in Africa.7This small yet detailed studyP falciparum has emerged in the Greater Mekong subregion and is spreading.P falciparumP falciparum.A single low dose of primaquine is easy to administer, safe, efficacious, and inexpensive. So is there need for further exploration of an alternative gametocytocidal agent? Reliance on a single therapeutic intervention to reduce mosquito infectivity is risky. Artemisinin-resistant"}
+{"text": "Malaria remains one of the most important causes of morbidity and mortality in pregnant women and their newborn babies in sub-Saharan Africa. Intermittent preventive treatment in pregnancy (IPTp) is recommended by the World Health Organization (WHO) to reduce the burden of disease and improve maternal and neonatal survival and general health. Due to the growing resistance to sulfadoxine-pyrimethamine (SP), the current WHO-recommended drug for IPTp, identification of new and effective drugs is an urgent priority.Plasmodium falciparum parasitemia in their first or second pregnancy. Parasitemia was determined by microscopy and molecular genotyping was performed to characterize parasites relative to the baseline infection. Weekly follow-up visits took place until day 42 after first dose and additional follow-up occurred after delivery. Systemic concentrations of azithromycin (AZ), chloroquine (CQ), and the CQ metabolite, desethyl CQ (DECQ) were evaluated at Day 0 (pre-dose), at Day 2  and randomly at Days 7 and 14. Systemic concentrations of CQ and DECQ were also measured randomly at Day 21 and Day 28. In total, 404 women were screened for eligibility and 168 were treated, 155 of whom completed the study. PCR-adjusted parasitological response in the modified intent-to-treat population at day 28 (the primary efficacy endpoint) was estimated by the Kaplan-Meier method as 99.35% . PCR-adjusted parasitological response remained high at day 42 . In general, the mean concentrations of serum AZ, plasma CQ, and plasma DECQ showed large CV% values . There were 157 live births, three stillbirths, and eight pregnancies of unknown outcome: 7 due to withdrawal of participant consent and 1 lost to follow-up. The most frequent treatment-emergent adverse events were vomiting (20.8%) and dizziness (19.6%).This was an open-label, non-comparative study (NCT01103713) in 5 countries in East and sub-Saharan Africa  to assess parasitological response and drug concentrations of a single, 3-day course of four tablets of a fixed-dose combination of azithromycin-chloroquine (AZCQ) 250/155 mg given during the second or third trimester to women with asymptomatic These results suggest that a 3-day course of AZCQ can lead to an adequate 28-day parasitological response. Plasmodium species that are transmitted when infected female anopheline mosquitoes bite a human host to consume a blood meal [Malaria is a serious infection caused by protozoan parasites of the ood meal . The chaood meal . Asymptoood meal .Although malaria cases occur in all the major tropical and sub-tropical regions of the world, Africa has the greatest number of people at risk of contracting the disease and has the greatest number of deaths related to malaria . The WorPregnant women are more susceptible to malaria, and susceptibility is greatest in the first and second pregnancy \u20137. Malarp\u22640.001) [Intermittent preventive treatment in pregnancy (IPTp) is an important means of reducing the detrimental health impact of malaria on pregnant mothers and their unborn children . The WHOp\u22640.001) . An addip\u22640.001) \u201315. TherNeisseria gonorrhoeae and Chlamydia trachomatis infections [in vitro and in animal model studies [The combination of azithromycin (AZ) and chloroquine (CQ) was evaluated as a potential alternative to SP for IPTp in another study (NCT01103063) . AZ and fections . The com studies ,18.Co-administration of AZ and CQ has demonstrated efficacy, safety, and tolerability in the treatment of symptomatic uncomplicated malaria in two multi-country Phase 3 clinical studies in non-pregnant adults (NCT00082576 and NCT00367653)  and one Plasmodium falciparum parasitaemia, in their first or second pregnancy, in sub-Saharan Africa. The aim of this study was to determine if AZCQ could clear the malaria parasites from the blood during pregnancy with a similar efficacy as that demonstrated in previous adult treatment studies. However, unlike typical treatment studies that rely on demonstrating adequate clinical and parasitological response as a primary endpoint [The study we describe here (NCT01103713) was designed to assess weekly parasitological response and drug concentrations of a single fixed-dose combination of AZCQ administered over 3 days in women with asymptomatic endpoint , the triP. falciparum parasitemia . Study participants received a single 3-day course of AZCQ IPTp therapy after providing informed consent.This was an open-label, non-comparative out-patient study in primi- or secundigravidae women \u226516 to \u226435 years of age in the second or third trimesters of pregnancy with asymptomatic peripheral Women were excluded if they were revealed to have multiple gestations as per ultrasound screening, had any chronic illness that might have adversely affected fetal growth or viability, or had evidence of current obstetric complications that could adversely impact the pregnancy or fetal outcomes, including presence of congenital anomalies, placenta previa, or abruption. Additionally, women were excluded if they had clinical signs and symptoms of malaria, history of fever within prior 24 h, baseline hemoglobin <8 g/dl, or had used antimalarial drugs in the previous 4 weeks. An inability to tolerate oral treatment in tablet form, or a known allergy to the study drugs  or to any macrolides or sulfonamides, use of any medication that may have interfered with the evaluation of the study drug or that was contra-indicated during pregnancy, a current history of smoking or alcohol/drug abuse, or severe acute or chronic medical or psychiatric condition or laboratory abnormality  were also reasons for exclusion.\u00ae 500 mg) and CQ  [A fixed-dose AZCQ combination (four fixed dose combination tablets of AZ 250 mg/CQ 155 mg) was used in this study . Two tab 300 mg) ). The fiThe study was conducted in accordance with the Declaration of Helsinki on Ethical Principles for Medical Research Involving Human Subjects, adopted by the General Assembly of the World Medical Association (1996), and was conducted in accordance with the International Conference on Harmonization (ICH) guideline on Good Clinical Practice (GCP), and applicable local regulatory requirements and laws. The study protocol was approved by the Independent Ethics Committee (IEC) of London School of Hygiene and Tropical Medicine and the following local Institutional Review Board (IRB)/IECs: the Comit\u00e9 National d\u2019Ethique pour la recherch\u00e9 en Sante in Cotonou, Benin; the Kenyatta National Hospital\u2014University of Nairobi Ethics Review Committee in Nairobi, Kenya; the College of Medicine Research and Ethics Committee in Blantyre, Malawi; the Medical Research Coordinating Committee in Dar es Salaam, Tanzania; and the Uganda National Council of Science and Technology, the School of Medicine Research and Ethics Committee of Makerere University, and the Mulago Hospital Research and Ethics Committee in Kampala, Uganda. An independent External Data Monitoring Committee (EDMC) oversaw the conduct of the study. Written informed consent (or assent from participants <18 years of age) was obtained from all study participants or their legally-acceptable representatives prior to any study-related procedure.P. falciparum count 80\u2013100,000 \u03bcl upon examination of 100 high power fields on thick smear. A blood slide was considered negative when the examination of 100 high power fields on the thick smear do not show the presence of any P. falciparum parasites.Parasitological response was assessed by analysis of peripheral blood smears (thick and thin) stained by standard Giemsa staining for parasite identification. Parasite numbers were counted using the white blood cell counting method on thick smears. Each blood smear was read by two microscopists blinded to the reading of each other. In the event of any discrepancies, a third microscopist read the blood smear and the average of the two most concordant results was used. A positive blood smear had a P. falciparum parasites [P. falciparum after parasitological cure of the baseline infection. Conversely, analysis of PCR-unadjusted data made no distinction between reappearance of the baseline infection or re-infection with a genotypically distinct parasite, both of which would be considered treatment failures. The primary endpoint was the PCR-adjusted parasitological cure rate on day 28 following a single 3-day dosing regimen of AZCQ in asymptomatic pregnant women with P. falciparum parasitemia. Secondary endpoints included PCR-adjusted and unadjusted parasitological cure rates on days 7, 14, 21, 28 (unadjusted), 35, and 42 post first dose of study drug, and parasite counts  at each visit.Molecular testing by polymerase chain reaction (PCR) was used to verify the genotype of asexual arasites . Analysiin utero were recorded. An AE with onset after the first dose of study drug was to be counted as a treatment emergent adverse event (TEAE) if it occurred on or before 35 days post last dose. If the onset was prior to the first dose of study drug and the severity increased thereafter, the event was also to be counted as a TEAE.Adverse events (AEs), reported in mothers and neonates, were analyzed. Additionally, temperature, physical examinations, hemoglobin concentrations, and exposure P. falciparum monoinfection. The modified ITT (MITT) comprised a subset of the ITT population who had P. falciparum monoinfection (confirmed by microscopy) with parasite counts in the range of 80\u2013100,000/\u03bcl on thick blood smears. The per protocol (PP) analysis population comprised a subset of MITT participants who received all 3 days of study drug. Safety analysis data included all mothers who received at least one dose of study drug, while for neonates it consisted of all liveborn babies. Parasite counts were summarized descriptively using the sample mean, standard deviation (SD), standard error (SE), median, minimum, and maximum. Primary and secondary parasitological endpoints were estimated from the Kaplan-Meier (KM) curve , with corresponding 95% confidence interval (CI), based on time to the first occurrence of parasitological failure. If, at a given time point, no parasitological failures had yet occurred, resulting in a missing variance estimate, an exact CI derived from the binomial distribution was computed. A subject was a parasitological responder if she had a zero parasite count on the day 7 visit without subsequent recurrence through the day of consideration (PCR adjusted or unadjusted as applicable), otherwise she was a parasitological failure. Participants who did not experience the defined event  were considered censored. Crude incidence of parasite cure was also calculated for days 28 and 42. A sensitivity analysis was also performed considering study discontinuations as failures. Data were pooled across all sites for analysis. No statistical hypothesis was tested.Study populations were defined as follows. The intent to treat (ITT) analysis population consisted of all participants who received at least one dose of study drug and who had baseline blood smears positive for Concentrations of serum AZ, plasma CQ, and the CQ metabolite, desethyl-CQ (DECQ) were evaluated with sparse pharmacokinetic sampling on day 0 predose, day 2 predose, 2 h (as close to 2 h as possible), and 8 h (time window: 4\u201312 h) post dose, and at a random time point on days 7 and 14. In addition, due to the long half-life of CQ, plasma concentrations of CQ and DECQ were measured at a random time point on days 21 and 28.Blood samples were centrifuged at 1700 rpm for 10 minutes at 4\u00b0C. Serum for AZ or plasma for CQ and DECQ samples were stored within 1 hour of collection at -20\u00b0C until shipped for assay. AZ, CQ, and DECQ concentrations were analyzed using validated, sensitive, and specific high-performance liquid chromatography tandem mass spectrometric methods in compliance with Pfizer standard operating procedures. AZ assays were performed by Bioanalytical Laboratory (BASi) (USA) and CQ/DECQ assays were performed by Bioanalytical Laboratory WuXi appTec (China).The study was conducted between March 2011 and October 2013 in six study sites in five countries: Benin, Kenya, Malawi, Tanzania, and Uganda. In total, 404 women were screened, of whom, 168 were treated and 155 completed the study . The sitP. falciparum monoinfection (confirmed by microscopy) with parasite counts between 80 and 100,000 per \u03bcl  and who received at least one dose of study drug. In addition, two participants were found to have been improperly consented and were, therefore, excluded from the efficacy analysis. The PP analysis set comprised 158 participants in the MITT population who received all three daily doses of study drug.Reasons for discontinuation included unwillingness to continue study participation (eight women), lost to follow-up (two women), termination of the study (two women), and another due to relocation (one woman). The safety analysis set comprised all 168 women who were treated and 157 live-born neonates. There were three stillbirths recorded, and eight pregnancy outcomes were not recorded because 7 participants withdrew consent and 1 was lost to follow-up. Drug concentrations were assessed from samples from 166 participants. The MITT population comprised 163 women who had a Baseline demographics for the safety population are shown in P. falciparum CQ resistance transporter (PfCRT) or P. falciparum multidrug resistance 1 (Pfmdr1) genes. The remaining 65 (39.4%) participants had wild-type infections.In the ITT population, 100 (60.6%) participants had infections with parasites carrying a mutant version of either one or both of the At day 28, 153 of 154 participants in the PP population had a PCR-adjusted parasitological response  . None of these were considered related to study drug.No mothers experienced serious AEs (SAEs) and nine SAEs were reported in neonates. TEAEs were reported in 92 (54.8%) mothers and 27 (17.2%) neonates. The most common TEAEs, those occurring in at least 5 mothers, are listed in Mean serum AZ concentrations were 194, 994, and 708 ng/ml at 0, 2, and 8 h on day 2, respectively, and 54.4 and 20.3 ng/ml on days 7 and 14, respectively . Mean plOf the 157 neonates, 66 (42.0%) were female and 91 58.0%) were male. The mean birth length (SD) was 46.3 cm (3.6) and the mean birth weight (SD) was 3022.2 g (494.5), while nine of the 137 with recorded birth weight (6.6%) had low birth weight . There were 144 (91.7%) normal newborns. Congenital malformations or anomalies were reported for two (1.3%) newborns (one hypospadias and one polydactyly), and other neonatal problems or abnormalities were reported for nine (5.7%) newborns. The outcome for two (1.3%) newborns was unknown  and dizziness (19.6%). After day 35, there were four neonate deaths due to SAEs, none of which were considered to be treatment-related.An accompanying Phase 3 pivotal clinical trial (NCT01103063) was designed to assess AZCQ versus SP for IPTp in East and Southern sub-Saharan Africa. This trial was terminated based on results of a pre-planned interim analysis, which showed that the futility boundary for efficacy had been crossed.Despite the failure of the pivotal Phase 3 trial, our findings showed that parasitological cure rates to a single, 3-day dose of AZCQ were high throughout the long duration (42 days) of the study, which was consistent with previous studies and further confirmed by sensitivity analyses in which study drop-outs were considered to be treatment failures.The reasons why this AZCQ IPTp treatment regimen failed to demonstrate superiority versus SP in the Phase 3 pivotal trial are beyond the scope of this article, but clearly, the data we present here did not appear to suggest that AZCQ IPTp treatment would not be a viable treatment option. Another potential alternative to SP IPTp, DHA-P, was recently described , 25. IntS1 File(DOCX)Click here for additional data file.S2 File(PDF)Click here for additional data file."}
+{"text": "Short Message Service (SMS) reminders have been suggested as a potential intervention for improving adherence to medications and health facility attendance.An open-label, randomized, controlled trial to test the efficacy of automated SMS reminders in improving adherence to artemether\u2013lumefantrine (AL) and post-treatment attendance in comparison with standard care was conducted at four health facilities in western Kenya. Children below five years of age with uncomplicated malaria were randomized to intervention (SMS reminders) or control groups. Within each study group they were further randomized to three categories, which determined the timing of home visits to measure adherence to complete AL course and to individual AL doses. A sub-set of caregivers was advised to return to the facility on day 3 and all were advised to return after 28\u00a0days. The primary outcomes were adherence to medication and return on day 3. The primary analysis was by intention-to-treat.Between 9 June, 2014 and 26 February, 2016, 1677 children were enrolled. Of 562 children visited at home on day 3, all AL doses were completed for 97.6% (282/289) of children in the control and 97.8% (267/273) in the intervention group . When correct timing in taking each dose was considered a criteria for adherence, 72.3% (209/289) were adherent in the control and 69.2% (189/273) in the intervention group . Sending SMS reminders significantly increased odds of children returning to the facility on day 3  and on day 28 .In this efficacy trial, SMS reminders increased post-treatment return to the health facility, but had no effect on AL adherence which was high in both control and intervention groups. Further effectiveness studies under the real world conditions are needed to determine the optimum role of SMS reminders.Trial registration ISRCTN39512726The online version of this article (doi:10.1186/s12936-017-1702-6) contains supplementary material, which is available to authorized users. The expansion of network coverage and mobile phone penetration in Africa  has offeMost SMS interventions in Africa have been assessed in the management of chronic diseases and long-term therapy, while their effects on the management of acute diseases, such as malaria, have been less commonly investigated . SMS remNon-adherence to anti-malarial medicines and lack of patients\u2019 follow-up compromises malaria case management and favours the emergence of anti-malarial resistance , 25. TheA randomized controlled trial was therefore undertaken in Kenya to assess whether SMS reminders sent to caregivers of children treated with nationally recommended ACT, artemether\u2013lumefantrine (AL), would enhance adherence to AL therapy and would increase the rates of post-treatment return to the health facility following completion of treatment.The trial was conducted at four public health facilities in Siaya County in western Kenya. Two trial sites are located in Bondo Sub-county  and two in Rarieda Sub-county . Malaria transmission in the study area is high with seasonal peaks in May\u2013July and October\u2013November . AL has The study was an open-label, randomized, controlled trial testing the additional effects of SMS reminders on patients\u2019 adherence to AL and their return to the health facility compared to the control group receiving standard care only. This was an efficacy trial where all children with uncomplicated malaria received care by study personnel in line with national guidelines , and witPlasmodium falciparum with parasitaemia between 500 and 200,000/\u03bcL; caregiver access to personal or shared mobile phone within household; caregiver ability to open and read SMS either themselves or through another person in the household; and, caregiver provision of written informed consent. The following exclusion criteria applied: presence of clinical danger signs, severe anaemia [haemoglobin (Hb)\u00a0<5\u00a0g/dl] or any other severe malaria criteria; severe malnutrition (weight for height\u00a0<70%); ongoing prophylaxis with drugs having anti-malarial activity such as cotrimoxazole; reported hypersensitivity to AL; presence of any concurrent illness; and, previous participation in the trial. Thick and thin blood smears were performed by the study microscopists, who counted asexual parasites per 200 white blood cells (WBCs) and calculated parasite density on the estimate of 8000\u00a0WBCs/\u03bcL. A blood smear was deemed negative only after examining 100 high-power microscopic fields. Hb levels were estimated using HemoCue.All children suspected of malaria were screened by study clinicians at outpatient departments in the study sites and enrolled into the trial if they met all of the following inclusion criteria: age six to 59\u00a0months; weight 5\u00a0kg and above; history of fever in the previous 24\u00a0h or presence of axillary temperature\u00a0\u226537.5\u00a0\u00b0C; microscopically confirmed infection of The randomization codes were generated by an offsite statistician and randomization numbers were applied in sequence of recruitment. The trial was open-label. Participants and nurses performing home visits could not be masked. The study personnel at health facilities were necessarily aware of categories for assigning day of home visits, but were blind to the intervention arm.In both arms children were treated with dispersible AL. The first dose was supervised at the health facility by a study nurse, and repeated if the child vomited within 30\u00a0min. The remaining five doses were taken at home. All caregivers received verbal instructions to give the second dose exactly 8\u00a0h after the first dose with the subsequent four doses given on the following 2\u00a0days at 08.00 in the morning and in the evening at 20.00, and illustrations on the AL blister packs that were taken home were used to support the explanations. Children weighing 5\u201314\u00a0kg were to take single tablet per dose while those weighing 15\u201324\u00a0kg two tablets per dose. The caregivers were advised to administer AL after a meal or with food, to complete all doses even if the child appeared better, and to return to the health facility immediately if the child\u2019s condition worsened.In total 11 text messages were used where content, timing, understanding, and distribution had undergone extensive pre-testing with community members, caregivers and patients at four facilities within the same county but outside of the study area . CaregivDuring recruitment, caregivers were informed that they would be visited at home but not informed of the specific day of the visit. They were advised to keep AL blister packs after completing the treatment course. Home visits were undertaken by a study nurse within 24\u00a0h of expected completion of the individual doses of interest . During the home visits, adherence to AL was assessed using pill counts and caregivers\u2019 reports to determine the number of pills taken prior to the visit and the timing of each dose. Caregivers were also asked whether they had received text message reminders. Caregivers who returned to the health facility on day 3 (category 1 and 2) were not financially compensated for transport costs as this was considered routine, however those who returned to the facility on day 28 received travel compensation of approximately 2 USD.Two primary outcomes were investigated in the trial. The first outcome was the proportion of patients adhering to the complete AL course (doses 2\u20136) measured in category 3 using the combination of pill count and self-reporting. The definition of correct AL adherence was based on two criteria of which both had to be met: (1) completion of all doses; and, (2) correct timing of all doses. To assess completion of all doses the evidence of empty AL blister pack during the home visit was used. In the absence of blister packs, caregivers\u2019 reports were used. To assess timing, caregivers\u2019 report of administered doses, within\u00a0\u00b11\u00a0h for dose 2 and\u00a0\u00b12\u00a0h for doses 3\u20136, compared to the instructions given at the time of recruitment, was classified as correct. The second primary outcome was the proportion of patients in combined categories 1 and 2 who returned to the health facility on day 3 after expected completion of AL treatment.Two secondary outcomes were also investigated in the trial. The first secondary outcome was the proportion of patients adhering to the individual AL doses measured within 24\u00a0h of expected dose administrations, i.e., for doses 2 and 3 in category 1, doses 4 and 5 in category 2, and for dose 6 in category 3. The adherence definitions for these patient groups followed the criteria of dose completion based on appropriate number of tablets found at the time of the visit and correct timing, using the same time allowances as described for the primary outcome. Finally, the last trial outcome was the proportion of patients across all categories that returned to the health facility on day 28.The sample size estimation was based on the first primary outcome  . AssuminThe primary analytic approach was intention-to-treat (ITT) including all randomized patients with available outcome data. The secondary analytic approach was per-protocol (PP) where adherence analyses excluded patients who were inadvertently recruited without meeting enrolment criteria and those who did not receive SMS reminders in the intervention group. SMS exposure was classified on the basis of a caregiver\u2019s report regarding delivery of any SMS reminder.p value. Participants\u2019 characteristics were tabulated by randomization group. To further assess potential confounders, multivariable regression was performed by examining each potential confounder as an independent variable with randomization group and retaining any of the covariates, which changed the unadjusted OR of randomization group by more than 5%.To estimate effects of the intervention on outcomes, mixed effects logistic regression models, with intervention arm as an independent variable adjusted for clustering by site, was used. The effect was expressed as an odds ratio (OR) with corresponding 95% confidence interval and All trial data were double entered, verified and cleaned in Access 2013 , and thereafter analysed in Stata version 12 .Between 9 June, 2014 and 26 February, 2016, 8600 children presenting with suspected malaria were screened, of whom 1677 children were enrolled into the trial. The most common reasons for screen failures were 5387 (78.6%) malaria test-negative patients, 1335 (19.3%) without access to SMS messages, and 788 (11.4%) unwilling to comply with study protocol over 28\u00a0days (Fig.\u00a0Adherence to complete AL course was measured among 562 patients in category 3 (i.e., having home visits on day 3, Table\u00a0Adherence to individual AL doses was measured in each of three categories of patients during home visits taking place within 24\u00a0h of expected completion of specific AL doses (Table\u00a0Patients\u2019 return to health facility was assessed by combining data in categories 1 and 2 for day 3 return and then by combined data across all categories for day 28 return Table\u00a0. Of 1032No covariate met the 5% inclusion criteria described above for either outcome of interest (Additional files This randomized controlled trial in western Kenya showed that SMS reminders significantly increased the rates of return to the health facility following anti-malarial treatment but did not have any effect on adherence to AL medications. The results of this efficacy trial comparing an SMS intervention to optimum clinical practices have several important implications.Nearly all children completed all AL doses in this trial, which indicates substantially higher adherence than previously observed in evaluations conducted under the routine conditions in the same area , 33, in Two other previous effectiveness trials tested effects of innovative SMS reminders on patients\u2019 adherence to anti-malarials and these showed discordant results, but lower overall adherence , 23. DueA recent review of anti-malarial adherence studies suggests that interactions between research teams or medical staff and patients is likely to influence adherence levels . SpecifiIn contrast to the findings on adherence, SMS reminders increased the rates of patients returning to health facilities for routine follow-up. Increasing the day 3 return from 74 to 81% and the day 28 return from 53 to 63% is a relatively modest but still significant impact on behaviour. Two trials in Kenya have shown similar effects of SMS on postpartum  and postWhen optimum care under the trial conditions is provided, text-message reminders can increase a child\u2019s return to the health facility following anti-malarial treatment, without an additional effect on already high levels of AL adherence that occur under trial conditions. Further effectiveness studies under varying real world conditions in different settings are needed to determine the optimum role of text-message reminders in improving patients\u2019 adherence to anti-malarial medicines."}
+{"text": "Plasmodium vivax relapse. Tafenoquine has a long half-life and the potential for more convenient dosing, compared with the currently recommended 14-day primaquine regimen.Tafenoquine is an investigational 8-aminoquinoline for the prevention of P. vivax were randomized (2:1) to tafenoquine 400 mg once daily for 3 days or 2500 mg total dose chloroquine phosphate (1500 mg chloroquine base) given over 3 days plus primaquine 15 mg daily for 14 days. Patients were followed to day 120.This randomized, active-control, double-blind trial was conducted in Bangkok, Thailand. Seventy patients with microscopically confirmed Day 28 adequate clinical response rate in the per-protocol population was 93% (40/43) (90%CI 83\u201398%) with tafenoquine, and 100% (22/22) (90%CI 87\u2013100%) with chloroquine/primaquine. Day 120 relapse prevention was 100% (35/35) with tafenoquine (90%CI 92\u2013100%), and 95% (19/20) (90%CI 78\u2013100%) with chloroquine/primaquine. Mean (SD) parasite, gametocyte and fever clearance times with tafenoquine were 82.5 h (32.3), 49.1 h (33.0), and 41.1 h (31.4) versus 40.0 h (15.7), 22.7 h (16.4), and 24.7 h (17.7) with chloroquine/primaquine, respectively. Peak methemoglobin was 1.4\u201325.6%  in the tafenoquine arm, and 0.5\u20135.9%  in the chloroquine/primaquine arm. There were no clinical symptoms of methemoglobinemia in any patient.P. vivax malaria. Also, monotherapy increases the potential risk of resistance developing to this long-acting agent. Clinical trials of single-dose tafenoquine 300 mg combined with standard 3-day chloroquine or artemisinin-based combination therapy are ongoing.Although there was no difference in efficacy in this study, the slow rate of parasite, gametocyte and fever clearance indicates that tafenoquine should not be used as monotherapy for radical cure of NCT01290601Clinicaltrials.gov  Plasmodium vivax causes around half of all malaria cases occurring outside Africa with an estimated 2.5 billion persons at risk of infection ). The World Health Organization recommended dosing regimen is standard blood schizonticidal therapy for 3 days plus primaquine 0.25\u20130.5 mg/kg bodyweight daily for 14 days . HoweverP. vivax relapse. The current investigational regimen is tafenoquine 300 mg single dose plus standard 3-day chloroquine [Tafenoquine is an 8-aminoquinoline discovered and developed by the Walter Reed Army Institute of Research, now being co-developed by GlaxoSmithKline and Medicines for Malaria Venture (MMV) for the prevention of oroquine .P. vivax treatment and prophylaxis [P. falciparum or P. vivax. This Phase II study evaluated the efficacy of high-dose tafenoquine monotherapy (1200 mg over three days) for the radical cure of P. vivax malaria. The study was conducted under an NIH Challenge Grant \u2018Tafenoquine, a Novel Drug for Malaria Prevention and Control\u2019.There have been many previous studies investigating tafenoquine at various doses and regimens for phylaxis \u201316. Howephylaxis , 18, andBoth 8-aminoquinolines, primaquine and tafenoquine, cause hemolysis in glucose-6-phosphate dehydrogenase (G6PD)-deficient individuals , 20. Botrd April 2003, the Ministry of Public Health, Nonthaburi, Thailand, and the Human Subjects Research Review Board, U.S. Army Medical Research and Material Command, Office of Regulatory Compliance and Quality, Fort Detrick, MD, USA. The study was done in accordance with Good Clinical Practice and written informed consent was obtained from all patients before participation. This study was monitored by the U.S. Army Medical Research and Material Command Quality Assurance Office,  and the Division of Microbiology and Infectious Diseases, National Institutes of Health . The protocol can be obtained from the corresponding author. The trial was registered at Clinicaltrials.gov (identifier NCT01290601) in February 2011. This study was conducted before clinical trial registration became a requirement for publication and was, therefore, registered retrospectively. The authors confirm that all ongoing and related trials for this drug/intervention are registered. All relevant data are within the paper.The protocol was approved by the Ethical Committee, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand on 23th September 2003 to 10th January 2005 at the Bangkok Hospital for Tropical Disease . Eligible patients were randomized to active treatment with either tafenoquine 400 mg for three days ; or chloroquine phosphate 1000 mg for two days  and chloroquine phosphate 500 mg for one day  followed by 15 mg primaquine base for 14 days . Matched placebos were given as shown in P. vivax infection, defined as the presence of P. vivax asexual stage parasites on a blood smear . Patients were excluded if they were pre-menarchal, pregnant or lactating, unwilling or unable to comply with recognized contraceptive methods, had mixed microscopically determined Plasmodium infection, severe vomiting or any medical condition that could interfere with drug absorption, clinically significant illness or an abnormal laboratory value, including evidence of renal dysfunction or G6PD deficiency. Because the study also assessed the potential ophthalmic effects of tafenoquine, subjects were also excluded if they had previous eye surgery, corneal or retinal abnormalities including risk for acute angle closure glaucoma, or used concomitant medications likely to affect renal or ophthalmic function. Patients allergic to 8-aminoquinolines, and those who had used other anti-malarial drugs or another investigational product in the previous 30 days were also excluded.Eligible participants were male or female patients 20\u201360 years old with microscopically confirmed Study participants were hospitalized from day 0 to day 28 with follow up on days 60, 90, 120 or at withdrawal, and were to remain in a malaria free region until day 90. A medical history was taken and vital signs recorded at screening. Vital signs were taken daily until day 7 then days 14 and 28 and at each follow-up visit. Body temperature was measured every 12 hours (\u00b12 hours) after the baseline measurement through day 7. All study medications were given by direct observation by study staff.P. falciparum malaria before day 28 were withdrawn from the study and were not considered evaluable for efficacy endpoints.Malaria blood smears and routine laboratory tests were performed at the Bangkok Hospital for Tropical Disease (HTD) and Armed Forces Research Institute of Medical Sciences (AFRIMS), Bangkok, Thailand. Thick and thin blood smears were obtained on day 0 and then every 12 hours (\u00b1 2 hours) up to and including day 7 until the blood smear became negative for two consecutive smears, then daily until day 7, then on days 14 and 28 and follow up visits at days 60, 90 and 120. Patients were asked to return to the clinic if they developed signs or symptoms of malaria during the follow-up phase. Additional smears were done each day a patient exhibited signs or symptoms of suspected malaria. Microscopically confirmed treatment failures were withdrawn from the study and given appropriate anti-malarial rescue medication as per the Bangkok Hospital for Tropical Diseases treatment practice. Patients who developed Microscopic examination of blood smears was conducted by two independent microscopists, blinded to treatment allocation and each other\u2019s results, who examined 200 oil-immersion fields (magnification x1000) on Field\u2019s-stained blood smears. Any discrepancies  were resolved by a third blinded, senior study microscopist. The third reading was considered final. Parasite densities were calculated based on a count of parasites per 1000 red blood cells in a thin film or per 200 white blood cells in a thick film.Adverse events were evaluated daily until day 29, then at each follow-up visit and were coded using MedRA dictionary version 8.1. Clinical chemistry was performed at days 0, 3, 7, 14, 21, 28, and 90. Blood samples for hematology were collected at days 0, 3, 7, and 14. Blood samples for methemoglobin were collected at days 0, 3, 7, and 21. Renal assessments included serum creatinine and urinalysis and were collected at days 0, 7, and 28. Two independent data monitoring committees (IDMC) were chartered; one for efficacy and routine safety, and one for interpretation of the detailed ophthalmic data (reported separately).Tafenoquine plasma concentrations were determined by Quintiles Limited  under the direction of Worldwide Bioanalysis Department, DMPK, GlaxoSmithKline Pharmaceuticals, UK. Plasma samples were collected from recipients according to a schedule designed for population pharmacokinetics.The primary efficacy endpoint was day 28 adequate clinical response, defined as parasitological clearance throughout the follow-up period until day 28 without previous early treatment failure (defined as parasitemia on day 7) or late treatment failure (parasitemia recurring after day 7 up to and including day 28).P. vivax re-emergence of parasitemia at days 60, 90, and 120; parasite clearance time and gametocyte clearance time, defined as the interval between starting treatment and the first of two consecutive negative blood smears for P. vivax parasites/gametocytes; fever clearance time, defined as the time taken from treatment start for the body temperature to decrease to 37.2\u00b0C and remain at or below this level for a minimum of 24 h. Note that gametocyte clearance time and fever clearance time calculations assumed that patients with fever or gametocytes at any time actually had fever or gametocytes present at baseline.Secondary endpoints were the proportion of patients without Tafenoquine safety was evaluated based on the frequency and severity of adverse events and abnormal values of clinical chemistry, hematology, methemoglobin and urinalysis parameters. Tafenoquine population pharmacokinetics was also reported.No formal comparison was planned between treatment groups. The primary objective of the study was to assess the efficacy of the tafenoquine doses alone. The chloroquine/primaquine treatment arm was included as a reference. For the purposes of estimation, of a one-sided 90% confidence interval (90%CI) was calculated using the Clopper-Pearson \u2018exact\u2019 methodology for the day 28 cure rate in each treatment arm. Secondary efficacy endpoints and all safety data were summarized descriptively.The day 28 per-protocol population was the primary population for efficacy analyses, including all randomized patients who completed all scheduled assessments to day 28, who received all drug treatments and who were compliant with the protocol up to day 28. All patients who were treatment failures, or who had withdrawn from the study, or experienced recurrence (recrudescence or relapse) by day 28 were not required to have attended subsequent visits for inclusion in this population. Definitions for the per-protocol population at days 7, 60, 90 and 120 were similar, though criteria were specific for that time point. The safety population and the intent-to-treat population included all randomized patients who received at least one dose of study medication.P. falciparum malaria (days 8 and 24). No study discontinuation was attributable to adverse events related to study medication. Baseline characteristics were similar between groups . These patients were asymptomatic, cleared their parasitemia by day 8 without additional anti-malarial treatment, and remained relapse free for the duration of their follow-up .In the tafenoquine group, the day 28 adequate clinical response rate in the per-protocol population was 93% (40/43) (90%CI 83\u201398) . The thrIndividual patient tafenoquine concentration\u2013time data from 46 patients are shown in P. vivax relapse. Tafenoquine prevented P. vivax relapse in all evaluable patients, while the chloroquine/primaquine arm had one patient with relapse on day 63  compared with chloroquine/primaquine. Also, a parasite clearance rate of 82 h for tafenoquine would provide a long window for the selection of resistance [P. vivax malaria treatment and prophylaxis. These studies eventually led to the formulation of the current clinical development plan for tafenoquine plus chloroquine or an ACT in P. vivax malaria radical cure. Recently a phase IIb study resulted in the selection of a dosing regimen comprising a single 300 mg tafenoquine dose plus standard 3-day chloroquine therapy\u2014at this dose, 6-month anti-recurrence efficacy was 89.2%  compared with 77.3%  with chloroquine plus 15 mg x 14 day primaquine, and 37.5%  with chloroquine alone [This small phase II study demonstrated that a three-day course of tafenoquine monotherapy was efficacious for the radical cure of sistance . Currentsistance , 25. ThiAlthough there were no clinical symptoms, the 1200 mg total dose given over three days in the current study increased methemoglobin levels in the tafenoquine arm to an extent that would be concerning if used in a less closely monitored population. With tafenoquine doses up to 600 mg, no adverse events have been reported for methemoglobinemia , 14, 15.A previous study of tafenoquine prophylaxis , reporteP. vivax malaria, showing effective, though slow, blood schizonticidal activity and providing high-dose safety data.In conclusion, this study adds to the published data on tafenoquine treatment of S1 CONSORT Checklist(DOC)Click here for additional data file.S1 Protocol(PDF)Click here for additional data file."}
+{"text": "Plasmodium knowlesi malaria; its excellent tolerability and rapid therapeutic response allowed earlier hospital discharge, and support its use as a first-line artemisinin-combination therapy treatment policy for all Plasmodium species in Malaysia.AL is highly efficacious for treating uncomplicated  Plasmodium knowlesi is reported increasingly across Southeast Asia and is the most common cause of malaria in Malaysia. No randomized trials have assessed the comparative efficacy of artemether-lumefantrine (AL) for knowlesi malaria.Plasmodium falciparum histidine-rich protein 2. Diagnosis was confirmed by means of polymerase chain reaction. Patients were block randomized to AL  or CQ (25 mg/kg). The primary outcome was parasite clearance at 24 hours in a modified intention-to-treat analysis.A randomized controlled trial was conducted in 3 district hospitals in Sabah, Malaysia to compare the efficacy of AL against chloroquine (CQ) for uncomplicated knowlesi malaria. Participants were included if they weighed >10 kg, had a parasitemia count <20000/\u03bcL, and had a negative rapid diagnostic test result for P = .06). Overall parasite clearance was shorter after AL than after CQ , with all patients aparasitemic by 48 hours. By day 42 there were no treatment failures. The risk of anemia during follow-up was similar between arms. Patients treated with AL would require lower bed occupancy than those treated with CQ . There were no serious adverse events.From November 2014 to January 2016, a total of 123 patients (including 18 children) were enrolled. At 24 hours after treatment 76% of patients administered AL  were aparasitemic, compared with 60% administered CQ (47%\u201372%; 39 of 65; risk ratio, 1.3 ; Plasmodium species in Malaysia. AL is highly efficacious for treating uncomplicated knowlesi malaria; its excellent tolerability and rapid therapeutic response allow earlier hospital discharge, and support its use as a first-line artemisinin-combination treatment policy for all NCT02001012. Plasmodium knowlesi is reported increasingly in humans infected across Southeast Asia [P. knowlesi from Plasmodium malariae or the early ring stage of Plasmodium falciparum, with misidentification as Plasmodium vivax also occurring [P. knowlesi has been associated with failure to recognize severe disease and death [P. falciparum or P. vivax infections. Rapid diagnostic tests lack sensitivity and specificity for P. knowlesi, and cannot be used to guide prompt and effective treatment [The simian parasite ast Asia , 2 and iast Asia  and areaast Asia , 7. Accuccurring , 9. Misind death , and inareatment .P. knowlesi reduces the antimalarial drug selection pressure, with P. knowlesi shown to respond rapidly to numerous antimalarials, including artemisinin-combination therapy (ACT) [P. knowlesi infection in CQ-susceptible areas or first-line ACT in areas with confirmed CQ-resistant P. vivax [The predominant zoonotic reservoir of py (ACT) , 17 and py (ACT) . To datepy (ACT) , with thpy (ACT) . In the py (ACT) . The curP. vivax , as founP. vivax .P. knowlesi malaria in both adults and children in areas where either AL or CQ may be currently used.Artemether-lumefantrine (AL) is widely used as the first-line ACT for malaria in endemic countries, including Malaysia , 24. AL P. knowlesi is the most common cause of malaria [P. knowlesi malaria presenting to the study hospital sites and meeting the following inclusion criteria: \u22651 year of age, weight \u226510 kg, microscopic diagnosis of P. knowlesi monoinfection, negative results of a rapid diagnostic test for P. falciparum malaria (histidine-rich protein 2), fever (temperature \u226537.5\u00b0C) or history of fever in the last 48 hours, and provision of written informed consent. Patients with clinical or laboratory criteria for severe malaria or warning signs according to WHO 2014 criteria [A 2-arm, randomized, open-label trial conducted at 3 sites in Sabah, Malaysia: Kudat, Kota Marudu, and Pitas district hospitals, where  malaria , 27. Parcriteria , parasitcriteria , pregnanAL , was given as a fixed-dose combination of artemether (20 mg) and lumefantrine (120 mg). Doses were administered at enrolment and at 8, 24, 36, 48, and 60 hours, with target total doses of 12 mg/kg for artemether and 60 mg/kg for lumefantrine. Chloroquine phosphate , consisting of 155-mg base tablets, was administered at enrolment and at 6, 24, and 48 hours, with a target total dose of 25 mg/kg. Dosages of AL and CQ were based on weight, consistent with current WHO  and MalaP. knowlesi asexual parasites at 24 hours. Secondary end points included microscopic parasite clearance as measured by time to the first of 2 negative blood films; the times to 50%, 90%, and 99% reduction in baseline parasite count in those with parasitemia counts >1000/\u03bcL [The primary end point was the difference between the 2 treatment arms in the proportion of patients with negative microscopic results for >1000/\u03bcL ; and the>1000/\u03bcL , risk of>1000/\u03bcL .A sample size of 58 participants in each arm would have 90% power to falsify the null hypothesis (no difference in parasite clearance between AL and CQ at 24 hours), with a significance level (\u03b1) of .05, assuming that 45% of study participants treated with CQ and 16% of those treated with ACT would remain parasitemic 24 hours after the start of treatment [Patient allocation codes were computer generated in blocks of 20 for each drug arm using Stata software (version 12) before the start of the study. Treatment allocation was provided in a sealed opaque envelope opened by a study nurse once the participant met all the study enrollment criteria and informed consent had been signed.The primary study end point was determined by microscopists who were blinded to treatment allocation. Research blood slides were labeled only with the study code, which was nonidentifiable for both demographic data and drug allocation. Staff conducting all biochemical and polymerase chain reaction (PCR) assays were blinded to results at microscopy and treatment arm.Venous blood was taken before enrollment for hematology, biochemistry, rapid diagnostic tests, and delayed species PCR, as described elsewhere , 32. Allt test or the Wilcoxon-Mann-Whitney test for continuous variables and \u03c72 or the Fisher exact test for categorical variables. Microscopic asexual parasite and gametocyte counts were calculated using thick blood smears [e) parasite counts versus time using the WorldWide Antimalarial Resistance Network (WWARN) parasite clearance method [I2).Data were double entered by separate individuals into Epidata software (version 3.1), and analyzed using Stata software (version 12). Primary analysis of parasite clearance at 24 hours was by modified intention to treat. Intergroup differences were compared using the Student d smears . Best-fie method . Treatmee method , using aP. knowlesi monoinfection microscopically diagnosed, with 141 (76%) enrolled in the study , 186 patients (65%) had he study . SeventeThere were no differences in baseline demographics of the patients enrolled between study arms . The medP = .06). OF patients with parasitemia counts >1000/\u03bcL at enrollment [P = .047; P = .02. All patients in the AL arm were aparasitemic) by 42 hours, compared with 48 hours for the CQ arm. Overall, the median parasite clearance time was 18 hours  after AL versus 24 hours (12\u201348 hours) after CQ (P = .02); the corresponding slopes of the log-normalized clearance curves were 0.27 and 0.22, respectively (P < .002). Of patients with parasitemia counts >20000/\u03bcL, parasite clearance times ranged from 24 to 36 hours for AL, and from 30\u201342 hours for CQ (P = .16). No patients had microscopic gametocytemia at any time point during follow-up, and there was no statistically significant difference in fever clearance time between treatment arms (The proportion of patients aparasitemic at 24 hours was 76% (44 of 58) in the AL arm and 60% (39 of 65) in the CQ arm . Of the patients without anemia at baseline, 65% (28 of 43) in the CQ arm became anemic by day 28, compared with 56% (23 of 41) in the AL arm (P = .68).The overall risk of anemia throughout the 28-day follow-up was 81% (46 of 57) in the CQ arm, compared with 67% (32 of 48) in the AL arm P = .10; . Of the P < .001).The actual durations of hospital stays were similar between treatment arms . When applying the national hospital discharge policy of 2 negative blood slides on consecutive days with medication to be completed after discharge, the predicted bed occupancy was 2414 days per 1000 patients treated with AL versus 2800 days per 1000 patients treated with CQ , although there were no differences in the reporting of cough, respiratory rate, or oxygen saturation. No other differences in adverse events were documented . Comparing direct treatment outcomes between AL and ASMQ, there was no statistically significant difference in proportions negative for parasites at 24 hours , bed occupancy (2414 vs 2426 days per 1000 patients), or risk of anemia at 28 days (29% vs 32%).A meta-analysis using individual patient data for the difference in proportional parasite clearance at 24 hours between AL and CQ arms 16%) and a previous trial of ASMQ versus CQ (29%) [% and a pAL and CQ were highly effective for treating uncomplicated knowlesi malaria, with both drugs well tolerated, all patients becoming aparasitemic within 48 hours, and no treatment failures by day 42 of follow-up. However, AL demonstrated faster parasite clearance and earlier predicted hospital discharge than CQ, with improved potential health cost benefits. The earlier parasite clearance for AL compared with CQ is consistent with a previous trial of ASMQ versus CQ for uncomplicated knowlesi malaria .P. knowlesi parasite counts and inappropriate initial treatment with oral rather than intravenous artesunate for severe disease has resulted in case fatalities [The difference in parasite clearance between AL and CQ was more pronounced in those presenting with higher parasite counts >1000/\u03bcL, although this may be due in part to methodologic difficulties in accurately evaluating parasite clearance at lower levels . Routinetalities . Howevertalities , 33, 34,talities . The saftalities .P. knowlesi isolates [Clinical outcomes with AL and ASMQ were similar despite differences in pharmacokinetic/pharmacodynamic properties between these medications. The proportion of patients with negative microscopy at 24 hours was only 1.1-fold greater in those treated with ASMQ  versus Aisolates . Howeverisolates . P. falciparum [P. knowlesi parasites from zoonotic transmission to humans. Patients in both clinical trials were aparasitemic within 72 hours, and both ACTs are administered for 3 days. Therefore, with the fast 24-hour life-cycle of P. knowlesi and high efficacy of the artemisinin component of both ACTs, the shorter half-life of the partner drug for AL, lumefantrine, of about 3 days, did not confer any disadvantage for eliminating any residual parasites compared with mefloquine, with a half-life of about 21 days [AL is also highly lipophilic, and optimal oral absorption requires coadministration of fatty foods . In routlciparum , fatty fBoth AL and CQ demonstrated high safety and tolerability, with minimal adverse events. The most commonly reported adverse event was nonsevere rash or itch, seen in 12% of patients treated with AL, with recognized drug-related gastrointestinal disturbance in <5%. AL and CQ were also both tolerated well by the 12 children \u226412 years of age. Suboptimal dosing of AL with children owing to administration as tablets or fractions of tablets in dose-weight bandings  had no dP. knowlesi gametocytes in about 25% of patients at presentation was probably underestimated by microscopy, with minimum estimates of 85% in a previous study using a sensitive Pks25 PCR assay [P. knowlesi, and risk of onward transmission after treatment is minimal, with other transmission-blocking agents, such as primaquine, not required.These findings are in contrast to comparisons of ASMQ and CQ , but theCR assay . HoweverP. knowlesi as P. falciparum or P. vivax [P. knowlesi infections with parasitemia underestimated by microscopy. AL has some advantage in safety and tolerability over ASMQ because of the small risk of severe mefloquine-related neuropsychiatric sequelae [In conclusion, our findings support the use of AL as a first-line treatment over CQ for uncomplicated knowlesi malaria, allowing a universal policy of ACT for all species of malaria in this region . AlthougP. vivax , both hiP. vivax , 37. In sequelae , despiteP. knowlesi infections, are also likely to be efficacious in the treatment of uncomplicated knowlesi malaria, although they require further evaluation.Results of this and the previous RCT seem generalizable to other areas of Malaysia and Southeast Asia where AL and ASMQ are used as first-line therapies for uncomplicated malaria , 24. Oth"}
+{"text": "Autosplenectomy, as a result of sickle cell disease, is an important risk factor for severe malaria. While molecular methods are helpful in providing rapid and accurate infection detection and species identification, the effect of hyposplenism on result interpretation during the course of infection should be carefully considered.Plasmodium falciparum. Parasitaemia was tracked over time by microscopy and nucleic acid tests to evaluate the therapeutic response in the setting of hyposplenism. The patient showed prompt resolution of patent infection by microscopy but remained positive by molecular methods for\u00a0>\u00a030\u00a0days after treatment initiation.A 32-year old autosplenectomized Nigerian male with severe sickle cell disease was referred to the National Institutes of Health for allogenic hematopoietic stem cell transplant. Despite testing negative for malaria by both smear and PCR 2\u00a0weeks after arrival in the USA, the patient developed fever and diffuse bilateral lower rib cage and upper abdominal pain 2\u00a0weeks later and subsequently tested positive for Plasmodium nucleic acid detection, the persistence of Plasmodium nucleic acids following adequate treatment in functionally asplenic patients can lead to a diagnostic dilemma. In such patients, clinical response and peripheral blood smears should guide patient management following treatment. Nonetheless, in pre-transplant patients at high-risk for pre-existing Plasmodium infections, highly sensitive molecular assays can be useful to rule out infection prior to transplantation.While molecular testing can provide sensitive  Malaria remains a prominent cause of infection and mortality worldwide with\u00a0~\u00a0212 million new infections in 2015 and almost 429,000 reported deaths \u20136. The pPlasmodium falciparum malaria, and clinical considerations when faced with conflicting diagnostic test results post-treatment due to the persistence of Plasmodium nucleic acids in blood samples.This case concerns a patient with SCD and autosplenectomy, who travelled from Nigeria to the USA to undergo allogenic haematopoietic stem cell transplantation (HSCT). Discussion points include the conundrum of initial negative malaria screening results, a subsequent diagnosis of Plasmodium multiplex PCR; all testing was reported as negative. On day-4, the patient developed fever as well as diffuse bilateral lower rib cage and upper abdominal pain. He was admitted on day-1 for a vaso-occlusive pain crisis. On day 0, the patient developed fevers to 39.1\u00b0C and a decrease in absolute haemoglobin from 8.2 to 5.8\u00a0g/dL. Possible intraerythrocytic parasites were observed initially from routine cell count and differential smears, and repeat malaria blood smears were positive for immature (ring form) and mature trophozoites and schizonts of P. falciparum at 2.9% parasitaemia  was started on day 1 after consultation with the clinical team and pharmacy. The patient was supported with RBC transfusions and defervesced by day 3 with 0.93% parasitaemia. Between days 5 and 6, his parasitaemia levels rose from 0.20 to 0.42%, respectively  for 7\u00a0days due to concerns for artemether\u2013lumefantrine resistance. Reflex testing by the Centers for Disease Control and Prevention did not reveal genetic markers on the kelch 13 gene that was consistent with artemisinin resistance. The patient remained clinically well with\u00a0<\u00a00.05% parasitaemia and was discharged after three doses of atovaquone/proguanil (day 8). Blood smears were negative by day 10, but the multiplex malaria PCR assay remained positive on day 17. Because of his scheduled transplant and slow parasite clearance, he received a course of doxycycline for 3\u00a0days. The multiplex malaria PCR assay became negative between days 25 and 31. The patient proceeded with the allogeneic HSCT on day 41 and engrafted fully with donor red cells without recurrence of malaria during the transplant process. He has returned to Nigeria and has remained clinically well.This is a case of a 32-year old Nigerian man with homozygous SCD. Despite receipt of hydroxyurea, he continued to experience monthly priapism and three to four hospitalizations for vaso-occlusive pain crises per year. He reported a history of several malaria infections including a severe episode that led to acute chest syndrome and heart failure requiring intensive care unit monitoring and red cell transfusions. The patient travelled from Nigeria to the USA (arriving on day-32) in preparation to receive a non-myeloablative allogeneic HSCT from his fully human leukocyte antigen-matched brother. As part of his pre-transplant evaluation on day-18 (2\u00a0weeks after entering the USA from Nigeria), blood was obtained for malaria thick and thin smears and for Plasmodium 18S rRNAs (LOD: 20\u00a0parasites/mL of whole blood [Retrospective testing was performed on 12 preserved whole blood samples using a quantitative reverse transcription PCR (qRT-PCR) that targets the highly expressed le blood , 11). qRPlasmodium parasites in peripheral blood specimens is important for effective management of patients with suspected malaria infection [P. falciparum  along with the rarely present mature trophozoites and schizonts were observed  was below the LODs for these assays . Regardless of splenic status, clinicians must consider whether ongoing molecular positivity in a treated patient is due to slower clearance of non-viable parasite-derived nucleic acids or represents a potential ongoing viable infection. Gradually-declining molecular assay-derived measurements of parasite density are most likely representative of non-viable, slow-clearing parasite biomass. In contrast, resurgent or clear positive persistence could represent impending treatment failure, which has been recognized for drug-resistant parasites manifesting as persistent PCR positivity in the week following initial treatment [Plasmodium infections, highly sensitive molecular assays such as the ultrasensitive qRT-PCR assay used herein may be useful for ruling out infection prior to transplantation. The LOD, range of species detected, and sample stability should be considered when selecting such tests.In functionally asplenic patients like the patient presented here, it is likely that rganisms . In contrganisms . It is prganisms . Additioreatment . Thus, a"}
+{"text": "Plasmodium falciparum malaria in the 'target' population of children\u2009\u2264\u20095\u00a0years of age in Africa as well as Asian patients of all ages.The clinical development of a single encounter treatment for uncomplicated malaria has the potential to significantly improve the effectiveness of antimalarials. Exploratory data suggested that the combination of artefenomel and piperaquine phosphate (PQP) has the potential to achieve satisfactory cure rates as a single dose therapy. The primary objective of the study was to determine whether a single dose of artefenomel (800\u00a0mg) plus PQP in ascending doses is an efficacious treatment for uncomplicated n\u2009=\u2009355) and Vietnam (n\u2009=\u200982) were included, with 85% of the total population being children\u2009<\u20095\u00a0years of age.Patients in six African countries and in Vietnam were randomised to treatment with follow-up for 42\u201363 days. Efficacy, tolerability, safety and pharmacokinetics were assessed. Additional key objectives were to characterise the exposure\u2013response relationship for polymerase chain reaction (PCR)-adjusted adequate clinical and parasitological response at day 28 post-dose (ACPR28) and to further investigate Kelch13 mutations. Patients in Africa  was 70.8% (61.13\u201379.19), 68.4% (59.13\u201376.66) and 78.6% (70.09\u201385.67) for doses of 800\u00a0mg artefenomel with 640\u00a0mg, 960\u00a0mg and 1440\u00a0mg of PQP respectively. ACPR28 was lower in Vietnamese than in African patients  respectively. Within the African population, efficacy was lowest in the youngest age group of\u2009\u2265\u20090.5 to\u2009\u2264\u20092\u00a0years, 52.7% (38.80\u201366.35). Initial parasite clearance was twice as long in Vietnam than in Africa. Within Vietnam, the frequency of the Kelch13 mutation was 70.1% and was clearly associated with parasite clearance half-life (PCt1/2). The most significant tolerability finding was vomiting (28.8%).In this first clinical trial evaluating a single encounter antimalarial therapy, none of the treatment arms reached the target efficacy of\u2009>\u200995% PCR-adjusted ACPR at day 28. Achieving very high efficacy following single dose treatment is challenging, since\u2009>\u200995% of the population must have sufficient concentrations to achieve cure across a range of parasite sensitivities and baseline parasitaemia levels. While challenging, the development of tools suitable for deployment as single encounter curative treatments for adults and children in Africa and to support elimination strategies remains a key development goal.NCT02083380. Registered on 7 March 2014.ClinicalTrials.gov, The online version of this article (doi:10.1186/s12916-017-0940-3) contains supplementary material, which is available to authorized users. Since 2000 the incidence of malaria has fallen by 41% and mortality rates have declined by 62% globally, due to increased deployment of new interventions including artemisinin-based combination treatments and insecticide treated bed nets (WHO 2016) [Numerous studies have suggested that in 'real-life' community settings, poor to moderate adherence to current standard 3-day treatment regimens is common, and this could impact morbidity and mortality as well as drive the development of resistance, although definitive data are difficult to obtain \u20134. AvailPlasmodium falciparum ranging from 0.9 to 1.88 [Exploratory clinical data suggested that artefenomel (OZ439) plus piperaquine phosphate (PQP) in combination could be efficacious as a single encounter cure. Artefenomel, a novel synthetic trioxolane, contains a similar peroxidic pharmacophore to artemisinins and has demonstrated rapid parasite clearance in patients, with a median parasite reduction ratio at 24\u00a0h (log10) post-treatment (PRR24) for We report the results of the first clinical efficacy study of the combination of artefenomel and PQP in a design which allowed rapid progression from adult African patient to children\u2009\u2264\u20095\u00a0years of age and Asian patients of all ages to ensure that dose finding for phase III is carried out in populations most likely to require the highest exposures to achieve cure. The study also employed interim futility analyses in order to drop doses with a low probability of success early.P. falciparum malaria.The primary objective of the study was to determine whether a single dose combination of artefenomel plus PQP is an efficacious treatment for uncomplicated Secondary and exploratory objectives included determination of the incidence of recurrence, recrudescence and new infection, estimation of parasite clearance kinetics and exploration of the relationship between Kelch13 genotype and parasite clearance half-life (PCt1/2) in Asian patients.An additional key exploratory objective was to characterise the dose/exposure\u2013response relationship for the combination for the primary efficacy endpoint across the patient population and to identify significant covariates influencing efficacy. Safety, tolerability and pharmacokinetics (PK) were also assessed. Details of the study objectives, design and endpoints are summarised in Additional file P. falciparum against chloroquine and sulfadoxine-pyrimethamine is widespread at all African sites, and evidence of artemisinin resistance was confirmed at the Vietnamese sites [The study was conducted at nine study sites across six African countries, Benin (Cotonou), Burkina Faso  , 10, Demse sites .P. falciparum malaria. The artefenomel dose of 800\u00a0mg was expected to deliver close to the maximum well-tolerated exposure, and PQP doses were selected to span a range of adequate clinical and parasitological response at day 28 (ACPR28) values, with the highest dose estimated to give a mean maximum placebo corrected change from baseline QTcF of 18\u00a0ms [This was a randomised, double-blind, single dose study to investigate the efficacy, safety, tolerability and PK of artefenomel 800\u00a0mg in loose combination with three doses of PQP  in male and female patients aged\u2009\u2265\u20096\u00a0months to\u2009<\u200970\u00a0years (body weight\u2009\u2265\u20095\u00a0kg to\u2009\u2264\u200990\u00a0kg) with uncomplicated of 18\u00a0ms .P. falciparum mono-infection in the range 1000 to 100,000 asexual parasites/\u03bcL of blood, and with fever (axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C) or history of fever in the preceding 24\u00a0h, were included following their submittal of written informed consent, and all eligible patients were randomised via an Interactive Web Response System (IWRS) in a ratio of 1:1:1 to one of the three treatment arms . Important exclusion criteria were the presence of severe malaria , haemogThe study was initiated in patients aged\u2009>\u200915\u00a0years, and following review of safety data by an Independent Safety Monitoring Board (ISMB), sequentially younger patients were recruited in a step-down procedure described in Additional file 0.75. Artefenomel was administered as a suspension formulation containing \u03b1-tocopherol polyethylene glycol (TPGS). PQP was included in the suspension (for patients\u2009<\u200924\u00a0kg) or was administered as separate tablets (patients\u2009\u2265\u200924\u00a0kg), with blinding maintained by administering matching placebo tablets. For the lowest weight band the dosing volume was 75\u00a0mL (plus 2\u00d7 15\u00a0mL rinses). Patients who vomited within 5\u00a0min of start of dosing were re-dosed once. Details of study drug treatments and administration are given in Additional file Fasted patients\u2009\u2265\u200935\u00a0kg received artefenomel 800\u00a0mg in loose combination with PQP doses of 640, 960 or 1440\u00a0mg at day 0. Patients who weighed\u2009<\u200935\u00a0kg received body weight-adjusted doses  within wFollowing drug administration, patients were followed for 42\u00a0days or 63\u00a0days at some centres (patients were consented separately for days\u2009>\u200942 to 63). Patients remained in the clinical unit for a minimum of 48 (African patients\u2009>\u20095\u00a0years old) or 72\u00a0h  and were discharged provided parasite and fever clearance had been achieved. Patients returned for assessment on days 3, 5, 7, 10, 14, 21, 28, 42 and 63 at selected centres. Blood films (thick and thin) were prepared and axillary temperatures were measured at screening/pre-dose, 6, 12, 18, 24, 30, 36, 48, 72\u00a0h and days 5, 7, 10, 14, 21, 28, 42 and 63. Assessments for safety included haematology, clinical chemistry, urinalysis and a triplicate 12-lead electrocardiogram (ECG). Clinical assessments were taken according to the schedule presented in detail in Additional file falciparum infections. A second microscopist, blinded to initial readings, re-read all slides, and a third resolved discrepant readings. A slide was considered negative in the absence of asexual parasites per 1000 counted leukocytes using a 100\u00d7 magnification oil immersion objective. Parasite density was calculated as follows: (number of counted parasites/counted leukocytes)\u2009\u00d7\u2009most recent absolute leukocyte count per microliter. Details of additional methods are given in the supplementary material.For screening, thick blood films were stained with 10% Giemsa for 10\u00a0min, and thick and thin films for baseline to follow-up were stained with 2% Giemsa for 30\u00a0min. Expert microscopists determined parasite densities and examined thick blood films for parasites and thin blood films for non-P. falciparum asexual parasitaemia at inclusion. The ITT subset defined for the Kaplan\u2013Meier (KM) estimates of recurrence, recrudescence and new infection rate included only those who consumed the entire dose.The intention to treat (ITT) and safety analysis sets were identical and included all patients who provided informed consent, received the study drug  and had a confirmed positive blood film for The per protocol (PP) set was the primary analysis set and included all patients comprising the ITT set who consumed the entire dose and were without major protocol deviations. The modified PP analysis set in addition excluded patients who vomited between\u2009>\u20095\u00a0min and\u2009\u2264\u20094\u00a0h after start of drug administration. Patients could be excluded from a population for more than one reason see Fig.\u00a0. The PK The study (MMV_OZ439_13_003) conformed to the Declaration of Helsinki and Standard Operating Procedures that meet current regulatory requirements and guidelines established by the International Conference on Harmonization for Good Clinical Practice in Clinical Studies. It was approved by the relevant Independent Ethics Committees (IECs), national Institutional Review Boards and, where relevant, local regulatory authorities at each of the participating sites . The study protocol was registered and the study results are reported on ClinicalTrials.gov (NCT02083380).The primary efficacy endpoint was polymerase chain reaction (PCR)-adjusted ACPR28 in the PP analysis set. We also report the following secondary and exploratory endpoints: PCR-adjusted ACPR at day 42 (ACPR42) and day 63 (ACPR63) and crude ACPR at days 28, 42 and 63 for the ITT and PP analysis sets and Kaplan\u2013Meier incidence rate of recrudescence over 63\u00a0days (ITT subset); in the PP analysis set, percentage of patients achieving parasite clearance at 72\u00a0h post-dose, PCt1/2, Kelch13 genotype and the relationship between Kelch13 genotype and PCt1/2 are also reported.ACPR was defined according to the WHO  as absenSafety and tolerability endpoints included incidence of adverse events (AEs) and serious AEs (SAEs), vital signs, physical measurements, laboratory safety measurements, liver function tests (LFT) increase, cases fulfilling the Hy's law definition and ECG abnormalities including absolute QTc value categorisation and change from baseline QTc. Further details are given in Additional file Hepatic: Hy's law definition cases; any alanine transaminase (ALT) or aspartate transaminase (AST)\u2009\u2265\u20095\u2009\u00d7\u2009the upper limit of the normal range (ULN); any elevation in total bilirubin\u2009\u2265\u20092.5\u2009\u00d7\u2009ULN (>\u200935% direct); any AST or ALT\u2009\u2265\u20093\u2009\u00d7\u2009ULN with the appearance of fatigue, nausea, vomiting, right upper quadrant pain or tenderness, fever, rash and/or eosinophilia (eosinophil percent or count above the ULN); or ALT\u2009\u2265\u20093\u2009\u00d7\u2009ULN that persisted for\u2009>\u20094\u00a0weeksCardiac: QTcF prolongation from baseline of\u2009>\u200960\u00a0ms; QTcF at any time; QTcF\u2009>\u2009450\u00a0ms; T-wave liability or T-wave morphologic changes during therapy; bundle branch block; and any arrhythmiaHaematological: Haemoglobin (Hb) drop\u2009>\u20092\u00a0g/dL from baseline; Hb drop\u2009<\u20095\u00a0g/dL; absolute neutrophil count (ANC)\u2009<\u20091000/\u03bcLPregnancy.Treatment emergent adverse events of special interest (TEAESIs), requiring rapid reporting, were also defined in the protocol to ensure careful monitoring:The aim of the study was to determine whether any of the treatment arms reached a target PCR-adjusted ACPR28 of\u2009\u2265\u200995% . The study was not powered for comparison between dosing arms. ACPR28 was categorised as Cure and Failure according to the WHO method . DescripTrial simulations suggested that a treatment arm size of 106 should be required if the dose combination is effective . RecruitIn adult patients (weighing\u2009>\u200935\u00a0kg) blood samples for pharmacological analysis were collected at 15 to 16 time points: pre-dose, 2, 4, 6, 12, 24, 48, 72\u00a0h and days 5, 7, 10, 14, 21, 28, 42, 63. In paediatric patients the number was between 3 and 10 samples. Artefenomel and piperaquine PK data were analysed separately using non-linear mixed effect modelling  in Monolix (version 4.3.3) or NONMEM (version 7.3 or later) respectively. For artefenomel, additional data from two mono-therapy clinical phase II studies in adult Asian patients were included in order to extend the dose range (100\u20131200\u00a0mg); see Additional file Subsequently, exposures of artefenomel and piperaquine for each patient were derived from the individual PK parameters estimated in the population PK analysis. Maximum plasma concentration (Cmax), time to reach maximum concentration (Tmax) and concentration on day 7 (Cday7) were obtained from the simulated profiles, and the area under the curve (AUC) extrapolated to infinity (AUCinf) was calculated directly from the estimated PK parameters. More details are provided in Additional file The relationship of the binary outcome of ACPR28 response to the estimated artefenomel and piperaquine Cday7 and other covariates was evaluated in a logistic regression model using the statistical software R (version 3.2.2). Within the single dose setting of the study, Cday7 is highly correlated with other exposure variables, such as AUCinf or Cday14. However, Cday7 was preferred to allow future extrapolation to multi-dose regimens as well for its scientific rationale . Additional covariates evaluated were presumed immunity status , region, baseline parasitaemia, age and Kelch13 genotype. All patients in the ITT set with ACPR28 values and exposure for both drugs were included in the analysis. More details are provided in Additional file The objective of the simulations was to evaluate the dose\u2013response relationship for single dose combination treatment with artefenomel and PQP based on the developed population PK and exposure\u2013response models. The simulations were performed for a range of single dose combination doses (for a TPGS formulation) for the African population\u2009\u2264\u20095\u00a0years of age. Actual doses assumed the same body weight bands and dose adjustments applied to this study. For further details see Additional file An interim futility analysis was carried out after recruitment of approximately 50 evaluable patients from the target population per treatment arm as planned. All doses were concluded to have reached the futility criteria (probability of ACPR28\u2009<\u200990% was 0.9999); hence, the study was stopped. Recruitment to the study continued during the futility analysis process.The analysis populations are shown in Fig.\u00a0n\u2009=\u2009355 in Africa and n\u2009=\u200982 in Asia (Vietnam) received the study drug  differed by region due to the enrolment structure. For the ITT/safety set, in Africa, 81.1% of patients were\u2009\u2264\u20095\u00a0years old, whereas in Asia 96.3% were\u2009>\u200915\u00a0years old. No patients\u2009\u2264\u20095\u00a0years old were recruited in Asia. There was also a difference in sex; 48.3% of patients in Africa and 93.9% in Asia were male.Median baseline asexual parasitaemia across all treatment arms was 12,913/\u03bcL , was similar in Asian and African patients\u2009\u2264\u20095\u00a0years, 13,140/\u03bcL  and 14,029/\u03bcL  respectively and was slightly lower in African patients\u2009>\u20095\u00a0years, 9714/\u03bcL .Of those randomised, 178 (39.7%) completed the study up to day 42 (or 63)  was 65% in the African population and 91.5% in the Asian population. Non-compliance was predominantly due to vomiting, with an overall incidence of vomiting of 28.8% (35% in Africa and 7% in Asia). Compliance and vomiting incidence were similar in the African population across age. There were anecdotal reports that, in some centres, young children were unable to ingest the full study dose, despite a reported success rate of administration of study drug (with or without subsequent vomiting) of\u2009>\u200995% in all populations.Crude and PCR-adjusted ACPR results are reported in Table\u00a0For the primary analysis set (PP) and endpoint (ACPR28), none of the treatment arms reached the target efficacy of\u2009\u2265\u200995% and there was no clear dose\u2013response relationship, although efficacy was highest for PQP 1440\u00a0mg  across the populations Table\u00a0.Fig. 2EfEfficacy in the mPP population (excluding patients who vomited) was similar to that in the PP population; e.g. for a PQP dose of 1440\u00a0mg, ACPR28 was  across the populations.ACPR28 appeared lower in Asian (Vietnamese) than in African patients; thus, for all treatment arms combined, ACPR28 was 66.2% (95% CI 54.6\u201376.6) and 74.5% (95% CI 7.81\u201379.7) respectively Fig.\u00a0. In the In the PP population, recurrence in African and Vietnamese patients was 37.0% and 31.6% respectively at day 28, with all but one determined by PCR to be recrudescence in the Vietnamese population, whereas in the African population, approximately one third of recurring parasites was determined to be a new infection at day 28.In Asia, in the PP population, the number and percentage of recrudescences and new infections, with 95% CIs, was 11/80  and 0/80 at day 14, 18/79  and 0/79 at day 21, 23/79  and 1/79  at day 28, 26/78  and 1/78  at day 42, and 27/75  and 2/75  at day 63.In Africa, in the PP population, the number and percentage of recrudescences and new infections, with 95% CIs, was 34/325  and 1/325 (0.3%) at day 14, 57/321  and 19/321  at day 21, 61/319  and 32/319  at day 28, 65/313  and 50/313  at day 42, and 62/277  and 49/277  at day 63.Kaplan\u2013Meier estimates of the fraction of patients with recrudescence and new infection over time by region and age group are presented in Figs.\u00a0A regional difference in the percentage of patients who cleared parasites by 72\u00a0h post-dose was evident, with 92.9% (95% CI 89.6\u201395.4) of African patients achieving parasite clearance by 72\u00a0h post-dose compared with 35.0% (95% CI 24.7\u201346.5) in Vietnamese patients. There was no clear difference in the percentage of patients who cleared parasites across the different piperaquine doses or in the African population\u2009>\u20095\u00a0years compared with those\u2009\u2264\u20095\u00a0years.The median time to 50%, 90% and 99% parasite clearance was approximately twice as long in Asia compared with Africa  h in Asian patients and 14.30 (IQR 10.90\u201317.60) h in African patients.R2\u2009>\u20090.75. PCt1/2 was longer in Vietnam versus Africa . Within Vietnam, PCt1/2 was similar across the four study centres.Initial clearance of parasites was rapid in the African population. Median parasite clearance half-life (PCt1/2) was calculated using the WWARN Parasite Clearance Estimator (PCE), details of which are published . PCt1/2 A total of 20 known Kelch13 genotypes were tested for  . In VietN\u2009=\u200924; minimum 2.4, maximum 12.3\u00a0h) and 8.1\u00a0h  respectively, versus 2.6\u00a0h  for wild type (WT)  was reported to have led to study treatment discontinuation.Six treatment emergent serious adverse events (TESAEs) were reported in four patients. One patient had severe anaemia (2\u00a0days post-dose), one patient had a reversible haemoglobin drop\u2009<\u20095\u00a0g/dL (26\u00a0days post-dose) and one presented with febrile convulsions (7\u00a0h post-dose). One patient had three TESAEs, two of reversible grade 3 transaminase elevations and one of neutropenia (28\u00a0days post-dose). None of these TESAEs led to premature study discontinuation. Five of the six TESAEs were considered to be potentially related to the study treatment, the exception being that of febrile convulsion which was considered not related. No AEs due to drug-induced liver toxicity  were reported.Table\u00a0The most frequently reported TEAESI was QT prolongation in ECG: 24 (16.8%), 37 (25.0%) and 38 (26.0%) of patients in the PQP 640\u00a0mg, 960\u00a0mg and 1440\u00a0mg treatment arms respectively. A first degree atrioventricular (AV) block reported as grade 1 at 48\u00a0h post-dose  was reported in one patient (PQP 640\u00a0mg). The event resolved at day 7 post-dose . One patient (PQP 1440\u00a0mg) had a mild reversible sinus bradycardia which resolved in 4\u00a0days. QTcF increase from baseline of 30\u201360\u00a0ms occurred in 55 (38.7%), 59 (40.4%) and 71 (49.7%) and of\u2009>\u200960\u00a0ms in 8 (5.6%), 11 (7.5%) and 27 (18 .9%) of patients respectively in the 640, 960 and 1440\u00a0mg PQP dosing arms. All but one QTcF value was\u2009<\u2009480\u00a0ms .One patient (PQP 1440\u00a0mg) experienced a reversible TEAESI of hyperbilirubinaemia  in the System Organ Class hepatobiliary disorders. This event was associated with a TESAE of anaemia (Hb drop\u2009>\u20092\u00a0g/dL from baseline). Other frequent TEAESIs were neutrophil count decreased\u2009<\u20091000/\u03bcL  and Hb decreased . The most significant tolerability finding was vomiting (28.8%) according to the compliance data. The high rate of vomiting is thought to be partly related to the 'high volume' TPGS formulation used in the study .The final artefenomel and piperaquine population PK models, including details of the analysis and model diagnostics, are provided in Additional file The PK of both artefenomel and piperaquine in adult and paediatric patients could be described by three compartment disposition models. All (apparent) clearance and volume parameters were related to body weight allometrically. Additional covariates identified for the PK of artefenomel were vomiting, artefenomel dose and age. In particular, relative bioavailability was a function of age; it was 40% lower for a patient of 1\u00a0year versus 20\u00a0years of age. For the PK of piperaquine, the only additional covariate identified was vomiting. No covariate effects of region, age (for piperaquine), sex, protocol-defined non-compliance or actual or adult equivalent PQP dose were identified.Individual artefenomel and piperaquine exposures were estimated for 427 and 426 patients respectively (including patients who vomited and were not successfully re-dosed). Summaries of the individual estimated exposures by region and age group are provided in Additional file Geometric mean artefenomel exposure was lowest in the African population 0.5 to\u2009\u2264\u20092\u00a0years of age (which includes body weight bands 5\u201314.9\u00a0kg). Thus, AUCinf and Cday7 were 7.6\u00a0\u03bcg*h/mL  and 2.0\u00a0ng/mL (CV 147%) respectively compared with 10.0\u00a0\u03bcg*h/mL (CV 111%) and 3.3\u00a0ng/mL (CV 141%) for African patients\u2009>\u20095\u00a0years. Asian patients\u2009>\u20095\u00a0years  had higher mean exposures than the African patients; AUCinf of 16.9\u00a0\u03bcg*h/mL (CV 66%) and Cday7 of 5.1\u00a0ng/mL (CV 95%) Fig.\u00a0. ArtefenPiperaquine exposures increased approximately proportionally with dose, although there was considerable overlap between the treatment arms. Exposures tended to be lower in the African population\u2009\u2264\u20092\u00a0years of age (including body weight band 5\u20139.9\u00a0kg) and in the Asian population Fig.\u00a0. ExposurWhen exploring the observed ACPR28 by artefenomel exposure bins  rather than dose, the relationship between exposure and ACPR28 is clearly visible  and region (interaction between region and artefenomel Cday7) (p\u2009=\u20090.002):The probability of achieving ACPR28 (pACPR) was found to be a function of artefenomel and piperaquine exposures, the baseline parasitaemia (p is the probability of ACPR28 and BasePar is the baseline parasitaemia (parasites/\u03bcL).where There was no statistically significant influence of either Kelch13 genotype or age on pACPR, and (once region was in the model) there was no additional influence of presumed immunity status. In addition, no interaction between the exposures of the two drugs was identified.Three-dimensional (3D) graphical representations of the exposure\u2013ACPR28 model are shown in Fig.\u00a0This difference in sensitivity to artefenomel (about five times lower in Asian versus African patients) is illustrated by the difference in the isoboles plotted in Fig.\u00a0For example, if each drug were administered by itself (Cday7\u2009=\u20090 for the partner drug), assuming a baseline parasitaemia of 100,000 parasites/\u03bcL in African patients, an artefenomel Cday7 of 8\u00a0ng/mL would be required for a 0.95 probability of achieving ACPR28, compared to 40\u00a0ng/mL in Asian patients. For each presented scenario, any exposure combination of artefenomel and piperaquine to the right of the isobole lines is associated with\u2009>\u20090.95 probability of achieving ACPR28.Figure\u00a0The projected ACPR28 simulations for various dose combinations, taking into account between-subject variability as well as model parameter uncertainty in both PK and exposure response, are shown in Table\u00a0P. falciparum malaria in adults and children in Africa and Asia (Vietnam).We report the results of the first phase II dose-ranging study to assess the potential of a single encounter curative treatment (artefenomel plus PQP) for uncomplicated None of the treatment arms reached the pre-specified target efficacy of\u2009\u2265\u200995% PCR-adjusted ACPR28 in the overall population or in any of the subpopulations, including African patients\u2009>\u20095\u00a0years of age, demonstrating that a single encounter treatment with artefenomel 800\u00a0mg plus up to 1440\u00a0mg PQP does not provide sufficient exposure for a sufficient duration to achieve the required efficacy. Efficacy appeared lower in Vietnamese than in African patients overall; however, the lowest efficacy was observed among the youngest African age group (>\u20090.5 to\u2009\u2264\u20092\u00a0years old).The study was not powered to compare outcomes between treatment arms, and no clear PQP dose trend was identified in the primary analysis for ACPR28. This was due to large exposure variability and the limited dose range studied, resulting in overlapping exposures between treatment arms, coupled with the binary nature of the clinical endpoint. However, this large exposure range allowed establishment of an exposure\u2013response relationship for both drugs in combination, which in turn resulted in identification of factors influencing efficacy, thereby providing a fuller understanding of the study results.Thus, the exposure\u2013response analysis demonstrated that both drugs contribute to efficacy (ACPR28) in a concentration-dependent manner, and as might be expected, higher baseline parasitaemia requires higher exposures to provide the same ACPR28.Within Africa, the concentration\u2013response relationship for artefenomel and piperaquine did not appear to differ with age (or presumed immunity). While age was not identified as a significant covariate for ACPR28, the numbers of African patients\u2009>\u20095\u00a0years old was relatively small and may have been insufficient to identify a difference. Instead, the lower efficacy in the youngest African patients appeared due to lower exposure to artefenomel (and to a lesser extent piperaquine). There was insufficient information to be able to identify the reason for the lower exposure. This may have been a consequence of incorrect dose adjustment to account for clearance/bioavailability differences across the age range; however, failure of the youngest children to consume the entire dose  or vomiting may have contributed.The exposure\u2013response analysis indicated that the lower efficacy (ACPR28) in the Vietnamese relative to the African population was due to lower sensitivity to artefenomel (but not to piperaquine), that is, a regional difference in the concentration\u2013response relationship. The mechanism of this lower sensitivity is not known.Kelch13 genotyping indicated a high frequency of patients infected with artemisinin resistant parasites within the Vietnamese sites. The most common Kelch13 genotypes were C580Y, the predominant validated marker of artemisinin resistance across the Greater Mekong Subregion, and P553L, a candidate marker of artemisinin resistance found in the Western Greater Mekong Subregion .Artemisinin resistance is characterised by a decrease in the rate of parasite clearance following artemisinin mono-therapy or artemisinin-containing combination treatments in patients infected with parasites with mutations in the Kelch13 gene . In vitrThe current study is the first to fully evaluate the efficacy of a combination containing a synthetic endoperoxide (artefenomel) in patients infected with artemisinin resistant parasites, and the association between PCt1/2 following artefenomel/PQP treatment and Kelch13 mutation suggests that these mutations may drive a similar decrease in the rate of parasite clearance for artefenomel-containing combinations as with artemisinin-containing combinations . In vitrP. falciparum parasites collected in this study is not currently known. Recent work to identify genetic markers of piperaquine resistance has confirmed that increased copy numbers of plasmepsin 2 and plasmepsin 3 genes, along with Pfmdr1 gene de-amplification, are independently associated with resistance to piperaquine, and that these markers of piperaquine resistance are prevalent in Cambodia and coexist with Kelch13 [High rates of DHA\u2013piperaquine treatment failures are now reported in the Greater Mekong Subregion, suggesting co-segregation, or at least coexistence of artemisinin and piperaquine resistance. Three of the four sites in Vietnam involved in the study  are located in provinces bordering Cambodia, and so conceivably artemisinin and piperaquine resistant genes could coexist. It is noteworthy that no difference was detected in the sensitivity (exposure\u2013response relationship) for piperaquine between Vietnam and Africa. However, the sensitivity to piperaquine of  Kelch13 , 32. We We are also currently investigating the efficacy of artefenomel in combination with ferroquine. Both parent drug and circulating active metabolite have significantly longer half-life values than piperaquine, and hence this combination has a greater probability of achieving the target efficacy. Ex vivo studies performed with Cambodian clinical isolates suggest negligible impact of piperaquine resistance on ferroquine potency .The significant rate of vomiting and the high dosing volume, particularly for young children, were problematic and may have contributed to the lower drug exposures in the youngest children. The development of age-appropriate formulations is key to the success of individual studies and to development programmes as a whole. Asymptomatic QTc increases from baseline were frequently reported as expected for PQP.The study results also illustrate the challenges in developing a single encounter combination treatments. Firstly, the administered dose needs to be higher to achieve the required duration of exposure compared to multiple day treatments; therefore, the ratio of Cmax to overall exposure will be greater. Secondly, high between-subject variability in drug exposures, due in part to a limited number of body weight bands for dosing , in addition to a large between-subject variability in baseline parasitaemia and potentially parasite sensitivity, means that the majority of patients will be required to be 'overdosed' if a very high cure rate is to be achieved with a single  dose level. Both factors mean that a wide therapeutic window is required.A significant limitation of the study was that, although the formulation used had been tested in adult healthy subjects, it had not been tested in adult and, more importantly, paediatric malaria patients. It is possible that the palatability of the formulation and/or volume of administration contributed to the higher than expected rate of vomiting in the study. In addition, although compliance data on drug consumption were collected, insufficient detail was recorded to truly capture compliance, since despite a high reported success rate of drug administration, there were anecdotal reports that young children were unable to ingest the full dose. The effect of this was that the study was unable to conclude on the best weight-based dose adjustment for patients weighing\u2009<\u200935\u00a0kg; i.e. drug exposure in young children was lower than in adults. However, it could not be concluded whether this was due to the weight-based dose adjustments or because the intended dosage was not successfully administered.However, the dose\u2013response simulations suggest that, even if the young children had similar exposures to the adults as well as no vomiting, the target efficacy would not have been achieved.None of the treatment arms reached the target efficacy of\u2009>\u200995% PCR-adjusted ACPR at day 28. Achieving very high efficacy following single dose treatment is challenging since\u2009>\u200995% of the population must have sufficient concentrations to achieve cure across a range of parasite sensitivities and baseline parasitaemia levels, meaning that a significant number of patients are 'overdosed'. Drugs with a substantial therapeutic window are therefore required. Projected single dose combination doses of artefenomel plus PQP that may achieve the target efficacy in the African population were not markedly higher than those tested in this study, but are most likely not clinically viable due to tolerability and practical dose size.A high frequency of patients in Vietnam were infected with artemisinin resistant parasites, and an association between PCt1/2 following artefenomel/PQP treatment and Kelch13 mutation suggests that these mutations may drive a similar decrease in the rate of parasite clearance for artefenomel-containing combinations as observed with artemisinin-containing combinations .While challenging, the development of tools suitable for deployment as single encounter curative treatments for adults, and particularly children in Africa, and to support elimination strategies remains a key development goal. There is currently no evidence that full artemisinin resistance has emerged in Southeast Asia; i.e. there is no evidence that reduced sensitivity of the early ring stage has progressed to full resistance . The resAdditional file 1: S1.Study Protocol. (PDF 1822 kb)Additional file 2: S2.Statistical analysis plan. (PDF 1515 kb)Additional file 3: S3.Pharmacokinetic analysis details. (DOCX 714 kb)Additional file 4: S4.Exposure\u2013response analysis details. (DOCX 274 kb)"}
+{"text": "Tafenoquine (TQ) is an 8-aminoquinoline anti-malarial being developed for malaria prophylaxis. It has been generally assumed that TQ, administered prophylactically, acts primarily on the developing exoerythrocytic stages of malaria parasites , and that polymorphisms in metabolic enzymes thought to impact the activity of other 8-aminoquinolines also inhibit this property of TQ. Furthermore, it has been suggested that a diagnostic test for CYP2D6 metabolizer status might be required. In field studies in which metabolic status was not an exclusion criteria, TQ has been shown to exhibit similar prophylactic efficacy as blood schizonticidal drugs (mefloquine). Also, its blood schizonticidal and anti-relapse efficacy is independent of 2D6 metabolizer status. The most reasonable explanation for the field study results, supported by other clinical and non-clinical data, is that TQ is not completely causal and exhibits substantial blood schizonticidal activity at the intended dose. Pharmacokinetic simulations demonstrate that trough concentrations of TQ exceed the proposed MIC of 80\u00a0ng/ml in\u00a0>95% of individuals. Based on these data a companion diagnostic for CP450 enzyme status is not required.The online version of this article (doi:10.1186/s12936-017-1862-4) contains supplementary material, which is available to authorized users. Plasmodium falciparum in individuals with certain enzyme deficiencies [Tafenoquine is a long\u2013half-life analog of primaquine being developed for travellers and community malaria prophylaxis by the United States (US) Army, 60 Degrees Pharmaceuticals, and their partners. Because of its long half-life (approximately 13\u00a0days), convenient dosing regimens, effectiveness against hypnozoites, broad spectrum activity against all species of malaria, and the absence of documented drug resistance, tafenoquine represents a potential improvement over the standard-of-care  in certain settings. However, the appropriateness of primaquine, and by inference tafenoquine, for prophylaxis in nonimmune subjects has been questioned, because of a perception that these agents may not arrest the development of 100% of the pre-erythrocytic stages of Plasmodium vivax but not those of P. falciparum [Primaquine is recommended for malaria prophylaxis at a dose of 30\u00a0mg/day . Primaqulciparum \u20136. Thus,P. vivax hypnozoites following primaquine administration. Similarly, in animal models, CYP2D6 knockout mice are not protected from Plasmodium berghei by primaquine following a sporozoite challenge [It has been hypothesized that a primaquine metabolite responsible for hypnozoite killing is generated by a CYP2D6-dependent pathway , and thahallenge , which shallenge  and somehallenge , 10. BecIn fact, as will be argued in the remainder of this review, the available evidence suggests that tafenoquine exhibits substantial blood schizonticidal activity, and by virtue of its long half-life, clears asexual blood stages in animals and humans, albeit more slowly than traditional blood schizonticidal drugs. Animal studies suggest that the blood schizonticidal effect of tafenoquine is independent of cP450 2D6 enzyme status. Furthermore, because tafenoquine, inherently, may not kill all developing exoerythrocytic parasites in all individuals at the intended dose, its blood schizonticidal effect makes an important contribution to its prophylactic efficacy at the intended dose (200\u00a0mg).P. vivax attack rate during the period of the deployment was retrospectively estimated to be 6.88% [In the pivotal Phase III study , 654 nonbe 6.88% . The subbe 6.88% . The subi.Tafenoquine exhibits no blood schizonticidal activity in humans,ii.Tafenoquine kills no developing exoerythrocytic stages in individuals with a low CYP2D6 metabolizer phenotype,iii.Tafenoquine kills all developing exoerythrocytic stages in a proportion of individuals with a normal CYP2D6 phenotype that is the same proportion of non-immune individuals that would be expected to be protected by a standard anti-malarial in this research setting (100% for mefloquine ).The following was assumed for the purposes of conducting a thought experiment:P. vivax attack rate was 6.88%. In such a study 1.7, 3.4 or 4.4 P. vivax cases should have been observed in the tafenoquine arm if the background rate of low metabolizer CYP2D6 phenotypes was 5, 10 or 13%, respectively.It can be further assumed that the pivotal study was repeated with the same sample size in the tafenoquine arm and included a placebo group where the However, in the actual Phase III study this was not the case: the point estimate of efficacy  for tafenoquine was: 100% (93\u2013100%).P. falciparum) 5.06 cases/person-year [In two Phase II studies from which the data were pooled, 519 African residents of high malaria transmission areas were randomly assigned to receive placebo (n\u00a0=\u00a0187), tafenoquine (n\u00a0=\u00a0190), or mefloquine (n\u00a0=\u00a0142) during a period in which the incidence density in the placebo group was .The following was assumed for the purpose of conducting a thought experiment:P. falciparum attack rate was 5.06 cases/person-year. In such a study, 22, 32 or 62 P. falciparum cases should have been observed in the tafenoquine arm if the background rate of low metabolizer CYP2D6 phenotypes was 5, 10 or 20%, respectively.It can be further assumed that the pooled Phase II was conducted with the same sample sizes and a However, this was not the case. The observed number of cases in the actual tafenoquine arm was only 14 and the prophylactic efficacy of tafenoquine was 93.5% (89\u201396%) .P. vivax and P. falciparum in addition to its substantial causal activity that is unaffected by cP450 metabolizer status. In the remainder of this section the literature was reviewed to determine which one of these hypotheses is most reasonable.Neither of the field studies discussed above excluded any individuals who may have had alternate metabolizer status associated with CYP2D6 or any other type of metabolic enzyme. There is no evidence from field studies that the efficacy of continuous treatment-compliant prophylaxis with tafenoquine is any different from treatment-compliant prophylaxis with mefloquine. Therefore, the assumptions made in the thought experiments above must be incorrect. The field study results can be explained only if continuous prophylaxis is completely causal or if tafenoquine exhibits substantial blood schizonticidal activity against both P. berghei parasite expressing the bioluminescent reporter protein luciferase. Drug activity against developing liver stages was assessed using a real-time in vivo imaging system and suppressive activity against erythrocytic stages was assessed using flow cytometry. When one dose of tafenoquine (5\u00a0mg/kg) was administered 1\u00a0day prior to sporozoite inoculation, merozoite leakage was not detected by flow cytometry through day 30 in 10 of 10 animals. However, in these animals real time imaging demonstrated that only 98.6% of exoerythrocytic parasites were eliminated compared to controls at 48\u00a0h after sporozoite challenge. Because parasites begin to emerge from the liver to enter the blood at 48\u00a0h in this model, some of the remaining 1.4% of liver parasites had entered the blood after that time and the 100% prophylactic efficacy of this regimen was due to elimination of a few blood parasites by tafenoquine remaining in the blood after 48\u00a0h. These data imply that at this particular dose in mice, both causal and blood schizonticidal activity are important for the prophylactic efficacy of tafenoquine. It is also true that at higher doses (e.g. 5\u00a0mg/kg/day for 3 days), 100% prophylactic efficacy was observed following 100% inhibition of liver schizogony. This means that it might also be possible in humans, assuming it could be tolerated, to administer a dose of tafenoquine at which prophylactic efficacy was mediated only via causal activity.In humans, it is not possible to definitively assess the relative contribution of causal and blood schizonticidal activity to the prophylactic efficacy of tafenoquine. However, animal models provide important clues. Li et al.  examinedP. falciparum challenge [The first challenge study involving tafenoquine attempted to position the drug as a causal prophylactic. Four nonimmune healthy volunteers were given a single 600-mg dose immediately prior to a mosquito-induced hallenge . The intPlasmodium species at relevant doses.Several lines of evidence suggest that tafenoquine acts as a slow-acting suppressive prophylactic against both animal and human Plasmodium cynomolgi in Macaca monkeys within 3\u20134\u00a0days and at doses of 6\u00a0mg/kg/day for 3\u00a0days there was no relapse or recrudescence [First, tafenoquine administered at doses\u00a0\u22652\u00a0mg/kg/day for 3\u00a0days cleared the erythrocytic stages of descence . Dow et descence . Assumindescence ), meaninAotus monkeys, doses of 1\u00a0mg/kg/day for 3\u00a0days cleared the erythrocytic stages of an established chloroquine-resistant P. vivax with recrudescence 20\u201325\u00a0days later [Macaca monkeys that is presented in Table\u00a0Aotus monkeys. Therefore, at an HED of 120\u00a0mg per day\u00a0\u00d7\u00a03, tafenoquine is able to clear an established chloroquine-resistant P. vivax infection, while at a dose of around 360\u00a0mg per day\u00a0\u00d7\u00a03, the drug is able to cure such an infection. This confirms that clearance of established P. vivax infections should be possible at the anticipated loading dose in humans.Second, in ys later . Doses oP.\u00a0falciparum challenge is not usually longer than day 11. A reasonable interpretation of this data is that tafenoquine at concentrations greater than 48 ng/ml are able to suppress the amplification of low-density P. falciparum infections.Third, Brueckner et al. , in the P. vivax cases with drug levels of 20 and 21\u00a0ng/ml and a single case of P. falciparum with a tafenoquine concentration of 38\u00a0ng/ml  blood stage infections at exposure levels equivalent to the intended loading dose, and to control sub-patent low density infections at relatively low drug concentrations (>50\u00a0ng/ml) in vivo.In a recent study, Milner et al.  demonstrThe most parsimonious explanation for all the available data is that continuous tafenoquine prophylaxis at the intended dose protects non-immune individuals from contracting symptomatic malaria by (i) eliminating most, but not all, developing asexual exoerythrocytic stages following an infectious mosquito bite, and (ii) slowly clearing the few erythrocytic stages that leak out of the liver in some individuals in a manner that is independent of cP450 2D6 metabolizer status. There is, therefore, no need for a cP450 2D6 companion diagnostic.Based on the association of symptomatic breakthroughs with measured plasma concentration discussed above, Edstein et al.  proposed50 of 97\u00a0ng/ml and a mean IC99 of 3108\u00a0ng/ml against 44\u00a0multiple-drug-resistant strains of P. falciparum from the Thai-Burmese border in a schizont maturation assay [50 of 64 to 110\u00a0ng/ml against 2 drug resistant strains of P. falciparum from South East Asia. Tafenoquine was found to exhibit an IC50 of 2041\u00a0ng/ml amongst 162 African isolates of P. falciparum [P. falciparum.However, this does not seem to make sense based on in vitro susceptibility data, since the IC50s, and most importantly the IC90/99s, reported for tafenoquine, are generally higher than the proposed MIC in vivo. Ramharter et al. reported that tafenoquine exhibits a mean ICon assay . Ohrt eton assay  reportedlciparum . Vennerslciparum  reportedThe counter argument is that endpoints measured in vitro, and the conditions utilized to generate them are vastly different from the relevant in vivo pharmacodynamics endpoints for malaria prophylaxis. First, the in vitro assays conducted to date all exhibit short exposure formats (48\u00a0h) relative to the parasite clearance time observed in vivo. Second, in vitro assays executed to date utilize much higher initial parasite densities than those after initial merozoite release from the liver in a nonimmune subject who has been administered prophylactic treatment. Third, the in vitro assays conducted do not account for either (i) any metabolic activation outside red blood cells (if this is required for activity) and (ii) cannot replicate possible deleterious effects of tafenoquine on parasites that survive exposure in the liver but escape into the blood stream. In fact, it may not be possible to replicate all the relevant in vivo parameters in an in vitro assay system.It should be stated that the mechanism(s) of action of tafenoquine against blood stage parasites are not known, and come from a single study reported by Vennerstrom et al. . In thatFirst, tafenoquine, but not primaquine, was shown to inhibit haem polymerization in vitro at concentrations lower than chloroquine (16\u00a0v\u00a080 microM ). HoweveSecond, Vennerstrom et al.  also proP. falciparum in non-immune subjects. The lack of availability of such data has largely been for ethical concerns due to the slow parasite clearance time of tafenoquine. However, two approaches that could generate such data can be envisioned. The first is utilization of a well-characterized blood stage challenge model. Such a model would avoid the confounding effect of the causal activity of tafenoquine that would be encountered with a sporozoite challenge. Additionally, the parasite density following inoculation is more in line with the density that would be encountered after merozoite release during prophylaxis. The second approach would be, in a treatment study enrolling P. falciparum patients, administration of tafenoquine in combination with a dose of artesunate that clears but does not cure. Tafenoquine should clear up the residual parasite burden in a manner analogous to that for mefloquine.The existing clinical efficacy database does not include any studies that directly show clearance of P. vivax in humans requires 2D6 activation. This implies a common mode of action of killing of P. vivax hypnozoites and the exoerythrocytic schizonts of P. falciparum by primaquine. Animal data suggest that 2D6 deficiency also arrests the causal activity of tafenoquine. However, St Jean et al. [P. vivax relapse and incidence of cP450 2D6 intermediate metabolizer status, implying different modes of causal and hypnocytocidal action for tafenoquine. However, these data should be treated cautiously until additional information becomes available. The St Jean et al. study contained few poor metabolizers [As discussed in the introductory paragraphs, it appears as if the causal prophylactic efficacy of primaquine in animals, and anti-relapse efficacy against n et al.  did not bolizers . Also, tbolizers , 29, 30,P. falciparum in non-immune subjects, and further mechanistic studies would be helpful, though not essential for a complete understanding of tafenoquine\u2019s prophylactic effect.Concern has been expressed in the literature regarding the use of tafenoquine for malaria prophylaxis due to the assumption that (i) it, like primaquine, is not 100% causal in individuals with certain cytochrome P450 enzyme deficiencies and (ii) that it exhibits little blood schizonticidal activity. However, although tafenoquine was not universally causally prophylactic in a challenge study following a single dose, it exhibits prophylactic efficacy following continuous dosing that is equivalent to that of mefloquine  in field studies in which study subjects were not excluded on the basis of metabolic enzyme status. It appears plausible based on non-clinical and clinical efficacy data and pharmacokinetics that the robust efficacy of the drug is due to a slow-acting blood schizonticidal effect in addition to its substantial although incomplete causal prophylactic effect. This effect, in animals, is independent of cP450 2D6 isoenzyme status, which suggests that, other than the routine testing required for G6PD deficiency, no companion diagnostic test is required. Additional studies to demonstrate directly clearance of It has been proposed that tafenoquine be administered by mass treatment to more rapidly eliminate malaria in endemic countries . Dow et"}
+{"text": "Prompt, effective treatment of confirmed malaria cases with artemisinin-based combination therapy (ACT) is a cornerstone of malaria control. Maximizing adherence to ACT medicines is key to ensuring treatment effectiveness.This open-label, randomized trial evaluated caregiver adherence to co-formulated artemether\u2013lumefantrine (AL) and fixed-dose amodiaquine\u2013artesunate (AQAS) in Sierra Leone. Children aged 6\u201359\u00a0months diagnosed with malaria were recruited from two public clinics, randomized to receive AL or AQAS, and visited at home the day after completing treatment. Analyses were stratified by site, due to differences in participant characteristics and outcomes.Of the 784 randomized children, 680 (85.6%) were included in the final per-protocol analysis . Definite adherence (self-reported adherence plus empty package) was higher for AL than AQAS at both sites . However, self-reported adherence (ignoring drug package inspection) was higher for both regimens at both sites and there was no strong evidence of variation by treatment . In Site 2, correct treatment (correct dose\u2009+\u2009timing\u2009+\u2009duration) was lower for AL than AQAS . In both sites, more caregivers in the AQAS arm reported adverse events .Self-reported adherence was high for both AL and AQAS, but varied by site. These results suggest that each regimen has potential disadvantages that might affect adherence; AL was less likely to be taken correctly at one site, but was better tolerated than AQAS at both sites. Measuring adherence to anti-malarials remains challenging, but important. Future research should focus on comparative studies of new drug regimens, and improving the methodology of measuring adherence.Trial registration: Clinicaltrials.gov, NCT01967472. Retrospectively registered 18 October 2013, https://clinicaltrials.gov/ct2/show/NCT01967472The online version of this article (10.1186/s12936-018-2370-x) contains supplementary material, which is available to authorized users. Universal coverage of diagnostic testing and prompt, effective treatment with artemisinin-based combination therapy (ACT) are key malaria control strategies , 2. HoweMalaria remains a major health problem in Sierra Leone, exacerbated by the Ebola outbreak in 2014, which overwhelmed an already fragile health system . In 2004Only three studies have compared the adherence to multiple ACT regimens in Africa; however, the primary outcome of all three studies was treatment effectiveness; adherence was evaluated as a secondary outcome \u201324. OnlyThe study was conducted at two government-run outpatient facilities in Freetown, Sierra Leone Fig.\u00a0. Both siChildren were enrolled in the study, if they met the following inclusion criteria: (1) age 6\u201359\u00a0months; 2) complaint of fever or history of fever; (3) living within 5\u00a0km of health facility; (4) no evidence of severe malaria or danger signs   complain, 25; (5)Following enrolment, study participants were subject to the standard of care provided at the health centres. All participants first underwent testing for malaria using rapid diagnostic tests (RDTs) at the health centre laboratory, and then saw a consulting health worker for a clinical evaluation. Study staff recorded RDT results onto case record forms, and observed patient-provider consultations for all participants. Children who had a positive RDT and were diagnosed with malaria by the health worker were randomly assigned to receive treatment with either AL or AQAS and were scheduled for a home visit on Day 4. If the RDT was negative for malaria, the child was excluded from the study and was provided standard care by the health worker.\u00ae: Novartis) was procured by the research team and provided to both sites for the purpose of this study. Fixed-dose AQAS (Winthrop\u00ae: Sanofi-Aventis) was available at both study sites through the standard government supply chain system. All study medications were produced by manufacturers pre-qualified by the World Health Organization (WHO) and approved by the Pharmacy Board of Sierra Leone, and were prescribed according to standard national and WHO treatment guidelines [Co-formulated AL  received 1 tablet per dose (infant dose), and those aged 12\u201359\u00a0months of age (or weighing >\u200915 to 20\u00a0kg) received 2 tablets per dose (child dose). Participants randomized to treatment with AQAS received one tablet dosed appropriately for age (or weight) once daily for 3\u00a0days. Children aged 6\u201311\u00a0months (or weighing 4.5\u20138\u00a0kg) received the infant dose (67.5\u00a0mg AQ/25\u00a0mg AS tablets) and children aged 12\u201359\u00a0months (or weighing 9\u201317\u00a0kg) received the child dose (136\u00a0mg AQ/50\u00a0mg AS tablets). Caregivers were responsible for administering the treatments to their child as instructed by the health worker.A computer-generated randomization list (in blocks of 10) was created for each site by a member of the team who was not directly involved in patient recruitment, consultation or follow-up. Prior to study initiation, individual treatment allocation slips were prepared from the randomization list. These were sealed into sequentially-numbered, opaque envelopes containing the treatment group assignments. The consulting health worker opened the envelopes and assigned the treatment number and corresponding treatment at the time of prescription. Health facility nurses were responsible for dispensing the study medications according to the assigned study number. Study medications were not identical in appearance or taste nor were the tablets per dose the same. The participants, health workers, and study team were not blinded to the treatment assignments.Study participants were visited at home 4\u00a0days after their clinic visit; the day after the last treatment dose should have been taken. The purpose of the follow-up visit was explained to the caregivers, and additional written informed consent for the interview was obtained; participation was voluntary . If the A semi-structured questionnaire was administered to caregivers to assess adherence to treatment, the characteristics of the child and caregiver, knowledge of malaria, household characteristics, and wealth indicators. Caregivers were asked to describe the treatment, including how the medication was administered and any adverse events, and to show the original medication packaging. If the packaging was available, any remaining tablets were tallied and recorded onto the questionnaire. If treatment had not been completed at the time of the follow-up visit, the caregiver was encouraged to complete the full treatment course or, if that was not possible, to return to the health facility.In 2010, the prevalence of adherence to co-packaged AQ\u2009+\u2009AS in Sierra Leone was estimated to be 50% , 28. At Using previous ACT adherence studies as a guide, the primary outcome of caregiver adherence was based on self-reports of completion of treatment and, when possible, verified by package inspection , 29, 30.As the main outcome variable was derived from two different measurements, the variable was recoded to produce two binary variables representing each component. These included: (1) \u201cself-reported adherence\u201d, which does not consider package availability ; and (2) \u201cpackage-based adherence\u201d, which does not consider self-reported adherence .Secondary outcomes were adherence to the prescribed number of doses, time-schedule and duration of treatment. The correct number of doses was defined as receiving the prescribed number of tablets, as indicated. Correct timing was defined as receiving the ACT at the prescribed intervals (twice daily for AL and once daily for AQAS). Correct duration was defined as receiving the ACT for the recommended number of days (3\u00a0days for both regimens). Correct treatment was defined as the composite of the three above indicators: correct dose\u2009+\u2009correct timing\u2009+\u2009correct duration, in which all three factors were met.Data were recorded on paper forms and entered into a database created in Epi Info\u2122 7.1.2.0 (Centers for Disease Control and Prevention (CDC), Atlanta, GA USA). Data were double entered into tablets using the Epi Info Companion for Android mobile application. Statistical analysis was performed using Stata 12 .Although intention-to-treat analysis is the preferred analytic approach for randomized controlled trials, a per-protocol analysis was favoured for this trial as the primary outcome was adherence. The main objective of this study was to assess the behaviours of caregivers related to the specific ACT received at the health facility. Therefore, the primary analyses were carried out using the per-protocol population, in which only children who received ACT as per the randomization schedule and had outcome data were included. Children who did not receive the correct ACT regimen based on the randomization list were excluded from this analysis. We also conducted and present the intention-to-treat analysis for comparison with the per-protocol findings, to assess whether results from the two analytic approaches were similar. Participant\u2019s characteristics and adverse events were tabulated by study site and randomization group in the per-protocol population. The wealth index was created using principle component analysis (PCA) .t-tests. Measures of effect (odds ratios-OR) were calculated using logistic regression for binary outcomes and ordinal logistic regression for multinomial outcomes along with the 95% confidence intervals and associated p-values. For the ordinal logistic regression model, the proportional odds assumption was tested with a likelihood ratio test.Chi squared or Fisher\u2019s exact tests were used to compare categorical data, and continuous data were tested using Student\u2019s or Welch\u2019s https://clinicaltrials.gov/ct2/show/NCT01967472). All participants provided written informed consent at the time of recruitment and again prior to administration of the follow-up survey in their homes.The study protocol was approved by the London School of Hygiene and Tropical Medicine (LSHTM) Research Ethics Committee and the Sierra Leone Ethics and Scientific Review Committee. The trial was registered at ClinicalTrials.gov ; most caregivers (>\u200970%) spoke Krio  at both sites. In Site 1, caregivers in the AL arm were more educated than those in the AQAS arm . At both sites, approximately half of caregivers reported that health workers instructed them to finish the anti-malarial treatment. Caregiver knowledge about ACT was low at both sites, more so in Site 1.The Muslim religion was practised by substantially more households at Site 1 than at Site 2 . Households at Site 2 were significantly poorer than at Site 1, with 149 (44.7%) households assigned to the poorest category at Site 2 vs 87 (25.1%) in Site 1 (p\u2009<\u20090.001). Otherwise, there were no additional differences in characteristics of participants, caregivers or households, between trial arms, at either site.At both sites, the odds of definite adherence (defined as self-reported adherence in the presence of an empty drug package) were higher for AL than AQAS Table\u00a0. Self-reOverall, the quality of treatment was high Table\u00a0. At bothA total of 106 caregivers reported that their child experienced an adverse event to treatment. Significantly more adverse events were reported at Site 2 than at Site 1 . At both sites, significantly more caregivers in the AQAS arm reported adverse events Table\u00a0, with voWith progress on malaria control slowing and resistance to artemisinin resistance emerging, it is vital that every effort is made to protect the efficacy of ACT . PatientCurrently, the available evidence on ACT adherence is limited by variation in study designs and outcomes, differences in drug regimens, and lack of comparative studies . In prioThe one other study that directly compared adherence to AL and AQAS in Benin found that \u2018full adherence\u2019 to the two regimens was not significantly different . HoweverThis study also identified specific characteristics which may impact adherence to AL and AQAS. AL was less likely to be taken correctly at one site, but was better tolerated than AQAS at both sites. The complexity of the AL dosing regimen, including the number of tablets, twice daily dosing, the requirement to give the second dose 8\u00a0h after the first, and to administer with fatty food , has beeIn this study, adherence to AL and AQAS varied depending on the outcome definition applied. Although definitions of adherence outcomes, based on self-report of treatment completion plus package inspection, which have been used in previous anti-malarial studies were adopted , 29, 30,A variety of approaches have been used to incorporate package inspection in the classification of adherence outcome , 59, 60.Although package inspections and pill counts serve as a gold standard for measuring adherence to treatment of other diseases , 65 the This study had several limitations, in addition to the challenges with the adherence outcome classification. First, the characteristics of the participants, caregivers, and households enrolled in the two sites varied substantially, which was unexpected. Specifically, variations in the age of participating children, level of caregiver education, household religion and socioeconomic position were found, all of which may influence treatment adherence , 67, 68.Maximizing adherence to anti-malarial drug regimens is essential for ensuring treatment effectiveness; however measuring adherence remains challenging. The results from this study suggest that although self-reported adherence to both AL and AQAS was high, the difference between the two regimens was not significant. However, potential disadvantages were identified for each regimen that might impact optimal treatment adherence. With the emergence of resistance to artemisinins in Southeast Asia fuelling the development of new drug formulations, information on adherence to different ACT regimens will become increasingly important to help guide drug delivery, improve treatment effectiveness, and inform drug policy. However, the methodology of measuring adherence in anti-malarial studies requires further advancement. This study highlights the limitations of package inspection, and suggests that an outcome measure based on correct treatment could have greater utility. Standardizing methodologies for evaluating adherence across diverse contexts would improve the evidence base on ACT adherence and effectiveness.Additional file 1:Table S1. Package availability by drug and site"}
+{"text": "Kelch13 (k13) propeller domain.In Myanmar, three types of artemisinin-based combination therapy (ACT) are recommended as first-line treatment of uncomplicated falciparum malaria: artemether\u2013lumefantrine (AL), artesunate\u2013mefloquine (AS\u00a0+\u00a0MQ), and dihydroartemisinin\u2013piperaquine (DP). Resistance to both artemisinins and ACT partner drugs has been reported from the Greater Mekong Sub-region, and regular efficacy monitoring of the recommended ACT is conducted in Myanmar. This paper reports on results from studies to monitor the efficacy of the three forms of ACT in sentinel sites in northern Myanmar, and investigations of mutations in the k13 propeller domain were examined in dried blood spots collected on day\u00a00. The primary endpoint was adequate clinical and parasitological response (ACPR) on day 28 for AL and on day 42 for DP and AS\u00a0+\u00a0MQ, corrected to exclude re-infection using polymerase chain reaction (PCR) genotyping. Safety data were collected through self-reporting.Seven therapeutic efficacy studies were conducted in 2011\u201312 and 2014 in three sentinel sites in Myanmar . Three studies were done for the evaluation of AL (204 patients), two studies for AS\u00a0+\u00a0MQ (119 patients) and two studies for DP (147 patients). These studies were done according to 2009 standard WHO protocol. Polymorphisms in the k13 mutations (>440) above 23% in the day-0 samples. The F446I mutation was the most common mutation, making up 66.0% of the mutations found. Seven out of nine day-3 positive patients were infected with k13 wild type parasites. The remaining two cases with day-3 parasitaemia had the P574L mutation.PCR-corrected ACPR was 97.2\u2013100% for AL, 98.6\u2013100% for AS\u00a0+\u00a0MQ and 100% for DP across the study sites and years. All studies found a prevalence of k13 mutations associated with delayed parasite clearance. This study showed that already in 2012 there was a high frequency of k13 mutations in Myanmar on the border with India. The high efficacy of the recommended ACT gives confidence in the continued recommendation of the use of these treatments in Myanmar.The efficacy of AL, AS\u00a0+\u00a0MQ and DP remains high in northern Myanmar despite widespread evidence of Trial registration numbers ACTRN12611001245987 (registered 06-12-2011) and ACTRN12614000216617 (registered 28-02-2014) Plasmodium falciparum. Overall, this is a 74% reduction from the number of cases reported in 2010 [P.\u00a0falciparum: artemether\u2013lumefantrine (AL), artesunate\u2013mefloquine (AS\u00a0+\u00a0MQ) and dihydroartemisinin\u2013piperaquine (DP) [In 2014, Myanmar reported 152,195 malaria cases, 72% of them due to ine (DP) .Myanmar is the only country in the GMS with more than one recommended first-line ACT in a given geographical area. While AL is the treatment that is most frequently used in the public sector and by most international non-governmental organizations, the other ACT are available in the private sector, making monitoring of all three recommended ACT a necessity. Therapeutic efficacy studies (TES) for anti-malarial medicine are the gold standard, providing the information needed to guide treatment policies. Monitoring of molecular markers for resistance can provide supplementary information and is helping to improve the surveillance of resistance.All ACT contain an artemisinin and a partner drug. Resistance to artemisinin was first identified in clinical studies in 2006 close to the Cambodia\u2013Thailand border. However, retrospective analysis of molecular markers indicates that artemisinin resistance likely emerged as early as 2001. Artemisinin resistance is defined as delayed parasite clearance following treatment with an artesunate monotherapy or with an ACT. In TES, this is measured by the proportion of patients found positive on day 3 after treatment .P. falciparum Kelch13 (k13) gene [k13 mutations have been reported, and these different mutations can have varying effect on the clearance genotype. Studies have shown a clear distinction between the k13 mutations that are most frequent in eastern GMS  and western GMS  [Artemisinin resistance has been shown to be associated with point mutations in the propeller region of the 13) gene . At preshailand) .k13 mutations extending through much of Myanmar [In Myanmar, delayed parasite clearance in patients treated with an ACT was first seen in 2009 in southern Myanmar . Recent  Myanmar , 8. Howe Myanmar .k13 propeller domain.This paper reports on the results of seven therapeutic ACT efficacy studies of the three recommended ACT in 2011\u201312 and 2014 in northern Myanmar, and results from investigations of mutations in the Two studies were conducted in September 2011\u2013September 2012 to evaluate AL and AS\u00a0+\u00a0MQ in Muse in northern Myanmar, close to the border with China. In Tamu, close to the Indian border, two studies were done in June\u2013October 2012 to evaluate AL and AS\u00a0+\u00a0MQ. Three additional studies were done in May\u2013November 2014: two in Tabeikkyin to evaluate AL and DP, and one in Tamu to evaluate DP. Location of the sites is shown in Fig.\u00a0The study design was an open-label, one-arm, prospective evaluation of clinical and parasitological responses to directly observed treatment for uncomplicated malaria. The WHO protocol for monitoring therapeutic efficacy was used . In the Estimation of the target sample size in each study was based on an assumption of treatment failure of 5%, with a 95% confidence level and precision of 5%. Allowing for a 10% loss to follow-up, the target sample size in each study was 80 cases. Ethical approvals were obtained from the Ethics Review Committee of the Department of Medical Research, Yangon, Myanmar, and the WHO Ethics Review Committee, Geneva, Switzerland. The studies were registered at Australian New Zealand Clinical Trials Registry [P. falciparum mono-infection, a parasite density of 500\u2013100,000 asexual parasites/\u03bcl, and a body temperature \u226537.5\u00a0\u00b0C, or a history of fever during the previous 24\u00a0h. Pregnant women were excluded from the study, as were patients with signs of severe malaria or with febrile conditions due to diseases other than malaria.Cases were enrolled after obtaining a patient\u2019s, or parent\u2019s/guardian\u2019s written consent. Eligible patients were aged six years and above  with microscopically confirmed, uncomplicated \u00ae, Novartis, Switzerland) was administered at a target dose of 1.3/8\u00a0mg/kg twice daily for 3\u00a0days. For treatment with AS\u00a0+\u00a0MQ, artesunate  was given at a target dose of 4\u00a0mg/kg once a day for 3\u00a0days. Mefloquine  was given at a target dose of 15\u00a0mg/kg once on the first day followed by 10\u00a0mg/kg on the second day. DP  was given at a target dose of 2\u20132.4/16\u201319.2\u00a0mg/kg once a day for 3\u00a0days. Quality-controlled medications for the studies were provided by WHO Global Malaria Programme.AL , and any other day if the patient returned spontaneously and parasitological re-assessment was needed. The blood smears were stained with fresh Giemsa, and examined at a magnification of 1000\u00d7 to identify parasite species and determine parasite density. Asexual parasitaemia was determined from Giemsa-stained thick blood smears against the number of parasites per 200 white blood cells on day 0, based on an assumed density of 6000 white blood cells per \u00b5l of blood. Gametocytes were similarly enumerated. A blood smear was declared negative after examination of 1000 white blood cells . Qualitymsp-1), merozoite surface protein-2 (msp-2) and glutamate-rich protein (glurp) polymorphism, as per WHO methods [Dried blood spots were obtained for polymerase-chain reaction (PCR) at enrolment (day 0) and on follow-up days 7, 14, 21, and 28 (and 35 and 42 for AS\u00a0+\u00a0MQ or DP). PCR genotyping was performed on paired dried blood spots in the case of parasitaemia detected on or after day 7 to distinguish between recrudescence and re-infection. This PCR genotyping determined polymorphisms in merozoite surface protein-l , late clinical failure (LCF), late parasitological failure (LPF), or an adequate clinical and parasitological response (ACPR) .The primary outcome measure was PCR-corrected ACPR. The patients were excluded from the analysis if PCR results were not available to check for re-infection. If PCR results suggested re-infection, then the patient was excluded from the pre-protocol analysis.Information on ACT-related side effects was collected through self-reporting and recorded in the case reporting forms. EpiData software was used for double data entry  and analysis .Seven studies were conducted in three sentinel sites in Myanmar in 2011\u20132012 and 2014 to assess the efficacy of AL (three studies), AS\u00a0+\u00a0MQ (two studies) and DP (two studies). A total of 4919 patients were screened and 470 enrolled. Of these, 20 patients were lost to follow-up before day 28/42, three patients were withdrawn due to PCR-confirmed re-infection, one patient was excluded from the analysis with parasitaemia without PCR being able to determine re-infection or recrudescence, leaving 446 patients for evaluation at day 28/42  of the 470\u00a0day-0 samples. The main reason for failure to do sequencing was that for some patients too little blood was collected on the filter papers. Among the sequenced samples, the prevalence of k13 mutations (>440) was 33.7% (97/288) (Table\u00a0k13 mutations was in Thabeikkyin in 2014 (49.2%).The k13 sequencing was not possible. Sequencing showed that k13 genotypes in seven of the nine cases with parasitaemia on day 3 were k13 wild type parasites. The remaining two cases with day-3 parasitaemia had the P574L mutation. While the numbers are small, there is evidence (p\u00a0<\u00a00.05 using a Fisher\u2019s exact test) of significant differences in the proportion of day-3 positive cases by genotype.For the three cases with treatment failure, This paper reports on the therapeutic efficacy of all three recommended first-line ACT from sentinel sites in northern Myanmar. The efficacy of all ACT in the studies were high >97%) in both PCR uncorrected and corrected analyses, meeting the WHO recommendation that cure rates for falciparum malaria should be at least 90% 7% in bot, 13.k13 mutations (position\u00a0>440) were recorded in the studies. Artemisinin resistance was first identified in southern Myanmar close to the border with Thailand where the most extensive data are from. In the 2012 study in Tamu on the Myanmar-India border, k13 mutations were found in 28.9% of the samples, mainly the F446I mutation (25 of 28 k13 mutations). These are the earliest data documenting k13 mutations in this part of Myanmar.A total of eight different k13 mutations in India. Four non-synonymous k13 mutations were identified among 384 samples but only at very low frequencies . Of these four mutations, three  were detected in the northeastern states [Data from the Indian side of the border show that AL is highly efficacious . AL was introduced as first-line treatment in northeastern India after increasing treatment failures with artesunate\u00a0+\u00a0sulfadoxine\u2013pyrimethamine (AS\u00a0+\u00a0SP) due to resistance to SP. Data from 2010\u20132013 showed very low prevalence of n states . Of thesk13 mutations in Cambodia [The most common mutation in all the Myanmar study sites was the F446I. In Tamu in 2012, the prevalence of F446I was 25.8%. This remained nearly constant with the prevalence of F446I in Tamu in 2014 at 24.6%. One mutation in Tamu not found in 2012, but found in seven patients (12.3%) in 2014 was the R561H mutation. In Tabeikkyin in 2014, the N458Y mutation was identified in seven patients (11.1%). Both the F446I and the R561H mutations have been reported in other studies from northern Myanmar , 12, 15.Cambodia .k13 wild type parasites. The overall percentage of patients positive on day 3 is only 3.1%. However, among patients identified as having the P574L mutation, 33% (2/6) was found to have day-3 parasitaemia. Despite the low number of patients, there is evidence of significant differences (p\u00a0<\u00a00.05) in the proportion of day-3 positive cases by genotype.The prevalence of molecular markers for artemisinin resistance found in the studies has not resulted in a high prevalence of day-3 positivity, or a fall in efficacy of the ACT. The majority (7/9) of the day-3 positive cases were k13 mutation. The F446I mutation may be associated with only an intermediate artemisinin resistance phenotype, not resulting in a slowing of the parasite clearance to the same extent as mutations most prevalent in eastern parts of the GMS [The overall low proportion of day-3 positive patients could be for several reasons. The patients had significant lower parasitaemia on day 0 than in other research studies done in the area, which makes it less likely that parasites would be found on day 3, even if there are relatively slow parasite clearance rates. These studies do not allow for estimation of parasite clearance half-life as blood was collected only once a day rather than every 6 or 8\u00a0h. Another possible explanation for the low proportion of cases positive on day 3 is the prevalence of  the GMS . In addiP.\u00a0falciparum. So far studies done in all sentinel site of Myanmar showed high level of efficacy (>90%) to the three recommended ACT. The role of triple ACT for treatment of uncomplicated falciparum malaria can be determined only after the completion of ongoing ACT versus triple ACT studies in Myanmar. Nevertheless, continued monitoring will be needed. While there is no evidence for ACT partner drug resistance resulting in ACT failures in these studies, partner drug resistance is leading to ACT failures in other GMS countries, emphasizing the risks and the need to continue monitoring ACT efficacy.A high efficacy was found for all the three recommended ACT, giving confidence in the continued recommendation of these ACT as appropriate first-line treatments for Reporting of adverse events showed that among patients receiving AS\u00a0+\u00a0MQ, there was a slightly higher proportion of patients reporting one or more adverse events. This could affect the effectiveness of the treatment, as a higher proportion of patients on AS\u00a0+\u00a0MQ may choose to discontinue treatment before completion of the full three-day treatment.k13 mutations associated with delayed parasite clearance after treatment with an artemisinin. This study showed that already in 2012 there was a high frequency of k13 mutations in Myanmar on the border with India. The high efficacy of the recommended ACT gives confidence in the continued recommendation to use these treatments in Myanmar. The threat of resistance to ACT partner drugs resulting in falling ACT efficacy elsewhere in the region highlights the need for continual monitoring of ACT efficacy.All the three recommended ACT in Myanmar have been shown to be highly efficacious despite widespread evidence of"}
+{"text": "Primary study endpoint was the rate of adequate and parasitological response (ACPR) to therapy on day 28 (PCR-corrected). Day 0 isolates were analyzed to assess the presence of the PfCRT-76T CQ resistance marker. A total of 52 and 27 male adults were included in the CQ and Placebo group respectively. PCR-corrected ACPR was significantly higher in the CQ arm 89.4% (95%CI 80\u201398%) compared to the placebo (p\u2009<\u20090.001). CQ cleared 49/50 infections within the first 72\u2009h while placebo cleared 12/26 (LRT p\u2009<\u20090.001). The PfCRT-76T mutation was present only in one out of 108 (0.9%) samples at baseline, well below the 84% prevalence found in 1999 in the same area. This study presents preliminary evidence of a return of chloroquine sensitivity in Mozambican Pf isolates, and calls for its further evaluation in community-based malaria elimination efforts, in combination with other effective anti-malarials. Trial registration: www.clinicalTrials.gov NCT02698748.Recent reports regarding the re-emergence of parasite sensitivity to chloroquine call for a new consideration of this drug as an interesting complementary tool in malaria elimination efforts, given its good safety profile and long half-life. A randomized (2:1), single-blind, placebo-controlled trial was conducted in Manhi\u00e7a, Mozambique, to assess the  In 2001 the World Health Organization (WHO) recommended artemisinin-based combination therapy (ACT) as the first line treatment for uncomplicated malaria in countries where Plasmodium falciparum (Pf) malaria had become resistant to CQ4, and since then the use of CQ for the treatment of Pf has progressively been abandoned.In recent years, interest in the potential use of Chloroquine (CQ) has re-emerged, partly on account of the observation that the prevalence of molecular markers associated with CQ resistance among circulating parasites has decreased after discontinuing the use of the drug. The emergence of CQ resistance was first documented in Southeast Asia and South America less than a decade after its introduction as first-line treatment and spread to the East of Africa and the rest of the continent by the end of the 1970\u2019s5, inflicting them a disadvantage when the drug pressure reduces. Such finding implies that resistance might be reversible if drug use is discontinued. A point mutation in the 76th position of the P. falciparum CQ-resistance transporter gene (PfCRT) from a lysine in sensitive parasites (K76) to a threonine in resistant parasites (76\u2009T) has been associated with CQ-resistant falciparum malaria6. Analysis of the population frequencies of the mutations in PfCRT known to be associated with CQ resistance7 and assessment of in vitro activity of the drug9 in Malawian parasite isolates have indicated that CQ resistance may revert to sensitivity within a decade of withdrawal of the drug10. Similar drops, have been shown in Zambia11, Tanzania12 or coastal Kenya (Kilifi)13, years after CQ had been withdrawn. Translation of in vitro sensitivity to in vivo efficacy has also since then been demonstrated in Malawi14, where CQ efficacy in patients has been confirmed as very high14, and its use as monotherapy in a clinical trial context -albeit not a WHO recommendation- has also been shown to be similarly efficacious to its combination with other partner drugs15.Drug-resistant microorganisms are generally thought to suffer from a fitness cost associated with their drug-resistant traitPfCRT 76\u2009T prevalence of 84%16. However, a recent study performed in Gaza Province showed also an important decline of PfCRT K76T prevalence from more than 90% in 2006 to around 30% in 201017. A similar study conducted in a different part of the country also confirmed this tendency towards an increase of wild type parasites and thus CQ sensitivity18. The last available efficacy results for CQ derived from an in vivo study performed in Mozambique in the year 2001 showed that the day 28 efficacy was 47.1%19.In Mozambique, which borders in the west with Malawi, the national policy moved away from using CQ in 2003\u20134. A study conducted among children from Manhi\u00e7a District in 1999 revealed a P. falciparum in certain malaria-endemic settings of Sub-Saharan Africa. While this does not support the reintroduction of CQ as first line therapy in such settings at this point, it does suggest that, if proven sensitive in a given area, CQ could be considered as a complementary tool to interrupt transmission in the context of malaria elimination efforts.Altogether, these data suggest that, in the absence of drug pressure, CQ may be regaining sensitivity against PfCRT K76T molecular marker of CQ resistance among parasites among the study population in 2015.With this rationale in mind, we conducted a randomized placebo-controlled single blinded clinical trial to assess the efficacy of CQ (vs. placebo) to treat asymptomatic infections among healthy adult Mozambican volunteers. We complementarily present prevalence estimates of the The protocol for this trial and supporting CONSORT checklist, are available and annexed as supporting information.in-vivo study to take place since 2001, when CQ had been shown to be poorly efficaciuos. A placebo comparator was consequently used to accurately account for the effect that the high levels of immunity expected in the study population would have on natural parasite clearance of low parasitaemic infections20.Between January and June 2015, a randomized, single-blinded, placebo-controlled trial was conducted in the district of Manhi\u00e7a, southern Mozambique, to treat asymptomatic infections among healthy adults from the community. This study population was considered as the most ethical option for the first Centro de Investiga\u00e7\u00e3o em Sa\u00fade de Manhi\u00e7a (CISM), which currently provides accurate demographic information on its circa 178,000 inhabitants. The region has two distinct seasons \u2013 a warm and rainy season from November to April, and a cooler and drier season the rest of the year. Malaria transmission is perennial but shows marked seasonality, with Plasmodium falciparum (Pf) being the predominant species, and Anopheles funestus the main vector. Study participants were selected from areas within Manhi\u00e7a were clinical malaria in children was historical known to be high (above 40%) and asymptomatic infections in adults were expected in the community21.The district of Manhi\u00e7a counts with a demographic surveillance system (DSS) set up in 1998 by the Pf malaria infection with an asexual blood density \u2265100\u2009p/\u00b5L but <10,000\u2009p/\u00b5L. Key exclusion criteria included presence of any other co-existing clinical condition or symptom that in the opinion of the recruiting physician would not allow the individual to be considered a \u201chealthy\u201d asymptomatic carrier; axillary temperature >\u2009=\u200937.5\u2009\u00b0C; intake of any medication which may interfere with antimalarial efficacy or antimalarial pharmacokinetics, such as cotrimoxazole; history of hypersensitivity reactions or contraindications to CQ; and known HIV concomitant infection under antiretroviral treatment. Individuals aged 18 years or more satisfying the inclusion criteria were enrolled if they signed a detailed written informed consent. Individuals aged 16 were offered participation provided their legal guardians also assented for participation.The study population comprised adult males with microscopically confirmed, asymptomatic malaria infections. In order to find these subjects, a random list was generated from the DSS databases, and individuals were visited in their household, and screened for malaria through finger-prick with an HRP2-based rapid diagnostic test (RDT) and a blood slide. Blood slides were only read at CISM\u2019s laboratory in those cases found to be positive by the RDT (\u201cpre-screened positive\u201d), providing a final confirmation of malaria infection and density of parasitaemia. On the following day, such individuals were again visited at home, and only if they remained symptomless were offered to be included in the study. Study staff enquired about the presence of symptoms by asking a series of questions following a standardised clinical questionnaire. Asymptomatic malaria was defined as the absence of any proactively referred symptom of disease as referred by the individual, together with a documented axillary temperature <37.5\u2009\u00b0C. Symptomatic patients, irrespective of being infected or not, were assessed by the study clinician and referred to the health system if needed. Females were not screened on account of the Mozambican Ethics Committee\u2019s recommendation to avoid exposing to placebo malaria-infected women of child-bearing age which may be pregnant and thus prone to developing severe disease. Other inclusion criteria included 22.After the pre-screening visit (Day \u201c\u22121\u201d), a screening evaluation was conducted 24 hours later (day 0) to check all inclusion and exclusion criteria, with a particular emphasis on ensuring that symptoms had not appeared. Eligible patients were randomly assigned to receive CQ (Arm 1) or Placebo , according to a 2:1 (CQ:PL) scheme, so as to have more patients in the CQ arm to provide better estimates for its cure rates. Patients having received on day 0 a study drug, but confirmed by the blood slide to be malaria infection negative or <100 parasites/\u00b5L were considered protocol violations, and excluded from the analysis. A randomization list was generated using the online available randomizer software (http://www.randomizer.org/). Study staff carried the study medication and directly supervised the three-day long treatment. Chloroquine sulphate  was administered at a total dose of 25\u2009mg/kg . A similarly-looking placebo not containing any active principle, and prepared at the department of pharmacology of the Hospital Clinic, Barcelona, Spain, was administered to those allocated this intervention following an identical scheme. All treatments were directly observed for a minimum of 30\u2009min. Any subject vomiting during this observation period was re-treated with the same dose of either CQ or placebo, and observed for an additional 30\u2009min. Repeated vomiting implied withdrawal from the study and the use of rescue treatment. Treatment of symptomatic malaria infections or rescue therapy in cases of early or late treatment failure followed national Mozambican guidelines and included the use of artemether-lumefantrine (AL)Follow-up visits were planned on days 1, 2, 3, 7, 14, 21 and 28 after enrolment, or at any time point should the enrolled individual develop any symptoms of sickness. Individuals who discontinued either study drug were excluded from the study. Vital signs and body temperature were assessed during each follow-up visit. Adverse events were recorded and assessed for severity and association with study medication.23 which counts parasites against an assumed known blood volume. Density of P. falciparum was assessed from blood-spots collected in filter paper through real-time quantitative polymerase chain-reaction (qPCR) assay targeting 18S ribosomal RNA (rRNA)25. Blood spots for PCR analysis were collected using 3M Whatman\u2122 filter papers at baseline and at days 7, 14, 21 and 28, on the day of treatment failure or at any other unscheduled visit, and subsequently stored at 4\u2009\u00b0C in plastic zip bags containing silica gel dessicant. PCR was performed centrally  for all cases of recurrent parasitaemia from day 7 onwards, or for whom evidence of parasite clearance had been documented, and including DNA extraction using a QIAamp DNA Mini Kit (Qiagen). The three polymorphic genetic markers MSP1, MSP2, and GluRP, were also investigated to distinguish recrudescence from new infections, according to WHO recommended procedures26. Recrudescence was defined as at least one identical allele for each of the three markers in the pre-treatment and post-treatment samples. New infections were diagnosed when all alleles for at least one of the markers differed between the two samples.Thick and thin Giemsa-stained blood slides were prepared prior to the administration of the drug and at every follow-up visit. Slides were examined by two independent microscopists and considered negative if no parasites were seen after examination of 200 oil-immersion fields in a thick blood film. Parasite density was estimated using the Lambar\u00e9n\u00e9 method27. However, as all recruited subjects were asymptomatic at baseline, some of the standard clinical endpoints such as fever clearance time were not applicable. The primary efficacy outcomes were the PCR-corrected early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) and adequate clinical and parasitological response (ACPR) at day 28. ACPR was defined as the absence of parasitemia at the end of the trial\u2019s follow-up period (Day 28), regardless of axillary temperature, without having previously met any of the criteria for early and late treatment failure. In the PCR-adjusted analyses, patients with recurrent infection were considered ACPR if this was classified as a new infection. Secondary outcomes included 28 day-uncorrected ACPR (crude efficacy), time to parasite clearance, parasite clearance curve during the first 72 hours of follow-up, and prevalence of chloroquine-resistance conferring PfCRT K76T mutation in pre-treatment infections.Treatment outcomes were classified on the basis of an assessment of the parasitological and clinical response to antimalarial treatment according to the latest WHO guidelinesPfCRT gene. In brief, a first round of amplifications was performed in 25\u2009\u03bcl reactions including 5\u2009\u03bcl of template DNA, 0.5\u2009\u03bcM of each forward (PfCRT_F-5\u2032tttaggtggaggttcttgtctt-3\u2032) and reverse (PfCRT_R-5\u2032 atacttaattgaagaacaaatgattgga-3\u2032) primers and 1x HOT FirePol Master Mix . The template DNA was denatured at 95\u2009\u00b0C for 15\u2009min in a thermocycler, followed by 25 cycles of amplification  and a final extension at 72\u2009\u00b0C for 10\u2009min. A reaction using 5\u2009\u03bcl of PCR-grade water instead of template DNA was included as a negative control. For the nested amplification, 5\u2009\u03bcl of the PCR product from the first amplification was used as the template in a PCR reaction  containing 0.5\u2009\u03bcM of each forward (PfCRT_F) and reverse (PfCRT_N_R-5\u2032ttggtaggtggaatagattctct-3\u2032) primers and 1x HOT FirePol Master Mix (Solis BioDyne). The reaction volume was make up by PCR-grade water. The template DNA was denatured at 95\u2009\u00b0C for 15\u2009min in a thermocycler, followed by 35 cycles of amplification  and a final extension at 72\u2009\u00b0C for 10\u2009min. PCR products were run on 2% agarose (Invitrogen) gels in 1\u2009\u00d7\u2009TBE buffer (Thermo Fisher Scientific) to determine the presence and size of the amplified DNA. PCR products were visualized using a UV trans-illuminator. Both PCR primer sets were also tested with human gDNA to check their specificity. Three positive  controls with known PfCRT_76\u2009T allele were also processed and amplified at the same time as the studied samples. Positive and negative controls were added in every run. The expected size of the nested PCR was 319bp covering amino acid positions 35 to 120 from PfCRT in the 3D7 strain. PCR products were quantified using EPOCH Biotech system. Approximately 1200ng of PCR products were sent to Genewiz, following safety instructions for the accurate shipment of PCR amplicons. PCR sequencing was performed in both directions using specific forward and reverse primers of studied genes. The variations in the test sequences were identified by sequence alignment against reference sequence of 3D7 (PF3D7_0709000) retrieved from PlasmoDB.Purified DNA templates were amplified using 2720 Thermal Cycler (Applied Biosystems) by nested PCR amplification followed by Sanger sequencing for Data were recorded using standardized questionnaires specifically designed for this study, which were doubled-entered into a study-specific database created using open clinica software . Statistical analyses were performed with Stata 14 , and the statistical significance level was set at 5%.Safety outcomes were assessed in the intent-to-treat (ITT) population, which comprised all patients who received one or more doses of study medication and underwent at least one post-baseline safety assessment. Efficacy was calculated in the according-to-protocol population (ATP), which included all patients fulfilling the protocol eligibility criteria, who completed the three-day course of study medication, with no protocol violations, accomplishing the day-28 assessment and having an evaluable PCR in case of recurrent parasitaemia. Kaplan-Meier estimates were computed to estimate the cumulative proportion of treatment success until day 28; and Log-Rank tests used to compare the number of events in each arm. For such an analysis, losses to follow-up and study withdrawals were censored on the last day of follow-up. Cases with re-infections were also censored from the analysis on the day of detection. Finally, linear regressions and Likelihood Ratio Tests (LRT) of the log-transformed parasite densities during the first 72 hours of treatment were estimated to better characterize the parasite clearance curve during treatment between study groups.PfCRT-76 alleles were calculated as the proportion of samples carrying the mutant form out of all samples processed. Samples carrying both wildtype and mutant forms, in which relative frequencies could not be determined, were excluded from the denominator and numerator13.Frequencies of mutations at 28, and assuming an unknown failure rate, a minimum sample of 50 patients was considered necessary to provide a meaningful evaluation of in vivo efficacy, regardless of the underlying rates of failure. For the placebo group, the proportion of parasites that are eliminated naturally by the host\u2019s immune system after 28\u2009days in Manhi\u00e7a is unknown, but expected to be less than one percent31. Thus, the sample size for the control group needed not be the same size as the treatment group in order to detect a significant difference in prevalence. If 25 controls were recruited (placebo arm), then, even if half of the controls were to clear their infections, we will still achieve a p-value of less than 0.001 from a Chi-square test with one degree of freedom . Therefore, our objective was to recruit 75 participants total, i.e. 50 to receive CQ and 25 to receive PL.Following WHO guidelinesth, 2015, at www.ClinicalTrials.gov (NCT02698748).This protocol, consent forms and questionnaires were approved by the CISM local ethics committee, the Ethics Committee of the Hospital Cl\u00ednic of Barcelona (HCB/2015/0122), the National Bioethics Committee of Mozambique  and the Mozambican pharmaceutical department (Ref./N\u00b04110/380/DF2014) before their implementation. The methodology used in this study was performed in accordance with the relevant regulations and guidelines from all committees. All participants signed an Informed consent form prior to the initiation of any study related activities. All samples for the separate analysis of trends in CQ resistance were obtained under an informed consent, and after individual ethical approvals for each of the parent studies by CNBS. The trial was registered on January the 13a priori inclusion criteria and were randomized (Day 0) to receive either CQ  or PL . Median age of randomized individuals was 21.5 years (IQR 17\u201335). No significant differences were observed between subjects randomized to receive either CQ or PL  for symptomatology and malaria infection. Of those, 112 (5.8%) fulfilled In terms of tolerability and safety of the interventions, the symptoms reported during the three-day treatment course did not significantly vary between study arms Table\u00a0. No seriThe day-28 PCR-uncorrected cure rate  was 89.4%  for CQ, and 18.1%  for PL (p\u2009<\u20090.001). PCR-correction did not change the D28 ACPR for CQ, which remained 89.4%  given that the only new infection identified in this group was lost to follow-up after day 21; but increased that of PL , as 8 of the treatment failures proved to be new infections, and were consequently excluded from the analysis (Table\u00a0Only one (2%) of the 50 participants in the CQ arm still presented an infection (24\u2009p/\u03bcL) 72 hours after the administration of the first dose. Conversely, 14 (54%) of the 26 individuals in the placebo group were infected at the same point in time, with parasite densities ranging from 22 to 28,089 p/\u03bcL. Finally, the linear regressions fitted to the log-transformed parasite densities at 0, 24, 48 and 72 hours revealed significantly different rates of parasite clearance  between study arms Fig.\u00a0.Figure 3PfCRT K76T mutation by bi-directional DNA sequencing. Further, these sequences were aligned, using NCBI-Blast online tool against the reference sequence of 3D7 (PF3D7_0709000) retrieved from PlasmoDB. Three positive control data were in accordance with existing data. Only one of the 108 (0.9%) samples analyzed was found to have the mutant allele  in both treatment groups as a surrogate of drug efficacy34. Second, low parasitaemic infections (approximately 500\u2009p/\u03bcL) could have potentiated drug efficacy, which needs to be assessed further in future studies involving clinical malaria cases with higher loads of parasite biomass. The evaluation of drug efficacy in low parasite density infections carried additional challenges, particularly when performing molecular analysis to comply with WHO standard analysis procedures35. Due to low parasitaemia, the sensitivity of the genotyping techniques was low, and therefore PCR-Corrected analysis was considered biased. As a result, PCR-uncorrected analysis should be considered as a more precise way of analyzing the efficacy of chloroquine in this particular study population. The standard PCR-Corrected analysis was also limited by the very nature of the control group, which consisted of natural, untreated, asymptomatic infections. Participants in the placebo group were consequently vulnerable to reinfections and parasite density oscillations around the microscopy detection threshold that could be interpreted as recrudescences or reinfections rather than as a chronic low parasitemic infection4.Considering the low in vivo efficacy found in Southern Mozambique through this clinical trial aligns well with the findings obtained molecularly that detected a sharp decline in the prevalence of drug-resistant parasites between 1999 (84%) and 2015 (0.9%) in the same area. Such a shift in the parasite population after the interruption of CQ use has been observed in several countries in Africa36. This reversal to the wild type form of the PfCRT gene indicates that parasites carrying the mutant PfCRT may have a substantial fitness cost in the absence of CQ, thus leading to their decline in frequency once drug pressure is removed14. In some instances, loss of fitness may be associated with the development of compensatory mechanisms, leading to persistence of the mutant parasite in the population despite the discontinuation of the drug. This feature may explain, at least in part, the persistence of PfCRT-76 mutant39, even if CQ had not been used in these areas for many years. There are a series of additional factors that may be involved in the presence of phenotypic CQ resistance \u2013 such as PfMDR1 mutations40 \u2013 that were not assessed in this study and could lead to an underestimation of the real molecular resistance profile in the area. These factors could be the cause of remaining CQ resistance unrelated to the presence of PfCRT, and should be considered when monitoring molecular resistance in the future.Despite all of the above, the high in vivo efficacy of CQ among adults, but this time enrolling only those with clinical symptomatology derived from their infections, and higher parasite densities on admission. Only if chloroquine shows good efficacy in this population, we will deem it possible to start evaluating CQ\u2019s real life efficacy among infected sick children.Overall, the encouraging results from this first exploratory trial and molecular analysis set the scene for subsequent studies to continue exploring the efficacy of CQ in clinical cases with higher parasitaemia, before any final recommendations are made with regards to the use of this drug in Mozambique. It could be argued that exposing children with clinical malaria to a drug that has only shown partial efficacy to clear infections in semi-immune adults with low-density, may be hard to justify based on the presented data. Indeed, this population, with allegedly lower levels of acquired immunity to malaria due to their age, may be more vulnerable to the infection, and at higher risk of responding poorly to a partially effective drug. Thus, as a next step, we aim to continue testing the 41 can provide a long post-treatment prophylactic effect. Thus, CQ could be a drug of choice in malaria elimination efforts such as in MDA campaigns, in combination with a highly effective anti-malarial. It could also be used in similar contexts for those populations who cannot receive ACTs, such as pregnant women in the first trimester, or very young children, as the safety of CQ in such populations has widely been demonstrated. Finally, through this innovative use of CQ, the drug pressure exerted to the circulating parasites would be minimal, and risk for re-introduction of resistant mutations would be very low.In the context of malaria elimination, CQ exhibits two conditions that make it attractive for elimination campaigns: 1) It has been demonstrated to have an excellent safety profile, allowing for its use in all age groups including pregnant women and young children; and 2) Its relatively long elimination half life (t1/2\u2009=\u20091\u20132 months)This study shows that CQ could play a role, on its own or in combination with other drugs or tools, as part of malaria elimination efforts at the population level, both on account of its therapeutic efficacy but more importantly by means of its chemoprophylactic activity and safety profile."}
+{"text": "This study describes the implantation of a community-based pharmacovigilance system in C\u00f4te d\u2019Ivoire and its use to document the safety of ASAQ WinthropThis prospective, longitudinal, descriptive, non-comparative, non-interventional study on the use of artesunate\u2013amodiaquine in real-life conditions of use was conducted in seven Community Health Centres of the Agboville district in C\u00f4te d\u2019Ivoire. Twenty trained Health Centre employees and 70 trained community health workers were involved in data collection in the field. All patients with suspected uncomplicated falciparum malaria, seeking treatment at one of the participating Health Centres, and treated with artesunate\u2013amodiaquine could be enrolled. Two visits were planned, one for inclusion at the Health Centre and a second at home, performed by a community health worker 3\u201310\u00a0days after the inclusion visit. Administration of artesunate\u2013amodiaquine was unsupervised. Adverse events (AEs) were documented at the home visit or during any unexpected visit to the Health Centre or to the hospital and coded and adjudicated by a local pharmacovigilance committee. Symptoms suggestive of hepatic failure, severe neutropaenia, extrapyramidal disorders and retinopathy were considered a priori as AEs of special interest.Some 15,228 malaria episodes in 12,198 patients were evaluated; 2545 AEs were documented during 1978 malaria episodes (13.0%). The most frequently observed events were asthenia (682 cases), vomiting (482 cases) and somnolence (174 cases). Most reported AEs were of mild or moderate intensity and resolved without corrective treatment. One-hundred and five (105) AEs reported during 100 episodes (0.7%) were considered as serious. Three serious cases of transient extrapyramidal disorders, identified as AEs of special interest were reported in three patients.\u00ae for the unsupervised treatment of uncomplicated falciparum malaria under real-life conditions of care in C\u00f4te d\u2019Ivoire is well tolerated. The study emphasizes the interest of involving properly trained community health workers to collect pharmacovigilance data in the field in order to document rare AEs.The fixed dose artesunate\u2013amodiaquine combination ASAQ WinthropThe online version of this article (doi:10.1186/s12936-016-1655-1) contains supplementary material, which is available to authorized users. The most frequent reason for non-compliance was treatment discontinuation due to the occurrence of an AE (180/425 cases). The dose of ASAQ Winthrop\u00ae prescribed was appropriate for the patient\u2019s age or weight in the majority of episodes (98.6% for age and 86.1% for weight).As assessed by questioning and by counting remaining tablets, ASAQ WinthropAll but 36 episodes benefitted from a follow-up home visit by a CHW. Overall, 2545 AEs were documented during 1978 of 15,161 malaria episodes (13.0%) during the course of the study  were reported during 837 episodes (5.5%). The most frequent of these are listed in Table\u00a0\u00ae.Regarding outcome, 2452 of the 2545 AEs resolved satisfactorily (96.3%) without sequelae. In addition, 39 AE were improving at the time of last follow-up. One patient presented a non-serious skin rash which left marks and scars. Twenty-four SAEs during 22 episodes (0.1%) were fatal and these are listed in Table\u00a0The incidence of AEs was compared between malaria episodes with parasitaemia detected on thick blood smears, unconfirmed malaria episodes without detectable parasitaemia, and episodes without available parasitaemia results . No statistically significant differences in the incidence or nature of AEs, SAEs or potentially ASAQ-related AEs were observed between the three sub-groups Table\u00a0.Table\u00a05I\u00ae under conditions of every day care in malaria treatment centres in C\u00f4te d\u2019Ivoire. The study was successfully implemented and the target sample size achieved. This illustrates the feasibility of such studies for monitoring anti-malarial treatments in West Africa and to complement \u2018passive\u2019 pharmacovigilance systems. The study population was principally composed of young children, with 65% of the sample being under 6\u00a0years of age, which reflects the epidemiology of malaria in many sub-Saharan African countries [\u00ae. AEs were documented in 13% of >15,000 individual treated malaria episodes in >12,000 patients over a period of 3\u00bd\u00a0years. Overall, treatment with ASAQ Winthrop\u00ae was found to be well tolerated and the nature of AEs reported was consistent with the known safety profile of ASAQ Winthrop\u00ae. Most reported AEs were not severe and resolved without the need for medical intervention.This large, naturalistic, observational study was designed to monitor the safety of ASAQ Winthropountries . The rel\u00ae in sub-Saharan Africa [In this study, a robust and structured reporting system was set up for AE reporting, with the HC staff systematically reporting safety observations in patients seen at HCs and a trained network of involved and motivated CHWs who were encouraged to report all AEs. Despite the high rate of follow-up home visits, the reporting rate of AEs in this study was lower than that reported in randomized clinical trials of ASAQ Winthropn Africa , 10\u201313. n Africa ). Nonethn Africa , 10\u201313, It is of note that the rate of AE reporting was highest when the interval between treatment initiation and the home visit was short and reduction of the mean interval before the home visit may increase the sensitivity of AE reporting. Although home visits were supposed to take place between 3 and 10\u00a0days following initiation of treatment, a small number of visits were made before 3\u00a0days, and documentation of SAEs was proportionally more frequent during these visits. It can be hypothesized that these early visits may have been triggered by the occurrence of AEs, leading to villagers contacting the CHW. Nevertheless, if follow-up is too early, there may be a risk of missing delayed effects, even if patients were strongly encouraged to re-contact their CHW in case of any problem. In these cases, a further later follow-up visit may be useful to ensure comprehensive documentation of all AEs.\u00ae) in Tanzania [\u00ae), conducted in four African countries between 2013 and 2014 [P. falciparum infections who were followed up by home visits from a CHW or by telephone. More recently, a study in Ghana of patients prescribed ACT [Comparison of this study with previous cohort-event monitoring studies (CEM) of anti-malarial drugs is complicated by differences in study design. No previous observational CEM study has evaluated the incidence of AEs in patients treated specifically with an identified ASAQ fixed-dose combination tablet, and several previous studies have evaluated different anti-malarial drugs in the same study. The present study is also much larger than previous studies, which included <5000 patients . In addTanzania  also reqand 2014 , which iibed ACT  has compibed ACT .\u00ae were reported during 837 episodes (5.5%). The most frequent of these potentially related AEs were asthenia, somnolence, vomiting, and dizziness, with an incidence of >0.5%. All these are known adverse drug reactions to ASAQ Winthrop\u00ae and listed as such in the prescribing information. No cases of retinopathy, symptomatic leukopaenia or symptoms suggestive of serious abnormalities in liver function, which are characterized AEs associated with amodiaquine, were observed. However, no laboratory blood testing was performed, so asymptomatic leukopaenia or perturbations of liver function cannot be ruled out. The incidence of SAEs (0.7%) and severe AEs (0.8%) was low and, of the 22 deaths reported in the study, none was considered potentially related to ASAQ Winthrop\u00ae. The nature and frequency of SAEs were consistent with previous clinical trials [\u00ae in C\u00f4te d\u2019Ivoire.Overall, 929 AEs potentially related to ASAQ Winthropl trials \u201313. Thes\u00ae without recurrence of symptoms. Prior to the initiation of the study, the study sponsor became aware of poorly characterized pharmacovigilance reports of extrapyramidal symptoms in patients taking amodiaquine or artesunate and amodiaquine combinations of different origins and of unknown causality. This was the reason why extrapyramidal symptoms were listed in the study protocol as an AESI, in an effort to better characterize any potential association. The identification of three characterized cases in this study prompted the Marketing Authorization Holder for ASAQ Winthrop\u00ae to carry out a comprehensive assessment of the association of extrapyramidal disorders with ASAQ Winthrop\u00ae or other anti-malarial drugs in 2012, which led to extrapyramidal disorders being listed as adverse drug reactions in the prescribing information for ASAQ Winthrop\u00ae. Around the same time, an analysis was published of some of the earlier pharmacovigilance reports documented in the WHO global pharmacovigilance database (VigiBase\u2122) [The large sample size of the study  was chosen in order to allow identification of rare AEs occurring with an incidence of one in 5000. In this context, three cases of moderate extrapyramidal disorders (incidence of 0.02%) were recorded and considered as possibly drug-related. All cases resolved without sequelae after treatment with diazepam. Causality could not be proven since all patients had also taken non-prescribed traditional medications; furthermore, one patient was re-challenged with ASAQ WinthropgiBase\u2122) , and thegiBase\u2122) , 24, 30.\u00ae observed in the study did not differ between treated patients with RDT-confirmed malaria and in patients without identified parasitaemia.At the beginning of the study, national malaria treatment recommendations did not require confirmation of malaria diagnosis by a blood test, and treatment could be initiated on the basis of clinical suspicion only. However, since malaria symptoms are relatively non-specific, this policy exposed patients to unnecessary anti-malarial treatment with a potential risk of toxicity, and was changed during the course of the study in 2011, such that laboratory confirmation of the diagnosis was required. Nonetheless, given the high sensitivity of RDT, false positives may still occur in patients with fluctuating low parasite densities or carrying residual parasite proteins from a previous infection \u201333, in p\u00ae, with a single intake of a low number of tablets (one or two) per day. In addition, these tablets can be either swallowed or dissolved in a small amount of water for people unable to swallow. The good compliance of ASAQ Winthrop\u00ae use with prescription recommendations at HCs should be noted, with the prescribed dose being correctly adjusted for a patient\u2019s age in 98.6% of episodes.Although treatment intake was unsupervised, the rate of reported compliance based on tablet count by CHWs at home visits was high (97.2%). It is possible that the fact that patients were aware that CHWs would be making home visits encouraged compliance (or hiding of tablets). For some patients, a home visit was made during the treatment period, when the CHW could check that the treatment was being taken correctly. Nonetheless, any such bias is likely to be limited and the good observed compliance rate may reflect the ease of administration of ASAQ Winthrop\u00ae in the Agboville region during the study period, which covered all seasons of the year.This study has a number of strengths and limitations. The principal strengths relate to the large number of patients included and the naturalistic treatment setting. The findings should thus be generalizable to every day malaria treatment practice in C\u00f4te d\u2019Ivoire. In particular, the study included essentially all patients with suspected malaria treated with ASAQ Winthrop\u00ae was unsupervised, which means that certain patients may not have respected the recommended dosage regimen. Finally, a large proportion of patients were prescribed or had taken other medications at the same time as ASAQ Winthrop\u00ae treatment, rendering unambiguous determination of causality very difficult. However, these limitations are necessary consequences of the naturalistic study design.The principal limitation related to the lack of systematic ascertainment of reported AEs by a physician. Safety evaluation was limited to clinical observation, and laboratory testing was only performed on an ad hoc basis. In consequence, cases of leukopaenia or impaired liver function may have gone undetected if these were asymptomatic. In addition, treatment with ASAQ WinthropThis type of study may help to extend awareness of the use of ACT for malaria treatment and on the importance of pharmacovigilance reporting by staff of local HCs and by CHWs, and thus contribute to effective long-term safety monitoring in the NMCP in C\u00f4te d\u2019Ivoire. A high level of communication was maintained between the different partners in the network, which ensured that CHWs and local HC staff received information and training from the study coordinators whenever the need arose. Following this study, the NMCP of C\u00f4te d\u2019Ivoire signed an agreement with the Drug and Pharmacy Board, which states that the latter should undertake training of healthcare workers in pharmacovigilance and analysis of pharmacovigilance report forms.This study underlines the added value of systematic involvement of CHWs in collecting pharmacovigilance data on anti-malarial drugs in a community setting in sub-Saharan Africa, at least when new therapeutic programmes are being implemented. Although the WHO has emphasized the need for post-approval safety information on new medicines in African countries , this is\u00ae for the unsupervised treatment of uncomplicated falciparum malaria under everyday conditions of care in C\u00f4te d\u2019Ivoire demonstrated that the safety of the drug was consistent with what had been seen in the clinical trials, allowed the identification of extrapyramidal disorders as rare AEs leading to a change in prescribing information, and emphasized the value of involving properly trained CHWs to collect pharmacovigilance data in the field.This large, naturalistic study of the tolerability of ASAQ Winthrop"}
+{"text": "Plasmodium falciparum malaria. The role of pharmacogenetic variation on anti-malarial drug disposition and efficacy during pregnancy is not well investigated. The study aimed to examine the effect of pharmacogenetics on lumefantrine (LF) pharmacokinetics and treatment outcome in pregnant women.Pregnancy has considerable effects on the pharmacokinetic properties of drugs used to treat uncomplicated CYP2B6 , CYP3A4*1B, CYP3A5  and ABCB1 c.4036A4G were determined. Blood smear for parasite quantification by microscopy, and dried blood spot for parasite screening and genotyping using qPCR and nested PCR were collected at enrolment up to day 28 to differentiate between reinfection from recrudescence. Treatment response was recorded following the WHO protocol.Pregnant women with uncomplicated falciparum malaria were enrolled and treated with artemether-lumefantrine (ALu) at Mkuranga and Kisarawe district hospitals in Coast Region of Tanzania. Day-7 LF plasma concentration and genotyping forP. falciparum. Parasites were detected during the follow up period in 11 (12%) women between day 7 and 28 after treatment and PCR genotyping confirmed recrudescent infection in 7 (63.3%) women. The remaining four (36.4%) pregnant women had reinfection: one on day 14 and three on day 28. The overall PCR-corrected treatment failure rate was 9.0% (95% CI 4.4\u201317.4). Day 7 LF concentration was not significantly influenced by CYP2B6, CYP3A4*1B and ABCB1 c.4036A>G genotypes. Significant associations between CYP3A5 genotype and day 7 plasma LF concentrations was found, being higher in carriers of CYP3A5 defective variant alleles than CYP3A5*1/*1 genotype. No significant influence of CYP2B6, CYP3A5 and ABCB1 c.4036A>Genotypes on malaria treatment outcome were observed. However, CYP3A4*1B did affect malaria treatment outcome in pregnant women followed up for 28\u00a0days (P\u00a0=\u00a00.018).In total, 92 pregnant women in their second and third trimester were included in the study and 424 samples were screened for presence of CYP3A4 and CYP3A5may influence LF pharmacokinetics and treatment outcome in pregnant women.Genetic variations in  Pregnancy-induced physiological changes alter the pharmacokinetic properties of a number of anti-malarial drugs, usually resulting in lower drug exposures and lower cure rates especially in advanced pregnancy stage compared to non-pregnant population , 2. PregLower drug exposure levels in pregnant women have been reported for artemether/dihydroartemisinin, artesunate/dihydroartemisinin, dihydroartemisinin and lumefantrine (LF) , 17\u201319. CYP3A4*1B allele frequency between the Tanzania and Cambodia populations presents a potential explanation for the lower efficacy of ALu in Cambodia and highlighted the importance of pharmacogenetic considerations in the decision-making process of first-line treatment policies for specific populations [Genetic variation in drug metabolizing enzymes and transporter proteins might predict plasma exposure and treatment failure and/or emergence of drug resistant pathogens on infectious and non-infectious diseases, such as malaria, HIV and tuberculosis \u201324. CYP3ulations .To date, there are limited studies that attempted to examine the role of pharmacogenetics on the pharmacokinetics of anti-malarial drugs and treatment outcomes in pregnant women. The aim of this study is to investigate the effects of pharmacogenetics on day 7 LF plasma concentrations and treatment outcome in pregnant patients treated with ALu in Tanzania. The findings might have implications for treatment policies of not only anti-malarial drugs, in particular the widely used artemisinin-based combinations, but also other drugs metabolized by these enzymes.Plasmodium falciparum infection and haemoglobin level of\u00a0>8\u00a0g/d were enrolled. The study received ethics approval from the institutional review board of Muhimbili University of Health and Allied Sciences (MUHAS). Participants were informed about the aim of the study and gave written consent before participating in the study. To ensure confidentiality, women's identification numbers were used when labelling samples and during data entry into confidential report forms (CRF).This was a one-arm prospective cohort study that included all pregnant women who gave consent to participate in the study when attending antenatal clinics at Kisarawe and Mkuranga district hospitals, northern Tanzania. Between May 2014 to April 2015, pregnant women attending the antenatal clinics (ANCs) were screened for malaria infection by using malaria rapid diagnostic test (MRDT). Pregnant women with uncomplicated Considering anticipated population proportion (P) of clinical failures in pregnant women being 18% , with 95\u00ae; Novartis Pharma AG, Basel, Switzerland) (20\u00a0mg artemether and 120\u00a0mg lumefantrine) over the course of 3\u00a0days at 0, 8, 24, 36, 48, and 60\u00a0h. For each patient, general physical examination was performed at enrollment (day 0) and on follow up visits on days 2, 7, 14, 21 and 28. Approximately 50\u00a0\u03bcl of blood was collected on filter paper (Whatman grade 3) for later PCR analysis. Each filter paper was dried and individually stored in a plastic bag and kept frozen at \u221280\u00a0\u00b0C. At enrollment day, 1\u00a0ml of whole blood was taken into an EDTA containing vacutainer tube and stored at \u221280\u00a0\u00b0C at MUHAS laboratory until further analysis. Additionally, 3\u00a0mls of venous blood were drawn from pregnant women in heparinized tubes on day 7 to determine plasma LF concentrations.The study participants received six doses of four tablets of ALu , which have been reported to be relevant for artemether and LF disposition [CYP2B6*6 , C__60732328_20 for CYP2B6*18 , C__26201809_30 for CYP3A5*3 , C__30203950_10 for CYP3A5*6  and C__32287188_10 for CYP3A5*7 (g.27131_27132insT rs41303343) and C__11711730_20) for ABCB1 c.4036A>G (rs3842), and C__11711730_20 for CYP3A4*1B . Genotyping was carried out using Quant Studio 12\u00a0K Flex Real-Time PCR system . The final volume for each reaction was 10\u00a0\u03bcl, consisting of TaqMan fast advanced master mix , TaqMan 20X drug metabolism genotyping assays mix (Applied Biosystems) and genomic DNA. The PCR profile consisted of an initial step at 60\u00a0\u00b0C for 30\u00a0s, hold stage at 95\u00a0\u00b0C for 10\u00a0min and PCR stage for 40 cycles step 1 with 95\u00a0\u00b0C for 15 and step 2 with 60\u00a0\u00b0C for 1\u00a0min and after read stage with 60\u00a0\u00b0C for 30\u00a0s.Genomic DNA was isolated from peripheral leukocytes in whole blood samples using QIAamp DNA Midi Kit  according to the manufacturer\u2019s instructions. Genotyping for the common functional variant alleles for position , 27 wereposition , 22. In Plasmodium parasites was performed using a species-specific PCR targeting the ssRNA gene [P. falciparum, Plasmodium malariae, Plasmodium vivax and Plasmodium ovale probes at a final concentration of 100\u00a0nM. All reactions were run on an ABI Prism 7000 sequence detection system (Applied Biosystems) with the default settings; each sample was initially denatured at 95\u00a0\u00b0C for 10\u00a0min and cycled 40 times, with each cycle consisting of 95\u00a0\u00b0C for 15\u00a0s and 60\u00a0\u00b0C for 60\u00a0s. Each reaction plate included four positive controls for P. falciparum, P. ovale, P. vivax, and P. malariae and a negative control with molecular-grade water in place of DNA.Dried blood spots on filter papers obtained at enrolment (day 0) and on follow-up days  were punched, and one circle 5\u00a0mm in diameter was used for DNA extraction using QIAamp DNA blood micro kit  following the manufacturer\u2019s recommendations. Detection of P. falciparum infections from reinfections were done according to World Health Organization (WHO) recommendations [msp2) in samples collected at day 0 and on the day recurrent parasitaemia was found. Recrudescence was determined when at least one msp2 allele of the same allelic type and of identical base pair size was found in samples collected on day 0 and on the day of recurrent infection. A reinfection was defined as all alleles were of different length in the sample collected on day 0 and that collected on the day of recurrent infection.PCR genotyping to differentiate recurrent ndations , by charBlood samples for the determination of LF plasma concentrations were collected on day 7 (corresponding to 168\u00a0h) following initiation of ALu treatment. Blood samples for quantification of LF levels were centrifuged and plasma was stored at \u221280\u00a0\u00b0C until analysis. Plasma LF concentration was analysed using a validated method of high performance liquid chromatography (HPLC) with ultraviolet detection at Sida/MUHAS bioanalytical laboratory in Dar-es-Salaam, Tanzania . The coe2 test was used to compare the observed and expected allele frequencies according to the Hardy\u2013Weinberg equilibrium. Influence of human genotype on malaria treatment outcome was analysed using Pearson\u2019s Chi square and Fisher\u2019s exact test. Haploview software package (version 4.2) was used to analyse linkage disequilibrium (LD) and haplotype construction. Statistical analyses were performed using Statistical Package for Social Sciences (SPSS) software, version 22.0 . P values <0.05 were considered to be statistically significant.LF plasma concentration data were log transformed to achieve normality of data distribution. Median (interquartile range) was used to describe day-7 LF plasma concentrations. Comparison of day-7 median LF plasma concentrations in pregnant women between the different genotypes were carried out using Kruskal\u2013Wallis one-way analysis of variance (ANOVA). XMalaria treatment outcomes were classified following the WHO protocol , as adeqIn total, 1835 pregnant women were screened using MRDT and a total of 92 pregnant women with malaria infection consented and were enrolled in the study. Baseline characteristics are presented in Table\u00a0P. falciparum was present in 11 (12%) patients during the follow up period. Two samples were detected on day 7, four on day 14, one on day 21 and four on day 28 after ALu treatment. PCR based genotyping of msp2 confirmed recrudescent infection in seven women (63.3%), two on day 7, three on day 14, one on day 21 and one on day 28. The remaining four (36.4%) pregnant women had reinfection; one on day 14 and three on day 28. The overall rate of reinfection was 4.3% (95% CI 0.15\u20138.45).In total, 424 samples were screened for presence of parasites and PCR uncorrected ACPR on day 28 was 88.9% (95% CI 82.06\u201395.74) and TF rates were 11.1% (95% CI 4.26\u201317.94). Rate of ACPR was calculated using per-protocol method, where patients were excluded due to lost to follow-up, protocol violations and TF due to reinfection. PCR-corrected ACPR rate as defined by absence of parasitaemia on day 28, irrespective of axillary temperature, in patients who did not previously meet any of the criteria of ETF, LCF or LPF was 91.0% (95% CI 82.62\u201395.58). The rate of PCR-corrected LPF, as defined by presence of parasitaemia on any day between day 7 and 28 in patients who did not previously meet any of the criteria of ETF or LPF, was 9.0% (95% CI 4.42\u201317.38). There was no ETF or LCF.CYP2B6*6, CYP2B6*18, CYP3A4*1B, CYP3A5*3, CYP3A5*6, CYP3A5*7, ABCB1 c.4036A>G genotype and allele frequencies in Tanzanian pregnant women is presented in Table\u00a0CYP3A4*1B followed by CYP2B6 c.516G>T . CYP2B6 c.983T>C (*18) had the lowest allele frequency which was 9.3%. Genotype of CYP450 enzymes and ABCB1 transporter was equally distributed among the age, gravida, trimester and PMTCT status of pregnant women (P\u00a0>\u00a00.05).The overall CYP3A4*1B (\u2013392A>G), CYP3A5*3 (g.6986A>G), CYP3A5*6 (g.14690G>A), CYP3A5*7 (27131_27132insT) is presented in Fig.\u00a0CYP2B6 variant alleles (c.516G>T and c.983T>C). Likewise, no LD between the three CYP3A5 SNPs  was found. Instead each SNP was inversely linked to each other . The haplotype frequency of CYP3A5*1, *3, *6 and *7 was 44.5, 22.6, 20.7 and 12.2%, respectively. Interestingly all the three CYP3A5 variant alleles  occur in high LD with CYP3A4*1B. The major CYP3A haplotype was CYP3A4*1B alone (34.2%) followed by its linkage with CYP3A5*6 (17.6%) and CYP3A5*3 (13.3%) , one  and zero  to investigate the effect of genotype on day 7 plasma LF concentrations.Based on the haplotype analysis result, subjects were grouped according to the numbers of functional CYP2B6, CYP3A4*1B or ABCB1 c.4036AG genotype on day 7 LF plasma concentrations. The median plasma LF concentration was significantly associated with CYP3A5 genotypes (P\u00a0=\u00a00.039). CYP3A5 defective allele carriers displayed a higher plasma LF concentrations compared to those homozygous for CYP3A5*1/*1 genotypes (P\u00a0=\u00a00.04). There was significant association between having CYP3A5 genotype and having plasma LF concentration\u00a0\u2265600\u00a0ng/ml . The proportion of subjects with two functional CYP3A5 genotype (*1/*1) was significantly higher among those with plasma LF conc\u00a0<600\u00a0ng/ml (34.6%) compared to those with\u00a0\u2265600\u00a0ng/ml (5.9%). On the other hand proportion of subjects with lacking functional CYP3A5  was higher among those with plasma LF conc\u00a0\u2265600\u00a0ng/ml (38.2%) compared to those with >600\u00a0ng/ml (19.2%).The median day 7 plasma LF concentration was 650 with the range of 2936 124\u20133059) ng/ml. Comparison of the median plasma LF concentrations between the different genotypes is presented in Table\u00a0\u20133059 ng/CYP2B6, CYP3A5 and ABCB1 on malaria treatment outcome was evaluated using Cox regression analysis. A log rank test was run to determine differences in the cumulative hazard distribution between the different genotypes. There were no associations between malaria treatment outcome and ABCB1 transporter and most of CYP450 genotypes with an exception to CYP3A4  and ABCB1 genes involved in the metabolism of anti-malarial drugs were assessed in pregnant malaria patients treated with ALu. The major finding includes: (i) CYP3A5 genotype has significant influence on plasma LF concentration and (ii) CYP3A4*1B is associated with malaria treatment outcome. A number of studies have speculated that lower artemether and LF concentration in pregnant women has been due to changes in catalytic activities of CYP450 metabolizing enzymes particularly the increased activity of CYP3A4 [CYP2B6*6 (c.516G\u2192T), CYP3A4*1B, and CYP3A5*3 are very similar to those previously reported from previous studies conducted in Tanzanian population [In the present study, the effect of pharmacogenetics on LF pharmacokinetics  and treatment outcome in pregnant women was investigated. For this purpose, functional variant alleles in CYP450  in Africa is metabolized by CYP3A4. Artemether is metabolized by CYP3A4 and CYP3A5. Therefore, information on the role of pharmacogenetics on drug disposition and treatment outcome in pregnant women using ALu is needed. CYP3A5 is mainly expressed in black population and its genotype contributes to variations in the total CYP3A enzyme activity as measured by 4beta-hydroxycholesterol, an endogenous CYP3A marker , 33, 34.CYP3A5 defective alleles than those without. Though not significant, a similar finding was observed in a recent HIV-malaria cohort study in Tanzania [CYP3A5*1/*1 genotype had significantly higher risk of having LF plasma concentration\u00a0<600\u00a0ng/ml. Plasma LF concentration <600\u00a0ng/ml is associated with risk of recurrent parasitaemia in pregnant women [There was significantly higher plasma LF concentration in patients carrying Tanzania . Pregnannt women . Howevernt women . This mant women .CYP3A4*1B on day 7 LF plasma levels. The findings are similar to previous reports from Cambodia and Tanzania [CYP3A4*1B genotype did not affect day 7 LF plasma levels unless it was induced by a potent CYP3A4 inducer [In this study, there was no influence of Tanzania . A studyTanzania . The decTanzania  where th inducer .CYP3A4*1B genotype and treatment outcome in pregnant women. It was observed that majority of pregnant women with CYP3A4*1B/*1B had attained ACPR compared to those with CYP3A4*1/*1. Inconclusive data on alterations of enzyme activity in CYP3A4*1B carriers are reported in the literature. Some investigators have suggested CYP3A4*1B is associated with increased CYP3A4 expression and enhanced drug elimination in carriers of CYP3A4*1B may lead to treatment failure [CYP3A4*1B with lower CYP3A4 enzyme activity in Tanzanians is reported previously, where carriers of CYP3A4*1B had a significantly lower enzyme activity than CYP3A4*1 [CYP3A4*1B in Tanzanians (77%) [CYP3A4*1B with low enzyme activity in blacks. CYP3A4*1B variant allele is absent in Asians and occurs at a much lower frequency (2\u20139%) in whites [The study also analysed patients with recrudescence and compared their pharmacogenetic profiles with those with an ACPR. There was a significant association between  failure . In contCYP3A4*1 . The finns (77%)  may indin whites .CYP3A4 and CYP3A5 influences the LF plasma exposure and malaria treatment outcome in pregnant women. Since CYP3A is responsible for the metabolism of artemether and LF, interplay between CYP3A4 and CYP3A5 genotypes may determine ACT plasma exposure and treatment outcome. CYP3A4 and CYP3A5 haplotypes are located in the same gene locus, effects initially considered to be due to a CYP3A4 allele might actually be due to a CYP3A5 allele in LD [CYP3A4*1B and CYP3A5*1Ais suggested as possible cause of inter individual variation in CYP3A metabolism [Preliminary finding indicates that pharmacogenetic variation in le in LD . LD betwtabolism , 42.CYP3A4*1B is linked with the functional CYP3A5*1 resulting in high enzyme activity [CYP3A4*1B is linked with CYP3A5 defective variant alleles  and hence may be associated with low enzyme activity. It is well known that sub-Saharan African population is the most genetically heterogeneous population globally, characterized by extensive population substructure and unique LD pattern compared to non-African populations [CYP2B6 variant alleles (c.516G>T and c.983T>C) and between the three CYP3A5 SNPs  in this study is similar to previously reports from Africa [Previous studies in white population reported that activity , 43. In ulations , 45. Lacm Africa , 23.CYP3A haplotypes on treatment outcome. However, the study finding highlights the importance of genetic variation in the CYP3A locus for LF pharmacokinetics and treatment outcome in pregnant women. This study also presents haplotype structure of the most common CYP3A functional variant alleles in African population for future genetic association studies.Limitation of this study includes that the sample size was calculated based on the previous study whereby treatment failure was reported to be 18% which is higher than the failure rate reported in this study. As a result, although the sample size is enough to investigate anti-malarial drug efficacy in the population, the study is under-powered to investigate the impact of CYP3A4*5 and CYP3A4*1B with high LF plasma exposure and better malaria treatment outcome, respectively. The effects of pharmacogenetics on LF pharmacokinetics were well characterized in pregnant patients with uncomplicated P. falciparum malaria. The importance of these findings is that in the future genotyping can be used to predict the need for anti-malarial drugs dosage adjustment to pregnant women with CYP3A4*1/*1or CYP3A5*1/*1. The impact of CYP3A haplotypes on the metabolism of anti-malarial drugs in a large sample size cohort needs to be further evaluated.In general, the study finding indicates association of the low enzyme activity genotype of"}
+{"text": "Plasmodium falciparum malaria remains high. Typical features at death are high parasite loads and obstructed micro- vasculature. Infected erythrocytes (IE) containing mature parasites bind to the host receptor heparan sulfate, which is also an important receptor for merozoite invasion. To block merozoite invasion has not previously been proposed as an adjunctive therapeutic approach but it may preclude the early expansion of an infection that else leads to exacerbated sequestration and death.Even with the best available treatment, the mortality from severe The drug sevuparin was developed from heparin because heparan sulfate and heparin are nearly identical, so the rationale was that sevuparin would act as a decoy receptor during malaria infection. A phase I study was performed in healthy male volunteers and sevuparin was found safe and well tolerated.A phase I/II clinical study was performed in which sevuparin was administered via short intravenous infusions to malaria patients with uncomplicated malaria who were also receiving atovaquone/proguanil treatment. This was a Phase I/II, randomized, open label, active control, parallel assignment study.P. falciparum malaria.Sevuparin was safe and well tolerated in the malaria patients. The mean relative numbers of ring-stage IEs decreased after a single sevuparin infusion and mature parasite IEs appeared transiently in the circulation. The effects observed on numbers of merozoites and throphozoites in the circulation, were detected already one hour after the first sevuparin injection. Here we report the development of a candidate drug named sevuparin that both blocks merozoite invasion and transiently de-sequesters IE in humans with NCT01442168ClinicalTrials.gov  P. falciparum claims 430,000 lives annually, despite the use of the best anti-malarial medications currently available [P. falciparum likely because the hosts, the parasites and the pathology that bring about severe malaria are only partially alike in animals and man [P. falciparum infected erythrocytes (IE) and inflammation are typical features of severe malaria but it is also known that patients with complicated disease carry higher parasite loads, a larger biomass, than those with mild malaria [P. falciparum infected humans is maintained by an asynchronous rupture of schizonts, the subsequent invasion of erythrocytes by merozoites and the development of the parasites into ring- and adhesive trophozoite stage IE. The time of development of a merozoite into an adhesive IE that may obstruct the micro-vasculature is about 18\u201320 hours and the majority of deaths in severe malaria occur the first day of hospitalization during an equivalent timespan [P. falciparum malaria if it was given as an adjunct to anti-plasmodial drugs, but its use was stopped because of the risk of hemostatic disorders [P. falciparum sequestration and merozoite invasion was found to be independent of its anti-coagulant activity [Severe malaria due to vailable . Improvevailable \u20134. Howev and man \u20138. Indee and man . Excessi malaria . We hypotimespan \u201313. In hisorders \u201316. Impoactivity \u201326. NotaP. falciparum merozoite invasion into fresh erythrocytes in vitro, and both disrupts and blocks the binding of IE to uninfected erythrocytes (rosetting) and binding to vascular endothelial cells (cytoadherence) in vitro. Furthermore, from in vivo studies sevuparin has been demonstrated to block IE from binding in the micro-vasculature of the rat but also to de-sequester IE, i.e. release already sequestered IE into circulation, both in rats and in a non-human primate [Sevuparin blocks  primate \u201327. In a primate .P. falciparum malaria.Here we report the results of both a phase I study in healthy volunteers and a clinical study with sevuparin in patients with Sevuparin, like other members of the chemical class of heparins, is polydisperse, encompassing a range of polysaccharide chain lengths. The predominant size is 6\u201316 disaccharide units of 2-N-sulfo-6-O-sulfo-glucosamine and iduronic-2-O-sulfate acid, which corresponds to molecular weights of 3.6\u20139.6 kDa. The weight average molecular weight is 6.5\u20139.5 kDa. The predominant chemical structure of sevuparin is illustrated in Sevuparin does not contain the specific binding sequence for AT which is the main contributor to prolonged coagulation and as a consequence it has minimal anticoagulant potency compared to other heparinoids . The antin vivo in humans an exploratory proof-of\u2013principle study was conducted. It was decided to conduct a study in uncomplicated falciparum malaria prior a proof-of- concept study in severe malaria patients after considering ethically limitations to go directly in to severely ill patients after only a phase I study. The phase I/II, randomized, open label, active control, parallel assignment study was conducted at Mae Sot Hospital and Mae Ramat Hospital, Tak Province Thailand between 2011 and 2014 [The clinical phase I/II study Treatment of Severe Malaria study 02 (TSM02) was conducted in compliance with International Conference on Harmonisation Good Clinical Practice guidelines and the Declaration of Helsinki . In ordeand 2014 . In the and 2014 . The stufalciparum malaria with parasitemia counts of 10 000\u2013100 000/ul at screening and admitted to the hospitals were enrolled in the study, provided that written informed consent was given by the patient. Patients with any sign of severe malaria, determined on the basis of World Health Organization criteria [Consecutive adult patients (18\u201365 years old) with slide-proven criteria ,36 were in vivo efficacy of sevuparin when administered as a short (five minutes) i.v. infusions as adjunctive therapy to standard care i.e. a fixed dose combination of 1000 mg atovaquone and 400 mg proguanil  once daily for three days, in subjects affected with uncomplicated falciparum malaria, as compared to sole atovaquone/proguanil  treatment (control). An artemisinin combination therapy was not used, because the broad stage specificity and the rapid onset of action of the artemisinins prevents maturation and subsequent sequestration IEs, thus obscuring the effects of sevuparin. Subjects were treated with sevuparin during three consecutive days . In part 1 of the study, the pharmacokinetic parameters of sevuparin were studied in cohorts of 3 patients receiving 12 consecutive i.v. infusions of 1.5 mg/kg, 3 mg/kg or 6 mg/kg sevuparin in addition to standard of care. Doses were selected following a safety evaluation of the single dose data from the phase I study. The dose 1.5 mg/kg every six hour for three days corresponds the no observable effect level (NOEL) of sevuparin at 2 hours after treatment and is 4-fold below the dose of the 360 mg, which was well tolerated when given every six hours to healthy volunteers. The selected starting dose is also in the range of the therapeutic dose of heparin and LMWH in adults as well as in children. Taken into account the significant decreased anticoagulant potency of sevuparin compared with heparin, the dose of 1.5 mg/kg is considered to have a sufficient safety margin for the intended patient population. This dose was therefore considered justified as a safe starting dose in 18\u201365 years old adults suffering from acute uncomplicated malaria disease. Subjects were enrolled in cohorts of three in each, and escalation of dose to the next cohort was determined based on evaluation of DLTs in the previous cohort, following a dose escalation strategy. After all three subjects in one cohort had been observed for a minimum of 14 days an evaluation of the dose limiting toxicity (DLT) was done and reported to the Data Safety Monitoring Board (DSMB). Part 2 of the study was open label and with patients randomized in a 1:1 ratio, in blocks of 10 , to receive either 12 consecutive i.v. infusions (6 hours apart) of sevuparin 3 mg/kg in addition to standard of care, or standard of care alone. This part of the study could only be initiated following a safety evaluation of part 1 data performed by the DSMB. The dose to be used was recommended by the DSMB due to increased, but not above stopping criteria, APTT, and no obvious improved efficacy with higher dose. Study design and dosing regimen of part 1 and part 2 are shown in The study was designed to determine the safety, tolerability, pharmacokinetics and The primary endpoint of part 2 was area under the curve (AUC) of the graph plotting the throphozoite stage peripheral blood parasitemia over time of sevuparin treatment as adjunctive to atovaquone/proguanil in comparison to atovaquone/proguanil treatment alone. Relative numbers of ring-, throphozoite- and schizont-IE were calculated from the number of IE at one time point related to the baseline number of IEs at time point 0 h (immediately prior to the first dose of sevuparin).The total parasitemia and the number of ring-IE, trophozoite-IE and schizont-IE were estimated from blood collected before treatment of sevuparin was initiated (H0) as well as at specified time-points after the first dose, including H1, H2, H3, H4, H6, H8, H10, and H11 thereafter every 6h until two sequential parasite negative slides were collected. Thin and thick films were prepared from 0.5 mL blood at each timepoint. One hundred parasites were counted. The slides were read for determination of numbers of parasites by two experienced independent microscopists at respective hospital and by one expert at Mahidol Oxford Tropical Medicine Research Unit (MORU), Mahidol University, Bangkok, Thailand for assessment of stages of the parasites in the blood ,37. Micr5 parasites/\u03bcL/h to log 3X105 parasites/\u03bcL/h between the sevuparin group and the control group, with an expected SD of 0.8 logs in both groups, significance level of 0.05 and a power of 80%. Due to slow recruitment, the study was prematurely stopped when a total of 44 patients were randomized in part 2 (out of 79 screened). Out of these, 21 patients were randomized to treatment with sevuparin and 23 were controls.For part 2, a sample size of 40 in the sevuparin treated group and 40 in the control group was estimated to allow the detection of a difference of 0.5 log units in terms of the Area Under the Curve (AUC) of the late-stage peripheral blood parasitemia over time curve from log 10Statistical analyses including tables, figures and data listings were produced using SPSS software (version 15.0), STATA (version 13) and R (versions 3.0.1 and 3.1.3). Data were log transformed to obtain a normal distribution, where necessary. Normally distributed data were compared using Student\u2019s t test. AUC was calculated using the trapezoidal rule, differences between groups were assessed using Mann- Whitney tests. Categorical variables such as gender, blood type etc was analyzed using the Fishers exact test. Differences in change from baseline in parasitemia, abundance of early and late stages of parasites were analyzed using the Mann- Whitney test. The Mann-Whitney U test has been used for non-paired nonparametric data.The sevuparin concentrations in the peripheral blood taken during TSM02 study were measured using a validated solid- phase competitive Enzyme-Linked Immunosorbent Assay (ELISA) kit for plasma samples  according to the manufacturer\u2019s instructions. This assay measures sevuparin directly using a sevuparin/heparin binding protein, conjugated to horseradish peroxidase (HRP). Calibration range for sevuparin was 500\u201310000 ng/ml. The range of quantitation for this method was 1000\u20135000 ng/ml, the calibration curve fitted with the 4PL logistic model spanned a concentration range between 500 and 10000 ng/ml. The Lower Limit of Quantification (LLOQ) for sevuparin was 1000 ng/ml and the Upper Limit of Quantification (ULOQ) was 5000 ng/ml.Trophozoite IE were placed at a haematocrit of 5% in 75 \u03bcL of culture medium with 5% human AB-serum in 96 well round-bottom microtiter plates  with the addition of 25 \u03bcL sevuparin  or 25 \u03bcL phosphate buffered saline (control).falciparum malaria diagnosed at Karolinska University Hospital, Sweden and established at Karolinska Institutet; three EBA-knock out strains  where the gift of Dr. Alan Cowman, four Cambodian isolates were from ATCC-MR4, Manassas, VA, USA ; IPC 5188 Rattanakiri; IPC 3663 Pailin; IPC 4912 artemisinin resistant (RSA 0\u20133 h: 49%)). P. falciparum laboratory strains were cultivated according to standard methods [Laboratory strains were grown for one cycle while patient isolates were grown for two, after which the cultures were stained with acridine orange and the parasitemia of each well was determined by flow cytometry using a FACSVerse, BD Bioscience with a plate reader. Data were analyzed using FlowJo software. Invasion inhibition was expressed as a percentage of the parasitemia in absence of sevuparin with PBS only. Parasite strains used were 10 lab strains , 11 isolates from Ugandan patients , two from Ethiopia/Eritrea, three from Kenya, one from Niger all of travellers with uncomplicated  methods . Patient methods , or in SIn a phase I study conducted in healthy male volunteers we found sevuparin to be safe and well tolerated at the dose levels studied, the data of which are to be found in P. falciparum malaria, we conducted an exploratory study, clinical phase I/II open-label trial . After the dose escalating part (part 1) safety data were reviewed by a data safety monitoring board (DSMB). Pre-defined stopping criteria included individual APTT \u22651.5 x ULN 5 hours after dose 1 or dose 5, mean APTT in one cohort \u2265 3 x ULN 2 hours after dose 1 or dose 5, mean APTT in one cohort close to, or \u22653 x ULN 2 hours after dose 12, or at the Investigator\u2019s discretion, but had not been reached in any dose group. However, as the dose 6.0 mg/kg 6 q6h did give higher APTT values than the lower doses and in one patient APTT was up to 120 seconds 1 hour after the dose (ULN = 33 sec), the DSMB considered the risk of adverse events was potentially increased and recommended that the study should continue to part 2 with 3.0 mg/kg sevuparin q6h. Forty-four patients were enrolled in the second part of this 1:1 randomized and open-label study, 21 were given i.v. sevuparin infusions in addition to the oral atovaquone/proguanil, and 23 patients were only given oral atovaquone/proguanil. All 53 subjects included in the trial completed the study treatment.There were minimal and clinically non-relevant changes in anti-Xa- and prothrombin-times, and the international normalized ratios (INR) associated to the sevuparin dose  Tables. No severe adverse event (SAE) or adverse event (AE) leading to withdrawal occurred in the study  Tables. falciparum malaria.There were no apparent changes from baseline or differences between dose groups with regard to physical examination findings and vital signs. There were no apparent changes in ECG or QTc in sevuparin treated subjects. To summarize, sevuparin administered as i.v. infusion for five minutes every six hours for three consecutive days  at different dose levels was safe and well tolerated in adult patients with uncomplicated P. falciparum infected patients was determined as change in numbers of mature throphozoit-IE, and number of ring-IE in the peripheral blood, as compared with control patients. The stages of the parasites in the peripheral blood and the total parasitemia were scored by microscopy. Sevuparin treatment was anticipated to both increase the number of circulating trophozoite/schizont IEs in the peripheral bllod, which would mean that sevuparin cause de-sequestration of previously bound/sequestered trophozoite/schizont IEs [P. falciparum malaria in the presence of sevuparin leads to fewer ring stage parasites in the peripheral blood.Efficacy of treatment with sevuparin in zont IEs ,19,21\u201326zont IEs ,20,25. IThe two patient groups in the clinical phase I/II open-label trial were also analysed in terms of de-sequestration, which is the increase of mature parasite IEs in the circulating blood after the initiation of treatment with sevuparin. Notably, more trophozoite and schizont IEs appeared in the circulating peripheral blood one hour after the first dose of sevuparin than in the controls who were not given sevuparin  for the first 11 hours after the initiation of sevuparin. In the sevuparin group, the AUC of late stage parasitemia was expected to increase over time as compared to the control group. No statistically significant difference between treatment groups was detected as regards cumulative AUC of late stage peripheral blood parasitemia at the time point 11 hours after first dose of sevuparin . Other tThe median parasite clearance time was evaluated in part 2 and was 53 hours  in the sevuparin group, versus 59 hours  in the control group (p = 0.720). The median parasite reduction rates (PRRs) at 24 and 48 hours were respectively 87.44%  and 99.96%  in the sevuparin group, versus 84.64%  and 99.78%  in the control group, both not significantly different . The median time to 50% reduction of peripheral blood parasitemia was 12.66 hours  in the sevuparin group, versus 12.87 hours  in the control group (p = 0.404), and the median time to 90% reduction was in the sevuparin group 25.59 hours  versus 30.71 hours  in the control group (p = 0.148). .In patients with uncomplicated malaria treated with sevuparin, systemic exposure in terms of Cmax and AUC increased in a dose-proportional manner. Mean Cmax in part 1 and part 2 were between 56.7\u2013149 \u03bcg/mL. In both parts and both after the first and last dose, plasma levels declined mono-compartmentally with an apparent terminal half-life of 0.7\u20131.3 hours. Systemic exposure in terms of Cmax and AUC did not show any notable differences on Day 3 vs. Day 1 at any dose level. Overall between-patient variability in terms of systemic exposure parameters was moderate and tended to decrease with increasing dose.in vitro with 34 parasites from various geographical origins, including 17 primary patient isolates from Ethiopia/Eritrea, Kenya, Niger or Uganda, four in vitro established parasites from Cambodia that were resistant or sensitive to artemisinin, three parasites that were deficient in invasion ligands  and 10 in vitro, long-term propagated parasites that were sensitive  or resistant to chloroquine and/or mefoloquine  [The rapid reduction in ring stage-IEs observed in the sevuparin treated subjects is likely a consequence of the capacity of sevuparin to block the earliest steps of merozoite invasion into erythrocytes ,25,43\u201348, TM284) . SevuparMost deaths in severe malaria occur within the first day of hospitalization when the characteristic features of high parasite loads and obstructed micro-vasculatures are present \u201314. It iAdhesive IE involved in causing severe malaria are unlike those of uncomplicated disease in that they are prone to bind excessively to the vascular endothelium and to form rosettes , processIn the sevuparin-treated individuals the mean ring-stage parasitemia decreased with 50% by 8 hours while it took another \u22485 hours for the group treated with solely atovaquone/proguanil . The sigS1 Supportive Information(DOC)Click here for additional data file.S2 Supportive Information(PDF)Click here for additional data file.S3 Supportive Information(PDF)Click here for additional data file.S4 Supportive Information(PDF)Click here for additional data file.S5 Supportive Information(PDF)Click here for additional data file.S1 Fig(PDF)Click here for additional data file.S1 Table(DOCX)Click here for additional data file.S2 Table(DOCX)Click here for additional data file.S3 Table(DOCX)Click here for additional data file.S4 Table(DOCX)Click here for additional data file.S5 Table(DOCX)Click here for additional data file.S6 Table(DOCX)Click here for additional data file.S7 TablePart 2.(DOCX)Click here for additional data file.S8 Table(DOCX)Click here for additional data file."}
+{"text": "There is general international agreement that the importance of vivax malaria has been neglected, and there is a need for new treatment approaches in an effort to progress towards control and elimination in Latin America. This open label randomized clinical trial evaluated the efficacy and safety of three treatment regimens using either one of two fixed dose artemisinin-based combinations or chloroquine in combination with a short course of primaquine  in Brazil. The primary objective was establishing whether cure rates above 90% could be achieved in each arm.A total of 264 patients were followed up to day 63. The cure rate of all three treatment arms was greater than 90% at 28 and 42\u00a0days. Cure rates were below 90% in all three treatment groups at day 63, although the 95% confidence interval included 90% for all three treatments. Most of the adverse events were mild in all treatment arms. Only one of the three serious adverse events was related to the treatment and significant drops in haemoglobin were rare.This study demonstrated the efficacy and safety of all three regimens that were tested with 42-day cure rates that meet World Health Organization criteria. The efficacy and safety of artemisinin-based combination therapy regimens in this population offers the opportunity to treat all species of malaria with the same regimen, simplifying protocols for malaria control programmes and potentially contributing to elimination of both vivax and falciparum malaria.Trial registration RBR-79s56sThe online version of this article (10.1186/s12936-018-2192-x) contains supplementary material, which is available to authorized users. Plasmodium falciparum as the leading cause of malaria mortality and morbidity worldwide. Nevertheless, Plasmodium vivax remains a major public health problem ) of the ASMQ group, 88% (95% CI [81\u201395%]) of the CQ group, and 84% (95% CI 76\u201392%]) of the AL group achieved an ACPR at day 63. The cure rate of all the three treatment arms was greater than 90% at 28 and 42\u00a0days in the PP population (Table\u00a0%) of theCure rates in the ITT analysis were slightly lower  meant that the ITT success rate at day 3 was 98% (95% CI [98\u2013100]).Fever clearance is an important surrogate for cure in malaria. Only one patient in the CQ\u00a0+\u00a0Pq arm and two in the AL\u00a0+\u00a0Pq arm had an axillary temperature higher than 37.5\u00a0\u00b0C at day 3. Four fever episodes were reported after day 7. One patient had fever at day 63 and was considered as a treatment failure. The other three patients completed 63\u00a0days follow up without evidence of parasites on microscopy. All patients cleared gametocytes by day 3. The five patients that had gametocytes present on day 2 had all been treated with CQ\u00a0+\u00a0Pq  were reported. One patient in the CQ arm had treatment suspended because of haemolytic anaemia on day 3. Qualitative calorimetric test confirmed glucose-6-phosphate dehydrogenase (G6PD) deficiency. Another in the AL arm had a rise in alanine aminotransferase (ALT) detected on day 12 after treatment. The third SAE reflected elective surgery and was not related to the trial. All patients recovered completely.The safety analysis was conducted in the ITT population. A total of 1593 adverse events were reported. 1379 of them (86.5%) were classified as grade I and 208 events (13.1%) were classified as grade II. The distribution of adverse events by age and weight range, and treatment arm is shown in Table\u00a0749 events (41.0%) were reported as possible, probable/likely or highly probably related to the treatment drug; 48.9% were grade I, 35.1% were grade II, none grade III and one SAE (grade IV). The distribution of all adverse events according with their intensity and causality are presented at Table\u00a0The distribution of all adverse events according with their intensity and causality per treatment group is provided  were grouped based on the main body system affected and treatment allocation , using Pearson\u2019s Chi squared test.Changes in haemoglobin (Hb) were a secondary outcome. There was a slight decrease in all groups at day 14 followed by recovery , the dependent variable was the count of possible, probable/likely or highly probable AE related to the study treatments per person. Adverse events were less common in males , but use of antipyretic or antihypertensive was associated with high rates of adverse events .This study was not powered to detect differences in cure rates between treatment regimens and no statistically significant difference could be detected between the treatment arms at any time point.A generalized linear model  was used to explore the influence of baseline variables and concomitant treatments upon failure at day 63, using a significance level of 5%. There was a strong association between the use of antipyretic medications and treatment failure . A trend towards an association between the use of anti-ulcerative drugs and treatment success was also observed .The effect of the treatment on the time to clearance of gametocytes was evaluated in the three arms. Patients in the ASMQ\u00a0+\u00a0Pq and AL\u00a0+\u00a0Pq arm had all cleared gametocytes by day 2. In the CQ\u00a0+\u00a0Pq arm, 5 out of 43 patients (11.6%) had gametocytes present of day 2. Gametocytes only reappeared in five of the 103 patients, making meaningful conclusions impossible.P. vivax to chloroquine may be due to the fact that standard CQ\u00a0+\u00a0Pq treatment has been an effective combination therapy [There has been recent international agreement that the importance of vivax malaria has been neglected , 23, 24, therapy . In addi therapy  and chlo therapy ; concomi therapy .P. vivax schizonticidal activity to chloroquine [P. vivax emerges. There are compelling arguments to look for a simpler and unified ACT treatment for both species of malaria. A single radical treatment would be useful in co-endemic areas where species diagnosis is difficult [P. falciparum. The use of ACT and primaquine for the treatment of P. falciparum infection also has the advantage of eradicating hypnozoites and preventing relapses from previous P. vivax infection that can happen following a P. falciparum infection [ACT has been recommended by the WHO to treat vivax since 2010 , as it aoroquine  and it iifficult  or mixednfection . There aIn this study, all three treatment arms demonstrated cure rates\u00a0>\u00a090% in all treatment arms at day 42 but by day 63, cure rates had dropped below 90% . It demonstrates the importance of a longer follow-up time in detecting failures, particularly in vivax when reinfection or relapse can occur . This stP. vivax malaria are developed [Drug interactions and the safety profile are important factors in choosing the optimum first line treatment, in addition to pill burden and food restrictions. This trial evaluated the efficacy and safety of three vivax treatment regimens, and provides additional reassurance about the safety of ASMQ and AL when given with primaquine; information that is considered important by WHO as strategies for control and elimination of eveloped . Most ofDespite this low level of adverse events, the ACT and primaquine combination still has significant drawbacks. Its use is limited in pregnancy, breastfeeding and G6DP-deficient population. The one SAE in this study was related to G6PD deficiency. These restrictions impose serious limitations to its use as an elimination tool in many parts of the world, although the primaquine regimen used in this study is the 7-day regimen currently recommended by the Brazilian National Malaria Control Programme. This regimen delivers a total dose from 3 to 4.2\u00a0mg/kg . This isThis study was not sufficiently powered to detect differences between the three arms and only followed up patients for 63\u00a0days, meaning that late relapses would not have been detected. The study population did not include children, limiting the conclusions of efficacy in this populations. The absence of PCR also meant that relapses could not be distinguished from reinfection; a critical issue in assessing vivax treatment. Nevertheless, this study demonstrates the efficacy and safety of two ACT and CQ in combination with 7\u00a0days of primaquine to treat uncomplicated vivax malaria in Brazil and demonstrated the feasibility and utility of a standardized treatment approach for all malaria cases.Additional file 1: Table S1. Reasons to not be included\u2014Consort Diagram.Additional file 2: Table S2. Distribution of use of concomitant medication (grouped in therapeutic class) in each study arm, parenthesis presents the percentage of the line. Figure S1. Distribution of most frequent medications used per study visit and treatment group.Additional file 3: Table S3. Proportion of treatment success per treatment arm in ITT population (n\u00a0=\u00a088 per arm) at day 28, 42 and 63. Table S4. Proportion of treatment success per treatment arm in PP population at day 07, 14 and 21. Table S5. Proportion of treatment success per treatment arm in ITT population (n\u00a0=\u00a088 per arm) at day 07, 14 and 21.Additional file 4: Table S6. Distribution of adverse events per causality and treatment group.Additional file 5: Table S7. Distribution of adverse events per causality and intensity (grade) in the treatment group ASMQ\u00a0+\u00a0Pq. Table S8. Distribution of adverse events per causality and intensity (grade) in the treatment group CQ\u00a0+\u00a0Pq. Table S9. Distribution of adverse events per causality and intensity (grade) in the treatment group AL\u00a0+\u00a0Pq.Additional file 6: Table S10. All adverse events (1593) per body system and treatment allocation.Additional file 7: Table S11. Adverse events with frequency higher than 3% in the ASMQ\u00a0+\u00a0Pq arm . Table S12. Adverse events with frequency higher than 3% in the CQ\u00a0+\u00a0Pq arm . Table S13. Adverse events with frequency higher than 3% in the AL\u00a0+\u00a0Pq arm .Additional file 8: Table S14. Haemoglobin mean per regimen allocation at day 0, 14, 28, 42, and 63."}
+{"text": "Plasmodium falciparum malaria in India. However, resistance against AS\u00a0+\u00a0SP is emerged in northeastern states. Therefore, artemether\u2013lumefantrine (AL) is the recommended first line treatment for falciparum malaria in north eastern states. This study investigates the therapeutic efficacy and safety of AL for the treatment of uncomplicated falciparum malaria in three malaria-endemic states in India. The data generated through this study will benefit the immediate implementation of second-line ACT as and when required.Anti-malarial drug resistance continues to be a leading threat to malaria control efforts and calls for continued monitoring of waning efficacy of artemisinin-based combination therapy (ACT). Artesunate\u00a0+\u00a0sulfadoxine/pyrimethamine (AS\u00a0+\u00a0SP) is used for the treatment of uncomplicated P. falciparum infection were administered six-dose regimen of AL over 3\u00a0days and subsequent follow-up was carried out up to 28\u00a0days. Molecular markers msp-1 and msp-2 were used to differentiate recrudescence and re-infection and K13 propeller gene was amplified and sequenced covering the codon 450\u2013680.This was a one-arm prospective evaluation of clinical and parasitological responses for uncomplicated falciparum malaria using WHO protocol. Patients diagnosed with uncomplicated mono k13 gene along with non-synonymous mutation at codon M579T in three (1.6\u00a0%) samples.A total of 402 eligible patients were enrolled in the study from all four sites. Overall, adequate clinical and parasitological response (ACPR) was 98\u00a0% without PCR correction and 99\u00a0% with PCR correction. At three study sites, ACPR rates were 100\u00a0%, while at Bastar, cure rate was 92.5\u00a0% on day 28. No early treatment failure was found. The PCR-corrected endpoint finding confirmed that one late clinical failure (LCF) and two late parasitological failures (LPF) were recrudescences. The PCR corrected cure rate was 96.5\u00a0%. The mean fever clearance time was 27.2\u00a0h\u00a0\u00b1\u00a08.2 (24\u201348\u00a0h) and the mean parasite clearance time was 30.1\u00a0h\u00a0\u00b1\u00a011.0 (24\u201372\u00a0h). Additionally, no adverse event was recorded. Analysis of total 186 samples revealed a mutation in the AL is an efficacious drug for the treatment of uncomplicated falciparum malaria. However, regular monitoring of AL is required in view of malaria elimination initiatives, which will be largely dependent on therapeutic interventions, regular surveillance and targeted vector control.The online version of this article (doi:10.1186/s12936-016-1555-4) contains supplementary material, which is available to authorized users. Plasmodium falciparum infection causes the most serious form of malaria and its proportion varies from 50 to 90\u00a0% in different states of the country. This study was undertaken in three out of five highly malarious states, i.e., Chhattisgarh, Madhya Pradesh and Jharkhand.Malaria is a major public health problem and a leading cause of mortality worldwide. The World Health Organization (WHO) estimated that 214 million cases of malaria occurred globally and approximately 438,000 deaths were recorded in 2015 . If any discordance was found, a third reading was performed by another senior microscopist. Quantification of parasitaemia was also performed by two independent microscopists with a third reading performed by a senior microscopist if the difference between the first two readings varied by more than 25\u00a0%. Each reader was blinded to the result of other reader.P. falciparum genes merozoite surface proteins (msp1 and msp2) [The PCR method was used to distinguish recrudescence from new infection in the case of treatment failure. Nested PCR was conducted to compare the genetic polymorphism of nd msp2) . RecrudeParents or guardians of children were instructed to return to the health centre at any time if they had any general danger signs as described under exclusion criteria. The study team made home visits as follow-ups for study participants that were late for their scheduled visits. Patients who failed to return on days 1 and 2 and missed one dose of the treatment or enrolled patients who could not attend scheduled visits were considered lost to follow-up (LFU) and excluded from the final analysis.On the basis of parasitological and clinical outcome of treatment with AL, patients were classified according to the WHO definition of therapeutic responses: early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) and adequate clinical and parasitological response (ACPR) .k13 propeller gene was amplified by the nested PCR method described earlier by Ariey et al. [k13 gene of P. falciparum isolate from the study site has been submitted to GenBank under Accession number KX121048, KX121049.The y et al.  with somData from both clinical and parasitological assessments for each participant were entered into the WHO standardized Microsoft Excel data collection sheet. This form was used for both data management and analysis. All data were independently entered double blind. Data were analysed by estimation of difference in proportion according to a 95\u00a0% confidence interval. Groups were compared using Chi Square test or Fisher Exact test for categorical variables and Student\u2019s t test for continuous variables where ever applicable. Otherwise, non-parametric tests  were used.A total of 402 eligible patients  were enrolled in the study from all four sites and the baseline demographics are presented in Table\u00a0P. falciparum recrudescence, while the remaining two LCF and one LPF patients were confirmed as having P. falciparum re-infection with one LPF shown as PCR negative. The PCR-corrected cure rate by Kaplan\u2013Meier analysis at Bastar was 96.5\u00a0%. Merozoite surface protein 1 (Pfmsp 1) and merozoite surface protein 2 (Pfmsp 2) genotyping was performed for all the treatment failure samples on day 0 and day of failure. Out of the seven cases, six were found positive for P. falciparum on both days (day 0 and day of failure). Two cases were recrudescence  whereas three patients having re-infection at the time of treatment failure. However, one patient had mixed msp2 allele at the time of treatment failure.Among a total of 402 patients  , which are classified as the second, third and fourth highest malarious states, due to their high contribution in total malaria cases  within the country . The premsp1 and msp2) and not all the three marker genes . All three cases of treatment failure were from Bastar, a highly malarious district bordering Odisha state, which contributes the highest number of malaria cases and malarial deaths in the country [P. falciparum causes cerebral and severe malaria with a case fatality rate of 32 and 9\u00a0%, respectively, in PCR-confirmed mono P. falciparum among hospitalized patients [P. falciparum proportion is more than 85\u00a0% [In this study, three cases of treatment failure on day 21 and day 28 were young children. It is not known whether this is due to actual resistance of parasites to drugs or due to inadequate blood levels caused by poor drug absorption or altered pharmacokinetics. The study has limitation as the characterization the recrudescence vs. re-infection is based on only two population marker gene ( country . Studiespatients . Since fhan 85\u00a0% . As malnP. falciparum parasites. As definite phenotype of ACT resistance is very rare, the association of polymorphism in marker genes is very difficult to correlate with the efficacy outcome. Ariey et al. have reported that polymorphisms C580Y, Y493H and R539T in the propeller region of k13 gene are strongly associated with artemisinin resistance [P. falciparum k13 propeller gene [In the present study, NS gene among three isolates was detected in the highly conserved K13 propeller gene of sistance . Followisistance \u201326. Thresistance . No mutaler gene , 27.The high cure rate and parasite clearance time in this study indicate no imminent threat to artemisinin resistance in the regions investigated. As artemisinin derivatives rapidly reduce the symptoms with an additional advantage of anti gametocidal activity to reduce malaria transmission . However"}
+{"text": "In Zambia, malaria is one of the leading causes of morbidity and mortality, especially among under five children and pregnant women. For the latter, the World Health Organization recommends the use of artemisinin-based combination therapy (ACT) in the second and third trimester of pregnancy. In a context of limited information on ACT, the safety and efficacy of three combinations, namely artemether\u2013lumefantrine (AL), mefloquine\u2013artesunate (MQAS) and dihydroartemisinin\u2013piperaquine (DHAPQ) were assessed in pregnant women with malaria.The trial was carried out between July 2010 and August 2013 in Nchelenge district, Luapula Province, an area of high transmission, as part of a multi-centre trial. Women in the second or third trimester of pregnancy and with malaria were recruited and randomized to one of the three study arms. Women were actively followed up for 63\u00a0days, and then at delivery and 1\u00a0year post-delivery.Nine hundred pregnant women were included, 300 per arm. PCR-adjusted treatment failure was 4.7% (12/258) (95% CI 2.7\u20138.0) for AL, 1.3% (3/235) (95% CI 0.4\u20133.7) for MQAS and 0.8% (2/236) (95% CI 0.2\u20133.0) for DHAPQ, with significant risk difference between AL and DHAPQ (p\u00a0=\u00a00.01) and between AL and MQAS (p\u00a0=\u00a00.03) treatments. Re-infections during follow up were more frequent in the AL  and MQAS  arms compared to the DHAPQ arm. PCR-adjusted treatment failure was significantly associated with women under 20\u00a0years [Hazard Ratio (HR) 5.35 ] and higher malaria parasite density [3.23 ], and still women under 20\u00a0years  had a significantly higher risk of re-infection. The three treatments were generally well tolerated. Dizziness, nausea, vomiting, headache and asthenia as adverse events (AEs) were more common in MQAS than in AL or DHAPQ (p\u00a0<\u00a00.001). Birth outcomes were not significantly different between treatment arms.As new infections can be prevented by a long acting partner drug to the artemisinins, DHAPQ should be preferred in places as Nchelenge district where transmission is intense while in areas of low transmission intensity AL or MQAS may be used. Malaria is a poverty-related disease and a major public health problem in many sub-Saharan African countries where over 90% of the cases worldwide are found. Pregnant women and children are at higher risk of malaria infection and of developing serious complications related to the disease. Malaria in pregnancy is associated with higher risk of maternal anaemia, low birth weight, spontaneous abortion, stillbirths and maternal mortality \u20133.There are few treatments with known safety and efficacy for the treatment of malaria in pregnancy. Some anti-malarials known to be efficacious e.g. quinine, are not well tolerated, resulting in poor compliance and higher risk of treatment failures . For othPlasmodium falciparum malaria infection. This study was part of a multi-centre trial carried out also in Burkina Faso, Ghana and Malawi. Each site tested 3 ACT medicines so that each country dataset could be analysed separately  to have in the final model. The proportion hazard assumptions for the Cox-regression model were evaluated using graphical approach .The hematological and biochemistry profiles by day of follow-up were assessed using box-plots plotted at each time point. Differences in these parameters between treatment arms at each day of follow-up were assessed using Kruskal\u2013Wallis test.Firth logistic regression was used to assess impact of placental malaria  on birth outcomes  for separation and \u2018empty cells\u2019 in the model. A \u201cstillbirth\u201d was defined as a baby born dead after 24\u00a0weeks gestation; a baby born dead before 24\u00a0weeks gestation or during the 24th\u00a0week was considered a \u201cmiscarriage\u201d. \u201cPreterm live born\u201d was defined as a delivery before 37\u00a0weeks of gestation following echography. This was calculated as date of delivery minus date of echography (in weeks) plus gestational age determined through echography. Or based on the Ballard score which determines gestational age based on the sum of neuromuscular and physical scores . A neonaA total of 1722 pregnant women were screened for malaria infection, regardless of symptoms. Out of these, 900 met the inclusion criteria and were randomized to one of the three study arms: 300 to AL, 300 to MQAS and 300 to DHAPQ. The ITT analysis included 900 pregnant women. The PP analysis included 729 women, i.e. 258 in the AL, 235 in the MQAS and 236 in DHAPQ arms  in AL, 1.3% (95% CI 0.5\u20133.4) in DHAPQ and 0% (95% CI 0.0\u20131.3) in MQAS. The range of the gametocyte density, if present, was 40\u2013440 gametocytes/\u00b5L. Gametocyte carriage remained low during the follow-up period and only appeared in AL  and DHAPQ  arms and none in MQAS. Use of preventive measures, i.e. ITN and IPTp, at recruitment was low  assumptions testing for treatment failure adjusted for several variables  were roughly parallel and met the PH assumptions. The day 63 PCR-adjusted treatment failure rate was 4.7% 12/258) (95% CI 2.7\u20138.0) for AL, 1.3% (3/235) (95% CI 0.4\u20133.7) for MQAS and 0.8% (2/236) (95% CI 0.2\u20133.0) for DHAPQ  and MQAS  arms compared to the DHAPQ arm. The risk of re-infections was higher in women between 15 and 20\u00a0years  than in women older than 20\u00a0years. Anaemic mothers had a higher hazard of new infection during follow up . Similarly, mothers with higher parasite density a higher hazard of new infection . All the other risk factors analysed were not significantly associated with new infection  was similar between the treatment arms (p\u00a0=\u00a00.47) Table\u00a0. TreatmeThe study drugs were generally safe with a total of 7 SAEs for mother. A woman treated with MQAS died 41\u00a0days after treatment, probably because of meningitis. There were three SAEs in DHAPQ . An additional SAE in the MQAS arm, severe vomiting, was considered related to study treatment and recovered completely. The other two in MQAS were asthmatic attack and pneumonia. They all recovered.The proportion of women with AEs in each treatment arm  were not significantly different (p\u00a0=\u00a00.19) (Table\u00a0There were 21 stillbirths: 8 (2.7%) in AL, 3 (1.0%) in MQAS and 10 (3.3%) in the DHAPQ arms and three miscarriages . The preterm delivery was 4.3% in AL, 2.0% in MQAS and 3.3% in the DHAPQ arm. There were 15 congenital malformations  with no significant difference between the arms (p\u00a0=\u00a00.54).With the range of 0.8\u20134.7% recrudescences, the three artemisinin-based combinations used for the treatment of uncomplicated malaria in the second and third trimester of pregnancy were efficacious, in an area of high endemicity in Nchelenge district, Zambia. Therapeutic equivalence could be shown for MQAS and DHAPQ but not for AL as compared to the other two treatments. In Nchelenge Zambia, there were significantly more treatment failures in the AL arm compared to the other two arms, though AL efficacy was still above the 90% cure threshold recommended by WHO for adopting new anti-malarial treatments as policy . In UganIn Zambia, ACT has been shown to have excellent cure rates among children and adults , 24. ThePregnant women have an increased susceptibility to malaria, and this susceptibility is greatest in the first pregnancy (primigravidae) . The decIn Nchelenge, pregnant women treated with AL had a higher risk of new infection than the other two treatments. This is probably due to the shorter post-treatment prophylaxis offered by lumefantrine which is eliminated more rapidly  than pipRecrudescence may easily occur in the context of emergence or spread of parasite resistance to a given anti-malarial when the partially efficacious anti-malarial may fail to clear the resistant strain or simply select for mutant parasites. In Zambia, artemisinin resistance has not been reported yet. Recrudescence can be caused by the parasites surviving the effect of a shorter-acting ACT , in thisOther studies have shown that high parasite density at presentation is associated with treatment failure , 36\u201338 aThe three artemisinin-based combinations tested are generally safe in second and third trimester of pregnancy in Zambia. Patients on MQAS had higher rates of treatment-related AE. Dizziness was the most common, followed by vomiting and weakness. Dizziness has been reported even in other studies as related to MQ treatment . On the This trial was done in an area where the majority of the population practice farming and fishing as a source of livelihood and they migrate to farming areas for a considerable period , possiblThe study has shown that both AL and DHAPQ were well tolerated in second and third trimester pregnant women, with low treatment failures. MQAS was less well tolerated than the other two treatments though it had similar low treatment failure. DHAPQ seems to be well tolerated and has low treatment failure with a longer post-treatment prophylaxis. As new infections can be prevented by a long acting partner drug to the artemisinins, DHAPQ should be preferred where transmission is intense as in Nchelenge while and in areas of low transmission intensity AL or MQAS may be used."}
+{"text": "DSM265 is a novel antimalarial that inhibits plasmodial dihydroorotate dehydrogenase, an enzyme essential for pyrimidine biosynthesis. We investigated the safety, tolerability, and pharmacokinetics of DSM265, and tested its antimalarial activity.Plasmodium falciparum induced blood-stage malaria (IBSM) and treated with DSM265 (150 mg) or mefloquine (10 mg/kg). Primary endpoints were DSM265 safety, tolerability, and pharmacokinetics. Randomisation lists were created using a validated, automated system. Both parts were registered with the Australian New Zealand Clinical Trials Registry, number ACTRN12613000522718 (part 1) and number ACTRN12613000527763 (part 2).Healthy participants aged 18\u201355 years were enrolled in a two-part study: part 1, a single ascending dose (25\u20131200 mg), double-blind, randomised, placebo-controlled study, and part 2, an open-label, randomised, active-comparator controlled study, in which participants were inoculated with max) ranged between 1310 ng/mL and 34\u2008800 ng/mL and was reached in a median time (tmax) between 1\u00b75 h and 4 h, with a mean elimination half-life between 86 h and 118 h. In part 2, the log10 parasite reduction ratio at 48 h in the DSM265 (150 mg) group was 1\u00b755 (95% CI 1\u00b742\u20131\u00b767) and in the mefloquine (10 mg/kg) group was 2\u00b734 (2\u00b717\u20132\u00b752), corresponding to a parasite clearance half-life of 9\u00b74 h (8\u00b77\u201310\u00b72) and 6\u00b72 h (5\u00b77\u20136\u00b77), respectively. The median minimum inhibitory concentration of DSM265 in blood was estimated as 1040 ng/mL (range 552\u20131500), resulting in a predicted single efficacious dose of 340 mg. Parasite clearance was significantly faster in participants who received mefloquine than in participants who received DSM265 (p<0\u00b70001).In part 1, 73 participants were enrolled between April 12, 2013, and July 14, 2015 . In part 2, nine participants were enrolled between Sept 30 and Nov 25, 2013 . In part 1, 117 adverse events were reported; no drug-related serious or severe events were reported. The most common drug-related adverse event was headache. The mean DSM265 peak plasma concentration (CThe good safety profile, long elimination half-life, and antimalarial effect of DSM265 supports its development as a partner drug in a single-dose antimalarial combination treatment.Wellcome Trust, UK Department for International Development, Global Health Innovative Technology Fund, Bill & Melinda Gates Foundation. Over the last 15 years the number of deaths from malaria has decreased by 63% globally as a result of better access to medicines and insecticide-treated bednets.The target product profile of an antimalarial drug is a combination therapy that would increase compliance, potentially allowing directly observed therapy, and prevent transmission.Evidence before this studyPlasmodium dihydroorotate dehydrogenase (DHODH), an essential enzyme for pyrimidine biosynthesis.Although the incidence of malaria has declined drastically in the last 15 years, WHO estimated that in 2015 there were more than 200 million cases worldwide, resulting in 429\u2008000 deaths. The WHO Global Technical Strategy goal is to reduce malaria case incidence and mortality rates by 90% by 2030. To achieve this, new treatments for malaria will be needed. Additionally, the emergence of parasite resistance to current first-line antimalarials is threatening progress towards malaria elimination. To reduce the development of resistance, new treatments for malaria formulated as combination therapies are required. Such combinations should include components with different, and preferably novel, mechanisms of action. DSM265 is a new antimalarial candidate that was discovered through a target-based research strategy aimed at inhibiting ClinicalTrials.gov and the Australian New Zealand Clinical Trials Registry (ANZCTR). DSM265 is currently the first DHODH plasmodium inhibitor in clinical development for the treatment of malaria. Preclinical studies predicted a safety profile for DSM265 supporting human studies, as well as a long half-life in human beings. The in-vitro antiplasmodial activity of DSM265 was comparable with mefloquine and chloroquine. Recently, further clinical trials have been done to test the antimalarial activity of DSM265, including a study to assess its chemoprophylactic activity and a phase 2 study in clinical malaria.We searched PubMed between Jan 1, 2013, and Dec 1, 2016, using the terms \u201cdihydroorotate dehydrogenase plasmodium inhibitor\u201d and \u201cDSM265\u201d, and the clinical trials registries Added value of this studyP falciparum minimum inhibitory concentration for more than 8 days, suggesting that DSM265 could potentially result in a single-dose cure when used with a partner drug. Additionally, pharmacodynamic analysis showed that DSM265 acts slowly against P falciparum. Therefore, DSM265 could be combined with a fast-acting drug to develop an antimalarial combination therapy.Integration of a malaria challenge cohort within a phase 1 human study allowed concurrent assessment of DSM265 safety, pharmacokinetics, and antimalarial activity. Our results show that DSM265 has a favourable safety profile in human beings, and a pharmacokinetic profile characterised by a long half-life. Plasma concentrations remained above the Implications of all the available evidenceThe design of this study allowed rapid determination of the key pharmacokinetic and pharmacodynamic parameters of DSM265, and estimated an efficacious dose for clinical malaria. This information, together with safety data, was crucial to enable the progression to a phase 2 study with malaria patients in Peru, accelerating the clinical development of DSM265. This phase 2 trial has already been completed and will be published shortly.This study showed that DSM265 is a promising drug combination partner for single-dose treatment of acute uncomplicated malaria. A single-dose treatment would improve patient compliance compared with the current first line of treatment, which requires multiple daily dosing, and would therefore result in a better treatment outcome.Plasmodium falciparum-infected severe combined immunodeficient (SCID) mouse model, DSM265 had potent in-vivo antimalarial activity with 90% effective dose (ED90) of 3 mg/kg per day, comparing favourably with chloroquine (ED90=4\u00b73 mg/kg) and mefloquine (ED90=7\u00b77 mg/kg).P falciparum DHODH, but shows much less antimalarial activity than DSM265.Dihydroorotate dehydrogenase (DHODH) is essential for pyrimidine biosynthesis in plasmodia because they lack pyrimidine salvage pathways and rely entirely on de-novo synthesis of pyrimidines. DSM265 was discovered through a target-based research strategy to develop an inhibitor of the plasmodial DHODH with high selectivity compared with the human orthologue.P falciparum-infected erythrocytes for preliminary assessment of antiparasitic activity.We undertook this first-in-human study to assess the safety, tolerability, and pharmacokinetic profile of DSM265 and its metabolite DSM450. A standard phase 1 safety and pharmacokinetic study protocol was integrated with an induced blood-stage malaria (IBSM) cohort, a controlled human malaria infection whereby healthy participants were inoculated with P falciparum and treated with a single dose of DSM265 or mefloquine. Preclinical efficacy studies done in the SCID mouse model estimated the plasma minimum parasiticidal concentration for DSM265 to be in the range of 1000\u20132000 ng/mL.This study consisted of two parts. Part 1 was a phase 1a, single ascending dose, double-blind, randomised, placebo-controlled study. Part 2 was a phase 1b, IBSM, open-label, randomised, active-comparator controlled study, in which participants were inoculated with blood-stage Healthy men and women  aged 18\u201355 years were eligible for the study. Participants were excluded if they had hyper-sensitivity to any study drug. In part 2, participants were not to live alone for the duration of the study, not to have had visited a malaria-endemic area for a period greater than 2 weeks in the last 12 months, and not to have received recent or current therapy with an antibiotic or drug with potential antimalarial activity. Full inclusion and exclusion criteria are listed in the Randomisation lists for parts 1 and 2 were created using a validated, automated system. Participants in part 1 were randomly assigned to receive either a single dose of DSM265 or placebo. The DSM265 to placebo allocation ratio was: 2:1 in the 25 mg sentinel sub-cohort, 4:1 in the remainder of participants in this cohort, 8:2 in the 250 mg cohort (fasted\u2013fed), and 6:2 in the other cohorts (75\u2013150 mg and 400\u20131200 mg). Participants and investigators were masked to group allocation. Treatment identity was concealed by providing placebo and drug doses in identical packaging and appearance.Part 2 took place in two subgroups, part 2a and 2b. Participants in part 2 were randomly assigned to receive either a single dose of DSM265 or treatment with mefloquine at a ratio 4:1 in part 2a and 3:1 in part 2b. Masking was not possible because of the different formats of the drugs given. However, the laboratory where parasite quantification assays by quantitative PCR (qPCR) were undertaken was masked to allocation.In part 1, single ascending doses of DSM265 (25\u20131200 mg) or placebo were given to participants, randomised in eight dose cohorts. A starting dose of 25 mg was chosen in agreement with guidelines on dose selection for first-in-human studies based on preclinical safety data,DSM265  was supplied as a 25% (250 mg/g) spray-dried dispersion of the active pharmaceutical ingredient in hydroxypropylmethylcellulose acetate succinate (HPMCAS-MF) as powder in a bottle . The powder was suspended in vehicle , and given as an oral suspension on site. Plasma concentrations during the absorption phase of DSM265 in the 400 mg cohort showed lower exposure than expected due to incomplete dispersion of the drug. Thus, the preparation instructions were modified to ensure adequate product dispersion, and the 400 mg cohort was repeated, with subsequent dose preparation using the amended instructions. The placebo  was supplied as HPMCAS-MF as powder in a bottle, and suspended according to the same protocol.P falciparum-infected human erythrocytes (about 1800 viable parasites). Parasite growth was monitored by qPCR targeting 18S DNA.pfs25 mRNA,In part 2, an IBSM cohort was inoculated as described previously.Safety assessment was done at screening and at protocol-specified times . Safety Blood samples to determine concentrations of DSM265 and its major metabolite DSM450 were taken before DSM265 dosing and at 0\u00b75, 1, 2, 4, 6, 8, 12, 24, 48, 96, 144, 216, 312, and 480 h post-dosing. To adequately capture exposure in the 1200 mg cohort, two extra pharmacokinetic timepoints were included in this cohort at 648 h and 816 h post-dosing. The pharmacokinetic profile of DSM450 was determined from the 150 mg cohort onward. Blood and plasma samples were analysed by liquid chromatography-tandem mass spectrometry  were measured by HPLC-MS/MS  as exploP falciparum dhodh (pfdhodh) genetic modifications that could confer DSM265 drug resistance was investigated in any recrudescent parasite population. Blood was taken from participants in part 2 at the time of recrudescence. Parasite DNA was extracted from blood samples, the pfdhodh genes were amplified by PCR, and their DNA sequence determined (The induction of termined .max), timepoint when Cmax was reached (tmax), maximum blood or plasma concentration 7 days after drug administration (C168h), area under the concentration\u2013time curve from 0 h extrapolated to infinity (AUC0\u2013\u221e), and elimination half-life (t1/2).The primary endpoints were safety and the pharmacokinetic profile of single ascending doses of DSM265, as well as establishing the maximum tolerated dose (MTD) in healthy participants. The pharmacokinetic parameters determined were maximum blood or plasma concentration (C10PRR48) and parasite clearance half-life, assessment of the effect of food on DSM265 pharmacokinetics and tolerability, and characterisation of the pharmacokinetics of DSM450.Secondary endpoints were the logarithm of the parasite reduction ratio per 48 h =ln(a)\u2008+\u2008b\u2008\u00d7\u2008ln(x), where ln(a) is the intercept and b is the slope. To estimate the proportionality coefficient, a proportional model was fitted to the data in the log-transform form, ln(a)=ln(y/x), where a is the proportionality coefficient. Phoenix WinNonlin version 6.3 was used for both models. Pharmacokinetic parameters were determined by non-compartmental analysis using Phoenix WinNonlin version 6.3 in part 1 and R version 3.1.1 in part 2. In part 1, population pharmacokinetic analysis was done using the NONMEM software and the non-linear mixed-effects modelling approach ; R versi10PRR48 and parasite clearance half-life were estimated using the slope of the optimal fit for the log-linear relationship of the parasitaemia decay.10PRR48 and corresponding 95% CI were calculated using the slope and corresponding SE of the optimal regression model. The log10PRR48 and parasite clearance half-life for a dose cohort were derived using the weighted mean of the optimal slope for participants with an adequate model fit (p<0\u00b7001). An omnibus test was used to investigate differences in log10PRR48 between groups. R version 3.0.2 was used for analysis of parasite clearance. The timepoint at which the parasitaemia nadir occurred was estimated from individual log-linear parasite clearance curves. The DSM265 concentration at nadir was assigned as the MIC. Single-dose pharmacokinetic profiles of DSM265 were simulated by the population pharmacokinetic model developed with part 1 pharmacokinetic data to identify a single curative dose, defined as a dose that would maintain DSM265 concentration above the maximum estimated MIC for a minimum of 7 days.In part 2, logIn part 1, the difference in concentrations of uridine and uridine nucleotides from timepoint 0 h to 96 h between treatment groups was analysed by a two-sample t test. Differences in changes over time between treatment groups were determined by a linear mixed model with main effects and an interaction for time and treatment group. Analyses were stratified by DSM265 cohorts 800 mg and 1200 mg. All analyses were done in Stata version 13 (5% significance level).t tests at the 5% significance level, that a cohort size of eight would identify a difference of 25% in the parasite clearance rate with 80% power. Mefloquine control participants were not included in sample size calculations because their purpose was to observe parasite growth and clearance curves in response to a drug with known activity.Previous IBSM studies have determined, based on two-sided The studies were registered with the Australian New Zealand Clinical Trials Registry as number ACTRN12613000522718 (part 1) and number ACTRN12613000527763 (part 2).The funders of this study had no role in study design, data collection, analysis, and interpretation or reporting. The corresponding authors had access to all data in the study and had final responsibility for the decision to submit for publication.P falciparum, and treated on day 8 with either DSM265 or mefloquine  or placebo n=18; . Part 1 floquine . Part 2 vs one [6%] of 18; relative risk 2\u00b73 [95% CI 0\u00b742\u201314\u00b70], p=0\u00b767). Other drug-related adverse events included dry mouth, malaise, nausea, vomiting, decreased appetite, thrombocytopenia, and increased reticulocyte counts  was detected on day 5 in one participant in the 1200 mg cohort, and was suspected to be drug-related. This participant presented a non-clinically significant decrease in haemoglobin (from 137 g/L at baseline to 122 g/L 10 days after DSM265 dosing), and a clinically significant elevated reticulocyte count on days 10 and 14 ; the elevated reticulocyte was suspected to be drug-related. This participant was diagnosed with a haemoglobinopathy (sickle cell trait), unknown to the investigators at study entry. Review of individual reticulocyte count and haptoglobin concentrations of participants across all dose cohorts did not reveal any other clinically significant changes.In part 1, two cases of thrombocytopenia were reported after administration of DSM265 (400 mg and 1200 mg). The lowest platelet count  showed an increase in alanine aminotransferase , and another participant  had an increase in aspartate aminotransferase . No cardiac arrhythmias or increase in QTc were observed up to the highest DSM265 dose tested.In part 2, eight of the nine participants reported at least one adverse event. Of the 30 adverse events reported, 18 were deemed as probably related to malaria . There wmax increased with dose (range 1310\u201334\u2008800 ng/mL) in a less than dose-proportional manner (max across dose cohorts (coefficient of variation [CV%] range 22\u201340%). Cmax was reached in a median range between 1\u00b75 h and 4 h (tmax). C168h increased in an approximately dose-proportional manner (range 217\u201310\u2008200 ng/mL). Likewise, plasma AUC0\u2013\u221e increased with dose (range 107\u2008000\u20134\u2008720\u2008000 ng\u00b7h/mL) in an approximately dose-proportional manner between 25 mg and 250 mg, and then between 250 mg and 1200 mg, except the 800 mg dose, where sub-proportionality was observed. Inter-participant variability in AUC0\u2013\u221e was similar across dose cohorts (CV% range 18\u201339%). DSM265 showed a long elimination half-life, ranging from 86 h to 118 h across the doses tested. Assessment of the effect of food in the pharmacokinetic profile of DSM265 (250 mg) indicated that mean Cmax decreased with food intake and tmax increased . The pharmacokinetic profile was best described by a two-compartmental model with zero-order absorption, dose-dependent duration of absorption and bioavailability, simple exponential independent random effect specification, and proportional residual variability .168h for this dose in part 1 was 1230 ng/mL, which was at the lower end of the predicted efficacious concentration range determined in preclinical studies in plasma (1000\u20132000 ng/mL). Pharmacokinetic analysis of DSM265 from the seven participants in the IBSM cohort indicated that low blood-stage parasitaemia levels had no effect on the pharmacokinetic parameters , which corresponded to a median blood MIC of 1040 ng/mL (range 552\u20131500). Simulations in the population pharmacokinetic model developed with part 1 data predicted that a dose of 340 mg would maintain DSM265 blood concentrations above 1500 ng/mL, the maximum estimated MIC, for 7 days.10PRR48 estimated in participants with significant regression models of the log-linear relationship of the parasite decay in the DSM265  and mefloquine  groups were 1\u00b755 (95% CI 1\u00b742\u20131\u00b767) and 2\u00b734 (2\u00b717\u20132\u00b752), respectively, corresponding to a parasite clearance half-life of 9\u00b74 h (8\u00b77\u201310\u00b72) and 6\u00b72 h (5\u00b77\u20136\u00b77), respectively. Parasite clearance was significantly faster in participants who received mefloquine than in participants who received DSM265 (p<0\u00b70001). Parasite clearance for participants in part 2a was significantly lower than in part 2b  in five participants who received 150 mg DSM265 did not show mutations compared with the canonical 3D7A gene sequence.DNA sequence analysis of the P falciparum is crucial to the pharmacodynamics of DSM265, supporting its further clinical development as a component of a novel combination antimalarial therapy.In this study, we integrated for the first time the assessment of antimalarial activity of a drug within a first-in-human study, thereby accelerating the development of DSM265, the first plasmodium-selective DHODH inhibitor tested in human beings. The results show that DSM265 has a good safety profile and a long half-life, with a plasma concentration that remained above the MIC for more than 8 days. This prolonged effect against blood-stage Although DSM265 showed a good safety profile, the drug-related thrombocytopenia and elevated reticulocytes reported in a participant diagnosed with sickle-cell trait will require further attention in patients with clinical malaria.max was observed when DSM265 was given with high-fat food but this effect was not clinically significant.The elimination half-life of DSM265 was the longest of any of the new-generation antimalarials; however, it is still shorter than 4-aminoquinolines or aminoalcohols that have been tested in the IBSM model, such as mefloquine (10 mg/kg)P falciparum was characterised by an initial lag phase, accompanied by an increase in parasitaemia immediately post-treatment. This increase is probably due to a combination effect of the drug's modest speed of action and the lifecycle stage at which the drug exposure occurred.P falciparum was slower (log10PRR48 1\u00b755 [95% CI 1\u00b742\u20131\u00b767]) than other long-acting drugs tested in the IBSM model, such as mefloquine 10PRR48 2\u00b721 [2\u00b715\u20132\u00b727]).10PRR48 of the two groups in part 2 (part 2a and 2b), which was not clinically significant, since both values were in the range characteristic of slow-acting drugs. The small size of the groups (n=3) might have contributed to the log10PRR48 variability between them. The log10PRR48 reported for mefloquine in this study is in accordance with previous reports, which validates the reproducibility of the IBSM model.The activity of DSM265 against The dose selected for the IBSM cohort resulted, after initial parasite clearance, in recrudescence in all participants, which allowed calculation of the MIC for DSM265. The design used in this study enabled both identification of the MIC and the prediction of the efficacious dose within 6 months from initiation of clinical development. Applying a conventional development process of separate phase 1 in healthy participants followed by phase 2a proof-of-concept studies in patients would take over 2 years. In the next step of DSM265 clinical development, it would be important to evaluate its efficacy in clinical malaria to confirm the pharmacokinetic and pharmacodynamic parameters. Data available from previous IBSM studies indicate that the pharmacodynamic parameters estimated in this study will correspond to those obtained in malaria patients.pfdhodh gene amplification.pfdhodh gene were identified in the IBSM cohort. pfdhodh gene amplification events could not be studied due to low parasitaemia. Antimalarial resistance to other antimetabolites, such as sulfadoxine\u2013pyrimethamine and atovaquone, can be more readily induced in vitro and in vivo than for drugs with other mechanisms of action.Combining DSM265 with a partner drug would be beneficial not only from a pharmacodynamic point of view, but also to reduce the probability of developing drug resistance.Generalisability of the study findings is limited by the recruitment of only men due to the contraceptive requirements of the study impeding recruitment of women. Evaluation in a wider population will be needed in further studies. Another limitation of this study was that the preparation of the DSM265 formulation was cumbersome, and drug reformulation will be required in future studies.In conclusion, DSM265 is a novel antimalarial with a good safety profile. Its long half-life suggests that it is a promising drug combination partner for single-dose treatment of acute uncomplicated malaria. Treatment with single-dose DSM265 could improve patient compliance and thus treatment outcome compared with current treatment options, which require multiple daily dosing."}
+{"text": "In 2013, mutations in the propeller domain of the kelch protein K13 was found to be associated with artemisinin resistance. A cross-sectional study was conducted to determine the prevalence of Day-3 parasitaemia, estimate the frequency of k13 mutation assay.Blood smears and filter paper blood spots were collected from febrile patients in Kravanh District, Pursat Province. The blood smears were examined by microscopy, and blood spots by a k13 assay were interpretable for 76 of the 92 samples. The findings were: wild type: 9 (12%), C580Y: 64 (84%), Y493H: 3 (4%). Therefore, despite the high prevalence of k13 mutants (67/76: 88%), only 1 of the 92 patients remained blood smear positive for Plasmodium falciparum on Day-3.Data from 92 patients were analysed. Only one was positive for Day-3 parasitaemia. Results of the k13 mutation in Kravanh, but the result is not necessarily representative of the western part of Cambodia. Further investigation should be made to determine if k13 marker remains useful as a tool for tracking artemisinin resistance and predicting the trend of the efficacy of artemisinin combination therapy once the mutant alleles have been well established in the population.These preliminary findings suggest good potency of artemisinin despite the dominance of  Plasmodium falciparum malaria in Cambodia since 2000. Within a few years, local malaria control staff began to observe frequent recurrences of infection among patients in western Cambodia especially in Pailin Province and nearby areas. Therapeutic efficacy studies (TES) independently conducted in Pailin and across the border in Thailand\u2019s Chanthaburi Province during 2002\u20132003 indicated A+M treatment failures [Artesunate in combination with mefloquine (A+M) has been the first-line regimen for the treatment of failures , 2. Arte\u00ae) during 2013\u20132014. In nearby Pursat Province, dihydroartemisinin\u2013piperaquine (DHA-PIP) replaced A+M in 2012 at the time when DHA-PIP was deployed countrywide. The Cambodian national therapeutics guidelines earlier adopted DHA-PIP (in principle) as the first line drug for both P. falciparum and Plasmodium vivax in 2010.The first-line therapy for falciparum malaria in Pailin was switched to atovaquone\u2013proguanil . Extra blood drops were collected for a malaria smear and onto filter paper. Patients positive for P. falciparum were treated with DHA-PIP according to the national treatment guidelines. The first dose was taken under the supervision of and observed by the pharmacy owner, who also provided the patients with clear explanation for consumption of the remaining two doses at home. All patients were asked again to confirm completion of DHA-PIP doses when they returned on Day-3 for clinical and parasitological follow-up. The blood smears and filter paper blood spots also allowed us to determine the prevalence of Day-3 parasitaemia, estimate the frequency of K13 molecular marker and assess their relationship. Patients\u2019 demographic data were collected along with interview data on recent exposure to malaria. A verbal consent was obtained from every patient, which is sufficient as part of operational research in Cambodia.The study utilized data that was collected as part of routine health service delivery. During 29 June 2014 to 18 July 2014, febrile patients presented to a local pharmacy in Kravanh District in Pursat Province were routinely screened with combo-rapid diagnostic tests (RDT)  by the reference microscopist.Among the 100 Day-0 blood smears, 8 were read as negative by the reference microscopist. These were excluded from data analysis. There was one false negative reading by the local microscopist; a Day-3 negative smear was reported as k13 assay were interpretable for 76 of the 92 samples. The findings were: Wild Type: 9 (12%), C580Y: 64 (84%), Y493H: 3 (4%). Therefore, despite the high prevalence of k13 mutants (67/76: 88%), only one of the 92 patients remained blood smear positive for P. falciparum on Day-3. The Day-3 positive sample was identified positive C580Y.Data from 92 patients were analysed. Only one was positive for Day-3 parasitaemia. Results of the pfmdr1 copies numbers (a marker of mefloquine resistance) [k13 mutants have almost replaced the wild type in Kravanh, Day-3 positive parasitaemia is rare. The results indicate a lack of correlation between k13 mutation and the presence of Day-3 parasitaemia. Although evidence of DHA-PIP resistance was detected in the study area in 2014, in reality the first-line treatment was still DHA-PIP in February 2016. The findings of this study supported the continuation of the use of DHA-PIP.In Cambodia, after mefloquine withdrawal, the in vitro sensitivity to mefloquine improved along with a decrease in the prevalence of multiple istance) . Howeverk13 mutation in Kravanh. This raises some concern over the relevance of highly prevalent k13 mutation to artemisinin resistance. Further investigation should be made to determine if k13 marker remains useful as a tool for tracking artemisinin resistance and predicting the trend of ACT efficacy once the mutant alleles have been well established in the population.These preliminary findings suggest good potency of artemisinin despite the dominance of k13 mutation and artemisinin resistance in western Cambodia.When interpreting the findings, there is need for caution since the data were obtained as part of an ongoing service delivery, not collected under a stringent research setting. The timing of Day-3 parasitaemia samples was not based on an exact 72-h duration from the time of administration of the first DHA-PIP dose (the Day-3 collection from patients was beyond 72\u00a0h and <96\u00a0h) and the second and third doses might not have been exactly 24\u00a0h apart. In addition, slides were primarily read by a laboratory technician from the local health centre. However, slide cross-checks by the reference microscopist showed <10% discrepancy suggesting that the local microscopist had adequate competency to serve in such a remote malaria endemic area as in Kravanh. The microscopist also detected very low-density parasitaemia in several smears that might have been missed by most field microscopists. Overall, these preliminary data are considered sufficient to warrant a comprehensive study to re-assess the relationship between"}
+{"text": "This study aimed to demonstrate the noninferior efficacy of TachoSil vs. TachoComb in Japanese patients undergoing liver resection and to assess the safety of TachoSil vs. TachoComb in these patients.This randomized, double-blind, noninferiority study (JapicCTI-090684) involved participants scheduled for liver resection/living donors (age\u00a0\u2265\u00a020\u00a0years). TachoSil or TachoComb  was applied to control persistent exudative bleeding after primary hemostasis during liver resection/removal for donation. The primary outcome was hemostasis 5\u00a0min after study treatment application. The 95% confidence interval (CI) for the difference in the proportion of participants with hemostasis 5\u00a0min after application of TachoSil/TachoComb was determined; noninferiority of TachoSil was indicated if the lower limit of the CI was \u2265\u221214%. Adverse events (AEs) were recorded.All participants in the efficacy analysis  achieved hemostasis 5\u00a0min after study treatment application. Therefore, TachoSil was noninferior to TachoComb. All participants experienced \u22651 AE; however, none discontinued because of an AE. Most (\u226597.8%) AEs were mild or moderate in severity.These findings confirm the safety profile and noninferior hemostatic efficacy of TachoSil compared with TachoComb. Hemostasis during liver resection is a critical determinant of surgical success. Indeed, the volume of blood loss during liver surgery is an established predictor of morbidity and mortality \u20133. Of noTachoComb\u00ae and TachoSil\u00ae are widely used fibrin sealants for tissue adhesion/closure during different types of surgery, including liver, lung, cardiovascular, gynecological, and urological. TachoComb comprises a collagen patch coated with human fibrinogen and bovine thrombin and aprotinin. This product version has been marketed and widely used in Japan since 1999. The newer product version, TachoSil, is currently marketed in more than 50 countries worldwide, and it comprises a collagen patch coated with human fibrinogen and human thrombin. TachoSil was developed after TachoComb to avoid potential immunogenic effects of bovine thrombin  and anapAccordingly, the aims of this study were to demonstrate noninferiority in efficacy of TachoSil compared with TachoComb in Japanese patients undergoing liver resection and to assess and compare the safety of the two products in these patients.This was a multicenter, randomized, double-blind, noninferiority (TachoSil vs. TachoComb) study carried out at 11 sites in Japan from 18th April 2008 to 19th August 2009. The trial was registered in the Japan Pharmaceutical Information Center Clinical Trial Information database (JapicCTI-090684).The study protocol was reviewed and approved by the Institutional Review Board at each study site. The study was carried out in accordance with Good Clinical Practice (GCP), based on the ethical principles outlined in the Declaration of Helsinki and the International Conference on Harmonization-GCP Guideline, and all applicable local Japanese laws and regulations. All participants provided written informed consent.Patients scheduled for elective liver resection  and living donors were considered for inclusion in the study if they were aged \u226520\u00a0years at the time of informed consent.Patients and living donors were excluded from the study if they had a Child\u2013Pugh Classification of C; had a history of hypersensitivity to any of the ingredients of the investigational products, or to bovine blood-derived, bovine lung-derived, or equine blood-derived preparations; had a history of TachoComb use; were females who were pregnant, lactating, suspected of being pregnant, or who planned to become pregnant during the study period; had participated in any other clinical study within 180\u00a0days before providing informed consent; or were judged by the investigator as not suitable to participate in the study.Prior to surgery, participants were randomly allocated (1:1 ratio) to TachoSil or TachoComb using a central allocation system. Randomization was stratified by study site, Child\u2013Pugh classification (A or B), and platelet count . Intra-operatively, patients with severe surgical complications and patients with persistent bleeding after completion of the primary hemostatic procedures were excluded from the study.2) and human thrombin (1.38\u00a0IU/cm2). Active ingredients in the TachoComb patches were human fibrinogen (5.5\u00a0mg/cm2), bovine thrombin (1.38\u00a0IU/cm2), and bovine aprotinin (128\u00a0IU/cm2). The two study treatments were indistinguishable in their appearance and physical characteristics.TachoSil and TachoComb were provided  in the form of identical 9.5\u00a0\u00d7\u00a04.8\u00a0cm equine collagen patches of a 0.5\u00a0cm thick spongy material with a dry yellow coating of active ingredients on one side. Active ingredients in the TachoSil patches were human fibrinogen  treatments were used to control bleeding from the wound.Adverse events (AEs) were recorded from the time of informed consent until day 28 after treatment and were defined as any unfavorable medical occurrence or worsening of subjective symptoms/objective findings, worsening of underlying diseases and complications, or clinically significant abnormal laboratory values. Adverse events were coded according to MedDRA, version 11.0.Serological tests for hepatitis B and C virus (HBV and HCV) and human immunodeficiency virus (HIV) were performed the day before study treatment and at day 28.Based on the results from European clinical studies of patients undergoing liver resection, 95% of Japanese participants were expected to achieve hemostasis at 5\u00a0min. A sample size of 100 participants (50 per study group) was determined to provide 90% power at a noninferiority margin of 0.14.The efficacy analysis population for demonstrating non-inferiority was the per protocol set and included all enrolled participants who received the allocated study treatment and from whom any data was collected , in accordance with the intention-to-treat principle . The safThe primary efficacy analysis, designed to demonstrate noninferiority of TachoSil compared with TachoComb, was the 95% confidence interval (CI) for the difference between TachoSil and TachoComb in the proportion of participants with hemostasis 5\u00a0min after application of TachoSil or TachoComb. Noninferiority of TachoSil was considered to be demonstrated if the lower limit of the 95% CI was greater than \u221214%. The 95% CI for the proportion of participants with hemostasis at 5\u00a0min in each group was also calculated using the score method by Wilson . The difSafety analysis comprised the number of AEs, the number of participants who experienced AEs, and the number of participants who were HBV, HCV, and HIV positive at day 28.A total of 130 participants were randomized to treatment, 64 to TachoSil and 66 to TachoComb . The 95% CI for the proportion of participants who achieved hemostasis at 5\u00a0min was 93.4 to 100% for both groups. The 95% CI for the difference in the proportion of participants who achieved hemostasis at 5\u00a0min between study groups was \u22124.9 to 4.9%, indicating that TachoSil was noninferior to TachoComb.All participants in the efficacy analysis population in both study groups  achieved hemostasis 5\u00a0min after the application of study treatment. There was no difference in the response rate between study groups . The SAEs of post-procedural bile leak and peritonitis were considered to be possibly treatment related. The participant who experienced intra-abdominal hemorrhage died; this SAE was not considered to be related to study treatment because the event was a rupture caused by hepatic artery aneurysm resulting from the surgery. In the TachoComb group, nine participants experienced a total of 10 SAEs, including post-procedural bile leak (n\u00a0=\u00a04), and ventricular fibrillation, gastritis, abdominal abscess, upper limb fracture, chylothorax, and arterial hemorrhage . The SAE of ventricular fibrillation was considered to be possibly treatment related. One participant in the TachoComb group died after the study period due to worsening of underlying disease; this death was not considered to be related to study treatment.A small number of participants experienced at least one serious AE (SAE), most of which were not considered to be related to study treatment. In the TachoSil group, seven participants experienced a total of eight SAEs, including ventricular flutter, peritonitis, intra-abdominal hemorrhage, pyrexia, post-procedural bile leak, atelectasis, aneurysm ruptured, and vascular pseudoaneurysm  was generally similar between study groups Table , with woChanges in laboratory values were similar between groups (data not shown). One participant in the TachoSil group experienced a clinically significant, but nonserious, abnormal laboratory value of C-reactive protein increase that was considered related to study treatment. There were no new cases of HBV, HBC, or HIV infection, nor were there any positive conversions for participants who had positive virus markers at the start of the study.This is the first randomized trial to compare the efficacy and safety of TachoSil and TachoComb in Japanese patients undergoing liver resection or living donors. We found that TachoSil has noninferior hemostatic efficacy compared with TachoComb and appears to have a similar safety profile. Our findings therefore suggest that the beneficial effects of TachoSil are similar to those of TachoComb and that TachoSil is an effective means of providing hemostasis in Japanese patients during surgery.The primary finding of our study was that TachoSil and TachoComb provided similarly effective hemostasis within 5\u00a0min of application in all participants. The assessment of hemostasis was judged by the surgeon; however, the binary nature of the endpoint avoids any grading and thus renders some objectivity. This endpoint reflects the immediate pharmacodynamic effect of the products, and although of minor clinical relevance, this kind of efficacy endpoint is widely used and acknowledged as basis for the assessment of hemostatic effect of treatments in surgery , 20\u201322. TachoSil was developed to reduce the potential for immunogenic effects in response to bovine thrombin and bovine aprotinin with repeated use, and to remove the potential for horizontal disease transfer (bovine to human). While no specific antibody testing was undertaken in the current analyses, a recent study reported that among 97 patients using TachoSil for the secondary treatment of local bleeding after hepatic resection, immunogenicity findings did not appear to be clinically significant . No treaWe did not aim to assess the long-term safety of TachoSil in this study; however, the safety profile of TachoSil in clinical practice has been confirmed in postmarketing settings , throughA key strength of this study is the prospective, randomized, double-blind, multicenter design with predefined statistical analyses based on the intention-to-treat paradigm. Limitations include the length of follow-up, which may have been insufficient to detect longer term postoperative morbidities, and the moderate number of participants. Additionally, one of the authors (M. Kobayashi) was sponsor-affiliated, which incurs a risk of bias . HoweverThis study demonstrated that TachoSil has noninferior hemostatic efficacy compared with TachoComb, and a similar safety profile, in patients undergoing liver resection."}
+{"text": "Plasmodium falciparum chloroquine-resistance transporter (pfcrt) and multidrug resistance gene 1 (pfmdr1) genes have been reportedly selected after artemether\u2013lumefantrine treatment. However, there is a paucity of data regarding in vivo selection of P. falciparum Kelch propeller domain (pfkelch13) polymorphisms, responsible for artemisinin-resistance in Asia, and six putative background mutations for artemisinin resistance; D193Y in ferredoxin, T484I in multiple resistance protein 2, V127M in apicoplast ribosomal protein S10, I356T in pfcrt, V1157L in protein phosphatase and C1484F in phosphoinositide-binding protein.Individual drug treatment may select resistant parasites in the human body, a process termed in vivo selection. Some single nucleotide polymorphisms in Artemether\u2013lumefantrine efficacy study with a follow-up period of 28\u00a0days was conducted in northern Uganda in 2014. The above-mentioned genotypes were comparatively analysed before drug administration and on days; 3, 7, and 28\u00a0days after treatment.pfkelch13 and in all isolates in the six putative background genes except I356T in pfcrt, which had 2.4% of isolates as mixed infections. In vivo selection study revealed that all isolates detected in the follow-up period harboured wild type alleles in pfkelch13 and the six background genes.In 61 individuals with successful follow-up, artemether\u2013lumefantrine treatment regimen was very effective with PCR adjusted efficacy of 95.2%. Among 146 isolates obtained before treatment, wild-type alleles were observed in 98.6% of isolates in pfkelch13 and the six background genes may not play an important role in the in vivo selection after artemether\u2013lumefantrine treatment in Uganda. Different mechanisms might rather be associated with the existence of parasites after treatment.Mutations in The online version of this article (doi:10.1186/s12936-016-1663-1) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria in nearly all malaria endemic countries [P. falciparum malaria in Western Cambodia [Since the mid-2000s, artemisinin-based combination therapy (ACT) has been deployed as first-line treatment for uncomplicated ountries . The widCambodia , geograpCambodia , 5. In ACambodia , there iCambodia \u201310.pfkelch13 (PF3D7_1343700) was identified as a useful molecular marker for tracking the emergence and spread of artemisinin resistant P. falciparum. Pfkelch13 encodes a 726 amino acid protein with a broad-complex, tramtrack, bric-a-brac/poxvirus and zincfinger (BTB/POZ) domain and a C-terminal 6-blade propeller domain [ferredoxin (fd), T484I in multidrug resistance protein 2+ (mdr2), V127M in the apicoplast ribosomal protein S10 (arps10), I356T in chloroquine-resistance transporter (crt), V1157L in protein phosphatase (pph) and C1484F in phosphoinositide-binding protein (pibp) [In 2014, r domain . Some mur domain . In addipfcrt and N86, 184F and D1246 in pfmdr1 [pfkelch13 [During the period of antimalarial treatment, less susceptible parasites can be selected in the human body, a process termed in vivo selection. This is because the treatment would create drug concentration circumstances that are sufficient to kill susceptible, but not less susceptible parasites. Previous investigations revealed that artemether\u2013lumefantrine (AL) treatment selected for parasites harbouring alleles with K76 in n pfmdr1 \u201317. Howepfkelch13 polymorphisms, and putative background SNPs for artemisinin resistance in P. falciparum recurrent infections was done to evaluate whether AL treatment selected for polymorphisms in a region where it has been used for a long time.In Uganda, AL was adopted as a first line treatment for uncomplicated malaria in 2004, but actual implementation was in 2006. So far, marked clinical efficacy has been reported to this regimen; 0\u20130.5% of cases with residual parasites by day 3 \u201328 and 1Anopheles funestus is the major mosquito vector and a few infections are due to Anopheles gambiae [P. falciparum positive results by rapid diagnostic test (RDT) were referred to the study physicians. The criteria for recruitment are shown in Additional file The study was conducted at the peak of malaria transmission between May\u2013July and October\u2013November 2014 at St Mary\u2019s Hospital Lacor in Gulu district, Northern Uganda. Malaria transmission in the study region is perennial with an estimated prevalence of >60%, and entomological inoculation rate (EIR) of 100 or more infective mosquito bites per person per year . Anophel gambiae . Malaria gambiae . SymptomBefore enrollment, written informed consent was obtained from the participants\u2019 parents or guardians, and children aged \u22657\u00a0years were assented. The study was reviewed and approved by Lacor Hospital Institutional Research and Ethics Committee (LHIREC) (Study protocol number LHIREC 008/05/2013 and 021/09/13) and regulatory approval was obtained from the Uganda National Council for Science and Technology (UNCST) (HS 1395).\u00ae, Novartis 20\u00a0mg artemether/120\u00a0mg lumefantrine tablets) was orally administered twice daily for 3\u00a0days and follow-up assessments were performed on days 1, 2, 3, 7, and 28 after initial drug treatment. Dosage of oral Coartem\u00ae was adjusted according to the participant\u2019s body weight: one (5\u201314\u00a0kg), two (15\u201324\u00a0kg), or three (25\u201334\u00a0kg) tablets. The drug was given as directly observed treatment (DOTS) for all patients by study nurses and physicians. After each treatment, patients were carefully observed for 30\u00a0min, and the same dose was re-administered if vomiting occurred. Rescue treatment regimen (dihydroartemisinin\u2013piperaquine) was administered daily for 3\u00a0days to any individuals who failed on the initial AL therapy. If the recruited patients developed severe malaria during follow-up, they were referred to the hospital for parenteral artesunate.For the efficacy study, AL  were obtained from the cubital vein before initial treatment except for children <2\u00a0years where finger prick sampling was performed. A finger-prick blood sample of 100\u00a0\u03bcL was obtained at each follow-up visit. Blood was spotted on chromatography filter paper . Haemoglobin (Hb) concentration was measured using a portable spectrophotometer Hemocue Hb 201  on days 0 and 28 or on the day of late clinical failure. Anaemic patients with Hb level <10.0\u00a0g/dL were treated with Ferrous sulphate tablets for 14\u00a0days. Plasma concentrations of artemether and lumefantrine were not measured. Treatment outcomes were classified according to WHO guidelines for areas of intense malaria transmission as: adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late clinical failure (LCF) and late parasitological failure (LPF) .P. falciparum infections were assessed by species-specific PCR as previously described [merozoite surface protein 1 (msp1), merozoite surface protein 2 (msp-2) and glutamate rich protein (glurp), was also performed to differentiate between recrudescence and new infections on day 0 and the day of positive infection [msp-1 and msp-2 and 1.5% agarose for glurp. Patient samples were run side by side. Gel images were digitised and molecular weights determined using imagej software [msp 2 and 20\u00a0bp for glurp.Thick and thin blood smears were stained with 2% Giemsa for 30\u00a0min. The number of parasites was counted per 200 white blood cells (WBCs), assuming 8000\u00a0WBC/\u00b5L. Parasite density was calculated by averaging independent counts made by two microscopists. Discordant results (difference in parasite density of >50%) were re-examined by a third microscopist and, parasite density calculated by averaging two closest counts. Slides were considered negative if after examination of thick smears, no parasite was detected in 100 high power fields. Parasite DNA was extracted from a quarter of dried blood spot (25\u00a0\u00b5L) using a QIAamp DNA Kit  . In all escribed . Genotypnfection , 37. Nessoftware , an openpfcrt (K76T) and pfmdr1  were determined as reported [P. falciparum 3D7 full-length sequence of K13-propeller domain (PF3D7_1343700) from PlasmoDB [Polymorphisms in reported , 40. K13reported . The seqreported  with P. fd, T484I in mdr2, V127M in arps10, I356T in crt, V1157L in pph and C1484F in pibp) were amplified by multiplex PCR using gene specific primers  as a dark quencher at 3\u2032 end.Six background mutations for artemisinin resistance  and a mutant-type (a strain from Thailand identified during preliminary experiments) were used as positive controls. Nucleotide sequence data are available in the GenBank\u2122, EMBL, and DDBJ databases under the accession numbers: LC193525\u2013LC193693.To evaluate performance of the SNP assay system in all 6 genes, two 2) test were used for comparison of categorical variables, and Wilcoxon rank sum test was used for the continuous variables. P values <0.05 were considered statistically significant. R statistical software  was used for all the statistical analyses.Kaplan\u2013Meier product limit formula was used for the estimation of day 28-PCR-adjusted cure rates, which was the primary efficacy endpoint. The uncorrected and PCR-corrected Kaplan\u2013Meier cumulative treatment success rates up to day 28 were calculated for all participants. Patients were censored when either lost to follow-up or withdrawn from the study. Either Fisher\u2019s exact test or Pearson\u2019s Chi square test . Other characteristics remained comparable between age groups.A total of 672 patients were screened, out of which 169 microscopically confirmed p\u00a0=\u00a00.0111) higher median parasitaemia at enrollment than the PCR negative group . This suggests that parasite biomass before treatment may also be associated with treatment success and PCR parasite-positive outcome on day 3  were used for genotyping of drug-resistance related alleles. Amino acid substitutions in Pfkelch13, which have been associated with artemisinin resistance in the Greater Mekong Sub region [fd, mdr2, arps10, pph and pibp. Mutant alleles were only observed in pfcrt as mixed alleles (I356\u00a0+\u00a0356T) (n\u00a0=\u00a03). Regarding the other polymorphisms in pfcrt, K76T, which is known to be the responsible genetic change for chloroquine resistance, was observed in 29.2% of the isolates. In pfmdr1, wild-type alleles were nearly fixed at all known polymorphic positions other than 184; position 86 (98.0%), 1034 (98.6%), 1042 (100%) and 1246 (93.7%), respectively. At position 184, the prevalence of wild type alleles was high (80.4%), but as above, was significantly lower than at the other positions (p\u00a0<\u00a00.0001 by Pearson\u2019s Chi square test).In addition to 64 patients for the therapeutic efficacy study, 105 patients who did not meet inclusion criteria for the therapeutic efficacy study were recruited into the molecular epidemiological study for drug resistance. In total, 146 b region  were notb region . All isoPlasmodium falciparum positivity after AL treatment was assessed by PCR in 61 individuals who were treated and successfully completed 28\u00a0days of follow-up. There were 23 PCR positive samples: 14 on day 3, 4 on day 7, and 5 on day 28. In pfkelch13 and the six background genes for artemisinin resistance , all isolates carried wild-type alleles on day 3, 7 and 28  and the follow-up period   as well. In contrast, position 184 remained polymorphic throughout the follow-up period without any trend of particular alleles except on day 7 where mutant alleles predominated  showed microscopically detectable parasites by day 3. Overall, similar excellent early response to AL treatment has been reported in other regions in Uganda \u201328. AccoPCR-confirmed parasite positivity after AL treatment was much higher than microscopically confirmed positivity; 91.8% on day 1, 54.1% on day 2, and 23% on day 3. These prevalences were similar to two previous studies that molecularly assessed parasite positivity in Kenya and Tanzania , 46. Bespfkelch13 at enrollment. The only mutation observed in Pfkelch13 was A578S, which has been widely distributed in Africa [Mechanisms of artemisinin resistance have been gradually uncovered albeit the overall picture has not been clarified. Enhanced stress response including activation of unfolded protein response and the PI3K/Pi3P/AKT pathway is thought to be the main mechanism for parasite survival in the presence of artemisinin , 48. Pfkn Africa . Computan Africa . Howevern Africa  and othen Africa , 18, 53.n Africa . This obn Africa , geneticpfkelch13 mutation was not observed in the parasite positive samples on day 3, 7 and 28, consistent with the recent observations [In vivo selection analysis revealed that rvations . Also, nrvations , 17. In rvations ; 68 versOne patient showed day 3 parasite positivity in the present study. Multiple factors are associated with delayed parasite clearance after AL treatment . QualityRole of the human immunodeficiency virus (HIV)-infection to malaria treatment efficacy with ACT remains to be elusive. Some studies showed a close association between HIV infection and lower ACT treatment outcome , but othUnlike previous studies with malaria recurrence rates of 35\u201360% after artemether\u2013lumefantrine treatment in Uganda \u201324, 26, P. falciparum malaria after 8\u00a0years of use in a region of high malaria transmission in Uganda. No clear evidence was obtained for the selection of mutant alleles in pfkelch13 and the six-background genes, all of which have been reported to be associated with artemisinin resistance in Southeast Asia. Close monitoring of AL efficacy is necessary as we seek to understand the influence of anti-malarial treatments and parasite persistence, malaria transmission setting and different genetic background of parasites and host interactions in the mutational process. This work was done where very low early treatment failure and recurrence as well as scarcity of mutant alleles in pfkelch13 and the six-background genes were observed; if the circumstances change, repeated in vivo selection analysis is recommended.This study demonstrated that AL treatment remains of high efficacy for the treatment of"}
+{"text": "Plasmodium falciparum in all endemic countries. In 2010, the combination dihydroartemisinin\u2013piperaquine (DP) was recommended for the treatment of uncomplicated P. falciparum malaria. DP is one of the first-line treatments used by the French army since 2013.Artemisinin-based combination therapy (ACT) introduced in the mid-1990s has been recommended since 2005 by the World Health Organization as first-line treatment against P. falciparum clinical failure with DP at day 20 was described in a 104\u00a0kg French soldier deployed in Djibouti. He was admitted to hospital for supervision of oral treatment with DP [40\u00a0mg dihydroartemisinin (DHA) plus 320\u00a0mg piperaquine tetraphosphate (PPQ)]. This corresponded to a cumulative dose of 4.6\u00a0mg/kg DHA and 37\u00a0mg/kg PPQ in the present patient, which is far below the WHO recommended ranges. No mutation was found in the propeller domain of the Kelch 13 (k13) gene, which is associated with artemisinin resistance in Southeast Asia. Pfmdr1 N86, 184F, S1034 and N1042 polymorphisms and haplotype 72\u201376 CVIET for the pfcrt gene were found in the present case. There was no evidence of resistance to DP.A case of This case confirms the risk of therapeutic failure with dihydroartemisinin\u2013piperaquine by under-dosing in patients weighing more than 100\u00a0kg. This therapeutic failure with DP by under-dosing highlighted the importance of appropriate dosing guidelines and the need of research data  in over-weight patient group. Plasmodium falciparum in all endemic countries. Combined with wide-spread use of insecticide-treated bed nets, ACT has led to a significant reduction in malaria incidence and mortality, especially in sub-Saharan Africa  at the maximal dosage of four pills once daily between meals for 3\u00a0days, recommended by the Sigma-Tau laboratory in absence of data on which to base a dose recommendation in patients weighing above 100\u00a0kg. He was discharged at the third day with no more fever and a negative QBC test. He took the correct doxycycline prophylaxis and used a bed-net. He was asymptomatic at day 12 with a negative QBC test. At day 20, he complained of fever (38.9\u00a0\u00b0C) and diarrhoea. Laboratory examination revealed recurrence of P. falciparum with 0.5\u00a0% parasitaemia and the absence of co-infection. A thin blood smear found late-stage trophozoites without gametocytes. He was successfully cured by the standard oral therapeutic course of a fixed atovaquone\u2013proguanil combination , four pills during meals once daily for 3\u00a0days. Clinical and parasitological follow-up were completed at day 22, 27, and 48 without recurrence.A 35-year-old French soldier was deployed to Djibouti. He had no medical history, no recent trips except a return to France during summer holidays and intermittent observance of doxycycline (100\u00a0mg/day) malaria prophylaxis. He presented with fever, headaches, and diarrhoea for 4\u00a0days with paracetamol self-medication. His weight was 104\u00a0kg. His blood pressure was 143/78\u00a0mmHg, and his pulse rate was 84\u00a0bpm. A rapid diagnostic test (RDT) for malaria was positive for P. falciparum isolate was sequenced and searched for polymorphisms involved in antimalarial drug resistance. The k13 gene was amplified by polymerase chain reaction (PCR) with the nested PCR method previously described   . These dk13 mutant parasites collected from recrudescent patients after a 3-day DP treatment were resistant in vitro by PPQ survival assay [k13 gene was found in the present patient\u2019s isolate. However, clinical failure by resistance cannot be excluded even if no mutations were identified in k13. Artemisinin failures seem not to be associated with k13 polymorphisms in Africa [However, clinical failure by resistance cannot be excluded. Multidrug resistance to DP is currently emerging in Cambodia, where recrudescent infections have increased from 15.4\u00a0% in 2011\u20132013 to 39\u00a0% in 2012\u20132014; 84\u00a0% of treated patients presented with parasite clearance half-lives longer than 5\u00a0h in 2012\u20132104, and 57\u00a0% were still parasitaemic at 72\u00a0h , 34. Allal assay . DHA resn Africa . Howeverpfcrt and pfmdr1 have been identified in recurrent infections with ACT partners like lumefantrine, mefloquine or amodiaquine [pfcrt or pfmdr1 genes in PPQ or DHA resistance [pftetQ gene, suggesting that the parasites were resistant to doxycycline [Decreased susceptibility or parasite resistance to artemisinin partner drugs has been observed in Africa, where single nucleotide polymorphisms in diaquine \u201341. Howesistance , 42. Theycycline \u201320. ThisSub-optimal dosing of either component of ACT can result in incomplete elimination of the parasite biomass and subsequent recrudescence, both of which are important driving forces for the selection of parasites with reduced drug susceptibility. ACT regimens should absolutely be deployed using optimal dosing strategies to maximize the likelihood of rapid clinical and parasitological cure, minimize transmission, and retard the onset and spread of resistances. Overweight patients are at particular risk of under-dosing. In some military units, the soldiers often weigh more than 90\u2013100\u00a0kg. Should physicians automatically exclude the use of dihydroartemisinin\u2013piperaquine as an anti-malarial treatment in these patients in the absence of data in patients weighing above 100\u00a0kg? Until studies assessing how to adapt treatment are performed (increase the duration or the daily dose or repeat a course later), patients weighing more than 100\u00a0kg should be aware of the higher risk of therapeutic failure and should be closely monitored at least until day 28, and beyond if possible. This therapeutic failure with DP by under-dosing highlighted the importance of appropriate dosing guidelines and the need of research data  in over-weight patient group."}
+{"text": "Plasmodium falciparum malaria. Reported here are the results of a large phase II, randomized, open-label, multicenter trial conducted in African infants (6 to 12 months of age) from Mozambique, Burkina Faso, The Gambia, the Democratic Republic of the Congo, and Tanzania. Primary efficacy endpoint was the PCR-corrected adequate clinical and parasitological response (ACPR) at day 28. Analysis was performed for the intention-to-treat (ITT) and per-protocol (PP) populations. A total of 201 patients received the dispersible-tablet formulation, and 99 received the conventional one administered as crushed tablets. At day 28, the PCR-corrected ACPRs were 86.9% (ITT) and 98.3% (PP) in the dispersible-tablet group and 84.9% (ITT) and 100% (PP) in the crushed-tablet group. At day 42, these values were 85.9% (ITT) and 96.5% (PP) in the dispersible-tablet group and 82.8% (ITT) and 96.4% (PP) in the crushed-tablet group. The comparison between survival curves for time to new infections showed no statistically significant differences (P = 0.409). The safety and tolerability profile for the two groups was similar in terms of type and frequency of adverse events and was consistent with that expected in African infants with malaria. A standard 3-day treatment with the new dispersible DHA/PQP formulation is as efficacious as the currently used tablet in African infants and has a comparable safety profile. Artemisinin combination therapies are considered the mainstay of malaria treatment, but pediatric-friendly formulations for the treatment of infants are scarce. We sought to evaluate the efficacy and safety of a new dispersible-tablet formulation of dihydroartemisinin/piperaquine phosphate (DHA/PQP) in comparison to the marketed tablet (Eurartesim) in the treatment of infants with uncomplicated  The last 15 years have witnessed impressive advances in the global fight against malaria, including in sub-Saharan Africa, where the highest burden of morbidity and mortality is still concentrated . HoweverEurartesim is a fixed-dose combination composed of dihydroartemisinin (DHA) and piperaquine phosphate (PQP) . This paPlasmodium falciparum malaria has been demonstrated in two phase III trials, respectively conducted in 1,553 African children and 1150 Asian adults and children  with uncomplicated Patient disposition as presented in the case report forms is summarized in P = 0.349 by the Fisher exact test). Among patients reported as \u201cother\u201d in The most common reasons for study withdrawal were malaria recurrence (classified as treatment failure) and informed consent withdrawal, occurring with a similar frequency in both treatment groups. Repeated vomiting after drug administration was more frequent in the dispersible arm compared to the crushed arm (5% versus 2%), although such a difference was not statistically significant . Two more patients were excluded from the PP population (one per each treatment group) since it was not possible to assess their outcome on day 28 not having the patients attended this visit and presenting malaria recurrence at day 42. There were also seven randomized patients presenting baseline parasitemia outside the range indicated in inclusion criteria, but all of these violations were judged as \u201cminor\u201d by the Data Safety Monitoring Board (DSMB), and according to the study protocol these patients were included in the PP population. Overall, there was a good balance for demographic and baseline characteristics between treatment arms , while the uncorrected ACPRs were nearly identical (80.9 and 80.8%). At day 42, the PCR-corrected ACPRs were to some extent lower than those observed at day 28 for both arms . SimilarP = 0.519 [chi-square test]) and 2.5 and 1.0% of recrudescences (P = 0.667 [Fisher exact test]). When estimated by Kaplan-Meier survival analysis, the cumulative risks of new infection at day 42 were 19.8 and 23.6% for the dispersible and crushed groups, respectively .The occurrence of malaria recurrences by time points, as well as other reasons for treatment failure, are shown in ectively , whereasP = 0.825 [chi-square test]; ITT population).One ETF occurred during the study due to the evolution of uncomplicated into severe malaria in a patient enrolled 2 days after initial malaria symptoms. The complication occurred 20 h after the first study drug intake and was probably due to the late starting of antimalarial treatment. At day 42, the overall proportions of treatment failures (ETFs plus LTFs) were similar between the treatment arms, corresponding to 42 cases (21.1%) in the dispersible-tablet group and 22 cases (22.2%) in the crushed-tablet group (P = 0.653).Parasite clearance was rapid in both treatment arms. In the ITT population, fewer than half of the patients were parasitemic at day 1 , with only few of them remaining parasitemic at day 2 (3.0% versus 2.0%). With the exception of one patient who was still parasitemic up to day 7 (dispersible-tablet group), all other patients were aparasitemic by day 3. No statistically significant differences in parasitemia clearance between treatment groups were found using the log-rank test  and 84 (84.9%) patients, respectively, treated with the dispersible or crushed formulations, had at least one AE during the study. Anemia, vomiting, diarrhea, rhinitis, and cough were the most frequently reported AEs and were often associated with malaria, influenza, and respiratory tract infections. The occurrence of laboratory AEs, e.g., altered liver enzymes (AST), was very rare and was similar between the treatment groups.P = 0.139 [chi-square test]) (P = 0.105 [chi-square test]). Cutaneous reactions were infrequent, with only one case of rash (crushed group). One patient in the dispersible-tablet group developed moderate anemia, which occurred 1 week after treatment and resolved 3 weeks later. Overall, none of the TEAEs required hospitalization.The proportion of patients experiencing at least one treatment-emergent adverse event (TEAE), for which the relationship with the study drug was suspected, was slightly more prevalent in patients treated with the crushed-tablet formulation than in patients treated with the dispersible-tablet formulation  . In partThere were no significant differences between treatment groups in ECG parameters at baseline and during follow-up. A relevant and comparable reduction of HR (heart rate) in both treatment groups was observed at day 2 and at day 7. Related with this reduction, an increase in PR, QT, and QTcF intervals was observed in both groups with a comparable extension. A change in QTcF interval from baseline was detectable at day 2 predose and increased after drug administration, with 22 patients in the dispersible-tablet group (11.1%) and 14 patients in the crushed-tablet group (14.1%) manifesting a QTcF prolongation over 60 ms, none of which included clinical translation. No cases of QTcF prolongation higher than 500 ms were reported. At day 7, the QTcF prolongation tended to normalization, with only one patient in the dispersible-tablet group and two patients in the crushed-tablet group still maintaining QTcF prolongation. Overall, no arrhythmias or any other cardiovascular AEs were reported.Two severe adverse events (SAEs) occurred during the study, both judged unrelated to the study treatment. The first SAE involved the death of a 6-month-old male patient enrolled in the crushed-tablet group. He received the full treatment course and was discharged from the hospital after malaria resolution (day 3). At day 27, the child was seen as an outpatient in a peripheral health post, with a history of fever, cough, and diarrhea. After a positive rapid diagnostic test for malaria, he was treated with artemether/lumefantrine. However, a malaria diagnosis was not confirmed by microscopy, and 2 days later the mother reported the child's death. The investigator considered sepsis the most likely cause of death . The second SAE occurred in a male of 11 months enrolled in the dispersible-tablet group. He received the first dose of the study drug, and 20 h later the patient condition deteriorated, with evidence of severe anemia  and polypnea. The study treatment was interrupted, and the patient received parenteral quinine for malaria and blood transfusion for anemia. After 4 days of hospitalization, followed by oral treatment with quinine for a further 5 days, the event was resolved.According to guidelines on the clinical investigation of medicinal products in a pediatric population, there is a need to develop new formulations that allow accurate dosing and enhance patient compliance in infants and young children . This rePresently, there is only one ACT reported in the last updated version of the WHO Model List of Essential Medicines for Children (last amended on August 2015), containing arthemether-lumefantrine, which was specifically developed as a dispersible formulation to improve the effectiveness and accuracy of ACT dosing in infants and young children  to allow drug concentrations in the blood to fall below the minimum therapeutic threshold . Prolongation of the QT interval was assessed using both Fridericia's and Bazett's correction methods but, notably, both formulas could be biased by the physiological high HR present in infants and exacerbated by the ill status and fever. In fact, baseline evaluations differed drastically (by \u223c60 ms) when the different correction methods were applied. The reduction in HR observed after drug administration could also induce a bias in the estimation of QTc. Accordingly, QTcF recorded at the screening visit was lower than that recorded at day 7 (335 \u00b1 16 ms versus 410 \u00b1 22 ms), when malaria was resolved and the blood level of piperaquine was below the concentration considered relevant for sustaining QT interval prolongation. These considerations suggest that the QTcF assessment in infants by DHA/PQP could be overestimated. Nonetheless, QTcF never exceeded the normal limits (450 to 470 ms), no cases evolved in rhythm disturbance, and all of the changes observed during therapy were never associated with clinical signs of cardiotoxicity.The dispersible formulation was well tolerated, and its safety profile was comparable to the crushed tablet. No new safety issues arose from the present study, and all findings were in line with the former ones. Most of the commonly reported AEs were symptoms of malaria. Similarly, the pattern of changes in laboratory variables was consistent with acute malaria and its resolution, with no differences between treatment groups. The most common drug-related AE was vomiting, and the corresponding frequency was similar to that previously reported in African children treated with crushed or uncrushed tablets . ImportaDispersible tablets are expected to contribute to ease of drug administration in subjects having difficulties in swallowing whole tablets, i.e., infants and young children. Since the dispersible formulations were similar in efficacy and safety to the conventional formulation in the infant population, we believe that this highly efficacious formulation should be made more readily available to all children below 5 years of age. Cost savings are also likely with its use, with the potential benefit of improving the acceptability of the combination once available on the market. It is noteworthy that a recent cost-effectiveness analysis of DHA/PQP for first-line treatment of uncomplicated malaria in African children showed that the use of this ACT, particularly in areas with moderate to high malaria transmission, is clearly justifiable from both clinical and economic perspectives , 19.The results presented here are part of a large study that was designed to evaluate also the population pharmacokinetics of piperaquine and dihydroartemisinin in African infants, since it appears important to demonstrate bioequivalence of the new formulation on PK grounds once similar efficacy and safety have been confirmed. The new dispersible formulation of Eurartesim is easy and safe to administer and provides better grounds for an enhanced compliance and effective treatment; hence, this should facilitate drug registration by Regulatory Authorities, prequalification by the WHO and, finally, full adoption in malaria control programs. Further studies are needed to clearly establish the improvement of the new formulation on compliance, hoping that the simplicity of its administration will improve substantially the adherence to the full 3-day course of treatment, therefore impacting the reduction of malaria morbidity and mortality in infants and young children and minimizing parasite resistance to ACTs.Between November 2013 and June 2015, a phase II, randomized, open-label, multicenter trial was performed in seven sites of five African countries: Centro de Investiga\u00e7\u00e3o em Saude da Manhi\u00e7a, Maputo, Mozambique; Kinshasa School of Public Health, University of Kinshasa, Democratic Republic of the Congo; Centre Muraz Bobo-Dioulasso, Nanoro, Burkina Faso; Centre National de Recherche et de Formation en Paludisme, Ouagadougou, Burkina Faso; Ifakara Health Institute, Bagamoyo, Tanzania; National Institute for Medical Research, Korogwe, Tanzania; and Medical Research Council Unit, The Gambia.The study protocol was approved by the Institutional Review Board of the Hospital Cl\u00ednic of Barcelona (Spain) and by the National Ethics Review Committee and/or Institutional Review Board at each trial site. The trial was conducted under the provisions of the Declaration of Helsinki and in accordance with GCP guidelines , 21. A dP values and confidence intervals. The inclusion criteria were age 6 to 12 months, a body weight of \u22655 kg, a P. falciparum monoinfection with asexual parasite densities between 1,000 and 200,000 parasites/\u03bcl of blood, fever (axillary temperature of \u226537.5\u00b0C), or a history of fever in the preceding 48 h. Exclusion criteria were previous treatment with antimalarials, acute malnutrition, severe malaria, danger signs, moderate/severe anemia (Hb < 7 g/dl), a family history of sudden death or known congenital prolongation of the QT interval, or treatment with QT prolongation inducers or strong cytochrome-P450 inhibitors/inducers or antiretroviral drugs . Patients satisfying the inclusion and exclusion requirements were enrolled if the parent or guardian signed a written informed consent.A targeted number of 300 infants  with uncomplicated malaria were enrolled in the study. Since the study was designed to evaluate also the population pharmacokinetics of piperaquine and dihydroartemisinin in African infants, the sample size has been estimated on the basis of previous PK studies , 23, andRandomization was centralized using an interactive web-based response system. A randomization list was generated by an independent contract research organization , with each treatment allocation concealed in sealed opaque envelopes that were opened by the investigators only after patient randomization. An allocation ratio of 2:1 (dispersible versus crushed formulation) was applied, balancing patients for sex, to recruit 200 patients to receive the dispersible DHA/PQP formulation and 100 patients to receive the marketed tablet. No stratification for countries or sites was applied.The DHA/PQP dispersible formulation was a coformulated, water-dispersible flat tablet, provided in two different strength dosages: 10/80 mg and 20/160 mg of dihydroartemisinin/piperaquine tetraphosphate (as cellulose-microencapsulated piperaquine tetraphosphate) and other components . The marketed Eurartesim formulation was a coformulated, film-coated tablet, provided in one strength of 20/160 mg of DHA/PQP  . Both foDHA/PQP administration has a food interaction effect resulting in an increase of piperaquine absorption when concomitantly administered with high-fat food . To reduPatients were kept at the health facility for the 3-day dosing period. Scheduled visits during follow-up were at days 7, 14, 21, 28, and 42 posttreatment. In case of recurrent parasitemia, rescue treatment was performed according to local guidelines.P. falciparum infection was made at each site by microscopy using standard methods .A 12-lead electrocardiogram (ECG) was recorded for each patient at enrollment (baseline) and then repeated at day 2 before the last drug administration, as well as after 4 to 6 h of drug intake. An ECG was also recorded at day 7 and repeated at day 28 if clinically relevant abnormalities were detected at day 7. All ECGs were digitalized using an ELI-150 cardiograph , and ECG reading and analysis were centralized by a cardiac laboratory .The main efficacy endpoint was the PCR-corrected adequate clinical parasitological response (ACPR) at day 28. Other efficacy and safety endpoints included the following: (i) day 28, PCR-uncorrected ACPR; (ii) day 42, PCR-corrected and -uncorrected ACPR; (iii) the proportion of patients with early and late treatment failure (ETF and LTF); (iv) asexual parasite density and clearance time; (v) fever clearance time and gametocyte carriage over time; (vi) Kaplan-Maier survival analysis for new infections and recrudescences over time; (vii) AE occurrence; and (viii) changes in hematology, blood chemistry, vital signs, and ECG parameters.in vivo efficacy definitions . Patients for whom the PCR indicated a new infection were considered failures in the PCR-uncorrected analysis and successes in the PCR-corrected analysis, whereas patients with PCR results indicating a recrudescence were counted as failures in both analyses. Patients for whom the PCR result was not interpretable or missing or not done were imputed as failures in the ITT population and excluded in the PP population. Patients lost to follow up were excluded from the PP population .The two-sided 95% confidence interval (CI) for the treatment difference in the PCR-corrected and uncorrected ACPRs at day 28 (or day 42) were generated using the normal approximation method . The limThe proportion of aparasitemic patients was computed for each study visit by treatment group. The parasite clearance time was defined as the time elapsed between the first study drug intake and the first of two consecutive evaluations with negative parasitemia. The proportion of afebrile patients was computed at each study visit by treatment group. Early and late treatment failures were summarized by proportion per each treatment group.The cumulative risks of new infections and recrudescences were estimated through survival analysis (Kaplan-Meier). In survival analyses of new infections, patients classified as new infections were considered as having reached the event, whereas patients who did not reach the event  were censored at the relevant time. In survival analyses of recrudescences, patients classified as recrudescence were considered to have reached the event, whereas patients who did not reach the event  were censored at the relevant time."}
+{"text": "An adaptive design and a PK-PD modeling approach were used to determine prospectively the MIC of the new antimalarial cipargamin (KAE609) in adults with uncomplicated Plasmodium falciparum malaria in an open-label, dose-ranging phase 2a study. Vietnamese adults with acute P. falciparum malaria were allocated sequentially to treatment with a single 30-mg (n = 6), 20-mg (n = 5), 10-mg (n = 7), or 15-mg (n = 7) dose of cipargamin. Artemisinin-based combination therapy was given after parasite densities had fallen and then risen as cipargamin levels declined below the MIC but before a return of signs or symptoms. The rates of parasite clearance were dose dependent, with near saturation of the effect being seen at an adult dose of 30 mg. The developed PK-PD model accurately predicted the therapeutic responses in 23/25 patients. The predicted median in vivo MIC was 0.126 ng/ml . Pharmacometric characterization of the relationship between antimalarial drug concentrations and parasite clearance rates following graded subtherapeutic antimalarial drug dosing is safe and provides a rational framework for dose finding in antimalarial drug development. The MIC of an antimalarial drug for a particular infection is the drug level associated with a net parasite multiplication rate of one per asexual cycle. To ensure the cure of malaria, the MIC must be exceeded until all parasites have been eliminated. The development of highly sensitive and accurate PCR quantitation of low-density malaria parasitemia enables the prospective pharmacokinetic-pharmacodynamic (PK-PD) characterization of antimalarial drug effects and now allows identification of the  The emergence of resistance to current antimalarial drugs threatens to reverse recent substantial gains in the control and elimination of malaria. Several factors contribute to the emergence and spread of resistance, notably, uncontrolled use of monotherapies, poor-quality medicines, and systematic underdosing , 2. Therin vivo, the MIC -pharmacodynamic (PD) relationships, although their direct relevance to the treatment of symptomatic malaria remains uncertain , disturbing parasite sodium homeostasis and causing osmotic dysregulation (The spiroindolone cipargamin (formerly KAE609) is a new antimalarial drug with potent activity against um vivax \u20137. Cipargulation , 8, 9. Tgulation , 6, 10.in vivo MIC of cipargamin in acute uncomplicated P. falciparum malaria. This was made possible by the development of sensitive and accurate quantitative PCR (qPCR) measurement of parasite nucleic acids, which permits tracking of parasitemia at densities well below those causing illness and, thus, the characterization of recrudescence without discomfort or risk for the patient , 20 mg (n = 5), and 10 mg (n = 7) (n = 7) was increased to 15 mg. Sixteen patients received early rescue treatment , four because of early treatment failure (ETF) and 12 because of later recrudescence .Twenty-five adult Vietnamese male patients with acute uncomplicated falciparum malaria were enrolled between 15 January 2014 and 12 March 2015. The first three groups received cipargamin in descending dose order: 30 mg ( (n = 7) . One pat1/2) estimates were 4.35 \u00b1 2.21, 3.79 \u00b1 1.22, 1.91 \u00b1 1.64, and 1.47 \u00b1 0.83 h for doses of 10, 15, 20, and 30 mg, respectively. An asymptotic nonlinear model described the relationship between PC1/2 and both the area under the plasma concentration-time curve (AUC) between 0 and 24 h (AUC0\u201324) and the observed maximum plasma concentration (Cmax) well and suggested saturation of the effect at higher doses of cipargamin (Rates of fever and parasite clearance were approximately dose dependent (Table S1). Mean \u00b1 standard deviation (SD) parasite clearance half-life (PCpargamin . DespiteNineteen (76%) patients reported at least one adverse event (AE) (Table S2). Three AEs in the 30-mg group were considered related to the study treatment . There were no serious AEs or AEs leading to study discontinuation. Most laboratory abnormalities did not exceed grade 2 severity; six patients had grade 3 abnormalities. One patient with a history of hypertension who received 30 mg had a rise in blood pressure to 160/100 mm Hg that resolved after treatment with furosemide at 20 mg.50) were \u22121.22  and 2.97 ng/ml , respectively, with the median individual estimated IC50 being 2.38 ng/ml .Population estimates for the slope and 50% inhibitory concentration (ICThe PK properties of cipargamin were well described by a flexible transit-absorption model followed by a one-compartment disposition model . AllowinKgrow) before drug administration are unknown, Kgrow was fixed to 10 per life cycle  (Kkill]). However, individual and population parasite clearance curves showed a clearly biphasic parasitemia decline that could not be adequately explained by gametocytemia. A small, refractory parasite subpopulation was considered the most likely explanation, and its incorporation significantly improved the model fit. The fraction of all asexual parasites that were refractory  was estimated, but interindividual variability was allowed. Their activation  and then removal was also estimated to explain cures. Higher doses and correspondingly higher plasma concentrations were associated with significantly faster maximum parasite killing rates . The final PK-PD model , occurring at a median interval of 7.45 days  after drug administration for patients correctly predicted as having recrudescent infections (n = 12). Time to MIC is a function of dose, as lower doses and, subsequently, lower drug concentrations reach MIC values more rapidly than higher doses, assuming similar PKs and parasite characteristics. Patients correctly predicted to be cured (n = 7) showed a maximum median MIC value of 0.236 ng/ml , defined as the cipargamin concentration when the predicted parasite numbers fell below 10.As individual patient malaria parasite multiplication rates  , 15. ThePD model  showed aPD model . The esthttp://www.who.int/malaria/publications/atoz/9789241549127/en/). Fixed-dose combinations (FDCs) are needed to ensure adherence to both components and prevent the use of monotherapies that could select for resistance to individual drugs. It is essential that the optimum dose of a newly developed compound be chosen before the drug is tied into a FDC. While patients with partial immunity may clear partially treated infections, to ensure a cure with the individual drug component in all patients, plasma concentrations must exceed the MIC for a sufficient time to eliminate all the blood-stage parasites. Identifying the dose and dosing strategy that achieves this requires characterization of two independent variables: the population PK properties of the antimalarial drugs in the target populations and the in vivo drug susceptibility of the infecting malaria parasite populations. The MIC can be estimated accurately in vivo only if there is recrudescence. As the MIC depends both on host factors, notably, immunity, and on the susceptibility of the infecting parasites, it is best assessed in patients from areas with low levels of malaria parasite transmission and, consequently, in patients with low levels of background immunity where multidrug-resistant parasites are prevalent, such as the patients evaluated in the present study. The final dosage in a combination regimen is determined by the predicted duration for which the MIC is exceeded in all relevant patient groups (on the basis of population PK modeling); global variations in malaria parasite susceptibility; practical considerations, such as weight banding; any interactions with the partner drugs; and safety and tolerability .In this study, the PK properties of the novel spiroindolone antimalarial cipargamin were described satisfactorily by a one-compartment disposition model with flexible absorption and were similar to results from previous studies in malaria patients . Exposurial drug . In the Cmax after single doses , were recruited if they could take oral medications reliably and if their axillary temperature or oral, tympanic, or rectal temperature was higher than \u226537.5\u00b0C or \u226538\u00b0C, respectively, at screening or during the previous 24 h. The patients gave fully informed written consent to the study procedures. Exclusion criteria included signs or symptoms of severe malaria, mixed Plasmodium infection, and use of an antimalarial drug within 2 months .Adult Vietnamese patients who were aged 20 to 60 years, who weighed between 40 and 90 kg, and who had acute An adaptive single-dose de-escalation design was employed. The study  was performed at a single center in Vietnam in accordance with the ethical principles of the Declaration of Helsinki. The study protocol was approved by the ethics committee of the Vietnamese Ministry of Health, the Oxford Tropical Research Ethics Committee (OxTREC), and the study center's institutional review board.P. falciparum and P. vivax malaria  until the MIC was identified were planned, and approximately eight patients were to be enrolled per treatment group. The option of dose adjustment on the basis of data arising during the study was available. The 30-mg adult dose has previously been shown to clear parasitemia (as assessed by microscopy) in less than 24 h in both in vivo MIC safely, it is necessary for parasitemia to fall below the level of detection by microscopy and then rise again as drug levels decline below the MIC. The antimalarial plasma concentration at the vertex of this approximate parabola is considered the MIC  conducted in real time on-site and were reported daily to the clinical investigators  mRNA transcripts were also taken and measured at the Queensland Institute of Medical Research (QIMR) Berghofer Medical Research Institute (http://evs.nci.nih.gov/ftp1/CTCAE/CTCAE_4.03_2010-06-14_QuickReference_5x7.pdf).Vital signs were recorded and blood samples were taken every 4 to 6 h until defervescence and two consecutive negative microscopy parasite counts, then once daily until day 8, then on alternate days until day 28, and on days 35 and 42. Serial parasite densities were measured from thick and thin blood film examinations at higher densities and then by a validated real-time qPCR of the tigators . Samplesnstitute . Blood snstitute . The valin vitro susceptibility of the P. falciparum parasites in pretreatment blood samples to cipargamin was measured using a 24-h trophozoite maturation assay  software . PK-PD characteristics were then evaluated with nonlinear mixed-effects modeling in the software NONMEM . The most appropriate model for PK characterization was developed by evaluating different absorption, disposition, and covariate models as well as random-effects components  , 25. IndP. falciparum malaria, as the density of asexual parasites declines, gametocytemia rises, because mature gametocytes are released from sequestration (Parasite densities (determined by microscopy and qPCR) were transformed to total parasite burdens assuming 80 ml whole blood/kg of body weight. In stration . DNA den0\u201324 or Cmax, and PC1/2 was modeled with a nonlinear regression model, which is illustrated as follows for AUC0\u201324: PC1/2 = 0\u201324\u2212k \u00d7 AUC, where aa + (b \u2212 a) \u00d7 e is the asymptotic maximal effect of cipargamin exposure on the PC1/2, b is the longest PC1/2, and k is the rate of decrease in PC1/2.The projected sample size was based on operational rather than statistical considerations . Parasite clearance and fever clearance times were analyzed using the Kaplan-Meier method . The relationship between cipargamin exposure, defined as AUC"}
+{"text": "Plasmodium falciparum malaria. A high number of treatment failures (14/40) was found, and piperaquine resistance in Vietnam was confirmed. A change in the malaria treatment policy for Vietnam is in process.We conducted a study in Binh Phuoc, Vietnam, in 2015 on the therapeutic efficacy of dihydroartemisinin/piperaquine for  Plasmodium falciparum malaria has been a growing concern in the Greater Mekong Subregion of Southeast Asia. High numbers of treatment failures were reported for dihydroartemisinin/piperaquine in Cambodia  copy number  years, and 93% (43/46) were male. Geometric mean parasitemia on day 0 was 17,759 /\u03bcL.Forty-six patients with uncomplicated On day 3, half (23/46) of patients were parasitemic. On day 42, a total of 65%  had an ACPR, and 35% (14/40) had recrudescence; 4 were withdrawn because they became reinfected.K13 propeller domain, the most prevalent being the C580Y mutation in the eastern Greater Mekong Subregion  from our study showed that 90.5% (38/42) were C580Y and 9.5% (4/42) were wild-type. This C580Y prevalence is higher than that reported in a previous study done in Binh Phuoc in 2014, in which 34.5% of samples had the C580Y mutation .Artemisinin resistance is defined as delayed parasite clearance and is associated with mutations in the PfPM2 showed that 25/46 (54.3%) samples had multiple copies of the gene. Of the 42 samples with known K13 types, 22 (52.4%) had both C580Y and PfPM2 amplifications. The remaining 3 had unknown K13 types. All 46 samples had a single copy of Pfmdr1, indicating that all parasites were sensitive to mefloquine  was 35.7 (range 11.1\u201371.0) ng/mL. In 57.1% (24/42) of patients, this concentration was at or above the cutoff value (30 ng/mL) associated with adequate piperaquine exposure  alone did not lead to dihydroartemisinin/piperaquine failures. The association between the presence of molecular markers and recrudescence is confounded by various factors, including parasite load, immunity, and drug levels. Of the 3 patients who had recrudescence and were infected with P. falciparum without PfPM2 amplifications, 2 had inadequate piperaquine levels. Of the 11 patients who had recrudescence and an infection with P. falciparum with PfPM2 amplifications, 7 had adequate piperaquine levels. Low piperaquine blood levels, irrespective of the presence of PfPM2 amplifications, might play a role in some treatment failures. Treatment failures in cases with PfPM2 amplification\u2013positive parasites and adequate piperaquine exposure support the presence of piperaquine resistance in Vietnam. Of the 14 patients who experienced recrudescence, 10 had parasites with the C580Y mutation and K13 mutation has become dominant and that piperaquine resistance is present in Vietnam. A change in the malaria treatment policy to treat with artesunate/mefloquine in Binh Phuoc Province is underway.Our results show that 1"}
+{"text": "Mutations in the cytochrome 1bc complex are causally associated with atovaquone resistance.Atovaquone/proguanil, registered as Malaroneb influences treatment failure and recrudescence based on published information.This systematic review assesses the clinical efficacy of atovaquone/proguanil treatment of uncomplicated malaria and examines the extent to which codon 268 mutation in cytochrome P. falciparum malaria (from 27 studies including between 18 and 253 patients in each case) and 20%\u201326% for Plasmodium vivax malaria (from 1 study including 25 patients). The in vitro P. falciparum phenotype of atovaquone resistance is an IC50 value >28\u2009nM. Case report analyses predict that recrudescence in a patient presenting with parasites carrying cytochrome b codon 268 mutation will occur on average at day 29 , 19  days longer than if the mutation is absent.Data suggest that atovaquone/proguanil treatment efficacy is 89%\u201398% for P. falciparum malaria is effective. Late treatment failure is likely to be associated with a codon 268 mutation in cytochrome b, though recent evidence from animal models suggests these mutations may not spread within the population. However, early treatment failure is likely to arise through alternative mechanisms, requiring further investigation.Evidence suggests atovaquone/proguanil treatment for  Plasmodium spp. is a major cause of mortality worldwide, causing 235\u202f000\u2013639\u202f000 deaths in 2015 and 148\u202f000\u202f000\u2013304\u202f000\u202f000 clinical cases of malaria. Most cases are in endemic countries, although malaria is also one of the most frequent causes of morbidity in travellers returning to non-endemic countries. Atovaquone/proguanil  is a fixed-dose combination often used as a first-line treatment for uncomplicated Plasmodium falciparum infections in non-endemic countries.,P. falciparum sporozoites and then take atovaquone/proguanil to treat pre-symptomatic infections and generate antimalarial immunity . Taken together with the recent expiry of patent protection for Malarone\u00ae, usage of atovaquone/proguanil is likely to rise in the future.Infection with 1bc complex of malaria parasites  (in particular leading to Y268S/C/N) cause atovaquone resistance both in vitro and in vivo.Plasmodium berghei CYTb are lethal during transmission of the parasite in the mosquito vector.P. falciparum in the field. Cycloguanil resistance in parasites is conferred by multiple mutations in DHFR. Polymorphisms in host CYP2C19 also affect proguanil metabolism and can lower cycloguanil concentrations.Several mechanisms can potentially influence the efficacy of atovaquone/proguanil for treatment. Mutations in PfCYTb. In this systematic review, we examine all original in vivo data where atovaquone/proguanil was used exclusively to treat malaria and relate findings on risk of recrudescence to mutations in PfCYTb and available results from in vitro assays. We also estimate clinical efficacy of atovaquone/proguanil treatment of uncomplicated malaria. Results may impact on existing guidelines for the treatment of uncomplicated malaria.Reports of frequencies of treatment failure associated with atovaquone/proguanil vary, although the risk of failure has not been systematically examined particularly with respect to mutations at codon 268 of This systematic review was registered at PROSPERO (number CRD42015020757) on 25 February 2015 and updated on 13 October 2017.JAC Online).PubMed (1966\u2013present) and ScienceDirect (1823\u2013present) were interrogated on the 19 May 2015 with the following search strategy {[(Atovaquone AND Proguanil) OR ] AND }. Records were assessed for eligibility using title, or title and abstract. Eligible records were screened for duplicates and full-text obtained for the remaining records that were then reassessed for eligibility. Data were extracted from these articles by two reviewers and tabulated. Inclusion and exclusion criteria and extracted data variables are summarized in the Two reviewers assessed group study eligibility and the risk of bias in the trials using the modified Cochrane risk of bias tool.For all group studies, the total numbers of patients enrolled into each treatment arm, those followed up to 28\u2009days and those with treatment failure or recrudescence were extracted and combined to obtain the proportion of patients for whom treatment had been successful in the ITT and PP populations. For randomized controlled trials (RCT), this information was also extracted for the comparator antimalarial arm(s) to allow meta-analyses .2 test and additional sensitivity analyses undertaken  minimum days to recrudescence , and (ii) parasitaemia at recrudescence with presence of mutation in th cases .in vitro/ex vivo data. The case reports and group studies were included in the meta-analysis.A total of 282 records were returned using PubMed and 966 using ScienceDirect . A total of 1640 patients were successfully treated up to 28\u2009days, 83.7% of the 1960 original patients and 96.8% of the 1695 treated and followed-up patients. The one P. vivax study began with 25 patients, of whom 19 were treated and followed up to 28\u2009days (76%). Five patients were successfully treated up to 28\u2009days, 20% of the original 25 patients, and 26.3% of the treated and followed up patients. The one study of P. ovale spp. and P. malariae began with six patients and all were successfully treated up to 28\u2009days.The 29 group studies Table  consisteOf note, only 14 of the studies were RCT designed to test the efficacy of atovaquone/proguanil or used atovaquone/proguanil as a control treatment and participants of these made up only 55% of the total participants included here. Most of the studies from which these data were gathered, including the RCT, were of low methodological quality, being small and having between 18 and 253 participants receiving atovaquone/proguanil. Risk of bias during selection was determined to be unclear in 10 of 14 RCT group studies, as methods for randomization and concealment of allocation were unclear  and 100% (in three patients each) for P. malariae and P. ovale spp. malaria.High-quality data for the efficacy of atovaquone/proguanil are scarce, but provide estimates of treatment success in RCT group studies of between 89% and 98% for P\u2009=\u20090.83) in treatment success between the use of atovaquone/proguanil and ACT  and statistical significance was maintained for the majority of scenarios during sensitivity analysis (Table P\u2009<\u20090.00001) and the sensitivity analysis was predominantly consistent with this outcome (Table pfcrt and pfmdr1 conferring resistance to chloroquine and amodiaquine in the regions of study.P\u2009=\u20090.001) emerged when analysing atovaquone/proguanil versus sulfadoxine/pyrimethamine  were chloroquine (two), amodiaquine (two), sulfadoxine/pyrimethamine (three), chloroquine/sulfadoxine/pyrimethamine (one), quinine (one), quinine/tetracycline (one), halofantrine (two), mefloquine (one), and the artemisinin-based combination therapies (ACT), artemether/lumefantrine (two), artesunate/mefloquine (one), artesunate/amodiaquine (one) and dihydroartemisinin/piperaquine/trimethoprim/primaquine (one). Nine of the 14 RCT presented here were analysed in a previous Cochrane Library systematic review from 2005.T Figure . Given tT Figure . Analysiis Table . As prevme Table . This cae Figure . This cain vitro/ex vivo clinical isolates exposed to atovaquone were available in 15 papers  IC50 value of 5.7\u2009nM (1.7\u20131216).in vitro, with median IC50 values between 4.7 and 5\u2009nM. Isolates of unknown genotype ranged in IC50 values from low nanomolar to low micromolar. The 38 P. vivax isolates had a pooled mean IC50 value of 29.4\u2009nM.Eligible data on rs Table . The amoP. falciparum infection and there was one case each of P. malariae, P. ovale spp. and P. vivax infection. Variables have been summarized, with means, standard deviations (SD), medians and IQR for continuous or count data and proportions for categorical or binary data types . In a subset of parasite isolates it was possible to define if there had been a change in codon 268 following treatment. A raw data plot of the minimum number of days to recrudescence versus this dataset suggested distributions may differ by codon 268 change  days shorter in duration if the mutation is absent. Note that although a slight departure from normality for the standardized residuals (P\u2009=\u20090.02) was calculated, we opted for model simplicity rather than introducing another quadratic term.Figure l Figure a and b. PfCYTb, using an interaction model , the overall efficacy of atovaquone/proguanil expressed as a weighted average based on study population sizes and heterogeneity is 89% and 83% in ITT analyses of RCT and observational studies, respectively, and is 98% and 99% in PP analyses. This is a reassuringly acceptable level of efficacy and to date there are no indications of treatment failures becoming associated with particular geographical areas that would preclude atovaquone/proguanil use to treat travellers or prevent infections from such areas. Furthermore, meta-analysis suggests that atovaquone/proguanil treatment success is equivalent to the use of ACT and amino alcohols and better than 4-aminoquinolines and sulfadoxine/pyrimethamine, although caution is required in some cases due to the grouping of different antimalarials within a class. This extends findings from a prior meta-analysis that concluded that atovaquone/proguanil is more effective than chloroquine, amodiaquine and mefloquine.in vitro phenotypic assays for atovaquone susceptibility and its relationship to target genotype suggest that WT amino acid (Y268) is uniformly associated with susceptibility. The threshold for defining susceptibility is an IC50 value \u226428\u2009nM, with most isolates in different studies having IC50 values <10\u2009nM. Although the aggregated IC50 values for P. vivax were 29\u2009nM, it is unlikely that this slightly higher value compared with P. falciparum susceptibility contributed to the higher treatment failure rates as these are most likely due to relapse because of the non-susceptibility of hypnozoite stages found in the liver to atovaquone/proguanil.The PfCYTb is strongly associated with a late recrudescing infection  in  Figures  and 5 whP. berghei CYTb are invariably lethal to the parasite during transmission in the mosquito vector.PfCYTb mutations may not be able to spread within a population. If true, this would preclude the requirement to monitor for these mutations in endemic areas. The available data are in general agreement with this, as codon 268 mutations are very rarely observed in parasites from patients that suffer later recrudescence, prior to drug pressure .A recent report has demonstrated that mutations in re Table  and no gPfDHFR mutation S108N, N51I, C59R  were caused by parasites carrying genotypes associated with resistance to cycloguanil. Therefore, atovaquone/proguanil treatment failures from day 7 onwards are most likely to be caused by parasites that are already resistant to cycloguanil.While not considered in detail, it is worth noting that there are 17 case reports that provide molecular markers for cycloguanil resistance, the triple 9R Table . Only 4 PfCYTb was available, it was of the 268C/S mutation. In four of six patients with second recrudescences, the time to recrudescence was \u226520\u2009days and all four genotypes bore mutant variants at position 268. These observations suggest that the proguanil component of atovaquone/proguanil has sufficient antimalarial efficacy to suppress parasitaemias for 2\u20133\u2009weeks and that the dynamics of late treatment failure are consistent with absence of atovaquone efficacy. These cases were incorporated into a secondary analysis of the case reports. Findings with regard to the relationship between pretreatment parasitaemia and minimum days until recrudescence in the absence or presence of a mutation in PfCYTb are consistent with those presented in Table After our database search was closed, an additional series of case reports that was not picked up was identified independently.P. falciparum infections.Overall, atovaquone/proguanil therapy is comparable in efficacy to ACT used in treating uncomplicated malaria. Detailed genotype\u2013phenotype analysis in this systematic review has illustrated several new findings. There are differences between early and late treatment failures because mutations in the target conferring resistance to atovaquone are identified most commonly in late and not early treatment failures. The mechanism of early treatment failure after atovaquone/proguanil treatment needs further investigation. Recent evidence is also reassuring that spread of the 268 mutations conferring atovaquone resistance may be limited by poor transmissibility in the insect stages of Supplementary DataClick here for additional data file."}
+{"text": "Plasmodium falciparum parasitemia within 6 weeks of treatment with no intervening travel to countries where malaria is endemic. Parasite isolates, all of African origin, harbored variants at some candidate resistance loci. No evidence of pfk13-mediated artemisinin resistance was found. Vigilance for signs of unsatisfactory antimalarial efficacy among imported cases of malaria is recommended.We present case histories of four patients treated with artemether-lumefantrine for falciparum malaria in UK hospitals in 2015 to 2016. Each subsequently presented with recurrent symptoms and  Plasmodium falciparum malaria has benefited from the availability of artemisinin-based combination therapies (ACT)  . In Asiaefficacy , 3. Howen Africa \u20134, whered pfubp1 \u20139.P. falciparum malaria in AL-treated patients occurring 17 to 43 days after the original episode were reported to the Malaria Reference Laboratory (MRL) (Artemether-lumefantrine (AL) is the recommended treatment for imported cases of falciparum malaria in the UK  and is try (MRL)  by two UThis work was undertaken under the statutory remit of the MRL set out by the UK Department of Health, which explicitly requires surveillance for evidence of emerging drug resistance among UK malaria cases, as previously described . All tesP. falciparum parasitemia following travel in Angola. He was treated with AL in the hospital, and all doses were observed. The first dose was consumed without food, and the patient vomited once 3 h later. All but one of the subsequent AL doses were consumed with food, and the patient was confirmed negative for parasitemia prior to discharge. Symptoms recurred more than 1 month later, and the patient presented with 0.02% P. falciparum parasitemia, confirmed by PCR and microscopy at the MRL. The patient responded well to administration of 4 tablets of 100/250 mg atovaquone/proguanil daily for 3 days. The pfmdr1 Tyr184Phe and pfap2mu Ser160Asn variants observed have both been associated with recurrent parasitemia in AL-treated African patients , UK, in 2015 with 1.4% P. falciparum hyperparasitemia estimated at more than 30% in early 2016 following a visit to Uganda. She responded well to initial treatment with intravenous (i.v.) artesunate. Parasitemia of 0.07% was recorded 60 h after admission, and the patient was discharged 3 days later, with a recorded parasitemia level of 0.0001%, on a full course of oral AL. Full compliance was reported under supervision of staff members at her place of learning. Symptoms recurred 4 weeks later. The patient was readmitted to HTD with 2.7% P. falciparum parasitemia, received 2 days i.v. artesunate as an inpatient until the parasitemia level fell below 0.0001%, and was discharged on a full course of AL plus 7 days of doxycycline (200 mg daily). This recurrent isolate was established in in vitro culture at the London School of Hygiene & Tropical Medicine (LSHTM), and standard 48-h 50% effective concentration (EC50) estimates were derived for chloroquine (102.6 nM), dihydroartemisinin (2.8 nM), and lumefantrine (38.7 nM) as previously described , with acute escribed , with th pfap2mu . HoweverP. falciparum parasitemia in early 2016, after travel to Liberia. He was treated with AL and was eventually discharged but returned with recurrent symptoms and 0.02% parasitemia 17 days later. He was successfully treated with atovaquone-proguanil. Genotyping identified mixed genotypes at both pfcrt and pfap2mu loci in the isolate from the primary episode but not in that from the recurrent episode  met the specified International Pharmacopeia tolerance limits  and doxycycline (200 mg daily). Data from the two episodes appeared identical at the parasite loci evaluated. Novel mutations at codons 226 and 326 of pfap2mu and codon 1500 of pfubp1 are of unknown significance, but, as this is clearly a recrudescent parasite genotype, both variants should be further evaluated in future studies.A female under 21 years of age who presented as febrile to the HTD in early 2016 following travel to Uganda was diagnosed with 0.08% pfk13 locus and thus did not exhibit reduced artemisinin susceptibility as seen in the Greater Mekong Region (\u2013pfk13-independent mechanisms for posttreatment survival in vivo. This suggests that, in countries where malaria is not endemic and in which imported malaria cases from Africa are seen, vigilance is required to protect the efficacy of our first-line ACT. Studies in countries where malaria is endemic are required to further validate potential genetic markers for monitoring in African parasite populations.Treatment failure cannot be unequivocally ascribed to parasite resistance in these four patients, although three harbored parasites with variant alleles of loci previously linked to reduced susceptibility to artemisinin or lumefantrine . Our fing Region \u20134. The pP. falciparum of African origin, but the cases presented here suggest that this favorable situation may deteriorate in the future.Artemisinins are very quickly metabolized and cleared from circulation, such that only a single full parasite life cycle is exposed to the highly potent action of these compounds under conditions of a 3-day regimen. The current practice of 3 days of treatment may therefore be insufficient to guarantee clearance of parasite genotypes under ACT pressure for a decade in many parts of Africa . The use"}
+{"text": "Plasmodium falciparum malaria transmission requires more safety data.The World Health Organization recommendation on the use of a single low dose of primaquine (SLD-PQ) to reduce We conducted an open-label, nonrandomized, dose-adjustment trial of the safety of 3 single doses of primaquine in glucose-6-phosphate dehydrogenase (G6PD)-deficient adult males in Mali, followed by an assessment of safety in G6PD-deficient boys aged 11\u201317 years and those aged 5\u201310 years, including G6PD-normal control groups. The primary outcome was the greatest within-person percentage drop in hemoglobin concentration within 10 days after treatment.Fifty-one participants were included in analysis. G6PD-deficient adult males received 0.40, 0.45, or 0.50 mg/kg of SLD-PQ. G6PD-deficient boys received 0.40 mg/kg of SLD-PQ. There was no evidence of symptomatic hemolysis, and adverse events considered related to study drug (n = 4) were mild. The mean largest within-person percentage change in hemoglobin level between days 0 and 10 was \u22129.7%  in G6PD-deficient adults receiving 0.50 mg/kg of SLD-PQ, \u221211.5%  in G6PD-deficient boys aged 11\u201317 years, and \u22129.61%  in G6PD-deficient boys aged 5\u201310 years. The lowest hemoglobin concentration at any point during the study was 92 g/L.SLD-PQ doses between 0.40 and 0.50 mg/kg were well tolerated in G6PD-deficient males in Mali.NCT02535767. Plasmodium falciparum malaria transmission.This phase 1, open-label, nonrandomized, dose-adjustment trial showed that single doses of primaquine between 0.40 and 0.50 mg/kg were well tolerated in G6PD-deficient and G6PD-normal males in Mali, supporting the World Health Organization recommendation on the use of a single low of dose primaquine to reduce  Plasmodium falciparum malaria from humans to anopheline mosquitoes.The success of malaria control and elimination programs supports global aspirations to eradicate human malaria in the coming decades . To achiA\u2212 variant whilst in Asia, the variants are heterogeneous, with the Mediterranean variant being the most severe [Primaquine (PQ) use must be informed by safety considerations, as this drug causes dose-dependent hemolysis in individuals with enzymatic glucose-6-phosphate dehydrogenase (G6PD) deficiency, the most commonly inherited enzyme deficiency worldwide . The saft severe .P. falciparum gametocytes, the World Health Organization (WHO) recommends that a 0.25 mg/kg single low dose of PQ (SLD-PQ) be used without G6PD testing, in conjunction with standard artemisinin-based combination therapy, in areas approaching malaria elimination and/or facing drug resistance [For treatment of sistance , 5. ThisImplementation of the WHO recommendation to use 0.25 mg/kg of SLD-PQ requires that each individual be weighed, PQ tablets be crushed and dissolved in water, and the corresponding amount of PQ solution for the 0.25 mg/kg dose be carefully measured. This process is labor intensive, introduces the possibility for dosing errors, and can be avoided by the establishment of age-based dosing bands. To develop age-based dosing bands, the therapeutic dose range of SLD-PQ, spanning the lowest efficacious dose to the highest safe dose in vulnerable individuals, must be established. This provides weight-based dose bands, which may then be converted to age-based dosing bands by using age-for-weight data . This stThis study was an open-label, nonrandomized, dose-adjustment followed by age de-escalation trial of the safety of single-dose PQ in G6PD-d and G6PD-normal (G6PD-n) males in Mali without microscopically detected malaria parasite infection. Participants were enrolled sequentially.In part 1, we investigated 3 single doses of PQ in adults: the first was a prespecified dose of 0.40 mg/kg in 7 individuals confirmed to be G6PD deficient according to a Carestart G6PD rapid diagnostic test ; this dose was demonstrated to be safe in a previous study . The remAfter enrollment of 3 dose groups, the DSMB reviewed study data and determined the highest dose of PQ tolerated in part 1, which was then administered to a control group of 7 G6PD-n individuals matched within 8 years of age to those enrolled in the group with the highest tolerable dose. Follow-up for the control group proceeded in the same manner as in the intervention groups. G6PD status was validated using semiquantitative spectrophotometry testing .In part 2, we investigated the safety of single-dose PQ in G6PD-d boys at a dose 0.1 mg/kg lower than the highest safe dose found in adults to be conservative. For the first age group (11\u201317 years inclusive), we enrolled 7 G6PD-d boys and 7 G6PD-n boys (the control group). Following DSMB review of data by use of criteria described for part 1, if PQ was safe in this age group, a second age group (5\u201310 years inclusive) comprising 7 G6PD-d and 7 G6PD-n boys would be enrolled.Participants were recruited from the town of Ou\u00e9less\u00e9bougou and its surrounding villages in Mali by the Malaria Research and Training Centre of the University of Bamako, Mali. Prior to recruitment, the study team met with village leaders to arrange for information sessions on the trial for prospective participants in the community, seeking informed consent from those interested in participation. All individuals who provided consent were tested for G6PD deficiency, using the Carestart G6PD RDT test, and had their name, address, and phone number recorded. Those for whom tests revealed G6PD deficiency scheduled a date to visit the study clinic in Ou\u00e9less\u00e9bougou for further screening, while those with a normal G6PD status revealed by testing were told they might be contacted at a later date for further screening.Eligibility criteria included males, to reduce the possibility of incorrect classification of G6PD status by qualitative G6PD tests, which is known to occur more commonly in heterozygous females ; provisiParticipants who reported any of the following conditions were excluded: known positive results of human immunodeficiency virus (HIV) and/or hepatitis B virus tests, allergy to PQ, current receipt of treatment for tuberculosis or HIV infection or of any drugs that have hemolytic potential in G6PD-d individuals , use of antimalarial drugs \u22642 weeks before contact with the study team, blood transfusion of >500 mL within the last 3 months, high alcohol intake  within the past 6 months, and/or reported use of illicit drugs  \u22646 months before study entry. All eligible participants could be enrolled in the study one time, and any enrolled participant who vomited \u22641 hour after ingesting PQ was excluded from analysis.The study was approved by the Ethics Committee of the Faculty of Medicine, Pharmacy, and Dentistry, University of Science, Techniques and Technologies of Bamako , and by the Committee on Human Research at the University of California, San Francisco . It also underwent human subjects review at the US Centers for Disease Control and Prevention and approved as non-engagement in human subjects research. The study was monitored by an external clinical trials monitor based in Bamako. Participants were compensated for time and travel. The trial was registered at Clinicaltrials.gov (registration no. NCT02535767).After collection of day 0 samples, each participant received an oral dose of PQ according to his group assignment, after a fatty snack (biscuits) to minimize gastrointestinal symptoms. The study pharmacist prepared the dose by crushing a 15-mg tablet of PQ  in 15 mL of drinking water and administered the dose to the nearest 0.1 mL under direct observation .Participants were evaluated at the study clinic at hours 1, 2, 4, 6, and 8 and days 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 14, and 28 following treatment. On all follow-up days, starting approximately 24 hours following PQ administration, a clinical examination was conducted, and blood samples were collected for measurement of the Hb concentration and for preparation of slides to determine malaria parasite and reticulocyte counts . To assess for oxidative stress, a noninvasive methemoglobin measurement was performed at all follow-up time points, using a Masimo Rad-57 pulse oximeter . Owing to technical issues, the pulse oximeter was not functional until the start of enrollment of the second dose group. Urine samples were also collected on each day of follow-up, to assess for hemolysis by using a urine color chart developed by Hillmen and Hall . If a \u22653AEs were assessed actively during all follow-up visits and passively through the availability of study clinicians 24 hours/day, 7 days/week. We recorded the duration and severity of AEs ; relationship to the study drug, according to the study physician(s) ; actions taken; and outcome. AEs were further assessed as serious or not serious, according to the treating physician. SAEs were prespecified in the study protocol and adapted by the DSMB prior to trial initiation and were defined as any of the following: development of clinical signs or symptoms of distress, including any requirement for hemodialysis; laboratory values indicating severe hemolytic anemia, including a drop in the Hb level of >40% from baseline levels; a need for blood transfusion; the development of hemoglobinuria, identified by black coloration in the urine associated with a rise in creatinine level; and any SAE, defined as an untoward medical occurrence or effect that, at any dose, results in death, is life threatening, requires hospitalization, or results in persistent or significant disability or incapacity that is clearly, probably, or possibly related to the study drug.A variant of both SNPs was known to be common among G6PD-d individuals in Africa at the time of study design [G6PD testing was conducted using 3 methods: qualitative phenotypic testing, semiquantitative testing, and genotyping. Qualitative phenotypic rapid testing using the Carestart G6PD test was conducted as a screening test throughout the study. Additionally, the field team conducted additional qualitative OSMMR-D G-6-PD tests in part 1 after enrollment of the first 3 individuals, excluding individuals with discordant results between the 2 qualitative tests . Semiquay design . G6PD-d y design .Cytochrome P450 2D6 SNPs were identified for adults, and polymerase chain reaction (PCR) methods were used to detect malaria parasites in all participants, using dried blood spots collected on filter paper prior to PQ administration .CYP2D6) SNPs are associated with hemolysis in G6PD-d individuals (The primary safety outcome was the largest within-person percentage decrease in Hb concentration between baseline levels (day 0) and day 10 following PQ administration. Prespecified secondary outcomes consisted of the incidence of AEs, graded by severity and relation to the study drug; the occurrence of acute hemolytic anemia at each PQ dose, including absolute and fractional changes in Hb concentration on day 7 and day 28 as compared to baseline; urine color assessment; reticulocyte count; total and direct bilirubin levels if a within-person Hb concentration drop of \u226530% from baseline levels was seen in an individual between day 0 and 7 (inclusive); methemoglobin concentration and development of physical signs or symptoms of hemolytic anemia; comparison of the change in the Hb concentration, the frequency and severity of AEs, and the occurrence of markers of acute hemolytic anemia between G6PD-d and G6PD-n participants; and exploratory studies among adults on whether cytochrome P450 2D6 , we used a regression model that adjusted for baseline Hb concentration, to determine whether there was a significant difference in the outcomes between G6PD-d and G6PD-n participants after treatment.AEs were summarized descriptively for each dosing group, using frequencies of observed events, severity, and proportion of participants experiencing at least 1 event.All analyses were conducted using Stata v12 .In part 1, 28 adult males were enrolled between 13 August and 19 December 2015 . The dosIn part 2, 28 boys were enrolled between 12 May 2016 and 10 January 2017 . Two boyP = .320; P = .199). Similarly, among boys aged 11\u201317 years , the mean largest within-person percentage change in Hb concentration among G6PD-d participants was \u221211.5% , whereas their G6PD-n counterparts experienced a change of \u22126.89% . There was no significant difference between the mean largest within-person percentage change in Hb concentration among G6PD-d adult men  and that among G6PD-d boys aged 5\u201317 years .Among men , the mean largest within-person percentage change in Hb concentration among G6PD-d individuals receiving the highest tolerable dose of PQ (0.50 mg/kg) was \u22129.7% , whereas their G6PD-n counterparts experienced a change of \u22127.89% ; this man had a Hb concentration of 147 g/L at baseline and 113 g/L 9 days following PQ administration, was positive for P. falciparum malaria at baseline by PCR, was positive by blood smear on days 1 and 3\u20137, and received a symptomatic malaria diagnosis on days 6\u20139 following treatment with PQ. At day 28, this participant\u2019s Hb concentration was 135 g/L. For all participants enrolled, the lowest absolute Hb concentration was 92 g/L, observed in a 6-year-old G6PD-d boy; he had a Hb concentration of 112 g/L at baseline, the nadir Hb concentration on day 10, no malaria parasites detected by blood smear, and a Hb concentration of 106 g/L by day 28.The mean largest percentage change in Hb concentration was a 16.8% decrease between day 0 and day 10, which occurred in the group of men who received 0.40 mg/kg of SLD-PQ. This group included the individual with the largest within-person percentage change in Hb concentration  had asymptomatic malaria parasitemia. During follow-up, 14 participants tested positive for malaria parasites on blood smear, with 4 being symptomatic . There wThis study found that a single dose of PQ as high as 0.50 mg/kg was well tolerated among G6PD-d adults and that a 0.40-mg/kg dose was well tolerated in G6PD-d adults and children. Our study is the first to systematically investigate the safety of SLD-PQ at doses of \u22650.40 mg/kg in G6PD-d individuals without malaria, addressing the evidence gap identified by the WHO Expert Review Group that issued the recommendation to use SLD-PQ. Despite consistent measures of reduced Hb concentrations in G6PD-d participants exposed to SLD-PQ, compared with G6PD-n participants, the reductions in Hb levels were small and not clinically relevant.This study also provides data to define the therapeutic dose range of SLD-PQ in West African settings, which will support field implementation of the WHO recommendation, currently based on patient weight and challenged by a requirement to dissolve tablets and carefully measure the correct amount of solution to give to the patient , 7. Age-Other studies on SLD-PQ to date have focused on the safety of the WHO-recommended dose of 0.25 mg/kg of SLD-PQ, which was demonstrated to be safe in Senegal, when given with artemether-lumefantrine, dihydroartemisinin-piperaquine, or artesunate-amodiaquine to 54 phenotypically G6PD-d patients with malaria ; in TanzAs malaria parasite infection can reduce hemoglobin concentration, this study was intended to examine the isolated effect of PQ on individuals without malaria. However, many individuals enrolled had asymptomatic malaria (40% during screening and 26% during follow-up), of whom 4 (8%) presented with symptomatic malaria on follow-up. While it is not possible to determine whether Hb level drops in these individuals were due to malaria parasite infection or to SLD-PQ, the results suggest that SLD-PQ is well tolerated in G6PD-d individuals with and those without malaria.The main limitations of this study include a small sample size, the exclusion of women and, as an ethical precaution taken to ensure the safety of study participants, and vulnerable individuals who were already anemic on enrollment. As SLD-PQ is now being rolled out in various countries in Africa, further assessment of its safety should be performed using pharmacovigilance mechanisms, with an emphasis on vulnerable individuals who are anemic, have HIV infection and/or tuberculosis, and/or are unaware of their pregnancy when they take PQ.In summary, these results provide evidence for extending the upper bound of the therapeutic dose range of SLD-PQ to 0.40 mg/kg in sub-Saharan Africa. This evidence may be used to facilitate the age-based dosing of SLD-PQ in sub-Saharan Africa.The Journal of Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at Supplementary MaterialClick here for additional data file."}
+{"text": "Plasmodium falciparum (k13), has outpaced containment efforts in South East Asia. For national malaria control programmes in the region, it is important to establish a surveillance system which includes monitoring for k13 polymorphisms associated with the clinical phenotype.Artemisinin resistance, linked to polymorphisms in the Kelch gene on chromosome 13 of P. falciparum treated with artesunate monotherapy followed by 3-day ACT in an isolated area on the Myanmar\u2013Thai border with relatively low artemisinin drug pressure. Molecular testing for k13 mutations was performed on dry blood spots collected on admission.Between February and December 2013, parasite clearance was assessed in 35 patients with uncomplicated k13 mutations in these patients was 41.7%, and only 5 alleles were detected: C580Y, I205T, M476I, R561H, and F446I. Of these, F446I was the most common, and was associated with a longer parasite clearance half-life (median) 4.1 (min\u2013max 2.3\u20136.7) hours compared to 2.5 (min\u2013max 1.6\u20138.7) in wildtype (p\u00a0=\u00a00\u00b701). The prevalence of k13 mutant parasites was much lower than the proportion of k13 mutants detected 200\u00a0km south in a much less remote setting where the prevalence of k13 mutants was 84% with 15 distinct alleles in 2013 of which C580Y predominated.The proportion of k13 mutations as well as allele diversity varies considerably across short distances, presumably because of historical patterns of artemisinin use and population movements.This study provides evidence of artemisinin resistance in a remote part of eastern Myanmar. The prevalence of The online version of this article (10.1186/s12936-017-2128-x) contains supplementary material, which is available to authorized users. Plasmodium falciparum, characterized by delayed parasite clearance in patients treated with artesunate, emerged in Western Cambodia in 2007 [k13) were found to be strongly associated with resistance to artemisinin derivatives [k13 mutations and the clinical resistance phenotype (delayed parasite clearance) has been demonstrated by a number of clinical [k13 mutations have been found, the distribution of different alleles has been variable. For example, the C580Y polymorphism is the most common mutation in Cambodia [P. falciparum are not only emerging \u201cde novo\u201d in different locations but also spreading contiguously.Resistance to the artemisinin derivatives in  in 2007 \u20133. This  in 2007  found to in 2007  and asso in 2007 . In 2014ivatives . The linclinical , in vitrclinical  and tranclinical . In the Cambodia , 16 and Cambodia , 17, 18.Cambodia . The C58Cambodia . IsolateCambodia . The C58k13 mutations have now spread to the China\u2013Myanmar [In 2011, the World Health Organization (WHO) proposed a strategy of containment of artemisinin resistant malaria in the region , but thi\u2013Myanmar  and Indi\u2013Myanmar , 23, 24.\u2013Myanmar .k13 alleles under strong selective pressure confer (or are associated with) slow parasite clearance after artemisinin treatment leaving a larger number of surviving (artemisinin resistant) parasites exposed to the partner drugs [More importantly, er drugs . The loser drugs , 27 and er drugs  have demk13 of parasites from a secluded relatively inaccessible region with limited access to ACT  and k13 mutations using the same protocol as a larger multicentre trial [This was an open label non-randomized trial designed to evaluate the association between the efficacy of artemisinin against\u00a0re trial . The stu\u00b0C) or history of fever within the last 24\u00a0h and who were between the ages of 6\u00a0months to 65\u00a0years were screened for malaria parasites using microscopy of thick and thin Giemsa-stained blood smears. Patients with uncomplicated P. falciparum malaria and asexual parasitaemia above or equal to 10,000/\u00b5L were enrolled. Pregnant women and patients with severe malaria [\u00ae 903 (W-903) filter paper for parasite genotyping. Haemoglobin colour scale Copack\u00ae was used for haemoglobin estimation. A clinical examination was also performed, and vital signs were recorded as well as demographic data.Patients who presented at Day Bu Noh clinic with fever  (4\u00a0mg/kg artesunate day 3\u20135 and mefloquine 15\u00a0mg/kg\u00a0day 4 & 10\u00a0mg/kg day 5) orartesunate-mefloquine  at a dose of 2.5\u00a0mg/kg dihydroartemisinin and 20\u00a0mg/kg piperaquine day 3\u20135 was administered.dihydroartemisinin-piperaquine , 4\u00a0mg/kg/day for 3\u00a0days. After 3\u00a0days of artesunate monotherapy, eitherAll treatments were supervised. Peripheral parasitaemia and temperature were measured at 0, 4, 6, 8, 12 and every 6\u00a0h thereafter until two consecutive negative slides were obtained. No further follow up was undertaken after the patient finished the full course of treatment and had a negative malaria slide on two occasions. Other clinical data and vital signs were recorded daily.Kelch genotyping was performed at the malaria molecular laboratory of Mahidol Oxford Tropical Medicine Research Unit using the method reported previously , 28 see.Patient data were recorded on individual Case Report Forms (CRFs) and later entered into a Microsoft Access database and analysed using STATA  and Graphpad prism (version 5). Normally distributed data were compared by Student\u2019s t test and non-normally distributed data by the Wilcoxon rank-sum (Mann\u2013Whitney) test. Parasite clearance half-lives were calculated using the World Wide Antimalarial Resistance Network (WWARN) Parasite Clearance Estimator , 30.Written informed consent was obtained after the screening process and the study was explained in detail through the use of a local interpreter. Consent was obtained from a parent or guardian for participants below 18\u00a0years.Ethical approval was given by the Oxford Tropical Research Ethics Committee (OXTREC 06-11). Verbal approval was given locally by existing health authorities KDHW  in the absence of a local ethics committee. However, a community advisory board composed of members of the local population also approved this study .P. falciparum malaria, 36 that met the inclusion criteria and gave consent were recruited into the study (including 3 mixed infections). The main reason for exclusion was low parasitaemia. The baseline demographic and clinical data of the patients are summarized in Table\u00a0Between February and December 2013, a total of 1736 febrile patient presented at the clinic, and 27.4% (475) of them were microscopically confirmed to have malaria  had fever on admission. By 48\u00a0h 30/35 (85.7%: 95% CI 74.1\u201397.3) of the patients were afebrile and all had cleared their fever by 72\u00a0h. After 3\u00a0days of artesunate, 13  patients remained parasitaemic but all were negative by microscopy by day 4. Three (8.6%) patients had gametocytes on admission that were cleared in 2 patients by day 6.k13 sequence. Five different non-synonymous polymorphisms were detected in 46.9%  of samples including 4 in the propeller region and 1 in the \u2018stem\u2019 region of the kelch protein. No sample had more than one mutation. Among these, the most common allele was F446I (31.3%)  parasite isolates were successfully genotyped for k13  compared to those infected with isolates carrying a mutation in the propeller region  (p\u00a0=\u00a00.04). Again, the patients infected with the most common allele, F446I cleared their parasites more slowly  (p\u00a0=\u00a00.04) compared to those infected with wild type K-13  of patients infected with wild type r region .1, 3.3\u00a0hk13 .5, 1.3\u00a0hThe 6-day treatment (3-day artesunate monotherapy follow by 3-day ACT) was well tolerated. None of the patients developed severe malaria. There were no serious adverse events.P. falciparum malaria, a pattern that is strongly associated with mutations in the k13 gene of P. falciparum and accompanying partner drug resistance. Until now, 108 non-synonymous polymorphisms of k13 gene have been found in all malarious regions of the world. Most of the mutations in the propeller region of the gene are associated with the slow parasite clearance phenotype, and in Asia show evidence of recent selection [k13 mutations has increased significantly, and among them the C580Y allele has out-competed others in recent years signifying that a \u201chard\u201d selective sweep has replaced an initially \u201csoft\u201d one [Throughout South East Asia parasite clearance rates are decreasing among patients with uncomplicated election , 19. HisThe main limitation of this study is the small sample size, which is related to both the remoteness of the study site, logistical difficulties related to working in this remote area, and the relatively high parasitaemia threshold for inclusion . The parasitaemia threshold was chosen in order to obtain reliable parasite clearance curves.P. falciparum with significant differences in allele frequencies across small distances. Even though the prevailing alleles may be different it is clear that artemisinin resistance has emerged even in geographically secluded areas and ultimately may be unstoppable. In the absence of alternative treatments ready to replace the failing ACT, elimination remains the best practical strategy to slow down or halt progression of artemisinin resistance in this region.However, this small study highlights the complexity of the spatial dynamics of artemisinin resistance in Additional file 1. Genetic markers of artemisinin resistance.Additional file 2. Parasite clearance curves for individual patients."}
+{"text": "Plasmodium falciparum and Plasmodium vivax infections in Papua, Indonesia, since March 2006. The efficacy of DHP was reassessed to determine whether there had been any decline following almost a decade of its extensive use. An open-label drug efficacy study of DHP for uncomplicated P. falciparum and P. vivax malaria was carried out between March 2015 and April 2016 in Timika, Papua, Indonesia. Patients with uncomplicated malaria were administered supervised DHP tablets once daily for 3 days. Clinical and laboratory data were collected daily until parasite clearance and then weekly for 6 weeks. Molecular analysis was undertaken for all patients with recurrent parasitemia. A total of 129 study patients were enrolled in the study. At day 42, the polymerase chain reaction-adjusted efficacy was 97.7%  in the 61 patients with P. falciparum malaria, and 98.2% [95% CI: 90.3\u2013100] in the 56 patients with P. vivax malaria. By day 2, 98% (56/57) of patients with P. falciparum and 96.9% (63/65) of those with P. vivax had cleared their peripheral parasitemia; none of the patients were still parasitaemic on day 3. Molecular analysis of P. falciparum parasites showed that none (0/61) had K13 mutations associated previously with artemisinin resistance or increased copy number of plasmepsin 2\u20133 (0/61). In the absence of artemisinin resistance, DHP has retained high efficacy for the treatment of uncomplicated malaria despite extensive drug pressure over a 9-year period.Dihydroartemisinin\u2013piperaquine (DHP) has been the first-line treatment of uncomplicated malaria due to both  Plasmodium falciparum has developed resistance to chloroquine (CQ), mefloquine, sulphadoxine\u2013pyrimethamine, piperaquine, and artemisinin, with the earliest evidence of resistance for all of these drugs documented in the Greater Mekong Subregion (GMS). By contrast, the epicenter of antimalarial drug resistance in Plasmodium vivax is on the island of Papua. In southern Papua, Indonesia, both P. falciparum and P. vivax malaria are equally prevalent.2 Clinical trials conducted in 2004 highlighted a very high risk of treatment failure following treatment with locally recommend antimalarial regimens. Within 28 days, 48% of patients with P. falciparum had failed treatment with CQ plus sulfadoxine\u2013pyrimethamine (CQ + SP) and 65% of those with P. vivax had failed treatment with CQ monotherapy.3 In view of these findings, the first-line treatment of uncomplicated malaria due to either P. falciparum or P. vivax was changed to dihydroartemisinin\u2013piperaquine (DHP) in March 2006. At the time of introduction of DHP, the risk of recurrent parasitemia within 42 days was 4.1% following P. falciparum and 10% following P. vivax malaria.4Over the last 50 years, 7 The spread of these parasites to other locations has potential to undermine the global policy of artemisinin-based combination therapy (ACT) for uncomplicated malaria. More recently, there have been reports of declining efficacy of the partner drug piperaquine in areas where the prevalence of artemisinin resistance is high.8 The spread of these multidrug-resistant parasites eastward across the Indonesian archipelago poses a significant threat to the national antimalarial policy. In Papua, where DHP has been used extensively over the last 9 years, a clinical study was undertaken to assess whether it still retained excellent efficacy.Over the last 10 years, there has been growing concern regarding the emergence of artemisinin-resistant parasites in the GMS.10 In brief, malaria is restricted to lowland areas where it is associated with three mosquito vectors: Anopheles koliensis, An. farauti, and An. punctulatus. The area has unstable malaria transmission. A household survey conducted in the same area in 2013 showed that malaria prevalence was 13.1%, with P. falciparum and P. vivax accounting for 5.9% and 6.0% of malaria, respectively.11The study was carried out in Timika, southern Papua, Indonesia, between March 2015 and April 2016. The epidemiology of this area has been described previously.12 CQ and SP were available in a few private pharmacies and their use has declined to less than 5% following policy change in March 2006.DHP is available, free of charge, from government health facilities and selected private-sector health facilities. Local protocol dictates that DHP is prescribed to patients with malaria confirmed by either rapid diagnostic test or microscopy. Local surveillance data showed that over 10 years, more than 485,000 doses of DHP have been administered to a population of 220,000 people. A cross-sectional survey undertaken in 2013 estimated that approximately 67% of individuals attend public health clinics, where DHP is available, for treatment of febrile illness.P. falciparum and P. vivax malaria in children and adults with uncomplicated symptomatic malaria. The study was based on the 2009 World Health Organization antimalarial drug therapeutic efficacy tests13 with 42 days of follow-up.This was a prospective open-label drug efficacy study of dihydroartemisinin\u2013piperaquine for P. falciparum and more than 250/\u03bcL asexual parasites for P. vivax. Pregnant and lactating women, patients with signs of severe malaria,14 or those presenting with severe malnutrition or significant comorbidities were all excluded. Informed consent was obtained from patients aged more than 18 years old and in those less than 13 years old, from their parent or guardian. Children aged between 13 and < 18 years were asked for their assent in the presence of their parent or guardian. If patients were illiterate, consent was obtained in the presence of a literate witness. All patients were provided with an information sheet in Indonesian language.Patients were eligible to enroll in the study if they were aged between 1 and 65 years, weighed more than 5 kgs, had a fever or history of fever in the preceding 24 hours, with slide-confirmed malaria with parasitemia of more than 1,000/\u03bcL asexual parasites for 15On admission, a standardized questionnaire was completed to record demographic information and details of the patients\u2019 symptoms, prior antimalarial medication, and their clinical examination. Axillary temperature was measured using a digital electronic thermometer. Venous or capillary blood was taken at enrolment for malaria smear, hemoglobin concentration, and parasite genotyping. Hemoglobin was measured using a portable photometer . Glucose 6 phosphate dehydrogenase (G6PDd) status was measured at enrolment using the Fluorescent Spot Test method.Patients were examined daily for the first 3 days of DHP treatment and then weekly for 6 weeks. Patients were also encouraged to return to the clinic on any other day if they felt unwell. At each visit, a physical examination was performed, the symptom questionnaire completed, and blood taken for blood film examination and measurement of hemoglobin concentration.Parasitemia was determined by microscopic examination of Giemsa-stained thick blood films, with parasites counted per 200 white blood cells (WBC) and peripheral parasitemia calculated assuming a white cell count of 7,300/\u03bcL. Slides were considered negative after review of 400 high-power fields. A thin smear was also examined to confirm parasite species and used for quantification if the parasitemia was greater than 200 per 200 WBC. All slides were read by a certified microscopist and cross-checked by a second experienced microscopist. In cases where readings were discordant, the slides were reread by a third microscopist and a consensus reached.\u2122 at enrolment and again on the day of failure was extracted using the QIAamp DNA mini kit  as per the manufacturer\u2019s instructions. Speciation was done by using an 18sRNA gene-based polymerase chain reaction (PCR) method for the detection of four different malaria species.17 For patients with recurrent P. falciparum infection, pre- and posttreatment isolates were genotyped using msp1, msp2, and glurp as described previously.19 The presence of polymorphisms in the propeller domains of the P. falciparum kelch (K13) gene, associated with artemisinin derivatives resistance, was determined by Sanger sequencing using the WWARN protocol.20Plasmepsin 2\u20133 copy number variants, associated with piperaquine resistance, were also examined in all samples as described previously.23DNA from venous or capillary blood samples collected in ethylene diamine tetraacetic acid-coated Vacutainers or Microtainers24 In addition, patients with P. vivax malaria were prescribed primaquine (0.5 mg/kg/day for 14 days) commencing on day 28 of the study follow-up. All doses of DHP, but not primaquine, were administered under direct supervision and the patients observed for 30 minutes for adverse reactions or vomiting. Any patient vomiting their medication within this period was re-administered the same dose of DHP and observed for an additional 30 minutes. If the patient vomited again, then they were withdrawn from the study and hospitalized for artesunate intravenous therapy.Dihydroartemisinin\u2013piperaquine (containing 40 mg dihydroartemisinin and 320 mg piperaquine) was administered once daily for 3 days as a dose-per-weight regimen with a target dose of 2.25 and 18 mg/kg of dihydroartemisinin and piperaquine, respectively, according to the National Guidelines.25 were treated according to local policy with a 7-day course of oral quinine  plus doxycycline  in patients older than 8 years or clindamycin  in children younger than 8 years. Patients with treatment failure after 28 days were retreated with DHP.Patients with therapeutic failure within 28 days of initial treatmentP. falciparum and the unadjusted risk of P. vivax recurrence. Secondary endpoints included the proportion of patients still parasitaemic on days 1, 2, and 3, posttreatment gametocyte carriage, and hematological recovery.The primary endpoints of the study were the PCR-adjusted risk of recurrence of U test or Kruskal\u2013Wallis method were used for nonparametric comparisons and Student\u2019s t test or one-way analysis of variance for parametric comparisons. For categorical variables, percentages and corresponding 95% confidence intervals (95% CI) were calculated using Wilson\u2019s method. Proportions were examined using \u03c72 with Yates\u2019 correction or by Fisher\u2019s exact test. The cumulative risk of failure was assessed by survival analysis using the Kaplan\u2013Meier method. Anyone lost to follow up or representing with a different outcome were censored on their last day of follow-up and regarded as not being treatment failures. For determining the risk of P. falciparum recrudescence, all early treatment failures and homologous recrudescences were considered failures.Data were double entered and validated using EpiData 3.02 software  and analysis performed using SPSS for Windows . The Mann\u2013Whitney http://www.clinicaltrials.gov/ct) as NCT 02353494.The study was approved by the Eijkman Institute Research Ethics Commission, Jakarta, Indonesia (Ref: 014/EI-IRB/XII/2014), and the Human Research Ethics Committee of the NT Department of Health & Families and Menzies School of Health Research, Darwin, Australia (HREC-2015-2342). Informed consent was obtained from all adult participants and from parents of children. The trial was registered with the clinical trials website . Of these patients, 329 (32%) were screened by the study team for enrolment criteria. A total of 281 patients were eligible for the study, of whom 129 (45.9%) gave consent to participate in the study   were positive for malaria (513 ria) see . Of 48 pP. falciparum was G6PD deficient and another male with P. vivax had intermediate G6PD activity; primaquine was not prescribed to either of these patients.Five patients (3.9%) vomited their medication after the first dose, but tolerated the repeat dose. The median total dose of dihydroartemisinin administered was 7 mg/kg (range 5.4\u201310.6) and that of piperaquine was 57 mg/kg (range 43.2\u201384.7). In total, one male patient with P. falciparum and one with P. vivax). The overall unadjusted efficacy at day 42 was 98.2% [95% CI: 90.3\u2013100].In total, four patients had recurrent parasitemia during the follow-up period  completed 42 days of follow-up and 41 had an adequate clinical and parasitological response. Two patients had asymptomatic recurrent P. falciparum parasitemia recorded on day 42: a 15-year-old female treated with 48.8 mg/kg of piperaquine with reinfection and a 52-year-old male treated with 53.3 mg/kg of piperaquine with recrudescent infection. The unadjusted efficacy for P. falciparum at day 42 was 95.3% [95% CI: 84.2\u201399.4] and after adjusting by PCR, this rose to 97.7% [95% CI: 87.4\u201399.9]. Of the 68 patients with P. vivax infection, 56 (82.3%) completed follow-up. Two patients had recurrent parasitemia on day 35: a 2-year-old boy with P. vivax infection in whom the parasite genotype could not be determined because insufficient blood was taken at enrolment and an 18-year-old female with symptomatic P. falciparum infection. The unadjusted efficacy for P. vivax at day 42 was 98.2% [95% CI: 90.4\u2013100]; Of the 61 patients with P. falciparum and P. vivax malaria. Within 24 hours, 62.7% (37/59) of patients with P. falciparum and 74.6% (50/67) of those with P. vivax were aparasitemic, and by 48 hours, this had risen to 98.2% (56/57) and 96.9% (63/65), respectively. None of these patients were parasitemic on day 3.Parasite clearance was rapid following treatment in both P. falciparum and 26.5% (18/68) with P. vivax malaria. The proportion of patients with patent gametocytemia in those with P. falciparum infection was 10.2% (6/59) at 24 hours, 12.3% (7/57) at 48 hours, and 5.9% (3/51) at day 7. In patients with P. vivax malaria, the proportion with gametocytes fell to 6% (4/67) by 24 hours, 4.6% (3/65) at 48 hours, and 1.5% (1/65) at day 7. One patient had P. falciparum gametocytes detected at day 14, and one patient had P. vivax gametocytes detected at day 21. None of the patients had gametocytes after day 21.Gametocytemia was present at enrolment in 3.3% (2/61) of patients infected with P. falciparum and 13.2 g/dL [95% CI: 12.6\u201313.8] in those with P. vivax .Mean hemoglobin concentration at enrolment was 13.8 g/dL [95% CI: 13.1\u201314.5] in patients with P. vivax . The meaNo severe adverse events were reported associated with treatment. Twenty-four hours following treatment, 21.8% (27/124) of the patients had nausea (8), dyspepsia (2), dizziness (13), urticaria (1), and weakness (3). At 48 hours, 9.9% (12/121) patients had mild nausea. All symptoms were resolved quickly. One case with urticaria was not associated with DHP, but due to mild food allergy.P. falciparum samples, with no mutations in the K13 gene (0/65) or increased copy number of plasmepsin 2\u20133 (0/74).Molecular analysis could be undertaken in all 4 Over the ensuing decade, DHP has been used extensively throughout the country. This recent therapeutic efficacy study in the eastern province of Papua reassuringly confirms that, despite the high prevalence of artemisinin and emerging piperaquine resistance in the GMS, DHP retains high efficacy against uncomplicated P. falciparum and P. vivax malaria after 9 years of use in this region.In March 2006, Indonesia was one of the first countries to adopt a unified first-line policy of ACT for the treatment of uncomplicated malaria due to any species of malaria. It was also the first country to adopt DHP as its preferred choice of ACT, a decision made in view of its high efficacy and prolonged posttreatment prophylaxis associated with piperaquine, the slowly eliminated partner drug.P. falciparum malaria was greater than 97%, similar to that reported in 2005.4 The efficacy of DHP against P. vivax was almost 100%, with only one of 56 patients (1.8%) having a recurrent infection on day 35, compared with an efficacy of 90% reported in 2005.4 Compared with our original efficacy study in 2005, the baseline parasitemias in the present study were 3\u20134 fold higher and the gametocyte carriage 2\u20137 fold lower. The high gametocyte carriage at enrolment in 2005 likely reflects the poor efficacy of the first-line treatment before policy change in 2006, with almost 20% of the patients reporting a history of malaria in the preceding month.3 With the wide deployment of highly efficacious treatment, the proportion of patients partially treated and reporting recent malaria has fallen substantially.12The PCR-adjusted efficacy in patients with 26 However, the proportion of parasitemic patients fell rapidly, with more than 98% aparasitemic by 48 hours and none of the patients remaining parasitemic at 72 hours. Molecular analysis confirmed that none of the parasites had K13 gene mutations that have been associated with artemsinin resistance or amplification of the plasmepsin 2\u20133 gene cluster that have been associated with piperaquine resistance.Patients were only assessed once per day precluding precise estimation of the slope of the parasite clearance curves.P. falciparum malaria was first described in western Cambodia in 200827 and has been reported subsequently in Thailand, Myanmar, Laos, and Vietnam.29 In Cambodia, where the prevalence of artemisinin resistance is greatest, recent clinical trials have revealed that the efficacy of DHP for P. falciparum has fallen to less than 80%, with almost half of the patients still parasitemic at 72 hours.30 Reassuringly, in the same study DHP retained good efficacy against P. vivax.30Resistance to artemisinin derivatives for the treatment of 31 The recommended combination has subsequently rationalized to DHA plus piperaquine which has been available outside of China since the start of the millennium. The significant mismatch in the pharmacokinetic profiles of these two drug components has raised concerns of exposing parasites to subtherapeutic concentrations of piperaquine monotherapy in the 2\u20136 weeks following treatment, particularly where the risk of reinfection is high. It is feared that this mismatch will foster the emergence and spread of drug-resistant parasites. Indeed in this respect, it is interesting to note that in an area where artemisinin resistance has emerged the efficacy of the combination fell quickly.30Piperaquine was first synthesized in the 1960s and used extensively in China and Indochina as prophylaxis and treatment over the next 20 years. Within a decade, high rates of treatment failure were observed and piperaquine monotherapy was discontinued. The drug was removed from the pharmacopeia until it was reintroduced in 1990s in combination with dihydroartemisinin, trimethoprim, and primaquine phosphate.plasmepsin 2\u20133 gene cluster has been identified as an important molecular determinant of piperaquine resistance in P. falciparum.22 Our molecular analysis reveals that neither of these polymorphisms is present in Timika.The results from this Papuan study are reassuring and suggest that in the absence of artemisinin resistance, the ACT regimen protects against de novo emergence of resistance. In Cambodia, amplification of the It is likely that the development of artemisinin resistance in Cambodia has been driven by high rates of self-treatment via private sector and unregulated use of antimalarial agents, including poor ACT quality. In Indonesia, DHP procurement is highly regulated by the Indonesian Ministry of Health, and the drug is only available at government health facilities and selected private-sector facilities, which are able to confirm the diagnosis of malaria by microscopy or rapid diagnostic test. Hence, DHP is generally only prescribed to those with confirmed malaria and this degree of control may have contributed to the sustained efficacy of the drug in this region.In conclusion, despite intense use of DHP over almost a decade in a region of mesoendemic malaria, there is no evidence for the presence of either artemisinin or piperaquine resistance and the combination regimen retains excellent efficacy. However, in view of the highly mobile population and potential spread of artemisinin resistance from the GMS, vigilance for declining efficacy is warranted."}
+{"text": "Plasmodium falciparum transmission to mosquitoes. We aimed to assess the efficacy and safety of primaquine and methylene blue in preventing human to mosquito transmission of P falciparum among glucose-6-phosphate dehydrogenase (G6PD)-normal, gametocytaemic male participants.Primaquine and methylene blue are gametocytocidal compounds that could prevent P falciparum malaria. G6PD-normal participants with gametocytes detected by blood smear were randomised 1:1:1:1 in block sizes of eight, using a sealed-envelope design, to receive either sulfadoxine-pyrimethamine and amodiaquine, sulfadoxine-pyrimethamine and amodiaquine plus a single dose of 0\u00b725 mg/kg primaquine, dihydroartemisinin-piperaquine, or dihydroartemisinin-piperaquine plus 15 mg/kg per day methylene blue for 3 days. Laboratory staff, investigators, and insectary technicians were masked to the treatment group and gametocyte density of study participants. The study pharmacist and treating physician were not masked. Participants could request unmasking. The primary efficacy endpoint, analysed in all infected patients with at least one infectivity measure before and after treatment, was median within-person percentage change in mosquito infectivity 2 and 7 days after treatment, assessed by membrane feeding. This study is registered with ClinicalTrials.gov, number NCT02831023.This was a phase 2, single-blind, randomised controlled trial done at the Clinical Research Centre of the Malaria Research and Training Centre (MRTC) of the University of Bamako . We enrolled male participants aged 5\u201350 years with asymptomatic Between June 27, 2016, and Nov 1, 2016, 80 participants were enrolled and assigned to the sulfadoxine-pyrimethamine and amodiaquine (n=20), sulfadoxine-pyrimethamine and amodiaquine plus primaquine (n=20), dihydroartemisinin-piperaquine (n=20), or dihydroartemisinin-piperaquine plus methylene blue (n=20) groups. Among participants infectious at baseline (54 [68%] of 80), those in the sulfadoxine-pyrimethamine and amodiaquine plus primaquine group (n=19) had a median 100% (IQR 100 to 100) within-person reduction in mosquito infectivity on day 2, a larger reduction than was noted with sulfadoxine-pyrimethamine and amodiaquine alone . The dihydroartemisinin-piperaquine plus methylene blue (n=11) group had a median 100% (IQR 100 to 100) within-person reduction in mosquito infectivity on day 2, a larger reduction than was noted with dihydroartemisinin-piperaquine alone . Haemoglobin changes were similar between gametocytocidal arms and their respective controls. After exclusion of blue urine, adverse events were similar across all groups .P falciparum transmission. Both primaquine and methylene blue were well tolerated.Adding a single dose of 0\u00b725 mg/kg primaquine to sulfadoxine-pyrimethamine and amodiaquine or 3 days of 15 mg/kg per day methylene blue to dihydroartemisinin-piperaquine was highly efficacious for preventing Bill & Melinda Gates Foundation, European Research Council. Since 2000, scale-up of interventions has led to substantial gains in malaria control across sub-Saharan Africa.Plasmodium falciparum in areas of low malaria transmission and emerging antimalarial drug resistance.Several malaria control and elimination strategies target the plasmodium parasite in human beings, including case management, seasonal malaria chemo-prevention, and mass drug administration. Drug combinations used in these strategies do not target the gametocyte stage of the parasite, allowing continued human to mosquito transmission despite clearance of the asexual blood stage infection. WHO recommends addition of a single 0\u00b725 mg/kg dose of primaquine (single low-dose primaquine) to standard artemisinin-based combination therapy to reduce onward transmission of Evidence before the studyClinicalTrials.gov and PubMed between Sept 18, 2015, and Sept 15, 2017, using the search terms \u201cprimaquine\u201d OR \u201cmethylene blue\u201d AND \u201cmalaria\u201d AND \u201cfalciparum\u201d, OR \u201cprimaquine\u201d OR \u201cmethylene blue\u201d AND \u201ctransmission\u201d, OR \u201cprimaquine\u201d OR \u201cmethylene blue\u201d AND \u201cgametocyte\u201d. No language restrictions were used in the search. Although numerous studies, including randomised controlled trials, systematic reviews, and meta-analyses, have reported on the efficacy and safety of single low-dose primaquine as a Plasmodium falciparum gametocytocidal drug, only four efficacy studies used membrane feeding assays, the gold standard for infectivity assays, to assess the transmission-blocking effect of primaquine. None of these studies assessed the efficacy of primaquine combined with sulfadoxine-pyrimethamine plus amodiaquine. One study in Colombia assessed the efficacy of adding a single dose of 0\u00b775 mg/kg primaquine to sulfadoxine-pyrimethamine plus amodiaquine on P falciparum gametocyte clearance. Using blood smear microscopy, authors found lower gametocytaemia on days 4 and 8, but no clearance of gametocytes by day 8. Since no direct measurements of membrane feeding assays were done, it is unclear if the persisting gametocytes would have been infectious to mosquitoes. Eight published trials of methylene blue were identified. Two studies assessed the effect of methylene blue with either artesunate, amodiaquine, or artesunate-amodiaquine on gametocytes. Groups receiving methylene blue had lower gametocytaemia by at least day 7. Neither of these studies assessed transmission using membrane feeding assays, nor was methylene blue paired with dihydroartemisinin-piperaquine, one of the major drugs used in community drug interventions, known for its long prophylactic period.We searched Added value of this studyP falciparum. Our findings show that antimalarial combinations of sulfadoxine-pyrimethamine plus amodiaquine with 0\u00b725 mg/kg primaquine and dihydroartemisinin-piperaquine with a 3-day treatment course of 15 mg/kg methylene blue are safe and block human to mosquito transmission by day 2 post-treatment.To our knowledge, this is the first study to use the membrane feeding assay to directly quantify the effect of primaquine (partnered with sulfadoxine-pyrimethamine plus amodiaquine) and methylene blue (partnered with dihydroartemisinin-piperaquine) on human to mosquito transmission of Implications of all the available evidencePotent gametocytocidal drugs primaquine and methylene blue are two promising transmission-blocking tools that can be used for falciparum malaria control and elimination. Future studies should focus on assessment of the community-level effect of adding gametocytocidal drugs to first-line treatments and large-scale, drug-based interventions, including seasonal malaria chemoprophylaxis (sulfadoxine-pyrimethamine plus amodiaquine) and mass drug administration (dihydroartemisinin-piperaquine).P falciparum transmission is unclear. The antimalarial combination sulfadoxine-pyrimethamine and amodiaquine has strong treatment and chemoprophylactic effects. Sulfadoxine-pyrimethamine and amodiaquine is the standard drug combination for seasonal malaria chemoprevention and has been highly effective in reducing malaria morbidity and mortality in the Sahel and sub-Sahel regions of Africa, where seasonal malaria chemoprevention is widely implemented.First, whether addition of single low-dose primaquine to non-artemisinin-based combination therapy can prevent P falciparum gametocytes, resulting in partial reductions in human to mosquito transmission.14The second question is whether addition of methylene blue  to artemisinin-based combination therapy reduces post-treatment transmission potential. Methylene blue has highly potent gametocytocidal effectsP falciparum gametocytes (the parasite stage infectious to mosquitoes), the addition of gametocytocidal drugs to these regimens might prevent transmission shortly after treatment and limit the spread of drug-resistant parasites.Since neither sulfadoxine-pyrimethamine and amodiaquine or dihydroartemisinin-piperaquine target the mature stage of The aim of this study was to establish the safety and efficacy of the addition of single low-dose primaquine to sulfadoxine-pyrimethamine and amodiaquine, and the addition of methylene blue to dihydroartemisinin-piperaquine in reducing human to mosquito transmission among glucose 6-phosphate dehydrogenase (G6PD)-normal male participants in Mali.This was a phase 2, single-blind, randomised controlled trial done at the Clinical Research Centre of the Malaria Research and Training Centre (MRTC) of the University of Bamako .P falciparum malaria at enrolment, had at least two P falciparum gametocytes per 500 white blood cells on thick film microscopy , and had an absence of other non-P falciparum species on blood film. Female participants were excluded to mitigate the risk of haemolysis through incorrect classification of G6PD-deficient female heterozygotes. Participants were also excluded if they had a serious or chronic illness , weighed 80 kg or more, reported antimalarial use within 7 days of screening, or reported allergies to study drugs.Study participants were recruited from the town of Ou\u00e9less\u00e9bougou, Mali, and its surrounding villages. Malaria transmission in this region is hyperendemic and highly seasonal with peaks from July to November. A study team, comprised of clinicians and lab technicians, met with community leaders, village health workers, and heads of households from each village to explain the study and obtained approval to conduct the study. Before participant screening, village health workers used a door-to-door approach to inform households of the date and location where consenting and screening would take place. Participants were enrolled if they were male, aged 5\u201350 years, G6PD-normal by Carestart G6PD rapid diagnostic test , had a haemoglobin concentration of at least 10 g/dL, had asymptomatic Participants aged at least 18 years provided written informed consent before screening and enrolment. Parental consent was obtained for participants younger than 18 years, and assent was obtained from children aged 12\u201317 years.Ethical approval was granted by the Ethics Committee of the Faculty of Medicine, Pharmacy, and Dentistry of the University of Science, Techniques, and Technologies of Bamako , the Committee on Human Research at the University of California San Francisco , and the Research Ethics Committee of the London School of Hygiene & Tropical Medicine . For the full study protocol see the Patients were individually randomised in a 1:1:1:1 ratio, in block sizes of eight, to receive either sulfadoxine-pyrimethamine and amodiaquine, sulfadoxine-pyrimethamine and amodiaquine with single low-dose primaquine, dihydroartemisinin-piperaquine, or dihydroartemisinin-piperaquine with methylene blue. A UCSF investigator (JMB) randomly generated the treatment assignment using Excel , which was linked to a pre-specified, unique participant ID number. The investigator assembled sealed, opaque envelopes with the participant ID number on the outside and treatment assignment contained within. The site pharmacist in Mali (AM) opened the envelopes and provided treatment. The study assignment was masked to investigators and staff involved in assessing all laboratory outcomes. Insectary technicians were masked to the treatment group and gametocyte density of study participants. The study pharmacist and treating physician were not masked. Participants could ask the study physician which treatment they received.Upon enrolment and collection of day 0 samples, treatment regimens were administered to participants after a fatty snack (biscuits) given to prevent gastrointestinal side-effects and vomiting , 2, and 7. Venous blood was collected in heparin tubes stored at 37\u00b0C and placed in a membrane feeding system within 1 min of collection. For the first three enrolled participants, about 70 Pfs25) and male  gametocyte mRNA, 100 \u03bcL of blood was collected in EDTA and transferred into 500 \u03bcL of RNAprotect Cell Reagent . Samples were stored at MRTC at \u221280\u00b0C until shipment on dry ice at a controlled temperature (\u221280\u00b0C to \u221270\u00b0C). Total nucleic acids were extracted by MagNAPure LC automated extractor . qRT-PCR was done as previously described using sex-specific trendlines of cultured gametocytes.Measurements of gametocyte density were taken before treatment (day 0), and on days 1, 2, 7, 14, 28, and 42 after enrolment. Gametocyte density was measured using blood smear microscopy and by real-time reverse transcription-PCR (qRT-PCR). Blood slides were stained with Giesma and independently read by expert research microscopists over 500 fields for quantification of gametocytes and asexual parasites. For qRT-PCR quantification of female  if at least one dissected mosquito had at least one oocyst present.Secondary outcomes included the prevalence, density, circulation time (days), and sex ratios of female and male gametocytes. Estimates of gametocyte sex ratio were restricted to samples with a total gametocyte density greater than 0\u00b72 gametocytes per \u03bcL (20 gametocytes per 100 \u03bcL of blood sample).Safety outcomes assessed included the number of adverse events and the mean within-person change in haemoglobin concentration per group after treatment, the proportion of participants who had a greater than 25% decrease in haemoglobin, and the proportion of participants with 8 g/dL or less haemoglobin at any point during follow-up. Adverse events were assessed at each follow-up visit and monitored passively by medical staff. All adverse events were graded by severity  according to the study's standard operating procedures and based on the Division of Microbiology and Infectious Diseases (DMID) toxicity tables. Adverse events were further categorised based on whether they were considered serious by the treating physician, and categorised by their causal relationship to the study drugs . Adverse events were considered causal if they were categorised as either possibly, probably, or definitely related to study drug.Based on preliminary data from a similar infectivity trial,vs treatment group). Our analysis of the change in mosquito infectivity included all randomised participants who had an infectivity measurement before treatment and at least one infectivity measurement after treatment. Additionally, we did an analysis of change in infectivity among the subset of participants who infected at least one mosquito after treatment.For the primary efficacy outcome, we calculated the median within-person percentage change in infectivity and accompanying IQRs for each group. Positive percentages indicated reductions in infectivity. We used a one-sided non-parametric Wilcoxon rank-sum test for pairwise comparisons of the change in infectivity after treatment .vs sulfadoxine-pyrimethamine and amodiaquine alone; and dihydroartemisinin-piperaquine plus methylene blue vs dihydroartemisinin-piperaquine alone), unless otherwise stated. Statistical analyses were done using Stata version 14.0 and SAS version 9.4.All other comparisons between proportions were done using two-sided tests, and we used Fisher's exact test for between-group comparisons and McNemar's test for within-group comparisons. Wilcoxon's signed rank tests were used for within-group comparisons and Wilcoxon rank-sum tests were used for between-group comparisons. All analyses compared treatment with or without gametocytocidal compounds .The trial was overseen by an independent data safety monitoring committee, monitored by an external clinical trials monitoring group , and registered with The funders of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit the publication.Between June 27, 2016, and Nov 1, 2016, 1749 male participants were screened for eligibility. 80 participants were enrolled and randomly assigned to receive either sulfadoxine-pyrimethamine and amodiaquine plus primaquine (n=20), dihydroartemisinin-piperaquine plus methylene blue (n=20), and their comparator groups, sulfadoxine-pyrimethamine and amodiaquine alone (n=20), and dihydroartemisinin-piperaquine alone n=20; . 72 90%.Figure 180 membrane feeding assays were done before treatment, 80 on day 2 after treatment, and 76 on day 7 after treatment. 20\u2008599 mosquitoes were used, 18\u2008026 (88%) of which survived to the day of dissection. Before treatment, 54 (68%) of 80 individuals infected at least one mosquito , and 908Before treatment, all study participants with blood samples available for molecular analysis (n=78) had detectable concentrations of both female and male gametocytes. Gametocyte sex ratios were female biased at enrolment, with no differences in gametocyte prevalence or density between study arms . The preThe mean circulation time of female gametocytes was longer than that of male gametocytes for all treatment arms . The sulCorrelations were found between qRT-PCR gametocyte density and mosquito infectivity rates . Fairly Absolute and within-person percentage change in mean haemoglobin concentration did not differ between the sulfadoxine-pyrimethamine and amodiaquine plus primaquine and sulfadoxine-pyrimethamine and amodiaquine alone groups or the dihydroartemisinin-piperaquine plus methylene blue and dihydro-artemisinin-piperaquine alone groups at any point during follow-up . One parOverall, 59 (74%) of 80 participants reported a total of 162 adverse events during follow-up, with 19 of these events being bluish colouration of urine . The numThis study shows the pronounced transmission-blocking effects of addition of gametocytocidal drugs to standard antimalarial treatments. Addition of single low-dose primaquine to sulfadoxine-pyrimethamine and amodiaquine or methylene blue to dihydroartemisinin-piperaquine resulted in almost complete blockage of human to mosquito transmission by day 2, and a larger reduction in transmission than that offered by sulfadoxine-pyrimethamine and amodiaquine or dihydroartemisinin-piperaquine alone. No differences in adverse events (excluding blue urine) or haemolysis were observed between groups, suggesting primaquine and methylene blue are safe in this population.P falciparum infection to prevent formation of mature gametocytes is crucial for malaria control.We examined gametocyte dynamics and infectivity to mosquitoes following treatment with sulfadoxine-pyrimethamine and amodiaquine plus primaquine or dihydroartemisinin-piperaquine plus methylene blue, two drug regimens that have important roles in malaria control and elimination efforts. The importance of our direct assessments of infectivity is illustrated by the observation that the transmission-blocking effect of gametocytocidal drugs precedes gametocyte clearance.P falciparum gametocyte carriers using mosquito infectivity assays, confirming the potency of methylene blue as suggested by previous in-vitroTo our knowledge, this study is the first to formally show the potent transmission-blocking effect of methylene blue in P falciparum gametocyte carriage and infectivity. Sulfadoxine-pyrimethamine and amodiaquine alone did not have an effect on reducing human to mosquito transmission in the week following treatment. Most individuals treated with sulfadoxine-pyrimethamine and amodiaquine carried gametocytes for at least 1 month following treatment with no reduction in mosquito infection rates achieved by day 7 after treatment. Sulfadoxine-pyrimethamine and amodiaquine is recommended by WHO for use in seasonal malaria chemoprevention. Although the primary objective of seasonal malaria chemoprevention is to reduce malaria morbidity and mortality and not to directly reduce transmission, adding single low-dose primaquine to seasonal malaria chemoprevention might confer indirect community benefits for malaria transmission by reducing post-treatment transmission potential. Transmission reduction by seasonal malaria chemoprevention could be further improved by expanding the age range to include adolescents because gametocyte carriage is high in this populationWhen combined with sulfadoxine-pyrimethamine and amodiaquine,Countries moving towards malaria elimination could consider adding a transmission-blocking drug to first-line treatments or community-wide mass drug administration to rapidly reduce population-level malaria transmission. Modelling studies suggest that, given adequate coverage and multiple rounds of mass drug administration, addition of gametocytocidal drugs  to antimalarial combinations with a long prophylactic period  might result in larger reductions in transmission than with dihydroartemisinin-piperaquine treatment alone.Our study revealed that gametocytocidal drugs might have different effects on male and female gametocytes. The strongly reduced circulation time of male gametocytes following methylene blue treatment supports hypotheses generated from in-vitro assessments that methylene blue disproportionally affects male gametocytes.P falciparum gametocyte densities. Therefore, our findings might not represent the effect of single low-dose primaquine and methylene blue in other malaria-infected populations, particularly those with lower gametocyte densities who have lower pre-treatment and post-treatment transmission potential and G6PD-deficient populations. Although studies in Burkina Faso showed that methylene blue is unlikely to be responsible for clinically relevant haemolysis in individuals with or without G6PD-deficiency,Our study has several limitations. First, the study population analysed was not as per intention to treat, as one patient was lost to follow-up and seven patients withdrew. This factor was unlikely to cause attribution bias since the reasons given for withdrawals were unrelated to study drug. Second, our study endpoint was the proportion of oocyst-positive mosquitoes, based on the assumption that even low oocyst densities are likely to render mosquitoes infective.P falciparum transmission in G6PD-normal male participants. Further work is needed to understand what effect these findings could have on malaria control and elimination strategies, using mathematical models and community trials to assess costs, risks, and benefits of adding single low-dose primaquine or methylene blue to existing regimens.Adding a single dose of 0\u00b725 mg/kg primaquine to sulfadoxine-pyrimethamine and amodiaquine or 3 days of 15 mg/kg per day methylene blue to dihydroartemisinin-piperaquine were safe and highly efficacious methods for preventing human to host Division of Microbiology and Infectious Diseases toxicity tables see https://www.niaid.nih.gov/research/dmid-safetyreporting-pharmacovigilanceFor"}
+{"text": "Plasmodium falciparum poses a great threat of increased fatalities in cases of cerebral and other forms of severe malaria infections in which parenteral artesunate monotherapy is the current drug of choice. The study aimed to investigate in a mouse model of human cerebral malaria whether a trioxaquine chemically synthesized by covalent linking of a 4,7-dichloroquinoline pharmacophore to artesunate through a recent drug development approach termed \u2018covalent bitherapy\u2019 could improve the curative outcomes in cerebral malaria infections.The emergence of multidrug-resistant strains of Plasmodium berghei ANKA and intravenously (iv) treated with the trioxaquine\u00a0from day 8 post-infection (pi) at 12.5 and 25\u00a0mg/kg, respectively, twice a day for 3\u00a0days. Treatments with the trioxaquine precursors , and quinine were also included as controls. In vivo safety evaluation for the trioxaquine was done according to Organization for Economic Co-operation and Development (OECD) guidelines 423, where female Swiss albino mice were orally administered with either 300 or 2000\u00a0mg/kg of the trioxaquine\u00a0and monitored for signs of severity, and or mortality for 14\u00a0days post-treatment.Human cerebral malaria rodent model, the C57BL/6 male mice were infected intraperitoneally (ip) with The trioxaquine showed a potent and a rapid antiplasmodial activity with 80% parasite clearance in the first 24\u00a0h for the two dosages used. Long-term parasitaemia monitoring showed a total parasite clearance as the treated mice survived beyond 60\u00a0days post-treatment, with no recrudescence observed. Artesunate treated mice showed recrudescence 8\u00a0days post-treatment, with all mice in this group succumbing to the infection. Also, 4,7-dichloroquinoline and quinine did not show any significant parasitaemia\u00a0suppression in the first 24\u00a0h post-treatment, with the animals succumbing to the infection.Covalent bitherapy proves to be a viable source of urgently needed new anti-malarials for management of cerebral malaria, and this polypharmacology approach could be a potential strategy to protect artesunate from parasite resistance and in potentially improving clinical outcomes in severe forms of malaria infections. Plasmodium falciparum, with a higher burden greatly felt in sub-Saharan Africa [Despite many years of research and great progress in the fight against malaria , cerebran Africa , a contin Africa . Children Africa . With non Africa . Extremen Africa . For decn Africa . Concernn Africa , 8. Despn Africa .Plasmodium falciparum, the most lethal pathogen among the Plasmodium species causing up to 90% of all malaria-related deaths in sub-Saharan Africa has shown multi-drug resistance [sistance . Reportssistance . Extremesistance  has necesistance . Currentsistance . Unfortusistance , thus caIn the long and difficult search to develop effective, cheaper and urgently needed anti-malarial drugs, many research groups have employed the emerging strategy in medicinal chemistry called hybridization or covalent bitherapy \u201314. ThisN-(7-chloroquinolin-4-ylamino)-ethyl-artesunate-19-carboxamide  [50, 6.89\u00a0ng/mL) and CQ-resistant (W2) \u00a0P.\u00a0\u200bfalciparum isolates [In this study, a trioxaquine termed ctively) . The duaisolates .Fig.\u00a01StPlasmodium berghei (ANKA strain) was used in the study. This parasite, maintained at \u221280\u00a0\u00b0C in Kenya Medical Research Institute (KEMRI) at the Centre for Biotechnology, Research, and Development (CBRD), was thawed and revived by intraperitoneal (ip) inoculation into male Swiss albino mice. The mice served as parasite donor for passaging into C57BL/6 male mice weighing 25\u00a0\u00b1\u00a02\u00a0g, used as a mouse model of CM. The mice were subsequently randomized into groups of five per cage and fed with pellets and water, ad libitum [ libitum .\u00ae, while the trioxaquine, a kind gift from Prof. Francis W Muregi, was previously synthesized by linking the 4,7-dichloroquinoline to a linker and coupling the product to artesunate [The test drugs, namely artesunate, 4,7-dichloroquinoline and quinine were purchased from Sigma Aldrichtesunate . On the tesunate . Quinine5 parasitized erythrocytes (pRBCs) at inoculum of 0.2\u00a0ml. The infected mice were then randomized in plastic cages in groups of five and observed daily for signs of experimental cerebral malaria (ECM), including rough coat, limb paralysis, ataxia, convulsions, and or coma [The donor mice were anaesthetized and the infected red blood cells collected using heparinized syringe via cardiac puncture. The parasitaemia was then adjusted downwards using sterilized phosphate-buffered saline (PBS). Male C57BL/6 mice were then inoculated ip with approximately 1\u00a0\u00d7\u00a010 or coma , 20. In  or coma , 20.\u00a0Gie\u00ae Excel (Microsoft Corp.). Percentage (%) parasite suppression (chemosuppression) at each drug dose was determined as described by Muregi et al. [The treatment of CM in experimental mice with established infection was carried out by the approach of Ryley and Peters\u00a0 in whichi et al. :\\documenp\u00a0<\u00a00.05) considered significant.The survival of mice was monitored for up to 60\u00a0days after the end of the 3-day treatment. All RBC counts and parasitaemia levels were expressed as mean values\u00a0\u00b1\u00a0standard deviations and the parasitaemia data were processed using the one-way analysis of variance (ANOVA). Mean parasitaemia between each experimental group relative to controls was compared using Student\u2019s t test with a probability of 5% (\u00ae) dye on day 8 pi after the experimental animals had exhibited clear signs for cerebral malaria. The animals were euthanized after 1\u00a0h for brain extraction to assess the extent of brain staining with the dye and the images documented. The principle behind this method is based upon the formation of bonds between the dye and plasmatic albumin and, in the event of brain blood vessels leakage due to compromised blood\u2013brain barrier (BBB), the dye\u2013protein complex migrates to the tissue, impregnating it in a tone of blue visible to the naked eye [C57BL/6 mice were injected iv with 200\u00a0\u03bcl of 2% (w/v) Evans Blue  were adopted. This is a protocol that makes use of three uninfected animals of same gender, especially females, at every dosage category and assessment based on death and or signs of severity observed with the dosed animals. Averagely, two to four dosage steps in the range of 5, 50, 300 and 2000\u00a0mg/kg are considered enough in making judgement on the acute oral toxicity of the test compound .Two fixed dose levels of 300 and 2000\u00a0mg/kg were used and dosing was done in a stepwise manner starting with the lowest dose upwards. Swiss albino female mice weighing 20\u00a0\u00b1\u00a02\u00a0g were used where the animals were first fasted for 4\u00a0h, after which they were dosed orally with the above dosages. The start, duration and severity of toxic signs of the test compound, such as changes in skin and fur, eyes, mucous membranes, tremors, convulsions, salivation, diarrhoea, coma, and even death\u00a0were monitored. The animals were observed individually at least once during the first 30\u00a0min, periodically during the first 24\u00a0h with special attention given during the first 4\u00a0h, and daily thereafter for the next 14\u00a0days.The use of mice in this research work was approved by KEMRI's Animal Care and Use Committee (ACUC)  and the\u00a0Scientific Ethical Review Unit (SERU) (Permit No: KEMRI/RES/7/3/1/2016).\u00a0All mice that were deemed to have completed their intended role in the study were euthanized using 150 mg/kg body weight sodium pentobarbital solution injected ip, placed in biohazard bags and incinerated.P value <0.05 was considered to be statistically significant.Data analysis was performed using SPSS version 16. All values are expressed in means (\u00b1SDs). Cumulative survival rates were calculated according to Kaplan\u2013Meier method and groups were compared using the log-rank test. Parasitaemia courses were compared by analysis of variance (ANOVA). Parasitaemia before and at 24\u00a0h after treatment were compared by paired Student t test. inter alia. Parasitaemia load reduction for each test drug within the first 24\u00a0h post-treatment is presented in Table\u00a0Treatment was initiated on day 8 pi after the experimental animals were clearly diagnosed with ECM or having presented with at least one or both well-recognized signs of ECM in animal models, including neurological perturbations such as ataxia, rough coat, limb paralysis, At the two dosages used, 12.5 and 25\u00a0mg/kg iv, the\u00a0trioxaquine showed a strong and promising antiplasmodial activity. At 12.5\u00a0mg/kg, parasitaemia decreased from 11.7 to 1.6 (86.6% clearance) and from 11.7 to 0.4 (96.4% clearance) at 25\u00a0mg/kg just after 24\u00a0h post-treatment and no parasite was observable under microscope at 48\u00a0h post-treatment. A potent anti-malarial activity was therefore exhibited by the artesunate\u2013quinoline hybrid drug at the two dosages, resulting in a rapid parasite clearance. The treated animals also survived beyond 60\u00a0days post-treatment except for the 12.5\u00a0mg/kg, where death of one animal was recorded on day 4 post-treatment even after no parasite was observed under microscope.Artesunate monotherapy at 12.5\u00a0mg/kg also showed a rapid parasitaemia clearance of 84.8% within the first 24\u00a0h post-treatment, and no parasite was observable under microscope at 48\u00a0h post-treatment. However, recrudescence was observed on day 8 post-treatment with all the animals succumbing to the infection by day 16 post-treatment Fig.\u00a0. QuinineLack of recrudescence observed with the trioxaquine-treated animals could be attributed to the 4,7-dichloroquinoline, the partner drug to artesunate in the\u00a0trioxaquine.Damage to the blood\u2013brain barrier (BBB) has been implicated and is believed to be one of the underlying mechanisms of pathophysiology of CM, as the leaky BBB would allow toxic or unwanted compounds to enter the brain, causing neurological dysfunction. Observations of the brain tissue in infected animals that were never injected with the 2% Evans Blue dye on day 8 pi showed a higher degree of brain whitening in C57BL/6 mice  could be within 2000\u00a0mg/kg resulting to a possible therapeutic index (TI) value of >400, considering the oral lethal dose-50 to that of effective dose-50 (ED50) (i.e. LD50/ED50) ratio. The ED50 had previously been determined orally during in vivo antiplasmodial evaluation for the\u00a0trioxaquine using the 4-day suppressive test [Acute oral toxicity of trioxaquine was evaluated by using uninfected female mice that had been starved for 4\u00a0h, administered with oral dosages of 300 and 2000\u00a0mg/kg once. The test animals were then observed for 14\u00a0days post-dosing, basing the assessment on signs of severity, and/or death observed with test animals. All the animals did not show any sign of severity or toxicity as no changes in skin and fur, eyes and mucous membranes, as well as aberant\u00a0respiratory activities were observed; also no tremors, convulsions, salivation, diarrhoea, sleep, and coma were observed. The mice survived past the 14\u00a0days of observation with 67% survival recorded with the dose of 2000\u00a0mg/kg. This is an indication that the exact lethal dose-50  was observed in the first 24\u00a0h post-treatment with artesunate and no parasite was observable under microscope 48\u00a0h post-treatment, recrudescence was observed in this treatment group with all the animals succumbing to the infection by day 16 post-treatment. Even though the quinoline moiety, on the other hand, did not show any significant parasite reduction in the first 24\u00a0h post-treatment, its contribution in trioxaquine was manifested by lack of recrudescence in trioxaquine-treated animals even beyond 60\u00a0days post-treatment. The quinoline pharmachophore  is believed to promote dual drug accumulation in the food vacuole, allowing the artemisinin-based partner (artesunate) to have a longer half-life , a possiHowever, albeit in blood-stage infection, it was earlier reported that trioxaquines were capable of inhibiting parasites in mice and also affording curative effect in both the 4-day suppressive test and in established infection studies .P. falciparum-infected RBCs (PfRBCs) cytoadhere to the brain endothelium by binding of P. falciparum erythrocyte membrane protein 1 (PfEMP1), a specific cell-surface ligand expressed by iRBCs [Two components have been implicated to be participating in the development of cerebral malaria, namely, the parasite-related factors and the host immune factors. Rupture of the hepatocytes leads to accumulation of the iRBCs within the brain microvasculature as the parasite invades the blood stream . On the by iRBCs . The seqby iRBCs . This cyby iRBCs . All theby iRBCs . The obsThe BBB is comprised of specialized endothelial cells (ECs) that function as a selective permeable membrane to control nutrients and ion transport into the brain and to bar unwanted molecules or compounds into the brain , 36. The50 could be within 2000\u00a0mg/kg. However, it should be noted that this is a dosage 40 times the curative dose of 25\u00a0mg/kg. Considering the possible LD50/ED50 ratio, a possible TI value >400 reflects a wider margin of safety for the\u00a0trioxaquine. The same safety profile was manifested in in vitro cytotoxicity studies where the selectivity index (SI) of\u00a0the trioxaquine was determined by comparing the IC50s of Hep2 cell line and that of the CQ-resistant parasite (W2) for the\u00a0trioxaquine [Good safety profile was exhibited as the drug was well tolerated in mice even at high dose of 2000\u00a0mg/kg administered in an acute oral toxicity test, with 67% survival. The remarkable mice survival indicates that the exact LDoxaquine . The higoxaquine . A similoxaquine . The higThe remarkable\u00a0efficacy and good safety profile of the\u00a0trioxaquine observed imply that if the findings could be replicated in clinical studies, then the drug has potential for use in management of CM.The remarkable in vivo results for the trioxaquine obtained by the established infection test, as well as the post-treatment survival data, clearly indicate that covalent bitherapy is a viable strategy for development of future anti-malarial agents for management of CM. An encouraging safety profile of the drug in mice with a therapeutic index (TI) value of >400 is not surprising since the precursors are part of drugs already indicated for clinical use. Overall therefore, the curative effect together with good safety profiles observed with\u00a0the trioxaquine clearly demonstrate its potential as a drug candidate for management of CM, especially in a time of shrinking anti-malarial armamentarium for management of CM."}
+{"text": "Recently, haemolytic anaemia occurring 1 to 3\u00a0weeks after artesunate treatment of falciparum malaria and parasite densities over 100,000/\u03bcL were randomly allocated to intravenous artesunate or quinine, hospitalized for 3\u00a0days and then followed for 42\u00a0days.To assess these potential safety concerns in African children, in whom most deaths from malaria occur, an open-labelled, randomized controlled trial was conducted in Kinshasa, Democratic Republic of Congo. 217 children aged between 6\u00a0months and 14\u00a0years with acute uncomplicated p\u00a0=\u00a00.009). This was explained by greater pitting then recirculation of once infected erythrocytes. Only 5% of patients (in both groups) had a\u00a0\u2265\u00a010% reduction in haemoglobin after day 7 (p\u00a0=\u00a00.1). One artesunate treated patient with suspected concomitant sepsis had a protracted clinical course and required a blood transfusion on day 14.The immediate reduction in haemoglobin was less with artesunate than with quinine: median (IQR) fall at 72\u00a0h 1.4\u00a0g/dL (0.90\u20131.95) vs. 1.7\u00a0g/dL (1.10\u20132.40) The online version of this article (doi:10.1186/s12879-017-2678-0) contains supplementary material, which is available to authorized users. This has raised general concerns about the safety of these life-saving compounds. Most cases followed intravenous artesunate, although some were reported after intramuscular artemether, intrarectal artesunate and oral artemisinin derivatives  and ACR assay .PfHRP2 by ELISA  and plasma lactate dehydrogenase  was performed in Bangkok, Thailand. Laboratory technicians were blinded to study treatments.Quantification of plasma The prespecified primary endpoint was the proportion of patients who developed late anaemia, defined as a reduction of \u226510% in haemoglobin compared with any previous measurement between days 7 and 42 after starting parenteral antimalarial treatment. Secondary endpoints included: the proportions of patients requiring blood transfusion, the fractional decreases in haemoglobin in the first week of treatment, plasma LDH as a measure of haemolysis, reticulocyte and pitted erythrocyte counts, parasite clearance times, and frequency of serious adverse events.The proportion of intravenous artesunate treated patients with delayed anaemia was assumed to be 20% . To provp) assessed associations between continuous variables. For more than two groups, the Kruskal-Wallis test was used. AUCs were estimated using the trapezoid rule. Hemoglobin, oi-RBC and parasite count values were censored on the day of malaria recrudescence or blood transfusion.Comparisons between groups used the Student\u2019s t-test or Mann-Whitney U test, depending on distributions. Kendall\u2019s Tau (\u03c4) or Pearson\u2019s correlation coefficient (rho50), the parasite clearance half-life (PC1/2)  reticulocytes. Fixed covariates included sex, age, homozygous/hemizygous G6PD deficiency, sickle cell disease trait, [log] parasitaemia on admission, splenomegaly on admission, malnutrition  and study drug treatment. Two models were considered: the first to evaluate associations with haemoglobin changes from day 0 to 7 (which included Hb at hour 0 as a covariate), the second to evaluate associations from day 7 to 28.To quantify associations with significant predictors of haemoglobin concentration, linear regression was used with random effects fitted for both the intercepts and slopes of Hb, [log] ClinicalTrials.gov; Identifier: NCT02092766).Safety reporting was performed according to the ICH Harmonized Tripartite Guideline for GCP (1996). The study was approved by the Oxford Tropical Medicine Ethics Committee and the University of Kinshasa, School of Public Health institutional review board. Data were entered using MACRO InferMed and analysed using STATA v14.0. (p\u00a0=\u00a00.80)  and correlated inversely with age . Parasites cleared significantly faster in patients treated with AS compared to QN: PC50 median (range) 5.1\u00a0h (0.08\u201315) vs. QN 15\u00a0h (0.04\u201328.6), p\u00a0=\u00a00.0001; PC1/2 median (range) AS 1.8\u00a0h (1.0\u20133.2) vs. QN 2.8\u00a0h (1.0\u20139.7), p\u00a0=\u00a00.0001. The median (IQR) PCT was 2\u00a0days in the AS arm (1\u20132) vs. 3\u00a0days in the QN arm (2\u20133), p\u00a0<\u00a00.001.Admission geometric mean parasitaemias were similar in the two treatment groups .The oi-RBC values: the initial decline was similar between treatments with a half-life of 2.0 (95% CI 1.7\u20132.3) days. The terminal elimination phase began around 2\u00a0days after peak values with a half-life which was shorter following AS than QN: 10.0 (95% CI 9.0\u201311.1) days vs.14.8 (95% CI 12.6\u201318.0) days (p\u00a0<\u00a00.001).A first-order biexponential decay best described the decline in p\u00a0=\u00a00.36, p<0.001). Malnourished children had a significantly lower mean (SD) Hb 9.7\u00a0g/dL (1.7) vs. 10.4\u00a0g/dL (1.6); p\u00a0=\u00a00.006. Children homozygous/hemizygous for G6PD deficiency (N=22) also had a lower mean (SD) values, 9.7\u00a0g/dL (1.4) vs. 10.3\u00a0g/dL (1.7), although the difference was not significant (p=0.12).On admission 74% of children were anaemic according to WHO cut-offs by age-group : 23% milFour patients received blood transfusions, two in the first 12\u00a0h (QN), one at day 5 (AS) and one at day 14 (AS). The single case given a late blood transfusion was a 6-year-old boy admitted with a parasitaemia of 145,068/\u03bcL which increased abruptly to 887,992/\u03bcL in the 25\u00a0min interval to the start of treatment (suggesting highly synchronous schizogony). The following day the patient deteriorated, was unable to sit unaided and became deeply jaundiced; antibiotics were started for suspected bacterial sepsis. During two-weeks\u2019 hospitalisation, his condition slowly improved, but fever persisted and the antibiotics were changed to ceftriaxone. His haemoglobin declined steadily from 11.0\u00a0g/dL on admission to 4.7\u00a0g/dL on day 14, with a reticulocyte count of 9.6% and plasma LDH of 2126\u00a0U/L. The patient was given a blood transfusion and discharged. He was clinically well when reviewed a week later.p\u00a0=\u00a00.036), and 1.7\u00a0g/dL (1.10\u20132.40) vs. 1.4\u00a0g/dL (0.90\u20131.95) by 72\u00a0h (p\u00a0=\u00a00.009). However, by day 7 the haemoglobin values were similar in the two treatment arms  1.5\u00a0g/dL (0.90\u20132.20) vs. 1.3\u00a0g/dL (0.80\u20131.90) by 48\u00a0h  and at day 7 in 10% . The median (IQR) interval to Hb nadir was 48\u00a0h (36\u201372) with AS and 60\u00a0h (48\u201372) with QN, p\u00a0=\u00a00.192. The mean (SD) Hb nadir values before day 7 were similar, AS\u00a0=\u00a08.3 (1.4) g/dL vs. QN\u00a0=\u00a08.1 (1.6) g/dL, p\u00a0=\u00a00.395, as were the nadirs observed at day 7, AS\u00a0=\u00a09.2 (1.1) g/dL vs. QN\u00a0=\u00a09.4 (1.2) g/dL, p\u00a0=\u00a00.736.The Hb nadir was observed before day 7 in 88% of patients (p\u00a0<\u00a00.001. There were no significant differences after day 3.Reticulocyte counts (haematocrit corrected) increased until day 7 and then decreased gradually in both arms Fig. . The coup\u00a0=\u00a00.103; Additional file The primary endpoint, defined a-priori as a reduction of \u226510% in the Hb level at any time between days 7 and 42, was observed in 19/207 patients: 10 children treated with AS, whose median Hb fractional reduction was 13.7%  and 9 treated with QN, whose median Hb fractional reduction was 12.8% . In 15 patients (7.3%) the Hb remained below the admission value throughout the follow-up  and in 4 patients (2%) the nadir occurred after day 7 .In the cohort of patients who completed the 42-days follow-up, the Hb concentration increased steadily and rose above the admission level in 92.7% of cases . The median (range) maximum LDH concentration was 688 (328\u20132215) U/L with AS and 703 (375\u20131664) U/L with QN.Between day 0 and 7 the median plasma LDH concentrations (U*days/L) decreased by similar amounts in the two groups Fig. : AUCs ASp\u00a0<\u00a00.05, Fig. p\u00a0=\u00a00.005; and between day 14 and 21, 24.1% in the QN arm (12.5\u201335.3) and 17.9% in the AS arm (6.4\u201332.4); p\u00a0=\u00a00.12. The median LDH AUC was slightly smaller with QN compared to AS  and was positively correlated with the maximum oi-RBC count with AS (p\u00a0=\u00a00.045), but not QN (p\u00a0=\u00a00.263).From day 7 onward the median LDH concentration decreased more in patients who received QN (p\u00a0=\u00a00.107).Some patients had transient fluctuations in the plasma LDH level during the follow-up period. A significant increase (above twice the paediatric reference range ULN ) was obsp\u00a0=\u00a00.045), had higher admission parasitaemias (p\u00a0=\u00a00.0042), higher maximum oi-RBC counts (p\u00a0=\u00a00.0150), higher reticulocyte counts (p\u00a0=\u00a00.0102) and lower Hb at days 14 and 21 . In the QN arm there were no differences in these parameters between patients with and without significant LDH elevations, although oi-RBC counts were higher in those with normal LDH (p\u00a0=\u00a00.0132).Children who received AS and later had a significantly elevated plasma LDH concentration were younger (p\u00a0=\u00a00.087) and only LDH, [log] oi-RBC and [log] reticulocytes were associated significantly with haemoglobin changes  lower Hb values from day 0 to 7, those with palpable splenomegaly had 0.431\u00a0g/dL (95% CI 0.113\u20130.749) lower Hb and malnourished children had 0.379\u00a0g/dL lower Hb levels (95% CI 0.024\u20130.734). Older children had higher Hb levels, 0.111\u00a0g/dL (95% CI 0.055\u20130.161) per year of age. The effect of treatment was borderline: Hb levels in children treated with AS were, on average, 0.312\u00a0g/dL (95% CI 0.002\u20130.627) higher from day 0 to 7, but thereafter the effect of treatment was no longer significant  have a much shorter survival time (7\u201314\u00a0days) compared with normal erythrocytes in healthy subjects (120\u00a0days) or in patients following severe malaria (44\u00a0days) [oi-RBC following artesunate probably reflects drug killing and then pitting of developed ring forms whereas background (and quinine associated) pitting may only occur for very young ring stages shortly after merozoite invasion with correspondingly less damage to the erythrocytes. The rapid and synchronous elimination of these oi-RBC from the circulation 1 to 2\u00a0weeks after the start of antimalarial treatment might result in haemolytic anaemia [Post-artesunate haemolytic anaemia has been attributed to the increased pitting of drug-damaged malaria parasites from infected erythrocytes . PittingRBC Fig. . These o44\u00a0days) , 19. The anaemia .oi-RBC to the initial parasitaemia (59%) was comparable to that observed in Malian children (60%) following artesunate, but lower than in returned travellers (78%) [oi-RBCs were left at this stage, thereby reducing the number of erythrocytes contributing to late haemolysis. These findings suggest that other, presumably immune mediated, mechanisms leading to infected cell removal contribute to splenic clearance of parasitaemia in African children. The erythropoietic response to malarial anaemia was rapid (peak 7\u00a0days) [oi-RBC, whilst more rapid clearance of pitted cells reduced haemolysis occurring after 7\u00a0days. Only one artesunate treated patient had haemolytic anaemia severe enough to warrant blood transfusion. This patient had a complicated disease course with suspected concomitant bacterial sepsis treated. Whether the haemolysis in this case was explained only by malaria and artesunate, or whether the presumed sepsis contributed cannot be determined.In this study, a similar proportion of artesunate and quinine treated patients had a\u00a0\u2265\u00a010% reduction in haemoglobin, but the degree of anaemia was moderate, although there are small but significant differences in the time course. Patients receiving artesunate had slightly lower initial reductions in haemoglobin and delayed reticulocyte responses. Previous studies also indicated a clinically insignificant temporary suppression of reticulocytosis by artemisinins in the treatment of malaria , 21. Morrs (78%) . Moreove 7\u00a0days) , and too 7\u00a0days) , 19, 23)falciparum malaria and high parasitaemias, which is similar to the frequency reported in West African children with severe malaria [PfHRP2 concentrations on admission, an indicator of parasite burden, were between the values reported for African children with uncomplicated and severe malaria .falciparum in African children.In conclusion, severe delayed haemolytic anaemia was rare in this population of African children with high parasitaemias. This reassuring evidence contrasts with recent reports form returned travellers and it strongly supports the wider deployment of life-saving parenteral artesunate for the treatment of severe Additional file 1: Table S1.Table S2. Median and range oi-RBCs/\u03bcL of blood by day and treatment arm. Table S3. Mean, SD and range of haemoglobin 0\u201342\u00a0days by treatment arm. Table S4. Linear regression using Hb (g/dL) as outcome and patient as random effect for days 0\u20137. Table S5. Linear regression using Hb (g/dL) as outcome and patient as random effect for days 7\u201328. (DOCX 24\u00a0kb) Efficacy outcome by treatment arm PCR uncorrected, Per Protocol. Additional file 2: Figure S1.n = 9 (TIFF 24\u00a0kb) Individual haemoglobin profiles of patients in the QN group with \u226510% reduction after day 7, Additional file 3: Figure S2.n =10 (TIFF 64\u00a0kb) Individual haemoglobin profiles of patients in the AS group with \u226510% reduction after day 7,"}
+{"text": "Plasmodium falciparum in mining areas of the interior of Guyana, a 7-day artesunate trial was conducted from March to December 2014. The day-3 parasite clearance rate, the efficacy of artesunate at day 28, and polymorphism of Kelch 13 (PfK13)\u2014the marker of artemisinin resistance\u2014were assessed. The study confirmed the continued sensitivity of P falciparum to artemisinin. A 7-day course of artesunate was 100% efficacious with only 2%  of enrolled subjects positive at day 3. All day-0 parasite samples were wild type. Continued resistance monitoring is nevertheless recommended, given the widespread availability and uncontrolled use of artemisinin drugs in mining areas of Guyana.Because of concerns about possible emergence of artemisinin resistance strains of  Plasmodium falciparum malaria in almost all malaria-endemic countries. In conjunction with long-lasting insecticidal bed nets, the use of these drugs has played a major part in the dramatic reduction in the global burden of malaria observed since 2000 [Over the past 15 years, artemisinin-based combination therapies (ACTs) have become the first-line treatment for PfK13) propeller region have been identified as a molecular marker of artemisinin resistance, being associated with delayed parasite clearance both in vitro and in vivo [Artemisinin resistance has been detected in 5 countries in the Greater Mekong Subregion . Artemisinin resistance is characterized by delayed parasite clearance after treatment with an artemisinin monotherapy, or ACT . For ACT in vivo . In the PfK13 mutations), a confirmatory study using artesunate monotherapy should be conducted, complemented by sequencing of the PfK13 gene of the parasite samples in patients enrolled on day 0 as recommended by the Technical Expert Group on Drug Efficacy and Response .The World Health Organization (WHO) recommends efficacy monitoring for first-line and second-line ACTs every 2 to 3 years in all endemic countries. Such studies aim to determine (1) the proportion of malaria cases with persistent parasitemia at day 3 after treatment with an ACT and (2) the rate of treatment failures at days 28 or 42. In the case of suspected artemisinin resistance  and one who traveled to region 1 (Barima-Waini) of Guyana that carried the C580Y mutant allele in the PfK13 propeller domain but with flanking microsatellites different from those observed in Southeast Asia [Recent studies have raised concerns regarding the possible emergence of artemisinin resistance in the Guiana shield region of South America. In Suriname, a study conducted in 2011 reported that 15 of 48 (31%) evaluable patients treated with artemether-lumefantrine were parasitemic on day 3, compared with 2% in 2005/2006, although therapeutic efficacy was 100% at day 28 and P falciparum malaria treated with a 7-day course of artesunate monotherapy. Samples from the enrolled patients were also analyzed to assess the association of PfK13 polymorphisms with clinical response.The primary objective of this study was to assess the possible emergence of artemisinin resistance in Guyana by determining the day-3 parasite clearance rate in patients with uncomplicated http://www.health.gov.gy/moph/), ACTRN12616000526471.This was a one-arm prospective survey conducted from March to December 2014 in patients recruited in the Malaria Clinic at the Georgetown Public Hospital Corporation, Georgetown, Guyana. Patients from all over the country can attend the Malaria Clinic, but the majority comes from gold mining areas in the country's interior. All subjects provided written informed consent. Ethical approval was provided by the Ministry of Health in Guyana, after review by its Institutional Review Board (no. 158). The study is registered at The Ministry of Public Health of Guyana  and were >2 years old and able to swallow oral medication. Exclusion criteria were severe malaria, severe malnutrition, febrile condition other than malaria, known drug hypersensitivity, and pregnancy or breast-feeding. A pregnancy test was required for all women of childbearing age.Eligible subjects were those attending the clinic with microscopically confirmed, uncomplicated To facilitate follow up, enrolled patients were housed in a hotel near the Malaria Clinic for days 0\u20137 and received directly observed treatment with artesunate (4 mg/kg body weight) once daily for 7 days followed by a single dose of primaquine (0.5 mg/kg) on day 8, with out-patient follow up to day 28. Artesunate was provided as 50-mg tablets  and primaquine as 15-mg tablets . Artesunate was quality controlled by North-West University in South Africa. Any patient vomiting within 30 minutes of dosing was retreated; repeat vomiting triggered study withdrawal and rescue treatment with injectable quinine 3 \u00d7 10 mg salt/kg per day. Failures were treated with artemether-lumefantrine.A complete medical history and demographic information were taken at enrollment. All patients received a clinical assessment at enrollment, daily on days 1\u20137 and on days 14, 21, and 28. Adverse events were recorded throughout the study.Duplicate Giemsa-stained thick and thin blood films were obtained at the time of initial screening (day 0), every 8 hours for 7 days after the initial dose of artesunate, and on days 14, 21, and 28. Parasite densities were determined as per WHO procedures by 2 independent microscopists [P falciparum strains was also explored by examining polymorphisms in the PfK13 propeller domain at day 0 using a nested PCR protocol followed by Sanger sequencing using primers specific for P falciparum. The amplicon used for sequencing covered 656 base pairs (amino acids 49\u2013705), which included the K13 propeller domain as recommended by WHO [Therapeutic efficacy was evaluated as per WHO guidelines using adequate clinical and parasitological response (ACPR), ie, absence of parasitemia without previous treatment failure . To diffThe required study sample size of 35 patients was calculated based on an estimated prevalence of parasitemic patients at day 3 after artesunate therapy >10% at a confidence level of 95%. Adding 20% to allow for study withdrawals gave a target sample size of 42 patients.http://www.who.int/malaria/areas/drug_resistance/en/.Study efficacy outcomes were the proportion of patients' who still had parasites in their blood smears on day 3 and ACPR at day 28. Parasite clearance time and slope half-life were calculated using the WHO parasite clearance estimator. Safety outcomes were the incidence of adverse events. Data analysis was performed using a Microsoft Excel spreadsheet provided by WHO. Both are available at P vivax infection. Most patients were male (41 of 50) and over 15 years of age (49 of 50). Mean age was 30.6 years , mean weight was 66.7 kg , mean temperature was 37.2\u00b0C , and geometric mean parasitemia was 6281 \u00b5L . The patients came from 3 of the 4 regions in Guyana where malaria remains endemic: region 1 (Barima-Waini) (26%), region 7 (Cuyuni-Mazaruni) (54%), and region 8 (Potaro-Siparuni) (16%). One patient had been living in region 9, and 1 patient originated from Venezuela. Parasitemia at day 3 was observed in 1 of 49 patients  with a median parasite clearance time of 38.5 hours . This patient did not have any outstanding feature that differed from the other patients. Median slope half-life (range) evaluated in 45 patients was 2.97 hours (1.09\u20135.9 hours), with 2 patients only presenting with a slope half-life >5 hours. In 47 evaluable patients, ACPR at day 28 was 100% . Because there were no recrudescences, PCR adjustment was not required. All 50 patients enrolled in the study, including the patient presenting with parasites at day 3, had day-0 P falciparum samples that were PfK13 wild type in the propeller domain. No serious adverse events were reported in the study. Two mutations were observed outside the propeller region: D80G in all the samples and K189T in 35.7% of them.Fifty patients were enrolled from June to November 2014. All enrolled patients complied fully with the study procedures and protocols. There were 3 withdrawals, 2 on day 21 and 1 on day 28; all were triggered by P falciparum in 2004. Efficacy of this combination was high after implementation and no delayed clearance was reported at that time. There are no recent therapeutic efficacy data for Guyana, as a trial conducted with artemether-lumefantrine in 2011\u20132012 failed to meet quality-control requirements [Guyana adopted artemether-lumefantrine plus primaquine as the first-line treatment for PfK13 gene.The Guiana shield region is an area of high population mobility due to mining activities where uncontrolled use of antimalarial drugs, including many artemisinin drugs, is widespread. Concern regarding the emergence of artemisinin resistance justified further investigation at the country level. This study conducted in 2014 by the malaria control program in Guyana did not confirm the presence of artemisinin resistance. None of the patients had delayed parasite clearance, and all parasites were wild type in the propeller domain of the PfK13 mutants in parasites collected in 2010 [PfK13 mutants in regions 7 and 8, because the results of the 2010 survey reporting C580Y mutants in very low frequency (5 samples of 98) were published after the 2014 clinical study was terminated. Although 70% of the patients included in the 2014 study were infected in regions 7 and 8, the probability of finding a C580Y mutant during this clinical trial including 50 patients only was very low. The working definition of suspected artemisinin resistance set by WHO considers that a \u22655% prevalence of a single PfK13 mutant signifies selection of the genotype in the parasite population, which requires further investigation. Clonal expansion under drug pressure in the presence of a mobile migrant population attracted by mining activities would have most probably led to an increase in the prevalence of mutants and regional spread over the last 5 years. So far, no C580Y mutants have been reported in neighboring countries [PfK13 mutants without clonal expansion and spread has been reported in Africa and seems to be related to natural polymorphism of the PfK13 gene in a parasite population rich in low-frequency alleles [PfK13 mutants. Previous antimalarial treatment is a well known factor that selects for resistant parasites for both artemisinin and partner drug [PfK13 mutation in Guyana.Several reasons could explain the discrepancy between this 2014 clinical study and the detection of  in 2010 . The obj alleles . Finallyner drug . An addiIn conclusion, this study did not confirm the presence of artemisinin resistance in Guyana, but continuous surveillance is necessary."}
+{"text": "Malaria remains one of the most important infectious diseases. Treatment options for severe malaria are limited and the choline analogue SAR97276A is a novel chemical entity that was developed primarily as treatment for severe malaria. Before starting clinical investigations in severely ill malaria patients, safety and efficacy of SAR97276A was studied in patients with uncomplicated malaria. Here, we summarize two open-label, multi-center phase 2 trials assessing safety and efficacy of parenterally administered SAR97276A in African adults and children with falciparum malaria.Study 1 was conducted in Burkina Faso, Gabon, Benin and Tanzania between August 2008 and July 2009 in malaria patients in an age de-escalating design . A total of 113 malaria patients received SAR97276A. Adults were randomized to receive a single dose SAR97296A given either intramuscularly (IM) (0.18\u00a0mg/kg) or intravenously (IV) (0.14\u00a0mg/kg). If a single dose was not efficacious a second adult group was planned to test a three dose regimen administered IM once daily for 3\u00a0days. Single dose SAR97276A showed insufficient efficacy in adults . The 3-day IM regimen showed acceptable efficacy in adults  but not in children . SAR97276A was well tolerated but no further groups were recruited due lack of efficacy. Study 2 was conducted between October 2011 and January 2012 in Burkina Faso, Gabon and Kenya. SAR97276A administered at a higher dose given IM was compared to artemether\u2013lumefantrine. The study population was restricted to underage malaria patients to be subsequently enrolled in two age cohorts . Rescue therapy was required in all teenaged malaria patients (8/8) receiving SAR97276A once daily (0.5\u00a0mg/kg) for 3\u00a0days and in 5 out of 8 teenaged patients treated twice daily (0.25\u00a0mg/kg) for 3\u00a0days. All patients (4/4) in the control group were cured. The study was stopped, before enrollment of children, due to lack of efficacy but the overall safety profile was good.Monotherapy with SAR97276A up to twice daily for 3\u00a0days is not an efficacious treatment for falciparum malaria. SAR97276A will not be further developed for the treatment of malaria.Trial registration at clinicaltrials.gov: NCT00739206, retrospectively registered August 20, 2008 for Study 1 and NCT01445938 registered September 26, 2011 for Study 2.The online version of this article (doi:10.1186/s12936-017-1832-x) contains supplementary material, which is available to authorized users. Plasmodium remains one of the most important infectious diseases. Plasmodium falciparum is the species responsible for nearly all severe malaria cases and deaths  or Group 2C (4 teenagers received artemether\u2013lumefantrine treatment). See Fig.\u00a0In All (8/8) patients in Group 2A and 5/8 in Group 2B received rescue treatment during the trial, whereas all patients (4/4) of the control Group 2C were cured. 7/8 patients in Group 2A were classified as treatment failures (4 early and 3 late failures) and one patient having hepatitis B could not be evaluated according to the predefined definitions. 5/8 participants were treatment failures in Group 2B group (4 early and 1 late failure), none in the control Group 2C group was classified as treatment failure. Median PRR at 72\u00a0h was 22.5 (0.4\u20136280.0) and 12.0 (0.3\u201383.1) in Group 2A and Group 2B, respectively. Group 2C patients had a PRR of 2562.2 (400.6\u20135711.8). Increase of parasitemia between 6 and 12\u00a0h after the start of treatment was observed in Group 2A, which was not seen in the other groups. Median parasite clearance time was 144\u00a0h (CI 30\u2013144\u00a0h) for Group 2A, 144\u00a0h (CI 30\u2013144\u00a0h) for 2B, and 27\u00a0h (CI 18\u201366\u00a0h) for the 2C group. The trial was stopped after recruitment of the first 16 participants due to lack of efficacy.The most frequent TEAE was malaria in both SAR97276A treatment groups and neutropenia in Group 2B followed by gastrointestinal disorders, especially diarrhea, in Group 2A , with interindividual variability on clearances and central volume of distribution.Descriptive statistics of the individual pharmacokinetic parameters for the different groups are presented in Table\u00a0In Study 2 AUC values were similar between the two different regimens of SAR97276A and Cmax was approximately 1.8-fold lower in Group 2A. Interindividual variability of pharmacokinetic parameters was low.Results of these two consecutive phase 2 studies showed that SAR97276A monotherapy is not efficacious to cure uncomplicated falciparum malaria in children. WHO recommends a cure rate of 95% assessed in clinical trials to advance clinical development of a new antimalarial towards licensure . The priSAR97276A showed a reasonable safety profile in both studies. Deterioration of malaria was indirectly due to SAR97276A as it was not efficacious enough to control parasitemia and malaria symptoms.The new mechanism of action of SAR97276A could make it an interesting partner for combination therapy with another strong malaria medication. However, as oral availability is low it has to be given parenterally and is therefore not a good candidate compound for treatment of uncomplicated malaria as this should be treated orally. Compounds against severe malaria should have a rapid onset of action, which cannot be seen by SAR97276A in the current studies. SAR97276A does not show a post-treatment prophylactic effect reflected by recurrent as well as new infections until day 28. As a secondary endpoint the parasite clearance time of SAR97276A was evaluated. When compared to artemisinins, the licensed drug class with the fastest clearance of parasites , 16, SARSAR97276A will not be further developed. However, compounds acting against phospholipid biosynthesis remain potentially interesting with a mode of action different to other antimalarials currently in use.Additional file 1. Detailed inclusion and exclusion criteria of Study 1 and Study 2.Additional file 2. Preparation of the investigational product SAR9727A (Study 1).Additional file 3. Definition of early and late treatment failure (Study 2).Additional file 4. Details on sample size calculation (Study 2).Additional file 5. Results of hematology, electrocardiography and vital signs (Study 1). Table S1. Electrocardiogram abnormalities during TEAE period (Study1).Additional file 6. ECG parameters (Study 2). Table S2. Electrocardiogram abnormalities during TEAE period (Study2). Table S3. Respiratory parameters (Study 2)."}
+{"text": "A recent randomized trial showed that artemisinin\u2013naphthoquine (AN) was non-inferior to artemether\u2013lumefantrine (AL) for falciparum malaria and superior for vivax malaria in young Papua New Guinean children. The aim of this study was to compare the cost-effectiveness of these two regimens.Plasmodium falciparum and/or Plasmodium vivax infections in an open-label, randomized, parallel-group trial. Recruited children were randomized 1:1 to receive once daily AN for 3\u00a0days with water or twice daily AL for 3\u00a0days given with fat. World Health Organisation (WHO) definitions were used to determine clinical/parasitological outcomes. The cost of transport between the home and clinic, plus direct health-care costs, served as a basis for determining each regimen\u2019s incremental cost per incremental treatment success relative to AL by Day 42 and its cost per life year saved.An incremental cost-effectiveness analysis was performed using data from 231 children with In the usual care setting, AN was more effective for the treatment of uncomplicated malaria in children aged 0.5\u20135.9\u00a0years. AL and AN were equally efficacious for the treatment of falciparum malaria, however AN had increased anti-malarial treatment costs per patient of $10.46, compared with AL. AN was the most effective regimen for treatment of vivax malaria, but had increased treatment costs of $14.83 per treatment success compared with AL.Whilst AN has superior overall efficacy for the treatment of uncomplicated malaria in PNG children, AL was the less costly regimen. An indicative extrapolation estimated the cost per life year saved by using AN instead of AL to treat uncomplicated malaria to be $12,165 for girls and $12,469 for boys (discounted), which means AN may not be cost-effective and affordable for PNG at current cost. However, AN may become acceptable should it become WHO prequalified and/or should donated/subsidized drug supply become available. Although malaria remains a major global health issue with nearly half the world\u2019s population still at risk and ongoing transmission in 91 countries, substantial progress has been made towards reducing the burden of the disease . This imPlasmodium species [Plasmodium falciparum and Plasmodium vivax infections [Artemisinin-based combination therapy (ACT) is the World Health Organization (WHO) recommended first-line treatment for uncomplicated malaria . Five di species . Papua Nfections . These cfections  and costfections  ACT for fections , 5. Givefections , a searc1/2\u00a0=\u00a023\u00a0days) [P. vivax emergence than the piperaquine component of DHA\u2013PQP.Of the few potential alternative ACT, artemisinin\u2013naphthoquine (AN) was one candidate. This ACT was already available in the private sector in PNG as single-dose treatment for uncomplicated malaria despite not being prequalified. Preliminary pharmacokinetic and safety studies in PNG children provided good evidence that the recommended single AN dose given daily for 3\u00a0days  would b23\u00a0days)  likely tP. falciparum and P. vivax infections in coastal Madang Province [A randomized comparative safety and efficacy trial of AL and AN was, therefore, performed in young PNG children with uncomplicated Province . The resProvince , 13, whiBetween 28 March 2011 and 22 April 2013, an open-label, randomized, parallel-group trial of AL and AN was conducted at the Mugil and Alexishafen Health Centres . The trial was approved by the PNG Institute of Medical Research Review Board, the Medical Research Advisory Committee of PNG, and the University of Western Australia Human Research Ethics Committee. Children aged 0.5\u20135.9\u00a0years presenting with blood slide positive uncomplicated falciparum or vivax malaria were eligible to participate. Full details of study procedures have been published [An initial standardized clinical assessment including measurement of the axillary temperature was performed and blood was drawn for blood film microscopy, measurement of haemoglobin (Hb) and blood glucose, hepatic and renal function, and a full blood count . A modifP. vivax recrudescence was determined by genotyping [Efficacy was assessed using WHO definitions , specifinotyping . No chilP. falciparum and remaining life expectancy.The perspective of the present analyses was societal. Direct health care costs  plus travel costs were estimated. An incremental cost-effectiveness analysis was performed in which the net costs and net effectiveness of AN were compared with those of conventional AL treatment and expressed as ratios. All analyses and comparisons were performed on both a per protocol (PP) and modified intention to treat (mITT) basis. The PP analyses included children with complete follow-up or confirmed treatment failure, and excluded those treated for malaria without confirmatory microscopy or who defaulted from follow-up. These excluded patients were retained in the mITT which utilized i) a worst-case approach  and ii) a best-case approach . In a secondary analysis, we extrapolated outcomes to estimate the increase in life expectancy of the most effective treatment based on estimated mortality associated with For each patient, standardized data were collected at each scheduled clinic visit and at extra, unscheduled \u201csick day\u201d visits. These included doses of all drugs used for treating malaria, its symptoms and its complications . Unit costs were obtained from the PNG National Department of Health , InterpaAll study participants were scheduled to attend eight times including Day 0 but excluding sick days. However, the frequency and type of visits are different in a usual care  situation. Therefore, a complementary analysis, in which costs of AL or AN treatment are based on a single clinic visit at which malaria is diagnosed and treated, was conducted. The same number of subsequent sick day visits was assumed except for ETFs where the scheduled Day 1\u20133 clinic visit was replaced by a sick day visit. Each patient\u2019s actual trial visit costs were replaced by the estimated standard practice cost depending on allocation. It was assumed that no-cost forms of fat  were consumed with AL.P. falciparum was performed by extrapolating the benefits using lifetables to estimate the potential life years saved. All comparisons were carried out separately for P. falciparum and P. vivax on both a PP and a mITT basis. All results are reported as means and SDs or mean differences and 95% CIs. The CIs for the key incremental cost-effectiveness ratios (ICERs) were estimated using the bootstrap approach with 1000 repeated random samples drawn with replacement from the original data. Bootstrap confidence intervals were constructed with the bias-corrected percentile method [The primary endpoint of the trial was treatment failure by Day 42 . A secone method . The effP. falciparum analysis were of mean\u00a0\u00b1\u00a0SD age 3.7\u00a0\u00b1\u00a01.3 (range 0.8\u20135.9) years and 53.8% were boys, while 41.3% of the 46 children in the PP P. vivax analysis were boys and the mean age was 3.1\u00a0\u00b1\u00a01.2 (range 0.5\u20135.8) years.The 186 children in the PP P. falciparum and P. vivax in both trial and usual care settings. For P. falciparum, AN had increased anti-malarial treatment costs for each patient of $10.46 (95% CI $9.77\u2013$11.16) on average compared with AL. There were no significant differences between AL and AN in other costs. Total usual care costs were significantly higher in the AN group compared with the AL group.Table\u00a0P. vivax infections, AN increased anti-malarial treatment costs for each patient by an average of $11.05 (95% CI $9.93\u2013$12.16) compared with AL  had an ACPR with the highest rate in the AN group (100%) compared with 97.8% in the AL group [P. vivax was 100% in the AN group compared with 30.0% in the AL group [In the PP AL group . For theAL group . Table\u00a03P. falciparum, the average excess cost per treatment success for AN when compared with AL was $436.82 with a 95% bootstrap bias-corrected confidence interval of $200.23\u2013$941.29 and a 68.3% chance that the incremental cost-effectiveness ratio was below $500 per treatment success. For P. vivax  and 0.055\u00a0\u00d7\u00a062.1\u00a0years for boys\u00a0=\u00a03.42\u00a0years (or 1.60\u00a0years when discounted at 3%). The extra cost associated with AN versus AL treatment was $19,590 per 1000 cases treated . Therefore, the cost per life year saved was $19,590/3.66\u00a0=\u00a0$5352.46  for girls and $19,590/3.42\u00a0=\u00a0$5728.07  for boys.A girl and boy aged between 1 and 4\u00a0years old in 2012 in PNG could expect to live another 66.6 and 62.1\u00a0years, respectively . In PNG Plasmodium species. The present data show that, while a 3-day course of AN had superior overall efficacy in uncomplicated paediatric malaria, the recommended first-line AL remained the more cost-effective treatment. In PNG children with falciparum malaria, AN was equally efficacious but far more costly than AL. In the case of vivax malaria, AN was significantly more efficacious than AL but only slightly more costly in the usual care setting. The most significant contributor to the difference in costs between AN and AL is the fact that AL is a subsidized prequalified first-line therapy provided with financial assistance from a Global Fund Affordable Medicines Facility Grant [This is the first economic analysis of AN for treatment of uncomplicated malaria in an area of intense transmission of multiple ty Grant , while A\u00ae. It satisfies WHO recommendations for universal combination therapy for uncomplicated malaria but has not yet met the WHO prequalification manufacturing standards [\u00ae as a single-dose treatment, a regimen which is inconsistent with WHO recommendations for 3\u00a0days of ACT [\u00ae is marketed in the private sector in a range of countries in Africa, Asia and Oceania, it is important for the cost-effectiveness implications of this form of ACT to be assessed.The only formulation of AN currently available is that manufactured by KPC under the brand name ARCOtandards . As a retandards , 20. As s of ACT , future \u00ae can be purchased as a single-dose therapy across the counter at an average cost of $1.84 per tablet, which is significantly more expensive than subsidized AL therapy ($0.19 per tablet) that is usually widely available through government hospitals and clinics [In PNG, ARCO clinics , 15. Eve clinics . Dose-raFor falciparum malaria in the present study, AN and AL had similar treatment efficacy  but AL was significantly more cost-effective therapy with a mean cost of US$7.66 per patient  compared to US$17.28 per patient for AN therapy. This finding is consistent with other analyses of AL in a usual care setting. A cost-effectiveness study comparing DHA\u2013PQP and AL for uncomplicated malaria in Tanzanian children completed in 2012 reported a mean cost of US$8.40 from the provider\u2019s perspective for successful treatment of a clinical case of uncomplicated malaria . ConsideP. vivax has increased by 13\u201336% [Plasmodium species causing infection and illness in young PNG children [P. vivax mainly because of the risk of haemolysis due to the relatively high prevalence of glucose-6-phosphatase deficiency in PNG. In this situation, selection of an ACT with a long half-life component is desirable so that there is an extended period of prophylaxis against relapses from hypnozoites during recovery [P. vivax infections in contrast to the marked disparity in the case of P. falciparum infections. The excess cost per treatment success with AN was $14.83 for P. vivax versus $436.82 for P. falciparum.A further consideration for countries such as PNG is that, despite a decline in the prevalence of falciparum malaria, the prevalence of y 13\u201336% \u201324 to thchildren . Primaqurecovery . In the recovery . This rarecovery \u201329. It iP. falciparum only, 14% to P. vivax only, and 6% to mixed P. falciparum and P. vivax infections. Projecting the trial PP usual care cost-effectiveness analysis to the general PNG population of 7,154,870 in 2012, and assuming that all suspected cases were confirmed as a worst case scenario, 213,358 and 28,499 more cases of P. vivax and P. falciparum, respectively, could be treated successfully every year if AN were used instead of AL, costing an extra US$14,663,345 per year. However, these estimates may have to be revised as, since the trial was conducted in 2012, two rounds of countrywide distributions of long lasting insecticide treated nets have resulted in a 6.7% reduction in the overall prevalence of malaria [per capita in PNG in 2012 was $1820 [There are population data from PNG detailed enough to provide estimates of the benefit of ACT regimens. In 2009, 1,431,395 suspected malaria cases were treated in PNG with a reported 604 malaria-attributable deaths. This represents 21.3% of the total PNG population assuming no multiple presentations. Furthermore, a study of febrile patients presenting at five sentinel health facilities across PNG in 2008 reported a malaria slide positivity rate ranging from 2.2 to 74.9% . The cli malaria . It was as $1820 , 34, unlThe present study had limitations. The analysis did not consider treatment compliance. All treatments are given over 3\u00a0days but AL requires two doses per day and the medication should be taken with supplementary fat , aspects which might reduce compliance. Although all children allocated AL in the trial had a therapeutic plasma lumefantrine concentration on Day 7, consistent with all six doses having been administered successfully , a cost-Plasmodium infections but these were a minor component of the burden of disease . Post-treatment gametocyte carriage and consequent malaria transmission was also not factored into the analyses. Opportunity costs, such as the time taken off work by parents needing to look after their sick child including transportation to the clinic or hospital, have not been addressed. Treatment that accelerates recovery might allow parents to return to work more quickly. Finally, there are sometimes unpredictable shortages of government supplies of approved drugs such as AL in PNG which means that parents have to fill prescriptions at commercial pharmacies at increased cost.Other potential limitations were that, while uncertainty has been accounted for in the outcomes of the trial via bootstrapping, the estimates of cost per life year are meant to be indicative and so a probabilistic model that would allow quantification of uncertainty around these estimates was not built. The current analysis also did not consider the cost implications of the mixed Plasmodium species in PNG but its low efficacy against vivax malaria remains a concern. While, for programmatic reasons, the use of a single first-line therapy that is effective against both P. falciparum and P. vivax infections is preferred in regions where both species are transmitted, the superiority of AN against vivax malaria [AL proved the most cost-effective ACT treatment for both infecting  malaria , 26 mean malaria  and rela malaria . Given i malaria , 40, DHA malaria . In the"}
+{"text": "Plasmodium falciparum malaria. Despite the success of ACT in reducing the global burden of malaria, the emerging of resistance to artemisinin threatens its use.Artemisinin combination therapy (ACT) is used worldwide as the first-line treatment against uncomplicated P. falciparum malaria diagnosed in Europe. It occurred in an Italian tourist returned from Ethiopia. She completely recovered after the DHA-PPQ treatment but 32\u00a0days after the end of therapy she had a recrudescence. The retrospective analysis indicated a correct DHA-PPQ absorption and genotyping demonstrated that the same P. falciparum strain was responsible for the both episodes.This report describes the first case of failure of dihydroartemisinin-piperaquine (DHA-PPQ) for the treatment of In consideration of the growing number of cases of resistance to ACT, it is important to consider a possible recrudescence, that can manifest also several weeks after treatment. Artemisinin combination therapy (ACT) is used worldwide as the first-line treatment against uncomplicated falciparum malaria . Dihydro9/L , haemoglobin (Hb) 12.5\u00a0g/dL , platelets 46\u00a0\u00d7\u00a0109/L , C-reactive protein 135\u00a0mg/L , procalcitonin 14\u00a0\u03bcg/L . The quantitative buffy coat (QBC) test, antigen malarial test and blood smears resulted positive for Plasmodium falciparum, with a parasitaemia of 0.3% . The patient was treated with DHA-PPQ 320/40\u00a0mg, three tablets/day for 3\u00a0days. The first day after treatment the parasitaemia dropped to 0.0023% (96/\u03bcL). After two days, the blood films and the QBC test resulted negative. Also, iv ceftriaxone 2\u00a0g/day was administrated for a left basal bronchopneumonia. The patient was discharged on 5 December, 2014.A 72-year-old Italian woman was admitted on 26 November, 2014 to the Centre for Tropical Diseases (CTD) of Negrar (Verona), for myalgias and arthralgias since 2\u00a0days, fever (up to 40\u00a0\u00b0C) and nausea since one day. She had visited Ethiopia  from 6 to 18 November 2014. She was vaccinated against yellow fever, hepatitis A and B, but had not taken any malaria chemoprophylaxis. Her travel history included Ethiopia, Niger, India, and Nambia, not South East Asia. Upon admission, her temperature was 38.3\u00a0\u00b0C, her weight 67.5\u00a0kg. Physical examination was unremarkable. The blood tests showed white blood cells (WBCs) 3.85\u00a0\u00d7\u00a0109/L, Hb 11.8\u00a0g/dL, platelets 233\u00a0\u00d7\u00a0109/L.At a follow-up visit on 23 December, the QBC test and blood smears were still negative. The blood tests showed WBC 7.98\u00a0\u00d7\u00a0109/L, Hb 9.5\u00a0g/dL, platelets 96\u00a0\u00d7\u00a0109/L. The patient was treated this time with atovaquone-proguanil 250/100\u00a0mg, four tablets/day for 3\u00a0days. The parasitaemia decreased to 0.36%  24\u00a0h after first dose of treatment, 0.16% (5017/\u03bcL) the second day, 0.0016% (46/\u03bcL) the third day. After 4\u00a0days, blood films resulted negative. The patient was discharged on 12 January, 2015.She was re-admitted on 7 January, 2015 complaining of fever, nausea and vomiting that had started 7\u00a0days before . The QBC test, antigen malarial test and blood smears all resulted positive again for falciparum malaria, with a parasitaemia of 0.4% . WBCs were 5.5\u00a0\u00d7\u00a010At follow-up visits 28 and 56\u00a0days after the second malaria episode, QBC and blood smears resulted negative and the main laboratory findings were normal.The malaria-PCR performed a posteriori on a blood specimen collected on 23 December, 2014 resulted positive. Serum DHA-PPQ concentrations were retrospectively evaluated on cryo-preserved (\u221280\u00a0\u00b0C) samples taken on different days, during and after the drug administration, coupled with tandem-mass spectrometry  responsible for the patient\u2019s infection was performed by amplification of three polymorphic markers, the merozoite surface protein 1 (msp1), merozoite surface protein 2 (msp2), and glutamate-rich protein (glurp) genes. These genes show a length polymorphism, allowing the detection of multiple infections by different P. falciparum genotypes [msp1/msp2 and for glurp as described by Wooden et al. [P. falciparum isolates showed the presence of a single isolate responsible for the first episode and for the recurrence  linked to resistance to the artemisinin derivatives, quinolines, antifolates-cycloguanil and atovaquone. The polymorphism of the propeller domain of the Pfk13 gene was assessed as described by Taylor et al. [Pfcrt/Pfdhps and Pfmdr1 and genes was performed as previously described in Menegon et al. [Pfdhfr gene spanning codons 51\u2013108 was analysed as described by Palmieri et al. [P. falciparum isolates showed the presence of mutations in Pfcrt and Pfmdr1 genes linked to resistance to the quinolines and mutation in Pfdhfr and Pfdhps genes correlated to antifolate/cycloguanil resistance. No mutations associated with artemisinin resistance and atovaquone were detected.Genotyping of enotypes , 13. Totn et al.  and Virir et al. . Analysin et al.  and Duran et al. , respecti et al. . The prei et al. . All PCRPfK13 gene suggest the involvement of an artemisinin-sensitive strain. Although it was not possible to analyse a specific molecular marker of resistance to PPQ  [This is the first case of failure of DHA-PPQ reported in Europe. The patient returned from Ethiopia, a country where DHA-PPQ failures have not been reported before. In this case, the rapid response to DHA-PPQ and the lack of mutations in the is drug) , the comIn the last 2\u00a0years (July 2014 to June 2016) DHA-P was administered to 36 patients attended at the CTD for falciparum malaria, observing no other failure. These data are in agreement with the literature. The efficacy of DHA-PPQ has been found very high, particularly in the African continent. There was a relevant delay between the onset of symptoms and the second diagnosis because the index of suspicion was low due to the negative laboratory tests performed at the 28-day follow-up visit. In consideration of the growing number of cases of resistance to ACT, it is important to consider a possible recrudescence, which can manifest several weeks after treatment."}
+{"text": "Innovative approaches are needed to complement existing tools for malaria elimination. Ivermectin is a broad spectrum antiparasitic endectocide clinically used for onchocerciasis and lymphatic filariasis control at single doses of 150 to 200 mcg/kg. It also shortens the lifespan of mosquitoes that feed on individuals recently treated with ivermectin. However, the effect after a 150 to 200 mcg/kg oral dose is short-lived (6 to 11 days). Modeling suggests higher doses, which prolong the mosquitocidal effects, are needed to make a significant contribution to malaria elimination. Ivermectin has a wide therapeutic index and previous studies have shown doses up to 2000 mcg/kg  are well tolerated and safe; the highest dose used for onchocerciasis is a single dose of 800 mcg/kg.The aim of this study is to determine the safety, tolerability, and efficacy of ivermectin doses of 0, 300, and 600 mcg/kg/day for 3 days, when provided with a standard 3-day course of the antimalarial dihydroartemisinin-piperaquine (DP), on mosquito survival.Anopheles gambiae s.s. populations fed on patients\u2019 blood taken at days 0, 2 (Cmax), 7 (primary outcome), 10, 14, 21, and 28 after the start of treatment. Safety outcomes include QT-prolongation and mydriasis. The trial will be conducted in 6 health facilities in western Kenya and requires a sample size of 141 participants (47 per arm). Sub-studies include (1) rich pharmacokinetics and (2) direct skin versus membrane feeding assays.This is a double-blind, randomized, placebo-controlled, parallel-group, 3-arm, dose-finding trial in adults with uncomplicated malaria. Monte Carlo simulations based on pharmacokinetic modeling were performed to determine the optimum dosing regimens to be tested. Modeling showed that a 3-day regimen of 600 mcg/kg/day achieved similar median (5 to 95 percentiles) maximum drug concentrations (Cmax) of ivermectin to a single of dose of 800 mcg/kg, while increasing the median time above the lethal concentration 50%  from 1.9 days (1.0 to 5.7) to 6.8 (3.8 to 13.4) days. The 300 mcg/kg/day dose was chosen at 50% of the higher dose to allow evaluation of the dose response. Mosquito survival will be assessed daily up to 28 days in laboratory-reared Recruitment started July 20, 2015. Data collection was completed July 2, 2016. Unblinding and analysis will commence once the database has been completed, cleaned, and locked.High-dose ivermectin, if found to be safe and well tolerated, might offer a promising new tool for malaria elimination. Onchocerca volvulus (the cause of river blindness) [Wuchereria bancrofti (the cause of lymphatic filariasis) [Strongyloides stercoralis  [Ivermectin is a potential new tool that is being considered in malaria transmission reduction strategies . Ivermecindness) , Wuchereariasis) , and Strelminth) . To dateelminth) .Anopheles spp. at concentrations present in human blood after standard doses. It reduces the re-blood feeding capacity, female fecundity, hatch rate of their eggs, the survival of progeny larvae, and importantly, it reduces the vector\u2019s lifespan [Ivermectin has secondary effects on ectoparasites, such as head lice, mites, bedbugs, and scabies, that feed on recently treated individuals ,7, and ilifespan ,8-11. Itlifespan . Ivermecin vivo studies assessed the long-term effect of ivermectin on mosquito survival by conducting feedings at least 7 days after administration of ivermectin [An. gambiae s.s. for up to 6 days, resulting in an estimated reduction of malaria transmission for at least 11 days as a result of a change in the age-structure of An. gambiae s.s. [An. gambiae survivorship by 33.9% for 1 week, their parity rates for more than two weeks, and sporozoite rates by more than 77% for 2 weeks [However, several studies have shown that the effects after the standard 150 to 200 mcg/kg doses of ivermectin are generally short-lived. Three ermectin ,13,14. Aermectin , while aiae s.s. . Similar 2 weeks .Modeling has also shown that adding 3 days of ivermectin (150 mcg/kg/day) to MDA with dihydroartemisinin-piperaquine (DP) would potentially provide an important boost to the effect of MDAs with artemisinin-based combination therapy (ACT) by allowing transmission to be interrupted faster and in areas with a higher malaria prevalence than MDA with ACTs alone . HoweverIvermectin 400 mcg/kg has been suggested as an improved treatment for head lice , and hasIvermectin has an excellent safety profile , and expLoa loa, possibly due to rapid lysis of parasite biomass [Loa loa is recommended before ivermectin administration in areas endemic for Loa loa filariasis [The only known severe adverse events have been in individuals with  biomass . Assessmlariasis .DP and ivermectin have, to the best of our knowledge, never been studied under simultaneous administration. Piperaquine, the long-acting component of DP, is metabolized by, and is an inhibitor of, cytochrome-P450 3A4 (CYP3A4) . There iIn vitro studies using human liver microsomes suggest that ivermectin does not significantly inhibit the metabolizing activities of CYP3A4, CYP2D6, CYP2C9, CYP1A2, and CYP2E1 [Ivermectin is primarily metabolized by CYP3A4 . In vitrd CYP2E1 . When DPWe will conduct a placebo-controlled dose-finding study to determine the safety, tolerability, and mosquitocidal effect of 3-day courses of ivermectin when given in combination with a standard 3-day course of DP to identify safe and practical regimens to boost the arsenal of available tools to reduce or interrupt malaria transmission. Pharmacokinetic data will be collected to facilitate the construction of a pharmacokinetic/pharmacodynamic (PK/PD) model to guide future study design.This is a double-blind, randomized, placebo-controlled, parallel-group, 3-arm, superiority trial to determine the safety, tolerability, and mosquitocidal effect of different doses of ivermectin . The primary endpoint will be mosquito survival 14 days after a blood feed from a patient who started ivermectin 7 days earlier; 5 days after the last dose of ivermectin with a 3-day regimen administering ivermectin at 0, 24, and 48 hours . Because mosquito feeding involves approximately 100 mosquitoes per feed, the study will use a clustered design with the patient as the unit of randomization and the mosquito as the unit of analysis. The study will have a nested rich pharmacokinetic component in the first 36 patients that give additional consent for rich/frequent sampling and a sparse sampling population pharmacokinetic component in the remaining patients. A second nested study will compare the effects of ivermectin when assessed by membrane feeding versus direct skin feeding in all patients who give additional consent for direct skin feeding.The primary objective of the study is to determine the safety, tolerability, and efficacy of ivermectin doses of 0, 300 and 600 mcg/kg/day for 3 days, when provided with a standard 3-day course of the antimalarial DP, on mosquito survival.The secondary objectives of the study are (1) to determine the effect of different doses of ivermectin on oocyst development; (2) to determine the pharmacokinetic profile of the different ivermectin regimens; (3) to determine if ivermectin interacts with the pharmacokinetics of piperaquine; (4) to determine whether the addition of ivermectin to DP affects the clinical and parasitological response to DP treatment; (5) to determine the role of genetic variants of CYP3A4 activity in metabolizing ivermectin; and (6) to determine the effect of direct feeding versus membrane feeding on mosquito survival.The goal was to design and evaluate a high-dose ivermectin regimen that could be given daily as adjunct therapy to a 3-day ACT regimen and that builds on the existing safety data available from previous studies. The highest dose of ivermectin used in studies for onchocerciasis is 800 mcg/kg given as a single dose . The pharmacokinetic profile of this 800 mcg/kg dose was used to design a 3-day regimen that would achieve a similar maximum drug concentration (Cmax) after the third dose. Since the highest dose of ivermectin used in humans that was tested and found to be well tolerated and safe is 2000 mcg/kg given as a single dose, this provides a large margin of safety allowing for inter-individual variation of pharmacokinetics. The middle group was chosen at 50% of the highest dose to allow for a dose response in terms of tolerance and efficacy.Using existing literature data ,35 we deThe proposed study uses a standard parallel design, comparing the 2 intervention arms with the placebo arm. This parallel design, instead of a dose-escalation design (when the lower dose group would be studied first prior to enrolling patients in the higher dose group), was considered appropriate because the Cmax levels and the 95th percentile concentrations in the proposed highest dose group of 600 mcg/kg/day will be equivalent to the Cmax found with single dose 800 mcg/kg, which has been administered to at least 402 patients before as treatment for onchocerciasis or as part of regulatory studies see . FurtherThe study will enroll patients with symptomatic uncomplicated malaria, instead of asymptomatic patients with malaria parasites (carriers) or malaria negative individuals who are the predominant target population in MDA campaigns. However, it is unlikely that the mosquitocidal effect of ivermectin will differ much amongst these groups. Preference is given to symptomatic patients based on the rationale that this study is labor intensive, requiring very frequent patient follow-up and blood sampling, and thus requires a major commitment from study participants. Symptomatic patients, aside from similarly requiring antimalarial treatment, are more likely to favor hospital admission and frequent outpatient visits than asymptomatic patients or other volunteers. The frequent follow-up is potentially also more beneficial to the patients with symptomatic malaria than asymptomatic patients.The cytochrome P450s (CYPs) are the major enzymes involved in drug metabolism. To be able to interpret variations in the pharmacokinetic drug profiles of piperaquine and ivermectin, and any drug interactions, we need to determine the genotypes of the genes encoding CYP enzymes (see above).The primary endpoint is based on membrane feeding of mosquitoes using blood obtained by venepuncture from patients recently treated with ivermectin. However, a nested sub-study, in all those that give additional consent, will compare mosquito mortality rates between clusters fed using standard membrane feeding versus clusters fed directly . Ivermectin feeding studies with direct feeding on humans , and catWe hypothesize that direct feeding could result in higher mosquito mortality due to potential differences between venous blood (used in membrane feeding) and blood in subdermal venuoles and arterioles (the main source of blood for mosquitoes during direct skin feeding) due to drug accumulation in subcutaneous fat, dermal, and facial tissue (2- to 3-fold higher concentrations than in venous blood ), or incThere have been no studies conducted directly comparing direct feeding versus membrane feeding on mosquito mortality following ivermectin administration. However, previous studies looking at infectivity  showed significant differences in terms of infectivity in favor of direct feeding (odds ratio 2.39) . AlthougMembrane feeding will be used as the primary outcome because direct skin feeding is labor intensive, may be unpleasant to the study participants, and result in higher refusal rates.The study will be conducted in the Jaramogi Oginga Odinga Teaching and Referral Hospital (JOOTRH) in Kisumu, western Kenya, a major tertiary care hospital. Almost 25,000 outpatients are treated for clinical malaria at JOOTRH annually, of which one-third are laboratory-confirmed. Approximately 20% of these patients are 18 to 50 years old. Malaria positive individuals will also be pre-screened at 5 nearby health facilities; those that pass pre-screening and give consent will be brought to JOOTRH for screening and all further study procedures.Inclusion and exclusion criteria for the study are shown in Inclusion Criteria:P. falciparum infectionSymptomatic, uncomplicated Positive malaria microscopy or malaria rapid diagnostic test Age 18 to 50 yearsProvide written informed consentAgree to be able to travel to clinic on days 1, 2, 7, 10, 14, 21, and 28Exclusion Criteria:Signs or symptoms of severe malariaUnable to provide written informed consentWomen who are pregnant or breast feedingHypersensitivity to ivermectin or DPRate corrected QT interval (QTc) of greater than 460 ms on electrocardiogram (ECG)Body mass index (BMI) below 16 or above 32 kg/m2Hemoglobin (Hb) concentration below 9 g/dLTaken ivermectin in the last monthTaken DP in the last 12 weeksLoa loa as assessed by travel history to Angola, Cameroon, Chad, Central African Republic, Congo, DR Congo, Equatorial Guinea, Ethiopia, Gabon, Nigeria and SudanHistory and/or symptoms indicating chronic illnessCurrent use of tuberculosis or anti-retroviral medicationPreviously enrolled in the same studyParticipants will be randomized to one of the following 3 arms: (1) the \u201c0 mcg/kg\u201d (placebo) arm consisting of DP plus ivermectin-placebo 600 mcg/kg/day for 3 days, (2) the \u201c300 mcg/kg\u201d arm consisting of DP plus ivermectin 300 mcg/kg/day and ivermectin-placebo 300 mcg/kg/day for 3 days, or (3) the \u201c600 mcg/kg\u201d arm consisting of DP plus ivermectin 600 mcg/kg/day for 3 days. Patients will receive their weight-based doses of DP and ivermectin/placebo. Each dose will be given as directly observed therapy by study staff, after which participants will be monitored for 30 minutes for any vomiting and adverse reactions. If vomiting occurs within 30 minutes, then the participant will be withdrawn from the study, DP will be re-administered, and no further ivermectin will be given.DP was selected as the drug of choice as it is the most likely candidate to be used in future MDA campaigns because of the longer prophylactic effect against malaria (4 to 6 weeks) compared with 2 to 3 weeks with artemether-lumefantrine (AL). Each participant will receive a weight-based dose of DP 320/40 mg  as per the product insert  once a day for 3 days.Ivermectin and/or placebo 6 mg tablets  will be administered per bodyweight. The 600 mcg/kg/day arm will receive only ivermectin tablets, the 300 mcg/kg/day arm will receive half the number of ivermectin tablets and an equal number of placebo tablets, and the 0 mcg/kg/day arm will receive only placebo tablets. All participants will receive the same total number of tablets once a day for 3 days based on their bodyweight .The primary efficacy outcome is mosquito survival at 14 days after feeding on blood taken from study participants who started the 3-day ivermectin and DP regimen 7 days earlier .Secondary outcomes include (1) mosquito survival at each day, up to day 21 or 28, after each feeding experiments performed at 0, 2 days plus 4h, 7, 10, 14, 21, and 28 days after start of treatment; (2) occurrence of oocysts from day 10 onwards after each feeding as determined by polymerase chain reaction (PCR); (3) malaria clinical and parasitological treatment response by day 28; and (4) plasma concentration profiles of piperaquine and ivermectin as described by standard pharmacokinetic metrics, for example area under the curve measurements from time zero to infinity (AUC0-\u221e), time zero to the time of the last measurable concentration (AUC0-tlast), Cmax, and plasma half-life, time to maximum plasma concentration, etc .Tolerability and safety endpoints are shown in TolerabilityAny adverse events assessed in general toxicity questionnaires asked at each study visitSafetyPrimaryMydriasis quantitated by pupillometry SecondaryCentral nervous system (CNS) effectsGeneral toxicitySerious adverse eventsHemoglobin concentrationsQTc interval (see below \u201cElectrocardiogram Monitoring\u201d)The study plan and schedule of assessment is provided in Loa endemic countries), physical examination, electrocardiogram (ECG), pupillometry, and laboratory tests . Those fulfilling all enrolment inclusion criteria and not meeting any exclusion criteria will be enrolled into the study, randomized, and treated with the appropriate tablets according to study arm (Patients presenting to the outpatient departments of the study clinics will be pre-screened to determine if they meet readily apparent study eligibility criteria including (1) age 18 to 50 years; (2) uncomplicated malaria; (3) in Kisumu next 4 weeks; (4) hemoglobin (Hb) less than or equal to 9g/dL ; (5) not pregnant or breast feeding; (6) no known chronic illness; and (7) not previously enrolled in IVERMAL. Patients passing pre-screening will be approached to obtain consent. For those consenting, study-specific screening procedures will take place, including demographics, full history, past medication use, travel history  and Hb will be taken if clinically indicated .The study requires a total of 141 participants . This is powered at 80% to detect a relative increase of 30% (RR 1.300) in the 14-day post-feeding mortality rate (primary outcome) from 24% in the control group (0 mcg/kg ivermectin) to 31.2% in the 300 mcg/kg/day group, and a 25% (RR 1.246) increase from 31.2% with 300 mcg/kg/day to 38.9% in 600 mcg/kg/day recipients, measured from blood taken 7 days after the start of intake of ivermectin and using clusters of 100 anopheline mosquitoes allowing for 10% non-feeders . The same sample size would give 90% power to detect a 35% (RR 1.348) increase from 24% (0 mcg/kg/day) to 32.4% (300 mcg/kg/day), and 27.7% increase (RR 1.285) from 32.4% (300 mcg/kg/day) to 41.3% (600 mcg/kg/day). The calculations assume an intracluster correlation coefficient (ICC) of .0622 and allow for 6.5% loss-to follow-up of participants by day 7  [2 and 21 kg/m2, for females and males respectively. Participants will be randomly assigned to 1 of the 3 study arms. The study statistician will computer-generate a randomization sequence using permuted block randomization with fixed block sizes.The study will use stratified randomization (4 strata) by body mass index (BMI: high/low) and sex  as these are important determinants of the pharmacokinetics of ivermectin . The higThe study will be double-blinded to participants and study staff. Allocation concealment will be achieved by use of sealed opaque envelopes. All study participants in all 3 arms will receive standard dose DP, and also active (600 mcg/kg/day arm), placebo (0 mcg/kg/day arm), or a combination of active and placebo ivermectin tablets (300 mcg/kg/day arm), such that each arm receives the same number of tablets in each weight strata.The first 36 patients to give additional consent for rich pharmacokinetics (approximately 12 per arm), will be enrolled in a rich pharmacokinetic study using frequent sampling per individual , including demographic, pathophysiological, such as BMI and gender, and other factors that might alter dose-concentration relationships. As this is a placebo controlled trial, the sampling methodology for the 47 patients in the ivermectin-placebo arm will be identical to that used for the 300 and 600 mcg/kg arms. The patients in the placebo-ivermectin arm will allow us to determine the piperaquine kinetic profile in the absence of ivermectin.Finger prick blood draws will be performed at a maximum of 4 time points in addition to the venous blood draws. The aim is to compare the capillary and venous drug concentration levels as it has been hypothesized that these might differ for ivermectin, similar to other drugs including piperaquine. A difference between capillary and venous drug concentrations could help further explain any observed difference in mosquito mortality between membrane and direct skin feeding .All of the rich pharmacokinetic participants (approximately 12 per arm) will have venous blood sampled  at baseline and each of 21 follow-up time points listed in Each of remaining patients (approximately 35 per arm), not enrolled in the rich pharmacokinetic sub-study, contribute to the population pharmacokinetic study, which consists of 13 sampling points see , page 15In anticipation of a 40% refusal rate or loss to follow-up, we estimate that the combined rich and population pharmacokinetic sub-studies will contribute 361 samples including 47 baseline samples (100%) and 314 (60%) follow-up samples out of a potential 524 follow-up samples across 22 sampling time points after baseline, 20 of which overlap, with a total of 12 to 47 observations per time point  and secondary entomological endpoints (sporogony). The section below describes the maintenance of the mosquito colonies.An. gambiae s.s. Kisumu strain, originally from Kisumu, Kenya. The strain is maintained at the KEMRI/Centre for Global Health Research (CGHR) insectaries and is susceptible to all insecticides approved by the World Health Organization (WHO). When performing membrane feeds on infected human blood, mosquitoes will be kept and fed in cages or paper cups. The cages or paper cups will be kept in a temperature- and humidity- controlled insectary. The feeding and the storage of live infected mosquitoes will occur in sealed rooms with at least 2 doors and/or barriers separating the inner rooms from the outside. Mosquitoes will not be removed from their enclosures, with exception of the cage for oocyst determination. During transportation, live infected mosquitoes will be transported within paper cups that are covered with a moist towel and enclosed within locked cool-boxes to remove any chances of escape. The cool-boxes will only be opened within the confines of a double door insectary.The mosquito colony used in this study will be The LC50 has been previously estimated using spiked blood (blood to which known concentrations of ivermectin are added) in membrane feeding assays ,15. We wHb will be tested using HemoCue  photometers.Thick and thin blood films for parasite counts will be obtained and examined. Malaria parasites will be counted against 200 high power fields before a slide is declared negative .Plasma will be stored locally on site at -20\u00b0C or in liquid nitrogen and shipped to a central laboratory for storage at -70\u00b0C prior to batch analysis at the Liverpool School of Tropical Medicine. Samples will be shipped in dry ice to the laboratories in Liverpool, United Kingdom where the plasma concentrations of ivermectin and piperaquine will be determined using assays validated to international Food and Drug Administration (FDA) standards. Plasma concentration-time data will be used to evaluate pharmacokinetic parameters including CL/F , V/F , and Ka (absorption rate constant) using population pharmacokinetic methods. Area under the curve (AUC) and half-life will also be calculated.A study statistical analytical plan for the final analysis, that supersedes the study protocol, has been drawn up during the course of the study before the unblinding of data at database lock see .An. gambiae s.s. mosquitoes will be presented to the membrane feeder for 20 minutes. The number of mosquitoes with an engorged abdomen will be counted and those with lean abdomens discarded. Each day up to day 28  or day 10 (oocyst cup), the number of dead mosquitoes will be counted and removed. After the initial feeding on human blood, the mosquitoes will be kept in an incubator and maintained on sugar feeds. Insectary staff assessing mosquito survival and oocyst development will be blinded to all characteristics of the cups .The following procedures will be conducted in accordance with a standard membrane feeding protocol . A 1 mL The primary outcome will be the survival of mosquitoes  at 14 days after feeding on blood taken from study participants who started the 3-day ivermectin and DP regimen 7 days earlier.Although the primary endpoint is assessed at day 14, the study will collect survival data of mosquitoes at each day up to day 21 or 28 for the mosquito survival cups and day 10 in the case of oocyst cups, after each feeding experiments performed at 0, 2 days plus 4h, 7, 10, 14, 21, 28 days after start of treatment. The methods will be identical to that described for the primary outcome where each day beyond day 14 the number of dead mosquitoes will be counted and removed until day 28 inclusive. The exact number of follow-up days (21 or 28 days) will be subject to logistical constraints of the laboratory, and mortality rates in the mosquito populations which will be further determined prior to the start of the study. The aim is to determine the median time to mortality, which requires that at least half of the mosquito population has died in each arm. It is anticipated that 21 days will be sufficient.A sub-study will determine the effect of direct feeding versus membrane feeding on mosquito survival, after feeding experiments performed at 7 days after the start of treatment. In direct skin feeding assays, 1 cup of 50 mosquitoes is placed directly on the skin of the human host and allowed to feed for 15 minutes . FurtherOn day 10 post membrane feeding, when residual DNA from the blood meal is highly unlikely ,41,42, aStandard methods will be used to assess the in vivo treatment response to DP using the microscopy and rapid diagnostic test (RDT) data collected at each scheduled follow-up visit and criteria described by World Wide Antimalarial Resistance Network (WWARN) .In animal studies, mydriasis has been shown to be a first sign of ivermectin toxicity. To monitor for possible toxicity, pupil diameter size will be measured at baseline and each scheduled visit using a portable, single-button activation, battery operated hand-held pupillometry device that very accurately measures pupil size requiring no calibration (NeurOptics VIP-200 Variable Pupillometer). This device measures the pupil 30 times per second over a 5-second period and provides the average pupil diameter and standard deviation (+/- 0.1 mm). The measurements will be taken in a dark room with standardized lighting conditions.0.33.Piperaquine can potentially lead to prolongation of the rate corrected QT interval (QTc) on an electrocardiogram (ECG). To exclude a possible interaction between ivermectin and piperaquine leading to an increased QTc interval, 12-lead ECGs will be performed to measure the QTc interval at baseline, day 2 pre last dose, day 2 at 4 to 6h post last dose and again at day 28. The day 28 sample is included as it can be difficult to assess a true baseline in patients with acute malaria, as malaria and fever are known to increase the heart rate and decrease the QTc interval. On day 28 most, if not all, patients will be malaria free and residual piperaquine levels low enough not to affect QTc intervals. A portable ECG machine  will be used with automated ECG interpretation. Patients with a QTc value of 480 ms or greater prior to the last dose of DP will not receive the last dose of DP, but receive a full course of artemether lumefantrine instead. Fridericia\u2019s correction will be used to calculate the QTc values for final data analysis using the following equation: QTc = QT/RRAdverse events and serious adverse events will be monitored, managed, and recorded during the course of the study. They will be recorded and tabulated for each treatment arm, overall, and per body system , LSTM (protocol #14.002), and JOOTRH. The Centers for Disease Control and Prevention  gave approval for reliance on the KEMRI institutional review board  are well tolerated, safe, and result in longer durations of mosquitocidal effects than standard 150 to 200 mcg/kg single dose treatments. This study requires major infrastructure and collaboration, as it brings together the disciplines of clinical medicine, entomology, parasitology, pharmacokinetics, and pharmacogenetics in a clinical trial. For this study, 141 patients and 150,000 mosquitoes will each be followed for 28 days. For this reason, this trial has been placed at the KEMRI, CDC, and LSTM collaboration in western Kenya, a research site, which in collaboration with its partners, has been conducting research for over 35 years and has the capacity to undertake such a trial. An important possible limitation of this study is that it will enroll participants with symptomatic malaria, whereas possible future applications of high-dose ivermectin may involve MDA with ACT\u2019s targeting asymptomatic carriers and uninfected individuals in addition to symptomatic patients. Should this study show promising results, then the next step will be to evaluate safety, tolerability, and efficacy in younger age groups with the ultimate goal of testing its effect on malaria transmission when applied at the population level through MDA.High-dose ivermectin, if found to be safe and well tolerated, could potentially complement existing tools for malaria elimination."}
+{"text": "Drug efficacy trials monitor the continued efficacy of front-line drugs against falciparum malaria. Overestimating efficacy results in a country retaining a failing drug as first-line treatment with associated increases in morbidity and mortality, while underestimating drug effectiveness leads to removal of an effective treatment with substantial practical and economic implications. in vivo data. The current WHO-recommended method for molecular correction and analysis of clinical trials should be reevaluated and updated.Drug efficacy trials monitor the continued efficacy of front-line drugs against falciparum malaria. Overestimating efficacy results in a country retaining a failing drug as first-line treatment with associated increases in morbidity and mortality, while underestimating drug effectiveness leads to removal of an effective treatment with substantial practical and economic implications. Trials are challenging: they require long durations of follow-up to detect drug failures, and patients are frequently reinfected during that period. Molecular correction based on parasite genotypes distinguishes reinfections from drug failures to ensure the accuracy of failure rate estimates. Several molecular correction \u201calgorithms\u201d have been proposed, but which is most accurate and/or robust remains unknown. We used pharmacological modeling to simulate parasite dynamics and genetic signals that occur in patients enrolled in malaria drug clinical trials. We compared estimates of treatment failure obtained from a selection of proposed molecular correction algorithms against the known \u201ctrue\u201d failure rate in the model. Our findings are as follows. (i) Molecular correction is essential to avoid substantial overestimates of drug failure rates. (ii) The current WHO-recommended algorithm consistently underestimates the true failure rate. (iii) Newly proposed algorithms produce more accurate failure rate estimates; the most accurate algorithm depends on the choice of drug, trial follow-up length, and transmission intensity. (iv) Long durations of patient follow-up may be counterproductive; large numbers of new infections accumulate and may be misclassified, overestimating drug failure rate. (v) Our model was highly consistent with existing  Plasmodium falciparum, and this study focuses on drug treatment of that species. Prompt treatment of malaria infections is an essential and effective public health tool, but drug resistance poses a constant threat to effective treatment of falciparum malaria. The World Health Organization (WHO) currently recommends that countries of endemicity test their first- and second-line antimalarial drugs every 2 years at sentinel sites to confirm their continued efficacy . If the profiles \u201cmatch,\u201d then the patient is considered to have a recrudescent infection, but if they do not match, the patient is considered to have a reinfection. This \u201cmatching\u201d is simple in principle, but in practice has substantial limitations. The main problem is that individual malaria infections may consist of several genetically distinct clones. Current genotyping techniques struggle to detect minority clones that are present in relatively low numbers and/or which carry alleles that do not amplify well during the genotyping process. These limitations were recognized early in the development of molecular correction methodology \u20138 and le\u20131.Recrudescent infections could be misclassified as reinfection if the recrudescent allele(s) were not detected during the genotyping of the initial infection .2.A recrudescent infection could be misclassified as a reinfection if the recrudescent allele(s) were not detected during the genotype of the recurrent infection .3.glurp\u201d algorithm was not affected by sharing an allele at glurp, and the \u201callelic family switch\u201d algorithm was sensitive to sharing an msp-1 or msp-2 family by chance, whereas the other algorithms were not).A reinfection could be misclassified as recrudescent if it shares (by chance) alleles with clones present at time of treatment. The exact number (or type) of alleles that must be shared depended on the molecular correction algorithm chosen  and have no reinfection, such that parasites were never detected during follow-up ; this obviously depends on the duration of follow-up.While not misclassification of recurrence, another source of bias affected the accuracy of failure rate estimates with respect to the true failure rate: a patient who failed to clear the initial infection may have had that infection persisting at a low level, below the limit of detection of detection of light microscopy .The noncorrected algorithm always produced a higher failure rate estimate than any of the four molecular correction algorithms . FailureThe ability of the four molecular correction algorithms to accurately estimate drug failure rates depended on their ability to correctly classify recrudescences and reinfections. This ability is shown  in \u2022The WHO/MMV-recommended algorithm consistently underestimated failure rates at all transmission intensities as shown in \u2022glurp algorithm produced slightly higher estimated failure rates than the WHO/MMV algorithm across all FOI settings  and increased the chance of a reinfection being misclassified as a recrudescence  . A complt sample  would beAlternate durations of follow-up length were simulated for DHA-PPQ, and their impacts on estimated failure rates are shown in Underestimation of the true failure rate occurred with all algorithms when a 28-day follow-up period was chosen. With a 42-day follow-up period, the allelic family switch algorithm produced the most accurate failure rate estimate, with an FOI of <7, and the \u22652/3 markers algorithm produced the most accurate failure rate estimate, with an FOI of \u22657. As length of follow-up increased to 63\u2009days, the \u22652/3 markers algorithm tended to slightly overestimate the failure rate. This effect was more apparent as the FOI increased. These patterns emerged because only a small number of initial clones recrudesced after 42\u2009days. Additional models for other ACTs are described in the supplemental material. In brief, these drugs differed from DHA-PPQ mainly in their persistence of active drug concentrations posttreatment (and hence in their prophylaxis against reinfections). Results for failing AR-LF and AS-MQ were highly consistent with those described above for DHA-PPQ, showing the same qualitative patterns .Different prophylactic profiles meant that the most effective duration of follow-up for AR-LF and AS-MQ (as would be expected) differed from that for DHA-PPQ; using the \u22652/3 markers molecular correction algorithm, a 28-day follow-up for AR-LF appeared to be most accurate at moderate to high FOI. A 49-day follow-up for AS-MQ appeared to be the most accurate with the \u22652/3 markers algorithm, but increased accuracy over the WHO/MMV algorithm was also seen with shorter follow-up periods.Models of nonfailing  pharmacokinetic/pharmacodynamic (PK/PD) calibration of AR-LF and AS-MQ showed that the \u22652/3 markers algorithm slightly underestimated the true failure rate, but this difference was small, and there is no evidence that this algorithm would incorrectly identify effective drugs as failing.glurp, \u22652/3 markers, and allelic family switch algorithms. Failure rate estimates are lower across all algorithms than with the shorter-prophylaxis two-compartment model, and a 63-day follow-up appears to be the most suitable under this calibration: the \u22652/3 markers algorithm produced an accurate failure rate estimate at all but the lowest FOI levels with this follow-up length. Crucially, the key message is the same: the WHO/MMV algorithm underestimates the true failure rate, and other algorithms can produce more accurate failure rate estimates. Perhaps the most interesting difference between the two DHA-PPQ PK/PD calibrations is that they suggested, given use of the same molecular correction algorithm, different optimal lengths of follow-up.Alternative parasite dynamics for DHA-PPQ were generated using a three-compartment model with PK parameters described in reference Failure rate estimates were calculated using the per protocol method rather than survival analysis. The per protocol method led to increased failure rate estimates with all algorithms, all ACTs, and all follow-up periods. These results are discussed in the supplemental material.glurp algorithm being the most accurate at a moderate to high FOI.Finally, the simulation was validated by varying the multiplicity of infection (MOI) at time of treatment, the relative detectability of alleles based on length, and the minority allele detection threshold. The results of these analyses are provided in the supplemental material and showed mostly the same qualitative patterns as the results presented above; the one key departure was that assumption of a minority allele threshold of 5% (reduced from the assumption of 25% presented above) led to slightly increased failure rates and the no-glurp algorithm, the \u22652/3 markers algorithm, then the allelic family switch algorithm. The pattern was quantitatively consistent: the WHO/MMV algorithm estimated failure rates to be around half that obtained by the \u22652/3 markers algorithm. Results are similarly consistent with reanalysis of a trial from low-transmission settings in Cambodia  were reanalyzed using the proposed molecular correction algorithms  and wereCambodia . The impFinally, we reviewed clinical trials that reported failure rates based on no correction and the WHO/MMV algorithm . The magmsp-1, msp-2, and glurp correctly classified all recurrent infections : hence, the approach of investigating multiple ACTs and various transmission intensities through FOI to assess if a single algorithm may be identified that gives robust and accurate estimates. Based on the results presented here, it appeared that the \u22652/3 markers algorithm was the most robust in areas of moderate to high transmission and provided estimated failure rates close to  the true failure rate  Note that a different DHA-PPQ parameterization  favored a longer follow-up period more in line with MQ, which also has longer prophylaxis posttreatment. The trends across all drugs modeled are clear: it is highly likely that use of the current WHO/MMV algorithm will generate substantial (near 2-fold) underestimates of failure rates and that switching to an alternative correction algorithm should be considered a matter of urgency.The other factor that can affect estimates of drug efficacy, given that molecular correction is imperfect, is the duration of follow-up. Recommended duration has gradually increased over the last 20 to 30\u2009years, with the objective of capturing all (or at least the majority of) recrudescences. However, the objective of clinical trials is not to capture every recrudescence, but to obtain accurate and robust estimates of efficacy. Technical problems with molecular correction approaches exist ; however, the purpose of reanalysis of these data was to investigate the change in failure rates from use of proposed algorithms on in vivo data\u2014analysis of trial results with these algorithms had not previously taken place. This reanalysis is not dependent on our model parameter space (nor vice versa), and all algorithms require the same data ; consequently, this reanalysis showing similar trends to our modeled results is encouraging.The evaluation of different classification algorithms relied on simulated data. This was not ideal, but there is no obvious alterative given that key parameters , cannot be directly observed msp-1, msp-2, and glurp and how they may be best used to distinguish recrudescences from reinfections; it would be straightforward to repeat these analyses for different types of molecular data, such as deep-sequenced amplicons, microsatellites, and single nucleotide polymorphism (SNP) barcodes, and this is discussed further in the supplemental material. Notably, reduction of the minority detection threshold to 5% increased the failure rate estimates and altered which algorithm produced the most accurate estimate. We are confident that the length-polymorphic markers do not have this level of sensitivity. We analyzed this assumption solely to test its effect on our results; however, this threshold emulates more closely the use of amplicon sequencing, where minority alleles are easier to detect, and we intend to test the accuracy of failure rate estimates with amplicon sequencing using a similar methodology in the future.Focus has been on the current WHO-recommended marker loci There is concern in the literature that reinfections may share alleles with the initial infection purely by chance and that subsequent misclassification of reinfections as recrudescence would lead to overestimation of failure rates . This coIn conclusion, both our modeling approach and reanalysis of clinical data suggest that more accurate and easily implemented algorithms are available to analyze clinical data and the field should consider implementing these methods. Which algorithm will perform best will depend on factors in the patient population/area\u2014our results demonstrate this explicitly for transmission intensity (FOI) and follow-up length. The four algorithms investigated here are not mutually exclusive and are based on the same data. Our firm recommendation is that initial and recurrent samples should be genotyped at all three loci: when using the current WHO/MMV algorithm, there is no need to genotype after a mismatch has occurred at one locus, so genotyping is often incomplete. These complete data would allow results obtained from all four algorithms to be presented; this maintains consistency with previous analyses based on the WHO/MMV algorithm, while also providing results that are likely to provide a substantially more robust estimate of malaria drug clinical failure rates.The World Health Organization (WHO) has published a standardized \u201cconsensus\u201d list of terms  that we 1.P. falciparum dynamics in these patients posttreatment.Use pharmacological modeling to simulate the parasite dynamics posttreatment in a population of patients enrolled in a clinical trial and track subsequent intrahost 2.Allocate genetic signals to each simulated parasite clone and calculate the genetic signals detected from a patient\u2019s blood sample at a given follow-up day , then analyze these signals using different algorithms  to classTo create a model with which to investigate the accuracy of molecular correction methods, we used a two-stage process implemented in the statistical programming language R (version 3.5.1) :1.Use phMalaria parasite dynamics were generated using pharmacological models of malaria drug treatment that have been developed over the last decade , and for both a two-compartment model and three-compartment model of DHA-PPQ . Thus, we are confident that PK/PD models of DHA-PPQ, AR-LF, and AS-MQ were appropriate means by which to generate parasite dynamics posttreatment for the purposes of this study. Alternative methods were available: i.e., arbitrarily constructing recurrent infections as containing a given proportion of recrudescent and/or reinfection and testing the algorithm\u2019s ability to correctly classify them  models; these models have been calibrated and validated for a range of ACTs and successfully used to investigate a variety of key research questions \u201318, 21. \u2013While it was obviously not feasible for us to simulate and present every possible parameterization to create parasite dynamics likely to occur in trials , calibrating the models to rerun a specific set of parameters for interested groups is a simple task upon provision of the parameters.10 to 1011 asexual parasites per person. Previous modeling approaches . Two MOI distributions were used in our models. A \u201chigh MOI\u201d is representative of the MOI in an area of intense transmission\u2014in this case, Tanzania, where MOI of 1 to 8 were assigned with probabilities of 0.036, 0.402, 0.110, 0.110, 0.183, 0.049, 0.061, and 0.049, respectively . A \u201clow-proaches  used 1015, and all drugs modeled were assumed to be inactive against the hepatic stages. The rate of emergence reflected the local intensity of malaria transmission and was quantified as the \u201cforce of infection\u201d (FOI). At the start of the model, each patient was assigned the number of reinfections that would emerge during a year. This number was drawn from a Poisson distribution whose mean value was the FOI. Values for FOI from 0 to 16 were used to reflect low-, medium-, and high-transmission areas; as a general guide, we regarded FOI \u2264 2 as representing a low-transmission setting, 2 < FOI \u2264 8 as indicative of moderate transmission intensity, and FOI\u2009>\u20098 as high transmission; the yearly value was then converted to the number of reinfections occurring during the follow-up period. Reinfections emerging from the liver are illustrated as the gray and orange dotted lines in Multiple lengths of follow-up are permitted in the WHO guidelines  and used12, then density-dependent effects, such as fever, set the growth rate of every clone to 0.The parasitemia of each clone in each patient was tracked and updated each day to reflect two factors. First was the extent of drug killing based on the PK/PD parameters ; second was the growth rate of each clone, which was assumed to be identical for every clone and set to 1.15/day as in previous modeling work , 28. The8 on that day. We note that variance in the limit of detection by light microscopy exists with respect to the skill of the microscopist  were also noted. The distributions we used gave msp-1 an expected heterozygosity (He) of 0.915, msp-2 an He of 0.963, and glurp an He of 0.956 . It was assumed that the genotypes of initial clones were independent of each other and independent of the genotypes of reinfections . Note that alleles at msp-1 and msp-2 exist in our distributions as members of three or two distinct families, respectively.Each clone, whether an initial clone present at treatment or a reinfection that emerged during the follow-period, was assigned a genetic profile based on three markers: Tanzania . glurp din vivo trials. Blood samples were taken from each patient immediately prior to treatment  and at predetermined days during the follow-up period. Samples were screened for the presence of Plasmodium falciparum by light microscopy, and any detected infection was labeled a \u201crecurrence.\u201d We simulated the genotyping that would be used in vivo to obtain the genetic profiles for all initial and recurrent infections. Recovering the genetic signal that would be observed at time of treatment and at any recurrence reflected the technical limitations of acquiring blood samples and genotyping as follows:\u20226 \u03bcl of blood. For a clone to be detected, a minimum of 1 parasite (which carries a single DNA template) would need to be present in 1 \u03bcl of blood, so there would need to be present at least 5 \u00d7 106 parasites representing a given clone for that clone to be physically sampled in the genotyping process. We also needed to allow for the fact that suboptimal storage conditions (such as temperature) frequently occur in the field and will lead to DNA template breakages, and there is periodical absence from the peripheral blood of sequestered parasites. Consequently, the limit of detection will be much higher than 1 parasite per \u03bcl of blood. We therefore assumed 10 to 20 parasites per \u03bcl would be required to reliably contribute a genetic signal and ensure its detection, corresponding to a total parasitemia of 5 \u00d7 107 to 5 \u00d7 108; we selected the upper limit  to ensure reliable detection of that clone and because it is consistent with the microscopy detection limit.A \u201csampling limit\u201d exists; a finite amount of blood is used for genotyping. A parasite clone  would not be detected if its density were so low that no parasites are included in the blood sample analyzed. Thus, the density and volume of the processed blood sample define the limit of detection. Obviously, this sampling limit differs between methods and laboratories. Typically, the equivalent of 1 \u03bcl of whole blood is introduced into PCR. An assumption there is 5 liters of blood in the human body gives a total of 5 \u00d7 10\u2022The magnitude of the genetic signal that will be produced by each malaria allele in the blood sample was proportional to the number of parasites carrying that allele.\u2022msp-1 and msp-2 were assumed to be amplified by separate reactions (i.e. are not multiplexed), so the effect only occurred between alleles within the same families.  The sensitivity of the results to this relative detectability was tested by shortening it to 0.1 ; we later show that it does not affect our results.An inherent feature of PCR is \u201ctemplate competition\u201d: i.e., the relative detectability of alleles at each marker depended on their length, with shorter alleles being more detectable due to their being better amplified in the PCR process . A lineaOnce the patient parasite dynamics were modeled (as described above), and genetic profiles at the three loci were assigned, our models followed the same process as The strength of the genetic signal contributed by an allele in a given blood sample was therefore the product of two factors: the number of parasites carrying the allele \u00d7 the detectability of the allele. Note that genotyping detects alleles, not parasites. Hence, if two (or more) clones within the infection shared the same allele, the signal for that allele was based on the total number of parasites in the two (or more) clones. The final step is to recognize that, in practice, if one allele makes up a large proportion of the genetic signal, then the smaller signals from \u201cminority\u201d alleles would be rejected as background \u201cnoise.\u201d We assumed this threshold to be 25% , although we test other values of this parameter .We do not explicitly incorporate the effect of malaria sequestration in our simulations. Sequestered stages are not detectable in blood, so if a malaria clone is asynchronous in its 48\u2009h of development, its detectability will differ over consecutive days , 32\u2014hencglurp algorithm that only considers msp-1 and msp-2, (iii) a \u22652/3 markers algorithm that considers msp-1, msp-2, and glurp but requires matching alleles at only two markers to classify a recrudescence, and (iv) an allelic family switch algorithm that considers only msp-1 and msp-2 and requires a family shift to classify a recrudescence if the markers are discordant . Full details of these algorithms are presented in (i)An early treatment failure (ETF) if a recurrence occurs on or before day 7. Note that all such recurrences are regarded as drug failures, and molecular correction is not required. In our simulations, on day 3, if total parasitemia exceeded 108 but was <25% of the total parasitaemia of the initial sample, the patient continued in the trial per the WHO protocol . If parasites were present at >25% of initial parasitemia, that patient was classified as an early treatment failure, consistent with the WHO procedure ((ii)A drug failure if a recurrence was classified as such by a PCR correction algorithm in (iii)A reinfection if a recurrence was classified as such by a PCR correction algorithm in (iv)\u201cCleared\u201d: i.e. no recurrent parasitemia was detected during follow-up. In these cases, the drug was assumed to have successfully killed all parasites present at time of treatment.For analysis of parasitemia during patient follow-up and, if required, application of molecular correction algorithms to recurrent infections, four molecular correction algorithms  were investigated: (i) the current WHO/MMV algorithm , (ii) a \u20228 parasites from a clone present at time of treatment.  This included patients who have a \u201cmixed\u201d infection on the day of recurrence . Note that all clones contributed to the genetic signal of the recurrence as described above.True recrudescence was defined as a recurrent infection that contained at least 10\u20228 parasites].)True reinfection was defined as a recurrent infection whose blood sample contained only parasites from a clone or clones that were reinfections.  on the final day of follow-up; such patients would be classified as \u201ccleared\u201d in vivo and thus a treatment success. However, simulated data have confirmed that it is possible for some patients to still harbor parasites below the detection level at the end of follow-up  that we can test against the various classification algorithms and reduces the uncertainty around results.The model was run for a cohort of 5,000 patients . This is an unrealistically high number for an F\u0302, in the same manner as outcomes reported in vivo. It was assumed, for simplicity, that no patients were lost to follow-up or removed from the trial for any reason other than recurrent parasitemia. There were three methods for calculating failure rates, which differed in how they processed patients with recurrent parasitemia that had been classified as reinfections, noting that all patients with recurrent parasitemia would, in vivo, be treated again with another antimalarial  and removed from the trial. The three methods were a non-PCR-corrected failure rate, a \u201cper protocol\u201d failure rate, and a failure rate obtained using survival analysis. The latter two methods are recommended by the WHO to analyze antimalarial drug trials  The non-PCR-corrected failure rate was obtained by considering all patients with recurrent infections as patients who had failed drug treatment. This method did not require distinguishing between reinfections and recrudescent infections. The estimated failure rate, (ii) The per protocol method, recommended by WHO , 3, 5, s(iii) Survival analysis, as recommended by WHO , used tht, \u015c(t), was obtained as t was a vector of all time points , ni was the number of individuals at time ti who remained uninfected, and di was the number of events (drug failures in this case) that occurred at time point ti. Plainly, what this method did was calculate the proportion of patients who remained free of recrudescence between consecutive days of follow-up, then multiply all these time periods to obtain the overall probability of \u201csurviving\u201d recrudescence free over the whole follow-up period. The advantage was that even those patients who are \u201ccensored\u201d (by acquiring a reinfection and leaving the study) will still contribute to the analysis through their inclusion prior to their removal.The Kaplan-Meier (KM) estimator of survivorship at time The estimator at the final time point  was the probability that their treatment was considered a \u201csuccess\u201d at the end of the trial. Consequently, its complement gave the probability that a given individual will fail treatment:The final methodological step was to interrogate the modeled data to determine the \u201ctrue failure rate\u201d\u2014i.e., the drug failure rate calculated directly from the parasitemia of each patient . For each patient in the simulation, an outcome on the final day of follow-up was determined: if, on the final day, the patient had any parasites from any initial clones , the patient was denoted as a drug failure. If no parasites had survived from the initial clones present at treatment, that patient was denoted as a treatment success.F, for the patient population was then calculated asf was the number of drug failures on the final day of follow-up and N was the total number of patients.The true failure rate, in vivo. It was compared to the estimated failure rates obtained from modeling the clinical trial and molecular correction process and allowed us to quantify the accuracy of different methods .This was the \u201cgold standard\u201d metric and cannot be obtained msp-1, msp-2, and glurp. In patients treated with AR-LF, 137 recurrences were observed, of which 110 could be classified as either a reinfection or a recrudescence. (It was not possible to classify 27 patients because they had incomplete genetic data.) In patients treated with DHA-PPQ, 48 recurrences were observed, of which 43 could be classified as either a reinfection or a recrudescence. (It was not possible to classify 5 patients because they had incomplete genetic data.) These data were initially presented internally to the National Malaria Control Program in Rwanda (unpublished data).Clinical data were obtained from Rwanda (a relatively high-transmission area) across 6 sites between 2013 and 2015, where patients were treated with either AR-LF or DHA-PPQ and genotyped at msp-1, msp-2, and glurp. In patients treated with AS-AQ, 12 recurrences were observed, of which 5 could be classified as reinfection or recrudescence. (Seven patients had incomplete genetic data.) In patients treated with AS-PYN, 14 recurrences were observed, of which 12 could be classified as reinfection or recrudescence (2 had incomplete genetic data). In patients treated with DHA-PPQ, 67 recurrences were observed, of which 48 could be classified as reinfection or recrudescence. (Nineteen had incomplete genetic data.) These data were initially presented internally to the National Malaria Control Program in Cambodia. A description of the AS-PYN trials has already been published  were obtained from 6 sites between 2014 and 2016. Patients were treated with either artesunate plus amodiaquine (AS-AQ), artesunate plus pyronaridine (AS-PYN), or DHA-PPQ and genotyped at ublished .msp-1, msp-2, and glurp alleles at the initial sample and any recurrent sample) were reinterpreted using the novel molecular correction algorithms described in For all data, the genetic signals (i.e., the The R code used to generate the results describe herein is available from the authors. The reanalyzed trial data sets are likewise available from the authors."}
+{"text": "Plasmodium vivax malaria requires treatment with a blood schizonticide and a hypnozoitocide (primaquine) to eradicate the dormant liver stages. There has been uncertainty about the operational effectiveness and optimum dosing of the currently recommended 14-day primaquine (PQ) course.Radical cure of A two centre, randomized, open-label, two arm study was conducted in South India. Patients were randomized to receive either high dose (0.5\u00a0mg base/kg body weight) or conventional dose (0.25\u00a0mg/kg) PQ for 14\u00a0days. Plasma concentrations of PQ and carboxyprimaquine (CPQ) on the 7th day of treatment were measured by reverse phase high performance liquid chromatography. Study subjects were followed up for 6\u00a0months. Recurrent infections were genotyped using capillary fragment length polymorphism of two PCR-amplified microsatellite markers (MS07 and MS 10).Fifty patients were enrolled. Baseline characteristics and laboratory features did not differ significantly between the groups. Mean age of the study population was 42\u2009\u00b1\u200916.0 years. Recurrences 80\u2013105\u00a0days later occurred in 4 (8%) patients, two in each the groups. All recurrences had the same microsatellite genotype as that causing the index infection suggesting all were relapses. One relapse was associated with low CPQ concentrations suggesting poor adherence.This small pilot trial supports the effectiveness of the currently recommended lower dose (0.25\u00a0mg/kg/day) 14\u00a0day PQ regimen for the radical cure of vivax malaria in South India.Trial registration Clinical Trials Registry-India, CTRI/2017/03/007999. Registered 3 March 2017, http://ctri.nic.in/Clinicaltrials/regtrial.php?modid=1&compid=19&EncHid=82755.86366. Plasmodium vivax infection is endemic , hypnozoitocidal drug- PQ regimen along with its initiation date. Cases were classified into severe and non-severe cases as per WHO severity criteria . The subPrimaquine 0.5\u00a0mg base/kg/day for 14\u00a0days .Primaquine 0.25\u00a0mg base/kg/day for 14\u00a0days .After completion of treatment of acute malaria with a schizonticidal drug [either chloroquine 25\u00a0mg base/kg or ACT (astesunate with doxycycline OR artemether-lumefantrine) as per the treating clinician\u2019s judgement of severity], patients were randomized into the study. A block randomization was used to allocate subjects to the groups. Five blocks of size 10 were used. The randomization sequence within each block was done using a lottery method. Eligible patients were allocated to one of the following two treatment groups.The subjects were advised to take the drug once a day in the morning after breakfast. Drug dosing was not observed. They were counselled about the importance of PQ for the effective cure of malaria and adherence to medication. Additionally, patients were counselled about personal protective measures against mosquito bite.The study subjects were followed up on day 7 of PQ treatment for PQ and CPQ estimation. Patients were asked to come for blood draw, 2\u00a0h after the intake of PQ, in a fed state. Self-reported time after intake of PQ to blood collection was noted. Three ml of venous plasma was collected in EDTA vacutainer for drug estimation. Participants were questioned about compliance to study medication, and confirmed by used blister packs of PQ.Any adverse events reported by the subject were noted. Patients were encouraged to return if ill. During the follow-up period, peripheral blood smears for parasitic forms were examined on day 28 from onset of illness, followed by a periodic examination by the end of every second month until the completion of 6\u00a0months, and if any fever episodes occurred in the follow-up period, the subjects\u2019 blood smears were examined for parasitic forms. Treatment adherence, mosquito repellent and bed net usage, intercurrent febrile illnesses and any other possible symptoms of malaria were recorded and documented. In cases with microscopy confirmed recurrence of malaria, patients were treated with CQ/ACT as per the severity of illness followed by PQ 0.5\u00a0mg/kg/day for 14\u00a0days.During the initial recruitment, 2\u00a0ml of blood sample was collected from all study subjects in heparinized vacutainers and preserved in frozen conditions until further testing. Two ml of heparinized blood was also collected from patients diagnosed (smear positive) with recurrence of malaria. DNA was extracted from the heparinized blood specimens of patients (n\u2009=\u20094) collected during initial and recurrent infections of malaria. For DNA extraction, HiPurA\u2122 Blood Genomic DNA Mini Purification kit meant for DNA isolation from fresh or frozen blood  was used as per the manufacturer\u2019s instructions.P. vivax was performed based on sequence repeats in the microsatellites markers MS7 and MS10 using primers and the method described by Karunaweera et al.  patients included in the present study, 2 patients in each arm. Tables\u00a0, 3. All Genotyping suggested all four recurrences were homologous (isogenic) relapses. All four primary infections were of a single genotype and were different on microsatellite typing, but all four recurrences had the same alleles as the primary infections. Results of the capillary fragment length analysis of the MS7 and MS10 microsatellite markers from patients with recurrent malaria are depicted in Fig.\u00a0This pilot study assessed the tolerability, safety and effectiveness of two radical cure PQ dosage regimens . Both proved very effective. PQ is readily absorbed from the gastrointestinal tract, undergoes high first pass metabolism and is extensively distributed into body tissues. The plasma concentration peaks within 1\u20133\u00a0h after ingestion. PQ has a plasma elimination half-life of 4\u20139\u00a0h in healthy adults . The metPlasmodium vivax cases recorded at health facilities during the non-transmission season likely arise from reactivation of dormant liver-stages. In the transmission season P. vivax cases are presumed to be arising both from new mosquito infections and relaps. [The present cohort consisted predominantly males; the gender imbalance is typical of patient population in the study setting and that of many others in malaria endemic areas of Asia , 17\u201319.  relaps. . The int relaps. \u201328.This was a small study, and one-third of recruited patients did not complete the full 6\u00a0months follow up. This comparison was, therefore, underpowered to show significant differences between the two treatment arms. Nevertheless the effectiveness results do support the current dosing recommendation of PQ 3.5\u00a0mg base/kg total dose radical cure regimen in South India.The recurrence rate of vivax malaria in this study was 8% at 6\u00a0months follow-up and it occurred equally in high dose as well as low dose PQ regimens. Recurrence cases were of non-severe phenotype and genotyping suggested all four recurrences were homologous (isogenic) relapses. This small pilot trial supports the effectiveness of the currently recommended lower dose (0.25\u00a0mg/kg/day) 14\u00a0day PQ regimen for the radical cure of vivax malaria in South India."}
+{"text": "Plasmodium vivax malaria relapses, but adherence to the standard 14-day regimen is poor. We aimed to assess the efficacy of a shorter course (7 days) of primaquine for radical cure of vivax malaria.Primaquine is the only widely used drug that prevents P vivax parasitaemia during the 12-month follow-up period, assessed in the intention-to-treat population. A margin of 0\u00b707 recurrences per person-year was used to establish non-inferiority of the 7-day regimen compared with the 14-day regimen. This trial is registered at ClinicalTrials.gov (NCT01814683).We did a randomised, double-blind, placebo-controlled, non-inferiority trial in eight health-care clinics . Patients (aged \u22656 months) with normal glucose-6-phosphate dehydrogenase (G6PD) and presenting with uncomplicated vivax malaria were enrolled. Patients were given standard blood schizontocidal treatment and randomly assigned (2:2:1) to receive 7 days of supervised primaquine (1\u00b70 mg/kg per day), 14 days of supervised primaquine (0\u00b75 mg/kg per day), or placebo. The primary endpoint was the incidence rate of symptomatic P vivax malaria was 0\u00b718 (95% CI 0\u00b715 to 0\u00b721) recurrences per person-year for 935 patients in the 7-day primaquine group and 0\u00b716 (0\u00b713 to 0\u00b718) for 937 patients in the 14-day primaquine group, a difference of 0\u00b702 . The incidence rate for 464 patients in the placebo group was 0\u00b796 (95% CI 0\u00b783 to 1\u00b708) recurrences per person-year. Potentially drug-related serious adverse events within 42 days of starting treatment were reported in nine (1\u00b70%) of 935 patients in the 7-day group, one (0\u00b71%) of 937 in the 14-day group and none of 464 in the control arm. Four of the serious adverse events were significant haemolysis (three in the 7-day group and one in the 14-day group).Between July 20, 2014, and Nov 25, 2017, 2336 patients were enrolled. The incidence rate of symptomatic recurrent P vivax malaria.In patients with normal G6PD, 7-day primaquine was well tolerated and non-inferior to 14-day primaquine. The short-course regimen might improve adherence and therefore the effectiveness of primaquine for radical cure of UK Department for International Development, UK Medical Research Council, UK National Institute for Health Research, and the Wellcome Trust through the Joint Global Health Trials Scheme (MR/K007424/1) and the Bill & Melinda Gates Foundation (OPP1054404). Plasmodium vivax malaria is a major cause of morbidity and a contributor to mortality in Asia, Oceania, the Horn of Africa, and the Americas.P vivax relapses.14Evidence before this studyPlasmodium vivax, WHO recommends a high-dose primaquine regimen of 7 mg/kg administered over 14 days . In 2012, at the inception of the study, a systematic review was done using the search terms \u201cvivax\u201d and \u201cprimaquine\u201d to identify all articles in the English language on PubMed, MEDLINE, Web of Science, Embase, and the Cochrane Database of Systematic Reviews between Jan 1, 1950, and Dec 31, 2011. The search identified 18 studies that compared the efficacy of schizontocidal treatment with and without primaquine. However, wide variation in study design and duration of follow-up confounded comparison of antirelapse efficacy across different endemic settings. The efficacy of an unsupervised 14-day primaquine regimen was significantly lower than that of a daily observed 14-day regimen (odds ratio 0\u00b718 [95% CI 0\u00b706\u20130\u00b757]). Only one study compared the efficacy of a high-dose primaquine regimen administered over 7 or 14 days, and although the results suggested equivalence, patients were only followed up for 28 days. A 2019 randomised controlled trial compared the antirelapse efficacy of high-dose primaquine administered over 7 or 14 days in Thai patients, and found non-inferiority at a 10% margin. However the generalisability of these findings across other endemic locations has yet to be confirmed.The primary determinant of primaquine radical cure efficacy is the total dose of primaquine administered. In areas with frequent-relapsing strains of Added value of this studyP vivax was done at eight sites in four countries with differing P vivax endemicity, to assess the safety and radical curative efficacy of a 7-day high-dose (1 mg/kg per day) primaquine regimen compared with a 14-day regimen (0\u00b75 mg/kg per day). The study shows that, in patients screened with the qualitative fluorescent spot test as glucose-6-phosphate dehydrogenase normal, the 7-day primaquine regimen was non-inferior to the 14-day regimen, with an acceptable safety profile.Our large multicentre clinical trial in patients with Implications of all the available evidenceP vivax malaria.In patients with normal G6PD, a 7-day high-dose course of primaquine is well tolerated and is as effective as a 14-day course. The shorter regimen has the potential to improve adherence and therefore the effectiveness of primaquine for the radical cure of P vixax are prevalent.WHO recommends primaquine (0\u00b75 mg/kg per day for 14 days) for patients with normal G6PD activity in areas where frequent relapsing strains of This was a randomised, double-blind, placebo-controlled, non-inferiority trial in patients with normal G6PD and uncomplicated vivax malaria presenting at two health-care clinics in each of the following countries: Afghanistan, Ethiopia, Indonesia, and Vietnam . EthicalAt the time of the study, primaquine radical cure was national policy in Indonesia, Vietnam, and Afghanistan, but not Ethiopia. The justification of a placebo control group in the Indonesia, Vietnam, and Afghanistan sites was based on clinical equipoise, considering the risks and benefits of different treatment regimens and the importance of controlling for variation in the risk of relapse. Rapid initial clinical cure was achieved with highly efficacious schizontocidal treatment, which was administered to all patients. Further details on the rationale for including a placebo control group have been published previously.Patients presenting with uncomplicated vivax malaria (mono-infection or mixed infection) who had fever or a history of fever within the past 48 h, were aged older than 6 months, weighed at least 5 kg, had a haemoglobin concentration of 9 g/dL or more, and had normal G6PD status as assessed by the fluorescent spot test were eligible for enrolment. Patients were excluded if they were pregnant or lactating, could not tolerate oral treatment, had previous haemolytic episodes or blood transfusion within the past 90 days, or had signs of severe malaria. Other exclusion criteria included any hypersensitivity to study drugs, or concomitant medication with the potential to cause haemolysis or interfere with the pharmacokinetics of the study drugs.Before enrolment, written informed consent was obtained from the patient or their guardian. Written assent was also obtained from patients who were 12\u201318 years old. Patients with G6PD deficiency were excluded from the randomised trial, but were enrolled into a parallel observational group and treated with chloroquine or dihydroartemisinin-piperaquine plus supervised primaquine (0\u00b775 mg/kg) once a week for 8 weeks.Eligible patients were treated with a blood schizontocidal drug and randomly assigned in a 2:2:1 ratio to a 7-day primaquine regimen, a 14-day primaquine regimen, or placebo. The allocation ratio of 2:2:1 was based on the sample size requirements for the non-inferiority comparison of a 7-day primaquine regimen with a 14-day regimen, and the superiority comparisons of the 7-day and 14-day primaquine regimens with the placebo. Randomisation was done using STATA version 14.1 , which generated blocks of 20 for each dosing band. The independent statistician who generated the randomisation list and selected code letters for primaquine or placebo was not otherwise involved in the conduct of the trial and did not visit any of the study sites. Identical primaquine and placebo tablets were produced by the same manufacturer  and packaged in blister packs . Randomisation in blocks of 20 was done at each site. Boxes of primaquine or placebo tablets for each of three bodyweight bands were provided and numbered sequentially. Patients were allocated randomly to a treatment group and given the next numbered box within the appropriate weight band of that group. Participants and all of the local study team were masked to treatment assignments.P vivax, which was chloroquine in Ethiopia, Afghanistan, and Vietnam and dihydroartemisinin-piperaquine in Indonesia, according to local guidelines  or subsequent symptomatic P vivax recurrence were treated with open-label supervised primaquine (0\u00b75 mg/kg for 14 days).At enrolment, a medical history was taken, a physical examination was done, and antimalarial treatment with chloroquine or dihydroartemisinin-piperaquine was initiated. Primaquine or placebo was usually started on the day of enrolment, although sites had clinical discretion to start the next day. After completion of treatment, patients were asked to return weekly until day 42 and then monthly for 1 year. At each visit, a medical history was taken, a symptom questionnaire was done, and any adverse events or serious adverse events were recorded. Patients were encouraged to report to the study centre if they became ill. Patients who missed their scheduled follow-up visits were contacted by study staff and encouraged to return to the study centre for review. Blood films were examined immediately for all symptomatic patients, but otherwise stored for later examination. Recurrent parasitaemia with any 6 erythrocytes per \u03bcL for thin film counts. Microscopists were trained in study laboratory procedures on-site and continuous quality control was implemented at all sites. Approximately 10% of slides, including all the slides from day 0, the day of recurrent parasitaemia, and the 6-month follow-up visits were assessed periodically over the course of the trial by expert malaria microscopists at the Mahidol-Oxford Tropical Medicine Research Unit in Bangkok, Thailand. Results of the quality control procedures are presented in the At the initial screening, venous blood was collected and tested for G6PD deficiency with the qualitative fluorescent spot test . Standard thick (6 \u03bcL blood on a 12-mm diameter template) and thin malaria films were prepared at each visit for Giemsa staining  according to the WHO Research Malaria Microscopy Standards . Blood fP vivax parasitaemia (mono-infection or mixed) over 12 months, with the primary analysis comparing this outcome in patients treated with 7-day and 14-day primaquine. The primary endpoint was calculated by site and then pooled across all sites. Secondary 12-month efficacy endpoints were the incidence rate and incidence risk  of any (symptomatic or asymptomatic) P vivax parasitaemia, Plasmodium falciparum parasitaemia, or all-species parasitaemia, comparing the treatment groups to each other and with the placebo group. Other efficacy endpoints were the incidence risks of P vivax at 28 and 42 days, the proportion of patients with P vivax parasitaemia on days 1, 2, and 3, and the proportion with fever on days 1, 2, and 3. The number of recurrences avoided per 1000 patients and the number needed to treat (NNT) to avoid one recurrence were derived from the rate difference.The primary endpoint was defined as the incidence rate of symptomatic Safety endpoints were the incidence risk of severe anaemia (haemoglobin <7 g/dL) or transfusion, an acute drop in haemoglobin of more than 5 g/dL within 7 days, haemoglobin concentration on day 3 and day 7, the median time to haemoglobin nadir, grade 3 or 4 adverse events within 42 days, and severe adverse events.Efficacy and safety outcomes from the patients with G6PD deficiency who were given weekly primaquine and cost data will be reported separately.P vivax recurrences per person-year in each group, a non-inferiority margin of 0\u00b707 recurrences per person-year, a projected loss of 20% of patients to follow-up or protocol violations, a one-sided significance level of 2\u00b75%, and a power of 80%. Two additional sites in Ethiopia were included to speed up recruitment. The 25% increase in sample size improved the generalisability of the study, and increased the power to detect non-inferiority between the 7-day and 14-day primaquine groups to 86%. Power calculations for superiority comparisons with the placebo group are provided in the The original sample size calculation of 750 patients per primaquine group was based on an assumed incidence rate of 0\u00b72 symptomatic P vivax recurrences per person-year between the 7-day and 14-day primaquine group was estimated using weighted least-squares regression with a robust standard error. For the secondary efficacy endpoints comparing 7-day primaquine versus placebo, 14-day primaquine versus placebo, and 7-day versus 14-day primaquine, incidence rate ratios were estimated using negative binomial regression . Cumulative incidence risks were estimated by Kaplan-Meier analysis. Hazard ratios (HRs) were estimated by Cox regression . The main analysis was done on the intention-to-treat population. Details of censoring for incidence risk analyses, sensitivity analyses, investigation of study site heterogeneity, and safety analyses are in the To assess the primary efficacy endpoint for radical cure, the absolute incidence rate difference (95% CI) in symptomatic ClinicalTrials.gov (NCT01814683).The Data and Safety Monitoring Board did a blinded safety review every 6 months. The trial was registered at The funders of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. All authors had full access to the data in the study and had final responsibility for the decision to submit for publication.Between July 20, 2014, and Nov 25, 2017, 11\u2008585 patients were screened for inclusion into the study, of whom 9197 did not meet the enrolment criteria and 50 were G6PD-deficient . 2338 paP vivax parasitaemia was 3440 parasites per \u03bcL  of 2309 patients becoming afebrile within 24 h of starting treatment. Overall, 2217 (96\u00b70%) of 2310 patients cleared peripheral parasitaemia within 2 days of starting treatment, with no significant difference between treatment groups .P vivax malaria was 0\u00b718 recurrences per person-year (95% CI 0\u00b715 to 0\u00b721) in the 7-day primaquine group, 0\u00b716 (0\u00b713 to 0\u00b718) in the 14-day primaquine group, and 0\u00b796 (0\u00b783 to 1\u00b708) in the placebo group (The incidence rate of symptomatic recurrent bo group . The incbo group . The numThe incidence rate of any (symptomatic or asymptomatic) recurrent vivax malaria at 1 year was 0\u00b723 recurrences per person-year (95% CI 0\u00b719\u20130\u00b727) in the 7-day primaquine group and 0\u00b719 (0\u00b716\u20130\u00b723) in the 14-day primaquine group p=0\u00b721; . These iP vivax at 1 year was 14\u00b728% (95% CI 11\u00b775\u201317\u00b729) in the 7-day primaquine group and 12\u00b772% (10\u00b719\u201315\u00b782) in the 14-day primaquine group  recurrences per person-year in the 7-day primaquine group, 0\u00b707 (0\u00b705\u20130\u00b709) in the 14-day primaquine group, and 0\u00b707 (0\u00b704\u20130\u00b710) in the control group.At the end of the 1-year follow-up, the incidence rate of symptomatic P vivax parasitaemia within 28 days. The incidence risk of any (symptomatic or asymptomatic) P vivax parasitaemia at day 28 was 0\u00b723% (95% CI 0\u00b706\u20130\u00b793) in the 7-day primaquine group, 0\u00b733% (0\u00b710\u20131\u00b700) in the 14-day primaquine group, and 1\u00b771% (0\u00b782\u20133\u00b756) in the placebo group. At 42 days, the incidence risk of any P vivax parasitaemia was 0\u00b787% (95% CI 0\u00b741\u20131\u00b781) in the 7-day primaquine group and 0\u00b782% (0\u00b739\u20131\u00b772) in the 14-day primaquine group (p=0\u00b778). Compared with the placebo group, the HR was 0\u00b711  for both treatment groups (14 (0\u00b76%) of 2336 patients had recurrent t groups .Reporting of serious adverse events and adverse events varied significantly by site . 27 seriAdverse events within 42 days with a severity grade of 3 or 4 were reported in eight patients, mostly in the 7-day primaquine group . Six of The median time to haemoglobin nadir was 3 days (IQR 3\u20137) in the placebo group and 3 days (3\u201313) in both primaquine groups. The mean absolute decreases in haemoglobin at day 3 were 0\u00b752 g/dL (SD 1\u00b719) in the 7-day primaquine group, 0\u00b762 g/dL (1\u00b709) in the 14-day group, and 0\u00b772 g/dL (1\u00b712) in the placebo group. The corresponding mean percentage change in haemoglobin between day 0 and day 3 was \u22123\u00b760% (SD 9\u00b717) in the 7-day group, \u22124\u00b753% (8\u00b747) in the 14-day group, and \u22125\u00b726% (8\u00b764) in the placebo group . The meaThis large, multinational, placebo-controlled trial confirms the high antirelapse efficacy of primaquine at a total dose of 7 mg/kg and demonstrates non-inferiority of 7-day primaquine compared with 14-day primaquine. A supervised 7-day course of high-dose primaquine prevented an estimated 781 recurrences per 1000 patients, equivalent to an NNT of 1\u00b728. By comparison, the 14-day course of primaquine prevented 800 recurrences per 1000 patients, equivalent to an NNT of 1\u00b725. The 7-day course was associated with a slightly higher frequency of adverse events than the 14-day course, which were mostly attributable to gastrointestinal symptoms.P vivax malaria for more than 60 years. WHO recommends the use of primaquine after screening for G6PD deficiency, but in most malaria endemic countries, reliable G6PD testing is unavailable. Therefore, particularly in areas of high prevalence of G6PD deficiency, primaquine is rarely used because of concerns over safety.24Primaquine has been used for the radical cure of For safety reasons, primaquine is widely recommended as a 14-day regimen; however, adherence to such a long course of treatment for an acute febrile illness can be low.vs 3\u00b70%). Ingesting primaquine with food can improve gastrointestinal tolerability considerably and should be encouraged in all patients reporting symptoms.Patients in the 7-day primaquine group in our study received twice the daily dose of those in the standard 14-day high-dose regimen (0\u00b75 mg/kg per day) and four times the dose of the 14-day low-dose regimen (0\u00b725 mg/kg per day) that is recommended by most national malaria control programmes.In malaria-endemic areas, the most widely used test to diagnose G6PD deficiency is the fluorescent spot test, a qualitative assay that requires laboratory facilities and can only identify individuals with less than 30% enzyme activity.P vivax radical cure.Tafenoquine, a slowly eliminated 8-aminoquinoline, was licensed in the USA and Australia in 2018. When combined with chloroquine, tafenoquine offers a single dose option for P vivax parasitaemia, was not compromised because the additional Ethiopian sites ensured that the total number of patient days of follow-up exceeded that required to achieve 80% power for non-inferiority. In addition, a significant difference was found in the risk of adverse events between sites, which probably reflected differences in recording and perception of symptoms rather than true differences in events between the patient cohorts. Although most adverse events were reported from the site in Hanura, Indonesia, the double-blinded treatment allocation means that this is unlikely to have affected the robustness of the comparison between treatment regimens.A major strength of our study is its multicentre design, with patients enrolled from four vivax-endemic countries with diverse patient populations and relapse periodicity. Therefore, our findings are likely to be generalisable to other vivax-endemic regions. Our study also has several limitations. Approximately one third of patients did not complete the 1-year follow-up, mainly because of the late start of the study in Ethiopia, where a high proportion of patients were unable to complete follow-up before study termination. However, the primary outcome, incidence rate of recurrent In conclusion, the efficacy of a shorter 7-day course of primaquine was non-inferior to the standard 14-day course in patients with normal G6PD and had an acceptable safety and tolerability profile. The reduced time of treatment has the potential to improve adherence and therefore the effectiveness of radical cure in vivax-endemic countries. The high-dose 7-day primaquine regimen was well tolerated, but it requires that G6PD deficiency can be excluded reliably. In malaria-endemic countries, G6PD testing at the 30% threshold can now be achieved using point-of-care qualitative and quantitative rapid diagnostic tests,applicants who provide a sound proposal to the Mahidol Oxford Tropical Medicine Research Unit Data Access Committee.Deidentified individual participant data will be available immediately after publication to"}
+{"text": "Plasmodium falciparum artemisinin drug resistance threatens global malaria public health gains. Limited data exist to define the extent of P. falciparum artemisinin resistance southeast of the Greater Mekong region in Malaysia.Spreading kelch13 polymorphisms associated with P. falciparum artemisinin resistance were also evaluated in P. falciparum isolates collected from patients presenting to health facilities predominantly within the tertiary referral area of western Sabah between 2012 and 2016.A clinical efficacy study of oral artesunate  daily for 3\u00a0days was conducted in patients with uncomplicated falciparum malaria and a parasite count\u2009<\u2009100,000/\u00b5L admitted to 3 adjacent district hospitals in Sabah, East Malaysia. On day 3 and 4 all patients were administered split dose mefloquine  and followed for 28\u00a0days. Twenty-one P. falciparum isolates evaluated were wild-type for kelch13 markers.In total, 49 patients were enrolled and treated with oral artesunate. 90% (44/49) of patients had cleared their parasitaemia by 48\u00a0h and 100% (49/49) within 72\u00a0h. The geometric mean parasite count at presentation was 9463/\u00b5L , with a median time to 50% parasite clearance of 4.3\u00a0h (IQR 2.0\u20138.4). There were 3/45 (7%) patients with a parasite clearance slope half-life of\u2009\u2265\u20095\u00a0h. All 278 P. falciparum in this pre-elimination setting in Sabah, Malaysia. Current guidelines recommending first-line treatment with ACT remain appropriate for uncomplicated malaria in Sabah, Malaysia. Ongoing surveillance is needed southeast of the Greater Mekong sub-region.There is no suspected or confirmed evidence of endemic artemisinin-resistant The online version of this article (10.1186/s12936-018-2593-x) contains supplementary material, which is available to authorized users. Plasmodium falciparum artemisinin resistance from Southeast Asia threatens global malaria elimination efforts  1], fo, fo1], fP. falciparum artemisinin resistance were evaluated using PCR to detect 21 single nucleotide polymorphisms on the kelch13 gene  and analysed using Stata (version 12). Microscopic asexual parasitaemia and gametocytaemia were calculated from thick blood smears . Best-fiver time .P. falciparum on screening microscopy were enrolled into the study .At 48\u00a0h after treatment 44/49  of patients were negative for parasites microscopically on blood smear. All patients were negative by 72\u00a0h, and remained negative up to 28\u00a0days post-treatment Table\u00a0. The med50) was 4.3\u00a0h (IQR 1.9\u20138.4), and time to 99% parasite clearance was 19.6\u00a0h (IQR 14.3\u201325.6). The mean slope of the loge normalised parasite clearance curve was 0.27 (95% CI 0.24\u20130.30). The median half-life of the clearance slope was 2.7\u00a0h (IQR 2.2\u20133.5), with 3 (7%) of patients having a clearance slope half-life greater than 5\u00a0h. There was no difference in the median clearance slope half-life when comparing patients enrolled from each year during the study period.Detailed parasite clearance estimation (using WWARN methodology ) could bP. falciparum isolates were wild-type for the 21 kelch13 SNPs evaluated (GenBank Accession Numbers: MH721937\u2013722214), including 98 (100%) of those subsequently tested for the F446I polymorphism (unpublished). Cases were predominantly distributed from the West Coast administrative division of Sabah (158 [58%]), followed by the northern Kudat division (111 [41%]), with 5 (2%) additional cases from the eastern Sandakan division  in\u2009\u2265\u20093% of a treated population [Ongoing therapeutic efficacy monitoring for artemisinin derivatives should incorporate the use of pulation . Howeverpulation . Conventpulation .kelch13 molecular study component. Falciparum malaria cases able to have kelch13 marker surveillance conducted were predominantly from the western Sabah catchment area of the tertiary referral hospital in the state capital Kota Kinabalu. Although it is possible more remote interior areas were underrepresented, this area has both the highest population density and falciparum malaria transmission in Sabah, as well as significant numbers of official and unofficial migrants from surrounding falciparum-endemic countries.The ongoing decline in falciparum malaria cases in Sabah over the study period limited both the number of those able to be enrolled in the in vivo parasitological assessment, and also the overall number evaluated in the P. falciparum artemisinin resistance in this population given the absence of kelch13 polymorphisms.Accurate evaluation of parasite clearance was possible by the study design utilising optimal dosing of oral artesunate (4\u00a0mg/kg) given in narrow weight bandings, with all patients receiving close to the target total dose of 12\u00a0mg/kg. In contrast, the first-line ACT used in Malaysia, artemether-lumefantrine, has an effective artemisinin derivative component dose of 1.8\u00a0mg/kg, given twice daily, with larger weight-bandings for standard WHO-recommended use known to result in lower efficacy among young children from Asia receiving\u2009\u2264\u200960\u00a0mg/kg total lumefantrine dose . HoweverPlasmodium falciparum artemisinin resistance was not evident in Sabah, Malaysia from in vivo parasite clearance, or surveillance of kelch13 molecular markers. Current ACT regimens should continue to be used as the first-line blood-stage treatment for all Plasmodium species in Malaysia. With Malaysia approaching the elimination of falciparum malaria, ongoing molecular surveillance for artemisinin and partner drug resistance will be an important adjunctive tool in achieving and maintaining this goal. Wider surveillance for artemisinin and partner drug-resistant P. falciparum in other countries to the south and east of the Greater Mekong region will also be an important component in the success of their malaria elimination goals.Additional file 1.kelch13 molecular marker reference table."}
+{"text": "Plasmodium is exposed to an effective concentration of the drug.A fixed-dose combination of mefloquine with artesunate was evaluated in cases of falciparum malaria in the Brazilian Amazon basin with acceptable efficacy, safety and tolerability. However, there are no data on the pharmacokinetics of mefloquine in this coformulation in Brazil, which is valuable to evaluate whether Plasmodium falciparum using two tablets of a fixed-dose combination of artesunate (100\u00a0mg) and mefloquine base (200\u00a0mg) once daily and over 3 consecutive days. Serial blood samples were collected at admission and throughout 672\u00a0h post-administration of the drugs. Mefloquine was measured in each blood sample by high-performance liquid chromatography. The pharmacokinetic parameters were determined by non-compartmental analysis.A prospective, single-arm study was conducted in male patients with slide-confirmed infection by max) was 2.53\u00a0\u00b5g/ml, the area-under-the-curve (AUC0\u2013\u221e) was 359\u00a0\u00b5g/ml\u00a0h, the observed clearance (Cl/f) was 0.045\u00a0l/kg/h and the volume of distribution (V/f) was 14.6\u00a0l/kg. Mefloquine concentrations above 0.5\u00a0\u00b5g/ml were sustained for a mean time of 9.2\u00a0days.A total of 61 patients were enrolled in the study and 450 whole blood samples were collected for mefloquine measurement. The mefloquine half-life was 10.25\u00a0days, the maximum concentration (CThe pharmacokinetic parameters of mefloquine determined in the study suggest an adequate exposure of parasite to mefloquine in the multiple oral dose regimen of the fixed dose combination of mefloquine and artesunate. Plasmodium falciparum is responsible for 15% of the burden of the disease in this endemic scenario, with approximately 13,000 cases each year [\u00ae) is recommended by the local health authorities as a first-line therapy for uncomplicated falciparum malaria in a 3-day course of 80 and 480\u00a0mg every 12\u00a0h in patients weighing 35\u00a0kg or more [\u00ae efficacy in the Brazilian Amazon basin, however, ongoing vigilance is needed to detect the emergence of resistance to artemisinin or partner drugs.Malaria remains one of the major public health issues worldwide. In Brazil, most of the cases occur in the Amazon basin, with about 148,000 cases reported each year. \u00ae in this endemic setting [Recent studies done with a fixed-dose formulation of artesunate with mefloquine (MQAS) manufactured by Farmanguinhos  in single 3-day doses of two tablets of 100\u00a0mg of artesunate and 200\u00a0mg of mefloquine base have shown acceptable efficacy, safety and tolerability of MQAS in uncomplicated falciparum malaria cases. Moreover, the coformulation reduces the pill number compared to the standard treatment. Thus, MQAS may be a potential alternative to Coartem setting .P. falciparum introduced in 1984 for clinical use in Thailand [Mefloquine, the partner drug, is a quinolone methanol compound effective against the asexual blood stage of Thailand . The phaThailand \u201314. The Thailand .As the clinical studies of efficacy and tolerability of a new anti-malarial drug formulation should be accompanied by the estimation of exposure of parasite to adequate concentrations of the anti-malarial drug, it was investigated in the present study the pharmacokinetics parameters of mefloquine given as a fixed-dose formulation with artesunate once daily in patients with uncomplicated falciparum malaria from the Brazilian Amazon basin.P. falciparum. The exclusion criteria included patients with signs or symptoms of severe disease , parasitaemia over 5%, mixed infections, history of psychiatric disorders, overweight and underweight, altered levels of creatinine, altered levels of aspartate aminotransferase (AST) or alanine aminotransferase (ALT), known hypersensitivity or allergy to mefloquine or artesunate and use of mefloquine in the previous 90\u00a0days.This was an open-label, single arm study carried out at Reference Centre for Tropical Diseases in the municipality of Macapa, AP, in the Brazilian Amazon Basin. Patients recruited for attendance in the health facility came from rural communities or gold mines located at the border between Brazil and French Guyana. Inclusion criteria for enrolment in the study were: adult male\u2009>\u200918\u00a0years of age with slide-confirmed mono-infection by Each patient received two tablets of a fixed-dose combination of mefloquine with artesunate  containing 100\u00a0mg of artesunate and 200\u00a0mg of mefloquine base per tablet, once a day and over 3 consecutive days [Serial venous blood samples (4\u00a0ml) were taken from each patient for mefloquine measurement before the commencement of the treatment (D0) and on days 1, 3, 5, 7, 14, 21, 28, and 35. On day 1, a blood sample was collected from each patient 30\u00a0min before anti-malarial drug intake. A similar time of blood sampling was adopted for all days of the study for all patients. After collection, whole blood samples were immediately stored at -80 C until analysis.Mefloquine was measured by a reversed-phase HPLC system with an ultraviolet detector  after liquid\u2013liquid extraction from the whole blood with methyl tert-butyl ether at pH 4.0. The separation was carried out on a reversed-phase column . The mobile phase consisted of acetonitrile\u2013phosphate buffer  (42:58). Quinidine (2.5\u00a0\u03bcg/ml) was used as internal standard. Whole blood spiked with mefloquine in concentrations of 0.05, 0.1, 0.5, 1, 2, and 4\u00a0\u03bcg/ml were used to estimate the within-day and day-to-day coefficients of variation as well as the recovery of the method. The limit of detection was considered as the lowest concentration of mefloquine that may be differentiated from the background noise of the ultraviolet detector and the limit of quantification was considered as the lowest concentration of mefloquine that was determined with a coefficient of variation below 10%. The assay was linear from 0.05 to 4\u00a0\u03bcg/ml. The within-day and day-to-day coefficients of variation were 6.7 and 8.1%, respectively. The limit of quantification was 0.05\u00a0\u03bcg/ml and the limit of detection was 0.03\u00a0\u03bcg/ml. The mean recovery was 90%. The stability of blank whole blood spiked with mefloquine (0.1\u00a0\u03bcg/ml) was 60\u00a0days at \u2212\u200980\u00a0\u00b0C. There was no significant interference of primaquine, quinine, chloroquine, desethyl-chloroquine, carboxy-mefloquine, and acetaminophen in the detection of mefloquine .Parasite count was done in Giemsa-stained thick films every day until it became negative and then on days 7, 14, 21, 28, 35, and 42. An experienced microscopist examined the blood films using 100\u00d7 (oil immersion) objectives. The parasite density was expressed as the number of parasites per \u03bcl of blood, which was derived from the number of parasites per 200 white blood cells, considering a total white blood cell count of 8000. The limit of detection of the parasites was 40/\u00b5L . Clinica0\u2212t was estimated from the time of drug administration to the time of the last measurable concentration by using the linear trapezoidal rule and the extrapolation to infinity AUC0\u2212\u221e was determined by dividing the last measurable mefloquine concentration by \u03bbZ. To obtain the terminal elimination rate constant (\u03bbZ), the mefloquine concentrations were log transformed and fitted a linear regression model to the terminal phase of the concentration\u2013time profiles by using the method of least squares. The terminal elimination half-life (t1/2) was estimated by dividing ln2 by \u03bbZ. The apparent oral clearance per fraction of drug absorbed (CL/f) was derived from the ratio of the dose to AUC0\u2212\u221e. The apparent volume of distribution (V/f) was estimated from CL/f divided by \u03bbZ. Data were analysed using WinNonlin .The data were analysed by non-parametric methods. A non-compartmental pharmacokinetic analysis was used to calculate the pharmacokinetics parameters, which were estimated separately for each patient enrolled in the study. The maximum concentration (Cmax) was obtained directly from the whole blood concentration\u2013time profile. The AUCThe study was approved by SEAMA Faculty Ethical Committee under the number 079/08. All patients enrolled in the study were informed about the goals as well as the risks and benefits of the study. All patients provided written informed consent before entering the study following national guidelines.A total of 61 patients were recruited for the study and 495 whole blood samples were collected and analysed for mefloquine with an average of 8 time points (range 5\u20139 time points) for each patient. A total of 54 samples were missing due to loss to follow-up of participants.The median total dose of mefloquine received by patients of 21.05\u00a0mg/kg (range 15.3\u201324\u00a0mg/kg) was sufficient to achieve curative concentrations of the drug. The geometric mean of parasites at admission was 1900\u00a0\u00b5l (range 800\u20135100\u00a0\u00b5l). Parasitaemia was cleared rapidly in all patients and there was no recrudescence by day 42, suggesting a high therapeutic efficacy of MQAS to treat uncomplicated falciparum cases in this endemic area. Furthermore, there were no reports of vomiting or diarrhoea, but 7 patients (17.5%) showed nausea or insomnia during the treatment. The baseline characteristics of participants are shown in Table\u00a0All pre-dose samples (d0) had undetectable levels of mefloquine. The median concentrations of mefloquine were 1.4\u00a0\u03bcg/ml (range 0.6\u20133.4\u00a0\u03bcg/ml), 2.53\u00a0\u03bcg/ml (range 1.5\u20133.88\u00a0\u03bcg/ml), 0.96\u00a0\u03bcg/ml (range 0.54\u20132.18\u00a0\u03bcg/ml), 0.8\u00a0\u03bcg/ml (range 0.5\u20131.4\u00a0\u03bcg/ml), 0.31\u00a0\u03bcg/ml (range 0.1\u20131.14\u00a0\u03bcg/ml), 0.23\u00a0\u03bcg/ml (range 0.06\u20130.6\u00a0\u03bcg/ml), 0.11\u00a0\u03bcg/ml (range 0.06\u20130.27\u00a0\u03bcg/ml) and 0.067\u00a0\u03bcg/ml (range 0.05\u20130.07\u00a0\u03bcg/ml) on days 1, 3, 5, 7, 14, 21, 28, and 42, respectively. Mefloquine concentrations above 0.5\u00a0\u03bcg/ml were sustained for a mean time of 9.2\u00a0days. The concentration\u2013time profile of mefloquine is shown in Fig.\u00a0The emergence and dissemination of ACT resistance in Southeast Asia constitute a threat to reduce the burden of falciparum malaria worldwide. The search for new compounds with anti-malarial action and the development of new ACT formulations has been a plausible strategy, adopted by several countries, to the fight against the disease. In this line, a fixed-dose formulation of MQAS was developed by Farmanguinhos, Brazil. In the current study, the pharmacokinetic parameters of MQAS were estimated. Furthermore, this is the first study to use a large data set to demonstrate the pharmacokinetic parameters of mefloquine in Brazilian patients with uncomplicated falciparum malaria. All patients had low parasite density and mild signs and symptoms of the disease, which is in line with the pattern of uncomplicated cases in the Brazilian Amazon Basin \u201320.In the study, the mean time for parasite clearance is aligned to studies in areas where MQAS remains effective. Moreover, there was no recrudescence of the infection by day 42. These results suggest a high therapeutic efficacy of the fixed-formulation of MQAS in this endemic area , 21, 22.The split-dose regimen of mefloquine was well tolerated by the patients with a low occurrence of side effects , 23\u201325. The non-compartmental modelling of mefloquine concentration\u2013time profile was used in the present study. This approach had been applied with success in studies of the pharmacokinetics of mefloquine \u20139, 25. TP. falciparum for a long period to effective concentrations of the drug in the blood.The coformulation MQAS involves a rapidly eliminated and fast-action artemisinin derivative and a slowly eliminated drug that kills the remaining parasites . MefloquThe comparisons of pharmacokinetic parameters of the present study with those derived from African and Thai patients presented some similarities. For instance, the mean half-life was comparable to mean half-life of 9.4\u00a0days found in Thai patients , 12. TheFurthermore, a considerable intra-individual and inter-individual variation of pharmacokinetic parameters of mefloquine was found in the study. A plausible explanation is that the data were normalized only by the weight of patients and not by other confounding factors, including age, parasitaemia at admission, time of fever clearance, and body mass index. This was considered the main limitation of the current study , 30, 31.P. falciparum to the fixed dose, which was corroborated by the efficacy of the treatment. Thus, artesunate-mefloquine is a reliable alternative treatment to Coartem\u00ae in this endemic area.The pharmacokinetic parameters of the coformulation of artesunate with mefloquine in a fixed-dose formulation suggest an adequate exposure of"}
+{"text": "Kelch 13 gene (pfk13) and dihydrofolate reductase/dihydropteroate synthase (pfdhfr/pfdhps) genes associated with artemisinin and SP resistance, respectively, and amplification in the pfplasmepsin2 gene for piperaquine resistance. A total of 295  eligible children were enrolled at three sites. PCR-corrected 100% adequate clinical and parasitological response and no parasitaemia on day-3 were observed for all patients in each treatment group. Of the 278 samples with interpretable molecular data, only 2.2% carried non-synonymous pfk13 mutants , which are not associated with artemisinin resistance. None of the 103 day-0 samples from the DAH/PPQ group had pfplasmepsin2 gene amplification, confirming the absence of piperaquine resistance. The prevalence of the triple pfdhfr mutant (N51I/C59R/S108N) was close to or reached fixation (97.4\u2013100%). For combined pfdhfr/pfdhps mutation, 55\u201371% carried the quadruple (N51I/C59R/S108N+A437G) mutant and about 10% the quintuple mutant N51I/C59R/S108N+A437G/K540E. Our findings confirm that ASAQ, AL and DHA/PPQ were highly effective for the treatment of uncomplicated malaria in the study areas, and neither pfk13 validated mutations nor pfplasmepsin2 multiple copies were found. The very low prevalence of the quintuple mutant in this study supports the NMCP\u2019s decision to introduce intermittent preventive treatment for infants with SP in the districts with high malaria transmission.Currently, the national malaria control programme (NMCP) of Sierra Leone recommends artesunate\u2013amodiaquine (ASAQ) and artemether\u2013lumefantrine (AL) as first- and second-line treatment for uncomplicated malaria, respectively, and artesunate\u202f+\u202fsulfadoxine\u2013pyrimethamine (SP) for intermittent preventive treatment during pregnancy and for infants. In 2016, the NMCP conducted a study to assess the clinical and parasitological responses of children under five years to ASAQ, AL and dihydroartemisinin\u2013piperaquine (DHA/PPQ) according to the WHO protocol. Day-0 samples were tested for mutations in the  Recent d spread . These d13) gene ; and speectively ; and the failure . The recsistance .Malaria is highly endemic in Sierra Leone, with 2.4 million estimated cases in 2016 . In 2004The NMCP of Sierra Leone has established sentinel sites throughout the country for routine monitoring of the efficacy of the recommended ACTs. A therapeutic efficacy study conducted in 2011 at four sites revealed that the recommended first- (ASAQ) and second-line (AL) treatments were highly efficacious . Five ye22.1The study was conducted between March and October 2016 at three sentinel sites Bo, Kenema and Makeni Government hospitals in Bo, Kenema and Bombali districts . The eff2.2The study was a one-arm prospective assessment of the clinical and parasitological outcomes of directly observed standard therapeutic regimens of ASAQ, AL and DHA/PPQ and the current WHO protocol for assessing the efficacy of antimalarial medicines was used . When twP. falciparum mono-infection and 500\u2013200 000 asexual parasites/\u03bcL of blood were recruited. Patients were not enrolled in the study if they presented with danger signs or severe malaria . Regular medication that might interfere with the pharmacokinetics of the study ACTs and a history of hypersensitivity reactions to the study medicines were further criteria for exclusion.Children aged 6\u201359 months with an axillary temperature \u2265 37.5\u202f\u00b0C or a history of fever during the previous 24\u202fh,  malaria , mixed o2.3Patients were treated with a daily dose ASAQ or DHA/PPQ for 3 consecutive days or twice daily doses of AL over 3 consecutive days according to the WHO weight-based dose regimen . The stuDay-0 was the day a patient received the first dose of the study medicine. Parents or guardians were invited to bring the study children on scheduled visits on days 1, 2, 3, 7, 14, 21 and 28 for those given ASAQ and AL, continued to days 35 and 42 for those treated with DHA/PPQ, or on an unscheduled day if symptoms occurred. Clinical and laboratory assessments were done during the follow-up visits.Adverse events, defined as a sign or symptom that was absent at enrolment but manifested during the follow-up period, were assessed by physical examination and by questioning caregivers. Similarly, serious adverse events, defined as conditions that result in death, a life-threatening condition, hospitalization or prolongation of hospitalization, persistent or significant disability or incapacity, were monitored. Both adverse and serious adverse events were recorded on individual case record forms.2.4Thick and thin blood smears were made during scheduled and unscheduled visits, stained with Giemsa and read independently by two qualified microscopists. The procedures for parasite counting and parasite density calculation were based on the WHO protocol . The fin2.5Treatment responses were classified as per the WHO protocol  as treat2.6For patient with recurrent parasitaemia on day-7 onwards, filter paper blood sample was taken on the day of recurrence. Genotypes from day-0 and day of recurrence parasites were compared to differentiate recrudescence from new infection. Specimens were labelled, dried and stored in individual plastic bags with desiccants, protected from light, humidity and extreme temperatures and sent to the Institut Pasteur, for analysis.msp1, msp2) and glutamate-rich protein (glurp) loci. The results were classified as recrudescence if the recurrent parasites were of the same parasite strain as those on day-0 or new infection if they presented with different genotyping profiles.Each dried blood spot was punched with a sterile puncher and placed in a 96-well plate in numerical order. Samples were lysed overnight in a saponin solution, and then DNA was extracted with Instagen Matrix resin, as described previously . DNA sam2.7pfK13 gene, which are associated with artemisinin resistance, whereby a portion of the pfK13 gene was amplified in a nested PCR assay . Amplicons were sent for sequencing, and DNA sequences were analysed to identify specific single nucleotide polymorphisms (SNPs) related to artemisinin resistance (pfplasmepsin2 gene (PF3D7_1408000) was determined in the day-0 samples of the DHA/PPQ-treated group, as described previously by pfplasmepsin2 copies.All day-0 DNA samples were tested for the presence of mutations in the propeller domain in the sistance . Brieflypfdhfr and pfdhps genes, as described previously by pfdhfr and 724 bp for pfdhps) were sequenced and DNA sequences were analysed with CEQ2000 genetic analysis system software (Beckman Coulter). The amino acid sequences were compared with the 3D7 wild-type amino acid sequences (PF3D7_0417200 for pfdhfr and PF3D7_0810800 for pfdhps). The presence of SNPs was confirmed by reading both the forward and the reverse strands. Parasites with mixed alleles were considered mutants.DNA extracts of day-0 samples were also analysed for the presence of mutations in the 2.8A minimum sample size of 50 was calculated after assuming 5% treatment failure of the study medicines, a confidence level of 95% and a precision level of 10%. With a loss to follow-up of 10% by day-28 (ASAQ and AL) or day-42 (DHA/PPQ), 55 patients per site and per drug were required.http://www.who.int/malaria/publications/atoz/9789241597531/en/).All clinical and laboratory data were recorded on standardized case record forms for each patient, and data were validated. Demographic, clinical and laboratory data were double-entered independently and analysed in an Excel\u00ae database specifically designed by WHO for studies of antimalarial drug efficacy  eligible children were recruited at the three sites. The baseline profiles of the enrolled patients given the different antimalarial medicines at each site are shown in 3.23.3pfk13 mutations, 15 did not give interpretable data due to too little or poor-quality DNA. Two hundred and seventy-eight (94.2%) samples gave interpretable sequences. Of these, majority  carried wild-type pfk13 gene and only six had non-synonymous pfk13 mutant: A578S  and I646\u202fT . The remaining two were with synonymous pfk13 mutation: Y493Y  and V510\u202fV . Among the 103 day-0 samples tested for pfplasmepsin2 amplification, 100 (97.0%) gave interpretable data, and all (100%) had pfplasmepsin2 single copy.The day-0 samples of two of the 295 patients were lost. Of the 293 day-0 samples tested for the presence of pfdhfr and pfdhps, respectively. dpfhfr, pfdhps separately and combined. All patients in Makeni and Kenema, and 97.4% (n\u202f=\u202f112) of patients in Bo carried parasites with a triple pfdhfr mutant. The most prevalent pfdhps mutation was A437G, accounting for 62.9% (56.5\u201371.1%) at the three sites, with the highest prevalence in Makeni. The double pfdhps A437G/K540E mutant occurred at a prevalence of 10% (8.7\u201311.3%), and all were part of the quintuple or sixtuple. Combined pfdhfr/pfdhps mutation was analysable for 291 patients of which 54.8%\u201371.1% of patients carried quadruple (N51I/C59R/S108N+A437G) mutant. The overall prevalence of the quintuple (N51I/C59R/S108N+A437G/K540E) mutant was 10%, with no difference among sites: 2X \u202f=\u202f0.37, p =0.83. Another quintuple mutant (N51I/C59R/S108N+A437G/A613S) was found in 14 cases, most of them in Kenema (11.3%) and Bo (5.2%), and a sextuple mutant (N51I/C59R/S108N+A437G/K540E/A613S) in four cases.Of the available 293 samples, 100% and 99.3% (n\u202f=\u202f291) gave interpretable data for 4Our findings confirm a high cure rate and efficacy (100% ACPR) of the nationally recommended ACTs (ASAQ and AL) for the treatment of uncomplicated falciparum infection in the study areas since their introduction almost a decade ago. A similar response was obtained for DHA/PPQ. Similar results for ASAQ and AL were observed in 2011 at the same sites, indicating that these ACTs have maintained their efficacy. The high cure rate of ASAQ and AL, particularly in children under five years, is encouraging, as treatment failure manifests easily in this age group because of low immunity .ASAQ and AL are the most commonly recommended ACTs for the treatment of uncomplicated falciparum malaria in African countries. Recent evidence suggests that these ACTs have maintained high efficacy (cure rate \u2265 95%) in many of these countries, despite their use for more than a decade . Howeverpfk13 mutations and a high piperaquine 50% inhibitory concentration. In Viet Nam, the rate of treatment failure rate with DHA/PPQ increased from 0 in 2012\u20132013 to 26% in 2015, with a background of increased molecular markers for artemisinin and piperaquine resistance  associated with artemisinin resistance  sistance . The A57 in vivo . The absountries  might suzambique , though pfdhfr (N51I/C59R/S108N) mutant in the study areas was close to or reached fixation, and that of the quadruple (N51I/C59R/S108N+A437G) mutant was >50%; however, that of the quintuple (N51I/C59R/S108N+A437G/K540E) mutant, which is associated with a high level of SP clinical failure  nor pfplasmepsin2 multiple copies (associated with piperaquine resistance) were found. Furthermore, the very low prevalence of the quintuple mutant in the current study supports the NMCP\u2019s decision to introduce IPTi with SP in selected high-malaria transmission districts.ASAQ, AL and DHA/PPQ were found to be highly effective for the treatment of uncomplicated malaria in the study areas, and neither SJS: conceived and designed the study, supervised the field work and contributed to writing the manuscript.ARYK: conceived and designed the study, conducted the field work and contributed to writing the manuscript.FS: contributed to the study design, study conduct and preparation of the manuscriptMS: contributed to the study design, study conduct and preparation of the manuscriptASS: participated in the field work, responsible for the quality assurance and quality control of the malaria microscopy and participated in the writing of the manuscript.DM: responsible for the analysis of parasite genotyping and molecular markers for artemisinin and piperaquine resistance and contributed to the write up of the manuscript.MW: contributed to the study design, validated and analysed data and contributed to preparation of the manuscriptAll the authors read and approved the final version of the manuscript.Marian Warsame is a staff member of the World Health Organization, and she alone is responsible for the views expressed in this publication, which do not necessarily represent the decisions, policy or views of the World Health Organization.The authors declare no competing interests.Trial registration number: ACTRN12618000517279."}
+{"text": "Intravenous artesunate and its follow on full course dihydroartemisinin\u2013piperaquine are the standard treatment for severe malaria in Indonesia. The current policy suggests that intravenous and oral quinine could be used when standard therapy is not available. Its pragmatic use of both treatment combinations in a field hospital is evaluated.A retrospective study among hospitalized malaria patients receiving intravenous anti-malarial treatments at Mitra Masyarakat Hospital, Timika from April 2004 to December 2013 was conducted. The length of hospital stay (LoS) and the risk of malaria recurrence within 28\u00a0days after hospital admission were compared between patients receiving intravenous artesunate and oral dihydroartemisinin\u2013piperaquine (Iv Art\u2009+\u2009DHP) and those receiving intravenous and oral quinine .Plasmodium falciparum infection accounted for 65.8% (6915), while Plasmodium vivax, Mixed infections, Plasmodium malariae and Plasmodium ovale were accounted for 17.0% (1789), 16.4% (1729), 0.8% (79) and 0.01% (2) of the infections, respectively. The majority of severe malaria hospital admissions were highland Papuans . In total 49% (5158) of patients were older than 15\u00a0years and 3463 (32.9%) were children under 5\u00a0years old. The median LoS was shorter in patients receiving intravenous artesunate compared to those treated with intravenous quinine . Patients treated with intravenous quinine had higher risk of being hospitalized longer than 2\u00a0days . The risk of recurrences within 28\u00a0days after hospital admission was 1.94 times higher  in patients receiving intravenous quinine with follow on oral quinine treatment than in patients treated with DHP after intravenous artesunate therapy.Of 10,514 patients requiring intravenous therapy, 2759 received Iv\u2009+\u2009Oral Qu and 7755 received Iv Art\u2009+\u2009DHP. Intravenous artesunate reduced the LoS of malaria patients and in combination with DHP reduced the risk of malaria recurrence within 28\u00a0days after hospital admission compared to those with Iv\u2009+\u2009Oral Qu treatment. Thus, ensuring continuous supply of intravenous artesunate and artemisinin-based combination therapy (ACT) should be a priority. Plasmodium falciparum and Plasmodium vivax infections are responsible for the greatest number of death among patients with severe malaria and this includes in Papua, Indonesia ) than those of normal nutritional status . Having Hb\u2009<\u20095\u00a0g/dl increased the risk of prolonged hospitalization . Severe anaemia patients had longer LoS of 3\u00a0days (IQR 1\u20136) versus 2\u00a0days (IQR 1\u20133) in those with Hb\u2009\u2265\u20095\u00a0g/dl (p\u2009<\u20090.0001).P. falciparum hospital admissions, other species of infections did not increase the risk of prolonged hospitalization . Median LoS of pregnant women was 3\u00a0days (IQR 1\u20135) and those of non-pregnant individuals was 2\u00a0days . Children aged less than 15\u00a0years old appeared to have lower risk of extended stay. Compared to The risk of having malaria representations to the hospital within 28\u00a0days after admission was 6.1% (169/2759) in patients receiving Iv\u2009+\u2009Oral Qu and 3.2% (249/7755) in those treated with IvArt\u2009+\u2009DHP . After treatment policy change, the risk of recurrence within 28\u00a0days after hospitalization remained significantly greater in patients receiving Iv\u2009+\u2009Oral Qu  compared to those treated with IvArt\u2009+\u2009DHP  with the adjusted HR of 4.91 (95% CI 2.02\u201311.89), p\u2009<\u20090.0001.Infants and children under-fives had significant risk of having recurrence  respectively compared to the 15\u00a0years older age group. Being highland Papuan also increased the risk of malaria recurrence  compared to non-Papuans. No recurrence was found in patients with severe malnutrition. Species at initial admissions and being pregnant did not increase the risk of malaria recurrence  with severe manifestations [Intravenous artesunate is highly effective for severe malaria treatment and has been widely used in malaria endemic areas in Africa, America and the Asia\u2013Pacific regions  The WHO stations , 21. Ensstations .The study findings highlight real life effectiveness of intravenous artesunate and intravenous quinine prescribed in a field hospital to patients with severe malaria. It was found that patients receiving intravenous artesunate had significantly shorter hospitalization days compared to those treated with intravenous quinine (median 2 versus 3\u00a0days). In addition, the risk of hospitalization longer than 2\u00a0days was 1.7 times higher in patients receiving intravenous quinine compared to those treated with artesunate. Consistent finding was also observed in subgroup analysis restricted to the period after treatment policy change suggesting minimal effect of possible shift in treatment practice to duration of stay. Artesunate is known for its rapid parasite clearance time compared to those of quinine , 24 and However, the association of LoS and the choice of intravenous anti-malarial drugs is not straightforward. Rapid parasite clearance in patients admitted already in an advance state of severe complications would not improve the overall patient\u2019s condition as further supportive therapy is still required to manage complications. The LoS in a controlled study in which strict severe criteria for admission was applied, intravenous artesunate had similar duration of hospitalization compared with quinine, which were 5 to 6\u00a0days , 23. IntThe threshold of severe criteria assessment in patients with parasitaemia in a hospital setting like in Timika, is likely to be lower than that in a controlled study. This could explain the greater reduction of LoS in patients receiving a highly effective treatment found in this study. Previous observational study in this region has shown that the majority of severe complications at the same hospital were severe anaemia, followed with respiratory distress and impaired of consciousness .Another significant risk factor for prolonged hospitalization was severe anaemia (Hb\u2009<\u20095\u00a0g/dl). Longer hospital stay in severely anaemic patients is very likely due to supportive treatment, which is multiple blood transfusions rather than delayed parasite clearance. Severely malnourished patients were more likely to stay longer in the hospital (aOR\u2009=\u20094.39) compared to those with normal nutritional status. Fluid, electrolyte and nutrient imbalance in patients with malnutrition are more likely associated with longer hospital days.Being highland Papuan also increased the risk of prolonged hospitalization compared to non Papuans. Both ethnic groups are known to be more susceptible to malaria compared to those of the lowland Papuans . MalariaA follow on oral anti-malarial agents after intravenous treatment should be given in patients with severe malaria to achieve optimum parasite clearance and prevent recrudescence . This stDHP, an artemisinin-based combination therapy, for 3\u00a0days is highly efficacious for both falciparum and vivax malaria . PiperaqInfant and young children were also at higher risk of having recurrence with the aOR of 3.97 and 2.99 respectively compared to those aged more than 15\u00a0years old. This group has been known to have less immunity and more vulnerable to malaria and recurrence episodes of malaria \u201333. CompInterestingly, none of patients with severe anaemia had recurrences. It has been shown that iron deficiency is protective to infections, including malaria . It is pIntravenous artesunate has been proven to significantly reduce the risk of mortality compared to quinine in multicenter controlled intervention studies , 8. ThisThis study has several important limitations. Firstly, due to the nature of this study, parasite clearance time (which is an indicator for treatment effectiveness) could not be assessed. LoS is used as a proxy indicator of effectiveness with the assumption that parasite clearance will improve clinical outcome and shorter duration of stay .Secondly, malaria recurrences in the population were not actively detected. This study only includes hospital malaria representations to examine recurrence. It is possible that some recurrence occur in the community or presents to other health facilities. However, RSMM provides free medical care for the local tribes and about 80% of malaria presentations in the district was to RSMM.Lastly, this study is not designed for clinical trial. The type and degree of severe manifestations could not be ascertained in this study. However, the hospital protocol suggests that intravenous treatment should only be given to malaria with severe complications and to some extent could be used as an indicator of presence of severity. In about 10% of the cases, intravenous treatment were given to those unable to receive oral treatment due to comorbidity with other illnesses such as surgical, neurological and metabolic diseases cases. Therefore, malaria with significant co-morbidity were excluded from the analysis.The ultimate goal of severe malaria treatment is to clear parasites rapidly and prevent death, provide standard management of complications and prevent recrudescence . This st"}
+{"text": "Pregnant women are more vulnerable to malaria leading to adverse impact on both mothers and fetuses. However, knowledge on the efficacy and safety of antimalarials in pregnancy is limited by the paucity of randomised control trials and the lack of standardised protocols in this special subpopulation. Pooling individual patient data (IPD) for meta-analysis could address in part these limitations to summarise accurately the currently available evidence on treatment efficacy and risk factors for treatment failure.To assess the treatment efficacy of artemisinin-based and quinine-based treatments for uncomplicated falciparum malaria in pregnancy, seven databases  and two clinical trial registries  were searched. Both interventional and observational cohort studies following up for at least 28 days will be included. IPD of the identified eligible published or unpublished studies will be sought by inviting principal investigators. Raw IPD will be shared through the web-based secure platform developed by the WorldWide Antimalarial Resistance Network using the established methodology. The primary objective is to compare the risk of PCR-corrected treatment failure among different treatments and to find the risk factors. One-stage IPD meta-analysis by Cox model with shared frailty will be conducted. A risk of bias assessment will be conducted to address the impact of unshared potential data and of the quality of individual studies. Potential limitations include difficulty in acquiring the IPD and heterogeneity of the study designs due to the lack of standard.This IPD meta-analysis consists of secondary analyses of existing anonymous data and meets the criteria for waiver of ethics review by the Oxford Tropical Research Ethics Committee. The results of this IPD meta-analysis will be disseminated through open-access publications at peer-reviewed journals. The study results will lead to a better understanding of malaria treatment in pregnancy, which can be used for clinical decision-making and conducting further studies.CRD42018104013. This study will be the first individual patient data (IPD) meta-analysis on the efficacy of currently recommended antimalarials in pregnancy incorporating IPD from both randomised control trials (RCTs) and single-arm cohort studies, overcoming the limitation of aggregated data meta-analysis that can only include RCTs.IPD that are standardised in the same format and analysed in a uniform way with adjustment of covariates will, in contrast to aggregated data, allow us to compare the efficacy of different treatments as well as to find risk factors for treatment failure in this vulnerable but understudied population.Limitations of this IPD meta-analysis include the potential difficulty in acquiring the IPD and the heterogeneity of the study designs, study population and parasite population. A risk of bias assessment will be conducted to address the impact of unshared potential data and of the quality of individual studies.About 60% of all pregnancies are estimated to take place in malaria-endemic areas.Plasmodium falciparum malaria for pregnant women in the first trimester by WHO.Due to the lack of evidence for both efficacy and safety of antimalarials in pregnancy, quinine (with clindamycin if available), rather than artemisinin-based combination therapy (ACT), has been recommended as the first-line treatment of uncomplicated The efficacy and safety of antimalarials in pregnancy can be different from the results from the non-pregnant populations because of altered immunity, physiological change in pharmacokinetics and sequestration of parasites to the placenta. The risk factors for treatment failure in pregnancy need to be assessed to improve clinical care in pregnancy. However, there are no agreed guidelines on how to assess the efficacy in pregnancy while it is standardised in the non-pregnant patients by WHO.The WorldWide Antimalarial Resistance Network (WWARN) has established a unique individual participant data (IPD)-sharing platform facilitating large-scale pooled meta-analyses. We plan to include both published and unpublished studies exploring the efficacy and safety of the treatment of malaria during pregnancy. We will conduct a one-stage IPD meta-analysis on the currently recommended antimalarial drugs, that is, artemisinin-based and quinine-based treatments, used for the treatment of uncomplicated falciparum malaria in pregnancy.The aim of this study is to evaluate and compare treatment outcomes of artemisinin-based and quinine-based treatment for uncomplicated falciparum malaria in pregnancy.Primary objectives areTo compare antimalarial efficacies among artemisinin-based and quinine-based treatments.To identify risk factors associated with treatment failure.Secondary objectives areTo assess the relationship between the dosing (dose per body weight) of artemisinin-based treatments and treatment efficacy.To evaluate the risk of gametocyte carriage following artemisinin-based and quinine-based treatments.To evaluate the safety and tolerability of artemisinin-based and quinine-based treatments.Prospective clinical efficacy studies with a minimum 28-day active follow-up.Both interventional and observational cohort studies regardless of the number of treatment arms .Genotyping conducted for distinguishing recrudescence and reinfection.The following studies will be excluded.\u226410 eligible pregnant women.Conducted in non-endemic countries .Pregnant women in any trimester.P. falciparum parasitaemia.Parasitologically confirmed Either asymptomatic or symptomatic.Treated with artemisinin-based or quinine-based treatments.Parasitological and clinical efficacy.Adverse events.A systematic literature review was conducted to identify the potential studies to be included in this IPD meta-analysis. Seven databases  and two clinical trial registries  were used. Both published and unpublished grey literature such as conference abstracts and registered trials were included. This systematic review and IPD meta-analysis is registered to PROSPERO, and the search terms and conditions are available there.Briefly, the search combined five components: malaria; pregnancy; treatment or names of antimalarial drugs; study design  and outcome types (efficacy) without limitation on publication year or language. The result of the literature search was published elsewhere.Principal investigators of the published and unpublished studies identified by the systematic literature review will be invited to share their IPD with WWARN. Emails will be sent to the corresponding authors on at least three occasions asking whether they are willing to join the study group. A secure web-based platform has been developed by WWARN, and IPD will be uploaded after agreeing to the terms and conditions of the submission, retaining the ownership and full control of their shared data.Raw data will be curated in a standardised format using the WWARN Clinical Module Data Management Plan to facilitate pooled IPD meta-analyses.Pregnant women will be eligible for the purpose of this analysis if the following information is available:Confirmed pregnancy status on day 0 of the treatment.Type, date and dose of antimalarial drugs (artemisinin-based or quinine-based treatments).Patient age and estimated gestational age (or trimester of pregnancy) on day 0.Date of the last day of follow-up or length of follow-up.The following patients will be excluded:P. falciparum infection at enrolment.No or missing data on parasitological confirmation of Presenting with severe malaria symptoms at enrolment as defined by WHO,P. falciparum treatment failure. Secondary outcomes will include any recurrence of malaria (PCR-uncorrected treatment failure); parasite clearance; gametocyte carriage during follow-up and adverse events that developed after drug administration. Pregnancy outcomes and placental malaria may be assessed if enough data are gathered.The primary outcome will be the PCR-corrected P. falciparum will be distinguished by PCR into recrudescence (treatment failure) and reinfection.Recurrences of Blood smears will be assumed negative between the two negative observations.If a patient came back to be followed up with a positive smear, the date of positive parasitaemia will be assumed to be the date of observation if this date is within 28 (\u00b13) days from the last observation.If parasite clearance is not recorded after treatment but the positive parasite count is recorded at least 7 days after starting the treatment, the day of the first positive count will be regarded as the day of recurrence.Definitions of status and other censorship are detailed in the Clinical Module Data Management PlanAdverse symptoms will include abdominal pain, dizziness, headache, body pain/myalgia, weakness/fatigue, vomiting, nausea, anorexia and tinnitus if data permit.The following baseline characteristics of patients will be included as appropriate if enough data are shared: age; estimated gestational age (or trimester); parity or gravidity; weight (weight before pregnancy and weight at treatment); body mass index; baseline parasitaemia; presence of fever (body temperature >37.5\u00b0C); haemoglobin (or haematocrit); anaemia (haemoglobin <110\u2009g/L or haematocrit <30% for anaemia and haemoglobin <70\u2009g/L or haematocrit <20% for severe anaemia)For each study, study locations and local transmission intensity will be considered. The study sites will be classified into three categories: low, medium and high malaria transmission based on the parasite prevalence estimates obtained from the Malaria Atlas Project for specific location and year of study.Plasmodium vivax intercalated infection  will be regarded as censored if the original study did not test PCR for falciparum recurrences after intercalated vivax infection, following the WHO guidelines.A summary of the studies and baseline characteristics of the patients included in the analysis will be presented. The number of available patients will be summarised for all variables listed above, proportion will be used for categorical or binary variables and mean and SD (or median and IQR) will be used for continuous variables.PCR-corrected and uncorrected outcomes will be used to compute the Kaplan-Meier (K-M) estimates for each study site. The efficacy of each treatment will then be summarised at fixed time points  by the aggregated meta-analysis approach.A one-stage IPD meta-analysis using the Cox model with shared frailty for study sites will be conducted to identify the risk factors for treatment failure as well as comparing different treatments. For repeated episodes, if any, multilevel mixed-effects model (if there are enough data) or the previous history of malaria will be used. If data permit, a non-linear relationship will be examined for continuous variables.Analysis of secondary outcomes will be carried out provided enough data are present; else, only summary statistics will be reported. Analyses similar to the primary outcome will be conducted for PCR-uncorrected treatment failure .Parasite clearance will be assessed as the proportions of patients cleared asexual falciparum parasitaemia on day 1, 2 and 3. Univariable and multivariable mixed-effects logistic regression models (or Cox models for the time to parasite clearance) will be used to identify the risk factors associated with parasite positivity status.P. falciparum gametocytes on day 0, 3, 7, 14, 21 or 28. Proportions after day 0 will be stratified by the presence of gametocytes at baseline. If enough data are available, mixed-effects logistic regression models will be used to assess the risk factors for gametocytes carriage after treatment stratified by the presence of gametocytes at baseline.Gametocyte carriage will be assessed as the proportion of patients with Adverse effects will be assessed as the proportion of patients who developed symptoms after the treatment initiation. Proportions of patients who developed symptoms after day 0 will be stratified by whether or not that symptom was present before the treatment initiation. If enough data are available, mixed-effects logistic regression models will be used to assess the risk factors for adverse symptoms developed after the treatment initiation. Symptoms on day 0 (before treatment) will be added as a covariate. Primarily, the symptoms developed in the first week will be included.For any regression models, the following strategy recommended by ColletThe multilevel logistic or Cox models would be used for explaining the study-site heterogeneity. Heterogeneity across study sites will be statistically assessed as the variance of the shared frailty term estimated in the Cox model or variance of the random intercepts in logistic regression. Additionally, the intraclass correlation in logistic regression model will be reported.Analyses will be conducted by malaria transmission intensity and by treatment (for assessing dose impact of each drug) if data permit.Two types of sensitivity analyses will be performed. First, a model will be refitted with excluding one study at a time to identify any influential studies. Second, to assess the impact of covariates with missing values, multiple imputation may be used.The risk of bias within and across the included studies will be assessed following the GRADE guidelines.The main analysis is planned as described above. Modification or additional analyses may be required as the data collection progresses. Updated statistical analysis plans will be available at the WWARN website if an amendment is required.Statistical analysis will be conducted using R  or Stata MP V.15.1 (StataCorp).This IPD meta-analysis will use existing secondary data. Patients and public were not involved in the design, recruitment or conduct of this IPD meta-analysis. The results of this study will be shared with the primary investigators of the shared studies and disseminated as publications in open-access journals.Findings will be reported following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA)-IPD statementThis IPD meta-analysis will update the previous aggregated data meta-analyses that included only four or five randomised control trials.WWARN has developed the secure and equitable data-sharing platform and the international collaborative network of malaria researchers worldwide over the last decade. With this unique collaborative effort, we hope that these findings will lead to the improvement of clinical management of this vulnerable but understudied population."}
+{"text": "Treatment adherence has been described as the process whereby patients take medications, follow diet, and effect other lifestyle changes that relate to agreed recommendations from healthcare providers. The determinants of such treatment adherence include patient, the health condition, therapy type, socioeconomic conditions, and the healthcare system. The study examined adherence in malaria patients treated with dihydroartemisinin-piperaquine in routine clinical care in northern Ghana. The study was conducted in Navrongo Health Research Centre in the Kassena-Nankana district of northern Ghana. Patients confirmed with uncomplicated malaria were prescribed dihydroartemisinin-piperaquine in blister packs to be taken daily for three days. Follow-up visits were made on days 3 and 28 after diagnosis to collect data on adherence, drug safety and therapeutic effectiveness. During follow-up visits, in-depth interviews were conducted and the blister packs directly observed for the number of tablets remaining. The in-depth interviews documented day-by-day account of doses taken, number of tablets taken during each dose, time of each dose, reasons for any leftover or missed dose, and whether or not there was vomiting. Treatment adherence was classified as definitely nonadherent, incomplete adherence, and completely adherent. A total of 405 patients were screened; 299 were positive by rapid diagnostic testing and 216 by microscopy. The average age was 12 years and females represented 54.0%. All participants completed day 3 follow-up but 12.7% had leftover pills. Treatment adherence was 50.9% , 36.1% , and 13.0%  for completely adherent, incomplete adherence, and definitely nonadherent, respectively. All completely adherent patients were free of parasitemia on day 28 of follow-up. A total of 49 adverse events related to malaria symptoms were documented. Effort to improve adherence should be individualized as it is dependent on a number of factors such as the patients' temperament, the disease, support at home, and complexity of treatment. Treatment adherence refers to the process whereby patients take medications, follow diet, and effect other lifestyle changes that relate to agreed recommendations from healthcare providers \u20134. The dThe most important therapy-related factors associated with treatment adherence are, however, adverse effects. The potential adverse effects associated with a specific drug can cloud the ability of a patient to adhere to a prescribed therapy whether the patient has experienced it personally previously or is from another source , 5. OtheSocioeconomic factors such as possession of health insurance, out-of-pocket expenses, and support from relations also influence treatment adherence. An increase in cost sharing makes patients less likely to adhere to a prescribed therapy, with the possibility of either delaying or never initiating it. This is because if a patient does not have the means, it is unlikely that he or she will seek the appropriate care necessary , 7. In aPrompt diagnosis, treatment, and adherence to dosage regimen of antimalarials provide individual benefits of curing infection and preventing progression to severe disease, and community-level benefits by reducing the infectious reservoir, avert reinfection, and the emergence and spread of drug resistance . AlthougThe study was conducted in Navrongo Health Research Centre located in Kassena-Nankana District (KND) of northeastern Ghana. The KND is on latitude 10\u00b030\u2032 and 11\u00b000\u2032N and longitude 1\u00b000\u2032 and 1\u00b030\u2032W and cover surface area of 1,674 square kilometers. . The NavThe study area has five demarcated zones  based on the Navrongo Health and Demographic Surveillance System. Two of the zones  were selected to represent the urban and rural areas, respectively, for this study. All the healthcare workers at the selected health facilities in the selected zones were retrained on current malaria treatment guidelines including dihydroartemisinin-piperaquine use for uncomplicated malaria. Project staff had training on study procedures and how to administer the questionnaire after it had been pretested. All patients who reported to the health facilities in the selected zones and who reported fever or history of fever in the previous 24 hours and who had positive malaria parasitemia were eligible for recruitment. A rapid diagnostic test (RDT) was used to screen for malaria parasites. Blood smears were also prepared for microscopic confirmation of the malaria before enrolment into the study. The two tests were done concurrently on the same samples. This is because persistence of the HRP2 antigen means that patients continue to test positive with HRP2-based RDTs long after the patient has been treated and the parasites had been eliminated. The two tests therefore helped to distinguish between lingering positivity and de novo infection in this high-transmission area where patients frequently experience repeated malaria infections and treatments. Adult patients, parents, or caregivers who accompanied the patient to the health facility responded to the interview questions where necessary. Recruitment of cases started in October 2014 and ended in November 2015.All patients had to satisfy the study selection criteria. Inclusion criteria were a positive malaria test by microscopy, a diagnosis of uncomplicated malaria, written informed consent, age \u22656 months and weight \u22655\u2009kg, capability of taking an oral medication, resident in the study area, and being available for follow-up. Exclusion criteria were unwillingness to provide informed consent, known pregnancy, lactating mothers, complicated malaria or severe disease, and nonresident in the study area or intention to move out within one month. Written informed consent was obtained from all 18-year-old or older patients prior to blood smear preparation. Parental consent was obtained for all patients below 18 years of age. In addition, patients aged 12\u201317 years gave assent to be eligible for enrolment into the study. A structured questionnaire was developed and administered in the form of an interview at enrolment and during follow up visits. At enrolment, baseline data including sociodemographic characteristics, place of residence, and the characteristic of the current medications prescribed were collected.Dihydroartemisinin-piperaquine brand , which was in the market at the time and was being prescribed, was used in the study. Treatment doses of dihydroartemisinin-piperaquine (Duo-Cotecxin\u2122) were as follows: 20/160\u2009mg (dihydroartemisinin-piperaquine) children dose was given as one tablet per day for those 5\u2009kg to <10\u2009kg and two tablets per day for those 10\u2009kg to <20\u2009kg and for the adults the dosage was 40/320\u2009mg (dihydroartemisinin-piperaquine) administered as two tablets per day for those between 20\u2009kg and <40 and 3 tablets per day for those \u2265 40\u2009kg. All the treatment doses were given once daily for three days. The non-artemisinin agent piperaquine is an oral active bisquinoline that is structurally similar to chloroquine. Both piperaquine and chloroquine have similar targets through the inhibition of the heme-digestion pathway in the food vacuole of the parasite [All patients prescribed with dihydroartemisinin-piperaquine (DP) were supplied with blister packs of the drug for the three-day treatment course for their current illness episodes and were taught how to take their medications at home. Participants were asked to repeat a dose anytime vomiting occurred within one hour of taking the scheduled medication. Follow-up of all enrolled patients was made to their homes 3 days after the health facility visit. During follow-up visits, consent was sought prior to in-depth interviews with patients or parents/guardians at home using a structured questionnaire. The DP blister packs were directly observed and the number of tablets remaining was counted. The in-depth interview included a day-by-day account of the number of doses taken, number of tablets taken during each dose, time of each dose, reasons for any leftover or missed dose, and whether or not there was vomiting. When study participants or caregivers were not available on the day of visit, attempts were made again to trace them within the next two days in order to minimize recall bias. Patients were considered lost to follow-up if still not found on rescheduled visits. Additional follow-up visits were made 28 days after initial diagnosis to collect further data on drug safety and to monitor therapeutic effectiveness of study drug. During the visit, if a patient was found to be still sick, s/he was immediately referred to the health facility for evaluation and further treatment.By combining the responses to the oral interviews and the physical tablet count from the blister pack, patient adherence to treatment was classified into three categories: definitely nonadherent, incomplete adherence, and completely adherent. A patient who did not take the tablets at all or as recommended by the time of follow-up with physical count of the pills showing a greater than expected number of pills was classified as definitely nonadherent. Any patient (s) who reported that they did not take all the doses as recommended at the time of follow-up or any case where a physical count of the pills showed a greater than or less than the expected number of pills was classified as incomplete adherence. Complete adherence was a case where a patient reported taking all doses as recommended at the time of follow-up visit and a physical count of the pills showed the expected number of pills was taken and no pill left in the pack.Ethical clearance was obtained from the Ghana Health Service Ethics Committee and Navrongo Institutional Review Board.A total of 405 participants were screened for inclusion into the study. Of this number, 299 (73.8%) tested positive by rapid malaria diagnostic testing (RDT). Of the 299 positive RDT patients, 216 (72.2%) were positive by microscopy, 36.6% for those <5 years, 47.5% for the 5-17 years old, and 16.2% for those \u2265 18 years. All the microscopically confirmed malaria patients were enrolled into the study. Overall female patients represented 54.0% (117/216) of the enrolled participants. The average age of the enrolled patients was 12 years; 11.6 years for males and 12.9 years for females, respectively. About 37% (79/216) of the participants were below 5 years of age, 47% (102/216) were 5 to 17 years old, and 16% (35/216) were 18 years old or older. The total geometric mean parasite density at enrolment was 6,908  parasites per microlitre of blood. The highest parasite burden, 10,053  parasites per microlitre of blood, was in children aged below 5 years, 9294  was in 5-17 years, and 1058  was in 18 years old and older. All (100%) the 216 participants recruited were met 3 days after initial visit to the health facility. The study completion rate at day 28 was therefore 99.1% (214/216). Two of the participants in the <5years old and 5 to 17 years old group were not met on day 28 visit and were therefore declared lost to follow-up. Total microscopy positive for malaria parasites on day 28 was 2.8% (6/214): 2.6% (2/78), 4.0% (4/101), and 0/0% (0/35) for the <5 years old, 5 to 17 years old, and those eighteen years and above, respectively. In all, participants with pills after day 3 were 13.0% (28/216). The total number of participants that took study drugs as instructed was 123 (56.9%) and those that did not take the study drugs as instructed were 93 (43.1%). The total number of participants that took the study drugs as instructed and repeated dose after vomiting was 110 (50.9%) and those that took the drugs as instructed but did not repeat dose after vomiting were 13 (6%).Overall, 43.1% of the participants did not take their medication as instructed and 28 participants (13%) had leftover pills three days after first visit to the health facility. Among the 123 (56.9%) participants who reported that they had taken their medication as instructed by the healthcare provider, 13 (10.6%) reported vomiting within an hour of their doses but did not repeat the dose. A total of 28 patients were classified as definitely nonadherent for not taking all the doses. Four of the 28 patients (14.3%) indicated that they felt well and so decided to keep the medication for future use. Similarly, 28.6% (8/28) felt well and so did not know it was necessary to continue the medication. Most of them, 85.7% (24/28), skipped their doses but had plans to continue taking the medication in subsequent days after the visit. Side effects experienced by 10.7% (3/28) were also mentioned as a reason for incompletion of the medication. Other less frequent reasons, forgetfulness, care-giver or participant not at home, and participant refusing to take medication, accounted for 17.9% (5/28). For the participants categorized as definitely nonadherent, they were asked what they would do with the remaining drugs. While 90% (25/28) reported they would continue with the medication, 10.7% (3/28) reported they would discard the remaining drugs. Only one participant reported that the drugs would be saved and used again when there is another illness episode.A total of 49 adverse events were associated with DP treatment as reported by the study participants. The most commonly reported events were weakness, 32.6% (16/49), dizziness, 18.4% (9/49), abdominal pain, 12.2% (6/49), itching, 10.2% (5/49), headache, 8.2% (4/49), and loss of appetite, 4.1% (2/49). There were other adverse events reported, which summed up to 14.3% (7/49). By age, most, 36.7% (18/49), of the adverse events were reported by patients aged 5-17 years followed by 32.6% (16/49) in those below 5 years of age. Most, 97.2% (210/216), of the patients reported that they would prefer the same treatment again if they experienced another episode of malaria. Among the reasons for this preference, 49.0% (103/210) of the patients perceived DP to be effective; 43.3% (91/210) of the patients reported that they experienced no side effects and so would like to have the same treatment if they had another malaria episode. Good taste, 5.2% (11/210), and the fewer number of tablets, 2.4% (5/210), were the other reasons given by patients for the choice of DP for their next malaria episode. Of the 6 patients who preferred a different treatment other than DP for their next malaria episode, 3 perceived DP as ineffective and the remaining 3 complained of the side effects experienced for their decisions.Adherence to malaria treatment is one of the cornerstones of the disease control as prompt treatment cures infection, delay drug resistance and reduces infectious reservoir and transmission . DetectiIn this study, we did not use self-reported approach as the most feasible method for assessing patient adherence to medication because it is prone to recall bias and social desirability bias , 23. DesFrom the study results, complete patient adherence to treatment was about 51%  of the patients and this increased with age and was higher in female patients than in male patients, although the difference was not significant. The proportion of complete adherence in the study was lower because higher adherence to treatment is often associated with acute and life-threatening diseases like malaria except that most of those findings have come from self-reporting studies . This stOverall, about two-fifths of the participants did not take their medication as instructed and about one in ten had leftover pills three days after first visit to the health facility. Even among the those participants who reported that they had taken their medication as instructed by the healthcare provider, about one in ten reported vomiting but did not repeat the dose. This resulted in about 36% of the patients being classified as incomplete adherence and 13% being classified as definitely nonadherent. Patients who were completely adherent to treatment were free of parasitemia at the end of the follow-up, while for patients who were parasitemic, majority of them were classified as nonadherent. However, there was no demonstrable relationship between parasite density and adherence to treatment.Treatment nonadherence is the most common cause of treatment failure. Failure of medical treatment may alternate treatment or cause a change in dose schedule and this may lead to the removal of effective drugs or subject patient to dangers of adverse effect of the alternative drugs , 29.Methods to improve treatment adherence include different packaging and new dosage forms. Others are health education, behavioral oriented programmes, tailoring medication taking to daily habits, and treatment contracts. The most significant finding for treatment adherence is the information given to the patient by the doctor as opposed to other healthcare providers, which increased the need required by the patient to take the medication , 31.Self-report is the most common and feasible method for assessing patient adherence to medication, but it can be prone to recall bias and social desirability bias. As such, this study had to verify the blister packs used by the participants directly to determine the number of tablets of the study drugs used by participants and this could have accounted for the low adherence compared to the self-reported studies. This could have also led to breach of patients confidentiality.Healthcare professionals should be aware of the importance of noncompliance to antimalarials because of the dangers associated with drug resistance and severe and complicated malaria. Patient education is a cornerstone in achieving compliance to malaria treatment and a good patient-clinician relationship is equally vital. Failure of malaria treatment due to noncompliance remains high and so prescribers must take time to improve adherence to malaria treatment."}
+{"text": "This study then recommends that improving patients' knowledge regarding the efficacy, benefits of completing ACT, and consequences of not completing ACT treatment may improve the likelihood of patients adhering fully to ACT.Despite increased support from government and other stakeholders for malaria control over the past decade, malaria burden remains high in many endemic countries, particularly in Sub-Saharan Africa. This study aimed to assess patients' knowledge of antimalarial treatment (ACT) and its association with patient adherence. A descriptive cross-sectional study design was employed in this study. Data were collected from April to May 2017. Both descriptive and inferential statistics in the form of frequencies, percentages, mean values, standard deviations, and Pearson's chi-square test were generated by use of Microsoft excel spreadsheet and IBM Statistical Package for Social Sciences (SPSS) version 23. The average age of the respondents surveyed for this study was 42.27 \u00b1 11.09. Adherence level to ACT was 47%. The results showed that there was a significant association between respondents' knowledge of the efficacy of antimalarial medication ( Despite increased support from government and other stakeholders for malaria control over the past decade, malaria burden remains high in most endemic nations, predominantly in Sub-Saharan Africa including Ghana . A key mGlobally, an estimated 216 million malaria cases occurred in 2016 , as compared to 237 million malaria cases in 2010 (95% CI: 218\u2013278 million) and 211 million cases in 2015 (95% CI: 192\u2013257 million). A higher percentage (90%) of all malaria cases reported in 2016 occurred in WHO African Countries . Out of Since 1998 till date, Ghana has been a committed member of the Roll Back Malaria (RBM) initiative of the World Health Organisation, which focuses on the Global Malaria Strategy with attention on Sub-Saharan Africa . FollowiArtemisinin-based combination therapies have been vital to the recent success of malaria control globally, and protecting the efficacy of the ACT for malaria treatment is a global health priority. Though multidrug resistance, including artemisinin  resistance and partner drug resistance, has been reported in 5 countries of the Greater Mekong Subregion (GMS) , none haPatients' knowledge of the correct intake of the proper dosage and timing is a key element in the success of ACT treatment . Poor paThough several studies have been done on patient adherence to antimalarial medication (ACT) and associated factors to nonadherence , 25\u201327, A descriptive cross-sectional study design was employed, using a pretested, reliable, and validated semistructured questionnaire to assess patients' knowledge of antimalarial treatment (ACT) and its association to adherence.The study was conducted in the Volta Regional Hospital (VRH). The VRH is in the Ho Municipality in the Volta Region of Ghana. The VRH has a bed complement of about 320, and it is the main referral center for facilities in the region and beyond. The target population for the study included all patients who were admitted at the Medical and Surgical wards and who had been on malaria medication for at least once for the past one year. The average number of cases in the two wards for a month is about 310 cases.Convenience sampling technique was employed in recruiting respondents for the survey. The convenience sampling was used because of its simplicity in recruiting study subjects and proximity to the researchers. The sample size for the study was determined by using Yamane sample size formula . Using aAfter permission and approval were obtained from the VRH and the University of Health and Allied Sciences Ethics Review Committee, the researchers commenced with data collection.Data were gathered over a period of 8 weeks, from April to May 31, 2017, between the hours of 8 am and 6 pm each day from Monday to Saturday. Trained research assistants were in charge of recruiting and orienting respondents for the study in the various units of the hospital. Written and verbal consent were sought from respondents before recruitment. On the day of recruitment, respondents were educated on the purpose and benefits of the study. Once patients consented, they received the questionnaire which consisted of patients' demographic data and patients' knowledge of antimalarial treatment (ACT). The respondents were informed that participation was voluntary and they could decide whether to participate or not. It was also explained that the respondents could withdraw without penalty at any time during data collection and this would not have any effect on the care they will receive at the facility. An explanation of the expected length of time (approximately 15 minutes) to complete the study questionnaire was provided. The research assistants were present while the respondents completed each questionnaire. Brief instructions for completing each questionnaire was reviewed with respondents to prevent undue burden on them. Most respondents were assisted to complete the questionnaire if they desired so, but respondents who could neither read nor write were assisted by the research assistant to complete the questionnaire. After completion of the questionnaires, the respondents received a copy of the signed consent form and were appreciated as well for taking time to respond to the study. Once completed, all the instruments were placed in an envelope and sealed. The researchers kept completed instruments in a locked desk drawer for safety and confidentiality.A pretested, validated, and reliable semistructured instrument was used for the survey. The instrument was divided into two sections (A and B). Section A had items that measured the demographic characteristics of the respondents; Section B measured patients' knowledge level on antimalarial treatment (ACT).P values obtained by comparing patient's knowledge scores and adherence were said to be significant if value was less than 0.05. To determine respondents' knowledge score on the treatment of malaria (ACT), respondents were given a score for each correct response to the following variables: name of ACT prescribed, dosage of ACT recommended, efficacy of antimalarial medication, side effects of antimalarial medication, benefits of completing treatment course, duration for taking antimalarial medication, time interval between each dose of antimalarial medication, and consequences of not completing the doses of antimalarial medication prescribed. Respondents who had each of the knowledge components correct were given a score of 1 mark. The overall score of the knowledge components for each of the respondents was 8 marks. The scoring was computerized to arrive at the knowledge score in a descriptive form regarding percentages, mean, and standard deviation.Both descriptive and inferential statistics in the form of frequencies, percentages, mean values, standard deviations, and Pearson's chi-square test were generated by the use of Microsoft excel spreadsheet and IBM Statistical Package for Social Sciences (SPSS) version 23. Cross tables of the frequency and percentages of the distribution were produced with regard to the various demographics. The inferential statistics was to establish the statistical significance of the association. To ensure validity and reliability of the instrument, the researchers were guided by the objective of the study and the reviewed literature in constructing and structuring of the various items. Two professional pharmacists at the VRH and an academic pharmacist face validated semistructured questionnaire. The questionnaire was pretested in the municipal hospital in Ho with 10 patients. The necessary corrections and reconstruction of the questionnaire were made to ensure that every part of the tool reflected the actual situation on the ground. The questionnaire was further retested to ensure its reliability. Different methods have been used to measure medication adherence. Such methods include electronic monitoring devices such as the Medical Event Monitoring System (MEMS), pill counts, self-report through interviews, and biological assays with each of the methods having both advantages and disadvantages. Thus, there is no clear gold standard that has been formalised for measuring adherence for treatment of acute disease like malaria , 19. TheApproval to conduct the study was sought from the Volta Regional Hospital and the Research Ethics Committee (REC) of the University of Health and Allied Sciences provided ethical clearance for this study. The purpose and goals of the study were explained to all the respondents. Written and verbal consent were obtained before each questionnaire was administered to the respondents. Respondents were assured of anonymity and confidentiality.The average age of respondents surveyed for this study was 42.27\u00b1 11.09. Majority of respondents, 107 (representing 62.2%), were with ages ranging from 26 to 45 years. Eighty-nine respondents were females representing 51.7% of the study population. About respondents' religion, the majority (132) were Christians representing 76.7%. With regard to respondents' level of education, the majority (51.2%) of respondents had attained tertiary education. Most respondents (88.4%) reported getting malaria infection about 1-4 times in a year. Also, 149 respondents representing 86.6% of the study population had attended hospital about 1-4 times, and the remaining 23 (13.4%) of the respondents visited the hospital about 5-8 times in a year due to malaria infection. Most respondents (68) representing 39.5% and 54 (31.4%) had their malaria treatment through a doctor's prescription and from licensed chemical sellers, respectively. Most respondents, 90 (52.3%), preferred the oral route of medication for treating malaria. See  P value of 0.001. Educational status was also significantly related to patients' adherence to antimalarial medication with P value of 0.003. Patients' usual malaria treatment and frequency of hospital attendance due to malaria were statistically significant with P values of 0.003 and 0.004, respectively. With the overall adherence level, the minority of, 80 (46.5%), respondents were adherent to antimalarial medications.From  p value of 0.003. Also, respondents' knowledge of the benefits of completing antimalarial treatment course was highly significant with p value of 0.001. Respondents' knowledge of the consequences of not completing the doses of antimalarial prescribed was statistically related to p value of 0.002. The overall knowledge score of the antimalarial treatment of respondent was 79% out of 100%, and respondents' knowledge of the \u201cbenefits of completing treatment course\u201d was the highly rated knowledge score of about 98%. Also, the mean for patients' knowledge score on malaria treatment was 6.282 (SD=1.718).From p< 0.001) was statistically related to patients' adherence to antimalarial medication. The current study's finding is consistent with other previous studies that reported a significant association between age of the respondents and the adherence level [ p value of 0.003. Similarly, educational status and literacy were both found to have a statistical relation with ACT adherence in five studies, with higher levels of education and/or literacy positively associated with adherence [This study assessed respondents' knowledge of antimalarial treatment (ACT) and its correlation with adherence. Adherence in this study was measured by patients' self-report of completing their ACT treatment, particularly with dosing and timing. The study found that age  of patients frequent hospital attendance due to malaria with adherence. It was established that the higher the amount of hospital attendance is due to malaria ailment, the more likely the patient would develop the habit of adhering to antimalarial medication (ACT). It was not therefore surprising that patients' \u201cusual malarial treatment\u201d was statistically associated with adherence. The more the patient attended hospital, the more the patient will be exposed to similar malaria medication prescription and will get an adequate education on malaria treatment, probably dosing and timing of taking the drug from the prescriber.There was also a significant association , which showed a high knowledge level within a range of 1-8. This high knowledge score can probably be due to the high level of health education on malaria prevention and control in the country . About 9The overall patient knowledge of antimalarial treatment (ACT) was 79%. It is quite surprising to note that, despite 79% of respondents having knowledge of antimalarial treatment, respondents did not have a high level of adherence to antimalarial medication (ACT). An adherence level of 47% was low as compared to respondents' high knowledge level of antimalarial treatment. Adherence level was measured by respondents self-reporting of completing the treatment course of antimalarial medication. The study findings revealed an adherence level of 47% which is consistent with Depoortere et al. where they found an adherence level as low as 39.4% in a Zambian refugee settlement , a littlThe lack of homogeneity in the results of the current study and the several other studies especially the large range in adherence levels of ACT can be attributed not only to the differences between the study settings, study designs, and ACT formulations, but also the differences in study implementation such as the study questionnaire and blinding participants/patients to follow-up . For exaSince adherence to artemisinin-based combination therapy was based on the patients' self-reports, though proved reliable, there was the possibility that respondents would have the difficulty to recall whether dosing and timing was followed as recommended by the prescriber.The study found a high level of knowledge among respondents. However, the overall high knowledge (79%) of ACT treatment did not reflect patients' antimalarial medication adherence as the study revealed adherence level below average (47%). The current study found a significant association between the following knowledge components: the efficacy of malaria treatment, benefits of completing treatment course and consequences of not completing the doses of antimalarial prescribed, and adherence. This study based on the findings recommends improvement in patients' knowledge regarding the efficacy and the benefits of completing treatment course as well as the consequences of not completing ACT treatment. These may improve the likelihood of patients adhering to ACT treatment."}
+{"text": "Plasmodium falciparum malaria in Greater Mekong Subregion has created impetus for continuing global monitoring of efficacy of artemisinin-based combination therapies (ACTs). This post analyses is aimed to evaluate changes in early treatment response markers 10 years after the adoption of ACTs as first-line treatments of uncomplicated falciparum malaria in Nigeria.The development and spread of artemisinin-resistant At 14 sentinel sites in six geographical areas of Nigeria, we evaluated treatment responses in 1341 children under 5 years and in additional 360 children under 16\u2009years with uncomplicated malaria enrolled in randomized trials of artemether-lumefantrine versus artesunate-amodiaquine at 5-year interval in 2009\u20132010 and 2014\u20132015 and at 2-year interval in 2009\u20132010 and 2012\u20132015, respectively after deployment in 2005.P\u2009=\u20090.002 and P\u2009<\u2009\u00a00.0001, respectively). Parasite clearance time increased significantly from 1.6\u2009days  to 1.9\u2009days  and geometric mean parasite reduction ratio 2\u2009days after treatment initiation decreased significantly from 11 000 to 4700 within the same time period (P\u2009<\u2009\u00a00.0001 for each). Enrolment parasitaemia >\u200975 000\u2009\u03bcl\u2212\u20091, haematocrit >\u200927% 1\u2009day post-treatment initiation, treatment with artemether-lumefantrine and enrolment in 2014\u20132015 independently predicted APPD1. In parallel, Kaplan-Meier estimated risk of recurrent infections by day 28 rose from 8 to 14% (P\u2009=\u20090.005) and from 9 to 15% (P\u2009=\u20090.02) with artemether-lumefantrine and artesunate-amodiaquine, respectively. Mean asexual parasitaemia half-life increased significantly from 1.1\u2009h to 1.3\u2009h within 2 years (P\u2009<\u2009\u00a00.0001).Asexual parasite positivity 1\u2009day after treatment initiation (APPD1) rose from 54 to 62% and 2\u2009days after treatment initiation from 5 to 26% in 2009\u20132010 to 2014\u20132015  contains supplementary material, which is available to authorized users. Please see Additional\u00a0file\u00a0Plasmodium falciparum to artemisinin in the Greater Mekong Subregion (GMS)  and 2012\u20132015 [n\u2009=\u2009106]), clinical and parasitological evaluations were done at the following times: pre-treatment (0), 1, 2, 4, 6, 8 and 24\u2009h and on days 2\u20137, 14, 21, 28, 35 and 42 post-treatment initiation. The kinetics of the time-course of the asexual parasitaemia was estimated using a non-compartment model as previously described  and AA [n\u2009=\u2009678]) (Table\u00a0P\u2009<\u2009\u00a00.0001), had significantly lower body temperature (P\u2009=\u20090.005), significantly higher haematocrit (P\u2009=\u20090.04) and significantly lower proportion of children with gametocyte carriage (P\u2009<\u2009\u00a00.0001) compared with those enrolled at the end of the first 5 years of deployment (2009\u20132010)  and it did not differ during the two study periods (35% [1826 of 5217 children] in 2009\u20132010 versus 35.9% [2410 of 6713 children] in 2014\u20132015). Gametocyte carriage declined significantly during the two study periods (see below). Data on other indices of transmission intensity during the period of observation were not evaluated.CI: 1.06\u20131.12, n\u2009=\u2009470) versus mean of 1.2\u2009days  respectively, P\u2009<\u2009\u00a00.0001]. Time to clear fever rose significantly (P\u2009=\u20090.002) from a mean of 1.1\u2009days  in 2009\u20132010 to a mean of 1.2\u2009days  in 2014\u20132015 (Table\u00a0P\u2009=\u20090.002) but not in AA-treated children (P\u2009=\u20090.36) compared with 2009\u20132010 (Table Treatment with AA cleared fever significantly faster than AL  and AA [n\u2009=\u2009240]) , and geometric mean asexual parasitaemia (P\u2009<\u2009\u00a00.0001) compared with those enrolled in 2009\u20132010 (Table\u00a0P\u2009<\u2009\u00a00.0001 for each) (Table P\u2009<\u2009\u00a00.0001) and proportions of children with asexual parasite clearance time of 2\u2009days increased significantly from 9.5% (24 of 254 children) to 33% (35 of 106 children) (P\u2009<\u2009\u00a00.0001) during the same time period   in 2009\u20132010 to 1.3\u2009h  in 2012\u20132015 (P\u2009<\u2009\u00a00.0001)  in 2009\u20132010 to 1.4\u2009h  in 2012\u20132015 (P\u2009=\u20090.004). In children older than 59\u2009months, terminal elimination half time of asexual parasitaemia also increased significantly from a mean of 1.1\u2009h  in 2009\u20132010 to 1.3\u2009h  in 2012\u20132015 (P\u2009=\u20090.001). There was no difference in mean terminal elimination half-times of asexual parasitaemia in the <\u20095 and\u2009>\u20095\u2009year-olds (P\u2009=\u20090.68). Mean estimated terminal elimination half-times of asexual parasitaemia was similar in the two treatment groups . There was a significantly positive correlation between parasite clearance time and terminal elimination half-time of asexual parasitaemia .Estimated terminal elimination half-time of asexual parasitaemia increased significantly from a mean of 1.1\u2009h  from 72.4% (184 of 254 children) to 50% (53 of 106 children) during the same time period , age\u2009<\u20095 or\u2009>\u20095\u2009years, body temperature\u2009<\u200937.4\u2009\u00b0C or\u2009>\u200937.4\u2009\u00b0C, presence or absence of fever 1 day after treatment initiation, haematocrit <\u200930% or\u2009\u2265\u200930%, enrolment parasitaemia \u2264100 000\u2009\u03bclP. falciparum infections to AL and AA. Parasite positivity 1\u2009day after treatment initiation increased insignificantly from 58 to 66% over time in children receiving AL and significantly from 50 to 58% over time in those receiving AA. In addition, parasite positivity 2 days after initiating both treatments increased significantly from 5 to 26% over time. These increases indicate declining or worsening of early response markers, which are thought to be primarily dependent on the artemisinin components of ACTs [P. falciparum [In these country-wide studies of the efficacies of artemisinin-based combination therapies during a 10-year period of deployment as first-line antimalarials in Nigeria, we report a significant decline of early responses of childhood  of ACTs , 29. Alt of ACTs , 30, theP. falciparum to ACTs [Increment in APPD1 over time is common to all areas with declining responsiveness in  to ACTs , 30. HowThere was remarkable similarity between some of the independent predictors of APPD1 and APPD2 such as haematocrit \u226527% and enrolment in 2014\u20132015 but there was no expansion of baseline asexual parasitaemia, stable baseline asexual parasitaemia, or less than 75% decrease in baseline asexual parasitaemia that would have indicated marked reduction of response to the two ACTs in any of the patients 1 day after treatment initiation. It is surprising that in contradistinction to the situation in some countries in the GMS, where declining responsiveness is associated with increased gametocyte carriage and transmission of parasites with reduced susceptibilities to ACTs , gametocPfK13 C580Y, R539T, Y493H are not frequently encountered in Africa [P. falciparum populations in Africa as has been recently described in a non-immune visitor to Equatorial Guinea [Parasites with alleles that confer slow clearance of parasitaemia in the GMS notably n Africa , 31. Howl Guinea .Four points may help explain the possibility of emergence with time of parasites with slow clearance phenotypes in endemic areas of Nigeria: First, an increase in APPD3 from a baseline of 0.9 to 2.3% over time is unlikely to be a random phenomenon. Second, the significantly increased risk of recurrent parasitaemia over-time, in the presence of the long acting partner drugs, may contribute to parasite survival in the presence of less than optimal parasiticidal drug concentrations , 34, eveEstimation of parasite clearance half-life is thought to be the best in vivo measure of artemisinin drug effect . AlthougIn order not to overestimate terminal elimination half-times of asexual parasitaemia or misinterpret the observed unimodal frequency distribution of terminal elimination half-times of asexual parasitaemia, frequent samples were obtained from the patients. The limitations of our study in this context are the non-estimation of concentrations of the artemisinin components of the administered ACTs and/or their metabolites, and apart from age, the non-evaluation of the many host factors that may influence parasite clearance times such as immune status. ACTs may mobilize asexual parasites from the deep tissue into peripheral blood in the early hours following initiation of treatments in drug sensitive infections in children from endemic areas of Africa , 22. ThiIndividual analysis of the two studies (2009\u20132010 and 2014\u20132015) showed spatial heterogeneity in clearance indices. For example, for both studies, recrudescence and residual parasitaemias on days 1 and 2 were significantly higher on the eastern compared with western flank of the study sites . In addiAlthough we did not evaluate immunity in the cohorts of children in the post hoc analyses, the limited data on transmission intensity suggested this did not change over time. The significantly increased rate of polyclonal infections and multiplicity of infection in the absence of presumed decrease in immunity, may support, in part, the emergence of a population of parasites with reduced susceptibility to the two ACTs. In a similar study of two ACTs in similarly aged Kenyan children, in an area of seasonal but intense transmission, Borrmann and others  found thP. falciparum isolates in Nigeria to artemisinin first reported in the 1992, 13 years before the adoption and deployment of artemisinin-based combination therapies [Overall, the critical questions are: are the present observations of post hoc analyses, an unmasking of innate reduced in vitro susceptibility of herapies ? Is a reherapies  an indirIn conclusion, the declining parasitological responses through time to artemether-lumefantrine and artesunate-amodiaquine raises concerns and may be due to predominance of parasites with reduced susceptibility or decreasing herd immunity in young children in endemic areas of Nigeria.Additional file 1:Multilingual abstracts in the five official working languages of the United Nations. (PDF 259 kb)"}
+{"text": "Non-malarial febrile illnesses comprise of almost half of all fever presenting morbidities, among under-five children in sub-Saharan Africa. Studies have reported cases of prescription of antimalarial medications to these febrile under-fives who were negative for malaria. The treatment of these children with antimalarial medications increases incidences of antimalarial drug resistance as well as further morbidities and mortalities, due to failure to treat the actual underlying causes of fever.To identify clinical and demographic factors associated with treatment type  of non-malarial febrile illnesses (NMFI) in children aged \u22645 at the Kenyatta National Hospital in Nairobi, Kenya.A positivist epistemological approach, cross sectional descriptive study design was used. A structured questionnaire was used on a sample of 341 medical records of children aged \u22645 years to extract data on clinical examinations (recorded as yes or no), diagnostic test results, and demographic data on the child\u2019s sex and age. Descriptive and inferential analysis was applied to the data.Prescription of antimalarial drugs despite negative microscopy results was found in 44 (12.9%) of the children, with mortality reported in 48 (14.1%). Assessment of respiratory distress was 0.13  times associated with less likelihood of prescribing an antimalarial in those with a negative microscopy. A male patient was 0.21  times less likely to receive an intravenous antimalarial after a negative microscopy. Patients aged \u02c21 with a negative microscopy result were more likely to receive an antimalarial than older children.There is a need to eliminate incorrect treatment of NMFI with antimalarial medication, while ensuring correct diagnosis and treatment of the specific illness occurs. This requires strengthening and adherence to diagnostic and treatment guidelines of febrile illnesses in under-fives, consequently reducing morbidities and mortalities associated with inadequate management of NMFIs. Malaria is a serious and sometimes life-threatening infectious disease that affects humans and other animals, caused by several species of the Plasmodium parasite and transmitted by the bite of an infected Anopheles mosquito. Malaria typically manifests as a febrile illness that can be fatal if unrecognized, especially in young children. Approximately 3.2 billion people live in malaria endemic areas, and, in 2015, there were an estimated 214 million cases and 438,000 deaths attributable to the disease. Most of these deaths occurred in sub-Saharan African countries in children under five years of age [Febrile illness is fever (defined as temperature >38\u00b0C ) withoutIn many malaria endemic areas, including Burkina Faso, more than half of these febrile children are considered not to be infected with malaria, as the incidence of this disease is declining due to increased control efforts over the last decade \u20136. This Non-malarial febrile illnesses (NMFI) are infectious diseases affecting patients who show signs of indistinguishable fever thus necessitate use of malaria diagnostic tests, but these tests turn out to be negative for malaria . LaboratNMFI accounts for about half of all fever presenting morbidities among under-five children in sub-Saharan Africa. A study of 70,666 fevers identified across 42 sub-Saharan countries estimated that there were 655.6 million reported fevers in 2007. Malaria did not appear to be the major driver of fever among the children, highlighting the importance of building a clearer picture of causes of NMFI to inform both patient management and prevention efforts. NMFI is known to carry case fatality ratios that equal or exceed malaria in low-resource areas.\u201310 with In Tanzania, the main clinical diagnoses among febrile under-fives who test negative for malaria include respiratory tract infections, pneumonia and urinary tract infections . In KenyLiterature supports several factors as predictors for prescription of antimalarial drugs to children tested negative for malaria. Clinical assessment findings, demographic characteristics of the child and in some cases, the healthcare workers characteristics have been associated with treatment type  of non-malarial febrile illnesses in children aged five years and under ,14,15. OSustainable Development Goals (SDGs) aim to significantly reduce childhood mortality to less than 25 from 43 per 1,000 live births globally. A major reduction in mortality due to infectious diseases in most low and middle income countries is necessary for the achievement of the SDGs . In KenyKenya sitting astride the equator is located in the eastern region of Sub-Saharan Africa, with a population of approximately 47 million and an under five mortality rate of 52 per 1000 . Four miMalaria still remains a major public health problem in Kenya and accounts for an estimated 16% of outpatient consultations based on data from the routine health information system . MalariaPrevalence data from a study using cross-sectional data extracted from the Kenya Malaria Indicator Survey (2015) showed that prevalence of malaria in children 10\u201314 years was 10.22%, while in children under the age of 5 it was 4.83%. Malaria prevalence was also found to be higher among male children (8.23%) than female children (8.04%) .This study was conducted at Kenyatta National Hospital (KNH), the largest public, tertiary referral hospital in Kenya and located in Nairobi, therefore the apex of health system in Kenya. The hospital receives most of the severe referred cases of childhood fevers from across the country. Approximately, 84% of all the children received in Kenyatta Hospital between July and August 2011 presented with fever .The study adopted a positivist epistemological approach with a cross-sectional, descriptive study design. The design was appropriate since the study tested the degree of association between and among variables at a specific point in time. This study design determines the status of phenomena without controlling or manipulating variables  and is cThe primary outcome variable in this study was measured through a proxy variable, which allows for a variable that cannot be confirmed to be included in the model to be estimated . NMFI isMedical records of children aged five years and under, admitted with febrile illness from March-August 2017, defined as the long rainy season in Kenya  was usedWhere Z is the standard normal variate, p is the proportion of children who test negative for malaria and were treated with malaria drugs, n is the desired sample size and d is the margin of error. Nevertheless, all the records during this period were used to eliminate selection bias.Data was obtained from the health information department at KNH. Data obtained on admission of the febrile patients including; medical history, clinical assessment and treatment was provided by attending medical doctors. Standardized training is provided in Kenyan medical schools on history taking and performance of a physical examination. These are supplemented with management guidelines of various illnesses by the health facilities, Ministry of Health and the WHO. Relevant data to the study including; clinical examinations (recorded as yes or no), diagnostic test results, and demographic data on the child\u2019s sex and age was extracted from patient records and entered electronically into study database using Epi-Data Manager software within the hospital premises. The database was password protected to ensure safety and confidentiality of data. Upon completion of data extraction and entry into Epi-Data Manager, the database file was exported to Statistical Package for Social Sciences (SPSS) version 20 for analysis.Children aged five years and under.Admission temperature of 38\u00b0 C and above.Children referred to KNH with a known diagnosis of malaria, for further management.Children aged five years and under whose medical records had missing information on more than two study variables.Admission temperature of below 38\u00b0C.Ethical approval was obtained from University of Liverpool Research Ethics Committee and Kenyatta National Hospital-University of Nairobi Ethics Research Committee  (P489/08/2017) and authorization provided by the Kenyatta National Hospital Health Information Department before the actual study commenced. The Epi-Data Manager entry template was developed in a way that de-identified the patients during extraction and ensured that none of the information could be linked back to the patients. Data will be kept securely for five years from the time of extraction from hospital.The data was collected and recorded in two parts, diagnosis and treatment sections. The diagnosis section contained eight, and treatment section had two elements of assessments. The first treatment element of assessment  was used as proxy to measure the variable of interest which is the children under-five who tested negative for malaria but were treated with antimalarial medication while postulated to be suffering from NMFI.All the responses to the questions were YES/NO except the age variable which was recorded as numbers. The gender variable was captured as Female/Male.The YES/NO responses were coded in SPSS IBM 20 as 1/0 respectively. While gender F/M was coded as 1/2 respectively.Age variable was recoded into a factor variable because there were cases of < 1 (less than one year) which was not specified in months; hence it was recorded as ordinal data.The variable which was indicating whether the patient is deceased or not deceased was created as Deceased/Not deceased and coded as 1/0 respectively.The primary outcome variable was the children with NMFI treated with antimalarial drugs despite negative microscopy results measured indirectly using correct/incorrect prescription of antimalarial after microscopy results responded to as YES/NO respectively and coded as 1/0 in SPSS.The secondary outcome Variable was whether the patient was Deceased/Not deceased at the time of discharge and was coded as 0/1 respectively.To capture the No response and Not Applicable (N/A) the codes assigned to these variables was 99 and 88 respectively.A total of 341 surveys were numbered from 1 to 341 as per standard rule for data entry. This helped to identify and track the surveys to avoid erroneous entry. The data was then entered to SPSS and screened for errors before being used for analysis.The data to assess the primary outcome was rigorously processed using R programming analytic software and SPSS IBM 20. The frequencies of the variables were run using SPSS to check if there were any missing data points as usually reported by the frequency output tables. There were two missing data points on patients\u2019 identification number. This variable would be important in tracing the patients\u2019 records in the database if required but had no effect on the analysis and results.There were two treatment questions in the data capture sheet; the primary outcome question  serving as the proxy variable and the isolating variable . Those who answered the first question with a YES/NO the second one was N/A, and vice versa. The data evaluated for the primary outcome was 72 cases (n = 72) and 269 cases (n = 269) for the second outcome variable.The descriptive statistics was done using SPSS. The descriptive analysis was done in two steps. The frequencies of all the variables were generated and tabulated in the predefined table. The crosstab analysis was generated for specific variables to assess the relationship between the primary outcome and the predictors displayed in the tables.The data set is categorical data comprising of the YES/NO responses. Male/Female and numerical age variables were transformed to ordinal data type. Non-parametric/categorical chi-square test was used to establish associations between outcome variable and exposure factors. On the other hand, binary logistic regression was used to model the categorical outcome variables against the categorical exposure factors.Chi square test was done to test the associations between two variables, outcome variable and a predictor (bivariate). The results from the chi square test were recorded in the tables.Binary logistic regression analysis was conducted using R to assess the association of the predictors and the binary outcome variables. The data was converted to comma separated values (CSV) and exported to R. The results between the primary variable and the predictor\u2019s variables were generated and recorded in the tables. The secondary variable was assessed against all the predictor variables to test if there were any associations. The binary logistic model yielded odd ratios, p-values and confidence intervals (CI) for drawing valid conclusions.The study used records from 341 children aged five years and under with fever of 38\u00b0C and above. The study findings were categorized into the following themes:Demographic characteristics;Clinical factors (exposures);Treatment outcomes;Relationship between outcomes and clinical factors (exposures);Relationship between outcomes and demographic characteristicsMost of the under-five patients (264) in this study who were admitted at the hospital were aged one and below, making 77.48% of the total admission . The stuExposures were reported based on clinical factors that were assessed as indicated in the files by the attending clinician on admission of the patient. Hours since fever onset were noted in 262(76.8%) children. Immunization cards were assessed among 196(57.5%) children. Temperature was taken for 337(98.8%) children. Danger signs which included \u2018at least 3 main symptoms \u2019 was assessed and noted in 253(74.2%) children. Presence or absence of respiratory distress  was noted in 261(76.5%) children. Pallor, jaundice, and sunken eyes were assessed in 320(93.8%) children. Lethargy, prostration, unconsciousness or either irritability and restlessness was assessed among 197(57.8%). .Treatment outcome was classified into two; primary and secondary outcome. Primary outcome was based on either correct or incorrect treatment used as a proxy to measure NMFI treated with antimalarial drugs. Among the 341 children sampled, a section of 72(21.1%) were prescribed with antimalarial medication. Among the 72 children, 28(8.2%) were correctly prescribed with antimalarial while 44(12.9%) were incorrectly prescribed with an antimalarial despite a negative microscopy result. The group of 44(12.9%) forms the variable of interest measured indirectly by first isolating those who received antimalarial medication (whether or not they had a positive microscopy result), then zeroed in to those who received the correct antimalarial prescription following positive microscopy result. Hence the primary outcome variable of the study was measured through proxy procedure.The secondary outcome was captured on basis of whether the patient was deceased/Not deceased at the time of discharge. This is a derived variable to describe whether the death of the patient can be linked to the incorrect treatment of the NMFI with antimalarial drugs. A proportion of 14.1% (48/341) of the patients in this study were deceased at the time of discharge . Of thisAssessment of hours since fever onset; immunization cards; temperature; presence or absence of respiratory distress; pallor, jaundice, and sunken eyes; lethargy, prostration, unconsciousness or either irritability or restlessness were not significantly associated with treatment outcome . Assessment of danger signs  significantly influenced treatment outcome , see The prescription of antimalarial drugs despite negative microscopy results was significantly associated with whether the patient was checked on presence or absence of respiratory distress . This implied that assessment of presence or absence of respiratory distress, influenced if the NMFI patient received antimalarial prescription even after negative microscopy results, see There was potentially good survival chance for the patients who were checked for presence or absence of respiratory distress (proportion of respiratory distress checked and survived (223/341) = 0.654 Vis a Vis proportion of those whose respiratory distress not checked and survived (70/341) = 0.205). This meant that 1 in every 5 patients checked for respiratory distress survived. Patients who were assessed for danger signs with main symptoms of  were 3.15 times more likely to survive than those who were not assessed .Gender and age significantly influenced the primary NMFI treatment outcome , see The gender of the patients was significantly associated with prescription of antimalarial treatment after negative microscopy results ). Data revealed that a male patient was 0.21 times less likely to receive an antimalarial after negative microscopy results compared to female patients, see The age of the patients was significantly associated with prescription of antimalarial treatment after negative microscopy results , see With reference to patients aged under 1 year (< 1 year), a patient aged 1 year was 0.05 times less likely to receive antimalarial treatment after negative microscopy results, those aged 2 years were 0.01 times less likely to receive antimalarial treatment after positive microscopy results, aged 3 years were 0.07 times less likely to receive antimalarial treatment after negative microscopy results while those aged 4 years were 0.09 times less likely to receive correct anti-malarial treatment after negative microscopy results.Gender and mortality. More females (29) than males (19) succumbed to death in under-five patients at KNH during this period of study. The gender of the patients was significantly associated with the mortality .The study sample comprised majority males and children aged one year and under.Majority of the clinical factors (exposures) were assessed at admission, including; hours since fever onset, immunization cards, temperature, danger signs, respiratory distress, pallor, jaundice and sunken eyes, and lethargy, prostration, unconsciousness or either irritability and restlessness.Outcomes included; 28(8.2%) of the children were correctly prescribed an antimalarial after positive microscopy results, while 44(12.9%) were prescribed an antimalarial despite a negative microscopy result. Mortality was the secondary outcome and was reported among 14.1% of the children.Among the clinical factors; assessment of presence or absence of respiratory distress significantly affected treatment of patients with a negative microscopy with an antimalarial, as it decreased the prescription of an antimalarial to those with a negative microscopy by 0.13 times. Assessment of danger signs was also noted to increase the chances of survival by 3.15 times compared to those who were not assessed.Demographic factors, gender and age significantly influenced the primary treatment outcome. A male patient was 0.21 times less likely to receive antimalarial treatment after negative microscopy result compared to female patients. Patients aged <1 year were more likely to receive antimalarial treatment after negative microscopy results compared to those aged \u22651 year. Children aged 2 years were less likely to receive antimalarial treatment after negative microscopy results compared to ages 1,3 and 4.It was also noted that a significant number of females succumbed to death than males.Of the 72 children in this study who were prescribed an antimalarial drug, 12.9% (44) had laboratory and treatment reports in their medical records that indicated that they had been prescribed with an antimalarial medication despite testing negative for malaria according to the microscopic examination.The prescription of antimalarial medication despite negative microscopy results is similar to other studies that reported cases of prescription of antimalarial medications to febrile under-fives who were negative for malaria. A study in Lagos, Nigeria  reportedEarly detection of children at the highest risk of dying and providing the correct diagnosis and early treatment, aids in the prevention of further morbidities and even mortalities. The clinical signs and symptoms of malaria can be confused with numerous other febrile causing etiologies such as pneumonia, typhoid, dengue, chikungunya and other bacterial and viral infections. Over-diagnosis and overtreatment of malaria leads to under diagnosis of NMFI. Lack of proper NMFI treatment may lead to increased morbidities and could have contributed to the 11.4% mortalities in the group that were prescribed an antimalarial despite negative microscopy in this study. A study in a tertiary referral center in Ghana did prospective data collection followed by retrospective analysis of blood culture. The findings concluded that patients who presented with WHO signs and symptoms of severe malaria but had a negative microscopy results had high mortality due to a higher risk of bacterial sepsis .Several factors have been reported to predict the prescription of antimalarial drugs to children despite testing negative for malaria. Children who have been established to show signs and symptoms of severe disease are more likely to be prescribed an antimalarial despite negative malaria test results as the clinicians try to avoid an opportunity to prevent further morbidity or mortality due to unavailability of diagnostics that could correctly identify the NMFI . A demogUse of a cross-sectional study enabled the study of multiple outcomes and exposures and adequately answered the generated hypotheses.The study assessed the patients\u2019 outcome data which was included in the secondary outcome variable as deceased or not deceased. It revealed a mortality rate of 14.1%, with 11.4% of these total mortalities being from the group that was incorrectly prescribed an antimalarial despite negative microscopy. This provides a better understanding on the need to provide correct diagnosis and treatment of NMFIs, to achieve the SDGs on reducing infant mortality rate.The study is not able to look at the reasons behind the health practitioners prescribing antimalarial medication to patients with negative microscopy tests. This would have been better achieved with a qualitative study.Factors such as the level of training of the clinicians, supervision, the laboratory capacity, expectations of the patients were not taken in account which could all possibly affect the prescription practices.The study was done at Kenyatta National Hospital, one of the tertiary referral hospitals in Kenya hence it receives many sick children, limiting the generalizability of the findings to the rest of the Kenya.The study did not collect any data on previous use of antimalarial or antibiotic medications before referral to KNH, and this could be a major influence on the clinicians\u2019 prescription practice.This study supports that clinical and demographic factors such as age, gender and assessment of presence or absence of respiratory distress are predictors of prescription of antimalarial drugs in under-fives. Tackling the NMFI challenge requires a concerted effort, to eliminate incorrect treatment with antimalarial medication, while ensuring correct diagnosis and treatment of the specific illness occurs.It has been repeatedly showcased that inadequate management of NMFIs, is a major cause of morbidities and mortalities among under-fives, especially in Sub-Saharan Africa. This study has shown that clinical and demographic factors play a role in treatment type of NMFI and survival outcomes; with a proportion of the 14.1% of the mortalities in this study postulated to be associated with delayed diagnosis and incorrect treatment of NMFI. There is a need for further exploration to ascertain factors that affect treatment type, together with factors that influence the prescribing physicians, and their knowledge and attitudes on NMFI treatment.Adherence to diagnostic and treatment guidelines in management of febrile children, will assist in the delineation of non-malaria febrile illnesses. This brings about a subsequent need to create and implement guidelines on the correct diagnosis and management of these NMFIs.S1 Appendix(PDF)Click here for additional data file."}
+{"text": "Vivax malaria relapses frequently even in low-transmission settings. Chloroquine delays but does not prevent recurrences. Adding primaquine to a slowly eliminated schizonticide significantly reduces recurrences and improves hematocrit, but this advantage is offset by hemolysis in G6PD-deficient females. Plasmodium vivax infections for >60 years, but resistance is increasing. To guide future therapies, the cumulative benefits of using slowly eliminated (chloroquine) vs rapidly eliminated (artesunate) antimalarials, and the risks and benefits of adding radical cure (primaquine) were assessed in a 3-way randomized comparison conducted on the Thailand-Myanmar border.Chloroquine has been recommended for P. vivax malaria were given artesunate (2 mg/kg/day for 5 days), chloroquine (25 mg base/kg over 3 days), or chloroquine-primaquine (0.5 mg/kg/day for 14 days) and were followed for 1 year. Recurrence rates and their effects on anemia were compared.Patients with uncomplicated P < .001), and 0.5% with chloroquine-primaquine . Median times to first recurrence were 28 days  with artesunate, 49 days  with chloroquine, and 195 days  with chloroquine-primaquine. Recurrence by day 28, was associated with a mean absolute reduction in hematocrit of 1% . Primaquine radical cure reduced the total recurrences by 92.4%. One-year recurrence rates were 4.51  per person-year with artesunate, 3.45  with chloroquine (P = .002), and 0.26  with chloroquine-primaquine (P < .001).Between May 2010 and October 2012, 644 patients were enrolled. Artesunate cleared parasitemia significantly faster than chloroquine. Day 28 recurrence rates were 50% with artesunate (112/224), 8% with chloroquine (18/222; Vivax malaria relapses are predominantly delayed by chloroquine but prevented by primaquine.NCT01074905. Plasmodium vivax, which is becoming the main cause of malaria outside sub-Saharan Africa, exerts considerable morbidity by causing repeated relapses. In Southeast Asia and Oceania, P. vivax relapses early and frequently ) were more likely than females (78 of 215 [36%]) to discontinue follow-up (P = .008). Adults (207 of 402 [52%]) were more likely to discontinue than children (73/241 [30%]) (P > .001).Between May 2010 and October 2012, 925 patients with acute enrolled . Patientenrolled  althoughn adults . The medP = .035) and 60% in the CQ + PMQ group (P = .002). More than 95% of patients were afebrile by day 2. More patients also cleared parasitemia by day 1 following AS (70%) compared to CQ  and CQ + PMQ   . Day 28 P. vivax parasitemia (n = 1349) occurred in 377 (59%) patients, peaking after the transmission season ended  over 1 year by 85.6%; annual rates per person-year (95% CI) were 1.85 (1.60\u20132.14) for AS, 1.60 (1.37\u20131.86) for CQ, and 0.23 (.17\u2013.32) for CQ + PMQ (P < .001). Age and gender did not affect either the risk or the number of P. vivax malaria recurrences. The day 6 mean (95% CI) capillary whole blood CQ drug concentrations in those with and without a first recurrence were 353 (330\u2013377) ng/mL and 397 (358\u2013435) ng/mL, respectively (P = .061). Patients with first recurrences between day 28 and 42 (n = 66) had lower day 6 CQ levels (330 [296\u2013365] ng/mL) compared to patients with no recurrence (379 [355\u2013404] ng/mL) (P = .028). Of the 47 patients who had levels measured at the time of this recurrence, all were <100 ng/mL. In the 18 patients who had a first recurrence before day 28, day 6 levels were similar to those without early recurrences , and 4 (22%) had levels \u2265100 ng/mL on the day of recurrence. Thus, only 4 of 216 infections  were CQ resistant. In vitro susceptibility assessments were successful in 95 cases; geometric mean CQ 50% inhibitory concentration (IC50) was 19.7  ng/mL. Seven isolates (7.4%) had an IC50 >50 ng/mL, suggesting resistance. Chloroquine susceptibilities were not available for the 4 patients with early (<28 days) recurrences following CQ.Overall, recurrences of on ended . Median CQ + PMQ . A histoP. vivax recurrence were similar in the AS (79% [72%\u201384%]) and CQ (78% [71%\u201384%]) groups, but were substantially lower in the CQ + PMQ group (14% [5%\u201330%]) (P < .001). The corresponding proportions with third recurrences were as follows: AS, 76% (68%\u201383%) and CQ, 71% (63%\u201379%) vs none in the CQ + PMQ group (P < .001). Over the 1-year follow-up, there were more recurrences per person-year in the AS (4.51) than CQ group , and substantially fewer in the CQ + PMQ group  (P = .013) and having a previous history of malaria . There were no significant risk factors for repeated recurrence in the CQ + PMQ group. Over all the recurrences (n = 1349), compared with CQ alone, adding PMQ radical cure reduced recurrences by 92.4%.The proportions (95% CI) of patients who had a second  < .001) . When th < .001) . If the  < .001) . IndepenP = .584), but in recurrent infections lower levels were neither associated with risk nor timing of subsequent recurrence. Chloroquine levels exceeded 100 ng/mL in 10 of 225 (4%) infections on the day of subsequent recurrence. Of the 58 patients with a subsequent recurrence before day 28, 16 (28%) had levels >100 ng/mL.The mean day 6 CQ drug level in recurrent infections  was similar to the first treatment . In the 43 patients who were anemic at enrollment (hematocrit \u226430%), the mean hematocrit actually increased by day 6 (P = .035) and CQ groups . Overall, the median fractional hematocrit recovery (day 6 to steady state) was 7.4%  (P = .009) (P < .001), age >15 years , and having a previous history of malaria   acute hematocrit reduction (day 0 to nadir at day 6) was similar in the CQ + PMQ (\u20130.8% [\u20131.4% to \u20130.3%]), AS (\u20130.8% [\u20131.2% to \u2013.3%]), and CQ groups  . Recurre = .009) . By weekAdverse event rates were similar in all treatment groups. Abdominal pain was more common in the CQ + PMQ than the CQ group but not the AS group . More paP. vivax malaria infections remain sensitive and CQ remains clinically efficacious in treatment. In areas with high-grade CQ resistance in P. vivax, national malaria programs now recommend artemisinin combination therapies. The main therapeutic challenge is the high relapse rate. In Southeast Asia and Oceania, P. vivax relapses frequently and repeatedly. Following rapidly eliminated drugs such as AS or quinine, relapses become patent approximately 3 weeks after starting treatment. More slowly eliminated drugs such as CQ suppress the asexual parasitemia of the first relapse, but there has been uncertainty whether the first relapse observed after these drugs derives from protracted suppression  of the first relapse, or clearance of this infection and emergence of the second relapse. This study, together with previous data [Although there is in vivo and in vitro evidence of CQ resistance on the northwestern border of Thailand , 13, mosous data   occurred by week 16. This suggests that studies evaluating antirelapse treatments in Southeast Asia and Oceania should have at least 16 weeks (~4 months) of follow-up to capture the majority of relapses. Primaquine  was highly efficacious with an estimated antirelapse  overall efficacy of at least 92%. Primaquine also has asexual stage activity [P. vivax malaria is enormous. In the Southeast Asia region, an estimated 4.6 million cases would have been prevented if all P. vivax infections were treated with PMQ radical cure [More than 90% of all activity  which woactivity . Approxiactivity , so 4%\u20135cal cure .P. vivax malaria and from PMQ treatment is anemia. In this low-transmission area with ready access to diagnosis and treatment, recurrent P. vivax malaria caused only small reductions in hematocrit; these reductions lessened with successive recurrences [The main clinical concern both from recurrent urrences . In conturrences . Even sourrences , 25.One of the study limitations was incomplete follow-up. This resulted from a combination of migration, environmental factors (flooding), the nature of forest work for young males, and long study duration.P. vivax malaria along the Thailand\u2013Myanmar border, although low-grade resistance has emerged. The benefits of slowly eliminated antimalarials in delaying early relapses are small in comparison with high-dose PMQ radical cure, which prevents nearly all relapses. However, when only G6PD qualitative screening is used, this regimen may cause significant hemolysis in G6PD-deficient heterozygous females. Despite this, PMQ should be used more widely. Greater availability of quantitative G6PD testing or development of safer radical curative regimens would ameliorate the risks.In conclusion, CQ continues to be an efficacious treatment of Clinical Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at Supplement FilesClick here for additional data file."}
+{"text": "Mobile phone short messaging services (SMS) have been investigated in health information reporting, provider performance, drug and diagnostic stock management and patient adherence to treatment for chronic diseases. However, their potential role in improving patients\u2019 adherence to malaria treatment and day 3 post treatment reviews remains unclear.A \u201cproof of concept\u201d open label randomised controlled trial will be conducted at four sites in Western Kenya. Principal research questions are: 1) Can mobile phone SMS reminders improve patient adherence to malaria treatment? 2) Can mobile phone SMS reminders improve day 3 post treatment reviews? Eligible caregivers (n=1000 per arm) of children under five years old with uncomplicated malaria will be randomly assigned (one to one) to: a) the current standard of care ; and b) the current standard of care plus SMS reminders. Within each arm, caregivers will be further randomized to three different categories. In categories 1 and 2, 300 caregivers per arm per category will be visited at home on day 1 and 2 of follow up respectively, to measure appropriate timing and adherence of the second Artemether-Lumefantrine (AL) dose and doses 3 and 4. Further, caregivers in categories 1 and 2 will be required to come to the health facility for the day 3 post treatment reviews. Finally, in category 3, 400 caregivers per arm will be visited at home on day 3 to measure adherence for the full AL course. Each category will be visited at home only once to avoid biases in the measures of adherence as a result of home consultations. Primary outcomes will be adherence to the full AL course (category 3), as well as, the proportion of patients reporting back for day 3 post treatment reviews (categories 1 and 2). The primary analysis will be intention-to-treat. Costs of the intervention will be measured over the period of the intervention, and a cost-effectiveness ratio will be estimated.If successful, evidence from this trial could improve malaria treatment adherence and offer pragmatic approaches for antimalarial drug resistance surveillance and risk mitigation in Africa.ISRCTN39512726 Plasmodium falciparum malaria has emerged in south East Asia (SEA), with possible foci in Western Cambodia, Western Thailand, Myanmar and possibly beyond . In brief the intervention development process was conducted through a six step process, namely:Step 1: Despite the high coverage of the network signal and mobile phone access from nationally representative surveys [ surveys ,27 facil surveys . Briefly surveys .Step 2: In this step, the various modalities of the intervention delivery were considered, including: the number, content, length, frequency and languages to be used for text messages. An intervention development workshop was conducted, involving clinicians and caregivers from the areas neighbouring the study sites, malaria epidemiologists, social scientists, mobile health programmers, and members of the district health management teams.Step 3: The understanding of the messages developed was pre-tested using focus group discussions (FGDs). FGDs were conducted among caregivers from the study sites to ensure that the messages were clearly understood by the target group. Twenty caregivers of sick children from the study health facilities were invited to participate in the FGDs during which they received SMS text messages in Dholuo, English or Kiswahili based on their language preference. Members of the groups discussed the meaning and understanding of the messages. Following the testing, the messages were amended as required.Step 4: A computerized SMS distribution system was developed by mobile phone programmers to ensure the automated SMS distribution to individual recipients upon their registration into the system. The system is a web-based application platform optimized for data collection and reporting. It is composed of: (1) an SMS data collection tool which uses a built-in SMS gateway to receive and send structured SMS messages according to the language option preferred by the caregiver, which are then stored in a local database and (2) an analytics tool that enables visualization and analysis of data sent via SMS to help real time tracking of the operational progress of the trial. Upon registration, the system creates a schedule of reminder messages to be sent at specific timings and generates log reports of all messages scheduled and those already sent.Step 5: Pilot testing of the automated distribution system and the selected messages was conducted, during a complete delivery of the intervention to 40 healthy recipients including all study investigators, and 20 malaria caregivers at health facilities not involved in the RCT. All malaria caregivers receiving the messages were visited by a social scientist to solicit their views and experiences of the pilot intervention.Step 6: Finally, based on the results of the pilot test, the content and modalities of the text-message distribution was finalized before being deployed in the RCT. The English version of the text-messaging content with timing of SMS distribution is presented in Trial design: Caregivers of eligible children with uncomplicated malaria will be randomly assigned (one to one) to two different arms: 1) the current standard of care based on provider counselling and health education alone, and 2) the current standard of care plus SMS reminders. Within each arm participants will be further randomly assigned to 3 different categories for the measurement of adherence to the different doses of AL. In the first category, 600 caregivers (300 per arm) will be visited at home on day 1 of follow up to measure adherence and timing of the second AL dose; in the second category, another 600 caregivers (300 per arm) will be visited at home on day 2 to measure adherence and timing of AL doses, 3 and 4, while, in the third category, a final 800 caregivers (400 per arm) will be reviewed at home on day 3 after they have completed the full treatment course to measure adherence for the full course of AL. All caregivers in each category will be visited at home only once to avoid biases in the subsequent measures of adherence as a result of the consultations at home. Caregivers in category 1 and 2 will be required to come to the health facility for the day 3 post treatment reviews, while those in category 3 will be visited at home on day 3.AL will be administered according to weight bands: 5-14 kg, 1 tablet twice a day for three days, 15-24 kg, 2 tablets twice a day for three days and will be directly observed for the first dose, with the remaining 5 doses being provided for administration at home. All caregivers will receive standard instructions about how to administer AL at home and when to return to the facility for scheduled visits or immediately if unwell. At first dose administration, the telephone numbers of all eligible caregivers will be registered into the automated system by the randomizing nurse who will also observe directly supervised therapy of the first dose. The registration will record the time when the first dose is administered. A procedure for guiding the correct registration of the random number that will act as a unique identifier has been put in place .In the intervention arm the registration generates a programme that automates SMS reminders from a central server, timed to eight hours after the first AL dose for the second dose and then every morning and evening (8 am and 8 pm) until the full AL course is completed. To limit the number of follow up SMS messages being received at very late hours of the night, patient recruitment will be stopped by 2 pm. Caregivers in category 1 and 2 in the intervention arm will in addition to the SMS dose reminders also receive an SMS reminder for the day 3 post treatment reviews at the health facility the evening before and on the morning of the appointment. Since caregivers in category 3 (intervention and control) will be reviewed at home on day, they will not receive SMS reminders for the day 3 post treatment reviews. Weekly SMS reminders  prompting caregivers to visit the health facility if their child is unwell will be sent to participants in the intervention arm. If such reminders result in a visit to the health facility, such a visit will be considered an un-scheduled visit. Finally, participants in the intervention arm will be sent final SMS reminders on the evening before and on the morning of the day 28 post treatment review appointment. Patients who do not return for post treatment review (intervention and control arms) will not be actively followed up.In order to ensure concealment of the intervention allocation to the trial staff, the randomization to the two study arms and subsequently to the 3 adherence measurement categories per arm was generated by an offsite statistician. Intervention allocation is concealed from the study staff reviewing the patients for treatment outcomes until evaluation is completed. However, such concealment could not be implemented for those conducting the home visits since they also collect data on SMS exposure, but we believe this lack of concealment will not affect the adherence outcome because the home visit will be conducted after the AL dose has been administered. Since caregivers will be visited at home only once, we will not measure adherence of the subsequent doses of AL in category 1 and 2. We designed the study to measure adherence to the full course of AL only in category 3 when all doses should have been completed to enable comparison with previous studies.The districts we have selected have very high malaria transmission intensity. In many malaria clinical trials recruitment rates of 5-7 caregivers per day has been demonstrated. Based on mobile phone ownership data, 50-65 % of the population is likely to own a mobile phone. So we anticipate recruiting 2-3 caregivers per day per site. Patients attending the outpatient department will be screened for eligibility criteria and those eligible will be randomized into the intervention and control arms as well as the adherence measurement categories. While recruitment at the four sites is cumulative we will try to ensure that participants at each of the four sites are as to close to equal as possible (approximately 500 participants per site). Potential challenges related to mobile phone and SMS use such as sharing of personal phones or not charging or switching off phones were determined during phase I and, if widely common, will be reinforced by the health workers to sensitize caregivers about the need to have mobile phones on and charged during the trial period.The major source of bias in adherence studies is reliance on participant recall and the possibility of false reporting. We have adopted an innovative approach of measuring adherence on different days during the dosing period, which is likely to reduce recall bias for the doses on days 1 and 2 in category 1 and 2 but not in category 3. The second primary outcome measure (day 3 post treatment review) requires objective assessment and we do not anticipate any biases in its measurement. The trial will be conducted under strict adherence to good clinical practice (GCP) standards.Plasmodium falciparum ; history of fever in the last 24 hours or presence of fever (axillary temperature \u2265 37.5\u00b0C); network coverage at patients\u2019 household; owning a mobile phone or having shared access to a mobile phone in household; ability to access a mobile phone on a daily basis for the period of follow up; ability to open and read SMS or presence of someone in the household who can open or read an SMS and written informed consent.Caregivers of male and female children aged <5 years old; body weight eligibility for AL; microscopically confirmed, mono-infection of Caregivers not owning a mobile phone; and have no shared access to any phone in the household; patients with severe malaria; danger signs: not able to drink or breast-feed, vomiting (>twice in 24 hours), recent history of convulsions (>1 in 24 h), unconscious state, unable to sit or stand; presence of concomitant illness or any condition which in the judgement of the investigator would place the subject at undue risk or interfere with assessment; severe malnutrition (weight for height<70% of the median NCHS/WHO reference); ongoing prophylaxis with drugs having antimalarial activity such as cotrimoxazole for the prevention of pneumocysti carini pneumonia in children born to HIV+ women; having taken AL in the previous one month and having participated in the current trial.The sample size estimation was based on the first primary outcome-adherence to the full course of AL treatment. Assuming a malaria treatment adherence of 65% with the current standard of care alone , a samplThe first primary outcome will be adherence to a complete AL course (doses 2-6) measured in category 3 only and defined as: a) adherent, if they present with empty blister packs and report taking the medications as recommended (\u00b1 1 hour for dose 2 and \u00b1 2 hours for doses 3-6); b) probably adherent if they do not present blister packs but report taking the medications as recommended, c) probably non adherent if they do not present blister packs and do not report taking the medications as recommended, and d) non adherent if they present blister packs with at least one tablet not used. The second primary outcome will be the proportion of patients reporting to the health facility for the day 3 post treatment review for the evaluation of clinical and parasitological cure at day 3 (only category 1 and 2).Secondary outcomes will include: adherence and timing for individual AL doses measured in category 1 for AL dose 2 and category 2 for AL doses, 3 and 4, and category 3 for all AL doses. The other secondary outcomes will be loss to follow up at day 3 defined as unable to return to the facility 24 hours after the scheduled appointment.At each of the time points ; caregiver reports and pill counts will be used for measuring adherence in the intervention and control arms. While the day 1 and day 2 visits could result into slightly better adherence for doses, 3 and 4 because they are visited at home in the early days of follow up, category 3 will provide closer to real life adherence as these patients will only be visited at home after completion of the treatment course.Measurement of the efficacy of the SMS intervention on day 3 post treatment reviewsAt enrolment, all caregivers will be informed about the importance of a day 3 post treatment review from a clinical and drug resistance surveillance perspective, to confirm a malaria free status at day, or anytime during follow up if unwell for a period of 28 days. The proportion returning for the day 3 post treatment reviews will be determined in the intervention and control arms.At enrolment and at all scheduled visits at the health facility or at home  and at all unscheduled visits, a blood smear will be taken for the estimation of parasite density. Screening thick blood smears will be stained with 10% locally prepared Giemsa for 10 minutes, while definitive thick and thin blood smears will be stained with 2% commercially prepared Giemsa for 30 minutes. The thick smears will be used to make the diagnosis of malaria, calculate parasite density and look for the presence of gametocytes, while the thin smears will be used for speciation. The number of asexual parasites per 200 WBCs will be counted, and if there are less than 10 parasites per 200 WBCs, the count will be continued until 500 WBCs. Parasite density will be estimated based on the assumption of a standard estimate of 8000 WBCs/UL. A blood smear will be deemed negative, if there are no asexual parasites after counting a hundred high power microscopic fields. In addition, haemoglobin level will be estimated at day 0 and day 28, using a hemocue for the quantitative determination of haemoglobin in blood using a specially designed photometer and micro-cuvettes.This study protocol has been reviewed by six independent peer reviewers nominated by the United Kingdom Medical Research Council (UK-MRC) before the grant was awarded. Further, the study protocol, the informed consent documents and all educational or recruitment material was further reviewed and approved by the Kenya Medical Research Institute ethical review committee and the University of Oxford ethical review board (OXTREC).The study will be conducted as multi-site trial with a field coordinator supervising all the study sites. After initiation of the trial, monitoring and supervision of the trial is being conducted to guarantee that all study procedures are being followed in accordance with GCP principles. An independent certified monitor from the East Africa Consortium for Clinical Research (EACCR) will be responsible for monitoring the trial. A Trial Steering Committee (TSC) was constituted before the study commenced to address policy and operational issues related to the study; and protecting the scientific conduct and integrity of the trial. In addition, a Data Monitoring Board (DMB) was constituted, whose main roles are: monitoring the recruitment rates of the trial, revising in blind a significant percentage of the primary outcome data and assessing the quality of the data.Individual patient level pooled analysis of the data from the four sites will be conducted, according to CONSORT standards for reporting RCTs. A random effects model will be used to adjust for site and other covariates. To analyse for effect differences for the primary outcomes, intention to treat (ITT) principles will be used and additional sensitivity and per protocol (PP) analyses will be conducted separately. ITT implies that all randomised patients will be considered in the outcomes as per randomization. Missing data will be imputed as intervention failure (non-adherent or not report for post treatment review), using multiple imputation for missing data not directly related to adherence. The latter is considered the gold standard for handling missing data as it leads to unbiased estimates and standard errors compared to single imputation methods . In the The quantitative study will provide information on the proportion of participants adhering to the treatment regime and the proportion returning for the day 3 post-treatment reviews. However, understanding the factors influencing this behaviour requires a more-in-depth qualitative approach. Following completion of the RCT, in-depth interviews will be conducted with a purposively selected sample of participants to explore their perceptions and experiences of the SMS intervention and to investigate the factors influencing their behaviours. Forty caregivers will be selected stratified by those who adhered to medicines and the day 3 post-treatment review and those who did not. A semi-structured interview guide with open-ended questions will be used to ensure that key topics are covered as well as allow flexibility to explore issues raised by participants during the interview. All interviews will be digitally recorded, with the audio files subsequently downloaded onto a computer and transcribed into Word. The data will be entered in NVivo software for data management and a framework approach will be adopted in the analysis. The process of analysis will involve familiarization with the data, development of initial codes based on the research questions and issues emerging from the data, refinement of codes, and their allocation to the primary and emerging themes. In addition, in-depth interviews will be conducted with eight health workers to get their views about the feasibility and anticipated burdens of the day 3 post treatment reviews in routine settings.Finally, a cost-effectiveness analysis of the intervention with respect to the primary outcomes will be conducted. Costs will be measured under trial conditions, from the provider\u2019s perspective and will include costs required for the Kenya ministry of health to expand the delivery of the intervention nationally. Cost data will be collected using standard procedures and unit costing menus  and willIn order to avoid long time lags between the generation of research evidence and its uptake into policy and practice, there is a need to think strategically about the impact of research. There are a few good examples where researchers have closely engaged policy makers to improve evidence-led malaria case management policies in east Africa. In the late 1990s, there was evidence from research demonstrating widespread malaria parasite resistance to monotherapy. However, despite the over whelming evidence, there was no change in treatment policy. In view of the failure of research to impact policy, the East African Network for Monitoring Antimalarial Treatment (EANMAT) was established and generated credible evidence that supported the change in policy from monotherapy to ACT. The success of EANMAT was attributed to bringing together national malaria control programmes and research stakeholders ,35. If tThe uptake of research evidence into policy can further be facilitated if policy makers/implementers are actively engaged from the design of research and agenda setting. This research was designed with the close involvement of the malaria control unit and the devolved districts in Kenya so as to get their buy in. If the trial is successful, the research team will work in partnership with the national malaria control programmes, the roll back malaria (RBM) partnership, and the World Health Organization\u2019s regional and country offices to ensure that the results are translated into policy and practice locally and regionally. Researchers on our team have established close working relationships with the Kenya ministry of health and other ministries of health in the sub-region and will use this experience to facilitate the uptake of our research evidence into policy in Kenya and the region.Policy makers and researchers by the nature of their motivations have different roles. However the two groups should talk closely to each other and strive for a better mutual understanding. Policy makers/implementers often have wider horizons, are curious about other solution options and may not rely on only one type of research. Moreover, there are other aspects-economic, cultural, behavioral and social, that impact policy uptake. The research described in this protocol is multi-disciplinary and involves investigating different aspects-efficacy, social, cost and cost effectiveness, so it is likely to generate diverse and critical information to enable policy makers to take informed decisions.Finally, the increasing use of new m-Health technologies to improve healthcare deliver in Africa is commendable. However, the introduction of m-Health technologies into health care delivery should be based on a robust evidence base of their efficacy, cost, effectiveness, feasibility and scalability. The scientific rigor required in assessing their impact, the assumptions that underpin their simplicity, low cost and effectiveness should not be less than would be required for any other new health intervention."}
+{"text": "P. falciparum malaria treated with parenteral artesunate followed by an oral artemisinin-combination therapy. We analyzed data from two independent studies in which children were followed on Days 7,14, and 28 after treatment with artesunate. Specific hematological parameters included the presence of hemoglobinopathies and erythropoietin. Presence of once-infected erythrocytes was assessed by flow cytometry in a sub-population. Of 143 children with a geometric mean parasitemia of 116,294/\u00b5L , 91 (88%) had anemia (Hb\u2009<\u200910\u2009g/dL) at presentation. Hemoglobin increased after Day 7 correlating with increased erythropoiesis through adequate erythropoietin stimulation. 22 children (24%) remained anemic until Day 28. Post-artesunate delayed hemolysis was detected in 7 children (5%) with only minor differences in the dynamics of once-infected erythrocytes. Hyperparasitemia and hemoglobin at presentation were associated with anemia on Day 14. On Day 28 only lower hemoglobin at presentation was associated with anemia. Most children showed an adequate erythropoiesis and recovered from anemia within one month. Post-artesunate delayed hemolysis (PADH) and hyperparasitemia are associated with early malarial anemia and pre-existing anemia is the main determinant for prolonged anemia.The pathophysiology of malarial anemia is multifactorial and incompletely understood. We assessed mechanistic and risk factors for post-malarial anemia in Ghanaian and Gabonese children with severe  Its use is not restricted by adverse events typical of quinine, such as QT prolongation, visual and auditory disturbances, and severe hypoglycemia5. Consequently parenteral artesunate is recommended as the treatment of choice for severe malaria by the World Health Organization6. The clinically most relevant adverse event of parenteral artesunate is delayed hemolysis occurring approximately two weeks after treatment8. Hemolysis affects between 20 and 30% of adult travelers from Europe with no or waning semi-immunity against malaria and high parasitemia treated with artesunate9. Delayed hemolysis has been reported in 5% of African children exposed to artesunate11.The superior efficacy of artesunate over quinine in the treatment of severe malaria has been shown in different clinical and geographical settings12. The causes of anemia in this population are manifold and range amongst others from iron-deficiency to parasitic infections and genetic traits13. Additionally, anemia is major clinical hallmark of malaria and related to direct destruction of red blood cells by the parasite but also indirect effects, such as disruption of erythropoiesis and immune mechanisms14. Here we describe the recovery from malarial anemia in children with severe malaria treated with artesunate and assess predictive and mechanistic factors contributing to prolonged anemia.Anemia is highly prevalent in African children under the age of 5 years16. The primary endpoint of this study was the incidence of post-artesunate delayed hemolysis. Exploratory results of this sub-study have been published10.This analysis is based on individual patient data from two studies: between April and September 2012 patients were recruited into a sub-study of the \u201cComparative, Open Label, Dose and Regimen Optimization Follow-up Study of Intravenous and Intramuscular Artesunate in African Children with Severe Malaria\u201d (PACTR201102000277177) at the Komfo Anokye Teaching Hospital in Kumasi, Ghana, and at the Centre de Recherches M\u00e9dicales de Lambar\u00e9n\u00e9, Gabon, conducted by the Severe Malaria in African Children Network (SMAC)Between January and August 2015 patients were recruited into a second study at the St. Michael\u2019s Hospital in Pramso and the Komfo Anokye Teaching Hospital in Kumasi, Ghana.P. falciparum malaria (>5000 parasites/\u00b5L on a thick blood smear) with signs or symptoms requiring hospitalization as judged by the treating physician as per the definitions of the Severe Malaria in African Children (SMAC) network reflecting standard practice in most African settings18. In both studies, patients were treated with parenteral artesunate followed by a course of weight-adapted oral artemether/lumefantrine. Within the first study, all patients received a total dose of 12\u2009mg/kg artesunate according to three randomly assigned regimens . Within the second study, patients received at least three weight-adapted doses of parenteral (i.m. or i.v.) artesunate (2.4\u2009mg/kg). If the patient was able to tolerate oral medication, treatment was switched to artemether/lumefantrine at this time point \u2013 else parenteral artesunate was continued. In both studies patients received folic acid supplementation for at least 14 days after treatment initiation.Children aged 6 months to 10 years were included in both studies if they presented to a study site with a primary diagnosis of 10. Visits were scheduled on Day 0 (first day of treatment), Days 7 (\u00b12), 14 (\u00b12), and 28 (\u00b14). The second study included an additional visit on Day 2.Procedures have been described beforeReticulocytes were assessed microscopically: after supravital staining with brilliant cresyl blue, reticulocytes per 1,000 erythrocytes were counted. Haptoglobin and erythropoietin was measured from serum stored at \u221280\u2009\u00b0C after the end of recruitment by a commercial quality-controlled laboratory in Hamburg, Germany.20.Genotyping of red cell polymorphisms was performed by high-throughput genotyping using fluorescent melting curve assays on 384-well microplate formats in a homogenous system 22. Erythrocytes were analyzed by flow cytometry . Erythrocytes were incubated with monoclonal mouse anti-RESA (ring-infected erythrocyte surface antigen) antibodies  followed by Peridinin chlorophyll protein (PerCP)-labelled anti-mouse-IgG to label RESA-positive erythrocytes. Nucleic acid staining was performed with Syto 16 (life technologies\u2122). RBCs were plotted in two-dimensional scattergrams and gated according to their logarithmic forward and side scatter properties. Syto16 (DNA staining) was detected in the FL1 channel, PerCP (RESA-staining) was detected in the FL3 channel.Samples from the second study were assessed for the presence of once-infected (\u201cpitted\u201d) erythrocytesDouble-positive (PerCP and Syto 16) erythrocytes were defined as infected, PerCP-positive, SYTO 16-negative erythrocytes as once-infected erythrocytes. Flow-cytometry data analyses was performed using FlowJo v10 software . Two representative plots of one patient showing the switch from infected to once-infected RBCs are available as Supplementary Fig.\u00a024.We defined anemia as a hemoglobin level lower than 10\u2009g/dL, corresponding to at least a moderate anemia as defined by the World Health Organization10. Patients had to present both any decrease in hemoglobin and any increase in LDH between Days 7 (\u00b12) and 14 (\u00b12) in combination with both an elevated LDH (>350 IU/L) and low haptoglobin (<0.3\u2009mg/dL) on Day 14.PADH was defined as previously reported by our groupStatistical analyses were performed with Stata IC 15 . Descriptive characteristics were reported using absolute and relative frequency for categorical variables, mean and 95% confidence interval for normally distributed, and median and interquartile range for non-normally distributed continuous variables. Continuous variables were represented by their mean and standard error of the mean in figures. The association between anemia level and transfusion status was assessed by a chi-square test. The difference in once-infected pitted erythrocytes between patients with PADH and those without was tested by the Mann-Whitney-U-test. To build models predicting anemia on Day 14 (\u00b12) and 28 (\u00b14) variables which were associated in the univariable analysis with anemia at a p-level of 0.1 were included in a multivariable logistic regression model.Study protocols have been reviewed and approved by the respective institutional review boards . Children were only included if informed consent was provided by the parent or legal guardian. All study procedures were performed in concordance with the Declaration of Helsinki.143 patients were eligible for this analysis, as represented in the patient flow diagram  were anemic (Hb <10\u2009g/dL). 37 children (26%) with a mean hemoglobin of 5.5\u2009g/dL (95% CI 5.1\u20136.0) received a blood transfusion. Children without transfusion had a mean hemoglobin of 9.8\u2009g/dL (95% CI: 9.4\u201310.2). Figure\u00a0All patients with a hemoglobin lower than 5\u2009g/dl at presentation received a transfusion, while no patient with a hemoglobin equal or higher than 9\u2009g/dl received a transfusion. Transfusion rates gradually decreased between these values . To explore the impact of transfusion on hemoglobin during follow-up, a restricted analysis was performed in children with hemoglobin levels between 6\u2009g/dL and 7\u2009g/dL (n\u2009=\u200913 transfused children and 19 non-transfused children). This restriction was chosen for three reasons: (a) to match hemoglobin levels at presentation; (b) as in this group there was the largest variability in transfusion patterns; (c) transfusions are not generally recommended over 7\u2009g/dL. Children who received a transfusion had higher hemoglobin levels at follow-up with a mean hemoglobin at Day 28 of 11.2\u2009g/dL (95% CI: 10.5\u201312.0) compared to those without a transfusion  only 4 were heterozygous for Hemoglobin S, 9 for hemoglobin C, and one had hemoglobin C disease. Due to the ensuing low power, hemoglobinopathies were not further assessed as risk factors for anemia. Similarly, the sample size was too low to detect any association between G6PD status in boys and anemia.10. The two new cases of PADH were only moderately affected with a nadir in hemoglobin of 10.0\u2009g/dL in both patients on Day 14. For these two patients, pitting data were available. Compared to 17 children with a similar parasitemia range  without PADH, there was some evidence that the median number of pitted erythrocytes was higher on Day 7 in children with PADH  but not on Day 2 . There was some evidence for a larger decrease in pitted erythrocytes between Days 7 and 14 in children with PADH .7 patients (5%) developed PADH, 5 of which have been described before9/L per unit increase in the logarithm of erythropoietin; 95% CI: 8.8\u201346.9; p\u2009=\u20090.005).Figure\u00a025. Additionally, malaria itself as well as the artemisinins impair erythropoiesis and early erythrocytic stages and reticulocytes are destroyed by erythrophagocytosis27. In combination with the potential destruction of infected reticulocytes, this translates to an inadequately low reticulocyte count at presentation, while erythropoietin is already elevated in response to anemia. There is a strong association between erythropoietin levels and hemoglobin levels at all time points, showing an adequate erythropoietin response. There are contradictory reports on whether the erythropoietin response is adequate in symptomatic malaria patients mainly due to methodological issues in older studies30. Our study showed an initially blunted response to erythropoietin but not a reduction in erythropoietin itself. Accordingly, after the initial lag phase, children showed a strong increase in reticulocytes and the erythropoietic response leads to a hematological recovery.In this population, children show a satisfactory hematological recovery after treatment with parenteral artesunate for malaria with a steady increase of hemoglobin after Day 7. The initial decline in hemoglobin up to the nadir on Day 7 is explained by the destruction of infected and uninfected red blood cells in acute malaria12. To explore the interplay of these covariates on malarial anemia and its recovery, larger cohort studies are needed. However, our data show that children recover from the early post-malarial decline in hemoglobin within one month after treatment. By raising the baseline hemoglobin through population-based measures, such as iron supplementation or mass treatment of helminthic infections, the duration and severity of malarial anemia on the individual child could be reduced. Similarly through reductions in malaria incidence and the subsequent reduction in malaria episodes per child, baseline hemoglobin values are likely to increase and the impact of each episode could be minimized31.The major risk factor for prolonged anemia is a low hemoglobin level on presentation with an influence of baseline parasitemia on Day 14 but not on Day 28. Baseline causes of anemia, such as iron deficiency, malnutrition, hemoglobinopathies, and chronic infections prevail in malaria-endemic countries6. Similarly, all children except one with a hemoglobin of less or equal than 6\u2009g/dL have been transfused. There was no consensus for transfusing those children with a hemoglobin level between 6 and 7\u2009g/dL, showing that the indication for transfusion in this group is mainly guided by individual patient factors. A conservative approach to transfusion has been historically propagated to ensure availability of blood transfusions and in the light of potential blood safety issues in African countries. However, there has been no solid evidence guiding transfusion thresholds in African children \u2013 especially not in severe malaria32. In our sample, transfusing children increased their hemoglobin up to 28 days after malaria compared to those without transfusion with the same starting hemoglobin. Whether this increase in hemoglobin also directly or indirectly reduces morbidity (such as fatigue or susceptibility to infections) is unclear. A recent randomized controlled trial in African children  did not show a difference in clinical outcomes after 6 months between aggressive and conservative transfusion approaches34.All children presenting with a hemoglobin level of less or equal than 5\u2009g/dL received a transfusion, as WHO recommends10. The overall prevalence and severity of PADH seems to be lower in patients living in malaria-endemic settings than in returning European travelers with severe malaria11. Available evidence supports an overall frequency of PADH in African children treated with artesunate of 5% - with severe PADH occurring in 1% of children35. While these results provide some reassurance, an adverse reaction with a frequency of 1% would still be defined as \u201ccommon\u201d by the standard categories as recommended by the Council for International Organizations of Medical Sciences (CIOMS)36. Children reported in our studies had only moderately severe malaria; only a small proportion had very high parasite counts. We cannot rule out a significantly higher frequency of PADH in children with parasite counts over 250,000/\u00b5L. More evidence in this patient population with high parasitemias is critical, especially as the life-saving benefit of artesunate is highest in this patient group1.In addition to five previously reported patients with PADH, we identified two additional patients with signs of PADH in the follow-up studyWith a lower than expected incidence of PADH (linked to lower than expected parasite counts), pathophysiological analyses can only remain exploratory due to a very low sample size.Most children with malaria show an adequate erythropoietic response to malarial anemia and show a full recovery one month after treatment with parenteral artesunate. The incidence of PADH was lower than expected, but it should be investigated as a risk factor for anemia in a higher risk population with higher parasitemia. The main risk factor for prolonged anemia in our cohort of African children treated with artesunate is pre-existing anemia upon presentation.Supplementary information"}
+{"text": "P. falciparum gametocyte prevalence and density were lower in the MB arm than in the PQ arm, which reached statistical significance on day 2 . However, it should be considered that PQ was given only on day 2. MB-ACT appears to be an interesting alternative to PQ-ACT for the treatment of falciparum malaria. While there is a need to further improve MB formulations, MB-ACT may already be considered useful to reduce falciparum malaria transmission intensity, to increase treatment efficacy, and to reduce the risk for resistance development and spread.Artemisinin resistance is threatening global efforts for malaria control and elimination. Primaquine (PQ) and methylene blue (MB) are gametocytocidal drugs that can be combined with artemisinin-based combination therapy (ACT) to reduce malaria transmission, including resistant strains. Children (6\u201359 months) with uncomplicated falciparum malaria in Burkina Faso were treated with artesunate-amodiaquine (AS-AQ) and randomized to MB (15 mg/kg/day for 3 days) or PQ (0.25 mg/kg at day 2) with the aim to show non-inferiority of the MB regimen with regard to haematological recovery at day 7 (primary endpoint). MB-AS-AQ could not be shown to be non-inferior to PQ-AS-AQ , however, haemoglobin recovery following treatment was alike in the two study arms . Occurrence of adverse events was similar in both groups, except for vomiting, which was more frequent in the MB than in the PQ arm . Adequate clinical and parasitological response was above 95% in both groups, but significantly more asexual parasites were cleared in the MB arm compared to the PQ arm already on day 1 . Moreover, Trial registration: ClinicalTrials.gov NCT02851108. Malaria remains the most important parasitic disease worldwide . Globallde novo in other endemic areas, including sSA [P. falciparum parasites, including drug-resistant parasites [Artemisinin-based combination therapy (ACT) is now the standard first-line treatment for uncomplicated falciparum malaria in all endemic areas , 5. Howeding sSA , 5. A poding sSA , 7. Combarasites \u201311.P. falciparum gametocytes but is not widely used in endemic areas because of concerns on the risk of haemolytic anaemia in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency [PQ is known to be effective against ficiency . Neverthficiency , 12. In ficiency , 14.P. falciparum and P. vivax asexual parasites, demonstrated synergy in combination with artemisinin derivatives, and confirmed a substantial gametocytocidal effect in P. falciparum infection [MB is the oldest synthetic antimalarial drug . In histnfection , 9, 16. nfection , 10.The study was designed as a mono-centre, two-arm randomised controlled phase II non-inferiority study and was conducted in the research zone of the Centre de Recherche en Sant\u00e9 de Nouna (CRSN) in north-western Burkina Faso . MalariaThe primary objective was to determine the safety of MB as compared to PQ when combined with AS-AQ for the treatment of uncomplicated falciparum malaria in children 6\u201359 month old. Secondary objectives were to determine the treatment efficacy and to assess its acceptance by the parents/care givers.The study was not blinded given that MB results in blue urines. Nevertheless, all lab technicians were blinded to the origin of samples to be analysed.Children attending Nouna Hospital with confirmed uncomplicated falciparum malaria were recruited if they fulfilled all the following inclusion criteria: 6\u201359 months old; parasite density: 2.000\u2013100.000 parasites/\u03bcl; axillary temperature \u2265 37.5\u00b0C or a history of fever during the last 24 hours; weight \u2265 6 kg; and written informed consent of parents or care givers. Children were excluded if they had at least one of the following criteria: Severe malaria ; mixed mFever measurement points equipped with rapid diagnostic tests (RDT) for malaria  were established in a rotating manner in different quarters of Nouna town. Eligible children were transferred to Nouna Hospital for further examination and final inclusion into the study. After informed consent, children were randomised by the study physician to one of the two study arms according to a pre-established randomization list using block randomization, with each treatment allocation concealed in opaque sealed envelopes that were opened only after the patient\u2019s recruitment.AS-AQ  was administered according to three weight groups . 2 mg MB mini-tablets  were administered once daily (together with AS-AQ) at a daily dose of 15 mg/kg for 3 days (days 0\u20132), and according to four weight groups . Depending on the age of the study children, tablets were dissolved or crushed and administered with water. MB was given on a spoon with local food . PQ tablets  were administered at the last day of AS-AQ treatment (day 2). Tablets were dissolved in water and given with a plastic cup or spoon, followed by a cup of orange juice, according to four weight groups . All treatments were directly observed. Full treatment was re-administered once if vomiting occurred within half an hour of treatment.Children with an axillary temperature \u2265 38.5\u00b0C received standard doses of paracetamol (10 mg/kg every 6 hours) until fever subsided. Study children developing severe malaria during the trial were admitted to the Nouna Hospital and treated with artesunate intravenously according to national guidelines.After treatment (days 0\u20132), the parent/care giver was asked to return with the child for scheduled visits on days 3, 7, 14 and 28, or if any symptoms occurred. For each visit, a physical examination was performed by the study clinicians, vital signs were recorded, and axillary temperature measured with an electronic thermometer. Adverse events (AEs) and serious adverse events (SAEs) were recorded and monitored throughout the study. Rescue treatment for recurrent malaria infections identified during the 28-day follow up as well as for severe malaria was according to local national guidelines.CRSN which is integrated in the Nouna Hospital. Quality control of all malaria slides was done at the malaria laboratory of the MRC Unit The Gambia. All molecular biology diagnostics were undertaken at the Institute of Tropical Medicine and International Health, Charit\u00e9, Berlin. DNA was extracted using commercial kits . Recrudescence was differentiated from re-infection by comparing PCR-generated P. falciparum msp1 and msp2 amplicons at day 0 (before treatment) and at the time of recurrent infection. G6PD typing  was performed by melting curve analysis applying commercial primers and probes .Dried blood spots on filter paper, for G6PD and parasite genotyping, were collected on days 0, 7, 14 and 28, and whenever a child presented to the study team with symptoms during follow-up. The laboratory work was conducted in the laboratory of the P. falciparum gametocyte prevalence and density by microscopy, the area under the curve (AUC) of P. falciparum gametocyte density, the incidence of observed and self-reported AEs and SAEs during follow-up, fever and parasite clearance rates as well as the rates of adequate clinical and parasitological response (ACPR), early treatment failure (ETF), late treatment failure (LTF) and late parasitological failure (LPF) [The primary outcome was haematological recovery (haemoglobin changes during follow-up); more specifically haemoglobin difference between day 7 and day 0 was the primary endpoint. Secondary outcomes were re (LPF) . The repThe AUC was calculated using the linear trapezoidal rule taking the measurements at day 0, 1, 2, 3, 7, 14 and 28 into account.The primary efficacy analysis was based on the full analysis set (FAS) according to the intention-to-treat (ITT) principle . AdditioThe sample size calculation was based on the primary comparison of the mean haemoglobin change between day 0 and day 7. Assuming a standard deviation of 1.1722 in both groups , 50 patiMB and \u03bcPQ are the mean differences in haemoglobin between day 7 and baseline for the MB and PQ group, respectively. Non-inferiority was tested at a significance level of \u03b1 = 0.025 using a one-sided t-test and the mean difference between groups was determined with a 95% confidence interval. Missing data for the primary outcome variable was replaced using multiple imputations [The hypotheses for the primary efficacy analysis wereutations . A complSecondary endpoints were assessed by means of descriptive statistics. Since the p-values derived are solely of an exploratory nature, no adjustment for multiple testing was done.Haemoglobin differences compared to baseline were analysed using a linear mixed model for repeated measures adjusting for baseline haemoglobin, treatment group, and time. In addition, separate linear models were fitted for each time point adjusting for baseline haemoglobin. The gametocyte densities and AUC were compared between groups using Mann-Whitney-U tests, and gametocyte prevalence using Cochrane-Mantel-Haenszel tests adjusted for baseline gametocyte prevalence. ACPR, ETF, LCF, LPF, fever and parasite clearance rates were compared using chi-squared tests. The acceptance of the different treatment regimens was compared using the Mantel-Haenszel chi-squared test for ordinal outcomes.AEs and SAEs were tabulated; absolute and relative frequencies with 95% confidence intervals were calculated. Possible differences between the treatment groups were tested using the chi-squared test.rd of October 2016, and the 7th of November, 2016, 100 children were recruited in the study out of 1,389 febrile children screened . Those enrolled were randomly assigned to receive, besides AS-AQ, either MB (n = 50) or PQ (n = 50) (Between the 3(n = 50) .P. falciparum gametocytes at baseline. Full G6PD deficiency (hemizygous or homozygous for GdA-) was observed in 14/100 (14%) patients and heterozygosity in 16/100 (16%).Overall, baseline characteristics between the two study arms were similar . The meaAll 100 study children were included in the ITT analysis. For the PP analysis, 42 in the MB arm and 49 in the PQ arm were included . In the All parents/care givers of children treated with MB confirmed the appearance of blue urines from the beginning of the treatment until day 3; none of the parents of children treated with PQ reported blue urines.The mean Hb difference between treatment groups on day 7 was -0.352  (p = 0.0767) for one-sided non-inferiority test, indicating that it was not possible to demonstrate non-inferiority according to the chosen non-inferiority margin of \u03b4 = -0.7. The complete case analysis of the ITT set and the PP set yielded comparable results for the mean difference . These results indicate a slightly worse haematological recovery for MB-AS-AQ compared to PQ-AS-AQ, which could neither be shown to be non-inferior nor inferior to PQ, since all confidence intervals contain both the non-inferiority margin of -0.7 and the value 0.The PP analysis provided similar results.One child in the MB arm developed repeated vomiting of the study medication on day 0 and was considered as a case of severe malaria as it was a rather young child (15 months) with high parasitaemia (14.600/\u03bcl), and as it was very weak . A 22 months old child  in the MB arm developed severe anaemia as defined in the present study , was hospitalized and received a blood transfusion, afterwards the child fully recovered. This was considered as a SAE; there were no further SAEs and no deaths in the study.By day 3, 97% of all patients were afebrile . It should be noted that 35 patients had only a history of fever at inclusion and not a raised body temperature; they were excluded from the fever clearance analysis.There were no differences in the ITT analysis between the two study groups in terms of PCR-uncorrected  (p = 0.677) and PCR-corrected  (p = 0.966) ACPR rate . AnalysiP. falciparum gametocytes in the MB and PQ arms was 10% and 18%, respectively. During follow-up and adjusting for baseline values, the gametocyte prevalence was consistently lower until day 14 in the MB than in the PQ arm, although it reached statistical significance only at day 2. At baseline, the mean density of P. falciparum gametocytes in the MB and PQ arm was 20.0\u00b170.1/\u03bcl and 27.2\u00b175.6/\u03bcl, respectively. During follow-up and adjusting for baseline values, gametocyte density was consistently lower until day 14 in the MB than in the PQ arm; the difference was statistically significant at days 1 and 2. The mean AUC of P. falciparum gametocyte density was lower in the MB than in the PQ arm, but the difference was not statistically significant (p = 0.165). However, it has to be considered that the differences between the two arms do not involve a direct comparison between MB and PQ on respective follow-up days as different application schemes were used in the two study arms.Overall, at day 14 more than 80% of parents or care givers reported a good acceptance of the study medication. However, acceptance was lower in the AS-AQ-MB compared to the AS-AQ-PQ schedule .P. falciparum parasites, and [P. falciparum gametocytes during follow-up.The main findings from this study are that treatment with MB-ACT compared to PQ-ACT is associated with  a highertes, and  a lower That MB is having a strong bitter taste is well known since it has been used as an antimalarial some 130 years ago . ProvidiP. falciparum transmission intensity [MB belongs to the large group of drugs\u2013many of them anti-malarials\u2013which are a risk factor for haemolysis in patients with G6PD deficiency. With regard to such risk in sSA, a synopsis of four RCTs comparing haemolysis risk of MB-based combination therapy (n = 844) with control therapy (n = 161) showed a small but significant effect of MB on the reduction of haemoglobin values in the treatment of children with malaria in West Africa, which was however considered not to be of clinical relevance . PQ is antensity , 12. In ntensity .in vitro to act synergistic with artemisinin derivates, which has already been supported by data from previous clinical studies in sSA [MB has been shown s in sSA , 26, 31.P. falciparum gametocytes, and this effect has been confirmed by a clinical study in Burkina Faso [P. falciparum transmission [P. falciparum gametocytes. If this effect would lead to a reduction of transmission intensity under malaria elimination program conditions needs to be studied in large community-based trials.MB has recently been shown in a preclinical study to be the most effective antimalarial for reducing ina Faso , 33. Morsmission . The finP. falciparum parasites and appears to have a very low potential for resistance development itself [Artemisinin resistance has rapidly been spreading in recent years in South-East Asia , 5. If at itself . Using tt itself , 35. ThuThis study has some limitations. First, MB was provided as uncoated mini tablets. If these mini tablets would have been given in a taste-masked formulation, the occurrence of early vomiting might have been much reduced. Second, as MB was given over three days from day 0 until day 2 and PQ was given as single dose treatment on day 2, the effects of the two regimens (including the different drug volumes applied) are not fully comparable and this may partly explain the observed differences in vomiting rates and acceptance as well as the tendency to a difference in haemoglobin development during follow-up. Moreover, at least the early effects of the different treatments on gametocyte prevalence and density are clearly influenced by MB and PQ being given in different schedules. Finally, non-inferiority could not be demonstrated for the primary comparison of the study. In the sample size calculation, we assumed no difference between the two treatment arms regarding the mean haemoglobin change between day 0 and day 7, while we observed a difference of -0.352 between MB and PQ in our trial. This indicates that we based our sample size calculations on a too optimistic treatment effect estimate, thus leading to a reduced power for the primary analysis of our trial.P. falciparum transmission intensity, to increase treatment efficacy, and to reduce the risk for development and spread of malaria parasites resistant against ACT.Although non-inferiority could not be demonstrated, adding MB to ACT has been shown to have a number of potential benefits regarding secondary outcomes of the trial. While there is a need to further modify the MB formulation to improve acceptability, adding MB to existing ACT should be considered as potentially useful to reduce S1 Checklist(PDF)Click here for additional data file.S1 Protocol(PDF)Click here for additional data file.S1 Dataset(ZIP)Click here for additional data file."}
+{"text": "Plasmodium vivax is the main cause of malaria in Nepal. Relapse patterns have not been characterized previously.P. vivax malaria were randomized to receive chloroquine  alone or together with primaquine  and followed intensively for 1 month, then at 1- to 2-month intervals for 1 year. Parasite isolates were genotyped.Patients with P < .0001). Microsatellite genotyping showed relatively high P. vivax genetic diversity  reflecting a low transmission preelimination setting. Of the 12 genetically homologous relapses, 5 (42%) occurred in a cluster after 9 months, indicating long latency.One hundred and one (49%) patients received CQ and 105 (51%) received CQ + PQ. In the CQ + PQ arm, there were 3 (4.1%) recurrences in the 73 patients who completed 1 year of follow-up compared with 22 of 78 (28.2%) in the CQ-only arm (risk ratio, 0.146 ; P. vivax malaria in Nepal.Although there may be emerging CQ resistance, the combination of CQ and the standard-dose 14-day PQ regimen is highly efficacious in providing radical cure of short- and long-latency  Plasmodium vivax malaria treatment in Nepal proved to be well tolerated and highly effective in preventing both short- and long-latency relapses.Chloroquine and a standard-dose 14-day primaquine regimen for  Plasmodium vivax ) of ent arms . The treent arms .P = .85) and 5 (5.0%) by day 3 (P = .006)  patients had persistent parasitemia by day 1 and 17 (17.2%) still had detectable parasitemia by day 3. In the CQ + PQ arm, the corresponding numbers were 88 (83.8%) by day 1 ( = .006) . In the P < .0001. The Kaplan\u2013Meier cumulative mean probabilities of recurrence in the CQ-only group were 1.5%  at 28 days, 3.1%  at 42 days, 18.6%  at 180 days, and 26.4%  at 1 year. By comparison, in the CQ + PQ group, the cumulative mean probability of recurrence at 1 year was 3.9% (0\u20138.1%). None of the sociodemographic characteristics were associated with recurrences  patients of the 151 patients who completed follow-up for 1 year had recurrences of vivax malaria . All werurrences .P. vivax isolates at day 0 was 0.843 . The majority (84%) of infections were apparently of a single genotype. The mean multiplicity of infection was 1.05 . If all alleles on day 0 (pretreatment) and the day of recurrence were the same, the recurrence was considered definitely to be the same genetically . Of the 25 paired P. vivax isolates, 10 were the same at each locus , 1 pair differed at 1 microsatellite locus, and the other differed at 2 loci. Thus, 12 (48%) recurrences were considered genetically homologous, and therefore assessed as very likely to be relapses. In the other 13 pairs (52%), the majority of alleles were different and so the recurrence isolates were considered heterologous . Following PQ radical cure, there were only 3 recurrences  and each was heterologous. As 2 of these occurred >4 months after enrollment, they were considered probable new infections. There was a clear temporal pattern to genetically homologous recurrences , which diverged from heterologous recurrences after 2 months with 7 of 12 relapses occurring within 5 months and the other 5 occurring in a tight cluster after 9 months  was detected by microsatellite marker MS8 and the lowest number of alleles was found in MS7 (n = 5). The mean heterozygosity of the 9 microsatellite markers in 25 9 months .Chloroquine remains the current first-line treatment for vivax malaria in Nepal. In general, CQ is still effective, although one-sixth of patients receiving CQ alone in this study had not cleared their parasitemias by day 3. There was 1 early treatment failure and 1 early relapse (day 21). These findings may well reflect emerging CQ resistance . Adding P. vivax is well described from South Asia [P. vivax [P. vivax strains similar to the Madagascar strain may have an early relapse, but more commonly relapse around 9 months after the initial infection [P. vivax in Nepal has this phenotype. This emphasizes the necessity of follow-up for 1 year when assessing vivax malaria in areas with long-latency strains. In recent years, this has been shown in Afghanistan [Genotyping cannot reliably distinguish relapses from newly acquired infections, as relapses can derive either from hypnozoites derived from sporozoites inoculated by the mosquito that caused the incident infection, or from previously acquired hypnozoites , 17. Oveuth Asia , 15. Extnfection . At leashanistan , Pakistahanistan , and Mexhanistan , where sP. vivax malaria. Both the G6PD-deficient patients identified were allocated to the CQ-only group. In nationwide studies conducted among a malaria-uninfected population, the overall prevalence of G6PD deficiency was approximately 6%. However G6PD deficiency in Nepal was more concentrated in particular ethnic groups in the southern plains [P. vivax infections [As in most areas, radical cure with CQ (25 mg base/kg given over 3 days) and PQ (0.25 mg/kg/day for 14 days) was well tolerated and highly effective in Nepal . In thisn plains , who werfections . As NepaSince 2009, the national malaria control program of Nepal has adopted the 3-day CQ and 14-day PQ regimen as first-line treatment for vivax malaria. As much of the remaining vivax malaria in the country is in the mobile, male migrant labor population, this poses specific challenges for adherence to lengthy treatment regimens such as 14 days of PQ, and consequently to control and elimination. Comprehensive community engagement with all relevant stakeholders, including both private and public healthcare providers, policy makers, health centers, members of the community, and patients attending the clinics, will be needed to ensure success in malaria elimination , 38.This study has a number of limitations. Although this was the first study in Nepal to assess relapse patterns in vivax malaria patients with a follow-up period of a year, supported by parasite genotyping, the number of recurrences was relatively small (n = 25), so the estimates are imprecise. We cannot exclude the possibility of transient asymptomatic recurrences, although this seems unlikely given that all the recurrences identified were symptomatic. Genotyping using 9 polymorphic loci provided satisfactory discrimination, but it does not separate heterologous relapses from new infections. Furthermore, in homologous recurrences, relapse cannot be distinguished with certainty from recrudescences , 17. UsiP. vivax parasites in Nepal. One year of follow-up is necessary to characterize relapse patterns in this and other areas where these strains are prevalent, otherwise relapse rates will be significantly underestimated.In conclusion, this randomized trial confirmed that the currently recommended radical treatment of acute vivax malaria in Nepal with CQ and a standard 14-day course of PQ is highly efficacious in preventing both short- and long-latency relapses despite possible emerging CQ resistance. Long-latency \u201cstrains\u201d comprise at least one-fifth of The Journal of Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at jiz126_suppl_Supplementary_Table_1Click here for additional data file."}
+{"text": "Plasmodium vivax and in Plasmodium ovale malaria, recurrent infection occurs from relapses which begin to emerge from the liver approximately 2 weeks after the primary illness. An important determinant of the interval from starting treatment of a symptomatic infection to the patency of these recurrent infections is the in vivo concentration-response relationship and thus the in vivo MIC. Using mechanistic knowledge of parasite asexual replication and the pharmacokinetic and pharmacodynamic properties of the antimalarial drugs, a generative statistical model was derived which relates the concentration-response relationship to time of reinfection patency. This model was used to estimate the in vivo MIC of chloroquine in the treatment of Plasmodium vivax malaria.The MIC is an essential quantitative measure of the asexual blood-stage effect of an antimalarial drug. In areas of high malaria transmission, and thus frequent individual infection, patients who are treated with slowly eliminated antimalarials become reinfected as drug concentrations decline. In the frequent relapse forms of  Careful characterization of the pharmacokinetic (PK) and pharmacodynamic (PD) properties of antimalarial drugs is necessary for the optimal design of malaria treatment regimens. Many antimalarial drugs have unusual pharmacokinetic properties, with very large apparent distribution volumes and long terminal elimination phases. The resultant protracted residual drug levels allow short-course treatments to be given ; they also provide a declining posttreatment prophylactic effect and thereby a disease-free interval during which malaria infections cannot become patent, and this affects the temporal pattern with which reinfections emerge , 2.in vivo in an individual infection is difficult, as it requires administration of inefficacious doses and serial measurement of submicroscopic parasite densities , all infections emerging from the liver following hepatic schizogony will be partially or totally suppressed; i.e., the asexual cycle (48-h) parasite reduction ratio is greater than 1, so parasite numbers decline. New infections usually emerge from the liver at a biomass of \u2248104 to 105 parasites (the progeny of between 1 and 10 successful sporozoites)  \u20138. As drer cycle , 9\u201311.tMIC plus 8 days . The precise timing depends on the terminal elimination half-life of the antimalarial drug and the slope of the concentration-response curve. If the drugs used do have liver-stage activity, then the bunching of the initial recurrent infections is delayed accordingly.In a population of individuals in whom times to recurrent infection are measured by the same method , this results in a \u201cbunching up\u201d or temporal clustering of initial recurrent infections reaching patency which are either newly acquired reinfections, relapses , or recrudescences (drug failures) , 13. If 3, and a half-maximal effect (EC50) of 250 \u03bcg/liter, with a slope coefficient of the concentration-response curve of 7. The top panels in A hypothetical example illustrates this bunching phenomenon for falciparum malaria. Suppose a drug has a terminal elimination half-life of 6 days, starting concentrations (day 0) of 1,000 \u03bcg/liter, a maximum PRR of 10n = 1,440 samples) , and 3% of the drug is cleared slowly .A randomized controlled trial comparing therapeutic options for the treatment of vivax malaria was conducted by the Shoklo Malaria Research Unit on the northwest Thailand-Myanmar border . In the samples) . The modP. vivax does not usually become patent within 4 weeks of treatment. This is a week earlier than reported in the first studies of chloroquine for the treatment of P. vivax . This corresponds to a concentration providing 50% of the maximum reduction of between 90 to 250 \u03bcg/liter and a slope coefficient of 5 to 30 , left. AP. vivax .If we make the assumption that chloroquine resistance can be quantified uniquely by a fold change in the MIC value (with no change to the slope parameter), then it is possible to predict how increasing resistance will affect the distribution of relapse intervals following chloroquine treatment. Our model predicts that a doubling of the MIC would result in the median time to relapse occurring approximately 6 days earlier . In a moPlasmodium vivax relapse was studied, but the same approach can be applied to Plasmodium falciparum infections in high-transmission settings provided that enough patients are studied. However, the following design aspects are important for the accuracy of the statistical estimation procedure. (i) There should be sufficient blood sampling to estimate the drug elimination profile. At a minimum this should be on day 7 and on the day of recurrent malaria if residual drug concentrations are still detectable. This depends on the drug studied. (ii) Follow-up should be for long enough to capture recurrent infections. A reasonable aim would be to observe 90% of the recurrences due to bunching. This depends on (a) the transmission rate  or relapse rate (P. vivax) and (b) prior estimates of the pharmacokinetic-pharmacodynamic relationship . (iii) Parasite clearance rates for the primary illness should be documented to estimate the maximum in vivo effect. (iv) Sample size estimations should be performed by simulation using a priori knowledge of the transmission rate or relapse rate, the terminal elimination half-life of the relevant antimalarial drug, and its pharmacodynamic parameters.The MIC of an antimalarial drug is a contextual parameter which is an important measure of drug efficacy and quantifies appropriate dosing. During the process of drug development, phase 2 trials specifically designed to estimate the MIC are already recommended, and their utility has been demonstrated , 5. We sThe antimalarial MIC is an individual value for a particular infection in a particular host. Ideally, in every treated patient antimalarial drug concentrations should exceed the MIC until the last blood-stage parasite has been killed. In practice, background immunity may clear infections which are not eliminated completely by the drug. But in any population background, immunity will vary widely and there will be some patients (notably young children) with little or no immunity. The MIC therefore provides a target allowing evidence-based rational design of treatment regimens . UnfortuP. vivax would not be a suitable candidate.This modeling exercise illustrates one approach to field measurement that could be used in areas of high malaria transmission . It exploits the phenomenon of bunching, whereby infections newly acquired, or in this case relapsing, during and after the period of antimalarial treatment for an incident infection (or in a mass treatment) cluster together in time as they become patent. The precision of individual MIC estimates depends on several factors, including the frequency of patient observations and sampling, the accuracy of the pharmacokinetic estimates, the terminal elimination rate constant, and the frequency of reinfection. The pharmacokinetic estimates for an individual can be derived from as little as a single blood level , with population estimates of the terminal elimination half-life obtained through weekly serial measurements. As many treatment studies involve weekly or less frequent outpatient follow-up and recurrent infections may be asymptomatic, the time to reinfection can be overestimated. If transmission is high, then partial immunity will result in many infections being cleared before they reach patency, and at lower transmission settings there will be substantial interindividual variation in the times when reinfections are acquired. Rapid reinfection  is necessary for the implementation of this methodology. For example, long-latency P. falciparum reinfection rates are high, or patients with high rates of P. vivax relapse, to provide accurate MIC estimates. Thus, the method may be more useful for postregistration dose adjustments  and for the interpretation of resistance. As the MIC reflects both host immunity and parasite susceptibility, it offers a method for assessing the former (by comparison between sites and ages) and calibrating the latter (by comparing in vivo and in vitro assessments from the same site).Although this approach could be incorporated in preregistration drug development, it is more suited to large studies of antimalarial drugs which have already been deployed. Phase 3 antimalarial drug studies evaluating asexual blood-stage activity may not enroll a sufficient number of patients in an appropriate setting, e.g., children, in whom in vivo whole-blood chloroquine MIC for Plasmodium vivax on the Thailand-Myanmar border was approximately 67 \u03bcg/liter , which may be compared with a recent in vitro geometric mean ex vivo 50% inhibitory concentration (IC50) estimate of 17.3 \u03bcg/liter . Given that whole-blood chloroquine concentrations are 8 to 10 times higher than in plasma samples, and that chloroquine is approximately 50% protein bound, these values are not very different  and so the minority of newly acquired or relapsing infections which do become patent are no different in terms of antimalarial drug susceptibility from those which emerged earlier and were cleared spontaneously. These sources of variability are compounded by variation in the duration of preerythrocytic schizont maturation, variation in the number of successful sporozoites inoculated, and variation in parasite growth rates. Nevertheless, if there are sufficient numbers of observations to observe bunching, MIC estimates can be provided from field studies in which patient follow-up is frequent and sufficiently long to capture recurrences and antimalarial blood concentrations have been measured.https://github.com/jwatowatson/bunching.All statistical analysis and modeling was done with R software (version 3.4.1). Reproducible stand-alone R code and data used for the chloroquine modeling can be found at This section presents the theoretical framework which elucidates the relationship between the temporal pattern of recurrent infections and the MIC, or time to reach MIC. The MIC is the estimated antimalarial drug concentration at which the parasite multiplication rate/cycle is one. If the parasite multiplication rate is 10 per blood-stage asexual cycle, then this would correspond to 90% inhibition of multiplication. This approach to MIC estimation relates to slowly cleared antimalarial drugs . The dynamics of the model show why the first recurrent infections to become patent after the administration of slowly eliminated antimalarial treatments cluster together temporarily and how this bunching effect is partially determined by the MIC.Many antimalarial drugs have multiphasic concentration profiles during the distribution and elimination phases. However, the terminal elimination profile is usually a first-order process characterized by a terminal elimination rate constant which is inversely proportional to the terminal elimination half-life. This varies in a population of treated patients because of population variation in pharmacokinetics.Plasmodium falciparum and P. vivax and are independent of the source of parasitemia, whether a reinfection, a recrudescence, or a relapse in the case of P. vivax.The pharmacokinetic-pharmacodynamic model defines the relationship between concentrations of the drug (as measured in whole blood or plasma) and the asexual parasite reduction ratio per 48-h cycle, i.e., the schizonticidal effect of the drug. Parameter estimation is done by discretizing into 24-h intervals. Because it is only the schizonticidal effect which is of interest, the setups of the PK-PD model are identical for both k), the drug's maximal effect Lmax , which we designate \u03c11/2. Other suitable choices would include the EC50 or the half-maximal inhibitory concentration (IC50). The choice of parameter should reflect best prior knowledge of the corresponding concentration-response curve. We argue that the EC50 or the EC90 will most often be points on the concentration-response with the most accurate a priori knowledge. These are all in vivo measures and have different values when assessed in vitro using different endpoints.In this exercise, the pharmacodynamic outcome was the parasite reduction ratio over the course of one asexual blood-stage life cycle (48 h). This is a positive real number which takes values greater than 1 when the parasite population declines and values less than 1 when the parasite population grows. Framing the problem in this way allows for population dynamics to be characterized with a single continuous outcome. We assume that the antimalarial concentration-effect relationship is a sigmoid function of the log concentration of drug in the blood or plasma. This sigmoid function is parameterized by its linearized slope coefficient .The EC50 is defined as the theoretical concentration of drug at which the parasite reduction ratio is Lmin/2 over the course of one life cycle. This is the concentration causing half the maximal growth per asexual cycle .The ICPD = {Lmax, Lmin, k, \u03c11/2}) uniquely define the pharmacodynamic properties of the drug. A commonly used mathematical formula for this sigmoid relationship is given by the logistic function. For example, the 48-h parasite reduction ratio as a function of the logarithm of the drug concentration \u03c1 can be written ask/4. Empirical data, both in vivo and in vitro, can usually give a robust characterization of the central aspect of this curve, i.e., around the \u03c11/2 and concentrations at which the effect is close to Lmax   . Howeverifficult . The MICLmin is known: this is the reciprocal of the normal parasite growth rate with negligible drug concentrations. In the estimation algorithm we set Lmin = 0.1 . This value is independent of the drug studied.In the following we assume that Lmax is known for chloroquine and is 103.We also assume that the in vivo MIC using data on intervals to recurrent infection. A Bayesian approach for the estimation of the MIC is appropriate, as there will be informative prior information regarding the key parameters in the model. For example, in vitro dose-response data can provide plausible values for the slope of the dose-response curve, although the EC50 (\u03c11/2) will in general not be the same.This section outlines the Bayesian statistical method we use for the estimation of the iT has to be greater than 8 days. This statistical model makes the following assumptions: (i) the drug lacks preerythrocytic activity and (ii) there are no recrudescences; i.e., the time to reinfection T1, \u2026,nT be the times of posttreatment malaria recurrences  for n patients, for a given antimalarial. Let i from treatment up to iT. In practice, these may either be simulated from a population PK model or extrapolated from patient measurements relevant to the terminal phase of the drug . The imputation of the drug concentrations over time is considered a separate inference problem. Thus, the logarithmic concentrations PD and the logarithmic drug concentrations iS be the (unknown) time of hepatic schizont rupture.Let iY conditional on a value of iY is computed by solvingpyrogenic is the threshold total number of blood-stage asexual parasites causing fever in an adult (the pyrogenic density), usually above 108, 104 is the number of parasites emerging from the liver for each inoculated sporozoite, and c indexes the number of parasite asexual cycles after hepatic schizont rupture. If iS occurs too early and the infection is cleared , then iY is set to 0.We define a generative model which computes the time of reinfection  PD.k = 1,..,K samples.Sample i \u2208 1..N:Sample latent variable iS until infection is cleared or total number of parasites goes above pyrogenic threshold.Simulate forward the fate of the infection (discretizing daily) as in Save time of reinfection For subjects L1 distance between observed and simulated data: Compute the Repeat for A.kL < \u03b5.Select all samples Select closest samples.We use the following approximate Bayesian computation (ABC)-type algorithm  to estimF(\u00b7) is the prior distribution over merozoite release times: the latent variables In this algorithm, \u03a0 is the prior distribution over the parameters t1/2 affect the precision of the posterior estimates of \u03c1MIC. It is easy to augment the statistical model with a hierarchical prior over values of ith patient individual elimination half-life. For example, the terminal elimination half-life of lumefantrine varies between 3 to 6 days, so one option would be a Gaussian prior with a mean of 4.5 days and standard deviation of 0.75 day , chloroquine , or chloroquine plus primaquine (0.5 mg base/kg/day for 14 days). All doses were supervised by the study staff. Informed consent was obtained from all patients. Ethical approval of the study was given by the Mahidol University Faculty of Tropical Medicine Ethics Committee (MUTM 2010-006) and the Oxford Tropical Research Ethics Committee (OXTREC 04-10). See clinicaltrials.gov trial number NCT01074905 , a reinfection from a new mosquito inoculation (termed reinfection), or activation of dormant parasites in the liver known as hypnozoites (termed relapse). In practice, most or all early recurrences are relapses.Subjects were followed daily for supervised drug treatment. Follow-up continued weekly for 8 weeks and then every 4 weeks for a total of 1 year. Sampling for chloroquine levels (capillary whole blood) was done on day 6 (\u00b11 day) and on the day of a recurrence. Recurrent episodes were detected actively at scheduled visits by microscopy (lower limit of detection is circa 20 parasites per \u03bcl). Patients were encouraged to come to the clinics between scheduled visits when unwell, so some recurrences would have been detected passively. When recurrence occurred with microscopy-confirmed For the purposes of this analysis, we used only the data from individuals randomized to the artesunate and chloroquine monotherapy arms. Times until relapse combined with estimated PK profiles for chloroquine treated patients are used to estimate the MIC of chloroquine. Times until relapse and total parasite biomass for artesunate monotherapy-treated patients were used to derive a prior distribution over liver schizont rupture times.The data used to estimate the whole-blood MIC of chloroquine consisted of 418 recurrences before day 60 occurring in 137 individuals . The number of recurrences ranged from 1 to 10, with a median number of recurrences of 2. The median time to recurrent infection in this subset of patients treated with chloroquine was 42 days. Patient ages ranged from 1.5 to 55 years (median age was 18 years).To estimate the chloroquine pharmacokinetic profiles of these patients, we pooled data from all chloroquine-treated patients, giving a total of 1,440 PK samples from 438 individuals. The median number of samples per individual was 1.P. vivax parasites depends on the latent time of merozoite release into blood. The following details how total parasite biomass is estimated for each individual and the data from the artesunate monotherapy arm are used to obtain a prior distribution of times of merozoite release.Our PK-PD model of chloroquine and 6 \u00d7 BV, where BV is the patient's blood volume, estimated as 2.78 liters/m2 for males and 2.44 liters/m2 for females (0.425 \u00d7 height0.725 \u00d7 71.84 \u00d7 10\u22124). The parasite count used is the one recorded on detection of each new episode.Total parasite biomass is estimated as follows: biomass = parasites per microliter \u00d7 10 females . Surface4 liver merozoites. The number of days since merozoite release is then estimated as5 merozoites, then the interval is 2 days shorter.Artesunate is a rapidly eliminated drug  and thus has no posttreatment prophylactic effect. Therefore, in patients randomized to artesunate monotherapy, the time (since last treatment) of relapse is indicative of when liver merozoites were released into the blood. We assume that in the absence of drug (or at negligible concentrations) the parasite multiplication rate is 10-fold per 48-h cycle. We assume that each new bloodstream infection is initiated by 10Outlier detection was done for samples taken around day 6 with very low concentrations (below 100 \u03bcg/liter). We also removed samples taken after day 60, as we consider recurrent infections only before day 60 to be informative for the MIC. Day 60 was chosen as a qualitative assessment based on the difference in the distribution to time to recurrence between artesunate-treated individuals (no postprophylactic effect) and chloroquine-treated individuals.nlme from package nlme). The concentration of chloroquine at time t is designated \u03c1(t) and is modeled asp is the proportion defining the biexponential decay and \u03b11 and \u03b12 are the two decay rates. To avoid overfitting, p was assumed to be a fixed effect. The intercept \u03b2 and the rates \u03b11 and \u03b12 were assumed to be random effects.A mixed-effects biexponential decay model was fitted to these PK samples , the estimated total parasite biomass, and the individual parameter fits from The following data and metadata on all subjects included in this analysis are available at iT (i = 1..418) is generated from a truncated Weibull distribution  where the lower bound is zero and the upper bound is the time of recurrent infection minus 6 days. The parameters of the Weibull distribution were estimated from the artesunate-treated individuals (see above).The latent merozoite release times for each reinfection time 3.The maximum PRR of chloroquine was fixed at 10PD was set as a bivariate normal distribution over the natural logarithm of the slope and the IC50, with means 1.5 and 5.5, respectively. For both parameters the standard deviation was set to 1.5  and plugged into the ABC algorithm.Total parasite biomass corresponding to patency of infection was taken as the individual estimated parasite biomass on detection of episode .We use the ABC algorithm outlined under \u201cStatistical inference\u201d above, with the following specifics.5 iterations of the algorithm. The posterior distributions of the PD parameters are shown in We ran 10"}
+{"text": "Plasmodium vivax malaria despite increasing reports of treatment failure. We did a systematic review and meta-analysis to investigate the effect of chloroquine dose and the addition of primaquine on the risk of recurrent vivax malaria across different settings.Chloroquine remains the mainstay of treatment for P vivax clinical trials published between Jan 1, 2000, and March 22, 2017. Principal investigators were invited to share individual patient data, which were pooled using standardised methods. Cox regression analyses with random effects for study site were used to investigate the roles of chloroquine dose and primaquine use on rate of recurrence between day 7 and day 42 (primary outcome). The review protocol is registered in PROSPERO, number CRD42016053310.A systematic review done in MEDLINE, Web of Science, Embase, and Cochrane Database of Systematic Reviews identified Of 134 identified chloroquine studies, 37 studies (from 17 countries) and 5240 patients were included. 2990 patients were treated with chloroquine alone, of whom 1041 (34\u00b78%) received a dose below the target 25 mg/kg. The risk of recurrence was 32\u00b74% (95% CI 29\u00b78\u201335\u00b71) by day 42. After controlling for confounders, a 5 mg/kg higher chloroquine dose reduced the rate of recurrence overall  and in children younger than 5 years . Adding primaquine reduced the risk of recurrence to 4\u00b79% (95% CI 3\u00b71\u20137\u00b77) by day 42, which is lower than with chloroquine alone .P vivax.Chloroquine is commonly under-dosed in the treatment of vivax malaria. Increasing the recommended dose to 30 mg/kg in children younger than 5 years could reduce substantially the risk of early recurrence when primaquine is not given. Radical cure with primaquine was highly effective in preventing early recurrence and may also improve blood schizontocidal efficacy against chloroquine-resistant Wellcome Trust, Australian National Health and Medical Research Council, and Bill & Melinda Gates Foundation. Plasmodium vivax for over 60 years.P vivax were published in 1989,P vivax was present in most vivax-endemic countries.8Chloroquine has been the mainstay of treatment for P vivax include optimising chloroquine regimens or changing policy to a more effective blood schizontocidal agent. In countries where high-grade chloroquine-resistant P vivax is prevalent, national treatment guidelines have been revised to a universal policy of artemisinin combination therapy (ACT) for all species of malaria.P vivax, the treatment regimen can potentially be optimised, either by increasing the dose or duration of chloroquine, or by combining chloroquine with an additional drug with blood schizontocidal activity or the ability to reverse chloroquine resistance.P vivax at doses below 25 mg/kg, higher doses are well tolerated and might provide increased efficacy.13Treatment options for chloroquine-resistant Evidence before this studyPlasmodium vivax malaria. A systematic review and meta-analysis showed that there was evidence of reduced chloroquine efficacy for P vivax present in most P vivax endemic countries. No reviews or pooled analyses had assessed the effect of chloroquine dose on the risk of recurrence.Using the search terms \u201cvivax\u201d and \u201cchloroquine\u201d, MEDLINE, Web of Science, Embase, and the Cochrane Database of Systematic Reviews were searched for articles published before Nov 29, 2017, that assessed the efficacy of chloroquine, with or without primaquine, for uncomplicated Added value of this studyP vivax clinical trials so far. Our findings highlight the substantial benefit of increasing the dose of chloroquine in children younger than 5 years and the additional benefit of adding primaquine to chloroquine.Our pooled analysis of individual patient data from 37 studies across 17 countries is, to our knowledge, the largest individual pooled analysis of Implications of all the available evidenceP vivax recurrence within 42 days in children younger than 5 years who are not given primaquine. The risk of P vivax recurrence was reduced by an even greater degree by the addition of primaquine to chloroquine in all age groups, through prevention of relapse and probably improvement in blood schizontocidal efficacy. These measures warrant consideration by regional and global policy makers to reduce the risk of early P vivax recurrence.Chloroquine is currently under-dosed in children younger than 5 years. Increasing the target dose of chloroquine from 25 mg/kg to 30 mg/kg could significantly reduce the risk of P vivax clinical trials to investigate the effect of chloroquine dose and primaquine co-administration on the risks of P vivax recurrence between day 7 and day 42.To explore alternative strategies for improving chloroquine efficacy, we did a pooled analysis of individual patient data from prospective P vivax malaria published in any language between Jan 1, 1960, and March 22, 2017, were identified using the following search terms:  AND . Further details are provided in the We searched MEDLINE, Web of Science, Embase, and Cochrane Database of Systematic Reviews, according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses statement . ProspecThe review process was done by two independent investigators (RJC and RNP), who also extracted the data. Disagreement was resolved through discussion. To ensure results were relevant to the current clinical landscape, only studies published after 2000 were included. Principal investigators were contacted and invited to share individual patient data and any additional unpublished data.P vivax monoinfection treated with chloroquine alone or chloroquine plus primaquine during the first 28 days after treatment were included. Individual patient data were uploaded into the WorldWide Antimalarial Resistance Network (WWARN) secure repository, anonymised, and processed according to a data management plan.Studies assessing patients with All data included in this analysis were obtained in accordance with ethical approvals from the country of origin. The data are fully anonymised and cannot be traced back to identifiable individuals; these do not require review from an ethics committee according to the guidelines of the Oxford Central University Research Ethics Committee.Chloroquine and primaquine doses were calculated from the number of tablets given to each patient. If these data were unavailable, doses were calculated based on the study protocol and assuming complete adherence. Individual patient records were excluded if the course of chloroquine was incomplete, the course of chloroquine or primaquine was intermittent, or if information on dose, parasitaemia, age, or weight were unavailable. Early primaquine was defined as the first dose of primaquine administered in the first 3 days of treatment .P vivax parasite rate <1\u00b75%), moderate (\u22651\u00b75% and <4\u00b70%), or high (\u22654\u00b70%) based on transmission estimates obtained from the Malaria Atlas Project and observed study site reinfection rates  when a linear association with outcome was not present. Figures of risk of recurrence were estimated according to chloroquine dose and primaquine co-administration, adjusted for other confounders and assuming no study site effect.The associations between chloroquine dose and microscopy-detectable parasite positivity in patients treated with chloroquine alone were analysed by logistic regression, with study sites included as a random effect. The association between the first day of parasite clearance and parasitaemia recurrence between day 7 and day 28 was assessed by Cox's proportional hazards regression.Heterogeneity of studies was assessed by removal of one study site at a time and calculation of the coefficient of variation around parameter estimates. Additionally, baseline characteristics of included studies were compared with targeted studies that were not included.Statistical analyses were done in Stata (version 15.0) and R (version 3.4.0), according to an a-priori statistical analysis plan.The funders of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. RNP had full access to all the data in the study and had final responsibility for the decision to submit for publication.P vivax clinical trials were identified, 134 of which included patients treated with chloroquine and were published between Jan 1, 2000, and March 22, 2017 , with 450 (8\u00b76%) aged younger than 5 years . 3773 7. The corIn the 2990 patients from 26 studies treated with chloroquine alone, the median total dose of chloroquine was 25\u00b74 mg/kg  with 1041 patients (34\u00b78%) receiving less than the WHO recommended target dose of chloroquine of 25 mg/kg . This prInformation on acute vomiting was available in eight studies (n=557), with 20 (4%) of 557 patients vomiting at least one dose of chloroquine within 60 min of administration. After adjusting for age, sex, baseline parasitaemia, presence of baseline fever, and relapse periodicity, chloroquine dose was not a significant risk factor for vomiting  of 298 recurrences within this period occurred by day 28. The cumulative risk of recurrence was 10\u00b74% (95% CI 9\u00b73\u201311\u00b76) at day 28 and 32\u00b74% (29\u00b78\u201335\u00b71) at day 42. The risks of recurrence for individual studies are presented in the After controlling for age, parasitaemia, regional relapse periodicity, and sex, a 5 mg/kg increase in chloroquine dose reduced the rate of recurrence between day 7 and day 42  of 2070 patients had cleared their detectable parasitaemia, increasing to 2095 (80\u00b79%) of 2590 on day 2 and 2369 (94\u00b78%) of 2499 on day 3. Low chloroquine dose (<25 mg/kg) was a risk factor for parasitaemia on day 1 in the univariable analysis , as were male sex, older age, and higher baseline parasitaemia . After cP vivax between day 7 and day 28, compared with 229 (9%) of 2657 who had already cleared their parasitaemia (p<0\u00b70001). After controlling for confounding factors, parasite clearance on or after day 3 was associated with an increased rate of recurrence between day 7 and day 28 compared with parasite clearance on day 1  of 139 who were parasitaemic on day 3 had recurrent In 17 studies, 1790 patients were treated with chloroquine and early primaquine. 917 (51\u00b72%) patients from 11 of these studies had a target dose of primaquine between 3\u00b75 mg/kg and less than 5\u00b70 mg/kg, and 873 (48\u00b78%) from six studies had a target dose of at least 5\u00b70 mg/kg. Overall, patients were administered a median dose of primaquine of 4\u00b77 mg/kg  at day 28 and 4\u00b79% (3\u00b71\u20137\u00b77) at day 42. When patients treated with chloroquine plus early primaquine were added to the previous Cox regression model, the addition of early primaquine was associated with a reduction in the rate of recurrent parasitaemia ; increasing the total mg/kg chloroquine dose reduced early recurrences if primaquine was not given, especially in children younger than 5 years; and, the risk of early recurrent parasitaemia was markedly reduced by co-administration of primaquine.P vivax.P vivax.P vivax relapses, with the added benefit of providing additional blood schizontocidal activity.P vivax recurrence substantially in children younger than 5 years.Increasing reports of declining chloroquine efficacy have highlighted the need for alternative treatment strategies for Previous pharmacokinetic studies have shown that chloroquine is under-dosed in children and have suggested that an increase in the chloroquine dose or dosing based on body surface area would be more appropriate and effective.Plasmodium falciparum were well tolerated in children younger than 15 years.Giardia lamblia (10 mg/kg twice daily for 5 days).Our data are in keeping with these findings and suggest that increasing the total chloroquine dose from 25 mg/kg to 30 mg/kg in children younger than 5 years would decrease the risk of early recurrence by more than 40% if chloroquine was used alone. Although increasing the target dose might reduce tolerability, substantial data support the safety of 30 mg/kg in children. In Guinea-Bissau, chloroquine doses of 50 mg/kg against drug-resistant P vivax parasitaemia: recrudescence, relapse, and new infections.Current molecular analyses cannot differentiate reliably between the three causes of recurrent The addition of primaquine to chloroquine reduced early recurrences before day 42 by 90% compared with chloroquine alone; probably in large part as a result of prevention of early relapse related to primaquine. However, addition of primaquine probably also reduces recrudescence through its blood schizontocidal activity, potentially in patients with low-grade chloroquine resistance. In the current pooled analysis, the reduction with chloroquine and primaquine did not vary before and after day 21, consistent with a reduction in both recrudescences and relapses.P falciparum malaria.P vivax.Delayed parasite clearance predicts treatment failure in Our study has several limitations. First, the analysis only included about 20% of patients from the clinical trials targeted. However, a sensitivity analysis in which one study site was removed at a time revealed no apparent bias relating to individual study sites that were included, and baseline characteristics of patients included had similar characteristics to those from all targeted studies . Second,P vivax after chloroquine monotherapy is high, it can be reduced by a modest increase in the dose of chloroquine, particularly in children younger than 5 years, and by the additional administration of primaquine. As reports of chloroquine treatment failure for P vivax increase, we recommend that the dose of chloroquine be increased to 30 mg/kg in children younger than 5 years, and health-care providers should be encouraged to provide adjunctive primaquine radical therapy to reduce the risk of both recrudescent and relapsing infections. Alternatively, a universal policy of ACT for uncomplicated malaria, with additional primaquine for vivax malaria, should be considered in regions where there is a high risk of recurrent P vivax after chloroquine treatment.In summary, although the risk of early recurrence of"}
+{"text": "Chloroquine or dihydroartemisinin-piperaquine with either 7- or 14-day primaquine regimens provided highly effective radical cure of vivax malaria on the Thailand-Myanmar border. Short course higher dose primaquine regimens for vivax malaria radical cure may improve efficacy by improving adherence. Plasmodium vivax malaria, but the optimum duration of treatment and best partner drug are uncertain. A randomized controlled trial was performed to compare the tolerability and radical curative efficacy of 7-day versus 14-day high-dose primaquine regimens  with either chloroquine or dihydroartemisinin-piperaquine.Primaquine is necessary for the radical cure of P. vivax malaria on the Thailand-Myanmar border were randomized to either chloroquine (25mg base/kg) or dihydroartemisinin-piperaquine (dihydroartemisinin 7mg/kg and piperaquine 55mg/kg) plus primaquine, either 0.5 mg/kg/day for 14 days or 1 mg/kg/day for 7 days. Adverse events within 42 days and 1-year recurrence rates were compared and their relationship with day 6 drug concentrations assessed.Patients with uncomplicated P. vivax recurrences  occurred in 80/654 (12%) patients; there was no difference between treatments. Compared to the 7-day primaquine groups the pooled relative risk of recurrence in the 14-day groups was 1.15 . Hematocrit reductions were clinically insignificant except in G6PD female heterozygotes, 2 of whom had hematocrit reductions to <23% requiring blood transfusion.Between February 2012 and July 2014, 680 patients were enrolled. Radical cure should be deployed more widely. The radical curative efficacy in vivax malaria of 7-day high-dose primaquine is similar to the standard 14-day high-dose regimen. Chloroquine and dihydroartemisinin-piperaquine are both highly effective treatments of the blood stage infection. Quantitative point of care G6PD testing would ensure safe use of the 7-day high-dose primaquine regimen in G6PD heterozygous females.NCT01640574. Plasmodium vivax is the most geographically dispersed of the human malarias, once extending to the Arctic circle and now as far north as the Korean peninsula ) was noninferior to 14-day primaquine (15.3% [10.9 to 19.5]) (een arms . The 4-mo 19.5]) . This coP. falciparum infections during the 1 year follow-up. The incidence of P. falciparum infections was not different either between the pooled dihydroartemisinin-piperaquine and chloroquine groups: 10/325 (3.1%) versus 15/329 (4.6%); P = .378, or between the pooled 7-day and 14-day primaquine groups: 8/327 (2.5%) versus 17/327 (5.2%); P = .073.There were 25/654 (4%) P. vivax recurrence within 8 weeks. No patients had \u201ctherapeutic\u201d blood chloroquine  but not primaquine were associated with a lower risk of subsequent recurrence (50 in the 36 successful in vitro tests was 17.9 ng/mL (11.5 to 27.7). Six isolates (16.7%) had an IC50 > 50 ng/mL, suggesting resistance.The majority of patients completed their primaquine treatment . There w10 ng/mL ) or pipe10 ng/mL ) concentcurrence . The geoP < .001. When pooled by primaquine treatment, more abdominal pain was reported in the 7-day 69/325 (21%) than the 14-day group 49/329 (15%):P = .032. As reported previously [P = .039. Hematocrit fell below 20% (~7 g/dL hemoglobin) in one heterozygous female in the DP7 group (requiring blood transfusion). Another heterozygote female in the DP7 group received a blood transfusion for symptomatic hemolysis. In G6PD wild-type females the mean (95% CI) absolute hematocrit change at day 6 was similar between the 7-day (\u22120.5% [\u22121.2 to 0.8]) and 14-day primaquine groups (\u22120.5% [\u22121.2 to 0.3]). The same was true for male patients: (\u22121.0% [\u22121.5 to \u22120.5]) and (\u22120.6% [\u22121.1 to \u22120.1]), respectively. The peak transcutaneous methemoglobin measurements were higher in the 7-day primaquine groups: mean (95% CI) difference 1.8% (1.7 to 1.9); P < .001 , haemolysis (n = 3), and presumed bacterial infection (n = 10). Four deaths occurred. None was considered related to the study drugs.Except for abdominal pain, there were no significant differences in reported adverse events between the 4 groups . When poeviously , hemolysP < .001 . Peri-orP < .001 . All resP. vivax infections within 8 weeks, whereas without primaquine, approximately 50% of patients experience relapse within 8 weeks of chloroquine treatment [Relapse of vivax malaria along the Thailand-Myanmar border was very effectively prevented by both the conventional 14-day high-dose regimen and the 7-day high-dose primaquine regimen. This is consistent with earlier studies with shorter follow-up periods , 25, 26.reatment , 28 in treatment . HoweverRadical cure efficacy is thought to depend mainly on the total exposure to the bioactive metabolites of primaquine. In this study, higher day 6 concentrations of the more slowly eliminated biologically inactive metabolite carboxyprimaquine were associated with lower recurrence rates . This maP. vivax is prevalent along the Thailand-Myanmar border, as evidenced by recent slowing of fever and parasite clearance, an increasing proportion of recurrences within 28 days of starting chloroquine alone and declining in vitro susceptibility [P. vivax [Low grade chloroquine resistance in tibility . DespiteP. vivax , remainsP. vivax malaria, so recurrence rates in this study could be underestimated.A limitation of this study was the high rate of premature discontinuation, which was predominantly due to migration. These patients may be at higher risk for P. vivax malaria. Radical cure needs to be deployed more widely. Quantitative G6PD testing would ensure the safe use of 7-day high-dose primaquine regimens in G6PD heterozygous females.In this region, both chloroquine and dihydroartemisinin-piperaquine combined with primaquine  provide highly effective radical cure regimens in Clinical Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at Supplementary TablesClick here for additional data file."}
+{"text": "HIV-infected individuals on antiretroviral therapy (ART) require treatment with artemisinin-based combination therapy (ACT) when infected with malaria. Artemether\u2013lumefantrine (AL) is the most commonly used ACT for treatment of falciparum malaria in Africa but there is limited evidence on the safety and efficacy of AL in HIV-infected individuals on ART, among whom drug\u2013drug interactions are expected. Day-42 adequate clinical and parasitological response (ACPR) and incidence of adverse events was assessed in HIV-infected individuals on efavirenz-based ART with uncomplicated falciparum malaria treated with AL.A prospective, open label, non-randomized, interventional clinical trial was conducted at St Paul\u2019s Hospital in northern Zambia, involving 152 patients aged 15\u201365\u00a0years with uncomplicated falciparum malaria, who were on efavirenz-based ART. They received a 3-day directly observed standard treatment of AL and were followed up until day 63. Day-42 polymerase chain reaction (PCR)-corrected ACPRs  were calculated for the intention-to-treat population.Enrolled patients had a baseline geometric mean (95% CI) parasite density of 1108 (841\u20131463) parasites/\u00b5L; 16.4% (25/152) of the participants had a recurrent malaria episode by day 42. However, PCR data was available for 17 out of the 25 patients who had malaria recurrence. Among all the 17 patients, PCR findings demonstrated malaria re-infection, making the PCR-adjusted day-42 ACPR 100% in the 144 patients who could be evaluated. Even when eight patients with missing PCR data were considered very conservatively as failures, the day-42 ACPR was over 94%. None of the participants, disease or treatment characteristics, including day-7 lumefantrine concentrations, predicted the risk of malaria recurrence by day 42. AL was well tolerated following administration. There were only two cases of grade 3 neutropaenia and one serious adverse event of lobar pneumonia, none of which was judged as probably related to intake of AL.AL was well tolerated and efficacious in treating uncomplicated falciparum malaria in HIV co-infected adults on efavirenz-based ART. However, a higher than anticipated proportion of participants experienced malaria re-infection, which highlights the need for additional malaria prevention measures in this sub-population after treatment with AL.Trial registration Pan African Clinical Trials Registry (PACTR): PACTR201311000659400. Registered on 4 October 2013. https://pactr.samrc.ac.za/Search.aspxThe online version of this article (10.1186/s12936-019-2818-7) contains supplementary material, which is available to authorized users. Malaria and human immunodeficiency virus (HIV) infections co-exist in most parts of sub-Saharan Africa . In thesFirst-line ART regimens in many parts of sub-Saharan Africa contain non-nucleoside reverse transcriptase inhibitors such as efavirenz. Efavirenz is metabolized by cytochrome (CYP) 450 enzymes, particularly CYP2B6 and to a lesser extent CYP3A4, which also metabolizes artemisinin-derivatives and lumefantrine, the longer-acting partner drug of AL  which maA prospective, open label, non-randomized, interventional clinical trial was conducted to estimate the day-42 polymerase chain reaction (PCR)-corrected, adequate clinical and parasitological response (ACPR) and incidence of adverse events in HIV+ adult patients with parasitologically confirmed, uncomplicated clinical falciparum malaria who were on efavirenz-based ART. Additionally, factors associated with malaria recurrence by day 42 of follow up, including day-7 plasma lumefantrine concentrations, were also assessed.The study was conducted from October 2014 to June 2015 at St Paul\u2019s Hospital in Nchelenge district in the northern province of Zambia, which borders the Democratic Republic of Congo. The district had an estimated population of 147,127 according to the 2010 population census . This isDuring the study period, the criteria for initiating ART in Zambia was WHO HIV disease stages 3 or 4, CD4 cell count\u2009<\u2009350, pregnancy or lactation. First-line ART regimen comprised of tenofovir/lamivudine/efavirenz. AL was the first-line treatment for uncomplicated malaria.pf malaria mono-infection with asexual malaria parasite densities\u2009<\u2009200,000/\u00b5L; ability to swallow oral medications and willingness and ability to comply with scheduled visits, supervised treatment administration, laboratory tests, and other study procedures. The following were the exclusion criteria: severe malaria as per WHO criteria \u00a0ng/mL. As shown in Fig.\u00a0In an exploratory analysis, none of the participant characteristics predicted risk of malaria recurrence neither did they predict risk of re-infection among those with available PCR results by day 42 of follow-up  of participants had predose QTcF\u2009\u2265\u2009450\u00a0ms. A change in QTcF interval of >\u200960\u00a0ms from baseline to day 2 occurred in 17.1% (26/152) of the patients (Table\u00a0The mean (SD) haemoglobin concentrations remained unchanged from baseline to day 7 (efavirenz-ART group: 11.3 to 11.2\u00a0g/dL (p\u2009=\u20090.592) but slightly increased from day 7 up to day 42, from 11.2 to 11.6\u00a0g/dL (p\u2009<\u20090.001). The median (IQR) CD4 cell count increased significantly from baseline to day 28, from 376 (248\u2013511) to 458 (324\u2013624) (p\u2009<\u20090.001).The World Health Organization recommends the use of first-line anti-malarial drugs with cure rates of >\u200995% and changing anti-malarial drugs with cure rates of less than 90% . This stA few studies have previously examined the efficacy of AL in HIV-infected non-pregnant adults on efavirenz-based ART. A Tanzanian study found that AL achieved a day-28 PCR-unadjusted cure rate of 82.5% in HIV-malaria co-infected patients on efavirenz-based ART . In thisPrevious pharmacokinetic studies have found lower lumefantrine and artemisinin concentrations in individuals on efavirenz-based ART than in ART-na\u00efve individuals \u201310. AlthThe finding of lack of evidence of an association between day-42 malaria recurrence and day-7 lumefantrine concentrations highlights that day-7 concentrations are unlikely to be predictive of malaria recurrence beyond 28\u00a0days following malaria illness. This is similar to what has been previously shown in a pooled analysis of data from HIV-uninfected individuals treated for uncomplicated falciparum malaria .In this study, AL was very well tolerated, as observed in studies involving HIV-negative individuals , 36, 37.Understanding other factors, besides lumefantrine concentrations, associated with malaria recurrent malaria infections in HIV-co-infected individuals treated with AL can inform the optimal management strategies for malaria in this sub-population. This study did not find evidence of any significant association between patient characteristics, including use of cotrimoxazole prophylaxis or malaria parasite clearance rate, and malaria recurrence by day 42 . Future studies should aim to assess this impact.This study found that AL was safe and efficacious when used to treat uncomplicated falciparum malaria among HIV-infected adults on efavirenz-based ART which provides evidence for maintaining the same AL dosing regimens used in HIV-uninfected individuals with uncomplicated malaria. Nevertheless, a higher than expected proportion of participants with malaria re-infections was observed, which warrants the use of additional malaria prevention interventions in HIV-infected adults on efavirenz-based ART.Additional file 1. Day-42 PCR-adjusted efficacy plot. PCR-adjusted ACPR by day 42 in the intention-to-treat population among malaria-HIV co-infected patients who were on efavirenz-based ART and were treated for malaria with artemether-lumefantrine. Participants with missing PCR results at day 42 (n\u2009=\u20098) are excluded from the plot.Additional file 2. Day-42 PCR-unadjusted efficacy plot. PCR-adjusted ACPR by day 42 in the intention-to-treat population among malaria-HIV-co-infected patients who were on efavirenz-based ART and were treated for malaria with artemether-lumefantrine.Additional file 3. Table on Cox regression analysis of risk of recrudescence by day 42 of follow-up."}
+{"text": "Plasmodium falciparum malaria, but its efficacy and safety has not been evaluated in HIV-infected individuals on ART, among whom drug\u2013drug interactions are expected. Day-42 adequate clinical and parasitological response (ACPR) and incidence of adverse events were assessed in HIV-infected individuals on non-nucleoside reverse transcriptase inhibitor-based ART (efavirenz and nevirapine) with uncomplicated P. falciparum malaria treated with dihydroartemisinin\u2013piperaquine.HIV-infected individuals on antiretroviral therapy (ART) require treatment with artemisinin-based combination therapy (ACT) when infected with malaria. Dihydroartemisinin\u2013piperaquine (DPQ) is recommended for treatment of P. falciparum malaria who were on efavirenz-based or nevirapine-based ART. They received a directly-observed 3-day standard treatment of DPQ and were followed up until day 63 for malaria infection and adverse events. Day-42 PCR-corrected-ACPRs  were calculated for the intention-to-treat (ITT) population.An open label single arm clinical trial was conducted in Malawi (Blantyre and Chikhwawa districts) and Mozambique (Manhi\u00e7a district) involving patients aged 15\u201365\u00a0years with uncomplicated The study enrolled 160 and 61 patients on efavirenz and nevirapine-based ART, with a baseline geometric mean (95% CI) parasite density of 2681 (1964\u20133661) and 9819  parasites/\u00b5L, respectively. The day-42 PCR-corrected ACPR (95% CI) was 99.4% (95.6\u201399.9%) in the efavirenz group and 100% in the nevirapine group. Serious adverse events occurred in 5.0% (8/160) and 3.3% (2/61) of the participants in the efavirenz and nevirapine group, respectively, but none were definitively attributable to DPQ. Cases of prolonged QT interval (>\u200960\u00a0ms from baseline) occurred in 31.2% (48/154) and 13.3% (8/60) of the patients on the efavirenz and nevirapine ART groups, respectively. These were not clinically significant and resolved spontaneously over time. As this study was not designed to compare the efficacy and safety of DPQ in the two ART groups, no formal statistical comparisons were made between the two ART groups.DPQ was highly efficacious and safe for the treatment of malaria in HIV-infected patients concurrently taking efavirenz- or nevirapine-based ART, despite known pharmacokinetic interactions between dihydroartemisinin\u2013piperaquine and efavirenz- or nevirapine-based ART regimens.Trial registration Pan African Clinical Trials Registry (PACTR): PACTR201311000659400. Registered on 4 October 2013, https://pactr.samrc.ac.za/Search.aspxThe online version of this article (10.1186/s12936-019-2909-5) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria infection than the HIV-uninfected population  to examine whether it exceeds 90%, the WHO recommended benchmark for an efficacious anti-malarial drugs [In view of the limited data on the efficacy and safety of DPQ in HIV-malaria co-infected patients, a single arm clinical trial was conducted to estimate the efficacy of DPQ when used to treat parasitologically-confirmed uncomplicated clinical This study was part of a multi-country single arm clinical trial aimed at assessing the efficacy and safety of two artemisinin-based combinations (DPQ and artemether\u2013lumefantrine) when used to treat malaria in HIV-infected adults on standard ART. One of the single arm trial assessing the efficacy and safety of AL was conducted in Zambia and findings have been reported elsewhere . In thisDuring the study period, the criteria for initiating ART in the two countries were WHO HIV disease stages 3 or 4, CD4 cell count\u2009<\u2009350, pregnancy or lactation . In OctoP. falciparum malaria monoinfection with asexual malaria parasite densities\u2009<\u2009200,000/\u00b5L; ability to swallow oral medications and willingness and ability to comply with scheduled visits, supervised treatment administration, laboratory tests, and other study procedures. The following were the exclusion criteria: severe malaria as per WHO criteria [Plasmodium species; haemoglobin (Hb) concentration\u2009<\u20097\u00a0g/dL; severe sickle cell disease or sickle-haemoglobin C anaemia; evidence of pregnancy or lactation; use of any anti-malarial treatment or drug with anti-malarial activity within the past 1\u00a0month, except cotrimoxazole; history of DPQ hypersensitivity reactions; gastrointestinal diseases that could alter gut absorption or motility; history of splenectomy; history of epilepsy or convulsions; pre-existing clinically-significant cardiac, liver, renal, neurological or psychiatric abnormalities; alternative clinical cause of fever other than malaria and participation in any investigational drug study in the past 30\u00a0days.This was a single arm clinical trial (Registration number: PACTR201311000659400). HIV-infected patients on nevirapine- or efavirenz-based ART suspected of having malaria were pre-screened through history taking and clinical examination to determine their eligibility for the study. The study inclusion criteria were as follows: age\u2009\u2265\u200915 to\u2009\u2264\u200965\u00a0years; weight\u2009\u2265\u200935\u00a0kg; documented fever (axillary\u2009\u2265\u200937.5\u00a0\u00b0C) or history of fever 24\u00a0h prior enrolment; smear positive criteria ; mixed iFinger-prick blood samples were taken from those who satisfied the preliminary eligibility criteria and tested for malaria using Rapid Diagnostic Test (RDT)  and for haemoglobin concentration using Hemocue Haemoglobinometer. Thick blood smear microscopy examinations were performed on patients with RDT positive malaria while clinical examinations were performed in those with confirmed malaria parasitaemia. Consenting participants were enrolled and scheduled for a 3-day hospital admission.\u00ae, Sigma Tau): 3 tablets for study participants\u2009<\u200960\u00a0kg or 4 tablets study for participants\u2009\u2265\u200960\u00a0kg. Each tablet contained dihydroartemisinin/piperaquine 40\u00a0mg/320\u00a0mg, respectively, administered at 0\u00a0h, 24 (+\u00a04) and 48 (+\u00a04)\u00a0h after the first dose. Participants\u2019 vital signs were measured at 6-hourly intervals and adverse events were monitored. A 12-Lead electrocardiogram (ECG) was performed before the first dose of DPQ and within 2\u00a0h after administration of the third dose DPQ. Any patient with Fridericia-corrected QT (QTcF) interval of\u2009\u2265\u2009450\u00a0ms or QTc increase of\u2009>\u200960\u00a0ms from the baseline underwent follow-up ECGs until resolution of the abnormality. Participants were discharged at least 24\u00a0h after taking the third (last) dose of DPQ (post-treatment day 3) and advised to come for follow up visits on post-treatment days 7 (\u00b1\u20091), 14 (\u00b1\u20091), 21 (\u00b1\u20092), 28 (\u00b1\u20092), 35 (\u00b1\u20092), 42 (\u00b1\u20092) and 63 (\u00b1\u20092). Participants were encouraged to return to the health facility any time they felt unwell (unscheduled visits). All adverse events were graded using the DAIDS criteria [The participants received dihydroartemisinin\u2013piperaquine (Eurartesimcriteria . Adverse\u00ae) at baseline and during recurrent malaria episodes. Parasite DNA was extracted from the DBS samples, amplified using polymerase chain reaction (PCR) and genotyped for merozoite specific protein (MSP) 1 and 2 to distinguish malaria recrudescence from re-infection, using methods previously described [During the admission period, thick blood slides were collected pre-dosing and at 6-hourly intervals until after obtaining two consecutive malaria negative smears. The slides were also collected at scheduled and unscheduled follow-up visits. The slides were Giemsa-stained and read by an experienced microscopist using standard protocols . For quaescribed . Samples\u00ae Chemistry analyzer, on days 0, 3, 7, 28, 42 and 63 for haematological tests using a Beckman Coulter\u00ae HMX Analyzer and on days 0, 28, and 63 for CD4 cell count measurement using a BD FACSCount\u2122 machine. Plasma samples collected on days 0, 28, and 63 were stored for future HIV viral load assays.Venous blood samples were collected on days 0, 3, 28, 42 and 63 for biochemistry tests using a Beckman CX5Blood samples for sparse pharmacokinetic (PK) assays were collected in sampling windows of 0\u20136, 6\u201348, 48\u201360\u00a0h, and on days 7, 21, 28 or 35 from first dose, as previously recommended . The PK P. falciparum malaria PCR markers (merozoite surface protein 1 and 2) with those at baseline, irrespective of axillary temperature, and who had not previously met any of the criteria of early treatment failure (ETF), late clinical failure (LCF) or late parasitological failure (LPF). Standard WHO definitions of ETF and LCF were used [The primary study endpoint was proportion of patients with PCR-corrected day 42 ACPR, defined as patients who did not have parasitaemia on day 42 that exhibited identical ere used .50) and 90% (PC90), fever clearance time and trends in haemoglobin concentrations and CD4 cell counts from baseline to day 28.The other primary study end points were grade 3 or 4 TEAEs of special interest  and serious adverse events (SAEs) as per standard definitions . Local sSample size calculation was based on estimates of total treatment failure rate (TTFR). The estimated day-42 PCR corrected TTFR was\u2009\u2264\u200910% . A preciFor the primary end-point, three analysis populations were used. Firstly, the Intention-to-treat (ITT) population included patients who received at least 1 dose of study medication. Secondly, the per-protocol (PP) population included all participants who had the primary endpoint data at day 42, received a full course of DPQ and adhered to the follow-up visit schedule. Thirdly, the safety population included all patients who received any amount of study medication and had at least 1 assessment after dosing. ACPR plus 95% CI in PP and ITT populations were calculated. Sensitivity analyses were performed using the ITT and PP populations that first considered all participants with missing data as having parasitological failure and then considered the same participants as having treatment success.50, PC90 and parasite clearance half-life) in the two ART groups were estimated using the WorldWide Antimalarial Resistance Network parasite clearance estimator, as described elsewhere [Statistical analyses for secondary ACPR endpoints were similar to the primary endpoints. In addition, the Kaplan\u2013Meier survival plots were used to summarize the time to PCR-corrected and uncorrected treatment failure. Parameters assessing post-treatment parasite clearance  on efavirenz-ART were lost-to-follow-up or withdrew consent.A total of 1864 patients presenting at the health facilities with symptoms suggestive of malaria were screened for trial eligibility  dosages of dihydroartemisinin and piperaquine administered to participants were 7.0\u00a0mg/kg (5.1\u20139.9) and 56.3\u00a0mg/kg (40.9\u201379.1), respectively in the efavirenz-ART group and 7.0\u00a0mg/kg (5.6\u20138.6) and 55.7\u00a0mg/kg (45.0\u201368.6), respectively in the nevirapine-ART group. The dosages were well tolerated: 3 participants in the efavirenz-ART group and none in the nevirapine-ART group vomited following intake of DPQ. Participants who vomited were re-dosed and none was withdrawn from the study due to persistent vomiting.As shown in Table\u00a0In the ITT analyses, the PCR-corrected day 42 ACPR was 99.4%  in the efavirenz-ART group and 100% in the nevirapine-ART group  in the efavirenz-ART group and 100% in the nevirapine-ART group. The day 42 PCR-uncorrected ACPR was 94.7% (95% CI 89.7\u201397.3%) in the efavirenz-ART group and 100% in the nevirapine-ART group.In sensitivity analyses which considered missed visits or samples, unavailable PCR results and loss to follow up as treatment failures, the day-42 PCR-corrected ACPR was 93.1% (95% CI 88.0\u201396.2%) in the ITT population and 98.8% (95% CI 95.1\u201399.7%) in the PP population. Details of the different scenarios accounting for the missing results are shown in Table\u00a050 and the median (range) PC90 in the ITT population were 3.1 (0.2\u201310.3)\u00a0h and 8.2 (2.4\u201319.7)\u00a0h, respectively, in the nevirapine-group and 4.2 (0.6\u201340.3)\u00a0h and 10.1 (3.2\u201363.1)\u00a0h, respectively, in the efavirenz-group. The median parasite clearance half-life (range) were 2.1 (1.1\u20136.8) and 2.2 (1.2\u20139.8)\u00a0h in the nevirapine- and efavirenz-ART groups, respectively  PCre 3.1 0.\u201310.3\u00a0h aOn day 7 of follow-up, blood samples for plasma piperaquine quantification were available from 179 of the retained 216 participants. Of these, 62.0% (111/179) had day-7 piperaquine concentrations that were below the LLOQ (<\u200925\u00a0ng/mL): 60.0% (81/135) in the efavirenz-ART group and 68.2% (30/44) in the nevirapine-ART group. The piperaquine values for these individuals were imputed to half the lower limit of quantification. However, since these values were more than\u2009>\u200910% of the data, they were excluded in calculation of median piperaquine exposure. Overall, the median (range) day-7 piperaquine concentration in participants with concentrations\u2009>\u2009LLOQ was 57 (25.5\u2013592.8) ng/mL in the efavirenz-ART group and 79.1 (25.4\u2013211.1) ng/mL in the nevirapine-ART group. After excluding\u2009<\u2009LLOQ values, participants who experienced malaria recurrence by day 42 (n\u2009=\u20093) had a median (range) day-7 piperaquine concentration of 63.9 (49.7\u201378.9) ng/mL and this value was 66.7 (25.4\u2013592.8) ng/mL in participants who did not experience malaria recurrence .At baseline, 58.1% (93/160) of participants in the efavirenz-ART group and 31.1%, (19/61) in the nevirapine-ART group were febrile (axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C). The median fever clearance time (IQR) was 6\u00a0h (6\u201360) in the efavirenz-ART group and 6\u00a0h (6\u201312) in the nevirapine-ART group. By treatment day 2, only 3.8% (6/160) participants in the efavirenz-ART group and 1.6% (1/61) in the nevirapine-ART group were febrile.From enrolment to follow-up day 63, there were 69 adverse events of special interest (AESIs)  and 12 serious adverse events (SAEs)  (Table\u00a0The mean (SD) haemoglobin concentration decreased from baseline to day 7 , but increased thereafter up to day 42, from 11.7 to 12.4\u00a0g/dL in the efavirenz-ART group (p\u2009<\u20090.001) and from 10.7 to 11.6\u00a0g/dL in the nevirapine-ART group (p\u2009<\u20090.001). Following DPQ treatment, the median (IQR) CD4 cell count increased from baseline to day 28, from 257 (140\u2013357) to 320 (216\u2013521) in the efavirenz-ART group (p\u2009<\u20090.001) and from 390 (204\u2013500) to 429 (204\u2013580) in the nevirapine-ART group (p\u2009=\u20090.133).Data on baseline QTcF interval were available for 218 participants; 158 and 60 in the efavirenz-ART and nevirapine-ART group, respectively. The proportions of participants with predose QTcF\u2009\u2265\u2009450\u00a0ms were 3.8% (6/158) in the efavirenz-ART group and 11.7% (7/60) in the nevirapine-ART group. Only 4 participants in the efavirenz-ART group had missing QTcF values on day 2 (last day of treatment). A change in QTcF interval of\u2009>\u200960\u00a0ms from baseline to day 2 occurred in 31.2% (48/154) and 13.3% (8/60) in the efavirenz and nevirapine groups, respectively  has been shown to be associated with resistance of the parasites to artemisinins . AlthougTreatment-emergent AESIs occurred in nearly a third and one-quarter of the participants on efavirenz- and nevirapine-based ART, respectively. However, only a few cases of AESIs in the efavirenz-group and none in the nevirapine-group were judged to be possibly associated with DPQ Table\u00a0. QTcF prThe major strengths of this study were the directly-supervised DPQ dosing, large sample size of participants on efavirenz-based ART and minimal loss to follow-up (<\u20095%). However, the required sample size was not achieved in the nevirapine-based ART group because the national ART programme phased out this regimen during the course of the study. Nevertheless, participants in this group had high baseline parasite densities which permitted accurate assessment of DPQ\u2019s parasite clearance and prophylactic efficacy.3 and nearly two-thirds were on cotrimoxazole prophylaxis which has some antimalarial effects [3 which is likely to have compromised their ability to clear parasites and prevent reinfections [Nearly one-half of enrolled study participants in the efavirenz group had parasite densities of\u2009<\u20092000/mm effects , 46. Thufections . In spitIn this study, the day-42 PCR corrected and uncorrected ACPR appeared to be higher among individuals on nevirapine-based ART than among those on efavirenz-based ART. However, this study was not designed to compare the efficacy and safety of DPQ between the two groups and the full sample size was not achieved in the nevirapine arm. It is therefore inappropriate to make direct comparison of DPQ efficacy between individuals taking the two ART regimens.P. falciparum malaria in individuals taking efavirenz or nevirapine-based ART. A higher than expected observed cases of QTc interval prolongation (>\u200960\u00a0ms from baseline to day 2) following treatment with dihydroartemisinin\u2013piperaquine were observed and thought to likely be due to resolution of fever as part of the malaria recovery process. Under the \u201cHIV Test and Treat\u201d approach, many HIV-infected individuals in high burden countries will initiate ART early before they are severely immunosuppressed and this study supports the use of dihydroartemisinin\u2013piperaquine for the treatment of uncomplicated malaria in such patients.This study found that dihydroartemisinin\u2013piperaquine was highly efficacious and safe when used to treat uncomplicated Additional file 1. Day 42 PCR-corrected efficacy plot. Kaplan\u2013Meier survival plot of participants who were treated with dihydroartemisinin\u2013piperaquine (DHA PPQ) in the efavirenz (EFV)- and nevirapine (NVP) based antiretroviral therapy (ART) groups according to polymerase chain reaction (PCR) corrected adequate clinical and parasitological response (ACPR) by day 42 in the intention-to-treat population with loss to follow up and indeterminate or unavailable PCR samples treated as treatment success.Additional file 2. Day 42 PCR-uncorrected efficacy plot. Kaplan\u2013Meier survival plot of participants who were treated with dihydroartemisinin\u2013piperaquine (DHA PPQ) in the efavirenz (EFV)- and nevirapine (NVP) based antiretroviral therapy (ART) groups according to polymerase chain reaction (PCR) uncorrected adequate clinical and parasitological response (ACPR) by day 42 in the intention-to-treat population with loss to follow up and indeterminate or unavailable PCR samples treated as treatment success.Additional file 3. Details of serious adverse events that occurred during follow up."}
+{"text": "Plasmodium falciparum infections. In Africa, artesunate\u2013mefloquine (ASMQ) is an infrequently used artemisinin-based combination therapy (ACT) because of perceived poor tolerance to mefloquine. However, the WHO has recommended reconsideration of the use of ASMQ in Africa. In this large clinical study, the pharmacokinetics (PK) of a fixed dose combination of ASMQ was investigated in an African paediatric population to support dosing recommendations used in Southeast Asia and South America.The World Health Organization (WHO) recommends combinations of an artemisinin derivative plus an anti-malarial drug of longer half-life as treatment options for uncomplicated \u00ae). The doses were one or two tablets containing 25/55\u00a0mg AS/MQ administered once a day for 3\u00a0days according to patients\u2019 age. A sensitive LC\u2013MS/MS method was used to quantify AS, DHA and MQ concentrations in plasma. An attempt was made to investigate the relationship between the absence/presence of malaria recrudescence and MQ area under the curve (AUC) using logistic regression.Among the 472 paediatric patients aged 6\u201359\u00a0months from six African centres included in the large clinical trial, a subset of 50 Kenyan children underwent intensive sampling to develop AS, its metabolite dihydroartemisinin (DHA) and MQ PK models. The final MQ PK model was validated using sparse data collected in the remaining participants  dosing recommendation. Besides body weight considerations, there is no indication that the dosage should be modified in children with malaria compared to adults.Trial registration Pan African Clinical Trials Registry PACTR201202000278282 registration date 2011/02/16The online version of this article (10.1186/s12936-019-2754-6) contains supplementary material, which is available to authorized users. Plasmodium falciparum infection, the predominant cause of malaria in Africa, recommended by the WHO since 2001 [The World Health Organization (WHO) estimates a significant 18% reduction in the incidence of malaria along with a considerable 28% decrease in the malaria mortality rate between 2010 and 2017 [nce 2001 . This conce 2001 , as wellnce 2001 , 6. Howence 2001 .initiative (DNDi) with the fixed-dose artesunate-based combination therapy (FACT) Consortium [i . Because MQ dose splitting into three equal daily doses has been shown to optimize treatment compliance and to improve MQ tolerability [The development of a fixed-dose combination (FDC) of AS and MQ was begun in 2002 by the Drugs for Neglected Diseases nsortium . This consortium \u20139, but trability , 11, FDCP. falciparum, as confirmed by microscopy , and with fever equal to or higher than 37.5\u00a0\u00b0C. Exclusion criteria were children with body weight less than 5\u00a0kg, signs of severe/complicated malaria, febrile conditions caused by diseases other than malaria, a known hypersensitivity to the study drugs, a mixed plasmodium infection, a history of anti-malarial treatment in the 2\u00a0weeks preceding the trial or 4\u00a0weeks in case of mefloquine and piperaquine, prior participation in a therapeutic trial within 3\u00a0months or inability to tolerate oral medication. Patients were followed up to day 63 after start of treatment or to the first recurrence of infection. The study protocol was reviewed and approved by national and independent ethics committees of all participating centres.The clinical trial was carried out in six African centres: three in Tanzania, two in Burkina Faso and one in Kenya. Written informed consent from a parent/guardian was required to enrol children younger than 5\u00a0years in the trial, who were infected by Of the 945 patients enrolled in the trial, 473 were randomized to the ASMQ arm (one of them was never dosed) and 472 were randomized to the AMLF arm. The pharmacokinetic analysis described here was performed on the 472 patients who received ASMQ.Administered doses for these patients were one or two dispersible tablets containing 25\u00a0mg AS and 55\u00a0mg MQ once a day for three consecutive days to children aged from 6 to 11\u00a0months and from 12 to 59\u00a0months, respectively. Clinical and parasitological examinations were scheduled at baseline, i.e. before drug administration, at day 0 (D0), D1, D2, D3, D7, D14, D21, D28, D35, D42, D49, D56 and D63 and on any other day if the patient spontaneously returned and parasitological reassessment was required (as per protocol). A margin of\u2009\u00b1\u20092\u00a0days to the assigned day of visit was allowed from D7 onward. In case of recurrence of parasitaemia on D7, D14, D21, D28, D35, D42, D49, and D56 the date was recorded and the type of recurrence was determined by PCR .According to the study protocol, the first fifty children from Kenya enrolled in the ASMQ arm underwent intensive blood sampling: at baseline, on D0 after drug administration (until 6\u00a0h after first dosing), D2 (until 6\u00a0h after the third dose), D3 (72\u00a0h after first dose), D7 and on one other occasion on day 28, 35, 42, 49, 56 or 63. Two blood samples, at baseline and on D7, were collected for all the other participants. Additionally, for all patients with recurrence of parasitaemia, a blood sample was taken on the day of failure.The mass spectrometry assay for AS, DHA and MQ used for the analysis of study samples is an adaptation of a previously published validated multiplex method . The assThe mobile phase was delivered at a flow rate of 0.3\u00a0mL/min on a 2.1\u00a0mm\u2009\u00d7\u200975\u00a0mm XSelect HSS 3.5\u00a0\u03bcm column , using solvent A (2\u00a0mM ammonium acetate\u2009+\u20090.1% FA) and solvent B (MeCN\u2009+\u20090.1% FA) distributed according to the following stepwise gradient program: 98% A: 0\u00a0min; 98% A\u2009\u2192\u200915% A: from 0.0\u00a0min\u2009\u2192\u200913.0\u00a0min followed by a re-equilibration step to the initial solvent proportions. The retention time of mefloquine/mefloquine-d9, DHA/DHA-13Cd4 and artesunate is 7.4\u00a0min, 8.2\u00a0min and 9.2\u00a0min, respectively. The chromatographic system was coupled to a triple stage quadrupole (TSQ) Quantum Ion mass spectrometer (MS) from Thermo Fischer Scientific  equipped with an Ion Max electrospray ionization (ESI) interface. The limits of quantification (LOQ) of the method are 2.5\u00a0ng/mL for MQ and 2\u00a0ng/mL for AS and DHA.Plasma samples were isolated by centrifugation and stored at \u2212\u00a020\u00a0\u00b0C until batch analysis. Briefly, 100 \u03bcL of plasma sample were mixed with 50 \u00b5L internal standard  and extracted with 600 \u00b5L of acetonitrile. The supernatant (700 \u00b5L) was evaporated under nitrogen at room temperature and was reconstituted in 150 \u00b5L of MeOH/ammonium acetate 2\u00a0mM (1:1) adjusted with formic acid at 0.1%, vortex-mixed and centrifuged again. The samples were maintained at +5\u00a0\u00b0C in autosampler racks throughout the analytical series. The injection volume was 20 \u03bcL.http://www.wwarn.org/toolkit/qaqc) organized by the World Wide Antimalarial Resistance Network WWARN (http://www.wwarn.org/).The method is precise (with mean inter-day CV\u00a0%\u2009<\u200910%), and accurate . Since its initiation, the laboratory has participated in the Pharmacology Proficiency Testing Programme for anti-malarial drugs  [\u00ae (PsN) toolkit (version 3.7.6) [http://www.r-project.org/).Non-linear mixed effects modelling program (NONMEMion 7.3)  with then 3.7.6)  was useda) estimation could not be made because of the small number of samples collected right after dose intake (one sample at maximum for each enrolled child on the first and third treatment day). Ka was thus fixed to 3.2\u00a0h\u22121, the mean of previously published estimates retrieved from papers using a first-order process to depict AS absorption [A stepwise modelling approach was undertaken to identify models that best described the MQ and AS/DHA pharmacokinetics. Multi-compartment dispositions with first-order absorption and elimination processes were compared for MQ. Due to the restricted amount of AS and DHA data, drug and metabolite pharmacokinetics were modelled simultaneously and directly described by means of a one compartment model with linear absorption and elimination. Moreover, since AS is rapidly and almost completely hydrolysed in DHA, its elimination was assumed to occur exclusively via irreversible conversion to DHA , 17. An sorption .M for metabolite), inter-compartmental clearance (Q), central (VC for drugs and VM for metabolite) and peripheral (VP) volumes of distribution and Ka. The metabolic conversion rate from AS to DHA was estimated by CL/Vc as previously discussed. AS and MQ relative bioavailability  were also tested for AS/DHA and MQ to account for dose variation with respect to the nominal value due to the administration of water dispersible tablets. Since the ASMQ combination is administered orally, the pharmacokinetic parameter estimates represent apparent values.Parameterization was performed in terms of clearances  variability. Finally, the correlation between AS and DHA concentration measurements was tested using the L2 function in NONMEMAvailable covariates were: body weight (BW), height/length, age, sex, creatinine, total bilirubin (BIL), aspartate (AST) and alanine (ALT) aminotransferases, haemoglobin (Hb), haematocrit (Ht), total parasitaemia and co-medications categorized as CYP3A4 inducers. Visual inspection of the correlation between post hoc individual estimates of the pharmacokinetic parameters and the available patients\u2019 characteristics was initially conducted to identify potential physiologically plausible relationships. Creatinine clearance was not evaluated since MQ elimination occurs mainly through non-renal processes and AS is completely converted into DHA, which is eliminated via glucuronidation . A stepw50 the AGE at 50% of maturation, MATmag, the maturation magnitude for age, and Kmat the age maturation rate constant [The influence of body weight on all MQ and DHA pharmacokinetic parameters (PAR) was tested using allometric scaling:ribution . A lineaconstant , 21. Thea and this was tested by integrating the effect of the treatment day (0 vs. 1 and 2) on Ka.The acute phase of malaria is associated with altered gastrointestinal motility and an increased likelihood of vomiting. In the three-daily dose ASMQ regimen, the second dose is administered when the patient is in an improved state of health, thanks to the first dose of AS, that kills most of the parasites . The pot1/2), maximum concentration (Cmax), and time to achieve Cmax (tmax) for all the three drugs, MQ area under the curve to infinite (AUC0\u2013inf) and AS and DHA AUC0\u201324 after the first and the third ASMQ intake were computed using final pharmacokinetic parameter estimates and classic pharmacokinetic equations or NONMEM integration, as appropriate.Terminal half-lives  method with interaction. AS and DHA non-zero concentrations measured more than a week after last drug intake were thought unreliable and thus omitted from the analysis. Other missing variables  were also omitted. Data below the quantification limit (BQL) of the assays were handled by setting the first of a series of BQL samples at LOQ/2 and as missing all the others (M6 method) .\u00ae objective function value (\u0394OFV), were employed to discriminate between nested models. Since a \u0394OFV between any two hierarchical models approximates a \u03c72 distribution, a change of more than 3.84 (p\u2009<\u20090.05) and 6.63 (p\u2009<\u20090.01) points was considered statistically significant for one additional parameter in model-building or forward insertion and backward-deletion covariate steps, respectively. Akaike\u2019s information criterion (AIC) was used for non-hierarchical models. Shrinkage was also evaluated. Sensitivity analyses removing outlying data with absolute conditional weighted residuals (CWRES) greater than 4 or potentially unreliable covariate values and concentration measurements were finally performed to avoid any potential bias in parameter estimation and covariate exploration.Diagnostic goodness-of-fit plots, along with differences in the NONMEM95%) were derived from 2000 replicates of the initial datasets and compared with the original estimates. Prediction-corrected visual predictive checks (pcVPC) were also performed using the PsN-Toolkit and the R package Xpose4 by simulations based on the final pharmacokinetic models with variability using 1000 children [The stability of the final MQ and AS/DHA models was assessed by means of the bootstrap method implemented in PsN-Toolkit . Median children , 24. Morchildren .90\u00a0%) of children and adult concentration\u2013time profiles were obtained through simulations (n\u2009=\u20091000) using the final pharmacokinetic model described above and published MQ pharmacokinetic models including BSV and intra-individual variability, respectively. A literature search allowed the identification of two pharmacokinetic models developed in adults receiving the same fixed dose formulation of ASMQ as the one administered to the children enrolled in this clinical trial [\u00ae, fixing simulated individuals\u2019 body weight to the corresponding median population value. Administered MQ doses were 110\u00a0mg and 400\u00a0mg over three consecutive days for children and adults, respectively. MQ drug exposure was quantified by computing median and PI95% AUC over the whole study period (AUC0\u2013day63) by NONMEM integration for all the simulated population/model.Median and 90% prediction interval  on study days 7, 28, 42, and 63 were calculated by concentration integration in NONMEM\u00ae. The relationship between recrudescence of infection  and model predicted AUC0\u2013dayx (independent variable) on study days 7, 28, 42, and 63 was inspected by means of logistic regression using STATA . The independent variable was log-transformed (using base 2) and cantered on its median value. The level of significance was set at 0.05.This exploratory analysis was carried out on MQ data collected from all children participating in the trial with complete dosing history information that did not drop out in the early days of the study. Model predicted MQ cumulative AUC  concentrations were available for the 48 Kenyan patients selected for the pharmacokinetic model development. Of note, none of the MQ concentrations were quantified as a BQL, while 71% and 57% of AS and DHA samples were BQL data. Median (range) treatment duration per study subject was 3\u00a0days (1\u20133) and the number of available non-BQL samples was 5 (1\u20137) for MQ, 1 (1\u20132) for AS and 2 (1\u20133) for DHA. MQ concentrations ranged between 0.17\u00a0ng/mL and 6552.51\u00a0ng/mL, AS (>\u2009BQL) between 2.1 and 8469.8\u00a0ng/mL and DHA (>\u2009BQL) between 2.9 and 2400.9\u00a0ng/mL.C did not improve data description  whilst assignment of BSV to CLM  and to VM  yielded a better fit of the data. Inclusion of relative F1 (fixed to 100% with estimated BSV) explained all the BSV on AS and DHA clearance and significant decreased the OFV . The estimates and variability (CV%) of the pharmacokinetic parameters obtained by the base population model were a relative F1 of 100% (67%), a CL of 180 L/h, a VC of 166 L, a CLM of 12.5 L/h and a VM of 13.8 L (57%).As previously described, a two-compartment model was used to simultaneously fit AS and DHA data with first-order absorption, drug exclusive elimination via irreversible conversion to DHA and first-order elimination of metabolite. Initially, BSV was assigned only on CL and a mixed error model was assumed for the intra-patient variability of both drug and metabolite. Model stability was achieved by integrating a correlation between AS and DHA concentration measurements . BSV on VM and VM markedly decreased the objective function . Maturation on CLM was adequately described using Eq.\u00a0M was significantly impacted by sex . Complete multivariate analyses allowed for the effect of sex on VM and F1 to be discarded, as well as that of maturation on DHA clearance. These results show that F1 is reduced by 68% upon doubling child age with respect to the population median (2.6\u00a0years), and is 29% higher in the first day of therapy than in the subsequent treatment days. The effect of BW on CLM and VM was also retained.Age, sex and BIL as well as the hepatic liver tests ALT and AST had a significant impact on F1 . Because of poor effect estimation , BIL was not kept for further covariate analyses. Sensitivity analyses revealed that the effect of ALT and AST on F1 were purely due to a single patient having the highest values for both hepatic enzyme tests. Whether this finding was a true or an incidental effect could not be validated and these covariates were thus not retained in the model. F1 was found to increase with the parasite counts , and to be higher at day 0 compared to days 1 and 2 of treatment . As shown in Table\u00a095% and differed less than 15% from their bootstrap estimations. Prediction corrected VPCs shown in fig.\u00a0The final model parameter estimates, together with their bootstrap estimations, are shown in Table\u00a0C  in addition to CL yielded a better fit of the data, which was further enhanced by inclusion of BSV on Ka . No improvement of the model fit was observed associating BSV on Q or VP . The inclusion of MQ F1 fixed to 100% with an estimated BSV significantly decreased the OFV whilst explaining all the BSV associated to VC . Finally, a proportional model was retained to describe the intra-patient variability. The estimates and variability (CV\u00a0%) of the pharmacokinetic parameters obtained by the base population model were an F1 of 100% (39%), a CL of 0.48 L/h (40%), a VC of 88 L, a Q of 0.41 L/h, a VP of 69 L, and a Ka of 0.15\u00a0h\u22121 (87%).A two-compartment model with first-order absorption and elimination described MQ data better than a one-compartment model . No additional benefit was observed using three compartments . BSV on Va . A decrease of 74% in Ka was observed while doubling the age with respect to the population median (2.6\u00a0year) and female children had 55% lower Ka than male children. Multivariate analysis showed that age accounted for the effect of sex on Ka and allowed for the discarding of the impact of age on F1. Finally, significantly different Ka at day 0 and 1/2 of ASMQ treatment were identified due to improvement in patient health following the first intake of AS . Multivariate and backward deletion step analyses performed using the reduced dataset, obtained by removal of the outlying patient, showed that the BW effect on clearances and volumes of distribution, as well as age and treatment day effect on Ka, should be retained in the final MQ pharmacokinetic model.The univariate analyses showed no association between the covariates tested and MQ bioavailability, clearances and volumes of distribution . However, the sensitivity analysis performed while removing the patient with extremely low concentrations after the second and third ASMQ dose revealed that this outlier masked the real impact of BW on clearances and volumes of distribution and of age on F1 , without inducing any modification in the MQ basic model. Sex and age were found to significantly influence K95% and differed less than 5% from the bootstrap estimations. The results of the pcVPC  with a precision of 16% at an individual level. A small bias of 18% (CI95% 13\u201324%) with a precision of 81% was calculated for population predictions. Non-significant or small  biases were calculated at each study site on an individual level .0\u2013day63 for children and adult volunteers and patients. Median (PI95%) AUC0\u2013day63 of 725\u00a0mg/L/h (310\u20131718) was computed through simulations of the final pharmacokinetic model for children weighting 12.2\u00a0kg and taking 110\u00a0mg of MQ once per day over three consecutive days. Adult patients had a median (PI95%) AUC0\u2013day63 of 1080\u00a0mg/L/h (599\u20131911) and 936\u00a0mg/L/h (570\u20131413) calculated using the model of Julien et al. and Reuter et al. respectively, while adult volunteers of 865\u00a0mg/L/h (555\u20131211) under the dosage regimen of MQ 400\u00a0mg once per day over three consecutive days. Median (PI90%) concentration time profiles for adult and children patient are shown in Fig.\u00a0Figure\u00a0L/h 570\u2013113 calcul0\u2013day7 were estimated to be 281\u00a0mg/L/h (70\u2013854\u00a0mg/L/h) in children with reported treatment success within the follow-up period, and 286\u00a0mg/L/h (167\u2013378\u00a0mg/L/h) and 286\u00a0mg/L/h (70\u2013579\u00a0mg/L/h) for children with or without malaria recrudescence, respectively. No significant associations were found through logistic regression between model-predicted AUC0\u2013day at day 7, 28, 42 or 63 and appearance/absence of recrudescence (p\u2009>\u20090.05) (data not shown for day\u2009>\u20097).Treatment failure was reported for 212 (56%) of the children enrolled in the study, of these failures, 81% (n\u2009=\u2009171) were due to new infections and 7% (15) to recrudescence during the 63\u00a0days of follow-up. In 2% of the enrolled individuals PCR information was missing and in 10% it was not possible to determine the nature of the treatment failure. Median (range) model-predicted AUCmax for AS and DHA agree with reported peak AS and DHA plasma concentrations within the 1st h and 2\u00a0h post-dose, also supporting the appropriateness of the value chosen for AS Ka in this work [The present analysis describes the pharmacokinetics of fixed-dose ASMQ in African children under the age of 5\u00a0years, with the aim of identifying the source of the significant variability in drug exposure and validating the recommended weight-for-age dosage regimen. The very short half-lives estimated for AS and DHA are in good agreement with reported values ranging from 22 to 72\u00a0min for the drug and from 30 to 186\u00a0min for the metabolite , 29. Thehis work , 29.A two-compartment model was used to describe mefloquine pharmacokinetics as already shown in previous analyses , 27. DruConsiderable between-subject variability characterized the pharmacokinetics of both anti-malarial drugs. Such variability remained largely unexplained by the inclusion of the available covariates. Body weight was associated to all the MQ and DHA pharmacokinetic parameters. The association between this demographic characteristic and AS and DHA dispositions has already been described , 33. RepTwenty-one percent of the initial AS relative F1 variability was explained by age and treatment day. It is worth realizing that F1 is intrinsically connected to AS and DHA pharmacokinetic parameters, apparent because of ASMQ oral administration. The decrease in F1 observed with age implies an increase in drug and metabolite eliminations. This effect might thus be related to organ maturation in the study population. F1 was significantly higher at day 0 than days 1 and 2 of treatment. This is a consequence of the rapid and efficacious therapeutic AS effect observed already after the first AS dose intake. Recently, the relationship between malaria disease and AS bioavailability has been described using parasitaemia variation , 35. An max has been reported in healthy fasting volunteers taking an MQ dose compared to those having a high-fat breakfast (36 vs. 17\u00a0h), meaning that food would increase MQ Ka [a than older ones. Of note, the impact of food on MQ Ka remains controversial [Age was found to markedly decrease MQ drug absorption rate. A significantly higher treakfast  vs. 17\u00a0hoversial , 38. Finoversial . The PK As already described in studies performed in Tanzania and Cambodia, more than half of the African paediatric participants had a residual concentration of at least one anti-malarial drug above the limit of quantification at baseline  , 40. TheThe MQ model developed in Kenyan children using intensive sampling was applied to data collected from children from Burkina Faso, Tanzania and Kenya. Similar non-significant or small biases and precision per study centre were estimated, suggesting comparable drug exposure among different populations. The relationships between therapeutic response and pharmacokinetics of MQ as monotherapy and in combination with AS have been previously compared in a Thai population . RecrudeP. falciparum malaria. The considerable variability characterizing the pharmacokinetics of these two anti-malarial drugs can be partly explained by children\u2019s body weight, justifying the current dosing recommendations based on weight-for-age considerations, to ensure similar exposure in children and adults.The study described provides the pharmacokinetic parameters for MQ and AS, administered as a FDC of AS/MQ, in African children under the age of 5\u00a0years with acute Additional file 1. Artesunate (upper panel) and dihydroartemisinin (lower panel) goodness-of-fit plots of observed vs. individual and population predicted concentrations, and conditional weighted residuals (CWRES) vs. population predicted concentrations and time after dose.Additional file 2. Mefloquine goodness-of-fit plots of observed vs. individual and population predicted concentrations, and conditional weighted residuals (CWRES) vs. population predicted concentrations and time after dose."}
+{"text": "Due to poor availability and affordability of the innovator\u2019s product, the government of Tanzania in 2013 prequalified the use of generic anti-malarial drugs, whereby Artefan\u00ae  was the first to be approved.In 2006, artemether\u2013lumefantrine (ALU), specifically Coartem\u00ae in comparison with innovator\u2019s product Coartem\u00ae. Patients aged 6 to 59\u00a0months with uncomplicated malaria were recruited and randomized to either receive Artefan\u00ae or Coartem\u00ae as a control. Participants were required to revisit clinic five times as follow up to monitor treatment outcome as per World Health Organization recommendations. On each visit, thick and thin blood smears, dried blood spot (DBS), haemoglobin concentrations and auxiliary temperature were performed and documented.This was an equivalence prospective study that aimed to determine the effectiveness of anti-malarial generic Artefan\u00ae and Coartem\u00ae. The overall PCR uncorrected cure rate were 80% for Artefan\u00ae and 75% for Coartem\u00ae (p\u2009=\u20090.44). Adequate clinical and parasitological response were 82.1% for Artefan\u00ae and 74.7% for Coartem\u00ae, and there was no early treatment failure (ETF) observed in both arms of treatment. Both drugs showed excellent early parasite clearance, whereby no participants had peripheral parasitaemia on day 3. Late clinical failures (LCF) were 3.6% for Artefan\u00ae and 1.3% for Coartem\u00ae (p\u2009=\u20090.31), and late parasitological failure (LPF) were 15.4% for Artefan\u00ae and 22.7% for Coartem\u00ae (p\u2009=\u20090.32). Mean haemoglobin (g/dl) concentrations observed on day 28 were higher compared to day 0 for both drugs, although not statistically significant. Only one (1.3%) participant on Artefan\u00ae had temperature\u2009\u2265\u200937.5\u00a0\u00b0C on day 3.Out of 230 recruited participants, 200 met inclusion criteria and were randomized equally to receive Artefan\u00ae and Coartem\u00ae are equivalent and effective in the management of uncomplicated malaria amongst children in the Coast part of Tanzania.The findings of this study indicate that both Artefan Malaria continues to be one of the major public health problems in many regions of the world, particularly the developing countries . AfricanAnopheles gambiae and Anopheles arabiensis are the two species of Anopheles mainly responsible for malaria transmission in Tanzania. Ninety-six percent of malaria in Tanzania is caused by Plasmodium falciparum and the remaining 4% due to Plasmodium malariae and Plasmodium ovale [Plasmodium determine appropriate treatment approach [A majority (90%) of Tanzanians (both mainland and Zanzibar) live in moderate to high malaria endemic region . About 1um ovale . Co-exisapproach  and is oP. falciparum to anti-malarial drugs has led to changes of drugs from chloroquine to sulfadoxine\u2013pyrimethamine (SP) and currently artemisinin-based combination therapy (ACT) [\u00ae  was prequalified by the Tanzanian government and the product was available in all public health facilities. Due to high cost and poor availability of the innovator\u2019s product, the government of Tanzania prequalified the use of generic ALU, which is now more widely available in public health facilities than Coartem\u00ae. Artefan\u00ae  was the first generic drug to be prequalified by the government of Tanzania and is still available in the supply chain.Proper diagnosis and prompt treatment with effective anti-malarial drugs are one of the major tools in the control of malaria. The emergence of resistance of py (ACT) , 5. In 2py (ACT) . Since tP. falciparum resistance to artemisinin. Proportion of patients with detectable parasitaemia on day 3, 72\u00a0h following initiation of anti-malarial therapy is used to predict resistance  recommends frequent monitoring of their effectiveness. Early parasite clearance after initiation of ACT is considered to be an indicator of treatment outcome , 8. Periemisinin , 9. AdeqGeneric artemisinin-based combinations play a great role in the management and control of malaria in developing countries as they are highly affordable and readily available. However, availability of substandard anti-malarial drugs in SSA, including Tanzania, has been reported and they pose a threat to the gained successes in malaria control , 8. PoorThis was an equivalence prospective study that aimed to determine the effectiveness of anti-malarial generic Artefan in comparison with innovator\u2019s product Coartem. The study was conducted at Kibiti health centre in Kibiti district, which is an area in the coastal region of Tanzania, with a malaria prevalence of about 10.2%. In this surveillance of anti-malarial drugs, a formula for sample size calculation in equivalence studies  was usedP. falciparum, (4) agreement to come to the study clinic for any febrile episode or other illnesses with the parent/guardian, (5) residence in Kibiti town perimeter, (6) absence of chronic diseases like HIV/AIDS, kidney disease and any active medical problem requiring hospitalization, and (7) provision of informed consent form signed by parents/guardians.Consecutive sampling technique was used to enroll patients aged between 6 and 59\u00a0months with uncomplicated malaria  after being examined clinically by physician). After being enrolled, patients were randomized to receive either the generic artemether\u2013lumefantrine (Artefan) or the innovator\u2019s product (Coartem) as a control group, and thereafter followed until day-28 as per WHO guidelines . EligibiDiagnosis of malaria followed current Tanzania standard treatment guidelines (STG) in which uncomplicated malaria is defined as symptomatic malaria without signs of severity or evidence  of vital organ dysfunction . But the clinical features listed above are not specific for malaria and can be found in several other febrile conditions. Therefore, diagnosis of uncomplicated malaria was done by using RDT  ) followed by confirmation by microscopy. Children who were malaria positive aged between 6 and 59\u00a0months were randomized to receive either Artefan or Coartem, as recommended by malaria treatment guidelines . Both ArThick and thin blood smears were stained with 2% Giemsa for 30\u00a0min and microscopic examination was performed by trained laboratory technologists who were not involved in direct patient care. Parasite densities were calculated from thick blood smears by counting the number of asexual parasites per 200 leukocytes , assuming a leukocyte count of 8000/\u03bcl. A blood smear was considered negative when the examination of 100 high power fields did not reveal asexual parasites. For quality control, all slides were read by two readers. In case of discordant readings, an independent third reader was used settled discrepancies between the first and second readings. Laboratory technicians were blinded to the study participants\u2019 treatment assignments. Thin smears were used to determine the parasite species , 15.+  microcuvette machine, from. The drop of blood was collected in an Hb 201 microcuvette and read using HemoCue Hb 201+ device and result was recorded in g/dl.Haemoglobin (Hb) was determined on D0, D3, D7, D14 and D28 and quantification of Hb was done using a Hemocue Hb 201Data were entered into an Excel sheet and analysis was done by using Statistical Package for Social Sciences (SPSS) software version 20 . Mean, geometric mean, range, standard deviation and percentage were used to summarize results. Chi square was used in finding association between variables of interest. Kaplan\u2013Meier using per-protocol approach was used to determine cure rate and malaria recurrent rate. p-value of\u2009<\u20090.05 was considered to be statistically significant.P. falciparum and P. malariae  on Artefan and male 51.6%) on Coartem with mean age of 33\u00a0months and 32\u00a0months, respectively. Baseline mean temperature was 38.4\u00a0\u00b0C on Artefan and 38.5\u00a0\u00b0C on Coartem. Baseline peripheral parasitaemia geometric means were 10,074.6% on CoThe treatment outcomes were defined as: early treatment failure (ETF)\u2014presence of danger signs or complicated malaria with a positive blood smear on day 3 of treatment, or day 3 parasite density\u2009>\u200925% of day 0 parasite density, or positive blood smear on day 3 with auxiliary temperature\u2009\u2265\u200937.5\u00a0\u00b0C. Late clinical failure (LCF)\u2014danger signs or severe malaria in the presence of parasitaemia on any day between day 4 and day 28 in patients who previously did not meet any of the criteria of early treatment failure, and presence of parasitaemia on any day between day 4 and day 28 with auxiliary temperature\u2009\u2265\u200937.5\u00a0\u00b0C in patients who previously did not meet any of the criteria of early treatment failure. Late parasitological failure (LPF)\u2014presence of parasitaemia on any day between day 7 and day 28 (day 42) with auxiliary temperature\u2009<\u200937.5\u00a0\u00b0C in patients who previously did not meet any of the criteria of early treatment failure or late clinical failure. Adequate clinical and parasitological response (ACPR)\u2014absence of parasitaemia on day 28 irrespective of auxiliary temperature, in patients who previously did not meet any of the criteria of early treatment failure, late clinical failure or late parasitological failure.None of the study participants had peripheral parasitaemia on day 3 on both Artefan and Coartem treatment arms. On day 3 only, 1 (1.1%) patient on Artefan treatment arm had temperature above 37.5\u00a0\u00b0C. Slightly decrease in Hb concentration was observed on day 3 in both arms of treatment but the difference was not statistically significant Table\u00a0. The meaAfter the introduction of artemisinin-based combination therapy (ACT) as the first-line and second-line option in the management of uncomplicated and complicated malaria by the WHO together with other methods of malaria prevention such as insecticide-treated nets (ITN) and indoor spraying, prevalence of malaria has decline in many countries. Protecting the efficacy of ACT for the treatment for falciparum malaria is among the top global public health priorities. The presence of falsified and substandard drugs has been reported in many parts of the world, including sub-Saharan Africa (SSA) . The WHOIn this study, the therapeutic efficacy of generic anti-malarial Artefan and innovator\u2019s product Coartem were compared. Early parasite clearance was excellent for both drugs with no detected peripheral parasitaemia on day 3  using microscopy. Temperature clearance was also outstanding for both Artefan and Coartem, with only 1 (1.1%) patient in the Artefan arm having temperature above 37.5\u00a0\u00b0C on day 3. The early parasite clearance in this study on day 3 was the same as what was reported in Uganda, in which early parasite clearance following treatment with artemether-lumefantrine was excellent .Plasmodium species to artemisinin has been reported in eastern and southern Asian countries [P. falciparum to artemisinin in Tanzania. This is in line with the WHO 2009 anti-malarial protocol, that if 10% of the study participants have peripheral parasitaemia on day 3, it is an indicator of emergence of artemisinin resistance to Plasmodium species [Resistance of ountries , but not species .Early treatment failure (ETF) was not observed in both Artefan and Coartem treatment arms, whereas late clinical failures (LCF) were 3.6% and 1.3% in Artefan and Coartem arms, respectively. However, late parasitological failures (LPF) were high in both Artefan (15.4%) and Coartem (22.7%) arms. Artemisinin products (for this study artemether) have a short half-life. Since most of the LCF and LPF occurred on day 21 and 28 with only one LPF occurring on day 14, its means the partner drug (lumefantrine) offers a prolonged protection against malaria of two to 3\u00a0weeks. From the survival analysis, cumulative hazard proportion curve shows that children using Coartem might be at higher risk of parasitological failure compared to children using Artefan. The observed differences in late treatment failures (LCF and LPF) between Artefan and Coartem were not statistically significant, an indication that the generic anti-malarial Artefan is effective for management of uncomplicated malaria in Tanzanian children.The observed LCF in this study is comparable to what was reported in Ethiopia . In the The WHO recommends the use of microscopy as a golden standard for diagnosis of malaria in middle and low income countries. In this study, microscopy was also used to assess the performance of anti-malarial drugs. Using microscopy, the observed adequate clinical and parasitological response (ACPR) were 83.3% in children in the Artefan and 77.3% in the Coartem arms. PCR uncorrected cure rate were\u2009>\u200980% for Artefan and 75% for Coartem. These results indicate that both generic Artefan and innovator\u2019s product Coartem are effective for the treatment of uncomplicated malaria in children.High adequate clinical and parasitological responses and malaria cure rate of ALU have been reported in different therapeutic efficacy studies. For instance, 100% adequate clinical and parasitological response were reported in South West Ethiopia in children under 5\u00a0years of age 23), and very low 45.4%) in Uganda 5.4% in U and sout, and verBased on mid upper arm circumference (MUAC), most of the participants in this study had good nutrition status. The difference in clearing malaria parasites between those with poor nutrition status and good nutrition status could not be established. Studies have shown conflicting conclusions on the role of nutrition in relation to malaria parasite clearance with some linking poor parasite clearance and malaria morbidity and mortality with poor nutrition status , 25, whiPlasmodium species. Therefore, the reported cure rate in both arms of the study are PCR uncorrected.The limitation of this study is that it was designed to mimic the routine standard of care for management of uncomplicated malaria in Tanzania. In this approach, patients are given anti-malarial drugs with instruction to take the prescribed doses at home. Therefore, information on the parasite density during day 1 and day 2 after drug administration were not collected. In addition, adherence to the prescribed drugs was based on self-reports from patients, parents or guardians. Also, microscopy instead of molecular techniques, such PCR, was used to identify This study revealed that Artefan as generic anti-malarial drug containing artemether and lumefantrine is equivalent to the innovator\u2019s product Coartem in the management of uncomplicated malaria in Coast region of Tanzania. Due to low cost and availability, the use of Artefan as the generic drug is advantageous in comparison to the relatively expensive innovator`s drug such as Coartem. Therefore, the findings of this study support continuing use of generic anti-malarial drugs in the management of uncomplicated malaria in children."}
+{"text": "The incidence of malaria in the Americas has decreased markedly in recent years. Honduras and the other countries of Mesoamerica and the island of Hispaniola have set the goal of eliminating native malaria by the year 2020. To achieve this goal, Honduras has recently approved national regulations to expand the possibilities of a shortened double dose primaquine (PQ) treatment for vivax malaria. Considering this new shortened anti-malarial treatment, the high frequency of G6PDd genotypes in Honduras, and the lack of routinely assessment of the G6PD deficiency status, this study aimed at investigating the potential association between the intake of PQ and haemolysis in malaria-infected G6PDd subjects.Plasmodium vivax infection was treated with 0.25\u00a0mg/kg of PQ daily for 14\u00a0days. Safety and signs of haemolysis were evaluated by clinical criteria and laboratory values before and during the 3rd and 7th day of PQ treatment. G6PD status was assessed by a rapid test (CareStart\u2122) and two molecular approaches.This was a prospective cohort and open-label study. Participants with malaria were recruited. Overall 55 participants were enrolled. The frequency of G6PD deficient genotypes was 7/55 (12.7%), where 5/7 (71.4%) were hemizygous A\u2212\u2009males and 2/7 (28.6%) heterozygous A\u2212\u2009females. Haemoglobin concentrations were compared between G6PD wild type (B) and G6PDd A\u2212\u2009subjects, showing a significant difference between the means of both groups in the 3rd and 7th days. Furthermore, a statistically significant difference was evident in the change in haemoglobin concentration between the 3rd day and the 1st day for both genotypes, but there was no statistical difference for the change in haemoglobin concentration between the 7th day and the 1st day. Besides these changes in the haemoglobin concentrations, none of the patients showed signs or symptoms associated with severe haemolysis, and none needed to be admitted to a hospital for further medical attention.The findings support that the intake of PQ during 14\u00a0days of treatment against vivax malaria is safe in patients with a class III variant of G6PDd. In view of the new national regulations in the shortened treatment of vivax malaria for 7\u00a0days, it is advisable to be alert of potential cases of severe haemolysis that could occur among G6PD deficient hemizygous males with a class II mutation such as the Santamaria variant, previously reported in the country.The online version of this article (10.1186/s12936-018-2564-2) contains supplementary material, which is available to authorized users. Malaria remains one of the diseases with higher rates of morbidity and mortality in the tropical world. According to the World Health Organization (WHO) an estimated of 216 million cases of malaria occurred worldwide in 2016, however the incidence rate of malaria is estimated to have decreased by 18% globally between 2010 and 2016, with the WHO Region of the Americas recording the second largest decline (22%) after the South-East Asia Region (48%) [Plasmodium vivax and Plasmodium falciparum, responsible for 90% and 10% of malaria cases, respectively . Due to persistent liver stages (hypnozoites) of P. vivax relapses of the disease may occur weeks or months after the first infection [P. vivax have their origin in the reactivation of hypnozoites [The only two parasite species causing malaria in Honduras are nfection , 4. It inozoites , that canozoites . Howevernozoites .G6PD deficiency is an X-linked genetic disorder that affects more than 400 million people worldwide, and its average prevalence in countries with endemic malaria is 8% (3\u201330%) . Recent Because of the new goals to eliminate malaria from the Mesoamerican/Hispaniola sub-region by 2020, the new national regulation of Honduras approved in 2017 opens the possibility of administering PQ in a shortened double dose scheme for 7\u00a0days for vivax malaria to improve the adherence to treatment. Although shortening treatment could improve patients\u2019 adherence in order to a radical cure of the parasite, it could also increase the risk of haemolysis in individuals with G6PDd, particularly those hemizygous males affected with severe class II variants.Therefore, when considering the high frequency of G6PDd genotypes in Honduras, the blind administration of PQ without G6PDd analysis at the point of care, and the change in the PQ treatment scheme from 14 to 7\u00a0days at double dose, it is pertinent to investigate if there is an association between the intake of primaquine and haemolysis in G6PD deficient subjects infected with malaria.This was a prospective cohort and open-label study carried out between February and June 2017. Participants were recruited from 4 malaria endemic municipalities located at the northern coast of Honduras: Tocoa/Saba (Department of Colon), Roatan (Department of Bay Islands), and Puerto Lempira (Department of Gracias a Dios) . A flowchart showing the procedures carried out in this study can be found in Fig.\u00a0The microscopic diagnosis of parasitaemia was performed by thick blood according to national standard procedures and confirmed by the immunochromatographic test RAPID 1-2-3P. falciparum received 25\u00a0mg/kg of CQ over 3\u00a0days plus 0.75\u00a0mg/kg of PQ in a single dose the first day of treatment.Immediately after the malaria diagnosis a phenotypic test was performed for the detection of G6PD deficiency . Patients were treated according to the National Malaria Standard valid since 2010 . UncomplP. falciparum were visited and monitored only the 3rd day of treatment. The clinical evaluation determined vital signs such as heart rate, respiratory rate, temperature and blood pressure. Symptoms and signs such as low back pain, abdominal pain, choluria, oliguria, jaundice, respiratory distress, pallor, and hepatosplenomegaly, were also recorded. Blood samples were sent to local laboratories to quantify levels of haemoglobin, haematocrit, red blood cells, reticulocytes, AST, ALT, bilirubin and creatinine. The values of reticulocytes, AST, ALT, bilirubin and creatinine could not be obtained for the participants of Puerto Lempira due to the lack of an appropriate clinical laboratory in the city.Patients were clinically assessed the first day of medical consultation, and during treatment. Two follow-ups were conducted to each participant by trained physicians, visiting them in their households. Blood samples were collected on the 1st day before treatment, and during the 3rd and 7th day of PQ treatment. Patients infected with A drop of blood was taken on filter paper from each participant on the first day of recruitment for further molecular analyses. Genomic DNA was extracted using a Chelex-100 based method . To conf\u00ae commercial kit enabled the detection of the following six variants: Class IV variant A+\u2009(376A\u2192G); 3 different class III variants A\u2212\u2009, , ; class II Santamaria ; and class II Mediterranean . This procedure was carried out according to manufacturer\u2019s instructions.In order to identify G6PD genotypes associated with enzyme deficiency, two molecular approaches were carried out, an in-house PCR\u2013RFLP and the DiaplexC\u2122 G6PD genotyping (African type) commercial kit . The PCR\u2013RFLP assay was designed to independently identify single nucleotide polymorphisms in the positions 202 (G\u2192A) and 376 (A\u2192G) within the gene corresponding to exons IV and V. This method was carried out according to protocols previously described and adapted , 17, 18.S) among the studied population. A PCR\u2013RFLP approach was conducted according to previous reports [Additionally, a genetic analysis of the \u03b2-globin gene was also performed to reveal the presence of the sickle cell trait . The Kolmogorov\u2013Smirnov (n\u2009\u2265\u200950) and Shapiro\u2013Wilk (n\u2009<\u200950) tests were used to evaluate whether the data were normally distributed for each laboratory variable  within each group (genotype). A p value\u2009<\u20090.05 was established for the rejection of the null hypothesis.The change in concentration for normally distributed data was expressed in absolute values. The difference among values recorded on the 3rd and 7th day of primaquine intake with respect to the baseline value of the 1st day before treatment was calculated for each group. The distributions of these changes were graphically represented by box plots. In all the statistical tests a \u00ae Kit) was assessed by the Cohen\u2019s kappa coefficient using the Epidat program, v. 3.1. A p value\u2009=\u20090.05 and confidence intervals\u2009=\u200995% were considered. The agreement between the RDT and the molecular approaches could not be calculated since the RDT is able to detect G6PD deficiencies with less than 10% of enzyme activity, and the most common class III mutations detected through the PCR\u2013RFLP and the multiplex PCR are associated with residual enzyme activity of 10\u201360%.The measure of agreement between G6PD molecular techniques . Thirty-three patients were males and twenty-two females. Ten participants were under 18\u00a0years old, seven of them from Puerto Lempira. Ninety-six percent of infections were caused by \u00ae) is able to detect six mutations  and revealed six subjects with genotype A\u2212\u2009 and one with genotype A+\u2009(376A\u2192G) . The phenotypic test detected 5 (9.1%) deficient patients, 4 of them males Table\u00a0. All patThe results of the two genetic tests matched on the diagnosis of the A\u2212\u2009genotype for 2 heterozygous women and 4 of 5 hemizygous males. However, one of the subjects with genotype A\u2212\u2009by PCR\u2013RFLP revealed a genotype A+\u2009with the commercial kit revealing a Kappa index\u2009=\u20090.723 , p\u2009<\u20090.001. When comparing the results of the genotypic and phenotypic approaches, we found that only 2 phenotypically deficient patients showed a class III A\u2212\u2009variant . In contrast, 3 patients with phenotypic G6PD deficiency by RDT showed a normal B genotype; and 5 patients  with a normal phenotype were classified as A\u2212\u2009by PCR\u2013RFLP. All discrepant genetic tests were repeated with the same results. The discordant phenotypic tests could not be repeated because the genetic results were obtained months later, and fresh blood samples were required.All the participants were clinically evaluated the first day of enrolment and the 3rd day of primaquine (PQ) treatment. Patients with vivax malaria (n\u2009=\u200953) were also evaluated on the 7th day of treatment. Signs and symptoms suggestive of haemolysis were recorded as present or absent. None of the patients showed signs or symptoms associated with severe haemolysis, and none needed to be admitted to a hospital for further medical attention. Mild or moderate abdominal pain was the most commonly reported sign, followed by hepatomegaly Table\u00a0. There wIn order to evaluate haemolysis levels during treatment with PQ, haematological analyses were performed to all participants (n\u2009=\u200955) during the 1st, 3rd and 7th follow-up days; however, biochemical analyses were performed only to participants who resided in Tocoa/Saba and Roatan (n\u2009=\u200945) due to logistic limitations in Puerto Lempira city.Table\u00a06/\u03bcL, 12.4\u00a0g/dL, and 37.5%, respectively, during the follow-up of days 3 and 7 and for both genetic groups. The mean of reticulocyte percentage was higher than 1% in all cases. The results of the biochemical analyses were more similar to normal values, with the exception of a slight elevation in bilirubin.The mean of red blood cells counts, haemoglobin, and haematocrit percentage were lower than 4.4\u2009\u00d7\u200910p\u2009\u2265\u20090.05) within each group and could be analysed by Student\u2019s t test. The rest of the laboratory data were not analysed by the Student\u2019s t test. The mean haemoglobin concentrations on the 1st day did not show a statistically significant difference between the A\u2212 and non-A\u2212 genotypes, however, there was a significant difference between the means of both groups in the 3rd and 7th days, with p values\u2009<\u20090.05 . Only haemoglobin values revealed a normal distribution of data (Kolmogorov\u2013Smirnov test: 05 Table\u00a0.Table\u00a06Mp\u2009=\u20090.040). The distributions of these changes are shown in Fig.\u00a0The difference was also calculated in the haemoglobin concentration among the values recorded during the 3rd day and 7th day with respect to the 1st day before PQ treatment for each genotype and compared the means of those changes by Student\u00b4s t test. As shown in Table\u00a0S/\u03b2A) women (3.6%), one of them heterozygous for the G6PD A\u2212\u2009mutation and the other one with a G6PD wild genotype.The results of the genetic analysis of sickle cell trait revealed two heterozygous (\u03b2P. vivax was 53 (96.4%) and the remaining 3.6% by P. falciparum. These results are consistent with the official data for the year 2017 in Honduras, which indicate 1155 cases (90%) for vivax malaria and the remaining 10% caused by P. falciparum.In order to evaluate the risk of haemolysis in patients infected with malaria and treated with primaquine in Honduras, fifty-five subjects were enrolled and treated according to the national scheme in force. The patients were recruited from three departments with the highest endemicity in the country, accounting for almost 70% of cases . The number of infections caused by According to the CareStart\u2122 rapid diagnostic test (RDT), 5 (9.1%) subjects were classified as deficient, however only 2 of these 5 subjects revealed a hemizygous A\u2212\u2009genotype through molecular analyses of the G6PD gene. In contrast, the other three subjects with putative phenotypic deficiency showed a wild genotype B.Although several authors report acceptable percentages of sensitivity, specificity, and predictive values for the detection of G6PD deficiency by the CareStart\u2122 test \u201325, therIn order to detect clinical signs associated with haemolysis, the 55 participants were evaluated before the start of anti-malarial treatment and on the 3rd day of treatment. Most of them (n\u2009=\u200953) were also evaluated on the 7th day of treatment. None of the patients showed signs associated with severe haemolysis and there was no notable difference between patients with wild genotype and those with deficient genotype. As established by the WHO , this reSimilar results were observed when analysing haematological values from patients during PQ treatment. As shown in Table\u00a0S alleles. These findings seem to rule out the influence of sickle cell disease as a contributor to mild anaemia in this study, since only 2 patients were heterozygous for \u03b2S.A previous report indicates that more than 23% of the Afro-descendant population living in malaria endemic areas in Honduras presents the sickle-cell trait, and an overall prevalence of anaemia of 47% in 2014 . Even thp\u2009=\u20090.034 for day 3, and p\u2009=\u20090.004 for day 7)  Table\u00a0. A stati Table\u00a0. These results could mean that patients with deficient genotype suffer a greater loss of haemoglobin than patients without G6PD deficiency due to the intake of PQ. According to the WHO , anaemiaP. falciparum infections receiving a single dose of PQ administered at different concentrations . Those P. vivax requiring longer treatments (7\u201314\u00a0days) are scarcer. A study conducted in Thailand compared the PQ safety (30\u00a0mg once daily for 7\u00a0days) in G6PD deficient and not deficient patients. A significant reduction in haematocrit was recorded only in patients with G6PDd after the 7th day of treatment, but without triggering severe anaemia [Studies evaluating the safety of PQ in G6PDd patients infected by  anaemia . Unfortu anaemia . With si anaemia  evaluateP. vivax constitutes 90% of annual cases, and its treatment requires 14\u00a0days of treatment with PQ in doses of 0.25\u00a0mg/kg [In the Honduran context, the malaria caused by 25\u00a0mg/kg , 41\u201343. 25\u00a0mg/kg . CircumsPoor adherence to the 14-day treatment with PQ scheme is common in many countries . For thaIn this study, the risk of haemolysis in patients infected with malaria was evaluated after receiving regular treatment with PQ in 3 endemic municipalities of Honduras. Seven individuals (12.7%) with genotype G6PDd A\u2212\u2009were detected, of which 5 were hemizygous males. It was confirmed that the intake of PQ during 14\u00a0days of treatment against vivax malaria is safe in both patients with and without G6PDd, however a statistically significant difference was demonstrated in the decrease of mean haemoglobin levels among the deficient and normal patients in the 3rd and 7th days after PQ intake. Because the new national treatment scheme for vivax malaria allows the use of double dose in a shortened PQ treatment of 7\u00a0days, it is advisable to be alert for the possible cases of severe haemolysis that could occur especially among G6PD deficient males with a class II mutation.Additional file 1: Table S1. Mean of haemoglobin concentration of individuals with G6PD genotype B and A+\u2009by age and sex."}
+{"text": "Low mefloquine exposure has been shown to contribute to treatment failure in patients with uncomplicated falciparum malaria following a 3-day artesunate\u2013mefloquine combination. The present study aimed to develop a population pharmacokinetic model for mefloquine based on whole blood concentration\u2013time profiles of this target population for further dose optimization.Plasmodium falciparum were included in the study. All were treated with the standard 3-day combination regimen of artesunate and mefloquine consisting of mefloquine for 2\u00a0days and artesunate for 3\u00a0days. Blood samples were collected before and at different time points after drug administration from different sub-groups of patients. Mefloquine concentrations were quantified in whole blood using high-performance liquid chromatography. A non-linear mixed-effect modelling approach was applied for population pharmacokinetic analysis using the NONMEM v7.3 software. Covariates investigated  were investigated in a step-wise manner using the SCM functionality in Perl-Speaks-NONMEM.A total of 129 Burmese patients aged above 15\u00a0years who presented with typical symptoms of malaria and had a blood smear positive for max) and elimination half-life (t1/2) were found in patients who had treatment failure (36 cases) compared to patients with successful treatment (107 cases).Population pharmacokinetic analysis of mefloquine was performed in all patients with a total of 653 samples. Whole blood mefloquine concentration\u2013time profiles were described by a two-compartment disposition model. Of the covariates investigated, none was found to have a significant impact on the pharmacokinetics of mefloquine. Significant differences in maximum concentration (CThe study successfully describes the pharmacokinetics of mefloquine following a 2-day treatment of mefloquine as a part of a 3-day artesunate\u2013mefloquine in patients with uncomplicated falciparum malaria from Thailand. A model has been developed which adequately describes the pharmacokinetics of mefloquine. More extensive clinical studies including both adults and children are needed to fully characterize the pharmacokinetics of mefloquine. Plasmodium falciparum [P. falciparum to monotherapy, artemisinin-based combination therapy (ACT) was recommended by WHO in 2006 as a strategy to counteract the increasing resistance of P. falciparum to anti-malarials, as well as to prevent disease transmission and reduce the risk of drug resistance [Malaria remains a significant infectious disease that kills over 1200 people every day [sistance . ACT consistance . These csistance . Unfortusistance .P. falciparum to ACT is of great concern to global malaria control programmes as alternative treatment options are limited.Among the recommended ACT regimens for falciparum malaria, artesunate\u2013mefloquine combination was introduced as first-line treatment of multi-drug-resistant, uncomplicated falciparum malaria in Thailand before WHO recommendation. The combination n was initially used in 1995 as a 2-day combination regimen of 25\u00a0mg/kg body weight of mefloquine and 12\u00a0mg/kg body weight of artesunate to ensure better compliance . The recIntroduction of the triple combination of anti-malarials instead of the standard two-drug combination has been suggested as a further strategy to control artemisinin resistance . ArtesunThis population pharmacokinetic study was a part of the study protocol to investigate the clinical efficacy of a 3-day artesunate\u2013mefloquine combination in patients with acute uncomplicated falciparum malaria in an endemic area along the Thai\u2013Myanmar border. The study was conducted at Mae Tao clinic for migrant workers, Tak Province, Thailand, from March 2008 to February 2009. Ethical approval of the study protocol was granted by the Ethics Committee of the Ministry of Public Health of Thailand. Detailed information of the study procedures and results of clinical efficacy assessment, including the relationship with drug concentrations were previously described in detail [P. falciparum diagnosed with blood smears, were included in the study. Inclusion criteria for enrolment in the study were patients with acute uncomplicated falciparum malaria according to the WHO protocol for areas with low to moderate malaria transmission [Burmese patients, aged over 15\u00a0years, who had typical symptoms of malaria and were positive for smission ; patientsmission ; historyAll patients received a 3-day combination regimen of artesunate and mefloquine. An initial dose consisting of 4\u00a0mg/kg body weight of artesunate  and 15\u00a0mg/kg body weight of mefloquine  was given on the first day of the study. On the second day, the patients received 4\u00a0mg/kg body weight of artesunate  and 10\u00a0mg/kg body weight of mefloquine . On the third day, mefloquine was not administrated; instead artesunate (4\u00a0mg/kg body weight) was given together with 0.6\u00a0mg/kg body weight primaquine . All doses were observed and patients were followed for 30\u00a0min. If patients vomited within 30 min of treatment, the dose was repeated. Anti-pyretic paracetamol and anti-emetic dimenhydrinate were administered if necessary. Patients were requested to return for follow-up on days 7, 14, 21, 28, and 42, or if the patient experienced fever or malaria symptoms. Parasites were counted (Giemsa-stain) at each visit, and general symptoms were recorded.P. falciparum DNA to distinguish between re-infection and recrudescence [P. falciparum multi-drug resistance 1 , P. falciparum chloroquine resistance transporter , and P. falciparum ATPase 6  [pfk13 sequencing was performed at K13-propeller domain  [pfmdr1 gene copy number was determined by SYBR Green I real-time PCR [Genotyping was performed on paired samples of descence . Polymer6-A623E) \u201315. The  720\u00a0bp) . The pfmtime PCR .Blood samples (1\u00a0ml of whole blood) were collected into sodium heparinized tubes before and after the first dose  for determination of mefloquine concentrations. Collected samples were stored at \u2212\u200920\u00a0\u00b0C until transported to Centre of Excellence in Pharmacology and Molecular Biology of Malaria and Cholangiocarcinoma, Chulabhorn International College of Medicine, Thammasat University. All samples were stored at \u2212\u200980\u00a0\u00b0C until drug quantification.Mefloquine concentrations were quantified using liquid chromatography according to a previously published method . The coeThe natural logarithm of quantified mefloquine concentrations was analysed to derive pharmacokinetic parameter estimates. A non-linear mixed-effect modelling approach was utilized using the NONMEM v7.3 software . Variousi is the individual parameter estimate, \u03b8P is the typical value of parameter P, and \u03b7i,P is the inter-individual variability for parameter P and patient is was drawn from a normal distribution with mean 0 and variance \u03c92.Individual parameter estimates were assumed to be log-normally distributed by adding between-subject variability with an exponential model according to Eq.  and the i,occ is the individual parameter estimate at occasion O, and \u03baO,P is the between-occasion variability of parameter P at occasion O drawn from a normal distribution with mean 0 and variance \u03a02.One-, two-, and three-compartment disposition modelswere evaluated to describe the absorption phase. A first-order absorption, a first-order absorption with the lag-time model, and a transit compartment model (1\u201313 transit compartments) with the absorption rate and rates between transit compartments were either assumed to be equal or estimated separately . BetweenThe model was parameterized as follows: apparent elimination clearance (CL/F), central volume of distribution (Vc/F), inter-compartmental clearance(s) (Q/F), peripheral volume of distribution(s) (Vp/F), and mean transit time of the absorption phase (MTT).Body weight was evaluated as an allometric function on all clearance and volume of distribution parameters. The exponents were fixed to 0.75 for clearance parameters and 1 for volume parameters based on previous reports , 26. OthDiscrimination between two hierarchical models was based on the difference in objective function value (OFV), which is proportional to \u2212\u20092 times the log-likelihood. The OFV was assumed to be Chi squared distributed resulting in a drop in OFV of 3.84 and 10.8 to be significant at statistical significance levels of 0.05 and 0.001, respectively.Model fit was also evaluated using goodness-of-fit plots by plotting observations against the population and individually predicted concentrations and conditionally weighted residuals against population predicted concentrations and time after dose . The perp\u2009<\u20090.05). Adding on one additional disposition model was not significant (p\u2009>\u20090.05). The absorption phase was described by 1 transit compartments; this absorption model was superior to the other tested absorption models (p\u2009<\u20090.05). Estimating the transit rate constant and the absorption rate constant separately resulted in a significantly improved model (p\u2009<\u20090.01). However, it also resulted in a model with a low precision in the estimation of the apparent volume of distribution of the central compartment (a relative standard error of 46.9% based on 300 bootstrap runs). The addition of relative bioavailability (F) fixed to 100% with an estimated inter-individual variability did not significantly improve the model and was excluded from the final model (p\u2009<\u20090.05). The addition of allometrically scaled body weight (exponent fixed to 0.75 and 1) did not improve the model and was not kept in the final model. Finally, the addition of between-occasion variability on the absorption parameters did not improve the model and was therefore excluded from the final model.Population pharmacokinetic analysis of mefloquine was performed in 129 Burmese patients with uncomplicated falciparum malaria with a total of 653 data points. Thirty-six patients had the re-appearance of parasitaemia during days 21\u201340 of the 42-day follow-up period, and 93 patients had the sensitive response. Patient demographic data are summarized in Table\u00a0kelch-13 mutations, only the pfmdr1-D12467 was found to have a significant impact on the pharmacokinetics of mefloquine. However, the increased relative bioavailability (before this parameter was excluded) found in patients with mutated parasites was deemed unlikely and was excluded from the final model. Significant differences in maximum concentration (Cmax) and elimination half-life (t1/2) were found in patients who had treatment failure (36 cases) compared to patients with successful treatment (107 cases).Covariates were investigated in a step-wise manner using the SCM functionality in Perl-Speaks-NONMEM. Of the covariates investigated , none was found significant. Of the investigated molecular markers of mefloquine resistance and max, time to maximum concentration (tmax), area under the concentration\u2013time curve (AUC), and t1/2 are presented in Table\u00a0A graphical representation of the final model is presented in Fig.\u00a00\u20137days). Drug concentrations capable of killing the parasite (above 500\u00a0\u03bcg/l) must be achieved for at least 3\u20134 parasite multiplication cycles (7\u00a0days) to be confident that cure will be achieved [Mefloquine has had a history of resistance development, but has been successfully used in Thailand in combination with artesunate , 31. Witachieved . In thisThe present study describes the pharmacokinetics of mefloquine with individual parameter estimates using a non-linear mixed effect model. The pharmacokinetics of mefloquine was well described with a two-compartment model. This is in agreement with that previously reported \u201338. AlthThe mefloquine dose of 25\u00a0mg/kg of body weight was split into a 15\u00a0mg/kg dose followed by 10\u00a0mg/kg body weight dose to improve oral bioavailability and tolerability . In the Between-occasion variability has previously been identified on elimination clearance of mefloquine . HoweverParameter estimates of mefloquine population pharmacokinetics were similar to that previously published, and all parameters could be estimated with reasonable precision, indicating a stable model \u201342. ShriP. falciparum, and the impact of mixed infection was not evaluated. Admission parasitaemia was however, evaluated on all parameters and was not found significant. Validated molecular markers for mefloquine resistance were investigated to evaluate if any of those would affect the pharmacokinetic properties of mefloquine. The only marker retained after the backward step was pfmdr 1-D12467, estimating an increased relative bioavailability in mutated patients. This was deemed as an unlikely scenario from a biological standpoint and was not included in the final model. In addition, the analysis showed that mutation of the Kelch-13 propeller did not affect the pharmacokinetic parameters of mefloquine. The significant influence of artesunate administration on mefloquine pharmacokinetics is unlikely [Several covariates were investigated in the model. Body weight with fixed allometric exponents was investigated but was not found significant. The pharmacokinetics of several anti-malarial drugs, including piperaquine are affected by the body weight of the patients . For mefunlikely , 46 and The model was also used to derive the secondary parameters, identifying a significant difference in maximum concentration and elimination half-life between patients who had treatment failure and patients with successful therapy. This could indicate a different volume of distribution in these two groups. However, none of the available covariates successfully described this difference and additional more extensive studies are needed to evaluate why this difference was seen.This study successfully describes the pharmacokinetics of mefloquine following a 3-day artesunate\u2013mefloquine in patients with uncomplicated falciparum malaria from Thailand. A model has been developed which adequately describes the pharmacokinetics of mefloquine. The developed model supports the previously published two-compartment models and a one-transit compartment absorption model. The differences in the maximum concentration and elimination half-life were also identified between patients with successful treatment and those with recrudescence. More extensive clinical studies including both adults and children are needed to fully characterize the pharmacokinetics of mefloquine."}
+{"text": "Children hospitalised with severe anaemia in malaria endemic areas in Africa are at high risk of readmission or death within 6\u00a0months post-discharge. Currently, no strategy specifically addresses this period. In Malawi, 3\u00a0months of post-discharge malaria chemoprevention (PMC) with monthly treatment courses of artemether-lumefantrine given at discharge and at 1 and 2\u00a0months prevented 30% of all-cause readmissions by 6\u00a0months post-discharge. Another efficacy trial is needed before a policy of malaria chemoprevention can be considered for the post-discharge management of severe anaemia in children under 5\u00a0years of age living in malaria endemic areas.We aim to determine if 3\u00a0months of PMC with monthly 3-day treatment courses of dihydroartemisinin-piperaquine is safe and superior to a single 3-day treatment course with artemether-lumefantrine provided as part of standard in-hospital care in reducing all-cause readmissions and deaths (composite primary endpoint) by 6\u00a0months in the post-discharge management of children less than 5\u00a0years of age admitted with severe anaemia of any or undetermined cause.This is a multi-centre, two-arm, placebo-controlled, individually randomised trial in children under 5\u00a0years of age recently discharged following management for severe anaemia. Children in both arms will receive standard in-hospital care for severe anaemia and a 3-day course of artemether-lumefantrine at discharge. At 2\u00a0weeks after discharge, surviving children will be randomised to receive either 3-day courses of dihydroartemisinin-piperaquine at 2, 6 and 10\u00a0weeks or an identical placebo and followed for 26\u00a0weeks through passive case detection. The trial will be conducted in hospitals in malaria endemic areas in Kenya and Uganda. The study is designed to detect a 25% reduction in the incidence of all-cause readmissions or death (composite primary outcome) from 1152 to 864 per 1000 child years  and requires 520 children per arm .Participant recruitment started in May 2016 and is ongoing.NCT02671175. Registered on 28 January 2016.ClinicalTrials.gov, The online version of this article (10.1186/s13063-018-2972-1) contains supplementary material, which is available to authorized users. Severe anaemia, defined as haemoglobin (Hb) concentration level below 5.0\u00a0g/dL or haematocrit below 15.0% , 3. ChilStandard in-hospital treatment of severe anaemia in many countries in sub-Saharan Africa consists of a blood transfusion and parenteral artesunate for severe malaria . For sevMalaria control strategies in endemic and epidemic-prone areas include intermittent preventive therapy (IPT). IPT is the administration of a full treatment course using long-acting antimalarials at pre-defined time intervals irrespective of a patient\u2019s malaria status to clear existing infections and to provide prolonged prophylaxis against new infections . The WorWe are conducting an efficacy trial in Kenya and Uganda to determine the efficacy and safety of 3\u00a0months of malaria chemoprevention post-discharge as a potentially cost-effective strategy to reduce all-cause readmissions and deaths in children admitted with severe anaemia. We hypothesise that, by creating a prophylactic time-window post-transfusion for malaria, more time is assured for bone marrow recovery, resulting in a more sustained haematological recovery post-discharge.We refer to this strategy as post-discharge malaria chemoprevention (PMC) to illustrate the similarities with SMC rather than with IPT in pregnancy as it aims to provide complete, rather than intermittent prophylaxis.ClinicalTrials.gov, NCT02671175; registered 28 January 2016). Randomisation to PMC-DP or placebo will occur at 2\u00a0weeks after enrolment, and PMC treatments will be administered at 2, 6 and 10\u00a0weeks. The primary outcome will be the number of all-cause deaths or all-cause readmissions between 2 and 26\u00a0weeks after enrolment (composite outcome). The study will be conducted in Uganda and Kenya, using randomisation stratified by weight and study centre. The study will include a total of 1040 children (520 per study arm) less than 5\u00a0years of age who have been admitted for severe anaemia and have successfully completed the standard in-hospital treatment.This will be a multi-centre, parallel group, two-arm, placebo-controlled, individually randomised, superiority trial with 1:1 allocation ratio comparing the safety and efficacy of three courses of monthly PMC with dihydroartemisinin-piperaquine (PMC-DP) or placebo post-discharge provided in addition to the standard single 3-day treatment course with artemether-lumefantrine given as part of routine in-hospital care .Use of clinical malaria as primary outcome would require a smaller study; however, the composite outcome is used because it is more likely to drive policy. We use a composite outcome rather than a single severe outcome, such as death, to keep sample size requirements manageable.The period 2\u201326\u00a0weeks instead of 0\u201326\u00a0weeks is used for the primary efficacy analysis because children will not be randomised until 2\u00a0weeks after enrolment. Prior to 2\u00a0weeks, all children, including those in the placebo arm, will receive a 3-day course of artemether-lumefantrine as part of standard in-hospital care, which will be started before discharge and completed at home after discharge. The duration of post-treatment prophylaxis in our previous trial with artemether-lumefantrine is about 2 to 3\u00a0weeks , and we The study will be conducted in hospitals in Kenya and Uganda located in areas with moderate to intense malaria transmission , 16. TheThe inclusion and exclusion criteria for pre-study screening, enrolment and for randomisation 2\u00a0weeks later are described in the following sections.Haemoglobin <\u20095.0\u00a0g/dL or packed cell volume\u2009<\u200915%, or requirement for blood transfusion for other clinical reasons on or during admission to the hospitalAged less than 59.5\u00a0monthsBodyweight\u2009\u2265\u20095\u00a0kgResident in catchment areaRecognised specific other cause of severe anaemia, e.g. trauma, haematological malignancy, known bleeding disorderKnown sickle cell diseaseChild will reside for more than 25% of the 6\u00a0months study period (i.e. 6\u00a0weeks or more) outside of catchment areaFulfilled the pre-study screening eligibility criteriaAged less than 59.5\u00a0monthsClinically stable, able to take oral medicationSubject completed blood transfusion(s) or became clinically stable without transfusionAble to feed (for breastfeeding children) or eat (for older children)Provision of informed consent by parent or guardianPrevious enrolment in the present studyKnown hypersensitivity to study drugUse or known need at the time of enrolment for concomitant prohibited medication including drugs known to prolong the QTc interval during the 14-week PMC treatment period  or eat (for older children) and able to sit unaided Used DP since enrolmentUse or known need at the time of randomisation for concomitant prohibited medication  a 3-day course of artemether-lumefantrine regardless of whether they were admitted with severe malarial anaemia or severe anaemia without evidence of malaria.The study will use a Good manufacturing practices (GMP) formulation of artemether-lumefantrine . The recommended treatment is a six-dose regimen over a 3-day period with dosing per bodyweight following WHO dosing recommendations as provided for in the latest WHO malaria treatment guidelines  (see AddThe study will use the Eurartesim\u00ae brand of DP from Alfasigma (formerly Sigma Tau), Italy, a co-formulated tablet containing 40\u00a0mg dihydroartemisinin and 320\u00a0mg piperaquine phosphate or as 20/160 (paediatric formulation). Dosing will be per bodyweight according to the schedule recommended by the current WHO guidelines  will be according to local  or national guidelines and therefore not subject to this study. Treatment for malaria in both Kenya and Uganda conforms with the current WHO malaria treatment guidelines , which iAll children will receive 28\u00a0days of iron and folate supplementation at 2\u00a0weeks post-discharge as part of routine care for severe anaemia. A standardised prophylactic dose of iron supplementation (about 2\u00a0mg/kg) will be given as monotherapy or as part of the fixed-dose formulation with folic acid .Readmission due to severe malaria  by 26\u00a0weeks from randomisationReadmissions due to severe anaemia  by 26\u00a0weeks from randomisationReadmission due to severe malarial anaemia  by 26\u00a0weeks from randomisationReadmission due to severe anaemia or severe malaria (composite outcome) by 26\u00a0weeks from randomisationAll-cause mortality by 26\u00a0weeks from randomisationAll-cause hospital readmission by 26\u00a0weeks from randomisationClinic visits because of smear- or malaria rapid diagnostic test (RDT)-confirmed non-severe malaria by 26\u00a0weeks from randomisationReadmission due to severe malaria-specific anaemia  by 26\u00a0weeks from randomisationReadmission due to severe disease other than severe anaemia and severe malaria by 26\u00a0weeks from randomisationNon-severe all-cause sick-child clinic visits by 26\u00a0weeks from randomisationNon-malaria sick-child clinic visits by 26\u00a0weeks from randomisationMalaria infection at 26\u00a0weeksHb at 26\u00a0weeksAny anaemia (Hb\u2009<\u200911\u00a0g/dL), mild anaemia (Hb 8.0\u201310.99\u00a0g/dL), moderate anaemia (Hb 5.0\u20137.99\u00a0g/dL) and severe anaemia (Hb\u2009<\u20095\u00a0g/dL) at 26\u00a0weeksWeight-for-age, height-for-age and height-for-weight Z-scores (standard deviation [SD] scores of reference population) at 26\u00a0weeksSerious adverse events, excluding primary and secondary efficacy outcomes, by 26\u00a0weeks from randomisationSerious adverse events within 7\u00a0days after the start of each course of PMC, excluding primary and secondary efficacy outcomesAdverse events by 26\u00a0weeks from randomisationAdverse events within 7\u00a0days after the start of each course of PMCQTc prolongation measured by electrocardiogram (ECG) 4\u20136\u00a0h after third dose of each course (in a subset of patients)The study timelines consist of an in-patient pre-study screening period while the patient is acutely ill visit 1), followed by a screening, consent and enrolment visit visit 2). During the convalescence phase in the hospital, patients receive artemether-lumefantrine (visit 3) prior to discharge. The patient returns to the study clinic 14\u00a0days later (visit 4) for randomisation. Home treatment visits are made at 6 (visit 5) and 10 (visit 6) weeks. The PMC period starts at 2\u00a0weeks and ends at 14\u00a0weeks, but participants receive passive follow-up for an additional 12\u00a0weeks and are then seen at 26\u00a0weeks (visit 7) for an end-of-study assessment  and filter paper dry blood spot (DBS) for parasite genetics are obtained. Verbal autopsy is conducted for children who die at home during the follow-up period. Adverse events and vital status are assessed during all scheduled or unscheduled visits.The initial estimate of the required sample size was 2212 children (1106 per arm) across both countries pooled. This estimate was designed to detect a 30% reduction in the incidence rate of the composite primary outcome  from 469 per 1000 child years in the control arm to 328 per 1000 child years in the intervention arm , which allowed for one interim analysis and 15.7% loss to follow-up. For these estimations, we assumed an average pooled event rate of 399 per 1000 child years across the two arms, with 328 and 469 events per 1000 child years in the intervention and control arms respectively (Rate Ratio (RR)\u2009=\u20090.70). We based this assumption on observations in western Kenya  and Malawi . HoweverFollowing recommendations from the Data Monitoring and Ethics Committee (DMEC) and the Trial Steering Committee (TSC), a blinded interim sample size re-estimation was conducted to take into account the lower than expected rate of loss to follow-up and the higher than expected pooled incidence rate of the composite primary endpoint . This was favoured over an interim analysis, because the available funding did not allow an extension of the recruitment period, even if the results of any interim analysis had suggested that this would be required.The revised sample size calculations show that a total sample size of 1040 children (520 per arm) is required to detect a 25% reduction in the incidence of the composite primary outcome from 1152 per 1000 child years (530 events per 1000 children) in the control arm to 864 per 1000 child years (398 per 1000 children) in the intervention arm , allowing for 10% loss to follow-up. The same sample size also provides 90% power to detect a 28.7% reduction in the primary endpoint from 1152 to 822 events per 1000 child years.Eligible children are randomly assigned 1:1) to either PMC-DP or placebo by a computer-generated randomisation schedule stratified by weight (per DP dosing schedule) and study site using permuted blocks of random sizes . The envelopes containing active DP or placebo look identical, and the appearance and consistency of the tablets are also identical.Hb is measured using HemoCue 201  photometers. Thick and thin blood films for parasite counts are obtained and examined. The films are read by two independent microscopists by counting any malaria parasites against 200 high-power fields before a slide is declared negative . Point-oA detailed study statistical analytical plan for the final analysis that will supersede the study protocol will be developed during the study before the unblinding of data.The intention-to-treat (ITT) population is defined as all randomised subjects allocated to one of the two treatment arms and will be analysed in the group to which they were randomised, regardless of the type (placebo or active PMC) or number of courses received. The per-protocol (PP) population is a subset of the ITT population, excluding participants with major protocol deviations.Every effort is being made to minimise the amount of missing data in the trial, and whenever possible, information on the reason for missing data is obtained. No adjustments will be made for missing outcome data, but missing data may be imputed for covariates.Primary analysis will be by ITT and will include all primary endpoint events (i.e. first and repeat events). The follow-up time will be measured as the time in days from the date of randomisation to the end of follow-up (around 26\u00a0weeks), death or drop-out. The incidence rate will be calculated per arm and the incidence rate ratio  and 95% confidence interval (CI)estimated using Poisson regression models with treatment (as randomised) as the only covariate. The results will also be expressed as the relative rate reduction (RRR) (95% CI).We will use stratified analysis to assess to what extent the effect of the intervention on the primary outcome is influenced by country, demographic parameters , clinical parameters, malaria transmission variables , season, insecticide-treated nets use, site), time of assessment and potential intervention modifiers. Because we did not power the study for sub-group analyses, we will interpret the results of the sub-group analysis cautiously. No adjustment will be made for multiple comparisons.A number of sensitivity analyses will be conducted to assess the robustness of the primary endpoint analysis. These include analysis of the PP subject population and a covariate adjusted analysis. Other regression models will also be explored. Additional post hoc analyses may also be conducted if deemed appropriate. In addition, we will compare the results of the covariate-adjusted analyses with and without imputation for missing values for covariate values at baseline.Adverse events and serious adverse events are monitored, managed and recorded during the study. They will be recorded and tabulated for each treatment arm, overall, and per body system. Treatment emergent adverse events are defined as adverse events that had an onset day on or after the day of the first dose of study medication. No formal statistical testing will be undertaken. Enrolled children who are clinically unstable\u00a02\u00a0weeks post-discharge (i.e. at the time eligibility is assessed for randomisation) and/or have rebound severe anaemia are re-admitted and become eligible for randomisation if they fulfil the entry criteria 2 weeks after the subsequent discharge.All-cause mortality will be assessed during visits 4 (2\u00a0weeks), 5 (6\u00a0weeks) and 6 (10\u00a0weeks) and at 18\u00a0weeks (by phone) and during the end-of-study assessment at 26\u00a0weeks.These readmissions will be assessed through passive case detection as well as a questionnaire administered during visits 4\u20137 at 2, 6, 10 and 26\u00a0weeks and during unscheduled sick visits. Details of admissions and treatment that the participants received are recorded including malaria diagnostic test results and use of antimalarials to allow for differentiation between malaria, severe anaemia and other syndromes.Secondary efficacy outcomes include all-cause and malaria-specific clinic visits. They will be assessed through passive case detection as well as questionnaires administered during visits 4\u20137 at 2, 6, 10 and 26\u00a0weeks and during unscheduled sick visits. Details of clinic visits are recorded including malaria diagnosis results to allow for differentiation between malaria and non-malaria clinic visits.We will adhere to the International Conference on Harmonisation (ICH) good clinical practice (GCP) principles in recording, reporting and managing adverse events and serious adverse events for all participants in both arms . This strategy builds on existing approaches used for seasonal malaria chemoprevention in west Africa and experience with IPT in pregnant women and infants , 49. ShoRecruitment started in May 2016 and is ongoing. Unblinding and analysis will begin after recruitment and follow-up are completed and the database has been completed, cleaned and locked.Additional file 1:Full study protocol (including SPIRIT figure): v4.0, dated 06 Feb 2018. (PDF 3510 kb)Additional file 2:Ethics approvals: KEMRI, SOMREC, LSTM, REK vest and CDC. (ZIP 1940 kb)"}
+{"text": "Follow-up for 28\u201342 days is recommended by the World Health Organization to assess antimalarial drug efficacy for nonpregnant populations. This study aimed to determine the optimal duration for pregnant women, as no specific guidance currently exists.The distributions of time to recrudescence (treatment failure), confirmed by polymerase chain reaction genotyping for different antimalarial drugs in pregnancy, were analyzed by accelerated failure time models using secondary data on microscopically confirmed recurrent falciparum malaria collected in prospective studies on the Thailand\u2013Myanmar border between 1994 and 2010.Plasmodium falciparum recrudescences were identified as eligible. With log-logistic models adjusted for baseline parasitemia, the predicted cumulative proportions of all the recrudescences that were detected by 28 days were 70%  for quinine monotherapy (n = 295), 66%  for artesunate monotherapy (n = 43), 62%  for artemether\u2013lumefantrine , 46%  for artesunate with clindamycin (n = 19), and 34%  for dihydroartemisinin\u2013piperaquine . Corresponding figures by day 42 were 89%  for AL and 71%  for DP. Follow-up for 63 days was predicted to detect \u226595% of all recrudescence, except for DP.Of 946 paired isolates from 703 women, the median duration of follow-up for each genotyped recurrence (interquartile range) was 129 (83\u2013174) days, with 429 polymerase chain reaction\u2013confirmed recrudescent. Five different treatments were evaluated, and 382 In low-transmission settings, antimalarial drug efficacy assessments in pregnancy require longer follow-up than for nonpregnant populations. Plasmodium falciparum malaria dramatically [Effective antimalarial medicines are required to limit the adverse effects of malaria in pregnancy, which are deleterious to both the mother and fetus. Global implementation of highly efficacious artemisinin-based combination therapies (ACT) has changed the treatment of atically . The effatically . A decliP. falciparum malaria [According to the current WHO guidelines, in evaluating the efficacy of antimalarial treatment, patients should be followed up actively with clinical and parasitological assessments by microscopy to detect recurrence of  malaria . Recurre malaria . The cur malaria .Currently, there is no standardized guideline for assessing antimalarial efficacy in pregnancy, and the guideline for nonpregnant patients is frequently adopted , 6. BothP. falciparum infections in pregnancy in a cohort followed until delivery and to predict what proportion of recrudescent infections would be detected by days 28, 42, and 63 for each treatment, after adjusting for other risk factors.The objective of this study was to characterize the factors that affect the time to recrudescence of msp1, msp2, and glurp) was done for P. falciparum recurrences regardless of the intervening interval or any intercalated Plasmodium vivax infections. If all 3-locus genotypes were the same before and after the treatment, the recurrence was considered a recrudescence [This study included secondary data collected prospectively in treatment efficacy studies on uncomplicated falciparum malaria in pregnancy , 14\u201321. descence . This andescence , 25.Recurrent malaria episodes  confirmed by PCR genotyping as recrudescence in pregnant women in any trimester who completed a quinine- or artemisinin-based treatment were included in the analysis. Only late treatment failure  , which rFor each treatment group, baseline characteristics were described as median and range for continuous variables and as proportions for categorical variables.To model the time to treatment failure, episodes of PCR-confirmed recrudescence were analyzed by accelerated failure time (AFT) models . These were compared without any covariates using the Akaike information criterion (AIC) .P <.05 by the Wald test in univariable analysis were all included in the multivariable model. Two variables, namely treatment group and baseline asexual parasite densities, were always included in the multivariable model as they were known a priori to affect recrudescence risk. P. vivax intercalated infections preceding PCR-confirmed P. falciparum recrudescence were included in the analysis but regarded as being censored on the day of P. vivax infection. A sensitivity analysis without censoring of P. vivax infections was also conducted. Each episode was regarded as unique in the analysis. A complete case analysis was conducted.The covariates with As a sensitivity analysis, flexible parametric models with 0\u20132 knots were constructed , but theCumulative probability densities with commonly used follow-up durations  were predicted using the final model. Probability density curves based on the final model were drawn for each drug to assess the distribution of recrudescence over time. Statistical analyses were done using R  and Stata MP 15.1 .Ethics approval for retrospective analysis of anonymized secondary data at SMRU was granted by the Oxford Tropical Research Ethics Committee (OxTREC 28-09) and the Tak Community Advisory Board (TCAB-4/1/2015).Of 40 662 pregnant women followed in this prospective cohort between December 1994 and January 2010, 3969 (10%) had at least 1 episode of falciparum malaria . During P. falciparum mono infections and the remainder were mixed infections with P. vivax  were P. vivax . The geoThere was no apparent difference in baseline characteristics among the treatment groups except for estimated gestational age and history of antimalarial use . DespiteP. vivax co-infection showed shorter times to recrudescence, neither fulfilled the inclusion criteria for multivariable models (P < .05).Among the different AFT models, a log-logistic model was chosen for the final analysis . In the In the multivariable analysis, the type of treatment was associated with time to recrudescence adjusted by baseline parasitemia. The adjusted interval to recrudescence compared with that after quinine was 1.05  times longer after AS, 1.09  times after AL, 1.29  times after AC, and 1.48  times after DP.10 scale. The modal interval to recrudescence was 20 days for quinine, 21 for AS and AL, 25 for AC, and 29 days for DP.Based on the final multivariable model, the predicted probability density functions  and the The proportion (cumulative probability) of total treatment failures detected by the end of currently recommended follow-up durations was derived from the final model . The preP. falciparum recrudescences detected by a model without P. vivax infection censoring and by a flexible parametric model were similar to the predictions of the final AFT model  times after mefloquine than after quinine. The predictions for other treatments were similar to those by the final AFT model .Highly efficacious treatment can limit the cumulative deleterious impact of recurrent malaria during pregnancy on the mother and fetus. This demands correct assessment of treatment efficacy, which in turn requires an adequate length of follow-up. This large study from an area of low and seasonal transmission suggests that recommended durations of follow-up to assess antimalarial drug efficacy for nonpregnant populations  are too short for pregnant populations.The duration of follow-up in a survival analysis needs to be long enough to capture the majority of events  so that appropriate statistical analysis can be performed , 28. ThiDrug elimination rate  is the main factor determining the interval to recrudescence . Immunity against malaria affects the survival and growth of parasites independently of the antimalarial drug . The prePrevious studies on nonpregnant populations, which assumed that 63 days of follow-up captured all recrudescences, concluded that patients should be followed up for at least 28 days for antimalarial drugs with a short half-life , for 42 days for antimalarial drugs with an intermediate half-life , and for 63 days for antimalarial drugs with a long half-life  , 28.These recommendations appear insufficient for pregnant women, at least in a low-transmission setting. This study shows that in pregnant women, a 28-day follow-up can miss \u226530% of the recrudescences for quinine or AS. For AL, 11%  of recrudescences are predicted to be missed with 42-day follow-up. For DP, around 10% of recrudescences are predicted to occur after 63 days, although in this cohort this prediction is based on very small observations (n = 6). Under the currently recommended dosing regimens, this study suggests longer follow-up periods, such as 53  days for AL and 72  days for DP, to capture 95% of all recrudescences in pregnancy. In the context of antenatal care, this length of time is shorter than 1 trimester and should not be too burdensome. This study did not extend to the postpartum period, and the rapidity of reversion to the nonpregnant adult therapeutic response is unknown.One previous study simulated the time to recrudescence based solely on drug half-life . In thatThe time to recrudescence was longer in AC than expected from the drug half-lives of artesunate (<1 hour) and clindamycin (2\u20134 hours). However, AC, AS, and quinine were given over 7 days, whereas other treatments (ACTs) were given over 3 days. Another possibility is that the higher efficacy achieved by adding clindamycin reduced the residual parasite burden. Similarly, an additional analysis including mefloquine monotherapy, which was used when high-grade resistance to mefloquine was already prevalent and well described in the nonpregnant population in this area , shows tThis is the first study to use AFT models to analyze the time to treatment failure in malaria. There are several reasons why the AFT model is better suited to this purpose than the Cox proportional hazards model. Biologically, the assumption of the AFT models is more plausible; the distribution of time to recrudescence is shorter (or longer) depending on factors such as the half-life of the drug. Clinically, the AFT models analyze the times to events directly (rather than hazard ratios); thus the interpretation of the output is clinically relevant and meaningful . AFT modDespite these advantages, there are also limitations. First, the number of observations included in this analysis was small for all drugs except quinine. However, modeling enables prediction of the temporal distributions for drugs with fewer observations based on the distributions for other drugs with more observations but with a proportional change in the time to events, as discussed above. The effects of covariates  might be different for each drug, but this was not assessed fully.Second, this study was based on the data collected on the Thailand\u2013Myanmar border, where falciparum malaria transmission is low and unstable. This limits generalizability. Nonetheless, available evidence from other studies is scarce . There aThese predictions need to be tested in high transmission areas, although this becomes more complex because of a higher risk of misclassification with genotyping and a higher level of immunity against malaria. In high-transmission areas, concomitant infection with multiple discrete genotypes is common  and the The possibility of predicting the \u201ctrue\u201d efficacy of antimalarial treatments by extrapolating the observed efficacy with a shorter follow-up needs cautious consideration. As was implied in this analysis, the shorter the duration of follow-up, the more uncertain the prediction of \u201ctrue\u201d efficacy becomes. This uncertainty becomes greater if the number of recrudescences is small. Efforts should be made to capture most of the recrudescences so that accurate assessments of the failure rate can be made.Insufficient length of follow-up in antimalarial therapeutic assessments underestimates recrudescence rates and thus overestimates treatment efficacy. This can lead to continuation of a less efficacious drug or insufficient dosage, resulting in more treatment failures at the individual level and a higher risk of selecting resistance. This study suggests that the currently recommended length of follow-up of 28\u201342 days in therapeutic assessments may be insufficient for pregnant women, at least in low-transmission areas. Follow-up of 63 days should be considered for AL, and even longer might be needed for DP. The optimal duration of follow-up for pregnant women, particularly in high-transmission settings, requires further investigation.Open Forum Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at ofz264_suppl_supplementary_materialClick here for additional data file."}
+{"text": "Plasmodium vivax infections in pregnancy are associated with maternal anaemia and increased adverse perinatal outcomes. Providing radical cure for women with 8-aminoquinolines  is hindered by gender-specific complexities.The Sustainable Development Goals (SDG) call for increased gender equity and reduction in malaria-related mortality and morbidity. P. vivax episode, severe intra-uterine growth restriction with no other identifiable cause and induction to reduce the risk of stillbirth. At 5\u00a0months postpartum a qualitative glucose-6-phosphate dehydrogenase (G6PD) point-of-care test was normal and radical cure with primaquine was prescribed to the mother. A 33% fractional decrease in haematocrit on day 7 of primaquine led to further testing which showed intermediate phenotypic G6PD activity; the G6PD genotype could not be identified. Her infant daughter was well throughout maternal treatment and found to be heterozygous for Mahidol variant.A symptomatic episode of vivax malaria at 18\u00a0weeks of gestation in a primigravid woman was associated with maternal anaemia, a recurrent asymptomatic P. vivax-endemic settings.Adverse effects of vivax malaria in pregnancy, ineligibility of radical cure for pregnant and postpartum women, and difficulties in diagnosing intermediate levels of G6PD activity multiplied morbidity in this woman. Steps towards meeting the SDG include prevention of malaria in pregnancy, reducing unnecessary exclusion of women from radical cure, and accessible quantitative G6PD screening in  In other words, one of the principles of gender equity requires that services for women not simply mirror services for men but differ from services for men in situations where women\u2019s needs or disease processes differ  and the infant\u2019s HCT was 36%; both blood slides were negative for Daily weight-based treatment with PMQ (0.5\u00a0mg/kg/day) for 14\u00a0days total was prescribed; the 4 tablets daily translated into an actual dose of 0.52\u00a0mg/kg/day (weight 58\u00a0kg). The first dose was supervised in the clinic. At follow-up on the 7th day none of the commonly observed PMQ side effects, such as abdominal pain, nausea or vomiting was reported  but the Biochemistry showed normal kidney function and, apart from a mildly increased direct bilirubin  and alkaline phosphatase (132 U/L [33\u201396]), normal liver function tests. At 236 U/L (115\u2013221) lactate dehydrogenase was slightly elevated. The direct Coombs test was negative and there was no haemoglobinuria. Otherwise, history and physical examination were unremarkable.Given the clinical picture, G6PD deficiency was suspected despite the normal G6PD FST result as the FST has poor sensitivity to detect intermediate G6PD activity levels. The G6PD activity was then quantified by spectrophotometry and was found to be normal at 7.7\u00a0IU/gHb (population median: 7.51\u00a0IU/gHb) , suggestThe patient\u2019s body weight was rechecked and found to be 56.5\u00a0kg. With no signs of severe haemolysis and no evidence of G6PD deficiency, the PMQ course was continued under supervision at a corrected dose of 3.5 tablets daily . The treatment was well tolerated from this point onwards and the woman\u2019s clinical condition improved. To treat the anaemia, the patient was provided with a 28-day course of ferrous sulfate, folic acid and vitamin B12. Since the mother was still breastfeeding, the infant\u2019s HCT was also followed and remained relatively stable . On day 12 of PMQ, the maternal HCT had increased to 30%, symptoms had resolved, and the patient requested to be discharged home. 11\u00a0weeks later, the HCT was 37%, and the patient was well. The child\u2019s vaccination schedule was completed, and the 6-month motor milestones were normal.Approximately 1\u00a0year after delivery, the mother returned with the infant for follow-up as part of the trial described earlier. Since the previous quantification of the G6PD activity during the haemolytic episode was considered unreliable (due to\u00a0increased reticulocyte count and low Hb level), the staff offered the woman to repeat quantitative G6PD testing while in steady state (HCT 36.1%). This time the G6PD activity was found to be 4.62\u00a0IU/gHb, which corresponds to 62% of the population mean and is highly suggestive of G6PD heterozygosity . FollowiRecognizing gender-specific aspects of disease acquisition, detection, treatment options, and response to treatment can help ensure that health policies are effective and equitable. As the historical legacy of gender-based inequality is deeply embedded in medical research and the health sector, action to prevent undue gender-specific disparities in health outcomes is an ethical imperative and mandated in the SDG .P. vivax in pregnancy, multiple layers of a gender-specific morbidity occur  without adverse effects on the infant. Given the pharmacokinetics of primaquine during lactation and the dose-dependent nature of primaquine-induced haemolysis, it is very unlikely that the clinically insignificant 8% fractional reduction in HCT seen in this infant is due to drug exposure via breastmilk. However, the fact that she was classified as G6PD normal by FST further highlights the need for improved G6PD testing modalities.Improving care and prevention of malaria in pregnancy, reducing unnecessary exclusion of women from radical cure, and providing quantitative G6PD screening that is as accurate for females as it is for males are achievable steps towards the SDG for a more equitable and malaria-free world."}
+{"text": "For many years, malaria was a major life-threatening parasitic infection in Iran. Although malaria elimination program is implementing in the country, still some cases annually are reported from malaria-endemic areas.vivax malaria and falciparum malaria infected patients were enrolled in the study from 2013\u20132014. All the cases were selected according to criteria of the WHO guideline for in vivo drug sensitivity tests in malaria parasites. Evaluation of drug sensitivity test was conducted with some modifications.This study was conducted in five malaria endemic districts of Sistan and Baluchistan Province, southeastern Iran, neighboring Afghanistan and Pakistan countries. Overall, 170 and 38 The patients with vivax malaria responded to the regimen of chloroquine in 37.4(\u00b115.9), 40(\u00b113.8) and 42(\u00b117.7) h for Pakistani, Iranian and Afghani nationalities respectively based on MPCT evaluation. The results showed a considerable difference between them and Iranian subjects. MPCT for the patients with falciparum malaria was calculated as 28(\u00b118.05), 26(\u00b112.03) and 36(\u00b116.9) h for Iranian, Pakistani and Afghani nationalities respectively. There was a marginally significant difference between Afghani and other nationalities and between males and females.P. vivax showed that the parasite became more sensitive to chloroquine than previous years in studied areas.Treatment of all the patients resulted in ACPR and MPCT of  P. vivax, P. falciparum and mixed cases respectively  in Iran, 1257 cases were recorded in 2014 including 1116 (88.8%), 117 (9.3%) and 23 (1.9%) ectively .P. falciparum and more recently in some strains of P. vivax , a 4-aminoquinoline antimalarial, is the drug of choice for the treatment of he world ,4, but sP. vivax  can makeectively . Such stectively \u201311.P. vivax and P. falciparum to chloroquine and artesunate-fansidar, currently used antimalarial drugs, after a few years of the last in vivo evaluating antimalarial drugs in Iran.This study was proposed to evaluate the susceptibility of P. vivax and P. falciparum were prevalent species in the areas with dominance of P. vivax  of 3 h for mean of parasite clearance time (MPCT) of P. vivax] were registered for the study, but 33 (19.4%) of them did not complete the study. All 38-falciparum malaria infected patient those referred to health centers of the districts were considered for the study. All patients aged more than 6 months and had the following criteria of WHO guideline: malaria positive P. vivax or P. falciparum with parasitaemia of 250\u2013100000 parasites/\u03bcl blood, axillary temperature equal or more than 37.5 \u00b0C or a history of fever during the last 24 h, ability to come for the stipulated follow-up visit. Patients with following criteria were excluded from the study; severe malnutrition, pregnancy, febrile disease other than malaria or other known underlying chronic or severe disease, severe or complicated malaria cases, children aged under 6 months, mixed or mono infections with another Plasmodium species detected by microscopy, regular medication which may interfere with antimalarial pharmacokinetics and history of hypersensitivity reactions or contraindications to any of the medicine(s) being tested or used as alternative treatment(s)  , 12. MosAt the enrollment time, a consent form was signed by each of the patients or their guardians. Implementation of the study was approved by TUMS Research Committee under the code number 92-02-160-23635.Study techniques: For evaluation of drug sensitivity in P. vivax and P. falciparum, two in vivo tests were employed according to the WHO guideline with some modifications (ications  as folloP. vivax: The included patients were treated with 25mg/kg of chloroquine (CQ)  divided into three doses over 3 d . Moreover, to achieve a radical cure amount to 0.75 mg/kg primaquine was administered weekly for 8 wk starting from day 2. Prior to treating the patients, a Giemsa stained thick and thin blood smear was prepared from each of the patients. The asexual parasites were counted against at least 200 WBCs on days 0, 1, 2, 3, 4, 7, 14, 21& 28 in the thick blood smears and then converted to the number of parasites per microliter of blood. Results of the counting, to understand drug sensitivity, were addressed as a mean of parasite clearance time (MPCT) and analyzed by means of a Microsoft Excel. Item of parasite clearance time (PCT) was employed for odd case.P. falciparum: Those patients with uncomplicated falciparum malaria were treated with a standard regimen of artesunate\u2013fansidar  combination including 12mg/kg artesunate over 3 d and 25mg/kg fansidar on day 0. The patients were given clinical examinations on days 0, 1, 2, 3, 4, 7, 14, 21& 28. The parasites were counted in thick blood smear as mentioned in P. vivax section for each patient on the same days. Therapeutic response to standard regimen of treatment by day 28 of follow up would be classified as early treatment failure (ETF), late treatment failure (LTF) [including late clinical failure (LCF) and late parasitological failure (LPF)] and adequate clinical and parasitological response (ACPR) , 40 (\u00b113.8) and 42(\u00b117.7) h for Pakistani, Iranian and Afghani nationalities respectively based on MPCT evaluation. P. vivax parasite was found in none of the patients, blood smears after day 3 at follow-up until day 28. MPCT in the blood of females is different from males, but not significant  (=\u22120.442) . The resP. falciparum: All of the 38-falciparum malaria-infected patients received standard artesunate-fansidar combination treatment after diagnosing the infection as mentioned in materials and methods section. MPCT for the patients with falciparum malaria was calculated as 28(\u00b118.05), 26(\u00b112.03) and 36(\u00b116.9) h for Iranian, Pakistani and Afghani nationalities respectively. There were some hours differences between Afghani and other nationalities  and also between males and females  with 30.7 h (1.3 d) and 24 h (1 day) respectively in the MPCT, but not significant . In a previous study conducted in Bandar-Abbas district of Hormozgan Province in southeastern Iran MPCT of P. vivax was calculated as 61.7 h (2.6 d) (P. vivax was recorded as 61.6 h (2.6d)  . Moreoveh (2.6d) . Compariern Iran .P. vivax in Mehrestan showed the maximum time with 49.7 h (\u00b12.1 d) and the minimum time in Sarbaz and Saravan with 36 h (1.5d). Interestingly MPCT of the parasite in Chabahar district, a malarious area, decreased from 58 in previous study (P. vivax in the areas (Among the studied districts, MPCT of us study  to 34.7 he areas \u201323.P. vivax in this study shows that the parasite has become more sensitive to chloroquine than previous years in studied areas, indicating that decrease of vivax malaria infection resulted in utilizing less antimalarial drugs against P. vivax strains and consequently less pressure of the drugs on the parasites. In this situation, the parasites become more sensitive to the previously used antimalarials due to being omitted many of the resistant mutants.MPCT of"}
+{"text": "Plasmodium sp., that still prevalence in some part of Indonesia. District of Pesawaran is one of malaria endemic area in the Province of Lampung. The purpose of this study was to evaluate the efficacy of the ACT treatment in the District of Pesawaran Province of Lampung, Indonesia from Dec 2012 to Jul 2013 and the genetic variation of the Plasmodium falciparum also studied.Malaria is an infectious disease caused by Puskesmas). DNA isolation was done with QIAmp DNA Mini Kit. Amplification of PfMDR1, MSP1, and MSP2 genes was done with appropriate forward and reverse primer and procedures optimized first. PCR Product of PfMDR1 gene was prepared for sequencing. Data analysis was done with MEGA 6 software.This study was observational analytic study of falciparum malaria patients treated with ACT and primaquine (DHP-PQ and AAQ-PQ) at Hanura Primary Health Centre  are low.The results of this research are DHP-PQ effectiveness was still wellness among falciparum malaria patients in District of Pesawaran, Province of Lampung, Indonesia. There is Single-nucleotide mutation of Therapy of DHP-PQ as an antimalarial falciparum in Pesawaran District, Lampung, Indonesia is still good. The genetic variation found was the SNP on the N86Y PfMDR1 gene, with dominant allele MAD20 and 3D7. Plasmodium falciparum is one of the most predominant causes of malaria diseases in the district of Pesawaran, the province of Lampung, in the south part of Sumatera island of Indonesia  gene is one of the transporter genes that play a role in regulating the pH in the food vacuole of Plasmodium. Polymorphisms in PfMDR1 gene has been shown to play a role in the change of Plasmodium susceptibility against amodiaquine, mefloquine, lumefantrine, halofantrine and artemisinin with different mechanism. Position N86 (wild-type) plays an important role in the increased resistance to arylaminoalcohol quinolines, such as mefloquine, lumefantrine, and halofantrine, while the position 86Y (mutant) plays an important role in the increased resistance to 4-aminoquinoline such as chloroquine and amodiaquine. Codon 86 of PfMDR1 gene, have an important role of fluids efflux, including drugs, from the parasite food vacuole into the cytoplasm  value > 1)  is one oytoplasm .Plasmodium sp. also affects its susceptibility to antimalarials. To investigate genetic variation, MSP1, MSP2 and GLURP genes were examined. These genetic variants are also associated with the nature of P. falciparum. On examination of this genetic variation will show K1, MD20 and RO33 alleles based on MSP1 gene and FC27 and 3D7 alleles based on MSP1 gene. Genetic variation data on P. falciparum in Pesawaran District has not been reported. Therefore, it is important to study genetic variation.In addition, genetic variation of PfMDR1, and genetic variation of P. falciparum will also bee studied.The purposes of this study were evaluate the efficacy of the ACT treatment in the District of Pesawaran; evaluate the nucleic acid changes of the This study was conducted at Pesawaran District, Lampung Province, Indonesia. Hanura public health center (PHC) which has the highest API  was selected  for parasite DNA analysis. After taking blood samples, the research subjects received ACT treatment according to the standard of malaria treatment issued by the Indonesian Ministry of Health (Day 0). A cohort of blood samples collection for evaluation of parasitemia was done on Day 1, 2, 3, 7, 14, 21 and 28 after ACT administration. Evaluation of the treatment responses of each case was evaluated using standard protocol of the WHO guidelines ,14.The process of selecting research subjects was done by referring to the inclusion criteria according to the WHO protocol in 2009 . At the Dry blood spot on Whatman paper was cut and put into 1.5 ml Eppendorf tube and DNA was extracted using QIAmp DNA Mini kit from Qiagen. The DNA extraction was carried out in accordance with the standard protocol provided within the kit .PfMDR gene was amplified using publish primer sequences of Humphreys et al.  Faculty of Medicine, Gadjah Mada University, Yogyakarta, Indonesia and the Medical and Health Research Ethics Committee (MHREC) Faculty of Medicine, University of Lampung, Lampung, Indonesia .P. falciparum of D14 on microscopic examination, while the D28 has 2 positive P. falciparum. The P. falciparum positive at D14 and D28 are categorized as late treatment failure.Artemisinin-Base Combination Therapy drugs used in District of Pesawaran for treat of malaria patients were consists of two types such as Dihydroartemisinin Piperaquine (DHP) and Artesunate-Amodiaquine (AAQ). Both of these drugs were combined with Primaquine (PQ) 0.75 mg/kgbw single dose. P=0.024). Adequate Clinical and Parasitological Response (ACPR) are more common in subjects treated with DHP-PQ compared to subjects treated with AAQ-PQ. Combination AAQ-PQ has a high failure rate (25.00%) compared with DHP-PQ (5.26%)  .Microscopic examination on Day 0 showed that asexual stage parasite density varied between 1.000 to 111.440 parasites/\u03bcL blood. Microscopic examination on the first day after treatment, 73.68% of the study subjects who received DHP-PQ treatment and 95.83% of the study subjects who received AAQ-PQ treatment were not found asexual stadiums. The third day observation has not found any asexual stage in all study subjects. Parasite density tends to decrease from day to day after treatment at day 0. In this study has not found signs of a shift in parasite clearance time even though it has been found late treatment failure (possibility recrudescence).PfMDR1 gene performed on 50 blood samples from 62 samples showed a change in nucleotide base sequence in the fifty samples. The nucleotide base of Adenine (A) at position 256 turns into Thymine (T). The nucleotide encoding codon 86, which causes amino acid changes from Asparagine (N) becomes tyrosine (Y). The nucleotide number 551, which encodes codon 184 reported to have undergone a change in previous research, found no changes in this study.Molecular analysis of the MSP1 and MSP2 genes , RO33  and K1  alleles, whereas MSP2 gene was found by FC27  and 3D7  alleles. Blood samples from subjects that fail therapy, performed genotyping to analysis of multiplicity of infection (MOI). Seven blood samples of eight blood samples of subjects failed therapy successfully performed molecular analysis. Blood samples from failed therapy subjects were taken at D0 and when the parasites reappeared (D14 and D28). Subjects with adequate therapy were examined for genetic variation in 5 blood samples taken at D0 for comparison. Alleles found based on the MSP1 gene are MD20, RO33, and K1, whereas in the MSP2 gene found 2 alleles that are FC27 and 3D7. Multiplicity of infection (MOI) based on allele is low .Genetic analysis was continued by genotyping on P2 genes . The resArtemisinin-based Combination Therapy in PHC Hanura has been used since 2004 after the Ministry of Health decided ACT as the standard treatment of malaria. Within 10 years after the first use of ACT, it is possible there has been a decrease in effectiveness. According to WHO criteria, antimalarial drugs are not used anymore, if the treatment failure rate \u2265 10% (WHO 2010). Based on this, the effectiveness of the AAQ-PQ has been reduced compared with the DHP-PQ.P. falciparum takes 24\u201372 h to develop from the ring stage to mature schizont stage (Subjects with therapeutic response late parasite failure (LPF) mostly parasites appeared at day 14 (75%) and day 28 (25%). The parasite was not found on examination D 2, 3 and 7 and reappeared on D 14 or 28. These conditions indicate recrudescence has arisen. Recrudescence is a re-invention of the parasite due to persistence of the parasite in the blood, but there is a decrease so it is not detected in microscopic examination. nt stage . Levels nt stage \u201321. A dePfMDR1 genes. Polymorphism at codon N86Y cause amodiaquine is not effective in eliminating parasites. This condition makes the high treatment failure in patients who received AAQ-PQ (AAQ-PQ treatment failure is 24.00%). This is due to an increase in pH and reflux of food vacuole so amodiaquine concentration in the food vacuole will decrease. High pH of the food vacuole will influence the amodiaquine to inhibit polymerization hem. The results of this study are consistent with another research which states that a single mutation of codon N86Y can cause changes in the function of the transport of fluids and drugs (amodiaquine) significantly  plays an important role in the increased resistance to arilaminoalkohol quinolines, such as mefloquine, lumefantrine, and halofantrine, while the position 86Y (mutant) plays an important role in the increased resistance to 4-aminoquinoline such as chloroquine and amodiaquine . Piperaquine as artemisinin combination still gives a good effect as a long-acting drug. In short, a half-time of artemisinin causes the concentration of drug in the body rapidly declined after the third day of treatment. The role of parasite elimination will be taken over by piperaquine. This can be seen in subjects who received treatment with DHP-PQ, still give good therapeutic results with failure rate 4.55%, although it has been found polymorphisms in codon N86Y, DR1 gene , so the PfMDR1, MSP1 and MSP2 genes, the genetic conditions of P. falciparum isolate Pesawaran classified as low genetic diversity with dominant alleles were MAD20 based on MSP1 gene and 3D7 allele based on MSP2 gene. This study is in line with the research in South Sumatra, with the dominant allele result being MAD20 , Pakistan 1.25) and India (1.38) , 32, 34, and Indifalciparum in Pesawaran District, Lampung, Indonesia is still good. The genetic variation found was the SNP on the N86Y PfMDR1 gene, with dominant allele MAD20 and 3D7.Therapy of DHP-PQ as an antimalarial"}
+{"text": "Plasmodium falciparum malaria worldwide. The benefits of artenimol\u2013piperaquine in children have been validated in endemic countries but experience remains limited in cases of imported malaria.Although malaria remains one of the major public health threats in inter-tropical areas, there is limited understanding of imported malaria in children by paediatricians and emergency practitioners in non-endemic countries, often resulting in misdiagnosis and inadequate treatment. Moreover, classical treatments  are limited either by lengthy treatment courses or by side effects. Since 2010, the World Health Organization (WHO) has recommended the use of oral artemisinin-based combination therapy for the treatment of uncomplicated P. falciparum positive on thin or thick blood smear; and the absence of WHO-defined features of severity.This prospective observational study in routine paediatric care took place at the Emergency Department, Robert-Debr\u00e9 Hospital  from September 2012 to December 2014. Tolerance and efficacy of artenimol\u2013piperaquine in children presenting with the following inclusion criteria were assessed: Among 83 children included in this study, treatment with artenimol\u2013piperaquine was successful in 82 children (98.8%). None of the adverse events were severe and all were considered mild with no significant clinical impact. This also applied to cardiological adverse events despite a significant increase of the mean post-treatment QTc interval.Artenimol\u2013piperaquine displays a satisfying efficacy and tolerance profile as a first-line treatment for children with imported uncomplicated falciparum malaria and only necessitates three once-daily oral intakes of the medication. Comparative studies versus artemether-lumefantrine or atovaquone-proguanil would be useful to confirm the results of this study. Plasmodium species, transmitted to humans by the bite of infected female Anopheles mosquitoes. The main species known to infect humans are Plasmodium falciparum, which causes the majority of severe cases, Plasmodium vivax, Plasmodium malaria, Plasmodium ovale and Plasmodium knowlesi. Malaria remains one of the major public health threats in inter-tropical areas. Although the incidence of malaria has fallen since 2010, there is no significant progress in reducing malaria cases for the period 2015\u20132017 [Malaria is a febrile illness caused by the protozoan parasite P. falciparum resistance to conventional anti-malarial drugs and to improve treatment efficacy. Qinghao (Artemisia annua) has been used by Chinese botanists for many centuries for the treatment of fever and the active component of the plant (qinghaosu or artemisinin) was purified in China in 1972 [P. falciparum malaria worldwide [Resistance to artemisinin is a major challenge and vaccine development is still far from providing long-lasting benefits . In the  in 1972 . Since 2orldwide . With 52orldwide , 7. Arteorldwide , 9, the orldwide  and noneorldwide . The resP. falciparum positive thin or thick blood smear and the absence of World Health Organization (WHO)-defined features of severity. Parasitaemia count >\u20094%, when it was an isolated finding, was not considered a criterion of severity, as suggested by the 2007 French recommendations [This prospective observational cohort study in routine paediatric care took place in the Emergency Department, Robert-Debr\u00e9 Hospital  from September 2012 to December 2014. All children presenting with a fever, or recent history of fever, and returning from a malaria endemic country within the last 3\u00a0months were screened for malaria (thin and thick blood smear). UFM was defined by fever or a recent history of fever, a Severe malaria or non-falciparum malaria patients were not included in the study and were treated with intravenous artesunate or oral chloroquine, respectively. Patients treated with another drug than AP were not included in the study. Following the manufacturer\u2019s recommendations, children received the 40\u00a0mg/320\u00a0mg AP tablets (the only dosage available in France) according to body-weight  on an empty stomach. When children were unable to swallow tablets, they were crushed and then given in very sweet yoghurt. In one case, nasogastric tube was used for drug administration. If the child vomited less than half an hour or between half an hour and 1\u00a0h after ingestion, a complete dose or half of the dose was re-administered, respectively. Nurses observed the child for 1\u00a0h and, in the absence of vomiting, significant asthenia or any symptoms or reason  requiring hospitalization, the child was discharged home. At the time of hospital discharge, the parents were given the rest of the treatment free-of-charge in order to administer it to their child at home for 2\u00a0days, every 24\u00a0h, on an empty stomach.-test was performed to determine whether there was a statistically significant difference in mean QTc before and after starting treatment with AP. Post-treatment clinical and laboratory follow-up (including thin and thick smear) were undertaken first between day 3 and day 8 and on a second occasion on day 28. Parents were informed of the study and gave their consent. The study protocol was approved by the Robert-Debr\u00e9 Hospital (Assistance Publique-H\u00f4pitaux de Paris) ethical committee.A 12-lead electrocardiogram (EKG) was performed before starting AP and at the first follow-up visit. The QTc interval was calculated by an experienced cardio-rhythmologist. A paired TDuring the study period, malaria was diagnosed in 123 children, 3 of them had severe malaria and 15 had non-falciparum malaria. 22 children with UFM were treated with drugs other than AP and were not included in the study  and 11.67\u00a0g/dL  on admission and on D30, respectively. Nine children (10.84%) vomited after one of the doses; all of them repeated AP according to the protocol without vomiting again. It is not possible to determine whether vomiting was a drug adverse effect or malaria symptom, or a combination of both.Evolution of QTc intervals was analysed in children who had EKG both before and after treatment (72.3%). EKG post treatment was performed within 24\u00a0h of the end of the treatment in 80% of the cases; 20% were registered between 24 and 72\u00a0h after the end of the treatment. The average pre-treatment QTc was 393\u00a0ms\u2009\u00b1\u20092\u00a0s.d. whereas the average post-treatment QTc was 405\u00a0ms\u2009\u00b1\u20092\u00a0s.d. (p\u2009<\u20090.01), children being afebrile after treatment. Before treatment, one febrile child had a QTc at 447\u00a0ms but post-treatment QTc was normal at 435\u00a0ms with no fever. Six patients with a normal QTc pre-treatment (less than 440\u00a0ms), had a prolongation of the QTc to 450\u00a0ms in 5 cases and to 490\u00a0ms in one case, without symptoms.It should be noted that none of the 83 included children in this study were taking concomitant medication other than paracetamol and, therefore, the risk of interaction with artenimol piperaquine could be ruled out.P. falciparum malaria. One child had a recrudescence probably related to immediate vomiting of two doses given at home and was subsequently cured after receiving three additional doses of AP. In conclusion, AP reached the WHO-recommended efficacy (>\u200995%) in this population [In this prospective observational study in routine paediatric care in France, one course of AP was successful in 82/83 children (98.79%) with uncomplicated pulation .Adverse events were rare and in no children was it deemed necessary to switch to another anti-malarial drug. The aryl-amino-alcohol compounds, of which piperaquine is a member, can prolong the QTc interval. The inhibitory concentration 50 (IC50), an indirect measure of this risk, is the average serum concentration of the molecule required to block 50% of potassium channels during phase 3 repolarization of cardiac cells. The IC50 of piperaquine (0.11\u00a0\u00b5mol/L) is six times greater than that of halofantrine (0.018\u00a0\u00b5mol/L), known to demonstrate cardiotoxicity, making it less likely to induce marked, life-threatening prolongation of the QT interval. It is, however, lower than that of chloroquine (1\u00a0\u00b5mol/L) or lumefantrine (2.6\u00a0\u00b5mol/L) , which iIn this study, moderate anaemia was infrequent, and severe anaemia was rare , possibly due the predominance of older children (95.2%) over children <\u200924\u00a0months. Furthermore, despite a follow-up period lasting 22 to 56\u00a0days after treatment initiation, no episodes of severe haemolytic anaemia were reported unlike that which has been recently reported after treatment of severe malaria with intravenous artesunate  or with The D3, D7 and D28 WHO-recommended follow-up visits are difficult to implement in this particular patient group. This seems mainly due to the fact that, even when parents are considered reliable, almost all patients display complex family dynamics resulting in missed or delayed appointments. Follow-up phone calls are often necessary to rearrange visits. Nevertheless, we have been able to follow up all our children with two controls, between day 3 and day 8 and between day 22 and day 56, respectively. Only one child had no second control; his parents informed us by phone from Ivory Coast\u2014where the family lived\u2014that the child was clinically well 2\u00a0months after the end of the treatment. The overall follow-up rate at least 1\u00a0month after treatment was 98%.There are some limitations to this study. The first is a lack of comparison treatment arm of the study. In France, two artemisinin-based combinations are recommended as treatment for UFM, artemether\u2013lumefantrine (AL) and AP. 30 children with AL were treated in 2011\u20132013; all children were cured, and adverse events were rare. However, the marked difference in the number of cases in each group rendered it inappropriate to compare the two molecules. A randomized study comparing AL and AP would have been difficult to perform, especially in paediatric practice. Additionally, comparative randomized studies often imply selection bias. This is the reason why a cohort study design was chosen to assess the efficacy and safety of AP in children in real-life conditions. Based on years of experience acquired at the Robert Debr\u00e9 Emergency Department, it appears that AP displays two advantages over AL. First, only one dose is required daily, instead of two doses for AL, which sometimes obliges parents to wake the child during the night. Secondly, and more importantly, it is recommended to give AP on an empty stomach while AL must be taken with food to improve lumefantrine absorption, which is a limitation given that food ingestion in children with acute malaria may induce vomiting. A second potential limitation of this study is the timing of the second EKG recording, between day 3 and day 8 (period of the first scheduled follow-up visit) compared to day 2 in the majority of other studies. It could be assumed that due to the long half-life of piperaquine, the impact of this delay in QTc prolongation is probably not clinically significant.Imported malaria in children is a rare illness, whose initial management by both paediatricians and general practitioners is frequently sub-optimal, necessitating both a simple and well-tolerated treatment. Artemisinin-based oral combinations have markedly improved the treatment of malaria for children who live in or travel to endemic countries and are nowadays the reference first-line drugs. Artenimol\u2013piperaquine has the advantage of being taken without food and with a simple dosing schedule. Moreover, this study did not show any cardiotoxicity.Based on the high observed cure-rate and good safety profile, DP appears as a valuable first-line option to treat children in the context of imported malaria."}
+{"text": "In 2009, the Papua New Guinea (PNG) Department of Health adopted artemether\u2013lumefantrine (AL) and dihydroartemisinin\u2013piperaquine (DHA-PPQ) as the first- and second-line treatments for uncomplicated malaria, respectively. This study was conducted to assess the efficacy of both drugs following adoption of the new policy.Plasmodium falciparum or Plasmodium vivax mono-infections were enrolled, treated with AL or DHA-PPQ, and followed up for 42\u00a0days. Study endpoints were adequate clinical and parasitological response (ACPR) on days 28 and 42. The in vitro efficacy of anti-malarials and the prevalence of selected molecular markers of resistance were also determined.Between June 2012 and September 2014, a therapeutic efficacy study was conducted in East Sepik and Milne Bay Provinces of PNG in accordance with the standard World Health Organization (WHO) protocol for surveillance of anti-malarial drug efficacy. Patients\u2009\u2265\u20096\u00a0months of age with microscopy confirmed P. falciparum and 70 P. vivax cases were enrolled. The day-42 PCR-corrected ACPR for P. falciparum was 98.1% (104/106) for AL and 100% (135/135) for DHA-PPQ. The day-42 PCR-corrected ACPR for P. vivax was 79.0% (15/19) for AL and 92.3% (36/39) for DHA-PPQ. Day 3 parasite clearance of P. falciparum was 99.2% with AL and 100% with DHA-PPQ. In vitro testing of 96 samples revealed low susceptibility to chloroquine (34% of samples above IC50 threshold) but not to lumefantrine (0%). Molecular markers assessed in a sub-set of the study population indicated high rates of chloroquine resistance in P. falciparum  and in P. vivax .A total of 274 AL and DHA-PPQ were efficacious as first- and second-line treatments for uncomplicated malaria in PNG. Continued in vivo efficacy monitoring is warranted considering the threat of resistance to artemisinin and partner drugs in the region and scale-up of artemisinin-based combination therapy in PNG.The online version of this article (10.1186/s12936-018-2494-z) contains supplementary material, which is available to authorized users. Plasmodium falciparum and Plasmodium vivax parasite strains [P. falciparum and DHA-PPQ most efficacious against P. vivax [The widespread resistance to commonly used 4-aminoquinolines continues to impede malaria control strategies in malaria-endemic countries. For decades the anti-malarial drugs chloroquine and sulfadoxine-pyrimethamine were highly effective against falciparum malaria. However, due to the emergence of resistant  strains , the Wor strains . By admi strains . Moreove strains . A compaP. vivax . The couP. vivax .Unfortunately, there is increasing resistance of parasites to ACT in the Greater Mekong Region , 8, 9, a50) of a range of anti-malarial drugs used in PNG and investigating the prevalence of molecular markers associated with in vivo failures.A therapeutic efficacy study was conducted in PNG with the primary objective of assessing the in vivo efficacy of AL and DHA-PPQ after their introduction. Secondary objectives included measuring the in vitro efficacy  from 2012 to 2014 following the standard WHO protocol for surveillance of anti-malarial drug efficacy . In MaprP. falciparum (\u2265\u20091000 parasites/\u03bcL) or P. vivax (\u2265\u2009250 parasites/\u03bcL) mono-infection. Patients with signs of severe malaria and other exclusion criteria were referred to health facility staff for clinical management according to routine practice. Patients were enrolled and allocated to either the AL or DHA-PPQ treatment arm. Due to a delay in the arrival of the second-line drug (DHA-PPQ) in the country, the AL treatment arm was started first followed by the DHA-PPQ arm a year later. Patients were initially enrolled on a provisional basis based on their RDT result and treated with AL or DHA-PPQ. After confirmatory diagnosis by light microscopy, only patients meeting the above inclusion criteria were retained in the study. RDT-positive cases that had parasite counts below the required threshold or mixed species infections were excluded while a complete treatment course was provided.Patients aged\u2009\u2265\u20096\u00a0months presenting with fever (\u2265\u200937.5\u00a0\u00b0C) or a history of fever in the previous 72\u00a0h at one of the study health facilities were initially screened and enrolled provided they had: (i) weight\u2009\u2265\u20095.0\u00a0kg; (ii) Hb\u2009\u2265\u20095.0\u00a0g/dL; (iii) easy access to the study facility to enable follow-up; (iv) positive malaria rapid diagnostic test ; and, (v) microscopy-confirmed Active follow-up was performed by study nurses on days 1, 2, 3, 7, 14, 28, and 42. During enrolment and follow-up visits, clinical and physical examinations were performed and the findings were recorded in case report forms. Blood slides and filter paper samples were collected in all visits while haemoglobin levels were measured on days 0, 7, 14, 28, and 42 using a hand-held Hemocue device .\u00ae tablets  containing 20\u00a0mg of artemether and 120\u00a0mg of lumefantrine over 3\u00a0days, administered with 250\u00a0mL of milk. Doses were allocated by weight group: 5.0\u201314.9\u00a0kg 1 tablet; 15.0\u201324.9\u00a0kg 2 tablets; 25.0\u201334.9\u00a0kg 3 tablets; and\u2009\u2265\u200935.0\u00a0kg 4 tablets. All doses were administered under direct supervision by a study nurse. For patients enrolled into the DHA-PPQ arm, Eurartesim\u00ae film-coated tablets  containing 40\u00a0mg of DHA and 320\u00a0mg of PPQ were administered with water under supervision once daily for 3\u00a0days. Doses were allocated according to the following weight groups: 7.0 to\u2009<\u200913.0\u00a0kg \u00bd tablet; 13.0 to\u2009<\u200924.0\u00a0kg 1 tablet; 24.0 to\u2009<\u200936.0\u00a0kg 2 tablets; 36.0 to\u2009<\u200975.0\u00a0kg 3 tablets; 75.0\u2013100.0\u00a0kg 4 tablets. The dosage schedules were in accordance with the National Malaria Treatment Protocol [Patients enrolled into the AL arm were treated with 6 doses of CoartemLight microscopic diagnosis was conducted independently by 2 WHO-certified Level 1 to Level 3 microscopists at the Papua New Guinea Institute of Medical Research (PNGIMR) in Madang. In case of discordant results, slides were examined by a third senior microscopist (WHO Level 1 certified). A minimum of 200 thick film fields were examined before a slide was declared negative and a patient was considered aparasitaemic if no parasites were found by two microscopists. The number of parasites was counted up to 200 white blood cells (parasite count\u2009>\u2009100/field) or 500 white blood cells (parasite count\u2009<\u2009100/field). Parasite counts for each species were converted to the number of parasites per \u03bcL of blood assuming 8000 white blood cells per \u03bcL .Pfmsp2 for P. falciparum and Pvmsp1 for P. vivax [Pfmsp2, Pvmsp1F3 and Pvms16 were determined using capillary electrophoresis at Macrogen (South Korea). The data obtained from GeneScan  were analysed using GeneMarker, version 2.4.0 . The method used for P. vivax allows discrimination between strains in post-treatment recurrent infections in a way analogous to that established for falciparum malaria. Although P. vivax genotyping cannot differentiate between a recrudescent infection and a relapse with the same genotype, based on previous observations, most relapses are genetically distinct from the primary infections [Polymerase chain reaction (PCR) assays were used to distinguish recrudescence from re-infection cases using P. vivax \u201317. Lengfections , 15, 17.P. falciparum chloroquine-resistance transporter gene (pfcrtK76T) and the P. falciparum multi-drug resistance-1  as well as the P. vivax multi-drug resistance-1 (pvmdr-1Y976F) genes. PfK13 for artemisinin and Pfplasmepsin 2/3 for piperaquine resistance were not assessed as this study was carried out before the discovery of these markers.A ligase detection reaction-fluorescent microsphere assay (LDR-FMA) method, which has been described in detail elsewhere , 16, 17,Plasmodium falciparum cultures were maintained using a modified candle-jar technique based on the Trager & Jensen method for culture of P. falciparum parasites [arasites . The antarasites , with mo50 values are the concentration of a particular drug at which 50% of the parasites is inhibited in vitro. The prevalence of molecular markers was calculated as the frequency of any of the single nucleotide polymorphisms (SNPs) within the key resistance molecular markers .The treatment outcomes were categorized according to WHO guidelines  as earlyP. falciparum, a 95% level of confidence and 5% precision. An additional 20% was added to account for patients who would be lost during follow-up, violate protocol or voluntarily withdraw, resulting in a total sample size of 88 patients per arm per site, allowing site-specific efficacy estimation. Due to the unpredictability of the incidence of P. vivax mono-infections, no formal sample size was performed for including P. vivax cases. Instead, the study would recruit all patients with P. vivax up until the total P. falciparum sample size was reached.The sample size calculation for this study was based on an expected 5% failure rate of http://project-redcap.org/). Analysis of study outcomes was based on a per-protocol analysis of pooled patient data from both study sites, which excluded cases with a Plasmodium species other than P. falciparum or P. vivax, protocol violations, and patients lost to follow-up. Statistical analysis was carried out using STATA 11.0 . Statistically significant differences in rates of treatment response outcomes, both for PCR-corrected and PCR-uncorrected, as well as in baseline characteristics were assessed using Chi square tests and non-parametric tests for not normally distributed data. Fever and parasite clearance rates were compared using Fisher\u2019s exact test. Fever was defined as axillary temperature of\u2009\u2265\u200937.5\u00a0\u00b0C for enrolment and a 37.3\u00a0\u00b0C cut-off was used to define fever clearance.Case report forms were entered using REDCap , or a recent history of fever, were screened at the health facilities in Maprik and Alotau between June 2012 and September 2014  and 4 between day 28 and 42 . Of the P. falciparum cases in the DHA-PPQ arm, 7 were excluded between day 0 and 28  and 3 were lost to follow-up between day 28 and 42. Of the P. vivax cases in the AL arm, 5 were excluded between day 0 and 28  and 4 between day 28 and 42 . Of the P. vivax cases in the DHA-PPQ arm, 1 was excluded due to protocol violation between day 0 and 28, while 2 were lost to follow-up between day 28 and 42. A comparison of the two study populations is provided in Additional file In total, 344 patients (age range 0.5\u201364\u00a0years) were finally enrolled and followed-up according to the study protocol (171 in Alotau and 173 in Maprik). Out of the P. falciparum and 20.3% (70/344) with P. vivax. Of the patients with falciparum malaria, 47.1% (129/274) were treated with AL while 52.9% (145/274) received DHA-PPQ. Of those treated with AL, 85.3% (110/129) and 82.2% (106/129) had clinical and PCR-corrected data on days 28 and 42, respectively. Of the DHA-PPQ-treated patients, 95.2% (138/145) and 93.1% (135/145) had clinical and PCR-corrected data on days 28 and 42, respectively. Of the patients with vivax malaria, 40% (28/70) were treated with AL and 60% (42/70) with DHA-PPQ. Of those treated with AL, 82.1% (23/28) and 67.9% (19/28) had clinical and PCR-corrected data on days 28 and 42, respectively, while 97.6% (41/42) and 92.9% (39/42) of DHA-PPQ-treated patients had the respective data available  of all enrolled patients were infected with Patients enrolled into the AL and DHA-PPQ treatment arms were comparable with regard to most baseline demographic and anthropometric characteristics  experienced a treatment failure by day 42. Of these, 8 were classified as LPF (3 in the DHA-PPQ arm and 6 in the AL arm) while 1 was a LCF in the AL arm . However, all fevers in both P. falciparum- and P. vivax-infected patients in the AL arm had cleared by day 3. One patient with P. vivax in the DHA-PPQ arm presented with fever at every time point post treatment (day 0\u2013day 7). One patient with P. falciparum in the DHA-PPQ arm had fever on day 2 and then on day 7. However, all these fevers were most likely not associated with the parasite infection as all parasites were cleared in these patients and there were no fevers on day 1 , followed by an increase on subsequent days. In all groups, day 42 mean Hb concentration was higher than on day 0 and no participant had developed severe anaemia (<\u20098\u00a0g/dL) over the course of the treatment.In the AL treatment arm, the mean Hb concentration increased from 9.6\u00a0g/dL in patients with Both treatments were generally well tolerated. The overall frequency of adverse events was 56.1% (88/157) in the AL arm and 60.4% (113/187) in the DHA-PPQ arm (P\u2009=\u20090.443)  was found in 96 (92.3%) samples, the polyclonal CVMNK/SVMNT mutant haplotype in 2 (1.9%), and the wild type (CVMNK) in 6 (5.8%) cases and 2 (1.9%) exhibited the polyclonal CVMNK/SVMNT mutant haplotype. Out of the two confirmed LPF in P. falciparum patients on day 42, one carried the SVMNT mutation. The genotype of the second patient could not be determined. Three cases treated with DHA-PPQ had a SVMNT mutation with 1 exhibiting a chloroquine IC50 value of 122\u00a0nM. The only polyclonal (CVMNK/SVMNT) isolate that was treated with AL had a chloroquine IC50 of 67.5\u00a0nM.Out of a total of 274 pvmdr1, the Y976F mutation was assessed. A total of 54 samples were successfully typed of which 19 (35.2%) were wild type (Y), 26 (48.1%) were mutant (F) and 9 (16.7%) were mixed wild type mutant (YF). Of the 6 P. vivax cases that had LPF at day 42 (3 from each treatment arm), 2 were wild type, 2 were mutant, 1 was mixed wild type/mutant and 1 was undetermined. The only LCF with P. vivax and treated with AL was a mutant.For P. falciparum parasite isolates from Maprik and 47 from Alotau were collected at enrolment were tested for in vitro sensitivity to chloroquine, lumefantrine, naphthoquine, piperaquine, and pyronaridine. The mean drug concentration required for 50% parasite growth inhibition in vitro (IC50) is provided in Table\u00a050 value above the\u2009\u2265\u2009100\u00a0nM cut-off, while all lumefantrine had IC50 values below their\u2009\u2265\u2009150\u00a0nM cut-off threshold. No cut-off values for piperaquine and pyronaridine were available in literature.A total of 49 50 to any anti-malarials tested . For the two PCR confirmed LPF with P. falciparum (day 42) in the AL treatment arm, no in vitro assays were performed.There were no significant differences observed between the two study sites in the geometric mean IC50 values for naphthoquine and lumefantrine differed significantly. Both drugs seemingly exhibited a lower susceptibility in the present study as compared to the previous trial in Madang.A further comparison was undertaken between in vitro drug susceptibility data collected in Madang between 2011 and 2013 [This therapeutic efficacy study conducted across all age groups in two sites of PNG confirms high efficacy of the current first- and second-line anti-malarial treatments, AL and DHA-PPQ, for uncomplicated malaria. No recrudescence was found within 42\u00a0days in patients with falciparum malaria treated with DHA-PPQ but 2 (1.9%) patients treated with AL experienced LPF. ACPR in AL-treated patients was comparable to two other studies carried out in children aged 0.5\u20135\u00a0years in Madang and East Sepik provinces  in patients treated with AL and 87.2% (95% CI 71.8\u201395.2) in patients treated with DHA-PPQ. ACPR was higher in this study than previously reported for AL  and DHA-PPQ  in the study by Karunajeewa et al. [P. vivax cases with recurrent parasitaemia on day 42, 33.3% in the AL group and 40% in the DHA-PPQ group were identified as infections with new genotypes suggestive of a re-infection or relapse rather than recrudescence. The resulting PCR-corrected ACPR of both treatment regimens in this study remained above 90% on day 28 and on day 42 in the case of DHA-PPQ; however, confidence intervals spanned across the 90% efficacy threshold generally applied for P. falciparum. As genotyping cannot differentiate between true recrudescence and relapses from hypnozoites with the same genotype, the PCR-corrected ACPR values for P. vivax need to be interpreted with caution. At day 28, there were 4.3% treatment failures seen in the AL group but none in DHA-PPQ. While these results may be indicative of a higher efficacy of DHA-PPA against P. vivax, as suggested in a previous study to be likely due to the longer half-life of the partner drug PPQ [The interpretation of the efficacy against P. falciparum. An increased parasite clearance time with\u2009\u2265\u200910% of cases with detectable P. falciparum parasites on day 3 is the current working definition of suspected artemisinin resistance [P. falciparum or P. vivax in both treatment groups, confirming the general observation that both, AL and DHA-PPQ are still highly efficacious treatments in PNG.This study further assessed parasite and fever clearance time over the first 7\u00a0days in order to detect any delayed treatment response. Day 3 parasite clearance after treatment with an ACT has been suggested by WHO as a proxy indicator for artemisinin resistance of sistance . This stP. falciparum patients treated with DHA-PPQ may be explained by the higher average parasite density in this study group . On the other hand, a decrease in Hb in patients treated with DHA-PPQ had previously been observed in other studies and further investigations may hence be warranted [The drop in Hb concentration observed in 50 values. When comparing the results from this study with a previous study carried out in Madang 5\u00a0years ago [50 in Madang and Alotau or Maprik, respectively, were: chloroquine (167\u00a0nM vs 69\u00a0nM), piperaquine (27.7\u00a0nM vs 21\u00a0nM), and lumefantrine (1.55\u00a0nM vs 6.92\u00a0nM). Further studies are required to determine if these differences are due to the use of a different assay (Sybr green in this study vs pLDH in the previous study). In addition, the lower IC50 values of chloroquine seen in this study could possibly be an early indication of decreased parasite resistance pressure due to the shelving of chloroquine in PNG over the past 6\u00a0years.There were no significant differences in in vitro efficacy of the tested drugs between the study sites (Alotau and Maprik). A recent study using the same assay conducted in Madang  found siears ago , differepfcrt gene however remained near fixation (>\u200995% of samples had mutant pfcrt). Mutations in the pfmdr gene were also widespread with 68% of isolates carrying the N86Y mutation with other mutations in pfmdr being rare or absent. These results are in accordance with the prevalence of molecular markers of resistance as shown in earlier studies [P. vivax chloroquine resistance in Melanesia [P. vivax chloroquine resistance is likely to be common in PNG, thus justifying the use of ACT as first-line treatments against both P. falciparum and P. vivax.The mutations in the  studies , 22 and elanesia . The higkelch 13 gene [Implementing this study was faced with operational challenges as only a few centres in PNG are equipped for conducting therapeutic efficacy studies. Furthermore, the start of enrolment coincided with a large-scale free distribution of long-lasting insecticidal nets in the study sites which led to a reduction in the number of malaria cases presenting to the health facilities . A consi 13 gene  and pipe 13 gene . These mThe results from this study have shown that AL and DHA-PPQ remain efficacious for the treatment of uncomplicated falciparum and vivax malaria in PNG. Based on the day-3 parasite clearance rate, there is no evidence of artemisinin resistance in the two study sites. Continued monitoring of anti-malarial drug efficacy is warranted considering increasing use of ACT in routine clinical practice, the persistence of artemisinin monotherapy in the system and the presence of artemisinin resistance in neighbouring regions. Regular molecular monitoring of resistance markers may be a more cost-effective alternative to the considerable investments necessary to conduct therapeutic efficacy studies in new or additional study sites in PNG.Additional file 1: Table S1. Study population and key outcomes by study site.Additional file 2: Figure S1. Haemoglobin concentration."}
+{"text": "The elimination of malaria depends on the blocking of transmission and of an effective treatment. In Brazil, artemisinin therapy was introduced in 1991, and here we present a performance overview during implementation outset years.Plasmodium falciparum malaria, under treatment with using injectable or rectal artemisinin derivatives, and followed over 35-days to evaluate parasite clearance, death and recurrence. It is a retrospective cohort (1991 to 2002) of patients treated in a tertiary centre of Manaus, with positive microscopic diagnosis of P. falciparum infections and received monotherapy. Considering patients who completed 35-day treatment, 98.2% (1527/1554) presented asexual parasitaemia clearance until D4 and 1.8% (27/1554) between D5-D10. It is important to highlight that had no correlation between the five treatment schemes and the sexual parasite clearance. Finally, it is noteworthy that we were able to observe also gametocytes carriage during all follow-up (D0-D35). This cohort outcome resulted 97.6% (1554/1593) of patients who completed the 35-day follow-up, 0.6% (10/1593) of death and 1.8% (29/1593) of follow-up loss. All patients that died and those that presented parasitaemia recurrence had pure falciparum malaria during first 12-years of use in north area of Brazil. Artemisinin derivatives remained effective in the treatment of  Plasmodium falciparum. In the Americas, malaria occurs in 21 countries, predominantly in South America, where four countries  account for more than 70% of cases. In Brazil, most of the malaria cases are concentrated in the Amazon states , an endemic area for the disease.Malaria is a major health problem in tropical and subtropical regions of the world. According World Health Organization (WHO) 91 countries around the world present active malaria transmission, with a total of 445,000 deaths worldwide, of which 99% are caused by ,Appropriate and early treatment is the basis of disease control, aiming at rapid cure in order to avoid the disease progress to severe forms and even death.Plasmodium are responsible for the clinical manifestation of the disease, but its transmission occurs through the ingestion of sexual forms, gametocytes, by a mosquito vector. Artemisinin-based combination therapies (ACTs) have recognised activity against immature gametocytes, but do not act against mature gametocytes.Asexual forms of P. falciparum to various compounds, such as chloroquine, sulfadoxine-pyrimethamine, mefloquine, quinine and amodiaquine.,falciparum malaria in all endemic countries and failure of this P. falciparum therapy was not reported in Brazil.In Brazil, studies published in 1992 and 2010 have reported resistance of ,P. falciparum to artemisinin. These studies report a delay in the parasite clearance time in the blood of some patients, which suggests a change in the parasite\u2019s susceptibility pattern to artemisinin and is probably the first sign of artemisinin resistance.,,The widespread diffusion of artemisinin treatment results in continuous pressure for the selection of the most resistant parasites and this is of great concern since there is no other pharmaceutical option as effective as artemisinin and its derivatives.P. falciparum malaria and absence of genetic markers associated with artemisinin resistance have been reported,P. falciparum treatment with five artemisinin regimens in the western Brazilian Amazon, by monitoring the parasitaemic clearance, death and recurrence during the follow-up period.However, the efficacy of ACTs against Study site - Our study was carried out at Funda\u00e7\u00e3o de Medicina Tropical Dr Heitor Vieira Dourado (FMT-HVD), Manaus, in western Brazilian Amazon. FMT-HVD is a reference hospital for the diagnosis and treatment of tropical, infectious, parasitic and dermatological diseases.Study design - This was a retrospective cohort study performed to assess the responses to artemisinin-based therapy regimens. Five groups of patients were constituted regarding the treatment regimens used, three of them as monotherapy  and two combined therapies (groups 4 and 5); group 1: artesunate (AS) IV 1.5 mg/kg/day until D3; group 2: rectal artesunate suppositories (RAS), called a Rectocap\u2122, at 50 mg/day D0 until D4 for children; group 3: artemether (ATM) IM 1.5 mg/kg/day until D3; group 4: artesunate IV 1 mg/kg/day followed by oral mefloquine 20 mg/kg for 12/12 hour at D2; and group 5: artemether IM 1 mg/kg/day until D3, followed by oral mefloquine 20 mg/kg for 12/12 hour at D2. Patients with mixed malaria, concomitance of P. falciparum and P. vivax, were treated with the same therapeutic schemes described above.The data were grouped into two 6-year periods (sexenium), to compare all the evaluated variables. Asexual and sexual parasitaemia were monitored at admission (DO) and in the standardised days of follow-up: D1 to D7, D14, D21, D28 and D35.Data collection - The data were collected by reviewing all the registration forms over an 11-year period (September 1991 to December 2002) and transferred to a protocol specifically prepared for the study. The extracted data from these forms were: Plasmodium species, parasite numbers, age, gender, artemisinin-based therapy and leukocytes count.Inclusion criteria - Cases with parasitologically confirmed malaria caused by the single-species P. falciparum or mixed , which received treatment with one of the five artemisinin-based regimens were enrolled in the study.Criteria of exclusion - Patients with incomplete or illegible information, losses to follow-up and treatments with other types of antimalarial compounds or combinations were excluded of the study.Evaluation of the asexual and sexual parasitaemia and recurrence analysis - The parasite densities were quanti\ufb01ed from thick blood \ufb01lms by counting the number of ring forms until 200 leukocytes were also counted, and then multiplying the number found by 30, which assumes an average number of 6,000 leukocytes/\u00b5L. During treatment, the parasitaemia was registered daily until day 7 (D0 to D7) throughout hospitalisation and, after that, weekly until day 35 . While, the gametocytaemia was evaluated in the diagnosis (D0) and at its appearance of gametocytes among the patients who did not have gametocytes at D0.,The \u201cday-4\u201d parasitaemia measure was considered a surveillance tool for artemisinin tolerance, guaranteeing the current WHO-recommended cut-off value (72 hours).The term recurrence was used to indicate the reappearance of the asexual parasitaemia after its remission, during the 35-day follow-up. Here, we considered recurrence as recurrent asexual parasitaemia, after having observed its negativation, during the 35-day follow-up. Recurrence cases were carefully evaluated, regarding artemisinin treatment regimens, time and age groups by Kaplan-Meier analysis.Statistical analysis - The enrolled patients were stratified into nine age groups (years): [a]: \u2264 9; [b]: 10 - 19; [c]: 20 - 29; [d]: 30 - 39; [e]: 40 - 49; [f]: 50 - 59; [g]: 60 - 69; [h]: 70 - 79; and [i]: \u2265 80. The distribution of patients by year of admission, gender and age range was performed by the chi-square test. Comparison of the time for clearance and recurrence regarding age range, gender and treatment was analysed by Kaplan-Meier survival curves and by the log-rank test to compare the curves. All statistical tests were 2-tailed, and significance was set up at p < 0.05. Statistical analysis was performed using SAS, version 9.2 (SAS Institute).Patients\u2019 characteristics and epidemiological data - The flowchart of the study showed that the 2702 patients previously screened, 1109 of them were excluded . The distribution regarding age groups showed significant differences (p < 0.0001), paying the attention the high prevalence of patients under nine years old. These data are present in The number of patients with mono-infection caused by 1 (one to nine years old) , with a substantial increase in coinfection cases in age group ars old) -A; mono-ars old) B.Progress of the asexual parasite during the 35-day follow-up - Considering patients who completed the 35-day treatment with one of five analysed artemisinin-based regimens, 98.2% (1527/1554) presented asexual parasitaemia clearance until D4 and only 1.8% (27/1554) between D5-D10  of enrolled patients, 26.2 % (196/748) of whom presented gametocytes at enrollment (day 0), compared to 73.7% (552/748) who exhibited sexual stage at least once during the 35-day follow-up  did not present reappearance of the parasitaemia during the follow-up. Although the statistical analysis revealed no association with treatment regimen , age  or time treatment .P. falciparum to artemisinin based-therapies among hospitalised patients to treatment in the Northern Brazilian region, after its introduction in 1991. Almost all hospitalised patients who completed the 35-day follow-up period presented good therapeutic response with negative parasitaemia, thus demonstrating the treatment regimen effectiveness. This finding is very important because the control and elimination of falciparum malaria depends not only on the blocking of transmission but also on the provision of an effective treatment.,This is a 12-year retrospective analysis on the susceptibility of This study found malaria occurrence was higher in the youngest age groups, with a substantial increase of mixed malaria between one to nine years old. In this sense, is important to highlights that usually, manuscripts about treatment of hospitalised patients with malaria presents high frequency of children. However, in our study there were no predominance of children among the analysed patients, because all of them were admitted to the hospital to treatment because the recommended protocol of the Service.falciparum malaria. This study showed more than 75% reduction in asexual parasitaemia clearance rates during the first three days of treatment, reaching 98.3% until D4. Delayed negative asexual parasitaemia, between D5 and D10, was observed in only 1.7% of the cases. The delay of the asexual parasitaemia clearance, by itself, can suggest resistance to this compound, but does not necessarily indicates failure of the artemisinin treatment;P. falciparum parasites from Brazilian endemic areas.,P. falciparum and treatment effectiveness remain high in Brazil.P. falciparum resistance to antimalarial compounds.Artemisinin-based therapies rapidly (~ 2 days) clear the asexual parasitaemia, which are responsible for the clinical picture, rendering this medication the choice for the treatment of P. falciparum in the Juru\u00e1 Valley, a Brazilian region with high transmission of P. falciparum, where the authors showed no molecular evidence of resistance to AS and MQ during the 2010 to 2013 period.Although artesunate monotherapy was highly effective in clearing asexual blood-stage parasitaemia, a good response was also observed with the ASMQ treatment. Similar clearance to this combination therapy was previously reported in the Peruvian Amazon.It is worth mentioning that these results reflect the first decade of artemisinin-based therapy use in northern Brazil, and current monitoring should be performed to evaluate the present situation in the region. Studies using artemetherP. falciparum resistance to artemisinin derivatives in southeast Asia, which is of great concern, as there is no alternative as effective as artemisinin, and the spread of resistance throughout the world would be catastrophic.,Plasmodium to artemether in a country outside the emerging region of resistance, Asia, which causes great concern because it is an area with a history of malaria infection, and also because it is a gold mining area.In addition, several studies have reported Although the main focus is the asexual parasitaemia, because it is responsible for the clinical symptoms of the disease, we also analysed the presence at admission and the emergence of sexual forms during the follow-up. The gametocyte is the transmission form from human host to mosquitoes.in vitro and increase the clearance of ring forms in vivo. The artemisinin derivatives are a first-line antimalarial compounds that act only on the early stage of the gametocytes. Thus, in combined therapies the sexual parasitaemia clearance depends on mefloquine, since this antimalarial compound is cleared slowly as its half-life elimination is two to three weeks.Plasmodium life cycle, artemisinin and its derivatives do not prevent this transmission.It is noteworthy that in our cohort we were able also to observe sexual parasitaemia after D3 until D35. Artemisinin and its derivatives do attenuate the growth of young parasites P. falciparum infection receiving artemisinin-derivative monotherapy. Thus, our results suggest that the combined therapy with mefloquine prevented recurrences and deaths, possibly because mefloquine remains longer in the bloodstream due to its greater elimination half-life, in contrast to artemisinin derivatives that are rapidly eliminated .Several factors may have contributed to recurrence, observed only in patients receiving artemisinin-derivative monotherapy. Noncompliance could explain these treatment failures, however the dispensing of drugs was assisted because the enrolled patients were hospitalised during their follow-up. In addition, the deaths were observed only in the group of patients with pure Among the patients who did not progress to death, 93% presented negative asexual parasitaemia after treatments. Differently, all patients who progress to death presented positive asexual parasitaemia until the last day of life and died until D7. So, even though the parasitaemia at D0 was lower in patients who died  than those who surveilled , it had been persistent. In addition, 30% of patients who died (3/10) presented gametocytes at admission compared to all enrolled patients (12.3%).P. vivax was detected in 10% of the enrolled patients, which increased over the years, since a greater number of cases were observed in the second sexenium. In the last decade, the world has seen a significant decline in the number of malaria cases, accompanied by an increase of malaria vivax / malaria falciparum ratio. Mohapatra et al.30) demonstrated that P. vivax infection may be protective when coexisting with P. falciparum, with less severe disease and a lower level of parasitaemia than it was observed in infections caused only by P. falciparum.Co-infection with Ethics approval and consent to participate - The study was in accordance with the ethical standards of the declaration of Helsinki and approved by the institution\u2019s ethics committee (CEP/FMT-HVD CAAE number: 46481215.2.0000.0005).In conclusion - In spite of the limitations, because it is a retrospective study involving observation for a long period of time and with information retrieved from medical record books, this study is the largest series of cases with patients treated with artemisinin-based therapies, which have already been described in the Amazon Region. This study showed stable levels of P. falciparum sensitivity to artemisinin derivatives in the Amazon Region over a period of 12 years. In conclusion, our findings do not evidence the occurrence of P. falciparum resistance to artemisinin derivatives in the assessed period; however, routine monitoring of artemisinin should continue, so that the first signs of resistance are detected early."}
+{"text": "Treatment for severe malaria must be prompt with effective parenteral antimalarial drugs for at least 24\u2009h to achieve fast parasite clearance, and when the patient can tolerate oral therapy, treatment should be completed with effective artemisinin based combination therapy (ACT) for complete parasite clearance and to prevent recrudescence. We evaluated piperaquine concentration and malaria treatment outcomes among Ugandan children treated for severe malaria with intravenous artesunate (AS) or quinine (QN) plus dihydroartemisinin-piperaquine (DP), in Tororo District Hospital in Eastern Uganda.Capillary blood piperaquine concentration data were obtained from a randomized clinical trial whose objective was to evaluate parasite clearance, 42-day parasitological treatment outcomes and safety, following treatment of severe malaria with intravenous AS or QN, plus artemether-lumefantrine or DP among children in Tororo District Hospital, in Eastern Uganda.p\u2009<\u20090.001), and lower than the recommended day 7 cut off level of 57\u2009ng/mL. There was no difference in median capillary piperaquine concentrations among participants with re-infection and recrudescence (35.3 (IQR) (17.9\u201355.2) vs 34.8 (IQR) (18.1\u201345.1), p\u2009=\u20090.847). The risk of treatment failure was two times higher among children with low (<\u200957\u2009ng/mL) day 7 capillary piperaquine concentration (relative risk: 2.1 CI 1.4\u20133.1), p\u2009<\u20090.001) compared to children with high day 7 capillary piperaquine concentrations (>\u200957\u2009ng/mL).Piperaquine concentration data from 150 participants who received DP were analyzed. Participants with unadjusted treatment failure had lower median day 7 capillary piperaquine concentration than those with treatment success (34.7 (IQR) (17.9\u201349.1) vs 66.7 (IQR) (41.8\u201381.9), Considering the low day 7 concentrations of piperaquine reported in the patients studied here, we suggest to adopt the recently recommended higher dose of DP in young children or a prolonged 5-day dosing in children living in malaria endemic areas who have suffered an initial episode of severe malaria in order to achieve adequate drug exposures for effective post-treatment prophylactic effects.PACTR201110000321348). Registered 7th October 2011.The study was registered with the Pan African Clinical Trial Registry ( Severe malaria is a life threatening emergency, responsible for 435,000 deaths annually, worldwide, with the greatest burden in sub-Saharan Africa . TreatmeP.falciparum malaria, with at least one of the laboratory or clinical features of severe malaria. Patients were excluded if they had obvious concomitant febrile illness, history of allergy to any of the study drugs, if they could not comply with study procedures and visits, or if they had received an antimalarial drug within 24 h before presenting to hospital.Study methodology has been previously described and published as a randomized single blind clinical trial conducted in Tororo District Hospital in Eastern Uganda , an areaIntravenous AS  was administered as a slow bolus into an indwelling cannula as 2.4\u2009mg/kg at start of treatment, repeated at 12 and 24\u2009h and every 24\u2009h till the switch to oral therapy. Intravenous QNN dihydrochloride  was administered over 4\u2009h as 10\u2009mg/kg body weight in 5% dextrose (10\u2009ml/kg) and repeated 8 hourly till the switch to oral therapy.Parenteral antimalarial therapy was administered for at least 24\u2009h, followed by a full course of the oral ACT when participants could tolerate oral therapy. Oral AL  was administered according to body weight as; one (5\u201314\u2009kg), two (15\u201324\u2009kg), three (25\u201334\u2009kg) and four (>\u200935\u2009kg) tablets 12 hourly, with a cup of milk or food, for 3 days. Oral DP  40\u2009mg\u2009+\u2009piperaquine (PQP) 320\u2009mg tablets) was administered targeting a total dose of 6.4 and 51.2\u2009mg/kg of dihydroartemisinin and piperaquine, respectively, given in three equally divided doses to the nearest quarter tablet. We used a pill cutter to ensure that the tablet fractions were as close to the nearest quarter tablet as possible.All participants received oral paracetamol in a dose of 15\u2009mg/kg at 8 hourly intervals. Adjunctive and supportive treatment for complications of malaria such as convulsions and hypoglycemia was given in accordance with the Uganda Ministry of Health guidelines. The study nurse provided information to caretakers about adherence to drugs, follow-up visits and potential drug side effects. Caretakers were instructed to observe the participants for 30\u2009min after drug administration and if vomiting occurred they were to administer another dose, for up to two extra doses, following which they were to bring back the participant to the study clinic for evaluation and treatment.We performed serial blood smears at 0, 1, 2, 4, 6, 8, 10, 12, 16, 20, 24\u2009h post start of intravenous therapy and every 6\u2009h until 6\u2009h post parasite clearance. Participants were initiated on oral ACT and discharged from hospital when they could tolerate oral ACT and the blood smear was negative for malaria parasites, and were followed up for 42\u2009days to ascertain parasitological outcomes and monitor adverse events on days 1, 2, 3, 7, 14, 21, 28, 35, 42, and any unscheduled day if the participant felt unwell. On each of these days we took medical history and performed physical examination, a finger prick was done to collect blood on slides for malaria diagnosis and on filter paper for genotyping and drug concentration measurements. Participants with positive malaria films were reassessed for severity and treated accordingly, those with severe malaria were re-admitted and treated with intravenous AS plus AL, and those with uncomplicated malaria were evaluated for treatment failure and treated according to national guidelines. Participants were discontinued from study follow up if they could not take study medication, missed a scheduled follow-up visit and could not be located at home, or if they received non study drugs.Thin smears were performed to determine the type of malaria parasite species and thick smears for parasite density. Thick blood smears were stained using 3% Giemsa for 30\u2009min and read by two independent experienced laboratory technologists, blinded to participants\u2019 treatment assignment. Any discrepant results were reviewed by a tie breaker.Parasite density was calculated by counting the number of asexual parasites (ring stages) per 200 white blood cells (WBCs) or per 500 if the count was less than 10 parasites per 200 WBCs, assuming a WBC count of 8000/uL of blood. A smear was considered negative if no parasites were seen after review of 100 high-power fields. Complete blood count and hemoglobin estimation were performed using the Coulter counter .Molecular genotyping of paired samples was conducted to distinguish re-infection from recrudescence, at the Makerere University-University of California San Francisco Molecular Biology laboratory in Mulago, Kampala. We used Whatman 3MM filter paper from Sigma. Parasite DNA was extracted from filter paper blood samples collected on the day of enrollment and the day of parasitological treatment failure using Chelex 100 Resin extraction  as previously described . The surThe primary study outcome was parasitological treatment failure unadjusted by genotyping classified as parasitemia detected by thick blood smear. This primary outcome was selected because it best represents the treatment outcome measure used in routine clinical care. The secondary outcomes were parasitological treatment failure adjusted by genotyping classified as reinfection or recrudescence. Adverse events were defined as any medical occurrence post study drug administration. They were graded as mild, moderate, severe and life threatening and their relationship to the study drug was classified as unrelated, possibly, probably or definitely related to study drug.On each follow up day ie days 1, 2, 3, 4, 7, 14, 21, 28, 35, 42 post the commencement of ACT administration, capillary blood samples were collected by finger prick and stored dry on Whatman 3MM filter paper. The fingers were disinfected and pricked with a lancet, following which the first blood drop of blood was discarded and the next three drops collected. Each drop filled a pre marked circle on filter paper. The filter papers were allowed to dry at room temperature and packed in sealed ziplock bags.The blood samples were transported at room temperature, to the Department of Clinical Pharmacology at the Mahidol-Oxford Tropical Medicine unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand. Piperaquine concentrations were measured using an LC-MS/MS based assay and validated according to U.S. FDA guidelines (unpublished data). Briefly, 3 discs of 3.2\u2009mm diameter were punched out from each dried blood spot sample and 375\u2009\u03bcL stable isotope internal standard in phosphate buffer 50\u2009mmol/L pH\u20092.0 was added followed by 150\u2009\u03bcL perchloric acid (0.3\u2009mol) and 75\u2009\u03bcL acetonitrile and then mixed for 60\u2009min. Approximately 500\u2009\u03bcL was transferred to a 96 well plate solid phase extraction column and extracted with a MPC-SD Empore 96-wellplate standard well 1\u2009ml . The extracted sample was then evaporated until dry and reconstituted in 250\u2009\u03bcL acetonitrile-ammonium bicarbonate 2.5\u2009mmol/L pH\u200910 (85\u201315\u2009v/v). The LC-MS/MS assay settings were the same as a previously published method . The lowThe study was approved by Makerere University School of Medicine Research and Ethics Committee (REC REF 2011\u2013175), Uganda National Drug Authority (369/ESR/NDA/DID-12/2011), Uganda National Council for Science and Technology (HS 1031) and registered with the Pan African Clinical Trial Registry (PACTR201110000321348). All study procedures were conducted according to Good Clinical Practice standards. Patients and parents or guardians of participants provided written informed consent prior to enrollment. Study related information was provided in the participants\u2019 local languages.Data were entered and verified using MS ACCESS and analyzed using STATA version 13.1 . Descriptive statistics were used to compare demographic and clinical characteristics among the four study arms. Continuous variables were compared using Wilcoxon test for non-normally distributed data. Categorical variables were compared using Chi-square test. Parasite density was normalized using logarithmic transformation. Intention-to-Treat analysis was used for comparison of treatment outcomes, which included all enrolled participants. Unadjusted treatment failure was classified as a positive blood smear on any of the follow-up days. Adjusted treatment failure was classified as either re-infection or recrudescence based on genotyping. The risk of treatment failure at 28, 35 and 42\u2009days of follow up (unadjusted and adjusted by genotyping) were estimated using the Kaplan-Meier survival method and compared using the Log Rank test. Time at risk was calculated from day one of ACT allocation to date of treatment failure among participants who failed, last day of follow-up for those who did not complete follow-up, or day 42 for the patients who completed 42\u2009days of follow-up. In the analysis for adjusted parasitological outcomes, only recrudescence was considered as true parasitological treatment failure. Safety data from all participants were analyzed.Piperaquine concentration data were compared using the Wilcoxon test. We compared day 7, 14, 35 and 42 piperaquine concentration among children with and without malaria treatment failure. Day 7 piperaquine concentration were also stratified above/below a previously reported cut off level of 57\u2009ng/mL, associated with an increased risk of therapeutic failure , and evaWe enrolled and followed up 300 participants between January 2012 and March 2013, of whom, 150 received DP. Baseline characteristics were similar across the four treatment arms  day 7 capillary piperaquine concentration was 42.1 (23.2\u201367.2) ng/mL, lower than the recommended cut off level of 57\u2009ng/mL , and 90 p\u2009<\u20090.001).The observed median (IQR) day 7 capillary piperaquine concentrations were significantly lower in patients with recrudescence (34.8\u2009ng/mL (IQR) (18.1\u201345.1) and re-infection (35.3\u2009ng/mL (IQR) (17.9\u201355.2) compared to patients with successful malaria treatment outcome (66.7\u2009ng/mL (IQR) (41.8\u201381.9), both p\u2009<\u20090.001) among children with low (<\u200957\u2009ng/mL) day 7 capillary piperaquine concentrations compared to children with high day 7 capillary piperaquine concentrations (>\u200957\u2009ng/mL). Figure\u00a0Furthermore, the risk of treatment failure (recrudescence and re-infection) was two times higher . Previous studies have reported day 7 piperaquine capillary concentration of 57\u2009ng/mL as a therapeutic target, with lower concentrations associated with an increased risk of recrudescence in patients treated for uncomplicated malaria . In 2015We found that children with piperaquine concentration below this target had an approximately 2-fold higher risk of malaria treatment failure. Despite this, we demonstrated low levels of recrudescence among our study participants with no difference in the risk of recrudescence across study arms .Previous studies have demonstrated high risk for readmission or death within 6\u2009months\u2019 post discharge among children hospitalized with severe malaria in malaria endemic areas. Therefore, researchers have recommended administration of malaria chemoprevention with DP at discharge in order to protect from novel infections in the period following the severe malaria episode \u201318. TreaConsidering the low day 7 concentrations of piperaquine reported in the patients studied here, we suggest to adopt the recently recommended higher dose of DP in young children  or a pro"}
+{"text": "Routine surveillance on the therapeutic efficacy of artemisinin-based combination therapy (ACT) has been ongoing in Ghana since 2005. The sixth round of surveillance was conducted between 2015 and 2017 to determine the therapeutic efficacy of artesunate\u2013amodiaquine (AS\u2013AQ) and artemether\u2013lumefantrine (AL) in 10 sentinel sites across the country.The study was a one-arm, prospective, evaluation of the clinical, parasitological, and haematological responses to directly observed treatment with AS\u2013AQ and AL among children 6\u00a0months to 9\u00a0years old with uncomplicated falciparum malaria. The WHO 2009 protocol on surveillance of anti-malaria drug efficacy was used for the study with primary outcomes as prevalence of day 3 parasitaemia and clinical and parasitological cure rates on day 28. Secondary outcomes assessed included patterns of fever and parasite clearance as well as changes in haemoglobin concentration.Day 3 parasitaemia was absent in all sites following treatment with AS\u2013AQ whilst only one person (0.2%) was parasitaemic on day 3 following treatment with AL. Day 28 PCR-corrected cure rates following treatment with AS\u2013AQ ranged between 96.7% (95% CI 88.5\u201399.6) and 100%, yielding a national rate of 99.2% (95% CI 97.7\u201399.7). Day 28 PCR-corrected cure rates following treatment with AL ranged between 91.3% (95% CI 79.2\u201397.6) and 100%, yielding a national rate of 96% (95% CI 93.5\u201397.6). Prevalence of fever declined by 88.4 and 80.4% after first day of treatment with AS\u2013AQ and AL, respectively, whilst prevalence of parasitaemia on day 2 was 2.1% for AS\u2013AQ and 1.5% for AL. Gametocytaemia was maintained at low levels (<\u20095%) during the 3\u00a0days of treatment. Post-treatment mean haemoglobin concentration was significantly higher than pre-treatment concentration following treatment with either AS\u2013AQ or AL.The therapeutic efficacy of AS\u2013AQ and AL is over 90% in sentinel sites across Ghana. The two anti-malarial drugs therefore remain efficacious in the treatment of uncomplicated malaria in the country and continue to achieve rapid fever and parasite clearance as well as low gametocyte carriage rates and improved post-treatment mean haemoglobin concentration.The online version of this article (10.1186/s12936-019-2848-1) contains supplementary material, which is available to authorized users. Malaria control efforts in Ghana have, over the years, yielded some positive results. The national parasite prevalence among children aged 6\u201359\u00a0months decreased from 27.5% in 2011 to 20.4% in 2016 , 4. MalaCurrently, case management remains one of the main malaria interventions in Ghana. There are three first-line anti-malarial drugs for treating uncomplicated malaria in the country. These are artesunate\u2013amodiaquine (AS\u2013AQ) combination, artemether\u2013lumefantrine (AL) combination, and dihydroartemisinin\u2013piperaquine (DHAP) combination . AL or DThe study was a one-arm, prospective, evaluation of the clinical, parasitological and haematological responses to directly observed therapy for uncomplicated malaria among children aged 6\u00a0months to 9\u00a0years in 10 sentinel sites across Ghana. Primary objectives were to assess prevalence of day 3 parasitaemia as well as the clinical and parasitological cure rates on day 28 following treatment with AS\u2013AQ and AL. Secondary objectives were to assess the patterns of fever and parasite clearance, changes in haemoglobin concentration, gametocyte carriage rates, and prevalence of adverse events. Five of the 10 sentinel sites were scheduled to first study AS\u2013AQ followed by AL whilst the other five were scheduled to first study AL followed by AS\u2013AQ.The study was conducted in all 10 sentinel sites for anti-malarial drug efficacy monitoring in Ghana  were able to study both AS\u2013AQ and AL during the study period whilst WRH and LEKMH studied only AS\u2013AQ, and YMH studied only AL.Plasmodium falciparum, parasite count ranging between 1000 and 250,000/\u00b5l, haemoglobin level\u2009>\u20095\u00a0g/dl, parental consent, ability and willingness to comply with study protocol, absence of signs/symptoms of severe malaria, absence of severe malnutrition, absence of other causes of fever such as pneumonia, otitis media, gastroenteritis, measles, and urinary tract infection, absence of chronic disease such as cardiac, renal and hepatic, and, history of allergy to test medicine.The study population was made up of febrile children aged 6\u00a0months to 9\u00a0years suspected to have malaria. Inclusion criteria were axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C or history of fever during the past 24\u00a0h, mono-infection with All anti-malarials used in the study were fixed-dose combinations, and were supplied by WHO, Geneva. The AS\u2013AQ used were from Sanofi Aventis whilst the AL used were from Ipca Laboratories Ltd, India. Treatment doses for AS\u2013AQ were as follows: children weighing: (i) 4.5 to\u2009<\u20099\u00a0kg received a tablet of the 25/67.5\u00a0mg product daily for 3\u00a0days; (ii) 9 to\u2009<\u200918\u00a0kg received a tablet of the 50/135\u00a0mg product daily for 3\u00a0days; and, (iii) 18 to\u2009<\u200936\u00a0kg received a tablet of the 100/270\u00a0mg product daily for 3\u00a0days. The administration of AL (20/120\u00a0mg), which was also by weight, was at 0, 8, 24, 36, 48, and 60\u00a0h. Treatment doses were as follows: children weighing: (i) 5 to\u2009<\u200915\u00a0kg received one tablet per treatment hour; (ii) 15 to\u2009<\u200925\u00a0kg received two tablets per treatment hour; and, (iii) 25 to\u2009<\u200935\u00a0kg received three tablets per treatment hour. All treatments were given under direct observation of a study nurse after satisfying himself/herself that the child had eaten. The child was then observed for 30\u00a0min to ascertain retention of anti-malarial. Children who vomited during the observation period were re-treated with the same dose of anti-malarial and observed for an additional 30\u00a0min. Children with repeated vomiting were withdrawn from the study and treated as severe malaria according to national standard treatment guidelines using the following options: (i) intravenous artesunate ; or, (ii) intramuscular artemether . Parenteral treatment of severe malaria cases was given for a minimum of 24\u00a0h. A full 3-day course of oral ACT was given when child was able to swallow oral medication [All children enrolled were scheduled to visit the outpatient clinic on days 1, 2, 3, 7, 14, 21, and 28 (day 0 being the day treatment was started). Enrolled children were clinically examined by the study physician any time they visited the clinic. Asexual parasite densities and presence of gametocytes were assessed on days 2, 3, 7, 14, 21, 28, and any unscheduled visit within the 28\u00a0days of follow-up.Thick and thin blood smears prepared on the days of parasitological assessment were stained with 3% Giemsa for 30\u201345\u00a0min. Asexual parasites were counted against 200 white blood cells using a hand tally counter whilst the presence of gametocytes was noted. Parasite densities were finally expressed per \u00b5l blood assuming white blood cell count of 8000/\u00b5l blood. A blood smear was declared negative after examination of 100 thick-film fields. All blood slides were read by two qualified independent microscopists, and discordant readings  were re-examined by a third qualified independent microscopist, whose reading was considered final. Filter paper blots were obtained on day 0 and at recurrence of parasitaemia for PCR genotyping using merozoite surface protein 2 (MSP2)-specific primers: FC 27 and 3D7 to distinguish between recrudescence and re-infection , 23. Sam\u00ae template for therapeutic efficacy tests [A minimum sample size of 50 was computed for each sentinel site based on an estimated treatment failure rate of 5% at 95% confidence level, 10% precision, and 20% loss to follow-up. Data were captured using WHO Microsoft Excelcy tests . Primarycy tests . Day 28 cy tests . SecondaOut of a total of 1463 children screened in nine facilities to receive AS\u2013AQ, 492 met the inclusion criteria and were enrolled\u00a0 under 5\u00a0years old. Mean weight ranged between 14.0\u00a0kg (\u00b1\u20096.0) in YMH and 17.8\u00a0kg (\u00b1\u20095.3) in HMH, yielding an overall mean of 16.0\u00a0kg (\u00b1\u20095.7). Mean axillary temperature ranged between 37.1\u00a0\u00b0C (\u00b1\u20090.8) in SMH and 38.5\u00a0\u00b0C (\u00b1\u20090.8) in NWMH, yielding an overall mean of 38.0\u00a0\u00b0C (\u00b1\u20091.1). Geometric mean parasitaemia ranged between 2941/\u00b5l blood in TAGH and 54,373/\u00b5l blood in BGH, yielding an overall mean of 23,534/\u00b5l blood. Gametocytaemia was prevalent in only one facility (BMH). Mean haemoglobin concentration ranged between 9.0\u00a0g/dl (\u00b1\u20091.5) in YMH and 10.9\u00a0g/dl (\u00b1\u20091.8) in TAGH, yielding an overall mean of 10.1\u00a0g/dl (\u00b1\u20091.6)  at a rate of 2.3% (1/43), yielding a national rate of 0.2% (1/462). ETF was reported in three sites: one site after treatment with AS\u2013AQ and two sites after treatment with AL  and 100% for AS\u2013AQ  and between 75% (95% CI 61.6\u201385.6) and 100% for AL . PCR-corrected cure rates ranged between 96.7% (95% CI 88.5\u201399.6) and 100% for AS\u2013AQ and between 91.3% (95% CI 79.2\u201397.6) and 100% for AL  or AL   Fig.\u00a0. This paThe overall mean haemoglobin concentration significantly increased on day 28 compared with day 0 following treatment with either AS\u2013AQ  or AL  showed no significant increase following treatment with AL ; 8.2% vs. 1.5% on day 1 (p\u2009<\u20090.001); and 6.5% vs. 0.4% on day 2 (p\u2009<\u20090.001). No serious adverse event was reported.Routine surveillance on the therapeutic efficacy of anti-malarials in sentinel sites across Ghana has been ongoing since the introduction of ACT in 2005. For five rounds of surveillance conducted between 2005 and 2013, each of the 10 sites representing the 10 regions of the country were scheduled to study either AS\u2013AQ or AL. For the purpose of obtaining site-specific efficacy data for both AS\u2013AQ and AL in one round of surveillance, each sentinel site was scheduled to test both anti-malarials during the 2015\u20132017 round of surveillance. Seven of the 10 sites successfully tested both AS\u2013AQ and AL whilst two sites tested only AS\u2013AQ and another site tested only AL. The primary outcomes assessed were: (i) prevalence of day 3 parasitaemia, which is an indicator of suspected artemisinin  resistance; and, (ii) a day 28 treatment outcome, which is an indicator of partner drug resistance . SecondaThe study showed no presence of parasitaemia on day 3 following treatment with AS\u2013AQ in all sites whilst only one person (0.2%) was parasitaemic on day 3 following treatment with AL. This finding is far below the WHO threshold of 10%, and therefore suggests that artemisinin  resistance following ACT treatment of falciparum malaria is not a problem in Ghana .The overall day 28 cure rates were 99.2% for AS\u2013AQ and 96.0% for AL, yielding ACT treatment failure rates of between 0.8 and 4.0%. The ACT treatment failure rates observed are below the WHO threshold of 10%, and therefore suggest that amodiaquine and lumefantrine are not failing as partner drugs in ACT use in Ghana . The higBoth AS\u2013AQ and AL showed rapid fever and parasite clearance during the first week of follow-up in all sites. Overall prevalence of fever declined by 88.4 and 80.4% after first day of treatment with AS\u2013AQ and AL, respectively, whilst prevalence of parasitaemia on day 2 was 2.1% for AS\u2013AQ and 1.5% for AL. Gametocytaemia was absent on day 7 following treatment with either AS\u2013AQ or AL. These findings suggest that ACT remains effective in rapidly clearing fever and asexual parasites as well as reducing gametocyte carriage rates in Ghana , 25\u201332.Generally, post-treatment mean haemoglobin concentration following treatment with either AS\u2013AQ or AL was higher than pre-treatment concentration. Only one of the nine sites that studied AS\u2013AQ and two of the eight sites that studied AL did not show a significant increase in post-treatment haemoglobin concentration. It is worth noting that all the three sites that did not show significant increase in post-treatment haemoglobin concentration following treatment with either AS\u2013AQ or AL showed increases of between 0.1 and 0.5\u00a0g/dl, suggesting that ACT treatment has a positive impact on post-treatment haemoglobin levels as shown in previous studies , 25\u201332.A commonly reported adverse event following treatment with either AS\u2013AQ or AL was vomiting. Prevalence of vomiting on each day of treatment was significantly higher among children treated with AS\u2013AQ compared with those treated with AL. This finding suggests that AL may be more tolerable than AS\u2013AQ influencing patient preference and use as reported by Chatio and colleagues .The main limitation of this study is the lack of pharmacokinetic data to better explain the recrudescence observed. The cure rates for AS\u2013AQ and AL may therefore be higher than the rates observed. Furthermore, in view of the fact that this study was not a randomized trial, it cannot be concluded that AS\u2013AQ is superior to AL even though its overall cure rate appeared to be higher than that of AL.The therapeutic efficacy of AS\u2013AQ and AL are over 90% in sentinel sites across Ghana. The two anti-malarial drugs therefore remain efficacious in the treatment of uncomplicated malaria in the country achieving rapid fever and parasite clearance as well as low gametocyte carriage rates and improved post-treatment mean haemoglobin concentration.Additional file 1. Participant flow showing number screened, enrolled and included in the per-protocol population."}
+{"text": "Plasmodium vivax malaria. Compared with chloroquine, dihydroartemisinin/piperaquine was faster in clearing asexual P. vivax parasites and blocking human-to-mosquito transmission. This drug combination was also more effective in preventing potential recurrences for >2 months.We assessed the efficacy of standard 3-day courses of chloroquine and dihydroartemisinin/piperaquine against  Plasmodium vivax is the most widespread human malaria parasite. Almost 2.5 billion persons are at risk for infection in >90 countries , which led to withdrawal of chloroquine and use of dihydroartemisinin/piperaquine (DHA/PPQ) as first-line therapy for uncomplicated P. vivax malaria in 2012  for non\u2013At enrollment, after we obtained written informed consent, patients were randomized to receive supervised standard 3-day courses of DHA/PPQ  or chloroquine . For each participant, medical histories were obtained and clinical and biological examinations performed. We followed up with patients according to an extended World Health Organization protocol on days 1, 2, 3, 5, and 7 and then weekly until day 63. At each visit, we performed clinical examinations and obtained an axillary temperature and a capillary blood sample. ,Malaria parasites were detected by microscopy (Giemsa-stained blood films) and PCR as described  before the first dose of DHA/PPQ or chloroquine, 2) on the same day at 9:00 pm , and 3) at 24h posttreatment for patients treated with chloroquine. We performed statistical analyses by using GraphPad Prism 5  and R software ; in each arm, 5 patients were lost to follow-up during days 2\u201335. A total of 40 patients (20 in each study arm) were followed up until day 63. Baseline patient characteristics were similar for both patient groups . We did ,,Within 2 months of follow-up, there were fewer patients with a recurrence (detected by PCR) in the DHA/PPQ-treated group than in the chloroquine-treated group  . Median P. vivax\u2013infected patients and the median proportion of infected mosquitoes fed on blood collected before the first dose of DHA/PPQ or chloroquine were similar for both patient groups  . Baseline patient characteristics were similar in both patient groups, except for day 0 parasitemia and gametocytemia, which were higher for the DHA/PPQ-treated group . The prot groups . Despiteed group . Overall 0.0001) . For the 0.0001) .<48 h) than chloroquine (\u224872 h) in eliminating sexual and asexual P. vivax parasites and that DHA/PPQ provides an excellent postexposure prophylaxis against potential recurrences for >2 months , which has a long terminal elimination half-life and is highly effective in preventing P. vivax recurrence for up to 56 days. Although the number of patients enrolled was small, we demonstrated that DHA/PPQ also acts faster (<5 h) than chloroquine in killing P. vivax sexual stages and thus prevents the risk for transmission of parasites to the mosquito vector the night after uptake of the first dose. This rapid clearance of gametocytes is a major benefit of DHA/PPQ in comparison with chloroquine, given that P. vivax gametocytes appear early in the course of disease and must be eliminated as soon as possible to limit risk of transmission (,We confirm that DHA/PPQ acts faster (P. falciparum and P. vivax uncomplicated malaria in regions to which these species are coendemic. These findings apply to areas in which chloroquine is still effective and no P. falciparum resistance to PPQ has been observed.In summary, our findings support the recommendation of DHA/PPQ as first-line treatment for"}
+{"text": "Plasmodium vivax malaria cases in Guragae zone, southern central Ethiopia.Malaria continues to be a public health problem and important cause of morbidity and mortality in Ethiopia. Due to continuous interventions to combat malaria in endemic regions, a decline in malaria related deaths and morbidity has been registered. These gains, however, are threatened with the emergency of antimalarial drugs resistant strains of plasmodium parasites. This study aimed to determine therapeutic efficacy of chloroquine for treatment of Plasmodium vivax malaria mono infection and eligible for study inclusion criteria were recruited. SPSS-21 used for data analysis and management. Kaplan-Meier survival probability analysis was estimated. Mean geometric parasitaemia and average haemoglobin concentration were calculated.A one arm prospective study with recurrence of parasitaemia and clinical conditions of patients were evaluated on days 0, 1, 2, 3, 7, 14, and 28. Patients with Among 87 total recruited subjects, 81 of them completed the 28\u2009days follow up. More than half of (57.5%) the study participants had a history of fever and 42.5% of them had fever at the time of enrollment. The mean body temperature on day of recruitment was 38.2\u2009\u00b0C and 36.8\u2009\u00b0C on day 28. Geometric mean parasitaemia calculated on day of enrollment was 2270 parasites/\u03bcl of blood. Recurrence of parasitaemia was registered from two subjects during entire follow up. The mean haemoglobin concentration of study participants on day of enrolment was 11.8\u2009g/ dl and 13.8\u2009g/dl on day 28.Plasmodium vivax malaria in the study area. However, there is a need to monitor chloroquine resistance by employing molecular tools for better evaluation of treatment outcome.This study registered a high chloroquine efficacy rate among the study participants. Therefore, chloroquine remains efficacious for the treatment of  Plasmodium vivax (P.vivax) is the most important and a major public health problem [P.vivax infection in east Africa, particularly in Ethiopia is higher [P.vivax distribution is mainly associated with altitude and rainfall. Vivax malaria accounts for up to 40% of malaria cases in Ethiopia [Malaria annually affects several hundred million people and malaria due to  problem . Studiess higher . MalariaEthiopia . AccordiP.vivax malaria as it benefit for both the patient and community. However, the effectiveness of such treatment strategy highly depends on the national policy of providing effective first line antimalarial drugs [P.vivax malaria need monitoring the anti vivax malarial drug for identifying emergence of multidrug resistant isolates [Prompting effective treatment is one of effective global strategies in fighting against al drugs , 4. Progisolates . Monitorisolates , 7.P.vivax isolates are reported from different parts of the world [P.vivax in malaria endemic areas of Ethiopia [P.vivax in malaria endemic areas like Ethiopia to have clear picture of the country wide CQ resistance distribution. The present study gives updated information about the status of CQ resistance for the treatment of P.vivax in the study area.Despite the fact that emergence of chloroquine(CQ) resistant he world \u201310. HoweEthiopia , 12. MorThe study was carried out from December, 2016 to May, 2017 in three selected health care centers located in Guragae zone, Southern Nations Nationalities, and People\u2019s region. Guragae zone is located 155\u2009k-meters south of Addis Ababa, the capital of Ethiopia. The zone has an annual rainfall of 800 to 1400\u2009mm.P.vivax mono-infection on blood film examination and fulfilled the inclusion criteria were recruited. Sample size was calculated using single population proportion formula with expected 5% failure rate, a desired precision of 5 and 95% confidence interval (CI), and assuming an additional 20% for loss to follow-up [A one-arm prospective study, with clinical and parasitological evaluation was done. 87 study subjects confirmed with ollow-up . ProportP. vivax mono-infection, non-pregnant or non-breast-feeding women, permanently living within the health center catchment area (10\u2009km radius) during the study period were recruited [Febrile patients (documented body temperature greater than 37.5\u2009\u00b0C or having history of fever within the previous 48\u2009h), who fulfilled other inclusion criteria and signed an informed consent was eligible for the study. Specifically, patients of aged >\u20096\u2009months, microscopically confirmed Study participants were excluded due to presence of other causes of fever at the time of screening, pregnancy, administration of any additional anti-malarial drugs during the study period, and subjects who vomit twice during drug administration .P.vivax including in our study area, CQ is the first line drug [P.vivax endemic areas of Ethiopia. Subjects were checked for vomiting for 30\u2009min after the drug ingestion. Blood smear microscopic examination was made in every scheduled visit. Moreover, haemoglobin concentration measurement was done on day 0, 28 and days of recurrence. Study participant failing to come to their scheduled appointment was traced to home assisted by the local guide. Study subjects were classified as having therapeutic failure and an adequate response [P.vivax after the initial clearance of parasite from circulation. Subjects are considered to have cleared parasitaemia if there are at least two sequential negative smears and the day on which the first such negative smear is observed defined as the day of clearance [Fixed schedule check-up visits were done on days 0, 1, 2, 3, 7, 14, 21 and 28. Corresponding clinical evaluation and laboratory examination were made. Baseline data on socio-demographic and clinical characteristics were recorded . In areaine drug . Hence, response . RecurreThe quality of CQ , used to treat all study participants was checked for its quality following the standard procedures before administration.P.vivax mono infection. Asexual stages of P.vivax were counted against 200 white blood cells, assuming the average total white blood cell count of 8000/\u03bcl. Parasite density, expressed as the number of asexual parasites per \u03bcl of blood, was calculated by dividing the number of asexual parasites by the number of white blood cells counted and then multiplying it by an assumed white blood cell density (8000 per \u03bcl). Blood film examination was recorded as negative when examination of 1000 white blood cells revealed no asexual parasites. Parasite reduction ratio(PRR) was calculated using the formula PRR\u2009=\u2009Po/P2 (where Po is parasite count on day 0 and P2 parasite count on day 2) [Thick and thin blood film microscopic examination was done to identify n day 2) .In pregnant females since Artesunate is recommended for treatment vivax malaria, female study participants aged 12\u2009years and older were tested for pregnancy . Haemoglobin concentration was also measured on day 0, on day of recurrence parasitaemia and on day 28 by using HemoCue Hb 301(Sweden) analyzer.All positive and 10% of negative slides were picked randomly for quality control and re-examined by expert microscopist at Wolkite University parasitology Laboratory.SPSS version-21  was used for data management and analysis. Kaplan-Meier survival probability analysis was used to calculate cumulative incidence of failure. Parasite reduction ratio, mean geometric parasite density and mean haemoglobin were calculated.Study variables include therapeutic efficacy of CQ, socio-demographic characteristics, clinical characteristics, haemoglobin concentration and parasite density.Subject that has an asexual parasitaemia of any level persisting to day 7 during follow up period .Presence of an asexual parasitaemia that becomes undetectable but reappears at any time between days 7 and 28 during follow up period .Absence of parasitaemia on day 28, irrespective of axillary temperature or subjects having no recurrence parasitaemia up to day 28 during follow period .P.vivax mono infected patients were screened in all selected health care centers, of which 87 were eligible to the 28\u2009days follow-up study. The rest 21 participants were not enrolled in the study due to different reason. Among 87 recruited participants 81 of them had completed the entire 28\u2009days follow up. While six were excluded because they vomited twice during the third dose of CQ, P. falciparum detection on day 7, and lost to follow up on day 7 and 14 as illustrated in Fig.\u00a0A total of 108 The median age of study participants was 19 (18\u2009months\u201342\u2009years). Males were higher in proportion than females (1.2: 1). More than half of the study participants had a history of fever (57.5%) and illness two days before the day of enrolment (55.4). Geometric mean parasite density of study participants at the day of enrollment was 2270 /\u03bcl and parasitaemia clearance time for all recruited participants was observed 48\u2009h after the day of enrollment. Mean body temperature registered on the day of recruitment was 38.2\u2009\u00b0C and 36.8\u2009\u00b0C on day 28. All study participants cleared fever (37.0\u2009\u00b0C) on day 2 following treatment as shown in Table\u00a0Two treatment failures  for CQ were registered. Parasitaemia persisted up to day 28 in treatment failure cases, while parasitaemia load was lower on day of admission as compared to day of enrollment (mean parasitic density on day 0 was 3700.5 parasites/\u03bcl and 139 parasites/\u03bcl on day of recurrence). PRR for treatment failure calculated was 2.7/\u03bcl  , DebrezeP.vivax is known to digest haemoglobin [The changes in parasite density that occur following treatment indicate therapeutic response to anti-malarial drugs. Parasite clearance from the blood reflects the stage specificity and intrinsic potency of the anti-malarial drugs used , 25, in moglobin , in thismoglobin , 18, 19 P. vivax. Because, the drug used was confirmed to have good quality and CQ can prevent the re-infection up to 35\u2009days, the possible causes of treatment failure in this study might be due to either mal-absorption of the drug or relapse [Relapse, mal-absorption of drug, poor drug quality and re-infection could be factors contributing to treatment failure of  relapse . Even thP.vivax malaria in the study area. However, the authors believe there is a need to monitor CQ resistance by employing molecular tools for better evaluation of treatment outcomes. Despite the fact that Ethiopia is in elimination phase malaria, it is worth mentioning that prevalence of P.vivax observed in the study area requires strong malaria intervention measures.It is important to survey and intervene CQ treatment failures before it become a public health problem. CQ remains efficacious for the treatment of"}
+{"text": "Plasmodium falciparum infections detected by rapid diagnostic tests in semi-immune adults fall and stabilize at low density levels during the first 4 days after detection, suggesting a rapid decline of potential transmissibility.Afebrile  Plasmodium falciparum infections usually remain undetected and untreated in the community and could potentially contribute to sustaining local malaria transmission in areas aiming for malaria elimination.Afebrile P. falciparum infections detected with rapid diagnostic test (RDTs) were followed for 28 days. Kaplan-Meier estimates were computed to estimate probability of parasite positivity and of reducing parasitemia by half of its initial level by day 28. Trends of parasite densities quantified by microscopy and real-time quantitative polymerase chain reaction (qPCR) were assessed using Poisson regression models, and the microscopy-to-qPCR positivity ratio was calculated at each time point. Three survival distributions  were used to evaluate their strength of fit to the data and to predict the median lifetime of infection.Thirty-two men with afebrile P < .001). The ratio of P. falciparum\u2013positive samples by microscopy to qPCR decreased from 0.9 to 0.52 from recruitment to day 28. The best model fit to the data was obtained assuming a Gompertz distribution.The cumulative probability of parasite qPCR positivity by day 28 was 81% . Geometric mean parasitemia at recruitment was 516.1 parasites/\u03bcL and fell to <100 parasites/\u03bcL by day 3, reaching 56.7 parasites/\u03bcL on day 28 (P. falciparum infections detectable by RDT in semi-immune adults fall and stabilize at low-density levels during the first 4 days after detection, suggesting a rapid decline of potential transmissibility in this hidden parasite reservoir.Afebrile NCT02698748 Plasmodium falciparum parasitemia is commonly defined as the presence of asexual parasites in the blood without symptoms or illness [afebrile infections. Although fever-less infections may still cause other less evident symptoms with a substantial burden on the infected individual\u2019s health [Asymptomatic  illness . These is health , individs health . Proactis health , indicats health .P. falciparum infections into adulthood through which the pathogenesis of the infection is minimized while the parasite densities remain relatively low [Albeit poorly understood, there are several parasite and host factors that could contribute to maintaining high community rates of afebrile malaria infections . In highvely low , thus levely low , the higvely low , 11 of tvely low , 13, mayvely low , 14.P. falciparum infections found that infections detected by microscopy lasted from a few weeks up to several years and that parasite densities and symptoms were reduced soon after the onset of infection [Several observational studies conducted in the early 1900s in malaria-endemic countries to quantify the durability of nfection . Howevernfection , 20, 21 nfection , 22\u201326, nfection , 26.P. falciparum infections is needed to design effective interventions that can successfully detect and tackle this parasite reservoir at community level [P. falciparum infections and contribute to the ongoing discussion of the relevance of afebrile infections as a barrier for malaria elimination.A better understanding of natural dynamics of afebrile ty level . Herein,P. falciparum infections recruited between January 2015 and June 2015 were treated with placebo and observed for 28 days in the context of a randomized, placebo-controlled clinical trial conducted in the district of Manhi\u00e7a (southern Mozambique) [Adult men with afebrile ambique) . This peHealthy men were screened at the household level using a Histidine Rich Protein 2 (HRP2)\u2013based rapid diagnostic test (RDT) from SD Bioline. All positive cases by RDT were subsequently confirmed by expert microscopy at the Centro de Investiga\u00e7\u00e3o em Sa\u00fade de Manhi\u00e7a (CISM). Asymptomatic malaria was defined as the absence of any proactively referred symptom of disease according to the screened individual, together with a documented axillary temperature <37.5\u00b0C. Enrolled individuals were randomly assigned to the CQ or placebo arm and followed up on days 1, 2, 3, 7, 14, 21, and 28 after enrollment. All individuals in the placebo group were treated at the end of follow-up according to national guidelines, and rescue treatment was administered to participants of either study arm that presented with clinical symptomatology during follow-up. Thick and thin Giemsa-stained blood slides and 50 \u00b5L of blood spotted onto filter papers were collected at every visit.P. falciparum was also assessed from filter paper blood-spots through real-time quantitative polymerase chain reaction (qPCR) assay targeting 18S ribosomal RNA [P. falciparum 3D7 in vitro culture ring stage. Samples without amplification (no Ct detected) were considered negative. A negative control with no template DNA was run in all reactions.Slides were examined by 2 independent microscopists and considered negative if no parasites were seen after examination of 200 oil-immersion fields in a thick blood film. A third read was conducted in cases where the first 2 readings were discrepant. Parasite density was estimated using the Lambar\u00e9n\u00e9 method , which comal RNA . ParasitData from participants assigned to the placebo group were analyzed using Stata 14  and R Statistical Software . Basic characteristics of participants at enrollment were summarized using arithmetic or geometric means and medians, as determined by the distribution of the data. Kaplan-Meier estimates were computed to measure the cumulative probabilities of parasite positivity by day 28 and of failure to reduce parasitemia levels by at least 50% of their initial value by day 28\u2014or half-life probability. An infection was considered to be cleared at the time of the first negative observation by qPCR and when parasites were no longer observed for the subsequent follow-up observations. For infections in which parasite densities were reduced by >50% of their initial value by the end of follow-up, the last time point at which parasite densities were observed to fall below this threshold was considered as the time at which half-life was reached. Geometric mean parasite densities (GMPDs) and 95% confidence intervals (CIs) were calculated for each day of follow-up. A Poisson regression random-effect model was used to measure the variation of the parasite densities measured by qPCR at each day of follow-up. Finally, the proportions of infections that were positive by qPCR or by microscopy were calculated, and a ratio of positive microscopy to qPCR samples was estimated.We evaluated the strength of fit to the data of 3 survival distributions considered by previous groups to model natural malaria infections  , 32 in owww.ClinicalTrials.gov (NCT02698748).The protocol, consent forms, and questionnaires were approved by the CISM local ethics committee, the Ethics Committee of the Hospital Cl\u00ednic of Barcelona (HCB/2015/0122), the National Bioethics Committee of Mozambique  and the Mozambican Pharmaceutical Department (Ref./No. 4110/380/DF2014) before their implementation. All participants signed an informed consent form prior to the initiation of any study related activities. The clinical trial was registered at This analysis used data from 32 of the 38 individuals originally enrolled in the placebo group of the clinical trial. Three individuals with only a single follow-up observation and 3 individuals who developed a fever after day 1 (n = 1) or day 3 (n = 2) of follow-up were consequently excluded from this analysis. Three individuals had missing qPCR data , and 9 skipped 1 day of follow-up  .The average age of participants at enrollment was 25 years. The median body temperature was 36.4\u00baC, and the average body weight was 55.6 kg. The geometric mean parasite density on day 0 was 733parasites/\u03bcL as measured by microscopy and 516.1parasites/\u03bcL as measured by qPCR .P < .001) , with only 5 of the 32 individuals becoming negative by qPCR during follow up: 1 on day 2, 1 on day 14, 1 on day 21, and 2 on day 28 . The GMP < .001) . Ten (31 < .001) , and 5 m < .001) .All infections detected by microscopy were also detected by qPCR. The ratio of positive samples by microscopy to qPCR decreased from 0.9 (n = 28/31) on day 0 to 0.52 (n = 11/21) on day 28 . OverallThe Akaike\u2019s information criterion values obtained after modeling the infection dynamics suggest that the best model fit to the data was obtained assuming a Gompertz distribution . The KolP. falciparum\u2013infected afebrile adults in southern Mozambique followed for 28 days after detection of infection by RDT shows that most study participants remained infected until the end of the study without developing fever and were able to control parasite densities at submicroscopic levels after the first few days of follow-up, following a Gompertz distribution. These findings suggest a rapid decline of potential transmissibility and detectability of recently acquired infections in semi-immune individuals in endemic areas, offering novel information about natural parasite dynamics that can be used to design malaria elimination interventions and model their impact.This longitudinal analysis of Despite not achieving full parasite clearance, most individuals were able to halve their parasite densities within the 28-day follow-up period. In fact, parasite densities drastically decreased during the first 4 days of follow-up to levels below the typical microscopy and RDT detection threshold (100 parasites/\u03bcL) and remained below this parasitemia level until the end of the study. Similar patterns have been reported by Bruce-Chwatt et al, who measured daily parasitemia by microscopy after the acquisition of infection among Nigerian adults . A studySeveral studies have observed a direct association between asexual parasitemia and gametocyte carriage . In thisThe ratio of infections detected by microscopy to those detected by qPCR rapidly decreased between day 0 and day 28 from 0.9 to 0.52, suggesting that approximately half of the afebrile infections may not be detected a few days after infection unless a more sensitive diagnostic tool is used. Blood samples that were positive by microscopy and qPCR had a GMPD of 351.1 parasites/\u03bcL, compared with 13.8 parasites/\u03bcL observed in blood samples that were only positive by qPCR. In addition, we show that afebrile infections that become chronic in semi-immune adults linger below 100 parasites/\u03bcL and above 5 parasites/\u03bcL based on the qPCR detection threshold.P. falciparum natural infections, in line with previous reports [Mathematical modeling of the data showed the Gompertz distribution as the best fit for  reports , 32. Thi reports . DiffereP. falciparum\u2013untreated infections for a longer period of time. Third, the study did not collect extensive clinical data, such as hemoglobin levels, to assess the adverse impact of these afebrile  chronic infections [This analysis is subject to a series of limitations that should be considered when interpreting the results. First, the exact time at which each participant acquired the infection could not be assessed. Thus, the survival analysis was performed assuming the same duration of infection since recruitment for each individual at every follow-up time point. A study using microscopy data had previously identified that untreated parasite densities in immune individuals tend to decrease in a short period of time, thus suggesting that infections that could still be identified through an RDT or microscopy could be considered recent and assumed to be of similar duration at the time of recruitment . Our obsfections . Additiofections , 36. Finfections , were noP. falciparum infections in semi-immune adults tend to transform into submicroscopic infections soon after being detected by RDTs or microscopy. Although the role of these low-density infections in sustaining local transmission is still widely unknown, the association of gametocytemia with parasite biomass suggests a short duration of potentially high transmissibility before reductions in parasite densities [In conclusion, this study suggests that afebrile ensities , 38. Nevensities , or on tClinical Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at Supplementary MaterialClick here for additional data file."}
+{"text": "Delayed haemolysis is a frequent adverse event after treatment with artesunate (AS). Removing once-infected \u201cpitted\u201d erythrocytes by the spleen is the most accepted mechanism of haemolysis in these cases. However, an increasing number of cases with positive direct antiglobulin test (DAT) haemolysis after AS have been reported.All malaria cases seen at Hospital Clinic of Barcelona between 2015 and 2017 were retrospectively reviewed. Clinical, parasitological and laboratory data from patients treated with intravenous artesunate\u2014specifically looking for delayed haemolysis and DAT\u2014was collected.Among the 36 severe malaria patients treated with artesunate at the hospital, 10 (27.8%) developed post-artesunate delayed haemolysis. Out of these, DAT was performed in six, being positive in four of them (at least 40%). DAT was positive only for complement\u2014without IgG\u2014suggesting drug-dependent immune-haemolytic anaemia of the immune-complex type. Three of the four patients were treated with corticosteroids and two also received blood transfusion, with a complete recovery.Drug-induced auto-immune phenomena in post-artesunate delayed haemolysis may be underreported and must be considered. The role of corticosteroids should be reassessed. Plasmodium falciparum severe malaria after demonstrating its superiority over quinine, in terms of reduction of mortality and neurological sequelae, in multicentre randomized clinical trials, with no serious drug-related adverse events initially reported [Intravenous artesunate (AS) has been established as the first-line treatment for reported \u20133. Howevreported .In spite of conflicting case-definitions , 6, postThe objective of this report was to review the clinical and immunohaematological characteristics of patients with positive DAT in the context of PADH.All the 83 consecutive patients diagnosed with malaria attended at Hospital Cl\u00ednic of Barcelona, Spain, between January 2015 and December 2017, were retrospectively reviewed. Thirty-nine per cent of them were migrants or visiting friends and relatives (VFR) and the remaining 61% were tourists or business travellers. Of these, specific clinical, parasitological and laboratory data was collected from the 36 patients diagnosed with severe malaria and treated with AS. Severe malaria was defined based on World Health Organization (WHO) malaria severity criteria [The direct antiglobulin test (DAT), also known as direct Coombs test, was performed by the tube method with polyspecific antiglobulin  and monospecific anti-IgG and anti-C3d/C3c .PADH was diagnosed in 10 of the 36 (27.8%) malaria patients treated with intravenous AS. Median age of the affected patients was 45\u00a0years (interquartile range (IQR): 33.25\u201354.25\u00a0years) with 80% being males. All patients lived in non-endemic areas and acquired the infection through travelling to malaria-endemic areas. No patients were considered semi-immune by the study investigators .Plasmodium falciparum malaria cases in which it was performed (see below), which represents at least 40% (4/10) of patients with PADH and 11.1% (4/36) of patients treated with AS. No IgG antibodies were found in plasma, and a cold agglutinin with limited thermal range was detected in one of these four patients. Three of the four patients with positive DAT were admitted to hospital and treated with high doses of corticosteroids, and two of them also required blood transfusion. All patients successfully recovered from the haemolysis and no relapses occurred. Due to the mildness of the anaemia, the remaining patient did not require any treatment. Interestingly, the four patients with positive DAT had initially presented with higher parasitaemia than the remaining six PADH patients .DAT was positive for complement and negative for IgG in four of the six Cases with Coombs positive haemolytic anaemia after AS are described below.P. falciparum (parasitaemia 14.8%). The patient was diagnosed with severe malaria based on hyperparasitaemia and treatment with intravenous AS (2.4\u00a0mg/kg) was started. However, in the blood smear performed 8\u00a0h after AS initiation parasitaemia increased to 28.6%. In spite of not fulfilling the WHO criteria of AS resistance, treatment was changed to intravenous quinine (10\u00a0mg/kg/8\u00a0h after a loading dose of 20\u00a0mg/kg) and doxycycline (100\u00a0mg/12\u00a0h), under the clinical suspicion of\u00a0decreased susceptibility of P. falciparum to AS. Blood smear performed 72\u00a0h after starting this treatment was negative. Quinine was interrupted after 5 doses due to ototoxicity and treatment was completed with atovaquone/proguanil for 3 more days.A 55-year-old European traveler presented to the hospital with a 2-day course of fever after travelling throughout Zambia, Malawi, Mozambique and South Africa without taking anti-malarial prophylaxis. A blood smear detected Plasmodium molecular screening by polymerase chain reaction (PCR) were negative. Laboratory tests showed haemolytic anaemia with hyperbilirubinaemia, elevation of LDH and undetectable haptoglobin. Haemoglobin levels decreased to 5.1\u00a0g/dL and DAT was positive only for the C3 fraction. Other causes of haemolytic anaemia were ruled out: no schistocytes were observed on the blood extension, glucose-6-phosphate dehydrogenase (G6PDH) deficiency and presence of antinuclear antibodies were discarded, as were other infectious aetiologies, such as common viral and bacterial infections. Treatment with corticosteroids with a loading dose of 250\u00a0mg of methylprednisolone and 1\u00a0mg/kg/day of prednisone was started. Transfusion of 4 erythrocyte concentrates was also required. The patient presented good clinical evolution allowing the discontinuation of corticosteroids after 2\u00a0months. Finally, the initial suspicion of AS resistance could not be proven by mutations on kelch13-propeller domain (k13).Eight days after being discharged (and 14\u00a0days after the first dose of AS) the patient was admitted again for a 4-day course of fever, jaundice and dark urine. Two consecutive blood smears and a P. falciparum malaria 11\u00a0days after returning from Sierra Leone. He did not properly take the anti-malarial chemoprophylaxis with atovaquone/proguanil as prescribed. The patient was diagnosed with severe malaria due to hyperparasitaemia (4.7%), acute kidney injury and hyperlactacidaemia, that prompted a rapid initiation of intravenous AS (2.4\u00a0mg/kg). Although clinical and analytical improvement were observed, malaria blood smears conducted at 12 and 24\u00a0h after AS initiation showed a steady increase in parasitaemia up to 8.7%. Under the suspicion of AS resistance, treatment was changed to intravenous quinine (10\u00a0mg/kg/8\u00a0h) and doxycycline (100\u00a0mg/12\u00a0h) after the 4th dose of AS. Ten days after admission, the patient was discharged asymptomatic and in good clinical condition. Sequencing of k13 showed no molecular markers associated to artemisinin resistance. A careful re-evaluation of malaria smears showed pyknotic non-viable forms due to the patient\u2019s inability to remove erythrocytes from the bloodstream by \u201cpitting\u201d. A blood smear on day 28 was finally negative. During follow-up, microcytic anaemia (haemoglobin nadir of 9.8\u00a0g/dL at day 20) with hyperbilirubinaemia, increased LDH and undetectable haptoglobin was observed. DAT was positive for complement and cold agglutinin was detected in plasma. Given the mildness of the anaemia and the absence of symptoms, no treatment was required. A complete recovery was confirmed by laboratory tests after 2\u00a0weeks.A 55-year-old splenectomized European traveller was admitted to the intensive care unit because of severe Plasmodium falciparum malaria based on hyperparasitaemia (45%), acute kidney injury and respiratory distress was made. The patient was admitted to the intensive care unit under mechanical ventilation, renal replacement therapy and 2.4\u00a0mg/kg of intravenous AS. After 5 doses a blood smear was negative. Anti-malarial treatment was completed with intravenous quinine and doxycycline for 5 more days due to oral intolerance. Despite the good parasitological response, renal disfunction progressed requiring renal replacement therapy. Thirteen days after AS initiation anaemia with haemolytic pattern was detected, with a nadir haemoglobin level of 6.7\u00a0g/dL at day 17. Glucose-6-phosphate dehydrogenase deficiency, schistocytes and other causes of haemolytic anaemia were ruled out and DAT resulted positive for the C3 fraction. The patient received a blood transfusion without anaemia improvement. Due to the kidney injury a renal biopsy was performed and reported as compatible with interstitial nephritis. Three pulses of 250\u00a0mg methylprednisolone were then prescribed. Regarding the anaemia, although a blood transfusion had previously been administered, haemoglobin concentration did not raise until corticosteroid treatment was initiated. The patient was finally discharged with a daily dose of 40\u00a0mg of prednisone and complete recovery was observed within the following month.Eight days after returning from Namibia, Botswana, Zimbabwe, Malawi and Mozambique, a 40-year-old European traveller was admitted to the hospital for a 6-day course of fever and chills. He did not take anti-malarial prophylaxis. Diagnosis of severe P. falciparum malaria based on hyperparasitaemia (25%), acute kidney injury, respiratory distress and hyperbilirubinaemia. Following the hospital protocols, after the first intravenous AS dose, red blood cell exchange was performed. Subsequently, four more doses of intravenous AS were administered. The patient had an excellent clinical, analytical and parasitological evolution with a parasite clearance time of 70\u00a0h. Anti-malarial treatment was then completed with a 3-day course of dihydroartemisinin/piperaquine. Seven days after being discharged the patient had to be readmitted due to haemolytic anaemia, with a nadir haemoglobin level of 7.7\u00a0g/dL at day 16. DAT was positive for the C3d fraction of the complement and daily treatment with 100\u00a0mg methylprednisolone was started. Finally, a blood transfusion was performed after the DAT was negative. The patient was finally discharged with haemoglobin levels of 8.4\u00a0g/dL. Two weeks after, the patient remained in good clinical condition and the haemoglobin levels raised up to 9.6\u00a0g/dL.A 49-year-old European man who frequently travelled to West Africa for business was admitted to the intensive care unit of the hospital after travelling to Liberia. He had not taken anti-malarial prophylaxis. The patient was diagnosed with severe Post-artesunate delayed haemolysis is increasingly being reported after anti-malarial treatment with artemisinins. In this series of 36 patients treated with parenteral AS, PADH occurred in over a fourth (27.8%) of patients treated with AS. Of these, DAT was positive in 4 of the 6 patients in whom the test was performed, which amounts to at least the 40% of patients with PADH. The true incidence of positive Coombs haemolysis after AS treatment is unknown. Since mechanical destruction of red blood cells (RBC) has been assumed as the main mechanism of haemolysis after AS, DAT has not been systematically performed and it has only been occasionally reported in most PADH case series. As a consequence, positive Coombs haemolytic anaemia after AS raises as a probable misdiagnosed entity.Interestingly, in these 4 cases, DAT was positive only for complement\u2014without IgG\u2014as it was in two previously reported cases with specific information about the test , 10\u201316  and concentration of once-infected erythrocytes after treatment, has been proposed as the main mechanism for PADH . These oFurthermore, in all but one of the 4 patients with a positive DAT, PADH was severe enough as to require hospital admission and RBC transfusion. Treatment with corticosteroids in these 3 patients was followed by a rapid resolution of the haemolytic crisis, an outcome already observed in 6 of the 8 previously reported patients with PADH and positive DAT . As a consequence, DAT positivity due to malaria itself, although less probable, cannot be completely ruled out. The inclusion of a control group could have contributed to assess this problem. On the other hand, other causes of haemolysis such as drug-induced haemolysis due to different drugs such as beta-lactams, which were prescribed in the four patients, or quinine, which was given to 3 of the 4 patients, cannot be excluded. This is a hypothesis-generating study highlighting the importance of performing DAT when haemolysis after AS is detected.These cases, together with those previously published, highlight the possible contribution of autoimmune mechanisms to haemolysis due to artemisinins in selected cases and reopen the debate about the possible benefit of corticosteroids \u201313, 16."}
+{"text": "The World Health Organization recommends regular therapeutic efficacy studies (TES) to monitor the performance of first and second-line anti-malarials. In 2016, efficacy and safety of artemether-lumefantrine (AL) for the treatment of uncomplicated falciparum malaria were assessed through a TES conducted between April and October 2016 at four sentinel sites of Kibaha, Mkuzi, Mlimba, and Ujiji in Tanzania. The study also assessed molecular markers of artemisinin and lumefantrine (partner drug) resistance.Plasmodium falciparum kelch 13 (Pfk13) (codon positions: 440\u2013600) and P. falciparum multi-drug resistance 1 (Pfmdr1) genes , markers of artemisinin and lumefantrine resistance, respectively.Eligible patients were enrolled at the four sites, treated with standard doses of AL, and monitored for 28\u00a0days with clinical and laboratory assessments. The main outcomes were PCR corrected cure rates, day 3 positivity rates, safety of AL, and prevalence of single nucleotide polymorphisms in Pfk13 and Pfmdr1 genes, respectively. Six (1.9%) had non-synonymous mutations in Pfk13, none of which had been previously associated with artemisinin resistance. For Pfmdr1, the NFD haplotype  was detected in 134 (39.0%) samples; ranging from 33.0% in Mlimba to 45.5% at Mkuzi. The difference among the four sites was not significant (p\u2009=\u20090.578). All samples had a single copy of the Pfmdr1 gene.Of 344 patients enrolled, three withdrew, six were lost to follow-up; and results were analysed for 335 (97.4%) patients. Two patients had treatment failure (one early treatment failure and one recrudescent infection) after PCR correction, yielding an adequate clinical and parasitological response of\u2009>\u200998%. Day 3 positivity rates ranged from 0 to 5.7%. Common adverse events included cough, abdominal pain, vomiting, and diarrhoea. Two patients had serious adverse events; one died after the first dose of AL and another required hospitalization after the second dose of AL (on day 0) but recovered completely. Of 344 samples collected at enrolment (day 0), 92.7% and 100% were successfully sequenced for Pfk13 mutations, but there was a high prevalence of a Pfmdr1 haplotype associated with reduced sensitivity to lumefantrine (but no reduced efficacy was observed in the subjects). Continued TES and monitoring of markers of resistance to artemisinin and partner drugs is critical for early detection of resistant parasites and to inform evidence-based malaria treatment policies.The study indicated high efficacy of AL and the safety profile was consistent with previous reports. There were no known artemisinin-resistance Trial Registration ClinicalTrials.gov NCT03387631 Despite a decline of malaria burden over the past decade, malaria remains a major public health threat , 2. An ePlasmodium falciparum [Plasmodium falciparum kelch 13 (Pfk13) gene mutations [Artemisinin-based combination therapy ACT is recommended by the World Health Organization (WHO) for the treatment of uncomplicated malaria caused by lciparum . The curlciparum . Tanzanilciparum , and it lciparum . Studieslciparum \u201312, Kenylciparum , Uganda lciparum , Rwanda lciparum , Burundilciparum , and in lciparum \u201319 have lciparum  or Plasmutations \u201323 are rutations , 24, yieP. falciparum genome as markers of resistance to artemisinins and partner drugs of the currently used artemisinin-based combinations. A number of single nucleotide polymorphisms (SNPs) in the Pfk13 gene have been shown to confer resistance to artemisinins [I, N458Y, M476I, Y493H, R539T, I543T, P553L, R561H and C580Y) have been validated as markers of parasite resistance to artemisinins in Southeast Asia [P. falciparum multidrug resistance 1 (Pfmdr1) gene appear to be associated with decreased susceptibility to lumefantrine [Pfmdr1 copy numbers has been suggested to be associated with reduced susceptibility to lumefantrine [Due to the threat of artemisinin drug resistance, WHO recommends regular surveillance  to monitor the performance of anti-malarials in malaria endemic countries. Furthermore, the WHO recommends molecular surveillance based on known polymorphisms (including copy number variations) in the misinins , 26; andast Asia . Polymorfantrine , 29. Genfantrine .In Tanzania, the National Malaria Control Programme (NMCP) and its partners have been implementing therapeutic efficacy studies (TESs) to monitor the efficacy and safety of different anti-malarials , 32, incThis study was carried out at four of the eight NMCP sentinel sites  between April and September 2016. The study sites , about 100\u00a0km west of Dar es Salaam . Kibaha has low malaria transmission (<\u200910%) as reported in previous population surveys \u201336. MalaMkuzi health centre is located in Muheza district of Tanga region, in northeastern Tanzania. The malaria epidemiological profile of Muheza district has been well characterized and a detailed description given elsewhere , 38. MkuUjiji health centre is located in Kigoma urban district of Kigoma region, on the eastern shores of Lake Tanganyika in the northwestern part of Tanzania. Despite a decline in malaria transmission reported from 2007 onwards \u201336, 39, Mlimba health centre is located in Kilombero district of Morogoro region and the areas around this site have been extensively studied under the Ifakara Health Institute demographic surveillance system over the past two decades \u201343. KiloThis was a single-arm prospective in vivo study designed to assess the therapeutic efficacy and safety of AL for the treatment of uncomplicated falciparum malaria and the markers of artemisinin and lumefantrine resistance. Malaria transmission has been decreasing in some of the sites compared to the past and, therefore, thresholds of parasitaemia for low transmission areas recommended by WHO  were used. The age of study participants (6\u00a0months to 10\u00a0years) was also broadened , 46.Per the WHO protocol, sample size estimates assumed 5% of the enrolled patients would have treatment failure after treatment with AL. At a confidence level of 95% and an estimate precision of 5%, a minimum sample size of 73 patients was required to detect a failure rate\u2009\u2264\u20095% . With a At each of the study sites, potential participants were screened at the outpatient departments using malaria rapid diagnostic tests and microscopy as previously described . PatientTwo blood slides were collected, and one of the slides was stained with 10% Giemsa for 10\u201315\u00a0min and examined by microscopy to detect presence of and an estimated density of malaria parasites. The second blood slide was stained with 3% Giemsa for 30\u201345\u00a0min and used to determine the actual parasite density, species, and presence of gametocytes. Parasitaemia was measured by counting the number of asexual parasites against 200 leucocytes in thick blood films and detection of the different parasite species was done on thin films. Parasite density per \u00b5l of blood was calculated by multiplying the total count by 40, assuming that 1\u00a0\u00b5l of blood had a mean count of 8000 leucocytes . When mo\u00ae, Beijing Novartis Pharma Ltd, Beijing China; provided by WHO) for 3\u00a0days. Weight-based dosing based on one of three ranges  was done using a fixed dose combination of 20\u00a0mg of artemether and 120\u00a0mg lumefantrine per tablet, thus patients were given one, two or three tablets from smallest to heaviest, respectively. A full course of AL consisted of 6 doses given twice daily (8\u00a0h apart on day 0 and 12-h on days 1 and 2). Patients were observed for 30\u00a0min to ensure they did not vomit the study drugs. If vomiting occurred, a repeat dose was given after vomiting stopped. Any patient who persistently vomited the study medication was withdrawn and treated with quinine (injection/intravenous) or artesunate injection according to the national guidelines for management of complicated and severe malaria [Enrolled patients were treated with AL  when patients felt unwell. Parents/guardians were informed and encouraged to bring their children back to the clinic whenever they were unwell without waiting for scheduled visits. Patients who did not show up for their scheduled visits by mid-day were visited at home by a member of the study team and asked to come to the health facility. If a patient had travelled and could not be traced for scheduled follow-up, he/she was classified as lost to follow-up. During the visits, both clinical and parasitological assessments were performed and DBS were also collected. Patients with recurrent infections occurring on day 7 and afterwards were treated with quinine  or artesunate injection based on clinical presentation as per WHO protocol .The safety of AL was monitored by both passive and active methods through interviews with parents/guardian and clinical/laboratory assessments during the 28\u00a0days of follow-up. This enabled investigators to capture and record adverse events (AEs) or severe adverse events (SAEs) that occurred after treatment. During scheduled visits, parents/guardians were directly interviewed and asked to report the occurrence, nature, and incidence of any events occurring at home between the follow-up visits. Clinicians took a history, observed patients, and performed a clinical examination during follow-up visits at the study sites. Laboratory tests were appropriately requested and done to determine and capture AEs/SAEs. The reported/captured events were recorded in respective case report forms for each follow-up visit. Any SAE occurring during the study was reported by the principal investigator to the sponsor , NMCP, and the Tanzanian Medical Research Coordinating Committee (MRCC) of NIMR  within 24\u00a0h of its occurrence. Reporting of SAEs was done regardless of whether the principal investigator considered the events to be related to the investigated drug or not. Patients with AEs or SAEs were thoroughly assessed and managed accordingly, and the events were also assessed to determine their association with the study drugs. According to the WHO protocol , an AE wPfk13 propeller domain (codon positions: 440\u2013600) and Pfmdr1  were analysed for SNPs. SNPs were called only if they fit the following criteria: (i) they were found on both the forward and reverse reads, (ii) they had a\u00a0p -\u00a0value of\u2009<\u20090.0001 , and (iii) they had a minimum strand bias p-value of\u2009<\u20090.0005 when exceeding 65% strand bias, as some errors from sequencing machines are more likely to happen on nearby upstream bases. Mixed-infection and/or heterozygous calls were excluded from the analysis. The 3D7 Pfk13 and Pfmdr1 were used as reference sequences. Detection of Pfmdr1 copy number variants was performed using an Agilent Mx3005 real-time PCR machine  according to previously described protocols [Molecular analysis was performed on all samples collected upon enrolment (day 0) and during follow up in the case of treatment failure. Parasite genomic DNA was extracted from DBS using QIAamp blood mini-kits  according to the manufacturer\u2019s instructions. Molecular markers of anti-malarial drug resistance and microsatellite markers were analysed at the Centers for Disease Control and Prevention (CDC) Malaria Laboratory in Atlanta, USA. Sanger sequences generated in this study were analysed using the Geneious software package . Raw sequence reads were cleaned using default settings, and reads with high-quality scores  below 70% were discarded from further analysis. The rotocols , 48.\u00ae V2.6.3  [Samples from 65 patients with recurrent infections were analysed to determine genetic diversity in the study populations using six neutral microsatellite markers  by nested PCR for all except C2M34-313 and C2M69-383 . Fragment size was measured by capillary electrophoresis on ABI 3033 (Applied Biosystems) and scored using GeneMarkerPA, USA) ). PairedPA, USA) . A recruPfmdr1 and Pfk13 genes. Treatment outcomes were classified as either early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), or adequate clinical and parasitological response (ACPR) before and after PCR correction; based on per protocol method and Kaplan\u2013Meier analysis.The primary end point was PCR corrected parasitological cure on day 28 as per WHO protocol , while sEthical clearance was obtained from MRCC of NIMR, while permission to conduct the study at the health facilities was sought in writing from the relevant regional and district medical authorities. Ethical clearance form CDC was not required because the assessments done at the CDC Malaria Laboratory, using samples without linked identifiers, were determined by the CDC Center of Global Health\u2019s Human Research Protection Coordinator to not constitute engagement in human subjects\u2019 research. Detailed information of the study and benefits as well as its risks were explained to each study participant. Oral and written informed consent were obtained from parents or guardians of all patients before they were screened for possible inclusion into the study. The study was retrospectively registered at ClinicalTrials.gov, No. NCT03387631 (on 2nd January 2018) because of problems within the system which did not allow to register it before the study was launched.10 transformed parasite density at enrolment and age, among the four sites were compared using t test or analysis of variance\u2014ANOVA  or Mann\u2013Whitney U/Kruskal\u2013Wallis test . The prevalence of different haplotypes or SNPs in the Pfmrd1 and Pfk13 genes as well as Pfmdr1 copy numbers were reported. For the Pfmrd1 gene, the analysis focused on the three SNPs  and their corresponding haplotypes (particularly NFD), which have been associated with reduced susceptibility to lumefantrine [YYY) to resistant haplotype (NFD) on day zero and day of recurrence with adjustment for age of patients and the study site . For all statistical tests and comparisons, a p-value\u2009<\u20090.05 (two tailed) was considered to be significant.Data were entered into a Microsoft Access database at the study sites followed by a second entry which was done centrally at NIMR Tanga Centre after the end of data collection. The data were later validated, cleaned, and analysed using STATA for Windows version 11 . Descriptive statistics such as percentages, mean, median, standard deviation, and range were reported as appropriate. In order to automatically generate the treatment outcomes , the data were appropriately formatted and transferred to the WHO Excel software template . Baselinfantrine , 29. LogA total of 963 children were screened between April and October 2016. All sites recruited 88 children except Kibaha, which enrolled 80, making a total of 344 enrolled at the four sites  were lost to follow-up and three (0.9%) withdrew, leaving 335 (97.4%) patients that reached the study end points who were used in per protocol analysis  had mutations in the Pfk13 gene and only 6 (1.9%) samples had non-synonymous mutations, of which none were similar to previously reported SNPs associated with artemisinin resistance. The six Pfk13 non-synonymous mutations included one patient from Mkuzi (R471S), two at Mlimba (A578S and E433D), and three at Ujiji (one with I416V and two with Q613E). All patients with parasitaemia on day 3 post-treatment had infections with Pfk13 wild-type at enrolment. For Pfmdr1, codons N86Y, Y184F and D1246Y were analysed and the prevalence of the different SNPs are presented in Table\u00a0F mutant was present between 34.1% (Mlimba) and 46.6% (Mkuzi) in all four sites  samples, and its prevalence ranged from 33.0% in Mlimba to 45.5% at Mkuzi, but there were no significant differences (p\u2009=\u20090.578) among the four sites  during recurrent infections. Although patients with the NFD haplotype at baseline had more recurrent infections, the difference was not significant, even after adjusting for age and study site . Both patients with PCR-corrected treat failure (one with ETF and the second with a recrudescent infection) had NFD haplotype at enrolment. The overall median day of recurrent infection was 21\u00a0days  and this was similar among the four sites. Recurrent infections among individuals with NFD haplotypes at enrolment had a median of 23\u00a0days compared to 21\u00a0days for NYD, but the difference was not statistically significant (p\u2009=\u20090.417). Analysis of Pfmdr1 copy number variants was done on all 65 individuals with recurrent infections; all samples had a single copy of the gene.Out of 344 samples collected at enrolment (on day 0), 92.7% (n\u2009=\u2009319) and 100% (n\u2009=\u2009344) were successfully sequenced for This study reports high efficacy of AL (PCR corrected ACPR\u2009>\u200998%), suggesting that the ACT has retained its ability to treat uncomplicated malaria despite its use in Mainland Tanzania for more than 10\u00a0years. Previous studies conducted at these and other sites in Tanzania reported similarly high efficacies of AL . HoweverThis study also showed that AL had a safety profile comparable to previous studies and was well tolerated with minimal AEs. Most of the AEs were minor and mainly reported at the two sites of Mkuzi and Ujiji. Studies conducted in Tanzania , 12 and Two patients had serious adverse events on the day of presentation (day 0), with one child from Mkuzi dying after the first dose of AL. The cause of death was not established and its possible association with AL treatment could not be ascertained. A similar incident of death was reported in a TES which was conducted at Nagaga in 2012 [Pfk13, a molecular marker of artemisinin resistance [Pfk13 non-synonymous mutations in this study was higher than in a previous study which reported a prevalence of 1.2% [Pfk13 at enrolment. These findings do not suggest that artemisinin resistance has emerged in Tanzania [Pfk13 mutations might be increasingly accumulating in Tanzanian parasite populations and thus continued surveillance will be required to monitor the trends of Pfk13 polymorphisms and their possible association with reduced efficacy of artemisinins.Analysis of sistance , reveale of 1.2% . All patTanzania \u201323 and oTanzania . HoweverPfmdr1 gene showed that 39.0% of the parasites tested had the N86/Y184F/D1246 (NFD) haplotype, a combination associated with reduced susceptibility to lumefantrine in some studies [Pfmdr1 SNPs had higher prevalence than what was reported by previous studies in Mainland Tanzania [Analysis of the  studies . This coTanzania  and ZanzTanzania ; howeverPfmdr1 copy number with reduced susceptibility to lumefantrine [Pfmdr1 gene. This could be related to the fact that mefloquine has never been used as a routine first-line therapy in the study areas.Although studies have associated elevated fantrine , 47, 57,Pfk13 mutations or amplification of the Pfmdr1 gene were identified. A high prevalence of a Pfmdr1 haplotype was detected but corresponding subjects with treatment failure were not found. Continued TESs and monitoring of markers of resistance to artemisinin and partner drugs is critical and should be sustained to facilitate early detection of resistant parasites and to inform evidence-based malaria treatment policies.The findings of this study showed that the efficacy of AL remains high in Tanzania despite the use of this combination for more than 10\u00a0years. The safety of AL was consistent with previous reports and no known artemisinin-resistance"}
+{"text": "Plasmodium falciparum has emerged and spread in Southeast Asia. In areas where resistance is established longer courses of artemisinin-based combination therapy have improved cure rates.Artemisinin resistance in The standard 3-day course of artemether\u2013lumefantrine (AL) was compared with an extended 5-day regimen for the treatment of uncomplicated falciparum malaria in Kayin state in South-East Myanmar, an area of emerging artemisinin resistance. Late parasite clearance dynamics were described by microscopy and quantitative ultra-sensitive PCR. Patients were followed up for 42\u00a0days.P. falciparum kelch13 propeller gene (k13) were found in 46% (70/152) of infections, with F446I the most prevalent propeller mutation . Both regimens were well-tolerated. Parasite clearance profiles were biphasic with a slower submicroscopic phase which was similar in k13 wild-type and mutant infections. The cure rates were 100% (70/70) and 97% (68/70) in the 3- and 5-day arms respectively. Genotyping of the two recurrences was unsuccessful.Of 154 patients recruited (105 adults and 49 children <\u200914\u00a0years) 78 were randomized to 3\u00a0days and 76 to 5\u00a0days AL. Mutations in the k13 mutations, the current first-line treatment, AL, was still highly effective in this area of South-East Myanmar. The extended 5\u00a0day regimen was very well tolerated, and would be an option to prolong the useful therapeutic life of AL.Despite a high prevalence of Trial registration NCT02020330. Registered 24 December 2013, https://clinicaltrials.gov/NCT02020330The online version of this article (10.1186/s12936-018-2404-4) contains supplementary material, which is available to authorized users. Plasmodium falciparum has emerged in South-East Asia and now extends over a large area of the Greater Mekong sub-region from the coast of Vietnam to the Eastern border of India , artefenomel OZ439] and KAF156 have shown promising results in phase 2 trials but are still under development \u201312. In t39 and KAversus time, or its surrogate the day 7 concentration, is the principal determinant of cure in acute falciparum malaria [Artemether\u2013lumefantrine (AL) is the first-line ACT in Myanmar. Artemether is mainly biotransformed to dihydroartemisinin , 15, the malaria . When AL malaria . Co-admi malaria . For pat malaria . This prFailure of AL in falciparum malaria has not been reported in Myanmar to date despite the emergence of artemisinin resistance and the decline in susceptibility to mefloquine, which shares resistance mechanisms with lumefantrine , but recP. falciparum (including mixed infections) and 3540 for Plasmodium vivax. Two VHV were trained to make blood-smears and to follow up patients with falciparum malaria for 3\u00a0days after treatment with AL. Of 78 patients followed up 11 (14%) were still positive by microscopy on day 3. Another study in this area has reported k13 mutations in 48% of 42 patients studied [This study was conducted between November 2013 and February 2015 in two remote village tracts in Kyainseikgyi Township in Kayin State. Kayin state is situated in Southeast Myanmar, along the Myanmar-Thailand border. In 2012, Medical Action Myanmar, an international NGO, started malaria control activities in villages in this township through a network of 114 Village Health Volunteers (VHV), who tested all fever patients for malaria at community level with a rapid diagnostic test (RDT). Falciparum malaria was routinely treated with 3\u00a0days artemether\u2013lumefantrine and a single gametocytocidal dose of primaquine according to national policy. In 2012\u20132013 these VHV tested 52,720 patients with a malaria RDT of whom 7541 (14.3%) tested positive; 4001 for 3. The main exclusion criteria were signs of severe or complicated malaria [Patients with fever (tympanic temperature >\u200937.5\u00a0\u00b0C) or a history of fever, aged between 6\u00a0months and 65\u00a0years, with microscopy confirmed uncomplicated falciparum or mixed malaria, were invited to take part in the study if they had a parasite count between 80 (\u2265\u20095/500 WBC on a thick film) and 175,000 asexual parasites per mm malaria , haemogl\u00ae) at standard doses twice daily3\u00a0days AL (Coartem\u00ae)3\u00a0days AL plus a one gram capsule of fish oil  was given to all patients on the 1st\u00a0day of treatment. The initial AL treatment dose was given under supervision. All subsequent doses were given to the patient to be taken at home. The importance of taking these drugs, even when the symptoms had subsided, was explained clearly. At initial treatment, if the patient vomited within 30\u00a0min, the full dose was repeated. If vomiting occurred after 30\u00a0min but within an hour, half the dose was repeated.Pfkelch13. Patients were then seen on days 3, 5, 6 and 7, when 3\u00a0mL (1\u00a0mL from children\u2009\u2265\u20096\u00a0months\u2009<\u20095\u00a0years old) of venous blood was taken for parasite DNA quantitation with uPCR [On enrolment, blood was taken for DNA extraction and full length sequencing of ith uPCR  . A symptom questionnaire including a list of specific questions related to adverse events was completed at each visit. Patients presenting again to the clinic with a microscopy confirmed fections .msp1, msp2 and glurp [After the first 39 patients had been enrolled, the uPCR collection time points were changed based on new information from another trial, which showed that 75% of patients with falciparum malaria still tested positive for parasite DNA with uPCR, on day 9 after ACT . To charnd glurp .P. falciparum positivity in the 1st\u00a0week detected by uPCR method or a difference in cure rates from 10 to 30% with 95% confidence and 80% power, 75 patients were required in each treatment arm. Data were entered into a web-based database Macro, (InferMed). Data cleaning and analysis were done using Stata14 (StataCorp) and GraphPad Prism 6 (GraphPad Software Inc.). Data were analysed by Student\u2019s t-test, Wilcoxon-rank sum test and chi-squared test as appropriate. Survival data were analysed using the Kaplan\u2013Meier method and Cox regression. Logistic regression was used to examine the relationship between treatment outcomes and mutation genotypes. A 5% significance level was used for all statistical tests.In order to detect a 20% difference in A total of 1311 patients were screened for eligibility criteria and 154 (105 adults and 49 children\u2009<\u200914\u00a0years) patients were enrolled  of all patients were still parasitaemic by microscopy on day 3 after starting treatment: 23% (18/78) of the AL3 treatment arm and 17% (13/76) in the AL5 treatment arm. On day 5, two of 78 patients were still positive after AL3 and 0 (0/76) after AL5. By day 7 all patients were microscopy negative.P. falciparum parasitaemia detected by microscopy during follow up, both after AL5. A 6\u00a0years old child who was symptomatic on day 28, and a 24\u00a0year old male who was asymptomatic on day 42. Neither patient received fish oil. DNA extraction from blood samples taken from both patients was unsuccessful so genotyping to confirm recrudescence or reinfection could not be carried out. Therefore, the recurrence rate was 2.9%  in the AL5 arm and 0%  in the AL3 arm).Only two patients had a recurrence of P. vivax at baseline. These P. vivax infections were all cleared by day 3. Four patients, one of whom had a mixed infection detected at presentation, had a P. vivax recurrence (presumed relapse) during follow up; 2 at day 35 and 2 at day 42 (1 after AL3 and 3 after AL5).Three patients had mixed infections with The profile of submicroscopic late phase parasite clearance estimated by uPCR showed a biphasic pattern in many patients with no difference between the two treatment arms at any time point Table\u00a0. During k13 gene was sequenced successfully in 152 (98.7%) of which 73 (48%) had a k13 mutation. There was no difference in k13 mutation prevalences between the 3 and 5\u00a0days treatment groups (48.7% (37/76) vs 47.4% (36/76), p\u2009=\u20090.87). Seventy (96%) of the mutations were concentrated within propeller blades 1\u20134 in the k13 gene. Fourteen patients had parasites with a \u2018confirmed\u2019 artemisinin resistance mutation according to the WHO classification: C580Y (8), R561H (6), while 38 patients carried parasites with so-called \u2018associated\u2019 artemisinin resistance mutations F446I (20), G449A (9), P553L (5) P441L (3), and P574L (1).Of the 154 patients\u2019 samples taken at enrollment, the k13 mutations (p\u2009=\u20090.49). On multivariable logistic regression >\u20091% parasitaemia (23/154) on admission was significantly associated with day 3 blood smear positivity , whereas infection with a k13 mutation was not (p\u2009=\u20090.25). k13 mutations did not affect the slow sub-microscopic phase of parasite clearance; at 21\u00a0days after treatment, 12.1% (7/58) patients with the kelch13 propeller mutation had detectable parasitaemia by uPCR compared to 12.8% (6/47) of those with wild type (p\u2009=\u20090.91)   had Gametocytaemia was detected by microscopy on admission in 17 of 78 (22%) of the AL3 arm and 16 of 76 (21%) of the AL5 arm (Table\u00a0The mean (SD) haemoglobin concentrations on admission in the AL3 and AL5 treatment arms were 12.2 (1.9) g/dL and 12.0 (2.2) g/dL respectively (p\u2009=\u20090.656). After 28\u00a0days of treatment, the mean (SD) haemoglobin values were 12.4 (1.7) g/dL for the 3-day and 12.5 (1.6) g/dL for the 5-day treatment arms.A total of 35 adverse events were reported, 18% (14/78) for AL3 and 13% (10/76) for AL5. There was no significant difference between the two treatment regimens (p\u2009=\u20090.39). Dizziness was the most commonly reported adverse event , followed by headache . All adverse events were only of mild or moderate severity and all patients recovered fully. No serious adverse events were reported.P. falciparum has spread across mainland South-East Asia and led to the failure of ACT in the eastern Greater Mekong subregion and along the Thailand-Myanmar border. There is natural concern over the continued efficacy of ACT in Myanmar, which bears the greatest burden of malaria in the region. However, in this study conducted in Eastern Myanmar the effectiveness, and thus the efficacy of the current first-line treatment artemether\u2013lumefantrine was excellent, despite a high prevalence of the k13 mutations which are associated with artemisinin resistance. These results are similar to those of a previous study in Myanmar conducted in 2009  [Artemisinin resistance in y 90.3%) . Artemety 90.3%) . Furthery 90.3%) , 9.Although lumefantrine efficacy was still preserved in this location at the time of the study, it is likely that resistance will worsen eventually. The recent reduction in malaria transmission in Eastern Myanmar as a result of active intervention programmes and substantially improved ACT availability will reduce population immunity and this could compromise therapeutic responses. The longer course and thus higher dose of artemether\u2013lumefantrine evaluated here was well tolerated and, if deployed, would be expected to provide greater efficacy if resistance does worsen, and thus prolong the useful therapeutic life of this important anti-malarial drug. In general anti-malarial drug policy changes occur late in the course of resistance emergence, and the delays result in increased transmission, increased morbidity, and compromise of the opportunity to modify the dose regimen to maintain efficacy. In Thailand in the early 1990s mefloquine resistance was already far advanced before the dose was increased from the original 15\u00a0mg/kg to the now generally accepted 25\u00a0mg/kg dose \u201330. The k13 parasites (artemisinin sensitive) and k13 mutant parasites (artemisinin resistant), and it was not affected by length of treatment. Persistent low density gametocytaemia is one possible explanation, but a single gametocytocidal dose of primaquine was given to all patients at the start of the treatment. Primaquine is rapidly gametocytocidal and this results in rapid clearance of gametocyte mRNA [Highly sensitive PCR methods (uPCR) allow quantitation of parasitaemia at densities one thousand times lower than microscopy , 31. Thiyte mRNA . It theryte mRNA , 36. It yte mRNA  althoughk13 mutations in the south-eastern part of Myanmar, the effectiveness of the current first-line treatment of 3\u00a0days artemether\u2013lumefantrine remained high. This may be explained, at least in part, by the predominance of the F446I k13 mutation causing an intermediate resistance phenotype [Although almost half the falciparum malaria infections in this region of Myanmar bore henotype . How lonk13 mutations, such as the C580Y mutation, in a \u201cfitter\u201d genetic background take over, as they have further east [If more resistant her east , then paAdditional file 1. Contains trial profile, Additional figures, Additional results tables, and a summary of ultrasensitive PCR quality control procedures."}
+{"text": "Plasmodium falciparum malaria. Its efficacy has been extensively assessed in clinical trials. In routine healthcare settings, however, its effectiveness can be diminished by delayed access to treatment and poor adherence. As well as affecting clinical outcomes, these factors can lead to increased transmission, which is the focus of this study.Artemether-lumefantrine (AL) is the most widely-recommended treatment for uncomplicated P. falciparum to include gametocytes, the parasite forms responsible for onward transmission. The model includes a pharmacokinetic\u2013pharmacodynamic model of AL, calibrated against both immature and mature gametocytes using individual-level patient data, to estimate the impact that delayed access and imperfect adherence to treatment can have on onward transmission of the parasite to mosquitoes.We extend a within-host model of Using survey data from seven African countries to determine the time taken to acquire antimalarials following fever increased our estimates of mean total infectivity of a malaria episode by up to 1.5-fold, compared with patients treated after 24\u2009hours. Realistic adherence behaviour, based on data from a monitored cohort in Tanzania, increased the contribution to transmission by 2.2 to 2.4-fold, compared with a perfectly-adherent cohort. This was driven largely by increased rates of treatment failure leading to chronic infection, rather than prolonged gametocytaemia in patients who have slower, but still successful, clearance of parasites after imperfect adherence to treatment. Our model estimated that the mean infectivity of untreated infections was 29\u201351 times higher than that of treated infections (assuming perfect drug adherence), underlining the importance of improving treatment coverage.Using mathematical modelling, we quantify how delayed treatment and non-adherent treatment can increase transmission compared with prompt effective treatment. We also highlight that transmission from the large proportion of infections which never receive treatment is substantially higher than those treated. Plasmodium falciparum worldwide, has been proven in clinical trials to be efficacious and well-tolerated.Artemether-lumefantrine (AL), the most widely-recommended treated for uncomplicated In routine healthcare settings treatment efficacy can be diminished by delayed access to drugs, or imperfect adherence to the dosing regimen, although this effect is difficult to quantify.Using individual-level adherence data collected in Tanzania as input to our within-host model of a malaria infection treated with AL, we find that the onward transmission from a treated cohort of patients is more than double that of a perfectly-adherent cohort.In our model delayed access to treatment, determined from data on the time taken to acquire antimalarials following fever in malaria-endemic countries, increased onward transmission by up to 50%, compared with patients treated 24\u2009hours after becoming febrile.We underline the importance of good adherence and prompt access to treatment, by showing how they can reduce onward transmission of the malaria parasite.Our modelling framework also allows us to compare the transmission capacity of treated patients with the much greater contribution to transmission made by untreated patients, emphasising the importance of extending treatment coverage.Although significant declines in malaria prevalence have been achieved this century, recent evidence suggests that progress in reducing the burden of disease has stalled. In 2017 there were an estimated 219\u2009million cases of malaria globally, compared with an estimated 217\u2009million the year before.Plasmodium falciparum, the human malaria species responsible for a large proportion of the global mortality. There are currently five WHO-approved ACTs available, all of which require a 3-day dosing regimen.Artemisinin-combination therapies (ACTs) are the frontline treatments for uncomplicated cases of P. falciparum malaria in order to obtain an estimate of the impact of imperfect adherence to AL on treatment outcomes.In previous work we developed a within-host model of uncomplicated Anopheles mosquito in order for onward transmission to occur. Treatment with an ACT can therefore reduce the probability that the patient will contribute to transmission after treatment. This is in contrast to other older treatments such as chloroquine and sulfadoxine-pyrimethamine, which have much weaker action against gametocytes.Although the primary goal of an ACT is to clear the asexual parasite population and, therefore, restore the patient to health, artemisinin-based drugs also have a partial killing effect against the sexual-stage parasites (gametocytes), clearing developing gametocytes and reducing the circulation time of mature gametocytes. Both a female and male gametocyte, in their mature forms,P. falciparum, the delay between the appearance of asexual parasites and mature gametocytes in the peripheral blood is longer than in the other human malaria species.In P. falciparum parasites to treat their syphilis infections.To assess the extent to which transmission increases due to delays to treatment and imperfect adherence, here we have extended our within-host model of asexual parasitaemiaP. falciparum infection  remain in the bloodstream. Using the malaria therapy dataset, detailed modelling work has been carried out to scrutinise these quantities, comparing models with a range of complexity.10.1136/bmjgh-2019-001856.supp1Supplementary dataThe malaria therapy dataset contains daily measurements, stated as an integer number of parasites per \u00b5l, of both asexual and sexual parasitaemia for each patient, measured by microscopy.et al.To describe the drug concentrations observed following treatment with AL, we used a population-PK model due to Hodel Here et al,To quantify the drug effect on gametocytes, we used data by Goncalves et al,We estimated the probability that a feeding mosquito would be successfully infected by using a recent study by Bradley To investigate the potential effect of delay to treatment in endemic settings, we used data from cross-sectional household surveys in subSaharan Africa in 2016 and 2017 and in which caregivers were asked to report the time taken to obtain antimalarial medication following fevers in children under 5\u2009years old.There was no patient or public involvement in the development or execution of this research study.Gametocytaemia in untreated infections is determined by the fraction of asexual parasites committing to the sexual pathway, the maturation time for gametocytes and the circulation time of the mature gametocytes  that is, the latter have no symptoms when parasites recur. Overall, 12 out of 46 patients (26.0%) were classified as clinical failures.In the model, the timing of treatment is determined by the parasite density at which fever commences (the pyrogenic threshold), and the subsequent time taken to seek treatment.To investigate the potential effect of delay to treatment in endemic settings, we used data from cross-sectional household surveys in sub-Saharan Africa in 2016 and 2017 and in which caregivers were asked to report the time taken to obtain antimalarial medication following fevers in children under 5\u2009years oldPoor adherence to a treatment regimen slows the clearance of gametocytes after treatment, as well as increasing the probability that treatment fails to clear the asexual parasite population. We estimated mean infectivity for patients who completed treatment with those who missed doses, starting by removing the sixth dose, then removing the fifth and sixth dose and so on . In eachWe observe large increases in average infectivity when the treatment regimen is not completed . For exaet al, and gametocytes produced following treatment failure (see We further estimated the impact of imperfect adherence using individual-level data collected in Tanzania in 2012.lure see . Only thThe efficacy of malaria treatment and its impact on transmission potential are typically measured under controlled conditions, rather than in routine healthcare settings. Here, we estimated that onward transmission from a treated cohort of patients with realistic adherence patterns is more than double that of perfectly-adherent patients. The major driver of this increased transmission in our analysis is treatment failure without retreatment, leading to chronic subsequent infections of long duration. Imperfect adherence also led to slower clearance of gametocytes following treatment, regardless of whether the asexual parasites were successfully cleared, but this had a relatively small impact on transmission potential.We also looked at how delays in obtaining antimalarials following fever affects onward transmission, based on data from seven endemic countries. Assuming patients were fully-adherent, we found that using these waiting times in the model increased patients\u2019 capacity to transmit malaria by up to 1.5-fold, compared with patients treated after 24\u2009hours. This effect stems from the long maturation time of gametocytes in falciparum malaria, and the high susceptibility of immature gametocytes to artemisinin derivatives. Our analysis quantifies the role that prompt and effective treatment has in reducing onward transmission, and of course it also has a critical role in alleviating clinical symptoms and reducing the risk of severe disease which we did not quantify here. A limited amount of data was available for some countries, such as Ethiopia and Malawi, and timing of treatment is self-reported. Therefore, the data should be interpreted as illustrative of the range of current access times, rather than painting a detailed picture of access to antimalarials in the countries considered. Furthermore, access to treatment within a country is likely to be highly heterogeneous and change over time. As more data becomes available, more detailed models for this variable can be developed, rather than assuming a single distribution for each country.While timing and adherence are important within cohorts of treated patients, our analysis suggests that in situations where a significant proportion of infections remain completely untreated, the contribution to transmission from these infections may be much greater than contributions from treated infections. This has been found to be the case in a number of modelling studies.et al.et alet al also used a second infectivity model, due to Carter and GravesSome published within-host models, largely informed by malaria therapy data, contain both asexual and sexual parasites, and consider effects of antimalarial drugs on both parasite populations.Although we have only considered treatment with AL in this work, the framework outlined here would enable similar PD models to be fitted for other ACTs given appropriate data. In addition, interventions that are designed to target mature gametocytes rather than the asexual parasites, such as single low-dose primaquine, could also be assessed. To do this, however, the model would need to reflect the sterilising effect of some gametocytocidal drugs precedes the gametocyte clearance effects.There are several limitations to our analysis. Adherence and treatment-seeking behaviour may vary with age, which we have not explored here. There is evidence that adults receive more mosquito bites than young children, which would influence their relative transmissibility.Incorporating naturally-acquired immunity into well-calibrated within-host malaria models remains a challenging undertaking, due to the lack of longitudinal data from untreated infections. One model used cross-sectional data from malaria endemic areas to assess how acquired immunity reduces parasite densities in infections.We have used within-host modelling to show how delays in obtaining antimalarials and imperfect adherence to the dosing regimen can lead to increased transmission of the parasite. These effects are not readily measurable in clinical trial settings for clear ethical reasons. As well as quantifying how delayed treatment and imperfect adherence to the dosing regimen can increase a treated individual\u2019s capacity to transmit malaria, our work suggests that the major priority should be reaching patients who do not currently have access to treatment. The WHO\u2019s Global Technical Strategy for Malaria aims to reduce the global malaria burden by 90% by 2030,"}
+{"text": "P. falciparum infected patients at health centers/hospitals treated over the period of 2\u00a0years (2013\u20132014).The national treatment guidelines of Nepal have adopted Artemisinin Combination Therapies (ACTs) for the treatment of uncomplicated falciparum malaria since 2004. Emergence of Artemisinin resistance in the Greater Mekong Sub-region (GMS) and beyond may become a threat for Nepal as well. The main objective of this study was to assess the therapeutic efficacy of antimalarial drug artemether-lumefantrine in uncomplicated Giemsa stained thick and thin smears, prepared from uncomplicated falciparum malaria patients who visited the selected sentinel sites in Nepal during 2013 to 2014 and met the inclusion criteria that included parasitemia , were evaluated until 28\u00a0days after ACTs treatment, following a World Health Organization (WHO) therapeutic efficacy protocol. Based on the re-occurrence of fever and resurge in parasitemia, the study patients were classified as resistant or susceptible. Blood specimens on filter papers were further analyzed by Polymerase Chain Reaction (PCR), specifically for the K13 propeller gene mutation (a recently identified molecular marker for ACT resistance).P. falciparum  were enrolled in the study, of which 24 cases completed the post-treatment follow up for 28\u00a0days. Only one case out of 24 (4%) was identified as a late treatment failure (LTF). K13 mutation, a proxy indicator for ACT resistance in parasites, was not detected on the day 1, which indicates resistance had not yet reached the molecular level.A total of 56,013 suspected malaria cases were screened for this study. Of which, 120 (0.21%) were infected with falciparum malaria. Out of 120, 28 cases of Only one case of late treatment failure was identified in this study. ACT combination using artemether-lumefantrine was still effective for the treatment of uncomplicated falciparum malaria in Nepal. A close monitoring and supervision for ACT resistance is essential for future malaria treatment in Nepal. Plasmodium species [P. falciparum, is the deadliest one [Malaria is a global public health problem caused by  species . Accordi species . However species . Among t species , falcipaiest one  on the bArtemisinin or its derivatives or Artemisinin in combination with other drugs, is used as a first line drug for the treatment of uncomplicated falciparum malaria in many countries . In NepaPlasmodium falciparum. Clinical symptoms monitoring is classified as early treatment failure (ETF), late treatment failure (LTF), late parasitological failure (LPF) and adequate clinical and parasitological response (ACPR).Assessment of therapeutic efficacy entails in-vivo measurements of parasitemia in blood, coupled with the monitoring of clinical symptoms in patients undergoing the treatment with ACTs for 28\u00a0days . This reETF is defined as the presence of danger signs or severe malaria on day 1, 2 or 3 including parasitemia. LTF constitutes late clinical failure and late parasitological failure. Late clinical failure is defined as presence of danger signs or severe malaria including parasitemia on any day between day 4 and day 28. Late parasitological failure is defined as the presence of parasitemia on any day between day 7 and 28. ACPR is defined as absence of parasitemia by the end of the treatment (day 28) irrespective of axillary temperature without previously meeting any of the criteria of early treatment failure or late clinical failure or late parasitological failure \u201321. AntiLate treatment failure by microscopy needs additional monitoring for Artemisinin resistance by PCR. Artemisinin resistance is specifically associated with mutation on Klech 13 (K13) propeller region of the parasite and therefore, identification of this is essential . Range oIn the current context of emergence of Artemisinin resistance around the region, and the burden of malaria being mostly (45\u201360%) imported in Nepal from migrant workers in south and south east Asia, it is critical to assess and monitor the efficacy of ACTs. There have not been any studies in Nepal to explore the efficacy and resistance of ACTs. The main objective of this study was to assess the efficacy and resistance pattern of ACTs currently recommended by national malaria control program.Seti and Mahakali- far western region, Janakpur- central region and Mechi- eastern region) with the catchment of patients from various districts   Fig.\u00a0. The stuThis study was an open-arm prospective exploration of clinical and parasitological responses based on the WHO therapeutic efficacy protocol-2009 and past research , 26. AllAll patients received standard treatment at the hospital. Treatment with artemether-lumefantrine (each Coartem tablet containing artemether 20\u00a0mg and lumefantrine 120\u00a0mg) was provided based on the national malaria treatment protocol. Coartem was provided for a total of 3\u00a0days based on the body weight of the patients .Clinical data included a standard physical examination report that included body weight and axillary temperature. Thick and thin blood films for parasite counts were obtained and screened on day 0 to confirm adherence to the inclusion criteria. Parasite counts were examined using thick smear on day 0, 1, 2, 3, 7, 14, 21 and 28\u00a0days (follow up period). Adverse events such as severe anemia, black water urine, hypoglycemia, abnormal bleeding, acidosis, and hemoglobinuria were monitored throughout the follow up period.Molecular analysis for K13 mutation, using PCR, was recorded and further analyzed to confirm the presence of this mutation in parasites. Dried blood samples were collected onto Whatman filter paper No.1 and stored at room temperature in zip-lock plastic bags containing silica gel desiccant beads for further molecular analysis. The collected filter paper spots were sent to Mahidol University through the World Health Organization South East Asia Regional Office (WHO SEARO), for molecular analysis of the resistance targeting towards K13 mutation, as an indicator of resistance development.P. falciparum. Among 120 cases, 14 were excluded because of severe malaria  were confirmed as malaria. Out of 1, 447, one hundred and twenty cases were confirmed with .5% were Kailali sentinel site and 8  were enrolled from Kanchanpur sentinel site\u00a0 of them were enrolled into the study with first 3\u00a0days\u2019 admission at hospital and 28\u00a0days of parasitological monitoring. Among them, total 19  enrolled patients were from A total of 24 cases  completed the study protocol. Of total 24 enrolled cases, 23  cases responded well with Artemisinin combination therapy and were classified as ACPR whereas one 4.17%; 1/24) case was microscopically found to have parasitemia at day 28 and therefore classified as LTF  showed no parasitemia in day 3 following administration of artemether-lumefantrine. Low level of treatment failures was consistent with the studies in Ethiopia [This is a first study in Nepal assessing the therapeutic efficacy of ACTs to inform the national malaria treatment guidelines. In this study, only one case with late parasitological failure was found, however, it did not show any resistance markers for Ethiopia  and IndiEthiopia .Nepal has achieved a significant progress in malaria control and treatment in recent years , 31, unlP. falciparum are readily cleared after the administration of artemether-lumefantrine (Coartem\u00ae) and was consistent with the standard cure rate. These results are consistent with the studies carried out in India [Artemisinin combination generally has a high cure rate, typically exceeding 95% . Our finin India , and varin India \u201340.In PCR analysis, no mutation was detected in Kelch 13 propeller region. K13 propeller region is associated with resistance to Artemisinin . In thisPlasmodium falciparum to test therapeutic efficacy compromised the statistical power. While the emergence and potential spread of Artemisinin resistance from the GMS can become a public health disaster, findings from this study can reassure the current treatment guidelines with ACTs in Nepal. Future studies to update on the resistance markers can become valuable to assess the efficacy of ACTs.This study was jointly conducted by WHO and Epidemiology and Disease Control Division (EDCD) of Nepal to determine the therapeutic efficacy of ACTs in malaria endemic districts of Nepal. Nevertheless, a low prevalence of malaria and Plasmodium falciparum in Nepal and should be continued as a first line drug against P. falciparum. Nevertheless, therapeutic efficacy using both microscopy and PCR in future are necessary to monitor K13 mutation, a hallmark of Artemisinin resistance in P. falciparum.A high percentage of cases  with uncomplicated falciparum malaria, enrolled in this study showed adequate clinical and parasitological response, with only one case of parasitological failure. This study also showed that Artemisinin combination therapy (artemether and lumefantrine in combination) at the dose prescribed by National malaria treatment protocol was still effective. However, further continued monitoring is required in the context of emerging Artemisinin combination therapy resistance in the Greater Mekong Sub-region. Based on this study, Artemisinin combination therapy (ACT) is still effective against"}
+{"text": "Malaria control largely depends on availability of highly efficacious drugs, however, over the years, has been threatened by emergence of drug resistance. It is, therefore, important to monitor the impact of recurrent anti-malarial treatment on the long-term efficacy of anti-malarial regimens, especially in sub-Saharan African countries with high malaria transmission. Evaluation of parasite clearance following treatment of severe malaria with intravenous artesunate among patients in Eastern Uganda, was performed, as a contribution to monitoring anti-malarial effectiveness.Parasite clearance data obtained from a clinical trial whose objective was to evaluate the 42-day parasitological treatment outcomes and safety following treatment of severe malaria with intravenous artesunate plus artemisinin-based combination therapy among patients attending Tororo District Hospital in Eastern Uganda, were analysed. Serial blood smears were performed at 0, 1, 2, 4, 6, 8, 10, 12, 16, 20, 24\u00a0h, followed by 6-hourly blood smears post start of treatment until 6\u00a0h post the first negative blood smear when parasite clearance was achieved. Study endpoints were; parasite clearance half-life (the time required for parasitaemia to decrease by 50% based on the linear portion of the parasite clearance slope) and parasite clearance time .One hundred and fifty participants with severe malaria were enrolled. All participants were treated with intravenous artesunate. All study participants tolerated artesunate well with rapid recovery from symptoms and ability to take oral mediation within 24\u00a0h. No immediate adverse events were recorded. The median (IQR) number of days to complete parasite clearance was of 2 (1\u20132). The median (IQR) time to clear 50% and 99% parasites was 4.8 (3.61\u20137.10) and 17.55 (14.66\u201320.66)\u00a0h, respectively. The median estimated clearance rate constant per hour was 0.32. The median (IQR) slope half-life was 2.15 \u00a0h.Parasite clearance following treatment with intravenous artesunate was rapid and adequate. This finding provides supportive evidence that resistance to artemisinins is unlikely to have emerged in this study area. Continuous monitoring of artemisinin effectiveness for malaria treatment should be established in high malaria transmission areas in sub-Saharan Africa where spread of resistance would be disastrous.Trial registration The study was registered with the Pan African Clinical Trial Registry (PACTR201110000321348). Registered 7th October 2011, http://www.pactr.org/)The online version of this article (10.1186/s12936-018-2552-6) contains supplementary material, which is available to authorized users. Plasmodium falciparum, which causes the most severe form of disease is the most prevalent [Anopheles mosquito which injects sporozoites into the human host. Sporozoites undergo pre-erythrocytic and erythrocytic stages of multiplication causing erythrocyte rupture with release of merozoites and pro-inflammatory cytokines into circulation, which are responsible for the symptoms. Patients seek medical attention when they begin to experience symptoms of malaria. Accurate diagnosis and prompt initiation of effective anti-malarial drugs is key to successful treatment of malaria, however, host immune mechanisms play a crucial role in parasite elimination [Worldwide, malaria ranks as one of the most important causes of morbidity and mortality with 216 million cases and 445,000 deaths in 2016 alone, 90% of cases and deaths occur in Africa and 80% in sub-Saharan Africa Uganda is one of the heavy malaria burden countries where In addition to vector control, malaria control largely depends on treatment with highly efficacious drugs, but over the years, this has been threatened by drug resistance. The artemisinin compounds currently recommended for malaria treatment are highly effective, however, there is significant risk for development of drug resistance if their use is not adequately controlled and monitored. Resistance to artemisinins which manifests as loss of the intracellular ring stage parasite susceptibility and slower parasite clearance , was repMonitoring of artemisinin efficacy is performed using parasite clearance curves constructed from sequential parasite densities following malaria treatment . PreviouData obtained from a randomized clinical trial whose objective was to evaluate the 42-day parasitological treatment outcomes and safety following treatment of children with severe malaria using intravenous artesunate plus artemisinin combination therapy (artemether\u2009+\u2009lumefantrine or dihydroartemisinin\u2009+\u2009piperaquine) in Tororo District Hospital, in Eastern Uganda, were analysed . EasternFollowing diagnosis of severe malaria, participants were immediately initiated on intravenous artesunate  administered as 2.4\u00a0mg/kg at start of treatment, repeated at 12\u00a0h and every 24\u00a0h from start of treatment until the switch to oral therapy. Each 60\u00a0mg vial of artesunic acid was dissolved in 1\u00a0mL of 5% sodium bicarbonate and mixed with 5\u00a0mL of 5% dextrose, injected as a slow bolus into an indwelling IV cannula. Intravenous artesunate was administered for a minimum of 24\u00a0h and when participants could tolerate oral therapy, they were given a full course of either artemether\u2009+\u2009lumefantrine or dihydroartemisinin\u2009+\u2009piperaquine. All participants received paracetamol 15\u00a0mg/kg at 8 hourly intervals. Adjunctive and supportive treatment, such as diazepam for convulsions and dextrose for hypoglycaemia, was given in accordance with the Uganda Ministry of Health Malaria Treatment guidelines.Thin blood smears were performed to detect the malaria species at diagnosis. Thick blood smears were performed to calculate parasite density serially at 0\u00a0h (before start of treatment), 1, 2, 4, 6, 8, 10, 12, 16, 20, 24\u00a0h, followed by 6 hourly blood smears post start of treatment until 6\u00a0h post the first negative blood smear when parasite clearance was achieved. Thick blood smears were stained using 3% Giemsa for 30\u00a0min and read by experienced microscopists. All slides were read by two independent experienced microscopists and any discrepant results were reviewed by a third experienced microscopist. Parasite density was calculated by counting the number of asexual parasites per 200 white blood cells (WBCs) assuming a WBC count of 8000/\u03bcL of blood. Participants were discharged from hospital with a full course of either artemether\u2009+\u2009lumefantrine or dihydroartemisinin\u2009+\u2009piperaquine, when the blood smear was negative for malaria.e parasite count versus time was described mathematically using a polynomial model and lag phase identification done from the mathematical model. If a lag phase was not identified, the clearance rate constant was estimated as the absolute value of the slope of the linear regression model to all data. If a lag phase was identified, the clearance rate constant was then estimated as the absolute value of the slope from the linear part of the predicted profile.Data were entered and verified using MS ACCESS and analysed using STATA version 13.1 . Descriptive statistics were summarized into medians and interquartile ranges (IQR). Parasite density was normalized using logarithmic transformation. Parasite clearance was estimated using the World Wide Anti-malarial Resistance Network (WWARN) parasite clearance estimator (PCE) algorithm , 12. TheThe study was approved by Makerere University School of Medicine Research and Ethics Committee (REC REF 2011-175), Uganda National Drug Authority (369/ESR/NDA/DID-12/2011), Uganda National Council for Science and Technology (HS 1031) and registered with the Pan African Clinical Trial Registry (PACTR201110000321348). All study procedures were conducted according to Good Clinical Practice standards. Patients and parents or guardians of participants provided written informed consent prior to enrollment. Study related information was provided in the participants\u2019 local languages.Data were obtained from 150 participants with severe malaria, treated with intravenous artesunate between December 2011 and April 2013. Baseline characteristics are shown in Table\u00a0All study participants tolerated artesunate very well, with rapid recovery from symptoms and ability to take oral medication within 24\u00a0h. No immediate serious adverse events were recorded. None of the participants required additional medication. The median (IQR) number of days to complete parasite clearance (PCT) was of 2 (1\u20132). A total of 1559 (42.5%) parasite density-time profiles were included in the parasite clearance analysis. The median estimated clearance rate constant per hour was 0.32. The median (IQR) clearance half-life was 2.15 \u00a0h. None of the patients had clearance half-life greater than 5\u00a0h. None of the patients had parasitaemia on day 3. The time to clear 50%, 90%, 95% and 99% parasites is shown in Table\u00a0P. falciparum to artemisinin. It is possible that parasite clearance times were shorter than what has been documented from other studies because we evaluated parasite clearance following intravenous administration of artesunate, while most previous studies from Asia evaluated parasite clearance following oral administration of artemisinin treatment, however, the current findings are comparable to those from studies performed in the African countries in which oral artesunate was administered [Parasite clearance following treatment of severe malaria using intravenous artesunate, among patients attending Tororo District Hospital in Eastern Uganda was evaluated. Time to clear parasites was significantly shorter compared to that proposed as indicator for artemisinin resistance. Artemisinin resistance is suspected to occur when\u2009>\u200910% of patients treated with ACT or artesunate monotherapy have positive parasitaemia on day 3 or a clearance half-life of\u2009\u2265\u20095\u00a0h \u201311. Thesnistered , 16.k13 resistance mutations in\u2009>\u20095% of patients confirms artemisinin resistance [The short parasite clearance times did not warrant assessment for resistance conferring mutations. Presence of sistance \u201311. Thessistance  and provArtesunate has been shown to clear malaria parasites fast from circulation and is currently the recommended drug of choice for treatment of severe malaria , 18. PreAlthough parasite clearance is currently used as an indicator of anti-malarial drug effectiveness and resistance, some authors suggest serious limitations with its use since parasite clearance is dependent on a combination of host immune mechanisms, parasite biology and drug pharmacology. Host immunity is said to be the primary determinant of parasite clearance, suggesting that parasite clearance is an insensitive tool that may lead to overconfidence and underestimation of reduction in drug sensitivity especially in areas where host immunity to malaria may be well developed .This study has some limitations that should be considered. Majority of study participants weighed less than 20\u00a0kg, however, all received 2.4\u00a0mg/kg of artesunate at start of treatment, repeated at 12\u00a0h and every 24\u00a0h from start of treatment until the switch to oral therapy. Currently this dose of artesunate is considered low for children weighing less than 20\u00a0kg, because of differences in pharmacokinetics, and lower exposure to the active metabolite dihydroartemisinin with lower body weight , howeverParasite density could have been overestimated by counting the number of asexual parasites per 200 WBCs assuming a WBC count of 8000/\u03bcL of blood because malaria parasitaemia based on assumed WBCs is generally higher than parasitaemia based on actual WBCs due to changes in the WBC count over time which could alter the ratio and introduce error in the estimation of parasite density .These findings contribute to the existing knowledge on parasite clearance properties of intravenous artesunate for treatment of severe malaria and provide evidence that artemisinin resistance is unlikely to have spread to this study area. Continuous monitoring of artemisinin effectiveness for malaria treatment should be established in high malaria transmission areas in sub-Saharan Africa where spread of resistance would be disastrous.Additional file 1. Study Data set."}
+{"text": "The intermittent preventive treatment in infants (IPTi) trial that took place in Papua New Guinea showed an overall reduction of 29% of the risk of malaria when delivering single-dose sulfadoxine-pyrimethamine (SP) associated to 3 days of amodiaquine (AQ) every three months to children during the first year of life. The aim of the present study was to assess if the last two doses of AQ were truly administered as prescribed by the parents at home based on drug level measurement and PK modelling, which is a good proxy of medication adherence. It provides also important information to discuss the efficacy of the intervention and on feasibility of self-administered preventive malaria treatment.During the three-arm randomized double-blinded IPTi trial, each child was prescribed one dose of SP (day 0) and 3 doses of either AQ or artesunate (AS) at day 0, 1 & 2 adjusted to weight or placebo. Treatments were given at 3, 6, 9 and 12 months of age. The first day of treatment was delivered by nursing staff (initiation under directly observed treatment (DOT)) and the two last doses of AQ or AS by parents at home without supervision. For this cross-sectional study, 206 consecutive children already involved in the IPTi trial were enrolled over a 2-month period. At the time of the survey, allocation of the children to one of the three arms was not known. Blood samples for drug level measurement were collected from finger pricks one day after the planned last third dose intake. Only children allocated to the SP-AQ arm were included in the present analysis. Indeed, the half-life of AS is too short to assess if drugs were given on not. Because of the short half-life of AQ, desethyl-AQ (metabolite of AQ (DAQ)) measurements were used to investigate AQ medication adherence. Two PK (PK) models from previously published studies in paediatric populations were applied to the dataset using non-linear mixed effect modelling (NONMEM) to estimate the number of doses really given by the parents. The study nurse reported the administration time for the first AQ dose while it was estimated by the parents for the remaining two doses. Out of 206 children, 64 were in the SP-AQ arm. The adjusted dosing history for each individual was identified as the one with the lowest difference between observed and individual predicted concentrations estimated by the two PK models for all the possible adherence schemes. The median (range) blood concentration AQ in AQ arm was 9.3 ng/mL (0\u20131427.8 ng/mL), (Quartiles 1\u20133: 2.4 ng/mL -22.2 ng/mL). The median (range) for DAQ was 162.0 ng/mL (0\u2013712 ng/mL), (Quartiles 1\u20133: 80.4 ng/mL-267.7 ng/mL). Under the assumption of full adherence for all participants, a marked underprediction of concentrations was observed using both PK models. Our results suggest that only 39\u201350% of children received the three scheduled doses of AQ as prescribed, 33\u201337% two doses and 17\u201324% received only the first dose administered by the study nurse. Both models were highly congruent to classify adherence patterns.Considering the IPTi intervention, our results seem to indicate that medication adherence is low in the ideal trial research setting and is likely to be even lower if given in day-to-day practice, questioning the real impact that this intervention might have. More generally, the estimation of the number of doses truly administered, a proxy measure of adherence and an assessment of the feasibility of the mode of administration, should be more thoroughly studied when discussing the efficacy of the interventions in trials investigating self-administered malaria preventive treatments. Plasmodium falciparum and Plasmodium vivax.In Papua New Guinea (PNG), malaria transmission is high and the disease is responsible for several thousands of deaths each year. The number of estimated cases in 2013 was between 800\u2019000 and 2\u2019000\u2019000[Intermittent preventive treatment in infants (IPTi) aims to reduce malaria risk by delivering several antimalarial drugs, three or four times, coinciding with immunization planned by the Expended Program on Immunization (EPI) to children younger than 1 year, particuet al., they reported that it was common for mothers to forget giving the tablets or keep them for later use.As the parents at home directly administer the second and third treatment doses without supervision, overall the number of doses truly administered remains largely uncertain, which might have a direct impact on the efficacy of the intervention. From a perspective of implementing IPTi, as with any therapy with self-administered treatment, knowing the adherence level is a key element. There are several reasons why infants might not receive the correct amount of drugs: nurses not giving the right dosage or crushing the pill, which might induce some loss of the active ingredient, children vomiting the drugs or the parents not administering drugs at home. The latter reason might be especially common in the case of IPTi, as it is a preventive intervention and parents might find it less useful or might prefer keeping the drugs for later use in case of malaria-like symptoms onset. In a qualitative study performed in children of the present IPTI trial, about 15% of the mothers mentioned not having given the drugs according to instructions[Medication adherence is defined as the process by which patients take their medications as prescribed by their healthcare providers. MedicatThe aim of the present study was to assess if the last two doses of AQ were truly administered as prescribed by the parents at home based on drug level measurement and PK modelling, which is a good proxy of medication adherence and an assessment of the feasibility of self-administred malaria preventive treatments. It provides also important information to discuss the efficacy of the intervention. It was performed in a random sample of 206 study participants included in the randomized controlled trial performed in Papua New Guinea.http://www.clinicaltrials.gov (number NCT00285662) and formed part of the IPTi Consortium . In concordance with MRAC regulations, written informed consent was obtained from the parents both for the main IPTi study and the present work.The IPTi randomized controlled trial was conducted in PNG in the Mugil area of the Sumkar District, Madang Province, between June 2006 and May 2010 and included 1125 children. Each study participant received four courses of treatment at 3, 6, 9 and 12 months. Each course consisted of three days of treatment and each time, nursing staff delivered SP (single dose) and the first dose of AQ or AS  by initiation under directly observed treatment (DOT) and parents at home without supervision the others doses of AQ or AS (day 1 and 2). The tablets were given with sweet syrup after being crushed by the nurse or parents. The first drug dose (D0) was given a second time if the child vomited within 30 minutes. A community reporter living in the village was in charge of visiting the children to check that no adverse events occurred. He/she had no responsibility for giving drugs or supervising drug intake. All details concerning the trial and efficacy results have been published elsewhere[For the present cross-sectional study, 206 consecutive children already part of the IPTi trial, were enrolled over an approximate 2-month period. At the time of sampling, allocation groups in the IPTi trial were unknown. Indeed, patients were assigned blindly into one of the three trial arms , so we enrolled a large number of children aiming to have approximately 60 patients in each arm.Fig 1. An experienced study nurse also collected at home parents\u2019 estimated time of drug administration on day 1 and day 2 using a scale that divides day-time into 2-hour slices. Parents specifically consented to this additional blood collection, not included in the main trial.Nurses of the study team visited each of the 206 study participants at home one day (D3) after the theoretical last-dose taken (D2). To avoid biasing the results of the study, parents were not informed in advance of the visit. Blood samples for drug level measurement were collected from finger pricks. The course of the study is detailed in The blood samples of the 206 participants were used to measure the levels of IPTi drugs . Drug level measurements were conducted at the laboratory of the Division of clinical pharmacology of the University hospital of Lausanne, Switzerland, using a chromatography-tandem mass spectrometry method (LC-MS/MS) .Children included in the PK analysis are those whose parents reported having given drugs and with drug levels available. Each study participant was at a different stage of the study and thus received the treatment course at 3, 6, 9 or 12 months. We performed the analysis on patients in the AQ arm only. More specifically, we used desethyl-amodiaquine (metabolite of AQ (DAQ)) measurements to if drugs were truly administered by parents as AS and AQ concentrations are low or undetectable one day after drug administration because of their short half-lives  or partial  medication adherence. We calculated individual residuals (IRES) as the difference between predicted (Cpred) and (Cobs) observed concentrations in each of the four adherence schemes and identified the true dosing history for each child as the one associated with the lowest IRES value among the four possible adherence patterns. Children were thus categorized into groups according to the identified dosing history: subjects receiving a dose on day 0 , on days 0 and 1 (D_12) on days 0 and 2 (D_13), and those who completed the treatment (D_123). We estimated bias and precision of the model assuming theoretical full adherence and the model-corrected dosage history according to IRES values by mean prediction error (pred and Cobs as previously defined and N the number of observations) with the associated CI95%, and root mean squared error (RMSE = http://www.r-project.org/) was used for statistical analyses and figures generation.Non-linear mixed effect modelling ) was employed to predict individual DAQ concentration (NONMEM option MAXVAL = 0) . This prCobs)2N) . R v3.2.et al. [et al. [et al.. As described in Table 1, both models provide similar PK parameters. For both models, a very long DAQ terminal half-life  was estFig 2). Age, weight and gender characteristics of the study participants are described in Table 2 and compared to those of patients in both studies used as models for the PK analysis.Out of 206 children, 68 belonged to the SP-AQ arm. One was removed because parents said they did not give the third dose and three because blood was not collected, thus 64 children fulfilled the inclusion criteria (The median (range) AQ blood concentration in the AQ arm was 9.3 ng/mL (0\u20131427.8 ng/mL), (Quartiles 1\u20133: 2.4 ng/mL -22.2 ng/mL). The median (range) for DAQ was 162.0 ng/mL (0\u2013712 ng/mL), (Quartiles 1\u20133: 80.4 ng/mL-267.7 ng/mL).In comparison, the median (range) AQ blood concentration in the placebo arm was 0 ng/mL (0\u2013137.0 ng/mL), (Quartiles 1\u20133: 0 ng/mL -0.2 ng/mL). The median (range) for DAQ was 0.7 ng/mL (0\u2013202.1 ng/mL), (Quartiles 1\u20133: 0 ng/mL-6.3 ng/mL). Overall, 16/65 patients had detectable concentrations of AQ and 34/65 of DAQ.Fig 3 illustrates the observed vs. individual Hietala and Stepniewska model-predicted concentrations for the theoretical and adjusted dosing datasets.Assuming full adherence, we observed a marked under prediction of concentrations using both the Hietala and the Stepniewska models. The results suggest that 11 children (17%) are in adherence configuration D_1, 8 (13%) in D_13, 13 (20%) in D_12 and 32 (50%) received full treatment according to the Hietala model. Applying the Stepniewska model provided similar results, with 15 children (24%) in configuration D_1, 13 (20%) in D_13, 11 (17%) in D_12 and 25 (39%) received all three doses. The two models agreed on the adherence schemes for 80% of the children. Discrepant results were observed in thirteen children. For seven of them, the adherence schemes were D_123 vs. D_13, for four of them, D_12 vs. D_1, and for two of them, D_13 vs. D_12, comparing the Hietala vs. Stepniewska models, respectively.95%) was 20% (CI95% 4\u201338%) with a precision of 79% for the Hietala model and was 82% (CI95% 40\u2013138%) with a precision of 236% using the Stepniewska model. Under the assumption of model-identified adherence, the initial bias and precision of the Hietala model decreased to -4% (CI95% -14-6%) with a precision of 50% and to 2% (CI95% -14-20%) with a precision of 97% using the Stepniewska model and was thus not significantly different from zero. The observed vs. individual predicted concentrations under the assumption of each adherence configuration for both models are shown in The bias  than non-ingestion of the middle dose 13\u201320%). It is interesting to note that adherence to the full IPTI treatment in this study was very close to the levels found in the literature for real life adherence to long-term medication for chronic diseases (around 50%), a totally different context[\u201320%. It These results show that, even in the frame of a well-controlled clinical trial, adherence is far from optimal. Furthermore,Even if parents at home were responsible to administer the last two doses, a community reporter was responsible for visiting families each treatment day, checking for serious adverse events and collecting empty drug bags (thus serving indirectly as a reminder for drug intake). This type of check procedure is possible only in the frame of a trial, and certainly not in routine care. Although impossible to make clear assumptions one way or another, we may still wonder what the real efficacy of SP-AQ would have been if the adherence had been 100% instead of 50%.et al., it was observed that only 47 to 76% children received the first dose of SP in an area where IPTi is carried out. They also assumed that children without health cards did not receive IPTi and approximately one third did not receive IPTi at the same time as immunization as staff forgot to give the drug or it was out-of-stock[The results of this study also question the potential impact of a preventive intervention with self-administered treatment such as IPTi if implemented in a real-life context. Indeed, the observed adherence of 37% to 50% for the full treatment course is certainly the highest that one could expect in a less controlled context. Furthermore, for an intervention such as IPTi, most of the protection is due to the prophylactic effect long-acting drugs as mentioned in the main trial publication.  SP is th-of-stock. Thus, iOn the other hand, one might argue that the overall risk reduction observed is under-estimated due to the fact that many children take non-prescribed \u201cover-the-counter\u201d antimalarial drugs, especially AQ. In the present study, even if the measures of AQ blood concentrations revealed that half of the children in the placebo arm took \u201cover-the-counter\u201d AQ and almost all SP (data not shown), the concentrations were much lower than in the SP-AQ arm. It is likely therefore that SP and AQ \u201cover-the-counter\u201d medications had only a marginal effect on the overall protection in this study population even more as resistance to these two drugs were low at the time of the study.The reasons for only 50% or less of the children receiving all three doses of IPTi may be multiple. First, it could be related to the preventive nature of the intervention. Indeed, parents may decide to keep the antimalarial drugs for later use, in case of illness rather than giving it to their healthy child. Second, since AQ tastes very bitter, children might have vomited or spat out the drugs. Third, and most probably, parents may just have forgotten to give the treatment, especially as no immediate benefit of the treatment is expected. Lastly, almost half the children aged 3-months received the three doses while only one third of those aged 12 months did. These results seem to indicate that parents are more likely to give a preventive treatment to younger children than to older ones. Overall, the results of the study point out that self-administration of malaria preventive treatments is probably a suboptimal approach.More generally, our study showed that it is feasible and reliable to assess objectively malaria treatment adherence using PK modelling, even in complex research settings. We are not aware of other studies that use this approach to investigate malaria adherence in developing, at least for preventive treatments. Few studies have compared drugs levels at the end of curative treatment courses to make broad assumptions on adherence, but not to measure precisely the number of doses taken., 15 Our A first limitation is that a full PK model could not be built due to the scarcity of data and we performed the analysis based on previously published models in children. These models were derived from older children, mostly from Africa, and were not validated in younger children in PNG. However, in order to control for an \u201ceffect model\u201d, we tested both published models and found very similar results, which reinforces the reliability of the results. Second, the relatively small sample size of the study might not reflect the adherence of the entire cohort of patients. On the other hand, even if some uncertainties may remain on the exact proportion of patients with poor adherence, the objective of this study was more to provide estimations rather than an exact figure. Furthermore, this is probably a conservative estimate of the adherence, mainly because we cannot exclude that some parents may have administered the treatment to their child at last minute because they heard that study staff were coming (even if our visit was not announced). Lastly, some uncertainties exist about the exact time at which the drugs were taken. However, the careful history taking performed by the nurses allowed a time estimate of around 2 hours, which is sufficient for the purpose of this analysis, especially considering the long half-life of DAQ.This study showed that less than half of the study participants in an IPTi randomized controlled clinical trial took the full three-day course of antimalarial preventive medication. This is very low considering that the first dose was directly administered by a study nurse and the study was closely monitored. These results seem to indicate that medication adherence is low in an ideal research setting and therefore might be even lower if implemented in day-to-day practice, questioning the impact that this intervention might really have. It also reinforces the hypothesis that AQ add little to the overall efficacy of the IPTi intervention, which relies mostly on the long lasting effect of a single dose of SP. Finally, these results question the self-administration approach for preventive malaria treatments in children. This is relevant information, not only for the specific IPTi intervention, but also more generally for drug trials of this kind. Indeed, researchers should consider using this approach when designing new studies and estimating effect sizes of short malaria treatment courses.S1 Fig(DOCX)Click here for additional data file.S2 Fig(DOCX)Click here for additional data file."}
+{"text": "In Ethiopia, medicinal plants have been used to treat different diseases, including malaria, for many centuries. People living in rural areas are especially noted for their use of medicinal plants as a major component of their health care. This study aimed to study treatment-seeking and prioritize plants/plant recipes as anti-malarials, in Dembia district, one of the malarious districts in Northwest Ethiopia.Parents of children aged under 5\u00a0years who had had a recent episode of fever were interviewed retrospectively about their child\u2019s treatment and self-reported outcome. Treatments and subsequent clinical outcomes were analysed using Fisher\u2019s exact test to elicit whether there were statistically significant correlations between them.Allium sativum (Liliaceae), Justicia schimperiana (Acanthaceae), Buddleja polystachya (Scrophulariaceae) and Phytolacca dodecandra (Phytolaccaceae) were the most frequently used. Justicia schimperiana was the one associated with the best clinical outcomes (69% self-reported cure rate). However, the difference in clinical outcomes between the plants was not statistically significant.Of 447 children with malaria-like symptoms, only 30% took the recommended first-line treatment (ACT) , and 47% took chloroquine (85% cured). Ninety-nine (22.2%) had used medicinal plants as their first-choice treatment. J. schimperiana was associated with the highest reported cure rate of the plants. Further research is warranted to investigate its anti-malarial properties.In this study, only 30% of children took the recommended first-line treatment. 22% of children with presumed malaria were first treated with herbal medicines. The most commonly used herbal medicine was garlic, but  Ethiopia has made tremendous progress in reducing incidence and mortality from malaria. There was a rapid scale-up of long-lasting insecticidal nets (LLINs) and artemisinin-based combination therapy (ACT) in 2007, associated with a 73% reduction in malaria admissions and a 62% reduction in malaria deaths in children aged under 5\u00a0years . The HeaPlasmodium falciparum infections. Chloroquine remains the first-line treatment for Plasmodium vivax. Although national treatment guidelines recommend that this should be followed by a 2-week course of primaquine, in practice it is not routinely used because there are no widely available tests for G6PD deficiency. It is only used under the supervision of healthcare providers for patients with limited risk of malaria infection in the future, such as those who are not living in malaria endemic areas. A recent review showed that in Ethiopia, 98.1% of patients with P. falciparum were successfully treated with AL and 94.7% of patients with P. vivax were successfully treated with CQ [P. vivax [P. falciparum has reduced rapidly since the introduction of AL, prevalence of P. vivax has risen slightly [Because of increasing chloroquine resistance, in 2004 Ethiopia adopted artemether\u2013lumefantrine (AL) as the first-line treatment for slightly .P. falciparum, but Ethiopia is home to the second highest number of cases and mortality due to P. vivax in the world (after India). There is still room for progress: use of insecticide-treated bed nets in children under 5 ranges from 45 to 69% [Therefore, malaria remains a leading public health problem in Ethiopia. It is estimated that about 75% of the total area of the country and 65% of the population is at risk of infection [5 to 69% , 9. The One possible explanation for this is the use of herbal medicines. Traditional medicinal plants are an integral part of the variety of cultures in Ethiopia ; up to 8The classical way of identifying medicinal plants for further research is through ethnobotanical studies. Yet conventional ethnobotanical studies rarely involve clinicians. They could and should provide much more clinical information if the ultimate goal is to know which one, among numerous treatments for a given ailment, has the best effects. Although identification of the plants is usually of a good standard, definition of the diseases which they treat is not. There is rarely sufficient questioning about the observed patient status and progress, perceived efficacy and limitations of the remedies, and whether these are indeed the \u2018treatment of choice\u2019. Many plants are \u2018supposed\u2019 to be good for one disease or another, but are not actually the preferred treatment used in everyday life. The \u2018Retrospective Treatment Outcome Study\u2019 (RTO) was designed circumvent these problems . This adThis study aimed to measure the frequency of use of different treatments, and associated outcomes, in children under 5\u00a0years of age with a recent episode of fever  in a rural district of Ethiopia.2 . According to demographic data of the district, Aberjeha, Chenker and Tezeba were the most malarious kebeles with populations of 10,490, 6520 and 6213, respectively, and with incidence rates of 7.7%, 7.0% and 6.6%, respectively . These were selected for the study. The proportion of under five children was estimated to be 14% (3251) of the total population.Dembia is a rural district in Northwest Ethiopia, covering a total area of 127,000\u00a0kmThe sample size for the study was determined using the formula for a single population proportion. It was estimated that 50% of the population would be using plants, and required a 95% level of confidence. Therefore, sample size was determined as follows:In order to allow for an estimated 15\u201317% non-response, the final sample size was increased to 451. The selection of respondents was performed in order to ensure that they were representative of the whole population, with a corresponding proportion of houses to be randomly visited until the desired sample size was reached.During the peak malaria season, from November to December 2013, the parents of children aged under 5\u00a0years who had had symptoms of uncomplicated malaria (mainly fever) within the previous 2\u00a0months were interviewed. Data was collected using a structured and pre-tested questionnaire which was developed by Graz et al.  and subsData were coded, checked for completeness and consistency, entered using EPI-INFO\u21227 statistical software and then exported to SPSS version 20 for further analysis. Descriptive statistics of the collected data  was done. Statistical correlation with reported clinical recovery was also computed using Fisher\u2019s exact test.The study was carried out after getting permission from the ethical review board of the University of Gondar (R/C/S/V/P/05/239/2013). Then a letter of permission was obtained from the district health office and local administrator, and the individual household heads were invited to give written informed consent before the interview. Confidentiality was respected by keeping the privacy of the respondents while filling the questionnaire. Seriously ill children were advised to visit a health facility. Any personally identifiable information  was not entered into the database.Among 451 patients who were approached for the study, 4 either did not complete the interview or were not willing to be included in the study. Finally, 447 were included, resulting in a response rate of 99.1%. 52.1% were girls and half of them (50.3%) were aged between 1 and 3\u00a0years. The mean age was found to be 34\u00a0months, with a range between 7\u00a0months and 5\u00a0years.Over three-quarters of respondents (75.6%) had sought treatment from a nurse, health extension worker, doctor or pharmacist, mostly 50.1%) from a nurse (Table\u00a0% from a Allium sativum (Liliaceae) was the most frequently reported plant . Although this seemed to be higher than the reported cure rate for all other medicinal plants (53%), the difference was not statistically significant (Fisher\u2019s exact test statistic\u2009=\u20090.284).According to their parents, all children who were treated with ACT were cured while 84.6% and 15.4% of children treated with CQ were cured and improved, respectively (p\u2009<\u20090.001). Of the herbal treatments, garlic was the most commonly used, but Most patients used leaves (41.9%) followed by roots (17.1%). The oral route was the most frequent form of administration (92.8%) whereas decoctions were the most common preparation. Respondents were also asked about the unit of measurement for medicinal preparations and the majority of them used teaspoons and coffee cups for liquid preparations such as decoctions. Water was the most widely used solvent and honey was commonly used as sweetening agent to mask unpleasant tastes.P. vivax is estimated to cause less than one-third of malaria cases in Ethiopia [It was found that much higher rates of treatment-seeking from formal health workers than reported in other sources. In spite of this, only a minority of children with malaria were treated with ACT, confirming results of other studies . ChloroqEthiopia . This maP. vivax in Ethiopia [P. vivax malaria in Ethiopia [Although there are a few reports of CQ resistant Ethiopia \u201322, CQ iEthiopia  and is sEthiopia . The lowThe use of herbal medicine was lower than expected. This may partly be due to the expansion of the Health Extension Programme (HEP) at the household level which increased treatment-seeking for malaria in some areas  althoughNone of the plants were associated with a very high reported \u201ccure\u201d rate, unlike studies in some other African countries . HoweverAllium sativum, was found to prevent the development of parasitaemia in mice infected with Plasmodium berghei and substantially improved the anti-malarial activity of chloroquine [J. schimperiana. Although not the most commonly used, it was associated with the highest reported cure rates in this study. Its root extract has been tested against P. falciparum in vitro and was not very active (IC50\u2009=\u200971 mcg/ml for the methanolic extract) [P. berghei in mice in the 4-day suppressive test by 41%, and the methanol leaf extract suppressed parasitaemia by 65% [Clerodendrum myricoides and Zehneria scabra also had good anti-malarial activity in mouse models, they were not widely used, possibly because they are widely regarded as poisonous [The anti-malarial activity of several of these plant extracts has been assessed in rodent models in vivo Table\u00a0. Ajoene,oroquine . Howeverextract) . Howevera by 65% . Althougoisonous .Table\u00a04PThis was a community-based study conducted in one of the most malarious areas of Ethiopia. There was a very high response rate so the results are likely representative of the population in this area, and it could be confident in the prevalence of use of the different treatments. Voucher specimens of plants were collected and identified by a botanist.P. falciparum or P. vivax. As the survey was retrospective, it is likely that some patients were not tested, and some may not actually have had malaria. Secondly, parents may not have been aware of the qualifications of the person they saw and may have said she was a \u2018nurse\u2019 when in fact she may have not been qualified. Thirdly, the data on outcomes is based on self-reporting by parents of the children. It seems that the question which discriminated best between treatments was whether a patient was \u2018cured\u2019, since almost all patients claimed to have at least \u201cimproved\u201d. Therefore, the % of patients \u2018cured\u2019 on each treatment was compared. This approach seems to be valid, because 100% of patients claim to have been \u2018cured\u2019 after taking an ACT (to which there is no documented resistance in Ethiopia) compared to 84.6% who took chloroquine, against which there is some resistance.The first limitation of this study is that patients were not asked whether they had been tested before receiving the treatment. With hindsight, it would have been useful to know the proportion of patients tested, and the proportion with J. schimperiana versus those who had taken Allium sativum.Lastly, because herbal medicine was less frequently used than predicted, the sample size was too small to be able to find statistically significant differences in outcomes between patients who had taken different herbal remedies. Sample size calculation was based on an estimation of 50% of patients having taken herbal medicines. If that had been the case, this sample would have included twice as many patients who had taken herbals, which would have increased the statistical power to find differences in outcomes between subgroups\u2014for example those who had taken P. falciparum rather than P. vivax. Since most of the anti-malarial medicines were provided by health workers, a qualitative study of health workers would help to understand this. Factors to explore would include availability of AL and use of diagnostic tests or microscopy to distinguish between malaria species.Further investigations are needed to understand why the majority of children with malaria in Ethiopia receive CQ rather than AL, although the majority of malaria cases are reported to be caused by Justicia schimperiana may have the potential to be a candidate for the development of efficacious and safe anti-malarial phytomedicines and/or compounds, using a \u2018reverse pharmacology\u2019 model [y\u2019 model . It woulJ. schimperiana was associated with the highest proportion of patients who said they were \u2018cured\u2019 (69%). Further research is warranted to understand reasons for the low use of AL, and to investigate the anti-malarial properties of J. schimperiana.In the most malarious villages of Dembia district, Ethiopia, only 30% of children with presumed malaria took the recommended first-line treatment (artemether\u2013lumefantrine), while 47% took chloroquine and 22% were treated with herbal medicines as the first-line treatment. The most commonly used herbal medicine was garlic, but"}
+{"text": "Malaria and schistosomiasis remain life-threatening public health problems in sub-Saharan Africa. The infection pattern related to age indicates that preschool and school-age children are at the highest risk of malaria and schistosomiasis. Both parasitic infections, separately or combined, may have negative impacts on the haemoglobin concentration levels. The existing data revealed that artemisinin derivatives commonly used to cure malaria present also in antischistosomal activities. The current study investigated the impact of Artesunate-Amodiaquine (AS-AQ) on schistosomiasis when administered to treat malaria in rural area of Lemfu, DRC. Schistosoma mansoni, and Plasmodium falciparum infections. The egg reduction rate and haemoglobin concentration were assessed four weeks after the treatment with AS-AQ, of all coinfected children of this series. A prospective longitudinal study including 171 coinfected children screened for anaemia, mansoni eggs. Out of 43 (25.6%) children who remained positives, 37 (22%) showed a partial reduction of eggs amount, and no reduction was noted in 3.6% of coinfected. The mean of haemoglobin concentration and the prevalence of anaemia were, respectively, 10.74\u00b11.5g/dl , 11.2\u00b11.3g/dl, and 64.8%, 51.8%, respectively, before and after treatment, p<0.001. One hundred and twenty-five (74.4%) out of 168 coinfected children treated and present during the assessment were found stool negative for S. S. mansoni in coinfected children and increasing the Hb level. For the future, the randomized and multicentric clinical trials are needed for a better understanding of the effectiveness of AS-AQ against Schistosoma spp. The trial registration number was 3487183. The AS-AQ commonly used against Plasmodium allowed curing Schistosomiasis is a parasitic disease world widely distributed, of which the most of the disease burden occurs in Africa . The mas S. mansoni or S. haematobium [ P. falciparum drug resistance, DRC has adopted the use of the Artemisin-based combination therapy with AS-AQ as the first-line treatment drug for uncomplicated malaria, while Artemether-Lumefantrine is being used too [ S. mansoni in rural area of Lemfu.The Democratic Republic of the Congo (DRC) is identified as schistosomiasis endemic country, where this parasite is present in almost all the provinces for over a century . In Kongmatobium \u20137. On thmatobium . The plaused too . Nowadayused too . Thus diused too \u201318. Despused too , 20, conused too . Neverth S. mansoni endemic area. Lemfu is located in Kisantu Health Zone, in Kongo central province, western part of DRC, at 150 km from Kinshasa, the capital city of DRC.The study was conducted among children aged from 3 to 15 years, living at Lemfu, a Plasmodium and S. mansoni infection, respectively, by microscopic examination of Giemsa-stained blood smears, and Kato-Katz technique for stool samples analysis during the study aimed to investigate the prevalence and comorbidity of both parasites. To determine the impact of Artesunate-Amodiaquine on S. mansoni infection, the coinfected children were treated according to the national recommended guidelines for the treatment of uncomplicated malaria [A prospective longitudinal, noncomparative, and nonrandomized study was conducted among the coinfected children, with a followup period of one month from July to August 2015. The included children have been screened previously for malaria . In addiExclusion criteria included a history of malaria infection according to WHO criteria, adverse reaction to artemisin derivatives, women with known or suspected pregnancy, and children who were absent during the study followup.Exclusion criteria included a history of blood transfusion, a current crisis or history of sickle cell crisis, or acute illness two months prior to the study. Pregnancy status was determined by details from the patient's history and/or by a positive pregnancy test. We excluded from the study all children who were absent during the results assessment. S. mansoni eggs from each stool sample were counted during microscopic examination and the number obtained was multiplied by the factor 24 in order to get the number of eggs per gram of feces (epg). Finally, from each individual we considered the average of egg counted for three stool samples. The infection intensities were defined following the guidelines established by WHO into light, moderate, or heavy infections, respectively, as follows: 1\u201399 epg, 100\u2013399 epg, and >400 epg [Stool samples were examined for the presence of Schistosoma eggs using the Kato-Katz method four weeks after the treatment with AS-AQ. To improve the sensitivity of the Kato-Katz technique, three stool samples were collected during three consecutive days from each participant after the treatment with AS-AQ has been administered. The>400 epg .\u00ae Hb 201 . Anaemia was defined by a haemoglobin concentration according to the age and sex of child as follows: <11, 11.5,12, and 13g/dl, respectively, for children of less than 5 years, 5\u201311.9 years, and 12\u201314.9 children, unpregnant women > 15 years, and men > 15 years. The anaemia was classified as severe (Hb=7 g/dl), moderate (Hb= 7\u20139.9 g/dl), and mild anaemia (Hb=10\u2013 11.4 g/dl) [Haemoglobin (Hb) levels were determined using a Hemocue.4 g/dl) .The coinfected children received a fixed oral dose of AQ-AS ( 4 mg/kg/ AS 10 mg/kg/ AQ) daily for three consecutive days according to the national treatment guidelines . The druThe medicines administration was performed by a nurse of the study team, under direct observation of a physician. The treated children were kept at study site for 30 minutes prior to be released. In the case of vomiting, a dose was repeated during the observation period.Signed informed consent for participation was obtained in French or in the local language of all participants' guardians or parents. Ethical clearance for the study was obtained from the Ethics Committee of the Kinshasa School of Public Health . mansoni egg-positive to negative after treatment and noncure or those whose status remained S. mansoni egg-positive after treatment. The rate of cure after treatment was calculated as the percentage of individuals who tested negative for S. mansoni eggs after treatment. Secondary, the gained Hb concentration level was assessed by comparing the mean and the prevalence of anaemia before and after treatment.Data were entered into Microsoft Office Excel 2007 spreadsheets and analyzed using Epi Info version 7 and SPSS software version 21 . The frequencies and percentages were calculated for categorical variables, paired t-test, and Wilcoxon test were used for statistical analysis of continuous data, according to the variable distribution. The a priori statistical significance level was 0.05. The primary efficacy endpoint was the number of participants cured 28 days after treatment. Treatment outcome was expressed as cure for participants whose status changed from S. Plasmodium and S. mansoni. Three were excluded from the study because of their absence at the assessment moment; then 168 were included in the current study (\u03bcl. The number of epg ranged from 24 to 8760 with a median of 192 (IQR: 60\u2013612). The S. mansoni infection was, respectively, light (37.6 %), moderate (28.9 %), and severe (33.5%). The overall prevalence of anaemia was 64.6%; its intensity was, respectively, light (56.2%), moderate (42.1%), and severe (1.7%). The mean level of Hb in the study population was 10.8\u00b11.4 g/dl (range: 3.8- 15.5).Among the 364 children screened for both parasites, 171 were coinfected withnt study . The age S. mansoni infection and haemoglobin concentration level was performed four weeks after the treatment (Assessment ofreatment . S. mansoni infection remained positive in 43 participants (25.6%), of which 37  children had partial reduction of eggs load; and the eggs load tended to increase in 6 children (3.6%) (95% CI: 1.32-7.6). An absence of eggs was observed in 125 children (74.4%)  . The med (74.4%) . A consiThe prevalence of anaemia passed from 64.8% to 51.8% (p<0.001) and the mean of Hb from 10.74\u00b1 1.5g/dl to 11.26\u00b11.3g/dl (p<0.001) after treatment .Plasmodium spp. and Schistosoma spp. coexist in many regions, and the prevalence of their coinfection varies according to the species of Schistosoma and the geographical location [location \u201327; ther Schistosoma was damaged [ S. haematobium [ S. hematobium egg reduction rates were 95.6% with PZQ compared to 92.8% with AS-SMP (p = 0.096)[Despite some cases of treatment failures with ACT against malaria reported in different provinces across DRC \u201329, data damaged  and this damaged \u201313. Indematobium , while Kmatobium . In Mali = 0.096). S. mansoni infection in group treated with PZQ compared to 14% found in group treated with AS-SMP (p<0\u00b70001). Later, after changing dose regimen, these authors found no difference in response between the two drugs, and 74% were cured in the PZQ group and 64% in the AS-SMP group (p = 0.4) [In earlier study from Kenya, Obonyo et al. reported 65% of cure rate ofp = 0.4) . S. mansoni infection [Moreover from studies carried out in Sudan and Ethiopia, the authors found 100% of cure rate after using Artemether-Lumefantrine to curenfection , 17.The difference in cure rate from those studies could be explained by the different treatment regimens administered, differences in the susceptibility of the parasite to the used medicine, rates of reinfection, and different severity of the infection according to the different geographic locations. In general, it has been reported that artemisinin acts better enough in light infections , 17, whe Schistosoma infections largely overlap geographically, the wide use of antimalarials might be beneficial in coinfected patients [ Schistosoma in Lemfu as well as in DRC and likely a new alternative therapeutic against schistosomiasis.Although this study has provided the useful data, budget constraints related to lack of adequate funding did not permit assessing malaria parasitaemia after the treatment of included children. However, other benefits of the treatment with AS-AQ were found as the clinical status of study participants improved after the treatment, as showed by the significant increase of the Hb level, therefore a significant decrease of anaemia in the study population. The use of antimalarials in deworming programs should be eyed with caution, as this could accelerate resistance to these drugs. Nevertheless, since malaria andpatients . In the  S. mansoni infection. This is of a capital importance in malaria and schistosomiasis endemic settlements. It is advantageous for individuals harboring Schistosoma parasites, as every time they will be treated with an ACT for malaria, they will be indirectly treated against schistosomiasis and its morbidity. Nevertheless, the heavy burden of the morbidity due to schistosomiasis is implying an integrated approach on pharmacological approaches, population-based chemotherapy, treatment of individual cases; the control of snails, intermediate hosts, with molluscicides, and improving the health education, behavioral changes, sanitation, and safe water supply.The present study has shown promising results about AS-AQ in the treatment of"}
+{"text": "Post-malaria neurological syndrome (PMNS) is a debated entity, defined by neurological complications following a post-malaria symptom-free period and a negative blood smear. Four cases of PMNS are hereby reported and a review the literature performed to clarify the nosological framework of this syndrome.A French teaching hospital infectious diseases database was investigated for all PMNS cases occurring between 1999 and 2016 and the PubMed database for cases reported by other institutions after 1997. A case was defined by the de novo appearance of neurological signs following a post-malaria symptom-free period, a negative blood smear, and no bacterial or viral differential diagnoses.Plasmodium vivax infection were found matching the definition. In the institution, the estimated PMNS incidence rate was 1.7 per 1000 malaria cases overall. Of the 52 patients (mean age 33\u00a0years), 65% were men. Malaria was severe in 85% of cases, showed neurological involvement in 53%, and treated with quinine in 60%, mefloquine in 46%, artemisinin derivatives in 41%, antifolic drugs in 30%, doxycycline in 8% and other types in 8%. The mean symptom-free period was 15\u00a0days. PMNS signs were confusion (72%), fever (46%), seizures (35%), cerebellar impairment (28%), psychosis (26%), and motor disorders (13%). Cerebrospinal fluid analyses showed high protein levels in 77% (mean 1.88\u00a0g/L) and lymphocytic meningitis in 59.5% (mean 48 WBC/mm3) of cases. Electroencephalograms were pathological in 93% (14/15) of cases, and brain MRIs showed abnormalities in 43% (9/21) of cases with white matter involvement in 100%. Fourteen patients were treated with steroids. The 18 patients with follow-up data showed no sequelae. The mean time to recovery was 17.4\u00a0days.Four patients from the database and 48 from PubMed, including 4 following Plasmodium falciparum and Plasmodium vivax should be added to the list of pathogens causing ADEM.PMNS is a rare entity englobing neurological signs after severe or non-severe malaria. It appears after a symptom-free period. PMNS occurred following treatment of malaria with a wide range of anti-malarials. The disease is self-limiting and associated with good outcome. MRI patterns underline a possible link with acute disseminated encephalomyelitis (ADEM) or auto-immune encephalitis.  Additionally, a return of neurological signs after cure has been reported since 1987 [Falciparum malaria remains a common cause of morbidity and mortality, with an estimated 212 million cases and 429,000 deaths in 2015 [A work by Senanayake et al.  publishePlasmodium falciparum and negative etiological investigations. In their study, the use of mefloquine for severe malaria was associated with PMNS (relative risk 7.4). Since then, a number of case reports and series have been published but a clear definition and pathophysiological hypotheses for this syndrome are still lacking. PMNS may be a part of acute disseminated encephalomyelitis (ADEM) or acute post-infectious encephalitis but this too remains controversial. Finally, the medical community knows little about the condition\u2019s underlying pathophysiology, the duration of its symptom-free period, or its outcome and prognosis, and furthermore has little in the way of diagnostic tools or treatment options. Four new cases of PMNS are herein reported and the characteristics of the cases reported since 1997 discussed further with the goal of contributing to a better understanding of this rare entity.In 1997, Nguyen et al.  providedPlasmodium spp. The disease of patients with imported malaria in France were classified as severe or non-severe using the French 2007 classification [Malaria was defined as the association of compatible clinical signs with a positive blood smear and/or antigens for a fication .Plasmodium species, i.e., P. falciparum Plasmodium vivax, Plasmodium ovale, Plasmodium malariae or Plasmodium knowlesi), associated with a negative blood smear and no retainable differential diagnosis. The symptom-free period was a crucial criterion.PMNS was defined as the occurrence of de novo neurological signs after a symptom-free period following acute malaria , the following data was assessed: age, gender, malaria severity, malaria treatment, symptom-free period duration, fever presence, neurological signs , C-reactive protein (CRP), cerebral spinal fluid (CSF) characteristics, electroencephalography (EEG) signs, brain magnetic resonance imaging (MRI) results, treatment and outcome when available.P. falciparum group.Rates and means were calculated on the available data. Continuous variables were expressed as medians with minimum\u2013maximum values (min\u2013max) and means with standard deviations (SD). Qualitative variables were expressed as percentages. 95% confidence intervals (95% CI) for sample proportions were calculated using Wilson\u2019s method. Statistical analyses were performed using STATA1 software, version 12 for Windows. Analysis was restricted to the Four of 2314 patients treated in the hospital for imported malaria during the study period fit the case definition of PMNS. Therefore, the estimated PMNS incidence rate for the hospital was 1.7 per 1000 malaria cases overall (95% CI 0.7\u20134.0 per 1000).P. falciparum with 5 parasites/2\u00a0\u03bcL, positive HRP2 antigen test) with hepatic impairment  but no neurologic involvement or any other severity criteria. The treatment regimen included intravenous artesunate  then atovaquone/proguanil (1000/400\u00a0mg per day for 3\u00a0days), and the patient improved quickly, both clinically and biologically . On day 7, he presented headaches and fever (38\u00a0\u00b0C) and on day 8 abdominal pain, nausea and vomiting. The renewed blood smear was negative. On day 10, the patient showed confusion, ataxia, tremor, and dysarthria, and his fever increased to 39\u00a0\u00b0C. On day 11, he was given ceftriaxone for presumed enteric fever. On day 12, he remained confused and started having visual hallucinations and urine incontinence. CSF analysis showed lymphocytic meningitis , anti-neutrophil cytoplasmic antibodies (ANCA), complement, anti-phospholipid (APL) antibodies, anti-neuronal antibodies including NMDA-receptor in CSF, rheumatoid factor, anti-CCP, angiotensin conversion enzyme and 16S RNA on CSF). The patient improved around day 54 without specific treatment. MRI on day 63 had returned to normal. On day 73, the patient was free of neurological symptoms.Case 2 was a 29-year-old Caucasian male who worked in Ivory Coast. He presented a first episode of acute falciparum malaria without severity criteria in July 2013. That episode was treated with a 3-day course of atovaquone/proguanil (standard treatment). The patient experienced a second symptomatic episode with a positive blood smear (0.18%) 3\u00a0weeks later and received a 3-day course of artemether/lumefantrine with good outcome. He then consulted on day 37 for vomiting and a 40\u00a0\u00b0C fever for which intravenous quinine was initiated despite a negative blood smear. On day 38, he presented convulsions and a severe alteration of consciousness requiring sedation and ventilation. He was evacuated to Paris where his anti-malarial treatment was changed to artesunate despite a still negative blood smear. On day 42, sedation was stopped but the patient presented visual hallucinations and generalized convulsions and consequently had to be re-sedated. The blood test was again negative for malaria but an HRP2 antigen test was positive. T2 and T1 gadolinium-enhanced MRI sequences on day 48 showed hippocampal lesions  falciparum malaria on 28 November and treated with a 3-day course of atovaquone/proguanil with good outcome. Thirteen days later (day 15) she became confused, aphasic and anosognosic. CSF analysis showed lymphocytic meningitis  in July 1999 after a trip to Ivory Coast. She was initially treated with intravenous quinine (25\u00a0mg/kg/day) and on day 5 was given an oral dose of quinine (500\u00a0mg tid) after which she developed hypoacusis. On day 6, she was treated with mefloquine (6\u2009\u00d7\u2009250\u00a0mg for 24\u00a0h). She was discharged after recovery but on day 18 she experienced dizziness, limb weakness, gait impairment, nausea, an increase in pre-existing headaches and an episode where she was unable to read. Tremor and ataxia were observed during a resulting physical examination. On day 21, the level of mefloquine in her blood was high . On day 27, her clinical status improved spontaneously and she was discharged without any specific treatment. Follow-up blood examinations showed that the half-life of mefloquine elimination for this patient was 9\u00a0days. On day 41, the patient had a normal clinical status and no P. falciparum alone, 4 P. vivax alone, and 2 P. falciparum and P. vivax together. Of these 48 patients, 16 were travellers  who acquired malaria while abroad and 32 were inhabitants of countries endemic for the infection . No cases were described with P. ovale, P. malariae or P. knowlesi.The systematic review of the literature found 48 PMNS patient cases of which 42 involved P. vivax infection alone [P. vivax infection [P. falciparum group, of the 42 CSF analyses available, 25 (60%) showed lymphocytic meningitis (mean of 48\u00a0cells/mm3), and 33 (78%) high protein levels (mean of 1.4\u00a0g/L) without low glucose levels. When abnormal, EEG showed diffuse or frontal slowing. Computed tomography was never abnormal when performed (9/9). MRI revealed brain abnormalities in 9 of 21 P. falciparum patients (43%), showing hypersignals, sometimes with gadolinium enhancement, in different localizations , usually with little extension. One child showed clinical and MRI characteristics of severe ADEM [P. falciparum and P. vivax infection, respectively, showed grey matter hypersignals.The main clinical findings are summarized in Table\u00a0on alone \u201326 and 2on alone , 28. Thrnfection , 24, 26.nfection . Table\u00a02ere ADEM . One casP. falciparum group, corticosteroids were used to treat 10 patients, among whom 6 had MRI abnormalities. Six of these 10 patients received intravenous high-dose methylprednisolone . Of the 4 patients in the P. vivax group, 3 received corticosteroids (missing data for one) and one in the mixed infection group. None of the 22 patients with follow-up data had sequelae. The mean time to recovery in the 21 patients with that datum was 17.4\u00a0days (12.3), . The characteristics of PMNS are summarized in Table\u00a0In the For the present work, the literature\u2019s 48 cases of PMNS published since 1997 were assessed and 4 local imported cases added thereto. From the database of the hospital, an incidence of 1.7 per 1000 was determined, comparable with that of 1.2 per 1000 reported in a previous study [P. vivax PMNS, appeared to show the same types of lesions  after severe falciparum malaria (85%), with cerebral malaria in 50% of the cases. It was seen most frequently in people who live in (two-thirds of reported cases), but also in those who travel to, malaria-endemic areas. PMNS was also seen after vivax malaria, notably in children in 50% of the cases , 24, 26.vailable , 24, 26)P. falciparum patients reported had received a range of treatments (including artemisinin combination therapy). Moreover, PMNS occurred after P. vivax infection for which mefloquine had not used. Considered together, these observations cast doubt on an association or causative effect for mefloquine and PMNS.A study from Vietnam had concluded that mefloquine was a risk factor for PMNS after severe malaria  [Cerebellar signs can be seen not only in severe malaria, where they respond to anti-malarial treatment \u201331, but sequelae [P. falciparum and P. vivax have not been identified as causative agents for ADEM, the great number of characteristics shared between this latter and PMNS suggest that the relationship is nonetheless highly plausible.PMNS shares many features with CNS post-infectious diseases such as ADEM and auto-immune encephalitis. ADEM \u201337 is a sequelae . Multiplsequelae , 40. AltN-methyl-d-aspartate-receptor antibody-NMDAR or P/Q type voltage-gated-calcium-channel antibody. The NMDAR antibody was described in 12 (24.5%) of 49 patients 3\u00a0months after HSV encephalitis [sequelae are largely absent.In 2007, a new group of neurological disorders mediated by neuronal antibodies (against ions channels and synapses) appeared in the literature under the term auto-immune encephalitis (AIE) . AIE cauphalitis . No neurphalitis . In AIE The clinical features of PMNS are compatible with post-infectious encephalitis, either ADEM or AIE. Symptom-free latency and negative extensive screening of possible infectious or systemic causes argue for a post-infectious immunologically mediated cerebral aggression. CSF analyses, inflammation characteristics and EEG are all compatible with post-infectious encephalitis Table\u00a0. For ADEP. falciparum clearance and even in non-neurological malaria [P. vivax infection could explain why most PMNS cases follow P. falciparum infections. The first case presented non-specific anti-nuclear factors and the second had neither anti-nuclear factors nor anti-neuronal antibodies. Recently, Sahuguet et al. [The mechanisms underlying PMNS are poorly understood. The delayed onset, negativity of blood smear, association with fever, and sometimes elevated CRP may suggest an inflammatory pathway, probably involving cross presentation like in AIE, but no demonstration of that has been made to date. The authors of the only study on inflammatory pathways of malarial post infectious neurological complications reported increased blood and CSF pro-inflammatory cytokines, like TNF, IL-6 and IL-2, in 12 patients with DCA (compared to 8 patients without it) . PMNS ha malaria . The fac malaria , 49 in Pt et al.  reportedP. vivax patients) and most without steroid treatment. However, steroids were given to the 14 most-severe cases . This observation casts a shadow on the \u2018self-limiting course\u2019 [sequelae or shorten time to recovery. An interventional study would be needed to respond to these questions. In analogy with AIE treatment, corticosteroids do nonetheless appear to be a first-line treatment for PMNS, at least in severe cases and at least for now. The prognosis of PMNS seems to be good and corticosteroid treatment should be discussed for the most severe cases. If corticosteroids are not effective, first the diagnosis of PMNS should be reconsidered, and thereafter intravenous immunoglobulin or plasma exchange discussed.The use of steroids in PMNS is another topic worth discussing. According to the follow-up data available, all patients fully recovered in a mean delay of 17.2\u00a0days .PMNS is a rare entity englobing a range of neurological disorders following malaria cure. There appears to be no direct role for the parasite but rather a putative immune-mediated aggression of the CNS. Most of its characteristics can fit into the diagnosis of post-infectious encephalitis, ADEM or AIE. The addition of malaria due to"}
+{"text": "Plasmodium vivax and Plasmodium ovale malaria only. We quantified the risk of P vivax parasitaemia after treatment of Plasmodium falciparum with commonly used antimalarial drugs to assess the potential benefits of radical cure for all patients with uncomplicated malaria in co-endemic regions.A 14-day course of primaquine is used for radical cure of P falciparum malaria in countries co-endemic for P vivax. Studies were included if the presence or absence of P vivax parasitaemia was recorded after treatment. The primary outcome was the risk of P vivax parasitaemia between day 7 and day 42 after initiation of antimalarial treatment for P falciparum, with the pooled risk calculated by random-effects meta-analysis. We compared the risk of P vivax parasitaemia after treatment with different artemisinin-based combination therapies (ACTs). This study is registered with PROSPERO, number CRD42017064838.In this systematic review and meta-analysis, we searched MEDLINE, Embase, Web of Science, and the Cochrane Database of Systematic Reviews for prospective clinical studies in any language, published between Jan 1, 1960, and Jan 5, 2018, assessing drug efficacy in patients with uncomplicated P vivax parasitaemia by day 42 was 5\u00b76% . The risk of P vivax parasitaemia was 6\u00b75% (95% CI 4\u00b76\u20138\u00b76) in regions of short relapse periodicity compared with 1\u00b79% (0\u00b74\u20134\u00b70) in regions of long periodicity, and was greater after treatment with a more rapidly eliminated ACT: 15\u00b73% (5\u00b71\u201329\u00b73) for artemether-lumefantrine compared with 4\u00b75% (1\u00b72\u20139\u00b73) for dihydroartemisinin-piperaquine and 5\u00b72% (2\u00b79\u20137\u00b79) for artesunate-mefloquine. Recurrent parasitaemia was delayed in patients treated with ACTs containing mefloquine or piperaquine compared with artemether-lumefantrine, but by day 63 the risk of vivax parasitaemia was more than 15% for all ACTs assessed.153 of 891 screened studies were included in the analysis, including 31\u2008262 patients from 323 site-specific treatment groups: 130 (85%) studies were from the Asia-Pacific region, 16 (10%) from the Americas, and seven (5%) from Africa. The risk of Our findings show a high risk of vivax parasitaemia after treatment of falciparum malaria, particularly in areas with short relapse periodicity and after rapidly eliminated treatment. In co-endemic regions, universal radical cure for all patients with uncomplicated malaria has the potential to substantially reduce recurrent malaria.Australian National Health and Medical Research Council, Royal Australasian College of Physicians, Wellcome Trust, and Bill & Melinda Gates Foundation. Plasmodium falciparum,P falciparum and Plasmodium vivax. In Thailand, the risk of P vivax parasitaemia after P falciparum infection is substantially higher than would be expected from background entomological inoculation rates and almost 1\u00b75 times greater than the risk of recurrent P falciparum.P vivax.P vivax parasitaemia is associated with substantial morbidity including a cumulative risk of severe anaemia.P vivax sexual stages (gametocytes) are commonly present with recurrent asexual parasitaemia, there is an increased risk of transmission of P vivax to the mosquito vector.6In 2016, more than 200 million cases of malaria were attributable to P falciparum and P vivax.P vivax hypnozoites. At the time of writing, primaquine is the only widely available hypnozoitocidal treatment, which is required to achieve the radical cure of vivax malaria.In some locations, declining chloroquine efficacy has led national malaria control programmes to adopt a universal policy of artemisinin-based combination therapy (ACT) for blood schizontocidal treatment of both Evidence before this studyPlasmodium vivax parasitaemia after Plasmodium falciparum infection. A pooled analysis of individual patient data from Thailand found that there was a 32% cumulative risk of P vivax parasitaemia by day 63 after treatment for P falciparum, greater than would be expected on the basis of local background entomological inoculation rates. A review of 19 studies in which patients with P falciparum were treated with artemether-lumefantrine identified a high risk of subsequent P vivax parasitaemia. To our knowledge, no systematic reviews and pooled analyses have assessed the risk of P vivax parasitaemia after treatment of P falciparum infection outside of Thailand.We searched MEDLINE, Web of Science, Embase, and the Cochrane Database of Systematic Reviews with the terms \u201cfalciparum\u201d, \u201cvivax\u201d, and \u201crecurrence\u201d, to identify all articles in any language published between Jan 1, 1960, and Jan 5, 2018, assessing the risk of Added value of this studyP vivax parasitaemia after treatment of P falciparum malaria. Our meta-analysis provides a comprehensive comparison of the risk of P vivax parasitaemia after treatment with different artemisinin-based combination therapies (ACTs). Our findings show a high risk of P vivax parasitaemia after P falciparum infection in co-endemic regions. The risk of P vivax parasitaemia is greatest after rapidly eliminated antimalarial regimens and in regions where P vivax has a short relapse periodicity. After day 42, the risk of P vivax is high after treatment with all of the major ACTs.In this systematic review, we identified 31\u2008262 patients from 153 studies across 21 countries to quantify the global risk of Implications of all the available evidenceP vivax parasitaemia and enhance malaria elimination efforts.In co-endemic regions, universal radical cure, such as an ACT plus primaquine or tafenoquine, in all patients with uncomplicated malaria has the potential to prevent subsequent P vivax parasitaemia after treatment of P falciparum with commonly used antimalarial treatments in a range of co-endemic regions.To determine the potential benefits of a universal policy of a 14-day regimen of primaquine radical cure, we quantified the risk of P falciparum malaria residing in areas co-endemic for P falciparum and P vivax. Co-endemic countries were defined as countries where indigenous P falciparum and P vivax cases were reported or suspected in 2016  and data extracted (RJC and MSH) by two authors, with discrepancies resolved following discussion with a third author (RNP). Findings are reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines . When data for P vivax relapse periodicity according to their geographical location and relapse criteria defined by the Malaria Atlas Project.P falciparum gametocytes present at enrolment, or if this was not recorded, within the first 3 days of treatment. When only the haematocrit was available, it was converted to haemoglobin using the formula: haemoglobin=(haematocrit\u20135\u00b762)/2\u00b76.Study sites were categorised into long or short P vivax parasitaemia before day 42 following initiation of treatment for P falciparum infection, defined as any P vivax parasitaemia detectable by microscopy between day 7 and day 42. Evaluable patients included patients with subsequent P vivax parasitaemia, P falciparum parasitaemia, treatment failure before day 7 (defined as failing to clear parasitaemia before day 7), or patients followed until day 42 with adequate clinical and parasitological response. Risk is the percentage of patients with P vivax parasitaemia out of the evaluable patients. Secondary outcomes were the risk of P vivax malaria before day 28 or 63 and recurrent parasitaemia due to any species before day 28, 42, or 63. If not stated explicitly, P vivax and P falciparum parasitaemia were assumed to have occurred in separate patients.The primary outcome was risk of Potential bias relating to individual studies was assessed using a tool developed by the Joanna Briggs Institute .13P vivax parasitaemia, any Plasmodium parasitaemia, and proportion of recurrent parasitaemia related to P vivax at day 28, 42, and 63 were estimated using random-effects meta-analysis with proportions pooled using the Freeman-Tukey double arcsine transformation without adjustment to observed values and with exact methods to calculate confidence intervals.I2 statistic and assessed with random effects meta-regression to investigate the association between the study-specific risks and baseline characteristics . Pooled risks of recurrent P vivax parasitaemia after treatment with artemether-lumefantrine, dihydroartemisinin-piperaquine, and artesunate-mefloquine were compared, with random effects meta-regression used to investigate these risks further. Analyses were done with Stata version 15 and R version 3.4.0 using the metafor package.The pooled risk of recurrent The study protocol is registered with PROSPERO, number CRD42017064838.The funder of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication.P vivax transmission met the inclusion criteria, mostly because they did not document P vivax parasitaemia. 153 (33%) of 465 studies from areas of stable indigenous transmission were included in the analysis  potentially relevant studies were reviewed. None of 426 studies from areas of unstable analysis . Of the analysis . Data foThe risk of parasitaemia was available for 323 records from the 153 studies, including 31\u2008262 patients from 21 countries. Of these records, 277 (86%) were from individual sites with a single treatment group and 46 (14%) were from aggregated sites or treatment groups, or both. 32 records included treatment with artemether-lumefantrine, 24 with dihydroartemisinin-piperaquine, and 38 with artesunate-mefloquine .P falciparum gametocytes at baseline ranged from 0% to 52% (104 records), and the mean or median baseline haemoglobin ranged from 7\u00b78 g/dL to 14\u00b73 g/dL (179 records).There was substantial heterogeneity between study populations . InformaI2=95\u00b74%; 213 estimates) by day 28, 18\u00b74%  by day 42, and 35\u00b77%  by day 63 (P vivax were 37\u00b74% (95% CI 30\u00b74\u201344\u00b76) by day 28, 37\u00b71% (28\u00b72\u201346\u00b72) by day 42, and 68\u00b76% (60\u00b76\u201376\u00b71) by day 63. By day 42, the risk of any parasitaemia was 28\u00b73% (95% CI 16\u00b78\u201341\u00b73) after treatment with artemether-lumefantrine, 16\u00b78% (7\u00b74\u201328\u00b78) after dihydroartemisinin-piperaquine, and 10\u00b79% (7\u00b74\u201315\u00b70) after artesunate-mefloquine  with no evidence of publication bias relating to small study effects  compared with regions with long periodicity . The risk of P vivax was also greater after treatment with rapidly and intermediately eliminated antimalarial regimens  compared with slowly eliminated regimens ; P vivax parasitaemia by day 42 was associated with the proportion of patients with baseline mixed P falciparum and P vivax infection (p=0\u00b70170), a lower mean age (p=0\u00b70010), higher baseline parasitaemia (p=0\u00b70149), and lower baseline haemoglobin  of 117 records for which data on all included covariates were available  risk after treatment with an intermediately eliminated regimen, and a 2\u00b70% (1\u00b72\u20133\u00b70) risk in patients treated with a slowly eliminated regimen  by day 28 and 24\u00b70%  by day 63 (The risk of y day 63 . Althougy day 63 . The majy day 63 .P vivax parasitaemia at both day 28 (p=0\u00b70069) and day 63 (p=0\u00b70381) compared with regions with long relapse periodicity; however, in multivariable analyses these effects were no longer apparent (P vivax parasitaemia at day 28 was greater in patients treated with rapidly eliminated drugs than in those treated with intermediately or slowly eliminated drugs (p<0\u00b70001), but this was no longer significant at day 63  for artemether-lumefantrine compared with 4\u00b75%  for dihydroartemisinin-piperaquine, and 5\u00b72%  for artesunate-mefloquine  after artemether-lumefantrine, 16\u00b72%  after dihydroartemisinin-piperaquine, and 15\u00b77%  after artesunate-mefloquine  was compared in 25 studies using multivariable meta-regression analysis controlling for age, baseline parasitaemia, presence of mixed infection at baseline, and regional relapse periodicity. The risk of P vivax parasitaemia was higher after treatment with artemether-lumefantrine  than with dihydroartemisinin-piperaquine or artesunate-mefloquine , accounting for more than half of all recurrent parasitaemias.Our systematic review and meta-analysis of 31\u2008262 patients treated for falciparum malaria shows a high risk of subsequent P vivax after treatment for P falciparum interaction has been described previously in a pooled analysis of patients from Thailand with a 32% cumulative risk of P vivax parasitaemia 63 days after treatment.P vivax was 24% and accounted for almost 70% of all recurrent parasitaemias. Although these observations could reflect simultaneous infection of patients by a mosquito carrying both species, entomological data suggest that this is unlikely.P vivax suggest that the vivax recurrences were attributable to reactivation of P vivax hypnozoites.21The high risk of P vivax infection after radical cure of primary vivax infection with primaquine and chloroquine, a slowly eliminated schizontocidal drug, is 0\u00b74 infections per person per year in EthiopiaP vivax recurrence by day 63 would be estimated conservatively to be 3\u00b78% in Ethiopia and 2\u00b75% on the Thailand\u2013Myanmar border. The pooled risks in our study were far greater, suggesting that new infections alone could not have accounted for these parasitaemias. Assuming that the excessive risk of P vivax parasitaemia after P falciparum infection is from reactivation of hypnozoites, the number needed to treat with radical cure to prevent one vivax recurrence by day 63 after P falciparum would be between 4\u00b77 and 5\u00b70.Previous studies have shown that the risk of P vivax parasitaemia varied significantly with the elimination half-life of the longer-acting drug of the antimalarial combination used.P vivax usually has a short periodicity, the first relapse occurring 21 days or more after the initial infection.P vivax emerging from reactivation of dormant hypnozoites. In our analysis, artemether-lumefantrine was associated with a four times greater risk of P vivax parasitaemia by day 42 compared with dihydroartemisinin-piperaquine and artesunate-mefloquine, which have substantially longer elimination half-lives, providing greater post-treatment prophylaxis against vivax parasitaemia.P vivax efficacy studies in which artemether-lumefantrine is associated with high risks of recurrent P vivax after treatment,27In the previous study from Thailand, the risk of P vivax parasitaemia in our meta-analysis, the risk of P vivax by day 63 was greater than 15% after treatment with all of the ACTs assessed. WHO antimalarial guidelines recommend changing antimalarial treatment policy if the risk of P falciparum recrudescence exceeds a 10% threshold.P vivax parasitaemia was 15\u00b73% after treatment with artemether-lumefantrine and the risk of any parasitaemia was 10\u00b79% or greater after treatment with any of the major ACTs. Hence, although slowly eliminated ACTs provide an early benefit over more rapidly eliminated ACTs, this effect is transient. Therefore, consideration should be given to co-administration of primaquine or tafenoquine to ensure eradication of P vivax hypnozoites in all patients presenting with malaria in co-endemic regions. However, our findings cannot necessarily be extrapolated to returning travellers in whom the risk of co-infection might be substantially lower.Although artesunate-mefloquine and dihydroartemisinin-piperaquine delayed P vivax, has blood schizontocidal activity against P vivaxP falciparum.P falciparum and P vivax, and decrease ongoing transmission in patients presenting with P falciparum monoinfection. Primaquine can induce substantial haemolysis in patients with glucose-6-phosphate-dehydrogenase deficiency;P vivax parasitaemia and preventing the cumulative morbidity associated with multiple relapses.P vivax parasitaemia and frequency of relapse are greatest, although this will need to be confirmed by prospective clinical trials. Additional modelling of the risks and benefits of universal radical cure will be needed to define priority locations where such a policy should be advocated.Primaquine, the only widely available hypnozoitocidal agent for P vivax, hence patients without any P vivax parasitaemia might have been under-represented if the study did not report this finding, leading to an artificial increase in the derived risk of P vivax parasitaemia after P falciparum infection. However, this bias is likely to be relatively minor, given that our results from studies from Thailand (P falciparum and P vivax parasitaemia occurred in separate patients unless specified in the report. Although this might have led to an elevated risk of parasitological failure, the risk of more than one recurrence within the 42 days after treatment is very low. Our study is also limited by heterogeneity of the studies and changes in epidemiology over the duration of the review, hence our findings might not necessarily be generalisable to all co-endemic regions. Finally, confidence intervals for pooling risk estimates include both between-study and within-study variation, and should be interpreted cautiously. Future meta-analyses pooling individual patient data will avoid many of these limitations and provide greater understanding regarding the factors associated with P vivax parasitaemia after P falciparum infection.Our study has some limitations. Comparison of studies with variable follow-up and the use of aggregated results could potentially bias our results. Studies were only included if they explicitly documented the occurrence or not of Thailand  were simP vivax parasitaemia after treatment for P falciparum infection in co-endemic regions. The risk is apparent for the major ACTs used in most P falciparum endemic regions and greatest in southeast Asia where P vivax relapse periodicity is short. Our findings suggest that in some regions co-endemic for both P falciparum and P vivax, the introduction of a universal policy of radical cure for all patients with uncomplicated malaria has potential to prevent recurrent parasitaemia, reduce ongoing transmission, and enhance malaria elimination efforts.In summary, there is a high risk of"}
+{"text": "Plasmodium falciparum malaria treated with an artesunate-mefloquine (AS-MQ) regimen. Relationship between mg/kg dose and therapeutic outcomes will be assessed through a one-stage individual participant data (IPD) MA.Antimalarial posology based on weight bands leaves patients at the margins vulnerable to receiving either lower or higher weight-adjusted (mg/kg) dosages. This article aims to describe the protocol for systematic review and individual patient meta-analysis (MA) for a study of the distribution of artesunate and mefloquine dosage administered in patients with uncomplicated Therapeutic efficacy studies with the AS-MQ regimen will be identified by searching the following databases: PUBMED, EMBASE and Web of Science. The corresponding authors of the relevant studies will be requested to share the IPD for the purpose of this MA to a secured repository. All available studies will be standardised using a common data management protocol and pooled into a single database. The relationship between mg/kg dosage and treatment failures will be assessed using a Cox regression model with study sites considered as a shared frailty term. Safety parameters will be explored where available.This IPD MA met the criteria for waiver of ethical review as defined by the Oxford Tropical Research Ethics Committee as the research consisted of secondary analysis of existing anonymous data. The results of this analysis will be disseminated at conferences, WorldWide Antimalarial Resistance Network website and any peer-reviewed publication arising will be made open source.CRD42018103776. With the exception of recent studies in Southeast Asia, the regimen artesunate-mefloquine has consistently demonstrated an efficacy greater than 95% and treatment failures in any single antimalarial study are few. This individual participant data (IPD) meta-analysis (MA) will allow a robust exploration of host, parasite and drug factors associated with therapeutic outcomes, which otherwise would not be possible.The proposed IPD\u00a0MA will allow exploration of variations in the weight-adjusted dosage received by patients, which is not possible with aggregate data MA.The IPD\u00a0MA will be carried out as a study group under the auspices of the WorldWide Antimalarial Resistance Network, which has championed responsible data sharing and advocates translational research.A limitation of this analysis will be heterogeneity between studies included in terms of design, patient population and the susceptibility of the parasites against the drug regimen.The combination artesunate-mefloquine (AS-MQ) was the first antimalarial regimen developed as an artemisinin-based combination therapy (ACT) when mefloquine (MQ)\u00a0resistance became rampant along the Thai\u2013Myanmar border in the early 1990s.Until recently ACTs have consistently demonstrated an efficacy greater than 95% and treatment failures in any single antimalarial study are few, thus limiting the ability to draw inferences regarding putative factors associated with therapeutic outcomes. Individual participant data (IPD) meta-analysis (MA) is being used increasingly to explore some of the putative factors which otherwise would not be possible through aggregate data MA.The overall aim of this study is to determine the mg/kg dosing range of artesunate (AS) and MQ adopted in clinical trials and investigate the effects of mg/kg dosing on clinical outcome.To investigate the effects of MQ\u00a0and AS mg/kg dosing on early and late clinical outcomes (treatment success or failure).To investigate the tolerability of AS-MQ across different study sites, population and age groups.The specific objectives are:Plasmodium falciparum  in patients of all ages.Prospective clinical efficacy study  of uncomplicated Assessing the efficacy of a fixed-dose AS-MQ combination, either as single tablet type, or co-blister pack of more than one tablet type, or assessing the efficacy of a loose combination of AS-MQ.Where AS was given over 3\u2009days (with any number of doses per day) with a target total dose of 6\u201330\u2009mg/kg.Where MQ was given over 1\u20133 days, on any of days 1\u20133 (with any number of doses per day) with a target total dose of 15\u201333\u2009mg/kg.Where all AS and MQ were administered orally.With a minimum of 28 days follow-up.With genotyping performed for late parasite recurrence.With individual patient data on dosage of MQ received  by patients .Studies identified will be deemed eligible for the purpose of this analysis if they meet the following criteria:Where other antimalarial drugs were given in addition to the initial AS-MQ treatment regimen, except for a single dose of primaquine of 0.25\u2009mg/kg in the first 3 days.P. falciparum malaria will be included in this IPD\u00a0MA. The following patients will be excluded from the analysis:P. falciparum malaria.Severe Pregnant women.Patients with uncomplicated Fixed-dose combination of AS-MQ, either as single tablet type, or co-blister pack of more than one tablet type, or loose combination of AS-MQ. AS given over 3\u2009days (with any number of doses per day) with a target total dose of 6\u201330\u2009mg/kg. MQ given over 1\u20133 days, on any of days 1\u20133 (with any number of doses per day) with a target total dose of 15\u201333\u2009mg/kg.Parasitological and clinical efficacy.Adverse events.We will carry out a systematic review and search PubMed, EMBASE and Web of Science to identify publications with AS-MQ between January 1990 and\u00a0July 2018; the full search terms are available from the PROSPERO registration and also presented as 10.1136/bmjopen-2018-027738.supp1Supplementary dataPlasmodium berghei, Plasmodium chabaudi), publications only including severe malaria, studies with follow-up\u00a0<28 days, data previously included in another published study, prophylaxis or mass drug administration studies, studies in healthy volunteers/challenge studies, or studies in asymptomatic patients or pregnant women.Any important protocol amendments will be documented in the PROSPERO registration. Studies will be included regardless of language and publication status. Study screenings will be carried by two independent reviewers who will screen title, abstract, full text as necessary. The following studies will be excluded: animal models  studies identified from the literature search will be invited to share IPD. At least three emails will be sent out in case of non-response. Researchers agreeing to the terms and conditions of the submission will be requested to upload anonymised IPD to the WorldWide Antimalarial Resistance Network (WWARN) repository through a secure web portal. Data will be fully anonymised and handled in compliance with the UK Data Protection Act to protect personal information and patient privacy. Original data will be stored on a secure server hosted by the University of Oxford.Raw data from individual studies will be standardised using an open and transparent data management and statistical analysis protocol.All the researchers who share individual patient data from eligible studies will become part of the study group; will have an opportunity to contribute to the analysis, interpretation of the results, manuscript preparation; and will be listed as coauthors on the publication arising from these analyses according to the WWARN publication policy.Information is available on drug dosage, either as exact number of tablets received, exact mg/kg dose received or number of tablets planned per protocol.Date of the last day of follow-up or length of follow-up.The following patients will be included in the analysis:P. falciparum treatment failure.Received other antimalarial drugs during follow-up before recorded Results of genotyping performed for late parasitological outcome are not available.P. falciparum infection on enrolment.Missing confirmation of Missing age or weight or gender.Haemoglobin (Hb) <50\u2009g/L on day 0.Haematocrit (Ht) <15% on day 0.Other deviations, as defined in the data management planThe following patients will be excluded from the analysis:P. falciparum recrudescence.Primary: PCR-corrected P. falciparum reinfection.Secondary: PCR-corrected P. falciparum recurrence.PCR-uncorrected Early parasitological responses on days 1, 2 and 3.Gametocyte carriage within 14 days of treatment initiation in patients without gametocytaemia at enrolment.Anaemia status on day 7.Adverse symptoms developed after the drug administration.P. falciparum recrudescence (treatment failure) on day 42. Day 42 was selected as the primary endpoint based on the current recommendations from the WHO as outlined in the 2009 protocol, which suggests that day 42 be the minimum follow-up period for the MQ regimen.The primary endpoint for this IPD\u00a0MA is PCR genotyping corrected risk of Acute drug vomiting within an hour of treatment administration, general vomiting within 7 days of treatment initiation, diarrhoea within 7 days of treatment initiation and neuropsychiatric adverse events (where available) are secondary endpoints.AS and MQ doses received will be calculated from the number of tablets administered to each patient daily. If the actual number of tablets received is not recorded, the total dose in mg or mg/kg recorded as administered to each patient will be used. If none of these are available, administration as per protocol will be assumed. The current recommended daily MQ dose is 8.3 (range 5\u201311)\u2009mg/kg.Nutritional status in children under 5 years of age will be assessed using standardised age, weight, height and gender specific growth reference standards according to the WHO 2006 recommendations using igrowup Stata package.The falciparum malaria transmission intensity of the study sites will be assessed using the prevalence estimates generated by the Malaria Atlas Project based on the latitude, longitude and the year the study was conducted.Anaemia will be defined as Hb\u00a0<100\u2009g/L or Ht\u00a0<30%. Severe anaemia is defined as Hb\u00a0<70\u2009g/L or Ht\u00a0<20%. Fever will be defined as body temperature\u00a0>37.5\u00b0C.Parasite resistance status will be defined (data permitting) for each patient from Southeast Asia region based on the reported prevalence of mutations of molecular markers  or the distribution of parasite clearance half-life for their study site and year of admission.Summary of included studies will be presented with respect to study location, years of study, study population, duration of follow-up, AS-MQ drug formulation, methodology for parasite quantification, methodology for PCR genotyping and\u00a0supervision of drug administration. PCR-corrected/uncorrected outcomes will be used to compute the Kaplan-Meier (K-M) estimates using the censoring rules outlined earlier. The K-M estimates will be presented graphically together with the number of patients in the risk set.P. falciparum or mixed infections); total mg/kg dose for each drug component; dosing strategies ; dose formulation (fixed or loose) and supervision of drug administration. The number of available patients will be summarised for all variables, proportion will be used for categorical or binary variables and mean and standard deviation (or median and range) will be used for continuous variables.Summary of baseline characteristics of the patients included in the analysis will be presented for each study, by region and in overall. The following baseline characteristics of patients will be presented: age; weight; parasitaemia on enrolment; presence of fever (body temperature\u00a0>37.5\u00b0 C); Hb (or Ht); anaemia (Hb\u00a0<100\u2009g/L) or severe anaemia (Hb\u00a0<70\u2009g/L); gametocytes on presentation; description of infection , WAZ, underweight for age termed underweight (defined as WAZ < \u22122), Hb, gametocytes on presentation , history of malaria (if available); description of infection: mixed species infections , presence of markers of drug resistance, eg kelch13 mutations or pfmdr1 amplification (if available), details of treatment received: total mg/kg dose of AS and MQ, regimen, drug supervision and vomiting of medication. Year of enrolment will also be included to account for changes in parasite susceptibility over time.All possible risk factors will be examined in a univariable analysis. The log-likelihood estimates All the variables identified in step\u00a0(i) will be fitted together in one model and variables that\u00a0are not significant in the presence of other variables based on the results of the Wald test will be identified.A likelihood ratio test (LRT)\u00a0will be used to assess the impact of omitting variables identified in step (ii). If the omitted variable does not significantly impact the model log-likelihood, then they will be dropped. Only those variables which lead to significant change in log-likelihood are retained.All variables excluded from step (i) will be added to the model identified in step (iii) one by one to check if they provide any improvement to the model.A final check of the model identified in step (iv) will be carried out to ensure that none of the variables in the model can be omitted without significantly increasing the model log-likelihood, and none of the excluded variables significantly reduce the model log-likelihood.A general strategy recommended by Collet (2015)Comparison of likelihood between nested models will be conducted using LRT. Akaike\u2019s information criterion will be used to compare non-nested models. Treatment dosage, drug formulation and baseline parasitaemia will be included in the multivariable model as a priori forced variables regardless of their statistical significance. Variables with more than 50% observations missing will not be included in multivariable analysis. Interactions will be assessed between dosing and the following variables: region, age group, transmission intensity, hyperparasitaemia (parasitaemia\u00a0>100\u2009000 parasites/\u00b5l), date of enrollment.The multilevel logistic or Cox models will be used for explaining study-site heterogeneity. Heterogeneity across study sites will be assessed by the variance of the shared frailty term estimated in the random effect Cox model or variance of the random intercepts in logistic regression. In\u00a0addition, intraclass correlation in logistic regression model will be reported.Analyses will be conducted by geographical region, drug regimen and resistance status if data permit.A model will be refitted with each study\u2019s data excluded, one at a time, and a coefficient of variation around the parameter estimates will be calculated. This would identify any influential studies, that is, studies with unusual results  that affect the overall pooled analysis findings. To assess the impact of missing data , sensitivity analysis will be performed to see if our main conclusion is affected or not by the exclusion of patients with missing data. Multiple imputation (MI) will be used for handling missing data for missing covariates and missing outcomes. MI will be carried out for covariates with missing proportion less than 50%.Two reviewers will independently assess risk of bias. The risk of bias within and across the studies included in the analysis will be carried out using the GRADE guidelines.Despite best possible efforts, it is anticipated that raw data from all the identified studies will not be available. Risk of potential bias in these studies will be assessed using a two-stage IPD\u00a0MA for the reported efficacy outcomes.The main analysis is planned as described earlier. Modification or additional analyses may be required as the data collection progresses. An updated statistical analysis plan will be available on the WWARN study group website.All statistical analyses will be carried out using R  or StataM\u00a015. Alternative statistical software will not require amendment of this SAP.This IPD MA met the criteria for waiver of ethical review as defined by the Oxford Tropical Research Ethics Committee as the research consisted of secondary analysis of existing anonymous data.Findings will be reported following the PRISMA-IPD guidelinesPatients were not involved in the development of the research question, outcome measure or study design.Large-scale deployment of highly efficacious ACT regimens such as AS-MQ has been the cornerstone of global malaria control for over a decade and this has contributed to the global reduction of mortality and morbidity associated with malaria.P. falciparum malaria, treated with AS-MQ. The WHO-recommended AS-MQ regimen is administered as a 3-day course, with a total of 12\u2009mg/kg of\u00a0AS and 25\u2009mg/kg of MQ split over 3\u2009days. Due to the poor tolerability of high-dose MQ, the dose of MQ is usually divided into either two doses (15 and 10\u2009mg/kg), or three as a fixed-dose combination (8\u2009mg/kg/day). The fixed-dose combination has been shown to provide better efficacy and improve treatment adherence for artesunate-amodiaquine.IPD MA is now considered the gold\u00a0standard for evidence synthesis and allows exploration of different risk factors which otherwise would not be possible through the aggregate data MA.In conclusion, this pooled analysis will provide critical information regarding the relationship between drug dosage and parasitological responses post-treatment with AS-MQ. The assessment of the host, parasite and drug determinants that influence the treatment response can provide evidence-based guidance for monitoring the early signs of artemisinin resistance and effective case management that will be critical in optimising malaria control and containment efforts."}
+{"text": "Plasmodium falciparum in pregnancy in India.In India, the recommended first-line treatment for malaria in the second and third trimester of pregnancy is artesunate\u2009+\u2009sulfadoxine-pyrimethamine (AS+SP). However, data on safety and efficacy of artemisinin-based combination therapy (ACT) in pregnancy is limited. This study assessed the safety and efficacy of AS+SP and artesunate\u2009+\u2009mefloquine (AS+MQ) for treatment of P. falciparum mono-infection of any parasite density with or without fever were randomized to receive AS+SP or AS+MQ. Blood slides and filter paper samples for Polymerase Chain Reaction (PCR) were collected on days 0, 1, 2, 3, 14, 21, 28, 42 and 63 post treatment. Women were followed up at delivery and at day 42 postpartum.This open-label, randomized clinical trial was conducted from October 2010 to December 2013 at three sites in India . Pregnant women in the second or third trimester who had P. falciparum mono-infection and were randomized to receive AS+SP (125) or AS+MQ (123) and all of these women were included in the intention to treat (ITT) analysis. The primary endpoint of an adequate clinical and parasite response (ACPR) on day 63 was not available for 9 women who were counted as treatment failure in the ITT analysis. In the ITT population, the ACPR was 121/125  92.0\u201399.1%) in the AS+SP group and 117/123  in the AS+MQ group. Among the 239 women (121 from the AS+SP arm and 118 from the AS+MQ arm) who completed the day 63 follow up  the ACPR was 100% in the AS+SP group and 99.2% (117/118) in the AS+MQ group. There were five serious adverse events (SAE) among pregnant women (4 in the AS+SP group and 1 in the AS+MQ group) and 13 fetal/neonatal SAEs (7 in the AS+SP group and 6 in the AS+MQ) but none of them were related to the study drugs. A higher proportion of women in the AS+MQ arm reported vomiting within 7\u00a0days post-treatment than did women in the AS+SP arm .Two hundred and forty-eight women of 7064 pregnant women (3.5%) who were screened at monthly antenatal clinics had a Both AS+SP and AS+MQ are safe and effective for treatment of uncomplicated falciparum malaria in pregnancy in India.Trial registrationCTRI This study is registered with Clinical Trial Registry India (CTRI), number CTRI/2009/091/001055. Date of Registration 11 January 2010, http://ctri.nic.in/Clinicaltrials/pmaindet2.php?trialid=1185&EncHid=&userName=anvikarThe online version of this article (10.1186/s12936-018-2393-3) contains supplementary material, which is available to authorized users. Plasmodium vivax and quinine for Plasmodium falciparum infections in all trimesters. Since 2011, artesunate\u2009+\u2009sulfadoxine-pyrimethamine (AS+SP) has become the recommended treatment for falciparum malaria in the second and third trimesters of pregnancy [P. falciparum malaria in pregnancy in two eastern states in India.Malaria continues to be a major public health challenge in India with about one million cases reported annually . The burregnancy . Althougregnancy , AS+SP wregnancy . Artemisregnancy . Evidencregnancy . AS+SP aregnancy . HoweverP. falciparum [This individually randomized, open label trial was conducted among pregnant women at three sites in India. Two sites (Ranchi and Jamshedpur) are in Jharkhand state and the third site (Rourkela) is in Odisha state. In all three sites, malaria transmission is perennial and the majority of the malaria cases are due to lciparum .After obtaining written informed consent, pregnant women were screened for malaria at monthly, public antenatal clinics (ANC) in the catchment area of the three study hospitals. At enrolment into the cohort and at each subsequent, monthly ANC visit, blood slides were collected from all women irrespective of whether they had a history of fever or not. In addition, women who had fever or a history of fever were tested with a malaria rapid diagnostic test (RDT) malaria Pf dipstick RDT (Guangzhou Wondfo Biotech Co. Ltd. China). If tested positive, they were referred to the study hospital for further assessment. All blood slides collected were examined on the same day. Women with a positive blood slide were invited to the study hospital the same day to assess eligibility. Women who fulfilled the inclusion criteria were enrolled into the trial and admitted to the study hospital for treatment.P. falciparum mono-infection were eligible for enrolment. The gestational age was determined by ultrasound. Women having any of the following conditions were excluded: (i) Hb\u2009<\u20097\u00a0g/dL; (ii) gestation of\u2009<\u200912 or\u2009>\u200936\u00a0weeks; (iii) age\u2009<\u200918\u00a0years (iv) abnormal liver or renal function tests; (v) a history of taking an anti-malarial within the past 7\u00a0days; (vi) a history of allergy to any of the study drugs; (vii) taking part in any other clinical trial of drugs or vaccines; (viii) severe malaria ; (ix) evidence of severe concomitant infection; (x) known chronic disease ; (xi) a history of convulsions during the present illness or a history of psychiatric disorder or seizures (xii) vivax malaria or a mixed infection.Pregnant women of the study cohort of any parity in the 2nd or 3rd trimester who had a P. falciparum malaria would be needed for inclusion in the trial based on the following assumptions: (1) day 63 adequate clinical and parasite response (ACPR) for AS+SP would be 85%, and that for AS+MQ it would be 95%; (2) loss to follow up would be\u2009<\u200920%; (3) the power of the study would be 90% to detect a 10% difference in ACPR between the treatment groups at a 95% significance level. Because of very low treatment failures in both arms and a lower than expected enrolment rate the sample size was subsequently revised to 300. Study enrolment was stopped at the end of the malaria transmission season in December 2013 when a total of 248 pregnant women were enrolled.Initially, it was estimated that 500 pregnant women with Treatment allocation envelopes were prepared by an independent statistician in permuted blocks of 12 for each site. Eligible participants were randomly assigned to either AS+SP or AS+MQ arm by selecting a sealed envelope that contained the treatment code; envelopes were labeled with unique identifier numbers and kept sequentially.Enrolled women were admitted to the study hospitals for 3\u00a0days for administration of the study drugs. They received either AS+SP (artesunate 200\u00a0mg daily for 3\u00a0days and sulfadoxine 1500\u00a0mg\u2009+\u2009pyrimethamine 75\u00a0mg on day one) or AS+MQ (artesunate 200\u00a0mg\u2009+\u2009mefloquine 440\u00a0mg daily for 3\u00a0days).Miscarriage was defined as loss of the fetus before 28\u00a0weeks of gestation. Abortion was defined as premature expulsion of the products of conception from the uterus. Intrauterine death defined as fetal death prior to complete expulsion or extraction from the mother. Stillbirth: baby born with no signs of life at or after 28\u00a0weeks\u2019 gestation.During the first 3\u00a0days of treatment, women were assessed in hospital. Study participants were then requested to come for follow up visits on days 7, 14, 21, 28, 42 and 63 post-treatment. Women who missed follow-up visits were visited at home by the study team. A blood slide and filter paper sample were collected on day 0, 1, 2, 3, 7, 14, 21, 28, 42 and 63. Venous blood samples for haemoglobin (Hb) measurement, renal and liver function tests were collected at enrolment and on days 14 and 63. Adverse events were assessed at each scheduled and unscheduled visit and treated in a study hospital when appropriate.After the initial 63-day follow-up period, women were seen at monthly ANC clinics and were encouraged to deliver at one of the study hospitals. They were given free transport to reach the study hospital for delivery and postnatal care.At delivery, peripheral blood samples and placental biopsy samples were collected for all institutional deliveries. When deliveries occurred outside the facility, home visits were conducted as soon as possible after birth by a study midwife. Birth-weight, gestational age , and the presence of congenital abnormalities were assessed at delivery or within 7\u00a0days of birth. All participants were followed up on day 42 post-partum to assess the health status of the mother and her baby.Rules for administration of rescue therapy with quinine were: (1) early or late treatment failure; (2)\u2009>\u20093\u00a0g/dL drop in haemoglobin or a haemoglobin level\u2009<\u20097\u00a0g/dL during treatment. The rule for stopping the trial was an unacceptable frequency of adverse events. The Data Safety and Monitoring Board (DSMB) reviewed data regularly and had the authority to recommend to the sponsor stopping the trial if they had major concerns.Thick and thin blood films were Giemsa-stained and examined by two microscopists for species identification. Parasite density was ascertained by counting asexual parasites against 200 white blood cells in the thick film. Parasites were counted against 500 WBCs if the count was less than 99 parasites per 200 WBCs. For determining the parasite density, a white blood cell count of 8000/\u03bcL was assumed. For discordant readings on parasite density the mean was taken if the difference was\u2009<\u200925%. For any other discordant results, the slides were read by a third microscopist whose findings were considered as the final result.P. falciparum infections following WHO PCR genotyping guidelines [msp2, msp1 and glurp. PCR analysis was done at the National Institute for Malaria Research (NIMR) in Delhi on paired blood samples collected on day zero and on the day of failure.In cases of treatment failures, PCR genotyping was carried out to differentiate between recrudescent and new idelines . GenotypCase Report Forms (CRFs) were filled manually. GCP-monitoring, including source data verification, was done for all CRFs before data was double entered and verified in OpenClinica. Data processing and analysis was done using the statistical software package STATA Version IC12.0 .t test were used to test the level of significance for categorical and continuous variables, respectively. The primary trial endpoint, adequate clinical and parasite response (ACPR), was defined as absence of parasite from day 3 until day 63 post-treatment. Treatment efficacy was assessed in the intention to treat (ITT) population, which included all women enrolled in the study, and in the per protocol (PP) population, which included all women who had completed treatment and the follow up on day 63. In addition, a Kaplan\u2013Meier (KM) survival analsysis was performed to compare the probability of treatment failure in both study arms, censoring the women when they were lost to follow up. Safety analysis was performed in the ITT population.Baseline characteristics at enrolment, parasite clearance rates and birth outcomes were summarized and compared across treatment groups. Binominal exact method was used to calculate confidence intervals. Chi square test and The protocol was approved by Ethics Committees of NIMR, Ispat General Hospital Rourkela and the London School of Hygiene and Tropical Medicine, London, UK (LSHTM). Written informed consent was obtained from all participants. An independent Data Safety Monitoring Board consisting of an obstetrician, statistician, pharmacologist and clinician monitored the safety and progress of the study.The study is registered with Clinical Trial Registry India (CTRI), number CTRI/2009/091/001055.P. vivax infection (11), severe malaria (9), RDT positive but blood slide negative (4), intrauterine death (3), jaundice (2), renal disease (1), gestation\u2009>\u200936\u00a0weeks (4) and gestation\u2009<\u200912\u00a0weeks (2).Between October 2010 and December 2013, a total of 7064 pregnant women were screened at public ANC facilities. A total of 248 women (3.5%) were enrolled into the trial . Following PCR correction, it was 100% . The results of the Kaplan\u2013Meier survival analysis showed no difference in the probability of treatment failure in both arms. The KM graph is available as Additional file Mean age, gestational age, Hb concentration and parasite density at enrolment were comparable between the two study arms Table\u00a0. On the There were five maternal SAEs, four in the AS+SP group and one in the AS+MQ group. In the AS+SP group, the SAEs were one maternal death, hospitalization due to oligohydramnios (2) and antepartum haemorrhage (1). The maternal death occurred on day Five due to complications of severe malaria. This woman had high parasitaemia on day 0  and, therefore, should have not been enrolled. In the AS+MQ arm there was one hospitalization due to a urinary tract infection. None of the serious adverse events were considered related to the study drugs. There were 13 fetal or neonatal SAEs, 7 in AS+SP and 6 in AS+MQ arm. The causes of fetal/neonatal SAEs in the AS+SP group were intrauterine death (4), neonatal jaundice (2), still birth (1). In the AS+MQ group they were intrauterine death (2), asphyxia (1), aspiration pneumonia (1) and still birth (2). None of these events were considered related to the study drugs.The observed and reported adverse events during the first week following treatment are shown in Table\u00a0Birth outcomes were available for 97.2% (241/248) women (121 in the AS+SP group and 120 in the AS+MQ group). The sex ratio was 843 females per 1000 males. The occurrence of low birth weight <\u20092.5\u00a0kg) was higher in the AS+SP arm than in the AS+MQ arm  . Due to these low patient numbers it was not possible to reach the original targeted sample size of 500 women. However, a clear result was obtained despite the reduced sample size.Results from this study suggests that AS+SP and AS+MQ are both efficacious for the treatment of MiP in India. The day 63 ACPR and PCR-corrected ACPR were 100% in both groups in the PP population and\u2009>\u200995% in the ITT population. Reasons for the high efficacy observed could be that very few people in the study area had been exposed to any ACT at the time the trial was conducted, and that the parasite density was relatively low  at recruitment. The low parasite density is partly due to the fact that the majority of the trial participants was detected by active screening and did not have clinical symptoms at enrollment; on the day of enrollment only 24.6% of women had fever or a history of clinical symptoms.dhps gene was observed [The efficacy of AS+MQ and AS+SP observed in this study is consistent with that reported from India in non-pregnant adult populations. The day 63 ACPR of AS+MQ was 100% in 2005\u20132007  and 98.3observed . The dayobserved . A revieobserved , AS+AQ [observed . These aIn this study, no serious adverse events related to AS+SP or AS+MQ were reported. However, as the sample size was just 248 subjects, the safety findings from this study need to be interpreted with caution. The incidence of vomiting was significantly higher in women who received AS+MQ than in those who received AS+SP which confirms the finding of a high incidence of vomiting when MQ was used for intermittent preventive treatment in pregnant women . The PREThe observed prevalence of low birth weight 22.4%), pre-term delivery (16.6%) and stillbirth (1.7%) was comparable to the results of a study carried out in Jharkhand state in 2009 [2.4%, preBoth AS+SP and AS+MQ are equally effective for the treatment of falciparum malaria in second and third trimester pregnancies in the central region of India. AS+SP was tolerated better than AS+MQ but it appears that AS+MQ may be slightly more effective than AS+SP at reducing adverse birth outcomes. Given that resistance to SP has emerged and is spreading in India, the risk benefit ratio of other artemisinin-based combinations (such as DHAPQ and AL) needs to be evaluated in pregnant women in India.Additional file 1. Kaplan\u2013Meier failure estimates."}
+{"text": "Plasmodium falciparum malaria since 2005 in Gabon. Since 2011, a rebound of malaria morbidity has been observed in this country, while no survey evaluating ACT efficacy was performed. During the same period, parasite resistance against artemisinin has been reported in Asia. The aim of this study was to assess the efficacy and tolerability of these two drugs in two sentinel sites of Gabon 10\u00a0years after their implementation.Artesunate\u2013amodiaquine (AS\u2013AQ) and artemether\u2013lumefantrine (AL) have been widely used for the treatment of uncomplicated Children aged from 12 to 144\u00a0months with uncomplicated malaria were recruited at the Regional Hospital of Melen, Libreville and in the Urban Health Center of Franceville between March 2014 and September 2015. The therapeutic efficacy was evaluated according to the WHO 2008 protocol of 28-day follow-up and PCR-uncorrected/corrected treatment outcomes were assessed.p\u2009=\u20090.2). Adverse events and serious adverse events were rarely observed with both treatments.One hundred and eighty-five children (98 ASAQ and 89 AL) were followed up until day 28. The PCR-corrected ACPR was 98.9% for AS\u2013AQ and 96.4% for AL. Late therapeutic failure rate was 3.6% and 1.1% for AL and AS\u2013AQ, respectively (AS\u2013AQ and AL are still efficacious and well-tolerated for the treatment of uncomplicated malaria in Gabonese children. Since 2000, the majority of malaria-endemic countries substituted monotherapies by artemisinin-based combination therapies (ACTs) for uncomplicated malaria treatment . ResistaP. falciparum, of which strains with molecular markers of artemisinin partner drugs are highly frequent in different areas of the country [In Gabon, malaria is the second leading cause of hospitalization in pediatrics wards after respiratory tract infections; it is responsible for more than one-third of all febrile patients . Almost  country , 13.At the time of artesunate\u2013amodiaquine (AS\u2013AQ) and artemether\u2013lumefantrine (AL) implementation in public health center as the first- and second-line treatments for uncomplicated malaria 2003\u20132005), respectively, their efficacy was estimated at 99\u2013100% \u20132005, re. In 2013This study presents the results of a clinical trial evaluating AS\u2013AQ and AL efficacy and tolerability in two cities of Gabon (Libreville and Franceville), where both molecular markers of resistance as well as a rebound of malaria morbidity have been previously described , 12.P. falciparum represents more than 93% of the parasite species diagnosed [This prospective study was carried out in two sentinel sites for malaria surveillance between March 2014 and September 2015 in Gabon. Precisely, the study sites were the Operational and Clinical Research Unit (OCRU) of the Regional Hospital of Melen (RHM), which is located at 11\u00a0km from Libreville (West part of Gabon) and the Urban Health Center of Franceville (south-east region of the country). In both cities, malaria transmission is perennial, with an entomological inoculation rate estimated at 33.9 infected bites per person per year [iagnosed , 15.P. falciparum mono infection with a parasite count between 2000 and 200,000 asexual forms per microliter of blood (p/\u00b5L). Criteria of exclusion were the presence of complicated malaria as defined by WHO 2000 [Plasmodium infection, concomitant disease, or other danger signs, known allergy to AS\u2013AQ and AL, presence of a treatment with an antimalarial drug in the previous 3\u00a0weeks.During the study period, children aged from 12 to 144\u00a0months with the following criteria were approached: febrile (tympanic temperature of\u2009>\u200937.5\u00a0\u00b0C) or with a 24\u201348\u00a0h history of fever before the day of consultation, ability to swallow oral medication, willingness to comply with the duration of the study, no signs of complicated malaria as per WHO guidelines , abilityWHO 2000 , a mixed\u00ae Novartis) and AS\u2013AQ  at the study site during three consecutive days. They were observed within 1\u00a0h after administration. Children who vomited less than 30\u00a0min after drug administration were given a second dose and were observed for 30\u00a0min more. They were monitored according to the 28-day-WHO protocol (WHO 2003).After enrollment, participants received appropriate dose of AL , late treatment failure (LTF), late clinical failure (LCF), late parasitologic failure (LPF), and adequate clinical and parasitological response (ACPR) as per WHO, 2003 protocol .In case of therapeutic failure, patients were retreated with dihydroartemisinin\u2013piperaquine phosphate combination (DHAP). Recrudescence and reinfection cases were distinguished using merozoite surface protein 1 (block 2) and 2 (block 3) gene genotyping by PCR as previously described .All data were entered into an Excel-based program and analyzed using the computer program developed by WHO for antimalarial drug efficacy. This software includes formula for the classification of treatment outcome with and without PCR. Patients who were excluded or withdrew from the study were not included in the per protocol analysis.The study was conducted in accordance with the principles of good clinical practices (GCP) and reviewed and approved by Gabonese National Ethics Committee and the World Health Organization (WHO) Ethical Review Committee. A written informed consent was obtained from parents or guardians of all study participants.n\u2009=\u200939/225): 17 from the AL group vs 22 from AS\u2013AQ. The reasons for exclusion are shown in the trial profile .A total of 222 of the 1425 screened children fulfilled the inclusion criteria. Among them, 116 patients received AS\u2013AQ and 106 AL. The excluded patients during the follow-up represented 17.3% (n\u2009=\u200989) and AL  treatment groups (p\u2009=\u20090.3).In both groups, the parasite clearance was obtained from the second day following the first dose administration. The results of the treatment efficacy before and after PCR correction are reported in Table\u00a0The microscopic gametocytemia was cleared from day 1 for AL and day 3 for AS\u2013AQ-treated children.n\u2009=\u20092) vs 5.6% (n\u2009=\u20095) (p\u2009=\u20090.2), cough 16.6% (n\u2009=\u200916) vs 6.7% (n\u2009=\u20096) (p\u2009=\u20090.004), asthenia 14.5% (n\u2009=\u200914) vs 5.6% (n\u2009=\u20095) (p\u2009=\u20090.05), and abdominal pain 2.0% (n\u2009=\u20092) vs 2.2 (n\u2009=\u20092) (p\u2009=\u20090.6). Hemoglobinuria (n\u2009=\u20091) and convulsion (n\u2009=\u20091) were found in only one patient treated with AS\u2013AQ and in one child who received AL.All the recruited patients recovered rapidly during the first 72\u00a0h of follow-up  and remain high and adequate according to WHO recommendations. Others studies reported cure rate varying from 99.3 to 100% after treatment with AS\u2013AQ and from 99.3 to 100% for AL , 24. In 6.5% and AL and AS\u2013AQ are still efficacious and well-tolerated in Gabon. Their use as first-line treatment is not compromised. Continuous monitoring throughout the country is necessary to allow early detection of decline efficacy of both drugs."}
+{"text": "Plasmodium falciparum has led to calls for malaria elimination on the Thai-Cambodian border. However, the optimal approach to elimination in difficult-to-reach border populations, such as the Military, remains unclear.Malaria remains a critical public health problem in Southeast Asia despite intensive containment efforts. The continued spread of multi-drug-resistant A two-arm, cluster-randomized controlled, open-label pilot study is being conducted in military personnel and their families at focal endemic areas on the Thai-Cambodian border. The primary objective is to compare the effectiveness of monthly malaria prophylaxis (MMP) with dihydroartemisinin-piperaquine and weekly primaquine for 12\u00a0weeks compared with focused screening and treating (FSAT) following current Cambodian national treatment guidelines. Eight separate military encampments, making up approximately 1000 military personnel and their families, undergo randomization to the MMP or FSAT intervention for 3\u00a0months, with an additional 3\u00a0months\u2019 follow-up. In addition, each treatment cluster of military personnel and civilians is also randomly assigned to receive either permethrin- or sham (water)-treated clothing in single-blind fashion. The primary endpoint is risk reduction for malaria infection in geographically distinct military encampments based on their treatment strategy. Monthly malaria screening in both arms is done via microscopy, PCR, and rapid diagnostic testing to compare both the accuracy and cost-effectiveness of diagnostic modalities to detect asymptomatic infection. Universal glucose-6-phosphate dehydrogenase (G6PD) deficiency screening is done at entry, comparing the results from a commercially available rapid diagnostic test, the fluorescence spot test, and quantitative testing for accuracy and cost-effectiveness. The comparative safety of the interventions chosen is also being evaluated.Despite the apparent urgency, the key operational elements of proposed malaria elimination strategies in Southeast Asian mobile and migrant populations, including the Military, have yet to be rigorously tested in a well-controlled clinical study. Here, we present a protocol for the primary evaluation of two treatment paradigms \u2013 monthly malaria prophylaxis and focused screening and treatment \u2013 to achieve malaria elimination in a Cambodian military population. We will also assess the feasibility and incremental benefit of outdoor-biting vector intervention \u2013 permethrin-treated clothing. In the process, we aim to define the cost-effectiveness of the inputs required for success including a responsive information system, skilled human resource and laboratory infrastructure requirements, and quality management. Despite being a relatively low transmission area, the complexities of multi-drug-resistant malaria and the movement of vulnerable populations require an approach that is not only technically sound, but simple enough to be achievable.NCT02653898. Registered on 13 January 2016.ClinicalTrials.gov, ID: The online version of this article (10.1186/s13063-018-2931-x) contains supplementary material, which is available to authorized users. Plasmodium falciparum continues to be endemic in border areas of Cambodia. Over the last several years, reports have highlighted the declining efficacy of artemisinin combination therapies (ACT) within a few years of their introduction as first-line therapies [P. falciparum outpaces the development of efficacious antimalarial drugs, the focus has shifted to elimination of MDR malaria on the Thai-Cambodian border [P. falciparum and fewer treatment options being available [Malaria remains a scourge to humankind with 270 million cases and over 600,000 deaths annually . Despiteherapies . As P. fn border . While tTo prevent the spread of multi-drug-resistant malaria, elimination is now being widely advocated in Cambodia, particularly in hard-to-reach mobile and migrant populations including the Military, an under-recognized reservoir of disease , 7. ElimHowever, the question remains: to effectively eliminate malaria in these populations, what are the optimal interventions? A potentially large, but as yet poorly defined, group of asymptomatic parasitemic persons poses a particular challenge to identifying the at-risk population \u201314. ThouUnfortunately, the only licensed, available drug able to prevent transmission of the parasite from human to mosquito, primaquine (PQ), can cause severe, life-threatening hemolytic anemia in persons with glucose-6-phosphate dehydrogenase (G6PD) deficiency, and requires effective screening tests that can be easily deployed at the point of care. To permit the greatest possible simplicity and cost-effectiveness during scale-up in resource-poor areas along the border, the ideal approach would require minimal medical monitoring or follow-up visits. Suitability for scale-up will also need to be assessed for operational feasibility, perception of risk among participants, cost and cost-effectiveness of interventions, and alignment with stakeholder values and policy.Primary objective: to compare the effectiveness of focused screening and treatment (FSAT) following current national treatment guidelines versus monthly malaria prophylaxis (MMP) with dihydroartemisinin-piperaquine (DHA-PIP) in combination with weekly primaquine (PQ) (22.5\u00a0mg) as transmission-blocking and radical curative agent to reduce the risk of malaria infection in personnel residing in military encampments on the Thai-Cambodian borderSecondary objectives:\u25e6 To estimate the incremental benefit of wearing a permethrin insecticide-treated uniform (ITU) over drug therapy and existing vector control interventions compared to a sham insecticide-treated uniform (sITU) treated with water. Civilian participants will have comparable selected outer garments treated\u25e6 To define the proportion of asymptomatic carriers of malaria in the population, including the cumulative incidence and incidence density of microscopic and submicroscopic blood-stage parasitemia and gametocytemia\u25e6 Additional secondary objectives can be found in Table\u00a01We present a pilot operational research protocol to investigate the effectiveness of monthly prophylactic drug administration compared to a focused screen-and-treat approach to inform a regional malaria elimination initiative by the Royal Cambodian Armed Forces (RCAF).Lastly, an overarching aim of this operational study is to increase The Cambodian Military\u2019s capability to diagnose, prevent, and treat malaria, and to understand the population\u2019s willingness to participate in malaria elimination campaigns. To do so, a scalable military malaria elimination \u201cunit of action\u201d is being established at the provincial level, staffed by RCAF personnel who are trained during the course of this study. Regardless of the outcome, the study will provide critically important information to inform elimination policy in Cambodia and the GMS.P. falciparum in provinces with known antimalarial resistance, with a single, low dose of PQ (15\u00a0mg) to prevent malaria transmission. For participants in the FSAT arm with Plasmodium vivax malaria, the recommended treatment for asexual parasitemia is DHA-PIP. All patients with P. vivax in the FSAT arm receive treatment with 15\u00a0mg of PQ daily for 14\u00a0days if G6PD normal, or 45\u00a0mg weekly if G6PD deficient.We are conducting a two-arm, cluster-randomized, open-label controlled trial of MMP with t3-day DHA-PIP and weekly PQ compared to focused screening and treating (FSAT) following current national treatment guidelines. Participants in the MMP arm receive a monthly 3-day course of fixed-combination tablet containing 40\u00a0mg of dihydroartemisinin and 320\u00a0mg of piperaquine phosphate for three consecutive months. Those aged 13\u00a0years and older receive low-dose weekly PQ (22.5\u00a0mg) therapy to prevent malaria transmission for 12 weeks. Participants in the FSAT arm are screened with highly sensitive polymerase chain reaction (PCR) methods, and all those testing positive for malaria receive therapy with first-line antimalarials following current national treatment guidelines. At the time of protocol development, artesunate-mefloquine combination therapy is the first-line treatment for The study takes place at multiple military encampments in malaria-endemic areas in border areas authorized by the Ministry of National Defense. Sites were chosen based on current malaria incidence estimates by the RCAF health services in consultation with the National Center for Parasitology, Entomology, and Malaria Control (CNM) in Cambodia and the Armed Forces Research Institute of Medical Sciences (AFRIMS). For reasons of operational security, locations or other descriptors will not be further described here. All sites will be evaluated as simultaneously as feasibly possible.Military volunteers aged 18\u201365\u00a0years of age and their dependents of \u2265\u20092\u00a0years of age, eligible for care at an RCAF facility, or otherwise eligible Cambodian civilians at risk for contracting malaria who live within close proximity, are enrolled if they meet the following criteria: are able to give informed consent/assent; reside in the selected study areas; are available for monthly follow-up for 6\u00a0months; agree not to seek outside medical care for febrile illness unless referred by study team; and are authorized by respective military unit commanders to participate in the study if on active duty. Volunteers who have known allergy or other contraindication to study medication; are pregnant, lactating or are women of childbearing age who do not agree to use an acceptable form of contraception during the study; or are judged by the investigator to be otherwise unsuitable for study participation are excluded from the study.Geographical areas were randomized prior to study start to a 1:1 allocation of MMP or FSAT using time- and region-blocked randomization to ensure that randomization is geographically distinct, and that malaria incidence is evenly distributed among groups given that high and low transmission areas exist. Volunteers meeting the enrollment criteria receive permethrin- or sham-treated (water-treated) uniforms according to their cluster\u2019s assignment in single-blind fashion (volunteer blinded to assignment but not investigator). Entomology staff conducting uniform testing are also blinded to treatment. Diagnostic microscopists are blinded to each others\u2019 readings and to study arm assignment. There is otherwise no blinding during the study.Volunteers randomized to MMP arm aged 13\u00a0years and older receive a monthly 3-day treatment course of  for 3\u00a0months. They will also receive a weekly dose of 22.5\u00a0mg of PQ for 12 weeks. Children of 12 years of age or less will receive only a monthly 3-day weight-based treatment dose of DP for 3\u00a0months without PQ treatment. In the MMP arm, symptomatic patients or microscopy-confirmed parasitemia in the first 3\u00a0months are considered a failure of prophylaxis and treatment is given according to the national treatment guidelines. If the volunteer has parasitemia detectable by PCR only and is symptom free at monthly follow-up, the study treatment with monthly DHA-PIP is continued for 3\u00a0months as outlined for the MMP arm. To reduce phlebotomy requirements for G6PD testing and potential hemolytic risk in children, PQ is not be administered to those less than 13\u00a0years old.Volunteers randomized to the focused screening and treatment (FSAT) arm are treated according to current national malaria treatment guidelines if they test positive for malaria based on blood smear and/or real-time PCR results during monthly follow-up. In the FSAT arm, all volunteers with parasitemia detectable with PCR receive antimalarial treatment, even if they are symptom- free.P. falciparum, or DP for blood-stage P. vivax infection. Elimination interventions are carried out monthly during the first 3 months after enrollment, and then followed by 3 months of additional malaria assessment by smear and PCR diagnosis with malaria treatment as necessary following current national treatment guidelines  and a single 15-mg dose of PQ for acute nes Fig.\u00a0, Fig.\u00a02.Following documentation of informed consent/assent and determination of eligibility, a study physician completes an initial medical history and physical examination Table . VolunteAll volunteers are followed monthly thereafter or at any time that they present with symptoms of suspected malaria, with a brief clinical evaluation and malaria diagnostics to include PCR assays for asexual and sexual parasites. Malaria-positive volunteers aged 13\u00a0years and older have additional tests for molecular markers of resistance. At these times, a swatch is taken from the volunteer\u2019s uniform or clothing for repellency monitoring.At the conclusion of the 180-day (\u2212\u20092 to +\u20093\u00a0days) follow-up period, volunteers have blood drawn for malaria smears, RDTs, asexual blood-stage and gametocyte PCR diagnosis. If infection is noted at discharge, volunteers will continue to be followed until resolution of infection.Stained thick and thin blood smears are examined by two microscopists blinded to each other\u2019s results and to the treatment status of the study volunteer. Two blood smears are made for every enrolled volunteer. Slide 1 is stained immediately and examination of Giemsa-stained thick and thin smears made. This slide is stored in a different box from slide 2, which is only read if there is a problem with the first slide.Parasite densities are calculated based on a count of parasites per 200 white blood cells (WBCs) (thick film) or for low parasitemias , per 500 WBC or 5000 red blood cells (RBCs) (thin film). Both asexual and sexual stages are enumerated. A total of 200 oil immersion fields are examined on the thick film before a blood smear is considered negative. The final count is determined by taking the geometric mean of the two counts. In case of a difference in results  between the two microscopists, the blood smear is re-examined by a third microscopist blinded to the results of the first two readers and the treatment regimen, and the third reading is accepted as the final result.P. falciparum and P. vivax using 18S ribosomal ribonucleic acid (RNA) (18S rRNA) genes unique to each species. Parasite deoxyribonucleic acid (DNA) will be isolated from approximately 200\u2013250\u00a0\u03bcL of venous or capillary blood collected in an EDTA microtube.Malaria microscopy results is confirmed using real-time PCR correction for the detection of msp1, msp2, and GLURP genes is performed to identify the unique fingerprint of the infecting parasite and any subsequent development of malaria after therapy in order to determine if it is a recrudescent infection or new infecting genotype [One milliliter of blood is drawn on day 1 if malaria-positive and at recurrence. PCR genotyping of genotype . Gametocgenotype \u201326.On enrollment, volunteers are assessed for G6PD deficiency with both fluorescence  and quantitative testing, and one or more commercially available point-of-care test kits, and correlated with single nucleotide polymorphism analysis for G6PD gene mutations. Approximately 1.5\u00a0mL per blood draw is collected in an EDTA (anticoagulant) tube for this purpose.Venous blood is tested for qualitative G6PD activity using the fluorescent spot test method, as recommended by the International Committee for Standardization in Hematology using commercially available kits . This method detects fluorescence of the enzyme NADPH under long-wave (365\u00a0nm) UV light. Reduction of NADP to NADPH occurs in the presence of G6PD. The rate and extent of NADPH formation is proportional to G6PD activity. Normal samples fluoresce brightly, whereas deficient samples show little or no fluorescence .Quantitative testing will be performed using an FDA-approved test kit  and results will be calculated based on same-day hemoglobin values from the CBC. Severe deficiency (WHO class I or II) will be defined as 10% or less of the lower limit of normal activity (in G6PD activity units per gram of hemoglobin) established for the quantitative assay system. Subjects with severe deficiency will be enrolled in the study as this is not an exclusion criterion; class I\u2013V deficiencies are permissible for enrollment. To evaluate the performance of a commercial G6PD deficiency RDT in malaria patients from Cambodia, the CareStart\u2122 G6PD test is being compared to the \u201cgold standard\u201d G6PD enzyme quantitative test for their potential use as point-of-care diagnostics for G6PD deficiency. However, no clinical decisions will be made based on the results of either point-of-care test in this study.Each uniform is treated by study staff according to the site\u2019s randomization prior to distribution with a single application of the 40% permethrin Individual Dynamic Absorption (IDA) kit from the US Army\u2019s supply system or water only for sham-treated uniforms. Permethrin treatment is designed to last for 30 washings, which should be suitable for the 6-month duration of the study. At enrollment, each volunteer has one to two field uniforms treated, and civilian volunteers have one to two sets of outer covering garments that they wear while in forested areas treated. The number of clothing sets treated will depend on the number in the volunteer\u2019s possession, and additional clothing sets may be treated during the course of the study if/when the volunteer acquires new clothing, or when treated clothing is damaged or becomes otherwise unserviceable.Monitoring of treated uniform repellency will be performed by taking uniform swatches numbered with the subject\u2019s study ID at monthly intervals for 6\u00a0months. The numbered swatches will be sewn into the uniforms at the time of treatment. Blinded entomology technicians will randomly select 5% of uniform swatches, and place them in cages of female mosquitoes to observe for repellency. The clothing swatches will be placed on the wall of the mosquito cages and proportional landings on the swatch side of the cage will be assessed.Up to 1200 persons are being enrolled and followed in this cluster-randomized study. The study compares estimates of the absolute risk reduction based on the proportion remaining malaria free at the end of 6\u00a0months. We assume that MMP administration combined with an insecticide-treated uniform (ITU) on the background of any pre-existing vector control methods that might be employed will need to reduce malaria incidence to zero by study conclusion to achieve elimination. The MMP approach will be using nearly every currently available intervention in Cambodia to eliminate malaria in this population. Current mathematical models show that multi-round mass drug administration (MDA) can reduce malaria prevalence to less than 10% if high coverage is achieved . FocusedBecause observations of individuals in the same geographical area  are correlated (non-independent), the effective sample size needed is larger than an individualized randomized trial. Given that the intracorrelation coefficient (ICC) is not known, we assumed a conservative value of 0.4. To detect a minimum of a two-fold difference in the effectiveness of MDA over FSAT in proportion of volunteers being malaria-free at 6\u00a0months with an \u03b1\u2009=\u20090.05 and 80% power, the number needed in each cluster is at least 78 individuals . Given tThe primary study endpoint will be the absolute risk reduction (ARR) with 95% confidence intervals (CIs) based on the proportion of volunteers in each arm with absence of PCR-positive parasitemia at the end of 6\u00a0months using an intention-to-treat analysis Table . In addiSecondary study endpoints are pilot endpoints to include incidence rates, molecular and parasitological endpoints, safety, vector control effectiveness , and comparison of malaria and G6PD diagnostic modalities. The calculation of incidence rates  for prophylactic and FSAT groups will be calculated adjusted and unadjusted with 95% CI. All volunteers completing at least one follow-up assessment will be included in the safety analysis. The safety analysis database will include those volunteers in the set of randomized volunteers who receive at least one dose of study drug. All parasitological data will be included in the parasitological analyses. A stratified analysis comparing the vector control arms within each group will also be compared as those remaining malaria-free at 180\u00a0days. Given the possibility of the highly variable nature of transmission at the geographical sites, existing pre-enrollment malaria surveillance data by RCAF and day 1 screening data by AFRIMS will guide if a pair-matched analysis is necessary.Any volunteer with G6PD deficiency who experiences grade 3 hemolysis following PQ administration will be discontinued from the use of PQ and monitored closely. If more than three subjects with G6PD deficiency are found to have grade 3 hemolysis following treatment with PQ for anti-relapse therapy, further treatment with PQ will be suspended for all G6PD-deficient subjects enrolled in the study.Clinical and laboratory data pertaining to drug efficacy are collected and managed by AFRIMS Immunology and Medicine staff and RCAF personnel. All data and medical information obtained from study volunteers are considered privileged and confidential. Volunteers enrolling in the study are issued a unique identification code (UIC), which is used on all study files and clinical sample labels. Individually identifiable volunteer information other than the UIC are not transcribed on other study documents to include laboratory sample labels, case report forms, nor included in the presentation of study results. Data is double-entered into a Microsoft Access database (Microsoft Corp) with source data verification for each entry.This trial has been approved by the Walter Reed Army Institute of Research (WRAIR) and the National Ethical Committee for Health Research (NECHR) in Cambodia.Elimination of malaria in mobile and migrant populations in border areas is of increasing importance as these transient populations carry a significant amount of MDR malaria burden in the GMS , 29, 30.While terms describing \u201cmobile populations\u201d are often used in reference to malaria elimination, they remain poorly defined, and encompass a wide range of persons including economic migrants, ethnic minorities, military and other government personnel, and tourists. Mobility among malaria endemic areas is their common shared trait, and for the most part, their mobility makes them more difficult to study. In this regard, military and government personnel may be the easiest of these varied groups to study because of the ability to provide comprehensive malaria prevention and treatment services reliably despite frequent movement. Military organization and chain of command provide frequent opportunities for structured intervention, suggesting that militaries may be among the easiest mobile groups to achieve early malaria elimination. Despite these advantages, the need to preserve operational and information security and the need to ensure that research does not interfere with the imperatives of military service remain important barriers. The Military is also considered a vulnerable population for medical research, and additional steps must be taken to ensure that study participation is never coerced through the chain of command. Potential transmission between surrounding communities and frequent personnel movements are additional challenges in designing interventions and assessing results .P. falciparum malaria and DHA-PIP for P. vivax malaria. While PQ use \u2013 either single-dose for P. falciparum transmission-blocking or 14-day anti-relapse therapy for P. vivax infections \u2013 is currently advocated by treatment guidelines, implementation has been limited and use officially restricted to facilities able to screen for G6PD deficiency, greatly limiting application outside controlled research studies. Perhaps more important is the question of which, if any of the drugs involved can be safely administered on a large scale, with minimal or no medical supervision. Such practice, termed \u201cmass drug administration,\u201d is often considered as a means to treat significant numbers of infected individuals in limited-resource settings [The study is effectively comparing a large-scale malaria prophylaxis approach using currently available interventions with a more systematic version of current public health efforts to identify and treat infected persons. The primary objective of this study is to directly compare the effectiveness of MMP with DHA-PIP and weekly PQ against an active screening and treatment strategy following the current national malaria treatment guidelines. In the province where the study was conducted at the time of study inception, these included artesunate and mefloquine combination treatment for all acutely infected persons with settings . Becausesettings .least unfavorable of currently available options in order to make current progress toward elimination rather than await alternatives. Though its efficacy has plummeted below accepted thresholds for treatment of acute P. falciparum infection, it is likely to retain efficacy against subclinical P. falciparum encountered in pre-elimination settings, and P. vivax infections [Selection of appropriate chemoprophylaxis requires a safe, tolerable, and effective drug that can be administered widely in the population. Such options, limited to begin with, are rapidly dwindling in the GMS as MDR malaria continues to gain a greater foothold . The invfections , 13, 44.P. vivax relapse in a cohort of Southeast Asian refugees in Australia in 1984 [While considered an essential component for malaria elimination \u201347, the  in 1984 . This wo in 1984 , 49. RisAnother component of the study is designed to measure whether using permethrin-treated clothing provides additional protection against malaria infection. The incremental benefit of an oft-proposed outdoor-biting vector intervention \u2013 permethrin-treated clothing \u2013 will be assessed. The results of limited prior clinical evidence suggested mixed outcomes. The intent of the present study is to evaluate potential additive benefits of permethrin-treated uniforms over the primary pharmacological interventions. Evidence for effectiveness of permethrin-treated clothing is largely limited to entomological studies given the scarcity of controlled clinical trials evaluating disease outcomes . While tIn the present pilot study it is necessary to make trade-offs between complete rigor in statistical calculations in order to conduct the study among military personnel actively guarding a national border and their families. The study protocol aims not only to assess the comparative clinical effectiveness of MMP versus FSAT interventions (the primary objective), but also the practical feasibility of conducting intensive interventions at scale. A key deliverable to public health authorities will be a candid assessment of the degree to which competing elimination strategies  could be achieved with limited medical supervision and safety monitoring in the field. As a result, clinical effectiveness and practical feasibility will be equally important outcome measures to determine next steps. It will be an ongoing challenge to conduct well-controlled studies in austere border areas, let alone rigorous elimination efforts. The study protocol design is logistically constrained by virtue of being primarily conducted in an active duty military population. The practical limits imposed on the number of clusters available, and finite follow-up periods are functions of the potential impact of the study on availability of RCAF personnel. While the limited number of clusters may effectively reduce power to draw definitive conclusions from the study, we accepted this compromise in order to assess operational feasibility. A randomized study of geographically isolated clusters also risks disease transmission from surrounding inhabitants and those choosing not to enroll in the study. This is another accepted risk. Because the present study cannot be confused with a public health directive by the government, it will be unethical to compel any inhabitant within or surrounding designated clusters to participate. This will be accounted for in the final interpretation of study results. The risks of transmission from unenrolled persons in the area will also be assessed by study team members during the trial  so as to incorporate into the final analysis to the best of our abilities.Analysis of the effectiveness of MMP versus FSAT is the primary intervention (and outcome) of interest based on absolute risk reduction at 24\u00a0weeks. While the MMP arm reverts to standard-of-care  after the 12-week intervention period, it would be unethical to not treat study participants who suffer new malaria infections during the follow-up period. This is intended to simulate a likely elimination program where an intervention is tried, and then the community defaults to standard of care while it is monitored afterward to determine effectiveness. While there is the potential for overlap between groups due to treatment during the follow-up period, the hypothesis is that the more intensive MMP intervention will reduce overall risk of disease further than the less intensive FSAT approach in the allotted study window. While we also wish to assess any additive effects of permethrin-treated clothing over pharmacological interventions in the treated clusters, the study is not powered based on permethrin treatment. In addition to estimation of additive effects, the intervention was performed to assess feasibility to effectively implement permethrin treatment and determine adherence to clothing wear.It is anticipated and often argued that the large effort required to effectively achieve malaria elimination will preclude medical and safety monitoring. The feasibility of this approach will be assessed through analysis of diagnostic and adverse event data. If determined that elimination objectives could not be met without the intensity of medical monitoring to be employed here, it is possible that the study may conclude that approaches to malaria elimination studied here are not currently feasible. Technical, organizational and/or resource constraints may favor a more gradual approach as patient safety must always remain a paramount consideration. Regardless of outcomes, this initial study will serve as a pilot for scale-up efforts, and inform future operational approaches. Lastly, in an effort to optimize safety, due to phlebotomy limits for younger children, pediatric volunteers aged less than 13\u00a0years will not be screened for G6PD deficiency and will not receive PQ. While not expected based on known demographics of military dependents living along Cambodia\u2019s borders, higher than anticipated pediatric enrollment could limit the study\u2019s ability to discern potential benefits of the selected PQ dose.P. falciparum malaria elimination by 2020, and all malaria eliminated by 2025, a multitude of research efforts are underway to develop best practices. While mathematical models provide some guidance, evidence from randomized controlled trials on the most effective practical interventions for elimination is lacking. To date, most trials of mass drug administration have been historical, poorly designed, non-randomized or controlled [As Cambodia approaches its stated goal of ntrolled . Use of Additional file 1:Standard Protocol Items: Recommendations for Interventional Trials (SPIRIT) 2013 Checklist: recommended items to address in a clinical trial protocol and related documents. (DOC 147 kb)"}
+{"text": "The purpose of this study was to compare the effect of 300\u2009mg of bupropion and 8\u2009mg of buprenorphine per day on the treatment of methamphetamine withdrawal cravings over a 2-week treatment interval.Diagnostic and Statistical Manual of Mental Disorders, 4th edition, text revision) criteria for methamphetamine dependence and withdrawal were randomly divided into two groups. Subjects randomly received 300\u2009mg of bupropion or 8\u2009mg of buprenorphine per day in a psychiatric ward. Of the 65 subjects, 35 (53.8%) received buprenorphine and 30 (46.2%) received bupropion. The subjects were assessed by using methamphetamine craving score, interview, and negative urine drug test.Sixty-five methamphetamine-dependent men who met the DSM-IV-TR (P = 0.011). Overall, a significant main effect of day (P <0.001) and group (P = 0.011) and a non-significant group-by-day interaction (P >0.05) were detected.There were no statistically significant differences between the two groups in regard to age, education, duration of methamphetamine dependency, marital status, employment, and income. The mean ages were 32.8\u2009years (standard deviation (SD) = 7.26, range = 22 to 59) for the buprenorphine group and 32.21\u2009years  for the bupropion group. All 65 patients completed the 2-week study. Both medications were effective in the reduction of methamphetamine cravings. Reduction of craving in the buprenorphine group was significantly more than the bupropion group  registration number:  Mental disorders have been growing problems around the world , 2. AmonMethamphetamine abuse induces an elevated mood associated with increased wakefulness, physical activity, and energy . ElongatMethamphetamine is abused globally. In the US, 18 million people over age 12 have used methamphetamine during their life [Previously in Iran, methamphetamine was illegally smuggled in from other regions of the globe, mostly the West , but curBuprenorphine administration is for the treatment of opioid withdrawal symptoms, and bupropion is for the treatment of nicotine dependence . Now we Buprenorphine, a semi-synthetic partial agonist at mu-opioid receptors and antagonist at delta- and kappa-opioid receptors, has been investigated largely for the treatment of opioid use disorder . It is cSubstances such as alcohol, methamphetamine, and cocaine stimulate release of dopamine from cells originating in the brain\u2019s ventral tegmental area (VTA) region, which is a component of a neuronal circuit called the mesolimbic dopamine system and is connected with behavioral reward and motivation. After exposure to alcohol, methamphetamine, or cocaine, dopamine released into the nucleus accumbens and prefrontal cortex reinforces alcohol-, methamphetamine-, and cocaine-seeking behaviors \u201320.To the best of our knowledge, based in part on a review of the literature, few studies on this matter (comparing the effects of buprenorphine with those of bupropion for the treatment of severe methamphetamine cravings) have been published , 10.Furthermore, we are administering bupropion as a new approach for the treatment of methamphetamine withdrawal craving because we hypothesize that  the biochemistries engaged in methamphetamine and bupropion use are more or less alike (both medications boost the level of dopamine) , 9, 11.Diagnostic and Statistical Manual of Mental Disorders, 5th edition (DSM-5) (American Psychiatric Association) [This study is a clinical trial that presents data obtained from comparing sublingual buprenorphine with oral bupropion in the treatment of severe methamphetamine withdrawal craving. The primary goal of the present research was to appraise the efficacy of 8\u2009mg of sublingual buprenorphine and 300\u2009mg of oral bupropion per day in the treatment of methamphetamine withdrawal cravings. Craving is a major feature of substance use disorders, including stimulant use disorders, as evidenced by its recent addition to the diagnostic criteria for these disorders in the ciation) , and it ciation) .Diagnostic and Statistical Manual of Mental Disorders, 4th edition, text revision (DSM-IV-TR) criteria by a board-certified psychiatrist through the Structured Clinical Interview for DSM-IV, clinical version (SCID-I).This study was a randomized, double-blind clinical trial of methamphetamine-dependent men who had been referred to the main psychiatric hospital affiliated with the Shiraz University of Medical Sciences. Severe methamphetamine dependence and withdrawal were diagnosed on the basis of the In order to have eligibility, subjects were examined and questioned by a board-certified psychiatrist at screening. Prior to each interview, we described the aims of the study, guaranteed confidentiality, and obtained written informed consent. The interviews and examinations were achieved on the premises of the treatment hospital because it appeared to be a non-threatening and suitable environment. Family members, friends, or relatives accompanied subjects to the hospital; this attendance provided a condition to confirm the data and information acquired from the patients.The inclusion criteria were (1) daily use of methamphetamine for at least 12 months, (2) being male , and (3) having a positive urine toxicology for methamphetamine.The exclusion criteria were (1) unstable medical conditions, organic mental disorders, major medical diseases , or any type of psychosis; (2) intolerable complications arising from the use of buprenorphine/bupropion; and (3) dependency to or abuse of drugs/substances other than methamphetamine (excluding tobacco).All subjects provided written informed consent before going into the clinical trial. The study was approved and monitored by the ethics committee of Shiraz University of Medical Sciences, which adheres to the Declaration of Helsinki Ethical Principles for Medical Research involving human subjects.We employed a standard randomization procedure generated by computer to have a random sample set for selecting participants from the patients who met the inclusion criteria and had a desire to participate in the study. In a double-blind manner, the individuals were allocated to groups of either buprenorphine or bupropion by using a random allocation method.The research team was adequately trained and includes an addiction psychiatrist, general psychiatrist, general physician, psychologist, nurse, and statistician. The pills had the same shape and color. The patients and the research staff were blind to the consuming medications for the period of the study. The ratings and interviews were carried out by a fully trained physician who was unaware of medications and adverse events. All of the pills, including buprenorphine/bupropion, had the same color and shape. During the trial, no other intervention was allowed.Patients were exactly and precisely monitored and assessed for 14\u2009days. Outcome was measured every day by scoring of craving based on the craving scale, a daily interview for side/adverse effects by a trained physician, and twice-a-week negative urine drug tests .Tolerability and safety of buprenorphine and bupropion were evaluated by daily interview for any side effects, spontaneously reported adverse event data, and rates of premature termination for side effects.A valid and reliable visual analogue scale that has been used previously , 14\u201316 wSubjects were randomly allocated onto 8\u2009mg/d of sublingual buprenorphine without naloxone (4 mg of buprenorphine twice a day) or 300\u2009mg/d of oral bupropion (150\u2009mg of bupropion twice a day). A trained nurse administered the medications. Subjects received treatment for 14\u2009days.It should be mentioned that, on the first day, patients who were blind to the medications received only 4 mg of buprenorphine once a day or bupropion 150\u2009mg once a day and, from the second to the 14th day, received 8\u2009mg/d of buprenorphine  or 300\u2009mg/d of bupropion .t tests analyses were used to test for differences in means, and chi-squared analyses were used to test for differences in frequencies. We used a mixed design \u201ctwo-way ANOVA\u201d; time (day) was a repeated measure factor, and group (bupropion versus buprenorphine) was the between-subjects factor. Then the crux of the analysis is testing the interaction term (time-by-group) and reporting the associated inferential statistics. All P values were two-sided, and statistical significance was set at a 5% level.Data analysis was performed by using SPSS version 18. Repeated measure two-way analysis of variance (ANOVA) and Student The CONSORT  flow and checklist of patients in the trial are shown in Figs. t test = \u22120.205, degrees of freedom (df) = 1, P = 0.838). The mean durations of education were 9.12 years (SD = 2.84) for the buprenorphine group and 9.97\u2009years (SD = 3.11) for the bupropion group . The mean durations of methamphetamine dependency were 9.69 years (SD = 6.28) for the buprenorphine group and 7.48\u2009years (SD = 4.65) for the bupropion group .All 65 subjects completed the 2-week period. Therefore, the data were collected from 65 methamphetamine-dependent men whose mean age was 32.98\u2009years (standard deviation (SD) = 7.9, range = 17 to 59). Of the 65 subjects, 35 (53.8%) received buprenorphine and 30 (46.2%) received bupropion. The mean ages were 32.8\u2009years  for the buprenorphine group and 32.21\u2009years  for the bupropion group . Furthermore, in the bupropion group, there are significant differences in craving scores from days 1 to 14 (P <0.001).Table P = 0.011)  and group (P = 0.011) and a non-significant group-by-day interaction (P >0.05) were detected.Overall, a significant main effect of day  had significant nausea, vomiting, or hypotension, which were treated by symptom therapy (anti-emetic medications or hydration). This nausea, vomiting, or hypotension was thought to be due to exceeding the patients\u2019 level of buprenorphine or bupropion tolerance. No other severe gastrointestinal, cardiovascular, respiratory, or adverse effects or drug intolerance were observed or reported in the other patients.Since there was a concern that such drugs could affect hepatic enzymes, blood tests for liver functioning were carried out at baseline and also at follow-ups. Outcome did not illustrate any significant hepatic enzyme changes.Opioid receptor, mainly mu-opioid receptor, a member of the opioid neuromodulatory system and of the large family of G protein\u2013coupled receptors, is the prominent pharmacological target for the treatment of moderate to severe pain and is of therapeutic value for the management of abuse of amphetamines, opioids, cannabis, alcohol, and other drugs \u201333.The mechanism of action by which opioids such as buprenorphine prevent methamphetamine craving and dependence is not fully understood; however, there are basic and essential interactions between dopamine and the endogenous opioid neuropeptide systems. Naltrexone (an opioid antagonist) decreases and interrupts the interactions between dopamine and the endogenous opioid neuropeptide systems \u201321. We tThe results of this study are supportive of the effect of buprenorphine and bupropion for the treatment of methamphetamine craving. There was superiority of buprenorphine compared with bupropion. We recommend these two medications as short-term and in-patient treatments to enhance retention or even as longer-term maintenance treatment to reduce relapse.P = 0.011).Although we had no control group, the fact that the two medications differed significantly in reduction of methamphetamine craving can compensate for this limitation; in the mean of craving between the buprenorphine and bupropion groups, there is a significant difference  prevent short- or long-term relapse.It can be considered that buprenorphine is a practical and safe medication for the cessation or reduction of methamphetamine withdrawal craving and is better than bupropion. Use of buprenorphine could be considered for the treatment of methamphetamine craving."}
+{"text": "Plasmodium falciparum K13-propeller (Pfk13) domain and amplification in Pfplasmepsin 2 (Pfpm2) gene in Somalia.Artemether\u2013lumefantrine (AL) and dihydroartemisinin\u2013piperaquine (DHA/PPQ) are the recommended first- and second-line treatments, respectively, for uncomplicated falciparum malaria in Somalia. The studies reported here were conducted to assess the efficacy of these artemisinin-based combinations and the mutations in One-arm prospective studies were conducted to assess the clinical and parasitological responses to DHA/PPQ and AL at two sites in 2016 and 2017, respectively, using the standard WHO protocol. The patterns of molecular markers associated with artemisinin and PPQ resistance were investigated for the first time in Somalia.Pfk13, only one (0.7%), from Bosaso, contained a non-synonymous mutation (R622I), which is not one of the known markers of artemisinin resistance. No Pfpm2 amplification was observed among the 135 samples with interpretable results.A total of 339 patients were enrolled with 139 for AL and 200 for DHA/PPQ. With AL, no parasite recurrence was observed among patients treated at either site, corresponding to 100% clinical and parasitological responses. For DHA\u2013PPQ, an adequate clinical and parasitological response rate\u2009>\u200997% was observed. All study patients on both treatments at both sites were parasite-free on day 3. Of the 138 samples with interpretable results for the polymorphism in AL and DHA/PPQ were highly effective in the treatment of uncomplicated falciparum malaria, and there was no evidence of resistance to artemisinin or PPQ. These two combinations are thus relevant in the chemotherapeutic strategy for malaria control in Somalia.Trial registration ACTRN12616001005448 (Jowhar DP study), ACTRN12616000553471 (Bosaso DP study), ACTRN12617001055392\u00a0(AL study in Bosaso and Jowhar) Plasmodium falciparum [P. falciparum k13-propeller (Pfk13) domain [Pfplasmepsin 2 (Pfpm2) gene amplification [Pfk13 mutations are so far rare in Africa, and the mutation most frequently observed is A578S, which is not associated with clinical or in vitro resistance to artemisinins [Malaria is still a major health problem: it caused an estimated 219 million cases and 435,000 deaths worldwide in 2017, most of which occurred in Africa [lciparum  and, morlciparum  in Southfication . A rapidfication , 11\u201313. misinins , 15.The World Health Organization (WHO) recommends routine monitoring of the efficacy of the recommended ACT at least every 2\u00a0years . This isPfdhfr/Pfdhps quadruple and quintuple mutations, while AL was highly efficacious [Pfk13 and amplification in the Pfpm2 gene.The National Malaria Control Programme (NMCP) of Somalia recommended artesunate\u2009+\u2009sulfadoxine/pyrimethamine (AS\u2009+\u2009SP) as first-line treatment in 2006 and artemether\u2013lumefantrine (AL) as second-line treatment in 2011 for uncomplicated falciparum malaria. Therapeutic efficacy studies conducted in 2011 and 2013 showed high AS\u2009+\u2009SP treatment failure rates (12\u201322%), with high levels of icacious , 19. On One-arm prospective studies were conducted with the standard WHO protocol  to evaluThe studies were conducted at two sentinel sites Fig.\u00a0: Jowhar P. falciparum mono-infection with a parasite density of 500\u2013200,000 asexual parasites/\u03bcL of blood and consented to participate in the study. There was no upper age limit. Patients were not enrolled if they presented with the following: severe malaria, mixed or mono-infection with non-falciparum species, known hypersensitivity to the study drugs, severe malnutrition, non-malaria febrile illness  or known underlying chronic diseases . In addition, patients on regular medication that might have interfered with the pharmacokinetics of the study ACT and those with a history of hypersensitivity reactions to the medicines were excluded.Patients aged\u2009\u2265\u20096\u00a0months, excluding girls aged 12\u201317\u00a0years and unmarried women aged\u2009\u2265\u200918\u00a0years, as the local customs and culture would not allow pregnancy testing), with suspected uncomplicated malaria infection (axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C or history of fever during the previous 24\u00a0h) were tested for malaria parasites and were recruited if they had \u00ae, Holley-Cotec Pharmaceuticals China) once daily for 3\u00a0days. AL (20/120\u00a0mg) was administered according to the following weight ranges: one tablet was given to patient weighing 5\u201314\u00a0kg; two tablets for 15\u201324\u00a0kg; three tablets for 25\u201334\u00a0kg and four tablets for\u2009\u2265\u200935\u00a0kg. DHA/PPQ was given as follows: half, three-quarters, one tablet, two tablets, three tablets and four tablets to patients weighing 5 to\u2009<\u20098\u00a0kg, 8 to\u2009<\u200914\u00a0kg, 14 to\u2009<\u200925\u00a0kg, 25 to\u2009<\u200936\u00a0kg, 36 to\u2009<\u200960\u00a0kg and 60 to\u2009<\u200980\u00a0kg, respectively. The study medicines were provided by the WHO, and all treatment doses were given under the direct supervision of a designated study team member. Each patient was observed for 30\u00a0min after administration; if he or she vomited during this period, the full dose was given again. Patients who vomited a second time were withdrawn from the study and referred or admitted to hospital, where parenteral artesunate was administered according to the national treatment policy.Eligible patients were given a standard therapeutic oral dose of either AL  twice daily for 3\u00a0days or DHA/PPQ (Duo-CortecxinPatients were followed up daily for the first 3\u00a0days after the first dose (day 0) and then weekly from day 7 onwards, up to day 28 for the AL-treated groups. Follow-up was extended to day 42 for DHA/PPQ-treated groups because piperaquine has a longer half-life compared to lumefantrine. Patients were also assessed on an unscheduled day if symptoms occurred. Clinical and laboratory evaluations were undertaken during the follow-up visits. Adverse events and severe adverse events, defined according to the WHO protocol for monitoring therapeutic efficacy of anti-malarial medicines , were moThick and thin blood smears were taken from finger-prick blood on day 0 and during follow-up visits and stained with Giemsa. Parasites were counted on Giemsa-stained thick films and recorded as the number of asexual parasites per 200 white blood cells . A smear was declared negative if no parasites were seen after 1000 white blood cells were counted. The presence of gametocytes at enrolment or follow-up days was recorded. In addition, 100 fields of the thick smear on day 0 were examined to exclude mixed infections; in case of any doubt, the thin film was examined for confirmation. Each blood smear was examined independently by two qualified microscopists, and parasite density (per \u00b5L) was calculated on the assumption of a leucocyte count of 8000 per \u00b5L blood. Final parasitaemia was calculated by averaging the readings of the two microscopists if they were in agreement (difference in parasite densities\u2009<\u200950%). If the two counts were discordant in terms of parasite positivity, species or density by\u2009>\u200950%, a third, independent microscopist re-examined the blood slides. For parasite species and positivity, two concordant results were considered the final result, while for parasite density, the average of the two closest estimates of parasitaemia was considered final.msp1, msp2 (merozoite surface proteins 1 and 2) and glurp (glutamate-rich protein) loci, as recommended by the WHO [In order to differentiate recrudescence from re-infection, blood samples were collected on filter paper from a finger-prick on day 0 and on the day of parasite recurrence (day 7 onwards) for genotyping. Specimens were dried, stored in individual plastic bags with desiccants and protected from light, humidity and extreme temperatures. The samples were analysed at the Institut Pasteur, Cambodia. Each dried blood spot was punched with a sterile puncher, and the spots were placed in a 96-well plate in numerical order. Samples were lysed overnight in a saponin solution, and then DNA was extracted with Instagen Matrix resin, as previously described . DNA sam the WHO . The resPfk13 propeller domain, which is associated with artemisinin resistance. The propeller domain was amplified in a nested-PCR assay, amplicons were sequenced according to Sanger\u2019s method , and DNA sequences were analysed to identify specific single nucleotide polymorphisms (SNPs) related to artemisinin resistance [Pfpm2 genes was assessed by reverse transcriptase-PCR (sybr green dye). The full method has been described by Witkowski et al. [Parasite DNA was analysed on day 0 for the presence of mutations in the sistance . The amii et al. .According to the WHO criteria , treatmeAssuming a treatment failure of 5% for both AL and DHA/PPQ with 95% confidence level and\u00a05% precision, a minimum sample of 73 patients per site per drug was estimated. With a 20% increase to allow loss to follow-up and withdrawals during the 28-day (AL) or 42-day (DHA/PPQ) follow-up period, 88 patients per site per drug were targeted.\u00ae database specifically designed for studies of anti-malarial drug efficacy .The clinical and laboratory data of each patient were recorded on standard case record forms, double-entered independently and analysed with an WHO ExcelThis software automatically computes treatment outcomes by both per-protocol and Kaplan\u2013Meier analysis. For the per-protocol analysis, patients were excluded if they were lost to follow-up or withdrawn because of withdrawal and protocol violation. In addition, patients with reinfections or undetermined or missing PCR were also excluded from PCR corrected per-protocol analysis. For the Kaplan\u2013Meier analysis, patients who were lost to follow-up or withdrawn were censored on the last day of follow-up according to the timetable. For the PCR corrected Kaplan\u2013Meier analysis, patients with reinfection were censored on the day of reinfection while those with undetermined or missing PCR were totally excluded.Baseline characteristics  at the two study sites were compared by study drug. Chi squared tests were used to compare categorical data. Differences in the baseline mean age and parasite density were evaluated in a two-sample Wilcoxon rank-sum (Mann\u2013Whitney) test for non-normally distributed data. Gametocyte positivity rates on enrolment and on follow-up days were computed. In addition, clearance of gametocytaemia after treatment among gametocyte carriers at recruitment was assessed by Kaplan\u2013Meier survival analysis. Confidence intervals were calculated for binomial proportions.The study protocol was approved by the Ministry of Health of the Federal Government of Somalia, the Ministry of Health of Puntland and the WHO Research Ethics Review Committee. Patients or parents/guardians of study children were informed about the study and provided written consent. If a patient, parent or guardian was illiterate, he or she chose a witness to co-sign the consent form. Informed assent was obtained from children aged\u2009\u2265\u200912\u00a0years. Travel costs for scheduled and unscheduled visits were reimbursed. In Jowhar, community leaders of the study villages were informed about the study objectives and procedures and gave their permission.Potential study patients were screened in Jowhar from August to October 2016 (DHA/PPQ) and 2017 (AL), and in Bosaso from January to February (DHA/PPQ), and from November 2017 to February 2018 (referred as 2017 in the rest of the document) for AL. A total of 2583 febrile patients were screened for eligibility criteria and 339 patients were recruited  higher than that in Jowhar for both DHA/PPQ and AL groups. The geometric mean parasite density at enrolment was higher among patients in Jowhar than in Bosaso, although the difference was significant (P\u2009<\u20090.01) only between the groups treated with DHA/PPQ. The proportion of patients with detectable gametocytaemia at enrolment was higher among patients in Jowhar than in Bosaso for both treatment groups  and not for the AL groups .The baseline age, axillary temperature and parasitaemia of the study patients are summarized in Table\u00a0Eight and six patients were lost to follow and withdrawn, respectively Fig.\u00a0. The reaP\u2009=\u20090.006) and in Bosaso , as shown in Fig.\u00a0Of the patients with detectable gametocytaemia at enrolment, clearance of gametocytaemia was significantly faster with AL than with DP in Jowhar  was recorded but it was not related to the study\u00a0drug but to a car accident. The child, a girl aged 11\u00a0years, presented with an axillary temperature\u2009\u2265\u200938.2\u00a0\u00b0C and a parasite density of 44,466 asexual parasites/\u03bcL. She completed medication, and no parasites\u00a0were detected on days 2 and 3 but did not attend the day 7 visit. Otherwise, no other adverse event was observed, and the drug was well tolerated.Pfk13 gene and amplification of Pfpm2 genes. For the Pfk13 analysis, 138 samples gave interpretable results, and only one sample (0.7%) from Bosaso showed a non-synonymous mutation at codon 622 (R622I). Among the 139 samples analysed for Pfpm2 gene amplification, 135 gave interpretable results, and none showed PfPM2 amplification.Day-0 samples from the 139 patients in the AL study were tested for the presence of polymorphism in the Pfk13 non-synonymous mutations (R622I) were present at a very low rate (1/138). The impact of this mutation on the parasite resistance phenotype is currently unknown, and it has also been detected in other locations in East Africa, such as Ethiopia [The findings of the studies demonstrate that AL and DHA/PPQ are highly effective, supporting the recent recommendation of the Somalia NMCP for use of these artemisinin-based combinations as first-line and second-line treatments for uncomplicated malaria in Somalia. With respect to artemisinin resistance, none of the patients was parasitaemic on day 3 after either DHA/PPQ or AL, indicating an adequate response of the parasites to the artemisinin component. Ethiopia . AdditioPfpm2 amplification in the parasites collected from the study areas. The same analysis could not be conducted for lumefantrine resistance because of the absence of reliable molecular markers.The very high ACPR rate with both artemisinin-based combinations indicates excellent therapeutic activity of both partner drugs. This was confirmed genetically for PPQ by the absence of AL, the most commonly recommended ACT medicine in malaria-endemic countries, remains highly efficacious in Africa, despite its use as first-line treatment of uncomplicated falciparum malaria for more than a decade \u201341. DHA/The post-treatment rate of gametocytaemia clearance varied significantly between AL and DP treated groups, with DHA/PPQ being less potent, confirming previous reports on slower gametocyte clearance with DHA/PPQ treatment than with AL \u201354. The The findings of these studies document high efficacy of AL and DHA/PPQ for the treatment of uncomplicated falciparum malaria as well as lack of evidence of resistance to artemisinins and PPQ, showing the relevance of these two combinations in the chemotherapeutic strategy for malaria control in Somalia. Regular monitoring of the efficacy of these artemisinin-based combinations and of markers of resistance for artemisinin and partner drugs should be continued in order to generate evidence to inform national malaria treatment policies."}
+{"text": "The decline in the efficacy of artemisinin-based combination treatment (ACT) in some endemic regions threatens the progress towards global elimination of malaria. Molecular surveillance of drug resistance in malaria-endemic regions is vital to detect the emergence and spread of mutant strains.Plasmodium falciparum infections in Lagos, Nigeria and determined the prevalence of drug resistant strains in the population. Parasite clearance rates were determined by microscopy and the highly sensitive var gene acidic terminal sequence (varATS) polymerase chain reaction for 65 patients with samples on days 0, 1, 3, 7, 14, 21 and 28 after commencement of treatment. The genomic finger print of parasite DNA from pre- and post-treatment samples were determined using 24 nuclear single nucleotide polymorphisms (SNP) barcode for P. falciparum. Drug resistance associated alleles in chloroquine resistance transporter gene (crt-76), multidrug resistance genes (mdr1\u201386 and mdr1\u2013184), dihydropteroate synthase (dhps-540), dihydrofolate reductase (dhfr-108) and kelch domain (K-13580) were genotyped by high resolution melt analysis of polymerase chain reaction (PCR) fragments.We observed 89 malaria patients for the efficacy of artemether-lumefantrine for the treatment of uncomplicated E) values across all barcodes were 0.50\u2009\u00b1\u20090.05 and 0.56\u2009\u00b1\u20090.05 on days 0 and 28 respectively. Barcode (\u03c0) pairwise comparisons showed high genetic relatedness of day 0 and day 28 parasite isolates in three (37.5%) of the eight individuals who presented with re-appearing infections. Crt-76 mutant allele was present in 38 (58.5%) isolates. The mdr1\u201386 mutant allele was found in 56 (86.2%) isolates. No mutation in the K-13580 was observed.By varATS qPCR, 12 (18.5%) of the participants had detectable parasite DNA in their blood three days after treatment, while eight (12.3%) individuals presented with genotypable day 28 parasitaemia. Complexity of infection (CoI) was 1.30 on day 0 and 1.34 on day 28, the mean expected heterozygosity (HPersistence of DNA-detectable parasitaemia in more than 18% of cases after treatment and indications of genetic relatedness between pre- and post-treatment infections warrants further investigation of a larger population for signs of reduced ACT efficacy in Nigeria.The online version of this article (10.1186/s12879-018-3314-3) contains supplementary material, which is available to authorized users. Plasmodium falciparum infection is a significant public health problem in Nigeria where over 30% of the global burden of the disease is borne despite recent mass scale-up of intervention measures [Plasmodium falciparum remains scarce, there have been emerging cases of potentially adaptive strains and slow parasite clearance following treatment [Malaria caused by measures . Artemismeasures . The effmeasures . Howeverreatment , 5. Thesreatment . Accuratvar gene family present in the sub-telomere of the parasite [var genes [mdr1), chloroquine resistance transporter (crt), dihydrofolate reductase (dhfr), dihydropteroate synthase (dhps) and kelch-13 (K13) genes provides molecular evidence of antimalarial resistant parasites [One recent technique for the ultra-sensitive detection of low parasitaemia involves the amplification of the parasite . Each paar genes , which par genes . Prompt arasites , 11. ForPlasmodium falciparum, first by rapid diagnostic test (RDT), followed by microscopy . The recruitment criteria included fever in the preceding two days, no history of antimalarial intake in the previous four weeks and P. falciparum parasitaemia >\u20092000/\u03bcl of blood on presentation [This observational study of a 28-day follow-up was conducted in Ijede and Agbowa General Hospitals (GH) in Ikorodu, a peri-urban settlement in Lagos, South-West Nigeria, from August \u2013 November, 2016 following the revised WHO drug efficacy protocol . Ijede Gentation . Individentation . Based oentation  and the entation . An addiThe primary endpoint was clinical and parasitological response within 28\u00a0days of commencement of treatment. The secondary endpoint was prevalence of drug resistance markers and genetic relatedness of pre- and post-treatment infections. Participants failing treatment, defined as the reappearance of parasitaemia after an initial complete clearance or failure of complete clearance on day 28, were retreated with AL whether or not they developed symptoms and after samples had been obtained for microscopy and molecular analyses. Before the first treatment was administered, dried blood spots from finger pricks were collected from individuals who gave informed consent and/or assent and also met the recruitment criteria.Plasmodium falciparum DNA was extracted from pre- and post-treatment dried blood spots using the QiaAmp DNA minikit . The var gene acidic terminal sequence (varATS) quantitative PCR was used to detect multi-copy genomic sequences of low-density infections in follow-up samples [ samples . The pricrt-76), multidrug resistance (mdr1\u201386 and mdr1\u2013184), dihydropteroate synthase (dhps-540), dihydrofolate reductase (dhfr-108) and Kelch-13 (K-13580),  or re-infection by other parasite strains. A panel of twenty-four unlinked neutral SNP barcodes with minor allele frequencies (MAF) ranging between 0.2\u20130.5 and representatively spread across the parasite genome . The varATS qPCR survival estimates of parasites at different time-points were determined using Kaplan-Meier survival analysis. Association between age and parasite counts on days 0, 3 and 28 was assessed using Spearman\u2019s correlation coefficient (rs). The relationships between enrollment parasite density on day 0 and parasitaemia on days 3 and 28 were also determined using Spearman\u2019s correlation coefficient. Bonferroni correction was used to adjust multiple testing of age and parasitaemia on days 0, 3 and 28. To determine the genetic identity of each pair of samples on day 0 and day 28, we calculated pairwise barcode similarity index (\u03c0), counting ambiguous or missing calls as mismatches [E\u2009=\u2009[P represents the frequency of each allele of the SNP. HE provides an indication of the probability that two samples will be different. It has a potential range from 0  to 1 . All statistical assumptions were made at 95% confidence limit.GraphPad Prism 7  software was used for statistical analyses. Parasite clearance rates were determined by the Parasite Clearance Estimator (PCE) tool  of participants presented with parasitaemia 3\u00a0days post-treatment while eight (12.3%) patients presented with genotypable day 28 parasitaemia . There was no linear association (P\u2009=\u20090.79) between the starting parasite density and day 3 parasitaemia. Similarly, there was no correlation (P\u2009=\u20090.69) between the enrolment parasitaemia and day 28 parasite count. There was a weak linear association between parasite densities on day 3 and reappearance on day 28 (rs\u2009=\u20090.12) even though the relationship was not statistically significant (P\u2009=\u20090.33). Analysis of parasitaemia and patient characteristics (Table\u00a0P\u2009=\u20090.25). There was a weak inverse association between the age of patients and (i) persistence of parasitaemia on day 3  and (ii) re-appearance of parasitaemia on day 28 . After adjusting for multiple testing using Bonferroni method and with a new cut-off of 0.0167 for the tests between age and days 0, 3 and 28 parasitaemia, we observed an inverse relationship between parasitaemia on day 28 and age of the patient. None of the patients with positive parasitaemia post-treatment reported with clinical symptoms of malaria.The relationship between parasite persistence after treatment and the levels of parasitaemia at enrolment was examined by varATS qPCR using Spearman\u2019s correlation coefficient (rP\u2009=\u20090.42). The mean HE values across all barcodes on day 0 was 0.50\u2009\u00b1\u20090.05 and 0.56\u2009\u00b1\u20090.05 on day 28. There was no evidence of significant linkage disequilibrium  isolates. The mdr1\u201386 mutant allele was found in 56 (86.2%) of isolates. We observed 94.4% mutant and 5.6% wildtype alleles of Dhps-540 and 100% mutant alleles of Dhfr-108 in the population. No mutant allele of the K-13 C580Y was observed  and crt K76\u00a0T mutations with persistence of parasitaemia on day 3  was reported in this study, but these mutations have not been directly associated with post- treatment re-appearance of infections. In addition, contrary to a previous report showing that polymorphisms in mdr1 genes were associated with decreased sensitivity to lumefantrine [Pfcrt 76\u00a0T is associated with reduced sensitivity to chloroquine [A high frequency of mutant alleles of fantrine , our pheoroquine  and the oroquine . The preoroquine . This maP. falciparum is thought to exhibit drug-induced developmental arrest until the activity of artemisinin in the host has diminished [We propose further investigations involving complete genome-wide associations between pre-treatment and persistent parasites to circumvent the limitation of our study which did not preclude the influence of re-infection with the same parasite type or residual parasite DNA and non-replicating mature gametocytes that can persist at low concentrations after treatment. Computational analyses that involve the combination of genomic information with epidemiological simulations will present complementary tools for interpreting population-level impact of drug interventions. In addition, since minished , detailsPlasmodium falciparum with Artemether/Lumefantrine (AL) in Nigeria. The genomic barcode analysis of pre-treatment and re-appearing infections suggested recrudescence despite the absence of mutation in K-13 C580Y. This finding implies that the Nigerian parasites could be deploying a more complex drug evasion mechanism beyond the C580Y polymorphism. Further investigation of a larger population of parasites in the country is required for signs of reduced efficacy of artemisinin-based treatment of malaria.We have reported in vivo re-appearance of parasites following appropriate treatment of uncomplicated Additional file 1:Table S1. Primer sequences and qPCR conditions for varATS assay. Table S2. High Resolution Melting Drug Resistance Assay. Table Table S4. Linkage disequilibrium analysis for pre- and post-treatment parasite populations. (DOCX 26 kb)Additional file 2:Datasheet S1. Allele frequencies at each of the barcodes in the day 0 and day 28 populations. Datasheet S2. Complexity of infection in day 0 and day 28 populations. Datasheet S3. Pairwise sequence comparison of day 0 and day 28 parasite isolates. (ZIP 94 kb)"}
+{"text": "Plasmodium dihydroorotate dehydrogenase. In a previous phase 1b study, a single 150-mg dose of DSM265 showed partial efficacy against experimentally induced blood-stage Plasmodium falciparum malaria (IBSM).DSM265 is a novel antimalarial drug in clinical development that acts as a selective inhibitor of  Plasmodium dihydroorotate dehydrogenase. In a previous phase 1b study, a single 150-mg dose of DSM265 showed partial efficacy against experimentally induced blood-stage Plasmodium falciparum malaria (IBSM). Pharmacokinetic/pharmacodynamic modeling predicted a human efficacious dose of 340\u2009mg. The primary objectives of the current study were to determine the safety and efficacy of a single oral 400-mg dose of DSM265 against P. falciparum in the IBSM model. Eight healthy participants were inoculated intravenously with 2,800 parasites and treated with DSM265 7\u2009days later. Unexpectedly, one participant did not develop parasitemia during the study. All other participants developed parasitemia, with the complete clearance of asexual parasites occurring following DSM265 treatment. All seven subjects also became gametocytemic. The secondary objectives were to investigate the gametocytocidal and transmission-blocking activity of a second 400-mg dose of DSM265, which was administered 23\u2009days after inoculation. Gametocytes were not cleared by the second dose of DSM265, and transmission-blocking activity could not be determined due to low gametocyte densities. Three DSM265-related adverse events occurred, including a cutaneous rash in one subject on the day of the second DSM265 dose. The results obtained in this study support the prediction of the efficacious dose of DSM265 and provide further evidence that DSM265 is generally safe and well tolerated. In addition, this study confirms preclinical data indicating that DSM265 permits the development and maturation of gametocytes and does not clear mature circulating gametocytes. DSM265 is a novel antimalarial drug in clinical development that acts as a selective inhibitor of  Despite advances in malaria prevention and treatment, in the last 5\u2009years global progress in malaria control has stalled . MalariaPlasmodium dihydroorotate dehydrogenase (DHODH). DHODH is an enzyme that is essential in Plasmodium species for pyrimidine synthesis, as the pyrimidine salvage pathway is absent in these organisms  of approximately 5\u2009days. Using the induced blood-stage malaria (IBSM) model, a single 150-mg dose of DSM265 was effective in clearing asexual P. falciparum parasites, with recrudescent infections developing in all participants and detection of gametocytes being reported after DSM265 treatment  profile, and efficacy of a single 400-mg dose of DSM265 in a volunteer infection study where the P. falciparum-infected red blood cells on day 0. In five participants, blood-stage parasitemia was first detected by 18S ribosomal DNA (rDNA) quantitative PCR (qPCR) on day 4 postinoculation, with a further two participants becoming qPCR positive on day 5. One participant did not develop detectable blood-stage parasitemia at any time during the study. This participant retrospectively reported an undisclosed history of azithromycin usage prior to inoculation. A retrospective assay for azithromycin from a plasma sample collected on day 0 did not reveal measurable drug. In addition, the participant\u2019s blood was shown to support the growth of P. falciparum when used for in vitro parasite culture, suggesting that this participant does not have red blood polymorphisms that inhibit parasite growth. All eight participants were treated with an initial 400-mg dose of DSM265 on day 7; the participant that did not develop parasitemia was dosed in order to assess the safety and pharmacokinetics of DSM265. The seven participants who developed blood-stage parasitemia received a second 400-mg dose of DSM265 on day 23 to investigate its activity against mature gametocytes. At the end of the study (day 28), all eight participants received systematic rescue treatment with artemether-lumefantrine and primaquine.This study was conducted from 11 November 2015 to 16 December 2015 in eight healthy malaria-naive adults. The majority of participants were Caucasian (7 participants), 5 were male, and the mean age was 24.8\u2009years . All eigOverall, the study showed that DSM265 was generally safe and well tolerated. No serious adverse events (SAEs) were reported. A total of 88 adverse events (AEs) which were mostly mild in severity (87.5%) were reported , and the majority were deemed related to malaria parasite infection (77.3%). Eleven AEs were moderate in severity, while none were classified as severe. There were no abnormal laboratory or electrocardiogram (ECG) findings that were considered clinically significant. Two participants reported AEs considered to be possibly related to DSM265 treatment. Mild abdominal tenderness was reported for one participant on the day of initial dosing with DSM265; this lasted approximately 3 days, before resolving spontaneously without intervention. One other participant developed a generalized rash and pruritus on the day of the second DSM265 dose, approximately 3 h after dosing. This skin reaction was of moderate severity and resolved after approximately 12 h with cetirizine and promethazine hydrochloride use.The severity of the malaria symptoms and signs experienced by the participants during the study was generally mild, as assessed by use of a clinical scoring tool. At the time the first dose of DSM265 was administered, 4 of the 7 participants who were parasitemic had no malaria symptoms or signs , while the remaining 3 participants each had a score of 3 (Table S2). The peak malaria clinical score recorded during the study was 8; this was recorded for a single participant approximately 36\u2009h after administration of the first dose of DSM265.0\u2013\u221e) was estimated to be 1,808\u2009\u03bcg\u00b7h/ml, and the mean terminal half-life was estimated to be 140.3\u2009h , thereby permitting calculation of the parasite reduction ratio (PRR). The overall cohort-specific ratio of the parasite density at admission and the parasite density 48 h after antimalarial treatment (log10 PRR48) was 2.78 , with a corresponding parasite clearance half-life of 5.20 h .Among the seven participants who became parasitemic, the geometric mean parasitemia upon initial dosing with 400\u2009mg DSM265 was 7,851 parasites/ml . A reduction in parasitemia was observed within 48 h of DSM265 administration, with complete clearance occurring in all seven participants . No recrarasites , 16\u2014werer\u2009=\u20090.964, P\u2009=\u20090.003; Following the clearance of asexual parasitemia, male and female gametocytes were detected in all seven participants, with low but relatively stable gametocytemia beginning from approximately day 16 after inoculation  were performed on days 20 and 28, while direct membrane feeding assays (DMFA) were performed on days 20, 23, and 28. The mosquitoes fed well on the blood, with a median feeding rate of 100% for the DFA and 97.2% for the DMFA. Mosquito colony health was confirmed by demonstration of low adult mosquito mortality at the time of midgut dissection . The rate of mosquito infection was evaluated 7 to 10\u2009days after the feeding assay by assessing the presence of midgut oocysts using 18S rDNA qPCR. Oocysts were not detected in any mosquito midgut either before or after DMS265 dosing. This is likely due to the gametocyte densities at the time of feeding being below the level known to reliably result in transmission to mosquitoes.P. falciparum parasitemia  following experimentally induced blood-stage infection in healthy malaria-naive participants. Pharmacodynamic analysis indicated that DSM265 administration resulted in a modest rate of parasite clearance, with an overall cohort-specific log10 PRR48 of 2.78 and a corresponding parasite clearance half-life of 5.2 h. This was faster than the clearance rate observed when 150\u2009mg of DSM265 was administered  and similar to the activity of other long-acting drugs that have been tested in the IBSM model, such as mefloquine      and ferr, 6.5\u2009h) . Importa, 6.5\u2009h) . Further, 5.0 h) , thereby, 5.0 h) \u201314.DSM265 was generally safe and well tolerated at the 400-mg dose level, with no SAEs or severe AEs being reported and with only 3 AEs that were considered related to DSM265 treatment  being reported in 2 subjects. The onset of a rash that had some features consistent with a drug reaction (due to the fact that it occurred soon after the second dose of DSM265) raised the possibility of an IgE-mediated immunoallergic reaction. A follow-up comprehensive skin prick test study was performed in this particular subject to investigate if this reaction was due to a hypersensitivity reaction to DSM265. The results of that study will be reported separately. The overall favorable safety profile for DSM265 observed in this study agrees with the safety findings from the previous clinical studies \u201313. In t\u2013P. falciparum 3D7 parasite line, where transmission to mosquitoes was also investigated   . While ietocytes  and is cetocytes . Althoug studies . However studies , and by  studies \u201324. The studies \u2013. Enrichmstudies \u2013 could heP. falciparum blood-stage inoculum in this study did not develop parasitemia represents a highly unexpected result. It is the first time that a participant has not developed blood-stage parasitemia in this model, with 326 subjects having developed PCR patency after intravenous inoculation with this isolate. With this one failure in 326 challenges, statistical analysis indicates a 95% confidence that the true failure rate does not exceed 1.4%. Although we have not been able to definitively determine the cause of this failure, the possibility of the refractoriness of the participant\u2019s erythrocytes has been excluded. While the possibility of an error in preparation or administration of the inoculum cannot be excluded, syringes for all participants in each cohort were prepared as a single batch, and intravenous injection in this participant took place without mishap. Finally, despite the fact that the participant retrospectively reported an undisclosed history of azithromycin usage prior to inoculation, this drug could not be detected in the plasma sample collected on day 0. However, azithromycin is known to be highly cell associated, and therefore, the possibility of drug-induced inhibition of parasite growth cannot be excluded. Two previous reports also exist of failure to induce experimental malaria parasite infection due to undisclosed suspected quinine consumption  are no longer effective. It is important to note that although extensive Kelch mutations are seen in Southeast Asia, the failure to respond clinically to ACT treatment is largely linked to the failure of the partner drugs: amodiaquine, mefloquine, and piperaquine. Both artemether-lumefantrine and pyronaridine-artesunate appear to maintain activity. A second goal, in addition to providing new options to fight resistance, is to simplify therapy, and a single-encounter therapy would clearly have significant advantages in terms of directly observed therapy. It is important to underline that a molecule such as DSM265 is capable of achieving a single-dose cure at a high dose, but this does not preclude the possibility of its use in combination therapy over multiple days at lower doses.P. falciparum. The second part investigated the antimalarial activity of another investigational drug, OZ439, against P. vivax. The results of the second part will be reported in a separate publication.This was a phase 1b, open-label, IBSM clinical trial conducted at Q-Pharm Pty. Ltd.  between October 2015 and June 2016 aiming to investigate the antimalarial activity of DSM265 against Healthy men and women  aged 18 to 55\u2009years were eligible for inclusion in the study. Individuals were excluded if they had visited an area where malaria is endemic for a period of longer than 2\u2009weeks in the past 12\u2009months or had received systemic therapy with a drug with potential antimalarial activity in the past 6\u2009weeks. Full inclusion and exclusion criteria for this study are included in the supplemental material. All participants gave written informed consent before being included in the study. This study was approved by the QIMR Berghofer Medical Research Institute Human Research Ethics Committee and was registered at ClinicalTrials.gov .P. falciparum-infected human erythrocytes  as previously described  from day 4 after inoculation . The threshold was reached on day 7 in this study. DSM265 was supplied as 400\u2009mg powder in a bottle . The powder was suspended in vehicle  to form a 100-ml suspension for oral administration.Pfs25 mRNA, a transcript preferentially expressed in mature female gametocytes, and PfMGET mRNA, a transcript preferentially expressed in mature male gametocytes  targeting etocytes . Interpoetocytes . To distP-1 mRNA , 16, 20.The gametocytocidal and transmission-blocking properties of DSM265 were assessed by administering a second single oral dose of 400\u2009mg DSM265 on day 23. The timing of this second dose was based on the appearance of gametocytemia observed during the study. All participants received compulsory rescue treatment with a 3-day course of artemether-lumefantrine  and a single 45-mg dose of primaquine  at the end of the study on day 28.Safety assessments were done at screening and at protocol-specified times. Safety parameters included adverse events reporting, physical examination, vital signs, clinical laboratory evaluation, electrocardiograms (ECGs), and malaria clinical score recording. The malaria clinical score served as a clinical indication of the severity of the induced malaria parasite infection; 14 signs and symptoms commonly associated with malaria were graded using a 3-point scale , and the values were summed in order to generate an overall score (the maximum possible score is 42).Blood samples to determine the concentrations of DSM265 were taken before initial DSM265 dosing and at the following time points postdosing: 1, 2, 4, 12, 24, 48, 96, 120, 168, and 384 h (before the second dose). Plasma samples were analyzed by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) as previously described .Anopheles stephensi mosquitoes was measured using both a direct feeding assay (DFA) and a direct membrane feeding assay (DMFA) on day 20, day 23 (DMFA only), and day 28, as previously described  was washed and added to different culture flasks at 5% hematocrit and 1% parasitemia in duplicate. Two days later, parasitemia was determined and all flasks were subcultured. Two days later, parasite growth was compared between conditions.A venous blood sample was collected in lithium heparin from participant R104 at the end of the study for use in an The concentration of azithromycin was determined by hydrophilic interaction ultraperformance liquid chromatography (HILIC-UPLC) coupled with mass spectrometry . Azithromycin was extracted from serum samples using a methanol-containing labeled internal standard in a 96-well plate. A 1-\u03bcl aliquot of the supernatant was injected into the HILIC-UPLC system. Azithromycin was measured against a 7-point calibration curve. The interrun imprecision (percent coefficient of variation) across 3 levels of quality control was <8%.Cmax), the time point when Cmax was reached (tmax), the area under the concentration-time curve from 0 h up to the last time point measure (AUC0\u2013last), the area under the concentration-time curve from 0 h extrapolated to infinity (AUC0\u2013\u221e), and the elimination half-life (t1/2). The PD variables of interest in this study were the parasite reduction ratio (PRR) and the parasite clearance half-life. PRR provides an estimate of the efficacy of an antimalarial treatment and is the ratio of the parasite density between that at admission and that 48 h after antimalarial treatment . Secondary endpoints were the incidence of transmission of gametocytes to Anopheles mosquitoes and the effect of a second dose of 400\u2009mg DSM265 on gametocytemia.The primary endpoints were the safety, pharmacokinetics, and pharmacodynamics (PD) associated with a single dose of 400\u2009mg DSM265. The PK parameters determined using noncompartmental analysis were the maximum plasma concentration  was comparable to that of previous P. falciparum IBSM challenge studies and, based on previously published experience, was considered sufficient for obtaining statistically meaningful data on the effects of DSM265 on malaria parasite kinetics.The sample size of the current study (0-last) and extrapolated to infinity (AUC0\u2013\u221e). The gradient of the elimination phase was determined by log-linear regression, leading to calculation of the elimination half-life (t1/2). The maximal concentration following DSM265 administration (Cmax) and the time that it was reached (tmax) were reported. All parameters were summarized using the geometric mean or median and range.Noncompartmental PK analysis was performed using R (version 3.3.1) and RStudio (version 0.99.903) software. The area under the curve was determined using a trapezoidal method up to the last time point measure (AUCP\u2009<\u20090.001).The PRR and parasite clearance half-life were estimated using the slope of the optimal fit for the log-linear relationship of the parasitemia decay . Individpost hoc analyses were performed. To determine if the level of gametocytemia correlated with the asexual parasitemia on the day of DSM265 dosing, Spearman\u2019s rank correlation coefficient was calculated. Furthermore, in samples where both male and female gametocytes exceeded 100 gametocytes/ml, an estimation of the gametocyte sex ratios was performed.Two"}
+{"text": "Plasmodium falciparum and chloroquine\u00a0(CQ)-resistant P. vivax malaria has been reported in Vietnam. Two therapeutic efficacy studies were conducted in Thuan Bac District  in 2015 and 2016 to determine the extent of reduced artemisinin susceptibility and ACT resistant falciparum malaria, and CQ-resistant vivax malaria were present.Reduced artemisinin susceptibility and artemisinin-based combination therapy (ACT)-resistance against Twenty-seven patients with falciparum malaria were randomized to receive artesunate alone (AS\u2009~\u20094\u00a0mg/kg/day) for 4\u00a0days followed by dihydroartemisinin (DHA) (2.2\u00a0mg/kg)\u2013piperaquine (PPQ) (18\u00a0mg/kg) daily for 3\u00a0days or artemether (AM) (1.7\u00a0mg/kg)\u2013lumefantrine (LUM) (12\u00a0mg/kg) twice daily for 3\u00a0days. Sixteen subjects with vivax malaria received CQ . The therapeutic efficacy study for treating falciparum malaria was complemented with molecular analysis for artemisinin and piperaquine resistance, and in vitro drug susceptibility testing. Patient\u2019s drug exposure following both falciparum and vivax treatment studies was determined.Plasmodium falciparum and Plasmodium vivax infection on day 0 by PCR. Of the vivax patients, 15 of 16 completed CQ treatment and two had a recurrence of vivax malaria on day 28, a failure rate of 13.3% (2/15). No mutations in the Pfkelch-13 gene for artemisinin resistance or exo-E415G gene polymorphism and amplification in plasmepsins 2 and 3 for piperaquine resistance were observed. In vitro testing of patient\u2019s falciparum parasites indicated susceptibility  to dihydroartemisinin, lumefantrine, piperaquine and pyronaridine. Patient\u2019s drug exposure to artesunate and lumefantrine was comparable to published data, however, blood CQ concentrations were lower.Twenty-five of 27 patients treated with the artemisinin regimens completed the 42-day follow-up period. None had parasites present on day 3 after commencing treatment with no incidence of recrudescence (100% curative rate). One patient on AS\u2009+\u2009DHA\u2013PPQ was lost to follow-up and one patient had Clinical findings, molecular analysis and in vitro testing revealed that the falciparum parasites at Phuoc Chien Commune were artemisinin susceptible. The clinical failure rate of the 15 vivax patients who completed CQ treatment was 13%. Further studies are required to determine whether CQ-resistant vivax malaria is present at the commune.The online version of this article (10.1186/s12936-019-2640-2) contains supplementary material, which is available to authorized users. Plasmodium falciparum malaria is made more difficult by the ongoing spread of anti-malarial drug resistance to standard drugs. Since 2006, Cambodia has experienced the emergence of artesunate (AS), artesunate\u2013mefloquine (AS\u2013MQ) and dihydroartemisinin\u2013piperaquine (DHA\u2013PPQ) resistance . Other than this study, there is little published information on the efficacy of Coartem\u00ae in Vietnam, an ACT recommended by the WHO for the treatment of uncomplicated P. falciparum in young children and adults worldwide.In 2012 and 2013, DHA\u2013PPQ was still highly effective in Vietnam (>\u200998% cure rate), but by 2014 the cure rate of DHA\u2013PPQ had declined to 93% in Binh Phuoc Province . This fiP. falciparum because of drug resistance, but pernicious P. vivax malaria is also a highly debilitating disease and is a major cause of morbidity -hypoxanthine uptake by parasites [P. falciparum were run as reference strains. Drug IC50 values  were determined using a nonlinear regression analysis .Adult patients with falciparum malaria were invited to provide a blood sample before starting treatment for in vitro drug susceptibility testing of their isolates. The heparinized venous blood sample (2\u00a0mL) was centrifuged at 295\u00d7k period . For in arasites , with a arasites . The druP. falciparum field isolates that were collected from patients prior to treatment with AS + DHA\u2013PPQ\u00a0as previously described [S (artemisinin-sensitive) and MRA1240R (artemisinin-resistant) strains (Cambodia), obtained from BEI Resources , were used as reference strains.The ring-stage survival assay (RSA) and PPQ survival assay (PSA) were carried out on all escribed , 27. MRAg for 5\u00a0min and plasma was separated. For patients with vivax malaria, heparinized venous blood (0.25\u00a0mL) was collected from the participants before CQ administration (day 0) and at day 28 after commencement of the CQ treatment regimen. Both blood and plasma samples were stored in liquid nitrogen at the field site. The samples were transferred to ADFMIDI on dry-ice and stored at \u2212\u00a080\u00a0\u00b0C until drug analysis.For determining drug exposure in patients treated for falciparum malaria a blood sample (1\u00a0mL) was collected at 1\u00a0h after the last dose of AS and on day 7 after commencing AM\u2013LUM treatment. Fluoride oxalate was used as the anticoagulant for the blood sample collections to minimize further hydrolysis of AS to DHA due to plasma esterases . The bloPlasma concentrations of AS and LUM and their active metabolites (DHA and LUMm) concentrations and blood CQ and CQm concentrations were measured by liquid chromatography\u2013mass spectrometry (LC/MS/MS) at ADFMIDI for the majority of patients. The limit of quantification for AS, DHA, LUM, and LUMm, CQ and CQm were 1.19\u00a0ng/mL, 1.96\u00a0ng/mL, 2.0\u00a0ng/mL, 1.0\u00a0ng/mL, 0.5\u00a0ng/mL and 0.5\u00a0ng/mL, respectively. For quality assurance, ADFMIDI participates in the WWARN proficiency testing/QC program for the measurement of plasma concentrations of the six analytes . The chr50) was assessed with\u2009\u2265\u20093 parasite counts using the parasite clearance estimator [p\u2009<\u20090.05.All data was analysed using SigmaStat . Descriptive statistics were used to summarize baseline values and demographic data. Demographic and efficacy data were assessed by means of a per-protocol analysis, with recipients of rescue treatment counted as failures and new infections as cured. In vitro drug susceptibility and drug concentrations data are presented as median values . The parasite clearance half-life (PCstimator . For detstimator  was applstimator  and is rstimator . StatistTwenty-seven subjects were enrolled into the study and randomized to receive either AS\u2009+\u2009DHA\u2013PPQ n\u2009=\u200913) or AM\u2013LUM (n\u2009=\u200914). At enrolment, the two treatment groups had similar demographic and clinical characteristics  for patients treated with AS  or AM\u2013LUM . The PC50 was significantly shorter (p\u2009=\u20090.002 Mann\u2013Whitney U test) in the AS\u2009+\u2009DHA\u2013PPQ group  compared with the AM\u2013LUM group .Both treatment regimens rapidly reduced parasitaemia, with only one patient on AM\u2013LUM requiring 60\u00a0h to be blood film negative. AS and AM\u2013LUM rapidly reduced the parasite load with a median 97.4% (range: 75.9\u2013100%) and 71.3% (range: 6.7\u201394.1%) reduction in parasitaemia after 12\u00a0h following the first dose, respectively. The proportion of patient positive by blood film microscopy on days 1, 2, and 3 were: 69.2% (9/13) on day 1 and 0% for days 2 and 3 for the AS\u2009+\u2009DHA\u2013PPQ group and 78.6% (9/14) for day 1, 7.1% (1/14) on day 2 and 0% on day 3 for the AM\u2013LUM group. No significant difference was seen between the median parasite clearance time  in the pfkelch-13 gene in 26 of the 27 patients. Also, no exo-E415G gene polymorphism on chromosome 13 or amplification in plasmepsins 2 and 3 was observed in the P. falciparum parasites obtained from the 27 patients.Sequencing of the 850\u00a0bp of P. falciparum isolates collected from 11 adults. All isolates were successful cultured at ADFMIDI, with median IC50 (nM) values derived from at least two independent experiments  for ATQ, 51 nM (8.5 to 99.5 nM) for CQ, 11.2 nM (4.1 to 21 nM) for DAQ: 1.0 nM (0.5 to 1.9 nM) for DHA; 34.5 nM (6 to 53 nM) for LUM; 19.4 nM (4.6 to 31 nM) for MQ; 10.4 nM (4.6 to 14.5 nM) for PPQ and 3.3 nM (1.8 to 5.4 nM) for PRN. For comparison purposes, the in vitro drug susceptibility profile of the laboratory control P. falciparum drug-sensitive D6 and multidrug-resistant MRA1240 lines and the 11 participant\u2019s field isolates are shown in Additional file P. falciparum are outlined in Additional file In vitro drug susceptibility testing was carried out on P. falciparum isolates that were collected from patients treated with AS\u2013DHA\u2009+\u2009PPQ, greater than those for MRA1239S (0.16%), but markedly less than that for MRA1240R (7.3%). In the PSA, the same five isolates and both MRA lines were sensitive to PPQ, with survival rates ranging from 1.8 to 4.0% (<\u200910% cut-off). The results for the RSA and PSA are shown in Additional file The in vitro findings showed that in the RSA the survival rates ranged between 0.7 and 1.5%  for all five Patients plasma concentrations of the parent drugs (AS and LUM) and their active metabolites exhibited wide interpatient variability. The median (IQR)\u00a0plasma concentration in 12 of 13 patients was 38.9\u00a0ng/mL (26.7\u201360.6\u00a0ng/mL) for AS and 777.4\u00a0ng/mL (225.2\u20131196.0\u00a0ng/mL) for DHA at 1\u00a0h after the last dose of AS on day 4. For determining exposure to AM\u2013LUM treatment a day 7 blood sample was collected from 14 patients. The median (IQR) plasma concentration was 613.8\u00a0ng/mL (474.7\u2013803.7\u00a0ng/mL) for LUM and 14.1\u00a0ng/mL (12.4\u201316.1\u00a0ng/mL) for LUMm. The mean (SD) day 7 metabolic ratio of LUMm/LUM was 2.4\u2009\u00b1\u20090.9%.p\u2009=\u20090.10 Mann\u2013Whitney U test) for children  and adults . CQ rapidly reduced the parasite load with a median 88.8% (range: 63.8\u201397.7%) reduction in parasitaemia 12\u00a0h after the first dose and by day 2 no parasites were detected in all participants blood films. The median PC50 in the CQ group was estimated at 4.7\u00a0h; with a range of 1.8\u201321.8\u00a0h. All patients were afebrile by 36\u00a0h after starting CQ treatment. The CQ treatment regimen was well tolerated, with no serious adverse events reported. The most common adverse events reported before treatment were rigors/chills, sweating, headache, nausea, loss of appetite, fatigue and myalgia  had a mono-infection of P. vivax malaria. One patient was confirmed to have P. vivax at day 28 and another patient determined co-infected with P. falciparum/P. vivax on day 28. Thus, when applying per-protocol analysis the cure rate was 86.7% (13/15) for CQ, with a 95% confidence interval of 60.9\u201397.5% and a best estimate of 82.5%.Sixteen patients enrolled in this study completed the CQ treatment with all but one completing the 28\u00a0days of follow-up and one patient diagnosed with P. vivax infection on day 28 had CQ and CQm concentrations of 5.2\u00a0ng/mL and 7.5\u00a0ng/mL, respectively. Corresponding concentrations for the patient that was diagnosed with a mixed infection of P. falciparum and P. vivax on day 28 were 34.3\u00a0ng/mL and 31.7\u00a0ng/mL.Of the 16 participants who enrolled in the study, 15 patients provided a blood sample before CQ administration (day 0) and of these 1/15 (6%) had measurable CQ and CQm concentrations of 2.1\u00a0ng/mL and 1.2\u00a0ng/mL, respectively. By day 28 the median (IQR) was 16.7\u00a0ng/mL  for CQ and 25.8\u00a0ng/mL  for CQm. The patient that had a recurrence of Reports of DHA\u2013PPQ treatment failures in Binh Phuoc Province in south Vietnam and reduced artemisinin susceptibility in five other provinces in Vietnam, including Ninh Thuan Province, in treating uncomplicated falciparum malaria is of immense concern as other treatment options are limited. Until new drugs with different mechanisms of action to ACT are developed there is a need to evaluate other artemisinin-based combinations and to continue monitoring for reduced artemisinin susceptibility and DHA\u2013PPQ failures nationwide and in the region.pfkelch13 mutations that are associated with delayed parasite clearance in vitro and in vivo [The WHO\u2019s definition of artemisinin resistance is a delay in parasite clearance following treatment with either AS monotherapy or with an artemisinin-based combination, which represents partial/relative resistance . The del in vivo , 34, 35. in vivo .In this study, both artemisinin and ACT resistance for the treatment of falciparum malaria at Phuoc Chien Commune in southern Vietnam was assessed. To evaluate artemisinin resistance, patients were treated with AS monotherapy alone for 4\u00a0days before receiving a standard 3-day course of DHA\u2013PPQ. The reason to evaluate AS alone prior to administering DHA\u2013PPQ was to determine the effectiveness of AS in clearing falciparum parasites without a partner drug. Four days of AS treatment was considered sufficient to assess the susceptibility of AS alone as it covers two blood stage cycles of drug exposure, which historically has been sufficient to clear sensitive parasites, even with a lower AS dose regimen  .P. falciparum-infected patients with parasitaemia on day 3 after starting DHA\u2013PPQ has been reported to be 8% for Phuoc Thang Commune (Bac Ai District) and 11% for Manoi Commune (Ninh Son District). These studies were carried out by the Oxford University Clinical Research Unit/Institute of Malariology, Parasitology and Entomology (IMPE) for Phuoc Thang Commune  and WHO/IMPE for Manoi Commune .Of the 13 patients that received the AS\u2009+\u2009DHA\u2013PPQ regimen all rapidly cleared their infections with a median parasite clearance time of 36\u00a0h. Only one patient had parasites present by microscopy at 36\u00a0h but was assessed as no parasites seen by\u00a048\u00a0h after commencing AS treatment. Similar parasite clearance was also seen in the patients administered AM\u2013LUM. These clinical findings suggest a lack of reduced susceptibility of AS and AM\u2013LUM in the treated patients. Although these results are encouraging for the residents of Phuoc Chien Commune this was not the case in nearby communes . In the same province in 2015 the number of pfkelch-13 and exo-E415G genes are molecular markers of P. falciparum parasite resistance to artemisinins and piperaquine, respectively. For the pfkelch-13 gene, polymorphisms associated with amino acid changes C580Y, R539T, Y493H and I543T in the eastern Greater Mekong Subregion  and F446L, N458Y, P574L and R561H in the western GMS  have been linked to artemisinin resistance [pfkelch-13 gene were seen in the participants P. falciparum parasites on either treatment regimens. Also, no polymorphism in exo-E415G and single copies of plasmepsins 2 and 3 were observed in the participants\u2019 P. falciparum parasites.Mutations in the 50 values for P. falciparum in vitro resistance or decreased susceptibility\u00a0to CQ, DAQ and MQ are\u2009>\u2009100\u00a0nM,\u2009>\u200960\u00a0nM and\u2009>\u200930\u00a0nM, respectively [50 values for reduced susceptibility to PRN and PPQ are\u2009>\u200960\u00a0nM and\u2009>\u2009135\u00a0nM, respectively [P. falciparum isolates collected from the 11 participants revealed that they were susceptible to CQ, DAQ, PRN, PPQ, MQ, LUM and ATQ. These in vitro findings further support the clinical data, particularly with the participants\u2019 P. falciparum isolates being susceptible to the long acting partner drugs of LUM and PPQ. However, with the hypoxanthine incorporation assay there is a substantial overlap in the distribution of IC50s for DHA between rapidly and slowly cleared parasites, with the later implicated in DHA resistance [P. falciparum isolates collected from patients before treatment with AS\u2009+\u2009DHA\u2013PPQ, all were susceptible to DHA and PPQ.In vitro drug susceptibility testing has routinely been used to monitor for anti-malarial drug resistance. Using the standard hypoxanthine incorporation assay the threshold ICectively ,\u2009>\u2009110\u00a0nectively , 38 and\u2009ectively . Cut-offectively . Based osistance . The RSAsistance , 27, 41.In the evaluation of artemisinin resistance a potential confounding factor that needs to be addressed is insufficient blood drug concentrations or reduced patient drug exposure. In the present study, a 1\u00a0h post last dose of AS was selected as this is close to the maximum plasma concentration of DHA after AS dosing and the levels obtained in the participants were comparable with published values \u201345.\u00a0 PlaP. vivax malaria tends to occur on or before day 28 following a standard 3-day course of CQ  [P. vivax malaria in Vietnam with other studies [P. vivax is persistent or recurrent parasitaemia within 14\u00a0days of starting a standard treatment course of CQ [P. vivax malaria is the presence of blood CQ concentrations at the time of an unremitting or recurrent parasitaemia in excess of the minimally effective concentration established for sensitive P. vivax strains. The putative minimal blood composite CQ\u2009+\u2009CQm concentration for CQ-sensitive P. vixax malaria is 100\u00a0ng/mL at day 28 after the commencement of a standard 3-day course of CQ [Unlike falciparum malaria whereby PCR methods are used to differentiate recrudescence and reinfection, distinguishing recrudescence from reinfection or relapse is still not possible for vivax malaria . The fir5\u00a0mg/kg) . The 3\u20134 studies , 11 repose of CQ . Additiose of CQ , 50, 51.P. vivax malaria in Binh Phuoc Province, with a median CQ\u2009+\u2009CQm concentrations of 46.1\u00a0ng/mL (n\u2009=\u200951) at day 28 [The first report of CQ-resistance in Vietnam for the treatment of vivax malaria was that by Phan et al.  with a ft day 28 . By contt day 28 .The low CQ and CQm blood concentrations measured in this study and that of Thuan et al.  is of coP. vivax infection on day 28 with CQ\u2009+\u2009CQm concentrations of 12.7\u00a0ng/mL and 66.0\u00a0ng/mL. As their concentrations were well below the minimal CQ\u2009+\u2009CQm concentrations of 100\u00a0ng/mL these P. vivax infections cannot be judged as CQ-resistant, particularly the patient that had the lowest concentration (indicating poor drug absorption). However, with the other 11 patients who did not experience a recurrence of parasitaemia at Phuoc Chien Commune it appears that the P. vivax parasites at the field site are still CQ-sensitive, despite having low blood CQ exposure. Because all patients treated with CQ did not experience a recurrence of vivax malaria within 14\u00a0days of starting CQ treatment, they were likely not infected with a highly CQ-resistant P. vivax strain.In the present study, two patients had a recurrence of A limitation of the two therapeutic efficacy studies was the low number of participants which was reflected in the overall marked decline in malaria cases in Vietnam from 9331 in 2015 and to 4161 in 2016 . NonetheP. falciparum malaria at the field site in Ninh Thuan Province substantiated by molecular analysis of drug resistance, in vitro drug susceptibility testing of field isolates and drug analysis providing evidence of adequate drug exposure suggest that the falciparum parasites were susceptible to AS and ACT. Vivax malaria also appears to be susceptible to CQ at the field site but the low patient blood CQ concentrations revealed in this study needs to be further evaluated to provide definitive data on the efficacy of CQ treatment.Despite the low number of participants in the two efficacy studies the clinical findings following AS\u2009+\u2009DHA\u2013PPQ or AM\u2013LUM treatment of patients with Additional file 1: Figure S1. Map of Vietnam showing location of Ninh Thuan Province (A) and location of Phuoc Chien Commune in Thuan Bac District, Ninh Thuan Province (B).Additional file 2. Summary of drug analysis of artesunate, dihydroartemisinin, lumefantrine, desbutyl-lumefantrine, chloroquine and desethyl-chloroquine.Additional file 3: Table S1. Adverse events before and after treatment of Vietnamese patients with AS\u2009+\u2009DHA\u2013PPQ or AM\u2013LUM for uncomplicated falciparum malaria.Additional file 4: Table S1. In vitro drug susceptibility data  of the laboratory control Plasmodium falciparum D6 and MRA1240 lines and the 11 field isolates of Plasmodium falciparum collected from study participants. Table S2. Admission parasitaemia and parasite clearance time of the 11 adult patients who provided a blood sample for in vitro drug susceptibility testing . Table S3. Determination of the proportion of viable parasites  in DHA-treated compared to untreated cultures by microscopy. Table S4. Determination of the proportion of viable parasites  in PPQ-treated compared to untreated cultures by microscopy.Additional file 5: Table S1. Adverse events before and after treatment of Vietnamese patients with CQ for uncomplicated vivax malaria."}
+{"text": "Plasmodium falciparum positivity were assessed on day 3 after initiation of treatment, pre-implementation and up to 8 years post-deployment of artemether\u2013lumefantrine as first-line treatment for uncomplicated malaria in Bagamoyo district, Tanzania. Samples originated from previously reported trials conducted between 2006 and 2014. Cytochrome b-nested PCR was used to detect malaria parasites from blood samples collected on a filter paper on day 3. Chi-square and McNemar chi-squared tests, logistic regression models, and analysis of variance were used as appropriate. Primary outcome was based on the proportion of patients with day 3 PCR-determined P. falciparum positivity. Overall, 256/584 (43.8%) of screened patients had day 3 PCR-determined positivity, whereas only 2/584 (0.3%) had microscopy-determined asexual parasitemia. Day 3 PCR-determined positivity increased from 28.0% (14/50) in 2006 to 74.2% (132/178) in 2007\u20132008 and declined, thereafter, to 36.0% (50/139) in 2012\u20132013 and 27.6% (60/217) in 2014. When data were pooled, pretreatment microscopy-determined asexual parasitemia \u2265 100,000/\u00b5L, hemoglobin < 10 g/dL, age < 5 years, temperature \u2265 37.5\u00b0C, and year of study 2007\u20132008 and 2012\u20132013 were significantly associated with PCR-determined positivity on day 3. Significant increases in P. falciparum multidrug resistance gene 1 N86 and P. falciparum chloroquine resistant transporter K76 across years were not associated with PCR-determined positivity on day 3. No statistically significant association was observed between day 3 PCR-determined positivity and PCR-adjusted recrudescence. Day 3 PCR-determined P. falciparum positivity remained common in patients treated before and after implementation of artemether\u2013lumefantrine in Bagamoyo district, Tanzania. However, its presence was associated with pretreatment characteristics. Trials registration numbers: NCT00336375, ISRCTN69189899, NCT01998295, and NCT02090036.Prevalence of and risk factors associated with polymerase chain reaction (PCR)-determined  Plasmodium falciparum malaria globally.1 Artemisinin-based combination therapy consists of fast-acting artemisinin derivatives and a long-acting partner drug with a different mode of action. Initially, artemisinin rapidly clears a large number of asexual parasites before the long-acting partner drug eliminates the residual parasitemia.2 For artemisinin-sensitive P. falciparum parasites, microscopy-determined parasite clearance usually occurs within 48 hours in most of the patients treated with ACT.3 Artemisinin-resistant P. falciparum parasites, on the other hand, are phenotypically characterized by a slow clearance rate.4 Microscopy-based P. falciparum positivity rate on day 3 after initiation of ACT treatment is considered an important determinant of delayed parasite clearance, and if the positivity rate exceeds 10%, this is considered an alert for artemisinin resistance.5 The resistance has been linked to polymorphisms in the Kelch gene located on chromosome 13 (k13) of P. falciparum parasites.6Artemisinin-based combination therapy (ACT) is the mainstay treatment for uncomplicated 8 In Africa, despite recent case reports of suspected resistance,10Pfk13 polymorphisms are rare and do not include those previously described and associated with delayed P. falciparum clearance in Southeast Asia.12 Artemisinin-based combination therapy has also remained highly efficacious in Africa, with rapid microscopy-determined clearance of the asexual parasites within 48 hours after initiation of treatment.15 However, few African studies have assessed the presence of residual polymerase chain reaction (PCR)-determined P. falciparum positivity on day 3 after initiation of ACT treatment,17 a phenomenon that may indicate reduced susceptibility of P. falciparum against artemisinin derivatives and thereby a potential early warning marker of artemisinin resistance.16Artemisinin resistance has been reported from four countries in the Mekong region, Southeast Asia.20 Higher initial microscopy-determined asexual parasitemia  and anemia, that is hemoglobin < 10 g/dL, are associated with slow parasite clearance.21 Conversely, in high malaria-endemic areas, malaria immunity increases with age, whereby children aged < 5 years have low acquired immunity against P. falciparum infection and are, therefore, prone to slower parasite clearance than older individuals.18 However, it is not well understood whether these factors are responsible for the previously observed day 3 PCR-determined P. falciparum positivity in Africa, rather than reduced parasite susceptibility against artemisinins.17Nonetheless, several pretreatment host and parasite characteristics including initial parasite density, age of the patient, and hemoglobin level influence parasite clearance after treatment with ACT.P. falciparum positivity on day 3 after artemether\u2013lumefantrine treatment initiation in clinical trials conducted between 2006 and 2014, covering pre-implementation and 8 years post-implementation of artemether\u2013lumefantrine as first-line treatment for uncomplicated P. falciparum malaria in Bagamoyo district, Tanzania.Therefore, the aim of this study was to evaluate the prevalence of, and risk factors associated with, PCR-determined 24 The district has a tropical climatic condition, with annual rainfall averaging 1,100 mm. Malaria transmission is high and stable throughout the year, with a slight variation between rainfall and dry season.25Plasmodium falciparum is the predominant malaria species and Anopheles gambiae sensu stricto is the main vector.26 Artemether\u2013lumefantrine was adopted in Tanzania as a first-line treatment for uncomplicated P. falciparum malaria in November 2006. The use of long-lasting insecticide-treated mosquito nets is the major vector control method in the study area.27The blood samples used for PCR analyses in this report originated from four clinical trials conducted in Fukayosi and Yombo dispensaries in Bagamoyo district, Tanzania, between 2006 and 2014.P. falciparum positivity on day 3 after initiation of artemether\u2013lumefantrine treatment among patients with acute uncomplicated P. falciparum malaria before and after the implementation of artemether\u2013lumefantrine treatment policy. Four clinical trials were included in the analysis: 1) an artemether\u2013lumefantrine pharmacokinetics and pharmacodynamics trial, 2006,22 2) the supervised arm of an artemether\u2013lumefantrine efficacy (supervised) versus effectiveness (unsupervised) trial, 2007\u20132008,23 3) an artemether\u2013lumefantrine efficacy trial, 2012\u20132013,15 and 4) both arms of an artemether\u2013lumefantrine versus artemether\u2013lumefantrine plus 0.25 mg/kg single dose of primaquine efficacy and safety trial, 2014.24 The primaquine arm was included in the analysis because no statistically significant differences in microscopy-determined parasite clearance, prevalence of PCR-determined P. falciparum positivity on day 3, prevalence of P. falciparum multidrug resistance gene 1 (Pfmdr1) N86 and P. falciparum chloroquine resistant transporter (Pfcrt) K76, and crude and PCR-adjusted cure rates were observed between the artemether\u2013lumefantrine and artemether\u2013lumefantrine plus primaquine arms.28This was a retrospective study designed to assess the prevalence of, and risk factors associated with, PCR-determined The 2006 trial was conducted in June, few months before deployment of artemether\u2013lumefantrine in Bagamoyo district, whereas the 2007\u20132008, 2012\u20132013, and 2014 trials were conducted 1\u20132, 6\u20137, and 8 years, respectively, after wide scale use of artemether\u2013lumefantrine as treatment policy in the study area.P. falciparum malaria attending the dispensaries as previously described.24 Patients with microscopically confirmed P. falciparum mono-infection and who fulfilled all the inclusion criteria and none of the exclusion criteria were enrolled and treated under supervision with standard weight-based six-dose artemether\u2013lumefantrine regimen according to the national treatment guidelines in Tanzania. For patients allocated with artemether\u2013lumefantrine plus primaquine treatment, an additional single 0.25 mg/kg bodyweight primaquine dose was administered concomitantly with the first artemether\u2013lumefantrine dose.24 Patients were observed for 30 minutes after the intake of each drug dose; and treatment was readministered in case of vomiting within this period. Patients with fever greater than 38.5\u00b0C were treated with paracetamol.The study subjects were recruited from patients with symptoms and signs compatible with acute uncomplicated 29 Patients with repetitive vomiting of the trial drug were managed according to national guidelines and followed up until recovery.27Follow-up for clinical and laboratory assessments was performed on days 0, 1, 2, and 3, for the 2006 trial; 0, 1, 2, 3, 7, 14, 21, and 28 or any day of recurrent illness, for the 2014 trial; 0, 1, 2, 3, 7, 14, 21, 28, and 42, for the 2012\u20132013 trial; and 0, 1, 2, 3, 7, 14, 21, 28, 35, 42, 49, and 56 for the 2007\u20132008 trial. Every clinical assessment included the history of clinical symptoms, possible adverse events, concomitant drug consumption, clinical examination, and measurement of axillary temperature. Patients who missed a scheduled follow-up visit and who could not be found despite efforts to trace them at their homes were considered lost to follow-up and were consequently withdrawn.P. falciparum, and parasite density thin films for the species identification for the 2012\u20132013 and 2014 cohorts, and 100 \u00b5L of blood on a filter paper  for molecular genotyping of asexual parasites.Laboratory assessment involved collection of a finger-prick blood sample from patients in all four studies for hemoglobin quantification, thick blood films for the microscopical determination of the presence of 24 The blood films were stained using 10% giemsa and read by qualified microscopists, as previously described.24 The filter papers were labeled, air-dried at room temperature for 3\u20134 hours, packed in an individual plastic bag, and transported to Karolinska Institutet, Sweden, for molecular analysis.Hemoglobin concentration was measured using a portable spectrophotometer HemoCue Hb 201+ , as previously described.P. falciparum infection during follow-up using ABI PRISM\u00ae 6100 Nucleic Acid Prep Station  for 2006, and on day 0 and the day of recurrent infection for 2007\u20132008 samples, whereas, a 10% chelex-100  method was used for the day 3 samples for 2007\u20132008 trial, and days 0, 3, and the day of recurrent infection samples for the 2012\u20132013 and 2014 trials.30 The detection limit of the ABI PRISM 6100 extraction method has been estimated to be 200 parasites/\u03bcL and that of the chelex-100 method to be 2 parasites/\u03bcL.31Genomic DNA was extracted from dried blood spots collected on a filter paper on days 0, 3, and, when applicable, on the day of microscopy-determined recurrent P. falciparum positivity using cytochrome-b nested PCR, as previously described.32 Briefly, 5 \u03bcL of the extracted DNA was amplified with 0.1 \u03bcL of 100 pmol/\u03bcL of each primer, 2.5 \u03bcL of 2 mM dNTPs , 2 \u03bcL of 25 mM MgCl2, and 0.125 \u03bcL of 5 U/\u03bcL Taq DNA polymerase (Sigma Aldrich) and 10.175 \u03bcL of dH2O, using the following cycling program: 3 minutes at 95\u00b0C, then 40 cycles of 15 seconds at 95\u00b0C, 60 seconds at 60\u00b0C, 70 seconds at 72\u00b0C, and a final extension of 5 minutes at 72\u00b0C. For the nested-PCR, 3 \u03bcL of primary PCR product was amplified with 0.05 \u03bcL of 100 pmol/\u03bcL of each primer, 2.5 \u03bcL of 2 mM deoxynucleoside triphosphates (dNTPs) 2 \u03bcL of 25 mM MgCl2, and 0.125 \u03bcL of 5 U/\u03bcL Taq DNA polymerase and 12.175 \u03bcL of dH2O, using the following cycling program: 3 minutes at 95\u00b0C, then 40 cycles of 15 seconds at 95\u00b0C, 60 seconds at 60\u00b0C, 60 seconds at 72\u00b0C, and a final extension of 5 minutes at 72\u00b0C. Pfmdr1 N86Y and Pfcrt K76T were genotyped using nested PCR followed by restriction fragment length polymorphism using ApoI  restriction enzyme.33Genomic DNA extracted from filter papers collected on day 3 after artemether\u2013lumefantrine treatment initiation from all four studies was used to detect PCR-determined P. falciparum block 3 of merozoite surface proteins (msp) 2, block 2 of msp 1, and region II (RII) of glutamate-rich protein (glurp).34Blood samples from patients with recurrent infection during follow-up  were further genotyped to differentiate recrudescence (treatment failure) from reinfection (new infection) by stepwise genotyping of \u2122  stained agarose, separated by electrophoresis, and visualized under Ultra Violet transillumination  using Image Lab\u2122 software (Bio-Rad). For merozoite surface protein (msp) 1 1, 2, and glurp, alleles in each family were considered the same if fragments\u2019 size was within 20 base pair intervals. Patients with microscopy-determined recurrent P. falciparum infection, but with negative PCR results, were considered to have uncertain PCR-adjusted outcome and were consequently excluded from the final analysis. Recrudescence was defined as the presence of at least one matching allelic band, and reinfection was defined as the absence of any matching allelic band at baseline and on the day of parasite recurrence.34The amplicons were loaded, respectively, on GelRedP. falciparum positivity on day 3 after treatment initiation with artemether\u2013lumefantrine between 2006 and 2014. Polymerase chain reaction\u2013determined P. falciparum positivity was predefined as parasites that were detectable by cytochrome b-nested PCR. The secondary outcomes were as follows: prevalence of day 3 PCR-determined P. falciparum positivity across years between 2006 and 2014, the proportion of Pfmdr1 N86Y and Pfcrt K76T on days 0 and 3, and their association with the presence of day 3 PCR-determined P. falciparum positivity across years; change in the proportion of Pfmdr1 N86Y and Pfcrt K76T between days 0 and 3 across years; the association between baseline characteristics including age < 5 years, microscopy-determined pretreatment asexual parasitemia , fever (\u2265 37.5\u00b0C), anemia (hemoglobin < 10 g/dL), Pfmdr1 N86 and Pfcrt K76, and PCR-determined P. falciparum positivity on day 3; and the association between PCR-determined P. falciparum positivity on day 3 and crude recurrent infection, as well as PCR-adjusted recrudescence during follow-up after artemether\u2013lumefantrine treatment.The primary outcome was the proportion of patients with PCR-determined Good Clinical Practice, Declaration of Helsinki, and applicable regulatory requirements in Tanzania were adhered to during the conduct of the trials. Approvals to conduct the respective trials were obtained from the Ethics Committees of the Muhimbili University of Health and Allied Sciences, National Institute for Medical Research, Tanzania Food and Drug Authority, and the Regional Ethics Committee, Stockholm, Sweden. Written informed consent was obtained from all patients and parents/guardians of patients aged < 18 years, before enrollment. The trials are registered with numbers: NCT00336375, ISRCTN69189899, NCT01998295, and NCT02090036.P. falciparum positivity, Pfmdr1 N86Y, and Pfcrt K76T across years was assessed using logistic regression model. Univariate and multivariate logistic regression models were used to assess the association of pretreatment risk factors with both residual parasites and recurrent infection. The association between pretreatment characteristics and day 3 PCR-determined P. falciparum positivity was assessed both for each trial individually and when data from all trials were pooled. The 2006 trial had a follow-up of 3 days only, and hence excluded in the analysis to assess the association of day 3 PCR-determined P. falciparum positivity and recurrent infection, including both crude microscopy-based recurrent infection and PCR-adjusted recrudescence, during follow-up after artemether\u2013lumefantrine treatment. Mean differences between groups were compared using the analysis of variance. Data were censored at the time of withdrawal in case of patient lost to follow-up, withdrawal of consent, and PCR-adjusted reinfection or uncertain PCR outcome. A statistical significance level was set at P \u2264 0.05.Data were double entered and analyzed using R version 3.2.3 . Independent proportions were compared using chi-squared test, whereas paired proportions were compared using McNemar chi-squared test. Age was negatively skewed and was, therefore, transformed using log. Trend of change in proportion of day 3 PCR-determined P. falciparum positivity on day 3, in which, 256 (43.8%) were PCR positive. Of the PCR-positive patients, only two (0.3%) had microscopy-determined asexual parasites on day 3. Pretreatment characteristics of the study participants are presented in A total of 584 patients were screened for PCR-determined P. falciparum positivity on day 3 increased from 28.0% (14/50) in 2006 to 74.2% (132/178) in 2007\u20132008 and declined, thereafter, to 27.6% (60/217) in 2014, P. falciparum positivity on day 3 by study year in patients aged < 5 years  in 2006 to 57.6% (19/33) in 2007\u20132008, 72.9% (27/37) in 2012\u20132013, and 71.4% (35/49) in 2014 , whereas that of Pfcrt K76 increased significantly from 54.5% (6/11) in 2006 to 70.9% (66/93) in 2007\u20132008, 87.5% (28/32) in 2012\u20132013, and 91.1% (51/56) in 2014 .A total of 131/256 (51.2%) and 192/256 (75.0%) samples were PCR positive on day 3 for Pfmdr1 N86 to 86Y or N86Y, Pfmdr1 86Y to N86 or N86Y, and from Pfmdr1 N86Y to N86 or 86Y  between pretreatment and day 3 across years of study. Similarly, the change of Pfcrt K76 to 76T or K76T, Pfcrt 76T to K76 or K76T, and from Pfcrt K76T to 76K or 76T between pretreatment and day 3 were not significantly different .There was no significant difference in the change of P. falciparum positivity are presented in P. falciparum positivity on day 3 with the OR of 6-folds (P < 0.001) and 3-folds (P = 0.017), respectively, as compared with data from 2006. Other predictors for PCR-determined P. falciparum positivity were microscopy-determined asexual parasite density \u2265 100,000/\u00b5L (P < 0.001), hemoglobin < 10 g/dL (P = 0.025), age < 5 years (P < 0.001), and fever (axillary temperature \u2265 37.5\u00b0C) at inclusion (P = 0.043). Both Pfmdr1 N86 (P = 0.71) and Pfcrt K76 (P = 0.72) were not significantly associated with PCR-determined P. falciparum positivity on day 3.Results for logistic regression of univariate and multivariate analyses for association between pretreatment characteristics and day 3 PCR-determined P. falciparum positivity on day 3 were included in this analysis. Twenty-seven of 534 (5.1%) were either lost to follow-up or withdrew their consent between days 3 and 28. Of the remaining 507 patients, 44 (8.7%) had microscopy-determined recurrent infection, of whom seven (1.4%) had PCR-adjusted recrudescence. Of the 507 patients, 227 (44.8%) had PCR-determined P. falciparum positivity on day 3, of which 24 (10.6%) had microscopy-determined recurrent infection  and four (0.018%) had PCR-adjusted recrudescence .A total of 534 patients screened for PCR-determined PfK13 polymorphisms associated with artemisinin resistance, showed that more than one-third of the patients tested positive for P. falciparum by PCR on day 3, whereas only two patients had day 3 microscopy-determined asexual parasites. Importantly, the prevalence of day 3 PCR-determined P. falciparum positivity across study years in patients aged < 5 years was slightly higher but not statistically significantly different from that of all ages and of those aged > 5 years. Parasite clearance following treatment is determined by many factors including the mode of action of the used antimalarial drug and pretreatment characteristics of both the parasite and host.36 The day 3 PCR-determined P. falciparum positivity observed in this study might have also been present during the chloroquine (CQ) and sulfadoxine\u2013pyrimethamine era because of their modes of actions, resulting in a relatively slow parasite clearance rate as compared with artemisinin.36 In the present study, the day 3 PCR-determined P. falciparum positivity might represent a remaining fraction of the parasite biomass that was then probably cleared by lumefantrine, the long-acting partner drug.37 Our observations are substantiated by the presence of a significant proportion of patients in the 2006 trial with PCR-determined P. falciparum positivity on day 3, despite being treated for uncomplicated P. falciparum malaria 6 months before the implementation of artemether\u2013lumefantrine policy in Bagamoyo district. However, a study in Kenya found an association of the presence of PCR-determined P. falciparum parasites on day 3 with parasite tolerance against ACT.16 Nevertheless, differences in parasite detection methods between the studies with the use of quantitative-PCR (qPCR) in the Kenyan study might explain the differences observed between these two studies.16 Further surveillance of PCR-determined P. falciparum positivity on day 3 to provide a better understanding of this phenomenon is, therefore, warranted. On the other hand, the proportion of PCR-determined P. falciparum positivity on day 3 increased between 2006 and 2007\u20132008, and, thereafter, declined toward 2014. Differences in methods used may have resulted in different findings between the individual trials in this report. However, with the exception of a different DNA extraction method used for the 2006 trial, the amount of blood collected, type and storage of a blotted filter paper, and amplification methods were the same for all the included trials. Therefore, it is not well understood why the 2007\u20132008 trial had a high proportion of day 3 PCR-determined P. falciparum positivity as compared with other trials included in the analysis.This study conducted in Bagamoyo district, Tanzania, where presently no data exist on presence of P. falciparum positivity on day 3. However, when the respective trials were analyzed separately, the same factors were significantly associated with PCR positivity on day 3 in some trials, but not in others. Contrary to our findings, a study in Kenya did not observe any association of pretreatment characteristics with qPCR-determined parasite positivity on day 3.16 However, previous studies have shown pretreatment characteristics of both the host and parasite, including existing immunity, pretreatment parasitemia, parasite strains, parasite developmental stage, and parasite drug susceptibility, to affect microscopy-determined parasite clearance time.39After data were pooled, higher initial microscopy-determined asexual parasitemia , hemoglobin < 10 g/dL, age < 5 years, body temperature \u2265 37.5\u00b0C, and the year of study were found to be associated with the presence of PCR-determined Pfmdr1 N86 and Pfcrt K76 proportions increased significantly across years, in line with previous reports.41 However, the presence of Pfmdr1 N86 and Pfcrt K76 alleles in this study was not associated with a statistically significant increase in the presence of PCR-determined P. falciparum positivity on day 3. Furthermore, the changes between days 0 and 3 of Pfmdr1 N86Y and Pfcrt K76T outcome were not significantly different across years. The absence of significant change of these alleles, particularly of Pfmdr1 N86 and Pfcrt K76, between days 0 and 3 probably indicates that more time than 3 days is required for the parasite to be exposed to the drug for such selection to occur. Conversely, the relatively high proportions of Pfmdr1 86Y and Pfcrt 76T in 2006 probably reflect previous exposure of the parasite population to amodiaquine and CQ.44P. falciparum positivity on day 3 was neither associated with the microscopy-determined recurrent infections nor the PCR-adjusted recrudescence during follow-up after artemether\u2013lumefantrine treatment. Furthermore, a large majority (83.3%) of the recurrent infections were due to reinfection based on PCR-adjustment. Similar findings have been reported in Uganda.45 A study in Kenya, however, reported the association of PCR-determined P. falciparum positivity with PCR-adjusted recrudescence.16 Nonetheless, differences in the methods used to determine parasites on day 3 and differentiate recrudescence from reinfection could probably account for the differences in the findings between the two studies.29 Unlike our study that determined day 3 P. falciparum positivity using conventional PCR, the study in Kenya used qPCR to determine both P. falciparum positivity and density.16 The qPCR is highly sensitive compared with the conventional PCR. The high qPCR-determined median parasite density in the Kenyan study, therefore, probably led to the observed association of day 3 parasitemia with PCR-adjusted recrudescence. Furthermore, in the Kenyan study, the standard WHO criteria to differentiate PCR-adjusted recrudescence from the new infection was modified, whereby samples obtained 1 day before and on the day of recurrent infection were compared with those of days 1 and 2 in addition to the baseline samples.29The PCR-determined P. falciparum parasites were cleared during follow-up between days 4 and 7. Moreover, neither was any evaluation of in vitro responsiveness to artemether\u2013lumefantrine carried out nor assessment of day 7 lumefantrine concentration levels.The strength of this study is that it included four studies conducted over different years, before and up to 8 years after implementation of artemether\u2013lumefantrine as first-line treatment of uncomplicated malaria in Bagamoyo district. However, study limitations include, for example, that no blood sampling was performed to assess whether the PCR-determined P. falciparum positivity remained common in patients treated with artemether\u2013lumefantrine before and after implementation of ACT as policy in Bagamoyo district, Tanzania. However, its presence was associated with pretreatment characteristics including high pretreatment asexual parasitemia , anemia (hemoglobin < 10g/dL), age < 5 years, documented fever at enrollment, and the year of study.Day 3 PCR-determined"}
+{"text": "Most epidemiological studies on the interplay between iron deficiency and malaria risk classify individuals as iron-deficient or iron-replete based on inflammation-dependent iron markers and adjustment for inflammation by using C-reactive protein (CRP) or \u03b1-1-acid glycoprotein (AGP). The validity of this approach and the usefulness of fibroblast growth factor 23 (FGF23) as a proposed inflammation-independent iron marker were tested.Conventional iron markers and FGF23 were measured in children with acute falciparum malaria and after 1, 2, 4, and 6\u00a0weeks. Children, who were transfused or received iron supplementation in the follow-up period, were excluded, and iron stores were considered to be stable throughout. Ferritin levels 6\u00a0weeks after admission were used as a reference for admission iron status and compared with iron markers at different time points.There were long-term perturbations in iron markers during convalescence from acute malaria. None of the tested iron parameters, including FGF23, were independent of inflammation. CRP and AGP normalized faster than ferritin after malaria episodes.Malaria may bias epidemiological studies based on inflammation-dependent iron markers. Better markers of iron status during and after inflammation are needed in order to test strategies for iron supplementation in populations at risk of malaria. The geographical distributions of iron deficiency and malaria overlap. Iron deficiency has a negative impact on child development \u20134, but mCorrespondingly, iron supplementation has been associated with increased risk of malaria, e.g., in the widely cited Pemba trial, which showed that routine iron supplementation resulted in increased morbidity and mortality among participating children . AlthougConventionally, serum ferritin has been used as the key iron marker as it reflects total body iron storage. However, ferritin is also an acute phase protein , so in aUsing this reference point, the possibility that other iron markers during acute malaria might be correlated with ferritin after normalization was studied. The main focus was on hepcidin and fibroblast growth factor 23 (FGF23). Hepcidin is a peptide hormone that is regulated by iron stores, inflammation and erythropoietic demand and controls iron efflux from most cells in the body, in particular enterocytes and macrophages. Hepcidin has been suggested as the best marker to guide iron treatment as it has been proposed to be a key determinant of iron utilization , 18. YetEwe, of the goals, benefits and risks of taking part in the study, and written consent was obtained prior to enrolment.The study was approved by the Noguchi Memorial Institute for Medical Research Institutional Review Board (NMIMR STC Number: STC Paper 5(1) 2013\u20132014) and by the Ethical Review Committee of the Ghana Health Service (file GHS-ERC 08/05/14). Parents/guardians of all study participants were informed in their local language, The study was conducted in Hohoe, a town located about 220\u00a0km northeast of Accra, in an area of tropical, semi-deciduous, forest vegetation and used a convenience sample of children included in the MAVARECA study  . MalariaPlasmodium falciparum rapid diagnostic test (RDT), microscopic finding of peripheral parasitaemia >\u20092500 infected erythrocytes (IEs)/\u00b5L, fever (\u2265\u200937.5\u00a0\u00b0C) within the first 24\u00a0h of admission or a history of fever in the preceding 24\u00a0h. Subjects were excluded if they had severe co-morbidity, including sickle-cell disease, or received a blood transfusion during admission due to severe anaemia (haemoglobin (Hb)\u2009<\u20095\u00a0g/dL) as the iron content of the transfused erythrocytes would otherwise distort the interpretation of the analysis of iron marker kinetics during follow up. During the\u00a0follow-up period, patients were excluded if they presented with malaria, other severe disease or new spikes in inflammatory markers (C-reactive protein (CRP)\u2009>\u20095\u00a0mg/L or \u03b1-1-acid glycoprotein (AGP)\u2009>\u20091\u00a0g/L), or had taken iron supplementation (only excluded after such an event). The reason for excluding these patients was that these events would have affected day-42 results and thus the final endpoint.Inclusion criteria were age 1\u201312\u00a0years, positive Upon enrolment, a project nurse and physician completed a standardized questionnaire and performed a clinical examination. Severe malaria was defined according to WHO criteria . UncomplThe patients were followed for 6\u00a0weeks and attended the research clinic 14 and 42\u00a0days post-admission (regular follow-up group). A sub-set of patients living near the hospital were also asked to come 7 and 28\u00a0days post-admission (frequent follow-up group). At follow-up visits, the patients or their parents were systematically interviewed about new symptoms and medicine use, including iron supplements, during the follow-up period. A missed follow-up appointment did not exclude the patient from future follow-up visits. Patients were encouraged to attend the research clinic at any time during the follow-up period if they developed any new symptoms.For the purpose of this study, inflammation was defined as either CRP\u2009>\u20095\u00a0mg/L or AGP\u2009>\u20091\u00a0g/L, while iron deficiency was defined as ferritin concentrations <\u200915\u00a0\u00b5g/L on day 42.\u00ae CM Weak Cation Exchange (WCX) beads  and ammonium acetate at pH 7.5. After washing, the analyte was eluted with 25\u00a0\u00b5L 2.5% Trifluoroacetic acid, 50% Acetonitrile LC\u2013MS Chromasolv solution (both from Sigma-Aldrich) and the WCX beads were separated from supernatant by centrifugation at 500g. Peptide spectra were then generated on a Microflex matrix-enhanced laser desorption/ionization (MALDI) TOF\u2013MS platform . Levels of FGF23  were measured in duplicates using a 2nd generation, C-terminal, two-site ELISA . Biochemical parameters (reference ranges), including AGP (\u2264\u00a01\u00a0g/L), bilirubin (4\u201322\u00a0\u00b5mol/L), high sensitivity CRP\u00a0(\u2264\u2009\u00a05\u00a0mg/L), fe (5\u201330\u00a0\u00b5mol/L), ferritin (15\u2013140\u00a0\u00b5g/L), haptoglobin (0.3\u20131.8\u00a0g/L), lactate dehydrogenase , transferrin (18.7\u201350\u00a0\u00b5mol/L), and transferrin saturation (20\u201350%) were analysed on a Cobas 8000, . Levels of soluble transferrin receptor  were measured (2014 samples only) in duplicates by BNII nephelometry . Sickle cell Hb phenotype was determined by electrophoresis, while glucose-6-phosphate dehydrogenase (G6PD) deficiency was determined by methylene blue reduction test  was mixed with Macro-Prepion test   was collected at enrolment and tested for signs of microscopic haematuria and urinary tract infections using Siemens MultistixCryptosporidium, Giardia lamblia and Entamoeba histolytica), while at least 2\u00a0mL were mixed with formalin (10%) and stored at room temperature for later formalin-ether concentration (a.m. Ridley) and microscopy for cysts, helminth larvae and eggs.Stool samples (>\u20094\u00a0mL) were collected during admission. At least 2\u00a0mL were stored frozen . All continuous variables are presented as geometric means\u2009\u00b1\u20092 standard deviations (SD), unless otherwise specified. Longitudinal data were analysed using log-transformed variables in a mixed effects model. P values <\u20090.05 were considered statistically significant. As some CRP values were 0, i.e., below the lower detection level of 1, \u201c1\u201d was added to the CRP value before log transformation. Pearson\u2019s correlation coefficient (R) was used to determine correlation between log-transformed iron markers.Only data collected \u00b1\u20092\u00a0days from the scheduled follow-up date were included, except for day\u00a042, where delays in data collection were tolerated .A total of 156 patients with a positive RDT were recruited to participate in the study. Three patients did not meet the fever criteria, one was excluded due to sickle cell disease, 40 had \u2264\u20092500\u00a0IEs/\u00b5L on presentation, and 14 patients received a blood transfusion during admission on clinical indication or because their Hb levels decreased to <\u20095\u00a0g/dL. The analysis is restricted to the 98 remaining patients  or moderately (Hb 5\u20138\u00a0g/dL) anaemic. (Patients with severe anaemia (Hb\u2009<\u20095\u00a0g/dL) were excluded). Hb levels decreased until day\u00a07 post-admission despite treatment, but had returned to normal levels by day 28 Fig.\u00a0a and lacFerritin levels were high on day 0 and gradually decreased during follow-up, including a statistically significant drop between day 28 and day 42  were iron-deficient (ferritin concentrations below <\u200915\u00a0\u00b5g/L on day 42). The acute phase proteins CRP ; MCH: R\u2009=\u20090.30 (P\u2009=\u20090.03), MCHC: R\u2009=\u20090.10 (P\u2009=\u20090.49)]. Furthermore, the slightly increased MCV values day 7 to day 28 , hepcidin levels were raised Fig.\u00a0a, which In the 2014 pilot study, sTfR, an indicator of erythropoietic intensity, was also measured on days 0, 14 and 42. Day 0-levels of sTfR levels were below the day 42 levels (P\u2009=\u20090.0004), whereas day\u00a014 levels were above day 42 levels (P\u2009=\u20090.03), consistent with post-malarial erythropoiesis. sTfR/log ferritin ratios were not associated with ferritin levels on day 42, why this parameter was omitted in the 2015 study.On admission, patients with severe malaria had lower Hb , lower haptoglobin , higher LDH [639 vs 445 U/L (P\u2009<\u20090.0001)], and higher bilirubin [26 vs 19\u00a0\u00b5mol/L (P\u2009=\u20090.005)], compared to patients with uncomplicated malaria, consistent with more pronounced haemolysis in the severe malaria group. Also, admission levels of ferritin and FGF23 were higher in patients with severe malaria , likely due to inflammation. By day 7, these differences were no longer detectable, except differences in haemoglobin levels that did not resolve until day 28.In this observational, longitudinal study, conventional iron markers as well as FGF23 were measured in children on admission with acute falciparum malaria and after 1, 2, 4 and 6\u00a0weeks. The most striking finding was the duration of iron marker perturbations following episodes of falciparum malaria. Ferritin was elevated for more than 4\u00a0weeks after the infection in children without evidence of recrudescence or re-infection , transferrin showed the best correlation with ferritin on day 42 [R\u2009=\u2009\u2212\u20090.45 (P\u2009=\u20090.001)]. However, transferrin was also markedly affected by inflammation, and the correlation between transferrin day 42 levels with plasma ferritin levels on day 42 was modest. In the pilot study, sTfR levels were also evaluated. This biomarker is less affected by inflammation than ferritin but increases in haemolytic anaemia , 38. LevThe possibility that artemisinin-associated haemolysis might have enhanced perturbations of markers of erythropoiesis in the follow-up period was also considered. This condition is associated with hyperparasitaemia at the initiation of treatment . In the The lack of an inflammation-independent marker of iron deficiency may restrict the possibility to develop innovative strategies for iron supplementation in malaria-endemic areas. Experimental studies suggest that the increased susceptibility to malaria in the course of iron supplementation is temporary . In ordeThe observed temporal changes in hepcidin levels are in line with previous studies . During Better biomarkers for iron stores in acute malaria are still needed to improve the understanding of the interplay between iron status and malaria and to develop safe strategies for iron supplementation in areas where malaria is endemic."}
+{"text": "The Tanzanian National Malaria Control Programme (NMCP) and its partners have been implementing regular therapeutic efficacy studies (TES) to monitor the performance of different drugs used or with potential use in Tanzania. However, most of the recent TES focused on artemether\u2013lumefantrine, which is the first-line anti-malarial for the treatment of uncomplicated falciparum malaria. Data on the performance of other artemisinin-based combinations is urgently needed to support timely review and changes of treatment guidelines in case of drug resistance to the current regimen. This study was conducted at two NMCP sentinel sites  to assess the efficacy and safety of artesunate\u2013amodiaquine (ASAQ) and dihydroartemisinin\u2013piperaquine (DP), which are the current alternative artemisinin-based combinations in Tanzania.This was a single-arm prospective evaluation of the clinical and parasitological responses of ASAQ and DP for directly observed treatment of uncomplicated falciparum malaria. Children aged 6\u00a0months to 10\u00a0years and meeting the inclusion criteria were enrolled and treated with either ASAQ or DP. In each site, patients were enrolled sequentially; thus, enrolment of patients for the assessment of one artemisinin-based combination was completed before patients were recruited for assessment of the second drugs. Follow-up was done for 28 or 42\u00a0days for ASAQ and DP, respectively. The primary outcome was PCR corrected cure rates while the secondary outcome was occurrence of adverse events (AEs) or serious adverse events (SAEs).Of the 724 patients screened at both sites, 333 (46.0%) were enrolled and 326 (97.9%) either completed the 28/42\u00a0days of follow-up, or attained any of the treatment outcomes. PCR uncorrected adequate clinical and parasitological response (ACPR) for DP on day 42 was 98.8% and 75.9% at Kibaha and Ujiji, respectively. After PCR correction, DP\u2019s ACPR was 100% at both sites. For ASAQ, no parasite recurrence occurred giving 100% ACPR on day 28. Only one patient in the DP arm (1.1%) from Ujiji had parasites on day 3. Of the patients recruited (n\u2009=\u2009333), 175 (52.6%) had AEs with 223 episodes (at both sites) in the two treatment groups. There was no SAE and the commonly reported AE episodes (with\u2009>\u20095%) included, cough, running nose, abdominal pain, diarrhoea and fever.Both artemisinin-based combinations had high cure rates with PCR corrected ACPR of 100%. The two drugs had adequate safety with no SAE and all AEs were mild, and not associated with the anti-malarials. Continued TES is critical to monitor the performance of nationally recommended artemisinin-based combination therapy and supporting evidence-based review of malaria treatment policies.Trial registration This study is registered at ClinicalTrials.gov, No. NCT03431714 A substantial decline in malaria incidence, over 20% between 2010 and 2015, was observed in the past decade . MalariaPlasmodium falciparum and other species, and it will potentially offer an additional effective anti-malarial drug on top of the current ACT formulations [Improved case management which involves early diagnosis (by microscopy or rapid diagnostic tests) and prompt treatment with effective anti-malarials is one of the current effective strategies to fight malaria . Artemisulations \u20139. Of thulations . Most ofulations \u201328, despulations \u201316.P. falciparum resistance to artemisinins [Due to high level of parasite resistance to SP and following WHO recommendations, Tanzania introduced AL as first-line drug for the treatment of uncomplicated malaria in 2006 . Studiesmisinins , 13, 14 misinins  in SEA, misinins . Thus, tmisinins .The Tanzania National Malaria Control Programme (NMCP), together with its partners, has been and are continuously implementing therapeutic efficacy studies (TES) to monitor the efficacy and safety of different anti-malarials including ACT that are being used or with potential future use in the country. The studies are conducted at eight sentinel sites located in regions with different malaria transmission intensities and also covering border areas, with high potentials of introducing parasites from neighbouring countries , 26. TheThis study was carried out between July and December 2017 at Kibaha and Ujiji sites, which are among the 8 NMCP sentinel sites for monitoring anti-malarial efficacy in Tanzania , 26  of the two health facilities were screened for possible inclusion in the study. Patients were first enrolled in the DP group between July and September 2017, until the sample size was attained before enrolment for ASAQ was initiated (September to November 2017). Enrolled patients were followed-up for 28 (ASAQ) or 42\u00a0days (DP) as per the WHO protocol of 2009 .The sample size was determined based on WHO standard protocol , with thChildren presenting to OPDs with age ranging between 6\u00a0months to 10\u00a0years and fever at presentation (axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C) or reported history of fever in the last 24\u00a0h were screened for possible enrolment, as previously described . RecruitLaboratory screening involved a finger prick to obtain a blood sample for quick detection of malaria parasites using rapid diagnostic tests (RDTs) and collection of thick and thin blood smears for microscopy . For patmsp1 and msp2), and glutamate rich protein (glurp) genes to distinguish true recrudescence from re-infection as previously described [From each patient, dried blood spots (DBS) on filter papers  were collected for PCR genotyping to distinguish recrudescent from new infections. Extraction of parasite DNA from DBS was done at the laboratory in Tanga using QIAamp DNA blood mini kit  according to the manufacturer\u2019s instructions. Genotyping of paired samples (day 0 and parasites collected on or after day 14) was done by analysis of the highly polymorphic loci of merozoite surface proteins 1 and 2  or DP  obtained from WHO. The co-formulated ASAQ tablets contained 25\u00a0mg or 67.5\u00a0mg of artesunate and 50\u00a0mg or 135\u00a0mg amodiaquine in a tablet. DP was also a co-formulated regimen with tablets containing 40\u00a0mg and 320\u00a0mg of dihydroartemisinin and piperaquine, respectively. The drugs were administered according to the recommended doses based on body weight of patients. For ASAQ (25/67.5\u00a0mg), 1 tablet was given to children weighing\u2009<\u200910\u00a0kg; and for the 50/135\u00a0mg tablets, 1 tablet was given to those with 10\u201320\u00a0kg; 2 tablets to children with 21\u201330\u00a0kg and 3 tablets to children weighing\u2009>\u200930\u00a0kg. DP (40/320\u00a0mg) was administered with a quarter, half, a full tablet or 2 tablets given to patients weighing 5 to\u2009<\u20097\u00a0kg, 7 to \u2009<\u200913\u00a0kg, 13 to \u2009<\u200924\u00a0kg and 24 to \u2009<\u200936\u00a0kg, respectively. A full course of either ASAQ or DP consisted of 3 doses given once daily after every 24\u00a0h. Patients were observed for 30\u00a0min to ensure that they did not vomit the study drugs. When vomiting occurred, a repeat dose was given after vomiting stopped. Any patient who persistently vomited the study medication was withdrawn and treated with intravenous quinine or intramuscular artesunate according to the national guidelines for management of severe malaria [Patients enrolled in the study were treated with either ASAQ  when patients felt unwell. Parents/guardians were asked to bring their children to the clinic at any time when they felt unwell without waiting for scheduled visits or taking them to other health facilities for medical attention. All patients who failed to turn up for their scheduled visits by mid-day were followed-up at their respective homes by a member of the study team and asked to come to the health centre for their visits. Patients who travelled away from the centre and could not be traced, were classified as lost and withdrawn from the study. During the visits, both clinical and parasitological assessments were performed; and follow-up samples (blood slides and DBS) were also collected.Both passive and active methods were used to assess the safety of the two drugs through interviews with parents/guardian to capture and report adverse events (AEs) or serious adverse events (SAEs). Parents/guardian were interviewed at each visit and asked to report any occurrence and nature of AE or SAE that occurred at home between follow-up visits. At the study facilities, clinical examination and/or laboratory tests were also used to determine and capture AEs and SAEs. The captured events were recorded on case report forms for each follow-up visit. An AE or SAE were defined and classified according to WHO protocol . ReportiThe primary end point was parasitological cure on day 28 for ASAQ and day 42 for DP as per WHO protocol of 2009 , while sThe protocol was reviewed and approved by the MRCC of NIMR and permission to conduct the study at the health facilities was sought in writing from the relevant regional and district medical authorities. Oral and written informed consent was obtained from parents or guardians of all eligible patients before their children were screened for possible inclusion into the study. Information about the study protocol, inclusion criteria, follow-up schedule, and benefits and risks of participating in the study was provided to parents/guardians during the consenting process. The study is registered at ClinicalTrials.gov, No. NCT03431714.10 transformed parasite density (at enrolment) and age of patients from the two sites were compared using t test  or Mann\u2013Whitney U test . Distinguishing recrudescent from new infections was done using msp2 followed by glurp and msp1 based on the WHO protocol [Single entry was concurrently performed at the study sites during data collection while the second entry was done by another data entry clerk after the end of fieldwork. The data were entered into a Microsoft Access database accessible online via internet, and was later validated, cleaned and analysed using STATA for Windows, version 13 . The data were also transferred to the WHO Excel software programme , for autprotocol , 34. IncA total of 724 patients were screened at both sites, 333 (46.0%) were enrolled into the study and 326 (97.9%) completed the follow-up visits or had an assigned treatment outcome Figs.\u00a0. ChildreThree patients (0.9%) were lost to follow-up at both sites while four 1.2%) (two from each site in the DP group) were withdrawn from the study for different reasons , 175 (52.6%) had one or more events of AEs, with 223 episodes of AEs at both sites. The commonly reported AE episodes included cough (39.0%), running nose 14.8%), abdominal pain (7.2%), diarrhoea (7.2%), fever (6.7%), vomiting (4.0%), skin itching (3.1%), painful micturition (2.7%), painful ear (2.7%), difficulty in breathing (2.2%) and others  was higher compared to what was reported in other studies done in both Mainland Tanzania and Zanzibar \u201338. The Previous studies conducted in Tanzania before and after the 2006 policy changes showed low PCR corrected cure rates among patients treated with ASAQ which ranged from 88 to 94%  and thisP. falciparum [According to WHO, the proportion of patients with parasitaemia on day 3 post-treatment (day 3 positivity rate) should be reported as an important indicator for identifying suspected artemisinin partial resistance in lciparum . In thislciparum \u201318 and olciparum \u201353, furtlciparum , 20. Simlciparum , 54\u201356 alciparum , 58 haveAnopheles funestus , which is a highly potent malaria vector, suggesting that more strategies and targeted malaria control interventions are urgently required to reduce the burden of malaria in this region. From the NMCP perspective, there is a need to ensure that preventive measures like insecticide-treated bed nets are made available and used by the population living in these areas.Although the current study reported day 42 PCR corrected ACPR of 100%, a high rate of recurrent infection was also reported in the DP group at Ujiji and all occurred after day 28. PCR uncorrected ACPR of both drugs on day 28 was 100% and all recurrent infections in the DP group were confirmed to be new infections after PCR genotyping. This is a major concern for effective case management since previous studies involving other artemisinin-based combinations , 19 alsoThis study also showed that the two drugs were well tolerated with minimal AEs and with no any SAE. Of the AEs, majority of the cases (39.0%) had episodes of cough and the rest had other mild symptoms. Studies conducted in Tanzania , 20 and This study showed that the two artemisinin-based combinations had high efficacy for the treatment of uncomplicated falciparum malaria with PCR corrected cure rate of 100% for both drugs. The drugs had adequate safety profile with no SAE and all the reported AEs were mild, and resolved on their own or after medical interventions. Further surveillance will be required to assess the efficacy and safety of these alternative forms of ACT in order to support regular review of malaria treatment guidelines in Tanzania."}
+{"text": "On Day 7, all the subjects had quantifiable levels of rifamycin SV in plasma at each sampling time. After a peak concentration attained 2 h post-dose (mean \u00b1 SD concentration: 10.94 \u00b1 16.41 ng/mL), rifamycin SV decreased in plasma to levels similar to those of Day 1. The amounts of rifamycin SV excreted in urine paralleled the plasma concentration at the corresponding times. On Day 1, the total amount excreted in urine was 0.0013%, and was 0.0029% on Day 7. The study results confirmed those of the previous Phase I study: the systemic absorption of rifamycin SV was also proved negligible after 7 days of the 600 mg t.i.d. dose regimen of the newly formulated tablets, currently under development for the treatment of several small and large intestinal pathologies, including diarrhea-predominant irritable bowel syndrome, hepatic encephalopathy, and others. Registered at ClinicalTrials.gov with the identifier NCT02969252, last updated on 26JAN18.The primary objective of this single- and multiple-dose pharmacokinetic study was the investigation of rifamycin SV\u2019s pharmacokinetic profile in plasma and urine. All the 18 enrolled healthy men and post-menopausal women received modified release tablets containing 600 mg of the oral non-absorbable antibiotic, rifamycin SV, according to a multiple dose regimen: one tablet three times a day (daily intake: 1800 mg) for 14 consecutive days. Blood sampling and urine collection were performed up to 24 h post-dose after the first dose on Days 1 and 7. On average, on Day 1, C Streptomyces mediterranei  [Rifamycin SV, used as sodium salt, is a semisynthetic antibiotic belonging to the class of ansamycins obtained from rifamycin B, which is produced in the fermentation broth of  n. sp.) ,2. RifamThe activity of rifamycin SV is predominantly bactericidal because of its interference with bacterial protein synthesis: the rifamycin family, like other ansamycins, inhibit protein synthesis by binding the \u03b2 subunit of bacterial DNA-dependent RNA polymerase.Escherichia coli, cholera, antibiotic-associated colitis due to Clostridium difficile proliferation, traveler\u2019s diarrhea, giardiasis, and amoebiasis [Antimicrobial drugs represent an important treatment approach and remain the current standard of care for some types of enteric infection , includioebiasis . Since toebiasis ,4 inasmuoebiasis ,6, offerThe efficacy and safety of sodium rifamycin 200 mg tablets administered four times a day (q.i.d.) for 3 days to patients with infectious diarrhea were previously investigated in comparison with rifaximin tablets administered according to the same dose regimen .\u00ae 200 mg modified release tablets were approved in the USA and several European countries for the treatment of patients with travelers\u2019 diarrhea caused by non-invasive strains of Escherichia coli.In addition, the pharmacokinetics of rifamycin SV was previously investigated in healthy male and female subjects after single and multiple administrations of the 200 mg tablets . Oral ri\u00ae modified release tablets, which contain a higher amount of sodium rifamycin SV. The test formulation differed from the recently approved 200 mg tablets, used in the previous studies [The present trial aimed at investigating the pharmacokinetics of the antibiotic after single and multiple doses, three times a day (t.i.d.) for 14 days, in healthy volunteers taking the newly formulated 600 mg MMX studies ,8, not o2\u20133 colony forming units (CFU) per mL [3 CFU/mL of proximal jejunal aspirate [The newly formulated tablets are currently under development for the treatment of diarrhea-predominant irritable bowel syndrome (IBS-D) and other intestinal pathologies with an infective component. Indeed, an increasing amount of data suggests that IBS is commonly associated with small intestinal bacterial overgrowth (SIBO), and that a bacterial overgrowth may be part of IBS pathogenesis ,10,11. U) per mL . A SIBO aspirate . Bacteriaspirate ,14. Indeaspirate ,16,17,18\u00ae tablets upon oral administration. The study included a screening visit, a confinement of 7 days and 8 nights, and 2 ambulatory visits for a minimum study duration of 17 days. All the study subjects received rifamycin SV 600 mg tablets according to the following multiple dose regimen: one tablet 3 times a day  for 14 consecutive days, starting from Day 1. The blood sampling and the urine collection schedule were planned up to 24 h post-dose after the 1st dose on Days 1 and 7, considering the previously published kinetic data [The present study was a single and multiple dose, open-label, pharmacokinetics and safety Phase I study. The primary objective was the investigation of the kinetic profile of rifamycin SV in plasma and urine, and the safety and tolerability of the new 600 mg MMXAccording to previous experience, the safety assessments of the present study were focused on the recording of adverse events, on clinical laboratory assays, and on the measurement of vital signs.The study was performed at the Phase I Unit of CROSS Research S.A., Arzo, Switzerland.2; (iii) good health based on medical history, physical examination, a 12-lead electrocardiogram (ECG), and routine hematology and blood chemistry tests; (iv) willingness to provide written informed consent.Healthy men and post-menopausal women were included into the trial according to the following main inclusion criteria: (i) age of 18 to 55 y; (ii) a body mass index inside the range 18.5\u201330 kg/mMain exclusion criteria were the standard criteria for bioavailability estimation of new drugs, namely: (i) intake of any medication; (ii) a history of drug, caffeine (>5 cups of coffee or tea/day) or tobacco (\u226510 cigarettes/day) abuse; (iii) history of alcohol consumption in excess of two drinks per day for men, and of one drink per day for women, as defined by the U.S.D.A. dietary guidelines .The study was descriptive and non-comparative. Therefore, the sample size was not derived from a statistical power calculation. A total of 18 subjects were planned to be included in the study.The planned population included healthy men, and healthy sterile or post-menopausal women. The exclusion of fertile women was aimed at preventing potential drug\u2013drug interactions of rifamycin SV with oral hormonal contraceptives as rifamycin SV, like the other ansamycins, is a metabolic inducer which could affect the bioavailability of other compounds.All the study subjects took 1800 mg of sodium rifamycin SV as one 600 mg tablet t.i.d. for 14 consecutive days at 08:00 \u00b1 1 h, 14:00 \u00b1 1 h, and 20:00 \u00b1 1 h, starting from Day 1. Each dose was administered with half a glass (\u2248150 mL) of still mineral water.The administrations from Day 1 to the morning of Day 8 were performed by the investigator or deputies at the Phase I Unit.After discharge on Day 8, the subjects continued the treatment according to the same dose regimen at home up to Day 14, for a total of 42 administrations.The planned dose regimen foresaw a daily intake of 1800 mg of sodium rifamycin SV, which is similar to the maximal dose recommended for various injectable preparations of sodium rifamycin available in Europe. The planned intervals between the 3 daily doses (6 h + 6 h + 12 h) aimed at standardizing the dosing conditions throughout each treatment day. In fact, maintaining a \u03c4 of 6 h between the first and the second dose, and also between the second and the third, allowed for each dose to be administered in fasting conditions before each meal. During the confinement, standardized meals were always served after each dosing.The study CRO-PK-15-294\u2013Sponsor code CB-01-11/27 was approved by the independent ethics committee of Canton Ticino on 24 February 2015. Ref. nr. 2888. The Swiss Federal Health Authorities (Swissmedic) authorized the study on 25 June 2015 and assigned the reference number 2015DR1084. The study was conducted in compliance with the Swiss ordinance on clinical trials in human research, and in accordance with the Declaration of Helsinki and the general principles of the ICH Harmonised Tripartite Guidelines for Good Clinical Practice (GCP). Subjects did not undergo any study procedure before signing the written informed consent form. The first subject was enrolled on 9 July 2015 and the last subject completed the trial on 17August 2015.Plasma concentrations of rifamycin SV were measured at: pre-dose (0), 1, 2, 3, 4, 6, 7, 8, 9, 10, 12, 13, 14, 15, 16, 18, and 24 h post-dose after the first dose on Days 1 and 7.Blood sampling at 7, 8, 9, 10 and 12 h post-dose followed the second dose on Days 1 and 7. Blood sampling at 13, 14, 15, 16, 18, and 24 h post-dose followed the third dose on Days 1 and 7.Urine concentrations of rifamycin SV were measured during the following time intervals: 0\u20133, 3\u20136, 6\u201312, 12\u201318, and 18\u201324 h post-dose after the first dose on Days 1 and 7.Blood samples were collected using an indwelling catheter with a switch valve into K2-EDTA tubes, and stored on ice for a maximum of 20 min. Then, the samples were centrifuged at 4 \u00b0C for 10 min at 2500 g to obtain plasma. Each plasma sample was immediately divided into 3 aliquots in pre-labelled amber polypropylene tubes, and stored frozen at \u2264 \u221270 \u00b0C pending analyses.During each interval, urine was collected into containers, and kept refrigerated. At the end of each collection interval, urine volume was measured, 1% of a 1% Tween-20 solution was added to prevent rifamycin SV absorption in the container material and, after thorough mixing, 3 aliquots were prepared in amber polypropylene tubes. The aliquots were stored at \u2264 \u221270 \u00b0C until analyses.The concentration of rifamycin SV in plasma and urine was determined at the ABL Analytisch Biochemisch Laboratorium, the Netherlands, by a validated LC\u2013MS/MS method, also used in the previous study . The low\u00ae 6.3 (Pharsight Corporation):The following pharmacokinetic parameters  were meaThe following parameters were measured and/or calculated for plasma rifamycin SV after the 3 multiple doses of Day 7.The following parameters were measured/calculated for urine rifamycin SV after the first 3 doses (Day 1), and after the 3 multiple doses of Day 7.The safety variables included: the recording of adverse events during the whole study duration; blood pressure, heart rate, and body weight measurements at screening, pre-dose and upon discharge; physical examinations; ECG; and routine blood chemistry, hematology, and urinalysis laboratory assays performed at screening, and upon the end of the study.Eighteen (18) Caucasians, aged 22 to 55 y were enrolled as planned. Baseline demographic data are summarized in All 18 subjects received the planned treatment and completed the study according to the protocol.Mean rifamycin SV concentration (ng/mL) vs. time profiles on Days 1 and 7 of treatment are shown in The plasma rifamycin SV concentrations vs. time curves could not be extrapolated to infinity for any subject at the established conditions.On Day 1, quantifiable levels of rifamycin SV were first found in plasma 3 h after the first dose. The plasma concentration of the antibiotic increased slowly over the second dose and attained a peak 9 h post-dose (3.51 \u00b1 4.55 ng/mL), i.e., 3 h after the second dose. After the sampling at 9 h post-dose, rifamycin SV decreased slowly in plasma over the third dose.On Day 7, as expected, all the subjects had quantifiable rifamycin SV concentrations at pre-dose (2.73 \u00b1 1.54 ng/mL). After the morning dose, rifamycin SV concentration increased rapidly in plasma, and attained a first peak concentration 2 h post-dose (10.94 \u00b1 16.41 ng/mL). At 2 h post-dose, one third of the subjects had plasma rifamycin SV concentrations >10 ng/mL, which in one case reached a concentration >70 ng/mL. Afterwards, the antibiotic decreased in plasma, and from 7 to 18 h post-dose ranged between 1.90 and 2.98 ng/mL. Pre-dose and 24 h post-dose concentrations showed that systemic rifamycin SV was at the steady state.max,0\u20136 of Day 1 vs. Cmax,12\u201318 of Day 7) increased by 1.32 times, and the extent of absorption  increased by 3.73 times. These data show that the rifamycin SV accumulation after 7 days of t.i.d. treatment was very limited. When AUC0\u201324 of Day 1 is compared with AUCss,0\u201324 of Day 7 an increase in extent of absorption of rifamycin SV by 1.83 can be observed. This increase in rifamycin SV AUC from Day 1 to Day 7 is partly due to the notable concentration peak at 2 h post-dose on Day 7, mentioned above. Otherwise, the curve of plasma concentration of rifamycin SV vs. time on Day 7 is almost superimposable on that of Day 1 from 8 h to 24 h post-dose.When rifamycin SV parameters of Day 1  are compared with those of Day 7 , a slight increase in the exposure to rifamycin SV can be observed after 7 days of treatment. In detail, the rate of absorption of the analyte . The peak observed in the 0\u20133 h interval is consistent with the peak observed in plasma concentrations at 2 h post-dose. Afterwards, excretion decreased in the 3\u20136 h interval and after 12 h had levels similar to those observed on Day 1 in the collections between 6 and 24 h post-dose. Between 6 and 24 h post-dose, the rate of rifamycin SV urinary excretion was on average constant.The total amount of rifamycin SV excreted on Day 7 of the treatment increased on average by 2.3 times, compared to the amount excreted on Day 1, which is consistent with the plasma concentration data.Adverse events occurred to 6/18 subjects (33.3%). The majority of reported adverse events were not judged to be related to the treatment.The reported adverse events are summarized by system organ class and preferred term in The most common event was headache. All the three reported episodes started within about 5 h of the previous treatment dose. One of the headache episodes was of moderate intensity, while the other two were mild and resolved spontaneously after about 12 h. None of the headache episodes was judged as related to the treatment.During the study treatment, some abnormalities in the measured clinical laboratory assays were judged as non-clinically significant by the investigator, with the exception of five urine parameters for one subject who suffered from cystitis on the last treatment day. In detail, the presence of hematic pigments and leukocytes in urine, and of leukocytes, erythrocytes, and bacteria in the urinary sediment found at the final visit analysis was abnormal and clinically significant. However, all five abnormalities were signs of the current cystitis with stranguria and were unrelated to the study treatment.No notable change in any other laboratory parameter was observed between the final laboratory analysis and the screening. As expected with non-absorbable drugs, no significant effect of rifamycin SV on any laboratory parameter was observed during the study.No significant change in vital signs, body weight, or ECG was observed after treatment with rifamycin SV for 14 consecutive days.The new dosage form of rifamycin SV was administered for 14 consecutive days for the first time in the present study, which also took advantage of a more sensitive LC\u2013MS/MS analytical method for the assay of the antibiotic in plasma and urine.Healthy post-menopausal women and men enrolled in the present study were exposed to a daily dose of 1800 mg of rifamycin SV for 14 consecutive days, thus allowing the investigation of the kinetic profile of the systemically available rifamycin SV, both after the first dose, and after 7 days of treatment, and the collection of extended safety information after further 7 days of treatment with the new formulation.0  measured 5 min after a single i.v. 250 mg dose of the antibiotic [The plasma concentrations of rifamycin SV observed in the present study were consistent with the results of the previous Phase I study  with a dNotably, even the tablet formulation, renewed in the core matrix composition, that allowed the active ingredient progressive release to start at pH 6 instead of 7, i.e., in the ileum, did not have any significant impact on the peculiar rifamycin kinetics, inasmuch as this change did not enhance the antibiotic systemic absorption.Safety data of the present trial were also in agreement with the results of the previous Phase I multiple dose study , in factThe results of the present study are in accordance with those of the previous studies, thus confirming an excellent safety profile. Furthermore, no severe adverse events were reported, and no subject discontinued the study due to safety concerns. Some abnormal urine parameters found after treatment were signs of an episode of cystitis, which was unrelated to the treatment. No significant change in any other laboratory parameter was observed. No significant change in vital signs, body weight, or ECG was observed after treatment. No significant change in liver or kidney functions was observed during the 14th day treatment.In conclusion, as evidenced in particular by the plots, which are unable to show any characteristic curves due to the reduced amount of the antibiotic in the systemic circulation, and by the pharmacokinetic parameters, the study results showed that the absorption of rifamycin SV was very low, and also after a multiple-dose treatment. With respect to the kinetics of any other drug claiming systemic efficacy, the feature of negligible systemic absorption is generally regarded as an inadequacy or a failure. On the contrary, it is indeed to be considered as beneficial and supportive in the case of rifamycin SV, which is expected to display high local efficacy and low systemic unwanted effects. The negligible rifamycin SV absorption shown herewith after a t.i.d. multiple-dose treatment for 14 days, makes the antibiotic also suitable for additional gastroenteric indications, such as several small and large intestinal pathologies, including diarrhea-predominant irritable bowel syndrome, hepatic encephalopathy, and others."}
+{"text": "P. falciparum. Artemether-lumefantrine  is the most widely used artemisinin-based combination therapy (ACT), for treating uncomplicated falciparum malaria globally. However, the development of resistance to antimalarial drugs is a major challenge for malaria control. In this review, the efficacy of AL for the treatment of uncomplicated falciparum malaria in Africa was evaluated.  Africa still bears the largest burden of malaria as the majority of infections in the continent are caused by  Articles published between January 2015 and July 2019 were systematically searched using comprehensive search strings from PubMed/Medline, SCOPUS, and grey literature from Google Scholar. Interventional studies that followed patients for at least 28 days were included. Two reviewers independently assessed study eligibility, extracted data, and assessed risk of bias. All the included articles were measured to be good quality. While computing the efficacy of AL, polymerase chain reaction (PCR)\u2013corrected cure rate  at day 28 was considered as the main endpoint. Meta-analysis was computed using STATA v 15 to calculate the pooled ACPR.  In this review, 39 articles that reported the treatment outcome of 8,320 patients were included. After 28 days of follow-up, the pooled PCR uncorrected and corrected APCR was at 87% (95% CI: 85-90%) and 97.0% (95% CI: 96-98%), respectively. Moreover, the proportion of early treatment failure (ETF) was almost 0%, while most of the included articles reported <8% late treatment failures. The reinfection and recrudescence rate was less than 10% and 2.6%, respectively, within 28 days. We noted rapid fever and parasite clearance in which greater than 93% and 94% patients were parasite and fever free at day three following AL treatment.  This review discovered that despite more than a decade since its introduction, Coartem\u00ae remains effective and thus could continue to be the drug of choice for the treatment of uncomplicated falciparum malaria for all age groups in Africa. However, the risk of new emerging resistance for this combination warrants regular monitoring of its efficacy across the continent. Althougdication , 3. Therglobally , 5. P. fn Africa . Althougn Africa .A patient who presents with symptoms of malaria and a positive parasitological test (microscopy or Rapid diagnostic test (RDT)) but with no features of severe malaria is defined as having uncomplicated malaria . UncomplWhile the exact numbers may be uncertain and under reporting is inevitable, according to the latest World Health Organization (WHO) malaria report, an estimated 219 million cases of malaria occurred worldwide in 2017. In same year, the African region was home to 92% of malaria cases and 93% of malaria deaths. Fifteen countries in sub-Saharan Africa and India carried almost 80% of the global malaria burden. Five countries accounted for nearly half of all malaria cases worldwide: Nigeria (25%), Democratic Republic of the Congo (11%), Mozambique (5%), India (4%), and Uganda (4%) . Similarfalciparum malaria [Malaria case management consists of early diagnosis and prompt treatment . In rece malaria . They ar malaria . The cho malaria . In Afri malaria  ACT, whe malaria .P. falciparum malaria. Both components are blood schizontocides. The dual mechanisms of action of AL provide rapid and sustained parasite clearance [AL or Coartem\u00ae  was the first fixed dose combination of an artemisinin derivative with a second unrelated antimalarial compound. It is a safe and effective treatment for children and adults with learance . Lumefanlearance .Resistance to antimalarial medicines is a great public health challenge. Antimalarial drug resistance may continue to be a leading threat to ongoing malaria control efforts and calls for continued monitoring of the efficacy of these drugs for drug policy input . RegularAs there is quite limited reviewed data on the topic of AL resistance in Africa, there is a need to better understand the dynamics of parasite clearance in patients treated with ACT in order to better detect the emergence of AL resistance for intervention . It is aThis systematic review stands with the following question: what is the therapeutic efficacy of AL for the treatment of uncomplicated falciparum malaria in Africa over the last five years?The main aim of the review was to summarize the latest five years data on the therapeutic efficacy of AL for the treatment of uncomplicated falciparum malaria in the African context.CRD42020142590.In accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guideline, the review protocol was registered on the International Prospective Register of Systematic Reviews (PROSPERO) with a registration number Study design: interventional studies that reported the therapeutic efficacy of AL for the treatment of uncomplicated malaria. Participants: P. falciparum-infected patients irrespective of gender and age group. Interventions: a standard six-dose regimen of AL over three days followed up for 28 days to measure its therapeutic responses. Setting: we included studies with the outcome of interest reported in Africa. Language and publication: we included peer-reviewed journal articles and unpublished findings reported in the English language in the last five years (January 2015 to July 2019).Studies were selected based on the following criterion. This review was done following PRISMA . A compuPlasmodium falciparum malaria\u201d, \u201cfaciparum malaria\u201d, \u201cantimalarial drug\u201d, and \u201cAfrica\u201d. We combined these terms using the Boolean operator \u201cOR\u201d and \u201cAND\u201d accordingly. Full search strategy for the two databases is attached separately as a supplement.The domains of the search terms were: \u201cefficacy\u201d, \u201ctherapeutic efficacy\u201d, \u201cartemether-lumefantrine\u201d, \u201dCoartem\u201d, \u201cStudies that have been published in the last five years (2015 to 2019) and reported the therapeutic efficacy of AL for the treatment of uncomplicated falciparum malaria in African context were included. Searched articles were directly imported and handled using EndNote X5 citation manager . Based on the PRISMA protocol, duplicated articles were excluded, and the titles and abstracts of the remaining papers were screened independently for inclusion in full text evaluation by the first two authors. Differences between the reviewers were resolved through discussion.The Joanna Briggs Institute (JBI) data extraction tool was adopted for data extraction. Relevant data such as the name of the first author, year of publication, country where the study was conducted, mean/median age of the study participants, proportion of male participants, type of the study design, total number of the study participants, follow-up period, baseline characteristics of study participants , and fever and parasite clearance rates were extracted from the included articles. Moreover, based on the WHO recommendation , the treValidity and methodological quality of all included studies were assessed using the national institute of health (NIH) study quality assessment tool for intervention studies . The tooThe data extracted from the included studies were fed into Microsoft Excel. Descriptive statistics, such as simple counts, ranges, and percentages, were used to present the synthesized data. A systematic narrative synthesis was provided in which summary results were presented using text and table. To compute the pooled ACPR with its 95% CI, meta-analysis was done using STATA v15  assuming a random effect model. In this review, following AL therapy, the primary endpoint  was cure rate (or ACPR), corrected to exclude reinfection using polymerase-chain reaction (PCR), at day 28.The definition of the following terms was adopted from cited references , 26\u201331.P. falciparum parasitological clearance at day 28 irrespective of axillary, oral, rectal, or tympanic temperature without previously meeting the criteria of early treatment failure or PCR corrected late treatment failure.Adequate clinical and parasitological response (ACPR): refers to P. falciparum parasitemia.Early treatment failure (ETF): signs of severe malaria/clinical deterioration requiring rescue medication on days 0, 1, 2, or 3, in the presence of P. falciparum parasitemia, without previously meeting any of the criteria of ETF.Late clinical failure (LCF): signs of severe malaria/ clinical deterioration requiring rescue medication after day 3 in the presence of P. falciparum parasitemia on any day from day 7 onward and the absence of fever without previously meeting any of the criteria of ETF/or LCF.Late parasitological failure (LPF): presence of PCR-corrected: refers to the use of molecular testing to differentiate recrudescence from reinfection when evaluating efficacy. Recurrent parasitemia classified as recrudescence if it was due to the same parasite strain as that on day 0  and as a new infection if it was due to a genetically different strain .From the systematically searched databases and other sources, a total of 605 articles were retrieved and sequentially screened for final inclusion. As depicted in The characteristic of the included articles is summarized in . The stuThe number of participants in each included study varied from 33 to 595. Majority of the included articles 31 (79.5%) measured the efficacy of AL in terms of ACPR at day 28, while in the remaining studies at 7 (17.9%) and 1 (2.7%), the follow-up period was 42 and 63 days, respectively. In total, the review contains reports of 8,320 patients. Most 31 (79.5%) of the studies included patients who were older than six months, while in the remaining studies at 7 (17.9%) included all age groups and 1 (2.6%) include only pregnant women.The methodological quality of all the included studies was assessed using the national institute of health (NIH) study quality assessment tool for intervention studies . Provide\u03bcl of whole blood) was estimated using the following formula; \u201cnumber of parasites counted\u201d/\u201cWBC counted\u201d multiplied with \u201ctotal WBC count/\u03bcl\u201d [The baseline characteristics of patients employed by the included articles are shown in ount/\u03bcl\u201d .The overall treatment outcome is summarized in . In the Likewise, the pooled PCR corrected APCR was at 97.0% (95% CI: 96-98%) .P. falciparum.In this review, the overall early treatment failure (ETF) rate was almost 0%, while the proportion of late treatment failures  was between 0% and 25.6% and 0% and 52.6%, respectively. The common type of treatment failure was late parasitological failure (52.6%) which was reported by only a study . The reiThe distribution of fever and parasite clearance rates reported by some of the included articles is shown in Finally, in this review, the most frequently reported adverse events associated with treatment with AL were cough, fatigue, weakness, anemia, GIT disorder , fever, and headache .This review paper provides the latest data on the efficacy of AL (Coartem\u00ae), one of the ACTs for the treatment of uncomplicated falciparum malaria in African, where the efficacy of this drug has been less frequently evaluated especially in the last five years. More than 40 malaria-endemic countries in Africa were using ACT as first-line treatment for uncomplicated falciparum malaria .P. falciparum in most African countries [Mekong Subregion), specifically Cambodia and Thailand, there is a fear that the resistance may spread globally and it may pose a significant threat to malaria elimination. This emergence of artemisinin resistance has raised concerns that the most potent antimalarial drug may be under threat. Therefore, as per the WHO recommendation, there is a need for routine monitoring of the effectiveness of artemisinin derivatives in Africa for early intervention [Six dose regimen of AL over three days is the standard treatment for uncomplicated ountries , 44. Howrvention , 22, 45.In this review, the primary efficacy endpoint was PCR-corrected ACPR (cure rate) at day 28. The pooled ACPR of AL for the treatment of uncomplicated falciparum malaria was at 87% (95% CI: 85-90%) before PCR adjustment and at 97.0% (95% CI: 96-98%) after adjustment. Except a study by Sondo et al. that reported 77.8% PCR corrected ACPR . Since aPlasmodium within short hours of treatment and prevention of maturation of the gametocytes by the partner drug (lumefantrine) offers the maximum performance of AL [A global pooled analysis by Makanga et al. (2011) showed that the 28-day PCR-corrected parasitological cure rate (primary efficacy endpoint) was >97% for all age groups. This paper also reported that AL had rapid resolution of parasitemia and fever and also showed an excellent safety . Anotherce of AL . Furtherce of AL .In this review, the overall treatment failure rate was low; <10%. The early treatment failure (ETF) rate was almost 0%, while the proportion of late treatment failures  was ranged between 0% and 52.6%. The common type of treatment failure was late parasitological failure (LPF) in which relatively higher proportion of such type of failure was reported by five studies; 12.3-25.4% , 59, 69 P. falciparum to artemisinins [Day 3 parasitemia after treatment with a full dose of AL shown to be delayed parasite clearance  or a goomisinins . The artmisinins . Delayedmisinins . Data onmisinins , the oveAmong 20 articles that reported data on fever clearance, about 13 reported a 100% clearance rate on the third day. Except a study by Yaka et al.  that repP. falciparum [In our review, during the 28-day follow-up, although AL almost cleared fever and parasitemia within 3 days, we have also noted some level of reinfection and recrudescence. Reinfection is the development of malarial signs and symptoms due to a new strain, while recrudescence specifies the infection that has recurred from persistent blood stages of lciparum . Except lciparum , 58, 59 lciparum  and 6.1%lciparum  recrudeslciparum . This melciparum . This relciparum .falciparum strains to lumefantrine [The reported recrudescence rate in our review may reflect a decrease in the sensitivity of some fantrine . In the fantrine . A recenfantrine . While dfantrine . Howeverfantrine . Therefofantrine .To the best of our knowledge, this systematic review reported the latest therapeutic efficacy of AL for the treatment of uncomplicated malaria in African. However, the review should be interpreted in light of a couple of drawbacks; the absence of data from some African countries might compromise the overall picture of the current efficacy of AL in the continent. The other pitfall of this review is the heterogeneity of the articles in terms of the study design and the included age groups of the participants. Yet another notable limitation of the review is that it primarily considered the ACPR data of the 28 days of follow-up, the minimum period recommended by WHO for drugs with elimination half-lives of less than seven days ; any addThis review discovered that despite its introduction for more than a decade, AL is effective and thus could continue to be the drug of choice for the treatment of uncomplicated falciparum malaria in Africa for all age groups. This would imply no imminent threat of AL resistance development in the region. There may be a need to further investigate the comparatively low efficacy (<90%) rate reported by a study  in order"}
+{"text": "Plasmodium falciparum malaria. ACT resistance is spreading in Asia but not yet in Africa. Reduced effects of ACT partner drugs have been reported but with little information regarding widely used artesunate/amodiaquine (ASAQ). We studied its efficacy in Zanzibar after 14 years as first-line treatment directly by an in vivo, single-armed trial and indirectly by prevalences of different genotypes in the P. falciparum chloroquine-resistance transporter, multidrug-resistance 1, and Kelch 13 propeller domain genes. In vivo efficacy was higher during 2017  than during 2002\u20132005  (p = 0.003). Molecular findings showed no artemisinin resistance\u2013associated genotypes and major increases in genotypes associated with high sensitivity/efficacy for amodiaquine than before ASAQ was introduced. Thus, the efficacy of ASAQ is maintained and appears to be increased after long-term use in contrast to what is observed for other ACTs used in Africa.Artemisinin-based combination therapies (ACTs) are first-line treatments for uncomplicated  Plasmodium falciparum malaria globally for the past 10\u201315 years and has contributed greatly to a reduction of malaria illnesses and deaths during 2005\u20132015 (Artemisinin-based combination therapy (ACT) has been first-line treatment for uncomplicated In Zanzibar, malaria transmission has been reduced substantially after new and reinforced malaria tools and interventions, including ACT for uncomplicated malaria . The in vivo and molecular results were then compared with findings from previous studies conducted during 2002\u20132013.In Zanzibar, 2 previous clinical trials in 2002\u20132003 , in accordance with World Health Organization  and primaquine (single low-dose) treatment for uncomplicated nization . We sele>37.5\u00b0C) or had a history of fever (past 48 hours) and confirmed microscopically with any level of asexual P. falciparum parasitemia. They were finally enrolled if considered able to comply with the study protocol  and if written informed consent was obtained from patient, parent, or guardian. Exclusion criteria were severe malaria signs, underlying disease, positive pregnancy test result, or suspected alternative reason for the febrile condition.The participants  were recruited among febrile patients attending the 14 selected health facilities. They were screened by using malaria rapid diagnostic tests (mRDTs), and if positive results were found, they were referred to the closest referral center. Participants were considered eligible for study inclusion if they were confirmed to be febrile (axillary temperature https://www.sanofi.com) once a day for 3 consecutive days. A single low-dose (0.25 mg/kg) of primaquine  was co-administered on the first day (D0). All doses were administered under direct supervision and observation for 30 min. If vomiting occurred, the patient was again given the same supervised dose and withdrawn from the study if a second vomiting occurred.The enrolled patients were given treatment and followed-up at the referral centers. They received the antimalarial standard treatment orally , the follow-up consisted of fixed appointments on D3, D7, D14, D21, and D28 and whenever the patient experienced any clinical symptoms. To ensure optimal compliance to the study protocol, an incentive of 5,000 Tanzanian shillings (US $2.20) was provided upon each visit, mainly to cover transportation costs.https://www.cytivalifesciences.com). These filter papers were packaged in desiccated individual plastic bags and sent to the Karolinska Institutet  for molecular analyses.At each follow-up visit, standard physical examination was performed and temperature was recorded. Finger-prick blood samples were collected for thick blood films and dried blood spots on filter paper  on D0, D3, D7, D14, and D28, or any day of clinical suspicion of (hemolytic) anemia.Microscopy reading of Giemsa-stained blood films was performed by 2 experienced microscopists at each visit. We quantified parasitemia by using the standard approximation method (40\u00d7 parasites/200 leukocytes). A blood film was defined as positive if >1 asexual parasite was found per 1,000 leukocytes and the final parasite density was the average of 2 independent reads. An independent examination by a third microscopist was performed in case of discordant reads. We measured hemoglobin levels by using a HemoCue B-Hemoglobin Photometer  and P. falciparum chloroquine-resistance transporter (pfcrt) genes, associated with amodiaquine resistance , associated with artemisinin resistance were analyzed by using nested PCR amplification with Q5 high-fidelity polymerase , followed by direct Sanger sequencing of the PCR amplicon  with uncomplicated P. falciparum malaria . Study procedures were similar to those in 2017. We performed paired PCR genotyping of the P. falciparum merozoite surface protein 2 gene in samples collected on D0 and for recurrent parasitemia days 14\u201328 .https://www.microsoft.com) and cleaned data by using GSPro (https://www.dji.com). We performed statistical analyses by using Stata (https://www.stata.com). We calculated 95% CIs for proportions of patients cured by D28 by using the exact method described by Fagan .We entered data into Microsoft Excel  were positive by mRDT, and 146 satisfied all inclusion criteria and thus enrolled at the 3 referral centers. We provide demographic, clinical, and laboratory characteristics for patients , along w<15 years  in 2002\u20132003, 16 (9%) in 2005, and none (0%) in 2017, confirming higher malaria transmission rates in 2002\u20132005.Parasite clearance rates by microscopy up to D3 were similar in the 3 trials ; Table 3PCR positivity and parasite density estimates by quantitative PCR were analyzed only for the 2017 study . Patient<5 years of age when compared with 2002\u20132003 and 2005 (Fever clearance rates were similar: temperatures <37.5\u00b0C by D3 in 92.8% (95% CI 88.4%\u201395.9%) of patients in 2002\u20132003, 98.8% (95% CI 95.9%\u201399.9%) of patients in 2005, and 97.9% (95% CI 94.0%\u201399.6%) of patients in 2017. Hemoglobin levels at enrollment (D0) were higher in all patients in 2017  but simiand 2005 . The avepfcrt K76T prevalence from 98.0% in 2003 to 4.9% in 2017 (p<0.001)   and nega in 2017 . The decpfK13 gene, 139 (98%) of 142 samples collected 2017 were successfully sequenced. No nonsynonymous SNP was identified. Two synonymous mutations were found, the SNPs C469C (5 samples) and S477S (1 sample). Five samples were from patients at the Bububu health center, and 1 (C469C) was from a patient in Uzini who had traveled to mainland Tanzania.Regarding the <3 days), and fever clearance was similar. These findings confirm maintained full efficacy of the artesunate compound (The high cure rate in 2017 (100%) was significantly higher than the combined cure rate in 2002\u20132003 and 2005 (94.7%) (p = 0.003). The microscopy-determined parasite clearance was as rapid as that in 2002\u20132003 and 2005 . In Zanzibar, our in vivo and molecular findings suggest that Resistance mutations confer an advantage in the presence of the drug, although such mutations often come with fitness costs in the absence of the drug  resistant parasites.Despite 14 years of widescale use of ASAQ as a first-line treatment for malaria in Zanzibar, there are no indications of increased tolerance/resistance to either drug. Our in vivo and molecular findings suggest an increased antimalarial activity by the partner drug amodiaquine. We believe that this finding might be primarily caused by fitness costs of the amodiaquine tolerance/resistance\u2013related mutations in the"}
+{"text": "Plasmodium falciparum malaria cases in Africa are treated with the artemisinin combination therapies artemether-lumefantrine (AL) and artesunate-amodiaquine (AS-AQ), with amodiaquine being also widely used as part of seasonal malaria chemoprevention programs combined with sulfadoxine-pyrimethamine. While artemisinin derivatives have a short half-life, lumefantrine and amodiaquine may give rise to differing durations of post-treatment prophylaxis, an important additional benefit to patients in higher transmission areas.The majority of n\u2009=\u20094214 individuals). The time to PCR-confirmed reinfection after treatment was used to estimate the duration of post-treatment protection, accounting for variation in transmission intensity between settings using hidden semi-Markov models. Accelerated failure-time models were used to identify potential effects of covariates on the time to reinfection. The estimated duration of chemoprophylaxis was then used in a mathematical model of malaria transmission to determine the potential public health impact of each drug when used for first-line treatment.We analyzed individual patient data from 8 clinical trials of AL versus AS-AQ in 12 sites in Africa , AS-AQ provided ~\u20092-fold longer protection than AL. Conversely, at a higher prevalence of 86Y and 76T mutant parasites (>\u200980%), AL provided up to 1.5-fold longer protection than AS-AQ. Our simulations found that these differences in the duration of protection could alter population-level clinical incidence of malaria by up to 14% in under-5-year-old children when the drugs were used as first-line treatments in areas with high, seasonal transmission.We estimated a mean duration of post-treatment protection of 13.0\u2009days (95% CI 10.7\u201315.7) for AL and 15.2\u2009days (95% CI 12.8\u201318.4) for AS-AQ overall. However, the duration varied significantly between trial sites, from 8.7\u201318.6\u2009days for AL and 10.2\u201318.7\u2009days for AS-AQ. Significant predictors of time to reinfection in multivariable models were transmission intensity, age, drug, and parasite genotype. Where wild type Choosing a first-line treatment which provides optimal post-treatment prophylaxis given the local prevalence of resistance-associated markers could make a significant contribution to reducing malaria morbidity. Plasmodium falciparum malaria. In each ACT, the artemisinin derivative is combined with a different antimalarial partner drug. There are currently five ACTs recommended by the World Health Organization (WHO): artemether-lumefantrine (AL), artesunate-amodiaquine (AS-AQ), dihydroartemisinin (DHA)-piperaquine, artesunate-mefloquine, and artesunate-sulfadoxine-pyrimethamine\u00a0(AS-SP) [Nearly all malaria-endemic countries use artemisinin-based combination therapies (ACTs) as first-line treatment for uncomplicated \u00a0(AS-SP) . In area\u00a0(AS-SP) .Each of the six drug regimens has different pharmacokinetic and pharmacodynamic properties, and these have implications for the public health benefit of the drugs in terms of their ability to reduce overall malaria transmission in the community, as well as cure disease . The artArtemether-lumefantrine (AL) is globally the most widely used ACT, followed by artesunate-amodiaquine (AS-AQ) . While rP. falciparum multidrug resistance transporter 1 (pfmdr1) gene and 76T in the P. falciparum chloroquine resistance transporter (pfcrt) gene are linked to reduced sensitivity to AS-AQ but increased sensitivity to AL, which is thought to be due to differential sensitivity to the amodiaquine and lumefantrine partner drugs rather than the artemisinin. Although the overall efficacy of each drug remains high in Africa, a meta-analysis found that the N86 wild type parasite was associated with a fourfold increased risk of recrudescence after AL treatment [There is evidence that the duration of protection after AS-AQ and AL treatment is affected by parasite mutations associated with reduced drug sensitivity , 11. Thereatment , 11. AllP. falciparum case management.The duration of protection can be estimated from clinical trials where reinfection rates are monitored. We previously estimated the mean protection provided by AL at 13.8\u2009days, and DHA-piperaquine at 29.4\u2009days , 24. HerTo assess the duration of post-treatment prophylaxis provided by AL and AS-AQ, we analyzed clinical trial data obtained from the WorldWide Antimalarial Resistance Network (WWARN) data sharing platform  with theP. falciparum malaria; AS-AQ and AL being compared; follow-up to at least day 28, with at least one follow-up visit at day 14 and another before day 28; 100 or more participants per study site or more than 28\u2009days follow-up; polymerase chain reaction (PCR)-adjusted efficacy available; at least 95% PCR-adjusted treatment efficacy in both study arms; PCR-unadjusted cure rates of <\u200995% in at least one trial arm by day 28 ; standard dose regimens of AL and AS-AQ (we included studies regardless whether AS-AQ was given as a fixed-dose combination or not); and known dosage taken for each patient. Individual patient data from eligible studies were shared, collated, and standardized using previously described methodology [WWARN invited investigators to contribute individual-level patient data for this meta-analysis  if theirhodology .glurp, msp1, and msp2 . Patients who experienced PCR-confirmed recrudescence were excluded. The majority of included trials did PCR correction using three molecular markers: In two studies  which in turn depends on the entomological inoculation rate (EIR), the number of infectious bites per person per year). More specifically, the time span between the end of the protected period and reinfection follows an exponential distribution with mean 1/ried out , 29. Wheried out , which hRi in host i asPdi is the duration of chemoprophylaxis of drug d in host i, Ii is the time until reinfection occurs in host i once at risk, and \u03b4 represents the time required for a blood-stage infection to become patent after hepatocyte rupture . P and \u03bb and shape parameter r are to be estimated, andi being the force of infection to which individual i was exposed during the trial follow-up. We assume that protection by the drug is all-or-nothing and that protection times in the population follow a gamma distribution, with a median for each drug that is constant in each trial site. The variance of this gamma distribution incorporates the effect of factors that are not specifically modeled, such as variation in pharmacokinetics, and potentially variation in sensitivity of different parasite clones to the drugs within each site. Individual-specific EIR values \u03b5i were determined, taking into account that young children are bitten less often due to their smaller body size, according to the formula\u03b5adult is the estimated site-specific EIR experienced by fully grown individuals, a is age and parameters a0\u2009=\u20092920\u2009days and \u03c1\u2009=\u20090.85 control the shape of the relationship [where the drug-specific scale parameter tionship . Pre-ery\u03b5adult for each site were estimated simultaneously with the other parameters, with moderately informative gamma priors with median as predicted by the Malaria Atlas Project [r, with hyperparameters (parameters of the prior distribution) determined using a fit of the hidden semi-Markov model with non-informative priors. This improved MCMC convergence. Non-informative gamma priors were chosen for all remaining estimated parameters. We ran the MCMC procedure for 1.25 million iterations, retaining 100,000 samples of the posterior after discarding 4000 adaptation steps, 4000 burn-in steps, and thinning.A number of hidden semi-Markov model variants were fitted via MCMC (Markov-Chain Monte Carlo), using the JAGS (\u201cJust Another Gibbs Sampler\u201d) software for Bayesian inference in conjunction with the \u201crjags\u201d package using R statistical software . The lik Project  . With data pooled across trials, the median duration of protection against reinfection after AS-AQ treatment, i.e., the time during which patients have drug levels which would prevent reinfections, was estimated at 15.2\u2009days (95% CI 12.8\u201318.4) and, after AL treatment, 13.0\u2009days (95% CI 10.7\u201315.7) Fig.\u00a0\u201d, since pfmdr1 86Y surveys matching 11 trial sites, and pfcrt 76T matching 10 sites . However, there were too few matched surveys of pfmdr1 1246Y to analyze this third mutation further. Local prevalence of the mutations pfmdr1 86Y and pfcrt 76T significantly altered the association between drug and time to reinfection. AS-AQ was associated with a significant 1.37 (95% CI 1.28\u20131.47)-fold increase in time to reinfection compared to AL when pfmdr1 86Y prevalence was 20% , but a significantly shorter time to reinfection than AL when pfmdr1 86Y was 80% (ratio of reinfection times AS-AQ vs AL\u2009=\u20090.89 95% CI 0.84\u20130.94). Similarly, AS-AQ was associated with a 1.54 (95% CI 1.38\u20131.71)-fold increase in time to reinfection compared to AL when pfcrt 76T prevalence was 20%, but a 1.06 (95% CI 1.03\u20131.10)-fold change when pfcrt 76T prevalence was 80%. Other factors that were significantly associated with longer time to reinfection when adjusting each factor only for log EIR were younger age and higher dose of lumefantrine (mg per kg)  after adjusting for EIR, age, and lumefantrine dose.We constructed multivariable models for each treatment arm separately. In the AL arm, EIR, age, lumefantrine dose (mg per kg), local pfmdr1 86Y prevalence remained significantly associated with time to reinfection overall, with 86Y associated with a 0.97-fold decrease in reinfection time per 10% increase in prevalence (p\u2009=\u20090.011). For sensitivity analysis, we repeated the regression model including only the trial sites which used the FDC formulation of AS-AQ, and here the effect of pfmdr1 86Y was no longer statistically significant although the effect size remained similar -fold change in reinfection times, p\u2009=\u20090.159). Again, we looked at pfcrt 76T in a separate multivariable model in the AS-AQ arm; here, it was no longer significantly associated with reinfection time after adjusting for EIR and age, although there was still a trend for shorter time to reinfection as 76T prevalence increased .In the AS-AQ arm, EIR, age, and pfmdr1 86Y and pfcrt 76T prevalence with prophylactic time by examining the site-specific estimates from the hidden semi-Markov model analysis. The median estimated duration of protection (adjusted for EIR and age) was 16.9\u201317.8\u2009days for AS-AQ in the trial sites with the lowest recorded 86Y and 76T prevalence (Bobo-Dioulasso and Gourcy in Burkina Faso), while it was 10.2\u201313.1\u2009days in the trial sites with the highest 86Y and 76T prevalence   Fig.\u00a0a, c. Conon) Fig.\u00a0b, d.FigPlasmodium falciparum transmission. Given the variation in prophylactic time between areas, we chose to use estimates from two of the trial sites with the most contrasting effects of the two drugs  and the pfcrt 76T mutation (25%), with a correspondingly long estimated median duration of protection by AS-AQ at 17.8\u2009days, approximately twice as long as the median duration of protection by AL in this site: 8.7\u2009days. Using the prophylactic profiles estimated in this trial site  it provides individual-level protection against reinfection and (b) prevention of reinfection reduces the total prevalence of infection in a population, and therefore onward transmission from infected individuals. Simulations comparing the public health impact of using either AL or AS-AQ as first-line drug were run using the existing individual-based age-structured mathematical model of ugs Fig.\u00a0. In the utation 2%, with autation 2%, with autation 2%, with autation 2%, with apfmdr1 86Y and pfcrt 76T were much higher at 69% and 95%, and the median duration of prophylaxis provided by AS-AQ was estimated at only 11.6\u2009days, while the median AL prophylactic time was 17.9\u2009days , the local prevalence of In both settings, there was minimal difference in impact on clinical episodes (<\u20091%) if we assumed that patients treated with AL were half as infectious as those treated with AS-AQ, compared with the scenarios where infectiousness was assumed to be equal after each treatment (results not shown). This is because even if there is some difference between treatments, both are estimated to have a high impact on gametocytes. Therefore, at a population level, transmission to mosquitoes is dominated by untreated infections which are thought to last on average about 6\u2009months, according to our model assumptions and parameters , 77, 78.pfmdr1 86Y and pfcrt 76T at the time of the trial, with AS-AQ providing better protection where wild type parasites with N86 and K76 genotypes were predominant, and AL performing better where 86Y and 76T mutants were common. This is consistent with previous studies demonstrating the collateral sensitivity of parasites with these different pfmdr1 and pfcrt genotypes to AL and AS-AQ. Our analysis extends previous work [In this analysis of clinical trials from 12 sites in Africa, we initially estimated that AS-AQ provided a slightly longer median duration of post-treatment prophylaxis than AL (15.2 versus 13.0\u2009days) when all data were pooled together. However, the duration of protection varied considerably between trial sites. In some locations, AS-AQ provided up to an estimated 19\u2009days of protection, ~\u20092-fold longer than AL, while in other trial sites the reverse was true, with AL providing up to 19\u2009days of protection, which was up to 1.5-fold longer than AS-AQ. This difference between sites appeared to be in part explained by the local prevalence of ous work , 79 by emdr1 86Y and crt 76T prevalence can have a public health impact, especially where malaria transmission is high and seasonal. We estimate that up to 14% of clinical episodes could be prevented in 0\u20135-year-old children by implementing first-line treatment with the drug providing optimal protection in a given setting, due to both individual protection from reinfection and population-level reduction in transmission . Countries with low (<\u200920%) or high (>\u200980%) prevalence of 86Y and 76T and intense transmission could consider the benefit of longer duration of protection if choosing between AL and AS-AQ policies. Using a first-line treatment with longer duration of protection is potentially a cost-effective way of reducing clinical cases and infections [Our transmission modeling suggests that the difference in duration of protection between the two drugs in areas with very low or very high fections .pfmdr1 86Y and pfcrt 76T mutations, initially driven through the parasite population by the previous widespread use of chloroquine, have been in decline in many parts of Africa. The decline has occurred fastest in countries using AL, consistent with the expected direction of selection [The election . The effelection . Our reselection , southerelection . Our reselection  and woulpfmdr1 86Y and pfcrt 76T mutations are associated with a longer time to reinfection after AL treatment and a shorter time after AS-AQ is consistent with a previous meta-analysis, where individual patient data on genotypes post-treatment were available [pfcrt and pfmdr1 haplotypes. Also, while we matched trials to the closest possible measures of mutation prevalence, these may not reflect the prevalence in the trial sites which can vary over space and time. We could not distinguish separate effects of 86Y and 76T in this analysis due to the close correlation of their prevalence. Other previous meta-analyses have examined the effect of dosing and other covariates on the probability of recrudescence after AL [Our finding that the vailable , 11. We after AL  and AS-Aafter AL . The tremdr1 and crt mutation prevalence, is reassuring.Our estimate of the mean duration of prophylaxis after AL at 13.0\u2009days is in good agreement with our previous estimate of 13.8\u2009days which was obtained from analysis of a completely different dataset of clinical trials in six sites in Africa  . When more than one molecular marker survey was matched to a trial site, a weighted average prevalence was taken. In some cases, these two molecular markers were assessed in the same matched survey(s), but in other cases matches from different surveys were found.Additional file 6: Figure S5. As Fig."}
+{"text": "Artemether-lumefantrine (AL) and artesunate-amodiaquine (ASAQ) are the first line therapy of uncomplicated malaria in Burkina Faso. We assessed the treatment efficacy, tolerability of these drugs 11 years following its adoption as first line treatment.In this opened randomized controlled trial carried out in 2016, participants with age over 6 months who consented to participate were randomly assigned treatment with artemether-lumefantrine or artesunate-amodiaquine and followed up for 28 days. Primary endpoint was the treatment efficacy over 28 days of follow up unadjusted by Polymerase chain reaction (PCR).Two hundred and eighty-one (281) participants were enrolled and the completion rate was 92.9%. No early treatment failure was found. Adequate clinical and parasitological responses were significantly higher in artesunate-amodiaquine group . On day 28, the risk of failure was 4 times higher in AL group 20.14%, 95% CI (13-30.47) against 5.16%, 95% CI (1.91-13.54) in ASAQ group. All treatments had a similar and good tolerability profile.Eleven years following artemether-lumefantrine and artesunate-amodiaquine adoption as first line therapy for uncomplicated malaria in Burkina Faso, artemether-lumefantrine retained fairly good efficacy even though its efficacy fell below WHO threshold of 90% considering uncorrected outcome. The cornestone of malaria treatment has been since the last fithteen years the artemisinin based combination therapy (ACTs) mainly artemether-lumefantrine and artesunate-amodiaquine. Procurement of ACTs faced severe shorted during the first five to ten years of its adoption. Currently, the procurement market has improved and ACTs are now widely distributed. Over 176 metrics tons of ACTs have been dispensed in endemic sub-Saharan Africa and Asia in 2017 and this demand is expected to growth up to 196 tons metrics in 2018, 205 MT in 2019 and 221 MT in 2020  or Asia Study site: the study was carried out in two peripheral health facilities, Colsama and Sakaby during the transmission season between June and December 2016. Each health facility was staffed by two to three nurses and one midwife. They were equipped with one outpatient department and one inpatient wall. All sites were located in Bobo-Dioulasso the second capital city of Burkina Faso at 365 km from Ouagadougou. Transmission of malaria is long but seasonal occurring over 6 months and containing at least 60-80% of clinical cases.Study participants: subjects presenting to the study health facilities with fever (axillary temperature \u2265 37.5\u00b0C) or history of fever in the previous 24 hours were screened against the following inclusion criterias in accordance with WHO guideline 2015: age at least 6 months, weight at least 5 kg, no evidence of concomitant illness, the provision of informed consent by the parents and the ability to participate in 28 days follow up, no history of antimalarial treatment within the last two weeks, no danger sign or evidence of severe malaria, P.falciparum mono infection, parasite density of 2000-200 000 parasites per \u00b5l of blood and hemoglobin \u2265 5 g/dL. During the screening process, participants satisfying the above criterias were enrolled while those who were excluded were referred to the health facility staff for standard care.Evaluation at entry and randomization: potential participants were assigned a number (ID), interviewed on past medical history, examined and then referred to the study nurses dedicated only to treatment allocation. Participants were randomly allocated to oral three days treatments based on a computer generated code provided by a staff not involving in the patient's evaluation. The treatment was either artemether-lumefantrine or artesunate-amodiaquine. The dosage was given on weight basis as per national malaria control program recommendation as this study is intented to evaluate the efficacy of first line treatments of malaria as implemented by the national malaria control program: (a) Artemether-lumefantrine (Coartem\u00ae Novartis administrated in tablet containing 20 mg of artemether plus 120 mg of lumefantrine); (b) artesunate-amodiaquine as given as tablet containing 67.5 mg plus 25mg respectively). The study as per its objectives is to inform straight away, it was not blinded to the study staff and patient. The treatment was administrated as DOT (directly observed treatment) and participants were retained for 30 minutes and in case of vomiting within this time period, a replacement dose was given. Any repeated vomiting led to the participant exclusion from further follow up and referral to health facility staff for standard care. Children presenting with hemoglobin less than 10 g/dL were treated according to the Integrated management of childhood Illness guidelines with ferrous sulfate for 14 days and anthelmintic treatment, if appropriate.Follow up and classification of treatment outcome: patients were asked to return to the study clinics on day 1,2,3,7,14,21 & 28 and at any time they were feel illness. Field workers visited those who failed to return to the clinics and each visit consisted on standard case report completion, physical examination, a finger pick for thin and thick blood smear. Blood smears were read to assess the parasite density and gametocytes. Patients were followed up for 28 days and their outcomes were assessed according to World Health Organisation (WHO) guidelines for antimalarial treatment efficacy  in AL group against 0.75%, 95% CI [0.11%-5.2%] in ASAQ group. On day 28, the risk of failure was 4 times higher in AL group 20.14%, 95% CI  in ASAQ group  (p = 0.1) .Treatment tolerability: over the study period, we found no serious adverse event. The registered adverse events were mild to moderate and none required active treatment or intervention's discontinuation. Common adverse events were weakness in either group. No pruritus or nausea were found  (ing day) . AL treaThis clinical trial was a study that is part of routine monitoring of first line therapies for uncomplicated malaria in Burkina Faso as per National Malaria Control Program recommendation. Eleven years following its introduction as first line therapies for malaria, AL and ASAQ remain effective in Burkina Faso even if efficacy of AL based on PCR uncorrected results was below 90% probably related with the fact that AL administration was done without fat food. Importantly, there is no early treatment failure or failures up to day 7; this reveled a good evidence that ACTs still work very well in this part of Africa and that currently the risk of tolerance to these first lines ACTs is limited. Also, all three daily doses were fully supervised by study team and a replacement was given once in case of vomiting. Previous studies in research settings have already reported good efficacy of AL and ASAQ in the central part of Burkina Faso , but alsAll treatment failures recorded were late after day 14. Late failures are mainly related to which extend the partner drug lasted in the bloodstream. Despite the absence of PCR correction, the relative short half-life of lumefantrine , 21 and This study completed in Burkina Faso as part of the regular monitoring of first line therapies of malaria has provided some up to date efficacy data for AL and ASAQ. The drugs retained a certain efficacy with good safety profile throughout follow up period. As of other studies, AL uncorrected results fall below WHO's threshold and merit more discussion to how to react in the context of limited available ACTs and the possibility to give a new life to some old drugs.Artemether-lumefantrine and Artesunate-Amodiaquine are first line therapies for malaria treatment in most countries and in Burkina Faso particularly;The apparition of resistant strains stressed the monitoring of drug efficacy.Overall these drugs remain effective even though the uncorrected efficacy is below the threshold of more than 90% recommended by the WHO;This study rises the discussion around the Polymerase Chain Reaction correction to consider the drug efficacy in real life.The authors declare no competing interests."}
+{"text": "Plasmodium falciparum in sentinel sites in Maferinyah and Lab\u00e9 Health Districts in Guinea in 2016.Anti-malarial resistance is a threat to recent gains in malaria control. This study aimed to assess the efficacy and safety of artesunate\u2013amodiaquine (ASAQ) and artemether\u2013lumefantrine (AL) in the management of uncomplicated malaria and to measure the prevalence of molecular markers of resistance of Plasmodium falciparum malaria in two sites. Children were followed for 28\u00a0days to assess clinical and parasitological response. The primary outcome was the Kaplan\u2013Meier estimate of Day 28 (D28) efficacy after correction by microsatellite-genotyping. Pre-treatment (D0) and day of failure samples were assayed for molecular markers of resistance in the pfk13 and pfmdr1 genes.This was a two-arm randomised controlled trial of the efficacy of AL and ASAQ among children aged 6\u201359\u00a0months with uncomplicated pfk13. All 9 observed pfk13 mutations were polymorphisms not associated with artemisinin resistance. The NFD haplotype was the predominant haplotype in both D0  and day of failure samples  successfully analysed for pfmdr1.A total of 421 participants were included with 211 participants in the Maferinyah site and 210 in Lab\u00e9. No early treatment failure was observed in any study arms. However, 22 (5.3%) participants developed a late treatment failure (8 in the ASAQ arm and 14 in the AL arm), which were further classified as 2 recrudescences and 20 reinfections. The Kaplan\u2013Meier estimate of the corrected efficacy at D28 was 100% for both AL and ASAQ in Maferinyah site and 99%  for ASAQ and 99% (97.1\u2013100%) for AL in Lab\u00e9. The majority of successfully analysed D0  and all day of failure  samples were wild type for This study observed high efficacy and safety of both ASAQ and AL in Guinea, providing evidence for their continued use to treat uncomplicated malaria. Continued monitoring of ACT efficacy and safety and molecular makers of resistance in Guinea is important to detect emergence of parasite resistance and to inform evidence-based malaria treatment policies. Worldwide, malaria represents a constant and persistent public health threat due to its morbidity and mortality especially among children , with suHowever, progress in providing effective treatment of malaria is facing challenges including parasite resistance to anti-malarial drugs, particularly in South East Asia . In respPlasmodium falciparum malaria and prevalence of artemisinin resistance molecular markers found high efficacy and safety of AL and no known artemisinin resistance pfk13 mutations  than for those in ASAQ arm 27,430 parasites/\u03bcl  in Maferinyah. In Lab\u00e9, median baseline parasite density was almost the same in both arms at 30,200 and 30,480 parasites/\u03bcl respectively. The median haemoglobin level of participants, was 9\u00a0g/dl for both arms in Maferinyah 10\u00a0g/dl for both arms in Lab\u00e9.Of the 421 included participants, 8 (1.9%) were lost to follow-up, including two deaths. This left 413 participants completing their follow-up. Treatment outcomes of participants who completed their follow-up are shown in Table\u00a0Of the 22 late treatment failures, 2 were classified as recrudescences and 20 were classified as reinfections  participants with 26 (44.1%) in the AL arm and 33 (53.9%) in the ASAQ arm. Two participants (one in the Maferinyah ASAQ arm and one in the Maferinyah AL arm) developed signs of severe malaria less than 24\u00a0h after inclusion in the study; one (in the Maferinyah AL arm) ultimately died. Both were excluded from analysis due to onset of severe symptoms less than 24\u00a0h after inclusion following WHO definitions. One additional participant in the Maferinyah ASAQ arm died from a car accident during follow-up, and was censored at day 14 in the analysis.The proportion of negative slides at D2 and D3 of follow-up is shown in Table\u00a0pfk13  were successfully amplified and sequenced for pfmdr1 gene at the 86, 184, and 1246 codons was successful in 380/443 samples (86%). The NFD haplotype was the predominant pfmdr1 haplotype at D0, present in 54% (197/362) of analysable D0 samples, followed by the NYD haplotype, present in 44% (158/362) of D0 samples. In late treatment failure samples, the NFD haplotype also predominated, at 61% (11/18).Amplification and sequencing of the This study marks the first round of anti-malarial resistance monitoring in Guinea, which has to date lacked regular and systematic surveillance since the introduction of ACT. The results showed high efficacy of ASAQ and AL (microsatellite-corrected D28 efficacies >\u200999%) to treat uncomplicated malaria, despite their use for more than a decade in Guinea. These results are consistent with the high ACT efficacies observed prior to introduction of ACT in Guinea and with other studies from Africa , 32, 33.pfk13 mutations associated with artemisinin resistance is further evidence that parasites remain susceptible to artemisinin derivatives in Guinea. This finding is similar to those studies in Africa [pfk13 mutations associated with artemisinin combination therapeutic resistance that have been identified in Southeast Asia [Rates of recurrent parasite density were relatively low in the Guinea study sites, below 8% in the 28-day follow up period. This low rate of recurrent parasite density may be explained by a high proportion of children <\u20095\u00a0years sleeping under an insecticide-treated net in Guinea as reported by the last demographic and health survey (2018) of the country . The resn Africa , 39\u201342 rast Asia , 43\u201345.pfmdr1 gene revealed that the majority of both pre-treatment samples and late treatment samples harbored the NFD haplotype, a molecular maker associated with reduced susceptibility of Plasmodium falciparum to lumefantrine [pfmdr1 haplotype is important.Sequencing of the fantrine , 47. As The current study only reports efficacy data from two sites in the country, and the results may not be representative of the whole country. Subsequent therapeutic efficacy studies from the remaining two sentinel sites in N\u2019Z\u00e9r\u00e9kor\u00e9 and Dabola health districts will further inform monitoring of anti-malarial resistance.This study found high efficacy and safety of ACT in Guinea, providing evidence that supports continued use of ASAQ and AL to treat uncomplicated malaria. Continued monitoring of ACT efficacy and safety and molecular makers of resistance in Guinea are important to detect parasite resistance and to inform evidence-based malaria treatment policies.Additional file 1: Table S1. Observed fragment lengths of neutral microsatellite loci from paired Day 0 (D0) and Day of Failure (DOF) samples from therapeutic efficacy studies in Guinea, 2016."}
+{"text": "Plasmodium vivax relapse prevention, when co-administered with 3-days of chloroquine or other blood schizonticide. Tafenoquine 200\u00a0mg weekly after a loading dose is also approved as travellers\u2019 prophylaxis. The development of tafenoquine has been conducted over many years, using various dosing regimens in diverse populations.Tafenoquine is an 8-aminoquinoline anti-malarial drug recently approved as a single-dose (300\u00a0mg) therapy for P. vivax relapse prevention is particularly examined.This review brings together all the preclinical and clinical data concerning tafenoquine central nervous system safety. Data were assembled from published sources. The risk of neuropsychiatric adverse events (NPAEs) with single-dose tafenoquine (300\u00a0mg) in combination with chloroquine to achieve P. vivax relapse prevention, nervous system adverse events, mainly headache and dizziness, occurred in 11.4% (36/317) of patients with tafenoquine (300\u00a0mg)/chloroquine versus 10.2% (19/187) with placebo/chloroquine; and in 15.5% (75/483) of patients with tafenoquine/chloroquine versus 13.3% (35/264) with primaquine (15\u00a0mg/day for 14\u00a0days)/chloroquine. Psychiatric adverse events, mainly insomnia, occurred in 3.8% (12/317) of patients with tafenoquine/chloroquine versus 2.7% (5/187) with placebo/chloroquine; and in 2.9% (14/483) of patients with tafenoquine/chloroquine versus 3.4% (9/264) for primaquine/chloroquine. There were no serious or severe NPAEs observed with tafenoquine (300\u00a0mg)/chloroquine in these studies.There was no evidence of neurotoxicity with tafenoquine in preclinical animal models. In clinical studies in P. vivax relapse prevention is favourable in the presence of malaria, with a low risk of NPAEs, similar to that seen with chloroquine alone or primaquine/chloroquine.The risk:benefit of single-dose tafenoquine/chloroquine in  In a chemoprophylaxis setting, a number of anti-malarial drugs have been associated with neurological or psychiatric adverse events (NPAEs), in particular, the quinoline-methanol derivative mefloquine \u20134. AlthoThe 8-aminoquinoline, tafenoquine, has been approved as monotherapy for travellers\u2019 prophylaxis, with an approved dose of 200\u00a0mg/day for 3\u00a0days followed by a 200\u00a0mg weekly maintenance doses thereafter , 7. TafePlasmodium vivax relapse prevention as a single-dose therapy (300\u00a0mg) in combination with the standard adult dose of chloroquine (1500\u00a0mg free base in staggered dosing over 3\u00a0days) [P. vivax relapse prevention required 3-days of chloroquine plus 14-days of primaquine 15\u00a0mg or 30\u00a0mg once daily. Primaquine is also an 8-aminoquinoline, but adherence to the dosing schedule is poor, and the clinical effectiveness of unsupervised primaquine is similar to that of placebo [P. vivax hypnozoites would potentially have a significant impact on reducing the burden of malaria and accelerating P. vivax elimination in endemic countries. This review considers the available preclinical and clinical safety data to assess the potential for tafenoquine to induce NPAEs in the context of findings for other anti-malarial drugs.Tafenoquine has also been approved for  3\u00a0days) \u201311. Prev placebo , 13. In  placebo \u201311. PrevNeurotoxicity is a complex area for preclinical investigation. In vitro studies of cytotoxicity against human neuroblastoma cells or similar models are suggestive of neurotoxic potential, but the relevance of such results depends on drug penetration into the CNS, as well as pharmacokinetic and other factors, for example, metabolic status. In animal models, cytotoxicity and morphological damage are obvious outcomes, but may be restricted to very specific brain areas. Neurological effects can also derive from purely functional disruption, with no morphological damage, for example, convulsions induced by fluoroquinolones \u201316. ThusFor older anti-malarial drugs, comprehensive neurotoxicological preclinical testing may not have been conducted. However, for new agents, supra-therapeutic or lethal drug doses are administered in animal models to attempt to induce observable lesions in CNS tissues. If lethal doses do not induce neurotoxicity, then the toxicities that caused death are deemed dose limiting. In such circumstances, a \u201csafety factor\u201d  is applied to the pharmacokinetic exposures in animals in order to predict the \u201chighest achievable exposure\u201d for humans. Consequently, the absence of neurotoxicity findings in pre-clinical animal models at supra-therapeutic or lethal doses is reassuring evidence that neurotoxicity in humans will be unlikely if the plasma concentrations in animals are at least tenfold greater than those achieved in humans following therapeutic or prophylactic dosing.P. falciparum malaria, as the artemether and dihydroartemisinin exposures exceeded the preclinical neurotoxicity safety margins [Even if drugs have the potential for neurotoxicity in animal models at supra-therapeutic doses, if the human dosing regimen does not achieve pharmacodynamically relevant exposures, and if the safety margins with CNS toxicity in preclinical species are acceptable, then NPAEs should not be expected. For example, although artemisinins are neurotoxic in animal models \u201321, the  margins . Neuroto margins . Note al margins . Thus, NThe MedDRA preferred term classification system is the standard method for assigning adverse events (AEs) to drug treatments used in clinical trials. Two system organ classes are relevant for neuropsychiatric risk: neurological disorders and psychiatric disorders Table\u00a0.Table\u00a01MNumerous papers of neuropsychiatric risk following anti-malarial treatment or during prophylaxis have been published, reaching various conclusions and promoting conflicting hypotheses . It is iA recent systematic meta-analysis of the published literature on \u2018mental and neurological manifestations\u2019 associated with anti-malarial drugs by Bitta et al.  providesIt can be seen immediately that there is no relationship between the prevalence of pooled NPAEs and the class of anti-malarial, except perhaps for the artemisinins, where prevalence is low for both artemether and artesunate. The lowest pooled prevalence of NPAEs was with the 8-aminoquinoline primaquine and the antibiotic dapsone and the highest was noted for the antibiotic minocycline and the 4-aminoquinoline amodiaquine. It can also be seen that for some drugs , the prevalence of events is far higher in prophylaxis studies than during malaria treatment, whereas for other drugs  there is little difference between prophylaxis and treatment, and for quinine, prevalence is higher during treatment than for prophylaxis. Thus, we can conclude that there is no clear pattern of predicting NPAEs based on class or clinical indication, but that the risk depends on the specific neurotoxic potential and pharmacokinetic/pharmacodynamic properties of the drug (i.e. whether toxicity is related to maximum drug concentrations or to cumulative exposure) under the different dosing regimens for treatment and prophylaxis.Plasmodium species, possess different immunity profiles, or have variations in transmission settings and socio-economic conditions that might affect non-drug related NPAE risk.The meta-analysis described above allows us to compare different drugs and identify outliers in treatment studies, which either by their nature or frequency, are suggestive of drug-related NPAEs. However, this introduces limitations in terms of making comparisons across different patient populations that may have different risk profiles for neuropsychiatric symptoms, be infected with different or multiple Malaria itself can produce varied and sometimes severe neuropsychiatric symptoms, the most serious being associated with cerebral malaria, including abnormal behaviour, impairment of consciousness, seizures, coma, or other neurologic abnormalities . HoweverP. vivax relapse prevention, new drug regimens are compared to placebo/chloroquine or placebo/artemisinin-based combination therapy (ACT) or the standard-of-care primaquine/chloroquine or primaquine/ACT. Thus, any adverse event signals outside the frequencies observed with these comparators would be concerning.In treatment studies, which are typically conducted across a single study population, the relative NPAE risk of a new anti-malarial versus the comparator is evaluated; the comparator is usually determined by the relevant National Malaria Treatment Guidelines. However, as most new anti-malarial drugs are administered in combination, the contribution of each component of the regimen to the incidence of adverse events cannot be determined. In the case of Prophylaxis studies remove the confounding effect of malaria symptomatology and allow a placebo-controlled comparison with a reference group. However, the dosing regimens and drug pharmacokinetics in prophylaxis are very different to those for treatment. In treatment, higher doses are generally given over a shorter time period, usually no more than 3\u00a0days, with the possible risk of adverse events related to maximum concentrations achieved. In contrast, prophylaxis generally involves lower doses regularly administered over a protracted time period, often months, with the risk of adverse events secondary to drug accumulation. For example, in the Bitta et al. meta-analysis outlined above, the difference between the NPAE risk for mefloquine in prophylaxis (25%) versus treatment 1%) was stark , we must first understand the underlying neuropsychiatric risk in military personnel versus civilians or in deployed versus non-deployed military personnel. If military personnel have a greater underlying risk for neuropsychiatric events than the general population, then trying to determine which NPAEs are drug-related, potentially over several months of treatment, becomes problematic, particularly if the underlying risk increases with deployment, i.e. over the same period that anti-malarial prophylaxis is taken.In general, examination of large military health databases indicates that serving military personnel are at greater risk of neuropsychiatric symptoms, particularly depression, post-traumatic stress disorder and other internalizing conditions than the civilian population \u201367. ThisThe activity\u2013structure relationships for the 8-aminoquinolines have been studied, with methyl substitution at position 4 of the quinoline ring conferring protection against neurotoxicity . As tafeSeveral pre-clinical studies to assess the potential CNS effects of tafenoquine have been conducted, including histopathological assessments in single- and repeat-dose studies in mice, rats and dogs, and detailed assessments of both neurobehavioural function and neurohistopathology in both single- and repeat-dose studies in rats . Across A comprehensive battery of preclinical neurotoxicological tests has been conducted on tafenoquine at supra-therapeutic and lethal doses in rats . BrieflyAlthough no pre-clinical neurotoxicity experiments were conducted in primates, several pharmacokinetic/pharmacodynamic studies of tafenoquine were conducted in Rhesus macaques. These studies were overseen by certified veterinarians and there was no evidence of any neurobehavioural disturbance or clinical evidence of neurotoxicity \u201379.P. vivax relapse prevention [P. vivax patients were randomized to tafenoquine 400\u00a0mg once daily for 3\u00a0days (N\u2009=\u200946) or standard 1500\u00a0mg (base) total dose chloroquine given over 3\u00a0days plus primaquine 15\u00a0mg daily for 14\u00a0days (N\u2009=\u200924) [Tafenoquine was previously investigated as monotherapy in evention . In a raN\u2009=\u200924) . This hi4 . This (N\u2009=\u200924) .Fig.\u00a02NeP. vivax relapse prevention examined tafenoquine doses of 300\u00a0mg/day for 7\u00a0days, 600\u00a0mg/day for 3\u00a0days and 600\u00a0mg single dose compared to chloroquine plus primaquine 15\u00a0mg/day for 14\u00a0days : a multicentre, phase 2b, double-blind, placebo-controlled, randomized, dose-selection study of single-dose tafenoquine  plus standard 3-day chloroquine versus placebo plus standard 3-day chloroquine or primaquine 15\u00a0mg for 14\u00a0days plus standard 3-day chloroquine .DETECTIVE Phase 3 (TAF112582 part 2): a multicentre, phase 3 pivotal, double-blind, double-dummy, randomized placebo-controlled clinical trial of single-dose tafenoquine 300\u00a0mg plus standard 3-day chloroquine versus placebo plus standard 3-day chloroquine or primaquine 15\u00a0mg for 14\u00a0days plus standard 3-day chloroquine .GATHER TAF116564): a multicentre, phase 3 supportive, double-blind, double-dummy, parallel group, randomized trial of single-dose tafenoquine 300\u00a0mg plus standard 3-day chloroquine versus primaquine 15\u00a0mg for 14\u00a0days plus standard 3-day chloroquine [6564: a mBased on the need to co-administer a blood schizonticide, three comparative clinical studies have been conducted with single-dose tafenoquine plus standard 3-day chloroquine in P. vivax relapse prevention in these studies. Firstly, patients were infected with P. vivax malaria and symptomatic. Secondly, tafenoquine was co-administered with standard 3-day chloroquine, and so evaluation of tafenoquine NPAEs is hampered by the absence of a true placebo arm with comparisons having to be made against either chloroquine alone or primaquine/chloroquine. Lastly, in all three clinical trials, P. vivax recurrence after day 29 resulted in retreatment with primaquine/chloroquine and so adverse events recorded after this point include adverse events associated with recurrence of malaria as well as those associated with the rescue therapy. As relapse was more frequent in the chloroquine alone group than with either of the 8-aminoquinolines, adverse events after day 29 were consequently more frequent with placebo/chloroquine than for the 8-aminoquinoline arms. Thus, only adverse events occurring before day 29 should be considered for comparison to the placebo/chloroquine arm.There are some key aspects that need to be highlighted in the assessment of NPAE risk with tafenoquine in To further investigate whether an NPAE signal might be present in the safety database from these studies, an expanded definition of NPAEs was applied. As well as including the standard terms included under \u2018nervous system disorders\u2019 and \u2018psychiatric disorders\u2019, the expanded definition added labyrinthitis, vertigo, vestibular disorder, asthenia, fatigue, and alcohol intolerance to include terms with possible reference to vestibulocochlear function, which has its seat in the brainstem. This was considered important, considering the eighth cranial nerve appeared to have been affected adversely by some earlier quinoline molecules , 83.P. vivax relapse prevention and so was conducted in malaria patients [The DETECTIVE 2b study was designed to select the clinical regimen for tafenoquine/chloroquine in patients . The mosDETECTIVE Phase 2b and DETECTIVE Phase 3 were placebo-controlled randomized studies designed as a seamless Phase 2b/3 programme, with similar protocols allowing data to be pooled for the single tafenoquine 300\u00a0mg therapeutic dose , 10. AllFor individual adverse events, only headache and dizziness occurred in more than 5% of patients, with a trend for a higher incidence of dizziness and lower incidence of headache with tafenoquine/chloroquine versus placebo/chloroquine, though 95% CIs overlapped in both cases Fig.\u00a0. PsychiaPrimaquine/chloroquine was included as a comparator in the DETECTIVE Phase 2b, DETECTIVE Phase 3 and GATHER studies \u201311. All As was seen for the comparison versus placebo, for both tafenoquine/chloroquine and primaquine/chloroquine, only dizziness and headache occurred in >\u20095% of patients, and with a similar prevalence for both comparators Fig.\u00a0. PsychiaWhen examining the frequency of all adverse events occurring over the 6-month study duration, there was no evidence of any increased risk for NPAEs with tafenoquine/chloroquine versus primaquine/chloroquine Table\u00a0. As woulP. vivax patients receiving tafenoquine/chloroquine and those receiving tafenoquine prophylaxis. Overall, 4131 tafenoquine-exposed subjects were included compared to 792 who received placebo/chloroquine or placebo [An analysis of all tafenoquine clinical safety data available to GlaxoSmithKline  was conducted to support review by the US Food and Drug Administration . This in placebo . CompariP. vivax patient; this individual had a history of depression. Three of the other four subjects with depression had no previous history of psychiatric illness or predisposing medical conditions. However, all four subjects with psychosis or suicidal behaviour had relevant previous psychiatric histories. Whether these events represented the underlying psychiatric conditions of the subjects or whether their symptoms were triggered or worsened by tafenoquine cannot be conclusively determined. Thus, for tafenoquine prophylaxis the precautionary recommendation is that the drug is not given to those with a history of psychotic disorders, or who currently have psychotic symptoms, including hallucinations , delusions , or disorganized thinking or behaviour [P. vivax relapse prevention, although it is recommended that a careful medical history be taken to identify any pre-existing mental health problems [The tafenoquine all studies safety database Table\u00a0, plus daehaviour . This isproblems .Table\u00a06DP. vivax relapse prevention, it is necessary to briefly discuss the prophylaxis studies as there are some features of these that should be explained in order to understand the safety signals for NPAEs at higher tafenoquine exposures.Although this review focuses on tafenoquine for In Table\u00a0A possible explanation for this increased reporting of NPAEs in ADF personnel is that soldiers experienced traumatic events, including danger of being killed or injured (71%); seeing dead bodies (49%); fear of exposure to a toxic agent, contagious disease, or injury (31%); and having a friend/associate killed or injured (30%) . AdditioThere was no evidence of neurotoxicity, neurobehavioural disorder, or clinical neurotoxicity with tafenoquine at supra-therapeutic doses in rodent and non-rodent models \u201379. ThesP. vivax relapse prevention, with single-dose tafenoquine (300\u00a0mg)/chloroquine, no serious or severe CNS events were reported with the observed events being mild-to-moderate and self-limiting [P. vivax malaria relapse prevention is anticipated to have a low risk of significant CNS effects [In limiting \u201311. Tafelimiting \u201311. Ther effects . HoweverP. vivax relapse by approximately 70% over 6\u00a0months\u2019 follow up compared with chloroquine alone, with a similar safety profile to primaquine [P. vivax relapse prevention and improve clinical effectiveness over the current 14-day primaquine regimen [P. vivax.In clinical trials, tafenoquine/chloroquine has been shown to reduce the risk of imaquine \u201311, 87.  regimen . TafenoqP. vivax relapse prevention is favourable. The potential of tafenoquine for both reducing the burden of P. vivax malaria and draining the hypnozoite malaria transmission reservoir represents the first major advance in the treatment and control of P. vivax since primaquine was introduced nearly 70\u00a0years ago.Coupled with advances in rapid quantitative diagnostics for screening of patients for glucose-6-phosphate dehydrogenase deficiency, a contraindication for 8-aminoquinoline therapy, the risk:benefit profile for tafenoquine in"}
+{"text": "CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017 compared with Artemether-Lumefantrine used as a positive control.  Considering the promising results of Phase I clinical trials with herbal medicine CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017 compared with Artemether-Lumefantrine used as a positive control. Methods. A single-blind randomized trial was conducted on 25 eligible males aged 18\u201340 years randomly assigned to two treatment groups: CoBaT-Y017 or Artemether-Lumefantrine. The first group received 35\u2009ml of CoBaT-Y017 in 1.5\u2009L mineral water administered daily for four consecutive days; the second group received oral Artemether-Lumefantrine, using WHO-recommended therapeutic dose regimens. For both drugs, efficacy for parasite clearance and safety were evaluated clinically, haematologically, and biochemically on days 1\u20134, 7, 14, 21, and 28. Clinical- and laboratory-adverse events (AEs) were recorded until day 28.  A single-blind randomized trial was conducted on 25 eligible males aged 18\u201340 years randomly assigned to two treatment groups: CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017 compared with Artemether-Lumefantrine used as a positive control. Methods. A single-blind randomized trial was conducted on 25 eligible males aged 18\u201340 years randomly assigned to two treatment groups: CoBaT-Y017 or Artemether-Lumefantrine. The first group received 35\u2009ml of CoBaT-Y017 in 1.5\u2009L mineral water administered daily for four consecutive days; the second group received oral Artemether-Lumefantrine, using WHO-recommended therapeutic dose regimens. For both drugs, efficacy for parasite clearance and safety were evaluated clinically, haematologically, and biochemically on days 1\u20134, 7, 14, 21, and 28. Clinical- and laboratory-adverse events (AEs) were recorded until day 28. Results. 13 and 12 patients were randomized into CoBaT-Y017 arm and Artemether-Lumefantrine arm, respectively. In all patients, parasitaemia was adequately neutralized with CoBaT-Y017 group patients' parasite clearance lagging slightly behind that of Artemether-Lumefantrine's group, but without a statistically significant difference . Physical and laboratory examinations did not show any significant changes in vital signs, biochemical, and haematological parameters. In the Artemether-Lumefantrine arm, 100% (12/12) of patients experienced, at least, one adverse event versus 61.5% (8/13) in the CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017 compared with Artemether-Lumefantrine used as a positive control. Methods. A single-blind randomized trial was conducted on 25 eligible males aged 18\u201340 years randomly assigned to two treatment groups: CoBaT-Y017 or Artemether-Lumefantrine. The first group received 35\u2009ml of CoBaT-Y017 in 1.5\u2009L mineral water administered daily for four consecutive days; the second group received oral Artemether-Lumefantrine, using WHO-recommended therapeutic dose regimens. For both drugs, efficacy for parasite clearance and safety were evaluated clinically, haematologically, and biochemically on days 1\u20134, 7, 14, 21, and 28. Clinical- and laboratory-adverse events (AEs) were recorded until day 28. Results. 13 and 12 patients were randomized into CoBaT-Y017 arm and Artemether-Lumefantrine arm, respectively. In all patients, parasitaemia was adequately neutralized with CoBaT-Y017 group patients' parasite clearance lagging slightly behind that of Artemether-Lumefantrine's group, but without a statistically significant difference . Physical and laboratory examinations did not show any significant changes in vital signs, biochemical, and haematological parameters. In the Artemether-Lumefantrine arm, 100% (12/12) of patients experienced, at least, one adverse event versus 61.5% (8/13) in the CoBaT-Y017 arm.  13 and 12 patients were randomized into CoBaT-Y017, a Phase II study was conducted with Plasmodium falciparum malaria-infected patients, for efficacy and safety evaluation of CoBaT-Y017 compared with Artemether-Lumefantrine used as a positive control. Methods. A single-blind randomized trial was conducted on 25 eligible males aged 18\u201340 years randomly assigned to two treatment groups: CoBaT-Y017 or Artemether-Lumefantrine. The first group received 35\u2009ml of CoBaT-Y017 in 1.5\u2009L mineral water administered daily for four consecutive days; the second group received oral Artemether-Lumefantrine, using WHO-recommended therapeutic dose regimens. For both drugs, efficacy for parasite clearance and safety were evaluated clinically, haematologically, and biochemically on days 1\u20134, 7, 14, 21, and 28. Clinical- and laboratory-adverse events (AEs) were recorded until day 28. Results. 13 and 12 patients were randomized into CoBaT-Y017 arm and Artemether-Lumefantrine arm, respectively. In all patients, parasitaemia was adequately neutralized with CoBaT-Y017 group patients' parasite clearance lagging slightly behind that of Artemether-Lumefantrine's group, but without a statistically significant difference . Physical and laboratory examinations did not show any significant changes in vital signs, biochemical, and haematological parameters. In the Artemether-Lumefantrine arm, 100% (12/12) of patients experienced, at least, one adverse event versus 61.5% (8/13) in the CoBaT-Y017 arm. Conclusion. CoBaT-Y017 exhibited similar antimalarial efficacy against P. falciparum to that of Artemether-Lumefantrine, with good tolerability and safety.P. falciparum The majority of antimalarial drugs have been derived from medicinal plants or made of chemical structures modelled on plant lead compounds.Plasmodium parasite's resistance against earlier antimalarials is now widespread. Moreover, artemisinins, currently the most potent and fast-acting antimalarial [CoBaT-Y017 was ranked as the leading antimalarial phytomedicine without marketing authorization. The second step was a preclinical study evaluating CoBaT-Y017 acute and subacute toxicity. Satisfactorily results obtained led to Phase I study to establish the safety and tolerability of CoBaT-Y017 and providing some indications on the antimalarial efficacy of the drug [Lessons from the past prove that progress towards \u201cMalaria Elimination\u201d entails both a judicious stewardship of existing treatments and the development of a steady stream of new drugs , 2. Plasmalarial , are assmalarial , 5. Thismalarial , 7 highlmalarial . Furthermalarial , to reduthe drug .CoBaT-Y017 efficacy and safety with the standard first-line treatment, AL.The aim of this study, as the third step of our \u201creverse pharmacology\u201d approach, was a randomized controlled trial to compare the CoBaT-Y017 is an herbal medicinal product used by the population for uncomplicated malaria self-treatment. It is presented in the form of coffee-brown syrup accommodated in 70\u2009mL bottles in a package including a measuring cup of 10\u2009mL, manufactured by COPHARBIOTECH Ltd., a Beninese local company which develops some pharmaceutical products of category 2 according to the WHO phytomedicines classification. CoBaT-Y017 has been formulated from two herbals mixture: Mentha piperita and Cinnamomum zeylanicum.CoBaT-Y017 efficacy and safety versus AL. It was conducted between March and September, covering the period June\u2013August, the peak transmission season. All the activities were performed at the internal medicine department of the Teaching hospital, Abomey-Calavi/S\u00f4-Ava.The study was consisted in a Phase II randomized, single-blind clinical trial in patients with uncomplicated malaria to evaluate P. falciparum detected by microscopy ; have axillary temperature higher than 37.5\u00b0C or history of fever during the past 24\u2009h; having consumed no antimalarial within the previous 14 days; been treated with any medical or herbal drug within the previous 7 days; being able to swallow oral medication; have no chronic pathology; have no gastrointestinal intolerance to oral medication, including nausea, vomiting, and/or diarrhea; have no allergy to CoBaT-Y017 and AL; have not undergone any substance abuse in the past 30 days; have not consumed any alcohol in the past week; being able and willing to comply with the protocol and visit the schedule of the study for its entire duration and sign an informed consent form.The following inclusion criteria were adopted: male gender; aged between 18 and 40; currently infected only by The following exclusion criteria were adopted: clinical or laboratory signs of severe malaria according to the WHO definition during the study ; severe On admission, patients were fully examined; blood samples were taken for full blood-cell count and routine blood biochemistry, confirmed of malaria by microscopy. Those patients who fulfilled the inclusion criteria were enrolled. The enrolled volunteers were randomized into one of two treatment arms. The investigator physicians and laboratory staff did not know the medical treatment administered to the patients, while the patients, the randomizer, and the nurse in charge of administering the drugs knew the exact treatment.The patients of both study arms were orally treated.CoBaT-Y017 arm, 35\u2009mL of CoBaT-Y017 diluted in 1.5\u2009L of mineral water was administered each day for 4 consecutive days.For the AL arm, patients were treated with oral AL, Bimalaril\u00ae80/480 (tablets), using the WHO-recommended therapeutic dose regimens [For the regimens .The study nurse supervised all the treatments and monitored the participants for 30\u2009min for adverse reactions or vomiting following intake. Patients with fever \u226538.5\u00b0C were treated with paracetamol.All participants were hospitalized during the four-day treatment and the fifth day to ensure strict monitoring. After five days in hospital, patients were medically checked at days 7, 14, 21, and 28, to ensure full recovery without complications while recording any adverse event. At each medically check, a clinical examination was performed, a parasitological evaluation according to the WHO protocol  was condThick and thin blood smears were performed before treatment (day 1) and days 2 to 5, 7, 14, 21, and 28 for both groups. Slides, using Giemsa stain, were examined through a microscope to determine parasite species and density according to the WHO protocol .\u00b5l) was calculated assuming a white blood cell count of 8000/\u00b5l. All slides underwent independent counts by two qualified microscopists, and the average of the two counts was taken as the parasite density. Parasite counts with discordant results  were re-read by a third microscopist, and parasite densities were calculated by averaging the two closest counts. Density was calculated using the following formula:Parasite density  and alanine transaminase (ALT), creatinine, and urea. All symptoms and adverse events were recorded. Incidence of all adverse events was scored from 1 to 5 , according to the Common Terminology Criteria for Adverse Events (CTCAEs) . AdverseCoBaT-Y017 for a noninferiority study. Assuming that the risk of erroneously rejecting an effective substance is limited to \u03b2\u2009\u2264\u20090.05, the size NA of one of the two arms of Phase II satisfies the relation: NA size\u2009\u2265\u2009ln 0.05/Ln (1-p), NA\u2009\u2265\u20096. Consequently by doubling the size, two cohorts (n\u2009=\u200912 or 13) were recruited.The sample size was calculated by determining the minimum size required for a 60% efficacy of P value\u2009\u2264\u20090.05 was considered as significant.Treatment efficacy was determined with the calculations of the proportion of positive thick blood film and the parasite density variation over time (follow-up days) in each treatment group using parasitaemia determined by microscopy. Graphical displays of the Kaplan\u2013Meier estimates for parasite clearance time and fever clearance time were generated. Chi-squared test or Fisher's exact tests were used for comparing differences in categorical data. A http://www.tga.gov.au/docs/pdf/euguide/ich/ich13595.pdf). All samples were coded with an ID number. All participants gave written informed consent and had the choice to withdraw from the study at any time.This study was approved by the Research Ethics Committee of Applied Biomedical Sciences Institute (CER-ISBA) of Cotonou with the reference no. N103 du 09/01/17. The approval was renewed by the Ethics Committee on August 20, 2018. The study was conducted in accordance with the Declaration of Helsinki principles the International Committee of Harmonization Good Clinical Practice Guidelines (n\u2009=\u20092) and herbal or traditional medicine product intake within the last 7 days (n\u2009=\u20094) and hyperparasitaemia (n\u2009=\u20092). The median age was 26 years (range 18\u201340 years), and the median weight was 60\u2009kg. Patients in both treatment groups had normal blood pressure at baseline with mean systolic arterial blood pressures of 11.61\u2009\u00b1\u20091.32 in the CoBaT-Y017 group and 10.75\u2009\u00b1\u20090.68 in the AL group; the mean diastolic pressures were 7.54\u2009\u00b1\u20091.27 in the CoBaT-Y017 group and 6.75\u2009\u00b1\u20090.62 in the AL group, respectively. All patients were feverish at the start of treatment with an average axillary temperature of 38.84\u2009\u00b1\u20091.06 and 38.5\u2009\u00b1\u20090.90 in the CoBaT-Y017 group and AL group, respectively.Among 33 males screened as potential study volunteers, 25 were deemed eligible and finally enrolled in two treatment arms . ReasonsCoBaT-Y017 became negative 24 hours after administration versus 25% of those for AL patients. After 48 hours following the administration, 76.1% thick blood films were negative for the CoBaT-Y017 arm versus 66.7% in the AL group. However, 72 hours after administration (day 3) 100% of the thick blood films were negative for the AL group against 84.6% for the CoBaT-Y017 group. Two patients (15.4% of treatment group) diagnosed at day 3 as coinfection with Salmonella sp. had their thick blood film positive. Upon co-treatment of the salmonellosia (15.4%), those two patients had their parasitaemia canceled on day 5, while it was already cleared in all other participants for both arms at day 3. The decline of the positive thick blood films reflected the reduction of the parasite density in the treatment arms. The decrease of parasite density is rendered by the decreasing number of positive thick blood films (53.8% thick blood films for patients treated with od films .CoBaT-Y017 had a slightly longer delay in parasite clearance (delayed parasite clearance) compared with those of the AL group, without a statistically significant difference . The parasite clearance in the two treatment arms is statistically best illustrated by the Kaplan\u2013Meier estimator . However, a slight haemoglobin increase was noted in the AL arm at D28, but this was not followed up due to the termination of the study. As for the mean absolute value of leukocytes, a nonsignificant increase was observed on D3, D14, and D21 in CoBaT-Y017 patients, while in the AL patients, this value remained quasi constant before and after treatment . After treatment. no statistically significant changes in these values were observed in both study arms during the follow-up days. Likewise. the values of these hepatic parameters remained within the normal limits for both study arms (reference range 12\u201342\u2009IU/L and 10\u201348\u2009IU/L for AST and ALT. respectively) until day 28. CoBaT-Y017 like AL did not alter the patients hepatic's functions  were within normal limits. our study subjects having not experienced any liver dysfunction prior to treatment. However. the baseline values of AST and ALT transaminases of patients randomized in unctions .P=0.32) with CoBaT-Y017 patients when compared with those of AL. Besides this. no statistically significant changes (P > 0.05) were observed during the follow-up time. The mean values of urea remained almost constant before and after treatment in both arms of the study (P > 0.05). The values of the renal parameters' remained within the normal limits in both study arms; neither CoBaT-Y017 nor AL altered the renal function of the patients  were within normal limits; patients of both arms had no renal dysfunction. In relation to creatinine. a nonsignificant increase was observed on D4 (patients .AL arm. 100% (12/12) of patients experienced. at least. one adverse event. while in the CoBaT-Y017 arm. only 61.5% (8/13) of patients have experienced adverse events. Headaches were the most reported adverse events in both treatment arms. with 66.7% and 30.8% for the AL and CoBaT-Y017. respectively; the second most frequent adverse event reported by 41.7% of patients only in the AL group was fatigue. Similarly. insomnia 8.3% (1/12). abdominal pain 8.3% (1/12). stiffness 16.7% (2/12). and rash 8.3% (1/12) were reported only in the AL arm. Adverse events reported only in the CoBaT-Y017 arm include increased appetite 15.4% (2/13). vomiting 15.4% (2/13). and nausea 15.4% (2/13) (In the % (2/13) .CoBaT-Y017 arm at D4. whereas persistence of the same was observed in some patients (8.3%) in the AL arm until D7.Spontaneous regression of adverse events was observed in the According to the CTCAE. all recorded adverse events recorded were categorized as grade 1 except headache reported as categorized grade 2 in both arms. No serious adverse event was reported in either treatment arm.According to information on hand. very few studies have been conducted on the validation of phytomedicines used for uncomplicated malaria . While hCoBaT-Y017 can be considered active and therefore be retained. More than one subject responded to this treatment. All thirteen patients enrolled in CoBaT-Y017 arm were completely cured after four days of treatment.The aim of this study was to determine. on as a few subjects as possible. whether or not the CoBaT-Y017 useful in the therapeutic arsenal. This allowed us to estimate the response rate observed with a good level of precision.Our investigation product. herbal medicine. was compared with a well-established first-line treatment recommended by the National Program of Malaria Control in Benin and by the WHO allowing to determine CoBaT-Y017 and AL with adequate clinical parasitological responses (APCRs) at day 28. No significant difference was observed for parasite clearance and rate of parasite regression between CoBaT-Y017 and AL groups (P=0.85) (The results showed a significant parasitaemia decrease in patients treated with (P=0.85) .Argemone mexicana decoction. an antimalarial phytomedicine for which a second-line treatment was not required for 89% of patients versus 95% of patients on AS-AQ [Similar results were obtained in another study. 87.9% APCR for PR 259 CT1. an antimalarial phytomedicine versus 96.9% APCR for Artesunate-Amodiaquine combination . Our reson AS-AQ . For theCoBaT-Y017 as the leading antimalarial phytomedicine used in Benin for uncomplicated malaria self-treatment and that of the Phase I study [This finding supports the clinical information on the most used antimalarial phytomedicines which was ranked  I study .CoBaT-Y017 that we used revealed the presence of six classes of chemical compounds including. alkaloids. flavonoids. and terpenes. Phenochemicals such as alkaloids. terpene. and its derivatives have been proven to be involved in the anti-Plasmodium activity of many plants [It should be pointed out that the phytochemical screening of the batch of y plants . 15. 16.After 28 days of follow-up. we did not observe any significant differences between the mean values of the different biological parameters in the two treatment groups.P > 0.05). Moreover. for the mean absolute value of leukocytes. a nonsignificant increase was observed on D3. D14. and D21 in CoBaT-Y017 patients. which probably conferred it. some immunostimulatory properties.For haemogram parameters. the mean value of haemoglobin. red blood cell. and platelet remained almost constant before and after treatment in both arms of the study . while some adverse events were still recorded with some AL group patients (8.3%) until D7. Headache was reported as the most frequent adverse event in both study arms but with a higher proportion in the AL group 66.7% for AL versus 30.8% for CoBaT-Y017. Fatigue was only recorded in the AL arm. which comes after the headaches in terms of frequencies. Similar data were obtained in another comparative study in which headache accounted for a higher proportion of adverse events followed by fatigue in patients treated with AL [More adverse events have been reported in the  with AL .The main interest of this work was that we started from the realities of field. remedy used informally. and without marketing authorization. to appreciate its safety and tolerability  and thenCoBaT-Y017 would be due to the synergic effect of its components. Further studies are planned to identify active compounds. which could be used as markers and demonstrate the mechanism(s) of action. That should be the next step of our simplified approach called \u201creverse pharmacology\u201d [The efficacy of acology\u201d  to develCoBaT-Y017 has a proven antimalarial efficacy and has good haematological. hepatic. and renal tolerance. The benefit exhibited by CoBaT-Y017 at the end of a 28 day follow-up outweighs by far the risk it presents. when compared with artemisinin-based combination therapies or other antimalarial phytomedicines available on the Benin market.We can conclude that"}
+{"text": "Plasmodium falciparum infection in sentinel sites in the Benguela, Zaire, and Lunda Sul Provinces were treated with artemether-lumefantrine (AL) or artesunate-amodiaquine (ASAQ) and monitored for 28\u2009days to assess clinical and parasitological responses. Molecular correction was performed using seven microsatellite markers.Biennial therapeutic efficacy monitoring is a crucial activity for ensuring the efficacy of currently used artemisinin-based combination therapy in Angola. Children with acute uncomplicated  Plasmodium falciparum infection in sentinel sites in the Benguela, Zaire, and Lunda Sul Provinces were treated with artemether-lumefantrine (AL) or artesunate-amodiaquine (ASAQ) and monitored for 28\u2009days to assess clinical and parasitological responses. Molecular correction was performed using seven microsatellite markers. Samples from treatment failures were genotyped for the pfk13, pfcrt, and pfmdr1 genes. Day 3 clearance rates were \u226595% in all arms. Uncorrected day 28 Kaplan-Meier efficacy estimates ranged from 84.2 to 90.1% for the AL arms and 84.7 to 100% for the ASAQ arms. Corrected day 28 estimates were 87.6%  for the AL arm in Lunda Sul, 92.2%  for AL in Zaire, 95.6%  for ASAQ in Zaire, 98.4%  for AL in Benguela, and 100% for ASAQ in Benguela and Lunda Sul. All 103 analyzed samples had wild-type pfk13 sequences. The 76T pfcrt allele was found in most  ASAQ late-failure samples but in only 16% (4/25) of AL failure samples. The N86 pfmdr1 allele was found in 97% (34/35) of treatment failures. The AL efficacy in Lunda Sul was below the 90% World Health Organization threshold, the third time in four rounds that this threshold was crossed for an AL arm in Angola. In contrast, the observed ASAQ efficacy has not been below 95% to date in Angola, including this latest round.Biennial therapeutic efficacy monitoring is a crucial activity for ensuring the efficacy of currently used artemisinin-based combination therapy in Angola. Children with acute uncomplicated  Plasmodium falciparum, and the cornerstone of malaria case management is prompt diagnosis and treatment with artemisinin-based combination therapy (ACT). Three ACTs are used as first-line treatments in the public health sector in Angola: artemether-lumefantrine (AL), artesunate-amodiaquine (ASAQ), and dihydroartemisinin-piperaquine (DP).Malaria is a major public health concern in Angola, with over 7 million cases registered in 2019. Most cases are caused by Antimalarial resistance has been a continuous threat to malaria control throughout the globe. The emergence of artemisinin resistance in Southeast Asia  presentsWhile initial pre-ACT-era trials in Angola showed 100% efficacies of AL and ASAQ , routine7\u2013pfk13 mutations associated with artemisinin resistance have been detected in Angola either as part of therapeutic efficacy monitoring or in separate molecular surveys  had the N86 allele present  entered the study . Twenty The rate of day 2 slide negativity was 68%  in the AL arm in Lunda Sul but was above 80% in all other arms . Day 3 sThe uncorrected Kaplan-Meier estimates of the day 28 efficacy were 84.2%  for the AL arm in Zaire, 84.7% (78 to 92%) for ASAQ in Zaire, 85.3% (79 to 93%) for AL in Lunda Sul, 90.1% (85 to 96%) for AL in Benguela, and 100% for the ASAQ arms in Benguela and Lunda Sul . After mpfk13  of AL day-of-failure samples had this allele. All (21/21) AL and most  ASAQ day-of-failure samples had the N86 pfmdr1 allele.All 103 sequenced samples from patients with early treatment failure at day 0, and day 0 and day-of-failure samples from participants with recurrent parasitemia, were wild type for pfk13 . Nearly For the third time in four rounds of Angolan therapeutic efficacy monitoring, one of the AL arms was observed to have a corrected efficacy below the key 90% threshold set by the WHO. While previous rounds have shown suboptimal AL efficacy in Zaire, the AL efficacy in Zaire for this round was 92.2%. In contrast, the AL efficacy in Lunda Sul for this round was 87.6%, compared to 96.5% in 2017. Consistent with previous findings, the ASAQ efficacy was uniformly high (above 95%) in all three sites.Recent reports of AL efficacy estimates below 90% from the Mikalayi site in the Democratic Republic of the Congo (DRC), less than 500 km from Lunda Sul and the closest efficacy monitoring site to Lunda Sul, provide additional context of potential warning signs for AL in the region. Moreover, the low day 28 AL efficacy in Lunda Sul in this study was observed in the backdrop of a relatively low rate of day 2 clearance, as only 68% of participants were microscopy negative on day 2, compared to 97% in the same site in 2017. Although day 3 slide negativity, the indicator used by the WHO to assess the ability of artemisinin to clear initial parasitemia, was 95% in this arm, the day 2 clearance rate is a notable finding. Finally, while the relatively short half-life of lumefantrine  is a likpfk13 sequences. The results of analyses of pfcrt and pfmdr1 sequences were consistent with literature showing an overrepresentation of the 76T pfcrt allele in amodiaquine treatment failures and a predominance of the N86 pfmdr1 allele in AL treatment failures. Interpretation of the latter finding should be done in the context of previous reports of the high population prevalence of the N86 allele in parasites circulating in Angola and the high prevalence of the N86 allele even in ASAQ treatment failures in this study. Molecular characterization of all day 0 samples, not just failures, will allow a better understanding of the population prevalence of these markers. Notably, fixation or near fixation of the N86 allele, together with the clinical evidence of reduced efficacy, might have implications for the future of AL use in Angola.There was no molecular evidence of artemisinin resistance, as all failure samples had wild-type The unavailability of quality-controlled DP precluded an assessment of its efficacy in this round of monitoring in Angola. The recent introduction of new prequalified formulations should ensure that DP returns to being evaluated in future rounds. The inability to directly observe evening AL doses means that lumefantrine drug level measurement might be warranted, as in previous rounds , especiain vivo protocol  or ASAQ  for 3 days. The dosage was weight based according to the manufacturers\u2019 recommendations. All ASAQ doses were directly observed by study staff and were administered with water or juice. For AL, morning doses were observed by study staff and were administered with yogurt or milk. Parents or guardians were given the evening dose and a yogurt or milk packet to give at home. Study staff telephoned parents or guardians in the evening to remind them to administer the evening dose, and compliance was further assessed by requesting parents or guardians to bring empty blister packets to the clinic the following day.Children were monitored daily for the first 4 days and then weekly thereafter for a total of 28\u2009days. At each visit, study staff performed clinical exams, performed blood smears (except for day 1), and collected blood on Whatman 903 filter paper . Hemoglobin was assessed fortnightly using HemoCue Hb301  or DiaSpect  hemoglobinometers.Blood smear microscopy was done using thick and thin smears  by two mP. falciparum, fragment lengths of the microsatellite markers were measured using an ABI3130 xl genetic analyzer  capillary sequencer and GeneMapper software . Day 0 samples from early treatment failures (before day 7) and day 0 and day-of-failure samples from cases of recurrent parasitemia (on or after day 7) were analyzed using a targeted deep-amplicon-sequencing workflow that included the full pfk13, pfcrt, and pfmdr1 genes  at National Institute of Health laboratories in Angola. Extracted DNA was transported to U.S. Centers for Disease Control and Prevention (CDC) Malaria Branch laboratories in Atlanta, GA. Molecular analysis was conducted by visiting Angolan study investigators under the President\u2019s Malaria Initiative-supported Antimalarial Resistance Monitoring in Africa Network capacity-building program . Paired r1 genes .Data were double entered into an Excel database . Baseline characteristics were summarized by treatment group. The proportions of participants with negative slides on day 2 and day 3 were calculated. Individuals finishing follow-up were classified as early treatment failures, cases of recurrent parasitemia, or cases of adequate clinical and parasitological response (ACPR) according to standard WHO definitions . For molThe study was reviewed by the human subjects review boards at the Angolan Ministry of Health, and the activity was approved as a nonresearch program evaluation by the Office of the Associate Director for Science at the CDC\u2019s Center for Global Health (protocol 2014-233c). Parents or guardians of study participants provided written informed consent.The full clinical data set has been uploaded to the WorldWide Antimalarial Resistance Network and WHO repositories. Full microsatellite genotyping data are available in Tables S1 and S2 in the supplemental material."}
+{"text": "Here, the persistence of ring-stage parasitaemia following ACT and non-ACT treatment was examined.Malaria control in sub-Saharan Africa relies upon prompt case management with artemisinin-based combination therapy (ACT). Ring-stage parasite mRNA, measured by Samples were used from naturally infected Malian gametocyte carriers who received dihydroartemisinin\u2013piperaquine (DP) or sulfadoxine\u2013pyrimethamine (SP\u2013AQ) with or without gametocytocidal drugs. Gametocytes and ring-stage parasites were quantified by qRT-PCR during 42\u00a0days of follow-up.day 14\u2009=\u20090.011 and pday 28\u2009=\u20090.068). No association of ring-stage persistence with gametocyte carriage was observed.At baseline, 89% (64/73) of participants had measurable ring-stage parasite mRNA. Following treatment, the proportion of ring-stage parasite-positive individuals and estimated densities declined for all four treatment groups. Ring-stage parasite prevalence and density was generally lower in arms that received DP compared to SP\u2013AQ. This finding was most apparent days 1, 2, and 42 of follow-up (p\u2009<\u20090.01). Gametocytocidal drugs did not influence ring-stage parasite persistence. Ring-stage parasite density estimates on days 14 and 28 after initiation of treatment were higher among individuals who subsequently experienced recurrent parasitaemia compared to those who remained free of parasites until day 42 after initiation of treatment (psbp1 as ring-stage marker. Lower persistence of ring-stage mRNA after ACT treatment suggests the marker may not reflect dormant parasites whilst it was predictive of re-appearance of parasitaemia.The current findings of lower ring-stage persistence after ACT without an effect of gametocytocidal partner drugs affirms the use of  Plasmodium falciparum malaria. Artemisinin-based combinations consist of an artemisinin derivative that rapidly reduces parasite burden and a partner drug with a longer half-life that clears remaining parasitaemia and provides prophylactic activity for weeks post-treatment. At present, artemisinin derivatives retain excellent efficacy in most of Africa despite decreased sensitivity to some of its partner drugs [Malaria is a leading cause of global morbidity and mortality. In 2018, an estimated 228 million cases and 405,000 malaria-associated deaths were reported worldwide [er drugs . Whilst er drugs , it is ier drugs .sbp1) were reported following ACT treatment [Plasmodium falciparum 3D7 parasites suggests that dormant parasites can be induced by artemisinin monotherapy and provides a plausible explanation for recrudescences [Post-treatment detection of parasite DNA may reflect (remnants of) asexual parasites and gametocytes , 8, the reatment , 11. Thireatment  which poreatment , 14 and escences . PublishThe aim of this study was to expand on earlier observations by examining ring-stage parasitaemia among trial participants that were followed for 42\u00a0days after being randomized to ACT or non-ACT anti-malarials with and without gametocytocidal drugs. This study allowed examination whether persisting ring-stage parasitaemia persists after administration of non-ACT, and whether the persistence of ring-stage parasites is associated with parasite recrudescence and/or continued gametocyte production.Ethical approval for the study was granted by the Ethics Committee of the Faculty of Medicine, Pharmacy, and Dentistry of the University of Science, Techniques, and Technologies of Bamako , the Committee on Human Research at the University of California San Francisco , and the Research Ethics Committee of the London School of Hygiene & Tropical Medicine .P. falciparum mono-infection, between 5 and 50\u00a0years of age, who were glucose-6-phosphate dehydrogenase (G6PD)-normal by CareStart G6PD rapid diagnostic test , had a hemoglobin concentration of\u2009\u2265\u200910\u00a0g/dL, and had a P. falciparum gametocyte density of\u2009\u2265\u20092 gametocytes/500 white blood cells by thick film microscopy. Participants were excluded if they had a serious or chronic illness , weighed 80\u00a0kg or more, reported anti-malarial use within 7\u00a0days of screening (artemether\u2013lumefantrine and artesunate\u2013amodiaquine being first-line treatments), or reported allergies to study drugs.This study used samples obtained from participants of a trial in Ou\u00e9less\u00e9bougou, Mali who were randomized 1:1:1:1 to receive either sulfadoxine\u2013pyrimethamine (SP\u2013AQ), SP\u2013AQ with single low-dose primaquine (SP\u2013AQ\u2009+\u2009PQ), dihydroartemisinin-piperaquine (DP), or dihydroartemisinin\u2013piperaquine with methylene blue (DP\u2009+\u2009MB) .\u00a0EligiblDetails on the study procedures are described in the original paper . In briesbp1 mRNA transcript using previously described methods [PfMGET and female Pfs25 mRNA transcripts as described elsewhere [P. falciparum parasites on days 7, 14, 28, and 42, PCR genotyping of\u00a0glurp (glurp2),\u00a0msp2 (Fc27), msp1  and lc1 alleles were performed on samples obtained at enrollment and day of post-treatment failure. Pre- and post-treatment pairs were analysed and classified as either recrudescent, re-infection, or indeterminate infections according to World Health Organization guidelines [Ring-stage parasites were quantified by qRT-PCR targeting the  methods . The lim methods , the limlsewhere . For samidelines .The sample size for this study was dictated by the original clinical trial, with 20 individuals per study arm to allow a within-person change in infectivity after treatment . It is ahttp://www.r-project.org/). Comparisons between proportions were conducted using Chi-squared or Fisher\u2019s exact test and Mann\u2013Whitney tests were used to compare differences in parasite densities, unless otherwise specified. Correlations between ring-stage and gametocyte parasite densities were assessed by Spearman\u2019s rank correlation coefficient using the log10 transformed versions of these variables. Generalized estimating equations were used to compare SBP-1 parasite prevalence and density between SP\u2013AQ and DP arms, accounting for repeated observations for individuals. Log-binomial regression was used to model relative risk ratios and linear regression was used to model mean differences in log10-transformed SBP-1 parasite density. Models adjusted for baseline log10-transformed SBP-1 parasite density. An interaction term between treatment and follow-up visit was included to assess whether participants of the DP group cleared parasitaemia at a more rapid rate than SP\u2013AQ. An overall F-test was used compute the p-value testing joint effect of the interaction terms. Log-binomial regression models were used to assess whether age, weight, treatment arm, or baseline SBP-1 parasite density were associated with persistent parasitaemia on day 7 post-treatment. All tests were two-sided with alpha\u2009=\u20090.05. p-values\u2009<\u20090.05 were considered statistically significant.All analyses were performed using Stata 14.0  and R  males from the original trial   (Table DP \u00d7 time\u2009=\u20090.005).Following anti-malarial treatment, the prevalence and density of ring-stage parasites reduced across all four treatment arms Fig.\u00a0, 3. RingDespite anti-malarial treatment, ring-stage parasite RNA was still detectable across all days of follow-up and all treatment arms Table . On day Spearman rank correlation tests were used to assess whether the persistence of ring-stage parasites was associated with later gametocyte density, which would be indicative of ongoing gametocyte production. Among those who had persistent ring-stage parasitaemia on day 7 (n\u2009=\u200914), no significant correlation was observed between their ring-stage parasite density on day 7 and gametocyte density on days 14 and 28   participants experienced recurrent parasitaemia detectable by microscopy . Conventional genotyping of polymorphic MSP-1, MSP-2 and GLURP genes  indicateP. falciparum carriage, ring-stage parasite mRNA was detected up to 42\u00a0days after anti-malarial treatment. Estimated densities of post-treatment ring-stage parasites reduced at a more rapid rate following receipt of ACT compared to non-ACT, arguing against the hypothesis that this signal reflects dormant parasites that tolerate can artemisinin treatment. This could, however, also be obscured by differences in treatment efficacy between DP and SP\u2013AQ. In this modestly sized population, few individuals experienced an episode of recurrent parasitaemia (n\u2009=\u200910), but these individuals tended to harbour higher ring-stage parasite densities prior to recurrence than those who were successfully treated.In this study of young Malian males with asymptomatic sbp1 ring-stage transcripts following ACT treatment in more detail [Whilst repeated assessments of parasite density shortly after initiation of treatment provide the most conclusive evidence on (changes in) parasite responsiveness , alternae detail , 11. Pree detail , 11 or ae detail . Along we detail  where pae detail . The stuFrom a public health perspective, it is important to examine whether persisting ring-stage parasites are predictors of recrudescent infections or the source of gametocyte production. The current study population was small and as the original study objective was to assess gametocyte clearance and infectivity , not allThe longer period of follow-up in this study also allowed the exploration of associations of ring-stage persistence with subsequent gametocyte carriage. Whilst data collection was not specifically designed for this, it was possible to relate ring-stage densities with gametocyte densities 7 and 14\u00a0days later, roughly the period needed for gametocyte production , and obsThe study is subject to several limitations. First, it is recognized that sample size was small, which may have limited the statistical power of the study. Thus, future studies  may be needed to confirm findings. Second, the study population consisted mostly of young males with high gametocyte densities at enrollment, which may limit the generalizability of findings to other parasitized populations. The study population is not representative of individuals with uncomplicated malaria; the average duration of infection will have been longer for the current study population who all had blood-stage infections sufficiently long to complete the 8\u201312\u00a0day maturation period of gametocytes . This wiIn summary, the current findings indicate that ring-stage parasites may persist at low concentrations following anti-malarial treatment. Whilst this parasite population is unlikely to reflect dormant parasites following artemisinin treatment, the association of ring-stage persistence with subsequent detection of recurrent parasitaemia by microscopy warrants further studies to examine whether they may reflect viable parasite populations that may recrudesce when the concentrations of anti-malarials become permissive during follow-up.Additional file 1: Table S1. Prevalence of SBP-1 on days of follow-up by treatment arm. Table S2. SBP-1 Parasite density on days of follow-up by treatment arm. Table S3. Recrudescence-reinfection genotyping."}
+{"text": "Plasmodium falciparum population.The World Health Organization recommends the provision of intermittent preventive treatment during pregnancy (IPTp) with sulfadoxine-pyrimethamine (SP) at 4-week intervals from gestational week 13 to delivery in areas of moderate to high malaria transmission intensity. However, the effect of IPTp-SP has been compromised in some areas due to parasite resistance, raising the importance of parasitological and chemoprophylactic surveillance, and monitoring SP-resistance markers in the P. falciparum population.Between November 2013 and April 2014 in Nchelenge, Zambia, 1086 pregnant women received IPTp-SP at antenatal-care bookings. Blood samples were collected on day 0, and on day 28 post-treatment to test for malaria parasites and to estimate SP parasitological efficacy in the treatment and prevention of parasitaemia. A random sample of 96, day 0 malaria-positive samples were analysed to estimate the prevalence of SP-resistance markers in the The overall parasitological and prophylactic failure among women who had paired day 0 and day 28 blood slides was 18.6% . Among pregnant women who had asymptomatic parasitaemia on day 0, the day 28 PCR-uncorrected parasitological failure was 30.0%  and the day 28 PCR-corrected parasitological failure was 15.6% . Among women who tested negative at day 0, 12.3%  developed parasitaemia at day 28. Among the 96 malaria-positive samples assayed from day 0, 70.8%  contained the DHPS double (Gly-437\u2009+\u2009Glu-540) mutation and 92.7%  had the DHFR triple (Asn-108\u2009+\u2009Ile-51\u2009+\u2009Arg-59) mutation. The quintuple mutation (DHFR triple\u2009+\u2009DHPS double) and the sextuple mutant (DHFR triple\u2009+\u2009DHPS double\u2009+\u2009Arg-581) were found among 68.8%  and 9.4%  of samples, respectively.The parasitological and chemoprophylactic failure of SP, and the prevalence of resistance markers in Nchelenge is alarmingly high. Alternative therapies are urgently needed to safeguard pregnant women against malarial infection. Plasmodium falciparum infection and related consequences that include stillbirth [Pregnant women in malaria-endemic areas are at high risk of illbirth , 2, smalillbirth , 4, pretillbirth , and lowillbirth . Pretermillbirth \u201310. The illbirth . IPTp-SPillbirth , 13. Theillbirth . Even thillbirth \u201321.This observational cohort study was conducted in Nchelenge, a rural northern district in the Luapula Province with a population of 173,680 . The obj\u00ae filter paper. Thick smear-slides and dried blood spots were, respectively used to diagnose malaria by microscopy and polymerase chain reaction (PCR). Participants received directly observed IPTp-SP during the same consultation. The follow up visit was conducted on day 28 post-treatment during which peripheral blood was collected for malaria diagnosis by microscopy and PCR. Participants verbally confirmed having not taken any anti-malarial treatment between day 0 and day 28.Details of enrolment procedures including inclusion and exclusion criteria have been previously reported . BrieflyIn alignment with the WHO treatment efficacy protocol , sample P. falciparum was conducted using a nested-PCR method as described by Snounou et al. [Laboratory staff stained thick blood films using 10% Giemsa which were then read by two independent microscopists. Details of slide-reading methods have been reported elsewhere . Parasitu et al.  with modu et al. .\u00ae Dream Taq PCR Master Mix (2X) and 0.5\u00a0\u00b5M of each primer and 2\u00a0\u00b5l of template. A 2\u00a0\u00b5l of the primary amplicon was used as a template in the secondary reaction. Negative and positive controls were included in every batch of sample processing, from extraction to amplification, and finally electrophoresis. Secondary PCR amplicons were analysed by gel electrophoresis to confirm amplification and band intensity before enzyme digestion. Restriction-digest assays were set up following manufacturer instructions; 4\u00a0\u00b5l of amplicon was used as substrate in the reaction mix. For samples showing faint bands in the nested PCR product, 6\u20138\u00a0\u00b5l was used as substrate. Amplicon and restriction fragments were analysed on ethidium bromide 2% agarose gels and visualized under ultraviolet transillumination on a Biosens (Genescope V1.76) digital imaging system.A sample size of 96 was needed to detect a 50% prevalence of quintuple mutation among pregnant women with 95% CIs \u00b1 10%. Parasite DNA template extracted for malaria detection by PCR was used for this part of the study. Nested PCR and restriction enzyme digestion methods were both used to detect antifolate drug resistance polymorphisms in the DHFR and DHPS genes . BrieflyIn vivo parasite-clearance efficacy and prophylactic effect of SP was established based on malaria microscopy  and mero\u00ae Dream Taq PCR Master Mix (2X) and 0.5\u00a0\u00b5M of each primer and 2\u00a0\u00b5l of template. A 2\u00a0\u00b5l of the primary amplicon was used as a template in the secondary reaction.Polymorphic regions of MSP2 were amplified by nested PCR. Primary PCR primers corresponding to the conserved sequence flanking this region  were useThe secondary amplicon from each sample was then analysed using electrophoresis on 2% ethidium bromide stained agarose gels. Samples from individual participants were loaded in adjacent lanes. In cases where there was no amplification, PCR was repeated using five-times the quantity of template DNA. In cases where no amplicon was detected after the second reaction, amplification was classified unsuccessful . Two indData were double-entered in EpiData version 3.1 software , cleanedParasitological failure was defined as presence of parasites on day 0 and day 28 post treatment that was deemed to be recrudescence and not a new infection. Prophylactic failure was defined as the presence of parasitaemia in previously aparasitaemic women on day 28 post-treatment, as well as parasitaemia deemed to be a new-infections in women who tested positive at day 0 and day 28. Overall parasitological and prophylactic failure was defined as the inability of SP to clear existing parasites and to prevent infection within 28\u00a0days of administration.Baseline characteristics among asymptomatic parasitaemic women were compared between paucigravidae (primi- and secundigravidae combined) and multigravidae using appropriate tests, namely Chi-squared test for proportions, t-tests for means and Mann Whitney for medians. Prevalence estimates of the DHFR triple, DHPS double; DHFR\u2009+\u2009DHPS quintuple and sextuple mutants and their 95% CIs were then calculated.Parasitological and prophylactic failure in paucigravidae and multigravidae women was compared using Chi-squared test, while the median age of women who experienced failure compared to those who did not was assessed with the Mann Whitney test.P\u2009<\u20090.001. Use of bed nets on the previous night was significantly higher among multigravidae than paucigravidae, respectively, 42.7% versus 27.5%, P\u2009=\u20090.003. Parasite count measured by geometric mean was lower in multigravidae compared to paucigravidae, 849 versus 1310, P\u2009=\u20090.001. Other characteristics of asymptomatic parasitaemic women in this cohort stratified by gravidity are shown in Table A total of 1086 ANC attendees were enrolled from November 2013 to April 2014. Among them, 343 women had asymptomatic parasitaemia at day 0  were lost to follow-up. The proportions of socio-demographic factors such as age, gravidity and marital status, did not differ between the lost to follow-up group and those who were retained with the exception of the number of years of schooling. Among women who were lost to follow-up, 54.4% reported spending 7\u00a0years and above in school versus 65.5% among those who were retained at day 28, The prevalence of malaria detected by polymerase chain reaction and microscopy have been reported elsewhere and were 57.8% (621 of 1074) and 31.8% (343 of 1079), respectively. The prevalence of submicroscopic malaria infection (microscopy negative but PCR positive) was 38.9% (285 of 731).Among women who tested positive for malaria on day 0 measured by microscopy, 60.3% 207 of 343) were tested again at day 28 post-treatment. Sulfadoxine-pyrimethamine cleared parasitaemia present on day 0 in 70.0% (145 of 207) of pregnant women confirmed by day 28 test  were also screened at day 28. Of these women, 12.3% (47 of 383) became malaria parasite positive at day 28 post-treatment  than multigravidae13.4% (37 of 277), P\u2009<\u20090.001. Younger women were more likely to experience parasitological failure. The median age of women who experienced parasitological failure was 19\u00a0years  compared to 22\u00a0years among those whose parasitaemia cleared , P\u2009<\u20090.001. The median age of women who experienced prophylactic failure was 24  years while that of women who maintained their malaria negative status was 26  years, P\u2009=\u20090.027.The proportion of parasitological failure observed among paucigravidae was much higher than that observed among multigravidae, 22.4% (26 of 116) versus 6.7% (6 of 89), P. falciparum positive samples. Mixed infection of Asn-108 occurring with the wild type Ser-108 was observed in 3.1% (3 of 96) of samples. High levels of the DHFR Ile-51 and Arg-59 mutants as shown in Fig.\u00a0There was near saturation of the pyrimethamine-resistant DHFR Asn-108 mutation at 94.8% (91 of 96) among the Overall, almost one-fifth (18.6%) of pregnant women in the study experienced a parasitological and prophylactic failure of SP at day 28 post-IPTp treatment in an area where 68.8% of malaria parasites carried the quintuple mutant and 9.4% expressed the sextuple mutant. Sulfadoxine-pyrimethamine retained partial efficacy in parasite clearance among pregnant women who had asymptomatic malarial infections. Similar observations have been made elsewhere despite a moderate to high prevalence of the quintuple mutant and presence of the sextuple mutant , 37.The risk of parasitological failure was higher among paucigravidae than multigravidae. This may be partially attributable to primigravidae and secundigravidae having acquired less semi-immunity to malarial infection in pregnancy than multigravidae , 38, 39 It is difficult to establish the precise prophylactic failure rate because it is unknown how many women who tested negative by microscopy at day 0 may have later been exposed to malarial infection post-treatment. This is because some women may have become infected after receiving IPTp-SP, cleared the parasitaemia to reflect important prophylactic effect but were indistinguishable from other women who may have never have been exposed to the parasite. Nonetheless, calculating the prophylactic failure based on the number of malaria positive women from those who tested negative at day 0 is a reasonable and common proxy for the estimate of true prophylactic failure, especially in areas with moderate to high malaria transmission like this study area .Only MSP-2 genotyping was conducted to distinguish recrudescence from reinfection in the current study. Although genotyping for genes MSP-1, MSP-2 and GLURP was not done in this study, MSP-2 results alone are reliable since the gene is known to provide a more accurate measure of treatment outcomes compared to MSP-1 and GLURP .Point mutations associated with SP resistance were very common in this study. The prevalence of both the quintuple and the sextuple mutants among pregnant women was moderate but higher than recorded in earlier studies, both of which were conducted in the same province, Luapula . The firOnly four studies conducted in Zambia have estimated the prevalence of SP resistance markers, two in the general population , 45 and The prevalence of submicroscopic malaria infection among pregnant women in the current study was considerable. Other studies have found wide spread submicroscopic malaria infections \u201349. ManyThe limitations in this study include the fact that samples were only collected at day 0 and day 28. These collection time points do not allow for determining early parasitological failure. Secondly, samples were not collected from the six women who verbally indicated that they had taken anti-malarial drugs before day 28, therefore requiring their exclusion from treatment-failure analysis at day 28. Another limitation is the fact that some of the women who tested negative on day 0 could have had submicroscopic infections which became positive by day 28 which may have affected the estimate of prophylactic failure.The number of women who were lost to follow-up was large. However, baseline characteristics did not differ between women lost to follow-up and those who were followed to day 28, with one exception: the number of years of schooling. This could have affected the results in terms of proportions of parasite clearance and parasitological failure if women lost to follow-up are different from those who were retained at day 28. However, given that these women are residents of the same geographic area where high malaria endemicity affects the community broadly , it is dThe quintuple and sextuple mutants were observed in this study population. Sulfadoxine-pyrimethamine retains partial efficacy in clearing parasites in pregnant women with moderate prevalence of the highly resistant quintuple mutation. These results suggest that continued monitoring is essential for future policy-making and there is a clear need to identify alternative regimens for use in IPTp."}
+{"text": "The prevalence of asymptomatic parasitemia was lower in the SP+AS compared to control (38 (7.5%) vs. 143 (28.7%); and 47 (12.7%) vs. 75 (21.2%); p < 0.002) in 2007 and 2008, respectively. Hemoglobin concentration was significantly higher in children receiving SP+AS  than in control children  in 2007, 2008, and 2009, respectively. No impact on clinical malaria was observed. Conclusion: IPTsc with SP+AS reduced the rates of all-cause clinic visits and anemia during a three-year implementation.Previous studies have shown that a single season of intermittent preventive treatment in schoolchildren (IPTsc) targeting the transmission season has reduced the rates of clinical malaria, all-cause clinic visits, asymptomatic parasitemia, and anemia. Efficacy over the course of multiple years of IPTsc has been scantly investigated. Methods: An open, randomized-controlled trial among schoolchildren aged 6\u201313 years was conducted from September 2007 to January 2010 in Kolle, Mali. Students were included in three arms: sulphadoxine-pyrimethamine+artesunate (SP+AS), amodiaquine+artesunate (AQ+AS), and control (C). All students received two full doses, given 2 months apart, and were compared with respect to the incidence of clinical malaria, all-cause clinic visits, asymptomatic parasitemia, and anemia. Results: A total of 296 students were randomized. All-cause clinic visits were in the SP+AS versus control (29 (20.1%) vs. 68 (47.2%); 20 (21.7%) vs. 41 (44.6%); and 14 (21.2%) vs. 30 (44.6%);  Intermittent preventive treatment (IPT) for malaria in children has recently been identified as an important tool in the fight against malaria, particularly in areas where malaria transmission follows a distinct seasonal pattern ,2,3. IPTIn 2009, we reported the results of a randomized controlled trial of artemisinin-based combination therapies (ACTs) used for intermittent preventive treatment in school-aged children (IPTsc) . The triIn areas of seasonal malaria transmission, IPTsc targeting the transmission season has decreased the rates of clinical malaria, asymptomatic malaria, all-cause clinic visits, anemia, and school absenteeism, but the efficacy of ACTs in the context of longitudinal IPTsc has been scantly investigated. Our study sought to understand the effects of longitudinal IPTsc among Malian schoolchildren. We hypothesized that IPTsc would continue to positively impact malaria morbidity reduction among school-aged children in an area with primarily seasonal transmission.Trial design: This open, randomized controlled trial was performed among schoolchildren aged 6\u201313 years in Kolle, Mali. The study began in September 2007 and completed follow-up in January 2010. This three-armed study compared two artemisinin combination-based seasonal chemoprevention therapies (SP+AS or AQ+AS) against no seasonal chemoprevention therapy (control). Seasonal chemoprevention consisted of two doses given eight weeks apart, with the first dose administered at the beginning of back to school during the high seasonal malaria transmission. Children were followed monthly throughout the transmission season with physical exams, blood smears, and hemoglobin levels. Changes in the initial design included a protocol amendment for the no therapy arm. In the first year, the control (no therapy) arm received a single dose of vitamin C. In the second and the third years, the control arm only received water. This recapitulated a more accurate and pragmatic comparison group.Plasmodium falciparum during the dry season is between 40% and 50%, which increases to 70\u201385% during the rainy season [Participants: Kolle is a rural village of 3000 inhabitants located 57 km southwest of Bamako . The study clinic was staffed 24 h a day, 7 days a week during the study follow-up period, serving as the only medical facility in the village. Based on our own epidemiological surveillance data, malaria is hyperendemic during the short transmission season from July to November. The prevalence of y season . Inclusi\u00a9, Hemocue Inc., Brea, CA, USA). Blood smears were read by two independent certified readers. Conflicting results entered arbitration and were read by a third certified reader. This was considered the final read. Clinical malaria cases were defined as fever and constitutional symptoms  and the presence of parasites on the smear. Asymptomatic parasitemia was defined as study subjects not having signs of illness but having at least one parasite per 100 fields on microscopy. Anemia was defined according to the World Health Organization (WHO) [Interventions: At the beginning of the study period, permission from village elders was obtained at a public meeting. Over a 5-day period, a village \u201ccrier\u201d was used to recruit interested volunteers. The study was explained to children and their parents or guardian in the local language. Upon receiving the informed consent, students were assigned a computer-generated random number assigning them to one of three study arms\u2014SP+AS arm: SP 25/1.25 mg/kg/day in a single dose plus artesunate 4 mg/kg once for 3 days; AQ+AS arm: amodiaquine 10 mg/kg/day plus artesunate 4 mg/kg/day for 3 days; control arm: vitamin C 250 mg tablet given once for 3 days in the year 2007. In the follow-up years 2 (2008) and 3 (2009), the control arm did not receive anything except water. Children remained in the same assigned study arm for all three years of the study. After randomization and arm assignments, children received an initial history and physical examination, where inclusion and exclusion criteria were reviewed. Any student with signs or symptoms of illness were treated according to the national guidelines. Finger-prick blood was collected for the preparation of thick and thin blood smears and the measurement of hemoglobin concentration (HemoCueon (WHO) . All-cau\u00ae) in years 2 and 3. Cases of severe malaria, defined by WHO criteria in 2000, were treated with quinine. Clinicians provided supportive care and treatment for all other diagnoses according to national guidelines.Outcomes: The primary outcome was the number of episodes of clinical malaria. Secondary outcomes were asymptomatic parasitemia, hemoglobin concentration, all-cause clinic visits, and school performance, as observed by their classroom teacher. In order to measure these endpoints, the team performed monthly follow-up visits for the 5 months following the initial treatment dose. Follow-up visits included history, physical examination, and finger-prick blood collection (parasitemia and hemoglobin concentration measurement). Students received a second dose of study medication 8 weeks after the initial course. In addition to the active monthly follow-up, participants were instructed that they should come to the clinic for any illness. All unscheduled visits by participants to the clinic during the study period were documented. Children with signs and symptoms of malaria underwent thick and thin smears and hemoglobin testing. In years 1, 2, and 3, students with clinical malaria were treated with the same IPTsc treatment. Students in the control arm received SP monotherapy in year 1 and were treated with artemether-lumefantrine  of participant study numbers and corresponding medication group. As children presented for enrollment, they were assigned a study number in the numerical order of presentation within a given grade. Participants were stratified by grade level such that there were approximately equal numbers of each study arm represented. Grade level was defined by the success of the new classroom. We had 6 classrooms . School grade average was the annual mean obtained by the sum of the all-month grade divided by the total number of months. Clinicians were aware of drug assignment at the time of initial randomization. Data were double entered using Microsoft ACCESS , and the statistical analysis was performed using R software. Baseline characteristics of the volunteers among treatment arms versus control were compared using the Fisher Exact test and Chi-square tests for categorical variables, Student for continuous variables, and the Mann\u2013Whitney U-test/Wilcox test for non-normally distributed continuous variables. The McNemar\u2019s test and the Paired sample t-test were used before and after each treatment arm. For binomial outcome, a weighted logistic regression was used for multivariate analysis , and for continuous outcomes . Statistical significance was set at p < 0.05.Randomization sequence generation and statistical methods: Participants were randomized using a computer-generated shuffle , Mali. Project Identification Code: 59_FMPOS.Out of a total of 475 students aged 6 to 13 years in the village, 305 were screened, and 296 were enrolled. The total loss to follow-up was two (0.7%) students at the end of the initial trial period in January 2008 . One stuBaseline characteristics, including age, hemoglobin concentration, and parasitemia, were similar between the study arms at the start of the study in 2007 . Cohort p < 0.001; 21.7%, p = 0.005; and 21.2%, p = 0.02 in the SP+AS arm and 32.6%, p = 0.009; 33.7%, p = 0.14; and 33.7%, p = 0.56 in the AQ+AS arm in 2007, 2008, and 2009, respectively , 36 (39.1%), and 15 (22.7%) had malaria confirmed by thick smear, respectively, in 2007, 2008, and 2009. There were no cases of severe malaria. The control arm experienced the most episodes of all-cause clinic visits, with 47.2%, 44.6%, and 44.6% during the season compared to 20.1%, ectively . Childrep < 0.001). In November, we observed an initial downward trend in parasite load for groups receiving IPT, with a subsequent convergent spike at 8 weeks post-initial IPT dose (p > 0.05). In December and January, this difference was significantly decreased after the second dose in SP+AS and AQ+AS arms compared to the control arm (p < 0.001). After the first dose in year 2 (2008), parasite prevalence was significantly decreased in the SP+AS arm only (p = 0.003). In November, the peak occurred at 4 weeks post-intervention (rather than 8 weeks post-intervention) in all three arms (p > 0.05). In December, this difference was significantly decreased in SP+AS and AQ+AS arms compared to the control arm (p = 0.003), and no difference was observed in January. In year 3 (2009), the beginning of the seasonal chemoprophylaxis dosing was postponed due to a delayed school start. Thus, volunteers began the season with higher initial parasite loads. IPT lowered parasite loads for the SP+AS arm (p = 0.007), but not the AQ+AS arm (p = 0.47) compared to the control arm; a slight rise in parasitemia rates was also noted. Besides, the significant decrease of parasite prevalence in the control arm compared to the AQ+AS arm (p = 0.04) before treatment was also noted. By the end of the season , however, all arms had lower parasite loads than at any point during the season. Overall, volunteers receiving SP+AS trended towards the lowest rate of parasite load compared to the AQ+AS and control arms at the end of the season, though this was not a statistically significant finding (p > 0.05) .Parasite detection in years 2 and 3 varied from our initial observations in year 1 of the trial . After t > 0.05) . Plasmodp < 0.001), and it significantly decreased in M4 and M5 (p < 0.001) compared to M1. Globally, PI was 48%, 36%, and 16% in 2007, 2008, and 2009, respectively. The difference was significantly decreased from 2007 to 2009 (p < 0.001) but not from 2008 to 2009 (p > 0.05) .Plasmodic index (PI) by month (M) for all the years was 14%, 29%, 30%, 15%, and 12% in month 1 (M1), M2, M3, M4, and M5, respectively . There was a slight increase in parasite prevalence in M2 and M3 (p = 0.012), significantly increased in M3 (p = 0.002), and significantly decreased in M4 and M5 (p < 0.001). For the AQ+AS arm, PI was significantly increased in M2 and M3 (p = 0.007 and p = 0.01) and decreased in M4 and M5 (p = 0.01) compared to M1 (p < 0.001) (p > 0.05). In M2, the PI was significantly different in the SP+AS and AQ+AS vs. control (p < 0.004), but this difference disappeared in M3 (p > 0.05) (p < 0.001).In ed to M1 . For the< 0.001) . In M1,  > 0.05) . This dip < 0.001 (p = 0.002 but not AQ+AS arm 62 (16.2%), p = 0.08 vs. control arm 75 (21.2%). No difference was observed between any of the three arms in 2009, p > 0.05  and AQ+AS arm 60 (11.7%) versus Control arm 143 (28.7%),  < 0.001 . In 2008p > 0.05 p > 0.05, p < 0.001) but increased in the AQ + AS arm compared to the control arm (p = 0.007) (p < 0.001) (p > 0.05), but this difference was significantly decreased from 2007 to 2009 (p < 0.001)  . In M3, < 0.001) . Globall< 0.001) .p = 0.001 versus control 42.1%, 44.6% and 25.3% in 2007, 2008, and 2009, respectively, and AQ + AS 30.6%, p < 0.001 and 35.5%, p = 0.02 vs. control 42.1% and 44.6% in 2007 and 2008, respectively, p = 0.04 and AQ+AS arm vs. control arm in 2007, p < 0.002; p = 0.001). At the end of the study, the control arm continued to have higher rates of anemia; this difference was significant in the SP+AS arm in 2007 (p = 0.01) and not significant in other years. From the first month to the last month, the incidence of anemia was significantly decreased in the SP+AS arm in 2007 and 2009 (p < 0.006), in the AQ+AS arm in 2009 (p < 0.003), and in the control arm in 2009 (p < 0.01), excluding January    , and AQ+ = 0.46)  vs. contp > 0.05, in 2007, 2008, and 2009, respectively. The success of different treatment arms was higher in all years (> 77.9%) (p > 0.05), respectively  and in the AQ+AS arm  was compared to the control arm ; > 77.9%) . The sucectively . The grap > 0.05 .p = 0.001 and p = 0.04, respectively. Breathing problems were significantly increased in the SP+AS arm (1.4%) vs. control arm (0.5%), p < 0.001 (p > 0.05 (Table not shown). No students elected to quit the study because of these events.Adverse events were defined as fever, headache, abdominal pain, respiratory symptoms, vomiting, diarrhea, lethargy, and pruritus. The most frequent were fever, headache, abdominal pain, and respiratory symptoms. Fever and headache were significantly decreased in the SP+AS arm  vs. control arm (2.7% and 3.8%);  < 0.001 . Across School-aged children suffer consequences of malaria but are generally neglected in malaria control strategies. One control strategy under investigation is IPT in school children (IPTsc) . IPT witOur results suggest that IPTsc administration of SP+AS would benefit schoolchildren living in areas with high malaria endemicity.In this controlled, randomized trial of school-aged Malian children living in a high-transmission setting, IPTsc with either SP+AS or AQ+AS was superior to control in reducing malaria incidence (year 1), all-cause clinic visits, anemia, and asymptomatic parasitemia , though Protection against anemia and all-cause clinical visits was observed in the SP + AS arm, but asymptomatic parasitemia appeared to be short-lived, though comparisons varied by year and regiment. Although each treatment arm was superior to the control one month after the first dose, this difference was lost after two months, suggesting that protection waned between 4 to 8 weeks post-treatment. For IPT, a drug should eliminate circulating parasites and persist at sufficient levels to prevent the multiplication of parasites acquired between doses. This time frame is consistent with previous findings of an IPT trial in Ghanaian infants, in which protection waned between 5 and 6 weeks post-treatment . In our Previous studies have shown that SP is highly efficacious for the treatment of malaria, with substantial post-treatment prophylactic efficacy in West Africa ,20,21. HSP resistance associated with AQ in SMC is a reality , while sThis study detailed the effects of longitudinal IPTsc with SP+AS and AQ+AS arms. During year 1, the effects were quite dramatic. While there continued to be a positive downward effect on the number of cases of malaria and asymptomatic parasitemia, the effect size declined over the two following years. This is likely attributable to the fact that the follow-up group consisted of older children, on average, as no new children were added to the study after initial enrollment. Another explanation could be that there were fewer children carrying parasite loads as much of the population had received IPT the years prior. Moreover, the declining efficacy could also be due to an overall decrease in transmission intensity in the area.Our most notable challenges were financial and political issues that arose during our trial. We were unable to obtain scholastic indicators before the final year of follow-up. Moreover, our dosing calendar was altered because of a teacher strike resulting in delayed dosing of several weeks. While this makes the data more difficult to compare year to year, it does give a more practical and real-world analysis of what a school-linked IPT program could expect to achieve under typical circumstances. Although protection from clinical malaria and reduced anemia is expected to result in improved school attendance and performance ,30, thes\u00ae); it is possible that ACT impacted the control group in 2008 and 2009. Finally, IPTsc implementation in our study was also impacted by the initiation of indoor residual spray (IRS), insecticide-treated nets (ITNs), and long-lasting insecticide-treated nets (LLINs); these strategies provide protection against morbidity and mortality attributable to malaria [One additional change to protocol occurred after year 1 (2007), in which the control group received SP alone for treatment of symptomatic malaria, but during the years 2 and 3, they were treated by artemether-lumefantrine (Coartem malaria ,32,33.IPTsc with SP+AS reduced the rates of all-cause clinic visits and anemia in three-year implementation together with IRS, ITNs, and LLIN strategies in Kolle."}
+{"text": "Plasmodium falciparum or Plasmodium vivax malaria were conducted in Myanmar between 2017 and 2019. Eligible subjects were aged at least 6 years, with microscopically confirmed P. falciparum (n = 196) or P. vivax mono-infection (n = 206). Patients received pyronaridine\u2013artesunate once daily for 3 days with follow-up until day 42 for P. falciparum or day 28 for P. vivax. For the primary efficacy analysis, adequate clinical and parasitological response (ACPR) in the per-protocol population at day 42 for P. falciparum malaria was 100%  in northern Myanmar (Kachin State and northern Shan State), and 100%  in southern Myanmar (Tanintharyi Region and Kayin State). Plasmodium falciparum day-3 parasite clearance was observed for 96.9% (190/196) of patients. Mutations in the P. falciparum Kelch propeller domain (K13) were detected in 39.0% (69/177) of isolates: F446I (14.7% [26/177]), R561H (13.0% [23/177]), C580Y (10.2% [18/177]), and P574L (1.1% [2/177]). For P. vivax, the day-28 ACPR was 100%  in northern Myanmar and 100%  in southern Myanmar. Across both P. vivax studies, 100% (206/206) of patients had day-3 parasite clearance. There were no adverse events. Pyronaridine\u2013artesunate had excellent efficacy in Myanmar against P. falciparum and P. vivax and was well tolerated. This study supports the inclusion of pyronaridine\u2013artesunate in national malaria treatment guidelines for Myanmar.Four single-arm, prospective, clinical studies of pyronaridine\u2013artesunate efficacy in uncomplicated  Plasmodium falciparum malaria is artemether\u2013lumefantrine, followed by single-dose primaquine (0.75 mg/kg). Dihydroartemisinin\u2013piperaquine or artesunate\u2013mefloquine are available as alternative ACT regimens. Plasmodium vivax malaria is treated with chloroquine complemented by radical curative treatment with primaquine (0.25 mg/kg/day for 14 days).The effective treatment of malaria is a priority health target in Myanmar, and the country has committed to eliminating falciparum malaria by 2025 and all kinds of malaria by 2030. Case management is used in areas targeted for elimination, and rapid diagnostic testing and treatment with artemisinin-based combination therapy (ACT) are provided free of charge by public and private clinics. The current first-line agent for P. falciparum.1 The artemisinin resistance phenotype is defined as an increase in day-3 parasite positivity following ACT treatment, that is, a delay in parasite clearance.2 Clinical efficacy is not affected unless resistance also develops to the partner drug, but this is more likely to emerge and to spread once the antimalarial activity of the artemisinin component is compromised. Unfortunately, multidrug-resistant P. falciparum is a serious problem across the Greater Mekong Subregion (GMS).4 Thus, potential alternative therapeutic options for Myanmar require evaluation in case current first-line treatments for falciparum malaria become ineffective, as observed in Cambodia.4Myanmar is one of the six countries with known artemisinin-resistant 12 Within the GMS, pyronaridine\u2013artesunate PCR-corrected day-42 adequate clinical and parasitological response (ACPR) rates against falciparum malaria in western Cambodia were 87.9%  in 2014\u20132015,13 but more recently > 98% efficacy was noted for this region (2018)14 and efficacy was > 96% in eastern Cambodia (1997),15 and > 96% in Vietnam (2017\u20132018).16 Thus, pyronaridine\u2013artesunate can remain a valuable treatment option for uncomplicated falciparum malaria in regions where the utility of other antimalarial drugs has been compromised by drug resistance.The ACT pyronaridine\u2013artesunate has shown good efficacy for uncomplicated falciparum and vivax malaria in large-scale clinical trials conducted in Asia and Africa.P. falciparum.17 However, P. vivax remains refractory to current interventions and has become the dominant parasite in some areas.19 Pyronaridine\u2013artesunate is the first ACT specifically registered for P. vivax malaria, with demonstrated high clinical efficacy across Southeast Asia.14 Chloroquine resistance has been reported in Myanmar,23 and to plan effective P. vivax elimination programs, the efficacy of alternative therapies requires confirmation.Intensive malaria control and treatment efforts in Myanmar have reduced the prevalence of P. falciparum and P. vivax malaria in southern and northern Myanmar to support a review of the national malaria treatment policy and to inform the design of malaria elimination programs in the context of artemisinin resistance.This study assessed the efficacy of pyronaridine\u2013artesunate for the treatment of uncomplicated Four single-arm, prospective, antimalarial treatment efficacy studies were conducted between July and December 2017 in two townships in northern Myanmar (Kachin State and northern Shan State) and between August 2017 and November 2019 in five townships in southern Myanmar (Tanintharyi Region and Kayin State) .24The studies complied with the Declaration of Helsinki, Good Clinical Practice guidelines, and all relevant local and international laws. The protocol was reviewed and approved by the Ethics Review Committee, Department of Medical Research, Ministry of Health & Sport, Myanmar, and the WHO Research Ethics Review Committee. All adult participants and parents or guardians of participants aged < 18 years provided written informed consent; in addition, assent was required from children aged \u2265 12 years. The trials were registered on the Australian New Zealand Clinical Trials Registry . The staP. falciparum  or microscopically confirmed P. vivax mono-infection   .25 body weight < 20 kg, mixed Plasmodium infection, severe malnutrition, the presence of febrile conditions other than malaria or other underlying chronic or severe diseases, regular medication which might interfere with antimalarial pharmacokinetics, a history of hypersensitivity reaction, or contraindications to the study medicine. All female participants aged 12\u201317 years were excluded because pregnancy testing before enrollment was not feasible owing to cultural conditions. Otherwise, women of child-bearing age were excluded if they had a positive pregnancy test, were breastfeeding, or were unwilling to use contraception. In addition, if patients had P. vivax infection, they were excluded if their hemoglobin was < 8 g/dL .General exclusion criteria were signs and symptoms of severe falciparum malaria,\u00ae, Shin Poong Pharmaceutical Co., Ansan, Republic of Korea) were given orally once daily for 3 days . Dosing was according to the body weight; 20 to < 24 kg, one tablet per day, 24 to < 45 kg, two tablets per day, 45 to < 65 kg, three tablets per day, and \u2265 65 kg, four tablets per day. Patients were treated as outpatients, but all doses were supervised and patients were observed for 30 minutes after dosing. In case of vomiting, patients were re-dosed and if the patient vomited again, they were withdrawn and administered rescue medication. Patients with P. falciparum malaria also received single-dose primaquine (0.75 mg/kg). In the case of P. vivax malaria, after testing for glucose-6-phosphate dehydrogenase (G6PD) deficiency using Carestart\u2122 rapid diagnostic test , primaquine was administered according to national treatment guidelines, that is, 0.25 mg/kg/day for 14 days if the patient was G6PD normal or 0.75 mg/kg/day once weekly for 8 weeks if they were G6PD deficient.Pyronaridine\u2013artesunate tablets  were examined by a third independent microscopist and parasite density calculated as the average of the two closest counts. In the case of P. falciparum recurrence, polymerase chain reaction genotyping (PCR) was used to distinguish between recrudescence and reinfection by comparing P. falciparum genes msp1, msp2, and glurp in baseline samples versus those obtained at failure, using published methods.27 Throughout the follow-up, patients were routinely asked about old symptoms and new symptoms emerging since the previous visit. The nature and incidence of adverse events and serious adverse events were recorded throughout the study.Patients were treated for 3 days , with follow-up on days 3, 7, 14, 21, 28, 35, and 42 for patients with K13). Studies were conducted at the laboratory of the Department of Medical Research, Yangon, using published methods.28 Briefly, nested PCR was used to amplify P. falciparum K13 genes (codon 432\u2013703) from extracted DNA samples, and purified PCR products were sequenced using a 3500\u00d7 L genetic analyzer . Sequence analysis of the K13 gene was performed using Sequencher\u00ae software  and BioEdit sequence alignment editor software . The K13 propeller gene sequence for P. falciparum 3D7 was retrieved from GenBank  and used as a reference strain. Fifteen percent of K13 mutant samples and 5% of wild-type samples were randomly selected and sent to the Pasteur Institute for external quality control; the results were found to be 100% concordant.Blood drops were collected on filter paper for assessment of molecular markers of artemisinin resistance in the Kelch propeller domain  and older than 16 years (84.7% [166/196]). For P. vivax, most patients were male (65.0% [134/206]) and 71.4% (147/206) were older than 16 years.In northern Myanmar, 90 patients were treated for P. falciparum in northern Myanmar, day-42 ACPR with pyronaridine\u2013artesunate was 100%  in the per-protocol analysis and 100% in the Kaplan\u2013Meier analysis  in the per-protocol analysis and 100% using Kaplan\u2013Meier analysis (n = 90), the day-3 parasite positivity rate was 6.7% . However, all of the patients with parasites at day 3 were from Kachin State, giving a day-3 parasite positivity rate of 13.3%  for this region. All of the 106 enrolled patients from southern Myanmar had parasite clearance at day 3.For patients with analysis . In soutanalysis . As therP. vivax, pyronaridine\u2013artesunate day-28 ACPR in the per-protocol analysis was 100%  in northern Myanmar and 100%  in southern Myanmar. In both regions, ACPR was 100% when estimated using Kaplan\u2013Meier analysis. All 206 enrolled patients across both studies had parasite clearance by day 3.For Plasmodium falciparum K13 sequences were obtained from 92.2% (83/90) of isolates collected in 2017 in northern Myanmar (K13 mutations was 41.0% (34/83); 33.3% (13/39) in Kachin State and 47.7% (21/44) in northern Shan State. In both states, the most common mutant was K13(R561H), with an overall prevalence of 27.7% (23/83). The C580Y mutant was only detected in northern Shan State (11.4% [5/44]). Myanmar . The proK13 sequences were available for 88.7% (94/106) of P. falciparum isolates (K13 mutation rate was 37.2% (35/94); 30.4% (14/46) in Tanintharyi Region and 43.8% (21/48) in Kayin State. K13(C580Y) was the most common mutant in Tanintharyi Region, whereas F446I was the dominant mutant in Kayin State  mutation, but the other four isolates were K13 wild type. Thus, there was no relationship between K13 mutations and day-3 parasite positivity rate.As pyronaridine\u2013artesunate clinical efficacy was 100%, no correlation could be made between P. falciparum- or P. vivax-infected patients.There were no adverse events of any cause recorded following treatment with pyronaridine\u2013artesunate in either P. falciparum. However, artemisinin-resistant P. falciparum is present in the region, and there is the potential for the emergence of partner drug resistance.30 Thus, the situation is fragile, and the impressive progress that Myanmar has made in reducing malaria prevalence and mortality over the last decade could be easily reversed should multidrug-resistant P. falciparum become established. Once a parasite emerges with a clear survival advantage under intense selective pressure from a narrow range of drug therapies, a selective sweep can cause a rapid loss of efficacy. This was seen for P. falciparum in Cambodia with a dominant artemisinin/piperaquine multidrug-resistant haplotype emerging within months of dihydroartemisinin\u2013piperaquine being introduced as first-line therapy.3 The most recent data from the GMS indicate that pyronaridine\u2013artesunate retains good efficacy in Cambodia and Vietnam against multidrug-resistant P. falciparum,16 and the current study confirms excellent efficacy in Myanmar. These findings are reassuring, potentially allowing the diversification of antimalarial therapy, and suggesting an alternative option should resistance undermine the efficacy of currently recommended antimalarial drugs.Malaria elimination in Myanmar is critically dependent on the continued efficacy of ACTs against P. falciparum K13 mutations suggests a genetically independent artemisinin-resistant parasite population in Thailand\u2013Myanmar\u2013China region versus Cambodia\u2013Laos\u2013Vietnam.31 An extensive geospatial mapping study reported the proportion of K13 mutants as 47.6% for Kayin State, 37.1% for Kachin State, and 66.7% for Shan State.32 This is broadly consistent with our findings . In the current study, four different K13 mutants were found: F446I, C580Y, and P574L in southern Myanmar and F446I, C580Y, and R561H in northern Myanmar. All of these are validated artemisinin resistance markers. However, in the current study, the majority of patients were aparasitemic by day 3, and delayed parasite clearance was associated with R561H in two cases and wild-type K13 in four cases. These findings are consistent with 2014\u20132015 data from Rakhine, Shan, and Kachin states, where artemether\u2013lumefantrine and dihydroartemisinin\u2013piperaquine retained high efficacy , despite the presence of artemisinin-resistant parasites in Shan and Kachin states.33 As seen in the current study, the day-3 parasite positivity rate was low following artemether\u2013lumefantrine (0\u20133.6%), and all parasites were cleared with dihydroartemisinin\u2013piperaquine.33Molecular surveillance of K13 mutations have limited impact on parasite clearance at present. This is in contrast to Cambodia\u2013Laos\u2013Vietnam, where a multidrug-resistant haplotype conferring high-level resistance to artemisinin and piperaquine has reached near fixation in some regions.16 Correspondingly, day-3 parasite positivity rates are relatively high, for example, 35.6\u201346.7% in eastern regions of Cambodia and 28.6\u201341.7% in the west.15 Moreover, high clinical failure rates are observed for dihydroartemisinin\u2013piperaquine.4 A limitation of this study was that molecular markers of P. falciparum resistance to piperaquine and mefloquine were not investigated. However, there is currently no clinical evidence of drug failure for ACT regimens containing these antimalarial drugs in Myanmar.34These studies indicate that in Myanmar, the circulating P. falciparum malaria in southern Myanmar was difficult, requiring two transmission seasons to achieve target recruitment. Falciparum malaria prevalence was low in the study areas, with only 1.6%  of patients screened having microscopically confirmed falciparum malaria. Social factors also hampered recruitment, for example, migrant workers in the Myanmar\u2013Thai border area were fearful and reluctant to engage, and a clinic set up by a nongovernmental organization also treated malaria patients in the region.The antimalarial resistance situation in Myanmar requires continued close monitoring. However, maintaining effective surveillance in a low-transmission setting is challenging. Finding eligible patients with P. falciparum has declined, P. vivax has emerged as a significant barrier to malaria elimination in Myanmar.35 There is also evidence of P. vivax chloroquine resistance in Myanmar,23 including high-level resistant strains circulating in northeast Myanmar.22 Although the prevalence of these strains is thought to be low, the success of elimination efforts based on chloroquine/primaquine could be jeopardized and intensification of P. vivax antimalarial treatment may further select and propagate resistant strains. Pyronaridine\u2013artesunate had 100% efficacy in the current study against P. vivax. This is consistent with data from Cambodia, Thailand, India, and Indonesia showing that pyronaridine\u2013artesunate was non-inferior to chloroquine, but with more rapid parasite and fever clearance.12 Thus, pyronaridine\u2013artesunate presents a clinically validated alternative to chloroquine for the treatment of P. vivax malaria and is the only ACT to receive approval for this indication by a stringent regulatory authority .12As the prevalence of 8 A limitation of the current study was that hepatic enzyme levels were not monitored. The standard protocol for therapeutic efficacy studies does not require laboratory monitoring, although there is provision for specific safety measures to be included if required. However, a recent extensive investigation showed no increase in the risk of hepatic transaminase elevations on repeated pyronaridine\u2013artesunate dosing, and no clinical manifestations of hepatic injury associated with the biochemical observations.7 There were also no clinical indications of any hepatic adverse effect of pyronaridine\u2013artesunate in the current study.Pyronaridine\u2013artesunate is known to cause asymptomatic mild-to-moderate transient increases in liver transaminases in some malaria patients.P. falciparum and P. vivax malaria in Myanmar. There were no cases of treatment failure for either parasite, and the therapy was well tolerated. Pyronaridine\u2013artesunate could be added to the national malaria treatment guidelines in Myanmar and can be considered as an additional tool for programmatic decisions aimed at achieving malaria elimination for both P. falciparum and P. vivax.In conclusion, this study evaluated pyronaridine\u2013artesunate efficacy against"}
+{"text": "Current FDA approved weekly chemoprophylaxis options have contraindications that can limit prescribing. The combination of chloroquine (CQ) with azithromycin (AZ) has previously been shown to be an efficacious treatment option for malaria, has pharmacokinetics compatible with weekly dosing, and has shown synergy when combined in vitro.Malaria remains the top infectious disease threat facing the U.S. military in many forward operating environments. Compliance with malaria chemoprophylaxis remains a critical component in preventing malaria in the deployed Service Member. Studies of previous military operations show that compliance is consistently higher with weekly Plasmodium falciparum. Volunteers that remained asymptomatic and had no evidence of parasitaemia continued to receive weekly post-exposure chemoprophylaxis for 3\u00a0weeks following malaria challenge. The primary endpoint was the number of volunteers that remained asymptomatic and had no evidence of parasitaemia 28\u00a0days after the malaria challenge.In this open label study, 18 healthy volunteers, aged 18\u201350\u00a0years (inclusive), were randomly assigned to receive either 300\u00a0mg CQ or 300\u00a0mg CQ and 2\u00a0gm azithromycin (CQAZ) of directly observed therapy, weekly for 3\u00a0weeks prior to undergoing mosquito bite challenge with chloroquine-resistant All 6 (100%) volunteers randomized to the CQ control group became symptomatic with parasitaemia during the 28-day post-challenge period. Only 1/12 (8.3%) of volunteers in the CQAZ group developed symptoms and parasitaemia during the 28-day post-challenge period. However, after chemoprophylaxis was discontinued an additional 6 volunteers developed parasitaemia between days 28\u201341 after challenge, with 4 of 6 experiencing symptoms. 80% of subjects in the CQAZ group experienced treatment related gastrointestinal adverse events (including 13% that experienced severe nausea) compared to 38% in the CQ group. A comparison of the pharmacokinetics in the CQAZ group demonstrated higher azithromycin Cmax (p\u00a0=\u00a00.03) and AUC (p\u00a0=\u00a00.044) levels in those volunteers who never became parasitaemic compared to those who did.Given the high rate of side effects and poor efficacy when administered for 3\u00a0weeks before and after challenge, the combination of weekly chloroquine and azithromycin is a suboptimal regimen combination for weekly malaria chemoprophylaxis.Trial registration ClinicalTrials.gov NCT03278808 Although the incidence rate has been declining since 2010, in 2018, there were still an estimated 228 million cases of malaria occurred worldwide [95% confidence interval (CI): 206\u2013258 million] . AlthougTafenoquine has the benefits of weekly dosing along with activity against all malaria stages that make it a promising prophylactic agent. However it is still new to the market and is contraindicated in those with unknown or deficient glucose-6-phosphate dehydrogenase activity . With reP falciparum isolates [P. falciparum field isolates from Mali showed some synergy between CQ and AZ at the IC90 level and an additive effect at the IC50 level [Azithromycin\u2019s anti-malarial activity has been known for several years and mechanistically is believed to work by inhibiting apicoplast function . Given tisolates . In addi50 level . FurtherP. falciparum.Azithromycin has been tried as a sole agent for malaria prophylaxis in clinical trials with modest effectiveness. Studies done in Kenya and Indonesia demonstrated modest results ranging from 72 to 84% efficacy with daily administration and 64% with weekly \u201312. The P. falciparum CQ resistance, no testing was reported [P. falciparum. Over 98% of subjects met the primary endpoint of being PCR confirmed parasite free at Day 28. In addition, in vitro analysis of study isolates of two countries showed rates of CQ resistance of 21% (Zambia) and 96% (Uganda), respectively. Serious adverse event (SAE) rates for CQAZ were four-fold lower than in the comparison treatment arm, mefloquine [P. falciparum, multi-country  treatment study in 2017 examined multiple AZ dosages  combined with 600\u00a0mg base CQ for 3\u00a0days. The 500\u00a0mg AZ arm was discontinued early due to poor parasite clearance rates by Day 28 (36% Colombia/Suriname & 66% India). The 1 gm AZ arm also showed clearance rates inferior to its comparator arms with day 28 parasite clearance rates of 59% (Colombia/Suriname) and 84% (India). The study conducted with 2 gm of AZ and 600\u00a0mg CQ base was a non-comparator study in India and Colombia that showed a parasite clearance rate of 97% (104/107) demonstrating evidence of a dose response relationship. In all three countries, high rates of P. falciparum CQ resistance were seen- 92.2% in India and 98.4% in Colombia & Suriname combined [In India, 63 subjects received a combination of AZ (1gm) and CQ 1500\u00a0mg) for 3\u00a0days and saw a 97% resolution of fever and parasitaemia by day 7 with no relapse by Day 28. Although performed in areas of high reported . Two stufloquine . A subse00\u00a0mg forThe United States military continues to engage in operations in malaria endemic areas. The current chemoprevention options utilized, while appearing to be effective with only 58 service members diagnosed with malaria in 2018, belie an underlying concern for the future. The 58 infected service members represents a 66% increase over 2017 . In addiP. falciparum infection utilizing a human malaria challenge model at the Walter Reed Army Institute of Research (WRAIR). Given the evidence of a possible dose response relationship and the prior success observed at 2\u00a0gm, the study utilized a weekly dosing regimen of 2\u00a0gm of AZ and 300\u00a0mg base of CQ.This study aimed to explore the use of the combination of CQ and AZ as a chemoprevention agent for preventing chloroquine resistant Plasmodium falciparum. Secondary objectives were to assess the tolerability and pharmacokinetics of the regimen.The primary objective of this study was to assess the safety and efficacy of a weekly CQAZ regimen for prophylaxis against CQ resistant https://clinicaltrials.gov/ct2/show/NCT03278808?cond=cq%2Faz&draw=2&rank=1.This study was an open-label, randomized controlled trial utilizing a human malaria challenge model. The investigation was conducted at the WRAIR Clinical Trials Center, Silver Spring, MD from 2018 to 2019. Healthy, non-pregnant, non-breastfeeding adults aged 18\u201350 (inclusive) were eligible for participation. Potentially eligible participants were screened utilizing medical history, physical examination and standard hematologic, renal and liver laboratory evaluations. Laboratory evaluations for human immunodeficiency virus, hepatitis B and C were also conducted. Cardiac risk factors and screening electrocardiogram were assessed. Main exclusion criteria included any chronic medical condition as determined by history, physical examination, or laboratory evaluation that would affect the study results or put the subject at an unacceptably increased risk. Female subjects had to have a negative urine pregnancy test at initial screening and prior to first treatment drug administration and malaria challenge. Women of child bearing potential were required to have been on some form of birth control from 1\u00a0month prior to study enrollment and agree to continue at least two forms of birth control for at least 56\u00a0days after the challenge. Prior to enrollment, study subjects could not have visited a malaria endemic country in the previous 3\u00a0months, received any malaria prophylaxis in the previous 2\u00a0months, been diagnosed with malaria within the past 3\u00a0years, or have ever received an experimental malaria vaccine. Concomitant medications that could potentially affect the pharmacokinetics of either treatment drug or the prescribed medication, such as cimetidine or other antacids, atorvastatin, or fluconazole or have potential anti-malarial activity were prohibited during the study. The study was registered on ClinicalTrials.gov\u2014NCT03278808 Registered 12 September 2017\u2014Retrospectively registered, \u00bd) for AZ, CQ and chloroquine\u2019s major metabolite, desethylchloroquine (CQm).The primary endpoint was symptomatic parasitaemia within 28\u00a0days of being challenged. Parasitaemia was determined by microscopic evaluation of thick blood smears. Symptomatic was defined as any one of the following solicited adverse events (AEs) that occurred concurrently with parasitaemia: fever (temperature\u2009>\u2009100.4\u00a0\u00b0F), chills, headache, arthralgia, myalgia, nausea, vomiting, or abdominal pain. Secondary safety endpoints included solicited, unsolicited AEs and electrocardiogram (ECG) findings. Secondary pharmacokinetic endpoints included area under the curve (AUC), maximum concentration (Cmax), time to maximum concentration (Tmax), and half-life  for challenges requiring chloroquine resistant arasites .The CHMI was conducted at the WRAIR, an institution where over 100 malaria challenge studies have been conducted since 1985. Each subject\u2019s forearm was exposed to five 7G8-infected mosquitoes for a period of 5\u00a0min. After this 5-min period, dissection of the mosquito midgut and salivary gland was used to determine if an infectious bite occurred. If none or fewer than 5 infectious mosquitoes bit a subject, then the subject\u2019s forearm was exposed to an additional number of mosquitoes to reach 5 infectious bites total.\u00ae) and 300\u00a0mg of CQ base (Natco Pharm Ltd) and the CQ only group received 300\u00a0mg of CQ base. Subjects received their treatment medication weekly for 3\u00a0weeks prior to the malaria challenge. All treatment medications were administered by study staff and directly observed. Although a standard meal was not provided, all participants were instructed to have eaten prior to taking the CQAZ or CQ, and were provided food if they had not eaten. The challenge occurred midway between weeks 3 and 4 of the study. Eight days after the malaria challenge, subjects checked into and were followed closely at a local hotel with 24-h on-site medical support. Malaria transmission was virtually eliminated by conducting the study during the fall and winter, the small number of subjects, counseling each subject to not leaving the local area and by providing rescue treatment as soon as symptomatic parasitaemia was encountered. Daily Giemsa-stained thick blood film smears for microscopy were obtained on post-challenge days 8\u201321, in addition to any time a subject reported possible malaria-related symptoms. Microscopists reading the smears were blinded to the treatment group. Parasitaemia was determined using Giemsa-stained blood slides using World Health Organization (WHO)-recommended methods [After enrollment, subjects were randomized into either the CQAZ group or CQ only control group. The CQAZ group received 2 gm of AZ , for any subject who had not yet been deemed a treatment failure, blood samples for drug concentrations were collected at 0, 1, 2, 4, 6, 10, 24, 48, 72, and 96\u00a0h post dosing. The same subjects also had an ECG performed approximately 6\u00a0h after this dose. QT measurements were corrected using Fridericia\u2019s (QTcF) formulas.1.00\u00a0mg/ml standard stock solutions of CQ, CQm and AZ  were prepared in dimethylsulfoxide (DMSO) and were used to make up a 10\u00a0\u00b5g/ml mixture of CQ, CQm and AZ in acetonitrile. The calibration standard curve and quality controls (QC) were prepared by spiking blank human plasma  with this 10\u00a0\u00b5g/ml stock. The calibration standard curve consisted of matrix and an internal standard, mefloquine, with analytes ranging from 0\u20131000\u00a0ng/ml concentrations, with QC samples covering the low, medium, and high concentration ranges of the standard curve. 100\u00a0\u00b5l of sample was placed in a microcentrifuge tube and 200\u00a0\u00b5l of acetonitrile with internal standard was added. Each sample was vigorously vortexed for 15\u00a0s and centrifuged at 13,000\u00a0rpm for 10\u00a0min at 4\u00a0\u00b0C. 220\u00a0\u00b5l of undisturbed supernatant was transferred to a 96-well plate for liquid chromatography\u2013mass spectrometry (LC\u2013MS) analysis.The human plasma samples were extracted in the same manner with 200\u00a0\u00b5l of internal standard added to 100\u00a0\u00b5l of sample. Concentrations of CQ, CQm and AZ in samples were interpolated from each corresponding standard curve. Samples with concentrations greater than the highest point of the calibration curve were diluted with blank human plasma and extracted in the same manner.A Waters  ACQUITY UPLC system was coupled with an AB Sciex  QTrap 4000 linear ion trap spectrometer equipped with a Turbo-V source. A Waters CORTECS C18  column was maintained at room temperature while the autosampler was maintained at 4\u00a0\u00b0C to minimize evaporation. Samples were eluted using a linear gradient going from 5% to 95% acetonitrile/0.1% formic acid in water over the course of 1.50\u00a0min followed by 1.75\u00a0min of isocratic gradient of 95% acetonitrile/0.1% formic acid in water at the flow rate of 0.400\u00a0ml/min.\u00ae software.The analysis was performed in multiple reaction monitoring in positive electrospray ionization mode by monitoring the ion transitions from m/z 320.200\u2009\u2192\u2009247.100 (CQ), m/z 292.120\u2009\u2192\u2009114.100 (CQm), m/z 749.601\u2009\u2192\u2009591.400 (AZ), and m/z 379.100\u2009\u2192\u2009361.100 (mefloquine). Compound parameters and source/gas parameters were optimized to obtain the highest intensity of the analytes. The instrument was controlled and data was collected using Analyst1/2, Tmax, Cmax, AUC from 0 to 96\u00a0h (AUC0\u201396), extrapolated AUC from 0 to infinity hours (AUC0\u2013\u221e) and the elimination rate constant (Kel).The measured plasma concentrations of AZ, CQ and the main chloroquine metabolite (CQm), from each study subject, based on 10 time points per subject, were evaluated. Using the Phoenix WinNonlin 8.1 software , non-compartmental analysis (NCA) with the linear up-log down trapezoidal method was performed to calculate the mean pharmacokinetic parameters. Calculated parameters included the tThe desired prophylactic efficacy for anti-malarial drugs in general is defined as approximately 95% compared to placebo. However, given that this was an exploratory proof of concept study efficacy rates >\u200990% were to be considered a success. The CQ control group was utilized as a measure of malaria challenge success with greater than 1/6 (17%) of subjects not getting study malaria defined as a malaria challenge failure. This number was based on the previous history of malaria challenges at the WRAIR. The study was designed to enroll 12\u201315 subjects in the CQAZ group with prophylactic success defined as equal to or greater than 91.6% (11/12), 92.3% (12/13), 92.8% (13/14) or 93.3% (14/15).All reported study data was recorded on the electronic Case Report Forms supplied by Statistics and Data Corporation (SDC) using an Electronic Data Capture clinical database called iMedNet. After data was entered into the clinical study database, electronic edit checks and data review were performed.Since the measure of success for the primary efficacy analysis was pre-set at greater than 1/12 treatment failures and the CQ group was a control for measuring challenge success, no inferential statistics were performed comparing the efficacy of CQAZ versus CQ. Group characteristics were compared between the two groups using the intention to treat population (ITT). The ITT population was defined as any subject that was enrolled, randomized and received at least one dose of either CQAZ or CQ. Variables were compared using the using Fisher\u2019s Exact Test or two sample T-test. Subjects in the CQAZ group were further analysed based on whether they were considered protected from malaria. Variables in each group (protected (P) vs. non-protected (NP)) were compared using Fisher\u2019s Exact Test or two sample T-test. Pharmacokinetic variables in these two groups were compared using two sample T-test. All inferential tests were performed at the \u03b1\u2009=\u20090.05 significance level.Forty (40) potential participants were consented and screened. Twenty-three 23) subjects met all screening criteria and were randomized into the CQAZ 15) or CQ (8) group . In the CQAZ cohort, 1/12 subjects presented with symptomatic parasitaemia while still receiving post-exposure CQAZ, on day 9 post-challenge. Two (2) of the remaining 11 subjects had positive smears during the post-exposure prophylaxis period but were asymptomatic. Both of those 2 subjects\u2019 subsequent smears cleared without rescue treatment and they remained in the study. After the 11 remaining subjects received all 6 weekly doses of CQAZ , 6 subjects had positive malaria smears, with 4 of those symptomatic. The mean post-challenge day of presentation for these subjects who had positive smears after finishing all doses of CQAZ was 35\u00a0days after challenge (28\u201341\u00a0days). Two subjects who had positive smears on day 28 were asymptomatic and therefore did not meet endpoint criteria but were treated with rescue A/P prior to symptom development. The mean number of days to a positive malaria smear after the last CQAZ dose was 17\u00a0days (10\u201323\u00a0days)  by post-challenge day 28. However, an additional 4 subjects had symptomatic malaria after day 28. Another 2 asymptomatic subjects presented with parasitaemia after the post-exposure treatment period (Day 28) that received rescue treatment and would have likely become symptomatic for malaria.Further analysis was conducted comparing those in CQAZ group that did not develop parasitaemia, (P) and those that did (NP). Given the low numbers in each group,  no statistical differences were found between each group Table\u00a0.Table\u00a03CBoth the CQAZ and CQ groups showed high rates of treatment-related AEs, (87% vs 63%) with trends towards higher rates in the CQAZ group . Both the P and NP groups had 1 subject with severe nausea.An analysis of the laboratory data showed one subject with a potentially relevant clinical abnormality. One subject in the CQAZ had an elevated aspartate aminotransferase (AST) of 126 U/L on Day 28. This subject was found to have a positive malaria smear on this visit. The subject had received all 6 CQAZ doses, with the last dose 10\u00a0days prior. While taking CQAZ, the subject\u2019s AST levels had been normal (65\u00a0U/l and 72\u00a0U/l). Subsequent AST levels normalized and no other liver function test was abnormal. The timing favours malaria infection as the cause of elevated AST rather than the CQAZ. No other subject had any clinically significant haematologic, electrolyte, kidney or liver function laboratory abnormality.An analysis of the CQAZ group ECG data showed at baseline, a mean QTcF of 406\u00a0ms (399\u2013413\u00a0ms)\u00a0. This trend continued with differences in AUC0\u2013\u221e  and Cmax  showing greater AZ exposure levels over time in the group that did not develop malaria  and AUC0\u2013\u221e   of subjects successfully avoided a symptomatic positive malaria smear while receiving weekly CQAZ, the drug combination was only able to keep the parasite levels to below the microscopic level of detection  while being actively taken . Once thdormancy . In addidormancy . RegardlP. falciparum malaria.Given that all control CQ subjects had symptomatic parasitaemia within 10-15\u00a0days after being challenged, this demonstrated that this was a viable challenge study for testing for prophylaxis of CQ resistant 0\u201396 and Cmax and appeared to have the strongest exposure\u2013response correlation with protection  to detect and provide rescue treatment before parasite load is high enough for symptoms to develop . This deWhile the study was powered successfully to determine the efficacy of CQAZ in this experimental model, it was insufficient to meet secondary objective goals. While strong trends existed for safety and pharmacokinetic data, larger numbers of participants would have further clarified these results. This study was a human challenge study that utilized mosquitoes to transmit the malaria rather than direct inoculation of sporozoites through venipuncture. Each method has its scientific merits and drawbacks. One drawback of using mosquitoes is the variable and unquantified number of sporozoites the mosquito injects into each subject . How sigP. falciparum mosquitoes, the weekly combination of CQAZ was not effective in preventing malaria at rates that would be acceptable to justify exploring in further larger scale studies. The GI AE rates from this study would also likely be unacceptable for travellers or a military deployed population.In conclusion, utilizing a malaria challenge model with chloroquine resistant Additional file 1. AE Listing: Listing of all adverse events in enrolled subjects.Additional file 2. EKG Data: Lisitng of electrocardiogram values at\u00a0 baseline and Day 11 after challenge.Additional file 3. PK Data Points: Table of CQ, CQm and AZ exposure levels in the CQAZ cohort."}
+{"text": "Plasmodium resistance to the available drugs alongwith the insurgence of multidrug- and particularly tuberculosis drug-resistantstrains are enough to justify efforts towards the development of novel medicinesfor both diseases. This literature review provides an overview of the state ofthe art of antimalarial and antituberculosis chemotherapies, emphasising noveldrugs introduced in the pharmaceutical market and the advances in research ofnew candidates for these diseases, and including some aspects of theirmechanism/sites of action.Malaria and tuberculosis are no longer considered to be neglected diseases by theWorld Health Organization. However, both are huge challenges and public healthproblems in the world, which affect poor people, today referred to as neglectedpopulations. In addition, malaria and tuberculosis present the same difficultiesregarding the treatment, such as toxicity and the microbial resistance. Theincrease of  It is estimated that there were228 million cases and 405,000 deaths in 2019, within a population of over 3 billion atrisk of infection.,Plasmodium resistance to the availabledrugs demonstrate the urgent need for developing new therapeutic options.Anopheles mosquito and five Plasmodium speciesinfect human beings: P. vivax, P. falciparum,P. malariae, P. ovale and P.knowlesi. The latter is mostly found in monkeys and, incidentally, caninfect people. Furthermore, P. falciparum infections account for mostdeaths, whilst P. vivax infections may induce severe chronicmalaria.,Malaria  is one oMycobacteriumtuberculosis (M. tuberculosis) and is still one of thegreatest problems for public health around the world today. It affects mainly the lungs;however, the brain, liver and other organ systems can be affected as well. In 2018,approximately 10.0 million people fell ill and 1.4 million died of TB, according to theWorld Health Organization (WHO) Global tuberculosis report released in 2019, as seen in,Tuberculosis (TB) is an infectious disease caused by Both diseases are huge challenges and public health problems in the world, which affectthe same populations, generally poor people. Furthermore, malaria and tuberculosispresent the same troubles regarding the difficulty in treatment, drug toxicity and themicrobial resistance to the available medicines. This scenario is enough to justifyefforts towards the development of novel therapeutic agents.Malaria chemotherapyCurrent status - The treatment regimen for malaria depends heavily onseveral parameters, such as age, pregnancy, species, severity and chronicity. For thisreason, there is a wide array of drugs employed and there are different drug regimenspossible, making malaria treatment heterogeneous and diverse. The main drugs employedfor malaria therapy are: artemisinins, endoperoxides and derivatives , primaquine, due to its gametocytocidal effect and itsactivity against the hypnozoite form , quinine and 4-aminoquinoline derivatives, such aschloroquine and amodiaquine, and the antifolate agents sulfadoxine/pyrimethamine, usedin combination. The WHO recommends artemisinin-based combination therapies (ACTs) forthe treatment of uncomplicated P. falciparum and P.vivax malaria,,,,Malaria chemotherapy has multiple biochemical pathways and enzymes as targets, some ofwhich are still unknown. Current chemotherapy explores mainly the formation of hemozoincrystals,Advances in the research of new antimalarial agents - Artemisinin andits analogs are endoperoxides, whose mechanism of action induces oxidative stressinflicted through ROS formation. The generation of the toxic free radical is dependenton the interaction of the drug with intraparasitic heme groups, present on the foodvacuoles of the parasite. It is proposed that, upon activation, the toxic radicalpromotes the alkylation of several protein targets, which leads to biological functionimpairment and parasite\u2019s death.\u00ae) is a synthetic1,2,4-trioxolane with a peroxidic pharmacophore, which exhibited invitro potency higher than most current chemotherapeutic agents againstP. falciparum, including against chloroquine resistantstrains.,,,P.vivax. The findings demonstrated that, even though there are signs ofCQ-resistant strains in Ethiopia, CQ treatment still exhibited smaller recurrence rates28 days and 42 days after starting the treatment. Nevertheless, both treatment regimenswere improved when co-administered with PQ, evidencing that the treatment schedule wouldfurther benefit patients at risk of relapsed infection and transmission.Several new combination therapy regimens for artemisinins and its derivatives are goingthrough clinical trials in order to establish the most efficient and safe therapeuticscheme, while also monitoring the development of local resistant strains. The ACTartesunate-amodiaquine (ASAQ) and AL regimens were tested in a 42 day trial in the IvoryCoast, and the study found successful treatment rates of 100% and 99.1% respectively,demonstrating that both ACTs are suitable for further use and no resistance is yetidentified in the country.Naphthoquine  is a 4-aP. falciparum, schizonts of all species andit is the only current drug used in chemotherapy, eliminating the latent hypnozoite ofP. vivax and P. ovale, thus providing a radicalcure.Plasmodium agent to mosquitoes, blocking furthertransmission to new vectors.Primaquine  is an 8-P. falciparum infection. However, resistance to this drugis widespread and CQ is no longer used for this species. In addition, this drug isapplied to uncomplicated malaria caused by chloroquine-sensitive strains of P.vivax, P. ovale and P. malariae.,Plasmodium parasites, occurring within the foodvacuole, in which the acidic environment aids the chloroquine accumulation, due to itsweak alkaline nature.,Chloroquine (CQ)  is the mP. vivax (91.9% of TQ versus 77.3% of PQ). Intotal, thirteen assays were performed to support the efficacy of TQ, with specialattention to three randomised, double-blind studies: DETECTIVE Part 1 and Part 2(NCT01376167) and GATHER. TQ was approved by the FDA in July 2018, and is currentlyproduced by GlaxoSmithKline\u00ae, under the brandKrintafel.,,Tafenoquine (TQ)  is anothP. falciparum strains whilst possessing lowtoxicity.,Ferroquine  is a 4-a\u00ae)is a spiroindolone in Phase II clinical trials. This compound inhibits a new moleculartarget, PfATP4, present in P. falciparum - the first new validatedmolecular target for malaria after 20-year research. PfATP4 is a Na+-ATPaseresponsible for maintaining a low concentration of cytosolic sodium, which, wheninhibited, causes a disturbance of parasitic sodium haemostasis, leading to its death.Cipargaramin derivatives can achieve an IC50 lower than 0.2 nM and displayactivity against oocytes, gametocytes and the asexual stage present inPlasmodium parasites blood. Phase I clinical trials used anestimated dose based on the PK/PD 30 mg model, demonstrating no major adverse events,with minor gastrointestinal and genitourinary events when high doses were administered.The drug showed a dose dependent effect, reaching a plateau of activity in doses around30 mg. The completion of the Phase II clinical trial is due in 2020.,,Cipargamin  drugs screened 226 FDA-approved drugsagainst P.falciparum. Its combination with clindamycin has already been trialed andhas presented initial success for malaria treatment, however, a more recent trialreported unfavorable results for this combination.,Fosmidomycin  is an anPlasmodiumparasites, ivermectin is being considered for mass administration to control malariavectors.,,,,,,Differently than other repurpose drugs that intend to kill ,There are at least 50 new bioactive compounds previously quoted and novel drugs currentlyunderway for malaria treatment, in lead optimisation, preclinical and clinicalevaluations.Tuberculosis chemotherapyCurrent status - Although there are established treatment regimens, rifampicin (R), pyrazinamide (Z) ,ethambutol (E) , preferaAdvances in the research of new tuberculostatic agents - Theintroduction of rifampicin and its first use in 1966 marked the last drug released for along period of time, as the therapeutic regimen was effective enough to treat infectedpatients.,,,Bedaquiline (TMC-207)  is adiain vitro and in vivowhen compared to the standard treatment protocol.,SQ109 is a diethylamine  related ,M.tuberculosis, comparatively to some fluoroquinolones.,,,Linezolid  is an oxSutezolid (PNU-100480)  is anox,,Rifapentine  is a rif,Pretomanid (PA-824)  is a bic,,,Delamanid (OPC-67683)  is anothM.tuberculosis. Clofazimine acts as a prodrug in M.tuberculosis by reduction through the NADH dehydrogenase (NADH2) enzyme,releasing reactive oxygen species upon reoxidation by O2.para\u2010aminosalicylic acid.Clofazimine  is a lip,M.tuberculosis, and retrospective studies on patients have shown thatmeropenem-clavulanate have added value to multidrug treatments, elevating the sputumconversion rate to levels as high as 87%.,,in vivo, presenting a 6- to 22-fold more efficient inhibition thanmeropenem.,,,,In the last few years, carbapenems such as meropenem and faropenem  have sur,in vitro assays, and is now under phase IIclinical trials.,,in vitro assays, witha MIC of 0.2 ng/mL, and is currently under phase II clinical trials.,in vitro, achieving a MIC ranging from0.001 to 0.000024 \u03bcg/mL on 40 different strains of M. tuberculosis,including MDR and XDR strains, and is now currently under phase II clinical trials.,BTZ043, a benzothiazinone, and OPC-167832, a carbostyril derivative, are members of anovel class of drugs  for TB tCurrently, there are more than fifteen clinical trials in Phase II or III, evaluating theefficacy of new tuberculostatic drugs in combination therapy regimens, alone or inassociation with another standard drug.invitro activity against M. tuberculosis, while other PPIssuch as omeprazole and pantoprazole showed no activity. A cohort study was performed inthe UK to assess the incidence of TB on users of PPIs, comparing lansoprazole users toomeprazole or pantoprazole users. Results demonstrated that lansoprazole users presenteda considerably lower TB incidence, when compared to omeprazole or pantoprazoleusers.The efficacy of a novel formulation for a low dose rifampicin regimen (200 mg compared tothe standard 450 mg), piperine (10 mg) and isoniazid (300 mg), named risorine, wasassessed in a Phase III clinical trial in comparison to the standard treatment regimen,conducted in India. The results showed a slightly higher sputum conversion rate and curerate in the risorine group, when compared to the control. Higher blood levels ofrifampicin were achieved in risorine, in relation to the standard regimen, despite thelower dose administered, hence an improvement on the safety profile, with a lower rateof adverse effects.The global distribution of tuberculosis is still undetermined, with the appearance of MDRand XDR-TB strains gradually spreading more widely. However, considering the history oftuberculosis treatment research, there has been a substantially greater advance in thediscovery of biologically active compounds, leads and repurposed drugs in the last 10years, when compared to the previous 40. In order to achieve global control of thisepidemic, some changes are necessary, such as decrease in treatment duration, targetingMDR or XDR and simplifying treatment by lowering dosing frequency. Most of theabove-mentioned compounds in this review achieved in some degree these aims.Drug repurposing has also been stimulated for TB, especially to overcome the severeproblem of multi-resistant and extensively drug resistant mycobacteria.There are some significant programs and institutions, such as the TB Alliance and theWHO, who focus on developing treatments, the political will and on providing orientationto countries with heavy burdens of tuberculosis. Considering these efforts, it ispossible that within the next decades, this disease could be brought under control.Researches for antituberculosis and antimalarial drugs from protein kinaseinhibitors - Protein kinases play as key controllers of signaltransduction, being responsible for regulating essential cellular processes, such asgrowth, development and replication.,,,,The emergence of drug-resistant strains highlights the need of new therapeuticapproaches, thereby kinase inhibitors should gain attention as new therapeutic choicesagainst tuberculosis and malaria.Protein kinases participate in phosphorylation processes involved in host-pathogeninteraction, being classified into three main groups: serine/threonine protein kinase(STPK), tyrosine protein kinase (TPK) and two-component regulatory system (2CRS)consisting of histidine kinase and response regulator. The 2CRS system responsible forprotein phosphorylation from prokaryotes occurs only on His and Asp residues.Tuberculosis - M. tuberculosis protein kinases haveshown to be critical targets for mycobacterial survival and proliferation.M. tuberculosis presents inits kinoma 11 STPKs number similar to 2CRSs, playing important roles for its survival,pathogenesis and virulence. These STPKs are named PknA to PknL.M. tuberculosis enzymes widelyinvestigated, and they are essential for growth and regulation of cell wall biosynthesisand cell division.,,,M. tuberculosis in cell culture.M. tuberculosis inside the macrophages.M. tuberculosis are essential for the mycobacterial growth,MtrA and MtrB perform in the regulation ofmycobacterial metabolism and adaptation to environmental changes.MtrA inhibitors and they found eight potential molecules.Biological assays revealed that the compounds 2IT4O  lower in macrophages, implying itcan also act in other pathways.2CRSs from Malaria - In antimalarial therapy, there are no protein kinaseinhibitor drugs. However, these enzymes present an essential role in the host and in theparasitic life cycle. In this context, protein kinase inhibitors are a promising fieldfor designing of antimalarial agents. P. falciparum protein and lipidkinases participate in the main signaling pathways at diverse steps of its lifecycle.P. falciparum kinome encodes 86 to 99 protein kinases and a small setof lipid kinases, though the function of most of them is still unknown. The mostadvanced studies involve calcium-dependent protein kinases (PfCDPKs),Protein kinase 7 (PfPK7), cyclin-dependent kinases(PfMRK), cGMP-Dependent Protein Kinase (PfPKG),Phosphoinositide lipid kinases (PIKs).,Plasmodium (PfCDPK) belongs to the STPKfamily and are composed of seven members named as PfCDPK1 toPfCDPK7. They are one of the most attractive targets for designingof new antimalarial agents due to the critical role in the life cycle ofPlasmodium and absence of their homologous in the human, which mayresult in higher selectivity against parasite and lower toxicity to the host.,CDPKs from PfCDPK1 phosphorylates important proteins of the parasitic motorcomplex involved in the invasion of host cells, especially in erythrocytes, beingconsidered a remarkable target for antimalarial therapy.,,PfCDPK1.Imidazopyridazine derivatives, 2, 6, 9-trisubstituted purines, and bisindolocarbazoleK252a  showed to be active in protein kinase and/or P.falciparum survival assays in host cells.,,,,P. bergheiranging from 46% to 51%, when administered orally once daily in doses of 50 mg/kg forfour days.,PfCDPK4 participates in the sexual cycle of Plasmodiumin Anopheles.P. berghei, PbCDPK4 regulates gameteformation mediated by xanthurenic acid and parasite transmission.PbCDPK4 knockout assay, male gametocytes were not able tomature into fertile male gametes in the mosquito gut. Thereby, this kinase prevents theparasite transmission, blocking the parasitic ex-flagellation in the gametogenesisprocess inside the vector.P. falciparum. In this context,pyrazolopyrimidine and imidazopyrazine compounds inhibited fromPfCDPK4, among them Bumped Kinase inhibitors, as the derivative ofBKI-1  plays atdifferent stages of the parasite life cycle, both in vector and in the human host. Itwas found that the suppression of the PfPK7 gene attenuates theparasite asexual growth in erythrocytes, thus PfPK7 acts a crucial rolein the transmission.PfPK7 inhibitors have been described, such as compoundsK510, K109, K497, imidazopyridazine, himenialdisine and staurosporine.,P. falciparum in blood-stage cells.PfPK7 and Plasmodium in in vivoassays.PfCDPK1, thereby they can play through two different pathways.,,,PfPKG  isa protein involved in multiple steps of Plasmodium life cycle, beingessential for the replication process in the host blood-stage.,,PfPKG important to the sexual stage of P. falciparum,blocking the gametocyte transmission to the vector.50 values of 130 and 160 pM, respectively, againstthe wildtype enzyme, and 2 and 102 nM against the wildtype Pf3D7strains. Furthermore, compound 14 displayed good results regarding cytotoxicity,moderate metabolic stability in vitro and high selectivity against apanel of 80 human kinases. The oral administration of analogue 14, twice daily withdoses at 100 mg/kg during four days in mice infected with P. falciparumled on the reduction of parasitaemia to undetectable levels.50 of 3.5 nM against recombinant PfPKG, similar to thenative strain.In vitro assay against chloroquine-sensitive strainPfNF54 and the chloroquine-resistant strain PfDd2, thecompound 15 displayed IC50 values of 0.49 and 1.3 \u03bcM, respectively. On theother hand, in mice infected with P. berghei, compound 15 was not ableto eliminate parasites at intraperitoneal doses of 50 mg/kg twice daily for eight days.In HepG2 cell culture, compound 15 decreased the infection of P.berghei by sporozoites.in vitro, compound 15 decreased the number ofparasites in the liver-stage below the detection limit, inhibiting host cells invasionwith an IC50 below 1 \u03bcM. However, sporozoites without PbPKGremained sensitive to compound 15, indicating that it affects other targets thanPbPKG. In mouse infected with P. yoelii, strainwith higher infectivity of the sporozoite than P. berghei, before theinfection was administered a single intraperitoneal dose of 50 mg/kg, which resulted inreduction of hepatic parasitic load by 1000 times. Thereby, three doses of 50 mg/kg wereadministered, the first 15 minutes before the infection and, second and third doses 6and 12 h after infection, all mice were free of parasites in the blood-stage during thethree week of the experiment.From a series similar to the imidazopyridine composed of 2,3-diaryl-pyrrole derivatives,compound 15  was founPfMRK protein plays an essential role in DNA replication andPlasmodium transcriptional control.,) From a series of quinolinones, the compound 16  and it showed higherselectivity than mammalian kinase (CDK1 - IC50 = 29 \u03bcM). However, insensitive PfD6 strains, the compound 17 showed moderate antiplasmodialactivity, which was attributed to the its low permeability.The pound 16  was iden,Plasmodium species arephosphoinositide 3-kinase (PI3K) and phosphatidylinositol 4-kinase (PI4K), both areessential for the survival of P. falciparum. PI3K acts on the growth ofthe parasite and was recently reported as one of the targets of dihydroartemisinin, inwhich dihydroartemisinin demonstrated to be a potent PfPI3K inhibitorin the nanomolar range.Phosphoinositide lipid kinases (PIKs) generate phosphorylated phosphatidylinositolderivatives, which are crucial for different cellular functions, such as messengersignaling, cell membrane remodeling and vesicular trafficking.,P. falciparum,P. yoelii and P. cynomolgi. However, KAI407exhibited poor physico-chemical properties. In this context, molecular modificationswere carried out in the core, which was changed to imidazopyrazine group, resulting inKDU691 compound less lipophilic  PI4K with IC50 of 3.4 nM,in vivo modelsof Plasmodium infections, for example, MMV048 presented ED90values of 0.80 mg/kg in mouses infected with P. berghei. MMV048 showedprophylactic activity, preventing infection in a monkey model infected with P.cynomolgi (2 mg/kg before infection). In addition, its pharmacokineticproperties were evaluated in mice, rat, dog and monkey models. From these assays, asingle dose ranging 80 to 100 mg for humans was established. MMV048 progressed frompreclinical development to Phase 1 clinical trials. Currently, it is in Phase 2a, thusMMV048 may be the first Plasmodium kinase inhibitor to reachtherapy.Other class of PI4K inhibitor, aminopyridine and pyrazine compounds were found in acollaboration between the University of Cape Town Drug Discovery and Development Centreand Medicines for Malaria Venture (MMV).Concluding remarks,,,,Although malaria and tuberculosis are not considered neglected tropical diseases by theWHO,Fortunately, many groups in academia, sometimes with the partnership of pharmaceuticalindustries - consortia maintained by DNDi and MMV are good examples - keep the searchfor new and better drug candidates alive, primarily for drug resistant diseases. In thissense, this literature review presents advances in the design and discovery of new andpromising molecules, some of them already in the clinical phase. It is worth noting thatthe repurposing approach has been explored, as this is much stimulated due to theadvantages it presents."}
+{"text": "Malaria -typhoid co-infection is associated with poverty and underdevelopment with significant morbidity and mortality with similarities in clinical features of the two diseases that often result in misdiagnosis and mistreatment of the febrile patients. The Co-administration of artemether lumefantrine (AL) with ciprofloxacin as treatment for malaria-typhoid co-infection is common in Nigeria and this increases risk of pharmacokinetic drug-drug interaction since ciprofloxacin is an inhibitor of CYP3A4 that metabolizes AL. In an open-label prospective three arm design with registration pactr201909811770922, one hundred and nineteen (119) febrile volunteers comprising 55 males and 64 females were distributed into three oral treatment regimen groups. Group 1 consist of sixty-five participants presenting malaria mono infection treated with AL only and fifty-four participants presenting malaria-typhoid co-infection randomly assigned to Group 2 treated with AL and ciprofloxacin concomitantly and Group 3 whose doses were staggered at 2 hours interval. Blood samples were collected from participants in the three groups on 3 different days: day 0 (before commencement of treatment); day 3 (after completion of AL); and day 7 (after completion of ciprofloxacin), The collected blood sample were used to determine parasite density, serum liver and kidney function parameters, haematological indices, and day 7 lumefantrine concentration. The data in this article provides the changes in PCR-uncorrected Early Treatment Failure (ETA), Late Clinical Failure (LCF), Late Parasitological Failure (LPF), Day 7 serum lumefantrine concentration, liver and kidney function parameters, axillary body temperature and PCV/Hb associated with the different treatment regimen. The dataset  The raw data files were deposited in Mendeley data repository 2The protocol for data collection is based an open-label prospective three arm design with registration pactr201909811770922. The main criterion for the classification of the participants into the two groups of Malaria and malaria-typhoid co-infection is clinical laboratory diagnosis based on Giemsa thick blood film microscopy and Widal antigen agglutination test. The malarial typhoid co-infected participants where further divided into two treatment sub-groups; those treated with AL orally co-administered with ciprofloxacin concomitantly (Group 2) and Group 3 treated with AL orally co-administered with ciprofloxacin at 2 hours interval. The doses were staggered by 2 hours in order assess the effect of staggering the dosage of the two drugs on the efficacy of AL on the hypothetical drug-drug interaction. This is based on the assumption that maximum gastrointestinal absorption and peak plasma concentration of artemether would have been achieved before the administration of ciprofloxacin since the short acting component of AL is absorbed within that time frame.30.53N and longitude 70.56E is the second largest city in Ebonyi State. The average temperature in the area is highest in the month of March  and lowest in the month of August with 78.10F (25.6\u00b0c). Annual rainfall ranges from 1600m to over 2000m and the driest month having less than 29m of rainfall. The population is estimated at 156,611 The dataset was collected between June and November, 2019 at the Akanu Ibiam Federal Polytechnic Unwana Medical Centre located in Afikpo North Local Government Area of Ebonyi state, Nigeria. The Centre serves the general medical needs of the entire polytechnic staff, students, staff dependents and patients living in and around the polytechnic community environment. Afikpo, situated on latitude 54Inclusion criteria were adults of both sexes between 18 and 60 years of age presenting symptoms like fever, head ache, vomiting, and weakness of the body, body pains and other symptoms indicating uncomplicated malaria and/or typhoid infection. Individuals who had taken antimalarial drug and/or antibiotics or herbal concoction within 2 weeks prior to the time of recruitment, those under treatment for any underlying diseases like diabetes, sickle cell anaemia, hepatitis, high blood pressure and those whose medical history suggest reduced susceptibility or sensitivity to ciprofloxacin were excluded. The goal was explained to the participants and they gave written consent before participating. The researchers took measures to ensure confidentiality to all individuals who participated. All collected samples and the filled confidential report forms were assigned codes as identification number.5Blood samples were collected from participants on 3 different days: day 0 (before commencement of treatment); day 3 (after completion of AL); and day 7 (after completion of ciprofloxacin), to determine changes in parasite density, serum liver function, kidney function, haematological indices, and lumefantrine concentration. On days 0 and 7, 5mls of blood were collected aseptically by vein puncture from each participant and labeled aseptically while on day 3, drops of peripheral blood were collected by finger prick method.6The drugs administered for the treatment of malaria mono-infection and malaria-typhoid co-infection is the generic artemether lumefantrine and ciprofloxacin dispensed at the Akanu Ibiam Federal polytechnic Medical Centre. The AL is fixed dose combination 80mg/480mg artemether lumefantrine. For the participants presenting uncomplicated falciparum malaria mono-infection Group 1, the first two doses of AL were administered at an interval of 8 hours while the subsequent four doses were taken at 12-hour interval for 3 days. For the volunteers presenting malaria-typhoid co-infection, the generic 500mg ciprofloxacin was orally co-administered with AL in Group 2 while in Group 3, ciprofloxacin was administered 2 hours after AL until the third day when the six dose 3 days artemether lumefantrine regimen was completed. Thereafter, the oral administration of the ciprofloxacin continued till day 7.7Malaria parasite density was determined from Giemsa-stained thick blood films prepared using peripheral blood collected from finger pricking. The prepared thick films were examined under the microscope using\u202f\u00d7\u202f100 oil immersion objective lens. Level of parasitaemia was calculated in microliter (\u03bcl) of thick blood film preparation and graded as: low\u202f+\u202f(1 to 999/\u03bcl), moderate ++ (1000 to 9999/\u03bcl) and high +++  according to WHO 8Salmonella typhi was taken as a cut of value The Widal agglutination test was performed on all blood samples by the rapid slide titration method using commercial antigen suspension for somatic (O) and flagella (H) antigens. Exactly 50\u03bcl of the blood serum was placed on eight rows of circles on the test tiles and a drop of positive and negative serum suspension were placed beside each sample on the test slide. Antibody titer of \u22651: 80 against O and H antigen of 9Randox Assay kits were used for determination of the biochemical markers of tissue toxicity and used according to manufacturer's instructions. The kits include Alanine amino Transferase (ALT) with Cat. No AL100, Aspartate Amino Transferase (AST) with Cat. No AS101), Alkaline phosphatase (ALP) with Cat. No AP 542, Bilirubin with Cat. No BR 411, Creatine with Cat. No CR 510 and Urea with Cat. No UR 1068. Serum ALT, AST, ALP, Bilirubin, creatine and urea kits were based on the methods by Reitman and Frankel 1018 column  and a gradient elution with acetronitrile/formic acid as mobile phase was used for quantification of lumefantrine. Samples were pretreated using a step protein precipitation by adding 50 \u00b5L of pyrimethamine (200 \u00b5g/mL) as an internal standard (IS) to a 250 \u00b5L aliquot of the plasma samples, vortex-mixed briefly, thereafter, 700 \u00b5L of cold acetonitrile was added to precipitate plasma protein. The mixture was vortex mixed for 1 min and centrifuged for 10 mins and 20 \u00b5L of clear supernatant from each sample was injected for analysis. The calibration curves fit linear regressions over the range of 50\u201320 000 ng/mL for lumefantrine based on the peak area ratios generated from analyte/IS peak response. The lower limit of quantification (LLOQ) for lumefantrine was 50 ng/mL. The average inter-day accuracy and precision of the lumefantrine assay in plasma were observed to range between 85.37%and 105.9%, and 4.58% and 7.21%, respectively. The average inter-day accuracy and precision of the lumefantrine assay in plasma were observed to range between 94.04% and 101.72%, and 9.38% and 11.27%, respectively.A HPLC-UV fitted with a RP Zorbax C11The primary treatment outcome is Late Parasitological Failure (LPF) defined as presence of parasitaemia between day 7 and 28 12Data were processed and analyzed using Statistical Package for the Social Sciences (SPSS version 17.0) software. Descriptive statistics such as frequency and percentages were used to summarize participants\u2019 demographic characteristics and treatment outcomes. Categorical data was analyzed using Chi squared statistics. Quantitative data on changes in blood chemistry, body temperature before treatment and on day 7 withing groups was done using paired t-test. The significance of disparity of means of the blood chemistry tests between the three groups and the day 7 serum lumefantrine concentration were analyzed using one-way analysis of variance (ANOVA). P < 0.05 was considered significant.Institutional ethical clearance AIFPU/REG/OR/102/VOL.4/416 was obtained after a review of the research protocol by the AIFPU Medical Centre board and the polytechnic Research and Ethics Committee. Written informed consent was also sought and obtained from each of the volunteer study subjects before inclusion in the study.Segun Solomon Ogundapo: Conceptualization. Segun Solomon Ogundapo, Soniran Olajoju Temidayo, Karian Chigozie Ngobidi, Ibukun Caroline Vining-Ogu, Ajuka Nwogo Obasi, Victor Uzochukwu Olugbue, Adebanjo Jonathan Adegbola, Foluke Adebimpe Ogundapo: Methodology. Segun Solomon Ogundapo, Foluke Adebimpe Ogundapo: Data curation, Writing - Original draft preparation. Soniran Olajoju Temidayo, Ajuka Nwogo Obasi, Adebanjo Jonathan Adegbola, Victor Uzochukwu Olugbue: Reviewing and Editing.The authors declare that they have no known competing financial interests or personal relationships which have, or could be perceived to have, influenced the work reported in this article."}
+{"text": "Plasmodium ATP4, is currently in clinical development. This study aimed to characterize the antimalarial activity of cipargamin in healthy volunteers experimentally infected with blood-stage Plasmodium falciparum.The spiroindolone cipargamin, a new antimalarial compound that inhibits  Plasmodium ATP4, is currently in clinical development. This study aimed to characterize the antimalarial activity of cipargamin in healthy volunteers experimentally infected with blood-stage Plasmodium falciparum. Eight subjects were intravenously inoculated with parasite-infected erythrocytes and received a single oral dose of 10\u2009mg cipargamin 7\u2009days later. Blood samples were collected to monitor the development and clearance of parasitemia and plasma cipargamin concentrations. Parasite regrowth was treated with piperaquine monotherapy to clear asexual parasites, while allowing gametocyte transmissibility to mosquitoes to be investigated. An initial rapid decrease in parasitemia occurred in all participants following cipargamin dosing, with a parasite clearance half-life of 3.99 h. As anticipated from the dose selected, parasite regrowth occurred in all 8 subjects 3 to 8\u2009days after dosing and allowed the pharmacokinetic/pharmacodynamic relationship to be determined. Based on the limited data from the single subtherapeutic dose cohort, a MIC of 11.6\u2009ng/ml and minimum parasiticidal concentration that achieves 90% of maximum effect of 23.5\u2009ng/ml were estimated, and a single 95-mg dose  was predicted to clear 109 parasites/ml. Low gametocyte densities were detected in all subjects following piperaquine treatment, which did not transmit to mosquitoes. Serious adverse liver function changes were observed in three subjects, which led to premature study termination. The antimalarial activity characterized in this study supports the further clinical development of cipargamin as a new treatment for P. falciparum malaria, although the hepatic safety profile of the compound warrants further evaluation. The spiroindolone cipargamin, a new antimalarial compound that inhibits  Despite efforts at progress toward eradicating malaria, it is still responsible for a large burden of morbidity and mortality worldwide . AchieviIn vitro studies indicated that cipargamin is active against both asexual and sexual stages of Plasmodium falciparum  was identified by high-throughput screening  and reprlciparum . The saflciparum . The comum vivax .The ability to investigate the activity of new antimalarial candidate compounds in healthy volunteers is a useful tool in the antimalarial drug development toolkit. Volunteer infection studies (VIS) using the induced blood-stage malaria (IBSM) model, where healthy adults are experimentally inoculated with blood-stage malaria parasites, allow for early characterization of a compound\u2019s pharmacological activity and tolerability in a controlled setting. Furthermore, determination of a compound\u2019s PK/PD relationship using VIS enables informed dose selection for efficacy trials in malaria patients, potentially reducing overall development timelines. VIS have been used successfully during the clinical development of several new antimalarial drug candidates \u201317.\u2013P. falciparum IBSM model in order to aid in optimizing future therapeutic dosing. A key component of this was to define the PK/PD relationship between cipargamin PK and parasite clearance in healthy volunteers. Additional objectives were to evaluate the safety/tolerability of cipargamin and investigate the ability of cipargamin to interrupt transmission of P. falciparum to mosquitoes.The primary objective of the current study was to characterize the antimalarial activity of cipargamin in a VIS using the P. falciparum-infected erythrocytes on day 0. All subjects were male, and the mean age was 27.9\u2009years  were reported, with all 8 subjects experiencing at least one AE . The mosThere were 3 serious adverse events (SAEs) reported in the study, which included the 2 cases of severe LFT abnormalities mentioned above. The third SAE was another case of abnormal LFT results , which was moderate in severity. Although a relationship between the SAEs and cipargamin was suspected, other administered drugs (paracetamol and piperaquine) and the induced malaria infection in previously malaria-naive individuals were also considered to be possible contributory factors. These serious LFT abnormalities are described in further detail below.There were no clinically significant abnormalities in biochemistry parameters other than the LFT findings mentioned above. Additionally, no clinically significant abnormalities were observed in hematology, urinalysis, or electrocardiogram (ECG) parameters. The severity of malaria symptoms and signs experienced by participants during the study assessed by the malaria clinical scoring tool are presented in Table S2 in the supplemental material. At the time of cipargamin dosing, subjects exhibited no or negligible malaria symptoms or signs . Six subjects developed mild to moderate symptoms or signs of malaria around the time of piperaquine administration in response to parasite regrowth .The subject with the most severe LFT abnormalities was a 52-year-old subject  who developed elevations in LFTs  8\u2009days after dosing with cipargamin, with a raised bilirubin level at 74\u2009\u03bcmol/liter , conjugated bilirubin at 22\u2009\u03bcmol/liter (3.1 \u00d7 ULN), aspartate transaminase (AST) at 106 U/liter (2.6 \u00d7 ULN), alanine transaminase (ALT) at 187 U/liter (4.6 \u00d7 ULN), and gamma-glutamyl transferase (GGT) at 215 U/liter (4.3 \u00d7 ULN). The subject received paracetamol as symptomatic treatment for malaria. Physical examination was notable for transient mild tender liver (duration of 3\u2009days), which began 5\u2009days after the onset of the LFT abnormality. The subject otherwise remained asymptomatic, with no clinical features of acute hepatitis (anorexia or nausea), or other evidence of decompensation . His liver tests completely normalized over a period of approximately 6\u2009weeks. Ultrasonography showed fatty liver but ruled out choledocholithiasis, a genetic test for Gilbert\u2019s syndrome was negative, and a liver FibroScan did not indicate evidence of advanced fibrosis or cirrhosis. Careful history taking by an external consultant hepatologist revealed a previous and undisclosed history of excessive alcohol intake. As the total bilirubin mostly comprised unconjugated bilirubin, the LFT elevations were inconsistent with drug-induced liver injury, being more likely explained by hemolysis or impaired bilirubin conjugation.The second case of severe abnormality in LFTs occurred in a 22-year-old subject  11\u2009days after dosing with cipargamin. A concomitant elevation in ALT, AST, and GGT was observed, with a peak ALT value of 964 U/liter (24.1 \u00d7 ULN), a peak AST value of 443 U/liter (11 \u00d7 ULN), and a peak GGT value of 159 U/liter (3.1 \u00d7 ULN). No elevation in bilirubin was observed. The subject received paracetamol for malaria symptoms, and the elevated liver function tests normalized over the course of approximately 2\u2009weeks. Tests for hepatitis, cytomegalovirus, and Epstein-Barr virus were negative, and liver and abdominal ultrasounds were normal. Furthermore, analysis of the subject\u2019s medical history did not reveal any conditions or medication use that may have contributed to the abnormal liver function tests.A third case of moderate abnormality in LFTs occurred in an 18-year-old subject  8\u2009days after dosing with cipargamin. Elevated ALT was observed (maximum of 208 U/liter [5.2 \u00d7 ULN]), along with a mild elevation in AST (maximum of 88 U/liter [2.2 \u00d7 ULN]); no elevation in bilirubin was observed. The subject remained asymptomatic, and the ALT normalized over the course of approximately 2\u2009weeks without treatment. Analysis of the subject\u2019s medical history did not reveal any conditions or medication use that may have contributed to the ALT elevation.Together, these three cases of elevated LFT levels resulted in the study being terminated by the sponsor.The intersubject variability in plasma cipargamin concentration-time profiles was low to moderate, with the coefficient of variation (CV) less than 40% for all PK parameters . Absorpt10 parasite reduction ratio over a 48-h time period postdose (PRR48) was 3.63 , which corresponded to a parasite clearance half-life of 3.99\u2009h . The individual subject log10 PRR48 and clearance half-life are presented in Table S3 in the supplemental material. Parasite regrowth occurred relatively rapidly in all subjects; piperaquine was administered to all subjects between 3 and 8\u2009days after cipargamin dosing to clear asexual parasites (The geometric mean parasitemia (measured using quantitative PCR [qPCR]) upon dosing with 10\u2009mg cipargamin was 9,687 parasites/ml . A reduction in parasitemia was observed within 24\u2009h of cipargamin administration  and B. Tarasites .Emax) model best characterized the effect of cipargamin on parasitemia . Visual predictive checks of the PK model  and PK/PD model  confirmed the adequacy of the models in characterizing the observed data. The efficacy parameters associated with administration of 10\u2009mg cipargamin are presented in Table S6 in the supplemental material. The PK/PD model estimated the median MIC of cipargamin to be 11.6\u2009ng/ml  and the median minimum parasiticidal concentration that achieves 90% of the maximum effect (MPC90) to be 23.5\u2009ng/ml . Simulations of various single doses of cipargamin  in patients with malaria  resulted in an estimate of 95\u2009mg  as the minimum effective dose needed to clear 109 parasites/ml.A two-compartment disposition model with first-order absorption and elimination and no lag time was developed to describe the pharmacokinetics of cipargamin . A maximum killing effect , although these elevations were not considered to be related to cipargamin because the values were already slightly elevated at screening . No clinically relevant increases in bilirubin level were observed in these patients. When cipargamin was administered to a total of 25 patients with P. falciparum malaria as single doses of 10, 15, 20, or 30\u2009mg, only one patient displayed increased ALT, although 40% of patients displayed increased blood alkaline phosphatase and 20% of subjects displayed hyperbilirubinemia  and to correlate the outcome with pharmacokinetics . In the first cohort of 11 patients treated with a single 75-mg dose, four patients had asymptomatic transient ALT elevations (>3 \u00d7 ULN). The sponsor terminated the study after the first dose cohort and initiated a dedicated dose escalation safety study to explore the hepatic profile of cipargamin in patients with acute uncomplicated P. falciparum malaria . This study has recently been completed, and the results will be published separately.A third phase 2 study (currently unpublished) in Thai and Vietnamese patients with P. falciparum or P. vivax infection indicated that LFT changes are not uncommon if testing is done in the 7- to 10-day window after treatment   was around 3-fold lower than was observed in the patients with naturally acquired malaria  and similar to the exposure observed in healthy subjects dosed with 10\u2009mg cipargamin in the first-in-human study    . However \u00b7 h/ml) . FurtherEmax) and to investigate a dose-dependent killing effect. Thus, the MIC, MPC90, and minimum effective single dose reported here are likely to be overestimations. Indeed, the estimated MIC of 11.6\u2009ng/ml was substantially higher than was estimated in patients with P. falciparum malaria (0.1\u2009ng/ml) where different doses  were tested , allowing the PK/PD relationship between parasitemia and cipargamin plasma concentration to be determined. However, the aforementioned limitation of the current study with respect to the enrollment of only a single-dose cohort is likely to have influenced the accuracy of these estimates. Since only one low dose was tested, it was not possible to achieve the higher exposure required to estimate the maximum killing effect , allowing gametocytemia development to be investigated. Gametocytemia was observed in all subjects following piperaquine treatment, indicating that a single low dose of 10\u2009mg cipargamin was not sufficient to prevent gametocyte development. Cipargamin was shown to be a potent inhibitor of both early- and late-stage gametocyte development in vitro . The inain vitro  since transmission is very unlikely to occur below 1,000 gametocytes/ml . It is pin vitro  warrantsin vivo antimalarial activity characterized in this study supports the further clinical development of cipargamin as a new treatment for P. falciparum malaria. The hepatic safety profile of cipargamin will be further evaluated in clinical development of the compound.In conclusion, the P. falciparum IBSM study conducted at Q-Pharm Pty. Ltd. . Healthy males and females  of ages 18 to 55\u2009years were eligible for inclusion in the study. Individuals were excluded if they had visited an area where malaria is endemic for a period greater than 4\u2009weeks in the past 12\u2009months or had received systemic therapy with a drug with potential antimalarial activity in the past 4\u2009weeks. A complete list of the protocol-specified inclusion and exclusion criteria is included in Text S1 in the supplemental material. All subjects gave written informed consent before being included in the study. This study was approved by the QIMR Berghofer Medical Research Institute Human Research Ethics Committee and was registered at ClinicalTrials.gov with registration no. NCT02543086.This was a phase 1b, open-label, P. falciparum 3D7-infected human erythrocytes  as previously described  was administered to subjects in response to parasite regrowth. The purpose of piperaquine treatment was to clear asexual parasites without clearing any gametocytes that may be present. This would allow the gametocytocidal activity of a second dose of cipargamin to be investigated, as well as the transmission of gametocytes to mosquitoes. Gametocytemia was monitored using quantitative reverse transcriptase PCR (qRT-PCR) targeting etocytes . Transcretocytes . All parThis study was planned to be conducted in two parts, with up to three-dose cohorts enrolled in part A following the procedures outlined above. Part B was planned to consist of single-dose cohort to specifically test the activity of a single dose of cipargamin against gametocytes. In this cohort, following inoculation, 480\u2009mg piperaquine would be administered to clear asexual parasites when parasitemia reached a threshold of \u22655,000 parasites/ml, and cipargamin would subsequently be administered at peak gametocytemia (expected to occur approximately 15\u2009days after piperaquine treatment). However, due to the occurrence of several serious adverse events (SAEs) following the initial dosing of cohort 1 in part A (see the \u201cSafety\u201d section in Results), the planned second dose of cipargamin was not administered, and the study was terminated without enrolling additional cohorts in part A or proceeding to part B.Common Terminology Criteria for Adverse Events, version 4.03 (https://evs.nci.nih.gov/ftp1/CTCAE/CTCAE_4.03/CTCAE_4.03_2010-06-14_QuickReference_5x7.pdf).Safety assessments were performed throughout the study from screening and included adverse event reporting, physical examination, vital signs, clinical laboratory evaluation , electrocardiography (ECG), and malaria clinical score recording. The malaria clinical score served as a clinical indication of the severity of the induced malaria infection; 14 signs and symptoms commonly associated with malaria were graded using a 3-point scale , and the values were summed in order to generate an overall score (with a maximum possible score of 42). The severity of AEs was graded in accordance with the 0\u201324 h), up to the last time point measure (AUC0\u2013last), and extrapolated to infinity (AUC0\u2013\u221e), the elimination half-life (t1/2), the peak plasma concentration (Cmax) and the time to Cmax (tmax), the apparent clearance (CL/F), and the apparent volume of distribution during the terminal phase (zV/F).Blood samples were collected at the following time points after dosing to determine the cipargamin plasma concentration: 1, 2, 3, 4, 8, 12, 16, 24, 36, 48, 72, 96, and 120\u2009h. Samples were analyzed by liquid chromatography-tandem mass spectrometry as previously described (10-transformed cohort-specific PRR over a 48-h time period [log10 PRR48]). Analyses were performed in R version 3.3.0. The PRR and parasite clearance half-life were estimated using the slope of the optimal fit for the log-linear relationship of the parasitemia decay as described previously  and parasite clearance half-life, with the former expressed as the ratio of the parasite density decrease over a 48-h time period (expressed as the logeviously .90) were derived from the PK/PD model. Simulations were performed to predict the minimum effective single dose, defined as the dose that clears 109 parasites/ml. Further detail on PK/PD modeling methods is presented in Text S1.Data preparation, exploration, model definition, model evaluation and simulations were performed in R (version 3.5.1) and the R package IQRtools  within the software package MonolixSuite 2018R2 . The MIC and the minimum parasiticidal concentration that achieves 90% of maximum effect (MPCAnopheles stephensi mosquitoes was measured using a direct membrane feeding assay (DMFA) as previously described  was comparable to those of previous P. falciparum IBSM challenge studies and based on previously published experience was considered sufficient for obtaining statistically meaningful data on the effects of cipargamin on malaria parasite kinetics.The sample size of the current study ("}
+{"text": "Background: Cocaine is a psychostimulant drug used as performance enhancer throughout history. The prolonged use of cocaine is associated with addiction and a broad range of cognitive deficits. Currently, there are no medications proven to be effective for cocaine-use disorder (CocUD). Previous preliminary clinical work suggests some benefit from repetitive transcranial magnetic stimulation (rTMS) stimulating the prefrontal cortex (PFC), involved in inhibitory cognitive control, decision-making and attention. All published studies to date have been limited by small sample sizes and short follow-up times.Methods: This is a retrospective observational study of 284 outpatients (of whom 268 were men) meeting DSM-5 criteria for CocUD. At treatment entry, most were using cocaine every day or several times per week. All patients underwent 3 months of rTMS and were followed for up to 2 years, 8 months. Self-report, reports by family or significant others and regular urine screens were used to assess drug use.Results: Median time to the first lapse (resumption of cocaine use) since the beginning of treatment was 91 days. For most patients, TMS was re-administered weekly, then monthly, throughout follow-up. The decrease in frequency of rTMS sessions was not accompanied by an increase in lapses to cocaine use. Mean frequency of cocaine use was <1\u00b70 day/month (median 0), while serious rTMS-related adverse events were infrequent, consistent with published reports from smaller studies.Conclusions: This is the first follow-up study to show that rTMS treatment is accompanied by long-lasting reductions in cocaine use in a large cohort. However, cocaine has been a focus of attention on a global scale for the serious harms related to its use, including addiction and cognitive dysfunctions. Currently, no medications have been proven to be effective for cocaine use disorder (CocUD). The traditional strategy has been to develop medications or psychological interventions to attenuate drug reward, which is mainly mediated by the dopaminergic pathway from the ventral tegmental area (VTA) to the nucleus accumbens. This approach has not resulted in effective therapeutic interventions for cocaine addiction. Recently, repetitive transcranial magnetic stimulation (rTMS) has received increasing attention as a potential treatment for CocUD . As we aClinical confirmation that rTMS is effective for CocUD awaits results from randomized controlled trials that are now in progress. Studies published to date are limited by small sample sizes and short duration of follow-up. In almost half the published studies using rTMS or a similar intervention for CocUD, there was, strictly speaking, no follow-up: responses  were assessed only within the laboratory on the day of stimulation \u20139. In 6 Here we report results from a cohort of 284 cocaine users who received rTMS and were then followed for up to 2 years, 8 months (median 164 days). This is a retrospective chart review, with all the attendant limitations thereof. However, it is also unprecedented in its size and duration. Until there are results from large randomized trials, the data we report here provide the strongest evidence to date that rTMS is well-tolerated and possibly effective in people with CocUD.Patients signed informed consent on the day of clinic intake and agreed that their data could be used for research. Patients were informed that the data collected would be processed in accordance with the law on privacy and in compliance with Legislative Decree No. 196 of June 30, 2003, \u201cPersonal Data Protection Code,\u201d ensuring anonymity. The data were extracted from patient clinical records and anonymized for analysis. All subjects gave their informed consent for inclusion before they participated in the study. This was a retrospective chart review of data from 284 men and women who were treated from 2013 to 2017 and followed for at least 12 weeks after the first week of rTMS sessions. The protocol, limited to the retrospective chart review, was approved by the Ethical Committee for the Psychological Research, Departments of Psychology, University of Padua (Protocol 2551) and the study was conducted in accordance with the Declaration of Helsinki. The total of 284 includes 58 patients who were lost to follow-up within the first 12 weeks  but excludes 44 patients for whom 12 weeks had not yet elapsed when data analysis started. All patients voluntarily underwent treatment for CocUD in an rTMS protocol at a clinic center for addiction treatment in Padua, Italy. At intake, each patient was assessed by a psychiatrist and a psychologist with expertise in the treatment of addiction. A complete family, physiological, remote pathological and near pathological history was collected, in addition to a detailed psychiatric, toxicological and clinical history. Patients were between 18 and 70 years of age and met DSM-5 criteria for CocUD. A published screening form was administered to all the patients to exclude contraindications to rTMS . Each paThe primary outcome measure was lapse to cocaine use during follow-up. Cocaine use was assessed through a combination of urine screening, self-report, and reports by collateral informants . As in our published pilot study , the \u201czeBecause this is a retrospective chart review, results are presented descriptively. For the sample as a whole, we used Kaplan-Meier survival analysis (SAS Proc Lifetest) to calculate the median number of days until the first lapse to cocaine use. Data were coded as right-censored for patients who were still abstinent at the end of monitoring (~44% of censored cases) or with whom the clinic lost contact (~56% of censored cases). To help contextualize the \u201cfirst lapse\u201d findings, we display them together with historical control data from an outpatient cohort of 173 cocaine users in the US who were undergoing group and/or individual psychotherapy after discharge from inpatient treatment 2. The twThe funders of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report. The corresponding author had full access to all the anonymized data in the study and had final responsibility for the decision to submit for publication.Demographic and drug-use data are shown in For the 284 patients in the whole sample, the duration of follow-up ranged from 4 to 989 days ; median was 164 days (just over 5 months). Time to the first lapse is shown in In the 147 closely followed patients, the duration of follow-up ranged from 84 days (12 weeks) to 974 days ; median was 217 days (just over 7 months). Time courses of rTMS and cocaine use are shown in n = 23), hypomania (n = 4), anxiety (n = 2) irritability (n = 2), dental pain (n = 2), scalp discomfort during the first 2 weeks of sessions (n = 1), angioedema and urticaria (n = 1), distractibility (n = 1), dizziness (n = 1), nausea (n = 1), nausea and numbness (n = 1), seizure (n = 1), and a hypomanic episode (n = 1). The seizure occurred in a 27-year-old woman 66 days after her first rTMS session. She has used cocaine shortly before; she had not recently undergone rTMS. The hypomanic episode occurred in a 37-year-old man, just under 90 days after his first rTMS session. His family reported that he had begun speaking aloud to himself without realizing it. rTMS was suspended for medical examinations, which did not show any abnormalities. Other AEs were transient and resolved spontaneously or with over-the-counter medications.Adverse events (AEs) were reported by 41 of the 284 patients. No patient reported more than one. AEs reported were: headache  for cocaine addiction, especially when considering large cohorts of patients and long period of observation. Nevertheless, we can argue that clinical outcomes, including lapse rate, may show a significant difference compared to conventional treatment for addiction. Compelling evidence from preclinical and clinical studies indicates that rTMS influences neural activity in the short and long term by mechanisms involving neuroplasticity and resulting in substantial behavioral changes , 26, 27.Human laboratory studies with rTMS suggest that the site we stimulated, left DLPFC, might also be an appropriate target for people with addictions to heroin , methampClinicalTrials.gov identifiers NCT03607591, NCT03333460, and NCT02986438), is represented by sham-controlled randomized trials with sufficient sample size and follow-up duration .The datasets for this article are not publicly available to protect proprietary information. Requests to access the datasets should be directed to LGa . The patients/participants provided their written informed consent to participate in this study.AT and LGa: conceptualization. GM, SC, and LG\u00f3: data curation. SC: formal analysis. SC, GM, and LG\u00f3: methodology. AT: project administration. LGa: supervision. GM, AT, SC, LG\u00f3, NC, MS, and LGa: validation. GM, SC, LG\u00f3, NC, MS, and LGa: visualization. GM: writing\u2014original draft. GM, AT, SC, LG\u00f3, NC, MS, AB, and LGa: writing\u2014review and editing.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Plasmodium falciparum represents an increasing threat to Africa. Extended ACT regimens from standard 3 to 6\u00a0days may represent a means to prevent its development and potential spread in Africa.Artemisinin-based combination therapy (ACT) resistant P. falciparum malaria, enrolled in Bagamoyo district, Tanzania. The study evaluated parasite clearance, including proportion of PCR detectable P. falciparum on days 5 and 7 (primary endpoint), cure rate, post-treatment prophylaxis, safety and tolerability. Clinical, and laboratory assessments, including ECG were conducted during 42\u00a0days of follow-up. Blood samples were collected for parasite detection (by microscopy and PCR), molecular genotyping and pharmacokinetic analyses. Kaplan\u2013Meier survival analyses were done for both parasite clearance and recurrence.Standard 3-day treatment with artemether\u2013lumefantrine (control) was compared to extended 6-day treatment and single low-dose primaquine (intervention); in a randomized controlled, parallel group, superiority clinical trial of patients aged 1\u201365\u00a0years with microscopy confirmed uncomplicated A total of 280 patients were enrolled, 141 and 139 in the control and intervention arm, respectively, of whom 121 completed 42\u00a0days follow-up in each arm. There was no difference in proportion of PCR positivity across the arms at day 5 (80/130 (61.5%) vs 89/134 (66.4%), p\u2009=\u20090.44), or day 7 (71/129 (55.0%) vs 70/134 (52.2%), p\u2009=\u20090.71). Day 42 microscopy determined cure rates (PCR adjusted) were 97.4% (100/103) and 98.3% (110/112), p\u2009=\u20090.65, in the control and intervention arm, respectively. Microscopy determined crude recurrent parasitaemia during follow-up was 21/121 (17.4%) in the control and 14/121 (11.6%) in the intervention arm, p\u2009=\u20090.20, and it took 34\u00a0days and 42\u00a0days in the respective arms for 90% of the patients to remain without recurrent parasitaemia. Lumefantrine exposure was significantly higher in intervention arm from D3 to D42, but cardiac, biochemical and haematological safety was high and similar in both arms.P. falciparum infections but, importantly, equally efficacious and safe. Thus, extended artemether\u2013lumefantrine treatment may be considered as a future treatment regimen for ACT resistant P. falciparum, to prolong the therapeutic lifespan of ACT in Africa.Extended 6-day artemether\u2013lumefantrine treatment and a single low-dose of primaquine was not superior to standard 3-day treatment for ACT sensitive Trial registration ClinicalTrials.gov, NCT03241901. Registered July 27, 2017 https://clinicaltrials.gov/show/NCT03241901 Plasmodium falciparum resistance to anti-malarial drugs, e.g., chloroquine and sulfadoxine\u2013pyrimethamine, has evolved in South-East Asia, and spread via the Indian sub-continent to Africa [P. falciparum malaria in Africa, and significantly contributed to the decline in malaria burden [Historically, o Africa  with devo Africa , 3. Duria burden . Howevera burden , 5. Morea burden \u20138. This P. falciparum clearance time after ACT (microscopy positivity rate on day 3 of \u2265\u200910%) [P. falciparum kelch 13\u00a0propeller gene (pfk13) [pfk13 mutations in Africa, although without affecting the efficacy of artemether\u2013lumefantrine, the most commonly used ACT in Africa [Artemisinin resistance, defined as partial resistance by the WHO, is phenotypically characterized by prolonged f \u2265\u200910%) . Resista pfk13) , 9\u201312. Sn Africa . From th , 9\u201312. P. falciparum malaria since 2006. Several observations from Bagamoyo district contribute to this concern. Firstly, there are reports of polymerase chain reaction (PCR) determined positivity rate on day 3 in the magnitude of 28\u201384% after supervised artemether\u2013lumefantrine treatment [P. falciparum sub-populations showed microscopy determined parasite clearance curves similar to artemisinin resistant parasites in Cambodia [P. falciparum population, without compromised cure rates [P. falciparum multidrug resistance 1 (pfmdr1) gene and, the K76 SNP in the P. falciparum chloroquine resistance transporter (pfcrt) gene [Although microscopy determined parasite clearance and overall average therapeutic efficacy rates has remained above 98% across Africa for artemether\u2013lumefantrine , 14, 15,reatment , 17. UsiCambodia . Secondlre rates . These Srt) gene \u201322. Durirt) gene . The selA potential strategy to protect the therapeutic life span of ACT, in an era of imminent risk of ACT resistance, would be to extend treatment duration. Dosage prolongation of artemether\u2013lumefantrine from 2 to 3\u00a0days has been previously implemented to prevent treatment failures and resistance development from suboptimal dosing , 25. PreP. falciparum gametocytocidal drug; for blocking transmission in low-transmission areas in combination with ACT irrespective of glucose-6-phosphate dehydrogenase enzyme status [P. falciparum drug resistance development and spread [In this study, the efficacy and safety of doubling the dose, i.e., from 3 to 6\u00a0days, of artemether\u2013lumefantrine was evaluated, since this is logistically pragmatic and feasible to implement with the current packaging of the drug. Additionally, a 6-day artemether\u2013lumefantrine treatment regime will cover three erythrocytic cycles with adequate concentration of the artemisinin component to kill parasites demonstrating a delayed clearance. Moreover, the World Health Organization (WHO) recommends use of single low-dose primaquine (0.25\u00a0mg/kg), a e status . Gametocd spread \u201333. Prevd spread . Howeverd spread . In thisd spread .P. falciparum malaria in Bagamoyo district, Tanzania.The overall aim of this study was to compare PCR-determined parasite clearance, cure rate, post-treatment prophylaxis and safety, of standard treatment (3\u00a0days) versus an extended treatment (6\u00a0days) of artemether\u2013lumefantrine with single low-dose primaquine for uncomplicated This was a two-armed, randomized controlled, parallel group, superiority clinical trial, with allocation ratio 1:1. Blinding to treatment arm allocation was done to investigators and staff who were not involved in study drug administration. The first day of screening and enrollment is referred to as day 0 (D0), with D0I referring to screening sample time point, and D0II as enrolment sample time point and D0III as second visit of day 0. For subsequent visits from day 1 (D1) onwards, AM was used to refer to the first visit of the day and PM referring to the second visit. The study was conducted between July 2017 and March 2018. The study has been registered at clinicaltrials.gov (identifier: NCT03241901).P. falciparum on D5 and D7 in the respective arms. Secondary outcomes included fever and microscopy parasite clearance times, crude and PCR corrected cure rates, post treatment prophylaxis, selection of genetic drug resistance markers and safety.The primary outcome was the proportion of PCR detectable P. falciparum positivity rate of 15% on D5 . There was no data available on PCR positivity rates beyond day 3 from the study site. PCR positivity rates were therefore predicted, based on the day 3 PCR positivity rates from previous studies [The sample size was calculated based on an assumed clinically meaningful difference between the arms in PCR determined  studies , to declPrior to commencement of the study, computer-based randomization was done using R studio program , version 1.1.456. A randomization list stratified by study site was generated by a statistician for both treatment arms. For allocation concealment, opaque envelopes were serially numbered and treatment arm allocation cards were inserted according to the list. The envelopes were sealed and the study nurse who was responsible for patient enrollment wrote the unique patient ID on top of the sealed envelope. This envelope was opened by the study clinician only, to determine the treatment arm allocation and for dispensing study drugs.P. falciparum is the predominant malaria species [The study was conducted in Bagamoyo district, Coast region, Tanzania. Malaria transmission in Bagamoyo district is considered moderate, occurring throughout the year with peaks related to the long rainy season in May to July, and short rains in November to December . P. falc species .Yombo and Fukayosi public primary health care facilities were purposely selected as study sites. These rural sites provide basic health care services for the residents in their respective catchment area and have served as research sites for previous studies , 19. TheAll patients presenting at the health care facilities with fever (defined as axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C) or history of fever in the last 24\u00a0h were screened using a malaria rapid diagnostic test (RDT)  Ag RDT, Access Bio, Inc. NJ, USA). Thick and thin blood films were obtained for microscopy determined parasite counts and species identification from RDT positive patients, who were then screened for study eligibility.P. falciparum mono-infection irrespective of parasite density, were between 1 and 65\u00a0years old, with a body weight >\u200910\u00a0kg, and electrocardiogram (ECG) determined QTc interval between 360 and 440\u00a0ms in males and 370\u2013460\u00a0ms in females.Participants providing written informed consent were eligible if they had microscopy confirmed Individuals were excluded from enrollment if they had symptoms/signs of severe illness, severe malnutrition, were pregnant, breastfeeding, or unwilling to practice birth control during participation in the study, had haemoglobin levels\u2009<\u20098\u00a0g/dL, were allergic to the study medications or on regular medication which may interfere with anti-malarial pharmacokinetics, reported anti-malarial intake within the last 2\u00a0weeks, or had had a blood transfusion within the last 90\u00a0days.\u00ae, Novartis Pharma, Basel, Switzerland), administered twice a day , as per Tanzania national treatment guidelines for uncomplicated P. falciparum malaria [\u00ae, Sanofi-aventis Canada Inc.) administered with the last dose of artemether\u2013lumefantrine. All patients received directly observed therapy for all drug doses. Children unable to swallow solid artemether\u2013lumefantrine tablets received dispersible tablets suspended in water. Primaquine was administered by suspending 15\u00a0mg tablets in 15\u00a0mL of water and measured using a sterile syringe as previously described [Patients allocated to the control arm received standard treatment, i.e., a weight-based, 3-day course of artemether\u2013lumefantrine 20/120\u00a0mg/kg  for initial reading. During enrollment (circa 1\u00a0h after screening), and all slides obtained at follow-up visits, were stained slowly with 2.5\u20133% Giemsa for 45\u201360\u00a0min. Parasites were counted against 200 white blood cells (WBC), and converted to parasite density per microliter (p/\u00b5L) assuming 8000\u00a0WBC/\u00b5L of blood. A blood smear was considered negative after examining 100 high-power fields or counting 500 WBC with no parasites seen. Each slide was read by two independent and experienced microscopists, and upon disagreement on presence of parasites or if density differed by more than 25%, the slides were subjected to a third independent and decisive reader. The mean parasitaemia of the two most concordant readings were used as final parasite densities.ECG machines  at Yombo and in Fukayosi  were used to measure the QTc intervals at D0, before treatment initiation, and at D5 for all participants, corresponding to 2\u00a0h after the 12th and final dose of artemether\u2013lumefantrine for patients allocated to the intervention arm. The ECG machines were set to provide heart rate-corrected QT interval (Bazzet\u2019s method).Haemoglobin concentration was measured on D0 and D7, using a portable spectrophotometer, HemoCue Hb 201+ . Urine samples were collected and evaluated for haematuria at D0 and D7, using the CYBOW  urinalysis reagent test strips.Venous blood (2\u00a0mL) were collected at enrollment and D7, to assess liver integrity by alanine aminotransferase (ALAT), aspartate aminotransferase (ASAT), serum bilirubin levels and kidney (creatinine) integrity. The samples were stored for a maximum of 48\u00a0h in the field refrigerator (4\u00a0\u00b0C) before transport to an ISO certified reference laboratory at the Bagamoyo Research and Training Unit  for analysis. The biochemistry analyses were done using COBAS INTEGRA 400 plus . The values were compared with age specific normal ranges .For artemether\u2013lumefantrine pharmacokinetics analysis, a total of 40 patients were randomly selected to contribute blood samples, 20 from each treatment group. Venous sample of 3\u00a0mL was collected in heparinized tubes, at two randomly selected time-points in 28\u00a0days of follow up. Venous samples were also collected from all patients with microscopy determined recurrent parasitaemia during follow-up, to assess lumefantrine plasma levels before re-treatment. Data were analysed by population pharmacokinetics modelling; the estimated parameters were used to simulate individual patient concentration time profiles from time 0 to 1008\u00a0h. The resulting profiles were used to compute maximum drug concentration in ng/mL (Cmax) and concentration at days 2, 3, 4, 5, 6, 7, 14, 21, 28, 35 and 42 for both lumefantrine and desbutyl-lumefantrine between treatment arms.Study withdrawal criteria included consent withdrawal, concomitant self-treatment with any medicine having anti-malarial properties outside the study protocol, and/or any other protocol violation. Study participants were categorized as lost to follow-up and eventually withdrawn if they missed a scheduled follow-up visit and did not attend on the successive 2\u00a0days despite efforts to trace them at their homes. A participant who missed a visit, but returned before the last day of scheduled follow-up was not considered lost to follow-up. Patients with symptoms/signs of severe disease or recurring vomiting of study medicine were withdrawn from the study and treated with parenteral artesunate 2.4\u00a0mg/kg 8\u00a0h for 24\u00a0h, followed by artemether\u2013lumefantrine for 3\u00a0days.Blood samples were collected on PerkinElmer 226 filter paper  during all patient visits for parasite detection and genotyping by PCR. Filter-paper blood samples were labelled, air-dried at room temperature for 3\u20134\u00a0h and then packed in individual Ziploc plastic bags with desiccants. The dried blood spots (DBS) were stored in room temperature until shipment to Karolinska Institutet, Stockholm, Sweden, in April 2018 where PCR analyses were conducted.\u00ae-100  boiling method as previously described [P. falciparum detection and quantification was conducted by 18\u00a0s quantitative PCR (qPCR) conducted in triplicate [P. falciparum detection for all 20 sampling time points during the 42-day study period. The subset included all patients that had recurrent parasitaemia, and 46 patients (23 from each arm) randomly selected among those that did not have recurrent parasitaemia.DNA extraction from DBS collected on days 3, 5, 7, 28 and 42 was done using the chelexescribed . P. falciplicate . SamplesP. falciparum clones present in the initial infection [glurp) PCR according to previously established protocols [All patients with microscopy determined recurrent parasitaemia during follow-up were subjected to stepwise genotyping from paired blood sampling. Blood samples collected on the day of recurrent parasitaemia were compared with two consecutive time points at D0 and D1, by genotyping the merozoite surface proteins msp-2 and msp-1 and the glutamate-rich protein (glurp), to distinguish recrudescence (treatment failure) from reinfection (new infection) , 47. Tworotocols .pfmdr1 N86Y SNP was done by PCR-restriction fragment length polymorphism based methods as previously described [pfk13 propeller region was done as previously described [pfk13 nested PCR. The 858 base pairs covering the six propeller domain were sequenced, with P. falciparum 3D7 as a Ref. [Genotyping of escribed , 50. Thiescribed , in a sus a Ref. .Analyses was done using both intention-to-treat and per-protocol approaches. Proportions were calculated with 95% confidence intervals (CI) and compared by Chi-square test, whilst medians were compared by the Mann\u2013Whitney U test. For parasite densities, geometric means were determined and compared using Wilcoxon rank-sum test. Survival analysis was done to compare post treatment prophylaxis between the treatment arms using Kaplan\u2013Meier survival curves, and mean survival time was compared by log-rank. Kaplan\u2013Meier analysis for PCR adjusted cure rates was also conducted. Analyses were conducted in STATA 15.0IC ; statistical significance was defined as p\u2009<\u20090.05.A total of 280 patients were enrolled in the study, 141 in the control arm and 139 in the intervention arm, of whom 121 completed 42\u00a0days follow-up in each arm  and 41.1\u00a0days (95% CI 40.4\u201341.8) in the control and intervention arm, respectively (log rank p\u2009=\u20090.17) . Among the microscopy determined recurrent infections, PCR success rate for pfmdr1 N86Y was 28/35 (80.0%), all of which were pfmdr1 N86  was higher in the intervention arm after D3 to D42, and the difference was highest by D7; control arm 734.8\u00a0ng/mL (95% CI 601.8\u2013867.8) and intervention arm 5432.1\u00a0ng/mL (95% CI 4403.9\u20136460.3), p\u2009<\u20090.0001  and in intervention arm 14,563.7\u00a0ng/mL , p\u2009=\u20090.76. For desbutyl-lumefantrine, mean Cmax in control arm was 53.5\u00a0ng/mL (95% CI 44.6\u201362.3) and in intervention arm 73.3\u00a0ng/mL (95% CI 58.3\u201388.2), p\u2009=\u20090.05. There was no difference in mean (95% CI) Cmax for both lumefantrine and desbutyl-lumefantrine between patients with vs without microscopy determined recurrent parasitaemia within arms . There were no reported adverse events related to cardiac disorders. The most commonly reported adverse events in both treatment arms, irrespective of cause, were abdominal pain, asthenia, fever, nausea, vomiting and headache Table\u00a0. Both trAt D7, the median (IQR) haemoglobin values were 11.5 (10.1\u201312.7) g/dL for the control and 11.4 (10.2\u201312.7) g/dL for the intervention arm, respectively (p\u2009=\u20090.80). The lowest recorded haemoglobin level at D7 was 6.8\u00a0g/dL from a 6\u00a0years old patient in the control arm whose baseline haemoglobin was 9.9\u00a0g/dL and baseline parasitaemia 168,720\u00a0p/\u00b5L. This patient had recovered fully by the end of follow-up period. Additional analysis of median change in haemoglobin concentration g/dL between D0 and D7 by age groups and sex for control and intervention arms was done, and demonstrated no difference  ms and 414 (SD 20.6) ms, respectively (p\u2009=\u20090.004). However, on D5, the mean QTc interval were similar in the control and intervention arm, 416 (SD 25.8) ms and 416 (SD 22.7) ms, respectively (p\u2009=\u20090.49). The difference in mean QTc change between the control and intervention arm, \u2212\u20096.4 (SD 25.6) ms and 3.1 (SD 22.1) ms, respectively, was statistically significant, 9.5\u00a0ms (p\u2009=\u20090.006).About 36% of the patients in each of the arms showed elevated total bilirubin levels before treatment initiation, which could be associated with disease process. By the second evaluation at D7, majority of the patients had normalized their high baseline levels after treatments. Proportion of patients with elevated liver enzymes on D7 was 6/114 (5.3%) in the control and 9/101 (8.9%) in the intervention arm (p\u2009=\u20090.30). The corresponding proportion of patients with increased creatinine levels on D7 was 2/118 (1.7%) and 2/110 (1.8%) in the control and intervention arm, respectively (p\u2009=\u20090.94). These elevated levels were within the Common Toxicity Criteria (CTC) Grade 1 and these patients had elevated enzymes levels at baselines. All the patients completed study follow-up with full recovery.P. falciparum parasite population in Bagamoyo district, Tanzania, the extended 6-day artemether\u2013lumefantrine treatment and a single low-dose primaquine administered on 6th day, was not superior to standard 3-day artemether\u2013lumefantrine treatment. However, microscopy determined parasite clearance, cure rates, and safety profiles were excellent and similar between the treatment arms.The results of this study did not reveal a significant difference in PCR positivity or PCR determined parasite clearance between the treatment arms at both, D5 and D7. Thus, in the present context of an ACT sensitive P. falciparum resistance preparation. This was done by testing a new, but simple treatment strategy with old tools, which potentially could play a future role in protecting/prolonging the therapeutic life span of artemisinin-based combinations as lifesaving drugs in Africa, by simply doubling the treatment duration of artemether\u2013lumefantrine. Given that the extended treatment had excellent efficacy and safety profile, it could be rolled out as policy in case ACT resistant P. falciparum enters East-Africa from South-East Asia or evolves independently, as has recently been reported in Rwanda [P. falciparum malaria. This strategy could be used while awaiting development of new efficacious anti-malarial drugs. This study\u2019s results are similar to a smaller study conducted in pregnant and non-pregnant women in Congo, where the efficacy of both 3 and 5\u00a0days artemether\u2013lumefantrine treatment was high, and despite increased exposure to the drug, the safety profiles were excellent [Importantly, this study used consecutive day blood sampling between D3 and D7, and, thus, provided a unique opportunity to understand PCR determined parasite clearance dynamics at timepoints normally not assessed in standard ACT trials. Moreover, this study was designed and conducted, for the first time in Tanzania, as a proactive measure and part of n Rwanda . Thus, txcellent .The PCR adjusted cure rates by D42 were similarly high between the arms, 97.1% and 98.2% in the control and intervention arm, respectively. However, after 42\u00a0days of follow-up, some non-significant differences worth highlighting were observed. First, it appears that more patients were returning with recurrent parasitaemia in the control arm compared to intervention arm, 60% and 40% respectively. Secondly, it took 34\u00a0days in the control arm, and 42\u00a0days in the intervention arm for 90% of the patients to survive without recurrent parasitaemia, i.e., a difference of 8\u00a0days despite having similar mean time to recurrent parasitaemia. The extended treatment seemed to prolong the post-treatment prophylaxis period compared to standard artemether\u2013lumefantrine treatment. The hypothesis to this is that the difference would probably have been even more pronounced with longer follow-up periods beyond 42\u00a0days. This may be explained by the fact that the extended treatment arm received higher lumefantrine exposure, as shown in Fig.\u00a0There was no significant difference observed in PCR positivity and PCR determined parasite clearance between the treatment arms at both D5 and D7. The D3 PCR determined positivity in this study (81.9%) is the highest recorded in Bagamoyo district since 2006 . MoreoveP. falciparum PCR positivity with the presence of asexual ring stages and mature gametocytes, but without increased risk to treatment failure [The highly sensitive PCR methods used in this study may partly explain the increase in D3 PCR positivity over time, as compared with previous studies from the same site . Other s failure . The Ken failure .P. falciparum PCR positivity requires further understanding as to whether this represents viable and metabolically active parasites that were not responsive to extended treatment. Analysing clearance times of individual parasite clones may give a better insight as to whether persistent PCR positivity is linked to resistance or not [Persistent e or not . Furthere or not , 57.P. falciparum PCR positivity. artemether\u2013lumefantrine works synergistically with the immunity in clearing parasites through the pitting in the spleen [P. falciparum PCR determined clearance rates.DNA debris in circulation after parasites are killed or damaged by the drug may be another contributing factor to persistent e spleen , 59. Dece spleen  could bee spleen . There ie spleen , which wpfmdr1 N86 genotype linked to lumefantrine tolerance [pfmdr1 N86 has been reported to increase significantly over time [pfk13 gene.The prevalence of the olerance , was higolerance , where tver time . No mutaP. falciparum with lumefantrine tolerant genotypes, implores being on high alert for clinical resistance in this parasite population. The parasite response to the drug depends on host immunity and pharmacokinetics of the drug. These, together with the lack of markers of artemisinin resistance, may explain the excellent cure rate of artemether\u2013lumefantrine despite the presence of lumefantrine tolerant genotypes in this study [Very high prevalence of is study . The satis study . CombinaThe results of the pharmacokinetics modelling showed that the extended 6-day treatment improved lumefantrine exposure in patients within the intervention arm during the 42\u00a0days of follow-up. This increased lumefantrine exposure has the potential to cure the possibly resistant parasites. These results are consistent with recent findings from Congo that showed pregnant women who received 5\u00a0days of artemether\u2013lumefantrine had higher drug exposure compared to those who received standard 3\u00a0days treatment . There wSafety and tolerability of the extended 6-day treatment of artemether\u2013lumefantrine and a single low-dose of primaquine was excellent. The adverse events reported were not perceived to be related with drug toxicity. Extended treatment with lumefantrine did not cause any clinically significant electrocardiographic changes, even though mean QTc change was higher in the intervention arm. These results are similar to a study in Congo of artemether\u2013lumefantrine treatment during 5\u00a0days, where they found a weak correlation of increase in QTc and lumefantrine concentration . SimilarP. falciparum positivity rate on D5. There was no data available from the study site on PCR positivity rates beyond D3, so the assumed PCR positivity rates on D5 were arbitrarily set at 20% and 5% in the control and intervention arms, respectively. However, the determined D5 PCR positivity rate (>\u200960% from both arms) was unexpectedly higher than assumed, indicating that PCR-determined parasite clearance requires further understanding as discussed above. Secondly, the inability to extract RNA from the DBS for distinguishing gametocytes and asexual parasites using stage specific markers, limited the understanding of the persistent PCR positivity. Thirdly, haemoglobin assessment beyond D7 was not done, and this limit thorough assessment of impact of single low-dose primaquine on extended artemether\u2013lumefantrine regimen. Finally, since the post treatment prophylaxis in the form of time to recurrent parasitaemia was also evaluated, and this parameter could be affected by increased use of insecticide-treated mosquito nets, coverage of ITNs warrants a cautious interpretation of the results; information on individual ITN ownership and use was not collected from the study participants, however, data on ITN use were available for the region [Firstly, since artemether\u2013lumefantrine still has excellent efficacy in the study area, a significant difference in parasite clearance by microscopy or PCR adjusted cure rate was not anticipated. Instead, the use of PCR based parasite clearance as the primary outcome was chosen. The study was powered to detect an assumed clinically meaningful difference of 15% in PCR determined e region .P. falciparum population in Tanzania, but importantly equally efficacious and safe. Thus, this study, as part of ACT resistance preparedness in Tanzania, provides evidence that extended artemether\u2013lumefantrine treatment could be considered as a future treatment regimen, if/when ACT resistant P. falciparum appears, in order to prolong the therapeutic lifespan of ACT in Africa. Moreover, the results of the study underscore the importance of studying the cause of prolonged PCR positivity after artemether\u2013lumefantrine treatment, and its potential role in P. falciparum tolerance/resistance development, as well as the influence of extended artemether\u2013lumefantrine therapy on post-treatment prophylaxis beyond day 42.Extended 6-day artemether\u2013lumefantrine treatment together with a single low-dose of primaquine was not superior to standard 3-day artemether\u2013lumefantrine for the treatment of uncomplicated malaria in the ACT sensitive Additional file 1: Fig. S1. Median change in haemoglobin concentration g/dL between D0 and D7 by age groups and sex for control and intervention arms."}
+{"text": "P. vivax malaria reported byChamma-Siqueira et al. from western Brazil complicates interpretation of thetrial result. Usually chloroquine and primaquine are given together to treatP. vivax malaria. Both drugs haveactivities at the asexual stage of parasite development, but only primaquinekills the dormant liver-stage parasites (hypnozoites) that cause relapse.Although long-latency P. vivax malaria occursin the Americas, most relapses emerge from the liver approximately 14 days afterpresentation of the initial illness. The slowly eliminated chloroquinesuppresses the relapsing blood-stage infection for several weeks.Chamma-Siqueira et al. delayed administration of primaquine for a median of 17days. Because each study group received the same dose of chloroquine, theauthors were therefore comparing the asexual-stage antimalarial effects ofprimaquine against early relapse and the hypnozoitocidal effect against laterrelapses. Primaquine has relatively weak activity against asexual-stageparasites, so, unsurprisingly, the 2-week course was superior to the 1-weekcourse. Clinical trials comparing radical curative activity inP. vivax and P.ovale malaria should assess concurrent \u2014 notsequential \u2014 treatment .The unusual sequential \u201cradical cure\u201d treatment regimen for"}
+{"text": "Plasmodium resistance to some of the current anti-malarial agents makes it imperative to search for newer and effective drugs to combat malaria. Therefore, this study evaluated whether the co-administrations of xylopic acid-amodiaquine and xylopic acid-artesunate combinations will produce a synergistic anti-malarial effect.Evidence of \u22121), artesunate , and amodiaquine  were evaluated in Plasmodium berghei (strain ANKA)-infected mice to determine respective ED50s. Artemether/lumefantrine was used as the positive control. XA/ART and XA/AQ were subsequently administered in a fixed-dose combination of their ED50s (1:1) and the combination fractions of their ED50s  to determine the experimental ED50s (Zexp). An isobologram was constructed to determine the nature of the interaction between XA/ART, and XA/AQ combinations by comparing Zexp with the theoretical ED50 (Zadd). Bodyweight and 30-day survival post-treatment were additionally recorded.Antiplasmodial effect of xylopic acid  below the additive isoboles showing XA acts synergistically with both ART and AQ in clearing the parasites. High doses of XA/ART combination significantly (p\u2009<\u20090.05) increased the survival days of infected mice with a mean hazard ratio of 0.40 while all the XA/AQ combination doses showed a significant (p\u2009<\u20090.05) increase in the survival days of infected mice with a mean hazard ratio of 0.27 similar to AL. Both XA/ART and XA/AQ combined treatments significantly (p\u2009<\u20090.05) reduced weight loss.EDP. berghei.Xylopic acid co-administration with either artesunate or amodiaquine produces a synergistic anti-plasmodial effect in mice infected with  Regardless of the efforts put in place in the twenty-first century to eradicate the staggering toll of malaria on human health, the global burden of the disease remains, especially, in several tropical countries. The World Health Organization (WHO) estimates that 40% of the world\u2019s population is susceptible to malaria infections . A recenPlasmodium falciparum has developed resistance to anti-malarial agents, such as chloroquine in the past and there are reports of the growing resistance of P. falciparum to artemisinin derivatives in South-east Asia [Plasmodium falciparum sporozoite vaccine (PfSPZ), Chemoprophylaxis vaccination (CVac), Genetically-attenuated parasite vaccine (GAP), RTS,S/AS01) are at various stages of development, but a clinically approved malaria vaccine is not available, therefore [Cinchona alkaloids (quinine and quinidine) and artemisinin derivatives  are the two classes of medicines available for the treatment of severe and uncomplicated malaria. ast Asia . An anecast Asia . Some maherefore  making tDrug combination therapies (DCTs) are pertinent to the optimum control of malaria in developing countries  because Xylopia aethiopica which is used to treat malaria by Ghanaian herbal practitioners [Natural products are essential in the drug discovery process, and there is no exception in anti-malarial agents. Medicinal plant extracts have been a source for anti-malarial drug discovery for long, and their treatment for malaria has been successful . About 1itioners .Xylopia aethiopica and has been reported to possess anti-malarial properties in Plasmodium berghei-infected ICR mice. Furthermore, it significantly reduced the lipopolysaccharide\u2014(LPS) induced fever in Sprague\u2013Dawley rats [P. berghei.Xylopic acid, a kaurene diterpene, is the major constituent of the fruits of ley rats . Thus, xley rats  on the sley rats . In the Xylopia aethiopica as previously described [X. aethiopica purchased from the Ho Central Market in Ghana were shade-dried and pulverized with a hammer mill. For every 100\u00a0g of plant material, 300\u00a0ml of petroleum ether was used as a solvent for maceration. The mixture of X. aethiopica and petroleum ether was left to stand for 3\u00a0days with continuous shaking every 24\u00a0h. Whatman filter paper was used to filter the mixture and left to stand overnight under dark conditions. The filtrate was then concentrated with a Rotary evaporator  at 120 revolutions per minute and 40\u201355\u00a0\u00b0C. The concentrate was left was to stand for 72\u00a0h and 3 drops of ethyl acetate added to facilitate the crystallization of crude xylopic acid crystals. Crude xylopic acid was washed several times with petroleum ether and dissolved in absolute ethanol for purification by recrystallization. The purity of the xylopic acid was assessed by thin-layer chromatography using petroleum ether and ethyl acetate (9:1) as the solvent system. Pure xylopic acid was used as a reference and both compounds gave Rf of 0.53.Xylopic acid was extracted from escribed \u201318. Fres3) with beddings made from softwood shavings. The animals were kept under appropriate laboratory conditions and fed with a normal commercial pellet diet purchased from Agricare,  and water ad libitum. The cages were kept in the Department of Biomedical Sciences animal holding facility, University of Cape Coast, and the wood shavings were replaced every 3\u00a0days and disinfected with 70% alcohol. The facility had a 12/12\u00a0h light/dark cycles and a mean temperature of 21\u00a0\u00b0C.Six to ten weeks-old female ICR mice purchased from the Centre for Medicinal Plant Research, Akuapim-Mampong, Ghana, were used for the study. They were housed in stainless steel cages (16.5\u2009\u00d7\u200911.0\u2009\u00d7\u200913.5\u00a0cmArtemether/lumefantrine combined tablets (20/120\u00a0mg), artesunate, and amodiaquine were acquired from Novartis Pharma AG Basel, Switzerland. Hydrochloric acid, Giemsa stain, absolute methanol, chloroform, petroleum ether, ammonium hydroxide, 96% ethanol, liquid paraffin, Tween 20, and ammonium chloride were also purchased from Sigma Aldrich. St Louis, MO, USA.Plasmodium berghei (strain ANKA) was acquired from Noguchi Memorial Institute for Medical Research (NMIMR), University of Ghana and by a continuous passage in mice intraperitoneally every 6\u00a0days [7P. berghei parasitized erythrocytes were prepared and each mouse was inoculated with 0.20\u00a0ml of PBS containing 1.2\u2009\u00d7\u2009106 parasitized red blood cells.A chloroquine-sensitive strain of rodent malaria parasite y 6\u00a0days . Once hiy 6\u00a0days . Blood wMice body weights were measured on days 0 and day 7 post-infection following a procedure described by Dikasso and colleagues  using a 50 values for the isobolographic analysis, the anti-plasmodial activity of each compound was assessed. After infection, mice were assigned to 18 groups (n\u2009=\u20095). Mice in all groups were inoculated with P. berghei except Group 18 mice which served as na\u00efve control. Seventy-two hours post-inoculation (day 3), all groups of animals were treated once daily by oral administration with a gastric gavage with either xylopic acid , artesunate , amodiaquine , artemether/lumefantrine (1.14/6.9\u00a0mg\u00a0kg\u22121), or vehicle, 10\u00a0ml\u00a0kg\u22121 (na\u00efve and sham control). The ED50 values obtained as fitted midpoints of XA, ART and AQ were determined by iterative curve fitting of log-dose responses of XA, ART, and AQ. Mice were observed at 12\u00a0h intervals for death and the median survival and hazard ratio over a 30-day period was computed.To confirm the reported anti-malarial properties of xylopic acid (XA), artesunate (ART), and amodiaquine (AQ), and also determine their EDP. berghei infection, 6 to 10\u00a0weeks female ICR mice were each inoculated with 1.2\u2009\u00d7\u2009106 in 0.20\u00a0ml PBS and assigned to 8 groups (n\u2009=\u20095). On day 3, each group received either fixed ratio (1:1) of the ED50s of XA and ART (9\u2009+\u20091.6\u00a0mg\u00a0kg\u22121) or combinations of fractions of the respective ED50 values: ED50 (XA/ART)/2, (4.5\u2009+\u20090.8\u00a0mg\u00a0kg\u22121), ED50 (XA/ART)/4 (2.25\u2009+\u20090.4\u00a0mg\u00a0kg\u22121), ED50 (XA/ART)/8, (1.13\u2009+\u20090.2\u00a0mg\u00a0kg\u22121), ED50 (XA/ART)/16 (0.6\u2009+\u20090.1\u00a0mg\u00a0kg\u22121. Positive control (AL) and negative control (sham) mice received 1.14/6.9\u00a0mg\u00a0kg\u22121 AL and 10\u00a0ml\u00a0kg\u22121 vehicle, respectively.To assess the antiplasmodial property of xylopic acid-artesunate (XA/ART) co-administration on established P. berghei infection, 6 to 10\u00a0weeks female ICR mice were each inoculated with 1.2\u2009\u00d7\u2009106 in 0.2\u00a0ml PBS and assigned to 8 groups (n\u2009=\u20095). Seventy-two hours later, each group received fixed ratio (1:1) or combinations of fractions of the respective ED50 values of (9\u2009+\u20093.1\u00a0mg\u00a0kg\u22121), (4.5\u2009+\u20091.6\u00a0mg\u00a0kg\u22121), (2.25\u2009+\u20090.8\u00a0mg\u00a0kg\u22121), (1.125\u2009+\u20090.4\u00a0mg\u00a0kg\u22121), (0.6\u2009+\u20090.2\u00a0mg\u00a0kg\u22121), ED50 (XA/AQ), ED50 (XA/AQ)/2, ED50 (XA/AQ)/4, ED50 (XA/AQ)/8, and ED50 (XA/AQ)/16, respectively. Positive control (AL) and negative control (sham) mice received 1.14/6.9\u00a0mg\u00a0kg\u22121 AL and 10\u00a0ml\u00a0kg\u22121 vehicle, respectively.To assess the anti-plasmodial property of xylopic acid-amodiaquine  test was used as a post hoc test. Two isobolograms which consisted of the ED50 of XA on the ordinate and ED50 of ART or AQ on the abscissa connected with a line of additivity were constructed. The ED50 of each drug was determined by linear regression analysis of the log dose\u2013response curve . Zadd was computed with the following formulae:All statistical analyses were computed with the windows version of GraphPad Prism 7.0 . Data were considered significant at add was computed as follows:and50 significantly lower) and higher than the theoretically calculated equieffect of a drug combination in the same proportion, it has a supra-additive or synergistic effect.SEMs were calculated from these variances and fixed according to the drug\u2019s ratio in the combination. If the effect of a drug combination was statistically different (EDP. berghei (p\u2009=\u20090.001). In XA-treated groups, loss in body weight was not statistically significant. High doses of artesunate (8\u00a0mg/kg) and amodiaquine (10\u00a0mg/kg) showed an increase in body weight similar to the na\u00efve and AL groups  for all groups. ED50s for xylopic acid, artesunate, and amodiaquine were 9.0\u2009\u00b1\u20093.2, 1.61\u2009\u00b1\u20090.6, and 3.1\u2009\u00b1\u20090.8\u00a0mg/kg. By these results, the artesunate was 1.9 times more potent than amodiaquine, and amodiaquine was 2.9 times more potent than xylopic acid  Fig.\u00a0.Fig. 4Bo50s for xylopic acid-artesunate and xylopic acid-amodiaquine were 1.98\u2009\u00b1\u20090.33 and 1.69\u2009\u00b1\u20090.83, respectively  of 5.3\u2009\u00b1\u20092.61, whereas the experimental ED50 (Zexp) was obtained as 1.98\u2009\u00b1\u20090.25  and the Zadd, which means the two drugs have a synergistic anti-plasmodial effect. The interaction index of 0.37, which is significantly less than 1, confirms a synergistic relationship [Combining xylopic acid with either artesunate or amodiaquine showed a remarkable suppression in parasite growth similar to the artemether/lumefantrine. Although, monotherapy of XA, ART, and AQ also suppressed parasite growth compared to artemether/lumefantrine it occurred at higher doses. An isobolographic analysis was employed to determine the enhanced or improved potency and efficacy of xylopic acid-artesunate, and xylopic acid-amodiaquine combination therapies. An isobolographic analysis gives a central basis for evaluating whether a biological response induced by a mixture of agents is smaller, equal, or greater on the concept of dose additivity and the basis of the components or agents\u2019 activities . The co-tionship  and a suP. berghei [Compared to a recent study by Ameyaw et al. , combini berghei  confirmiPlasmodium haemoglobin digestion of parasite glycolysis might contribute to the enhanced antiplasmodial activity of ART and XA.The observed increased antiplasmodial activity of the XA/ART combination could also be attributed to the two drugs interacting with several targets in the parasite. XA inhibits plasmodium dehydrogenase , an enzy2). XA inhibited albumin denaturation, and also maximal edema, and average paw thickness induced by the phlogistic agents for both prophylactic and therapeutic studies. It also inhibited the arachidonic acid pathway [P. berghei infection, splenic dendritic cells, CD8\u03b1+ and Clec9A+ phagocytose, and cross-present parasite antigens which lead to the priming of parasite-specific CD4+ and CD8+ T cells. Circulating parasitized red blood cells (pRBC) adhere to the endothelium of blood vessels releasing inflammatory ligands such as hemozoin crystals which contain parasite DNA. These stimuli are responded to by the release of cytokines and chemokines leading to the upregulation of adhesion molecules  and receptors (CXCR3) capable of presenting antigens [Furthermore, the anti-inflammatory properties of xylopic acid may have contributed to the limiting survival of the parasite. Osafo and colleagues recently reported the anti-inflammatory properties of xylopic acid against various phlogistic agents  and uninfected RBCs to agglutinate promoting the appropriate microenvironment for sequestration [+ T-helper cells have been reported to be involved in malaria conferring protection. However, they have also been implicated in immune evasion and malaria pathogenesis [stration \u201343. The stration . CD4+ T-ogenesis . Despiteogenesis , 46 mighA combination of xylopic acid and amodiaquine showed enhanced activity due to their synergistic interaction. Like the XA/ART combination, XA/AQ interaction also showed an interaction index of 0.13, which is significantly different from . PreviouP. berghei [In malaria treatment, like any other infectious disease, it is crucial not only to pay attention to the pathogen but also the reduction of the symptoms of the infection which independently increases the pathogen burden . Among t berghei , 18. It All the characteristics of an ideal anti-malarial agent should be able to prevent eventual death caused by parasites by suppressing the growth of parasites, thereby reducing the risk of death. An increase in parasite growth causes various symptoms of malaria which eventually leads to the death of the hosts . The hazThe findings of this study is heartwarming in the light of the report of growing resistance to current artemisinin . The com"}
+{"text": "P. vivax, is constrained by concerns over its safety. We used routinely collected patient data to compare the overall morbidity and mortality in patients treated with and without PQ without prior testing of Glucose-6-Phosphate-Dehydrogenase (G6PD) deficiency in Papua, Indonesia, where there is a low prevalence of G6PD deficiency. Records were collated from patients older than 1 year, with P. vivax infection, who were treated with an artemisinin combination therapy (ACT). The risks of re-presentation, hospitalization, major fall in haemoglobin and death within 30 days were quantified and compared between patients treated with and without PQ using a Cox regression model. In total 26,216 patients with P. vivax malaria presented to the hospital with malaria during the study period. Overall 27.56% : 26.96\u201328.16) of 21,344 patients treated with PQ re-presented with any illness within 30 days and 1.69% (1.51\u20131.88) required admission to hospital. The corresponding risks were higher in the 4,872 patients not treated with PQ; Adjusted Hazard Ratio (AHR) = 0.84  and 0.54  respectively. By day 30, 14.15% (12.45\u201316.05) of patients who had received PQ had a fall in haemoglobin (Hb) below 7g/dl compared to 20.43% (16.67\u201324.89) of patients treated without PQ; AHR = 0.66 . A total of 75 (0.3%) patients died within 30 days of treatment with a mortality risk of 0.27% (0.21\u20130.35) in patients treated with PQ, compared to 0.38% (0.24\u20130.60) without PQ; AHR = 0.79 . In Papua, Indonesia routine administration of PQ radical cure without prior G6PD testing, was associated with lower risk of all cause hospitalization and other serious adverse clinical outcomes. In areas where G6PD testing is not available or cannot be delivered reliably, the risks of drug induced haemolysis should be balanced against the potential benefits of reducing recurrent P. vivax malaria and its associated morbidity and mortality.The widespread use of primaquine (PQ) radical cure for  Plasmodium vivax. Primaquine (PQ) is currently the only widely available drug to kill hypnozoites, but its use is hampered by concerns over haemolysis in patients with Glucose-6-Dehydrogenase (G6PD) deficiency. At a recent round table discussion of the Asia Pacific Malaria Elimination Network (APMEN) policy makers prioritised further studies to determine the risk-benefit of PQ in different endemic settings.The radical cure of vivax malaria requires killing the blood stages and the dormant liver stages (hypnozoites) of We used a large dataset of more than 26,000 patient treated in a routine care setting in Papua, Indonesia over a 9-year period to explore the risk of adverse outcomes following PQ. Even though patients were treated with PQ without G6PD testing, those patients receiving PQ had a 20% lower risk of all cause re-presentation and 50% lower risk of hospitalization than those not treated with PQ. The risk of mortality was 20% lower, although this didn\u2019t reach statistical significance.Great emphasis is placed on the risks of PQ induced haemolysis, inhibiting its widespread use. It is critical that these risks are interpreted in the context of the substantial benefits of reducing the recurrence of episodes of malaria and associated morbidity and mortality. Our analysis provides reassuring evidence that, in an area with a high burden of malaria and low G6PD prevalence the beneficial effects of PQ outweigh its risks. P. vivax than for P. falciparum. Unlike P. falciparum, P. vivax forms dormant liver stages (hypnozoites) that can reactivate weeks to months after an initial infection, causing recurrent parasitaemia (relapses). Whereas schizontocidal drugs are effective in killing blood stage parasites and reducing acute symptoms, they have no efficacy against the dormant liver stages of the parasite. The parasite\u2019s propensity to recur weeks or months after initial infection results in recurrent febrile illness and a cumulative risk of anaemia and associated morbidity and mortality [In the last decade major gains have been made in malaria control, but successes have been less apparent for ortality \u20133. Relaportality .The only widely available drug to kill hypnozoites and therefore prevent relapsing infection is primaquine (PQ), which is administered with schizontocidal treatment in a combination known as radical cure. The effectiveness of PQ is limited by the prolonged treatment course (14 days) recommended by most national malaria control programs, and concerns over its safety, particularly the risk of acute haemolysis in individuals with glucose-6-phosphate-dehydrogenase (G6PD) deficiency . G6PD deP. vivax malaria in Papua, Indonesia, an area endemic for P. vivax strains with a short relapse periodicity and low G6PD prevalence [We undertook a retrospective cohort study of patients with evalence . The aimThe study is a retrospective analysis that was conducted on routine data collected from a healthcare facility in Papua, Indonesia. Although individual consent was not collected from patients, all data were anonymised. Ethical approval was obtained from the Human Research Ethics Committee of the Northern Territory (HREC 10.1397) and the Health Research Ethics Committees of the University of Gadjah Mada (KE/FK/544/EC), Indonesia.The study was designed as a retrospective cohort study of routinely collected patient data to compare the morbidity and mortality of patients treated with and without PQ.P. vivax [The climate, geography, malaria endemicity and demographics of the study site have been described previously \u201310. In bP. vivax .Plasmodium species [P. vivax malaria. Before March 2006 the recommended total dose was 3.5mg/kg administered over 14 days. In March 2006 local guidelines were changed to recommend a total dose of 7mg/kg administered over 14 days. For patients presenting to the hospital outpatient clinic, PQ was commenced at the same time as schizontocidal treatment, whereas in inpatients, PQ was administered once clinical recovery had begun and patients were able to take oral medication.The study was undertaken in patients presenting to the Rumah Sakit Mitra Masyarakat (RSMM) hospital, in Timika, Papua province, Indonesia, between April 2004 and December 2013. RSMM is the largest health care facility in Timika and one of two public hospitals in the district. Since March 2006 local guidelines have recommended dihydroartemisinin-piperaquine (DHP) as the first line treatment for uncomplicated malaria due to any  species . PQ radiIn Indonesia, treatment guidelines do not recommend routine testing for G6PD deficiency to guide the administration of PQ. In a cross-sectional survey in 2013, the prevalence of severe or intermediate G6PD deficiency was 2.6% and was slightly lower in individuals of highland Papuan ethnicity (2.1%) compared to lowlanders and non-Papuans (2.6% and 2.8%) respectively [Data from all patients presenting to the RSMM between 2004 and 2013 were recorded in a Q-Pro database by hospital administrators. The following data were recorded: the patients\u2019 unique identification number, demographic details, date of visit and duration of hospital admission, clinical diagnosis and laboratory investigations. Details on prescriptions at the hospital pharmacy were recorded in a separate database and linked to the same unique hospital record number (HRN).falciparum malaria.Hospital protocols require that all patients attending the outpatient clinic with fever or symptoms consistent with malaria and all inpatients regardless of their clinical presentation, were tested for malaria by microscopic examination of Giemsa-stained blood films. In some instances, HRP2 based rapid diagnostic tests were used for additional confirmation of P. vivax malaria are associated with a cumulative risk of anaemia and both direct and indirect mortality [P. vivax, alone or mixed with another Plasmodium species, and all associated clinical events occurring within the ensuing 30 days. Pharmacy records reported the number of tablets prescribed and the actual mg per kg dose of PQ prescribed were derived for each individual patient by applying estimated mean body weights within each age, sex, and ethnicity strata derived from a cross sectional survey [The HRN was used to link clinical and laboratory data with the pharmacy records. Since recurrent episodes of ortality ,12, the l survey . The totl survey : (i) No P. vivax parasitaemia, either mono-infection or part of a mixed infection, were included in the analyses if they were treated with an artemisinin combination therapy (ACT). Patients treated with non-ACT regimens prior to policy change in 2006 were excluded, since chloroquine resistant P. vivax is highly prevalent in Papua and its poor efficacy is associated with a high risk of adverse events attributable to parasitological failure [P. vivax [Patients with microscopically confirmed  failure . In thisP. vivax ,17. PQ iThe analysis was conducted according to an a priori statistical analysis plan  and is rThe cumulative risks of each of the outcomes were estimated using Kaplan Meier survival curves. Patients with an event on day 1 or 2 were censored on that day under the assumption that this event was related to the initial vivax episode rather than the drug treatment. Hazard Ratios for the association between treatment with/without PQ and each of the outcomes were calculated using a Cox regression model. In view of significant changes in morbidity and mortality of malaria in this area , all mulP. vivax malaria . A total 2,140 patients were excluded because they had no matching pharmacy records and 5,575 were excluded because they received schizonticidal treatment other than ACT or injectable artesunate. A further 1,455 only received a single dose PQ and for 474 patients the PQ dose was missing  received a high dose of PQ, 2,065 (7.88%) were prescribed a low dose of PQ, and 4,872 (18.58%) received no PQ. The baseline characteristics of patients treated with or without PQ were similar . The maiOverall 24,148 (92.1%) patients were treated with DHA-Piperaquine and 1,720 (6.6%) with artesunate-amodiaquine. A total of 2,482 (9.6%) of patients were treated with intravenous artesunate alone or in combination.P. vivax , 457 (6.26%) with falciparum malaria, 17 (0.23%) with other malaria species and 6,048 (82.89%) were due to non-malarial illness. The cumulative risk of re-presentation at 30 days was 27.56% (95%CI: 26.96\u201328.16) for those with any PQ dose compared to 30.30% (95%CI: 29.01\u201331.64) in patients not treated with PQ (adjusted Hazard Ratio (AHR) = 0.84 (95%CI: 0.79\u20130.91); p<0.001)  patients re-presented to hospital between 3 and 30 days after their initial presentation; 774 (10.61%) of these re-presentations were associated p<0.001) , Table 2P. vivax mono-infection and those requiring hospital admission were significantly more at risk of re-presenting to hospital within 30 days ; p<0.0001), but there was no difference between treatment arms in the risk of representing with non-malarial illness (HR = 0.98 (95%CI: 0.92\u20131.05); p = 0.551). Females, children, those presenting with  30 days . After c 30 days . After c with PQ .a priori subgroup analyses, the risk of hospitalization in patients treated with a high dose PQ was also lower than that in patients not treated with PQ  of 23,426 patients initially treated as outpatients, required admission to hospital between days 3 and 30. The cumulative risk of hospitalization at 30 days was 1.69% (95%CI: 1.51\u20131.88) in those treated with any dose of PQ compared to 2.71% (95%CI: 2.25\u20133.27) in patients not initially treated with PQ; HR = 0.62 (95%CI: 0.50\u20130.77), p<0.001 , Table 2 with PQ .a priori subgroup analyses, the rate of death in non-Papuans after high dose PQ was significantly lower than that in patients not treated with PQ; AHR = 0.05 (95%CI: 0.006\u20130.43); p = 0.006  patients died between day 3 and 30 days, of whom 18 (24.00%) were not treated with PQ, 8 (10.67%) were treated with low PQ and 49 (65.33%) were treated with high dose PQ. The median number of days before death was 8  and did not vary with PQ treatment (p = 0.887). The cumulative risk of death at 30 days was 0.27% (95%CI: 0.21\u20130.35) in those treated with any dose of PQ and 0.38% (95%CI: 0.24\u20130.60) in patients not treated with PQ; AHR = 0.79 (95%CI: 0.43\u20131.45); p = 0.448. The rate of death was not significantly lower in patients treated with high dose PQ; AHR = 0.77 (95%CI: 0.41\u20131.43); p = 0.403 , Table 2 = 0.006 .P. falciparum, 26 (52.00%) with P. vivax infection and 19 (38.00%) with had a mixed species infection. Of those who died a total of 4 (5.33%) were diagnosed with acute renal failure, three of whom had received high dose PQ and one didn\u2019t receive any PQ. The clinical details of the 75 patients who died are summarized in At time of death 50 (66.67%) of 75 patients had represented with malaria: 5 (10.00%) with Data on full blood count were available in 8,964 (34.19%) patients at their initial presentation, of whom 20.25%  had a subsequent Hb measure between day 3 and 30. Among those a total of 253 (13.94%) patients presented with an Hb below 7g/dl at first visit. The risk of the Hb concentration falling below 7g/dl was 14.15% (95%CI: 12.45\u201316.05) in patients treated with any dose PQ compared to 20.43% (95%CI: 16.67\u201324.89) in those not treated with PQ; AHR = 0.66 (95%CI: 0.45\u20130.97); p = 0.033. Overall the NNT with PQ to prevent one patient dropping their Hb below 7g/dl within 30 days was 16 (95%CI: 9\u2013183). The comparative AHR for PQ treatment versus those not treated with PQ for the Hb falling below 7g/dl before day 14 were 0.61 , and between day 15 and 30 it was 0.75  . The treA total 36 patients  had a drop in Hb greater than 25% to an absolute Hb <7g/dL within the first 30 days. These included 4.05% (29/716) of patients treated with PQ, and 3.72% (7/188) of patients not receiving PQ. In the multivariable analysis there was no difference in the rate of a clinically significant fall in Hb between those treated with PQ and those who were not administered PQ; AHR = 1.16 (95%CI: 0.43\u20133.10); p = 0.769 . The cliOur study quantified the morbidity and mortality in more than 26,000 patients presenting with vivax malaria treated with or without PQ in a routine healthcare setting. Although patients were prescribed PQ according to local guidelines, without prior testing for G6PD deficiency, patients treated with PQ had a significantly lower risk of re-presenting to hospital, admission to hospital and developing severe anaemia compared to those patients not treated with PQ within 30 days. The overall risk of dying was lower after high dose PQ, although confidence intervals were wide.P. vivax relapse periodicity such as East Asia and Oceania [Current WHO treatment guidelines recommend a 14 day course of PQ at 0.25mg/kg/day, but also state that a higher dose (0.5mg/kg/day) may be required in areas with high  Oceania . In PapuProviding safe and effective radical cure is important at both an individual and community level. Recurrent episodes of vivax malaria result in a cumulative risk of severe anaemia and associated morbidity and mortality ,19. TheyP. vivax re-presentations in the year following initial presentation [P. vivax relapses by the provision of effective radical cure would also reduce vulnerability to other non-malaria illnesses [Our analysis shows that in Papua, Indonesia, providing PQ radical cure without prior testing for G6PD reduced the risks of re-presentation within 30 days. A previous analysis from the same location also showed a modest, but statistically significant, reduction in entation . In the llnesses ,19,25. ROur study has a number of strengths. The very large sample size reduces the possibility of chance findings. Furthermore, the patients included in this analysis represent a sample of the population seeking treatment for vivax malaria in a real world setting as compared to carefully selected patient cohorts enrolled into comparative clinical trials. This increases the applicability of the results to the patient population of interest.a priori subgroup analysis in which only outpatients were included, the risk of re-presentation was also lower in the PQ group. The remaining a priori subgroup analyses confirmed similar trends in reduction of the primary and secondary outcomes. The subgroup of patients presenting in 2007, was to include a pharmacy stock-out of PQ which lasted for 7 months, with patients also included from 6 months intervals before and after this period. The objective was to focus on patients who didn\u2019t receive PQ due to logistical rather than clinical reasons. Whilst the trend for reduction in re-presentation and hospital admission remained, the confidence intervals were wide due to reduced sample size.Our study also has a number of important limitations. The decision regarding whether or not to prescribe PQ was made by the attending clinician and patients were not allocated randomly to a treatment regimen. Although clinicians may have been hesitant to prescribe PQ to patients who were very unwell or had significant anaemia, discussions with senior clinicians at the hospital suggest that omission of PQ was more often related to junior doctors being less aware of the need to offer PQ radical cure. The baseline characteristics between the treatment groups were generally comparable, however inpatients were less likely to receive PQ than outpatients, but are known to have a higher risk of re-presentation, hospitalization and death. In the P. vivax malaria occurred on day 2, even in patients not treated with PQ and thus is likely to be predominantly related to malaria [In our analysis, patients with an event on day 1 or 2 were censored, assuming that these are related to deterioration from clinical malaria rather than its treatment. In a recent meta-analysis the nadir Hb in patients with uncomplicated  malaria . Reports malaria ,27. HoweA further limitation of the study was that it was not possible to confirm from the hospital records whether patients took the full course of PQ treatment or not. Previously we have postulated that poor adherence to PQ treatment at the same location may limit effectiveness of radical cure . The modOur analysis included all severe adverse events within 3 to 30 days of patients\u2019 initial presentation with vivax malaria. Hb concentration was not measured routinely, although it is hospital practice to check the Hb of inpatients and those with clinical anaemia. Our primary analysis therefore focused on all cause re-presentation and hospital admission as indirect measures of adverse outcomes rather than direct measures of haemolysis. The latter was addressed in a subgroup of patients in whom Hb was measured at presentation, and reassuringly showed similar trends to a lower risk of anaemia in patients receiving PQ compared to those who did not.P. vivax infection and its associated morbidity and mortality. Our analysis in Papua, suggests that irrespective of the aetiology, one re-presentation to hospital can be prevented by treating 24 patients with PQ even without G6PD testing. The corresponding NNT with PQ was 16 to prevent one patient dropping their Hb below 7g/dl and 81 to prevent one patient requiring admission to hospital.We were also unable to discriminate whether the reported clinical endpoints were attributable to malaria, non-malaria or drug-induced aetiologies. It is possible that some of the severe adverse events could have been due to PQ-induced toxicity in G6PD deficient individuals, and that these could have been prevented if prior G6PD testing had been offered. Whilst all efforts should be made to diagnose patients with G6PD deficiency prior to PQ administration, this is often unavailable or unreliable due to logistical, financial and infrastructural limitations in remote and inaccessible locations where the main burden of malaria resides ,5. The oThe relevance of our findings for patients with vivax malaria in other endemic regions needs to be considered with caution. In southern Papua less than 3% of the population are G6PD deficient , a likelS1 Text(PDF)Click here for additional data file.S1 Data(CSV)Click here for additional data file.S1 Table(PDF)Click here for additional data file.S2 Table(PDF)Click here for additional data file.S3 Table(PDF)Click here for additional data file.S4 Table(PDF)Click here for additional data file.S5 Table(PDF)Click here for additional data file.S6 Table(PDF)Click here for additional data file."}
+{"text": "Amodiaquine is a 4-aminoquinoline antimalarial used extensively for the treatment and prevention of malaria. Orally administered amodiaquine is largely converted to the active metabolite desethylamodiaquine. Oral amodiaquine can cause bradycardia, hypotension, and electrocardiograph (ECG) QT interval prolongation in animals and humans but the relationship of these changes to drug concentrations is not well characterised.We conducted a secondary analysis of data from a pharmacokinetic study of the cardiac safety of amodiaquine (10mg/kg/day over 3 days) in 54 Kenyan adults (\u226518 years) with uncomplicated malaria. Non-linear mixed effects modelling was used to assess amodiaquine and desethylamodiaquine concentration-effect relationships for vital sign  and ECG interval  outcomes. We also measured the spontaneous beating heart rate after cumulative dosing of amodiaquine and desethylamodiaquine in isolated mouse atrial preparations.Amodiaquine and desethylamodiaquine caused concentration-dependent mean decreases in pulse rate (1.9beats/minute per 100nmol/L), supine systolic blood pressure (1.7mmHg per 100nmol/L), erect systolic blood pressure (1.5mmHg per 100nmol/L), and erect diastolic blood pressure (1.4mmHg per 100nmol/L). The mean QT interval prolongation was 1.4milliseconds per 100nmol/L irrespective of correction factor used after adjustment for residual heart rate dependency. There was no significant independent effect of drug concentration on postural change in blood pressure or PR and QRS intervals. In mouse atria, the spontaneous beating rate was significantly reduced by amodiaquine (n=7) and desethylamodiaquine (n=8) at 3\u03bcmol/litre  and 10\u03bcmol/litre  concentrations with no significant difference in potency between the two compounds.Amodiaquine and desethylamodiaquine have concentration-dependent effects on heart rate, blood pressure, and ventricular repolarisation. In 2005, the World Health Organization (WHO) recommended that artemisinin-based combination therapy (ACT) should become the first-line treatment for all falciparum malaria. In 2008, artesunate-amodiaquine (ASAQ), the fixed-dose formulation of amodiaquine and artesunate (AS), became the first major development product of a public-private partnership, the Drugs for Neglected Diseases initiative (DNDi). ASAQ is now recommended by the WHO for the treatment of uncomplicated Plasmodium falciparum and P. vivax malaria3 and is the first-line oral antimalarial in more than 20 African countries4 where malaria is endemic. Since 2012, WHO has recommended use of seasonal malaria chemoprevention (SMC) for young children (aged 3-59 months) living in areas of seasonal high-intensity malaria transmission in the Sahel subregion of Africa. SMC comprises a single dose of sulfadoxine-pyrimethamine together with amodiaquine (SP + AQ) divided over three days monthly during the rainy season for up to four months annually. Millions of children are now protected with SMC every year5.Amodiaquine (AQ), a 4-aminoquinoline structurally similar to chloroquine, is an important antimalarial drug which has been deployed extensively in the treatment and prevention of malaria over the past 70 years. First synthesised by the United States World War II (WWII) antimalarial research programme1. During its development, pulsus bigeminus was noted in anaesthetised dogs receiving high doses of parenteral amodiaquine1. Like chloroquine, amodiaquine exhibits anti-arrhythmic properties, terminating experimental atrial arrhythmias in both decentralised and innervated canine hearts6 but, unlike chloroquine, does not appear to protect against experimental ventricular arrhythmias7. Electrocardiograph QT interval prolongation7, bradycardia8, and hypotension8 have also been observed after parenteral amodiaquine administration to anaesthetised dogs and cats. We have reported recently that amodiaquine prolongs the QT interval less, but is more bradycardic and hypotensive than chloroquine at current standard oral malaria treatment doses when given to adolescents and adults9. The clinical significance of these cardiovascular effects with standard malaria treatment dosing is unclear9 although bradycardia and hypotension may contribute to the higher incidence of mild asthenia and asthenia-like reactions after amodiaquine compared to other antimalarials12. Direct multiple ion channel blockade of cardiac17 and vascular18 myocytes along with altered autonomic tone20 may both be relevant. There are few data on the cardiovascular pharmacology of amodiaquine and information on its main metabolite desethylamodiaquine is especially limited, as the metabolite desethylamodiaquine was only identified in the 1980s21. This was around the time reports of the fatal toxicity of amodiaquine in chemoprophylaxis24 led to its temporary withdrawal in 199025 from the list of WHO-recommended antimalarials.The cardiovascular effects of amodiaquine have been recognised from the earliest studies in animal modelsWe conducted a secondary analysis of a clinical pharmacokinetic study of ASAQ in adult malaria patients focusing on electrocardiographic interval  and cardiovascular vital sign (pulse rate and blood pressure) outcomes. This analysis of the clinical study was complemented by laboratory assessment of the concentration-heart rate response in murine atrial preparations with intact sino-atrial node (SAN).P. falciparum monoinfection with an asexual parasitaemia of >1,000 parasites/\u03bcL and either a history of fever or a measured temperature of \u226537.5\u00b0C in the preceding 24 hours. Pregnant or lactating women, those with significant known comorbidities , individuals with an ECG abnormality requiring urgent treatment, or those who had taken an artemisinin derivative and/or sulfadoxine-pyrimethamine in the previous three or seven days respectively were excluded. Further clinical details are reported in full elsewhere27.This was an open-label randomised controlled trial conducted between 2007 and 2008 in the Chulaimbo Sub-district Hospital of Kisumu, Kenya. The trial compared the fixed-dose ASAQ combination with loose (i.e. non-fixed dose) AS + AQ in adult patients (aged 18 to 60 years inclusive) presenting with acute uncomplicated Patients received either two fixed-dose ASAQ tablets  daily for a total dose over three days of 600mg of AS and 1620mg of AQ, or four tablets each of non-fixed dose AS  and AQ  daily for a total dose over three days of 600mg of AS and 1836mg of AQ. No concomitant food was given. All treatments were directly observed. Patients who vomited a dose within 1 hour after drug administration were retreated. Exact dosing times were recorded and used in modelling.Medical and drug histories were taken and physical examinations performed at baseline (D0). Basic clinical assessments including measurement of vital signs  were conducted and malaria thick blood film slides were prepared at D0, D1, D2, D3, D7, D14, D21, and D28. A blood film was considered negative if no asexual parasites were seen after examination of 1000 white blood cells. Patients were asked about adverse events of headache, weakness, anorexia, nausea, abdominal pain, itching, vomiting, diarrhoea, rhinitis, cough, vertigo, and rash at each clinical assessment. Standard 12-lead ECGs were performed on D0 , D2 (+2 and +4 hours), and D28. ECGs were assessed centrally by cardiologists for arrhythmias and measurement of ECG intervals .Full blood counts were performed on D0 (pre-dose), D7, and D28. Blood samples for drug concentration measurements were taken from all patients at fixed time points on D0 (pre-dose), D7, D14, D21, and D28. Each patient also had additional samples taken at random combinations of the following time points on D0  and D2 . Exact sample times were recorded and used in the pharmacokinetic modelling.Venous plasma samples were analysed using liquid chromatography-mass spectrometry (LC-MS/MS) methods. Amodiaquine, desethylamodiaquine, and the internal standard (AST-D4) were analysed by reversed-phase liquid chromatography  and MS/MS (Sciex API3000) detection in the Turbo Ion Spray positive mode.The trial protocol was approved by the Kenya Medical Research Institute (KEMRI) Ethical Review Committee. Additional ethical approval for this secondary analysis of fully anonymised individual patient data was not deemed necessary in keeping with University of Oxford Central University Research Ethics Committee guidance.28. It is generally accepted that artesunate does not have a significant effect on the QT interval29.Trial data were standardised and checked according to a pre-specified data dictionary 34 version 4.8.0 were used for automation, model evaluation, and diagnostics during the modelling process. The NONMEM $PRIOR functionality was used to stabilise model performance.Observed amodiaquine and desethylamodiaquine concentrations, transformed into their natural logarithms, were analysed using non-linear mixed-effects modelling implemented in NONMEM version 7.4P. vivax malaria in pregnant women with rich (i.e. more intensive) pharmacokinetic sampling35  and electrocardiographic intervals  were measured were used in the concentration-effect analyses.36 and expert consultation37. The drug effect was evaluated using the total predicted concentration of amodiaquine plus its metabolite desethylamodiaquine based on prior evidence that both amodiaquine7 and desethylamodiaquine38 affect cardiovascular physiology as well as lack of evidence for any substantial difference in their activities from descriptive analyses. In the ECG interval models, the other fixed effects were body temperature change, age, and sex, with the addition of RR interval change to adjust for residual heart rate changes. For cardiovascular vital signs measured at multiple time points after recovery from malaria, the malaria disease effect was incorporated as a binary categorical fixed effect variable present during days 0-2 of treatment, with the addition of body temperature change and sex for the change in the pulse rate model only.Multivariable linear mixed effects modelling was performed with the corrected QT, QRS, and PR intervals as well as the change from baseline of pulse rate and blood pressure  as response variables  identifiad libitum access to standard diet and sterilised water. Mice were culled by cervical dislocation in accordance with the United Kingdom Home Office Guidance on Animals (Scientific Procedures) Act 1986. The heart was excised rapidly and washed in heparin-containing Physiological Salt Solution . Ventricles were dissected away, the atria were separated at the atrial septum and the area adjacent to the SAN was cleared of connective tissue. The spontaneously-beating right atrial preparation was mounted in a 37\u00b0C organ bath containing PSS (continuously oxygenated with 95% O2/5% CO2) and connected to a force transducer  with a resting tension of 0.2\u20130.3 g. The tension signal was low-pass filtered at 20Hz and the beating rate calculated from the time interval between contractions . After rate stabilisation in PSS (variation in average rate of a 10-second sample of no more than 2 beats/minute over a 10-minute period), cumulative concentrations of amodiaquine dihydrochloride  or N-desethylamodiaquine dihydrochloride solution  were pipetted directly into the bath. Preparations were excluded if, under control conditions (PSS only), stabilised beating rate was less than 300 beats/minute or arrhythmic.Adult male CD-1 mice  were housed maintained in a 12-hour light-dark cycle with Data are presented as mean \u00b1 standard error of the mean (SEM). Repeated measures analysis of variance with Tukey\u2019s or Dunnett\u2019s correction as appropriate was used to assess the isolated heart findings.54 patients aged between 18 and 60 years were randomised. Seven patients had received antimalarial pre-treatment: five patients chloroquine and two artemether-lumefantrine. None were receiving any cardiovascular concomitant medications. Baseline characteristics are presented in 53 completed the full treatment course. One patient in the fixed-dose ASAQ arm withdrew consent. Four patients were subsequently lost to follow-up and censored in analyses at the time of drop-out. All patients recovered uneventfully. There were no serious cardiovascular events reported in any of the 54 patients.A total of 363 post-dose venous plasma samples were collected from the 53 participants who received full treatment courses. Amodiaquine concentrations were measurable in 115 (31.7%) samples and 352 (97.0%) had measurable concentrations of desethylamodiaquine .Population pharmacokinetic parameter estimates were reliable, with small relative standard errors. Secondary pharmacokinetic parameters of maximum concentration, time to maximum concentration, terminal elimination half-life, and total exposure were also computed . Goodnesmax) of amodiaquine and desethylamodiaquine was approximately 750nmol/L (or 250ng/mL)  . This wa50ng/mL) .max was associated with a mean fall of 12.4mmHg (95% CI: 8.9 to 15.9) in supine systolic blood pressure and 11.0mmHg (95% CI: 7.4 to 14.7) in erect systolic blood pressure. Corresponding reductions in supine diastolic blood pressure were 4.7mmHg (95% CI: 1.9 to 7.4), and 10.3mmHg (95% CI: 7.4 to 13.1) in erect diastolic blood pressure . Unlike the study-specific corrected QT interval, the Fridericia- and Bazett-corrected QT intervals retained clinically and statistically significant heart rate dependency after adjustment although these were in opposite directions . Thus, without adjustment, use of the Fridericia heart rate correction overestimated amodiaquine-related QT prolongation from baseline while the Bazett correction underestimated it  .14. The application of cumulative doses of amodiaquine (n=6) and N-desethylamodiaquine (n=8) to spontaneously beating mouse atrial preparations produced concentration-dependent reductions in beating rate which were statistically significant (p < 0.05) at concentrations of 3 \u03bcmol/litre  and 10 \u03bcmol/litre  but not 1 \u03bcmol/litre  compared to time-matched controls of the vehicles dimethyl sulfoxide (n=2) and methanol (n=2). There was no statistically significant difference between the response to amodiaquine and N-desethylamodiaquine at any concentration studied Despite its widespread use, amodiaquine has been relatively little studied in recent years. To our knowledge, this is the most detailed investigation to date of the cardiovascular concentration-effect relationships of amodiaquine and its active metabolite desethylamodiaquine, incorporating both clinical data in malaria patients and a pre-clinical study of the cardiac pharmacology of desethylamodiaquine.41. It is observed more in adolescents and adults than in children42 for reasons which are not fully understood, although modulation of cardiac ion currents by sex hormones may play a role43.Bradycardia is a common cardiovascular effect after antimalarial treatment with amodiaquine and mefloquine14. The amodiaquine-induced bradycardia in both innervated  and decentralised (mouse) hearts supports a direct pharmacological effect on cardiac myocyte ion channels through modulation of the pacemaker If current at the SAN, as observed previously with hydroxychloroquine14. Autonomic tone may also be relevant as both amodiaquine and mefloquine are associated with reversible inhibition of human acetylcholinesterase (AChE), with amodiaquine having a much higher potency than mefloquine20.Our murine atrial studies show that both amodiaquine and desethylamodiaquine have direct concentration-dependent bradycardic effects of similar potency. These effects are greater than that of hydroxychloroquine at the concentrations evaluated, measured using the same method in our previous study44 and chloroquine45 are known to cause lethal hypotension when injected rapidly but can be used safely with rate-controlled continuous intravenous infusion. As proposed for amodiaquine18, these hypotensive effects are likely due to vasodilation and negative inotropy from multiple ion channel blockade46. Orthostatic hypotension is a feature of acute malaria which is exacerbated by the quinoline antimalarials quinine and mefloquine47. However, in this study of uncomplicated malaria infections, there was no significant effect of the total plasma concentration of amodiaquine and desethylamodiaquine on postural changes in systolic or diastolic blood pressure after adjustment for malaria recovery.The quinoline antimalarials quinine9. While amodiaquine and desethylamodiaquine cause concentration-dependent bradycardia and hypotension in adult malaria patients, the clinical impact of these effects appears to be mild26, although they may contribute to the commonly reported asthenia.Amodiaquine is the most bradycardic of the front-line antimalarials48. Despite their QT-prolonging potential, the frontline quinoline and structurally-related antimalarials recommended currently by the WHO all have excellent track records of cardiac safety. They have not been associated with increased risk of sudden cardiac death or cases of TdP in their extensive use at standard doses for the treatment or prevention of malaria over seven decades49.Drug-induced QT interval prolongation is the most widely-used surrogate marker of the risk of development of torsades de pointes (TdP), a polymorphic ventricular tachycardia that can degenerate in some cases into ventricular fibrillation and cause sudden cardiac death50, particularly in malaria where there is further confounding from disease recovery which occurs as antimalarial concentrations peak36. As before9, a study-specific correction provided the best heart rate correction of the QT interval (QTcS) in our analysis while use of the Fridericia (QTcF) and Bazett (QTcB) corrections respectively overestimated and underestimated amodiaquine-related QT prolongation in malaria patients. Once any residual heart rate dependency had been adjusted for the drug-attributable QT prolongation from amodiaquine and desethylamodiaquine was comparable regardless of correction factor used and consistent with QT prolongation similar to that observed with piperaquine52 but less than with chloroquine53 at standard malaria doses.The QT interval lengthens as heart rate decreases. Correction formulae modelling this inverse and non-linear relationship are used to attempt to minimise the heart rate dependency of measured QT intervals to allow for comparisons across different heart rates. However, the commonly used Bazett and Fridericia corrections are both known to retain significant heart rate dependency55 which may predominantly reflect recovery from malaria rather than a direct drug effect. This differs from chloroquine53 which prolongs both the PR and QRS intervals with over one quarter of the QT prolongation following chloroquine resulting from QRS widening.In contrast, we found no significant effect of total amodiaquine and desethylamodiaquine concentration on PR or QRS intervals after adjustment for demographic factors  and malaria recovery . Small increases in unadjusted PR and QRS intervals after amodiaquine for malaria have been previously reportedThe widely used 4-aminoquinoline amodiaquine, like other quinoline and quinoline-like drugs, has transient effects on cardiac and vascular physiology. We characterised the concentration-dependency of the bradycardic, hypotensive, and QT prolonging effects of amodiaquine and its main metabolite desethylamodiaquine providing further evidence of their causal role. Further characterisation of the cardiovascular effects of amodiaquine and desethylamodiaquine in adult healthy volunteers and other pre-clinical models may help to improve the tolerability of this important medicine in the treatment and prevention of malaria.Supplementary Appendix"}
+{"text": "The oral IP receptor agonist selexipag is approved for the long-term treatment of pulmonary arterial hypertension (PAH). Treatment interruptions should be avoided due to the progressive nature of the disease. An intravenous (IV) formulation of selexipag was developed to provide a treatment option for short-term interruptions to oral selexipag. In this prospective, multicenter, open-label study, the safety, tolerability, and pharmacokinetics of temporarily switching between oral and IV selexipag were investigated .PAH patients receiving stable oral selexipag doses were enrolled. Following three consecutive IV selexipag infusions patients resumed oral selexipag. Corresponding IV and oral doses were selected to achieve comparable exposure to the active metabolite of selexipag. Safety outcomes were monitored throughout, and pharmacokinetic samples were obtained after oral and IV administration.All 20 patients completed the study. Fifteen patients had adverse events (AEs), most were mild, and none resulted in discontinuation. Headache was the most common AE throughout the study (four patients). Three serious AEs occurred in two patients with underlying comorbidities when oral dosing had resumed. There were no changes in WHO functional class for any patient and no clinically symptomatic changes in blood pressure were observed. Comparable exposure to the active metabolite of selexipag was demonstrated following corresponding oral and IV selexipag doses.Temporarily switching between corresponding doses of oral and IV selexipag was well-tolerated with no unexpected safety findings and comparable exposure to the active metabolite. Treatment with IV selexipag is a feasible option to bridge temporary oral selexipag treatment interruptions. Pulmonary arterial hypertension (PAH) is a progressive, debilitating and potentially fatal disease . SeveralUninterrupted treatment is considered important to maintain the therapeutic effect of PAH-specific therapies . In clinFollowing administration, selexipag is hydrolyzed by carboxylesterases to the active metabolite , which is approximately 37-fold more potent than selexipag and therefore the main contributor to the pharmacological effect . TherefoThe objective of this study was to assess the safety, tolerability, and pharmacokinetics of temporarily switching between corresponding oral and IV doses of selexipag in patients with PAH on stable oral selexipag treatment.This was a prospective, multicenter, open-label, single-sequence, cross-over, phase 3 study in patients with PAH NCT03187678). The study comprised a 28-day screening phase, followed by a 12-day treatment and observation phase, and a subsequent 30-day safety follow-up phase , discontinuations due to AEs, and serious AEs (SAEs). SBP and diastolic blood pressure (DBP) were monitored before oral administration on Day 1 (Period 1), before (pre-dose), during (25 and 87\u00a0min after infusion start) and after  the morning IV infusions on Day 2 and Day 3 (Period 2), and on Day 12 (Period 3). Electrocardiograms (ECG) were recorded on Day 1 before oral selexipag administration (Period 1), immediately before infusion start and within 30\u00a0min of completing each IV selexipag infusion on Day 2 and 3 (Period 2), and on Day 12 (Period 3). Hematology and blood chemistry samples were collected at screening, on Day 1, 3, and 12. WHO FC was assessed on Day 1, 3 (at the end of the third IV infusion), and 12.\u03a4, ss), the maximum plasma concentration at steady state , the time to reach maximum plasma concentration at steady state , and combined AUC\u03a4, ss  of selexipag and ACT-333679. cAUC\u03a4, ss was defined as the potency-weighted average of AUC\u03a4, ss, selexipag and AUC\u03a4, ss, ACT-333679, and calculated as cAUC\u03a4, ss\u2009=\u20091/38\u2009\u00d7\u2009AUC\u03a4, ss, selexipag\u2009+\u200937/38\u2009\u00d7\u2009AUC\u03a4, ss, ACT-333679.Pharmacokinetic sampling was performed pre-dose, and at 1, 2, 4, 6, 8, and 12\u00a0h(s) post-oral morning administration on Day 1 (Period 1), and at pre-dose, 25\u00a0min, 87\u00a0min, 4, 6, 8, and 12\u00a0h post-IV selexipag infusion start of the morning administration on Day 3 (Period 2). The pharmacokinetic endpoints assessed for selexipag and the active metabolite were the area under the plasma concentration\u2013time curve (AUC) during a dose interval at steady state . For all analyses SAS\u00ae version 9.4  was used. Results were dose normalized to the lowest selexipag dose , by dividing the respective parameter by the actual dose for that patient and multiplying by 200  or 225 (IV).\u03a4, ss and dose proportionality were performed to assess whether the exclusion of those three samples in the primary analysis affected the overall conclusion.Three plasma concentrations of selexipag (two patients) and ACT-333679 (one patient) were excluded in the primary analyses as the concentrations were considered implausible when compared with the remaining measurements from the same patients. As the underlying root cause could not be identified, sensitivity analyses for AUCTwenty patients with PAH were enrolled and completed the study  to Day 3 of\u2009\u2212\u20095.7\u00a0g/L was observed. At Day 12\u2009\u00b1\u20092\u00a0days, the mean change was\u2009+\u20091.2\u00a0g/L. The changes between the visits were suggestive of normal variability of this parameter rather than a trend. None of the post-baseline individual changes in hemoglobin levels were reported as an AE. Changes from baseline in other hematology and clinical chemistry variables were unremarkable.\u03a4, ss and Cmax, ss) of selexipag were approximately two-fold higher following IV versus oral administration . While AUC\u03a4, ss and Cmax, ss of the active metabolite were comparable   was 0.82 (Additional file The tested corresponding doses of oral and IV selexipag resulted in comparable exposure to the active metabolite of selexipag (Fig.\u00a0max, ss was similar following oral and IV selexipag administration, for both selexipag and the active metabolite (Fig.\u00a0tIn this study, 20 patients with PAH who were receiving oral selexipag at a stable maintenance dose were switched to corresponding IV selexipag doses for three infusions. The switch from oral to IV selexipag and back was well tolerated with comparable exposure to the active metabolite of selexipag.The overall safety results reported in each period in this study are consistent with previous studies of oral \u20135 and IVBased on pharmacokinetic assessments, the tested IV doses that were 12.5% higher than the corresponding oral doses provided comparable exposure to the active metabolite after oral and IV selexipag administration. The exposure to selexipag increased 2.13-fold following IV versus oral administration, as expected based on the previous bioavailability study [The pharmacokinetics of selexipag and its active metabolite are known to be dose-proportional for multiple oral doses up to 1800\u00a0\u00b5g bid . In agreThe combined pharmacokinetic and safety results reported support the use of the IV formulation of selexipag as a suitable bridging option for temporary treatment interruptions of oral selexipag and may benefit patients by allowing them to remain on selexipag treatment and maintain exposure to the active metabolite. In addition, this avoids the need to either repeat up-titration of oral selexipag or switch to a parenteral formulation of another IP-receptor agonist, which will require a route for continuous infusion.This study has some limitations due to the small sample size, the short treatment duration and the potential reporting bias of AEs during IV selexipag treatment because of the open-label nature of the study. Vital signs were not monitored during or following the evening IV infusion on Day 2, therefore changes in blood pressure during this time period may not have been observed, however, no AEs of hypotension were reported at any time during the study. Finally, the number of pharmacokinetic samples collected per patient was reduced to a minimum in consideration of the patient burden. However, the pharmacokinetic data from the available time points demonstrate comparable exposure to the active metabolite after IV and oral selexipag administration.The switch from oral to IV selexipag and back was well tolerated and the safety findings are consistent with those known following treatment with oral selexipag. Exposure to the active metabolite was comparable following oral and IV selexipag administration when applying an IV selexipag dose that is 12.5% higher than the corresponding oral dose. The study results support the use of IV selexipag as a temporary option for PAH patients to avoid treatment interruptions during clinical scenarios in which the administration of oral selexipag is not possible.Additional file 1: Additional figures and tables."}
+{"text": "Plasmodium vivax VISs where subjects were treated with chloroquine (n = 24) or artefenomel (n = 8) and compared them with studies in Thailand (n = 41) and Malaysia (n = 76). In the VISs, alanine transaminase (ALT) increased to \u2265 2.5 \u00d7 upper limit of normal (ULN) in 11/32 (34%) volunteers, peaking 5\u20138 days post-treatment. Transaminase elevations were asymptomatic, were not associated with elevated bilirubin, and resolved by day 42. The risk of an ALT \u2265 2.5 \u00d7 ULN increased more than 4-fold  for every log10 increase in the parasite clearance burden (PCB), defined as the log-fold reduction in parasitemia 24 hours post-treatment. Although an elevated ALT \u2265 2.5 \u00d7 ULN was more common after artefenomel than after chloroquine , this risk disappeared when corrected for PCB. Peak ALT also correlated with peak C-reactive protein . Elevations in ALT (\u2265 2.5 \u00d7 ULN) were less common in malaria-endemic settings, occurring in 1/41 (2.5%) Thai patients treated with artefenomel, and in none of 76 Malaysians treated with chloroquine or artemisinin combination therapy. Post-treatment transaminase elevations are common in experimental P. vivax infection but do not appear to impact on participant safety. Although the mechanism of these changes remains uncertain, host inflammatory response to parasite clearance may be contributory.Liver transaminase elevations after treatment in malaria volunteer infection studies (VISs) have raised safety concerns. We investigated transaminase elevations from two human  Plasmodium falciparum1 and Plasmodium vivax2 malaria volunteer infection studies (VISs), otherwise known as controlled human malaria infection studies. These studies entail intentional infection of healthy volunteers with Plasmodium parasites, by mosquito bites,3 direct injection of sporozoites,4 or by intravenous inoculation of parasitized red blood cells.5 The latter bypasses the liver stage of infection and is referred to as the induced blood-stage malaria (IBSM) model. VISs are increasingly being used to evaluate antimalarial vaccine and drug candidates6 and have been established for P. falciparum,7P. vivax,8 and Plasmodium malariae.5 However, recent reports of elevated liver enzymes post-treatment have raised concerns related to the safety of healthy volunteers and have the potential to impede antimalarial drug development because of difficulties distinguishing drug-induced liver injury (DILI) from the effect of malaria.Liver enzyme elevations have been recently reported in both 9 DILI can be divided into intrinsic injury, which is related to a direct drug effect and is typically dose dependent, and idiosyncratic injury, which occurs in specific susceptible individuals. The latter is typically not dose related and is, therefore, more difficult to predict. Most drugs that cause severe DILI do so infrequently, typically requiring at least a few thousand exposed subjects to demonstrate the effect.9 Therefore, close attention must be paid to lower degrees of DILI during preclinical and clinical drug development. Hence, liver function test (LFT) abnormalities must be carefully evaluated during malaria VISs where an NCE is under development. Incorrect attribution of liver injury to an NCE can lead to cessation of further development; conversely, attributing the liver injury solely to malaria may risk failure to detect DILI. Currently, there is no reliable biomarker to distinguish DILI from other causes of liver injury. Therefore, better understanding of the factors associated with post-treatment LFT abnormalities in malaria VISs is required.DILI is the most common cause of cessation of development of a new chemical entity (NCE) for safety reasons.P. falciparum and P. vivax infections. It is defined as a bilirubin > 2.5 \u00d7 upper limit of normal (ULN) and an increase in transaminase > 3 \u00d7 ULN. These LFT elevations generally peak around the time of diagnosis and before commencement of antimalarial chemotherapy.14 By contrast, a distinct pattern of LFT abnormality has been reported in P. falciparum1 and P. vivax VISs.2 This is characterized by transaminase elevation without an increase in bilirubin, with a peak observed approximately 2\u20136 days post-treatment.17 Similar changes have been described in natural P. falciparum19and P. vivax infections,18 although they have been reported less commonly than malaria hepatopathy. The differences between the malaria hepatopathy and post-treatment LFT changes suggest that they may be mediated by different mechanisms. The mechanisms underlying post-treatment liver injury are incompletely understood. Possible contributing factors include an increased inflammatory response following malaria treatment, release of hepatotoxic iron products from red cells, oxidative damage, and, potentially, a hepatotoxic effect of antimalarial drugs or other concomitant drug treatment (including acetaminophen).15Malaria hepatopathy is a common phenomenon in natural malaria infection, occurring in both P. vivax IBSM studies and to evaluate possible risk factors. One of these studies entailed treatment with the investigational antimalarial artefenomel, and the other with the licensed drug chloroquine. Neither artefenomel nor chloroquine has been associated with hepatotoxicity when used as monotherapy in standard dose regimens in healthy adults23 or in children with malaria.24 However, post-treatment LFT abnormalities have been reported in malaria patients treated with artefenomel in malaria-endemic settings.26 We compared findings from the P. vivax VISs with data from a published phase IIA open-label study of artefenomel treatment of malaria in Thailand25 and from studies of chloroquine or artemisinin combination therapy conducted in Malaysia.28The aim of our study was to describe in detail the LFT abnormalities observed in two P. vivax IBSM analysis were obtained from 32 malaria-naive subjects who participated in two clinical trials at the Queensland Institute of Medical Research (QIMR) Berghofer between March 2016 and April 2017. The results of one of these studies, which used chloroquine29 and artefenomel30 as antimalarial treatment have been published. An article describing the results of the other study (NCT02573857), which used the experimental compound artefenomel as antimalarial treatment, is currently under review. Key details are included in P. vivax isolate HMP-013.29 In the artefenomel study, eight subjects were treated with a single 200 mg dose of the drug 10 days post-inoculation.30 Data from previous studies had indicated that a 200 mg dose would be subcurative,31 and as expected, 7/8 subjects had recrudescent parasitemia; they subsequently received a standard course of artemether/lumefantrine on day 21\u201328. In the chloroquine study, 24 subjects were treated with a standard 3-day course of chloroquine, totaling 25 mg/kg, beginning on day 8 (n = 8 subjects), day 9 (n = 1 subject), or day 10 (n = 15 subjects). All subjects in the chloroquine study cleared parasites.Data for the 25 This study had enrolled patients at the Hospital for Tropical Diseases in Bangkok and Shoklo Malaria Research Unit located on the northwestern border of Thailand between October 2010 and May 2012. Data from 41 adult patients with microscopically confirmed P. vivax mono- (n = 40) or mixed infection (n = 1) were analyzed. LFTs were measured before artefenomel treatment, 4 hours post-treatment, and on days 1, 2, and 7.Individual patient-level biochemistry results were made available from a previously published phase IIA open-label study.P. vivax monoinfection were enrolled from July 2013 to November 2015 as part of a randomized controlled trial27 or a prospective observational study,28 both conducted at Kota Marudu, Kudat, and Pitas district hospitals. LFTs were undertaken on samples from 85 patients. All had ALT levels tested on days 0 (before treatment) and 7, whereas AST was measured in 19/85 and 85/85 subjects on day 0 and day 7, respectively. Total bilirubin levels were tested in 76/85 subjects on day 0 and 85/85 subjects on day 7.In the study undertaken in Malaysia, adults and children > 1 year of age with 32 and analyzed on the log10 scale. The parasite clearance rate was reported using the parasite clearance half-life (PCt1/2) in hours, as previously reported.33Parasitemia was measured before treatment and at multiple time points throughout the VISs by 18S-qPCR1/2 was censored at parasitemia nadir, \u223c36 hours post-treatment, and before recrudescence. Although chloroquine was administered over 3 days, the initial parasite PCt1/2 was assumed to represent a continuous pharmacodynamic effect.29For the artefenomel VISs, the PCtpeak-24) was calculated by subtracting the parasitemia at 24 hours posttreatment initiation from the peak parasitemia on the log10 scale and is reported as a log-fold change. The 24-hour cutoff was selected because most parasites were cleared over this interval, yet parasitemia remained measurable. The peak parasitemia occurred at or around the time of treatment for all subjects.A variable was calculated to capture the biomass of parasites cleared in the first 24 hours after treatment. This variable, termed as the parasite clearance burden . The maximum temperature and clinical scores were taken from values recorded within 3 days pre- and post-treatment to ensure the changes were due to malaria infection and treatment and not to an alternative cause.Temperature, malaria clinical score, and C-reactive protein (CRP) were measured as markers of host response. Temperatures < 37.5\u00b0C were considered normal. The malaria clinical score has been developed as a tool for assessing the clinical severity of malaria in IBSM studies35 Peak ALT was categorized into \u2265 2.5 \u00d7 ULN and < 2.5 \u00d7 ULN  in line with the World Health Organization guidelines.36LFT and hematology results from the IBSM subjects were taken at screening, pre-inoculation, pre-treatment, 72 hours post-treatment, 5\u20138 days post-treatment, and at the end of study (EOS). CRP levels were retrospectively measured on a total of 31 stored plasma and 16 stored serum samples in the artefenomel cohort, and 62 stored plasma and serum samples in the chloroquine cohorts. Additional CRP results were available from tests undertaken for clinical assessments. The peak ALT level was assessed as the primary measurement of hepatocyte injury because of its greater liver specificity than AST.37 and figures were produced using GraphPad Prism v.8, San Diego, CA.38 Descriptive statistics of frequency and percentage were given for categorical measures, and continuous variables were presented as mean and 95% CI or median and interquartile range (IQR), depending on whether the variable was normally distributed or not. For the VIS participants, maximum and minimum values for all parameters  were identified from inoculation to drug treatment and from inoculation to the EOS. Correlations between continuous variables were assessed by Pearson\u2019s correlation of log10-transformed data. Logistic regression analysis was used to test for associations between explanatory parameters and peak ALT \u2265 2.5 \u00d7 ULN. A fixed cohort effect was added to the regression models to account for inherent differences between cohorts. The Mann\u2013Whitney test was used to assess for difference in ALT values between the two drug treatments.Data were analysed using Stata v.15 ,P < 0.05) in the univariate logistic regression was included in the backward stepwise regression. In the first multivariable model, the parasite clearance rate was forcibly retained into the final model; in the second multivariable model, the PCB was included. For the Thailand and Malaysia datasets, continuous measures were described using mean (95% CI), and paired t-tests on the log10 scale of the measure were used to test the differences in LFT results between day 0 (pre-treatment for Thailand artefenomel) and day 7.A backward stepwise regression was used to create two multivariable logistic models for predictors of developing a peak ALT \u2265 2.5 \u00d7 ULN. Any variable that was significant  of subjects were males. There was no difference in the age of subjects recruited in the artefenomel and chloroquine studies .R = 0.99; P < 0.001). Levels typically began to increase 3 days after treatment, peaking between days 5 and 8 after treatment and resolving between day 11 and day 32 after treatment  subjects. Alanine transaminase elevations generally exceeded AST (correlation peak ALT to peak AST reatment . The higP = 0.002) (P = 0.064).The peak ALT was greater among the eight subjects administered with artefenomel (median 5.3 [IQR: 2.2\u20139.1] \u00d7 ULN) than that among the 24 subjects who were given chloroquine  . LikewisR = 0.39; P = 0.027), with a nonsignificant trend also observed between peak parasitemia and risk of developing ALT \u2265 2.5 \u00d7 ULN  for a one log10 increase in PCB. A significant correlation between log10 peak ALT and log10 PCB  was also observed. In keeping with its more rapid antimalarial effect, the PCB was higher in the artefenomel-treated subjects than that in chloroquine-treated subjects .There was an association between CRP and the likelihood of ALT elevation: with each increase in the maximum CRP by 4 mg/L (ULN = 4 mg/L), the likelihood of a subject developing peak ALT \u2265 2 \u00d7 ULN increased by 1.43-fold (95% CI: 1.10\u20131.86); P = 0.064] to 0.84 ). No other pre-treatment laboratory parameter predicted the development of an ALT \u2265 2.5 \u00d7 ULN after controlling for the cohort effect .On multivariable analysis, peak CRP remained a significant risk factor for developing an ALT \u2265 2.5 \u00d7 ULN after controlling for the PCB but not after controlling for the cohort effect. After controlling for the PCB, artefenomel drug treatment no longer predicted the development of peak ALT \u2265 2.5 \u00d7 ULN , time to maximum drug concentration (Tmax), or area under the parasitemia curve at 96 hours post-treatment, for artefenomel or chloroquine, and the timing or severity of ALT elevations.There was no association between maximum drug concentration . Review of the source documents did not reveal any alternative causes (including exercise) for the observed LFT abnormalities.In the chloroquine study, the median peak ALT did not differ between cohort 2 (1.63 \u00d7 ULN [IQR: 0.82\u20133.46]), where ibuprofen was administered for symptom relief, and cohort 3 (1.39 \u00d7 ULN [IQR: 1.05\u20132.5]), where acetaminophen was the preferred treatment .25The median  age of the 41 participants was 26  years, and 32 (78%) were male. The mean (range) parasite count for P < 0.001). One patient had an ALT elevation of \u2265 2.5 \u00d7 ULN, with a maximum absolute increase of 156 U/L . This patient was treated with a 200 mg dose of artefenomel, the lowest dose used in the study  age was 16  years, and 63 (74%) were male. The median (IQR) parasite count at day 0 was 4,048/\u03bcL .P < 0.001), whereas in those treated with artemether/lumefantrine, the median ALT decreased from 17 (IQR: 11\u201335) at day 0 to 10 (IQR: 7\u201313) at day 7 (P < 0.001). No patient developed an ALT \u2265 2.5 \u00d7 ULN (Forty-four (51.8%) of the 85 subjects were treated with artesunate/mefloquine, 32 (37.7%) with chloroquine, 7 (8.2%) with artemether/lumefantrine and primaquine, and 2 (2.4%) with chloroquine and primaquine. Among those treated with chloroquine, the median ALT decreased from 14 (IQR: 7\u201329) at day 0 to 7.5 (IQR: 5\u201311) at day 7  subjects in the two P. vivax IBSM studies were rare in the clinical P. vivax studies, occurring in only 1/41 (2.4%) Thai patients treated with artefenomel and in none of the 76 Malaysian patients treated with chloroquine or artemisinin combination therapy. Consistent with these findings, in a previous phase II study of artefenomel (800 mg) in combination with piperaquine  for treatment of P. falciparum malaria, only 1/448 (0.2%) subjects experienced a grade 3 (5.1\u201310 \u00d7 ULN) transaminase elevation.26 Transaminase elevations were also not observed in a phase I study of artefenomel in 26 healthy volunteers without malaria.23Transaminase elevations mirroring those seen in the Supplemental Table 7). Of significance, artefenomel has a more rapid parasiticidal effect than chloroquine.31In the VISs, although the risk of developing elevations of ALT \u2265 2.5 \u00d7 ULN was higher among subjects treated with artefenomel than that among those treated with chloroquine, this association disappeared when corrected for other factors, notably the PCB (1/2) and peak parasitemia around the time of treatment, and thus, it represents the biomass of parasites cleared immediately post-treatment. As such, it distinguishes between artefenomel (a fast-killing drug) and chloroquine (a drug with a slower effect on parasite clearance), while still accounting for the overall reduction in parasite biomass post\u2013antimalarial treatment  subjects in a 46 However, even when testing has occurred 5\u20138 days post-treatment, transaminase elevations mirroring those observed in P. vivax IBSM studies appear to be uncommon in P. falciparum\u2013infected patients treated with artemisinin combination therapies (0.2\u20137.8%).53 For example, in a recent randomized clinical trial in West Africa where four different artemisinin combination therapies were evaluated in 4,710 patients experiencing 8,640 malaria episodes , the LFT was undertaken on days 0 (pre-dose), 3, 7, and 28. In this study, LFT changes resembling those we report here were rare, being reported after treatment with any of the four drug combinations in 0.9\u20133.9% of the 8,640 treatment encounters, and were more commonly seen in children < 5 years of age.19 All patients were treated with artemisinin-based regimens characterized by rapid parasite clearance, and thus more likely to have a high PCB.In clinical trials of antimalarials, clinically significant transaminase elevations occurring after treatment have not been commonly reported. Assessments of liver function during the day 5\u20138 window where ALT changes were observed in the IBSM studies may not have been universally undertaken.54 with resulting modulation of the host immune response.55 This may also explain why ALT increases post-treatment occur more often in returned travelers,47 in whom most malaria episodes will be a primary infection. Alternatively, it is also possible that genetic polymorphisms or other host characteristics such as iron deficiency could influence susceptibility to transaminase elevations following malaria treatment.57One possible explanation for the lower frequency of post-treatment increases in ALT in malaria-endemic settings is that there may be a reduced susceptibility to hepatic injury due to previous exposure to malaria,39 Rodent malaria studies have suggested that TNF-\u03b1 generated during malaria infection and free heme, produced following rapid hemolysis, act together to induce hepatocyte apoptosis and transaminase leakage.61 Testing for additional markers of hemolysis, free heme, and TNF\u03b1 would also be worth undertaking in future studies. In addition to the effects of systemic inflammation and hemolysis, it is possible, though speculative, that liver injury in primary infections may also result from killing of P. vivax parasites potentially sequestered in the liver, as now recognized to occur in the spleen,62 possibly exacerbating localized hepatic inflammation.Higher maximum CRP was associated with an increased risk of a peak ALT of \u2265 2.5 \u00d7 ULN, suggesting that systemic inflammation likely contributes to the development of post-treatment LFT abnormalities in VISs, as previously postulated.The relatively small number of subjects in this study limits statistical power, and a larger dataset would be required for a more robust multivariable analysis. Differences between cohorts meant comparisons across cohorts was difficult and could confound the analysis. In the case of PCB, it is strongly affected by both the day of treatment and the antimalarial used. In addition, as it is calculated solely from two data points, the measure may be susceptible to bias and should be prospectively evaluated in future studies.P. vivax IBSM than natural infection. Although treating physicians should remain vigilant, these changes have to date been asymptomatic, self-limiting, and not associated with adverse clinical outcomes. Researchers and pharmaceutical companies should become familiar with these findings so that the mechanisms underlying this phenomenon can be better understood and alternate explanations of the causation of LFT changes can be considered in the development of antimalarials. LFT on days 3, 5, and 7 post-treatment initiation should be incorporated, where possible, into the study design of antimalarial clinical trials.Post-treatment LFT changes appear to be more common in Supplemental materials"}
+{"text": "Plasmodium falciparum. Additional interventions are needed to control and eliminate malaria. We aimed to assess whether preventive treatment of malaria might be an effective means of reducing P falciparum infection and anaemia in school-aged children and lowering parasite transmission.The burden of malaria infection in sub-Saharan Africa among school-aged children aged 5\u201315 years is underappreciated and represents an important source of human-to-mosquito transmission of Clinicaltrials.gov for intervention studies published between Jan 1, 1990, and Dec 14, 2018. We included randomised studies that assessed the effect of antimalarial treatment among asymptomatic school-aged children aged 5\u201315 years in sub-Saharan Africa on prevalence of P falciparum infection and anaemia, clinical malaria, and cognitive function. We first extracted data for a study-level meta-analysis, then contacted research groups to request data for an individual participant data meta-analysis. Outcomes of interest included prevalence of P falciparum infection detected by microscopy, anaemia , clinical malaria (infection and symptoms on the basis of study-specific definitions) during follow-up, and code transmission test scores. We assessed effects by treatment type and duration of time protected, and explored effect modification by transmission setting. For study-level meta-analysis, we calculated risk ratios for binary outcomes and standardised mean differences for continuous outcomes and pooled outcomes using fixed-effect and random-effects models. We used a hierarchical generalised linear model for meta-analysis of individual participant data. This study is registered with PROSPERO, CRD42016030197.In this systematic review and two meta-analyses, we searched the online databases PubMed, Embase, Cochrane CENTRAL, and P falciparum prevalence , anaemia , and clinical malaria ; results for cognitive outcomes are not presented because data were only available for three trials. In our individual participant data meta-analysis, we found treatment significantly decreased P falciparum prevalence , anaemia , and subsequent clinical malaria  across transmission settings. We detected a marginal effect on cognitive function in children older than 10 years  although we found no significant effect when combined across all ages.Of 628 studies identified, 13 were eligible for the study-level meta-analysis (n=16 309). Researchers from 11 studies contributed data on at least one outcome (n=15 658) for an individual participant data meta-analysis. Interventions and study designs were highly heterogeneous; overall risk of bias was low. In the study-level meta-analysis, treatment was associated with reductions in P falciparum prevalence, anaemia, and risk of subsequent clinical malaria across transmission settings. Policy makers and programme managers should consider preventive treatment of malaria to protect this age group and advance the goal of malaria elimination, while weighing these benefits against potential risks of chemoprevention.Preventive treatment of malaria among school-aged children significantly decreases  However, since 2014 progress has slowed and the number of malaria cases has even increased in some countries.20 infants,21 and preschool children (younger than 5 years) in some malaria-endemic areas.22 There are no recommendations, however, for school-aged children, despite mounting evidence that preventive treatment of malaria among school-aged children decreases P falciparum infections, malaria-related anaemia, and improves cognitive performance.31 We aimed to do two meta-analyses of malaria treatment trials among asymptomatic school-aged children: one drawing on summary study-level data, and the other involving individual participant data that allows for subanalyses of treatment type, frequency of treatment, and intervention strategy. We aimed to use the findings to discuss the effect of school-based preventive treatment on P falciparum infection, anaemia, subsequent clinical malaria, and cognitive function, as well as the optimal treatment, regimen, target-age group, and transmission setting.WHO recommends providing intermittent preventive treatment or chemoprevention to asymptomatic pregnant women,ClinicalTrials.gov to identify malaria studies in school-aged children ,32 clinical malaria (infection and symptoms on the basis of study specific definitions) during follow-up, and code transmission test scores.We extracted study-level data without masking to author or publication, and assessed risk of bias in each study using RevMan 5.2 software. For the study-level meta-analysis, we extracted the number of participants, treatment used, dosing interval, and timing of outcome measurement. We then contacted each research group to request individual-level data, including participant age, sex, treatment group, specific geographical locations of the trial, To evaluate the effect of treatment regimens, we grouped interventions by drug class and pharmacokinetic features: sulfadoxine\u2013pyrimethamine alone, sulfadoxine\u2013pyrimethamine combined with an aminoquinoline (either amodiaquine or piperaquine), sulfadoxine\u2013pyrimethamine plus artesunate, artemisinin-based combination therapy including an aminoquinoline (artesunate\u2013amodiaquine or dihydroartemisinin\u2013piperaquine), and artemether\u2013lumefantrine. We constructed a variable to estimate the proportion of follow-up time protected by treatment for each trial , 48 to a33 Malaria Atlas estimates reflect the average prevalence of P falciparum infection among children aged 2\u201310 years (PfPR2\u201310) to within 5 km of any location in sub-Saharan Africa. In studies where fieldwork straddled multiple years, we weighted the Malaria Atlas estimates by the number of months of each year that each contributed to the study. We divided areas into WHO transmission settings based on parasite prevalence and further divided mesoendemic into two categories: low (<10%), low\u2013moderate (10% to <30%), moderate\u2013high (30% to <50%), and high (\u226550%).34To assess whether treatment effect varied by transmission setting, we extracted site-specific and year-specific malaria parasite prevalence estimates from the Malaria Atlas Project for the geocoordinates of each school or cluster midpoint involved in each study.I\u00b2 statistic, and meta-regression was done to determine whether any between-study heterogeneity of effect could be explained by study characteristics, including drug class, region of study, prevalence of P falciparum infection according to Malaria Atlas estimates, proportion of follow-up time protected, and study design.As a first step in our study-level meta-analysis, we calculated risk  ratios for binary outcomes and standardised mean differences for continuous outcomes and pooled these outcomes using using random-effects and fixed-effects models. Counts from cluster-randomised studies were divided by the design effect due to clustering before pooling them with individually randomised studies. Between-study heterogeneity was estimated using the P falciparum infection and anaemia, whereas age, but not sex, was associated with clinical malaria and code transmission test scores  when the intervention was given to both the intervention and the control groups. For multigroup factorial studies, all groups receiving antimalarial treatment were combined and compared with all groups not receiving antimalarial treatment. Age and sex were independently associated with t scores . Thus, fP falciparum infection, anaemia, and clinical malaria. There was insufficient variation in transmission setting to do the same among studies measuring code transmission test scores. We compared the effects of different treatment types with control (placebo or no treatment), and the effect of follow-up time protected by treatment type. Because malaria immunity increases with cumulative exposure to P falciparum parasites, we stratified results by age to explore the effect of intervention among children aged 5 years to less than 10 years versus children aged 10 years or older to less than 15 years. Because two of the larger trials37 fundamentally differed from others, we also did sensitivity analyses excluding these datasets. We used Stata/IC 15 software for all analyses. This study is registered with PROSPERO, CRD42016030197.To determine whether the effect of treatment varied by local transmission, we fitted an interaction term between treatment and transmission setting for The funders of the study had no role in study design, data collection, data analysis, data interpretation, or writing the report. The corresponding author had full access to all the data in the study and had final responsibility for the decision to submit for publication.PfPR2\u201310) ranged from 3% to 67%  and outcomes were measured at a median of 60 days after the last treatment dose (0\u2013180). The median proportion of follow-up time protected by treatment was 49% (2\u2013100%). Summaries of each study are provided in the 39Of 628 studies screened, 13 trials met inclusion criteria . These t% to 67% . 11 diff% to 67% . Nine trP falciparum infection . As the duration of follow-up time protected by treatment increased, the risk of P falciparum infection decreased , but there was no consistent pattern to this interaction  in the study-level meta-analysis  of six trials were published with results presented in a way that was amenable to analysis, and pooled estimates from individual participant data did not show an improvement following treatment , with a modest increase in test scores among children aged 10\u201315 years  to very high (67%) parasite prevalence. Combination drug regimens and increasing the duration of time protected by treatment improved the protection conferred by preventive treatment. While fewer studies measured the impact of treatment on clinical malaria, those that did showed a substantial benefit. The 50% reduction in clinical malaria episodes is similar to the reduction observed in early studies of bednets treated with insecticide\u2014a widely implemented malaria control measure\u2014as well as seasonal malaria chemoprevention in children younger than 5 years, which is now standard in the Sahel.41These two meta-analyses provide strong and consistent evidence that preventive malaria treatment among school-aged children decreases 42 One trial included in the study-level meta-analyses did not provide data for the individual participant data meta-analysis, and there were differences in how the effects from cluster randomised controlled trials were included in the two meta-analytic approaches: cluster-adjusted effects if available, or raw counts scaled by design effect due to clustering, were used in the summary data meta-analysis, whereas a random effect for clusters (in addition to one for studies) was used in the individual participant data meta-analysis. Although this effect appears relatively modest, even daily or weekly iron supplementation reduces anaemia in this age group by only 50%.44 In Mali, a one-time antimalarial treatment was associated with a larger reduction in the prevalence of anaemia than were weekly-doses of iron supplementation given for 10 weeks in a previous study in the same area.45 Anaemia in most malaria-endemic areas is attributable to multiple factors including malaria, helminth infections, chronic inflammation, chronic undernutrition, and micronutrient deficiencies.46 School-based antimalarial treatment addresses only the fraction of anaemia attributable to malaria, which might explain why most treatment studies had a dramatic effect on P falciparum infection, but less effect on anaemia. Thus, our meta-analysis might underestimate the benefit of preventive treatment on anaemia.Preventive treatment was also associated with a statistically significant reduction in anaemia, although the magnitude of the reduction was greater in study-level analysis (23%) compared with individual participant data analysis of all data (15%) and sensitivity analysis (21%). These differences might be due to the included studies and the analytic methods applied.14 factors such as poverty, insufficient stimulation at home, poorly resourced schools with large class sizes, poor general health, and inadequate nutrition have all been linked to decreased cognitive function.49 These factors also interact and contribute differentially to decreased cognitive function and complicate the interpretation of our results. However, several trials reported results of cognitive function improvement after malaria treatment.28 Our results might differ from these studies due to the exact methods of analysis and the conservative methods we employed to adjust for clustering. Treatment did, however, improve test scores in children aged 10\u201315 years in stratified analysis. Children aged 10\u201315 years should have higher proficiency in numeracy and writing, resulting in a better understanding of the test instructions and the ability to record their responses; therefore, results in this age group should be more reliable. For more definitive evidence, additional studies with age-appropriate outcomes sensitive to assessing the effects of preventive treatment on cognitive function in younger school-aged children might be needed.Similarly, the absence of improvement in cognitive test scores after treatment might be explained by the complexity of influences on cognitive function in this age group and the duration of interventions. While decreases in cognitive function have been linked to both cerebral malaria and to asymptomatic parasitaemia,35 which was the only study to use a screen-and-treat approach, used short-acting artemether\u2013lumefantrine as treatment, that took place in a low transmission setting showed no effect. This study design resulted in participants being protected by treatment for an estimated average of 2% of their follow-up. The inferiority of screen-and-treat interventions, compared with chemoprophylaxis or intermittent preventive treatment, is consistent with interventions to prevent malaria in pregnancy and mass treatment to interrupt transmission.51 Currently available screening methods do not detect low-density infections, which make up a larger proportion of the infections in low transmission settings.52 An additional shortcoming of screen-and-treat approaches is that only children who test positive benefit from the chemoprophylactic effect of treatment against future P falciparum infections in the near term.Our study-level analyses showed that some intervention designs did not do as well as others. Notably, the trial by Halliday and colleagues,P falciparum infection by transmission setting, the key finding is that treatment was effective in all regions and transmission settings.The antimalarial drugs used in each of these trials are well studied formulations known to be safe and well tolerated. None of the studies reported deaths related to the intervention and no unusual adverse events were reported . The var17 Three studies,37 reported that intervention significantly decreased the prevalence of gametocytes, the parasite life-stage required for transmission. Staedke and colleagues36 found that treating school-aged children decreased infection in the surrounding community by a small but statistically significant proportion. This, despite low coverage, was consistent with seasonal malaria chemoprevention among school-age children where other age groups not target for intervention have experienced concomitant reductions in parasitaemia.53Although these analyses focused on the effect of treating asymptomatic school children at the individual level, school-based malaria treatment can confer an additional community-level effect by decreasing local transmission. School-aged children are significant reservoirs of human-to-mosquito transmission.The primary limitation to these analyses is the variability between studies, including differences in the intensity and seasonality of malaria transmission in study sites, differences in the intervention drugs and frequency of dosing, as well as differences in the timing of measuring outcomes. A random-effects regression model of individual participant data with adjustment for study and participant characteristics was used to account for some sources of heterogeneity and incorporate any residual heterogeneity into the pooled effect estimates. Ultimately, this variability limits our ability to define the optimal intervention strategy for each setting. Based on our results, we can suggest guiding principles for initial policy and programmatic interventions, as well as studies to further optimise interventions.40 whereas in areas with year-round transmission, treatment each school term would be most practical. However, monthly treatment might be required in areas with high perennial transmission.27 In the studies included in these analyses, artemisinin-based combination therapies or sulfadoxine\u2013pyrimethamine with an aminoquinoline were effective drug combinations. However, because large scale chemoprevention efforts might increase drug pressure and resistance, the local first-line treatment should not be used for school-based treatment.51 Therefore, artesunate\u2013amodiaquine and artemether\u2013lumefantrine for preventive approaches should be avoided in some settings. Dihydroartemisinin\u2013piperaquine is favourable as it is frequently used as first-line treatment in Africa, has a long-half life, and has been used effectively in multiple studies. However, the only study to measure directly the effect of school-based treatment on drug resistance showed that recent treatment with dihydroartemisinin\u2013piperaquine was associated with higher prevalence of molecular markers of drug resistance.54 Sulfadoxine\u2013pyrimethamine in combination with another compound should be weighed carefully given changing resistance patterns. The ideal drug characteristics for this purpose include combination therapies with well matched half-lives, drugs with competing resistance mechanisms from first-line treatment drugs, or a rotation of drugs. The benefits of preventive treatment in this population must be weighed against the potential risk of drug resistance.Our results are robust and provide evidence for development of policy and programmatic interventions. School-based preventive treatment is effective across a wide range of transmission settings in sub-Saharan Africa. In areas with higher parasite prevalence and thus more intense transmission, higher proportions of time protected are required, either by using drugs with longer half-lives above therapeutic efficacy levels, or by more frequent dosing of shorter-acting drugs. In highly seasonal settings a single treatment at the end of the transmission season provides substantial benefits,55 Vector control has been widely applied and benefits the total population. Yet, the prevalence of infection in school-aged children remains high and additional interventions are needed to target this population. School-based preventive treatment merits further consideration as an addition to standard malaria control interventions in these areas.School-based preventive treatment should be considered for implementation alongside vector control and other interventions to increase protective effects, reduce community transmission, and limit opportunities to select for resistant parasites.58 Importantly, as transmission declines in malaria-endemic areas, evidence suggests that the prevalence of P falciparum infection and malaria disease will likely increase in school-aged children.61 Thus, developing interventions to target this age group will help to counter this epidemiological shift in infection and disease burden.9 Additionally, access to primary school and school attendance rates are increasing in sub-Saharan Africa, providing an efficient delivery point for preventive treatment similar to school-based deworming campaigns and nutrition programmes.63 Indeed, providing preventive treatment alongside other interventions could yield synergistic effects to decrease anaemia, improve cognitive function, and educational attainment. Initial policy support for this intervention would facilitate operational and implementation research to evaluate alternative drugs or drug strategies, assess the effect of combined interventions, investigate the community-level effect of school-based treatment on transmission, monitor for rebound morbidity and mortality, and determine cost-effectiveness under operational conditions. Moreover, these results would enable policy makers to weigh up the risks and benefits of the intervention.Providing preventive treatment to school-aged children could theoretically hinder the acquisition or maintenance of immunity. This, however, has not been widely observed in studies of intermittent preventive treatment among infants or chemoprevention in children.Despite historic strides towards malaria elimination over the past 15 years, progress hangs in the balance. Additional interventions are urgently needed, particularly ones that target populations responsible for human-to-mosquito transmission. Our analysis supports preventive treatment of malaria among school-age children that will decrease the burden of disease in this vulnerable age group.mmc1"}
+{"text": "Plasmodium falciparum malaria in south-western Ethiopia is poorly documented. Regular monitoring of drug efficacy is an important tool for supporting national treatment policies and practice. This study investigated the therapeutic efficacy of AL for the treatment of P. falciparum malaria in Ethiopia.The efficacy of artemether-lumefantrine (AL) for treatment of uncomplicated P. falciparum malaria. Real-time polymerase chain reaction (PCR) and nested PCR reaction methods were used to quantify and genotype P. falciparum. A modified protocol based on the World Health Organization 2009 recommendations for the surveillance of anti-malarial drug efficacy was used for the study with primary outcomes, clinical and parasitological cure rates at day-28. Secondary outcomes assessed included patterns of fever and parasite clearance. Cure rate on day-28 was assessed by intention to treat (ITT) and per protocol (PP) analysis. Parasite genotyping was also performed at baseline and at the time of recurrence of parasitaemia to differentiate between recrudescence and new infection.The study was a one-arm, prospective, evaluation of the clinical and parasitological, responses to directly observed treatment with AL among participants 6\u00a0months and older with uncomplicated Of the 80 study participants enrolled, 75 completed the follow-up at day-28 with ACPR. For per protocol (PP) analysis, PCR-uncorrected and-corrected cure rate of AL among the study participants was 94.7% (95% CI 87.1\u201398.5) and 96% (95% CI 88.8\u201399.2), respectively. For intention to treat (ITT) analysis, the cure rate was 90% (95% CI 88.8\u201399.2). Based on Kaplan\u2013Meier survival estimate, the cumulative incidence of failure rate of AL was 3.8% (95% CI 1.3\u201311.4). Only three participants 3.8% (95% CI 0.8\u201310.6) of the 80 enrolled participants were found to be positive on day-3. The day three-positive participants were followed up to day 28 and did not correspond to treatment failures observed during follow-up. Only 7.5% (6/80) of the participants were gametocyte-positive on enrollment and gametocytaemia was absent on day-2 following treatment with AL.The therapeutic efficacy of AL is considerably high (above 90%). AL remained highly efficacious in the treatment of uncomplicated malaria in the study area resulted in rapid fever and parasite clearance as well as low gametocyte carriage rates despite the use of this combination for more than 15\u00a0years. Plasmodium protozoan parasites, threatens over half the world\u2019s population  which was manufactured by Ipca Laboratories Ltd  was provided by the Ethiopian FMoH through WHO support. Drug dosage was determined according to the revised WHO weight-based guideline . BrieflyOn day 0 (enrollment day), participants who were successfully treated with the first dose of AL were given an appointment card bearing patient name and identification code and next scheduled visit date, and the evening dose to be administered at home by health extension workers. Participants were then advised to come back for treatment the next 2 days . Scheduled follow-up visits were on day-3, day-7, day-14, day-21, and day-28. There were unscheduled visits as well when a participant felt sick. Microscopy was performed during each subsequent visit to determine infection status, species, and parasite density.Finger-pick blood samples were used to measure haemoglobin using a portable spectrophotometer (Haemocue).Thick and thin blood films were collected from each patient at screening. Blood films were also obtained on days 1, 2, 3, 7, 14, 21, 28 and any other day, if the patient returned due to some complaints spontaneously . Giemsa Plasmodium spp.  genomes was assayed for the in vitro quantification of all Plasmodium species genomes by targeting the 18S ribosomal RNA (18S) gene according to the protocol of Primerdesign\u2122 Ltd [Primerdesign\u2122 Genesig standard kit for ign\u2122 Ltd . Each rePlasmodium falciparum genome was analysed for the in vitro quantification of P. falciparum genomes by targeting the plasmepsin 4 gene according to the protocol of Primerdesign\u2122 Ltd [ign\u2122 Ltd .P. falciparum polymorphic genes msp1 and msp2 was performed as per WHO protocol [Dried blood spots were obtained for PCR analysis at enrolment (day 0) and on follow-up days 7, 14, 21, and 28. PCR genotyping was performed on paired dried blood spots in the case of parasitaemia detected on or after day-7 to distinguish between recrudescence and re-infection for all treatment failures. PCR genotyping of protocol \u201331. The Treatment outcomes were classified based on parasitological and clinical outcomes as recommended by the WHO . Efficacp\u2009<\u20090.05 was considered significant during the analysis.Data entry and analysis was done by using the WHO designed Excel spreadsheet  and SPSSA total of 282 febrile participants were screened and of these, 80 participants were included in this study. The rest who were not fulfilling the WHO revised protocol for malaria drug therapeutic efficacy study  criteriaThe age, axillary temperature, Hb level, body weight, parasitaemia and gametocyte carriage of the study participants are summarized in Table Four treatment failures, three LPF (one case at day-14 and two cases at day-21) and one LCF case at day-14 were observed, giving PCR-uncorrected failure rate of 5.3% (4/76) (95% CI 0.8\u201318.2). By PCR correction, only the LCF was confirmed as a reinfection case with a failure rate of 4% (95% CI0.8\u201311.2). There was seen no early treatment failure (ETF). For per protocol (PP) analysis, PCR-uncorrected cure rate of AL among the study participants was 94.7% (95% CI 87.1\u201398.5) and the PCR corrected cure rate was 96% (95% CI 88.8\u201399.2). For intention to treat (ITT) analysis, the cure rate was 90% (95% CI 88.8\u201399.2) (Table p\u2009<\u20090.05) in age group less than 15\u00a0years (702.4\u2009\u00b1\u20091390.46) compared to age group greater than 15\u00a0years (408.3\u2009\u00b1\u20091041.09) implying that study participants age greater than 15\u00a0years cleared the parasite faster than study participants age less than 15\u00a0years.Based on qPCR quantitative parasite assessment, parasitaemia detection rate was 38.8% (31/80) on day-1, and declined to 18.8% (15/80) on day-2 and 3.8% (3/80) on day-3. Accordingly, parasite clearance rate was high that 61.2% of the participants cleared parasitaemia on day-1, 81.2% on day-2 and 96.2% on day-3  at the day of recruitment and decreased to 47.8% (38/80) on day 1, 13.8% (11/80) on day-2 and 2.5% (2/80) on day-3. Accordingly, fever clearance rate was 87.2% (69/80) on day-2, 97.5% (74/80) on day-3  gametocyte carriers, among all study participants, were detected at enrolment: Of these, 6.2% 2/30) detected in\u2009<\u200915\u00a0years and 10% (5/50) was detected in\u2009\u2265\u200915\u00a0years. Of the day-0 gametocyte carriers detected, three cases on day-1 and four cases on day-2 were cleared giving gametocytaemia clearance rate of 42.9% (3/7) and/or 57.1% (4/7) respectively. The proportion of gametocyte carriage per total study participants was declined from 8.8% (7/80) on day 0 to 5% (4/80) on day 1, 3.8% (3/80) on day 2 and totally disappeared on day 3 , vomiting (3.4%), Shortness of breath (2.5%), cough (1.3%), diarrhea (1.3%) and joint/muscle pain (2.5%). Most of these probable AEs disappeared with the clearance of parasitaemia except cough. Cough persisted for some time beyond parasite clearance.. and 99.3% in Tanzania by Ishengoma et al. were demonstrated [In this study, the PCR-corrected cure rate 96% (95% CI 88.8\u201399.2), which showed the high therapeutic efficacy of AL since its introduction for the treatment of uncomplicated falciparum malaria in the study area, meeting the WHO recommendation that cure rates for falciparum malaria should be at least 90% . The obsnstrated , 34. Mosnstrated .Early treatment failure was not observed in this study, whereas three LPF were 3.9% (95% CI 0.8\u201311.1) in PCR uncorrected and 4% (95% CI 0.8\u201311.2) treatment failure was observed in PCR corrected data. Several studies \u201339 in whAccording to the Kaplan Meier PP survival analysis in the current study, the PCR-corrected AL failure rate was 3.8% (95% CI 1.3\u201311.4) and the cumulative incidence of success rate of AL among study participants was 96.2% (95% CI 88.6\u201398.7) . AL remains highly efficacious in the treatment of uncomplicated malaria in the study area achieving rapid fever and parasite clearance as well as low gametocyte carriage rates despite the use of this combination for more than 15\u00a0years. Day-three parasitaemia warrants a close monitoring of the efficacy of AL in the future and should be continued in order to generate evidence to support national malaria treatment policy and practice."}
+{"text": "Artemisinin-based combination therapy (ACT), such as artemisinin-piperaquine (AP), dihydroartemisinin-piperaquine (DP), and artemether-lumefantrine (AL), is the first-line treatment for malaria in many malaria-endemic areas. However, we lack a detailed evaluation of the cardiotoxicity of these ACTs. This study aimed to analyze the electrocardiographic effects of these three ACTs in malaria patients.  We analyzed the clinical data of 89 hospitalized patients with falciparum malaria who had received oral doses of three different ACTs. According to the ACTs administered, these patients were divided into three treatment groups: 27 treated with AP (Artequick), 31 with DP (Artekin), and 31 with AL (Coartem). Electrocardiograms and other indicators were recorded before and after the treatment. The QT interval was calculated using Fridericia\u2019s formula (QTcF) and Bazett\u2019s formula (QTcB).  Both QTcF and QTcB interval prolongation occurred in all three groups. The incidence of such prolongation between the three groups was not significantly different. The incidence of both moderate and severe prolongation was not significantly different between the three groups. The \u0394QTcF and \u0394QTcB of the three groups were not significantly different. The intra-group comparison showed significant prolongation of QTcF after AL treatment.  Clinically recommended doses of DP, AL, and AP may cause QT prolongation in some malaria patients but do not cause torsades de pointes ventricular tachycardia or other arrhythmias.  According to the World Malaria Report published by the World Health Organization (WHO) in 2019, the global incidence rate of malaria has decreased from 71 cases per 1000 people in 2010 to 57 cases per 1000 people in 2014, but this rate of decline has begun to slow down since 2014Plasmodium falciparum malaria in more than 80 countries,,,-ACT has been used as the first-line treatment for uncomplicated ,The WHO redefined mass drug administration (MDA) in 2017,With reports of many antimalarial drugs involving QT prolongation, the cardiotoxicity of such drugs has received renewed attention. Prolongation of ventricular repolarization can lead to effective refractory period prolongation, and an electrocardiogram (ECG) will reflect the QT interval prolongationWe conducted a retrospective analysis of clinical data of 89 hospitalized falciparum malaria patients who had received oral doses of three different ACTs in Cambodia in 2005. Positive results for rapid diagnostic tests (RDTs) and microscopy were used as the basis for laboratory-verified cases. Depending on the ACTs administered, we divided the patients into three treatment groups: 27 treated with AP, 31 treated with DP, and 31 treated with AL. ECGs and other indicators were recorded before and after treatment. The QT interval was calculated using Fridericia\u2019s and Bazett\u2019s formulas.The inclusion criteria were as follows: (1) malaria symptoms; (2) malaria falciparum found in peripheral blood smears on microscopic examination, and (3) no antimalarial medications taken 7 days before enrollment.The exclusion criteria were as follows: (1) pregnant or breastfeeding, (2) age < 7 or > 65 years, (3) nonmalarial febrile disease, (4) taken antimalarial medicine in the past 7 days; and (4) history of allergic reactions to DP, AL, AP, or similar drugs.We defined fever as a body temperature > 37.3\u00b0C. Body temperature of febrile patients was measured every 8 h until it remained normal for 24 h, and then measured once a day. Thick and thin blood smears were taken at 07:00 and 17:00 from the day of the first dose to the day of parasite clearance. Negative blood smears were defined as the examination of 200 fields, each of which had parasites less than 1/200 white blood cells. Parasite clearance was defined as three consecutive negative blood smears.ECGs were obtained before treatment and 4 h after the final dose. All ECG recordings were sent to the First Affiliated Hospital of Guangzhou University of Chinese Medicine for blinded manual adjudication. QTcF and QTcB were corrected using Fridericia\u2019s and Bazett\u2019s formulas, respectively, as follows: \u0394QTc was defined as the difference in QTc before and after drug intervention. Moderate and severe prolongation were defined as the QT interval prolongation > 30 ms and > 60 ms, respectivelyPlasmodium species and its corresponding load. Adverse drug reactions were monitored and recorded. If vomiting occurred within 1 hour of taking the drug, the drug was readministered. Patients were hospitalized for 7 days or until malaria symptoms disappeared or the parasites cleared. All patients were followed up weekly from the start of treatment to the 28th day. If any discomfort occurred during the follow-up, patients could return to the hospital for additional follow-up. At enrollment, all patients underwent comprehensive physical examinations, including assessment of clinical symptoms of malaria and neurological examinations. Microscopic examinations of thick and thin blood smears confirmed the presence of P< 0.05 was considered significant. After performing normality and variance homogeneity tests, the rank sum test (Stata 13.0) or chi-square test was used to count the medians or constituent ratios. We recruited 89 patients: 31 in the DP group, 31 in the AL group, and 27 in the AP group. Among them, 67 (75.28%) were men, 22 (24.72%) were women and 35 (39.33%) were aged \u2264 20 years, 31 (34.83%) were aged 21-30 years, 13 (14.61%) were aged 31-40 years, 7 (7.87%) were aged 41-50 years, and 3 (3.37%) were aged > 50 years. At baseline, the DP group had 31 patients with fever, AL group had 30 patients with fever and 1 with fever but normal baseline body temperature, and AP group had 23 patients with fever and 4 with fever but normal baseline body temperature. The time of recovery from fever was not recorded for 1 patient in the AP group. Body temperatures of all febrile patients returned to normal during the 7-day hospitalization. Blood smears of all patients exhibited parasite clearance during this period. The time to parasite clearance for one and three patients in AL and AP groups, respectively, could not be calculated owing to failure of collection of blood smears at a specific time. Because of the influence of fever, the difference in heart rate between patients before and after treatment is significant, necessitating the use of corrected QT. All DP, AL, and AP groups exhibited a prolonged QT interval. In the DP group, 22 patients (70.97%) had QTcF prolongation, with 7 (22.58%) and 3 (9.68%) having moderate and severe prolongations, respectively. Similarly, 14 patients (45.16%) had QTcB prolongation, with 2 (6.45%) patients each having moderate and severe prolongations. In the AL group, 24 patients (77.42%) had QTcF prolongation, with 7 (22.58%) and 5 (16.13%) having moderate and severe prolongations, respectively. Similarly, 16 patients (51.61%) had QTcB prolongation, with 2 (6.45%) and 5 (16.13%) having moderate and severe prolongations, respectively. In the AP group, 15 patients (55.56%) had QTcF prolongation, with 6 (22.22%) and 2 (7.41%) having moderate and severe prolongations, respectively. Similarly, 13 patients (48.15%) had QTcB prolongation, with 3 (11.11%) and 1 (3.70%) having moderate and severe prolongations, respectively. We did not observe drug-related QTc > 500 ms in the AL and AP groups; one case of QTcF and one case of QTcB > 500 ms in the DP group were considered to be drug-related. The comparison revealed that the incidence of QT interval prolongation among the three groups was not significantly different. We did not observe TdP or other arrhythmias in this study. Additionally, \u0394QTcF and \u0394QTcB values of the three groups were not significantly different. Comparing the QTc before and after the treatment, we found that QTcF in the AL group was significantly prolonged after the treatment, and there was no significant difference in QTc before and after the treatment in the other two groups. Plasmodium species and transmitted from humans to humans or from animals to humans by mosquitoes. The WHO guidelinesMalaria is an infectious disease caused by The WHO Malaria Policy Advisory Committee (MPAC), which met on March 22-24, 2017, expressed that DP may account for 1 cardiac death among 200,000 treated patients. However, AL was not associated with such sudden deaths. The MPAC mentioned that the cardiotoxicity of halofantrine, in addition to other antimalarial drugs, is negligible but not investigated in detail. Cases of TdP or life-threatening arrhythmias caused by drugs are rare, but there is currently no simple screening method to identify high-risk individuals.k), especially those affected by the human fibroblast subunit encoded on chromosome 7 ,-,P. falciparum can be isolated from cardiac microvessels, though severe malaria rarely causes significant myocardial dysfunction or arrhythmia; therefore, we can exclude the impact of Plasmodium species on cardiac activityThe mechanism of cardiac ventricular myocyte repolarization is mainly related to the outward current of potassium ions  during acute infection may also affect the QT interval by inhibiting the function of cardiomyocyte ion channelsIn this study, we retrospectively studied the effects of three artemisinin complexes on the ECG findings of malaria patients by conducting a case-series analysis. This study has some limitations. First, there was a certain bias with respect to the study subjects. To avoid serious adverse events, only patients with non-severe malaria were included in the study. Because local men have more exposure to the field labor environment than women, there were more cases of malaria in men; hence, there is a sex bias among the cases in this study. Second, because of the medical conditions at that time, the ion changes before and after treatment were not monitored, making it impossible for us to evaluate the effect of the drugs on the QT interval through ion channels. Third, the ECGs of patients with prolonged QT intervals were not continuously tracked; thus, we cannot evaluate the prognostic ability of patients\u2019 ECGs.Though DP, AL, and AP treatment can cause QT prolongation in some malaria patients, we did not observe TdP or other types of arrhythmias in this study. Therefore, we can consider that the use of these three ACTs at their clinically recommended doses does not produce significant QT interval prolongation and that they can be safely used for malaria treatment. However, this study did not represent all patients with malaria. Patients with a history of heart disease or factors that may cause QT interval prolongation should be closely monitored during medication."}
+{"text": "Day\u00a07 plasma lumefantrine concentration is suggested as a predictor for malaria treatment outcomes and a cut-off of \u2265\u2009200\u00a0ng/ml is associated with day\u00a028 cure rate in the general population. However, day\u00a07 lumefantrine plasma concentration can be affected by age, the extent of fever, baseline parasitaemia, and bodyweight. Therefore, this study assessed the usefulness of day\u00a07 lumefantrine plasma concentration as a predictor of malaria treatment outcome in under-fives children treated with generic or innovator drug-containing artemether-lumefantrine (ALu) in Tanzania.\u00ae) in comparison with the innovator\u2019s product (Coartem\u00ae). Children with uncomplicated malaria aged 6\u201359\u00a0months were recruited and randomized to receive either generic or innovator\u2019s product. Children were treated with ALu as per World Health Organization recommendations. The clinical and parasitological outcomes were assessed after 28\u00a0days of follow up. PCR was performed to distinguish recrudescence and re-infections among children with recurrent malaria. Analysis of day\u00a07 lumefantrine plasma concentration was carried out using a high-performance liquid chromatographic method with UV detection.This study was nested in an equivalence prospective study that aimed at determining the effectiveness of a generic ALu (ArtefanThe PCR corrected cure rates were 98.7% for children treated with generic and 98.6% for those treated with the innovator product (p\u2009=\u20091.00). The geometric mean (\u00b1\u2009SD) of day\u00a07 plasma lumefantrine concentration was 159.3 (\u00b1\u20092.4)\u00a0ng/ml for the generic and 164 (\u00b1\u20092.5)\u00a0ng/ml for the innovator groups, p\u2009=\u20090.87. Geometric mean (\u00b1\u2009SD) day\u00a07 lumefantrine plasma concentration between cured and recurrent malaria was not statistically different in both treatment arms . Nutritional status was found to be a determinant of recurrent malaria \u2009=\u20093(1.1\u20138.2), p\u2009=\u20090.029.Using the recommended cut-off point of\u2009\u2265\u2009200\u00a0ng/ml, day\u00a07 plasma lumefantrine concentration failed to predict malaria treatment outcome in children treated with ALu in Tanzania. Further studies are recommended to establish the day\u00a07 plasma lumefantrine concentration cut-off point to predict malaria treatment outcome in children. Malaria is still a burden disease in sub-Saharan African countries including Tanzania . The preChildren under 5\u00a0years of age are the most affected group because of lower immunity against malaria . PartialALu is recommended as the first-line anti-malarial drug for the treatment of uncomplicated malaria in mainland Tanzania . The com\u00ae, which is a product from the innovator company Novartis, was prequalified to be used in the country. Due to poor availability and high cost of innovator\u2019s product, since 2013 generic ACT medicines were approved by the government to be used in the management of uncomplicated malaria .A linear regression analysis was conducted to determine factors associated with day\u00a07 lumefantrine plasma concentration for children treated with ALu. The factors included in the analysis were gender, age, body weight, mid-upper arm circumference (MUAC), baseline temperature (fever on admission), baseline haemoglobin and parasitaemia. None of the variables was associated with day\u00a07 plasma lumefantrine concentration.Several variables were analysed against treatment outcome to assess determinants of recurrent malaria in children under 5\u00a0years of age treated with ALu. Factors assessed were gender, age, weight, baseline temperature, baseline parasitaemia, baseline haemoglobin, nutritional status (MUAC) and day\u00a07 lumefantrine concentration. Only nutritional status (MUAC) was found to be a determinant of recurrent malaria on multivariate analysis with adjusted hazardous ratio  of 3 (1.1\u20138.2) at a p-value of 0.029  . Indeed,Plasmodium species to artemisinins, therefore supporting the observed high cure rate. Despite these findings, it was observed that there were re-infections in some children. This could be explained by the endemicity of malaria in the study area as well as the vulnerability of children under 5\u00a0years of age, due to inadequate immunity to malaria [In the current study, most of the observed recurrent malaria was due to re-infection and only 1 recrudescence was observed in each group of treatment arms on day\u00a028. The observed small number of recrudescence indicates that the artemisinin derivative (artemether) is still effective in Tanzania. Recrudescence after being treated with ALu is highly observed in areas with reported emergence of artemisinin resistance , 40. ThiLinear regression analysis indicated that age, gender, weight, and MUAC, baseline haemoglobin, temperature, and parasitaemia have no association with day\u00a07 plasma lumefantrine concentration. These results are contrary to the findings from the general population where factors such as baseline parasitaemia, unsupervised administration of ALu, and baseline haemoglobin levels have been reported to affect day\u00a07 plasma concentration and malaria treatment outcome .In this study, weight, baseline temperature, baseline parasitaemia, baseline haemoglobin, day\u00a07 plasma lumefantrine concentration and nutritional status (MUAC) were assessed against recurrent malaria using Cox regression analysis. All factors except nutritional status were not determinants of recurrent malaria. This finding is in line with previously reported observations in which children with malnutrition were found to be at a high risk of acquiring malaria infection . MalnutrThe sample size calculation did not take into account different age groups for children aged <\u20095\u00a0years. This probably impaired the power of the study to establish the effect of age on day\u00a07 plasma lumefantrine concentration and treatment outcome in young children. Because it takes up to 2\u00a0years for children to acquire premunition immunity against malaria following several exposures, the larger sample size is recommended in future studies. The trial was unsupervised and parents/guardians were asked to bring medicine blister packs with children on day\u00a03 for assessment of adherence. Due to limitations of pill count as a method for assessment of adherence, it could not be verified with certainty whether all children were administered ALu tablets by parents or guardians as prescribed. In addition, the nutritional status of the study participants was measured using MUAC only.Using the recommended cut-off point of \u2265\u2009200\u00a0ng/ml, day\u00a07 plasma lumefantrine concentration failed to predict malaria treatment outcome in children treated with generic or innovator products containing ALu for uncomplicated malaria in Tanzania. Nevertheless, day\u00a07 plasma lumefantrine concentration levels were comparable in both generic and innovator product treatment arms. The study to establish the day\u00a07 plasma lumefantrine concentration cut-off point to predict malaria treatment outcome in children, as well as the role of DBL in the treatment of malaria, is recommended. Taken together, the findings of this study indicate that the use of ALu generic products should be continued for the management of uncomplicated malaria in children in Tanzania."}
+{"text": "Plasmodium vivax malaria, the changing trend of parasite clearance time, and fever clearance time during 2000\u20132016 in South Korea. Median parasite clearance time and fever clearance time increased significantly over the study period. Chloroquine was mostly underdosed when used to treat P. vivax malaria. We reviewed the clinical efficacy of chloroquine for  Plasmodium vivax is the only form of malaria indigenous to South Korea . For PCT and FCT, we reported median and interquartile ranges (IQR) because Shapiro-Wilk test results showed that outcome measures did not follow a normal distribution. We compared trend analyses for PCT and FCT by enrollment periods using a linear-by-linear association. This study was approved by the local institutional review board. Statistical analyses were performed with SPSS Statistics 20  age of patients was 41.7 +14.2 years. None of the patients died of malaria during the study period; 44 relapsed. We divided the years of the study period into 3 ranges. During 2000\u20132005, 404 malaria cases occurred; that number increased to 566 during 2006\u20132010, then decreased to 229 during 2011\u20132016. . The proportion of patients whose PCT was \u02c348 hours significantly increased over the 17 years from 76.3% during 2000\u20132005 to 86.6% during 2011\u20132016 (p = 0.03). In addition, the proportion of PCT at \u02c372 hours also significantly increased from 30.5% during 2000\u20132005 to 52.4% during 2011\u20132016 (p = 0.005). P. vivax malaria was diagnosed. Among those patients, median FCT increased from 28.0 hours during 2000\u20132005 to 48.0 hours during 2011\u20132016 (p<0.001). The proportion of patients whose FCT was \u02c348 hours increased from 17.6% during 2000\u20132005 to 48.1% during 2011\u20132016 (p<0.001). The proportion of patients whose FCT was \u02c372 hours also increased significantly from 5.7% during 2000\u20132005 to 16.0% during 2011\u20132016 (p = 0.001). We found fever in 1,074 (89.6%) of 1,199 patients at the time All of the study patients were treated with either hydroxychloroquine or chloroquine phosphate. The mean total dose of chloroquine base administered was 23.0 mg/kg body weight in 838 patients (69.9%), which is less than the WHO-recommended target dose of 25 mg/kg. The trend of the mean of the total dose of chloroquine base decreased from 23.5 mg/kg during 2000\u20132005 to 22.1 mg/kg during 2011\u20132016 (p<0.001). In addition, the proportion of patients receiving less than the WHO-recommended dose increased from 68.8% during 2000\u20132005 to 77.3% during 2011\u20132016 (p = 0.02).>25 mg/kg or <25 mg/kg, we found that men were more likely than women to be in the <25 mg/kg group   . Median P. vivax susceptibility to chloroquine that could be seen before the emergence of WHO-recognized drug resistance. Mass chemoprophylaxis of military personnel with hydroxychloroquine started in 1997 in South Korea (P. vivax emerging. This finding may explain the failure of the prophylactic hydroxychloroquine treatment to widely protect South Korean army soldiers from P. vivax malaria by 2000, chloroquine-resistant cases occurring by the early to mid-2000s (In this study, we made 2 important findings. First, the trend of increased PCT and FCT in South Korea might be ascribed to decreased P. vivax, WHO recommends oral chloroquine at a total dose of 25 mg base/kg. Lower total doses are not recommended because they encourage the emergence of resistance (Second, in areas with chloroquine-sensitive This study has several limitations. First, parasite clearance was identified in only one third of study patients, but malaria smears were taken once daily, so clearance of parasites could have occurred at any hour in the interval between slides being taken, which could have affected the calculation of PCT. Second, temperature measurements might have been affected by the use of antipyretics and patients were not checked for fever hourly during hospitalization, either of which could have affected the calculation of FCT. Third, due to the retrospective nature of the study, we could not obtain other information that might affect estimates of PCT, such as patient vomiting, which indicates that the medication may not have been absorbed normally, or FCT, such as duration of fever before malaria diagnosis. Prospective studies are needed to more accurately assess whether PCT or FCT has increased. P. vivax malaria in South Korea; thus, clinicians should adhere to weight-based treatment guidelines. In summary, increased PCT and FCT might be ascribed to reduced parasite susceptibility to chloroquine that could be seen before emergence of drug resistance in South Korea. Therefore, a proactive and continuous surveillance system for PCT and FCT is needed. In addition, chloroquine is mostly underdosed in the treatment of"}
+{"text": "Malaria is a major public health concern in Togo. The Est-Mono district of Togo has a population of 150,000. Accordingly, the Guangzhou University of Chinese Medicine, China and the Ministry of Health and Social Security, Togo launched a nationwide Mass Drug Administration Project with artemisinin\u2013piperaquine (AP) in Est-Mono. Before launching this project, the sensitivity test of AP was conducted in a general clinic in Elawagnon, Togo. With this background, we evaluated the efficacy and safety of AP for the treatment of uncomplicated falciparum malaria in children under the age of 5 years.merozoite surface protein-2 (msp-2) polymorphism. The primary end point was a 28-day cure rate and polymerase chain reaction (PCR)-corrected reinfection and recrudescence. This research followed the standardized World Health Organization (WHO) protocol for the assessment of the efficacy of antimalarial drugs.Children aged 6\u201359 months with uncomplicated falciparum malaria were enrolled in this study. The selected patients were treated with a combination regime of artemisinin\u2013piperaquine. The patients were followed up for 28 days, during which signs of the following were observed for: the duration for fever clearance, parasitemia density, gametophyte generation, cure rate, hemoglobin level, and msp-2 polymorphism study of 24 children treated for parasite infection, one case of msp-2 with 3D7 haplotype and FC27 haplotype was noted, indicating its recrudescence, with a frequency of 4%. The remaining 23 cases could have been of reinfection, with a frequency of 96%. No serious adverse reactions occurred, and AP was well-tolerated by all patients.A total of 91 children with uncomplicated falciparum malaria were enrolled in this study. Adequate clinical and parasitological responses (ACPRs) before and after PCR-correction were 66 (73%) and 90 (99%), respectively. The average hemoglobin level in the patient increased by 0.05 g/dl per day (p < 0.0001) after the treatment. The gametophyte generation did not decline at the beginning of the treatment; however, after 14 days, it declined . In the Plasmodium falciparum remains sensitive to artemisinin\u2013piperaquine, necessitating its trial in this region.Artemisinin\u2013piperaquine was found to be an effective combination for treating uncomplicated falciparum malaria in children aged <5 years in Togo, and the drugs were well-tolerated. In Togo, http://www.chinadrugtrials.org.cn/eap/main.Trial registration: ECGPHCM No. B2017-054-01; MHSST AVIS N\u00b0 0001/2016/CBRS du 07 janvier 2016. Registered 17 March 2014,  Malaria remains a major public health concern in Togo . In 2018The Ministry of Health and Social Security of Togo (MHSST) decided to implement chemoprophylactic measures for handling seasonal malaria in children under the age of 5 years in the Grassland, Kara, and Central regions . In 201The efficacy and safety of AP have not yet been tested in children under the age of 5 years in Togo. Therefore, sensitivity test for AP was conducted at a general clinic in the Elawagnon county before launching the MDA program universally. With this background, we evaluated the efficacy and safety of AP for the treatment of uncomplicated falciparum malaria in children under the age of 5 years in Elawagnon, Prefecture on Est-Mono province, Togo.2 in Togo and has a population of 89,060, with a population growth rate of 1.03% in 2010  were included in the study. Moreover, the participants should be with a single infection caused by Plasmodium falciparum parasite at densities of 2,000\u2013250,000/\u03bcl and with the ability of intaking oral drugs. Informed consent of the patient's parent or legal guardian was obtained after confirming their convenience to attend clinic examinations and confirming no history/allergic reaction/contraindications to AP in the patients.Children aged 6\u201359 months with body temperature \u226537.5\u00b0C, no signs or symptoms of severe malaria, and without obvious symptoms of fever or other symptoms of severe malnutrition  were administered in 1/2 tablet dosage at every 24 h over 2 days for children aged 6\u201324 months and 3/4 tablet dosage at every 24 h over 2 days for children aged 25\u201359 months. The medicines were administered under adult supervision. All study participants were provided with acetaminophen treatment every 6 h until the fever symptoms resolved completely.AP tablets  of P. falciparum in order to distinguish between incidence of recrudescence and reinfection . The henfection .Clinical outcomes were evaluated with reference to the classification system suggested by the WHO  analysis was performed as the primary end-point. The PP population included all participants whose primary endpoint data was at day 28, who received a full course of AP, and who adhered to the follow-up visit schedule.The GraphPad PRISM 7.0  was employed for all statistical analyses. The Shapiro\u2013Wilk normality test was used to test the normality of the data. For normally distributed data, t-test was used for comparison between two groups in combination with Welch's t-test; and one-way analysis of variance (ANOVA) and Tukey's test were used for multiple comparisons among multiple groups. For nonparametric data, Mann\u2013Whitney test was employed to compare between two groups, while Kruskal\u2013Wallis and Dunn test were used to compare among multiple groups. Bidirectional ANOVA was employed for two-dimensional grouping data. Additionally, parasite clearance time and fever clearance time were represented by the mean \u00b1 standard error of the mean (SEM).P. ovale and P. malariae, 33 (16%) tested positive for parasitemia with density <2,000 parasites/\u03bcl, two (1%) could not fulfill the 28-day follow-up, one (0.5%) received other antimalarial treatment on the first day of the study duration, and one (0.5%) was lost to follow-up. Finally, 91 (44%) patients who met the inclusion criteria were included in this study  tested negative for malaria by microscopy, 7 (3%) tested positive for mixed infections by The baseline clinical and laboratory characteristics of the study subjects are shown in After AP treatment, most patients experienced rapid remission of clinical symptoms. For instance, their average body temperature of 38.5\u00b0C on day 0 fell to 36.5\u00b0C on day 2 (p < 0.0001), with 79 (87%) patients reporting a temperature of <37\u00b0C  for AP (3D7 and FC27) were detected in 24 patients who remained positive for malaria despite AP treatment by nested PCR. In these patients, msp-2 of all isolates was analyzed before and after treatment. The condition of only one patient, whose 3D7 and FC27 haplotypes of msp-2 were the same before and after taking the drugs, suggested recrudescence, with a frequency of 4% (1/24). The remaining 23 patients may have been reinfected, with a frequency of 96% (23/24).Two allele genes  . Some researchers  [Grant Number 2018ZX09303008] to CD and QW, Project of Traditional Chinese Medicine Bureau of Guangdong [Grant Number 2019(43)] to JS and QW, and YangFan Innovative And Entrepreneurial Research Team Project[Grant Number 2014YT02S008] to CD and QW.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Accurate measurement of anti-malarial drug concentrations in therapeutic efficacy studies is essential to distinguish between inadequate drug exposure and anti-malarial drug resistance, and to inform optimal anti-malarial dosing in key target population groups.A sensitive and selective LC\u2013MS/MS method was developed and validated for the simultaneous determination of amodiaquine and its active metabolite, desethylamodiaquine, and used to describe their pharmacokinetic parameters in Ghanaian patients with uncomplicated falciparum malaria treated with the fixed-dose combination, artesunate-amodiaquine.0-\u221e) was significantly higher in infants (4201\u00a0ng\u00a0h/mL) and children aged 1\u20135\u00a0years (1994\u00a0ng\u00a0h/mL) compared to older children and adults , even though infants received a lower mg/kg amodiaquine dose (median 25.3 versus 33.8\u00a0mg/kg in older patients). Desethylamodiaquine AUC0-\u221e was not significantly associated with age. No significant safety concerns were identified.The day-28 genotype-adjusted adequate clinical and parasitological response rate in 308 patients studied was\u2009>\u200997% by both intention-to-treat and per-protocol analysis. After excluding 64 patients with quantifiable amodiaquine concentrations pre-treatment and 17 with too few quantifiable concentrations, the pharmacokinetic analysis included 227 patients . Increased median day-3 amodiaquine concentrations were associated with a lower risk of treatment failure . Amodiaquine exposure (median AUCEfficacy of artesunate-amodiaquine at currently recommended dosage regimens was high across all age groups. Reassuringly, amodiaquine and desethylamodiaquine exposure was not reduced in underweight-for-age young children or those with high parasitaemia, two of the most vulnerable target populations. A larger pharmacokinetic study with close monitoring of safety, including full blood counts and liver function tests, is needed to confirm the higher amodiaquine exposure in infants, understand any safety implications and assess whether dose optimization in this vulnerable, understudied population is needed. Plasmodium falciparum malaria, which has contributed to significant decreases in the malaria burden  and by per protocol (PP) analysis was 95.2% (280/294) . The overall day 28 PCR-adjusted ACPR rate was 98.0% (299/305)  by ITT and 98.0%  and those aged 5\u00a0years or older (98.3%), p\u2009=\u20090.8  patients, many of which were consistent with features of malaria. All adverse events occurring within 28-days of\u00a0follow up\u00a0(Table 0-\u221e) was 1318  ng\u00a0h/mL overall. Infants had a higher median (IQR) AUC0-\u221e of 4,201  ng\u00a0h/mL, when compared to 1994  ng\u00a0h/mL in children aged 1\u20134\u00a0years and 875 (400\u20131434) ng\u00a0h/mL in those aged\u2009\u2265\u20095\u00a0years (p\u2009<\u20090.001). Overall, the median maximum amodiaquine concentration (Cmax) was 18.8 (IQR 9.8\u201350.7) ng/mL, which was reached (Tmax) in a median of 2 (IQR 1\u20133) days. The median observed Cmax also decreased with age, from 49.6 (21.7\u2013107.0) ng/mL in infants, to 27.7 (14.7\u201370.0) ng/mL in children aged 1\u20134\u00a0years and 11.6 (6.7\u201318.8) ng/mL in older children and adults (p\u2009<\u20090.001). Median apparent clearance was 25.9 (IQR 8.6\u201356.0)\u00a0L/kg/h overall, which increased with age, p\u2009<\u20090.001. The effects of age on amodiaquine AUC0-\u221e and on Cmax remained statistically significant when the two study sites were analysed separately, and the effect of age on apparent clearance was significant among the 198 patients enrolled in the Navrongo site, but not the smaller sample of 25 patients enrolled in the Kintampo site  5.42 (95% CI 1.20\u201324.57), p\u2009=\u20090.029] and a trend towards a higher Cmax  in infants compared to those aged 5\u00a0years and older. There was a 4% increase in AUC0-\u221e, , and a 3% increase in Cmax  for each 1\u00a0mg/kg increase in total dose administered. Fever at enrolment was associated with a significant increase in AUC0-\u221e , and Cmax  with a lower apparent clearance . There was an 84% lower AUC0-\u221e  and an 82% lower Cmax  in patients from Navrongo compared to patients from Kintampo. This is consistent with the almost sevenfold increase in apparent clearance  and fourfold increase in the apparent volume of distribution  in patients from Navrongo compared to Kintampo. Given that the elimination half-life was similar between the study sites, the differences observed by site could reflect differences in bioavailability.After adjusting for pre-defined covariates Table , there w0-\u221e of desethylamodiaquine were much larger than those for amodiaquine. The desethylamodiaquine terminal elimination half-life of 196.4\u00a0h was four-fold longer than that for amodiaquine. Maximum desethylamodiaquine concentrations increased with age (p\u2009=\u20090.046), with a trend towards AUC0-\u221e (p\u2009=\u20090.057) decreasing with age and apparent clearance increasing with age (p\u2009=\u20090.071). There was a strong linear correlation between the day 7 desethylamodiaquine concentrations and the AUC0-\u221e; rs\u2009=\u20090.881, p\u2009<\u20090.001 , p\u2009=\u20090.041], which is substantial given the bodyweight adjusted total dose range between 23 and 45\u00a0mg/kg. Although high baseline parasitaemia was associated with a 40% reduction in the elimination half-life compared with patients with parasite densities\u2009<\u2009100,000/\u00b5L , no other pharmacokinetic parameters were affected by baseline parasite density . The median amodiaquine concentration on day 3 was significantly lower in patients with parasite recurrence than in those with an ACPR [4.8 (IQR 3.0\u20138.2) vs. 12.5 (IQR 5.9\u201324.2) ng/mL; p\u2009=\u20090.002, Fig.\u00a0P. falciparum malaria burden is carried by children under 5\u00a0years of age [Amodiaquine is rapidly and extensively metabolized to desethylamodiaquine, its main active metabolite that has a much longer elimination half-life than amodiaquine and generally provides most of its anti-malarial effect. This study demonstrates that desethylamodiaquine exposure in whole blood is remarkably consistent across all age groups, which is reassuring since the highest s of age . This iss of age \u201339. EquaThe inclusion of infants with uncomplicated malaria in this study provides preliminary evidence that they may have greater exposure to amodiaquine than older children and adults. There was a significant decrease in total amodiaquine exposure and maximum concentration with age, even when study sites were analysed separately Table . After aAge-dependent changes in body composition or maturational effects may contribute to the pharmacokinetic differences by age group , and amo0-\u221e in this study is similar to those obtained in whole blood spots from malaria patients aged 1\u201310\u00a0years in Papua New Guinea [0-\u221e in this study was substantially higher than the few published amodiaquine AUC0-\u221e values reported, all of which were measured in plasma [Although there was a trend towards total desethylamodiaquine\u00a0exposure and apparent clearance decreasing with age, its\u00a0pharmacokinetic parameters did not differ significantly between age categories after adjusting for pre-defined covariates. The median capillary whole blood desethylamodiaquine AUCw Guinea , but abow Guinea . Similarn plasma , 29, 46.n plasma \u201326,\u00a0and n plasma , 47, 48.No safety concerns were identified in this study, with all adverse events mild to moderate and many consistent with malaria features. Although safety was assessed clinically at all scheduled and unscheduled study visits, it was not feasible to perform laboratory tests to monitor for neutropenia or hepatotoxicity, which have been associated with amodiaquine use particularly when taken as chemoprophylaxis or with interacting medicines , 49, 50.P. falciparum in Africa [In 2005, Ghana replaced chloroquine with the artemisinin-based combination treatment, artesunate-amodiaquine, given as loose tablets or co-blister packs for the treatment of uncomplicated malaria . This stn Africa \u201357.. [All patients in this study had quantifiable concentrations of desethylamodiaquine on day 28, showing that longer follow up would be needed to detect all treatment failures, as recrudescent parasitaemia may be suppressed by these quantifiable drug concentrations , 30, 54.. , amodiaq. , 48, 59.Assessments of amodiaquine/desethylamodiaquine concentrations should be included in these therapeutic efficacy studies wherever possible, at least on day 7 and before starting treatment . In holoIn sub-Saharan Africa, malaria and malnutrition often co-exist and are important public health conditions. It is estimated that 1 in 3 children under 5\u00a0years of age in sub-Saharan Africa suffer from malnutrition . MalnutrStudy site appeared to be an important factor affecting the pharmacokinetic parameters of amodiaquine, despite adjustments for other predefined covariates. However, the pharmacokinetic properties of the active metabolite responsible for most anti-malarial activity, desethylamodiaquine were similar between sites. While the exact reason for the disparities observed by site of sample collection is not known, these differences may be consistent with higher amodiaquine bioavailability in Kintampo than Navrongo. Although it is possible that more patients Kintampo had non-protocol high-fat intake before dosing, this alone would not fully explain the apparent site effects. Compared with the fasted state in healthy adult volunteers, a high fat meal given before dosing had relatively modest effects, delaying absorption slightly and increasing both Cmax (by 22% for amodiaquine and 21% for desethylamodiaquine) and AUC (by 59% for amodiaquine and 13% for desethylamodiaquine) . It has A simple liquid\u2013liquid extraction method coupled with LC\u2013MS/MS detection that uses a small volume of whole blood (20\u00a0\u00b5L) was developed and fully validated in this study for the simultaneous determination of amodiaquine and its active metabolite, desethylamodiaquine. The method achieved lower limits of detection relative to the small sample volume, with good sensitivity and reproducibility. Small sample volumes may be necessary for field studies to ease sample collection, particularly from infants and small children. This method can readily be used for pharmacokinetic studies and therapeutic drug monitoring in patients and at-risk groups, including children to whom amodiaquine based combinations are given for the treatment or prevention of falciparum malaria.This large study of the pharmacokinetic properties of amodiaquine when used with artesunate for the treatment of uncomplicated malaria provides reassuring evidence of high cure rates with desethylamodiaquine exposure remarkably consistent across all age groups, including in underweight-for-age children and those with hyperparasitaemia. The inclusion of infants with uncomplicated malaria in this study provides preliminary evidence that they may have greater exposure to amodiaquine than older children and adults. Although no safety concerns were identified in this study, there is the potential for more adverse events in infants given their higher amodiaquine exposure and observed maximum concentrations (Cmax), particularly with increasing deployment of seasonal malaria chemoprophylaxis (SMC) in the Sahel region where 81 million courses of SP-amodiaquine were delivered in 2018 . In addiAdditional file 1. Liquid chromatography tandem mass spectrometry assay methods.Additional file 2. Pharmacokinetic parameters  of amodiaquine and desethylamodiaquine following artesunate-amodiaquine fixed-dose combination treatment of Ghanaian patients with uncomplicated P. falciparum malaria, by study site and age category."}
+{"text": "Favipiravir is an oral broad-spectrum inhibitor of viral RNA-dependent RNA polymerase that is approved for treatment of influenza in Japan. We conducted a prospective, randomized, open-label, multicenter trial of favipiravir for the treatment of COVID-19 at 25 hospitals across Japan. Eligible patients were adolescents and adults admitted with COVID-19 who were asymptomatic or mildly ill and had an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1. Patients were randomly assigned at a 1:1 ratio to early or late favipiravir therapy . Favipiravir is an oral broad-spectrum inhibitor of viral RNA-dependent RNA polymerase that is approved for treatment of influenza in Japan. We conducted a prospective, randomized, open-label, multicenter trial of favipiravir for the treatment of COVID-19 at 25 hospitals across Japan. Eligible patients were adolescents and adults admitted with COVID-19 who were asymptomatic or mildly ill and had an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1. Patients were randomly assigned at a 1:1 ratio to early or late favipiravir therapy . The primary endpoint was viral clearance by day 6. The secondary endpoint was change in viral load by day 6. Exploratory endpoints included time to defervescence and resolution of symptoms. Eighty-nine patients were enrolled, of whom 69 were virologically evaluable. Viral clearance occurred within 6\u2009days in 66.7% and 56.1% of the early and late treatment groups . Of 30 patients who had a fever (\u226537.5\u00b0C) on day 1, times to defervescence were 2.1\u2009days and 3.2\u2009days in the early and late treatment groups . During therapy, 84.1% developed transient hyperuricemia. Favipiravir did not significantly improve viral clearance as measured by reverse transcription-PCR (RT-PCR) by day 6 but was associated with numerical reduction in time to defervescence. Neither disease progression nor death occurred in any of the patients in either treatment group during the 28-day participation.  The new coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread throughout the globe since late 2019 and caused an unprecedented pandemic. The virus has infected at least 28 million people and caused over 900,000 deaths due to COVID-19. While the majority of those infected with SARS-CoV-2 develop mild, self-limiting disease, with even some remaining fully asymptomatic throughout the course, some patients progress to severe pneumonia, multiorgan failure, and death (in vivo by its triphosphorylated derivative (T-705RTP), which translates to broad-spectrum inhibition of RNA viruses  population. The infected ITT population, defined for the primary outcome analysis of the viral clearance, consisted of 36 and 33 patients in the early and late treatment groups after excluding 8 and 11 patients whose reverse transcription-PCR (RT-PCR) result on the first day was already negative. Finally, the safety population included 44 patients in the early treatment group and 38 patients in the late treatment group after excluding 7 patients who did not take any favipiravir dose . The dayn\u2009=\u200988). A total of 32 patients (36.4%) had a body temperature of 37.5\u00b0C or greater at enrollment , and 54 (61.4%) patients were men in the ITT population .In the infected ITT population, the likelihood of viral clearance by day 6 was not significantly different between the early treatment and late treatment groups . Viral cThe proportions of patients with 50% logarithmic reduction in the SARS-CoV-2 viral load by day 6 in the infected ITT population were 94.4% and 78.8%, respectively . The change in viral load in the infected ITT population was more negative in the early treatment group than the late treatment group, but the difference was not statistically significant .Mean times to defervescence defined by the highest temperature of a given day being below 37.5\u00b0C were 2.1\u2009days  in the early treatment group and 3.2\u2009days  in the late treatment group  in the ITT population . Mean ti4.3 copies/ml versus 107.7 copies/ml). The early treatment group was also more likely to achieve viral clearance than the late treatment group for patients who were randomized \u22654\u2009days (median) after the first positive RT-PCR .Prespecified subgroups for the primary outcome analysis included age of \u226565 years, viral load on the first day, time from the first positive RT-PCR, and presence of any symptom on the first day. Viral clearance by the sixth day was significantly more likely to occur in the early treatment group than the late treatment group for patients with age \u226565 (8/9 [88.9%] versus 0/5 [0%]). However, there was an imbalance in the initial viral load between the two groups despite stratified randomization . Of 32 patients who had serum uric acid level determined on the 16th or 28th day (early treatment group) or on the 28th day (late treatment group), 24 patients had the level normalized to below 7\u2009mg/dl, with the highest being 8.8\u2009mg/dl. Other reported adverse events included serum triglyceride elevation (9/82 [11.0%]) and serum alanine aminotransferase elevation (7/82 [8.5%]).P value computed by the generalized Wilcoxon test indicated statistical significance  targeting SARS-CoV-2 for molecular diagnosis of COVID-19 in Japan . The N2 8\u2013The most common adverse event associated with favipiravir is hyperuricemia (4.79% according to the package insert), which is associated with inhibition of OAT1, OAT3, and URAT1 by this agent . The incSeveral limitations are notable in our study. First, the sample size of this study was based on an earlier report of a nonrandomized study from China, which reported highly significant viral clearance rates in the first 5 days of therapy with favipiravir plus interferon alpha compared with lopinavir-ritonavir plus interferon alpha. The effect size in our randomized setting was smaller, which along with an unexpected high frequency of a negative RT-PCR at the time of enrollment likely underpowered the study. Second, the open-label study design may have biased assessment and management outside the trial. Third, the staggered treatment design where all patients eventually received favipiravir, adopted due to the unavailability of placebo at the time of study conception, made it difficult to interpret outcome differences beyond the sixth day. Fourth, the study only recruited asymptomatic to mildly symptomatic COVID-19 patients, and the findings cannot be extrapolated to patients with moderate to severe disease. Enrolling only patients without symptoms or with mild disease also made it difficult to assess symptoms, possibly underestimating potential clinical benefits of favipiravir. Finally, virological endpoints were assessed by RT-PCR only, and it is not known whether early treatment had any impact on replication-competent viruses.In summary, in this randomized trial of patients with asymptomatic to mildly symptomatic COVID-19, administration of favipiravir did not significantly improve viral clearance in the first 6 days, but there was a trend toward earlier viral clearance with the agent. Favipiravir was associated with numerical reduction in time to defervescence, and a significant improvement in fever was observed the day after starting therapy, compared with findings with no therapy. None of the patients experienced progression of disease or death. While limited by the small sample size, the findings suggest antiviral activity of favipiravir in this patient population. Further studies are required to demonstrate whether this effect translates to prevention of disease progression and mortality.This was an investigator-initiated, individually randomized, open-label trial to assess the efficacy and safety of oral favipiravir in adolescents and adults (aged \u226516\u2009years) admitted to hospital with asymptomatic to mildly symptomatic COVID-19.The study was centrally approved by the certified review board of Fujita Health University, which served as the coordinating center, and subsequently approved by the director of each participating hospital prior to site initiation. Written informed consent was obtained from all study participants. The trial protocol is available in the supplemental material.Patients were recruited at 25 hospitals across Japan. The study recruitment period was from 2 March 2020 to 18 May 2020. The follow-up was completed on 14 June 2020.Inclusion criteria were the following: (i) age of 16 years or older, (ii) inpatient status, (iii) positive RT-PCR for SARS-CoV-2 from a pharyngeal or nasopharyngeal swab specimen collected within 14\u2009days, (iv) Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1 , (v) abiExclusion criteria were the following: (i) performance status of 2 or greater, (ii) severe hepatic disease, (iii) need for dialysis, (iv) altered mental status, (v) pregnancy, (vi) female patients who did not agree to use effective contraceptive methods, (vii) male patients with female partners who did not agree to the use of effective contraceptive methods, (viii) hereditary xanthinuria, (ix) hypouricemia or history of xanthine urolithiasis, (x) uncontrolled gout or hyperuricemia, (xi) immunosuppressive conditions, and (xii) receipt of systemic antiviral agent against SARS-CoV-2 within 28\u2009days.Potential study participants were screened for eligibility within 24\u2009h prior to the study randomization. Patients were randomly assigned via computer-generated random numbering (1:1) to start favipiravir either on day 1 (early treatment group) or on day 6 of study participation (late treatment group). An open-label design was adopted, as placebo was not readily available at the time of study initiation. Treatment was staggered by 5\u2009days between the two groups to allow for a sufficient window to directly compare effects of treatment versus no treatment while keeping the delay within a clinically reasonable period in the late treatment group. The randomization was stratified based on age (\u226565 or <65) and days between collection of the SARS-CoV-2-positive specimen and enrollment (<8\u2009days and \u22658\u2009days). Patients and clinicians were not masked to treatment assignment.Favipiravir was dosed at 1,800\u2009mg twice orally at least 4 h apart on the first day, followed by 800\u2009mg orally twice a day, for a total of up to 19 doses over 10\u2009days. This regimen achieves plasma concentration of approximately 60\u2009\u03bcg/ml and higher in healthy individuals . If the patients met the discharge criteria  which were sanctioned by the government during the study period, and they had reached at least the sixth day of study participation, they were allowed to discontinue favipiravir, be discharged home, and followed up at the end of the study visit. Use of other medications with antiviral activity was prohibited during the course of study participation.Nasopharyngeal swabs were collected daily between day 1 and day 6 and then every other day through day 16 if the patients remained in hospital. RT-PCR was conducted at a centralized study laboratory using the protocol that was developed at the National Institute of Infectious Diseases and widely adopted in Japan . BrieflyThe primary outcome endpoint was time to SARS-CoV-2 clearance and presence or absence of SARS-CoV-2 clearance by RT-PCR of nasopharyngeal specimens by day 6. For the late treatment group, the specimen was collected prior to initiation of favipiravir on day 6.Secondary outcome endpoints were as follows: (i) SARS-CoV-2 clearance by day 10, (ii) 50% logarithmic reduction in the SARS-CoV-2 viral load by day 6, (iii) changes in logarithmic SARS-CoV-2 viral load, and (iv) time until SARS-CoV-2 clearance by local RT-PCR. Exploratory outcome endpoints were as follows: (i) duration of fever (\u226537.5\u00b0C or \u226537.0\u00b0C), (ii) change in body temperature, (iii) change in symptoms, assessed by the number of symptoms, including subjective fever, cough, sore throat, headache, myalgia or arthralgia, chills or diaphoresis, fatigue, disturbance of consciousness, shortness of breath, rhinorrhea, chest pain, diarrhea, and nausea or vomiting, (iv) disease progression or death, and (v) death.The sample size was calculated upon the assumption that the virus would be undetectable by day 6 in 80% of the subjects in the early treatment group and 50% of the subjects in the late treatment group, based on a prior study  was 50 or greater, which was defined as [/] \u00d7 100 where the limit of detection was set to 120 copies/ml. Logistic regression was used to compute the odds ratio of the 50% logarithmic viral load reduction.To assess changes in viral load, body temperature, and symptoms, intergroup comparisons were conducted with the repeated measures regression model using the mixed-effect (MIXED) model. In order to assess change from the baseline in the model with the logarithmic viral load as an outcome variable, the baseline logarithmic viral load was further adjusted in the model. Similarly, Cox proportional hazard regression was used to compare time to defervescence, which was defined as the day body temperature became less than 37.5\u00b0C among patients whose body temperature was greater than 37.5\u00b0C at baseline. To compare average change in body temperature from the baseline, the mixed-effect regression model was used where the body temperature at each time point was the outcome variable, and the regression model further adjusted for the baseline body temperature. Change in symptom score was analyzed using the generalized estimation equation (GEE) regression. All regression analyses mentioned above were adjusted for age and days between collection of the SARS-CoV-2-positive specimen and enrollment.a priori-decided exploratory subgroup analyses, subgroups were generated based on the initial SARS-CoV-2 viral load, presence or absence of symptoms, age of \u226565 and <65, and days from the first positive RT-PCR result prior to study enrollment to examine if efficacy was observed in these subgroups. Cox proportional hazard regression models were used in the subgroup analysis without covariate adjustment due to small sample size. Statistical analyses were performed with SAS software, version 9.4 .As"}
+{"text": "More sensitive methods such as qPCR might better characterize the clearance phenotype in sub-Saharan Africa.PCT is explored in Mali using light microscopy and qPCR after artesunate for uncomplicated malaria. In two villages, patients were followed for 28 days. Blood smears and spots were collected respectively for microscopy and qPCR. Parasitemia slope half-life was calculated after microscopy. Patient residual parasitemia were measured by qPCR.Uncorrected adequate clinical and parasitological responses (ACPR) observed in Faladje and Bougoula-Hameau were 78% and 92%, respectively (p\u00a0=\u00a00.01). This reached 100% for both after molecular correction.Proportions of 24H microscopy positive patients in Faladje and Bougoula-Hameau were 97.2% and 72%, respectively (p\u00a0<\u00a00.0001).Slope half-life was 2.8\u00a0h in Faladje vs 2H in Bougoula-Hameau (p\u00a0<\u00a00.001) andProportions of 72H patients with residual parasitemia were 68.5% and 40% in Faladje and Bougoula-Hameau, respectively (p\u00a0=\u00a00.003). The mean residual parasitemia was 2.9 in Faladje vs. 0.008 in Bougoula-Hameau (p\u00a0=\u00a00.002).P. falciparum resistance to artemisinins.Although artesunate is efficacious in Mali, the longer parasite clearance time with submicroscopic parasitemia observed may represent early signs of developing  Artemisinin-based Combination Therapies (ACT) are first line treatment for malaria case management . Combiniin vivo was reported in South-east Asia and defined as a delay in clearance of parasite as measured by light microscopy  in SE-Asia . Malaria transmission in Faladje is intense between July and October . BougoulThe trial enrolled patients of 6 months of age and older in both Faladje and Bougoula-Hameau.P. falciparum asexual forms per microliter, hemoglobin levels greater than 8.0\u00a0g/dl and no declared allergy to artemisinins. Volunteers with other acute illnesses or those with severe/complicated malaria were not included in this study. Participants were included after written informed consent, assent or parental consent for minors was obtained. A treatment course of 7 days of artesunate was administered at a first single dose of 4\u00a0mg/kg at inclusion day followed by a daily single dose of 2\u00a0mg/kg for the remaining of the treatment course. Patients were actively followed for 28 days according to modified standard protocols  adapted protocols . After tDried blood spots (DBS) were obtained at inclusion and every 8\u00a0hours until three consecutive negative slides were obtained, then at days 7, 14, 21, 28 and/or at unscheduled days of clinical visit.msp1, msp2 and Ca1, as described elsewhere , Late Clinical Failure (LCF), Late Parasitological Failure (LPF) and Adequate Clinical and Parasitological Response (ACPR) . Treatme2.2Light microscopy: the well-established parasite clearance time parameter was determined as the time from the first positive blood slide at inclusion to the time of the first negative slide followed by two consecutives negative slides. The WWARN parasite clearance estimator (PCE) was used to calculate parasite clearance slope half-life and the clearance rate constant . Other pPlasmodium tRNA methionine (pgMET) and a human gene coding for the human \u03b2 tubulin (HumTuBB). From the initial parasitemia, Parasite Reduction Rate (PRR) was calculated at different time point  for each patient . DNA was extracted from DBS collected prior to treatment, and at 24, 48 and 72\u00a0hours afterwards as previously described . Parasit patient . PatientFor each time point, a mean was calculated on patient's residual parasitemia per village.2.3Pfmdr1, Pf K13, Pfdhfr and Pfdhps as well as Pfmdr2, arps10, and ferredoxin . Polymorphisms for und, PGB ) Pfcrt Kescribed . A basel2.4Data was included in the final analysis when the patient received the full dose of treatment, had not received another antimalarial treatment and attended all follow up visits. The sample size was calculated based on the in vivo clinical and parasitological failure rate around 17% on Day 28 using artemether-lumefantrine as the reference drugs in Mali . Assumin2 test, the student's t test or the Wilcoxon-Mann Whitney test as appropriate. The main parameters affecting parasite clearance such as patient age and the initial parasitemia were included in the model as confounding factors as well as sex, hemoglobin levels, gametocyte density and fever at enrolment. Data were entered in excel 2011 and STATA version 11.2 for statistical analysis. The genotyping data for each molecular marker was extracted from the MalariaGEN genetic report card based for these locations. Missing data were removed and the frequency of variations in each molecular marker was calculated using the dplyr package implemented in R Proportions, medians and means were measured by descriptive statistics and compared by the \u03c7The study protocol was reviewed and approved by the Ethics Committee of the Faculty of Pharmacy and the Faculty of Medicine and odonto-stomatology, University of Sciences Technics, Technologies of Bamako, Mali.33.1Overall 221 volunteers were included in this study, 121 from Faladje and 100 from Bougoula-Hameau. One exclusion case for persistent vomiting after drug administration was observed in Faladje. No other losses to follow up happened for the two study sites . At enroThere were 2 LCF and 24 LPF cases (78.3% ACPR) registered for Faladje versus 8 LPF (92% ACPR) for Bougoula-Hameau (p\u00a0=\u00a00.01). After PCR correction, all recurrent infections were new infections, bringing the artesunate PCR corrected cure rate to 100% in both villages.3.2At eight hours after treatment initiation, only 1.7% of Faladje patients had an increase in the baseline parasitemia while in Bougoula-Hameau 39% of patients had an increase in the baseline parasitemia (p\u00a0<\u00a00.0001) . HoweverUsing the WWARN PCE tool, the parasite clearance slope half-life was significantly longer in Faladje with a median of 2.8\u00a0hours than in Bougoula-Hameau with 2\u00a0hours (P\u00a0<\u00a00.0001) . There w3.3Complete qPCR results were obtained for 104  out of 111 analyzed samples.For the first 24 and 48\u00a0hours following artesunate treatment, the proportions of patients with residual parasitemia was comparable for the two study sites . At 72\u00a0h3.4PfK13 mutation was found in Bougoula-Hameau (A578S) (N\u00a0=\u00a098), while no such mutation was found in Faladje (N\u00a0=\u00a0118). The prevalence of Pfcrt K76\u00a0T, Pfmdr1 N86Y, Pfdhfr-Pfdhps quadruple mutations in Faladje and Bougoula-Hameau were comparable  was significantly longer in Faladje than in Bougoula-Hameau, using both light microscopy and qPCR. Although, the nearly 3-hour slope half-life of parasitemia clearance of Faladje did not reach the current 5-hour threshold for artemisinin resistance described in SEA , this obA number of hosts, parasite and environmental factors might affect parasite elimination from the bloodstream. Differences in malaria transmission levels could impact the PCT . In our P. falciparum MSP1-42 and P. falciparum AMA-1 antibodies were similar between the two areas where the respective villages are located . Nevertheless, prevalence of current molecular markers of antimalarial drug resistance was comparable for the two villages . Malaria Research and Capacity Development (MARCAD), a DELTAS Program [DEL-15-010] supported Aminatou Kone through a Post-doctoral fellowship. The Developing Excellence in Leadership and Genetics Training for Malaria Elimination in sub-Saharan Africa (DELGEME) a DELTAS Program [DEL-15-002] supported Aoua Coulibaly for a PhD fellowship. The DELTAS Africa Initiative is an independent funding scheme of the African Academy of Sciences (AAS)\u2019s Alliance for Accelerating Excellence in Science in Africa (AESA) and supported by the New Partnership for Africa's Development Planning and Coordinating Agency (NEPAD Agency) with funding from the Wellcome Trust [grant 107741/A/15/Z] and the UK government. The views expressed in this publication are those of the author(s) and not necessarily those of AAS, NEPAD Agency, Wellcome Trust or the UK government.A.K received support from the World Academia of Sciences (TWAS) for providing financial support through the grant number Ref.: 17-346 RG/BIO/AF/AC_I\u2013FR3240297741.Corresponding author: Professor Abdoulaye A. Djimde, adjimde@icermali.org, Malaria Research and Training Center, Faculty of Pharmacy, University of Science, Techniques and Technology, Bamako, MaliPr. Djimde has nothing to disclose."}
+{"text": "Plasmodium falciparum infections following household mass administration. Adherence information was reported for 181,534 of 336,821 DHAp (53.9%) treatments administered during four rounds of MDA/fMDA, of which 153,197 (84.4%) reported completing the full course of DHAp. The proportion of participants fully adhering to the treatment regimen differed by MDA modality (MDA versus fMDA), RDT status, and whether the first dose was observed by those administering treatments. Among a subset of participants receiving DHAp and selected for longitudinal follow-up, 58 were positive for asexual-stage P. falciparum infection by microscopy at baseline. None of the 45 participants followed up at days 3 and/or 7 were slide positive for asexual-stage parasitemia. For those with longer term follow-up, one participant was positive 47 days after treatment, and two additional participants were positive after 69 days, although these two were determined to be new infections by genotyping. High completion of a 3-day course of DHAp and parasite clearance in the context of household MDA are promising as Zambia\u2019s National Malaria Programme continues to weigh appropriate interventions for malaria elimination.Mass drug administration (MDA) with artemisinin combination therapy is a potentially useful tool for malaria elimination programs, but its success depends partly on drug effectiveness and treatment coverage in the targeted population. As part of a cluster-randomized controlled trial in Southern Province, Zambia evaluating the impact of MDA and household focal MDA (fMDA) with dihydroartemisinin\u2013piperaquine (DHAp), sub-studies were conducted investigating population drug adherence rates and effectiveness of DHAp as administered in clearing  Previous studies comparing DHAp with other artemisinin-based combination therapies (ACTs) have shown DHAp to be highly efficacious at clearing asexual 16 When adherence to the full treatment regimen is incomplete, the antimalarial may not achieve or maintain therapeutic levels of the drug for a sufficient time to clear all parasites in an individual, placing selective pressure on surviving parasites.17 Low adherence rates across the treated population could fail to achieve maximum levels of effect and promote drug resistance, despite high treatment coverage in MDA.18 Studies across diverse settings in malaria and neglected tropical diseases have demonstrated wide variation in adherence during MDA activities.19 In a systematic review of adherence to malaria treatment, Bruxvoort et al.15 found adherence proportions varying as much as 11\u2013100% across 30 descriptive studies. Therefore, it is reasonable to assume that a percentage of the population will not adhere fully to an MDA treatment program.20Using a highly efficacious drug such as DHAp, MDA effectiveness is dependent on the individual\u2019s adherence to the medication regimen.2 we evaluated population acceptance and adherence to the DHAp regimen and parasite clearance during household MDA. Specifically, this study assesses population treatment adherence to DHAp under both community-wide MDA and fMDA modalities. We further examine reasons for adherence and nonadherence to treatment. Finally, to assess the effectiveness of DHAp as administered in this setting, we examine 3-day and 7-day parasite clearance among participants accepting treatment, as well as parasitemia at approximately 40 and 70 days following treatment.As Zambia\u2019s National Malaria Programme weighs the impact of MDA, additional insight into mediating issues, such as population-wide treatment adherence and the drug effectiveness in clearing parasites, would be useful for shaping expectations of MDA impact moving forward. To this end, this study presents results of analyses exploring issues of drug uptake and effectiveness. As part of the aforementioned mass treatment community-randomized controlled trial,2 In summary, the trial assessed the impact of community-wide MDA with DHAp or household-level focal MDA (fMDA), where DHAp was given to all eligible household members if anyone in the house had a positive malaria rapid diagnostic test . The trial engaged all households and residents of 60 health facility catchment areas covering a population of approximately 330,000 people in a region of heterogeneous transmission, with malaria prevalence in children ranging from < 1% to > 25% before the trial.21 The region is characterized by high coverage of vector control  and good access via health facilities and community health workers (CHWs) to confirmed malaria case management with RDTs and artemether\u2013lumefantrine (AL). The trial interventions included house-to-house campaigns of four rounds of RDT testing and DHAp treatment during both peak and nonpeak transmission seasons; rounds 1 through 4 occurred during November/December 2014 (nonpeak), February/March 2015 (peak), September/October 2015 (nonpeak), and February/March 2016 (peak), respectively.A full description of the trial has been published elsewhere.During each campaign round, a pair of trained surveyors visited each household: a CHW who performed the testing and treatment for malaria and an enumerator who administered a standardized questionnaire to the head of the household or parental representative. Dihydroartemisinin\u2013piperaquine formulations of 20 mg/160 mg and 40 mg/320 mg were provided per treatment modality to participants based on age according to the manufacturer\u2019s recommendations. Pregnant women and children younger than 3 months were excluded from receiving DHAp. The full course of a single treatment was taken over three consecutive days and monitored using a modified directly observed therapy (DOT) strategy. On day 0 (baseline), the first dose of medication was administered and observed by the CHW and noted by the enumerator. The recipient was instructed to take the second dose at the same time the following day (day 1) but was unobserved. On day 2, a separate adherence officer visited the household to observe the third  dose, if possible, and to administer a follow-up questionnaire, irrespective of observing the final dose.Initial questionnaires conducting during day 0 household visits recorded participant demographics, recent history of fever, household IRS exposure, recent travel, and bednet usage. At the end of each day, enumerators transferred name, age, gender, and a personal identifier for individuals who received DHAp to the adherence officer, blinded to testing status of individuals, for follow-up. The adherence questionnaire conducted on day 2 sought to ascertain the recipient\u2019s testing and treatment recall, whether the dose was taken on the correct day, the number of tablets taken daily, reasons for not taking all doses, reasons for treatment refusal, and whether the recipient had heard MDA sensitization messages before the campaign visit. Visual inspection of the blister pack noted the number of remaining tablets, if any. These data were self-reported by the participants if aged 10 or older, or by the caregiver for those 9 years and younger. The person responding to the questions for the listed DHAp recipient was also recorded.Assessment of treatment adherence used aggregated data from the four treatment rounds. An individual\u2019s adherence status for a single course of drugs was classified into three categories: full adherence, partial adherence, and nonadherence. Full adherence was defined as taking all three doses and verifying that no tablets were remaining in the blister pack. Partial adherence was considered as taking one or two doses and/or having tablets remaining in the blister pack. Nonadherence was reporting having taken no doses. Although many people within the MDA treatment arm received up to four separate courses of DHAp over the study period, an individual\u2019s adherence to treatments over multiple rounds was not assessed because it was not possible to link participants across treatment rounds.Overall, adherence was compared across treatment arms by examining the proportion of full, partial, and nonadherence against key characteristics . Reasons provided for nonadherence were assessed to determine the relative frequency of these responses. If responses coded as other were thematically similar to prelisted survey responses, these were recoded into the primary categories.Multivariable logistic regression analysis was performed to assess whether demographics and/or household characteristics were associated with reporting of treatment adherence. A similar regression analysis was performed to assess factors associated with full adherence to DHAp treatment. Factors associated individually with full adherence in crude logistic models were added to a final multilevel regression model that also included random effects for catchment and treatment round.For trial evaluation purposes, a subset of households in each of the control, MDA, and fMDA arms (260 households each) were enrolled in a concurrent longitudinal malaria incidence cohort beginning with the first treatment round in November 2014. These households were followed for 18 consecutive months\u2014a period including all four treatment rounds. Cohort-enrolled households received the mass treatment intervention that coincided with their respective study arm at the same time as all other households within the area; however, visits to administer questionnaires, provide testing and treatment, and ascertain adherence to treatment were administered by local CHWs separately from campaign round households. In addition to RDT testing provided to campaign participants, all cohort participants had thick smear duplicate slides prepared for microscopy diagnosis and confirmation. Initial day 0 household questionnaires and day 2 adherence questionnaires were otherwise similar to those administered to households within the campaign areas but not enrolled in longitudinal follow-up.As per protocol, participants in the longitudinal cohort with a positive RDT at enrollment  and receiving an age-appropriate dose of DHAp were recruited to participate in a further sub-study to assess the effectiveness of DHAp, as administered via household MDA, in clearing parasites. To reduce the number of household visits, consenting participants were asked to present at the nearest health facility 3 and 7 days following the first dose of DHAp\u2014corresponding to 1 and 4 days after completing the full course of DHAp, respectively.P. falciparum. Other species were not consistently recorded. Results were fed into Microsoft Excel  spreadsheets. Discrepant readings were resolved by a third microscopist blinded to previous readings. Thin smear slides were not prepared to reduce the total number of slides prepared in the field.At each follow-up facility visit, capillary blood was collected by finger stick and thick smear slides were prepared in duplicate. All slides were linked to respective cohort participants and sent to the National Malaria Elimination Centre for review. Two independent microscopists examined slides, specifically noting the presence and density of asexual- or sexual-stage P. falciparum infection at baseline and receiving a course of DHAp were ultimately included in analyses. Short-term parasite clearance was estimated as the percentage of individuals present at follow-up who received a course of DHAp, had a microscopically confirmed P. falciparum infection at enrollment, and were negative for asexual P. falciparum (by microscopy) 3 and 7 days after the initial dose of treatment. Similarly, longer term parasite clearance was estimated as the percentage of individuals present at follow-up negative for asexual P. falciparum (by microscopy) approximately 1 and 2 months after treatment meeting the same criteria and also not receiving further treatment with antimalarials between baseline treatment and follow-up. For participants slide positive at long-term follow-up, molecular barcode analysis was performed on samples collected at baseline and follow-up (if available), as previously described,22 to determine whether P. falciparum infection at follow-up was likely due to infection with a new parasite or recrudescence of the initial infection. The study protocol was reviewed and ethical approval provided by the Research Ethics Committee of the University of Zambia, the Zambian Medicines Regulatory Authority, the Tulane University Institutional Review Board (IRB), and Western IRB.In addition to the general exclusion criteria for receiving MDA mentioned earlier, individuals were excluded from parasite clearance assessment if questionnaires could not confirm receipt of DHAp. Following processing of blood slides, individuals with at least one thick blood smear positive for n = 1,309). Adherence teams completed follow-up visits for 181,534 (53.9%) of all 336,821 DHAp treatments. The proportion of treatments receiving follow-up visits ranged from 33.3% to 67.6% over the four rounds. There was no significant difference in adherence reporting between treatment arms overall .During the four campaign rounds, 336,821 DHAp treatments were provided to 383,768 eligible individuals, with a mean RDT positivity across the four campaign rounds of 5.1% . IndividComparing characteristics of participants with and without adherence follow-up data, in adjusted analyses, individuals who reported fever in the last 2 weeks, were RDT positive, and had heard sensitization messages, and men were more likely to have been followed up and report adherence information . Control2, P = 0.001).Among participants with adherence follow-up data, 153,197 of 181,534 (84.4%) across trial arms reported taking all three doses of DHAp , whereas 22,438 (12.4%) took only one or two doses  of the DHAp regimen . Only 5,2, P < 0.05). If the dose was taken the same day but after the CHW visit, full adherence was lower (78.0% in MDA and 86.5% in fMDA). If the treatment began any day after the initial visit, full adherence was 7.2% in MDA and 37.8% in fMDA.Adherence status also differed by DOT status during the first DHAp dose. When the first dose was directly observed by the CHW, full adherence was 83.6% in the MDA arm compared with 68.0% when it was not (Pearson\u2019s \u03c7P < 0.001).A positive RDT result, residing in a cluster receiving fMDA, younger than 18 years, reporting hearing MDA sensitization messages, taking the first dose in front of a CHW, and self-responding to the adherence questions were significantly associated with an increase in the odds being fully adherent to treatment . ConversAmong the 27,841 individuals classified as partially or completely nonadherent, 10,800 (39.3%) of these also reported reasons for their nonadherence. The primary reasons provided were as follows: 1) 8,909 (32.4%) individuals forgot, 2) 4,637 (16.8%) felt better, or 3) 2,558 (9.3%) lost medication. Only 1,369 (5.0%) reported having experienced side effects as the reason for stopping the treatment course early.P. falciparum malaria infection at the time of treatment. Questionnaires indicated that 58 (93.5%) of these confirmed cases received DHAp. Baseline treatment was not reported or confirmed by adherence follow-up for the remaining four (6.5%) individuals who were excluded. Reasons for missing treatment data were not specified by data collectors, although two of these participants were from fMDA households where all household members were negative by RDT and, therefore, likely did not receive treatment as per trial protocol.During the first treatment round, 1,567 participants from cohort households were reported as receiving a course of DHAp. Laboratory testing of 1,353 baseline thick blood smear slides identified 62 individuals with an asexual- or sexual-stage The number of days between baseline and follow-up household visits in later months varied among participants because of field-worker schedules for the cohort follow-up visits. First and second follow-up visits ranged from 21 to 63 (median: 44) days and 59 to 83 (median: 70) days after baseline, respectively. Three individuals tested positive by RDT during the first household visit and received further treatment with AL as per protocol for nonintervention months. Subsequent samples for these participants were excluded from analysis. Another participant reported having had a fever within the 2 weeks before the first follow-up visit but did not report taking an antimalarial during that period.Among those included in parasite clearance assessment at each time point, 45 of 58 (77.6%) and 39 of 58 (67.2%) presented to a local clinic 3 and 7 days after treatment for follow-up, respectively . Blood sP. falciparum parasitemia (Microscopy results noted only gametocytes present at baseline for two of 58 (3.4%) participants. Parasite densities at baseline ranged from 142 to 13,960  parasites per \u03bcL. All available samples from days 3 and 7 were clear of asexual asitemia , althoug12 were distinct between pairs of samples, indicating that these were likely new parasite infections.Earlier parasite clearance could not be confirmed for the participant slide positive at day 57, as this individual did not appear for any short-term follow-up before this time. Polymerase chain reaction (PCR) confirmed infection at day 57, although lack of a baseline PCR sample precluded comparative genotyping to determine if this was indeed persistent infection or newly acquired. As mentioned earlier, this person was treated with AL at day 57 and subsequently clear of parasites by microscopy at day 80 ,25 we examined the uptake and adherence to a 3-day drug regimen across multiple treatment rounds and explored factors associated with higher or lower levels of adherence. We also assessed the effectiveness of DHAp as administered in household MDA/fMDA for both short-term and durable clearance of P. falciparum infection. Overall, our evidence suggests acceptance and adherence to the DHAp regimen were high and maintained at these levels across all four mass treatment rounds, and that the drug was highly effective for short-term clearance of asexual parasite infection.The effectiveness of an infection control intervention is a function of whether the treatment is efficacious against the infection, the intervention reaches the target group, its users and providers adhere to intervention guidelines, and a high level of coverage is sustained over the necessary interval of time.Adherence to a full course of DHAp observed across campaign rounds was high, with more than eight in 10 (84.4%) participants taking the full course of DHAp. Individuals in the fMDA areas, where treatment was provided to all members of a household when at least one person tested positive, were more than twice as likely to fully adhere as compared with those in the MDA arm. This difference was consistent across all four campaign rounds, irrespective of individual RDT status. These results could suggest that individuals in the fMDA trial arm recognized that if their household qualified for treatment they were at increased risk for malaria when they themselves were negative or sought to clear their infection when positive, whereas in MDA, where all individuals received treatment regardless of RDT positivity, full adherence was lower but constant across four rounds of MDA with declining RDT positivity in each successive round.26 in a modeling study of the optimal use of AL and DHAp in programmatic settings, which estimated parasite clearance rates at partial courses of DHAp. The predicted effects of taking all doses, missing the second dose, missing the third dose, and missing the second and third doses of DHAp had 91%, 70%, 71%, and 28% parasite clearance rates, respectively, for age-based dosing, but delaying a dose did not alter the treatment outcome.26 In areas where parasite transmission is low, such as the MDA trial arm in this study, the effect of not taking all doses may be attenuated in the context of a robust malaria control program with high vector control coverage for attaining elimination.27 Given that few participants eligible for clearance assessment reported incomplete adherence, this study did not have the power to produce clearance rate estimates for each category of adherence and, therefore, was not able support or contest the position of these modeling studies. Notwithstanding this limitation, efforts to improve full treatment adherence in MDA settings through improved community and individual sensitization of the parasite clearance and prophylactic benefits are necessary.Of particular note, among campaign respondents in either of the MDA or fMDA trial arms with reported adherence data, 96.8% reported taking at least one dose of DHAp. The importance of taking all doses was emphasized by Hodel et al.Pf parasites after the initial MDA/fMDA round. DHAp contains two antimalarial drugs with differing elimination half-lives, and our assessment of parasite clearance considered both short- and long-term clearance, particularly important for longer lasting drugs.28 Among individuals with thick blood smear slides positive for P. falciparum infection and treated with an age-appropriate course of DHAp, all who were present for follow-up at day 3 and/or day 7 were free of asexual parasitemia. Two individuals (4.3%) visited 69 days following treatment were positive by microscopy. One individual was positive at day 57, although it was unclear whether this was due to late treatment failure or a new infection. These results are consistent with high clearance rates previously reported in sub-Saharan Africa.30Integration of a parallel longitudinal cohort and early follow-up posttreatment allowed us to further assess clearance of 13 Distinguishing reinfection from recrudescence via genotyping has its limitations, however, particularly in higher transmission areas where infection with multiple P. falciparum genotypes are more likely.28 In cases of multiple infections before treatment, standard msp-1, msp-2, and glurp genotyping may miss genotypes present at baseline and misclassify the case as a reinfection at follow-up.32 The genome-wide SNP-based method used here aimed to provide better detection of mixed infections,22 with clear genotyping differences between alleles consistent with distinct infections. Genotyping findings were also supported by negative thick blood smears observed in the interim between baseline treatment and follow-up at 69 days. Both cases reported taking the full course of DHAp at baseline and occurred in higher transmission clusters where DHAp was only provided to households with a member testing positive by RDT . fMDA may have left local infections untreated if undetected by RDTs during household visits, increasing the risk of reinfection for these individuals.Genotyping indicated that the two late follow-up infections were likely newly acquired infections. The timing of these infections, between months 1 and 2 following treatment, is consistent with the waning prophylactic effect reported elsewhere.20 Here, we relied on self-reported histories (which is the most common assessment method), and to mitigate potential recall bias, we verified pill counts to ascertain physical proof of adherence. This too is prone to error because the absence of remaining medication is not confirmation that it was taken or taken correctly.15 However, there is limited evidence to suggest that the reporting is unreliable, and studies of adherence and accuracy of recall suggest that individuals do accurately recall their treatment during MDA scenarios.33Although adherence data were collected for slightly more than half of the treatments provided, differences in demographic data between respondents were minor; consequently, it is believed that the adherence reporting and results herein provide a relatively unbiased estimate of adherence for the study area for the trial. Assessing individual adherence to medication that relies on self-reported histories may be prone to recall and social desirability bias. Evidence here suggested that individuals who were reporting on their own behavior were twice as likely to report full adherence compared with those who were not present and whose adherence was reported by another person. Furthermore, there are differences in how studies define full adherence.34 Inclusion of high transmission areas meant the possibility for a large number of potentially eligible participants. To balance the feasibility of collecting multiple samples across a rural area within a short period of time, recommended blood sample collection at 1, 2, and 14 days after treatment was dropped. As a result, the study was neither able to assess differences in clearance before day 3 nor identify late treatment failures before 1 month.Challenges with conducting longitudinal follow-up across a large rural population limited availability of data and the ability to replicate the conditions of strict efficacy studies. The number of follow-up time points was fewer than those recommended in guidelines for efficacy studies.P. falciparum parasitemia, as these infections are the most prevalent in this region. Although Plasmodium malariae, Plasmodium ovale, and Plasmodium vivax have been observed in this region, 97.5% of infections within this population during this time period were with P. falciparum.35Recruitment of participants based on RDT results conducted during household visits affected availability of short-term follow-up data. Because microscopy results, needed to determine eligibility, were not available at the time of baseline household visits, individuals were invited to participate on RDT results alone and excluded those who were negative by RDT but later found to be positive by microscopy. Consequently, a relatively high number of slide-positive cohort individuals did not have follow-up results at days 3 and 7. This also resulted in differences in the proportion of participants in the clearance study available for assessment of short-term and long-term parasite clearance, as long-term follow-up used samples collected during monthly longitudinal cohort household visits. There was no evidence of statistically significant differences in demographic characteristics or study arm participation throughout follow-up, however. Additionally, the study only assessed clearance of It is possible that long-term clearance follow-up participants sought additional treatment for malaria outside of the MDA campaign activities between baseline and follow-up visits. Data collectors recorded any treatments given during their visits and asked about recent history of fever and malaria treatment. The three participants receiving treatment for malaria roughly 1 month after baseline were excluded from analysis at later time points. Results reported here assume that no other malaria treatment was received during the period under analysis other than that already noted.37; however, our estimates of determinants of adherence were consistent with other MDA studies and help characterize adherent from nonadherent individuals in this trial.38Despite challenges with conducting household treatment and adherence assessment over a large rural population, we were able to investigate components of mass treatment important to the overall impact of MDA conducting in an area of heterogeneous malaria transmission. The results reinforce the importance of community sensitization and mobilization before an MDA campaign, as well as directly observing the first dose. If testing is performed before drug administration, it appears that the household members will be significantly more likely to adhere to their treatment regimen if they or their family member test positive. Of note, the large sample size in our study likely led to finding statistically significant relationships with low effect estimatesP. falciparum parasite infections, with high PCR-adjusted clearance between 62 and 83 days after the DHAp treatment. Despite evidence of DHAp resistance outside of Africa, these results support the current utility of DHAp administered through MDA or fMDA rounds for effectively clearing asexual P. falciparum parasite infections in this setting.In the context of a community-randomized controlled trial in Southern Province, Zambia, community-wide MDA and fMDA with DHAp both demonstrated high treatment adherence and high and durable clearance of malaria infections. Differences in adherence between MDA and fMDA trial arms suggest that the specific strategy for deploying an MDA intervention can influence the reception of treatment. In households where testing is performed and at least one person is positive , this may be sufficient encouragement for other household members to complete a course of DHAp. In settings where no testing or selective testing is performed and no one is identified as positive , lower adherence may occur, especially where malaria risk is seen to be low. In such settings, additional efforts, such as community sensitization and full DOT, may be needed to ensure high treatment adherence. For microscopy-confirmed cases followed over time, DHAp provided a 100% short-term clearance of asexual"}
+{"text": "Plasmodium falciparum, there are usually some patients who fail treatment even in the absence of drug resistance. Treatment failures, which can be due to recrudescence or re-infection, are categorized as \u2018clinical\u2019 or \u2018parasitological\u2019 failures, the former indicating that symptoms have returned. Asymptomatic recrudescence has public health implications for continued malaria transmission and may be important for the spread of drug-resistant malaria. As the number of recrudescences in an individual trial is often low, it is difficult to assess how commonplace asymptomatic recrudescence is, and with what factors it is associated.In clinical trials of therapy for uncomplicated A systematic literature review was carried out on clinical trials of artemether-lumefantrine (AL) in patients seeking treatment for symptomatic uncomplicated falciparum malaria, and information on symptoms during treatment failure was recorded. Only treatment failures examined by polymerase chain reaction (PCR) were included, so as to exclude re-infections. A multivariable Bayesian regression model was used to explore factors potentially explaining the proportion of recrudescent infections which are symptomatic across the trials included in the study.Across 60 published trials, including 9137 malaria patients, 37.8% [95% CIs (26.6\u201349.4%)] of recrudescences were symptomatic. A positive association was found between transmission intensity and the observed proportion of recrudescences that were asymptomatic. Symptoms were more likely to return in trials that only enrolled children aged\u2009<\u200972\u00a0months . However, 84 studies had to be excluded from this analysis, as recrudescences were not specified as symptomatic or asymptomatic.P. falciparum in Africa, remains a highly efficacious drug in most endemic countries. However in the small proportion of patients where AL does not clear parasitaemia, the majority of patients do not develop symptoms again and thus would be unlikely to seek another course of treatment. This continued asymptomatic parasite carriage in patients who have been treated may have implications for drug-resistant parasites being introduced into high-transmissions settings.AL, the most widely used treatment for uncomplicated  Plasmodium falciparum, remains highly effective in Africa [Artemisinin combination therapy (ACT), the World Health Organization-recommended treatment for uncomplicated n Africa . ACT reqn Africa . Howevern Africa . Pharmacn Africa \u20138. Theren Africa , and than Africa , 11. In n Africa , 13. Somn Africa .Treatment failures have a number of undesirable consequences: most immediately for the patient concerned, who may feel better initially but become symptomatic again as the parasites which have survived treatment multiply once more. In addition, the patient may be able to infect a feeding mosquito thus contributing to malaria transmission. Furthermore, a parasite population being exposed to, but not completely cleared by, anti-malarial drugs has implications for the development of drug resistance. During the evolution and spread of drug resistance, drug-sensitive parasites coexist with more resistant parasites. Some patients have genetically distinct sub-populations of parasites within their infection, either because mutations have occurred de novo as the parasites multiplied during the current infection, or because mosquitoes have injected more than one parasite clone in a single or multiple bites. If there are any parasites with some degree of resistance to the drug within a patient\u2019s infection, they have a survival advantage over more sensitive parasites. Those which survive treatment are more likely to be transmitted to others. In this regard, an important consideration is whether patients who have failed treatment become symptomatic again. The reoccurrence of symptoms, although unpleasant for patients, may have positive implications at the community level. If a patient\u2019s recrudescent infection is treated again, the capacity for further transmission is diminished . Even ifOne can gain insight into the proportion of treatment failures that become symptomatic from clinical trials. Typically, treatment failures that are recorded in a trial are stratified into different categories. Early treatment failures (ETFs) indicate that the patient did not respond to treatment, and falls into one of 4 categories: (i) indications of severe malaria on day 1, 2 or 3 in a patient who is still parasite positive; (ii) parasitaemia on day 2 that is higher than observed on admission; (iii) parasitaemia on day 3 with axilliary temperature\u2009\u2265\u200937.5\u00a0\u00b0C; (iv) parasitaemia on day 3\u2009\u2265\u200925% of that observed on admission . Late trHere, a systematic review of clinical trials was conducted to quantify the proportion of PCR-corrected LTFs that were recorded as being symptomatic failures (symptomatic recrudescences). Intuitively, this proportion will depend on the drug treatment administered: drugs that are eliminated more slowly from the body should suppress parasite densities for a longer period of time. Therefore, the systematic review is restricted to clinical trials of a single therapy, the ACT artemether-lumefantrine (AL). As of 2017, AL is a first- or second-line treatment for uncomplicated falciparum malaria in 59 countries across the world, including 32 in Africa . SpecifiFor pre-2009 trials an existing Cochrane systematic review of clinical trials of ACT for treatment of uncomplicated malaria was used . For posP. falciparum infection, who sought treatment for symptoms of malaria.Included studies were clinical trials of AL for treatment of uncomplicated malaria which reported PCR-corrected treatment failure rates and whether these failures were clinical or parasitological . Only studies which used the standard dosing of AL were included: 6 doses taken over 3\u00a0days of tablets comprising 20\u00a0mg of artemether and 120\u00a0mg of lumefantrine. The number of tablets per dose is determined by weight, with one, 2, 3, and 4 tablets for individuals weighing 5\u201315\u00a0kg, 15\u201325\u00a0kg, 25\u201335\u00a0kg, and\u2009>\u200935\u00a0kg, respectively [Studies that did not report any recrudescences  were excluded. Studies in which multiple drug therapy was evaluated but results not sufficiently stratified by therapy were also excluded.From these studies, data for the number of subjects enrolled, the number and type of recrudescences observed, the duration of follow-up, and the location of the trial was extracted. Information on the inclusion criteria (if any were reported) for baseline parasite density in each trial was also extracted, as was the age range eligible for inclusion in each trial.Titles and abstracts were reviewed against the inclusion criteria. Any discrepancies regarding inclusion/exclusion of an article was agreed upon with the consultation of fellow reviewers.PfPR2-10) by the Malaria Atlas Project (MAP) at the time and location of each trial [www.latlong.net [Estimates were used for falciparum malaria prevalence by microscopy in children between 2 and 10\u00a0years of age  at the time and location of each trial (expressed in terms of the MAP-estimated PfPR2-10), the duration of follow-up in each trial (x2), the age range of the cohort (x3), and the minimum baseline parasite density that was required for patients to be included in the original trial (x4). For the model containing all 4 variables, the regression model is written asBayesian regression modelling was used to explore the variation observed in the proportion of recrudescences for which symptoms returned, which is denoted by PfPR2-10, expressed as a proportion, is retained in the regression models, and 3 categorical variables for x2, x3 and x4 are constructed. For duration of follow-up, the categorical variable was set to 1 for trials with a follow-up greater than 28\u00a0days and set to 0 otherwise. For age of participants, of particular interest were studies that only included young children, so x3 was set equal to 1 for trials that enrolled participants with a maximum age of 72\u00a0months or less, and equal to 0 for all other trials. For baseline parasite density, a categorical variable was set equal to 1 for trials that allowed individuals with a baseline parasitaemia of 5 parasites per \u00b5l to be enrolled, and equal to 0 otherwise. The intercept, The regression models were fitted in R using RStan , via theThe 2009 Cochrane review, artemisinin-based combination therapy for treating uncomplicated malaria , identif5 parasites per \u00b5l, whilst 53% allows individuals with a baseline parasitaemia of up to 2 \u00d7 105 parasites per \u00b5l to be enrolled. One study permitted patients with a baseline parasite density of up to 5 \u00d7 105 parasites per \u00b5l to be enrolled.Across the included studies, 14 ETFs and 398 recrudescences were recorded out of 9137 patients who were enrolled and remained in the trial until at least the first day of follow-up. This results in an overall failure rate of 3.7% (95% CI (2.9\u20134.7%), based on a random effects meta-analysis of proportion) for this population. Of the recrudescences, 162 were recorded as symptomatic recrudescences , based on a random effects meta-analysis of proportion). The number of recrudescences in each trial was low ) The majority of studies (81%) reported PCR-corrected results at one time point, with the remaining studies (19%) reporting results at 2 time points. The most common time point (81% of studies) for reporting results was 28\u00a0days after treatment commenced, although a substantial number (33%) reported results at day 42. A number of trials enrolled only young children, especially in high transmission areas. For example, 37% of the trials included here had a maximum age for enrolment of 72\u00a0months or less. Of the studies included, there were no trials that enrolled only adults. The transmission intensity at the trial location, as estimated from the MAP, varied widely with slide prevalence in 2\u201310\u00a0year olds ranging from 0.2 to 86.5%. Not all studies reported information on whether they used patient inclusion criteria based on parasite densities upon presentation. For those that did, the most frequent minimum parasite density required for inclusion in a trial was 1000  and 2000  parasites per \u00b5l. For the maximum parasite density, 17% of trials set a value of 10Among the studies that only enrolled young children (aged 72\u00a0months or less), 50 out of 111 recrudescences were symptomatic , by meta-analysis of proportion). In the remaining trials, 113 out of 287 recrudescences were symptomatic , by meta-analysis of proportion). Among studies which reported results until day 28, 104 out of 244 recrudescences were symptomatic , by meta-analysis of proportion). In the remaining studies, 59 out of 154 recrudescences were symptomatic , by meta-analysis of proportion).PfPR2-10), duration of follow-up, age range of the cohort, or whether patients with very high baseline parasitaemia were enrolled in the study  if the same trial was carried out in a location with a PfPR2-10 of 50%.The goodness of fit for each model (containing combinations of these 4 variables) was assessed, with a penalty for the number of parameters included. The leading 5 candidate models, ordered by goodness of fit, are summarized in Table\u00a0PfPR2-10 outperformed the intercept-only model (the WAIC score of the latter was 520.1).As indicated by Table\u00a0This work carried out a systematic review of clinical trials in which AL was used to treat uncomplicated falciparum malaria. Among individuals whose infections recrudesced following treatment, it was found that 37.8% (95% CIs (26.6\u201349.4%)) were classified as symptomatic recrudescences. An association was found between a higher proportion of recrudescences being symptomatic and younger age as well as lower transmission settings, consistent with lower prior exposure and immunity in these study populations. The number of recrudescences observed in an individual trial is usually very low ), which means that the results presented here could only be obtained via a pooled analysis.The lack of recurrent symptoms in the majority of patients when an infection recrudesces suggests they would be unlikely to seek further treatment despite having persistent parasitaemia. These parasites have passed through drug pressure, giving any parasites with newly emerging or spreading drug resistant mutations a survival advantage. It is not known how long these recurrent infections would last, but untreated infections are often chronic and last on average about 6\u00a0months , potentiOne interesting avenue for further work would be to repeat the analysis for another ACT. In particular, it would be interesting to see if different results were obtained for dihydroartemisinin-piperaquine, as piperaquine has a much longer half-life than lumefantrine , and migOne limitation of using this approach to estimate the proportion of recrudescences that become symptomatic is that it is restricted by the duration of follow-up in each trial. One cannot be sure if a patient would become symptomatic beyond the end of the follow-up period. Equally, an asymptomatic infection detected during follow-up and treated within the trial setting may have later developed into a symptomatic infection in a non-trial setting. Within the follow-up period, some trials will assess patients regularly , whereas other trials will assess the patients at the end of the evaluation period. In a trial in which patients are assessed more frequently, recrudescent infections will be detected earlier, which could prevent some of them from becoming symptomatic. In the regression modelling, no association was found between the duration of follow-up and the likelihood of symptoms returning after treatment failed. However, the dataset has very little temporal resolution (83% of studies had only reported results for one time point), which made it difficult to scrutinize this in any detail. Outside trial settings, AL failure rates are likely to be somewhat higher due to imperfect adherence to treatment , 33. It Not all studies provided information on the inclusion criteria for baseline parasite density in each trial. For studies that did not report it, it was assumed that any individual with uncomplicated symptoms who was positive for parasites by microscopy could be enrolled in the trial. Furthermore, the connection between the range of parasitaemia permitted in each trial and the distribution of parasitaemia measured in the cohort is not a clear one, but this was not available for all trials either.In recent within-host modelling work , 32 the In this study, by pooling together data from a large number of clinical trials, insight was gained into the likelihood of patients becoming symptomatic again following recrudescence of a falciparum malaria infection. It was found that this is more likely in younger children and in lower transmission settings.Additional file 1. Supplementary Figure 1 and details of the search strategy for the systematic review.Additional file 2. Details of the studies included in the systematic review."}
+{"text": "Malaria in pregnancy affects both the mother and the fetus. However, evidence supporting treatment guidelines for uncomplicated (including asymptomatic) falciparum malaria in pregnant women is scarce and assessed in varied ways. We did a systematic literature review and individual patient data (IPD) meta-analysis to compare the efficacy and tolerability of different artemisinin-based or quinine-based treatments for malaria in pregnant women.ClinicalTrials.gov) were searched. The final search was done on April 26, 2019. Studies that assessed PCR-corrected treatment efficacy in pregnancy with follow-up of 28 days or more were included. Investigators of identified studies were invited to share data from individual patients. The outcomes assessed included PCR-corrected efficacy, PCR-uncorrected efficacy, parasite clearance, fever clearance, gametocyte development, and acute adverse events. One-stage IPD meta-analysis using Cox and logistic regression with random-effects was done to estimate the risk factors associated with PCR-corrected treatment failure, using artemether-lumefantrine as the reference. This study is registered with PROSPERO, CRD42018104013.We did a systematic review of interventional or observational cohort studies assessing the efficacy of artemisinin-based or quinine-based treatments in pregnancy. Seven databases  and two clinical trial registries . Risk of PCR-corrected treatment failure was higher for the quinine monotherapy  but lower for artesunate-amodiaquine , artesunate-mefloquine , and dihydroartemisinin-piperaquine  than artemether-lumefantrine (n=1278) after adjustment for baseline asexual parasitaemia and parity. The risk of gametocyte carriage on day 7 was higher after quinine-based therapy than artemisinin-based treatment .Efficacy and tolerability of artemisinin-based combination therapies (ACTs) in pregnant women are better than quinine. The lower efficacy of artemether-lumefantrine compared with other ACTs might require dose optimisation.The Bill & Melinda Gates Foundation, ExxonMobil Foundation, and the University of Oxford Clarendon Fund. The physiological changes that occur during pregnancy mean that pregnant women are more susceptible to malaria than women who are not pregnant. Symptomatic and asymptomatic infections affect both mother and fetus.Over the past 30 years, pregnant women have been systematically excluded from standard antimalarial treatment efficacy studies.13Evidence before this studyClinicalTrials.gov). The final search was done on April 26, 2019, without restrictions on publication year or language. Previous aggregated data meta-analyses showed a lower efficacy of quinine-based treatment than artemisinin-based treatments, though the strength of evidence was low as the number of randomised control trials (RCTs) was very small, and different study designs and outcome measures were used. It was impossible to compare efficacy of different ACTs because of the paucity of RCTs.We did a systematic literature search for studies assessing treatment efficacy of artemisinin-based or quinine-based treatments for uncomplicated falciparum malaria  in pregnancy using seven databases  and two clinical trial registries  of data identified in the systematic literature review. Quinine is recommended to treat malaria in the first trimester of pregnancy and is still commonly used for all trimesters. Our study shows that quinine was less preferable than ACTs because of higher treatment failure, unless combined with clindamycin, higher occurrence of acute adverse events , and higher risk of gametocyte development . Artemether-lumefantrine (AL), the most commonly used ACT, shows the best tolerability but a lower efficacy than other standard ACTs. In moderate-to-high falciparum malaria transmission areas, the risk of treatment failure was higher in nulliparous women. Dose optimisation of AL for pregnant women should be further investigated. Although the numbers of first trimester pregnancies studied were small, these findings support recommendations in 2017 that ACTs should replace quinine as the treatment of choice for falciparum malaria in all trimesters.Implications of all the available evidenceThis meta-analysis, together with previous research on the safety of ACTs in the first trimester, provides compelling evidence that both efficacy and safety of ACTs in pregnant women are better than quinine-based treatments. Suboptimal dosing of lumefantrine with pregnant women might explain a slightly lower efficacy of AL than other ACTs. Dose optimisation of antimalarial drugs in pregnancy, supported by pharmacokinetic studies, will be required to achieve the highest treatment success in pregnancy to protect both mother and fetus from the adverse effects of malaria infection.Considering the variability of study designs and the fact that most antimalarial studies in pregnancy were single-arm studies, conclusions from conventional aggregated meta-analyses were not reliable.By including single-arm studies and more studies done after the release of the treatment guidelines, this study aims to contribute to the body of evidence by pooling individual patient data from studies to assess the efficacy of recommended antimalarial drugs for uncomplicated falciparum malaria during pregnancy, including patients who were asymptomatic. Our objective was to compare the efficacy and tolerability of artemisinin-based treatments (ABT) and quinine-based treatments (QBT), and also assess the difference between ACTs using artemether-lumefantrine (AL), the most used ACT, as the reference standard.ClinicalTrial.gov) were used. Prospective treatment efficacy studies of uncomplicated falciparum malaria, including pregnant women in any trimester, were identified using a combination of five components: malaria, pregnancy, treatment or names of antimalarial drugs, study design , and outcome types (efficacy), up until the final date of April 26, 2019.We did a systematic review on the efficacy of ABT and QBT on uncomplicated falciparum malaria, including asymptomatic parasitaemia  in pregnancy without any restrictions on language or publication year, and this was published elsewhere.Plasmodium falciparum parasitaemia was confirmed by microscopy before treatment, regardless of the patient's symptoms, the length of active follow-up was 28 days and over, and PCR was used to differentiate the recurrence of infections with P falciparum during follow-up.Studies were included in this meta-analysis if P falciparum within 28 (\u00b13) days. Acute adverse events were regarded as present if the symptom was actively assessed and recorded on any day between days 1 and 7.Pregnancy outcomes are reported elsewhere.All individual patient data from each study were pooled together for statistical analyses. For the descriptive analysis, treatment efficacy, and gametocyte positivity for each treatment at fixed timepoints  were estimated by the Kaplan-Meier method in each study at each site. Then, after complementary log-log transformation,We did a one-stage IPD meta-analysis using the Cox proportional hazards regression with shared frailty for study sites to identify the risk factors for recrudescence and to compare different treatment drugs. We used univariable and multivariable mixed effects logistic regression models to model risk of parasite positivity, gametocyte positivity, and acute adverse events. We used hazard ratios (HRs) for PCR-corrected efficacy and ORs for the other outcomes.Plasmodium vivax intercalated infection  was regarded as a time-dependent covariate if the original study genotyped falciparum recurrences regardless of intercalated infection with P vivax. Otherwise, intercalated infection with P vivax was regarded as censored following the WHO guidelines.We did a complete case analysis because of the small proportion of missing data for the main variables of interest. For all regression models, we identified independent risk factors by backward elimination.Statistical analysis was done using R  or Stata MP15.1 .The funders did not participate in the study design, the writing of the paper, decision to publish, or preparation of the manuscript.Our search identified 30 studies with PCR-corrected efficacy. With the exception of two unpublished studies, individual patient data from 4968 (92%) of 5360 episodes in 19 studiesThe included studies were done between 1995 and 2014 in ten different countries : nine stTen antimalarial treatments were included in this analysis: AL (1278 [89\u00b77%] of 1425 episodes in the literature), artesunate-amodiaquine , artesunate-mefloquine , dihydroartemisinin-piperaquine , artesunate-sulfadoxine-pyrimethamine , artesunate monotherapy (230 [100%] of 230 episodes), artesunate with clindamycin , artesunate-atovaquone-proguanil , quinine monotherapy (244 [81\u00b76%] of 299 episodes), and quinine with clindamycin . The fixed dose formulation was used in all participants who took ASAQ and in 962 (93\u00b76%) of the 1028 participants who took ASMQ .P falciparum mono-infections. The geometric mean parasite density before treatment was 1189 (range 1\u2013447\u2008638) asexual parasites per \u03bcL, with 79 (1\u00b76%) of 4968 episodes considered to be hyperparasitaemic (defined as >100\u2008000 parasites per \u03bcL). Only 504 (10\u00b74%) of the 4854 women were febrile (>37\u00b75\u00b0C) at presentation.Of the 4968 episodes in 4745 women, the mean age was 23\u00b75 years (SD 6\u00b70), and the median parity was one . Compared with nulliparous women, the risks of treatment failure were decreased in primiparous women in moderate  and high transmission areas ; and in multiparous women in moderate  and high transmission areas ; but not different in low transmission areas.P vivax intercalated infection. The estimates for these models were similar to the main analysis above  in the multivariable analysis might be due to its under-dosing in pregnant women, particularly if the immunity level is low , although not significant. ASSP efficacy was essentially equivalent to artesunate monotherapy (but given only for 3 days) in areas of sulfadoxine-pyrimethamine (SP) resistance. Considering the spread of SP resistance,Higher baseline asexual parasite density was associated with a higher risk of PCR-corrected treatment failure, as described in non-pregnant populations.The risk of recurrent falciparum malaria was high after AL and quinine monotherapy, as was expected from the shorter half-life and post-treatment prophylactic effect. Considering the cumulative effect of malaria recurrences on the fetus,The use of QBT leads to a higher risk of gametocyte development than ABT because artemisinin derivatives have a gametocytocidal effect but quinine does not.Quinine was associated with higher risks of abdominal pain, anorexia, dizziness, nausea, tinnitus, and vomiting than AL, limiting its practical use in real-life settings. Tolerability is important because pregnant women infected with malaria are generally less symptomatic than non-pregnant patients and are therefore less likely to accept adverse drug events. Poor adherence and patient discomfort could even be further exacerbated by morning sickness in the first trimester when QBT is recommended.Among ACTs, adverse events were considered to be mainly due to the partner drug, and AL and DP were the better tolerated regimens. The risk of anorexia, nausea, vomiting, and dizziness was higher in ASAQ and ASMQ than in AL or DP, although none of the studies included in this pooled analysis were double-blinded. Pregnancy outcomes are described in detail by Saito and colleagues,P vivax intercalated infection also did not affect our conclusions (This study has some limitations. Firstly, although this meta-analysis includes over 90% of patients that are available in the literature, some non-standard treatments were documented in only a small number of women. Nine studiesclusions , therebyIn conclusion, this meta-analysis, together with the evidence of safety shown in previous research,The WorldWide Antimalarial Resistance Network data repository.De-identified data are available from"}
+{"text": "Plasmodium falciparum malaria to interrupt its transmission, especially in low-transmission settings of multidrug resistance, including artemisinin resistance. Policy makers in Cambodia have been reluctant to implement this recommendation due to primaquine safety concerns and a lack of data on its efficacy.Since 2012, a single low dose of primaquine  with artemisinin-based combination therapies has been recommended as the first-line treatment of acute uncomplicated  Plasmodium falciparum malaria to interrupt its transmission, especially in low-transmission settings of multidrug resistance, including artemisinin resistance. Policy makers in Cambodia have been reluctant to implement this recommendation due to primaquine safety concerns and a lack of data on its efficacy. In this randomized controlled trial, 109 Cambodians with acute uncomplicated P. falciparum malaria received dihydroartemisinin-piperaquine (DP) alone or combined with SLDPQ on the first treatment day. The transmission-blocking efficacy of SLDPQ was evaluated on days 0, 1, 2, 3, 7, 14, 21, and 28, and recrudescence by reverse transcriptase PCR (RT-PCR)  and membrane feeding assays with Anopheles minimus mosquitoes (gametocyte infectivity). Without the influence of recrudescent infections, DP-SLDPQ reduced gametocyte carriage 3-fold compared to that achieved with DP. Of 48 patients tested on day 0, only 3 patients were infectious to mosquitoes (\u223c6%). Posttreatment, three patients were infectious on day 14  and on the 1st and 7th days of recrudescence ; this overall low infectivity precluded our ability to assess its transmission-blocking efficacy. Our study confirms the effective gametocyte clearance of SLDPQ when combined with DP in multidrug-resistant P. falciparum infections and the negative impact of recrudescent infections due to poor DP efficacy. Artesunate-mefloquine (ASMQ) has replaced DP, and ASMQ-SLDPQ has been deployed to treat all patients with symptomatic P. falciparum infections to further support the elimination of multidrug-resistant P. falciparum in Cambodia. Since 2012, a single low dose of primaquine  with artemisinin-based combination therapies has been recommended as the first-line treatment of acute uncomplicated  Plasmodium falciparum lineage resistant to both artemisinin and piperaquine (the KEL1/PLA1 lineage) in the eastern Greater Mekong Subregion (GMS) threaten this remarkable global achievement  and are active against early gametocyte stages \u201310 and r\u2013like GMS .To date, several clinical studies conducted in Africa, Colombia, and Cambodia have assessed the safety and efficacy of SLDPQ \u201324, main\u201315\u201321\u2013Anopheles minimus mosquitoes, one of the main malaria vectors in Southeast Asia.In Cambodia, glucose-6-phosphate dehydrogenase deficiency (G6PDd) is a common X-linked disorder of the red blood cells, with frequencies ranging from 10.8% to 29.6% in males  and withP. falciparum recrudescences; 24 occurred within the 28-day follow-up on days 12 (n\u2009=\u20091), 14 (n\u2009=\u20091), 17 (n\u2009=\u20092), 18 (n\u2009=\u20091), 21 (n\u2009=\u20092), 22 (n\u2009=\u20092), 23 (n\u2009=\u20091), 24 (n\u2009=\u20092), 25 (n\u2009=\u20091), 26 (n\u2009=\u20092), 27 (n\u2009=\u20091), and 28 (n\u2009=\u20098), and 4 occurred on days 44, 46, 52, and 124. Of the 28 patients with recrudescences, 10 were retreated with the standard 3-day DP regimen and 18 were treated with quinine or quinine plus tetracycline; 6 of the 10 patients retreated with DP experienced a second recrudescence.One hundred nine patients with uncomplicated falciparum malaria were enrolled and treated with the standard 3-day regimen of dihydroartemisinin-piperaquine (DP) alone  or DP-SLDPQ . Seven patients were lost to follow-up (6.4%). Of the remaining 102 patients, 28 (27.4%) had PCR-proven P\u2009=\u20090.3057).At day 0, gametocytes were detected in 47/109 patients (43.1%) by TaqMan RT-PCR  .A total of 55 patients were included in the direct membrane feeding assays (DMFAs). Among the 292 feeding assays, only 113 of 14,444 (0.78%) dissected mosquitoes were infected . OverallAnopheles minimus infectivity by DMFA.In 2012, the WHO recommended adding 0.25-mg/kg SLDPQ to first-line treatments (ACTs) to kill mature gametocytes and block human-to-mosquito transmission . As no dOur results showed that SLDPQ, when given on the first day of the 3-day standard ACT treatment, significantly decreased gametocyte carriage over time, starting on day 3, and that gametocyte carriage was the lowest on day 7, resulting in a 5.59-fold reduced risk relative to that with treatment with DP alone over 28\u2009days in the univariate analysis. This finding is consistent with recent reports on the efficacy of PQ in reducing gametocyte carriage 15\u2013\u201325, 31 a\u2013P. falciparum infections in western Cambodia (\u223c19%) but which is 3-fold higher than the 6% reported earlier in Ratanakiri Province, Cambodia  in our study and the very high recrudescent rate (~28%), our high gametocyte carriage rate is consistent with the presence of artemisinin-resistant P. falciparum, which has also led to piperaquine resistance , which is similar to that reported previously in 2010 in patients with artemisinin-resistant Cambodia . Given tsistance . This islciparum , 35.P. falciparum infection who, nevertheless, had low rates of human-to-mosquito transmission  at baseline and three infectious events during follow-up. This is a major limitation of DFMAs, as previously reported by Lin et al.  at basele et al. . In anote et al.  were alse et al. , 19. Howe et al. . Male gae et al. , and theectivity \u201341.P. falciparum infections. Although we were unable to reconfirm the findings of the elegant studies of Dicko et al. , as described by Dysoley et al. (Anopheles minimus mosquitoes were fed on blood samples collected from a subset of 55 patients and on the days of recurrent (Drec) falciparum parasitemia (defined by the WHO as late treatment failures), depending on mosquito availability, using direct membrane feeding assays (DMFAs).This open-label randomized controlled trial assessing the tolerability and the safety of SLDPQ was carried out in Banlung, Rattanakiri Province, in northeastern Cambodia in 2015 and 2016 , a two-sided alpha value of 0.05, and a power of 80%, the sample size was 48 patients per arm, and this number was rounded up to 50. G6PD status was initially diagnosed in the field using a fluorescent spot test to allow for the allocation of SLDPQ. The results of the qualitative Carestart  rapid diagnostic test were assessed in parallel.The sample size calculation was based on the primary outcome of a reduction in the mean day 7 hemoglobin (Hb) concentration of 1 g/dl in the DP-SLDPQ arm with G6PDd patients versus that in the DP-SLDPQ arm with patients with normal G6PD levels. Assuming a mean \u00b1 standard deviation Hb concentration of 11.27 \u00b1 1.74 g/dl  with constant access to a 10% sucrose solution. Patient infectivity was determined by assessing infection prevalence  and infection intensity . Midguts were dissected in a 1% mercurochrome stain, and the presence and the number of oocysts were determined under a microscope (\u00d720 magnification). Dissections were performed on 7 day after the blood meal. The mean number of dissected females was 49 .DMFAs were carried out to assess individual malaria infectivity as described previously . BrieflyP. falciparum-positive samples were analyzed for the presence of gametocytes by a TaqMan RT-PCR, using primers spanning an exon-exon junction and targeting the Plasmodium falciparum meiotic recombination protein DMC1-like protein gene (GenBank accession number AF356553), as described previously -conserved whole-blood samples using a QIAamp RNA blood minikit , following the protocol recommended by the manufacturer. A two-step semiquantitative real-time PCR was performed to detect malaria parasites, as previously described . Followieviously . GametocClinicalTrials.gov . The protocols conformed to the Helsinki Declaration on ethical principles for medical research involving human subjects (version 2002), and informed written consent was obtained from all volunteers.Ethical approvals were obtained from the Cambodian National Ethics Committee for Health Research , and the trial is registered at Initially, univariate generalized linear mixed models (GLMMs) with a binomial distribution were fitted to model the PCR-measured gametocyte prevalence rates. In these models, follow-up days , treatment (DP versus DP-SLDPQ), the presence of gametocytes prior to treatment on day 0, recrudescence status (recrudescent versus cured patient), and the hemoglobin concentration prior to treatment on day 0 were included in the model as fixed factors. The patient identifier was coded as a random factor to account for repeated measures on the same individual. The univariate analyses were followed by analyses with a multivariable GLMM to model the dynamics of gametocyte prevalence posttreatment by including all the significant factors from the univariate analysis.n\u2009=\u20091, DP-SLDPQ arm). P values of <0.05 were deemed significant. All analyses were performed in R (v3.5.1) software  of the 21 patients with microscopically detected gametocytes on day 0, only 7 had complete follow-up microscopy data, (ii) TaqMan RT-PCR-measured gametocyte densities were highly variable during the 28\u2009days of follow-up, and (iii) by day 28, 10/34 patients with gametocytes on day 0 by TaqMan RT-PCR were still gametocytemic (software ."}
+{"text": "Artemether-lumefantrine antimalarial efficacy in pregnancy could be compromised by reduced drug exposure. Population-based simulations suggested that therapeutic efficacy would be improved if the treatment duration was increased. P=0.005). Pregnant women had lower exposures to artemether and dihydroartemisinin than nonpregnant women, resulting in 1.2% decreased exposure for each additional week of gestational age. By term, these exposures were reduced by 48% compared to nonpregnant patients. The overall exposure to lumefantrine was improved with the extended regimen, with no significant differences in exposures to lumefantrine or desbutyl-lumefantrine between pregnant and nonpregnant women. The extended artemether-lumefantrine regimen was well tolerated and safe and increased the overall antimalarial drug exposure and so could be a promising treatment option in pregnancy in areas with lower rates of malaria transmission and/or emerging drug resistance. Artemether-lumefantrine antimalarial efficacy in pregnancy could be compromised by reduced drug exposure. Population-based simulations suggested that therapeutic efficacy would be improved if the treatment duration was increased. We assessed the efficacy, tolerability, and pharmacokinetics of an extended 5-day regimen of artemether-lumefantrine compared to the standard 3-day treatment in 48 pregnant women and 48 nonpregnant women with uncomplicated falciparum malaria in an open-label, randomized clinical trial. Babies were assessed at birth and 1, 3, 6, and 12\u2009months. Nonlinear mixed-effects modeling was used to characterize the plasma concentration-time profiles of artemether and lumefantrine and their metabolites. Both regimens were highly efficacious (100% PCR-corrected cure rates) and well tolerated. Babies followed up to 1\u2009year had normal development. Parasite clearance half-lives were longer in pregnant women  than in nonpregnant women (2.43 h [1.05 to 6.00 h]) ( In sub-Saharan Africa, malaria is a major cause of maternal and newborn morbidity and mortality. Efficacious antimalarial preventive and treatment regimens can reduce this significantly, but therapeutic choices are limited by concerns over possible toxicity to the developing fetus. Because of these concerns, pregnant women are commonly excluded from clinical trials . This, c2\u20135\u20138\u2013Artemether is rapidly absorbed and rapidly cleared from plasma . Its major active metabolite, DHA, is formed rapidly and has a clearance pattern similar to that of the parent drug , 14. ArtP\u2009=\u20090.971), whereas the median hematocrit was significantly lower in pregnant women  than in nonpregnant women (36% [25 to 43%]) (P\u2009<\u20090.001). Pregnant women, as expected, had higher median body weights  and higher median body mass index (BMI) values . The median doses of artemether and lumefantrine administered by treatment arm are presented in Table S1 in the supplemental material.Ninety-six patients were enrolled between June 2013 and April 2014  and 2. BTables 1 to 1/2) were significantly longer in pregnant women than in nonpregnant women: 3.30 h (1.39 to 7.83 h) versus 2.43 h (1.05 to 6.00 h) (P\u2009=\u20090.005). There was no significant relationship between predicted exposure (area under the concentration-time curve [AUC] from time zero to infinity) to artemether or DHA or the sum of the molar exposure to both drugs and parasite clearance half-life  parasite clearance half-lives  in pregnant patients and 14.6% (7/48) in nonpregnant women; in 6 women, gametocytes were observed on day 1 but not at inclusion (3 in each arm). There were no differences in gametocyte positivity rates between the two arms (data not shown). Gametocytes persisted in the peripheral blood for up to 2\u2009weeks posttreatment.P\u2009=\u20090.92). In the nonpregnant group, the uncorrected cure rates by day 42 were 87.5%  following 3\u2009days of treatment and 95.7%  following 5\u2009days of treatment (P\u2009=\u20090.32)  following 3\u2009days of treatment and 90.0%  following 5\u2009days of treatment (\u2009=\u20090.32) . The PCRP\u2009=\u20090.111). In addition, 62.5% (30/48) of nonpregnant patients had a history of fever prior to admission, compared to 46% (22/48) of patients in the pregnant group (P\u2009=\u20090.101). Fever resolution was rapid and similar in the two groups. Within 24\u2009h, all women were afebrile.At admission, 12.5% (6/48) of nonpregnant patients had fever, compared to 2.1% (1/48) of pregnant patients (P\u2009=\u20090.32), and those in pregnant women were 4.5 (8.4) in the 3-day arm and \u22121.3 (8.6) in the 5-day arm (P\u2009=\u20090.08). By day 28, the mean (SD) relative differences in hematocrit in nonpregnant women were 0.70 (5.71) in the 3-day arm and 0.03 (6.06) in the 5-day arm (P\u2009=\u20090.96), and those in pregnant women were 6.9 (9.7) in the 3-day arm and 3.2 (11.6) in the 5-day arm (P\u2009=\u20090.16) (absolute hematocrit differences are shown in Table S2).By day 7, the mean (standard deviation [SD]) relative differences from the baseline in hematocrit in nonpregnant women were \u22120.70 (6.81) in the 3-day arm and 0.24 (6.27) in the 5-day arm (P\u2009=\u20090.16) in the pregnant group and in 3 cases in the 5-day arm in the nonpregnant women. AST levels were above the normal range in 2 patients in the 5-day arm  and 3 patients in the 3-day arm  (P = 0.72) in the pregnant group and in 3 women  in the 5-day arm and 5 women  in the 3-day arm (P = 0.44) in the control group. None of the transaminase elevations were more than three times above the upper limit of normal, and all cases normalized within a month.Median serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), creatinine (milligrams per deciliter), and albumin (grams per deciliter) were similar at baseline and at discharge between groups and treatment regimens . The ALTP\u2009=\u20090.82), but all values returned to normal within a month.The serum creatinine levels were comparable and within the normal range between the two treatment arms at baseline and at discharge in pregnant and nonpregnant women. Creatinine was above the normal range in 5 cases  in the 5-day treatment arm and in 4 cases  in the 3-day treatment arm in the pregnant group (n\u2009=\u200918), dizziness (n\u2009=\u20098), and gastrointestinal disorders (n\u2009=\u20098). Gastrointestinal disorders occurred more frequently in women who received 5-day AL than in those who received the 3-day treatment (P\u2009=\u20090.03).Artemether-lumefantrine was generally very well tolerated. At least one adverse event of mild or moderate intensity was reported in 65.6% of patients (63/96) . All advFour serious adverse events, all in pregnant women, were reported, 3 in the 3-day arm and 1 in the 5-day arm. All were classified as being unrelated to the study treatment .The electrocardiogram (ECG) values  were normal at admission in all patients, and none of the women had observed QTc intervals exceeding 500\u2009ms during the trial.P\u2009=\u20090.0017) correlation was found between predicted lumefantrine concentrations and the Fridericia-corrected QT intervals  . Median peak concentrations were approximately 6,000 to 7,000\u2009ng/ml in the four study arms (see Tables 7 and 8), resulting in a maximum Fridericia-corrected QT (QTcF) interval prolongation of 7.02 to 8.19\u2009ms. A weak but significant (0\u2009ng/ml) . No sign0\u2009ng/ml) .P\u2009=\u20090.61). A positive correlation between birth weight and BMI was observed (P\u2009=\u20090.032); the mothers of the babies with low birth weight (n\u2009=\u20097) had the lowest BMI values. There were no significant relationships between parasitemia, fever, or anemia and birth weights. One case of polydactylism  of both hands was observed in a baby of a woman treated with the 5-day regimen at 31\u2009weeks of gestation. This malformation was a family trait, and the exposure to the study drug occurred after the formation of fetal digits was complete; it was therefore classified as being unrelated to the study treatment. Two infant deaths at 3\u2009months of age in the 5-day arm were attributed to severe diarrhea and suspected respiratory infection, respectively. These were also assessed as being unrelated to the study treatment. All other infants had normal physical and neurological development according to Denver developmental milestones (Forty-five women gave birth to a live baby: 41 women (85.4%) had a full-term pregnancy (between \u226537 and 40\u2009weeks), and 4 (8.3%) had a preterm pregnancy (between \u226528 and\u2009<37\u2009weeks). Three women (6.3%) had a stillbirth: two in the 3-day group and one in the 5-day group (lestones  when assP\u2009=\u20090.672). There was no significant difference in the median times between study treatment and delivery between women without placental malaria  and those with placental malaria  (P\u2009=\u20090.673). However, some women might have had a second episode of malaria between the study treatment and delivery. All pregnant women in this study received intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP), administered by the study nurses.The median times between study treatment administration and delivery were not significantly different between women who received 3-day  and those who received 5-day  treatments (n\u2009=\u200922) and 9 in the 5-day arm (n\u2009=\u200922). Eight women (47%) with a malaria-positive biopsy specimen also had a malaria-positive peripheral blood slide at delivery. There were more cases of active placental malaria in the 3-day arm  than in the 5-day arm , although the difference was not significant (P\u2009=\u20090.105). In one case , the mother had a positive peripheral blood smear at delivery, and malaria parasites were found in the cord blood . There was no significant difference in the median birth weights between those with and those without placental malaria: 3,300\u2009g  versus 3,070\u2009g  , respectively.Placental biopsy specimens were available from 44 patients. The remaining four patients delivered at home, and the placenta was discarded. Parasite-infected red blood cells (RBCs) were observed in 38.6% (17/44) of the placental biopsy specimens: 8 in the 3-day arm (P\u2009<\u20090.001). In nonpregnant women, the median values were 541\u2009ng/ml  in the 3-day arm and 1,995\u2009ng/ml  in the 5-day arm (P\u2009<\u20090.001). Only one pregnant woman in the 3-day arm had a lumefantrine plasma level at day 7 below the 280-ng/ml cutoff. On day 14, the median (range) plasma lumefantrine levels in pregnant women were 151\u2009ng/ml (68 to 265\u2009ng/ml) in the 3-day arm and 247\u2009ng/ml (99 to 541\u2009ng/ml) in the 5-day arm (P\u2009<\u20090.001). In nonpregnant women, the median day 14 levels were 146\u2009ng/ml (78 to 409\u2009ng/ml) in the 3-day arm and 312\u2009ng/ml (87 to 699\u2009ng/ml) in the 5-day arm (P\u2009<\u20090.001).The observed median (range) lumefantrine concentrations at day 7 in pregnant women were 597\u2009ng/ml (216 to 928\u2009ng/ml) in the 3-day arm and 1,545\u2009ng/ml  in the 5-day arm (P\u2009<\u20090.05). A transit absorption model with four transit compartments performed better than all other absorption models (P\u2009<\u20090.05). Estimating the transit rate constant trk and the absorption rate constant (ak) separately did not improve the model significantly, and they were assumed to be identical in the final model. None of the covariates evaluated  had a significant impact on the pharmacokinetic properties of lumefantrine. However, as a recent pooled analysis showed that body weight influences lumefantrine pharmacokinetic properties (P\u2009<\u20090.05) . Body weight was included in the model based on biological plausibility and improved the model fit. No other covariates were evaluated for the metabolite. Between-subject variability was estimated on all parameters. However, to stabilize the model, the variability in pharmacokinetic parameters associated with one peripheral compartment was fixed to zero. Final primary parameters, parameter precision, shrinkage, and secondary parameter estimates for the parent drug and metabolite are presented in A total of 816 samples were analyzed for plasma lumefantrine concentrations, and 712 samples were analyzed for desbutyl-lumefantrine concentrations. A two-compartment disposition model was used to describe the pharmacokinetics of lumefantrine . This was superior to other investigated disposition models  but resulted in improbable parameter estimates . A transit compartment model with six transit compartments was superior to all other absorption models (P\u2009<\u20090.05). Estimating trk and ak separately did not improve the model significantly, and they were assumed to be identical in the final model. Only 5.7% and 7.2% of artemether and DHA samples, respectively, were below the lower limit of quantification (LLOQ). Similar pharmacokinetic parameter estimates were obtained for both the one- and two-compartment disposition models when using the M1 or M3 method for LLOQ data . Thus, for parsimony, censored data (drug measurements below the LLOQ) were omitted in the final pharmacokinetic model . Estimated gestational age was a significant covariate affecting the relative artemether bioavailability, resulting in a 1.2% decrease in the relative bioavailability for each additional week in estimated gestational age. This resulted in a maximum difference of 48% between nonpregnant and pregnant women . Estimating just a categorical difference between pregnant and nonpregnant women resulted in a 29.2% reduction in the relative bioavailability, while estimating a difference for each trimester resulted in a 25.2% reduction for trimester 2 and a 32.9% reduction for trimester 3 compared to nonpregnant women. The impact on DHA exposure was proportional to that established for artemether. No other covariates evaluated (including body weight and treatment duration) had a significant impact on the pharmacokinetic properties of artemether or DHA. However, body weight was retained in the model due to its strong biological prior, as argued above for lumefantrine. The full covariate approach, investigating the effect of pregnancy on secondary DHA pharmacokinetic parameters, showed significant reductions in exposure to artemether and DHA . Two-compartment disposition models resulted in improved model fits for both AM and DHA  worldwide for the treatment of uncomplicated malaria in pregnancy. Previous studies suggested that exposure to these drugs is reduced in pregnancy , 22, 23.election , 13. A rIn this prospective randomized study, the tolerability and safety of the extended regimen were excellent, with no evidence of drug-related toxicity. There were no adverse effects in the pregnant woman, nor were any significant electrocardiographic, biochemical, or hematological changes detected. A weak correlation was found between predicted lumefantrine concentrations and the Fridericia-corrected QT, showing an increase of 1.17\u2009ms with an increase in the lumefantrine concentration of 1,000\u2009ng/ml, resulting in a median QTcF prolongation of <10\u2009ms in all treatment groups. Birth and infant (up to the age of 1\u2009year) outcomes did not differ significantly between the two regimens, both of which achieved 100% cure rates in pregnant and nonpregnant women. This high cure rate in western Democratic Republic of the Congo (DRC) was observed 2 years after the introduction of artemether-lumefantrine as a second first-line treatment (in addition to amodiaquine-artesunate) for uncomplicated malaria as part of the revised National Guidelines of Treatment of Malaria . The parasite clearance times were significantly shorter in pregnant women  than in nonpregnant women (2.43\u2009h [1.05 to 6.00 h]). This was not associated with the early exposures to artemether and DHA and suggests that pregnancy may reduce splenic clearance function. Malaria parasites were detected in the placentas of 38% of women. In the study area, malaria transmission is high and perennial, so it is likely that these placental infections resulted from prior or subsequent malaria episodes, not treated during the study, which remained subpatent, and/or reduced efficacy of sulfadoxine-pyrimethamine used as a preventive treatment. Pregnant women had lower exposures to both artemether and DHA than nonpregnant women, resulting in 1.2% decreased exposure per week increase in gestational age. By term, these exposures were reduced by 48% compared to nonpregnant patients. This finding was also confirmed with a full covariate model, which showed that exposure to dihydroartemisinin was significantly reduced in pregnant women. These results are comparable with previous findings in pregnant Congolese women, in the same study area, treated with artesunate , 25. LowIn the present study, the results of the nonlinear mixed-effects modeling showed that pregnancy did not affect significantly any of the pharmacokinetic parameters for either lumefantrine or the desbutyl metabolite. The overall exposure, as expected, was substantially improved in the 5-day regimen . The observed median day 7 lumefantrine plasma concentrations were not different between pregnant and nonpregnant women. The median levels in our study  were comparable to those found in Uganda   after thThese results contrast with findings from previous studies in Asia and Africa investigating the pharmacokinetic properties of lumefantrine in pregnant women. In Asia, a reduced exposure (AUC) to lumefantrine was observed in pregnant Karen women , 38% of whom had day 7 lumefantrine venous blood concentrations of <280\u2009ng/ml. This was associated with increased lumefantrine elimination . In a laThe present study was not powered to detect low frequencies of adverse events, and by study design, women in the 5-day arm were observed for a longer period than were those in the 3-day arm. However, the drug tolerability and safety results in this small cohort are reassuring. Longer courses risk reduced adherence. In this study, the treatment was administered under direct observation and with milk to maximize drug absorption. In daily clinical practice, adherence to a 5-day treatment and coadministration with fatty foods might be suboptimal. A new solid-dispersion formulation of lumefantrine is currently under development by Novartis . The preIn conclusion, both the current and extended regimens of artemether-lumefantrine were highly efficacious, well tolerated, and safe in Congolese pregnant women with uncomplicated falciparum malaria. The 5-day treatment increases drug exposures without apparent toxicity and is therefore a promising option for the treatment of malaria in pregnancy in areas where efficacy has declined.This study was conducted by the Kinshasa School of Public Health-University of Oxford Medical Research Unit, Kinshasa, Democratic Republic of the Congo, at the Maternity Hospital of Kingasani, located in a suburban area of Kinshasa. Malaria transmission is high and perennial in this region.Plasmodium falciparum parasitemia of between 100 and 200,000 parasites/\u03bcl and a hematocrit of \u226521%. Pregnant women were included if the estimated gestational age was \u226514\u2009weeks and they had a singleton viable fetus confirmed by ultrasound using Hadlock\u2019s method  according to their status . The randomization lists were computer generated by an independent statistician, with sampling time points randomized within each of the 6 blocks.In the 3-day regimen, patients received 6 doses , and in the 5-day regimen, patients received 10 doses . Each dose  was administered under direct observation with 200\u2009ml of milk. If vomiting occurred within 30 min, a full dose was repeated, and if vomiting occurred between 30 min and 1 h, half the dose was repeated. Recurrent episodes of malaria were to be treated with oral quinine sulfate , three times daily for 7\u2009days. All patients were admitted for the duration of the treatment, and doses were supervised.The outcomes were the therapeutic efficacy , physicaData on clinical symptoms were collected daily during hospitalization  and then weekly until day 42 or at delivery, whichever occurred first. Babies were assessed at delivery and at 1, 3, 6, and 12\u2009months according to Denver developmental milestones .Hemoglobin was measured from capillary blood at screening using Hemocue Hb301 . Hematocrit was measured at screening, at baseline, every 12\u2009h during hospitalization, and weekly using a Hawksley Haematospin 1400 instrument . Total and differential white blood cell counts were performed by Sysmex at baseline and discharge. Plasma alanine aminotransferase (ALT), aspartate aminotransferase (AST), albumin, and creatinine levels were measured by a SEAC Screen Master from blood collected in lithium heparin tubes at baseline and 48\u2009h in the 3-day group and 96\u2009h in the 5-day group. Blood films were prepared and read using standard procedures at screening; at 0, 6, and 12\u2009h; and then every 12\u2009h until two negative blood smears were observed. Blood samples from recurrent episodes of malaria during follow-up and at delivery from peripheral blood in the mother and from cord blood and heel prick in the newborn were analyzed by PCR to distinguish recrudescence from new infections using a standard parasite genotyping methodology .A 12-lead electrocardiogram was recorded before or within 28\u2009h after the first dose and at discharge  using a Cardiofax ECG-9620 electrocardiograph , which provides automated Fridericia-corrected QT intervals (QTcF). The instrument-generated QT and QTcF values were used to calculate Bazett-corrected QT (QTcB) values. The correlation between QT, QTcF, and QTcB and heart rates was evaluated with unweighted ordinary linear regression using GraphPad Prism 8.2.0.g for 7 min at +4\u00b0C, and plasma was then stored in cryovials at \u221280\u00b0C until shipment on dry ice to the Department of Clinical Pharmacology at the Mahidol-Oxford Research Unit in Thailand.Venous blood for drug measurement was drawn into prechilled lithium heparin tubes at 3 fixed time points  and taken at random 7 times within the following time windows, after the first drug intake (hour zero): 0 to 3\u2009h, 3 to 6\u2009h, 6 to 12\u2009h, 12 to 60\u2009h, 60 to 72\u2009h, 72 to 144 h, and 192 to 336\u2009h. Blood samples were centrifuged at 1,400\u2009\u00d7\u2009A placenta biopsy was performed within 4\u2009h of delivery, and the specimen was fixed in 10% buffered formalin before routine processing and hematoxylin and eosin (H&E) staining for microscopic examination to determine the presence of active placental malaria and any other pathology.Safety outcomes were adverse events as defined by International Council for Harmonisation good clinical practice guidelines .Artemether and DHA plasma concentrations were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS) . Qualityz-test, and Fisher\u2019s exact test were used to compare proportions. Analysis of variance (ANOVA) was used for normally distributed continuous data, and a Kruskal-Wallis test was used for continuous data with a skewed distribution. Paired t tests were used for comparisons within the same patient over time. All statistical analyses were performed using STATA IC software  or GraphPad Prism 8.2.0. The parasite clearance half-life (PC1/2) was estimated using the WorldWide Antimalarial Resistance Network (WWARN) parasite clearance estimator, modified to allow for a lower threshold of parasitemia at time zero  .The study was approved by the Oxford Tropical Research Ethics Committee, the Kinshasa School of Public Health Institutional Review Board, and the Ministry of Public Health of DRC. The study was conducted in accordance with good clinical practice guidelines .The data that support the findings are available from the authors upon reasonable request and with permission of the University of Oxford and the Kinshasa School of Public Health."}
+{"text": "Data was entered and analyzed using the WHO Excel-based application. A total of 496 patients were enrolled. In Diourbel, PCR non-corrected/corrected adequate clinical and parasitological responses (ACPR) was 100.0% in both the AL and ASAQ arms. In Kedougou, PCR corrected ACPR values were 98.8%, 100% and 97.6% in AL, ASAQ and DP arms respectively. No Pfk13 or Pfcoronin mutations associated with artemisinin resistance were found. This study showed that AL, ASAQ and DP remain efficacious and well-tolerated in the treatment of uncomplicated P. falciparum malaria in Senegal.In 2006, Senegal adopted artemisinin-based combination therapy (ACT) as first-line treatment in the management of uncomplicated malaria. This study aimed to update the status of antimalarial efficacy more than ten years after their first introduction. This was a randomized, three-arm, open-label study to evaluate the efficacy and safety of artemether-lumefantrine (AL), artesunate-amodiaquine (ASAQ) and dihydroartemisinin-piperaquine (DP) in Senegal. Malaria suspected patients were screened, enrolled, treated, and followed for 28 days for AL and ASAQ arms or 42 days for DP arm. Clinical and parasitological responses were assessed following antimalarial treatment. Genotyping ( This policy review was adopted as ACTs were proven to be the most effective treatment in the context of resistance to chloroquine and other antimalarial drugs2. The scaling up of ACTs at national level has largely contributed to the reduction in morbidity and mortality linked to malaria in Senegal3. Artemether-lumefantrine (AL) and artesunate-amodiaquine (ASAQ) are used as first-line treatment but dihydroartemisinin-piperaquine (DP) has been widely used to compensate for antimalarial drug shortages in 2010 and 2011.Since the abandoning of chloroquine, due to high rate of resistance, the Senegal national malaria control program (NMCP) adjusted its national guidelines for the treatment of uncomplicated malaria by introducing sulphadoxine-pyrimethamine (SP) in 2003, and artemisinin-based combination therapy (ACT) in 2006, complying with the World Health Organization (WHO) recommendations4. Since May 2010, ACTs were offered free of charge in the public sector5.In 2008, ASAQ combination was subsidized by the government and its partners, and was made available at a very low price in public health facilities and private pharmaciesPlasmodium falciparum populations over time. Recent studies in Southeast Asia (SEA) have shown a decrease in the effectiveness of ACTs such as artesunate used as monotherapy7. Although this situation is not yet reported in Africa, it is likely that this resistance could potentially spread in Africa as history showed for chloroquine8. This is particularly important as a previous ex vivo study in Senegal showed that parasites became less sensitive to amodiaquine, artemisinin and chloroquine over time9.However, the wide use of ACT may exert selective pressure on 10.Thus, simple method for monitoring antimalarial drugs is crucial for proper management of clinical cases and early detection of resistance. In accordance with WHO recommendations, the efficacy of first and second-line antimalarial drugs should be evaluated at least once every two years at all sentinel sitesIn vivo drug clinical trials are the gold standard for assessing the therapeutic efficacy of antimalarials11. In addition, in vitro assays for the sensitivity of human malaria parasites to antimalarial drugs provide useful complementary data from drug-efficacy surveillance12.In Senegal, the NMCP and its partners regularly conduct therapeutic efficacy study (TES) and molecular markers of antimalarial drug resistance are monitored.Pfkelch13 (Pfk13) propeller domain were found to be associated with delayed parasite clearance in vitro and in vivo in SEA. These mutations were discovered in the laboratory by long-term in vitro selection using culture-adapted Plasmodium falciparum isolates by stepwise increases in artemisinin exposure13. Although the frequency of mutant alleles strongly correlated with the resistance to artemisinin, the relevance of these mutations on artemisinin resistance in other endemic areas has been subject of debates15. Using the same methodology as above13, but with P. falciparum isolates from Senegal, a recent study found no selected mutations on the Pfk13; instead, genetic variants  of the gene encoding the actin-binding protein P. falciparum coronin (Pfcoronin) were found to reduce the susceptibility of the parasite15. Thus, Pfcoronin, which is structurally similar to Pfk13, is believed to be a strong predictor of potential artemisinin resistance in Senegal and even probably elsewhere in Africa. However, for those mutations to be validated as a marker for artemisinin resistance there should be, at least, a correlation with delayed clearance in clinical studies.Mutations in the Pfk13 and Pfcoronin on samples that were subject to treatment failure, along with the routine TES of ACTs.Thus, we aimed to analyze simultaneously the sequences of A total of 796 patients suspected for malaria were screened during the study period. Of these, 496 62.3%) met the inclusion criteria and were enrolled accordingly as shown in Fig.\u00a0.3% met tAt baseline (day 0), the male to female sex ratio was largely dominated by males in Diourbel 5.1) while in Kedougou this was balanced (0.9). Thirty-six (12.2%) were under-five years old children, while this age group was not found in patients enrolled in Diourbel. Age group 5\u201315 years represented 117 patients (58.5%) and 163 patients (55.1%) in Diourbel and Kedougou respectively. Adult patients represented 83 (41.5%) and 97 (32.8%) in Diourbel and Kedougou respectively. Mean weight was 39.3\u2009kg and 36.1\u2009kg in Diourbel and Kedougou respectively. Mean body temperature was 38.3\u2009\u00b0C and 38.1\u2009\u00b0C in Diourbel and Kedougou respectively. Mean parasitemia was estimated at 11939 parasites/ul and 12991 parasites/ul in Diourbel and Kedougou respectively  in both AL and ASAQ arms.In Kedougou, PCR non-corrected ACPR were 96.5% (95%CI: 90.1\u201399.3), 100% (95%CI: 96.0\u2013100.0) and 96.4% (95%CI: 89.9\u201399.3) in AL, ASAQ and DP arms respectively; when PCR-corrected, ACPR became 98.8% (95%CI: 93.5\u2013100.0), 100% (95%CI: 96.0\u2013100.0) and 97.6% (95%CI: 91.6\u201399.7) respectively.Treatment failures (TF) were noted in Kedougou, as following: a) one ETF at day 3 in the ASAQ arm; b) three TF in the AL arm: 1 LCF at day 7 and 1 LPF at day 21, giving PCR-uncorrected failure rate of 3.5% (95% CI: 0.0\u20136.3%); c) three TF in the DP arm: one LPF at day 14, one LPF at day 35, and one LPF at day 42. After PCR correction, one sample in the AL arm and two in the DP arm (at day 14 and 35) were confirmed to be recrudescence cases.Considering the 10% of AL and ASAQ patients followed up to 42 days, there was three LPF in the AL arm and when PCR-corrected this yielded to one confirmed recrudescence day 35) as shown in Table\u00a05 as showmsp1 and msp2 genotyping and 24 SNP-based barcoding results were in agreement on all tested samples. Among the nine treatment failures, one was classified as ETF (recurrence at day 2), six samples were successfully genotyped and two samples gave negative PCR results on the day of recurrence. Thus, the successfully genotyped samples showed three cases of recrudescence and three re-infections. Out of the three recrudescence cases, two samples were successfully sequenced and gave interpretable sequences at day 0 and the day of respective recurrence for both Pfk13 and Pfcoronin. As well, the sequencing of the ETF sample was successful for the two molecular markers. For one recrudescence sample, the sequencing was obtained only at day 0 and this was unsuccessful for the day of recurrence.Pfk13 allele and wild-type Pfcoronin allele for both day 0 and day of recurrence , artesunate-amodiaquine (ASAQ) and dihydroartemisinin-piperaquine (DP), following a modified WHO 2009 protocol11, a minimum of 73 patients should be recruited to detect a failure rate \u22645%, at confidence level of 95% and an estimate precision of 5%. An additional of 20% was added to take into account lost to follow-up and withdrawals. By applying this guideline, 88 patients were targeted for each drug combination in each site.According to the WHO protocolP. falciparum mono-infection parasitemia of 1,000 to 100,000 asexual forms/\u03bcl, ability to swallow oral medication, ability and willingness to comply with the study protocol and visit schedule for the duration of the study and written informed consent from the patient or from the parent or guardian for enrolled children. Infected patients who did not meet inclusion criteria were treated according to the NMCP guideline.Malaria suspected patients were screened using both malaria rapid diagnostic test (RDT) (SD Bioline Pf/HRP2) and microscopy. Patients were enrolled if they were aged 6 months or above, had an axillary temperature \u226537.5\u2009\u00b0C at presentation or history of fever during the last 24\u2009hours, a positive In addition, dried blood spots (DBS) were collected on Whatman filter paper for molecular analysis including parasite genotyping (PCR correction) to differentiate re-infections from recrudescence and to determine molecular markers of artemisinin resistance on samples that were subject to treatment failure.41, mixed or mono-infection with another Plasmodium species detected by microscopy, positive pregnancy test, severe malnutrition, febrile conditions due to diseases other than malaria or other known underlying chronic or severe diseases, regular medication, which may interfere with anti-malarial treatment, history of hypersensitivity reactions or contraindications to any of the medicine(s) being tested or used as alternative treatment(s).Key exclusion criteria were any of the following: presence of general danger signs in children aged under 5 years or signs of severe falciparum malaria according to the definitions of WHO11. The allocation sequence was generated using a randomization table created by an Excel-based application. Patients were observed for 30\u2009min after the drug administration for adverse events or vomiting the study drugs. Any patient who vomited was re-treated with the same dose of medicine and observed for an additional 30\u2009min. If vomiting occurred again, the patient was withdrawn and treated with IV quinine and if necessary, the patient was referred to the nearest hospital. Paracetamol was offered to all feverish patients.Enrolled patients were randomly allocated into the three drug arms: oral administration of AL, ASAQ or DP using the WHO recommended therapeutic dose regimensAll daily doses were administered at the health facility level under the supervision of the medical staff.Follow-up visits were scheduled on days\u00a01, 2, 3, 7, 14, 21, and 28 for treatment with AL and ASAQ and additional follow up on days 35 and 42 for DP treatment . During the follow-up visit, the patients were subject to clinical assessment and parasitological examination of blood smear to seek for malarial parasites, and DBS was also collected.A physician from the research staff was responsible for monitoring the safety of ACTs. Adverse events were documented through interviews about previous symptoms and about symptoms that have emerged since the previous follow-up visit. A clinical examination was performed to determine any adverse event. The reported events were recorded and any serious adverse event was to be reported to the sponsor.Thick and thin smears were performed and slides were examined on days 0 to confirm the meeting criteria and during subsequent scheduled follow-up day or any unscheduled day if the patient returned for being unwell.42. Parasite density (per \u03bcl) was calculated assuming a white blood cell count of 8000/\u03bcl. All slides were read independently by two level 1 malaria microscopists and the average of the two counts was calculated. If any discrepancy was noted (either in species identification or difference of parasite density of >30%), slides were checked by a third independent reader, and parasite densities were calculated by averaging the two most close counts.Microscopy examination of Giemsa-stained thick and thin blood films was performed to identify parasite species and determine density according to the WHO proceduremsp1 and msp2 loci, as recommended by WHO11. In addition, a 24 SNP-based molecular barcode using High Resolution Melting qPCR44 was used. The WHO 2009 protocol requires sequential genotyping of msp1, msp2 and glurp genes to discriminate re-infections from recrudescence11. The latter is confirmed only when all three markers yield to the same genotype. However, comparable results from msp1 and msp2 genotyping and the 24 SNP-based barcodes have been reported. Unlike msp genotyping, the SNP-based barcodes present the advantage to be faster and less intensive45.In order to differentiate a recrudescence (same parasite strain) from a newly acquired infection (different parasite strain), a genotype analysis was performed. DNA was extracted from the DBS on day 0 (before treatment) and during recurrence of parasitemia on day 7 onwards (cases of treatment failure). All DNA samples from patients undergoing parasitemia recurrence were analyzed for genotyping of the highly polymorphic regions Pfk13 propeller domain (codon positions: 440\u2013600) and a portion of the Pfcoronin gene (codon positions: 31\u2013186) were amplified using a nested PCR assay. Amplicons were sequenced using Sanger method as previously described15 from extracted DNA on day 0 (before treatment) and during recrudescence of parasitemia on day 7 onwards.In the case of confirmed recrudescence as described above, DNA sequences were analyzed using the Geneious Prime software (version 2020.1.2) to identify specific single-nucleotide polymorphism (SNP) related to artemisinin resistance. Sequences generated in this study were submitted into the European Nucleotide Archive (EBI) database under the accession numbers LR782242, LR782250, LR782259, LR782260, LR782261 and LR782262.11.Treatment outcomes were classified according to the WHO protocol as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) and adequate clinical and parasitological response (ACPR)11.Data for each study participant were entered into the WHO standardized Microsoft Excel data collection form. This WHO excel sheet was specifically design for the classification of treatment outcome with and without PCR correctionThe study protocol received ethical clearance from the Ethic Committee of the Senegalese Ministry of Health. Written informed consent was obtained from all participants or parents/guardians of the children. The authors have complied with all relevant regulations for work with human participants. This study was registered at the Pan African Clinical Trials Registry on 09 March 2020 under the number PACTR 202003802011316."}
+{"text": "Plasmodium falciparum remains sensitive to artemether-lumefantrine in Mali.In Mali, since 2007, artemether-lumefantrine has been the first choice against uncomplicated malaria. Despite its effectiveness, a rapid selection of markers of resistance to partner drugs has been documented. This work evaluated the treatment according to the World Health Organization\u2019s standard 28-day treatment method. The primary endpoint was the clinical and parasitological response corrected by a polymerase chain reaction. It was more than 99.9 percent, the proportion of patients with anemia significantly decrease compared to baseline ( Plasmodium falciparum malaria, which has positively contributed to reducing the global burden of malaria [In the past two decades, we have witnessed a revolution in the diagnosis and treatment of malaria worldwide. In sub-Saharan Africa, classical antimalarial drugs have been gradually abandoned in favor of faster and more effective molecules, mainly because of the increasing resistance of parasites ,2. Indee malaria .In Mali, malaria remains a major health problem, with more than 2 million cases and more than 1050 deaths recorded in 2017, a case-fatality rate of 0.50\u2030 . In addiIn order to promote evidence-based policy decisions, the WHO stressed the need to monitor regularly the efficacy of antimalarial drugs using standardized protocols .Plasmodium falciparum are essential to validate the treatment and guarantee a rapid response to the infection emergence of parasite resistance. In Mali, some studies showed a rapid selection of molecular markers of P. falciparum resistance to artemisinin partner drugs [In vivo efficacy of antimalarial as well as the evaluation of the prevalence of molecular markers associated with resistance of er drugs ,9,10,11.In Southeast Asia, reports have shown the emergence and potential spread of resistance to artemisinin ,13,14, rPlasmodium falciparum to artemether-lumefantrine in two sentinel sites of Mali\u2019s National Malaria Control Program (NMCP) during two malaria transmission seasons 2017\u20132018.In an open and global world with the movement of populations, it seems appropriate to strengthen the monitoring of the effectiveness of ACTs. This study is part of the monitoring of the sensitivity of Study sites and malaria in Mali: The study was conducted according to WHO standard in vivo efficacy protocol  in two NThe work took place in three community health centers in Bougouni district  and a district hospital in the Bandiagara district between July 2017 and December 2018.Plasmodium falciparum mono-infection with asexual blood density \u22651000/\u03bcL and <200,000/\u03bcL, and the absence of severe signs of complicated malaria as defined by WHO [Patients: The study population included patients aged six months or older with uncomplicated acute malaria confirmed by microscopy. Other inclusion criteria included body weight \u22655 kg, presence of fever (axillary \u2265 37.5 \u00b0C) or a history of fever in the previous 24 h, Treatment: Eligible patients were treated with Artemether Lumefantrine (each tablet containing 20 mg artemether and 120 mg lumefantrine). The study drug used was provided to the local health facilities by the Malian Ministry of Health. The dosage and method of administration of the drugs followed the manufacturer\u2019s instructions and the Mali NMCP malaria treatment guideline. One dose at baseline (time 0), a second dose 8 h later, the third dose 24 h later and the last three doses 12 h apart  for 3 days. The number of tablets to be taken was determined according to body weight: one tablet for children from 5 to <15 kg, two tablets for children from 15 to <25 kg and three tablets for subjects of 25 to <35 kg and four tablets for subjects over 35 kg. Drug administrations were directly observed by the investigators. When vomiting occurred within 30 min of dosing, a new dose was re-administered. For febrile subjects (fever > 37.5 \u00b0C), paracetamol was used. In the event of signs of danger or severe malaria, the patient was hospitalized and given injectable artesunate, in accordance with the National Malaria Treatment Policy [Follow-up: Follow-up visits took place on days 1, 2, 3, 7, 14, 21 and 28 after enrollment or at any time when the child was ill. Patients who withdrew their consent or participation were stopped prematurely for various reasons were followed through local guides until the end of the study. Adverse events that occurred were recorded, treated, and assessed by severity and drug-study relationship. Parasite clearance was monitored by microscopy. Microscopy slides were obtained and then stained with Giemsa later prior to each dose of AL and at each follow-up visit on days 2, 3, 7, 14, 21, and 28. The slides were examined by certified microscopists and were considered negative in the absence of parasites after examination of 200 fields in a thick smear of blood according to MRTC standard operating procedures. The parasite density was estimated by counting the number of asexual parasites in 200 white blood cells, assuming a standard count of 8000/\u03bcL. The determination of the species (and thus the confirmation of the mono-infection) was carried out on the basis of an evaluation of the thin films. As part of quality control measures, a second independent microscopist from the project team read 10% of all smears from all visits.Plasmodium merozoite surface protein 2 genes (msp2) and microsatellite (CA1 and TA87) to discriminate reinfection from recrudescence as described previously [Dried blood spots (DBS) for Polymerase Chain Reaction (PCR) analysis were collected from each patient using 3MM Whatman\u2122 filter papers at enrolment (day 0) and at 7, 14, 21 and 28 days, on the day of treatment failure or any other unplanned visits with suspected malaria. After drying, they were stored in plastic bags containing silica gel (desiccant) and were used to distinguish the recrudescence of a new infection using the procedures recommended by WHO . For pareviously ,23. RecrStudy outcomes: The main measured outcome of the efficacy of AL was the adequate clinical and parasitological response (ACPR) at day 28 corrected by PCR. Then, early treatment failure (ETF), late clinical failure (LCF) and late parasitological failure (LPF) as defined by the WHO  were alsData management and statistical analysis: The data were recorded using electronic case report forms with the open data kits software (ODK). All survey questionnaires were captured using a digital tablet and sent to a database hosted in a server at the Malaria Research and Training Center at the University of Science, Techniques and Technologies of Bamako (USTTB), Mali. Tablets were secured by an individual password and kept under seal outside working hours. Efficacy was calculated in the protocol-treated population, which includes all patients meeting the eligibility criteria of the protocol, having completed the three-day treatment of the study drug, having adhered to all study procedures until a possible failure or at the final evaluation of 28th day. Efficacy rates were calculated by dividing the number of patients with clinical and parasitological cure at day 28 by the total number of patients that could be assessed. In addition, statistical analyses  were performed with the R version 3.4.3 software and the statistical significance level was set at 5%.Sample size calculations: Previous studies in Mali found artemether-lumefantrine efficacy of at least 95% in the different research sites ,25. The Ethical considerations: The protocol was approved by the Ethics Committee of the Faculty of Medicine and Odonto-Stomatology and Pharmacy (FMOS-FAPH) of Mali before the start of all activities (Ref N\u00b0 2017/108/CE/FMPOS) and at the amendment for the extension of the study to 2018 (Ref N\u00b0 2018/108Bis/CE/FMOS). The trial was conducted in accordance with the guidelines of good clinical practice. Written informed consent was obtained from all participants. Informed assent was also obtained from children aged 12 to 17 years.Trial profile and baseline characteristics: A total of 432 febrile patients or patients with a history of fever within 24 h were screened between July 2017 and December 2018, of whom 375 (86.8%) were recruited and 330 (95.7%) completed the study, with or without recurrent parasitemia .Plasmodium falciparum mono-infection or the presence of danger signs such as vomiting or the notion of taking other antimalarials within two weeks or during follow-up and withdrawal of consent were the main causes of exclusion. The absence of During the 28-day follow-up during the two malaria transmission seasons 2017 and 2018, 13 patients  did not successfully complete the study due to loss of follow-up, withdrawal of consent or other protocol violations such as the accidental or deliberate taking of antimalarial drugs, incoercible vomiting after inclusion or noncompliance with the visit schedule. Reported fever was the main recorded sign at the enrollment with 79.2% of the patients. This frequency decreased rapidly during the first 48 h of follow-up, to 2% or less throughout the rest of the follow-up .The treatment results by locality according to season are summarized in For both locations, 50.9% of treated patients eliminated their parasitemia at the end of the first 48 h of follow-up, while 93.1% (22/439) did it at the end of the first 60 h of follow-up, and only 0.3% did not eliminate parasites at the end of the first 72 h of follow-up .We also used a parasite clearance estimator from The Worldwide Antimalarial Resistance Network (WWARN) . The estp < 0.0001) between the study sites  who had reinfection is provided in Fourteen patients reported vomiting  during the three days of treatment in both locations . Twenty-The proportion of patients with hemoglobin below 11 g/dL at inclusion  was significantly improved on day 28 . This improvement was observed in all localities and in both seasons for the Bougouni site . HoweverThis study describes the 28-day in vivo efficacy of AL under field conditions at two NMCP sentinel sites in Mali. These two sites, one Guinean Sudanese area, the other Sahelian area, provide a good geographical representation of the variability of malaria endemicity and transmission patterns in the country. The standard protocol for evaluating the in vivo efficacy of WHO ACTs was used at both sites , includiFrom the point of view of efficacy, the cure rate corrected by PCR (>99.9%) remains high and adequate according to WHO recommendations. These results confirm those of previous studies in Mozambique and Myanmar, all of which have documented the efficacy and safety of the combination of artemether-lumefantrine ,29.The study was not designed to compare sites, so caution should be exercised in interpreting results such as late parasitological failure (LPF) levels, Es identified and the effect of treatment on hemoglobin recovery. Parasitic clearance was obtained at the sites within the first 72 h after treatment, apyrexia without analgesia was obtained 48 h after treatment as documented in 2016 in Mali , a reassDifficulties encountered were on the respect of the AL dose taking schedules, which often occurred late at night. In order to minimize the inconveniences linked to this situation, the investigators took the participants\u2019 telephone number with their agreement and called them an hour or thirty minutes in advance or went directly to the participants\u2019 homes with the help of the local guide if they encountered a problem in coming to the health center to take the doses. The same strategy was used for parasitological follow-up (every 12 h) until two consecutive negative thick blood smears were obtained. On the other hand, the 28-day follow-up time was also difficult because it was considered long and contributed to a relatively high number of people lost to follow-up.The molecular genotyping with msp1, msp2 and The Glutamate-Rich Protein (GLURP) is commonly used to distinguish recrudescence from reinfection in the monitoring of antimalarial drug efficacy ,34,35,36Plasmodium falciparum to antimalarial using the 53 episodes of recurrent parasitemia samples and the results will be published.In addition, efforts are underway to perform the parasite molecular resistance of Continuous monitoring of the in vivo efficacy of artemether-lumefantrine across the country is necessary to detect early signs of decreased efficacy of artemether-lumefantrine.Plasmodium falciparum malaria after more than a decade of use of this drug and was safe and well-tolerated. The findings of this study advocate for the continuous use of AL as first-line therapy for uncomplicated malaria in Mali. However, a genotyping of the parasite molecular resistance to antimalarial is needed to support these findings, and the monitoring of AL efficacy should continue as recommended by WHO.This study showed high efficacy of AL against"}
+{"text": "Background: Traditional medical treatments are not effective for some patients with Tourette syndrome (TS). According to the literature, repetitive transcranial magnetic stimulation (rTMS) may be effective for the treatment of TS; however, different targets show different results.Objective: To assess the efficacy and safety of low-frequency rTMS in patients with TS, with the bilateral parietal cortex as the target.Methods: Thirty patients with TS were divided into two groups: active and sham groups. The active group was subjected to 0.5-Hz rTMS at 90% of resting motor threshold (RMT) with 1,200 stimuli/day/side, whereas the sham group was subjected to 0.5-Hz rTMS at 10% of RMT with 1,200 stimuli/day/side with changes in the coil direction. Both groups were bilaterally stimulated over the parietal cortex (P3 and P4 electrode sites) for 10 consecutive days. The symptoms of tics and premonitory urges were evaluated using the Yale Global Tic Severity Scale (YGTSS), Modified Scoring Method for the Rush Video-based Tic Rating Scale (MRVBTS), and Premonitory Urge for Tics Scale (PUTS) scores at baseline, the end of the 10-day treatment, 1 week after treatment, and 1 month after treatment.Results: At the end of the 10-day treatment, the YGTSS total, YGTSS motor tic, YGTSS phonic tic, MRVBTS, and PUTS scores in the active group significantly improved and improvements were maintained for at least 1 month.Conclusions: Low-frequency bilateral rTMS of the parietal cortex can markedly alleviate motor tics, phonic tics, and premonitory urges in patients with TS. Tourette syndrome (TS) manifests as a variety of motor tics and at least one phonic tic lasting for more than 1 year . The preTranscranial magnetic stimulation (TMS) is a non-invasive technology that was developed in 1985 . RepeateThe pathological processes of tics remain unclear. The combination of neuropathology and neuroimaging findings strongly supports the conclusion that there is a cortico-striatal-thalamo-cortical (CSTC) network dysfunction in patients with TS , 22, 23.The involvement of the sensory part of the CSTC loop in TS has not been sufficiently valued. The parietal cortex can integrate different sensory information and influence the production of movement . In a fuIn the present study, we performed low-frequency rTMS in patients with TS for the following reasons. First, as mentioned earlier, before and during the onset of a tic, the parietal cortex is activated , 27. Secn = 15 each): active rTMS and sham groups.Thirty subjects with TS from the Neurology Clinic of Xuanwu Hospital were included. The subjects met the diagnosis criteria of TS stated in the fourth edition of the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV)  and had All patients or their guardians provided informed consent. This study was approved by the Ethics Committee of Xuanwu Hospital.All assessments were performed by trained raters who were blinded to the study and did not know whether the patients belonged to the active or sham groups. All assessments were completed by the same raters. All patients were assessed using the Yale Global Tic Severity Scale (YGTSS) , ModifieThe above scales were scored at baseline, the end of the 10-day treatment, 1 week after treatment, and 1 month after treatment.The patients' RMT values were measured prior to treatment. The patients were made to sit in a comfortable chair in a relaxed manner and were asked not to suppress tics. Magstim  and a figure-of-eight coil were used. The bilateral RMT of the abductor pollicis brevis was measured. The lowest intensity that produced five motor-evoked potentials (\u226550 \u03bcV) in 10 trials is defined as RMT .During the treatment, both the active and sham groups used Magstim  and a figure-of-eight coil. In the active group, the frequency was 0.5 Hz, the intensity was 90% of RMT (independent of hemisphere), and the targets were the P3 and P4 electrode sites. Stimulation comprised three trains of 400 pulses per side  for 10 consecutive days. First, the figure-of-eight coil was used to stimulate three trains  at P3 and then used to stimulate three trains at P4 . The inter-train interval was 10 min. The cortical regions underlying the P3 and P4 electrode sites in the international 10\u201320 EEG system include the Brodmann area (BA) 40, BA 7, and BA 39  for 10 consecutive days. However, the intensity in the sham group was 10% of RMT (independent of hemisphere) . The stit-test and categorical variable Fisher's exact test were used to compare demographic and clinical characteristics of the groups at baseline. The Kolmogorov\u2013Smirnov test of normality was used for the scores of each scale. The group- and time-dependent effects of rTMS on YGTSS total scores, YGTSS motor tic scores, YGTSS phonic tic scores, MRVBTS scores, and PUTS scores were evaluated via repeated-measures analysis of variance (ANOVA) with adjustments for non-sphericity. The Greenhouse\u2013Geisser correction results were used whenever necessary. A P < 0.05 was considered statistically significant.Statistical analysis was performed using SPSS version 16.0 . The statisticians were not aware of the grouping of the patients. The continuous variable All 30 patients completed the entire experiment without dropping out or missing the follow-ups. The demographic and clinical characteristics, including age, sex, course of disease, and baseline scale scores of all patients, are summarized in F = 192.555, df = 1.828, P < 0.001) as well as a significant difference between the active rTMS and sham group ; the improvement continued from the end of treatment to 1 month after treatment   as well as a significant difference between the active rTMS and sham groups , with a marked improvement from the end of treatment to 1 month after treatment   as well as a significant difference between the active rTMS and sham groups ; the improvement continued from the end of treatment to 1 month after treatment   as well as a significant difference between the active rTMS and sham group , with a marked improvement from the end of treatment to 1 month after treatment   and a significant difference between the active rTMS group and sham group ; the improvement continued from the end of treatment to 1 month after treatment   . Further< 0.001) . Moreove< 0.001) . There w< 0.001) . Lastly,< 0.001) .The results suggest that for all patients in this study, rTMS treatment was safe and well-tolerated. During the study, no patient reported any signs of headaches, seizures, memory, attention impairments, or other side effects.Our study demonstrated significant improvements in YGTSS motor tic, YGTSS phonic tic, YGTSS total, MRVBTS, and PUTS scores after parietal stimulation. The results of our study suggest that bilateral rTMS of the parietal lobe at 0.5 Hz and 90% RMT could significantly improve motor tics, phonic tics, and premonitory urges without complications. The effect was significant and was observed after 10 days of treatment and lasted for at least 1 month.As mentioned earlier, the effectiveness of rTMS treatment highly depends on the stimulation target. The bilateral parietal lobe is an effective target for rTMS stimulation. However, the underlying mechanism of bilateral rTMS of the parietal lobe used to treat patients with TS remains unclear. The parietal lobe plays a role in the generation of tics and requires targeted treatment. It is part of the association cortex that can integrate various sensations, select the most appropriate movement, and participate in final precise execution . TherefoPrevious studies have suggested that the function and structure of the parietal lobe are abnormal in patients with TS. Bohlhalter et al.  and Neun15O]H2O-positron emission tomography in patients with TS and found that for different types of tics, the activated cortex and subcortical regions were different, as were the clinical manifestations. Coprolalia was not only associated with activation in the region of the Broca's area and frontal operculum but also with the other language regions, including the posterior superior temporal gyrus and supramarginal gyrus. Motor tics were associated with activation in a region deep within the inferior parietal, sensorimotor cortex, superior temporal gyrus, and somatosensory cortex. Therefore, both motor tics and phonic tics are related to parietal lobe activity. This may be the reason for the improvements in both motor and phonic tics in this study.In this study, low-frequency bilateral rTMS of the parietal lobe improved both motor and phonic tics. Stern et al.  performeAlthough the main clinical manifestations of TS are motor tics and phonic tics, sensation plays an important role in the pathophysiology of TS. At present, the most studied sensory symptom is premonitory urges. Studies have confirmed that premonitory urges are associated with the occurrence and severity of tics. Using fMRI, Wang et al.  discoverThere were possible confounding effects of medications in our study. To minimize the confounding effects of medications, all patients remained on their usual medications at the same doses for at least 2 months. Each patient either took no medication or only one medication; the type of medicine taken was small and the dose was low. Nevertheless, we still cannot completely rule out the influence of medications because these medications may alter brain excitability and therefore have an effect on rTMS .In summary, with bilateral stimulation of the parietal lobe, 0.5-Hz rTMS is effective in patients with TS. The potential mechanism may involve the regulation of the parietal cortex activity, enhancement of parietal lobe suppression, reduction of sensory system activity, reduction of sensory motor cortex output, and disruption of tic preparation and execution.This study has some limitations. This study did not explore the effective mechanism of bilateral rTMS of the bilateral parietal cortex at 0.5 Hz for the treatment of TS. There are no neurophysiological measures; therefore, future research could use fMRI and electrophysiology to detect changes in the activity and excitability of the cortical and CSTC loops generated by rTMS. In addition, the bilateral parietal cortex could not be located using a neuronavigation system. There are individual differences in the positioning of the international EEG 10\u201320 system. It cannot be ruled out that the difference in the efficacy of patients with TS is related to differences in the precise location of the stimulation. Moreover, future studies should use a control with other stimulation sites to better illustrate the role of the parietal lobe as a stimulation target. The sample size of this study was small; therefore, future studies with larger samples are warranted to confirm our conclusions.Our research suggested that with the left and right parietal cortex as target sites, 0.5-Hz rTMS was effective and safe in the treatment of TS patients and significantly improved motor tics, phonic tics, and premonitory urges. The parietal lobe could be a new and effective target for rTMS in the treatment of TS. The mechanisms underlying the therapeutic effect may involve regulation of parietal cortex activity, enhancement of parietal lobe suppression, reduction of sensory system activity, reduction of sensory motor cortex output, and disruption of tics preparation and execution.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Xuanwu Hospital Ethics Committee, Xuanwu Hospital, Capital Medical University. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.MF, YS, and YW contributed at all stages of manuscript preparation. MF wrote the manuscript. All authors contributed to manuscript revision, read, and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest."}
+{"text": "Single Encounter Radical Cure and Prophylaxis (SERCAP) describes an ideal anti-malarial drug that cures all malaria in a single dose. This target product profile has dominated anti-malarial drug discovery and development over the past decade. The operational advantage of a single encounter has to be balanced against the need for a high dose, reliable absorption, little variability in pharmacokinetic properties, slow elimination  and a very low rate of vomiting. The demanding aspirational target may have hindered anti-malarial drug development. Aiming for three-day regimens, as in current anti-malarial treatments, would be better. Between 2008 and 2011 the Bill and Melinda Gates Foundation (BMGF) sponsored the Malaria Eradication Research Agenda  initiative. This was described as a \u201crigorous scientific consultative process to identify knowledge gaps and new tools needed to eradicate malaria globally\u201d. From this process came an ideal. \u201cThe ideal malaria eradication drug is a co-formulated drug combination suitable for mass administration that can be administered in a single encounter at infrequent intervals and that results in radical cure of all life cycle stages of all five malaria species infecting humans\u201d . No-one Malaria is currently treated with three-day regimens . The varProviding radical cure for vivax and ovale malaria adds an additional and difficult challenge. The only drugs providing radical cure are the 8-aminoquinolines, but these compounds cause haemolysis in glucose-6-phosphate dehydrogenase (G6PD) deficiency, which is common in most tropical areas. The recently registered tafenoquine is a slowly eliminated 8-aminoquinoline which does provide a single dose radical cure . However, the price of the operational advantage of the single dose is the operational disadvantage of requiring a quantitative  G6PD evaluation before treatment. This is because the slowly eliminated 8-aminoquinoline causes protracted haemolysis in G6PD deficiency, and this could be clinically significant even in female heterozygotes who may test as \u201cnormal\u201d with the current rapid G6PD screens (i.e.\u00a0the \u2018spot\u201d test or recently introduced rapid diagnostic tests) . So tafe4) or four asexual cycles (parasite reduction ratios >\u2009103) i.e., 6 or 8\u00a0days respectively [Single dose treatment has to cure >\u200995% of non-immune patients reliably without incurring toxicity. For mass treatment  it must be very safe and very well tolerated, as nearly all the recipients will be healthy and well. The current three-day artemisinin-based combination therapy (ACT) regimens are all spaced \u201cloading doses\u201d for the artemisinin partner drugs as they have to provide parasiticidal concentrations in blood for four asexual cycles (>\u20096\u00a0days) to ensure cure Fig.\u00a0. SERCAP ectively . Howeverectively . With thP. falciparum), developed by Sanofi, was selected instead. The two were co-developed in the hope of providing a single encounter cure. Unfortunately, the extended phase 2B studies conducted between 2017 and 2019 were stopped early because of poor efficacy [P. falciparum, but it also failed to provide >\u200990% cure rates in African children. The poor efficacy in African children was driven by vomiting . Overall, one third of patients vomited and, unsurprisingly, these patients had low drug levels following the single dose administration  because it was considered to be too unstable. Development switched to a more stable and slowly eliminated peroxide compound . Since then, arterolane-piperaquine has been developed by Indian manufacturers into a successful 3-day ACT . Initialefficacy . The sinThis is an unusual example having progressed so far. The influence of the SERCAP TPP is likely to have been greater at an earlier stage of drug discovery and development in lead optimization and candidate selection. There is nothing intrinsically wrong with aspiring to overcome a very difficult challenge . That is unless it prevents lesser, but still valuable, developments or improvements. We cannot say with certainty that the SERCAP TPP has been counterproductive, only that we suspect it may have been. We suggest that the TPP for new anti-malarial drugs is for a three-day regimen. If, in the future, drugs with suitable properties are discovered, and a 3-day regimen can be shortened safely without reducing efficacy, then that would be a welcome bonus."}
+{"text": "Plasmodium falciparum malaria is associated with anaemia-related morbidity, attributable to host, parasite and drug factors. We quantified the haematological response following treatment of uncomplicated P. falciparum malaria to identify the factors associated with malarial anaemia.P. falciparum malaria, available through the WorldWide Antimalarial Resistance Network data repository prior to August 2015, were pooled using standardised methodology. The haematological response over time was quantified using a multivariable linear mixed effects model with nonlinear terms for time, and the model was then used to estimate the mean haemoglobin at day of nadir and day 7. Multivariable logistic regression quantified risk factors for moderately severe anaemia (haemoglobin < 7 g/dL) at day 0, day 3 and day 7 as well as a fractional fall \u2265 25% at day 3 and day 7.Individual patient data from eligible antimalarial efficacy studies of uncomplicated p < 0.001).A total of 70,226 patients, recruited into 200 studies between 1991 and 2013, were included in the analysis: 50,859 (72.4%) enrolled in Africa, 18,451 (26.3%) in Asia and 916 (1.3%) in South America. The median haemoglobin concentration at presentation was 9.9 g/dL (range 5.0\u201319.7 g/dL) in Africa, 11.6 g/dL (range 5.0\u201320.0 g/dL) in Asia and 12.3 g/dL (range 6.9\u201317.9 g/dL) in South America. Moderately severe anaemia (Hb < 7g/dl) was present in 8.4%  of patients from Africa, 3.3%  from Asia and 0.1% (1/916) from South America. The nadir haemoglobin occurred on day 2 post treatment with a mean fall from baseline of 0.57 g/dL in Africa and 1.13 g/dL in Asia. Independent risk factors for moderately severe anaemia on day 7, in both Africa and Asia, included moderately severe anaemia at baseline (adjusted odds ratio (AOR) = 16.10 and AOR = 23.00, respectively), young age , high parasitaemia  and delayed parasite clearance . In Asia, patients treated with an artemisinin-based regimen were at significantly greater risk of moderately severe anaemia on day 7 compared to those treated with a non-artemisinin-based regimen (AOR = 2.06 [95%CI 1.39\u20133.05], P. falciparum malaria, the nadir haemoglobin occurs 2 days after starting treatment. Although artemisinin-based treatments increase the rate of parasite clearance, in Asia they are associated with a greater risk of anaemia during recovery.In patients with uncomplicated The online version contains supplementary material available at 10.1186/s12916-022-02265-9. Malaria remains a major cause of anaemia in malaria endemic countries, with a complex pathogenesis attributable to red cell destruction and haematopoietic suppression  that canP. falciparum infection and the clinical and demographic factors that underlie this. The aim of this study was to determine the pattern of haematological recovery following uncomplicated falciparum malaria and define the risk factors for moderately severe haematological outcomes at baseline and during early follow-up.To assess the comparative benefits of different antimalarial treatment regimens, it is critical to quantify the haematological impact attributable to P. falciparum transmission intensities. Data in the repository have been standardised and collated using methodology described previously in the WWARN Clinical Module Data Management and Statistical Analysis Plan [Haemoglobin concentrations are often not reported in antimalarial trial publications, even if these data are collected. Since a review of published literature would not provide sufficiently comprehensive information, the focus of this individual patient data meta-analysis was on studies identified in the WWARN repository. The WWARN repository contains data from 451 antimalarial efficacy studies in which patients were enrolled from locations in 69 countries, with a diverse range of P. falciparum malaria in non-pregnant patients that followed subjects prospectively for a minimum of 28 days and reported haemoglobin concentration (or haematocrit) at least at baseline (day 0). Investigators of the identified studies were invited to participate in this study group and information was made available on the WWARN website [P. falciparum malaria was defined as microscopy-proven falciparum malaria without features of severe malaria [The WWARN repository was searched in August 2015 for all antimalarial efficacy studies of uncomplicated  website . Uncompl malaria . PatientThe primary outcome of the analysis was the risk of moderately severe anaemia (Hb < 7 g/dL) on day 7 after initiation of treatment. Secondary outcomes included the mean fall in haemoglobin at day of nadir and day 7, the timing of nadir haemoglobin, risk of moderately severe anaemia at days 0 and 3, and the risk of a large reduction in haemoglobin from baseline, defined as a fractional fall in Hb of \u2265 25% on day 3 or 7.All statistical analyses were done using R  or Stata MP 15, based on an a priori statistical plan shared with data contributors .Haematocrit measurements were converted to haemoglobin concentrations using the following formula: Haemoglobin = (Haematocrit \u2212 5.62)/2.60 . The timChanges in mean haemoglobin over time were examined, after stratifying by region, using linear mixed effects models. Fractional polynomial terms for time were fitted as random effects for patients to capture the nonlinear associations and random intercepts for patients and study site. All available haemoglobin measurements were included in these analyses. Additional analyses of mean haemoglobin over time within each region were undertaken, stratified by age group (< 5 years and \u2265 5 years).Plasmodium species infection, underweight (defined as weight-for-age Z-score <\u22122 for children younger than 5 years) [P. falciparum prevalence rate (PfPR) according to enrolment year and location [For all regression models, independent risk factors were identified following the strategy recommended by Collet . Covaria5 years) , high pa5 years) ), presenlocation , assuminAll data included in this analysis were obtained in accordance with ethical approvals from the countries of origin. The data are fully anonymised and cannot be traced back to individuals. This analysis did not require separate ethical approval according to the guidelines of the Oxford Central University Research Ethics Committee.P. falciparum clinical trials undertaken between 1991 and 2013 met the inclusion criteria and were available for analysis , with 2 studies (483 patients) excluding patients with a haemoglobin < 6 g/dL, 9 studies (5964 patients) excluding patients with a haemoglobin < 7 g/dL and 3 studies (566 patients) excluding patients with a haemoglobin < 8 g/dL. In 60 studies, haematological exclusion criteria were not stated; Additional file The median haemoglobin at enrolment was 9.9 g/dL (range 5.0\u201319.7 g/dL) in Africa, 11.6 g/dL (range 5.0\u201320.0 g/dL) in Asia and 12.3 g/dL (range 6.9\u201317.9 g/dL) in South America  and day 7  versus 2.3%  respectively; p < 0.001).Overall, 9.1%  of patients had moderately severe anaemia (Hb < 7 g/dl) on day 3 and 4.4%  on day 7. The risk of falling below 7 g/dL in people who did not have moderately severe anaemia at baseline was greater in Africa than in Asia on both day 3  versus 6.5% ; p < 0.001 for Africa and 23.00 (14.27\u201337.06), p < 0.001 for Asia), younger age (for age < 1 year compared to patients \u2265 12 years AOR = 12.81 (95%CI 6.79\u201324.17), p < 0.001 and AOR = 6.79 (95%CI 2.36\u201319.58), p < 0.001 and for age 1 to 4 years compared to patients \u2265 12 years AOR = 6.09 (95%CI 3.33\u201311.13), p < 0.001 and AOR = 2.87 (95%CI 1.84\u20134.47), p < 0.001) and parasitaemia > 100,000/\u03bcL (AOR = 1.78 (95%CI 1.42\u20132.24), p < 0.001 and 1.58 (1.10\u20132.27), p = 0.013) (Table p < 0.001) was an independent predictor of moderately severe anaemia whilst female sex (AOR = 0.80 (95%CI 0.69\u20130.93); p = 0.004) was protective. In Asia, female sex was an independent predictor (AOR = 1.51 (95%CI 1.15\u20131.99), p = 0.003) and mixed infection was protective (AOR = 0.44 (95%CI 0.24\u20130.80), p = 0.007). The effect of sex was explored by evaluating the models separately in children compared with adolescents and adults. In adolescents and adults (age \u2265 12 years), female sex was associated with moderately severe anaemia, and this reached statistical significance in Asia (AOR = 2.55 (95%CI 1.65\u20133.94), p < 0.001), but not Africa (AOR = 1.98 (95%CI 0.71\u20135.49), p = 0.191). In contrast, in children (age < 12 years) female sex was associated with a lower risk of moderately severe anaemia, reaching statistical significance in Africa (AOR = 0.78 (95%CI 0.67\u20130.91), p = 0.001), but not Asia (AOR = 0.98 (95%CI 0.73\u20131.32), p = 0.903).The following independent predictors of moderately severe anaemia at day 7 in both Africa and Asia were identified: moderately severe anaemia at baseline (AOR = 16.10 95%CI 12.59\u201320.60), .59\u201320.60d 23.00 1.27\u201337.06p < 0.001), but this was not the case in African patients (AOR = 1.01 (95%CI 0.56\u20131.82); p = 0.987). In Asia, the difference in risk between artemisinin- and non-artemisinin-based treatments remained when only the 8570 patients enrolled before 2007 (when artemisinin resistance was first described in the Greater Mekong Subregion) were included in the model (AOR = 1.98 (95%CI 1.39\u20132.82); p < 0.001). Overall, the risk factors for moderately severe anaemia on day 3 were similar to those at day 7, with patients treated with artemisinin-based therapy at significantly greater risk of moderately severe anaemia on day 3 in Asia (AOR = 3.27 (95%CI 2.42\u20134.42); p < 0.001), but not in Africa (AOR = 0.69 (95%CI 0.32\u20131.49); p = 0.343) ; 9\u20133.05; pr = 0.47; p < 0.001 adjusted for clustering of study site). A high baseline haemoglobin was associated with a greater risk of a large fractional fall (\u2265 25%) on day 7 in Africa (AOR for every 1 g/dL increase in baseline haemoglobin = 1.52 (95%CI 1.40\u20131.65); p < 0.001) and in Asia (AOR for every 1 g/dL increase in baseline haemoglobin = 1.43 (95%CI 1.35\u20131.52); p < 0.001)  and Asia (AOR = 2.59 (95%CI 1.20\u20135.58); p = 0.015)  were parasitaemic on day 2 and 2.6% (358) were parasitaemic on day 3. The corresponding proportions in Asia were 14.9% (1339/8960) and 4.2% (375/8960). After controlling for confounding factors, the risk of moderately severe anaemia at day 7 was greater in patients with delayed parasite clearance in both Africa (AOR = 2.44 (95%CI 1.59\u20133.75); Methodological factors potentially contributing to bias are presented in Additional file Our study provides a detailed analysis of haemoglobin concentration kinetics in patients with falciparum malaria, enrolled across geographically diverse regions. The available data, exceeding > 70,000 individual data from patients of all ages, provides unprecedented power to define the factors associated with the acute fall in haemoglobin before and after treatment. Malaria is due to an intraerythrocytic infection which results in a reduction of red blood cells, intra- and extravascular haemolysis, bone marrow suppression and sequestration . The admAlthough the greatest fall in haemoglobin occurred before treatment\u00a0in Africa, our analysis focused primarily on factors associated with the subsequent fall and recovery which may\u00a0be more amenable to clinical intervention. Consistent with a recent pooled analysis from Africa , our stuThe baseline haemoglobin in patients with falciparum malaria varied substantially with age and parasite density at presentation. After controlling for confounding factors, significant site to site variation remained, likely reflecting variations in transmission intensity, host immunity and factors unrelated to malaria. Patients from Asia tended to be older than those enrolled in Africa, but after controlling for age there were minimal differences in haemoglobin between regions, either at baseline or during follow-up. Following treatment, the absolute and proportional reductions in haemoglobin were greater in patients from Asia compared to Africa and were correlated with the higher baseline haemoglobin in Asian patients. Hence, patients presenting with a low haemoglobin concentration were less likely to experience a further fall in their haemoglobin.Plasmodium parasitaemia in these regions will have an alternative diagnosis, such as bacterial sepsis, which is also associated with anaemia [The relationship between level of parasitaemia and degree of anaemia is complex , 179. In anaemia . Second, anaemia , 181.In Asia, the risk of anaemia at presentation increased with rising parasitaemia, peaking at 10,000 parasites/\u03bcL before decreasing thereafter. As transmission intensity in endemic parts of Asia is generally significantly lower than in Africa, immunity is less robust and a much greater proportion of infections will be symptomatic and of short duration. In this setting, anaemia will be related primarily to acute destruction of both parasitised and unparasitised red cells, the severity of which is correlated with the level of parasitaemia.Treatment with artemisinin-based therapy in Asia was associated with a twofold higher risk of moderately severe anaemia  within 7 days compared with non-artemisinin-based therapy, whereas in Africa, artemisinin-based treatment was not associated with an excess risk of early anaemia. This relationship was not attributable to the presence of artemisinin resistance. We hypothesise that rapid killing of intraerythrocytic parasites by artemisinins in non-immune Asian adults likely leads to more rapid clearance of whole red blood cells from the circulation than that occurring after slower acting drug treatments. In immune African patients, a greater proportion of infected red cells undergo targeted intraerythrocytic parasite removal (pitting) followed by a return to circulation, thus ameliorating the early development of anaemia . ReticulOur study has a number of limitations. The analysis focused on the acute haematological impact of malaria and the early recovery phase and did not address the influence of late treatment failure on subsequent recovery to baseline haemoglobin concentrations. This will be addressed in a subsequent analysis. Our estimates of the pattern of haemoglobin changes during the first few days after diagnosis may have been influenced by selection bias, as only a small subset of patients had multiple haemoglobin measurements during the first 7 days of follow-up. Although we did not employ a traditional systematic review to identify eligible studies, our analysis is the largest meta-analysis to date of patients treated for malaria in both Africa and Asia. This unprecedented data collection ensures robust parameter estimates and minimises the risk of inclusion bias. Furthermore, a systematic review would not preclude bias, since some studies recorded haemoglobin/haematocrit measurements but did not present these data in published manuscripts. Whilst the results of the current study are likely to be generalisable to Africa and Asia, the small number of patients from the Americas prevents the generalisability of our findings to this region. An additional potential cause of bias is the exclusion of patients from the original studies, prior to pooling, according to variable definitions of severe anaemia. Almost two-thirds of studies excluded patients with a haemoglobin < 5 g/dL, with a few studies excluding patients based on higher cut-offs and the remaining 30% having an unknown cut-off. Additional limitations of our study include the use of various methodologies to measure haematocrit or haemoglobin, a lack of a robust conversion factor to adjust haematocrit to haemoglobin in different studies\u2019 populations and no reliable data on the following confounding factors that can influence haemoglobin and its recovery: the duration of prior parasitaemia (which has been shown to correlate with anaemia at presentation ), host gP. falciparum infection.In conclusion, the majority of patients with uncomplicated falciparum malaria had a modest fall in haemoglobin following treatment, before subsequent improvement in haemoglobin during recovery. Despite highly effective treatment, some patients remained at significant risk of moderately severe anaemia. Young children had a particularly high risk, likely related to lower immunity and high initial peripheral parasitaemia. The risk of anaemia is exacerbated by prolonged parasitaemia prior to presentation  or delayAdditional file 1: Table S1. Describes studies included in the analysis. Table S2. Describes assessment of bias by included study.Additional file 2: Figure S1. Describes study sites. Figure S2. Describes the relationship between haemoglobin on enrolment and continuous covariates. Figure S3. Describes the relationship between the predicted probability of moderately severe anaemia on day 3 and continuous covariates. Figure S4. Describes the relationship between the predicted probability of a large fractional fall in haemoglobin on day 7 and continuous covariates.Additional file 3: Table S3. Describes the overview of antimalarial treatments. Table S4. Describes the risk factors for moderately severe anaemia at enrolment (univariable logistic regression). Table S5. Describes the risk factors for moderately severe anaemia at enrolment (multivariable logistic regression). Table S6. Describes the risk factors for moderately severe anaemia at day 7 (univariable logistic regression). Table S7. Describes the risk factors for moderately severe anaemia at day 3 (univariable logistic regression). Table S8. Describes the risk factors for moderately severe anaemia at day 3 (multivariable logistic regression). Table S9. Describes the risk factors for a large fractional fall in haemoglobin by day 7. Table S10. Describes the sensitivity analysis for risk factors for moderately severe anaemia at enrolment (multivariable logistic regression). Table S11. Describes the sensitivity analysis for risk factors for moderately severe anaemia at day 7 (multivariable logistic regression)."}
+{"text": "In the control group of second malaria attacks, the difference between the two groups was not statistically significant ; there was no significant difference in treatment failure during follow-up . There were also very few serious adverse events in both groups. ACTs showed good therapeutic effects in preventing gametocythemia but did not control the recrudescence rate and overall cure, which indicated the effectiveness of the combination of antimalarial drugs. Further research is required to explore which compatibility method is most conducive to the development of clinical malaria control.The WHO recommends Artemisinin-based combination therapy (ACTs) as the first-line treatment for malaria. This meta-analysis aims to analyze the effects of artemisinin and its derivatives as well as non-artemisinin drugs on the gametophytes in the host during the treatment of falciparum malaria. Fourteen studies were included in this analysis, and the artemisinin combination drugs involved were: artemether-lumefantrine (AL), artemisinin (AST), artemether-benflumetol (AB), dihydroartemisinin-piperaquine + trimethoprim + primaquine (CV8), amodiaquine + sulfadoxine-pyrimethamine (ASP), pyronaridine-phosphate + dihydroartemisinin (PP-DHA), dihydroartemisinin (DHA), and mefloquine + artesunate (MA), with 1702 patients. The control intervention measures involved the following: sulfadoxine-pyrimethamine (SP), mefloquine (MQ), atovaquone-proguanil (AT-PG), chloroquine + sulfadoxine-pyrimethamine (C-SP), quinine (Q), pyronaridine-phosphate (PP), pyronaridine (PN), and mefloquine + primaquine (MP), with 833 patients. The effect of ACTs was more obvious (OR = 0.37, 95%CI: 0.22\u20130.62,  Plasmodium falciparum (P.f)\u2014mainly causes severe malaria, Plasmodium vivax, Plasmodium malariae, Plasmodium ovale, and Plasmodium knowlesi , Chinese Biomedical Database (CBM), and Cochrane Library. The search keywords were \u201cuncomplicated falciparum malaria,\u201d \u201cgametophyte,\u201d and \u201carteether OR dihydroartemisinin OR artemether OR artemisinin OR artesunate OR artemether-benflumetol OR artemether-lumefantrine.\u201d We included trials that compared artemisinin combination drugs with non-artemisinin antimalarial drugs, and subsequent references were analyzed to find potentially eligible literature.P falciparum/\u00b5L; 2) Fever \u226537.5\u00b0C or a history of fever in the past 3\u00a0days; 3) The onset of malaria did not exceed 1\u00a0week. The exclusion criteria were as follows: 1) Patients with symptoms of severe malaria; 2) Pregnancy or breastfeeding patients; 3) Patients who had consumed antimalarial drugs and antibiotics in the past week or had an antimalarial history in the past 2\u00a0weeks; 4) Patients who had been infected with other malarial parasites; 5) Patients with severe malnutrition or a history of chronic diseases of the heart, liver, and kidney; 6) Patients with allergic reactions to antimalarial drugs.All patients infected with falciparum malaria without other complications were included in this study. There are no special requirements for age and gender. The inclusion criteria for patients were as follows: 1) Parasitemia \u22655000\u00a0This study used ACTs as the main intervention measure, including artemether-lumefantrine (AL), artemisinin (AST), artemether-benflumetol (AB), dihydroartemisinin-piperaquine + trimethoprim + primaquine (CV8), amodiaquine + sulfadoxine-pyrimethamine (ASP), pyronaridine-phosphate + dihydroartemisinin (PP-DHA), dihydroartemisinin (DHA), and mefloquine + artesunate (MA). Although the drug brands used in the included studies were different, all drugs were manufactured by regular manufacturers with product numbers and were used in strict accordance with the recommended drug dosage.The non-artemisinin antimalarial drugs (nACTs) were treated as the control group, including sulfadoxine-pyrimethamine (SP), mefloquine (MQ), atovaquone-proguanil (AT-PG), chloroquine + sulfadoxine-pyrimethamine (C -SP), quinine (Q), pyronaridine-phosphate (PP), pyronaridine (PN) and mefloquine + primaquine (MP). Similar to the intervention group, the dosage, and brand of the drugs were not required to be the same, but all drugs were manufactured by regular manufacturers with product numbers and were used in strict accordance with the recommended drug dosage.The transmission rate of falciparum malaria is known to be related to many factors. The primary endpoint of this study was the rate of patients with gametophytes after the completion of drug administration, and the secondary endpoints were the treatment failure rate and the rate of second malaria attacks. Gametophyte clearance was defined as follows: when under the microscope, no gametophyte (\u2265500 white blood cells) was found in the thick blood film. Clinical treatment failure was defined as follows: after the end of the treatment course, if the protozoan asexual body was still detected on the thick blood smear twice, the treatment was deemed invalid. The second episode of malaria was defined as follows: the second episode, including new infections and recurrences, i.e., the patient after the peripheral blood smear was identified as negative , and parasitemia was again observed under the microscope during the follow-up period.Randomized controlled trials (RCTs) evaluating ACTs and nACTs were included in this meta-analysis. We also included references, abstracts, and conference reports that provided relevant data.Two independent examiners screened the titles, abstracts, and full texts generated by the electronic database search. Any differences of opinion arising during the period were resolved by mutual consensus. The reviewers collected the following information from each included study: first author, year of publication, age and characteristics of participants, confirmation of malaria, the dosage of experimental drugs used, the sample size of the intervention group and the comparison group, duration of treatment, the number of patients with gametophytes in their bodies, clinical cure rates, recurrence rates, adverse events, and follow-up time. \u201cClinical cure rates\u201d was the 28-days parasitological cure rate. Which describes the proportion of patients with clearance of asexual parasitaemia within 7\u00a0days of initiating study treatment without recrudescence at day 28, based on blood smears. \u201cRecurrence rate\u201d refered to the proportion of the recurrence of asexual parasitaemia after treatment of the infection with the same infection that caused the original illness. This results from incomplete clearance of parasitaemia due to inadequate or ineffective treatment. It differs from a new infection or re-infection (as identified by molecular genotyping in endemic areas). If necessary, the original author was contacted for more information.The Cochrane System Deviation Risk Assessment Tool was used to assess the quality and deviation risk of the included clinical trials. The assessment tool specifically included the following seven aspects: 1) whether it was a generated random sequence; 2) whether it was randomly assigned; 3) whether the participants and personnel were double-blinded; 4) whether the result evaluation was blinded; 5) whether the result data were complete; 6) whether the report was selective; 7) whether there were other biases . The qua2 test and the I2 test were used to evaluate the statistical heterogeneity between the combined experiments .The analysis results of the primary and secondary endpoints were expressed using relative risk and 95% confidence interval (CI). The \u03c7eriments . If I2 wThis meta-analysis included 14 RCTs. The Cochrane bias risk assessment tool was used to evaluate the methodological quality of the included studies. The evaluation method included a comprehensive evaluation of the following seven aspects: 1) whether the random sequence was generated; 2) whether the allocation was hidden; 3) whether the result evaluation was blinded; 4) whether the patients and researchers were blinded; 5) whether the incomplete result data was processed; 6) whether the data was deliberately selected for reporting; 7) whether other potential biases existed. All evaluations were derived from the original text of each study. For the original text without a clear description, we evaluated it as low-risk or unclear based on the content of the full text. Based on these seven evaluation criteria, five or more studies were found to be low risk. We classified them as \u201clow risk of bias\u201d trials. The 12 experimental studies included were all low risk of bias .14 Since the follow-up period of this study was 28\u00a0days. We divided it into the 28th day gamete appearance rate. Based on the analysis results, we found that on the third day of treatment, the difference between ACTs and nACTs was not statistically significant . However, after 1\u00a0week of treatment, there was a significant difference in the gametophyte carrying between the two groups of patients . At the second week of follow-up, the difference between ACTs and nACTs became insignificant . On the 28th day of follow-up, there was a statistically significant difference between ACTs and nACTs . Thus, we found that there was a statistically significant difference between ACTs and nACTs . . .p < 0.001). After correction, the overall effect size was still statistically significant; thus, the possibility of publication bias was ruled out. Sensitivity analysis was conducted on 11 studies that reported the rate of gametophyte carrying conditions after 7\u00a0days of treatment, and the results showed that good stability of the analysis results  (We use funnel plots to determine the publication bias of the primary endpoint data. There was one study on the left side of the funnel chart and two studies on the right side . Since t< 0.001) .2 = 76% > 40%, a random-effects model was used, and the analysis result obtained showed the absence of significant difference between the two groups of ACTs and nACTs in the efficacy of curing patients . . .p > 0.05). The results showed the absence of a statistically significant difference. In the sensitivity analysis , we adopted a random-effects model, and the results showed that there was no significant difference in the recurrence rate between the two groups   was reported . The rem < 0.01) .p = 0.6082 > 0.05). When analyzing the sensitivity , we found that the results of one study were not stable  is a 4-aminoquinoline drug that mainly acts on the large and small loops and trophozoites of mature malaria parasites. It has a heat-clearing effect, and the activity has early specificity. Although the protozoan clearance rate is slower than that of artemisinin derivatives, it is well-tolerated, and its combination with SP significantly improves the fever clearance time and protozoan clearance time. Also, a study found that the initial response of C-SP was significantly better than that of AL . HoweverAmodiaquine (A) combined with SP has been found to be more effective than AL in reducing the overall incidence of malaria after treatment. Also, AL has a little protective effect on malaria recurrence after treatment, while ASP has a good protective effect on new infections .Quinine (Q) is the first proven antimalarial drug and is administered for approximately 7 days. It is well known that it has poor tolerance and poor compliance , and a sP. vivax to prevent recurrence and acts on the gametophytes of P. falciparum to prevent its spread. It inhibits all activities of Plasmodium in the liver stage, which is the primary basis of preventive drugs. PQ interferes with heme polymerization (Primaquine (PQ) is an 8-aminoquinoline drug with antimalarial activity similar to other 4-aminoquinoline drugs. It acts on the dormant seeds of rization , and comrization .P. falciparum  is a relatively slow-acting anti-schizophrenia drug with a half-life of 4\u20136\u00a0days . Long haThe mechanism of action of mefloquine (MQ) is similar to that of Q. It acts on falciparum malaria with multidrug resistance. It is an effective plasmodium asexual killer but has no direct effect on gametophytes. The gametophyte carrying rate is high after a large single dose . AlthougP. falciparum.Pyronaridine (PN) is a highly effective and low-toxicity red endogenous schizont killer. It has a short curative effect with a low re-combustion rate after treatment, but it has no effect on the gametophytes in the body, and the time to clear the patient\u2019s fever is slightly longer. Also, the gametophyte rate after single drug use is high . PyronarHere, we divided the treatment methods into ACTs and nACTs. In terms of the overall cure rate, since we adopted an intention-to-treat method, the loss of people involved in the evaluation during the follow-up period was relatively large, which had a relatively large impact on the overall results . AdditioHowever, the results of the primary endpoint showed that, compared with previous non-artemisinin-based first-line antimalarials, ACTs could significantly reduce the body\u2019s gametocytes, although they could not completely prevent the second malaria attack. As more and more areas use ACTs as the primary antimalarial drug, and families have adopted a variety of defensive measures, such as using soaked mosquito nets to eliminate nearby mosquito breeding sites, etc., the transmission capacity of malaria might be effectively controlled.As a parasitic infectious disease, malaria has affected humans for centuries. Although the cause of the disease might be insignificant, the consequent problem is big. About half of the world\u2019s population is at risk of developing malaria, and most of these infections occur in sub-Saharan Africa. With the development of drug resistance and considering the economic impact, antimalarial drugs have gradually changed from nACTs to ACTs.We systematically analyzed the effects of ACTs and nACTs on host gametophytes during the treatment of falciparum malaria, the failure rate of clinical treatment, and the rate of secondary malaria attacks, which are considered essential for judging the efficacy of drugs. The analysis results showed that compared with the previous non-artemisinin first-line antimalarial drugs, although ACTs did not show a better trend in preventing secondary attacks and clinical cure rates. But ACTs could significantly reduce the body\u2019s gametocytosis. It is a work which deepened the understanding mechanism of mediating malaria parasite human-mosquito-human transmission, and offered great potential for new targeted interventions to block parasite transmission from humans to mosquitoes to aid the elimination of malaria. And it was essential for controlling the further spread of malaria and reducing the overall incidence of an area.Influencing factors on the transmission of malaria are not only related to the carrying density and time of gametophytes in the body but also to the sex ratio of gametophytes in the peripheral blood and the infectivity of protozoa to mosquitoes . Therefo"}
+{"text": "Naturally acquired immunity to malaria is incompletely understood. We used controlled human malaria infection (CHMI) to study the impact of past exposure on malaria in Kenyan adults in relation to infection with a non-Kenyan parasite strain.3 aseptic, purified, cryopreserved Plasmodium falciparum sporozoites  by direct venous inoculation and undertook clinical monitoring and serial quantitative PCR (qPCR) of the 18S ribosomal RNA gene. The study endpoint was met when parasitemia reached 500 or more parasites per \u03bcL blood, clinically important symptoms were seen, or at 21 days after inoculation. All volunteers received antimalarial drug treatment upon meeting the endpoint.We administered 3.2 \u00d7 10One hundred and sixty-one volunteers underwent CHMI between August 4, 2016, and February 14, 2018. CHMI was well tolerated, with no severe or serious adverse events. Nineteen volunteers (11.8%) were excluded from the analysis based on detection of antimalarial drugs above the minimal inhibitory concentration or parasites genotyped as non-NF54. Of the 142 volunteers who were eligible for analysis, 26 (18.3%) had febrile symptoms and were treated; 30 (21.1%) reached 500 or more parasites per \u03bcL and were treated; 53 (37.3%) had parasitemia without meeting thresholds for treatment; and 33 (23.2%) remained qPCR negative.We found that past exposure to malaria, as evidenced by location of residence, in some Kenyan adults can completely suppress in vivo growth of a parasite strain originating from outside Kenya.ClinicalTrials.gov NCT02739763.Wellcome Trust. Plasmodium falciparum malaria following repeated exposure , administered by needle and syringe has facilitated CHMI studies among nonendemic and endemic populations  have been enrolled in published CHMI studies using Sanaria PfSPZ Challenge to either assess vaccine efficacy or infectivity , 10\u201312. For endpoint assessment in CHMI studies, the conventional method for monitoring parasitemia has been thick-blood smear microscopy, which does not detect low-density parasitemia . MoleculTo systematically explore qPCR outcomes following CHMI in semi-immune adults, we conducted the largest CHMI study to date, recruiting volunteers with previous exposure to malaria from specific rural areas in Kenya. As adults in malaria-endemic regions of Kenya are frequently found to be asymptomatic despite significant blood parasitemia , we adopThe aim of this study was to investigate how the in vivo parasite growth in CHMI would be modified by preexisting immunity, where qPCR was used to quantify parasite growth and PfNF54 parasites were used for challenge. NF54 is a laboratory-adapted parasite line that is of West African origin and is genetically distinct from East African parasites . Moser en = 37), February 2017 (n = 64), and February 2018 (n = 60), representative of volunteers from Ahero (n = 15), Kilifi North (n = 34), and Kilifi South (n = 112). All 161 volunteers were inoculated with 3.2 \u00d7 103 PfSPZ of Sanaria PfSPZ Challenge NF54 by direct venous inoculation (DVI) and monitored for outcomes by clinical assessment and qPCR. All volunteers completed CHMI and were successfully treated and discharged from inpatient care (https://doi.org/10.1172/jci.insight.146443DS1) and were excluded from further analysis; a further 12 volunteers were excluded on the basis of antimalarial drug concentrations  the first 6 days from day of administration of PfSPZ (days 1\u20137), when there were few grade 1 local adverse events relating to the injection site and some isolated unsolicited events; (b) day 8 after administration of PfSPZ up to the day of treatment, when systemic febrile events  became more common; and (c) the day of treatment to the exit from in-patient stay, when systemic febrile adverse events were most common . UnsolicThe most common abnormalities on safety blood tests during CHMI were grade 1 and 2 elevated alanine aminotransferase (ALT) and creatinine values, all of which resolved to within the normal range during the follow-up period . The cutP = 0.25; Antimalarial drugs were detected in some volunteers . Two volr = 0.93, P < 0.0001).One volunteer had high concentrations of both pyrimethamine and sulfadoxine and did not meet criteria for treatment. Two further volunteers had detectable concentrations of sulfadoxine without pyrimethamine, and both met criteria for treatment before day 22. There was no detectable artemether or dihydroartemisinin in any sample. The prevalence of detectable lumefantrine concentrations was higher in Kilifi South  compared with that in Kilifi North . Lumefantrine concentrations measured at the 2 independent laboratories were closely correlated , 26 (18.3%) had febrile symptoms and were treated, 30 (21.1%) reached 500 or more parasites per \u03bcL and were treated, 53 37.3%) had parasitemia without meeting thresholds for treatment, and 33 (23.2%) remained qPCR negative, 10 of these with no detectable antimalarial drug levels . Of the .3% had pIn contrast, in the high transmission locations of Kilifi South and Ahero it was more common to observe volunteers that experienced limited parasite growth or with negative qPCR results. The presence of fever was more common in Kilifi North than in Kilifi South or in Ahero. Fever was less frequent in the Nairobi study, in which treatment was given based on microscopy . Similar results were seen after excluding all volunteers with detectable lumefantrine concentrations .r = 0.65, P < 0.0001) and UW  and between MU and UW assays , were shared with 2 external laboratories at Mahidol University (MU), Thailand, and at the University of Washington (UW), Seattle, Washington, USA, for their analytically sensitive qPCR and reverse-transcription PCR (RT-PCR) assays, respectively , 16, 22.Z = 0.79, P = 0.43; Finally, we reanalyzed the ALT elevations observed after CHMI according to qPCR outcomes and found that ALT elevations did not vary according to qPCR status. Volunteers who were qPCR negative for malaria parasites following CHMI had a median ALT of 46 IU/L (IQR 27\u2013120 IU/L) at day 10, compared with a median ALT of 43 IU/L (IQR 26\u201380 IU/L) among those who were qPCR positive . It is likely that prior malaria exposure rather than human genetic differences explains the observed variations in CHMI outcomes. In relation to more recently published CHMI studies in endemic populations , 10, 11,P. falciparum . Volunteers were recruited from differing malaria-endemic regions in Kenya: Ahero in Western Kenya (moderate-to-high transmission at community age\u2013adjusted parasite rates [PfPR] of 40%); Kilifi South ; and Kilifi North on the Kenyan Coast , 75,000 (n = 4), or 25,000 (n = 4) PfSPZ. In the Kilifi study, real-time qPCR results were used for malaria diagnosis, whereas criteria for malaria diagnosis in Nairobi were based on thick-blood film microscopy with retrospective analysis of qPCR results. A positive thick film corresponds to a qPCR quantification of approximately 50 parasites per \u03bcL  parasitemia reached 500 parasites per \u03bcL; (b) clinically important signs and/or symptoms were observed, with any evidence of malaria parasites by blood film positivity; or (c) at day 22 (21 days after infection). For criteria b, thick- and thin-blood films were prepared when requested by the assessing clinicians and made from whole blood by experienced microscopists who examined 100 high-power fields. In Nairobi, endpoint criteria were met when thick-blood film microscopy was positive with retrospective analysis of qPCR results. A positive thick film corresponds to a qPCR quantification of approximately 50 parasites per \u03bcL .18S ribosomal RNA P. falciparum gene, was used in real time where 500 \u03bcL whole venous blood was collected twice every day from day 8 to day 15 after infection and then once every day from day 16 to day 22 after infection  and 8 days after challenge (C+8). Plasma samples were sent to 2 independent laboratories to determine plasma concentrations, Strathmore University, Nairobi, Kenya, and Mahidol Oxford Tropical Medicine Research Unit (MORU), Bangkok, Thailand. Artemether and dihydroartemisinin concentrations were measured at MORU; sulfadoxine, pyrimethamine, and chloroquine at Strathmore University; and lumefantrine and desbutyl-lumefantrine at both laboratories. Antimalarial drug concentrations were measured by liquid chromatography\u2013tandem mass spectrometry using previously published procedures  and adapMSP2 genotyping and antimalarial drug detection was carried out. MSP2 genotyping was performed on blood samples collected on the day of treatment initiation for all volunteers regardless of parasite outcome during CHMI. This was done by a nested PCR, with amplification specific for FC27 and IC/3D7 allelic types, with fragment size analysis by capillary electrophoresis as previously described .r2 value of 0.3, with a power of 80% to detect a single variable to account for 0.15 of the variability in parasite growth. Nonparametric correlation test and Spearman\u2019s rank correlation were used for comparisons. All analyses were performed using STATA . All tests were performed at 5% significance level, where P < 0.05 was considered significant.The sample size for the study was based on considering analysis of outcome of CHMI in a multivariable model considering parameters other than PCR outcome, assuming an The study was conducted at the Kenya Medical Research Institute-Wellcome Trust Research Programme in Kilifi, Kenya, and received ethical approval from the Kenya Medical Research Institute Scientific and Ethics Review Unit (KEMRI/SERU/CGMR-C/029/3190) and the University of Oxford Tropical Research Ethics Committee (OxTREC 2-16). The study is registered on ClinicalTrials.gov (NCT02739763) and was conducted based on good clinical practice and under the principles of the Declaration of Helsinki.MCK, PFB, and CHMI-SIKA Study Team members contributed to the study design. Protocol development was performed by MCK, PN, and PFB. Conduct of the clinical trial was performed by PN, MH, JM, ON, and CHMI-SIKA Study Team members. Experiments and data acquisition were conducted by MCK, DK, JMN, and CHMI-SIKA Study Team members. EO and CHMI-SIKA Study Team members acquired and curated data. CHMI-SIKA Study Team members were involved in manufacture of Sanaria PfSPZ Challenge. MCK, EO, and PFB analyzed the data. MCK and PFB wrote the manuscript. All the members of the CHMI-SIKA study team read and reviewed the manuscript."}
+{"text": "Plasmodium falciparum gametocytes. Primaquine is metabolized by a highly polymorphic cytochrome P450 2D6 (CYP2D6)\u00a0enzyme. Mutations in the gene encoding this enzyme may lead to impaired primaquine activity. This study assessed if 0.25\u00a0mg/kg single-dose primaquine is safe and sufficient to reduce transmission of gametocytes in individuals with no, reduced, or increased CYP2D6 enzyme activity.Primaquine is a pro-drug and its active metabolite is potent against mature P. falciparum malaria were enrolled in the study. The enrolled patients were treated with a standard artemether-lumefantrine regimen plus 0.25\u00a0mg/kg single-dose primaquine and followed up for 28\u00a0days for clinical and laboratory assessment. Primaquine was administered with the first dose of artemether-lumefantrine. Safety assessment involved direct questioning and recording of the nature and incidence of clinical signs and symptoms, and measurement of haemoglobin (Hb) concentration. Blood samples collected from 100 patients were used for assessment of post-treatment infectiousness on day 7 using mosquito membrane feeding assays. Molecular methods were used to determine CYP2D6 and glucose-6-phosphate dehydrogenase (G6PD) status. The primary outcome was the safety of 0.25\u00a0mg/kg single-dose primaquine based on CYP2D6 status.Between June 2019 and January 2020 children aged 1\u201310\u00a0years, attending at Yombo dispensary, Bagamoyo district, with confirmed microcopy-determined uncomplicated t\u2009=\u20090.012, p\u2009=\u20090.990). The day 3 mean absolute Hb concentration reduction in G6PD deficient, G6PD normal and heterozygous female was 1.82\u00a0g/dL (95% CI 1.32\u20132.32), 1.48\u00a0g/dL (95% CI 1.30\u20131.67) and 1.47\u00a0g/dL (95% CI 0.76\u20132.18), respectively . Sixteen percent (16/98) of the patients each infected at least one mosquito on day 7, and of these, 10.0% (2/20) and 17.9% (14/78) had reduced and normal CYP2D6 enzyme activity, respectively .In total, 157 children [median age 6.4 (Interquartile range 4.0\u20138.2) years] were recruited, of whom 21.0% (33/157) and 12.7% (20/157) had reduced CYP2D6 and deficient G6PD activity, respectively. Day 3 mean absolute Hb concentration reduction was 1.50\u00a0g/dL [95% confidence interval (CI) 1.10\u20131.90] and 1.51\u00a0g/dL (95% CI 1.31\u20131.71) in reduced and normal CYP2D6 patients, respectively (P. falciparum gametocytes regardless of CYP2D6 or G6PD status.Single-dose 0.25\u00a0mg/kg primaquine was safe and sufficient for reducing transmission of Trial registration Study registration number: NCT03352843.The online version contains supplementary material available at 10.1186/s12936-022-04100-1. Plasmodium falciparum gametocytes in many malaria-endemic settings and this has contributed significantly to the global decline of malaria transmission [P. falciparum gametocytes, and, therefore, not sufficient to prevent transmission [The scale-up of malaria control efforts particularly using insecticide-treated bed nets and artemisinin-based combination therapy (ACT) has led to the global decline of malaria transmission, consequently renewing the interest in the possibility of eliminating the infection \u20133. Artemsmission , 4. HoweP. falciparum resistance against artemisinin in Southeast Asia [Anopheles resistance against pyrethroid in Africa [The emergency of ast Asia \u20137, and An Africa \u201310, is an Africa .Plasmodium vivax and Plasmodium ovale infection [P. falciparum gametocytes, a parasite stage responsible for onward transmission of the infection [P. falciparum gametocytes [Primaquine is an 8-aminoquinoline pro-drug indicated for radical cure of nfection \u201314. It infection , 15, 16.nfection . It may etocytes \u201323, and etocytes . A failuetocytes \u201323.P. vivax and P. ovale hypnozoites, and P. falciparum transmission [P. falciparum gametocytes probably at a lower concentration than that needed to kill and clear them from the circulation [P. falciparum gametocytes in patients with uncomplicated malaria based on CYP2D6 alleles common in African Tanzanian.Primaquine gametocytocidal effect and haematological toxicity are thought to depend on cytochrome P450 isoenzyme 2D6 (CYP2D6) conversion of a pro-drug into active phenolic metabolites \u201329. But smission . The nulsmission , 35. Redsmission , hence rsmission . Howeverculation . Of noteculation \u201340, howeculation , 31, whiculation , 33. Incculation . This stThe study was carried out at Yombo primary health facility, Bagamoyo district, Tanzania, between June 2019 and February 2020. The health facility is located southwest, about 20\u00a0km from Bagamoyo town. It serves approximately 7000 people and can carry out routine malaria microscopy and rapid diagnostic tests . The inhPlasmodium falciparum and Anopheles arabiensis are the predominant malaria parasite species and vector, respectively [Malaria transmission in the Bagamoyo district occurs throughout the year with peaks corresponding to the long and short rainy seasons from May to July and November to December, respectively. ectively . Malariaectively . Artemetectively , and priectively . Insectiectively . Previouectively .P. falciparum gametocytes. Patients with microscopically confirmed uncomplicated P. falciparum mono-infection were enrolled in the trial, treated with standard artemether-lumefantrine regimen plus 0.25\u00a0mg/kg single-dose primaquine, and then followed-up until day-28 after treatment initiation for clinical and parasitological assessment. Individuals with reduced CYP2D6 activity were defined as those with CYP2D6*17 and CYP*29 alleles, whereas those with no activity were those with CYP2D6*4 and CYP2D6*5, and normal individuals were those with CYP2D6*1, while those with CYP2D6*2 were defined as having increased activity [There is no rapid diagnostic test at the moment that could be used to evaluate and group participants based on their CYP2D6 status at the enrolment. Thus this study was a single-arm clinical trial to assess the safety and efficacy of 0.25\u00a0mg/kg single-dose primaquine added to standard artemether-lumefantrine treatment in individuals with CYP2D6 reduced or no activity compared to those with normal or increased activity for clearance and reducing transmission of activity , 46. AddP. falciparum mono-infection, parasitaemia level of 2000\u2013200,000/\u00b5L, ability to swallow oral medication, ability and willingness to abide by the study protocol and the stipulated follow-up visits, and a written informed consent from a parent/guardian and assent from children aged \u2265\u20097\u00a0years. Children aged below 1\u00a0year were not included in the study because primaquine is contraindicated in this group, and children up to the age of 10\u00a0years were included to increase the recruitment rate. Exclusion criteria were evidence of severe malaria or danger signs, known allergy to trial medicines, reported anti-malarial intake \u2264\u20092\u00a0weeks, haemoglobin <\u20095\u00a0g/dL, blood transfusion within last 90\u00a0days, febrile condition other than malaria, and known underlying chronic or severe disease .Patients presenting at the health facility with suspected acute uncomplicated malaria were screened for eligibility. Inclusion criteria were age from 1 to 10\u00a0years, weight \u2265\u200910\u00a0kg, body temperature \u2265\u200937.5\u00a0\u00b0C or history of fever in the last 24\u00a0h, microscopy confirmed At the health facility patients and their parents/guardians were informed in detail about the trial, purpose, the risks and benefits of participating in the trial, and their rights, and encouraged them to participate in case they are diagnosed with malaria. Thereafter, patients were evaluated clinically, and those with signs and symptoms of malaria were directed to the laboratory for parasitological evaluation.P. falciparum malaria infection, and met all the inclusion and none of the exclusion criteria their parents/guardians were requested to allow their children to participate in the trial. The parents/guardians were then administered with the consent form. The consent form was self-administered to the literate parents/guardians and for illiterate one\u2019s the trial clinician explained to them. All the procedures were done in the presence of a witness to ensure that there is no coercion. Parents/guardians were given time to ask questions to their satisfaction. All consented parents/guardians signed the form either by a written signature for literate or a thumbprint together with a written signature of their witnesses for the illiterate.For the children who were found to have uncomplicated P. falciparum malaria [Enrolled patients were treated using a standard 3-day course of artemether-lumefantrine  according to Tanzanian national treatment guidelines for uncomplicated  malaria . The art malaria . Accurac malaria , 25. AllPatients were withdrawn from the study in case of vomiting the study drug >\u20093 times, withdrawal of consent, the onset of a serious febrile illness, intake of any drug with anti-malarial properties outside the study protocol, or any protocol violation. Patients who missed scheduled follow-up visits and did not show up on the successive days despite efforts to trace them at their residences were considered lost to follow-up and were consequently censored from the analysis on the last day they were seen. Participants who returned before the last day of follow-up were not considered as lost to follow-up. Patients with symptoms/signs of severe disease (including repetitive vomiting of study drug) were managed according to the Tanzania national guidelines and followed up until recovery, but were censored from the analysis on the day of withdrawal . PatientClinical and laboratory assessments were performed on days 0, 1, 2, 3, 7, 14, 21, and 28 or any day of recurrent illness as follows:The assessment involved taking a history of clinical symptoms, possible adverse events (AEs), concomitant drug consumption, and clinical examination including measurement of axillary temperature. Fever was defined as a body temperature \u2265\u200937.5\u00a0\u00b0C. All the clinical and laboratory data were recorded in a case record form (CRF).Safety assessment for AE and serious adverse events (SAE) was performed using the Primaquine Roll Out Monitoring Pharmacovigilance Tool (PROMPT) . An AE wThe safety assessment involved direct questioning and recording the nature and incidence of AE and/or serious adverse events (SAE). Signs and symptoms such as fatigue, weakness, dizziness, headache, palpitations or allergic drug reactions (rash), diarrhoea, abdominal pain, dizziness, itching, fever, and haemoglobin level  were recorded and determined to whether they were related to the trial medicine. Patients with AE were kept under observation, and those requiring treatments were treated accordingly. Patients with SAE were admitted and if necessary referred to the Bagamoyo district hospital.All the patients received an information card with instructions on how to identify signs and symptoms of commonly reported AEs including monitoring of urine colour. In addition, patients were given clear containers and asked to provide urine samples on day 0 before the first drug dose and days 1, 2, and or 3. The urine colour estimation for haemoglobinuria was gauged against the Hillmen colour chart with a colour score ranging from 1 to 10 . A colouLaboratory assessment involved collection of finger-prick blood samples that were used to measure haemoglobin concentration, prepare the thick film for microscopy to assess presence and density of asexual parasitaemia and gametocytaemia, thin-film for asexual parasite species determination, and also blotted on filter paper (Perkin Elmer 266) for molecular analysis of CYP2D6 and G6PD status. On day 7, 3\u00a0mL of venous blood were collected in vacutainers from 100 patients randomly selected using research randomizer software   and usedHaemoglobin concentration was measured using a portable spectrophotometer, HemoCue Hb 201\u2009+\u2009, with a precision of \u00b1\u20090.3\u00a0g/dL. The HemoCue was calibrated every morning using a control cuvette at 16.0\u2009\u00b1\u20090.3\u00a0g/dL according to the manufacturer\u2019s instructions. Anaemia was classified as haemoglobin level <\u200911\u00a0g/dL (mild), <\u20098\u00a0g/dL (moderate) and <\u20095\u00a0g/dL (severe).Thin blood smears were prepared only at enrolment, whereas thick blood smears were prepared at all sampling time points. At the enrolment, two pairs of thin and thick smears were collected, and the first pair was quickly stained using 10% Giemsa for 15\u00a0min to accelerate the enrolment of patients, whereas the second pair was stained slowly using 3% Giemsa for 1\u00a0h. Thick smears collected during follow-up were stained slowly using 3% Giemsa for 1\u00a0h. Asexual and sexual parasites on thick smears were counted against 200 white blood cells (WBC). This number was then multiplied by 40, assuming 8000 leukocytes per microlitre of blood, to gain an approximate parasite density per microlitre of blood. A blood slide was considered negative if no parasite was seen after examining 100 fields. Two, independent microscopists read all the microscopy slides. In case they disagreed on the presence of parasitemia or if the density differed by more than 25%, a third independent reading was performed. In the case of positive versus negative results, a third independent reading was used to confirm the reading of the first two readers.The genomic DNA was extracted from 100\u00a0\u03bcL of dried blood spots on filter papers using Quick-DNA\u2122 Miniprep plus Kit Catalog Nos. D4068 and D4069 .To detect CYP2D6 point mutations, the extracted DNA was amplified using mass array VeriDose CYP2D6 CNV Panel . The CYPThe G6PD analysis was performed through amplification of the extracted genomic DNA using a single-round PCR followed by gel electrophoresis. Amplicons were loaded on agarose gel containing GelRed\u2122 (Biotium) and then visualized under ultraviolet- transillumination. The molecular weight of the DNA fragments was sized using Gene Ruler TM 100\u00a0bp molecular weight marker. Amplicons with an expected band size of 1061\u00a0bp for Exon 3\u20135 were sequenced using the Sanger sequencer. The interested two most common polymorphisms associated with G6PD deficiency in Africa i.e., A376G (rs1050829 A\u2009>\u2009G/T\u2009>\u2009C) and G202A (rs1050828 G\u2009>\u2009A/C\u2009>\u2009T) were found in Exon 3\u20135 after analysis of sequences data by Geneious prime software. The outcomes were classified as follows; For males A was defined as wild-type/normal and A\u2212 as hemizygous/deficient G6PD status, whereas for females A\u2212A\u2212 was defined as homozygous/deficient, AA\u2212 and BA\u2212 as heterozygous/intermediate and AA and BA as wild-type/normal G6PD status .P. falciparum block 3 of merozoite surface protein (MSP) 2, block 2 of MSP 1, and region II (RII) of glutamate-rich protein using a nested PCR method [To differentiate recrudescence from new infection, the extracted DNA from patients classified during the 28\u00a0days follow-up period as late clinical failure or late parasitological failure was genotyped using a stepwise genotyping of R method . BrieflyAnopheles gambiae sensu stricto mosquitoes through an artificial membrane. These mosquitoes were starved for a minimum of 5\u00a0h before the experiments. The membrane feeder setup was maintained at 37\u00a0\u00b0C using a temperature-controlled water bath and was run for 15\u00a0min. Only fully fed mosquitoes were transferred into a holding cage in the insectary and kept on 10% glucose, at 28\u00a0\u00b0C for eight days for oocyst maturation, whereas the partially fed and unfed ones were destroyed. A minimum of 25 mosquitoes that survived to this period was dissected in 0.5% mercurochrome, and their midguts were examined using microscopy for oocyst prevalence and density. The proportion of infected mosquitoes and the density of oocysts in infected mosquitoes were determined. An infected mosquito was defined as any mosquito with oocysts detected after dissection. Membrane feeding experiments were performed at the IHI, Bagamoyo, Tanzania.Infectivity was assessed using mosquito membrane feeding experiments. Blood samples collected on day 7 from a subset of 100 randomly selected participants were used in the experiment to assess the biological evidence of gametocyte sterilization \u201358. BrieP. falciparum gametocytes in patients with uncomplicated P. falciparum malaria based on CYP2D6 status defined as mean maximal absolute Hb (g/dL) fall during follow-up (day 0\u201328). Secondary outcomes included: follow-up day of haemoglobin nadir defined as mean day of follow-up (day 0\u201328) per group with lowest Hb measurement; maximal percentage fall in haemoglobin defined as the percentage relative change of maximal haemoglobin drop during follow-up (day 0\u201328) from baseline; percentage of participants with anaemia during the days 0\u201328 of follow-up; percentage of patients receiving blood transfusion during days 0\u201328; proportion of patients with urine colour change score \u2265\u20095 using Hillmen Urine Colour Chart during days 0\u201328; incidence of serious adverse events by sign, symptom, laboratory parameter during days 0\u201328; proportion of participants with gastrointestinal symptoms within day 0\u20137 after taking study drug; proportion of patients with G6PD deficiency; proportion of patients who were infectious to mosquitoes on day 7 determined by direct membrane feeding assay; and proportion of patients with PCR-adjusted adequate clinical and parasitological response (ACPR) by day 28.\u00a0All the evaluated study outcomes were compared between CYP2D6 or G6PD status. The primary outcome was the safety of 0.25\u00a0mg/kg single-dose primaquine for reducing transmission of The study was conducted following Good Clinical Practices (GCP), the Declaration of Helsinki, and applicable regulatory requirements in Tanzania. Ethical clearances to conduct the trial were obtained from the Muhimbili University of Health and Allied Sciences Institutional Ethics Review Board and the National Health Research Ethics Review Committee of the National Institute for Medical Research. Permission to import and use primaquine was sought from the Tanzania Medicines and Medical Devices Authority. An independent data and safety monitoring board (DSMB) was established, visited the trial site, and performed the interim analysis of safety data. Written informed consent was obtained from parents/guardians of the participants whereas children aged \u2265\u20097\u00a0years also provided assent before their enrollment. The trial is registered at clinicaltrials.gov with the number NCT03352843.The study was powered to measure the efficacy of primaquine (0.25\u00a0mg/kg primaquine dose) in maximum reduction of gametocytaemia from day one through day 28. A sample size of 155 evaluable subjects was chosen to estimate a prevalence of gametocytes carriage measured by QT-NASBA on day 7 of 10% among the individuals with normal/increased activity compared to 35% in reduced activity at a power of 80% . All theIn this trial, a similar prevalence of CYP2D6 genotypes was assumed to be as described by Wennerholm et al. ; 63% norNo sample size calculation was performed for the number of patients required for membrane feeding experiments but based on other studies \u201323 100 pData were double entered in an electronic database, cleaned, and analysed using R version 3.2.3  software. The data were analysed based on intention to treat and per-protocol analyses.t-test. In addition, a one-sample t-test was used for comparisons of the mean maximal fall Hb with that reported in Uganda by using 0.4\u00a0mg/kg single-dose [Chi-square or Fisher exact test. Data were censored at the time of withdrawal for patients lost to follow-up, withdrawal of consent, and PCR determined reinfection or uncertain PCR outcome. A p\u2009<\u20090.05 was defined as statistically significant.The primary safety end-point, the mean maximal haemoglobin concentration (Hb) reduction during 28\u00a0days of follow-up was assessed and compared between the CYP450 2D6 status using independent two paired samples gle-dose . ProportP. falciparum infection. Of the infected patients, 157 were enrolled in the study after fulfilling all the inclusion and none of the exclusion criteria  had microscopy-confirmed % had micThe pre-treatment characteristics of the enrolled patients are presented in Table At the enrolment, 69.4% (109/157) of the patients had a fever. Following the initiation of treatment, fever clearance was rapid with only 22.9% (36/157), 3.2% (5/157), and 0.6% (1/157) of the patients remaining febrile on days 1, 2, and 3, respectively. Microscopy-determined asexual parasites decreased to 81.5% (128/157) on day 1 and 13.4% (21/157) on day 2, after the initiation of the treatment, and none of the patients had parasitaemia on day 3. Only one (0.64%) patient had microscopy-determined gametocytes at the enrolment, and these gametocytes were cleared on day 2 after the initiation of treatment. Another patient had gametocytes detected on day 1 after the initiation of treatment, but was not detected again on day 2 and onward.Fisher\u2019s test\u2009=\u20090.736, p\u2009=\u20090.513). In addition, there was no statistically significant difference in the infectiousness between children under five years of age (17.6% (6/34) and those aged 5\u00a0years and above (15.6% (10/64) .One hundred patients were randomly selected to provide blood samples for mosquito membrane feeding experiments on day 7 after the initiation of treatment, and of these, 98 (98.0%) provided the blood sample. Sixteen out of 98 (16.3%) patients were infectious to at least one mosquito on day 7. Of the infectious patients, 10.0% (2/20) and 17.9% (14/78) had reduced and normal CYP2D6 enzyme activity, respectively, and the difference between the groups was not statistically significant  patients had late clinical and parasitological failure, 6.9% (10/143) had a late parasitological failure, and 6.9% (10/143) had PCR adjusted recrudescence. The day 28 artemether-lumefantrine per-protocol PCR-adjusted adequate clinical and parasitological response (ACPR) was 93.0% (133/143), , whereas, 25.0% (5/20), 20.7% (24/116) and 23.8% (5/21) of them were G6PD deficient, normal and heterozygous females, respectively .Furthermore, during follow-up, acute haemolysis defined as haemoglobin reduction of \u2265\u200925% occurred in 21.7% (34/157) of the patients. Of the patients with acute haemolysis, 18.2% (6/33) and 22.6% (28/124) had reduced and normal CYP2D6 activity, respectively .On the other hand, 45.0% (9/20) of G6PD deficient patients had reduced CYP2D6 enzyme activity. Out of 11 participants with G6PD deficiency and normal CYP2D6 enzyme activity, 5 (45.5%) had acute haemolysis, whereas none (0/9) of participants with G6PD deficiency and reduced CYP2D6 had acute haemolysis . Likewise, the prevalence of anaemia at the enrolment was 55.0% (11/20), 53.4% (62/116), and 57.1% (12/21) in G6PD deficient, G6PD normal, and heterozygous patients, and was not significantly different between the status . During follow-up, the prevalence of anaemia increased significantly from 54.1% (85/157) at the enrolment to 89.8% (141/157) on day 3  and 82.8% (130/157) on day 7 . Prevalence of anaemia was not significantly different between CYP2D6 status on days 3  and day 7 , but was significantly different between G6PD status with higher prevalence occurring in G6PD normal patients on day 3 (93.1% (108/116), trend x2\u2009=\u20096.136, p\u2009=\u20090.047) and in G6PD deficient patients (100% (20/20), trend x2\u2009=\u20097.994, p\u2009=\u20090.018) on day 7.At the enrolment 39.4% (13/33) and 58.1% (72/124) of patients with reduced and normal CYP2D6 enzyme activities had anaemia, but the prevalence of anaemia was not significantly different between the status (About 5.7% (9/157) of the patients were both reduced CYP2D6 and G6PD deficient. The haematological changes in patients with both reduced CYP2D6 and G6PD deficiency compared to other CYP2D6 and G6PD status are presented in Table x2\u2009=\u20091.29, p\u2009=\u20090.257) or G6PD  status was not statistically significant. Likewise, all other AEs were not statistically significantly different between the CYP2D6 or G6PD status.Other adverse events (AEs) occurred in 65.6% (103/157) of the patients. In total 124 AEs were reported during follow-up and were all mild and self-limiting, Table P. falciparum gametocytes [P. falciparum gametocytes, to probably be influenced by the polymorphic nature of the gene encoding this enzyme [Primaquine is the only approved anti-malarial drug potent against mature etocytes , 59, 60.s enzyme , 36, 61.s enzyme , 63. This enzyme , 25, 64 s enzyme , 65, 66.s enzyme , 25, 67.s enzyme \u201370. In aP. falciparum gametocytes. Following mosquito membrane feeding assay, 16.3% of the patients were infectious to at least one mosquito on day 7. However, there was no statistically significant difference in the prevalence of infectiousness between CYP2D6 status. In addition, there was no statistically significant differences in the rate of infectiousness between the underfives and those aged above five years. Contrarily, a previous study has reported a higher prevalence of gametocytes carriage in CYP2D6 poor/reduced activity [Anopheles mosquitoes. Importantly, sterilization of P. falciparum gametocytes occurs at a lower primaquine concentration and earlier than it is required for them to get killed and cleared from the circulation [P. falciparum gametocytes transmission; second, other factors such as poor intestinal absorption especially during the acute clinical malaria infection may be responsible for the failure in reducing the transmission; and third, the CYP2D6 reduced activity has a more significant role in P. vivax hypnozoites clearance rather than P. falciparum gametocytes sterilization.Reduced CYP2D6 activity is thought to reduce primaquine efficacy probably by impairing the production of phenolic metabolites that determines the drug\u2019s efficacy , 61. Theculation , therefoP. falciparum parasite clearance is a hallmark of artemisinins [P. falciparum K13 gene.Rapid fever and asexual misinins . In thismisinins , 73, othmisinins \u201376, and misinins \u201379 usingP. falciparum gametocytes was not assessed.Despite its strength, the study had limitations including, at the enrolment patients were not tested and then grouped based on the CYP450 2D6 status as there is no rapid test for the condition, thus it was not possible to recruit an equal number of participants based on their CYP2D6 status, and therefore, it was not possible to have a double-blinded randomized clinical trial with artemether-lumefantrine alone versus artemether-lumefantrine plus SLD primaquine. However, the attrition rate of 20% was used to ensure that the required proportions of patients with CYP2D6 normal/increased and reduced/null are recruited. Another limitation is that the sample size calculation was based on previous studies that assessed PQ efficacy for reducing gametocytes transmission using QT-NASBA, but during the implementation, the efficacy was assessed using mosquito membrane feeding assays only, and the study was not powered to assess efficacy using membrane feeding assay as the primary outcome. Furthermore, the available primaquine concentration in the plasma of treated patients was not measured, therefore, its relationship with sterilization of P. falciparum gametocytes, as assessed by mosquito membrane feeding assays, regardless of the CYP2D6 or G6PD status, and the observed adverse events were mild and self-limiting.The SLD primaquine was safe and sufficient for reducing transmission of Additional file 1. Dataset."}
+{"text": "Plasmodium falciparum malaria. This study was designed to measure the efficacy of artemether-lumefantrine and to assess the selection of the P. falciparum chloroquine resistance transporter (pfcrt) and P. falciparum multi-drug resistance 1 (pfmdr1) genotypes that have been associated with drug resistance. Methods: A 28-day follow-up efficacy trial of artemether-lumefantrine was conducted in patients aged 6 months and older suffering from uncomplicated falciparum malaria in four different Malian areas during the 2009 malaria transmission season. The polymorphic genetic markers MSP2, MSP1, and Ca1 were used to distinguish between recrudescence and reinfection. Reinfection and recrudescence were then grouped as recurrent infections and analyzed together by PCR-restriction fragment length polymorphism (RFLP) to identify candidate markers for artemether-lumefantrine tolerance in the P. falciparum chloroquine resistance transporter (pfcrt) gene and the P. falciparum multi-drug resistance 1 (pfmdr1) gene. Results: Clinical outcomes in 326 patients (96.7%) were analyzed and the 28-day uncorrected adequate clinical and parasitological response (ACPR) rate was 73.9%. The total PCR-corrected 28-day ACPR was 97.2%. The pfcrt 76T and pfmdr1 86Y population prevalence decreased from 49.3% and 11.0% at baseline (n = 337) to 38.8% and 0% in patients with recurrent infection (n = 85); p = 0.001), respectively. Conclusion: Parasite populations exposed to artemether-lumefantrine in this study were selected toward chloroquine-sensitivity and showed a promising trend that may warrant future targeted reintroduction of chloroquine or/and amodiaquine.Background: Artemether-lumefantrine is a highly effective artemisinin-based combination therapy that was adopted in Mali as first-line treatment for uncomplicated  Plasmodium falciparum to chloroquine is present in almost all malaria-endemic countries [P. falciparum [Resistance of ountries . Resistaountries ,3,4. Artlciparum . The Worlciparum . In Malilciparum . Althouglciparum . Studieslciparum ,9,10.Artemether-lumefantrine combines the rapid-acting, short half-life synthetic artemisinin with the long-acting, more slowly cleared aryl amino alcohol, lumefantrine . EffortsPlasmodium falciparum multi-drug resistance 1 (pfmdr1) gene on chromosome 5 encodes a digestive vacuole transmembrane glycoprotein . Point mutations at codons 86, 184 (predominant in Asia and Africa) and 1034, 1042, and 1246 (predominant in South America) and increased gene copy numbers of pfmdr1 are associated with resistance to many drugs, including chloroquine, quinine, mefloquine, and the aryl amino alcohols  [The antrine) ,17,18.P. falciparum chloroquine resistance transporter (pfcrt) gene on chromosome 7, which also encodes a digestive vacuole transmembrane protein, are associated with chloroquine resistance [Mutations in the sistance . Point msistance .pfcrt and pfmdr1 SNPs, but just a few were from West Africa [pfmdr1 and pfcrt polymorphisms three years after the wide use of artemether-lumefantrine in Mali.Several countries in West Africa, including Mali, use artemether-lumefantrine as a first-line treatment for uncomplicated malaria. A number of other studies from Africa have investigated whether artemether-lumefantrine administration selects for particular t Africa  and Malin = 77), 88 in Faladje (n = 88), 100 in Bandiagara (n = 100), and 72 in Pongonon (n = 72). After 28 days of follow-up, 327 (97%) participants completed the study: 70 in Kolle (91%), 88 in Faladje (100%), 99 in Bandiagara (99%), and 70 in Pongonon (97.2%), as shown in Of 400 recruited patients, 337 were enrolled: 77 in Kolle (p > 0.78).The clearance of fever was not statistically different among the four sites (data not shown). By Day 2, nearly all fevers had resolved. Parasitemia was statistically different among sites and highest in Pongonon on Day 1, but parasites were undetectable in all patients by Day 3 . Hemoglopfcrt 76T and pfmdr1 86Y mutations in all participants at baseline (Day 0) and re-treatment (Day R). Carriage of the pfmdr1 N86 allele and/or the pfcrt K76 allele (wild types) was not associated with blood smear positivity after initial parasite clearance in participants at any site within the 28-day follow-up period (p > 0.05). Additionally, we conducted an analysis of directional selection acting on alleles pfmdr1 codon 86 and pfcrt codon 76 from infected patients following therapy with artemether-lumefantrine. The frequencies of the pfmdr1 N86 and the pfcrt K76 alleles in recurrent infections increased from 84.7% to 100% (p = 0.0002) and 43.5% to 61.2% (p = 0.02), respectively  were selected in vivo after treatment with artemether-lumefantrine. Statistically significant pfcrt allele-selection has not been observed as often as pfmdr1 allele-selection. Our study, in addition to those by Sisowath, et. al. [pfmdr1 and pfcrt.We found that both  et. al. , and Oti et. al. , confirm et. al. . All thepfcrt K76 and pfmdr1 N86 are selected for under the same mechanism remains to be determined. The reason for recurrent infections is primarily reinfection and allele selection occurs in reinfecting parasites. The concentration of lumefantrine is high enough to eliminate mutated parasites, but insufficient to eliminate wild-type parasites without the combined protection of artemether. Hastings and Ward have suggested that the selection of N86 may represent a marker of tolerance to lumefantrine [Whether fantrine .pfcrt K76 and pfmdr1 N86 alleles (wild types) among recurrent infections. In this analysis, allele prevalence on Day 0 was compared with allele prevalence on Day R among the 85 patients who had recurrent infection. Recurrent parasitemia was statistically similar among three of the four sites , though recurrent malaria was most commonly observed in Bandiagara (15.2%). Recurrent malaria (4.3%) and parasitemia (2.9%) were both statistically lower in Pongonon.In our study, treatment with artemether-lumefantrine (AL) was selected for pfmdr1 N86 allele has been associated with treatment failure in patients given artemether-lumefantrine [pfmdr1 N86 allele also increased in all isolates, which shows a selection in favor of the wild-type allele, as seen previously with the use of artemether-lumefantrine [pfmdr1 allele prevalence change was similar to that reported recently by Dokomajilar [pfcrt K76 allele (wild type) increased in all isolates after treatment. We found evidence for significant selection of both pfcrt K76 and pfmdr1 N86 SNPs (wild types) after treatment with artemether-lumefantrine.Carriage of the fantrine ,45,46,47fantrine . The patomajilar  from theomajilar , and froomajilar . Prevalepfmdr1 N86 allele-selection (wild type) are consistent with previous studies; however, the risk of new infections among these studies has varied. In areas of Tanzania with intermediate transmission, the risk of new infection after artemether-lumefantrine treatment was only 5%; a similar study performed in an area of high transmission in Tanzania showed new infection rates of about 50%. In our study, risk of reinfection also varied by site, from Pongonon to Faladje/Bandiagara/Kolle. As transmission intensity increased from one site to another, so did the proportion of treated patients selecting for parasites harboring a particular allele. In vitro studies propose that gene selection occurs during the elimination phase of lumefantrine (t \u00bd = 3\u20136 days) [The selection of a SNP previously associated with quinoline antimalarial drug resistance was first documented in 2005. Our results regarding pfmdr1 or pfcrt selection alone, albeit the sample size may have been insufficient to identify any associations. We thus relied on the combined outcomes of reinfection and recrudescence, as previous studies have done. However, grouping these outcomes involves the risk of seeing relationships that may not exist. Unless an increased frequency of recrudescence occurs in patients treated with artemether-lumefantrine, insights regarding tolerance and resistance will remain limited.The eight (2.5%) recrudescent infections in this study could not be explained by pfmdr1 haplotype of N86-184F-1246 with treatment failure [pfcrt 76T and pfmdr1 86Y (mutant types)) against artemisinin partner molecules [pfcrt K76 and pfmdr1 N86 alleles (wild types), suggesting a potential benefit of using different ACTs simultaneously as first-line treatment to reduce selective pressures by each regimen. Alternating the use of AL versus ASAQ could maintain drug effectiveness, while decreasing the risk of tolerance/resistance. Larger studies examining this hypothesis could inform NMCPs to adjust malaria treatment policies.The presence of these wild-type alleles has been associated with reduced sensitivity to lumefantrine in vitro . Other s failure ,53. We holecules ,55. AL aP. falciparum was previously resistant to these drugs.In conclusion, parasite populations exposed to artemether-lumefantrine in this study were selected toward chloroquine-sensitivity and showed a promising trend that may warrant future targeted reintroduction of chloroquine or/and amodiaquine in areas where pfcrt K76T mutation prevalence was previously recorded in 1999 and 2002 at 28.3% and 85%, respectively [pfcrt K76T mutation prevalence was previously recorded in 2003 at 80% [pfcrt K76T mutation prevalence was previously recorded in 2002 at 39% [pfcrt K76T mutation prevalence was previously recorded in 2002 at 13% (unpublished data).This study was conducted during the malaria transmission season, from October 2009 to January 2010 in four different Malian areas: Kolle, Faladje, Bandiagara, and Pongonon. Kolle is a rural village of ~3000 inhabitants located 57 km southwest of Bamako. The climate is Sudanian savanna with two distinct seasons, the dry season (January\u2013June) and the rainy season (July\u2013December). Malaria is hyperendemic during 3 to 4 months of the rainy season with a parasite index (PI) of 70%\u201385%, and of 40%\u201350% during the dry season . The entectively . FaladjeP. falciparum at a density \u22652000 asexual parasites/\u00b5L to 200,000/\u03bcL of blood, (iii) axillary temperature \u226537.5 \u00b0C or reported fever in the preceding 24 h, (iv) ability to swallow oral medication, and (v) willingness to consent and comply with the study protocol for the duration of the study . Exclusion criteria consisted of the following: (i) signs of severe malaria according to 2003 WHO definitions, (ii) a concomitant febrile condition , (iii) severe malnutrition, (iv) disclosed or clinically patent pregnancy, (v) other known underlying chronic or severe diseases, or (vi) history of hypersensitivity to artemether-lumefantrine or the rescue treatment, artesunate-amodiaquine.Inclusion criteria consisted of the following: (i) age \u22656 months, (ii) microscopic detection of Finger-prick blood samples were collected from enrolled patients to assess the parasite density and hemoglobin level on-site, and to be preserved on the filter papers , which were stored in self-sealing plastic bags with desiccant for later analysis. Patients received a twice-daily dose of artemether-lumefantrine of 20 mg/120 mg as indicated by weight for three consecutive days: one tablet per dose for patients weighing 5\u201314 kg, two tablets per dose for those weighing 15\u201324 kg, three tablets per dose for those weighing 25\u201334 kg, and four tablets per dose for those weighing \u226535 kg. All doses were administered on-site under supervision. A full dose was re-administered if a participant vomited within 30 min of the initial drug administration. After treatment initiation, patients were examined and finger-prick blood samples were collected routinely on Days 1, 2, 3, 7, 14, 21 and 28 of the treatment and on any day of recurrent illness during the 28-day follow-up period. In cases of treatment failure, quinine was administered if the patient was re-diagnosed with uncomplicated malaria or parasitemia. As per the national guidelines, patients were referred to a hospital and intravenous quinine was administered if severe malaria was diagnosed at any time during the follow-up period. Treatment was also given for any other conditions diagnosed .Specimen analysis was completed at the Malaria Research and Training Center (MRTC) lab in Bamako, Mali. Parasite density and rate of gametocyte carriage were measured, as number of parasites per microliter, from Giemsa-stained thick blood smears. A positive smear was defined as a thick smear with at least one asexual parasite per 100 fields under 1000\u00d7 magnification . Blood was collected for hemoglobin measurement (HemoCue AB) on Day 0 prior to treatment and on Days 14 and 28 of treatment. Children with hemoglobin <7 g/dL were treated with oral iron therapy according to national policy . For quaThree methods were used for DNA extraction. (1) Methanol was used to extract parasite DNA from all filter paper blood samples. Samples that did not yield DNA by this method were extracted using (2) the Chelex method . For a sP. falciparum isolates, in order to distinguish between episodes of recrudescence and reinfection [P. falciparum isolates were denoted by the number of different-sized PCR products [MSP2, MSP1, and Ca1 were used to compare genotypic profiles between pre- and post-treatment nfection ,62,63. Tproducts . Length pfcrt and pfmdr1 were assessed for all baseline infections (Day 0) and all recurring infections post-treatment (Day R) to examine any association between the artemether-lumefantrine treatment and the development of tolerance/resistance, as has been previously suggested [pfcrt K76T SNP was analyzed by restriction fragment length polymorphism (RFLP)-PCR and mutation-specific MS-PCR [P. falciparum isolates Dd2 (containing the 76T mutation) and 3D7 (lacking this mutation) were used as the positive controls, while a DNA-free reaction mix was used as negative control. The pfmdr1 N86Y SNP was analyzed by PCR-RFLP, as described previously [P. falciparum isolates, representing wild-type (N86) and mutant genotypes (86Y), respectively, were used as the positive controls. PCR was performed in a tetrad 2 thermocycler . Primers for pfcrt were synthesized by Integrated DNA Technologies, Coralville, IA, USA. Primers for pfmdr1 were synthesized by the UMD Biopolymer core facility. RFLP and MS-PCR products and digestions were separated by electrophoresis in 2% agarose gels with ethidium bromide and were visualized under UV transillumination .Mutations in uggested . The pfcc MS-PCR . DNA froeviously . The negpfcrt 76T + pfmdr1 86Y) were calculated with 95% confidence interval (CI). Chi-square or Fisher exact probability tests and McNemar\u2019s \u03c72 test were used for comparisons as appropriate with statistical significance set at p value < 0.05.Data were collected on case report forms and double entered using MS Access and analyzed using Stata (Stata Corp 11). The prevalence of single mutations and of various genotypes including the double mutant (o9/63/FMPOS#. Community permission was obtained from each locality prior to the study. Written informed consent was obtained from all adults and parents/guardians of children enrolled prior to screening.The study was approved by the ethical committee of the Faculty of Medicine, Pharmacy and Dentistry of Bamako, University of Sciences, Techniques, and Technologies of Bamako, Mali #N"}
+{"text": "Plasmodium-mixed infection may lead to aggravated morbidity and increased mortality. The present study aimed to quantify the pooled proportion and risk of malarial recurrences after the treatment of Plasmodium-mixed infection. The results of the study may provide benefits in the management of Plasmodium-mixed infection in co-endemic regions.Malaria mixed infections are often unrecognized by microscopists in the hospitals, and a delay or failure to treat Plasmodium-mixed infection. The primary outcome was the pooled prevalence of Plasmodium parasitemia after initiating antimalarial treatment for Plasmodium-mixed infection. The secondary outcome was the pooled risk ratio (RR) of malarial recurrence in Plasmodium-mixed infection compared with those in Plasmodium falciparum and Plasmodium vivax mono-infection. The pooled analyses were calculated by random-effects meta-analysis. After the initial treatment in different days of recurrences (\u2264\u200928\u00a0days or >\u200928\u00a0days), the risk of Plasmodium parasitemia was compared in subgroup analysis.This systematic review and meta-analysis searched the international Prospective Register of Systematic Reviews , MEDLINE, Web of Science, and Scopus for potentially relevant studies in any language published between January 1, 1936, and July 20, 2020, assessing drug efficacy in patients with Plasmodium-mixed parasitemia was 30%  16\u201343; I2: 99.2%; 11 studies). The RR of malarial recurrence within 28\u00a0days after the initial treatment  of Plasmodium-mixed parasitemia compared with the treatment of P. falciparum was 1.22 , while there was no significant difference in the risk of recurrence 28\u00a0days after initial treatment compared with the treatment of P. falciparum . The subgroup analysis of antimalarial drugs showed that significant malarial recurrence within 28\u00a0days was observed in patients treated with artemisinin-based combination therapies (ACTs) with no significant heterogeneity .Out of 5217 screened studies, 11 were included in the meta-analysis, including 4390 patients from six countries. The pooled prevalence of all recurrences of Plasmodium-mixed infection. Moreover, significant malaria recurrence of mixed infection occurred within 28\u00a0days after treatment with ACTs.The present findings showed a high prevalence of malarial recurrence after the initial treatment of The online version contains supplementary material available at 10.1186/s13071-021-04792-5. Plasmodium are recognized as the cause of malarial diseases in humans, Plasmodium falciparum and Plasmodium vivax are the most common Plasmodium species infecting humans worldwide [P. falciparum infection is the most common cause of death from malaria, particularly in endemic areas of stable transmission or high malaria endemicity [P. vivax is the most common cause of benign malaria in Central America, South America, and Asia [P. vivax infection can occur [P. malariae and P. ovale are recognized as benign malaria parasites, but may cause severe malaria [P. ovale parasites are divided into P. ovale curtisi and P. ovale wallikeri according to the dimorphism in defined genes [Malaria remains important to global health, as it is related to severe disease morbidity and mortality . Althougorldwide \u20133. P. fademicity . P. vivaand Asia \u20138. Howevan occur , 10. P.  malaria . P. ovaled genes , 13.Plasmodium species in a single-host or mixed-species infection can occur in an endemic area [Plasmodium-mixed infection may be acquired by simultaneous inoculation of sporozoites from multiple infected anopheline mosquitoes [Plasmodium species in a single host remains controversial, a previous study demonstrated that a mixed infection of P. falciparum and P. vivax in a single human host can exhibit clinical signs in two ways: (1) suppressing each other and therefore reducing the severity of malaria, or (2) enhancing each other and therefore leading to an increased risk of severe malaria [Plasmodium-mixed species due to very low parasitemia in mixed infections. Recently, the polymerase chain reaction (PCR) method has been conducted to detect Plasmodium mono-infection [Plasmodium-mixed infection with high evidence of sensitivity and specificity for detecting low parasitemia [Plasmodium-mixed infection may lead to aggravated morbidity and increased mortality rate. Therefore, the consideration of Plasmodium-mixed infection and its management have important clinical and therapeutic implications, particularly in patients who are co-infected with P. falciparum and P. vivax. The present study aimed to quantify the pooled proportion and risk of malarial recurrences after treatment of Plasmodium-mixed infection. The results of this study may provide benefits to the management of Plasmodium-mixed infection in co-endemic regions.The coexistence of two or more mic area , and thisquitoes . Althoug malaria . Mixed infection  and Plasasitemia \u201320. FurtThe protocol was registered in the International Prospective Register of Systematic Reviews (PROSPERO) (ID\u2009=\u2009CRD42020199709).Plasmodium-mixed infection in any language published between January 1, 1936, and July 20, 2020. The search strategy was used as previously described [MEDLINE, Web of Science, and Scopus were searched for potentially relevant studies on drug efficacy in patients with escribed , with soPlasmodium species after treatment for Plasmodium-mixed infection. The following studies were excluded: studies reporting the prevalence of Plasmodium-mixed infection, in vitro studies, genetic studies, case reports or series, studies with animal models, reviews, studies on co-infection of malaria and other agents, studies where all data on recurrence could not be extracted, studies with pregnant women, and studies where full-text manuscripts were unavailable. Studies were selected by two authors (AM and MK), and discrepancies were resolved by a discussion with a third author (KUK). The results of this study were reported according to the preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines [Studies in any language published between January 1, 1936, and July 20, 2020, were included in the analysis if they explicitly reported the presence of recurrent parasitemia with any idelines .Plasmodium-mixed infection at baseline, number of patients with Plasmodium falciparum and Plasmodium vivax infections at baseline, day of recurrence after the initial treatment, antimalarial drugs used, and number of any Plasmodium species after treatment. Antimalarial drugs used in the included studies were categorized as artemisinin-based combination therapies (ACTs) or chloroquine for subgroup analysis. Based on 2015 [The data from the included studies were extracted independently by two authors (AM and MK); any discrepancies between them were resolved by discussion for consensus. The following details were extracted into a standardized pilot datasheet (Excel form) before further analysis: authors, year of publication, study site, year of the experiment, and information on patients, including age, gender, clinical signs, number of patients with recurrence, number of patients with  on 2015  and 2021 on 2015  World HeThe quality of the included studies was assessed using a tool developed by the Joanna Briggs Institute  .Plasmodium parasitemia after treatment was estimated using random-effects meta-analysis with proportions pooled using the Freeman-Tukey double arcsine transformation. The risk ratio (RR) of malarial recurrence after treatment of Plasmodium-mixed infection compared with those after treatment of P. falciparum and P. vivax were estimated using random-effects meta-analysis. The subgroup analysis of days of recurrence (\u2264\u200928\u00a0days and >\u200928\u00a0days) was conducted to determine whether recurrence was caused by clinical treatment failure or by other causes. The subgroup analysis of antimalarial drugs (ACTs or chloroquine) and clinical signs  was also conducted. Data heterogeneity among the included studies was assessed using Cochran's heterogeneity statistic and quantified by the I2 statistic. All analyses were performed using Stata version 15 software .The pooled prevalence of recurrence of any t distribution (t test) instead of the normal distribution (z-test). If a small-study effect was found by Egger\u2019s test, the contour-enhanced funnel plot was further analyzed to explore the source of publication bias.Publication bias across the included studies was assessed using funnel plots and Egger\u2019s test. In the funnel plot, the precision of the estimated intervention effect increases with the size of the study. Therefore, the effect estimates from small studies will scatter more widely at the bottom of the graph, with the spread narrowing among larger studies. Without bias, the plot should approximately resemble a symmetrical (inverted) funnel. For Egger\u2019s test, the significance of the coefficients is based on a Student Plasmodium infection, 82 in vitro studies, 58 genetic studies, 24 case report/series, 19 animal models, 17 studies that reported mixed infection at enrollment but with no data on its recurrence during follow-up, 16 studies without mixed Plasmodium infection, 16 review articles, 8 studies with no full text, 8 studies that reported malaria and other pathogens, 8 studies in pregnant women, 5 knowledge/attitude/practice studies, 3 mathematic models, and 2 guidelines  was demonstrated in six studies [Of the 11 included studies  was demonstrated in a study by Smithuis et al. [Plasmodium parasitemia after treatment of Plasmodium-mixed infection was 30%    . A higher risk of malarial recurrence after 28\u00a0days of treatment was demonstrated in the studies by Douglas et al. [A subgroup analysis of the days of recurrence (\u2264\u200928\u00a0days or\u2009>\u200928\u00a0days) was conducted for the 11 studies. A significantly higher risk of malarial recurrence within 28\u00a0days after the treatment of mixed s et al. , while as et al. .Fig. 4SuPlasmodium infection and P. falciparum infection   after treatment with chloroquine. A subgroup analysis of antimalarial drugs was further performed in six studies [Plasmodium parasitemia within 28\u00a0days after treatment. The results showed that the malarial recurrence within 28\u00a0days was significantly observed in patients treated with ACTs, with no significant heterogeneity , while there was no significant difference in the risk of malarial recurrence within 28\u00a0days in patients treated with chloroquine  . In addition, a significantly higher risk of malarial recurrence after treatment of Plasmodium-mixed infection was also observed in patients with uncomplicated malaria, with no significant heterogeneity across the studies analyzed   was conducted. The results showed that a significantly higher risk of malarial recurrence after treatment of Plasmodium-mixed infection was compared with malarial recurrence after treatment of P. vivax infection. Overall, no significant difference in the risk of any recurrent parasitemia was observed when the recurrence risks from the seven studies were pooled  , indicating that the publication bias might be due to other causes, such as the variability in included studies, especially concerning the prevailing Plasmodium species and resistance pattern driven by drug pressure.The publication bias is shown in Fig.\u00a0Plasmodium-mixed species, if left untreated or managed poorly, can lead to severe malaria [Plasmodium-mixed infection or P. falciparum mono-infection showed a similar trend of complications in which severe anemia, pulmonary failure, and renal impairment were the three most common complications found [Plasmodium parasitemia after treatment of Plasmodium-mixed infection (30%). This result indicated that treatment failure, relapse, or recrudescence might have occurred in these patients. The present study also demonstrated clearly that a significantly higher risk of malarial recurrence occurred within 28\u00a0days after treatment of Plasmodium-mixed infection compared to those with P. falciparum infection, while no significant recurrence was observed between the two groups after 28\u00a0days. This implies that the cause of malarial recurrence in patients with mixed infection might be due to early or late clinical treatment failure. In comparison to the treatment of P. falciparum, the treatment of Plasmodium-mixed infection significantly led to malaria recurrence within 28\u00a0days as observed in patients treated with ACTs, while no significant malarial recurrence within 28\u00a0days in patients treated with chloroquine was observed. This indicated that patients with Plasmodium-mixed infection who are treated with ACTs similar to the treatment of patients with P. falciparum can still experience treatment failure, particularly patients with severe malaria. This result was supported in studies by Sikora et al. [Plasmodium-mixed infection, malarial recurrence can occur in patients with uncomplicated malaria, as demonstrated in the subgroup analysis of clinical signs. The risk of malarial recurrence was not observed in patients with mixed infections compared to those with P. vivax mono-infection. The significance of malarial recurrence in patients with uncomplicated malaria was observed in studies by Douglas et al. in Thailand [Plasmodium-mixed infection might be caused by many factors, including drug resistance or inappropriate antimalarial drugs [P. vivax transmission substantially [Infection by  malaria . A previns found . The prea et al.  and Ahmea et al. , which rThailand . This real drugs , severital drugs , 39, or antially .Plasmodium species and disease severity. In uncomplicated malaria, chloroquine and primaquine drugs are administered to treat P. vivax, while ACTs are administered to treat P. falciparum malaria since P. falciparum is resistant to chloroquine [Plasmodium-mixed species, WHO suggested that ACTs (except artesunate\u2009+\u2009sulfadoxine\u2013pyrimethamine) are effective against all malaria species and are the treatment of choice against mixed infection in co-endemic areas of P. falciparum and P. vivax [Treatment decisions on malaria infection are different based on oroquine . For theP. vivax . In all P. vivax . Since mP. vivax , intraveP. vivax , 33.P. vivax parasitemia after the treatment of P. falciparum or mixed Plasmodium species in Southeast Asia [P. vivax parasitemia is an important consideration for clinical drug trials of malaria control strategies in this region.Most of the studies included in this meta-analysis were performed on the Asian continent because of the emergence of multidrug resistance in driving the treatment policy changes, but fewer malaria treatment options are available in Asian countries. It was also due to the potential spread of ACT resistance in sub-Saharan Africa because of the progression of chloroquine resistance and sulfadoxine\u2013pyrimethamine resistance from Asia to sub-Saharan Africa in the past, which contributed to millions of childhood deaths , 41. Preast Asia , 28, 42.Plasmodium infection, which might result in a low statistical power to be applied over a large population. Some potentially eligible studies did not report the exact number of Plasmodium recurrences after treatment with antimalarial drugs, and no studies reported in non-English language were found for the present study, leading to a limited number of included studies. Second, the baseline characteristics of patients, such as age, could not be extracted in all the included studies since the treatment efficacy of malaria was improved as the age increased [Plasmodium species and resistance pattern driven by drug pressure. Therefore, there is a need for a careful interpretation of the RR of malarial recurrence in patients with mixed Plasmodium infection after treatment with antimalarial drugs. Fifth, the present meta-analysis included the publication of Douglas et al. [This study had limitations. First, there was a limited number of studies reporting the treatment of mixed ncreased . Therefos et al.  that pooPlasmodium-mixed species. Moreover, significant malaria recurrence of mixed infection occurred within 28\u00a0days after treatment with ACTs. Therefore, in regions where more than one Plasmodium species are endemic, the use of appropriate antimalarial drugs with increased vigilance is required and should be strengthened during and after treatment. Further research is required to gain a better understanding of the mechanisms involved in the recurrence observed after treatment with ACTs.The present findings showed a high prevalence of malarial recurrence after initial treatment of Additional file 1: Table S1. Search terms."}
+{"text": "Plasmodium falciparum. Transient asymptomatic liver function test elevations were previously observed in cipargamin-treated subjects in two trials: one in malaria patients in Asia and one in volunteers with experimentally induced malaria. In this study, the hepatic safety of cipargamin given as single doses of 10 to 150\u00a0mg and 10 to 50\u00a0mg once daily for 3\u00a0days was assessed. Efficacy results, frequency of treatment-emerging mutations in the atp4 gene and pharmacokinetics have been published elsewhere. Further, the R561H mutation in the k13 gene, which confers artemisinin-resistance, was associated with delayed parasite clearance following treatment with artemether\u2013lumefantrine in Rwanda in this study. This was also the first study with cipargamin to be conducted in patients in sub-Saharan Africa.The novel anti-malarial cipargamin (KAE609) has potent, rapid activity against This was a Phase II, multicentre, randomized, open-label, dose-escalation trial in adults with uncomplicated falciparum malaria in five sub-Saharan countries, using artemether\u2013lumefantrine as control. The primary endpoint was\u2009\u2265\u20092 Common Terminology Criteria for Adverse Events (CTCAE) Grade increase from baseline in alanine aminotransferase (ALT) or aspartate transaminase (AST) during the 4-week trial.Overall, 2/135 patients treated with cipargamin had\u2009\u2265\u20092 CTCAE Grade increases from baseline in ALT or AST compared to 2/51 artemether\u2013lumefantrine patients, with no significant difference between any cipargamin treatment group and the control group. Cipargamin exposure was comparable to or higher than those in previous studies. Hepatic adverse events and general safety and tolerability were similar for all cipargamin doses and artemether\u2013lumefantrine. Cipargamin was well tolerated with no safety concerns.This active-controlled, dose escalation study was a detailed assessment of the hepatic safety of cipargamin, across a wide range of doses, in patients with uncomplicated falciparum malaria. Comparison with previous cipargamin trials requires caution as no clear conclusion can be drawn as to whether hepatic safety and potential immunity to malaria would differ with ethnicity, patient age and or geography. Previous concerns regarding hepatic safety may have been confounded by factors including malaria itself, whether natural or experimental infection, and should not limit the further development of cipargamin.Trial registration ClinicalTrials.gov number: NCT03334747 (7 Nov 2017), other study ID CKAE609A2202The online version contains supplementary material available at 10.1186/s12936-021-04009-1. Plasmodium species by inhibiting PfATP4, a plasma membrane Na+-ATPase [Plasmodium falciparum and gametocytocidal activity [There is a need to develop new anti-malarials as emerging resistance to artemisinin derivatives threatens to undermine progress in malaria control. Cipargamin (KAE609/NITD609) is a novel anti-malarial spiroindolone analogue compound with promising and potent activity in clinical studies \u20133. Spiroactivity .Eight clinical trials with oral cipargamin have been conducted. A total of 152 healthy volunteers received single doses between 1 and 300\u00a0mg or 10\u2013150\u00a0mg daily doses for 3\u00a0days, and 57 patients received single doses between 10 and 75\u00a0mg or 30\u00a0mg daily for 3\u00a0days. In addition, 8 healthy volunteers were treated in a human challenge study ) at 10\u00a0mg single dose.P. falciparum and Plasmodium vivax infections, and was well tolerated in malaria patients [Cipargamin was well tolerated in healthy volunteers in Phase I trials. In Phase II trials, cipargamin showed rapid parasite clearance and fever resolution in both patients , 6, 7. FP. falciparum, transient Grade 2\u20133 LFT elevations were observed in 4/11 subjects treated with a 75\u00a0mg single dose. These were as follows: one case of alanine aminotransferase (ALT) elevation up to\u2009>\u20095\u2009\u00d7\u2009upper limit of normal (ULN) (Grade 3) with elevated baseline alkaline phosphatase (ALP), aspartate transaminase (AST) and ALT values; one case of ALT elevation\u2009>\u20095\u2009\u00d7\u2009ULN (Grade 3) combined with Grade 2 AST elevations, with normal AST/ALT baseline levels; and, two cases of Grade 2 (>\u20093\u2009\u00d7\u2009ULN) AST/ALT elevations in patients with normal baseline values. All elevations were transient and asymptomatic.In two of these trials, participants treated with cipargamin experienced transient abnormalities of liver function tests (LFTs). In a trial evaluating patients in Asia infected with In a human challenge trial, with healthy volunteers in Australia, transient Grade 3\u20134 LFT elevations were observed in 3/8 subjects following a single oral dose of 10\u00a0mg cipargamin \u201311, and \u00ae/Riamet\u00ae).A Phase II trial was designed to systematically address the potential hepatotoxicity of cipargamin in an actively controlled setting by comparison to the current standard of care, i.e., artemether\u2013lumefantrine  criteria [Eligible patients were men and women (\u2265\u200918\u00a0years old and\u2009\u2265\u200945\u00a0kg) with microscopic confirmation of acute uncomplicated falciparum malaria . Exclusion criteria included mixed criteria , active Open-label cipargamin treatment was administered in five sequential cohorts, using ascending single or multiple doses of cipargamin with a pause between cohorts for a review of safety data. Within each cohort, patients were randomized in parallel to treatment groups using interactive response technology (IRT). After being contacted by the investigator, the IRT assigned a randomization number  that linked the patient to a treatment arm and specified a unique medication number for the study drug to be dispensed to the patient. Randomization could be suspended within a cohort if patients experienced safety events. The study or a specific treatment arm could be stopped in the event of serious safety observations especially related to hepatic safety.A cipargamin starting dose of 10\u00a0mg (cohort 1) was chosen, as this was the minimum dose at which LFT elevations were observed in the IBSM trial. Following acceptable safety results in cohort 1, the dose was increased stepwise in subsequent cohorts up to a single dose of 150\u00a0mg and multiple doses of 50\u00a0mg (once daily (QD) for 3\u00a0days) according to the dosing schedule  and an optional sixth cohort in which patients would receive 110\u00a0mg or 225\u00a0mg single doses, depending on hepatic safety in cohort 5. If, based on risk\u2013benefit assessment, there was no added benefit expected to patients from using the 225\u00a0mg dose, the trial could be stopped after cohort 5. After completion of cohort 5, the safety review committee approved further dose escalation. However, the optional sixth cohort was not initiated as no added benefit was expected from dosing 225\u00a0mg due to expected overlapping exposures with 150\u00a0mg dose in previous cohort.Decision criteria  was used as an active comparator in each cohort. As the cipargamin dose in the first two cohorts was potentially sub-therapeutic, a minimum number (N\u2009=\u200910) of patients were included to detect possible changes in liver function parameters. In subsequent cohorts, higher numbers (N\u2009=\u200920) of patients were included. All patients were followed up for 28\u00a0days, with close monitoring in an inpatient setting for at least the first 3\u00a0days, followed by frequent outpatient monitoring. To be discharged, patients were required to yield two consecutive trial assessments with negative blood smears for The primary endpoint was at least two Common Terminology Criteria for Adverse Events (CTCAE) grades increase from baseline in ALT or AST during the 4-week trial period. Secondary endpoints reported here are standard safety and tolerability assessments of cipargamin . Two categories of LFT abnormalities/adverse events were considered during the course of the trial: (1) liver laboratory triggers, which required repeated assessments of the abnormal laboratory parameter, and, (2) liver events, which were considered as medically significant events, i.e., were to be reported as serious adverse events which consisted of marked elevations of LFTs and/or pre-specified events: ALT or AST\u2009>\u20095\u2009\u00d7\u2009ULN, ALP\u2009>\u20092\u2009\u00d7\u2009ULN (in the absence of known bone pathology), total bilirubin (TBL)\u2009>\u20092\u2009\u00d7\u2009baseline value, ALT or AST\u2009>\u20093\u2009\u00d7\u2009ULN and International Normalized Ratio\u00a0\u2009>\u20091.5, potential Hy\u2019s Law cases (defined as ALT or AST\u2009>\u20093\u2009\u00d7\u2009ULN and TBL\u2009>\u20092\u2009\u00d7\u2009ULN (mainly conjugated fraction) without a notable increase in ALP to\u2009>\u20092\u2009\u00d7\u2009ULN), any clinical event of jaundice , ALT or AST\u2009>\u20093\u2009\u00d7\u2009ULN accompanied by  malaise, fatigue, abdominal pain, nausea, or vomiting, or rash with eosinophilia, any adverse event potentially indicative of a liver toxicity. These events required close observation and follow-up monitoring. Samples for LFT were collected at baseline and 24, 48, 72, 96, 168, 240, 336, 504, and 672\u00a0h post-first dose.Triplicate electrocardiograms (ECGs) were performed at baseline, during and following treatments at multiple time points . Initial manual read-out was done locally in order to detect significant safety findings and allow for immediate response if needed. Additionally, all ECGs were assessed centrally by an independent and blinded (with age of patient identified) cardiologist.The primary (hepatic safety) and secondary  analyses were based on the safety analysis set . Data were summarized by cohort and treatment. For artemether\u2013lumefantrine, data from all cohorts were pooled and summarized. As the significance of safety related results was determined by both the frequency and severity of events, there was no pre-defined hypothesis for the primary variable . The following analyses were performed for the primary variable: proportion of patients with occurrence of the primary variable by cohort and treatment group and 95% confidence interval based on the exact confidence interval (Pearson\u2013Clopper method); and, a 2-sided Fisher exact test for each cipargamin treatment group compared to the pooled artemether\u2013lumefantrine group. General safety variables (adverse events (AEs), laboratory parameters, vital signs, electrocardiography) were summarized by treatment.For a cipargamin treatment group in each cohort, the probability of observing at least 2 safety events in 10 patients or 3 safety events in 20 patients is at least 80% if the true event rate is\u2009\u2265\u200930%, and less than 5% if the true event rate is\u2009\u2264\u20093%. Given the sample size of 10 to 20 patients per treatment group within a cohort, the power to detect treatment difference is low even with large treatment differences using a 2-sided statistical significance level of 5%. If the true rate is 50% for a cipargamin treatment group and 3% for the artemether\u2013lumefantrine treatment group, the power to detect between treatment difference is about 80% using a 2-sided test at the 20% significance level for n\u2009=\u200910 per treatment group or at the 10% significance level for n\u2009=\u200920 for a cipargamin treatment group and 10 for the artemether\u2013lumefantrine treatment group. With 4 cohorts for a total of 40 patients treated with artemether\u2013lumefantrine group, the power to detect treatment difference using 2-sided test at the 5% significance level is 77% and 91% for n\u2009=\u200910 and 20 patients treated with cipargamin, respectively, if the true rate is 40% for cipargamin and 5% for the artemether\u2013lumefantrine.The trial started on 16 November, 2017 and completed on 23 November, 2019. A total of 188 patients were randomized  were male. Cohorts were balanced in terms of baseline characteristics, except for baseline parasite counts, which tended to be higher in cohorts 4 and 5  than in cohorts 1 to 3  patients treated with cipargamin had at least two CTCAE grade increases from baseline in ALT or AST compared to 2/51 (3.92%) patients treated with artemether\u2013lumefantrine (detailed LFT results for these 4 cases are available in supplement). There was no significant difference in the proportion of patients with two CTCAE grade increases between any cipargamin cohort and the pooled artemether\u2013lumefantrine group  by day 8, with normalization by day 29. AST and TBL remained within the normal range.One patient treated with 150\u00a0mg single-dose cipargamin had an asymptomatic increase in ALT from Grade 1 at baseline  to Grade 3  on day 4, returning to Grade 1 on day 11 and normalized by day 15. Per-protocol, this event was reported as a serious adverse event. Cipargamin exposure for this patient was the lowest in the cohort Fig.\u00a0. This paTotal bilirubin exceeded 2\u2009\u00d7\u2009ULN in one patient treated with cipargamin and in one artemether\u2013lumefantrine patient. TBL normalized in both patients before day 29. Hy\u2019s law criteria were not met for any patient , 82.6% (19/23) of patients experienced no AST or ALT elevation, while 17.4% (4/23) had a Grade 1 ALT or AST increase. There were no Grade 2 or 3 events. In comparison, 74.1 (100/135) and 64.7% (33/51) of patients treated with cipargamin or artemether\u2013lumefantrine alone had no ALT or AST grade elevation.All cipargamin doses were tolerated well. There were no discontinuations due to AEss. AE rates were similar across treatment groups, with no obvious relationship to cipargamin dose. Gender did not appear to impact the incidence of AEs. Most AEs were considered not related to the study drug. Nineteen out of 135 (14.1%) patients treated with cipargamin and 5/51 (9.8%) patients treated with artemether\u2013lumefantrine had at least one AE suspected to be related to the study drug. None of these events was considered serious except for one event of increased ALT, as described above.The most common AEs were disease-related signs and symptoms, including headache, malaria and treatment failure. Malaria reported as an AE was defined as worsening malaria symptoms or recrudescence/re-infection. AEs of Grade 3\u20134 severity were reported for four cipargamin patients  and two artemether\u2013lumefantrine patients . These events were transient, not dose-related and resolved without treatment. Haematological abnormalities, including anaemia and thrombocytopaenia, of varying severity, are common in malaria patients .Hepatic AEs were reported in 10/135 (7.4%) patients in cipargamin groups as compared to 6/51 (11.8%) in the pooled\u00a0artemether\u2013lumefantrine group.Five non-fatal serious AEs were reported; four in cipargamin patients (Grade 2 bilirubin increase from day 15 to day 21 (50\u00a0mg QD for 3\u00a0days), Grade 2 ALP increase from day 2 (75\u00a0mg single dose), Grade 3 ALT increase from day 4 to day 11 (150\u00a0mg single dose), Grade 4 thrombocytopaenia from day 9 to day 35 (75\u00a0mg single dose) and one in an artemether\u2013lumefantrine patient (Grade 2 bilirubin increase from day 2 to day 8). Four of these events met protocol-defined serious AE criteria for elevations of LFTs. All were asymptomatic, transient and resolved within a few days without treatment. None was suspected to be related to the trial drug except for the Grade 3 ALT elevation (Table Minor elevations (>\u200930 to\u2009\u2264\u200960\u00a0ms) in QT interval corrected by Fridericia\u2019s formula (QTcF) occurred in all treatment groups, with frequencies from 9.1 to 36.8% in the cipargamin treatment groups and 27.5% in the pooled Coartem controls. Asymptomatic increases of\u2009>\u200960\u00a0ms were noted on day 29 in two patients treated with cipargamin 25\u00a0mg QD for 3\u00a0days, and on day 2 in one patient treated with 75\u00a0mg single-dose cipargamin. These did not appear to be related to cipargamin exposure. No patient had QTcF\u2009>\u2009500\u00a0ms. An increase of\u2009>\u200925% in heart rate (>\u2009100\u00a0bpm) was reported for one patient in the cipargamin 150\u00a0mg single dose group and in one artemether\u2013lumefantrine patient; while a heart rate decrease of\u2009>\u200925% (<\u200950\u00a0bpm) was reported for one patient each in the cipargamin 50\u00a0mg single-dose, 75\u00a0mg single-dose, and 25\u00a0mg multiple-dose groups. None of the abnormalities was considered clinically significant by the investigators, and hence not reported as AEs  (\u2265\u2009180\u00a0mmHg or increase from baseline by\u2009\u2265\u200920\u00a0mmHg) or diastolic BP (\u2265\u2009105\u00a0mmHg or increase from baseline by\u2009\u2265\u200915\u00a0mmHg). Clinically notably low SBP (\u2264\u200990\u00a0mmHg or decrease from baseline by\u2009\u2265\u200920\u00a0mmHg) or DBP (\u2264\u200950\u00a0mmHg or decrease from baseline by\u2009\u2265\u200915\u00a0mmHg) were observed in two patients treated with a single 75\u00a0mg dose of cipargamin compared to four patients treated with artemether\u2013lumefantrine.P. falciparum and one IBSM trial. No significant LFT elevations were identified in healthy volunteers given a single dose of up to 300\u00a0mg [There is an urgent need for the development of new non-artemisinin-based anti-malarial drugs. Cipargamin is a promising new anti-malarial that showed rapid parasite clearance in Phase II trials. Some transient, mainly asymptomatic, LFT elevations occurred in two previous trials with cipargamin; one in patients with o 300\u00a0mg .The trial reported here evaluated the hepatic safety of cipargamin in adult patients with uncomplicated falciparum malaria, applying a dose escalation design, using 10\u00a0mg to 150\u00a0mg single doses and 10\u00a0mg to 50\u00a0mg QD for three days, and a control arm. The low rate of LFT abnormalities in patients treated with cipargamin was comparable to that in artemether\u2013lumefantrine patients. There was no obvious relationship between LFT abnormalities and cipargamin dose or exposure. Exposure to cipargamin in this trial was higher or comparable to previous trials [The SRC did not observe any serious hepatic safety concerns after cohorts 1, 3, 4, and 5, and approved further dose escalation after each review. No further escalation was made as no increased benefit was expected for the patients, so the trial was considered complete after completion of cohort 5.As cipargamin will be further developed as a combination treatment for uncomplicated malaria, good tolerability, including liver safety, when administered with other anti-malarials is important. The current trial included only adults from sub-Saharan African countries with falciparum malaria. Comparison with the two previous cipargamin trials  therefore requires caution as no clear conclusion can be drawn as to whether hepatic safety and potential immunity to malaria would differ with ethnicity, patient age and or geography. The differences in baseline parasite counts in the current trial may complicate comparisons between cipargamin doses and between cipargamin and artemether\u2013lumefantrine in terms of safety. In addition, patients with known liver abnormalities or disease were excluded from the current trial, thereby the effects of cipargamin on those with pre-existing liver abnormalities cannot be determined from this trial.Abnormalities in liver function parameters are common in malaria patients and have also been observed in volunteers with experimental infection. In IBSM studies, LFT elevations appear to be specific to the experimental model as similar results were seen with multiple tested compounds of different chemical classes , 12, 15.In malaria patients, changes in LFT results are likely to be an inherent, although variable, aspect of the disease; individual-specific factors may confer particular susceptibility to hepatocyte injury. These abnormalities are transient and may vary widely in severity (up to\u2009>\u200925 ULN) . ElevateCipargamin was well tolerated in this study without any hepatic safety concerns at a wider dose range and higher exposures than previous studies. This trial, the first conducted in Africa and the largest carried out to date with cipargamin, showed the hepatic safety of cipargamin was comparable to that of artemether\u2013lumefantrine in adult patients with uncomplicated falciparum\u00a0malaria. This indicates that elevations in liver function tests observed in previous trials with cipargamin are likely to be malaria-related in patients and confounded by the experimental model in the human challenge trial and lack of an active control.\u00a0This study paves the way for the further development of this novel anti-malarial. In addition to the good tolerability of cipargamin, this study has confirmed its potency and rapid onset of parasiticidal activity with parasite clearance times of around 8\u00a0h for doses of 50\u00a0mg or higher . FurtherThis example of monitoring liver safety of clinical candidates in a disease that itself is causing increases in liver function tests and the challenge of dissecting the potential drug related causality from disease background should inform future development programmes in related areas.Additional file 1. Supplement Hepatic safety and tolerability of cipargamin (KAE609)."}
+{"text": "Plasmodium vivax recurrence in patients treated for Plasmodium falciparum malaria in co-endemic areas. Primaquine radical cure has the potential to reduce P vivax recurrences in patients presenting with P falciparum as well as P vivax malaria but is undermined by poor adherence to the currently recommended 14-day regimen. We aimed to assess the efficacy and safety of supervised versus unsupervised primaquine radical cure in patients presenting with uncomplicated malaria.There is a high risk of P falciparum or P vivax malaria with oral dihydroartemisinin\u2013piperaquine plus either a supervised or unsupervised 14-day course of oral primaquine (0\u00b75 mg/kg per day). Patients in the supervised group were supervised taking their primaquine dose on alternate days. Patients were followed-up for 6 months and those who presented again with malaria were retreated with the same drug regimen. Masking was not possible due to the nature of the study. The primary outcome was the incidence risk of P vivax malaria over 6 months, assessed in the modified intention-to-treat population . This trial is now complete, and is registered with ClinicalTrials.gov, NCT02787070.We did a cluster-randomised, controlled, open-label superiority trial in Papua, Indonesia. 21 clusters of village health posts, matched by annual parasite index, were randomly assigned (1:1) to treat patients (age >12 months and body weight >5 kg) presenting with confirmed uncomplicated P vivax recurrence in the supervised group was 29\u00b77% (95% CI 16\u00b74\u201349\u00b79) versus 55\u00b78% (32\u00b73\u201381\u00b78) in the unsupervised group . The incidence rate for P vivax recurrence was 539 (95% CI 390\u2013747) infections per 1000 person-years in the supervised group versus 859 (673\u20131096) in the unsupervised group . The corresponding rates in the 224 patients who presented with P falciparum malaria were 346 (95% CI 213\u2013563) and 660 . Seven serious adverse events were reported , none of which were deemed treatment-related, and there were no deaths.Between Sept 14, 2016, and July 31, 2018, 436 patients were screened for eligibility and 419 were enrolled; 223 (53%) patients in 11 clusters were assigned to supervised primaquine treatment and 196 (47%) in ten clusters to unsupervised primaquine treatment. 161 (72%) of 223 patients in the supervised group and 151 (77%) of 196 in the unsupervised group completed 6 months of follow-up. At 6 months, the incidence risk of P vivax recurrence. This finding was apparent for patients presenting with either P falciparum or P vivax malaria. Further studies are warranted to investigate the safety and efficacy of radical cure for patients presenting with uncomplicated falciparum malaria in other co-endemic areas.In this area of moderate malaria transmission, supervision of primaquine radical cure treatment reduced the risk of The Bill & Melinda Gates Foundation, Wellcome Trust, and Department of Foreign Affairs and Trade of the Australian Government.For the Indonesian translation of the abstract see Supplementary Materials section. Plasmodium vivax malaria reported worldwide each year from 49 endemic countries.P vivax is the predominant cause of malaria.P vivax is more difficult to eliminate than Plasmodium falciparum, because it forms dormant liver stages (hypnozoites) that can reactivate weeks to months after an initial infection, causing recurrent symptomatic illness (relapses). The risk and frequency of relapse varies considerably with geographical location.P vivax parasitaemia results in a cumulative risk of anaemia, and both direct and indirect morbidity and mortality, particularly in young children and pregnant women.There are between 7\u00b75 million and 14\u00b75 million cases of Evidence before this studyPlasmodium vivax recurrence following Plasmodium falciparum infections, using the terms \u201cvivax\u201d, \u201cfalciparum\u201d, and \u201crecurrence\u201d. The risk of P vivax recurrence within 63 days of treatment for P falciparum malaria exceeded 15% across a range of co-endemic areas, highlighting a potential benefit of primaquine radical cure for P falciparum malaria. The risk of P vivax recurrence within 12 months was less than 10% following supervised high-dose primaquine  treatment plus dihydroartemisinin\u2013piperaquine, but increased to more than 88% when primaquine treatment was unsupervised. As of Feb 28, 2021, no trials were identified that quantified the efficacy or effectiveness of supervised 14-day primaquine or single-dose tafenoquine for the treatment of both vivax and falciparum malaria.We searched PubMed, MEDLINE, Web of Science, Embase, and the Cochrane Database of Systematic Reviews from Jan 1, 1960, to Feb 28, 2021, for relevant clinical trials and systematic reviews, published in English, on the risk of Added value of this studyP vivax recurrence in patients presenting with either P vivax or P falciparum malaria.This cluster-randomised, controlled, open-label, superiority trial showed that supervision of primaquine radical cure treatment on alternate days reduced the risk of Implications of all the available evidenceP vivax or P falciparum, has potential to reduce the risk of recurrent P vivax parasitaemia, which could reduce ongoing transmission and facilitate malaria elimination.In areas of moderate malaria transmission, active measures to ensure patient adherence to a complete course of primaquine radical cure treatment, and extending its use to all patients presenting with uncomplicated malaria due to either P vivax liver stages and prevent relapse. Supervised administration of artemisinin-based combination therapy and a high dose of primaquine reduces the risk of recurrence, even in areas with high relapse periodicity,The radical cure of malaria refers to a combination of drugs to kill both the blood and liver stages of the parasite. Primaquine, an 8-aminoquinoline, is currently the only widely available drug that can kill P vivax or Plasmodium ovale, the only species causing human malaria that form hypnozoites.P vivax parasitaemia in patients following treatment for P falciparum malaria.P vivax originated from reactivation of occult hypnozoites in patients exposed to both species, potentially triggered by the acute febrile illness of P falciparum malaria.P vivax or P falciparum monoinfection or a mixed infection with both species.Radical cure of malaria is currently restricted to patients presenting with P falciparum infection, 45% to P vivax, and the remainder due to Plasmodium malariae or mixed species infections.15We did a cluster-randomised, controlled, open-label superiority trial in Papua, Indonesia. The climate, geography, malaria endemicity, and demographics of the study site have been described previously.The unit of randomisation was the village health post. 21 village health posts (clusters) were included in the study and randomly assigned to one of the study treatment groups. Each health post was associated with one of eight public clinics where initial recruitment and treatment occurred. Clinics were selected to be located within 1\u00b75 h drive from the research office and from those that treated more than 200 cases of malaria per year .Written informed consent, or consent from the legal guardian, was required for participation. Ethical approval was obtained from the Human Research Ethics Committee of the Northern Territory Department of Health, Australia (HREC 15.2517) and the Health Research Ethics Committees of the University of Gadjah Mada, Indonesia (KE/FK/522/EC/2016).P falciparum or P vivax malaria (including either monoinfection or mixed infections), who were older than 12 months and with body weight greater than 5 kg, were eligible for enrolment. Patients were excluded if they were pregnant (determined by history and urinary \u03b2-human chorionic gonadotropin test), lactating, G6PD deficient , anaemic (haemoglobin <9 g/dL), or had signs or symptoms of severe malaria using the criteria defined by WHO.Febrile patients living in one of the study cluster villages and attending one of the eight public clinics with suspected malaria were screened for malaria by microscopic examination of Giemsa-stained peripheral blood film by a laboratory microscopist at the clinics. All slides were re-read the same day by an expert trial microscopist. Patients with confirmed uncomplicated The 21 clusters were selected according to location, size, and malaria transmission. Individual clusters with similar annual parasite index were then randomly assigned (1:1) to either the supervised or unsupervised primaquine treatment groups using Stata version 15.1. The independent statistician who generated the randomisation list and allocated clusters to treatment groups was not otherwise involved in the conduct of the trial. Masking was not possible due to the nature of the study.After obtaining informed consent, eligible patients were enrolled and a baseline clinical questionnaire and examination were completed at the public clinic. All participants were immediately commenced on a 3-day regimen of oral dihydroartemisinin\u2013piperaquine provided by the national malaria control programme according to national guidelines . All parPatients residing in the clusters assigned to unsupervised treatment were prescribed a course of oral primaquine according to local treatment guidelines, at a daily dose of 0\u00b75 mg/kg per day, and were instructed to take the prescribed tablets once daily for 14 days . In the +; HemoCue, \u00c4ngelholm, Sweden). Blood film microscopy during follow-up was done by the study laboratory technicians, who were masked to treatment allocation.At each study visit, a medical history and symptom questionnaire were completed and any adverse events or serious adverse events were recorded by a research nurse, and referred to the study clinician as necessary. Patients were encouraged to present to the study centre if they became unwell at any time during the study. At each routine review or upon presentation with symptoms compatible with malaria, a capillary blood sample was taken for peripheral blood film examination and measurement of haemoglobin concentration  or blood transfusion over 6 months, and an acute fall in haemoglobin greater than 5 g/dL or fractional fall of greater than 25% to a concentration of less than 7 g/dL within 14 days of starting primaquine treatment.P falciparum parasitaemia at 6 months was compared between supervised and unsupervised clusters.In a post-hoc analysis, the incidence risk and rate of P vivax recurrence over 6 months from 30% in the unsupervised group to 10% in the supervised group.The required sample size was calculated to detect an absolute reduction of 20% in the incidence risk of The combined data from all clusters were analysed to provide a pragmatic comparison of the different treatments using a modified intention-to-treat strategy; analyses were done per the assigned treatment groups, regardless of whether participants were actually supervised or not, but excluding patients who were lost to follow-up after their first visit. Descriptive statistics of patient and disease characteristics at baseline were calculated by treatment group with frequency and percentage presented for categorical variables, and median with corresponding IQR for continuous variables. A clinically relevant decrease in haemoglobin was defined as an absolute decrease from baseline of more than 5 g/dL or a fractional fall of more than 25% to a concentration of less than 7 g/dL. Overall adherence to primaquine was evaluated by calculating the proportion of participants in each group who received a total dosage of at least 5 mg/kg of primaquine.P vivax recurrence over 6 months in each treatment group. For the primary endpoint, hazard ratios (HRs) with 95% CIs were estimated using mixed-effects Cox proportional hazards regression with a time-varying coefficient for treatment effect  and a frailty term for clustering for time to first P vivax recurrence analyses. Incidence rate ratios (IRRs) were estimated using negative binomial regression with robust SE estimation (to account for clustering) for analysis of all P vivax recurrences. Details regarding censoring for the first P vivax recurrence, and safety analyses are provided in an a priori statistical plan  had P vivax monoinfection, and 12 (3%) had a mixed species infection. Baseline characteristics were similar between the supervised and unsupervised treatment groups  patients in 11 clusters were assigned to supervised primaquine treatment and 196 (47%) in ten clusters to unsupervised primaquine treatment . At enrot groups . Seven pThe initial response to schizontocidal treatment was rapid. Within 48 h, 399 (99%) of 405 patients had become afebrile and 375 (93%) had cleared their peripheral parasitaemia, with no significant difference in fever or parasite clearance between treatment groups. Clearance times could not be calculated in seven patients due to missed visit on day 2. On day 2 review, 213 (96%) of 223 patients in the supervised group and 191 (97%) of 196 in the unsupervised group with haemoglobin of at least 9 g/dL were prescribed primaquine . During the period of primaquine treatment, 184 (83%) of 223 patients in the supervised group completed all required visits. 161 (72%) of 223 patients in the supervised group and 151 (77%) of 196 in the unsupervised group completed 6 months of follow-up.P vivax recurrence was 29\u00b77% (95% CI 16\u00b74\u201349\u00b79) in the supervised group versus 55\u00b78% (32\u00b73\u201381\u00b78) in the unsupervised group . The HR was 0\u00b778  at day 60, and 0\u00b743  at day 120.At 6 months (day 180), the cumulative incidence risk of ed group , table 2P falciparum, one with P vivax, and none with mixed infections. By 6 months, 95 patients had recurrence with P falciparum, 110 with P vivax, and six with mixed infections (P vivax recurrence was 539 (95% CI 390\u2013747) infections per 1000 person-years observed in the supervised group versus 859 (673\u20131096) in the unsupervised group . In the cluster-level analysis, the incidence rate of P vivax recurrence was 538 (95% CI 312\u2013765) infections per 1000 person-years observed in the supervised group versus 862 (623\u20131100) in the unsupervised group  of 95 P falciparum recurrences were symptomatic at the time of presentation to the clinic.At day 28, four patients had recurrent parasitaemia , three of whom were infected with fections . The incp=0\u00b7053; . OverallP vivax infection (monoinfection or mixed species), the risk of the first P vivax recurrence at 6 months was significantly lower in the supervised group than in the unsupervised group . In the 224 patients who initially presented with P falciparum infection, the corresponding HR at 6 months was 0\u00b733  and the IRR was 0\u00b752 .In the 195 patients who initially presented with P falciparum recurrence, the cumulative incidence risk of P falciparum parasitaemia at 6 months was 27\u00b71% (95% CI 21\u00b72\u201334\u00b72) in the supervised group and 36\u00b74% (21\u00b78\u201356\u00b75) in the unsupervised group; the corresponding incidence rates for all P falciparum infections were 646 (95% CI 503\u2013830) infections per 1000 person-years and 544 (405\u2013730), respectively. Neither the incidence risk nor the rate of P falciparum recurrence differed significantly between treatment groups.The cumulative incidence risk of any recurrent parasitaemia at 6 months was 50\u00b70% (95% CI 35\u00b71\u201367\u00b71) in the supervised group versus 80\u00b76% (49\u00b79\u201398\u00b70) in the unsupervised group . In post-hoc analyses that examined the comparative risks of The total dose of primaquine administered was calculated in all 223 patients in the supervised group and 155 (79%) of 196 in the unsupervised group. Although the median total dose of primaquine prescribed was similar between groups (7\u00b73 mg/kg [IQR 6\u00b71\u20138\u00b72] in the supervised group and 7\u00b71 mg/kg [5\u00b77\u20138\u00b70] in the unsupervised group), only 126 (64%) patients in the unsupervised group took 5 mg/kg or greater, compared with 199 (89%) in the supervised group.P falciparum infection compared with 4\u00b72% (2\u00b74\u20135\u00b79) in patients with P vivax infection (p=0\u00b715). On day 16, after patients had completed primaquine treatment, the mean haemoglobin among patients in the supervised group was 11\u00b75 g/dL (95% CI 11\u00b73\u201311 \u00b77) and 11 (6%) of 195 patients had a fractional fall of more than 25% from baseline; the mean haemoglobin among patients in the unsupervised group was 11\u00b76 g/dL (11\u00b74\u201311\u00b78) and eight (5%) of 171 patients had a fractional fall from of more than 25% from baseline . One male patient aged 15 years, who presented with falciparum malaria and was treated with supervised primaquine, had a clinically significant fall in haemoglobin, from 9\u00b75 g/dL at baseline to 6\u00b77 g/dL on day 16; he remained asymptomatic and did not require hospitalisation, and by day 28 his haemoglobin had risen to 9\u00b70 g/dL without the need for blood transfusion. The haematological profiles during follow-up by treatment group are presented in Overall, the mean haemoglobin concentration fell from 11\u00b79 g/dL (95% CI 11\u00b77\u201312\u00b71) at baseline to 11\u00b72 g/dL (11\u00b71\u201311\u00b74) on day 2, before the first dose of primaquine. The fractional fall in haemoglobin on day 2 was 5\u00b79% (95% CI 4\u00b73\u20137\u00b75) in patients with baseline . Three pThe analysis of early primaquine tolerability between days 2 and 16 was confined to patients in the supervised group; participants in the unsupervised group were assessed on days 2 and 16 only. None of the patients in the supervised group reported vomiting their primaquine dose within 1 h of administration. 23 (11%) of 215 patients in the supervised group had gastrointestinal symptoms during their course of primaquine treatment , although the difference didn't begin to emerge until after 90 days , the high-dose 14-day primaquine regimen was well tolerated and was not associated with severe adverse complications. However, six patients had a fall in haemoglobin greater than 5 g/dL, four of whom presented with nfection , which mGastrointestinal intolerance is an acknowledged side-effect of primaquine and although it is dose-related, it can be reduced by concomitant administration of food. Less than 5% of patients vomited their dose of primaquine within 1 h of administration, and subsequent rates of vomiting after clinical recovery were even lower. Two patients with normal G6PD levels reported dark urine, but subsequent investigation suggested that the dark colour was due to dehydration rather than haemoglobinuria. None of the patients required admission to hospital due to primaquine treatment .P vivax relapses, and the corresponding dose of primaquine required to achieve high anti-relapse efficacy, vary considerably between geographical regions.P vivax relapse, with the risk at 12 months ranging from 7% to 20%,P vivax following P falciparum also varies with the background prevalence of malaria and relapse periodicity.P falciparum malaria. Further studies addressing these issues in different endemic settings are warranted.The risk and frequency of P falciparum infections in the supervised group than in the unsupervised group, potentially reflecting greater heterogeneity between clusters than expected. A higher intracluster correlation coefficient than predicted might have led to the study being underpowered, particularly for the secondary analyses.Our study has some limitations. Follow-up was incomplete, with only 72\u201377% of participants contributing to the primary endpoint. The loss to follow-up was attributable to the high patient mobility of migrant workers. Loss to follow-up might have selected patients with a more positive attitude towards treatment adherence and thus led to underestimation of the true effect of the intervention. In the unsupervised treatment clusters, the study provided more consultation time with clinic or study staff than patients would have in real-world settings,P vivax infection significantly, in patients presenting with either P vivax or P falciparum malaria. Extending the use of safe and effective primaquine radical cure to patients presenting with non-vivax malaria should be considered, but will need to be tailored to areas with a high risk of P vivax recurrence after P falciparum infection.P vivax substantially, paving the way for ambitious malaria elimination targets to be met.In conclusion, partial supervision of 14-day primaquine treatment enhanced adherence and reduced incidence and rates of recurrent malariaDAC@iddo.org via the data access form available at https://www.wwarn.org/working-together/sharing-accessing-data/accessing-data. The WWARN is registered with the Registry of Research Data Repositories.The data are available for access via the WorldWide Antimalarial Resistance Network (WWARN). Requests for access will be reviewed by a data access committee to ensure that use of data protects the interests of the participants and researchers according to the terms of ethics approval and principles of equitable data sharing. The study protocol and statistical analysis plan are provided in the We declare no competing interests."}
+{"text": "The current first-line treatments for uncomplicated malaria recommended by the National Malaria Control Programme in Mali are artemether\u2013lumefantrine (AL) and artesunate\u2013amodiaquine (ASAQ). From 2015 to 2016, an in vivo study was carried out to assess the clinical and parasitological responses to AL and ASAQ in S\u00e9lingu\u00e9, Mali.Plasmodium falciparum infection and 2000\u2013200,000 asexual parasites/\u03bcL of blood were enrolled, randomly assigned to either AL or ASAQ, and followed up for 42\u00a0days. Uncorrected and PCR-corrected efficacy results at days 28 and 42. were calculated. Known markers of resistance in the Pfk13, Pfmdr1, and Pfcrt genes were assessed using Sanger sequencing.Children between 6 and 59\u00a0months of age with uncomplicated Pfk13 gene. Overall, for Pfmdr1, the N86 allele and the NFD haplotype were the most common. The NFD haplotype was significantly more prevalent in the post-treatment than in the pre-treatment isolates in the AL arm (p\u2009<\u20090.01) but not in the ASAQ arm. For Pfcrt, the CVIET haplotype was the most common.A total of 449 patients were enrolled: 225 in the AL group and 224 in the ASAQ group. Uncorrected efficacy at day 28 was 83.4% (95% CI 78.5\u201388.4%) in the AL arm and 93.1% (95% CI 89.7\u201396.5%) in the ASAQ arm. The per protocol PCR-corrected efficacy at day 28 was 91.0% (86.0\u201395.9%) in the AL arm and 97.1% (93.6\u2013100%) in the ASAQ arm. ASAQ was significantly (p\u2009<\u20090.05) better than AL for each of the aforementioned efficacy outcomes. No mutations associated with artemisinin resistance were identified in the The findings indicate that both AL and ASAQ remain effective for the treatment of uncomplicated malaria in S\u00e9lingu\u00e9, Mali.The online version contains supplementary material available at 10.1186/s12936-021-03760-9. Integrated strategies to control malaria have led to a decrease in the number of malaria cases and deaths. The incidence rate of malaria declined globally between 2010 and 2018, from 71 to 57 cases per 1000 population at risk though the statistical significance of this decline has not been reported [Plasmodium falciparum malaria [In 2001, treatment failure using chloroquine (CQ) was linked to an increase in child mortality in Africa . In 2006 malaria . ACT is  malaria . TherefoDeclining efficacy of ACT in Southeast Asia  has led to heightened concern for the spread of resistant parasites to other malaria endemic countries , 6 and tA handful of studies over the past decade have investigated the efficacy of AL in Mali and reported PCR-corrected efficacies greater than 98%. These include: a 2010\u20132014 study in Sotuba, Koll\u00e9, and Banambani . AlthougP. falciparum parasites to anti-malarial drugs provides an additional way of assessing whether efficacy may be threatened. For instance, certain mutations in the propeller domain of the P. falciparum kelch 13 (Pfk13) gene are associated with artemisinin partial resistance [I, N458Y, M476I, Y493H, R539T, I543T, P553L, R561H, P574L,and C580Y. To date, there have been no reports of Pfk13 polymorphisms associated with artemisinin partial resistance in Mali. Polymorphisms in P. falciparum multidrug resistance-1 (Pfmdr-1) and P. falciparum chloroquine resistance transporter (Pfcrt) genes are associated with recurrent parasitaemia after treatment with AL and ASAQ [Pfmdr1 are found at codons N86Y, Y184F, and D1246Y [Y mutation has been associated with decreased chloroquine and amodiaquine sensitivity while the N86 has been implicated in decreased sensitivity to lumefantrine. The N86, 184F, and D1246 (NFD) haplotype is associated with decreased sensitivity to AL, while the 86Y, Y184, and 1246Y (YYY) haplotype is reported to be associated with decreased sensitivity to ASAQ [pfcrt mutations are observed in codons 72, 74\u201376 [IET haplotypes are associated with chloroquine resistance. The Pfcrt 76\u00a0T mutation has also been shown to play a role in amodiaquine resistance in some studies in Africa [Surveillance of molecular markers associated with the resistance of sistance , in partsistance : F446I, and ASAQ . In Afrid D1246Y . The 86Y to ASAQ . The mosn Africa . Prior sn Africa  or ASAQ n Africa , 20\u201322.P. falciparum endemic areas, a therapeutic efficacy study of AL and ASAQ for the treatment of uncomplicated malaria was carried out. In addition, the study investigated the prevalence of molecular markers associated with resistance and reduced susceptibility to AL and ASAQ.To fulfill the WHO recommendation of performing biannual monitoring of parasite susceptibility to anti-malarial drugs in The study was conducted in S\u00e9lingu\u00e9, a commune in the southern part of Mali in the Sikasso region , weight less than 9.0\u00a0kg, the presence of any acute comorbidity , or with a history of taking anti-malarial drugs in the four past days were not included in the study and were referred to the health centre for management. In addition, those children whose parents refused to participate or lived outside the radius of 5\u00a0km from S\u00e9lingu\u00e9 Health centre were excluded from the study. After consent was obtained from parents, enrolled patients were randomly assigned to either the AL or ASAQ arm, with a ratio of 1:1, using a random number generator with PASS11 . The study was single blinded and the randomization was implemented by the research pharmacist. Dosage was weight-based according to manufacturer recommendations. Clinic staff, including physicians and laboratory personnel involved in the study, were blinded to the allocation except for the research pharmacist who provided the treatment. The research pharmacist administered all treatment drugs to each patient on days 0 (day of enrollment), 1, and 2. Blood samples were collected on the day of enrollment (pre-treatment); days 1, 2, and 3; and weekly from days 7 to 42. Participants were followed up with physical and clinical exams after anti-malarial administration to evaluate for adverse medication effects or recurrent parasitaemia. Enrolled patients were requested to return to the clinic if they experienced any symptoms of malaria during the follow-up period. The study schedule was available to all parents of each study participant. A blood slide for microscopy and dried blood spots on filter paper were prepared any time a blood sample was collected.The efficacy of AL and ASAQ was assessed in a prospective two-arm study conducted from November 2015 to November 2016, as per the WHO protocol for surveillance of anti-malarial drug efficacy . ParticiPlasmodium species, a third slide reading was performed by another senior microscopist and the closest readings averaged for a final parasite count.Thick and thin malaria blood smears were prepared on glass slides and stained with 3% Giemsa  in phosphate buffer (pH 7.0). The slide reading was performed independently by two experienced microscopists at the study sites. Parasite density was calculated according to well-established methods , brieflyA sample size of 240 patients per arm was targeted in order to provide adequate power to estimate an efficacy of 95% with a confidence interval of \u00b1\u20095%, assuming an expected efficacy of 95% and a maximum loss to follow-up and withdrawal rate of 20%. This was based on the ability of the drug to clear parasitaemia at day 7 or before and give enough sample size to compare the efficacy of the two treatment arms.Each enrolled child received a three-day course of either AL  or ASAQ . All doses administered were given under the direct observation of the study pharmacist at the S\u00e9lingu\u00e9 health centre and both AL and ASAQ were given with a peanut-based paste and milk. For ASAQ-Denk fixed dose, the dosage was adjusted based on the patient\u2019s body weight (as recommended in the package\u00a0insert), 2 to 10\u00a0mg of artesunate per kg; 7.5 to 15\u00a0mg of amodiaquine once daily for 3\u00a0days. Enrolled participants randomly assigned to AL , received a weight-adjusted dosing of 20\u00a0mg of artemether and 120\u00a0mg of oral lumefantrine twice a day for 3\u00a0days as recommended in the insert package, 1 tablet for 5\u201314\u00a0kg, 2 tablets for 15\u201324\u00a0kg, and 3 tablets for 25\u201334\u00a0kg. All children were observed for a minimum of one hour after each dose to monitor for vomiting or other side effects. In case of vomiting, another dose of the drug was then administered. After the three days of dosing, each enrolled child was followed on days 7, 14, 21, 28, 35, and 42. During each visit, the following were performed or collected: physical exam , temperature, thin and thick blood smears, and dried blood spots. Haemoglobin levels were measured at the time of enrollment and on day 42, at the completion of the study. All blood specimens were collected by finger prick. In cases of recurrent parasitaemia, quinine was administered according to the guidelines of the Mali National Malaria Control Programme.Treatment responses were classified following the WHO guidelines . Treatmemsp1 and msp2), and glutamine-rich protein (glurp) markers, and Sanger sequencing of P. falciparum parasites was performed on samples obtained from participants on enrollment day (pre-treatment) and on the day of recurrent parasitaemia (post-treatment). Genomic DNA was extracted from all collected samples using the QiAamp mini kit  following the manufacturer\u2019s instructions. Primers designed to amplify three allelic families from block 2 of msp1 , two allelic families from msp2 (FC27 and IC/3D7), and the polymorphic region of glurp were used in PCR amplification and analysis as previously described [msp1 and msp2, bands with lengths within 10 base pairs were considered a match; for glurp, lengths within 50 base pairs were considered a match. If there was at least one matching band in any allelic family for all three markers, the recurrence was classified as a recrudescence .\u00a0If there were no shared alleles for at least one marker, the recurrence was classified as a reinfection. If there were no amplification products resulting in sharp, defined bands in both the pre-treatment and day of recurrence samples for a gene, that gene was not used to distinguish between recrudescence and reinfection, but the aforementioned classification criteria were applied for the genes that were amplified.Genotyping using the merozoite surface proteins 1 and 2 , and Pfmdr1 codons 86, 184, and 1246 were analysed for SNPs. The P. falciparum laboratory strain 3D7 Pfcrt, Pfk13, and Pfmdr1 were used as reference sequences for the analysis. Molecular analyses were performed in collaboration with the U.S. Centers for Disease Control and Prevention (CDC) Malaria Laboratory in Atlanta, USA, as part of the President\u2019s Malaria Initiative (PMI)-supported Anti-malarial Resistance Monitoring in Africa Network [Paired pre-treatment and day of recurrent parasitaemia samples were assessed for known markers of resistance in the  primers , 29. Dir Network .P. falciparum. Uncorrected and PCR-corrected ACPR for AL and ASAQ were compared at both day 28 and 42 using a chi-square test. The capacity of the two ACTs to clear parasites by day 7 and the post-treatment prophylactic effect of the two treatments was evaluated using a previously described approach by Koita et al. [Both per protocol and Kaplan\u2013Meier analyses were conducted to evaluate the study outcome data. The main difference between the two is the Kaplan\u2013Meier approach takes into account patients who were withdrawn from the study, such as for acquiring a reinfection, and censors them at the day of reinfection, whereas the per protocol approach removes those with reinfection from the analysis altogether in the PCR-corrected calculations. Treatment responses were classified following the WHO guidelines . Uncorrea et al. . This apa et al. ), and tha et al. ). CorresPfk13, Pfmdr1, and Pfcrt genes were reported as single or mixed (wild-type and mutant) infections. For samples with mixed infections and SNP variations at multiple sites, each possible haplotype constructed from the observed SNPs was reported for Pfmdr1. In reporting Pfcrt haplotypes in samples with a mixed infection and SNP variations at multiple sites, the wild-type (CVMNK) and most likely mutant type  were reported. The prevalence of the Pfmdr1 mutant alleles and haplotypes was calculated stratifying by the treatment arms (AL and ASAQ) and compared between the pre-treatment samples (samples with ACPR and samples from subjects that later had reinfections) and post-treatment samples (reinfections and recrudescent infections). Likewise, the prevalence of the Pfcrt mutant alleles and haplotypes was compared between pre-treatment and post-treatment samples in the ASAQ arm; only samples from the ASAQ arm were used because of the known association between Pfcrt and amodiaquine efficacy [versus mutant alleles were compared between groups . Mixed infections were excluded. To compare haplotypes, the sum of samples with the predominant haplotype was compared to the sum of those without that haplotype . Statistical significance was defined as p\u2009<\u20090.05 for all statistical tests. Analyses were performed using Graph Pad Prism version 6.00 for Windows  and R .Point mutations in the efficacy . For theEthical review and approval was obtained from the Internal Review Board of the Institut National de Recherche en Sant\u00e9 Publique . Work performed at the Centers for Disease Control and Prevention was deemed to not constitute engagement in human subject research . Parents or guardians of study participants were asked to provide written informed consent.A total of 3732 patients were screened for the study and, of these, 449 were enrolled, 225 in the AL group and 224 in the ASAQ group . Both AL and ASAQ were able to clear parasitaemia before or on day 7 with 99.5% and 100% clearance rates, respectively (post-treatment prophylaxis). A total of 75 patients had recurrent infections during the 42-day follow-up period, 34 of whom had recurrent infections during the first 28\u00a0days. The uncorrected efficacy rates on days 28 and 42 were higher in the ASAQ group than in the AL group (Table The day 28 uncorrected efficacy was 83.4% (95% CI 78.5\u201388.4%) for AL and 93.1% (95% CI 89.7\u201396.5%) for ASAQ, which was significantly different . Any of the known mutations associated with artemisinin partial resistance in the Pfk13 gene was not observed. However, we observed synonymous polymorphisms in some pretreatment samples at codons: T478T, Y493Y, K503K, and Q613Q, all in one sample each. The synonymous polymorphism C469C was found in two samples and the A578S mutant allele was observed in two samples (pre-treatment and day 28) collected from the same patient.A total of 296/386 (76.6%) samples were successfully sequenced (23.4% of the samples did not give interpretable data likely due to low parasite density) for polymorphisms in the pfmdr1 gene (314 pre-treatment and 74 post-treatment samples). Overall, the wildtype N86 allele was found in a majority of the successfully sequenced pre-treatment  and post-treatment  isolates. Similarly, the overall prevalence of the 184F allele was high in pre-treatment  and in the post-treatment isolates  but not in the ASAQ arm . A similar trend was observed for the prevalence of the N86 allele, which was higher in the post-treatment (97.2%) than the pre-treatment (81.7%) isolates in the AL arm, although the difference was not statistically different (p\u2009=\u20090.195). The prevalence of the N86 allele was similar in the pre-treatment (80.8%) and post-treatment (81.5%) isolates in the ASAQ arm (p\u2009=\u20090.766)  samples were successfully sequenced. Mutations were observed in the M74I, N75E and K76T codons. The Pfcrt 76T allele, associated with amodiaquine resistance, was found in 57.7% of pre-treatment and 40.0% of post-treatment isolates,  and 2 (6.7%) of pre-treatment and post-treatment isolates, respectively. All isolates sequenced harboured either the mutant CVIET or wild-type CVMNK haplotypes.The Even though this study investigated efficacy results through 42\u00a0days, WHO recommends reporting efficacy for AL and ASAQ at day 28, with 90% being the threshold where a change in first-line treatment should be considered . The dayAlthough there were 14 and 3 ETFs in the AL and ASAQ arms, respectively, nearly all subjects cleared parasitaemia by day 7, implying that both treatments were able to clear parasites early in the infection. Compared with AL, the higher day 28 and 42 uncorrected efficacies of ASAQ may be a result of the longer half-life of amodiaquine compared with lumefantrine [Pfk13 mutations associated with artemisinin resistance were found in any of the parasites investigated, consistent with other studies carried out in Africa where malaria parasites are largely wildtype in the Pfk13 propeller domain and remain sensitive to artemisinin derivatives [Pfk13 polymorphisms were detected at six positions. The A578S polymorphism has been found throughout Africa and so far has not been associated with resistance to artemisinin derivatives but appears as a common polymorphism observed in African parasites [H) is one of the PfK13 mutations associated with artemisinin resistance [; however, we found a silent mutation at this codon, Y493Y (TAC\u2009\u2192\u2009TAT), in one sample. While it is possible this is a transitory mutation that could lead to the artemisinin resistant allele Y493H, no data currently exists to support this notion.No ivatives , 38. Howarasites , 38, 39.sistance ; howeverPfcrt and Pfmdr1 genes are associated with decreased sensitivity to lumefantrine and amodiaquine [Pfmdr1 N86 allele and NFD haplotype have been shown to be associated with reduced susceptibility to lumefantrine in some studies. In agreement with previous studies conducted in other parts of Africa [FD haplotype in the post-treatment isolates in the AL arm was observed compared to pre-treatment isolates. This was not observed in the ASAQ arm, suggesting that parasites harbouring the NFD haplotypes are likely to be more tolerant to AL treatment, consistent with previous observations. Overall, the prevalence of the N86 allele was more than 80% in the pre-treatment isolates in our study, similar to what was recently reported in two study sites in Mali, Dangassa and Nioro-du-Sahel [F allele, which can be contrasted with previous reports from Dangassa (39.9%) and Nioro-du-Sahel (48.2%). The significance of the high prevalence of N86 is unclear and may simply reflect the withdrawal of CQ drug pressure that selected for the 86Y polymorphism as previously indicated [Y allele in Mali, however, provides further support for the use of amodiaquine, a drug combined with sulfadoxine-pyrimethamine and used for seasonal malaria chemoprevention in children under five years in Mali.Equally important in the efficacy of an ACT is the partner drug. While delayed clearance (within 3\u00a0days post treatment) of parasites is associated with resistance to the artemisinin component, parasite recrudescence may occur when resistance to the partner drug exists, leading to an inadequate clinical and parasitological response. It is, therefore, imperative that resistance markers to partner drugs are evaluated. Point mutations in the diaquine . In partf Africa , 29, 40,f Africa , a highedu-Sahel . In addindicated  and/or aPfcrt 76T mutation has also been shown to play a role in amodiaquine resistance in several studies in Africa [Pfcrt 76T allele in the post-treatment was observed compared to pre-treatment isolates in this study, in contrast to previous reports. These unexpected findings can be explained by the fact that a majority (27) of the post-treatment isolates in our study were reinfection isolates and only four recrudescent isolates (among which three had the 76T mutation). The withdrawal of CQ for the treatment of malaria was followed by a steady decline of the prevalence of the Pfcrt 76T allele in many African countries [The n Africa . Prior sn Africa  or ASAQ n Africa , 20\u201322. ountries . Howeverountries . The basountries  who founAlthough investigation of molecular markers was performed on post-treatment samples (by combining samples with reinfections and recrudescent infection), association between observed parasite genotypes and recrudescent infection was not possible due to the small number of recrudescent infections (n\u2009=\u20099) which, when analysed alone, did not provide sufficient power. Nonetheless, monitoring of molecular markers of resistance to partner drugs during a therapeutic efficacy study is becoming increasingly feasible and provides a cost-effective tool for the early detection of decreases in parasite susceptibility to the drugs.This study demonstrated that day 28 PCR-corrected efficacy of both AL and ASAQ for uncomplicated malaria exceeded 90%, a WHO-recommended cut-off below which alternative treatments should be considered. Both uncorrected and PCR-corrected results suggested that ASAQ may be more efficacious, although the study was not designed to provide a comparison; however, this artemisinin-based combination is not employed routinely as a first-line malaria treatment because amodiaquine is paired with sulfadoxine-pyrimethamine and used throughout the country for seasonal malaria chemoprevention. Future studies in Mali should continue to monitor the efficacy of the current first-line ACT and consideration should also be given to evaluating artemisinin-based combinations, such as DP, with longer acting partner drugs.Additional file 1: Table S1. Kaplan Meier efficacy estimates and hazard ratios between AL and ASAQ, therapeutic efficacy monitoring, Mali, 2015\u20132016. Table S2.msp1, msp2, and glurp genotyping data )."}
+{"text": "Plasmodium falciparum resistance to artemisinin and its derivatives poses a threat to the global effort to control malaria. The emergence of anti-malarial resistance has become a great public health challenge and continues to be a leading threat to ongoing malaria control efforts. The aim of this review was to synthesize available evidence on the efficacy of dihydroartemisinin-piperaquine (DHA-PQ) compared to artemether-lumefantrine (AL) for the treatment of uncomplicated falciparum malaria among children in Africa.Emergence of A systematic literature search was done to identify relevant articles from online databases PubMed/ MEDLINE, Embase, and Cochrane Central Register of Controlled Trials\u2019 database (CENTRAL) for retrieving randomized control trials comparing efficacy of DHA-PQ and AL for treatment of uncomplicated falciparum malaria in African children. The search was performed from August 2020 to April 2021. Using Rev-Man software (V5.4.1), R-studio and Comprehensive Meta-analysis software version 3, the extracted data from eligible studies were pooled as risk ratio (RR) with 95% confidence interval (CI).2\u2009=\u20090%, high quality of evidence). Consistently, the PCR-adjusted treatment failure was significantly lower with DHA-PQ treatment group on day 28  and on day 42 . However, the efficacy was\u2009\u2265\u200995% in both treatment groups on day 28.In this review, 25 studies which involved a total of 13,198 participants were included. PCR-unadjusted treatment failure in children aged between 6\u00a0months and 15\u00a0years was significantly lower in the DHA-PQ treatment arm on day 28 than that of AL (RR 0.14, 95% CI 0.08\u20130.26; participants\u2009=\u20091302; studies\u2009=\u20094; IFrom this review, it can be concluded that DHA-PQ reduces new infection and recrudescence on days 28 and 42 more than AL. This may trigger DHA-PQ to become a first-line treatment option.The online version contains supplementary material available at 10.1186/s12936-021-03873-1. Plasmodium falciparum parasite was responsible for an estimated 24 million malaria cases in African children ) OR \u201cAcute malaria\u201d [Supplementary Concept]) OR \u201cArtemether, Lumefantrine Drug Combination/therapeutic use\u201d [Mesh]) OR \u201cLumefantrine\u201d [Mesh]) OR \u201cdihydroartemisinin\u201d [Supplementary Concept]) OR \u201cpiperaquine\u201d [Supplementary Concept]) OR ) AND ) AND  AND \u201cThe Cochrane Handbook for Systematic Reviews of Interventions  was follTitles and abstracts from the electronic search were independently screened by two authors based on randomized control trials (RCTs) that assessed human falciparum malaria. The information collected were trial characteristics, including methods, participants, interventions, outcomes, data on dose, and drug ratios of the combinations. Relevant information such as title, journal, year of publication, publication status, study design, study setting, malaria transmission intensity, follow-up period, sample size, funding of trial or sources of support, baseline characteristics of study subjects, treatment failure, fever clearance, and parasite clearance were extracted from each article using an extraction format in the form of a table adapted from the Cochrane online training materials repository and modified for this study. The numbers of participants randomized and analyzed in each treatment group for each outcome were also collected. One author independently extracted data and the information collected was cross-checked by another investigator. Missing data were requested from authors whenever necessary. The numbers of participants experiencing the event and participating in each treatment group were documented.The risk of bias for each trial was evaluated by two review authors independently using the Cochrane Collaboration\u2019s tool for assessing the \u2018risk of bias\u2019 . The risThe main outcomes in this review were total treatment failure on day 28, 42 and 63, PCR-adjusted and PCR-unadjusted. The numbers of patients with adequate clinical and parasitological response (ACPR) from the studies were combined and presented using risk rations accompanied by 95% confidence interval (CI).Participants were included according to the treatment group of the randomized clinical trials.2 statistic used to measure heterogeneity among the trials in each analysis, the Chi2 test with a P\u2009<\u20090.10 to indicate statistical significance, and the results were interpreted following Cochrane Handbook for Systematic Reviews of Interventions Version 6.0, Chapter 10: analysing data and undertaking meta-analyses [0\u201340%: might not be important;30\u201360%: may show moderate heterogeneity;50\u201390%: may show substantial heterogeneity;75\u2013100%: considerable heterogeneity.Heterogeneity among the included trials was assessed by inspecting the forest plots (to detect overlapping CI) and the Cochrane Q and Ianalyses .0\u201340%: mTo assess the distortion in the pooled effect estimate caused by one or more studies with extreme effect sizes, the pooled effect was checked again by removing outliers from the analysis . To asse2\u2009>\u200950%) was identified, it was reported and possible causes by sub-group analyses were explored.To further investigate the pattern of effect sizes and heterogeneity in the data, GOSH plot analysis was used. Using this plot, different sub-clusters with different effect size that were candidate for sub-group analysis were identified. When substantial heterogeneity  were used. When the Egger's test showed publication bias, the P-curve was used to estimate the presence of a \u201ctrue\u201d effect size behind the findings, and that the results were not the product of publication bias and P-hacking alone , 44. To The meta-analyses was done in consistency with the recommendations of Cochrane . To aid The potential sources of heterogeneity were investigated through the following sub-group analyses: participants under 5\u00a0years old compared with participants between 6\u00a0months to 15\u00a0years old.Meta-regression was used to investigate the association of study characteristics which cause heterogeneity with treatment effect. The covariates were age, HIV status, malaria transmission intensity in publication included, risk of bias, region, and drug administration (night dose). The results were presented in Figures and Tables.To investigate the strength of the methodology used in the primary analysis, a series of sensitivity analyses were conducted. To restore the integrity of the randomization process, the following steps were used: adding and excluding trials which were classified as high risk for bias back into the analysis in a stepwise fashion. To assess the influence of small-study effects on the results of the meta-analysis, fixed-effect and random-effect estimates of the intervention effect were compared. The robustness of the meta-analysis results explored the using influence analyses and the leave-one-out method.Quality of evidence was assessed using GRADE criteria and GRADE pro software . The resA total of 3211 studies from databases were searched, of which 49 full-text trials for eligibility were assessed; 25 fulfilled the inclusion criteria for meta-analysis and for qualitative analysis ; I2\u2009=\u200990%, moderate quality of evidence). The result could not be pooled. As there was high heterogeneity, it was more useful to consider the individual trial results. In 12/13 studies, the risk of treatment failure unadjusted by genotyping in children under five was significantly lower in the DHA-PQ treatment group than that of AL. Statistically significant difference was found between the two sub-groups [Chi2\u2009=\u20095.13, df\u2009=\u20091 (P\u2009= 0.02); I2\u2009=\u00a080.5% . The risk of treatment failure varied across the studies, meaning that some studies had high treatment failure and others had low treatment failure. Of the total variance in treatment failure, only 32% were explained by participant age and malaria transmission intensity within the countries. The funnel plot shows that all studies lay symmetrically around the pooled effect estimate , P\u2009=\u20090.39 ; I2\u2009=\u200974, moderate quality of evidence , P\u2009=\u20090.021). The P-curve evaluation shows that 13 studies were included into the analysis, of which 12 had a P-value lower than 0.025. The power of the analysis was 95% . The result shows that the evidential value was present, and there was a true effect size behind the findings, and that the results were not the product of publication bias and P-hacking alone Fig.\u00a0.Fig. 6P-2\u2009=\u20090%, high quality of evidence ; I2\u2009=\u200989%, moderate quality of evidence) and the result could not be pooled. It is more useful to consider individual trial results. On day 63, the PCR-unadjusted treatment failure in participants treated with DHA-PQ was significantly lower than those treated with AL in the two studies. However, the relative risk of PCR-unadjusted treatment failure was not significantly different between the two treatment groups ; I2\u2009=\u200975% . There were 19/7,115 early treatment failures in the DHA-PQ group arm versus 30/6,083 in the AL arm. The 28 and 42-days PCR-corrected treatment failure in patients receiving DHA-PQ were significantly lower than those patients receiving AL . On day 28, the PCR-corrected treatment failure was below 5% in both treatment arms and a similar result was seen in the DHA-PQ treatment arm on day 42. On the contrary, a study from Burkina Faso  reportedThe observed lower PCR-unadjusted treatment failure on days 28 and 42 in the DHA-PQ treatment arm was similar with that of former reviews from Africa , 7. As sHowever, no significant difference was seen on PCR-adjusted treatment failure between the two treatment groups on days 28 and 42 , 68. SimP. falciparum remains sensitive to artemisinin derivatives. Other studies conducted in Papua New Guinea [In this review, many children under the age of five who were treated with DHA-PQ had parasite clearance on day 1. Consistently, on day 2, participants from the DHA-PQ treatment group had faster parasite clearance than patients who were treated with AL. On day 3, the majority of the children in both treatment arms had parasite clearance. According to WHO , suspectw Guinea  and elsew Guinea \u201372 reporkelch-13 (k-13) gene mutations associated with artemisinin resistance in Southeast Asia [k-13 gene.Some molecular studies in Africa show absence of the known ast Asia , 73, 74,ast Asia , 75; AfrIn African settings, several risk factors were associated with persistent parasitaemia on days 1 and 2 after commencement of therapy . ConsideP. falciparum is effective in Africa at present. However, insufficient drug levels, ineffectiveness of anti-malarial drugs, and drug resistance could lead to treatment failure. Further studies should be conducted in different African countries with different malaria transmission intensity to identify the risk factors for treatment failure.The current malaria treatment guideline for This study has two limitations. Most of the studies were not blinded and the efficacy assessments had potential for bias. This review could not provide strong evidence for the long-term, post- treatment prophylactic effect of the two drugs up to day 63.From this review, it can be concluded that DHA-PQ reduces new infection and recrudescence at days 28 and 42, more than AL. This may trigger DHA-PQ to become the first-line treatment option. Continuous studies to measure the efficacy of DHA-PQ and AL within 42 and 63\u00a0days follow-up are needed.Additional file 1. Detailed search strategy.Additional file 2. Commands used for P-curve.Additional file 3. Characteristics of excluded studies.Additional file 4. Characteristics of included studies.Additional file 5. Meta-regression of PCR-unadjusted treatment failure at day 28, association between age of the children and treatment failure.Additional file 6. Meta-regression of PCR-unadjusted treatment failure at day 28, association between malaria transmission intensity within the countries and treatment failure.Additional file 7. Funnel plot of comparison: dihydroartemisinin-piperaquine versus artemether-lumefantrine for treatment of uncomplicated falciparum malaria in African children, outcome: PCR-unadjusted treatment failure at day 28.Additional file 8. Funnel plot of comparison: dihydroartemisinin-piperaquine versus artemether-lumefantrine for treatment of uncomplicated falciparum malaria in African children, outcome: PCR-adjusted treatment failure at day 28.Additional file 9. Funnel plot of comparison: dihydroartemisinin-piperaquine versus artemether-lumefantrine for treatment of uncomplicated falciparum malaria in African children, outcome: PCR-adjusted treatment failure at day 42.Additional file 10. Forest plot of comparison between dihydroartemisinin-piperaquine and artemether-lumefantrine for treatment of uncomplicated falciparum malaria in African children on fever clearance.Additional file 11. Forest plot of comparison: dihydroartemisinin-piperaquine versus artemether-lumefantrine for treatment of uncomplicated falciparum malaria in African children, outcome: Parasite clearance day 2 and 3.Additional file 12: Summary of finding tables."}
+{"text": "Plasmodium vivax are critical for the future control and elimination of malaria. We conducted a P. vivax volunteer infection study to characterize the antimalarial activity of artefenomel, a new drug candidate.Interventions that effectively target P. vivax and subsequently received a single oral 200-mg dose of artefenomel. Blood samples were collected to monitor the development and clearance of parasitemia, and plasma artefenomel concentration. Mosquito feeding assays were conducted before artefenomel dosing to investigate parasite transmissibility.Eight healthy, malaria-naive participants were intravenously inoculated with blood-stage 10 parasite reduction ratio over 48 hours, 1.67; parasite clearance half-life, 8.67 hours). Recrudescence occurred in 7 participants 11\u201314 days after dosing. A minimum inhibitory concentration of 0.62 ng/mL and minimum parasiticidal concentration that achieves 90% of maximum effect of 0.83 ng/mL were estimated, and a single 300-mg dose was predicted to clear 109 parasites per milliliter with 95% certainty. Gametocytemia developed in all participants and was cleared 4\u20138 days after dosing. At peak gametocytemia, 75% of participants were infectious to mosquitoes.Initial parasite clearance occurred in all participants after artefenomel administration (logP. vivax malaria.The in vivo antimalarial activity of artefenomel supports its further clinical development as a treatment for NCT02573857. Plasmodium falciparum is responsible for the majority of deaths, Plasmodium vivax is the predominant human malaria parasite species outside of Africa and has become recognized as a considerable cause of severe morbidity and a contribution to mortality ) and were mild or moderate in severity . No seri9/L). The elevations in alanine aminotransferase and aspartate aminotransferase levels were not accompanied by an elevation in bilirubin; thus, Hy\u2019s law was not reached. All severe AEs were considered related to malaria and resolved without treatment or with the use of standard antipyretic medication.Eleven AEs were classified as severe: 4 cases of elevated alanine aminotransferase , 1 case of elevated aspartate aminotransferase , 3 cases of pyrexia , and 3 cases of lymphopenia  and contributed to the PD analysis of parasite clearance. The log10 PRR48 was 1.67  and the corresponding parasite clearance half-life was 8.67 hours (8.11\u20139.31 hours). After the initial period of rapid parasite clearance occurring within 24 hours of artefenomel dosing, a second period of slower parasite clearance occurred over the course of 2\u20135 days, likely reflecting the clearance of gametocytes (see gametocytemia results below). Recrudescence was observed in 7 participants, 11\u201314 days after artefenomel administration, which was treated with artemether-lumefantrine  model best described the relationship between artefenomel plasma concentration and parasite killing rate (90 to be 0.83 ng/mL (.55\u20131.05 ng/mL). Modeling simulations of various single doses of artfenomel in malaria patients estimated 300 mg to be the minimum effective dose needed to clear 109 parasites per milliliter with 95% certainty. This dose was predicted to maintain artefenomel plasma concentrations above the MPC90 for a median of 11.8 days .Artefenomel concentration-time profiles were adequately described by a 3-compartment disposition model with first-order absorption, a lag time, and linear elimination . A direcing rate . The PK/The profile of gametocytemia development was similar to that of total parasitemia, steadily increasing until administration of artefenomel on day 10 . For gamAnopheles mosquitoes was evaluated with direct skin feeding and membrane feeding assays on day 8, and day 10 before artefenomel dosing. The mosquito feeding rates were high  and mortality rates were low , indicating good colony health  [The PK profile of artefenomel observed in this study was consistent with the results reported previously in both healthy participants and malaria patients . A singl1 hours) .P. vivax parasitemia to be determined. The estimated MIC of 0.62 ng/mL was considerably lower than the MIC for P. falciparum (4.1 ng/mL) calculated using the IBSM model [9P. vivax parasites per milliliter was 300 mg. Single doses between 200 and 1200 mg were found to be equally effective in clearing both P. falciparum and P. vivax malaria in Thai patients [P. falciparum; chloroquine plus primaquine for P. vivax) was given 36 hours after artefenomel dosing; thus, the potential for recrudescence was not evaluated.As expected, recrudescence occurred in most participants (7 of 8), allowing the PK/PD relationship between artefenomel plasma concentrations and SM model , perhapsP. vivax in malaria patients at doses of 800 and 1200 mg [P. vivax gametocytes in the circulation, it is not possible to be certain whether the slight delay in clearance of gametocytes compared with asexual parasites that was observed in the current study indicates that the dose tested is less effective, or that gametocytes disappeared from circulation at the normal rate. Gametocytocidal activity is a highly desirable property for new antimalarials to prevent parasite transmission to vector mosquitoes, important for progress toward malaria eradication.The current study also enabled in vivo assessment of the gametocytocidal activity of a 200-mg dose of artefenomel, because all participants developed gametocytemia together with the development of asexual parasitemia. Gametocytes were cleared in all participants within 8 days of dosing, supporting previous data indicating that artefenomel has activity against sexual stages of P. vivax to transmit parasites to Anopheles mosquitoes. The ability of artefenomel to prevent transmission was not assessed. Transmission was successful from the majority of participants (75%) in feeding assays performed on day 10 before artefenomel administration, but was not observed in feeding assays performed on day 8. This is likely due to the considerably higher level of gametocytemia on day 10 compared with day 8 . Indeed the participant in which the prevalence of transmission was highest had the highest gametocytemia. These results further validate the reproducibility of a P. vivax transmission model that was recently developed using IBSM [An exploratory objective of the current study was to determine the potential for healthy participants infected with blood-stage ing IBSM , and theP. vivax and exhibits antimalarial activity toward both asexual parasites and transmissible gametocytes. A single dose of 300 mg is predicted to clear parasitemia in patients with P. vivax malaria. Finally, we further validated the use of P. vivax IBSM as a model for evaluating transmission-blocking interventions.In conclusion, the current study has demonstrated that a single 200-mg oral dose of artefenomel is safe when administered to healthy participants experimentally infected with The Journal of Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at jiaa287_suppl_Supplementary_methods_and_resultsClick here for additional data file."}
+{"text": "Plasmodium falciparum parasitemia by day 42 than those receiving other regimens and experienced, on average, a shorter interval before the next clinical episode. Haplotypes of pfcrt and pfmdr1 were also evaluated in persisting parasites. These data identify a possible threat to the parasitological efficacy of artemether-lumefantrine in West Africa, over a decade since it was first introduced on a large scale.A recent randomized controlled trial, the WANECAM  trial, conducted at seven centers in West Africa, found that artemether-lumefantrine, artesunate-amodiaquine, pyronaridine-artesunate, and dihydroartemisinin-piperaquine all displayed good efficacy. However, artemether-lumefantrine was associated with a shorter interval between clinical episodes than the other regimens. In a further comparison of these therapies, we identified cases of persisting submicroscopic parasitemia by quantitative PCR (qPCR) at 72 h posttreatment among WANECAM participants from 5 sites in Mali and Burkina Faso, and we compared treatment outcomes for this group to those with complete parasite clearance by 72 h. Among 552 evaluable patients, 17.7% had qPCR-detectable parasitemia at 72 h during their first treatment episode. This proportion varied among sites, reflecting differences in malaria transmission intensity, but did not differ among pooled drug treatment groups. However, patients who received artemether-lumefantrine and were qPCR positive at 72 h were significantly more likely to have microscopically detectable recurrent  Plasmodium falciparum infections in Africa remains high, with >90% efficacy for all current regimens  failed treatment  than did pretreatment isolates for both artemether and lumefantrine in ex vivo susceptibility testing  against regimens . Despitereatment . Similardy sites . However testing , suggestdy sites 0 than di testing . Relevan  mutation, with 86N showing significant directional selection in a number of African studies of AL-treated malaria patients  amplification of parasite DNA from the WANECAM study. PCR-corrected drug efficacy findings for this study of 4,710 ACT-treated participants, carried out between October 2011 and December 2013, have been previously published . For the remaining 83 infections classed as slow clearing for episode 1 (15.0%), day 2 parasite density ranged from 0.11% to 92.1% of day 0 parasite density . Thirty-eight of 276 evaluable infections in episode 2 were slow clearing (13.8%), with day 2 parasite density ranging from 0.11% to 155.8% of day 0 density .Extracted DNA from all time points up to day 3 was analyzed by duplex qPCR, with each time point run twice. A total of 405 treated participants at episode 1 were qPCR negative by day 2 (73.4%), and a further 64 participants displayed a relative parasite density between 0 and 0.1% of that on day 0; all 469 met the criteria for good clearance  with AL than with PA or DP among patients recruited at Kolle , but this difference was not seen among those recruited at Sotuba or Bobo Dioulasso. Similarly, only in Kolle was there a substantially higher prevalence of qPCR-positive day 3 parasitemia in the AL group than in the PA and DP groups . In paired analyses on an individual level, slow clearance in episode 1 was not associated with a higher chance of slow clearance occurring in episode 2 , and being qPCR positive on day 3 in episode 1 was not associated with a higher chance of being qPCR positive on day 3 in episode 2 .Two binary parasite clearance parameters were derived from the analysis: slow clearance, defined as PRRP\u2009=\u20090.002; Wilcoxon rank sum test). However, after stratification by treatment arm, this relationship was maintained for ASAQ  and PA  but not for AL  or DP .The detection of persistent parasite carriage over the first 72\u2009h posttreatment could plausibly be associated with higher parasite densities at enrollment, which may take longer to clear. We therefore compared parasite densities by microscopy (as previously reported ) for indpfmdr1 86 Tyr and pfcrt 72 to 76 CVIET genotypes that had previously been favored by the use of chloroquine  or decreases (1 site) by day 3 were small , but in contrast, we did find evidence of directional selection against pfcrt 72 to 76 CVIET .Previous studies suggest that ACT, and artemisinin in particular, selects against the oroquine \u201315. The 48h below 1,000  was not a risk factor for subsequent treatment failure among these participants; in contrast, qPCR positivity at day 3 was significantly associated with treatment failure on or before day 42  sampling for microscopy as used for clinical studies of artemisinin efficacy in the Mekong region , 19, 21.efficacy , 35, parefficacy , 37 and There are a number of aspects of our study that may reduce the generalizability of our findings. Most importantly, the subjects of the work described here are a subset of 552 from a larger study of 4,710 individuals . More imin vivo parasite susceptibility to AL is waning in Africa. Our Kenyan studies  and day 1, day 2, and day 3 of posttreatment follow-up for the primary episode of falciparum malaria that led to the inclusion of the individual in the study. Blood samples taken at day 0, day 1, day 2, and day 3 from individuals with a second episode occurring 28\u2009days or more after the primary malaria episode were also included in the analysis if available.pgmet (a genus-specific target) normalized to the human \u03b2-tubulin gene humtubb as previously described (48h) and by the binary outcome positive or negative by qPCR on day 3. This method has now been successfully deployed in a number of field studies in Africa and Asia and has an estimated limit of detection of 0.05 parasite per \u03bcl of whole blood  and 95% confidence interval were estimated for each test of association between binary variables, and statistical significance was estimated from the \u03c7"}
+{"text": "In the Greater Mekong Subregion, adults are at highest risk for malaria. The most relevant disease vectors bite during daytime and outdoors which makes forest work a high-risk activity for malaria. The absence of effective vector control strategies and limited periods of exposure during forest visits suggest that chemoprophylaxis could be an appropriate strategy to protect forest goers against malaria.The protocol describes an open-label randomised controlled trial of artemether-lumefantrine (AL) versus multivitamin as prophylaxis against malaria among forest goers aged 16\u201365 years in rural northeast Cambodia. The primary objective is to compare the efficacy of the artemisinin combination therapy AL versus a multivitamin preparation as defined by the 28-day PCR parasite positivity rate and incidence of confirmed clinical malaria of any species. The sample size is 2200 patient-episodes of duration 1\u2009month in each arm. The duration of follow-up and prophylaxis for each participant is 1, 2 or 3 consecutive 28-day periods, followed by a further 28 days of post-exposure prophylaxis, depending on whether they continue to visit the forest. Analysis will be done both by intention to treat and per protocol.All participants will provide written, informed consent. Ethical approval was obtained from the Oxford Tropical Research Ethics Committee and the Cambodia National Ethics Committee for Health Research. Results will be disseminated by peer-reviewed open access publication together with open data.NCT04041973; Pre-result. Malaria is a major health problem for forest workers in Cambodia. Preventing malaria will provide major health as well as socioeconomic benefits for participants in the first instance and, if rolled out, for forest goers more broadly.The trial intervention was designed together with the Cambodian government to be potentially implementable depending on the results.Broad engagement with healthcare workers and communities in the study area preceded enrolment and this is continuing throughout with local healthcare workers and forest goers assisting with running the trial including identifying potential participants and supporting follow-up.The trial is open label so participants can know which study drug they are taking.The outcomes are dependent on the incidence of malaria during the trial follow-up period.In the Greater Mekong Subregion (GMS), adults are at highest risk for malaria. The most relevant disease vectors bite during daytime and outdoors which makes forest work a high-risk activity for malaria. The absence of effective vector control strategies and limited periods of exposure during forest visits suggest that chemoprophylaxis could be an appropriate strategy to protect forest goers against malaria.Anopheles dirus reduces the impact of the two most commonly employed control measures, long-lasting insecticide treated bednets (LLIN) and indoor residual spraying. Several studies have also demonstrated poor use of personal protection measures against malaria transmission.An. dirus), which tend to bite outdoors in daytime. LLIN have a high protective efficacy against nocturnal, indoor malaria transmissionAn. dirus. The improvised housing of forest workers is frequently poorly suited to hanging bed nets.13In the GMS, a large proportion of malaria transmission occurs in forested areas, which serve as perpetual sources of transmission.Plasmodium falciparum infections, but there are increasing treatment failures of clinical malaria casesP. falciparum, P. vivax and P. knowlesi.We propose to evaluate the feasibility and protective efficacy of antimalarial prophylaxis during forest work. It has been demonstrated in sub-Saharan Africa that seasonal malaria chemoprophylaxis (SMC) of children, the highest risk group for malaria in tropical Africa, can reduce malaria cases by 75%, is cost effective and safe and can be given by community health workers. Chemoprophylaxis of forest workers could protect this high-risk group and could reduce or even interrupt transmission in villages. The highly encouraging results of SMC in selected regions of sub-Saharan Africa provide hope that targeting another high-risk group, forest workers, could reduce malaria transmission in Cambodia and the wider GMS. In sub-Saharan Africa, children remain the main risk group for Plasmodium infections. In Southeast Asia, the main risk group are adults working and sleeping outdoors hence we propose to provide chemoprophylaxis for these adults. A major challenge for this strategy is the choice of an appropriate chemoprophylactic regimen in the GMS. The chemoprophylactic regimen of choice in Africa is sulfadoxine/pyrimethamine (S/P) plus amodiaquine despite high level resistance against the S/P component of the regimen. Similarly, we propose the use of AL, a drug whose efficacy remains high in the GMS, unlike, for example DHA/piperaquine. The study of AL versus a multivitamin preparation will be a two-arm randomised open-label comparative study. Laboratory assessments of malaria infection at baseline and day 28 post forest will be performed blind to treatment allocation and clinical cases during follow-up will be recorded.The main activity proposed is an in vivo clinical assessment of prophylaxis to prevent malaria in 4400 participant episodes in 50 villages in Stung Treng Province, Cambodia. The subjects will be randomised in a one-to-one ratio between the artemisinin combination therapy (ACT) AL and a multivitamin preparation with no antimalarial activity.The study site has been chosen based on current information on incidence of malaria, known predominance of malaria among forest goers, presence of an established clinical research programme and feasibility to perform the proposed research activities.Efficacy of AL ACT will be assessed through follow-up visits 28 days after returning from the forest on completing each course of prophylaxis when temperature, symptom questionnaires, brief physical examinations and malaria parasite PCR, and, in selected individuals, parasite genetics will be performed. Episodes of confirmed clinical malaria among study participants at any time point between enrolment and follow-up will also be recorded.All the organisations in this collaboration will work closely with local counterparts including the NMCP, non-governmental and other relevant organisations. Training is an integral part of this collaborative working relationship, and the building of local research capacity is an essential component of all research plans.All research-related activities, from study design, planning, implementation through to analysis and writing of reports will be performed jointly with local counterparts. Both on-the-job training and formal training will be provided when needed, in particular for good clinical practice (GCP) skills.The close interaction between WHO and its regional offices will ensure that new knowledge is disseminated efficiently and effectively throughout the region.To compare the efficacy of the ACT AL versus a multivitamin preparation as defined by the 28-day PCR parasite positivity rate and incidence of confirmed clinical malaria of any species.To compare the efficacy of the ACT AL versus a multivitamin preparation as defined by the 28-day, 56-day and 84-day PCR parasite positivity rate and incidence of confirmed clinical malaria for each species.To quantify the impact of the ACT AL as prophylaxis for forest goers on overall malaria transmission using mathematical modelling.Kelch13 mutations) and partner drug resistance.To assess the impact of AL prophylaxis on the spread of genetic markers of artemisinin  mapping on a subset of participants and their peers relating to the behaviours and risk factors associated with malaria infection in order to improve understanding of local malaria transmission among forest goers.To determine the prevalence of asymptomatic Plasmodium infections in high-risk populations at varying seasonal time points.To determine the prevalence of other infectious diseases that affect the study population.The study will take place at up to 50 villages in selected malaria endemic districts in Stung Treng Province, Cambodia. As the malaria situation in this area is dynamic, the villages will be identified prior to the start of the trial from analysis of up to date malaria incidence from passive surveillance collected by the Cambodia National Center for Parasitology, Entomology and Malaria Control (CNM). The rationale for choosing these areas include high forest cover and ongoing malaria transmission among forest goers. Malaria transmission in this area is generally low but varying over time.An open-label randomised parallel group superiority trial among forest goers comparing the ACT AL with a multivitamin with no antimalarial activity to evaluate the efficacy of prophylaxis, and to better understand high-risk groups and locations of malaria transmission. Follow-up will be for 1\u20133 consecutive periods of 28 days depending on whether the participant continues to visit the forest.The recruitment phase of the study is expected to last 12 months. Training and community sensitisation will precede study execution for 3 months. Data management and analysis, sample analysis , mathematical modelling and report writing are expected to take about 5 months. The total time to complete the study will be about 20 months.28-day PCR positivity rate* of Plasmodium infections of any species and/orProportion of participants with confirmed clinical malaria of any species reported between day 0 and day 28.28-day, 56-day and 84-day PCR Plasmodium positivity rate for each Plasmodium species.Proportion of participants with confirmed malaria reported between day 0 and day 28 for each species.Description of epidemiological situation of malaria in the study areas from passive surveillance data.**Kelch13 mutations and other genetic markers of antimalarial drug resistance of known functional significance.Prevalence of Incidence of adverse events (AEs) and serious adverse events (SAEs) by study arm during the course of prophylaxis.Data on the place of residence, work, recent travel history and mobile phone use.Detailed data and GPS mapping on a subset of participants and their peers relating to the behaviours and risk factors associated with malaria infection.Overall prevalence of Plasmodium at baseline, stratified by season and risk factors.Day 0, 28, 56 and 84 capillary blood levels of lumefantrine.Prevalence of serological diagnostic markers of other infectious diseases.*PCR positivity rate as determined from the proportion of blood samples that were PCR positive.**This will include the number of cases per village and demographics of those cases.Male and non-pregnant female participants aged between 16 years and 65 years planning to visit the forest within 72 hours are the target study population. The upper age limit was chosen as people over 65 years in the study area rarely travel to the forest and are at low risk of malaria. All pregnant women will be excluded as a conservative measure to minimise risk because of insufficient evidence for safety of AL in the first trimester together with frequent uncertainty about the stage of pregnancy, as well as lack of consensus about the required dose in pregnancy. All study participants must meet the applicable inclusion and exclusion criteria.Male or female, adults aged between 16 and 65 years.Planning to travel to the forest within the next 72 hours and stay overnight.Written informed consent.Willingness and ability of the participants to comply with the study protocol for the duration of the study.For females: known pregnancy or breast feeding.Participants who have received artemisinin or a derivative or an ACT within the previous 7\u2009days.History of allergy or known contraindication to artemisinins, lumefantrine or multivitamins.Documented or claimed history of cardiac conduction problems.Severe vomiting or diarrhoea on the day of screening.Signs/symptoms of clinical malaria (febrile or history of fever in the previous 24 hours) confirmed by rapid diagnostic test (RDT).Study procedures will be performed according to the schedule of assessments . This wi10.1136/bmjopen-2020-045900.supp1Supplementary dataPrior to the start of enrolment we will conduct community mobilisation and sensitisation activities in each village community where the trial will recruit participants. During the trial, the participant (or witness if illiterate) must personally sign and date the latest approved version of the informed consent form before any study specific procedures are performed. Written and verbal versions of the participant information and informed consent in the local language will be presented to the participants by trained study staff detailing no less than: the exact nature of the study; the implications and constraints of the protocol; and the known side effects and any risks involved in taking part. It will be clearly stated that participation is voluntary and that the participant is free to withdraw from the study at any time for any reason without prejudice to future care, and with no obligation to give the reason for withdrawal.The participant will be allowed as much time as possible to consider the information and take the opportunity to question the investigator, or other independent parties to decide whether they will  participate in the study. Written informed consent will then be obtained by means of participant dated signature or thumb print (if unable to write) and dated signature of the person who presented and obtained the informed consent. Examples of the patient information sheet and consent form for this study are provided in English in 10.1136/bmjopen-2020-045900.supp2Supplementary data10.1136/bmjopen-2020-045900.supp3Supplementary dataA copy of the signed informed consent document(s) will be given to the participants.Children aged 16 to\u00a0<18 years will be required to sign the latest approved version of the written informed assent form in addition to their parent or guardian signing a consent form.Participants who present at the participating sites will be screened to assess eligibility. Full consent will be obtained before any enrolment procedures are conducted. It will be made clear from the outset that refusal to participate will not jeopardise subsequent antimalarial treatment (if applicable). A screening log will be kept. As detailed below, participants may be enrolled a maximum of three times during the study period. People that cannot return for follow-up as per the schedule will not be enrolled. Known pregnancy will be identified by self reporting.Basic demographic and epidemiological data , and a full medical history will be recorded by the study staff.Brief physical examination and vital sign will be conducted by a qualified study team member. Weight and temperature will be documented. A symptom questionnaire will be performed.All prescribed or over-the-counter and traditional antimalarial medications used within the last 7\u2009days will be recorded. Any drug allergies will be recorded.Participants who are screened and are found to be febrile or have a current history of fever will not be enrolled (as per exclusion criteria) but will be tested for malaria and, if positive, given antimalarial treatment by the village malaria worker or local clinic. All this will be done in accordance with the current national malaria treatment guidelines in Cambodia. Individuals treated for malaria in this way will not be enrolled in the study as per the exclusion criteria. Such individuals may be enrolled later following recovery provided they meet the inclusion and exclusion criteria.Participants who fulfil all the inclusion criteria and have none of the exclusion criteria will be randomised 1:1 to one of the two treatment arms according to a randomisation schedule. Randomisation will be in permuted blocks of size that will be determined by the trial statistician and the block size will not be revealed to the investigating team. Randomisation will be stratified by village and villages combined for the analysis. Allocation will be done by trained study staff drawing the next sequential numbered opaque envelope , which contains the study number and treatment allocation.The participants will be assigned a study arm through a computer-generated randomisation schedule. Individual, sealed and sequentially numbered envelopes will be provided for each trial site with one envelope per participant, indicating the treatment allocation.This is an open-label study so the blinding of investigators and participants is not applicable. However, the randomisation procedure allows for adequate drug allocation concealment before envelopes are opened. All laboratory investigations will be performed without knowledge of the treatment allocation.On study enrolment, immediately before drug administration, blood will be collected for the following:Parasite PCR (up to 1\u2009mL).Storage for later identification of other causes of fever (2\u2009mL).In case of difficulties with venipuncture on enrolment  or loss of cold chain during transport from remote villages, three dried blood spots will be collected on enrolment for PCR and the other sample collected at follow-up.Participants will be treated with weight-based doses according to the schedule in The study drugs will be administered by trained study staff.If the participant vomits within half an hour after intake of the antimalarial drugs, the dose will be repeated. If vomiting occurs between half and 1\u2009hour, half of the dose will be repeated. If vomiting occurs more than 1\u2009hour after drug administration, no repeat dosing will be done. Repeat doses will be recorded on the case report forms (CRFs). If vomiting within 1\u2009hour occurs more than one time, no repeat dosing is allowed. The participant will then be treated at the discretion of the investigator.The prophylaxis will start with a 3-day course of two times per day AL. This will be followed by two doses 8\u2009hours apart on 1\u2009day per week during the time that the person is travelling in the forest and for 4 weeks after leaving the forest.Participants will be asked to return for a follow-up assessment any time from 28 to 35 days after commencing prophylaxis. Twenty-eight days was chosen as the upper limit of the time from infection to detectable parasites in the blood. Ongoing studies in the area found the duration of forest visits varied from a day to several weeks, with very few people being away for more than 28 days. This will be regardless of the duration of their visit to the forest or the number of times they visit it in that period. At this assessment, they will be interviewed about how long they spent in the forest, where they went, why, who they travelled with and about risk factors for infection. Brief physical examination, vital signs and symptom questionnaire will be performed. They will also be asked to report any diagnostic tests and/or treatment for malaria during the preceding 28\u201335\u2009days.At each follow-up visit, the following blood will taken:All individuals:Parasite PCR (up to 1\u2009mL).In those from whom sufficient blood could not be collected at baseline:Storage for later identification of other causes of fever (2\u2009mL).From minimum 100 randomly selected individuals:Lumefantrine level (0.2\u2009mL).In those with confirmed clinical malaria at any time point between enrolment and follow-up:Dry blood blots  collected on filter papers for:Parasite PCR and DNA genotyping for genetic markers of antimalarial resistance.Parasite whole genome sequencing and barcoding to identify geographical origin of parasites and compare genotypes to identify persistent infections.In individuals who are planning to return again to the forest within the following 28 days after the follow-up visit, they will be asked to continue their weekly prophylaxis according to the original treatment allocation on enrolment. They will then be asked to return for a second follow-up visit a further 28\u201335 days later when the above procedure will be repeated. This will be repeated one more time. If the person cannot be followed up within the scheduled period, for example, because they do not return from the forest in time, then they will be followed up at the first opportunity and this will be recorded in the CRF.Thus individuals may take prophylaxis continuously for a maximum of three periods of 28\u201335\u2009days in the forest plus 4 weeks after returning totaling 112 days. The choice of study medication for each individual will follow the initial assignment on enrolment throughout the follow-up period.In those who do not declare an intention to return to the forest within 28 days at any follow-up visit, no further follow-up visits will be offered at that time but they will be asked to complete 4 weeks of prophylaxis following their last day in the forest as post-exposure prophylaxis.Individuals who have been enrolled in the study may be enrolled into the study up to two more times during the 12 months study period only if a minimum period of 28 days (4 weeks) has elapsed following their last dose of prophylaxis. Thus they can be enrolled in the study up to three times. If an individual is enrolled again in this way, they will be re-randomised following the same procedure as enrolment. The rationale for this re-enrolment was that malaria transmission and forest travel are seasonal at this location and this allows detection of malaria positive episodes in people who continue to visit the forest throughout the year while minimising the period of follow-up for the majority of people who visit the forest only during a particular season; in addition, it allows a wash-out period between episodes of taking prophylaxis.The time-window for the follow-up visits is 28\u201335\u2009days. If a participant does not attend, the study team will try to locate the participant and conduct the necessary examinations and tests.Participants presenting to the village malaria worker, mobile malaria worker or clinic with a fever or other symptoms at any time after enrolment that is not a scheduled study follow-up visit will be assessed and treated by the healthcare workers in the local healthcare system as per routine clinical practice in Cambodia.On enrolment, participants will be encouraged to attend a village malaria worker or government clinic for the assessment of fever or other symptoms and to report this to the study team as soon as possible. Information on these healthcare encounters including malaria test result and treatment will be recorded in the study CRF.Participants who have an episode of confirmed clinical malaria at any time after enrolment up to the last follow-up visit and for 1\u2009month afterwards will have blood taken for parasite genetic analysis. As clinical malaria at follow-up is part of the composite primary endpoint, and the participants and field staff will not be blinded as to study arm, there is potential for bias if, for example, people in the AL arm choose not to attend for a malaria test. However, extensive efforts will made through community engagement and individual counselling to advise participants against this.The blood volumes for the protocol mandated tests are as follows:PCR: up to 1\u2009mL.Lumefantrine level: 0.2\u2009mL.Dried blood spots for parasite genetics: 0.4\u2009mL.Storage for serology at baseline 2\u2009mL.Bulletin of the World Health Organization 2011:89:46\u201353).Maximum blood volumes are presented below for adults for the maximum of three periods (84 days) of follow-up. The maximum blood volume is the total amount taken if the participants returns for follow-up on three consecutive occasions and had all blood samples taken. The maximum blood volume will be approximately 10.2\u2009mL . Access to research samples will be limited using either a locked room or a locked freezer.This is required for the primary study objective. Blood samples will be analysed in the Molecular Tropical Medicine Laboratory, Bangkok, Thailand using PCR to identify which individuals have malaria parasites of any species. It is anticipated that results will be available around 3\u20136 months after collecting each sample, thus they will not be used to guide antimalarial treatment at the time of testing. The study teams will be informed which samples were positive for malaria and they will follow-up positive participants to conduct a brief clinical assessment. Any individuals who are symptomatic will be referred to the village malaria worker or clinic for testing and treatment. The laboratory will be blinded to the study arm of the patient.Blood samples (dried blood spots) for parasite genetic analysis will be obtained and stored from all subjects recruited with subject\u2019s consent. In individuals in whom parasites are found by PCR, samples will be processed for parasite genetic analysis. Genetic samples (in the form of dried blood spots or extracted DNA) will be stored (for a maximum of 10 years) at the Molecular Tropical Medicine Laboratory, Bangkok, Thailand. In those with confirmed clinical malaria, parasite genotyping will be performed at the Wellcome Trust Sanger Institute in Hinxton, UK or other suitable laboratory using a set of informative single nucleotide polymorphisms (SNPs) selected from whole genome sequencing. The subject will be asked for consent for this transfer during the initial informed consent process. A material transfer agreement will be in place if required before any samples are shipped. The results of the parasite genotyping will not be reported back to the subjects. This analysis will only be done for those with confirmed clinical malaria as it is anticipated that there will be insufficient genetic material in samples taken from those with asymptomatic infection due to the low parasite burden in these individuals.Blood samples for lumefantrine level will be taken at follow-up visits from a minimum of 100 randomly selected participants, where logistically possible, to assess adherence with the study drugs. These will be analysed in the Pharmacology Laboratory at Mahidol-Oxford Tropical Medicine Research Unit (MORU) in Bangkok, Thailand.Among those who specify by written consent, the serology samples will be analysed for diagnostic markers of other infectious diseases.AL is currently available as standard tablets containing 20\u2009mg artemether and 120\u2009mg of lumefantrine in a fixed-dose combination formulation. It is included in this formulation on the WHO Model List of Essential Medicines. 22The AL is administered as a two times per day dose for 3\u2009days for a total of six doses  followed by two times per day once a week according to the treatment schedule in The multivitamin preparation will be HEXA CMP  or suitable equivalent alternative administered as a once daily dose using the treatment schedule in All efforts will be made to store the study drugs in accordance with the manufacturers' recommendations in a secure area. This may be difficult at some sites where air-conditioned storage rooms are not available. The ACT should be stored between 15\u00b0C and 30\u00b0C (59\u00b0F\u201386\u00b0F).Where this is not possible and monitored storage conditions do not meet the recommendations, the artemisinin-derivatives and partner drug content of batches of ACT will be retested at the end of the study.Study drugs will be administered as directly-observed-therapy (DOT) on the first day. Where possible, study drugs will also be administered as DOT on days 2 and 3. Where DOT is not possible, the participant will be contacted by the study team by telephone or in person to ensure they take the second and third days of medication and to ensure they follow the correct procedure in case of vomiting. If the participant vomits, and is re-dosed; this will be recorded in the CRF. If vomiting within 1\u2009hour occurs again after retreatment, no repeat dosing is allowed. All drug doses will be recorded in the CRF. To maximise adherence to the study medication, the study will be preceded by a period of community sensitisation and engagement including information sessions on the importance of taking all doses of medication. The participants will be requested to take each dose with food to maximise absorption of the lumefantrine.All movements of study medication will be recorded. Both study medication of individual participant and overall drug accountability records will be kept up to date by the study staff.Throughout the study, investigators may prescribe concomitant medications or treatments deemed necessary  to provide adequate supportive care except for antibiotics with antimalarial activity unless unavoidable . If these are required, the participants will be kept in the study and this will be noted as a protocol deviation. Anti-emetics should not be prescribed as a prophylaxis if no nausea or vomiting is present.Antimalarials for symptomatic, confirmed malaria infections will be prescribed as described above. Any medication, other than the study medication taken during the study will be recorded in the CRF.In order to have a greater understanding of the possible sites of malaria transmission, and to relate genetic diversity to geographical location, participants will be asked a short set of questions on their place of residence, place of work and their history of travel plus possible risk factors for malaria. This is to obtain a detailed understanding of the behaviours and risk factors for malaria infection. We will collect GPS coordinates of the places of residence of all participants. In a subset of participants, GPS coordinates will be collected for their travel patterns during follow-up including place of work, forests, forest camps, farms or plantations to identify places where their infection may have occurred. The size of this subset will be determined by the availability of GPS devices with the number being limited to 50 participants at any one time. The GPS devices will be offered to unselected consecutive trial participants whenever they are available, being returned on completion of follow-up for that individual. We will collect all available local malaria treatment records to describe how the study population compares to the overall population who receive treatment for malaria and this will allow us to better understand local malaria epidemiology and transmission patterns. All personal information will be anonymised so that no individual can be identified from their treatment records, through interviews or from mapping data.Passive surveillance data from all available sources for the study province collected by CNM will be analysed to identify any changes in malaria incidence rate in study villages before, during and after the study where AL prophylaxis was administered compared with non-study villages.Enrolled participants who experience an episode of confirmed clinical malaria during follow-up will be linked back to their individual case records to quantify the incidence of clinical malaria in each study arm.PCR will be used to identify which individuals have parasites at enrolment (prior to taking the study medicine) and at each follow-up visit and is required for the primary study objective.Parasite DNA will be used for genomic studies including but not limited to parasite species confirmation, microsatellite typing to identify parasite clones and SNPs typing/whole genome sequencing to generate data for studies of the geographic origins of the parasites.Lumefantrine levels will be used to assess adherence in a random sample of study participants.The serology sample will be used for anonymised investigation of the prevalence, incidence, association with fever, and risk factors for other common infectious diseases affecting the study population. Samples will be stored for later analysis.All efforts will be made to retain as much data as possible. The main strategy that is being re-enforced for data retention includes study staff reminding participants of the upcoming data collection. This will be emphasised during management training. However, each participant has the right to discontinue the study drug or the study at any time. Data accrued up until the time of discontinuation will be used in the analysis.In general, the investigators will be required to make every effort to perform the study procedures until completion of follow-up (maximum three visits over 84 days), including in the following situations:Significant non-compliance with treatment regimen or study requirements.An AE which requires discontinuation of the study medication or results in inability to continue to comply with study procedures.Disease which requires discontinuation of the study medication or results in inability to continue to comply with study procedures.Loss to follow-up (every attempt should be made to re-contact the participant).However, the investigator may discontinue participation in the study of a participant if he or she considers it necessary.In addition, the participants always have the right to withdraw consent in writing or verbally.The reason for withdrawal or discontinuation, if available, will be recorded in the CRF. If the study drug or participation in the study is discontinued due to an AE, the investigator will arrange for follow-up visits at least until the AE has resolved or stabilised.If a participant does become pregnant during participation in the study, they will be withdrawn from the study immediately on it being reported to the study team. Any pregnancy must be reported to the principal investigator (PI) within one working day of awareness. The PI must take all reasonable efforts to discover the outcome of the pregnancy and fill out the pregnancy form. If there is a congenital abnormality or a stillborn baby, this needs to be reported as an SAE.Source documents are original documents, data, and records from which participants\u2019 CRF data are obtained. These include, but are not limited to, village malaria and clinic records , clinical and office charts, laboratory and pharmacy records, diaries, microfiches, radiographs and CRFs.CRF entries will be considered source data if the CRF is the site of the original recording (e.g. there is no other written or electronic record of data). In this study, the CRF will be used as the source document for most of the data points.All documents will be stored safely in confidential conditions.This trial will use drugs that have either been registered or evaluated extensively. To add to the evidence base for safety of AL as prophylaxis, we will record and review all AEs and SAEs that are reported to occur in the study.A symptom questionnaire will be performed on enrolment and at each subsequent follow-up visit to the healthcare centre, to aid in the identification of AEs. In addition, enrolled individuals will be encouraged to promptly report any unexpected symptoms or illnesses between follow-up visits to the study team.The investigator is responsible for the detection and documentation of events meeting the criteria and definition of an AE or SAE, as provided in this protocol.All SAEs and AEs will be promptly documented from the moment of drug administration in the study to discontinuation of the participant from study participation. Any events occurring between screening and drug administration will be considered as baseline, preexisting conditions.All AEs must be recorded in the AE/SAE CRF. To avoid colloquial expressions, the AE should be reported in standard medical terminology. Whenever possible, the AE should be evaluated and reported as a diagnosis rather than as individual signs or symptoms. If a definitive diagnosis is not possible, the individual symptoms and signs should be recorded. Whenever possible, the aetiology of the abnormal findings will be documented on the CRF. Any additional relevant laboratory results obtained by the investigator during the course of this study will be recorded on the CRF.If the event meets the criteria for \u2018serious\u2019, the SAE must be reported to the PAL-Cambodia safety team within 24 hours of the time that the event was identified. If further data are required, additional documentation can be submitted. All SAEs must be followed until resolution, or until the SAE is deemed permanent or leads to death.Samples will be shipped for PCR to a molecular laboratory where they will be analysed in batches. Quality control results will be available approximately 3\u20136 months from the time of collection. The list of positive tests will be returned to the field sites. If a participant is found to have a Plasmodium infection, and has not already received antimalarial treatment subsequent to the sample being collected, then these individuals will be contacted by a local health worker, and if a participant reports fever or illness they will be offered appropriate diagnosis and treatment.An AE is any undesirable event or clinical deterioration that occurs to a study participant during the course of the study; that is, from the time of administration of study drugs until study ends (i.e. until the follow-up visit) whether or not that event is considered related to the study drugs, or to a concomitant drug or procedure: for example,Any unfavourable and unintended symptom.Physical sign.Abnormal laboratory result.An illness.Any new clinical sign or clinical deterioration that occurs between signing the consent form and the administration of study drugs is not an AE. This information will be recorded in the medical records, as a pre-existing condition.An SAE is an AE that:Results in death.Is life-threatening that is, the participant was at risk of death at the time of the AE.Requires in participant hospitalisation or prolongation of existing hospitalisation.Results in persistent or significant disability/incapacity.Is a congenital anomaly/birth defect.Any other significant medical condition.All of the above criteria apply to the case as a whole and should not be confused with the outcomes of individual reactions/events. More than one of the above criteria can be applicable to the one event. Important medical events that may not be immediately life-threatening or result in death or hospitalisation may be considered an SAE when, based on appropriate medical judgement, they may jeopardise the participant or require medical or surgical intervention to prevent one of the outcomes listed in the definition above. Examples of such medical events include allergic bronchospasm requiring intensive treatment in an emergency room or at home, blood dyscrasias or convulsions that do not result in hospitalisation, or development of drug dependency or drug abuse.All SAEs must be reported by the site investigator to the Study PI and PAL-Cambodia safety and medical monitor, within 1\u2009day of his or her awareness of the SAE. The SAE report, should be emailed to the email paltrial@tropmedres.ac.Further reports should be submitted, if required, until the SAE is resolved.The site investigator must also report the SAEs to the local ethics committee in accordance with local requirements.Each AE will be graded according to the Common Terminology Criteria for Adverse Events (CTCAE) V.5.0 November 2017.If an AE is not listed in the CTCAE table, the investigator will assess the severity using the following guidelines:1=Mild; asymptomatic or mild symptoms; clinical or diagnostic observations only; intervention not indicated.2=Moderate; minimal, local or noninvasive intervention indicated; limiting age appropriate instrumental ADL.*3=Severe\u2009or medically significant but not immediately life-threatening; hospitalisation or prolongation of hospitalisation indicated; disabling; limiting self care ADL**4=Life-threatening consequences; urgent intervention indicated.5=Death related to AE.Activities of Daily Living (ADL).*Instrumental ADL refer to preparing meals, shopping for groceries or clothes, using the telephone, managing money and so on.**Self care ADL refer to bathing, dressing and undressing, feeding self, using the toilet, taking medications and not bedridden.The term \u2018severe\u2019 is often used to describe the intensity (severity) of a specific event ; the event itself, however, may be of relatively minor medical significance (such as severe headache). This is not the same as \u2018serious\u2019, which is based on the outcome or criteria defined under the SAE definition. An event can be considered serious without being severe if it conforms to the seriousness criteria, similarly severe events that do not conform to the criteria are not necessarily serious. Seriousness (not severity) serves as a guide for defining regulatory reporting obligations.The investigator is obligated to assess the relationship between study drug and the occurrence of each AE/SAE using the following categories of relatedness:Definite: clear-cut temporal association.Probable: clear-cut temporal association, with improvement on drug withdrawal, and not reasonably explained by the participant\u2019s known clinical state or other aetiology.Possible: less clear temporal association; other aetiologies are possible (other possible aetiologies should be recorded on the CRF).Not related: no temporal association with the study drug; assessed as related to other aetiologies such as concomitant medications or conditions, or participant\u2019s known clinical state.The investigator will provide the assessment of causality as per the AE/SAE data collection tool.The investigator will follow-up the AE and SAE until resolution or until no further medically relevant information can be expected. AE and SAE outcome will be classified as follows:Continuing/ongoing.Resolved.Resolved with sequelae.Permanent.Fatal.P. falciparum incidence in forest workers.The target population for this study will be adult Cambodians who work and sleep in the forest . Two thousand two hundred study participant episodes are required in each arm to have sufficient power to detect a statistically significant difference between the treatment arm and a control arm. An episode is defined as a follow-up period of 28 days with each enrolled individual contributing 1, 2 or 3 episodes. The estimate of the required sample size is complicated by the scarce data on P. falciparum positive without receiving prophylaxis will be around 5%. A total of 1605 participant-episodes per arm are enough to detect a difference of at least 40% in the proportion of episodes with a P. falciparum positive result as defined by the 28-day PCR parasite positivity rate, that is from 5% positivity in participants without receiving antimalarial prophylaxis  to 3% positivity in participants receiving AL prophylaxis. This has been estimated with 80% power and 5% significance level. However, we also anticipate that we will likely observe multiple episodes being recruited into the study that can reduce power of the study if not accounted for. To compensate for the multiple episodes and any losses to follow-up, we plan to recruit approximately 600  additional episodes in each group on top of the required 1605 single episodes. This gives an additional 27% episodes to account for the multiple episodes and losses to follow-up. Thus, the overall sample size will be 4400 episodes . The sample size calculations have been performed in Stata V.15.Formally, we anticipate that the risk of being The main analysis strategy for the primary outcome will be the intention to treat (ITT) principle followed by the per protocol (PP) analysis. Thus, we will first analyse the ITT population in which all participants recruited in the trial will be included in the analysis according to the randomisation arm irrespective of what they actually got. These ITT analyses will be followed by the analysis of the PP population in which participants who did not adhere to the protocol will be excluded from analysis.The composite primary endpoint will be analysed as follows. For 28-day PCR Plasmodium positivity and parasite positive clinical episode rate analysis, each arm will be summarised using crude proportions and binomial exact 95% CIs. The risk differences in Plasmodium positivity between AL versus multivitamin will be reported along with the corresponding 95% CIs. Robust standard errors will be used to handle multiple episodes. These analyses will be complemented by the use of the crude Kaplan-Meier estimates of cumulative PCR Plasmodium positivity and parasite positive clinical episode probabilities as recommended by WHO. The incidence of confirmed clinical malaria between day 0 and day 28 analysis will be modelled using the mixed effects Poisson regression model to obtain incidence rate ratios comparing AL versus multivitamin arms. The mixed effects models will take into account the correlation of multiple episodes from the same participant. Tests of significance will be performed at 5% significance level. Analysis of all endpoints will be described in detail in a Statistical Analysis Plan finalised prior to locking the database. A brief overview is given below.These will be compared using chi squared or Fisher\u2019s exact test, as appropriate. Crude proportions will be calculated with the exact 95% CIs, where relevant.These will be summarised by medians  and means , as appropriate, and will include the parasite counts and laboratory parameters. Comparisons of continuous data will be assessed using the paired/unpaired t-tests or the signed rank/Mann-Whitney U tests, as appropriate.Safety analyses will be based on the whole population that get administered the study drug. Safety and tolerability of ACT versus multivitamin will be assessed by comparing the frequency (%) of AEs and SAEs, with particular attention to abdominal pain and appetite perturbation, using the Fisher\u2019s exact test. Safety data will be presented in tabular and/or graphical format and summarised descriptively. Any clinically relevant abnormalities or values of potential clinically concern will be described. Participants will be analysed according to an ITT and a PP method where appropriate.For analyses of proportions, missing outcomes will be imputed using plausible values. For example, worst-case scenario may be deemed appropriate and in that case sensitivity analysis will be performed with the best-case scenario. In the ITT Kaplan-Meier/survival analysis, participants who are lost to follow-up, or who have Plasmodium reinfections or inconclusive PCR correction, will be censored from the moment of occurrence of one of these events. This survival analysis approach is the best way of handling missing data because participants with partial information are included in the analysis up to the time when they are lost/withdraw from the study.AEs will be graded according to CTCAE V.5.0, November 2017.All AEs that are newly started or increased in intensity after the study drug administration will be reported. AE reports will be generated for all AEs that occurred after study drug administration, until the end of the study.The impact of the ACT AL as prophylaxis for forest goers on overall malaria transmission will be quantified using mathematical modelling. For this we will develop a population dynamic village level individual-based model of malaria transmission and treatment parameterised with published data, results from the analysis of data from the trial and fitted to surveillance data from the study area.Direct access will be granted to authorised representatives from the sponsor and host institution, the regulatory authorities, and ethical committee (if applicable), to permit trial-related monitoring and inspections.The study will be conducted in accordance with the current approved protocol, International Conference on Harmonization (ICH) GCP, any national regulations that may apply to this study and standard operating procedures. The WorldWide Antimalarial Resistance Network (WWARN) will be engaged in assuring quality assurance (QA)/quality control (QC) of study execution in collaboration with the MORU Clinical Trials Support Group (CTSG). Their role will include but not be limited to monitoring adherence to standard operating procedures (SOPs) for collection of laboratory specimens and quality checks (curation) of laboratory data according to standard methodologies.Study sites may have in place a system for internal monitoring. In addition, regular external monitoring of all sites will be performed by the MORU CTSG according to ICH GCP and a Monitoring Plan. Data will be evaluated for compliance with the protocol and accuracy in relation to source documents. The monitors will check whether the clinical trial is conducted and data are generated, documented and reported in compliance with the protocol, GCP and the applicable regulatory requirements. Evaluation of on-site monitoring schemes, such as a reciprocal monitoring scheme, may be undertaken at selected sites by CTSG.During 2018, extensive consultations were held with local authorities, patients, and study communities regarding the design and organisation of the trial. This included the district health authorities and Governor\u2019s office. The Siem Pang field station conducted malaria treatment studies and as part of these studies interviews and questionnaires were done with patients to better understand the local risk factors for malaria, travel histories and the nature of forest work. 26The Investigator will ensure that this study is conducted in compliance with the current revision of the Declaration of Helsinki.The investigators will ensure that this study is conducted according to any National Regulations and that it will follow the principles of the ICH Guidelines for GCP.The study protocol and its associated documents will be submitted to the Oxford Tropical Research Ethics Committee and the appropriate local ethics committees for written approval.The investigator will submit and, where necessary, obtain approval from the above parties for all substantial amendments to the original approved documents.This study will use drugs that have been studied thoroughly and their toxicities are well described. In general, they are all well tolerated. In the event of any serious or severe AE participants will be referred to the local referral hospital where best available care will be provided.The safety of artemether and lumefantrine for treatment of malaria has been evaluated in clinical trials and, post licensing, widespread use for treating malaria in hundreds of millions of patients per year. Reported AL side effects have generally been mild. Reported adverse reactions in clinical trials have been similar or lower in frequency and magnitude to other ACTs. The most common (>=3%) reported AEs in clinical studies with AL in adults were headache, anorexia, dizziness and asthenia. AL is not known to cause harmful prolongation of the corrected QT interval (QTc). 27The main side-effects of multivitamin are upset stomach, unpleasant taste or headache which are mild to moderate in nature. Very rarely, these may cause an allergic reaction.The primary risks of phlebotomy include local discomfort, occasional bleeding or bruising of the skin at the site of needle puncture, and rarely haematoma or infection. Phlebotomy will be performed by suitably qualified and trained staff using appropriate hygiene measures including gloves and alcohol swabs to clean the skin.Due to the potential unique nature of the GPS tracking data, it may be possible to identify individuals from their tracks. This will be minimised by the GPS tracking data being kept separately from any personally identifiable information and linked to the data collected on the study CRF only through a unique study code. The GPS tracking data will also be stored anonymously on the tracking device during collection and moved to an encrypted hard drive on completion of collection.There are no anticipated direct benefits to the participants in this study. However, knowledge gained from this study is expected to help to assess the efficacy and feasibility of prophylaxis to prevent malaria in forest workers, and to predict its efficacy in reducing overall transmission. The proposed study is a critical step for future use of chemoprophylaxis to protect forest workers in the GMS against malaria.Participants are able to decline freely participation in this study. If so, they will receive standard care for their malaria (if applicable).Study participants will be compensated for time lost from work as a result of trial activities, the cost of local transport to attend for the follow-up visits and will receive a per diem to cover the costs of meals on those days. The amounts in monetary terms will be determined by CNM in accordance with local norms.The study will pay for treatment for drug-related SAEs or other research-related injuries. The study cannot pay for long-term care for disability resulting from complications of the illness.The trial staff will ensure that the participants\u2019 anonymity is maintained. The participants will be identified only by initials and a study number on the CRF and electronic databases. All documents will be stored securely and be accessible to trial staff and authorised personnel only.Samples collected will be used for the purpose of this study as stated in the protocol and stored for future use no longer than 10 years. Consent will be obtained from participants for sample storage and/or shipment of specific samples to collaborating institutions for investigations that cannot be performed locally. Any proposed plans to use samples other than for those investigations detailed in this protocol will be submitted to the relevant ethics committees prior to any testing. Material transfer agreements will be arranged and signed where appropriate/needed.Study data will be recorded on the CRF at the study sites and stored in a secure database. Validation checks will be built into the study database to identify missing values, inconsistencies or invalid data. Additionally, study data will be profiled using statistical software to check for outliers and errors not detected by the database. All tasks related to data management will be carried out in accordance with the study data management plan.De-identified, individual participant data from this study will be available to researchers whose proposed purpose of use is approved by the data access committee at MORU. Enquiries or requests for the data may be sent to datasharing@tropmedres.ac.The University of Oxford has a specialist insurance policy in place: Newline Underwriting Management, at Lloyd\u2019s of London\u2014which would operate in the event of any participant suffering harm as a result of their involvement in the research.Results will be published in the open access peer-reviewed medical literature. Any data published will protect the identity of the participants. This trial will be registered in a web based protocol registration scheme. All those who have made a substantial contribution will be coauthors on publications. The sites have the right to publish their data individually and to include members of the sponsor\u2019s team who have made a significant contribution. There will also be publications of pooled data which will be coordinated by the MORU group. All sites will have the opportunity to contribute to these publications.All the research findings from the programme and from relevant research outside the Programme will be analysed and integrated, and through the WHO Global Malaria Programme will be disseminated to policy makers, NMCPs and other researchers.A completed Standard Protocol Items: Recommendations for Interventional Trials checklist for the protocol is provided in 10.1136/bmjopen-2020-045900.supp4Supplementary data"}
+{"text": "Plasmodium falciparum infection and the patterns and predictors of treatment failure in Vietnam.Drug-resistant falciparum malaria is an increasing public health burden. This study examined the magnitude of Medical records of all 443 patients with malaria infection admitted to the Hospital for Tropical Diseases between January 2015 and December 2018 were used to extract information on demographics, risk factors, symptoms, laboratory tests, treatment, and outcome.Plasmodium falciparum infection of whom 21.9%  had severe malaria, while 7.2%  and 19.2%  developed early treatment failure (ETF) and late treatment failure (LTF) respectively. Among 58 patients with severe malaria, 14 (24.1%) acquired infection in regions where artemisinin resistance has been documented including Binh Phuoc (11 patients), Dak Nong (2 patients) and Gia Lai (1 patient). Under treatment with intravenous artesunate, the median (IQR) parasite half-life of 11 patients coming from Binh Phuoc was 3\u00a0h (2.3 to 8.3\u00a0h), two patients coming from Dak Nong was 2.8 and 5.7\u00a0h, and a patient coming from Gia Lai was 6.5\u00a0h. Most patients  recovered completely. Four patients with severe malaria died. Severe malaria was statistically associated with receiving treatment at previous hospitals (P\u2009<\u20090.001), hepatomegaly (P\u2009<\u20090.001) and number of inpatient days (P\u2009<\u20090.001). Having severe malaria was a predictor of ETF . No predictor of LTF was identified.More than half  of patients acquired Plasmodium falciparum remains the prevalent malaria parasite. Despite low mortality rate, severe malaria is not rare and is a significant predictor of ETF. To reduce the risk for ETF, studies are needed to examine the effectiveness of combination therapy including parenteral artesunate and a parenteral partner drug for severe malaria. The study alerts the possibility of drug-resistant malaria in Africa and other areas in Vietnam, which are known as non-endemic areas of anti-malarial drug resistance. A more comprehensive study using molecular technique in these regions is required to completely understand the magnitude of drug-resistant malaria and to design appropriate control strategies. Plasmodium species causing malaria in humans, Plasmodium falciparum is responsible for the most severe forms of malaria [P. falciparum is the most prevalent malaria parasite followed by Plasmodium vivax [P. falciparum infection since 2007 [P. falciparum infection in Dak Nong and Binh Phuoc which are the two provinces mostly affected by DHA-PPQ resistance [Malaria is a mosquito-transmitted infection that affects 219\u00a0million people and causes 435 thousand deaths worldwide [um vivax . In lineum vivax , artemisnce 2007 . Howevernce 2007 . Moreovence 2007 . Recentlsistance .P. falciparum to DHA-PPQ combination therapy in southern Vietnam has declined rapidly with the increase in the proportion of patients with parasite clearance time of more than 72\u00a0h (from 38% to 2012 to 57% in 2015) [-resistant P. falciparum in Vietnam and other countries in the GMS, it is important to know the current burden of P. falciparum infection and the effectiveness of anti-malarial drugs in Vietnam. The Hospital for Tropical Diseases (HTD) in Ho Chi Minh City is a tertiary hospital which receives patients with malaria from Central Highland and southern Vietnam including the Vietnam\u2013Cambodia border area. The presenting study was conducted at this hospital to examine the magnitude of P. falciparum infection, severe malaria and the response to anti-malarial treatment including the patterns of and predictors for treatment failure.Diagnostic tests used to identify anti-malarial drug resistance such as molecular assays are not always available, especially in low-resource countries . TherefoP. falciparum infection was extracted and included demographics , risk factors , travelling to malaria endemic areas domestically and internationally within 14 days before the onset of illness [Medical records of all patients with malaria admitted to the HTD between January 2015 and December 2018 were retrieved for review. Information derived from medical records of patients with  illness , outcomeP. falciparum infection was classified into different types of severe malaria in accordance with the WHO guidelines for the treatment of malaria and included shock, acute kidney failure, impaired consciousness, jaundice, anaemia, haemoglobinuria, acidosis, hyperparasitaemia, prostration, convulsion, hypoglycaemia and bleeding [All laboratory tests including microscopy were performed at the HTD and in line with the national laboratory performance standards. According to the HTD policy, all cases were diagnosed using microscopy. Once the diagnosis is confirmed, patients must be screened for all clinical and laboratory signs of severe malaria. Blood smear for the presence of parasites and parasite density is performed at an interval of six  or 12\u00a0h  until blood smear shows a negative result for at least two times. Based on the course of the disease, bleeding . Regardibleeding . Since Sbleeding . ETF andbleeding  and the bleeding , 19. TheP. falciparum infection was treated with a three-day DHA-PPQ age-based dosing therapy. Severe malaria was treated with intravenous artesunate 2.4\u00a0mg/kg on admission, at 12\u00a0h, and then every 24\u00a0h until oral therapy can be given [The presence of ETF among study participants was examined based on the clinical symptoms and laboratory tests. Regarding LTF, although study participants were not followed up on day 28 and day 42 after anti-malarial treatment due to the nature of a retrospective cohort study, information on previous malaria infection within 28 days was documented in the medical record as required by the HTD policy. Therefore, patients with LTF in the presenting study were those who met the WHO definition of LTF within 28 days before admission. In detail, LTF includes (i) danger signs or severe malaria in the presence of parasitaemia on any day between day 4 and day 28 in patients who did not previously meet any of the criteria of early treatment failure and (ii) presence of parasitaemia on any day between day 4 and day 28 (day 42) with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C in patients who did not previously meet any of the criteria of early treatment failure, or (iii) presence of parasitaemia on any day between day 7 and day 28 with axillary temperature\u2009<\u200937.5\u00a0\u00b0C in patients who did not previously meet any of the criteria of early treatment failure or late clinical failure. Before September 2016, uncomplicated be given . Since Sbe given . Given tbe given , 20, theP. falciparum infection was performed based on the number of subjects with malaria infection and the point estimate of the prevalence of P. falciparum infection. Similar calculations were performed for the prevalence of severe malaria, ETF and LTF. Chi-square, chi-square for trend and Fisher\u2019s Exact tests were used to compare categorical data. Student\u2019s t-test was used to compare continuous data. Multinomial logistic regression models were developed to test predictors of ETF and LTF. Independent variables including age, severe malaria, and acquiring malaria before 2017 were entered into the ETF model. Similarly, age, severe malaria, and patients with parasitaemia 72\u00a0h of treatment were entered into the LTF model. The significant level was set at P\u2009\u2264\u20090.05.Data were stored and analysed using SPSS version 22 (IBM). For comparison purposes, a calculation of 95% confidence interval (CI) for the prevalence of P. falciparum malaria accounted for 59.8% . Plasmodium vivax and Plasmodium malariae accounted for 35.7%  and 3.6% , respectively, while mixed P. vivax\u2013P. falciparum infections accounted for 0.9% . Among 265 P. falciparum infected patients, nearly half  of them acquired infection before 2017 and most of them (83.4%) were male with the mean age of 35.3\u2009\u00b1\u200913.1 years  had fever on admission, while only 0.8% had anaemia, 3% had splenomegaly, and 10.2% had hepatomegaly  had severe malaria, of whom single manifestation accounted for 77.6%  and 4 patients died. Eleven patients were from Binh Phuoc, 10 from Africa, 9 from Cambodia, two from Dak Nong, and one from each of the provinces including Binh Thuan, Dong Nai and Gia Lai. The remaining 23 patients had an unknown history of coming from or travelling to malaria endemic areas. Among 58 patients with severe malaria, 82.8% (48/58) were male and the mean age was 37.38\u2009\u00b1\u200914.17 years old. One patient (1.7%) had a history of injecting drug use. The mean number of days of illness at the time of admission was 6.9\u2009\u00b1\u20094.4. Fifteen patients (25.9%) had a number of days of illness\u2009>\u20097 days at the time of admission. The most common manifestations included jaundice , impaired consciousness , and shock . All five patients developing shock were adults, of whom three admitted to our hospital before September 2016 and all had a systolic blood pressure of \u2264\u200980 mmHg at the time of shock. Just under half  developed splenomegaly, hepatomegaly, or both, 13 (22.4%) required mechanical ventilation, 8 (13.8%) required haemodialysis, 12 (20.7%) received red blood cell transfusion. ETF and LTF developed in 12 (20.7%) and 8 (13.8%) patients with severe malaria, respectively. In the 58 patients presenting with severe falciparum malaria, 14 (24.1%) patients acquired malaria infection in regions of Vietnam where artemisinin resistance has been documented including Binh Phuoc (11 patients), Dak Nong (2 patients) and Gia Lai (1 patient). Under treatment with intravenous artesunate, the median (IQR) parasite half-life of 11 patients coming from Binh Phuoc was 3\u00a0h (2.3 to 8.3\u00a0h), while the parasite half-life of two patients coming from Dak Nong was 2.8 and 5.7\u00a0h and the parasite half-life of a patient coming from Gia Lai was 6.5\u00a0h. One  had a fatal outcome and was from Binh Phuoc. None of them required rescue treatment with parenteral quinine.Having fever and parasitaemia after 3 days of anti-malarial treatment were documented in 18 (31%) and 11 (19%) patients with severe malaria, respectively. Among 4 patients with severe malaria died, one patient was a 41-year-old male with an unknown history of travelling to malaria endemic areas who presented with impaired consciousness, acute kidney failure, and jaundice. Another patient was a 16-year-old female coming from Binh Phuoc with impaired consciousness, anaemia, and jaundice. The remaining two patients were those coming from or travelling to Africa with disease manifestations outlined the section listing the characteristics of patients coming from or travelling to Africa below. The mean number of inpatient days was 7.9\u2009\u00b1\u20094.3. Eleven patients developed hospital acquired infections which were described in the section listing the characteristics of patients with hospital acquired infections below. Based on the availability of the parasite count results among 58 patients with severe malaria, the mean parasite density was documented at 183,287\u2009\u00b1\u2009321,304 at the start of ACT among 58 patients, 153,095\u2009\u00b1\u2009205,081 at the 6th hour among 45 patients, 88,029\u2009\u00b1\u2009142,219 at the 12nd hour among 58 patients, 46,226\u2009\u00b1\u200991,477 at the 24th hour among 46 patients, 2552\u2009\u00b1\u20092272 at the 36th hour among 46 patients, and 1040\u2009\u00b1\u20091,180 at the 48th hour among 35 patients. The mean parasite half-life (50% reduction in parasitaemia during the linear phase of the log-parasitaemia over time) was 4.7\u2009\u00b1\u20092.1\u00a0h.ESBL)-producing Escherichia coli (two patients), Klebsiella pneumoniae (one patient), and Pseudomonas aeruginosa (two patients) were detected. The infectious agent was not detected in the remaining six patients.In addition to anti-malarial treatment, 5.7% (15/265) of patients received red blood cell transfusion, 4.9% 13/265) received mechanical ventilation, and 3% (8/265) undertook haemodialysis (Table\u00a0/265 receThirty-five (13.2%) and 40 (15.1%) patients had fever and parasitaemia after 3 days of anti-malarial treatment, respectively. The mean fever clearance time was 2.4\u2009\u00b1\u20091.5 days and the mean parasite clearance time was 53\u2009\u00b1\u200930.8\u00a0h. ETF and LTF were documented in 19  and 51  patients, respectively. ETF accounted for 4.7% (CI 2.3\u22129.4%) of patients coming from or travelling to Binh Phuoc and Central Highland, 12.9% (CI 6.7\u221223.5%) from other areas in Vietnam, 7.7% (CI 2.1\u221224.1%) from Cambodia and 6.9% (CI 1.9\u221222.0%) from Africa. Similarly, the proportion of LTF was 16.2% (CI 11.2 \u2212\u200923.0%) among patients coming from or travelling to Binh Phuoc and Central Highland, 22.5% (CI 14.0\u221234.4%) from other areas in Vietnam, 19.3% (CI 8.5\u221237.9%) from Cambodia and 27.6% (CI 14.7\u221245.7%) from Africa. The proportion of patients with parasitaemia on day 3 with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C was 3% (8/265). The mean number of inpatient days was 5.7\u2009\u00b1\u20093.5. There were 29 (10.9%) patients required ICU admission and the mean ICU length of stay was 2.9\u2009\u00b1\u20091.5 days. Most patients  recovered completely and 6.9% (4/58) of patients with severe malaria died. There was no association between having parasitaemia 72\u00a0h of treatment and response to treatment (P\u2009=\u20090.48). Among 207 patients with uncomplicated malaria, no one died or needed mechanical ventilation and haemodialysis, and one (1%) developed hospital acquired infection. Among 58 patients with severe malaria, eight required haemodialysis, 13 required mechanical ventilation, and 10 developed hospital acquired infection.Among 29 patients coming from or travelling to Africa, 96.6% (28/29) was male with a median age of 35.1\u2009\u00b1\u200910.2 years Table\u00a0. The meaSevere malaria was statistically associated with receiving treatment at previous hospitals (P\u2009<\u20090.001), hepatomegaly (P\u2009<\u20090.001) and number of inpatient days (P\u2009<\u20090.001) Table\u00a0. There wPatients with severe malaria were significantly more likely to develop ETF  and less likely to develop LTF  compared with those who did not have severe malaria Table\u00a0. Gender,No predictor for ETF was identified other than having severe malaria  Table\u00a0. No predP. falciparum infection. This rate is lower than the rate of 98% (CI 97.5\u221298.5%) reported from a provincial hospital in southern Vietnam in 1990 [P. falciparum in Vietnam [A total of 433 patients with malaria admitted to the HTD between 2015 and 2018. Of these 433 patients more than half  acquired  in 1990 . It is i Vietnam , 23. Add Vietnam . At this Vietnam .P. falciparum infection was recorded throughout the year and peaked during the months from November to March of the following year. In Vietnam, Lunar New Year holiday which is the most important holiday usually occurs in the second half of this period. It is observed that to financially prepare for the holiday many people travel to malaria endemic areas to work in this period in response to the increase in the number of seasonal jobs in these areas.In the presenting study, Blood transfusion and IDU are considered as malaria risk factors , 27. OnlAccording to the WHO definition of severe falciparum malaria  and the P. falciparum infected patients, while hepatomegaly was not mentioned in another study [Plasmodium infection, particularly in P. falciparum infection, this condition is usually transient [Regarding the clinical symptoms, similar to previous reports , 37, fever study , 36. Altransient . About 3ransient . One-fouransient . This coransient . In addiransient .Having a prolonged fever after receiving anti-malarial treatment is considered as a predictor of treatment failure , 42. In P. falciparum infection is prevalent in Africa [The rates of ETF and LTF in other areas in Vietnam were found to be not different to those in Binh Phuoc-Central Highland where DHA and PPQ resistance has been confirmed, this raises a big concern about the spread of malaria parasites that are resistant to both DHA and PPQ to other areas in Vietnam. It has been documented that artemisinin resistance has emerged in the GMS, followed by ACT failure, because both artemisinin and partner drug have decreased their susceptibility . Accordin Africa . The pron Africa  and resin Africa  have beeP. falciparum infection in many countries including Vietnam [P. falciparum [DHA-PPQ combination has been widely used as a first-choice therapy for  Vietnam . However Vietnam  and subs Vietnam . In Viet Vietnam . However Vietnam , 51. It  Vietnam . Indeed, Vietnam . A pharm Vietnam . In addi Vietnam \u201355. To olciparum . In Septlciparum . Howeverlciparum . This inA study on African children with uncomplicated falciparum malaria demonstrated the association between age less than 2 years and delayed parasite clearance . In addiThis study had some limitations. First, this is a single center study, and thus we may have missed patients with falciparum infection receiving treatment at other hospitals but were not referred to the HTD. However, the HTD is the only major tertiary teaching hospital for infectious diseases including malaria in southern Vietnam and receives not only local patients, but also patients from other countries. This would enhance the generalizability of the study findings. The study interval included the period when the WHO\u2019s new treatment guidelines were adopted. This allowed us to examine the change in the patterns of treatment failure in response to the utilization of the new guidelines. Second, it was unable to perform molecular tests to further examine the magnitude of parasites carrying mutations due to the nature of a retrospective cohort study. Third, examining the presence of parasitaemia on day 28 for all patients was not performed due to the same reason. Some cases who had parasitaemia on day 28 but did not exhibit any clinical symptom may have been missed. Therefore, the burden of LTF documented in this study may be underestimated.Plasmodium falciparum remains the prevalent malaria parasite in Vietnam. Despite the low mortality rate, severe falciparum malaria is not rare and having severe malaria is a significant predictor of ETF. To reduce the risk for ETF, studies are needed to examine the effectiveness of combination therapy including parenteral artesunate and a parenteral partner drug for severe malaria. The study alerts the risk of the spread of P. falciparum that is resistant to both DHA and PPQ to other areas in Vietnam and Africa which are currently known as non-endemic areas of anti-malarial drug resistance. A more comprehensive epidemiological survey using molecular technique in these regions is required to completely understand the magnitude of anti-malarial drug resistance and to design appropriate control strategies."}
+{"text": "Plasmodium falciparum kelch13 propeller gene mutations in sub-Saharan African pose the greatest threat to global efforts to control malaria. This is a critical concern in Uganda, where artemisinin-based combination therapy (ACT) is the first-line treatment for uncomplicated falciparum. The objective of this study was to compare the efficacy and safety of dihydroartemisinin\u2013piperaquine (DHA\u2013PQ) and artemether\u2013lumefantrine (AL) for the treatment of uncomplicated falciparum malaria in Ugandan children.The emergence of artemisinin resistance in Southeast Asia and A search of PubMed and the Cochrane Central Register of Controlled Trials for retrieving randomized controlled trials comparing the efficacy and safety of DHA\u2013PQ and AL for treatment of uncomplicated falciparum malaria in Ugandan children was done. The search was performed up to 31 August 2020. The data extracted from eligible studies and pooled as risk ratio (RR) with a 95% confidence interval (CI), using Rev Man Software (5.4). The protocol was registered in PROSPERO, ID: CRD42020182354.2\u2009=\u200993%, low quality evidence) and at day 42 . The PCR adjusted treatment failure at day 42 was significantly lower with DHA\u2013PQ treatment group , and it was below 5% in both arms at day 28 . AL showed a longer prophylactic effect on new infections which may last for up to 63\u00a0days . Compared to AL, DHA\u2013PQ was associated with a slightly higher frequency of cough . In both treatment groups, the risk of recurrent parasitaemia due to possible recrudescence was less than 5% at day 28. The appearance of gametocyte between 29 and 42\u00a0days was also significantly lower in DHA\u2013PQ than AL .Eleven trials were included in this review and two of them only included under safety outcome. Total 3798 participants were enrolled. The PCR unadjusted treatment failure was significantly lower with DHA\u2013PQ at day 28 (RR 0.30, 95% CI 0.19\u20130.49; participants\u2009=\u20097863; studies\u2009=\u20095; ICompared to AL, DHA\u2013PQ appeared to reduce treatment failure and gametocyte carriage in Ugandan children. This may trigger DHA\u2013PQ to become the first-line treatment option. Both treatments were safe and well-tolerated.The online version contains supplementary material available at 10.1186/s12936-021-03711-4. Plasmodium falciparum was the predominant and life-threatening parasite in Africa, causing 99.7% of estimated malaria cases in Africa ), but with no sign of severe malaria , 6. If ikelch13 (k13) gene, was detected in the Greater Mekong Sub-region and dozens of k13-propeller mutations have been detected at very low frequency in 18 countries in sub-Saharan Africa  OR 'falciparum malaria in children' [MeSH Terms] OR 'asymptomatic malaria in children' [MeSH Terms] AND 'artemisinin based combination therapy' [MeSH Terms] OR 'artemisinin' [MeSH Terms] OR 'artemether lumefantrine' [MeSH Terms] OR 'coartem' [MeSH Terms] OR 'dihydroartemisinin piperaquine' [MeSH Terms] OR 'Duocotecxin' [MeSH Terms] OR 'Eurartesim' [MeSH Terms] OR 'D-Artepp' [MeSH Terms])) AND 'randomized controlled trial' [MeSH Terms] OR 'controlled clinical trial' [MeSH Terms] OR 'randomized' [MeSH Terms] OR 'drug therapy'[MeSH Terms] OR 'trial' [MeSH Terms] OR 'groups' [MeSH Terms] OR 'humans' [MeSH Terms]).The search strategies in PubMed for the MeSH terms and text words was 'uncomplicated malaria in children' [MeSH Terms] OR 'uncomplicated The Cochrane Handbook for Systematic Reviews of Interventions , the Revfalciparum malaria. The information collected were trial characteristics including methods, participants, interventions, and outcomes as well as data on dose and drug ratios of the combinations. Relevant information such as title, name of the journal, year of publication, publication status, study design, study setting, follow-up period, sample size, funding source, baseline characteristics of study subjects, fever clearance, parasite clearance, treatment failure, and gametocyte carriage were extracted from each article using a structured data extraction format adapted from Cochrane. The number of participants randomized and the number analysed in each treatment group for each outcome were also captured. Two authors independently extracted the data and cross-checked. For dichotomous outcomes, the number of participants experiencing the event and the number of participants in each treatment group were documented. For continuous outcomes, the arithmetic means and standard deviations for each treatment group collectively with the numbers of participants in each group were extracted.The title and abstract were produced from the electronic search and independently screened by two authors based on RCTs that were assessed human The risk of bias for each trial was evaluated by two authors independently using the Cochrane Collaboration's tool for assessing the 'Risk of bias' . The risThe main outcomes in this review were total treatment failure at days 28, 42, and 63; PCR-adjusted and PCR unadjusted. Dichotomous data were combined and presented using risk ratios. Continuous data were summarized by arithmetic means and standard deviations, and then data were combined using mean differences. Risk ratios and mean differences were accompanied by 95% CIs. In the forest plot, the upper and the bottom tips of the diamond (the centre of the diamond) represents point estimate and the left and right tips of the diamond represents confidence interval. Also, the treatment arm is on the left side and the one in the right side is comparator arm.2 statistic were used to measure heterogeneity among the trials in each analysis, the Chi2 test with a P\u2009<\u20090.10 to indicate statistical significance was used, and the results were interpreted following Cochrane Handbook for Systematic Reviews of Interventions Version 6.0, Chapter 10: Analyzing data and undertaking meta-analyses [2\u2009>\u200950%) was identified, it was reported, and explored the possible causes by subgroup analyses.Heterogeneity among trials was assessed by inspecting the forest plots (to detect overlapping CI) and the Cochrane Q and Ianalyses . When suThe meta-analyses were done consistent with the recommendations of Cochrane . To aid The potential sources of heterogeneity were investigated through the following subgroup analyses: the known studies with HIV negative participants were compared to studies with both HIV negative and positive participants in the overall assessment because HIV infection has an effect of parasite clearance .Studies only with low risk of bias were included and to assess the small study effect, the fixed-effect and random-effect estimates of the intervention were compared.Quality of evidence was assessed using GRADE criteria and the GRADE pro software . The resA total of 488 trials through the databases were searched, of which 52 full-text trials for eligibility were assessed and found 10 of them fulfilled the inclusion criteria for meta-analysis and an additional one for qualitative analysis ; I2\u2009=\u200978%). Relative risks for the individual studies were: 0.33 , I2\u2009=\u200993.1%, Fig.\u00a0Consistently, in three studies to 0.46, ); and 0.2\u2009=\u20090%, Fig.\u00a0At day 28, the PCR adjusted treatment failures was below 5% in both treatment arms without significant difference between the two treatment groups . The result had considerable heterogeneity and we couldn't pool the result. The PCR unadjusted treatment failures for the individual studies were: 0.79 ; 0.36  and 0.5, 0.97 ; 2\u2009=\u20093%, Fig.\u00a0The overall PCR adjusted treatment failures was lower for participants treated with DHA\u2013PQ than those treated with AL . The result had considerable heterogeneity. It is more useful to consider individual trial results. At day 63, in one study [The PCR unadjusted treatment failure was not statistically different between the two treatment groups . Four trials with HIV negative participants, in two studies [Six studies with 2978 were reported in this outcome, but the pooled result showed considerable heterogeneity between studies. The result of studies was heterogeneous , I2\u2009=\u200979.6%).Two studies , 35 with2\u2009=\u200969%). Relative risks for the individual studies were: 0.35 ; 0.91 ; 1.10 ; 0.70 ; and 0.21, 0.59 , partici2\u2009=\u20090%, Additional file The prevalence of fever was similar over 3\u00a0days of follow up in both treatment groups in two trials , 39. The2\u2009=\u200966%, Additional file 2\u2009=\u200921, Additional file 2\u2009=\u20090%, Additional file The percentage of patients with parasitaemia at day one in two trials was significantly lower in the DHA\u2013PQ treatment group than AL  at baseline in both treatment groups . The quality of evidence was assessed on comparative adverse effects; cough slightly more frequent in DHA\u2013PQ arm was of high quality of evidence. Generally, the quality of evidence of safety of the two treatments ranges from low to high quality.The evidence that DHA\u2013PQ is more effective than AL at day 28, 42, and 63 unadjusted by genotyping was of This systematic review and meta-analysis focused on the safety and efficacy of DHA\u2013PQ and AL for the treatment of uncomplicated falciparum malaria in children. The main finding of this meta-analysis is that the PCR unadjusted risk of recurrent falciparum parasitaemia at day 28 and 42 was significantly lower for participants treated with DHA\u2013PQ than those treated with AL .Early treatment failure was observed in three patients from the DHA\u2013PQ group and 16 from the AL group had an early treatment failure in the three trials. At day 28, the PCR adjusted treatment failure was below 5% in both treatment arms without significant difference between the two treatment groups were observed . The PCR adjusted treatment failure day 42 was significantly lower for participants treated with DHA\u2013PQ than those treated with AL . Nevertheless, at day 63 the PCR adjusted treatment failure in participants treated with AL was significantly lower than those who are treated with DHA\u2013PQ .The appearance of gametocyte at day 29\u201342 was significantly lower in patients treated with DHA\u2013PQ than AL . In addition, In this review, most of the adverse events were mild or moderate severity and consistent with symptoms due to malaria. However, some adverse events like cough, anorexia, diarrhoea, and vomiting were the most common adverse events. In most studies, no significant difference was found in the proportion of study participants who experienced an adverse event of moderate and great severity between the DHA\u2013PQ and AL treatment groups. But, cough was significantly more frequent in patients treated with DHA\u2013PQ than AL .Pfk13 non-synonymous mutations (R622I) with unknown impact on the parasite resistance phenotype have been seen at a very low rate.The observed high efficacy of DHA\u2013PQ was similar to that of other studies conducted in Africa , 44 and Recent studies conducted in Mali, Somalia, Angola, and Papua New Guinea also reported that both DHA\u2013PQ and AL were highly effective in the treatment of uncomplicated falciparum malaria , 47\u201349. Similarly, a recent study in Rwanda reported that 42\u00a0day PCR corrected efficacy was significantly better in patients with falciparum malaria treated with DHA\u2013PQ . Hence, In this review several studies using microscopic detection of gametocytes have shown no difference , 60\u201362 aIn this systematic review, significant haematological recovery from the baseline has been observed among patients treated with AL than DHA\u2013PQ at day 42. Recent studies in Africa have reported that there was significant haematological recovery from the baseline in both treatment groups , 56. OneA recent study in Papua New Guinea reported a high frequency of cough without significant difference between the two treatment groups . A formeIn this systematic review, four studies reported 21 serious adverse events in DHA\u2013PQ and 13 in the AL treatment group. However, the distributions of serious adverse events were not significantly different in the two treatment groups. All serious adverse events were not related to study medications. No death has occurred in any of the studies. This might be justified by the fact that these studies were conducted among participants with uncomplicated malaria rather than the severe form which can lead to death.P. falciparum is endemic molecular surveillance may also play an important role in detecting genetic markers.Treatment failure could be occurred due to the drug\u2019s ineffectiveness or development of resistance, as it may be due to insufficient drug levels . FurtherThe study has some limitations. A majority of included studies were conducted in Tororo District Eastern region Uganda where malaria transmission intensity is high. The result of this study might not be representative of other regions in Uganda where malaria transmission intensity is low and moderate. Most studies reported treatment failure at 28 and 42\u00a0days, this review might not provide strong evidence about the long-term post-treatment prophylactic effect of the two drugs.This systematic review provides comprehensive evidence about the treatment efficacy and safety of ACT in children in an area of malaria-endemic areas in Uganda. The overall parasite clearance, drug efficacy, and safety were good enough. Compared to AL, DHA\u2013PQ appeared to reduce treatment failure and gametocyte carriage in Ugandan children. This may trigger DHA\u2013PQ to become the first-line treatment option. Both treatments were safe and tolerable. As ACT resistance is emerging in different parts of the world, continuous studies that measure the efficacy of DHA\u2013PQ and AL with 42 and 63\u00a0days follow-up are needed.Additional file 1:\u00a0S1. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: PCR-unadjusted treatment failures at day 63.Additional file 2:\u00a0S2. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: PCR-adjusted treatment failures at day 63.Additional file 3:\u00a0S3. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: Fever clearances on day 1.Additional file 4:\u00a0S4. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: Fever clearances on day 2.Additional file 5:\u00a0S5. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: Fever clearances on day 3.Additional file 6:\u00a0S6. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: Parasite clearances.Additional file 7:\u00a0S7. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: Gametocyte carriages at baseline.Additional file 8:\u00a0S8. Forest plot of comparison: Dihydroartemisinin-piperaquine versus artemether-lumefantrine, outcome: Anemia.Additional file 9. Additional Tables: GRADE Summary of finding tables."}
+{"text": "This research aims to study the efficacy of tannins co-supplementation on disease duration, severity and clinical symptoms, microbiota composition and inflammatory mediators in SARS-CoV2 patients.This is a prospective, double-blind, randomized, placebo-controlled, parallel-group trial to evaluate the efficacy of the administration of the dietary supplement ARBOX, a molecular blend of quebracho and chestnut tannins extract and Vit B12, in patients affected by COVID-19.https://www.argentina.gob.ar/salud/coronavirus-COVID-19/definicion-de-caso).18 years of age or older, admitted to Hospital de Clinicas Jose de San Martin, Buenos Aires University (Argentina), meeting the definition of \"COVID-19 confirmed case\" Participants are eligible to be included in the study if the following criteria apply:Exclusion CriteriaPregnant and lactating patientsPatients who cannot take oral therapy Hypersensitivity to polyphenolsPatients already in ICU or requiring mechanical ventilationPatients already enrolled in other clinical trialsDecline of consentParticipants are excluded from the study if any of the following criteria apply:TREATED ARMExperimental: Participants will receive a supply of 28 -- 390 mg ARBOX capsules for 14 days. Patients will be supplemented with 2 capsules of ARBOX per day.CONTROL ARMPlacebo Comparator: Participants will receive placebo supply for 14 days. The placebo will be administered with the identical dose as described for the test product.All trial participants will receive standard therapy, which includes: Antipyretics or Lopinavir / Ritonavir, Azithromycin and Hydroxychloroquine, as appropriate . In addition, if necessary: supplemental O2, non-invasive ventilation, antibiotic therapy.Primary Outcome Measures:Time to hospital discharge, defined as the time from first dose of ARBOX to hospital discharge [ Time Frame: Throughout the Study (Day 0 to Day 28) ]Secondary Outcome Measures:28-day all-cause mortality [ Time Frame: Throughout the Study (Day 0 to Day 28) ]-proportionInvasive ventilation on day 28 [ Time Frame: Throughout the Study (Day 0 to Day 28) ]-proportionLevel of inflammation parameters and cytokines [ Time Frame: day 1-14 ] -mean differenceDifference in fecal intestinal microbiota composition and intestinal permeability [ Time Frame: day 1-14 ]Negativization of COVID-PCR at day 14 [ Time Frame: day 14 ]-proportionPotential study participants were screened for eligibility 24 hours prior to study randomization. Patients were randomly assigned via computer-generated random numbering (1:1) to receive standard treatment coupled with tannin or standard treatment plus placebo (control group).Study personnel and participants are blinded to the treatment allocation, as both ARBOX and placebo were packed in identical containers. Thus, all the used capsules had identical appearance.Considering an alpha error of 5%, a power of 80% a sample size of 70 patients per branch was estimated. 140 patients in total.The protocol version is number V2, dated May 23, 2020.The first patient, first visit was on June 12, 2020; the recruitment end date was October 6, 2020.The protocol was not submitted earlier because the enrollment of some patients took place after the closure of the recruitment on the clinicaltrials platform. In fact, due to the epidemiological conditions, due to the decrease of the cases in Argentina during the summer period, the recruitment stopped t before reaching the number of 140 patients (as indicated in the webpage). However, since there was a new increase in cases, the enrolment was resumed in order to reach the number of patients initially planned in the protocol. The final participant was recruited on February 14, 2021.NCT04403646, registered on May 27th, 2020.ClinicalTrials.gov, number: The full protocol is attached as an additional file, accessible from the Trials website (Additional file The online version contains supplementary material available at 10.1186/s13063-021-05281-x. Additional file 1. Full study protocol."}
+{"text": "Plasmodium chabaudi, to test whether the daily rhythms of hosts, parasites and their interactions affect sensitivity to the key antimalarial, artemisinin.Circadian rhythms contribute to treatment efficacy in several non-communicable diseases. However, chronotherapy (administering drugs at a particular time-of-day) against infectious diseases has been overlooked. Yet, the daily rhythms of both hosts and disease-causing agents can impact the efficacy of drug treatment. We use the rodent malaria parasite Asexual malaria parasites develop rhythmically in the host\u2019s blood, in a manner timed to coordinate with host daily rhythms. Our experiments coupled or decoupled the timing of parasite and host rhythms, and we administered artemisinin at different times of day to coincide with when parasites were either at an early (ring) or later (trophozoite) developmental stage. We quantified the impacts of parasite developmental stage, and alignment of parasite and host rhythms, on drug sensitivity.We find that rings were less sensitive to artemisinin than trophozoites, and this difference was exacerbated when parasite and host rhythms were misaligned, with little direct contribution of host time-of-day on its own. Furthermore, the blood concentration of haem at the point of treatment correlated positively with artemisinin efficacy but only when parasite and host rhythms were aligned.in vivo. The hitherto unknown modulation by alignment between parasite and host daily rhythms suggests that disrupting the timing of parasite development could be a novel chronotherapeutic approach.Parasite rhythms influence drug sensitivity We reveal that chronotherapy (providing medicines at a particular time-of-day) could improve treatment for malaria infections. Specifically, parasites\u2019 developmental stage at the time of treatment and the coordination of timing between parasite and host both affect how well antimalarial drug treatment works. That circadian rhythms affect drug treatment outcomes  has been known for decades. Chronotherapeutic approaches aim to optimise the timing of drug administration by considering how daily changes in gene expression and physiology influence the absorption, metabolism, toxicity and half-life of drugs, the availability of drug targets, and co-active immune effectors . For exaPlasmodium) parasites replicate synchronously in the host\u2019s blood, with the duration of the intra-erythrocytic developmental cycle (IDC) taking multiples of 24\u2009h depending on the species . The IDiewed in , 12). Deiewed in , such asiewed in , 15, whiin vitro efficacy tests and in vivo treatment outcomes, reveal how temporary arrest (quiescence) at certain IDC stages confers tolerance to antimalarials and allow malaria treatment to be optimised by chronotherapy. Here, we use the rodent malaria parasite Plasmodium chabaudi to investigate whether both host and parasite rhythms contribute to antimalarial efficacy. We used the short-acting drug artemisinin to test for proof-of-principle, whilst acknowledging the ongoing importance of artemisinin-based therapies in malaria management. By manipulating whether IDC rhythms are aligned with host rhythms or not, and treating infections at different times of day with artemisinin, we tested whether drug efficacy depends on the parasite\u2019s IDC stage, host time-of-day or their interaction. Additionally, we assessed putative correlates (haem and blood glucose concentration) of host rhythms and drug efficacy. Haem is an essential molecule for IDC progression, which parasites can biosynthesise, or scavenge from host haemoglobin [Understanding the relative contributions of host and parasite rhythms to antimalarial efficacy could resolve the disconnect between moglobin . Intracemoglobin , 47, andmoglobin . Thus, dmoglobin  and is amoglobin , suggestWe reveal that parasites early in their IDC (rings) are less sensitive to artemisinin than mid-IDC parasites (trophozoites). Host time-of-day did not directly affect drug efficacy but the coordination between parasite and host rhythms did. Specifically, when the timing of the IDC was misaligned (mismatched) with host rhythms, rings exhibited even lower drug sensitivity and trophozoites became more sensitive. Whereas glucose levels did not correlate with drug efficacy, haem concentration at the point of treatment correlated positively with drug efficacy, but only in matched infections.We carried out two experiments to test whether host and parasite rhythms affect artemisinin efficacy. The first experiment revealed intra-erythrocytic developmental cycle (IDC)-stage-specific efficacy and suggested an impact of host time-of-day. The second experiment tested whether the apparent role of host time-of-day was better explained by the alignment between parasite and host rhythms.P. chabaudi chabaudi (genotype CW) originally isolated from Grammomys poensis . Hosts were 7- to 11-week-old male C57BL/6 mice which had access to food ad libitum and drinking water was supplemented with 0.05% para-aminobenzoic acid [Mus musculus is a natural host for some rodent malaria species [We used the synchronous species oic acid . Mus mus species . All micBoth experiments followed the same general set-up. We expanded cryopreserved parasites through 2\u20133 passages in donor mice kept in a 12 h light\u201312 h dark photoschedule . This ensured parasites\u2019 IDC schedule was aligned with host rhythms. To initiate experimental infections, we diluted blood from donor mice in citrate saline  and passaged parasites by intravenous injection of ring stage infected red blood cells (iRBCs) into two groups of experimental mice simultaneously. One group of experimental mice were in the same photoschedule as the donor mice , so that parasite and host rhythms were aligned in experimental hosts (\u2018matched\u2019 infections). The other group of experimental mice was in the opposite photoschedule , such that parasites transferred from donor to experimental mice experienced an instantaneous 12-h shift in host time-of-day  . To ensun\u2009=\u200940) with 108 ring stage iRBCs and drug treated on Day 2 post infection (p.i.). At this point, the IDC of mismatched parasites was misaligned to host rhythms by \u223c7\u2009h , or during the trophozoite stage (ZT8 for n\u2009=\u20098 matched infections and ZT15 for n\u2009=\u20098 mismatched infections) (t0) to quantify parasites and measure haem and glucose levels, and also quantified parasites after 24\u2009h (t24). We quantified parasites from 5-\u00b5l blood samples using a semi-automatic Kingfisher Flex Magnetic Particle Processor and MagMAX\u2122-96 DNA Multi-Sample Kit (Thermo Fisher Scientific) with slight modifications from the standard protocol 4413021DWblood [\u00ae Performa Nano device (Roche). For the haem assay , absorbance was measured at 405\u2009nm using a Multiskan Ascent 96/384 Plate Reader (MTX Lab Systems) in a single plate, following manufacturer\u2019s instructions. We were unable to measure one haem sample .We initiated infections ( by \u223c7\u2009h . Based oections) . We took1DWblood  and enum1DWblood . We measn\u2009=\u200930) at a 10-fold lower parasite density (107 ring stage iRBCs) than Experiment 1, followed by drug treatment on Day 3 p.i. This allowed mismatched parasites an additional IDC to reschedule, so that parasite and host rhythms in mismatched infections were only misaligned by \u223c4\u2009h at t0. Therefore, each IDC stage could be treated in the same host phase across matched and mismatched infections , and trophozoites were treated in the resting (light) phase (ZT8 for n\u2009=\u20095 matched infections and ZT11.5 for n\u2009=\u20095 mismatched infections)  and determined the proportion of iRBCs by microscopic counting from thin blood smears stained with 20% Giemsa.The aim of the first experiment was to target rings and trophozoites in both matched and mismatched infections. This resulted in parasites in matched and mismatched infections being treated at times coupled to their hosts\u2019 active or resting phase . Therefofections . Specifiections) . We tookhttps://www.R-project.org/) using the base R system unless otherwise indicated. To assess any differences between treatment groups at t0, we compared parasite densities, as well as glucose and haem levels for Experiment 1, between treatment groups using linear models with IDC \u2018Stage\u2019 (rings or trophozoites), \u2018Alignment\u2019 (matched or mismatched infections) and their interaction as explanatory variables. Parasite densities were log10-transformed to meet assumptions of normality and homogeneity of variance. Because the time-of-day of drug administration varied between groups, parasites in some groups had gone through an additional part-cycle of replication, resulting in differences in parasite densities at t0 ) by fitting parasite density at t24 (ParDens(t24)), in linear models including an offset for parasite density at t0 (ParDens(t0)). Results are presented as the corresponding fold change:Data were analysed with R v.3.6.3  (AICc) for small sample sizes; \u2018MuMIn\u2019 package). We report results for the minimally adequate models in the main text, and full statistical outcomes in t0 based on the minimised model shown in To confirm artemisinin-treated infections had fewer parasites than placebo-treated controls regardless of whether infections were matched or mismatched, we tested a linear model including the terms \u2018Drug\u2019 (drug-treated or placebo-treated infections), \u2018Alignment\u2019 (matched or mismatched infections) and their interaction . To testt0)  = 5.48, P\u2009=\u20090.027; Experiment 2, Stage \u00d7 Alignment interaction: F = 1.90, P\u2009=\u20090.187, Stage: F = 11.14, P\u2009=\u20090.004) ).Parasites were treated at either ring or trophozoite stage and, as expected, the intended intra-erythrocytic developmental cycle (IDC) stage predominated in the blood in all treatment groups just prior to treatment (t0) . Because=\u20090.004) . This coF = 29.94, P\u2009<\u20090.001; Experiment 2, Drug: F = 28.64, P\u2009<\u20090.001)  \u00b1 SEM 4.22\u2009\u00b1\u20090.93 for matched and 3.54\u2009\u00b1\u20091.64 for mismatched infections in Experiment 1, and 1.47\u2009\u00b1\u20090.18 for matched and 1.65\u2009\u00b1\u20090.07 for mismatched infections in Experiment 2). In contrast, parasite densities decreased or remained unchanged in artemisinin-treated infections  \u00b1 SEM 0.52\u2009\u00b1\u20090.10 for matched and 0.98\u2009\u00b1\u20090.30 for mismatched infections in Experiment 1, and 0.44\u2009\u00b1\u20090.05 for matched and 0.50\u2009\u00b1\u20090.12 for mismatched infections in Experiment 2)  . Specifiiment 2) .rel) in Experiment 1 varied significantly according to whether we treated rings or trophozoites and whether host and parasite rhythms were aligned  = 10.19, P\u2009=\u20090.003)  \u00b1 SEM 0.69\u2009\u00b1\u20090.18 for matched rings and 0.35\u2009\u00b1\u20090.07 for matched trophozoites). When parasite and host rhythms were mismatched, drug efficacy was enhanced for trophozoites and weakened for rings  \u00b1 SEM 1.73\u2009\u00b1\u20090.48 for mismatched rings and 0.23\u2009\u00b1\u20090.03 for mismatched trophozoites) (Drug efficacy (\u0394ParDens=\u20090.003) . In matcozoites) . Thus, mF = 1.53, P\u2009=\u20090.226, Phase: F = 10.01, P\u2009=\u20090.004, Stage: F = 38.11, P < 0.001)  or resting (light) phase because upon misalignment, the appearance of each IDC stage in the blood also shifted to the opposite host phase compared to aligned infections . Therefo< 0.001) . Hence, F = 13.52, P\u2009=\u20090.002)  \u00b1 SEM 0.56\u2009\u00b1\u20090.05 for matched rings and 0.31\u2009\u00b1\u20090.03 for matched trophozoites), and this difference was exacerbated by mismatch  \u00b1 SEM 0.85\u2009\u00b1\u20090.08 for mismatched rings and 0.15\u2009\u00b1\u20090.04 for mismatched trophozoites). Therefore, alignment between host and parasite rhythms has a greater influence on IDC-stage-specific drug efficacy than host phase per se.Administering treatment 1\u2009day later in Experiment 2 enabled parasites to reschedule further. Consequently, rings in Experiment 2 could be treated during the host\u2019s active phase for both matched and mismatched infections, whilst all trophozoites were treated during the resting phase . If the =\u20090.002) , revealit0 in Experiment 1 ranged from 2.95 to 10.38\u2009mM, but did not differ significantly between treatment groups  = 1.30, P\u2009=\u20090.264, Alignment: F = 0.01, P\u2009=\u20090.941, Stage: F = 0.45, P\u2009=\u20090.508)  = 6.24, P\u2009=\u20090.019) . Haem concentrations correlated positively with artemisinin efficacy for both rings and trophozoites, but only in matched infections  = 5.87 P\u2009=\u20090.024) (Haem concentrations at =\u20090.508) . Despite=\u20090.508)  was sign=\u20090.019) . Includifections . When us=\u20090.024) . However=\u20090.024) . This sut0 in Experiment 1 ranged from 7.7 to 12.4\u2009mmol/L. The alignment of parasite and host rhythms did not affect blood glucose concentrations  = 0.14, P\u2009=\u20090.709; Alignment: F = 3.10, P\u2009=\u20090.089), but blood glucose was 9.6% higher when trophozoites were treated (mean concentration = 10.77\u2009\u00b1\u20090.23\u2009mmol/L) compared to when rings were treated (mean concentration = 9.83\u2009\u00b1\u20090.36\u2009mmol/L)  = 5.00, P\u2009=\u20090.033)  of infections treated as rings or trophozoites, irrespective of using \u2018Alignment\u2019 or \u2018Phase\u2019 in the analyses (t0.Blood glucose concentrations at =\u20090.033) . Howeveranalyses  and 3C, in vivo and used a fast-acting antimalarial to test their independent and interacting effects on drug efficacy. Our experiments revealed three phenomena. First, mid-ring stages of P. chabaudi are less sensitive to artemisinin than mid-trophozoites. Second, alignment between parasite and host rhythms differentially affects drug sensitivity of rings and trophozoites; mismatch causes rings to become less sensitive and trophozoites to become more sensitive. Third, unlike blood glucose concentrations, haem levels in the blood at the time of drug administration (t0) positively correlate with artemisinin efficacy (regardless of the IDC stage treated) but only in infections where host and parasite rhythms are aligned.We experimentally separated parasite intra-erythrocytic developmental cycle (IDC) rhythms and host rhythms That we find rings are less sensitive to artemisinin treatment than trophozoites, is in keeping with other studies that also challenge mid-rings, rather than hypersensitive very early rings , 32, 33 P. falciparum [The differential impact of the alignment between IDC stages and host rhythms on drug sensitivity of rings versus trophozoites cannot simply be explained by the effects of the host\u2019s active versus resting phase on drug activity. When the alignment between parasite and host rhythms was lost, regardless of whether treatment was administered during the host\u2019s active or resting phase, trophozoites became more sensitive and rings less sensitive to artemisinin treatment, thus aggravating the stage-specific differences in drug efficacy. Artemisinin is an important currently used antimalarial, and if our findings translate to human malaria infections, then timing administration of this short-acting drug against trophozoites would maximise efficacy. The World Health Organisation recommends artemisinin combination therapy to be administered for 3\u2009days against the human malaria parasite lciparum , 63, whilciparum ). Whilstlciparum ), then aP. chabaudi parasites [in vitro [P. falciparum lines selected for artemisinin resistance show transcriptional changes for similar pathways [Finding targets for IDC-schedule-disrupting drugs will be facilitated by understanding why mismatch exacerbates stage-specific differences in drug sensitivity. Clues could lie in the altered gene expression patterns recently uncovered in mismatched arasites . This inarasites . Many ofarasites , 65; . Care should be taken when extrapolating results to human malaria, among others because P. falciparum has a 48-h IDC and humans are not nocturnal. However, like P. chabaudi, many genes of P. falciparum are transcribed with 24-h periodicity [P. chabaudi, and have been implicated in artemisinin resistance in both rodent malarias and P. falciparum croscopy  or qPCR croscopy  to infercroscopy , 82, andcroscopy , 58, 82.eoab013_Supplementary_DataClick here for additional data file."}
+{"text": "Since the introduction of artemisinin-based combination therapy (ACT) in Ghana in 2005 there has been a surveillance system by the National Malaria Control Programme (NMCP) and the University of Ghana Noguchi Memorial Institute for Medical Research (UG-NMIMR) to monitor the therapeutic efficacy of ACTs for the treatment of uncomplicated malaria in the country. We report trends and determinants of failure following treatment of Ghanaian children with artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL) combinations.Per protocol analyses as well as cumulative incidence of day 28 treatment failure from Kaplan Meier survival analyses were used to describe trends of failure over the surveillance period of 2005\u20132018. Univariable and multivariable cox regression analyses were used to assess the determinants of treatment failure over the period.Day 28 PCR-corrected failure, following treatment with ASAQ, significantly increased from 0.0% in 2005 to 2.0% (95% CI: 1.1\u20133.6) in 2015 (p\u2009=\u20090.013) but significantly decreased to 0.4% (95% CI: 0.1\u20131.6) in 2018 (p\u2009=\u20090.039). Failure, following treatment with AL, decreased from 4.5% (95% CI: 2.0\u20139.4) in 2010 to 2.7% (95% CI: 1.4\u20135.1) in 2018, though not statistically significant (p\u2009=\u20090.426). Risk of treatment failure, from multivariable cox regression analyses, was significantly lower among children receiving ASAQ compared with those receiving AL ; lower among children with no parasitaemia on day 3 compared with those with parasitaemia on day 3 ; and higher among children who received ASAQ and had axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C on day 1 compared with those with axillary temperature\u2009<\u200937.5\u00a0\u00b0C .Treatment failures for both ASAQ and AL have remained less than 5% (below WHO\u2019s threshold of 10%) in Ghana since 2005. Predictors of treatment failure that need to be considered in the management of uncomplicated malaria in the country should include type of ACT, day 3 parasitaemia, and day 1 axillary temperature of patients being treated.The online version contains supplementary material available at 10.1186/s12879-021-06961-4. Since the discovery of malaria parasites by Charles Louis Alphonse Laveran in 1880 . About 6Plasmodium falciparum malaria [Prompt and effective treatment of uncomplicated malaria remains one of the key interventions within the Global Technical Strategy for Malaria 2016\u20132030) because it has the advantage of preventing the progression to severe illness and death . Since A\u20132030 bec malaria . The ACT malaria , 9.Ghana adopted the use of ACTs in 2004 when it had become clear that the therapeutic efficacies of chloroquine and sulphadoxine/pyrimethamine, which were the first- and second-line drugs for uncomplicated malaria, were less than 70% . In 2005Following the introduction of ACTs in 2005, a surveillance system was established by the National Malaria Control Programme (NMCP), in collaboration with the University of Ghana Noguchi Memorial Institute for Medical Research (UG-NMIMR), to continuously monitor the therapeutic efficacies of ACTs to inform antimalaria drug policy in the country using the recommended WHO criteria, which has the proportion of patients parasitaemic on day 3 as indicator of suspected artemisinin partial resistance and the proportion of treatment failure by day 28 or 42  as indicator of partner drug resistance Fig.\u00a0. We repoSince 2005, ACT efficacy studies have been conducted in purposively selected ten (10) sentinel sites across Ghana. These sites represent the previously ten administrative regions of the country and the three main ecological zones: savannah, forest, and coastal. Three of these sites  are located within the northern belt, which is savannah; four  are located within the middle belt, which is forest; and three are located within the southern belt, which is forest  and coastal   approach using WHO 2003 and 2009 protocols , 15.P. falciparum detected by microscopy; ability to swallow oral medication; parental consent; parent/guardian willing to comply with study protocol for the duration of the study; and parent/guardian willing to comply with follow-up schedule.A summary of specific inclusion criteria adopted for surveillance activities from 2005 to 2009 using the WHO 2003 protocol include: children aged 6\u00a0months to 59\u00a0months; axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C; and asexual parasitaemia of 2,000\u2013200,000/\u00b5L. A summary of specific inclusion criteria adopted for surveillance activities from 2010 to 2018 using the WHO 2009 protocol include: children aged 6\u00a0months to 9\u00a0years (extended to 14\u00a0years in some studies)  and glutamate-rich protein (glurp) were used to distinguish between re-infection and recrudescence as recommended by WHO [msp2 was used based on the observation that msp1 was less discriminatory and that glurp was prone to \u201cartefact bands\u201d [All treatment given followed the DOT approach. There were fixed and non-fixed combination therapies Table . The fold by WHO \u201319. Fromt bands\u201d , 20, 21.The main outcome variable was treatment failure from data collected during routine surveillance years  and years when in-vitro/ex-vivo studies were being conducted in some sentinel sites Table . As per Per protocol analyses as well as cumulative incidence of treatment failure from Kaplan Meier survival analyses were used to describe trends of PCR-uncorrected and PCR-corrected day 28 treatment failure over the surveillance period. The estimates of treatment failure were obtained from the WHO Excel\u00ae data template used in the studies . Data foUnivariable analysis was performed using the Cox Proportional Hazard Model (CPHM) to assess the association between each covariate and treatment failure and to determine variables to be considered in a multivariable analysis based on a significance level of less than or equal to 20%. Covariates considered for treatment failure, using pooled data from all ten (10) study sites, were gender, age, ecological zone, drug type, parasite density at enrolment (day 0), parasitaemia on day 3, vomiting at least once during the three days of treatment, axillary temperature on day 0, and axillary temperature on day 1. Variables that were not statistically significant but considered to be clinically important were included in the adjusted or multivariable model.Multivariable cox regression analysis was used to determine the simultaneous effect of multiple risk factors on treatment failure. Analyses were done for the overall pooled ASAQ and AL data as well as separately pooled data for ASAQ and AL. The Hazard ratio, which is the exponent of each regression coefficient was used in the interpretation of model results. All tests were conducted with 95% confidence interval. P-values\u2009\u2264\u20090.05 were considered statistically significant. A cluster term was included in the model to account for clustering of data by study site. All cox regression analyses were carried out using R software (Version 4.0.2). Proportional hazard assumption test performed showed no violations at 5% significance level , did not vomit during treatment (86.7%), had parasite density\u2009<\u200950,000/\u00b5L (55.1%), had no parasitaemia on day 2 post-treatment (97.0%), and no parasitaemia on day 3 post-treatment (99.7%) ,\u2009<\u20095\u00a0years old (60.7%), exposed to ASAQ treatment (69.9%), had axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C on day of enrolment (79.7%), had axillary temperature\u2009<\u200937.5%\u00a0\u00b0C on day 1 post-treatment (91.5%), had axillary temperature\u2009<\u200937.5%\u00a0\u00b0C on day 2 post-treatment (91.5%), had axillary temperature\u2009<\u200937.5%\u00a0Following treatment with ASAQ, PCR-uncorrected per protocol failure on day 28 (post-treatment) significantly declined from 7.9% (95% CI: 5.6\u201311.0) in 2005 to 3.5% (95% CI: 2.3\u20135.4) in 2015 (p\u2009=\u20090.003), and significantly declined further to 1.2% (95% CI: 0.5\u20132.8) in 2018 (p\u2009=\u20090.025). PCR-corrected treatment failure on day 28 (post-treatment) significantly increased from 0.0% in 2005 to 2.0% (95% CI: 1.1 -3.6) in 2015 (p\u2009=\u20090.013), but significantly decreased to 0.4% (95% CI: 0.1\u20131.6) in 2018 (p\u2009=\u20090.039). Similar patterns were observed with KM analysis  declined from 11.8% (95% CI: 7.5\u201317.9) in 2010 to 6.8% (95% CI: 4.6\u20139.9) in 2018 but this was not statistically significant (p\u2009=\u20090.073). Similarly, PCR-corrected treatment failure declined from 4.5% (95% CI: 2.0 -9.4) in 2010 to 2.7% (95% CI: 1.4\u20135.1) in 2018 but this was not statistically significant (p\u2009=\u20090.426). Similar patterns were observed with KM analysis ; significantly lower among patients without no parasitaemia on day 3 compared with those with parasitaema ; significantly lower among patients who received ASAQ treatment compared with those who received AL treatment ; and significantly lower among patients who vomited at least once during the three days of treatment compared with those who did not vomit . PCR-corrected treatment failure was significantly higher in the coastal zone compared with the savannah zone ; significantly lower among patients with no parasitaemia on day 3 compared with those with parasitaemia ; and significantly lower among patients who received ASAQ treatment compared with those who received AL treatment  ; significantly higher among patients with axillary temperature on day 0\u2009\u2265\u200937.5\u00a0\u00b0C compared with those with temperature\u2009<\u200937.5\u00a0\u00b0C ; and significantly lower among patients with no parasitaemia on day 3 compared with those with parasitaemia . PCR-corrected ASAQ treatment failure was significantly higher among patients with axillary temperature on day 1\u2009\u2265\u200937.5\u00a0\u00b0C compared with those with temperature\u2009<\u200937.5\u00a0\u00b0C ; and significantly lower among patients with no parasitaemia on day 3 compared with those with parasitaemia   and , respectively ; significantly higher in the coastal zone compared with the savannah zone ; significantly lower among patients with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C on day 1 compared with those with axillary temperature\u2009<\u200937.5\u00a0\u00b0C ; significantly lower among patients with no parasitaemia on day 3 compared with those with parasitaemia ; and significantly lower among patients who received ASAQ treatment compared with those who received AL treatment  ; and significantly lower among patients with no parasitaemia on day 3 compared with those with parasitaemia  ; and significantly lower among patients with no parasitaemia on day 3 compared with those with parasitaemia  ; lower among patients with no parasitaemia on day 3 compared with those with parasitaemia ; and significantly lower among patients who received ASAQ treatment compared with those who received AL treatment   to distinguish between reinfection and recrudescence during the surveillance period between 2014 and 2018 [PCR-corrected treatment failures over the years have remained less than 5% for both ASAQ and AL suggesting that Ghana, as other countries in the sub-region, has not reached the failure threshold of 10% necessary for treatment policy change  appears to be a hotspot for the spread of drug resistant parasites. The relatively high cosmopolitan nature of the sites within the coastal zone is likely to have resulted in a higher level of human population movement which has the potential of facilitating genetic recombination and subsequent phenotypic traits of reduced drug susceptibility within the zone \u201337.Generally, risk of PCR-uncorrected and PCR-corrected treatment failures were respectively, 92 and 98% higher among children with parasitaemia on day 3 compared with those without parasitaemia. The high risk of treatment failure, either PCR-uncorrected or PCR-corrected, associated with parasitaemia on day 3 was observed after treatment with both ASAQ and AL, suggesting that parasitaemia on day 3 is a key predictor of either PCR-uncorrected or PCR-corrected treatment failure following ACT treatment.Risk of treatment failure following treatment with ASAQ was about four times higher among children with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C on day 1 post-treatment compared with those with temperature\u2009<\u200937.5\u00a0\u00b0C. On the contrary, risk of failure following treatment with AL was 79% greater among children with temperature\u2009<\u200937.5\u00a0\u00b0C on day 1 post-treatment compared with those with temperature\u2009\u2265\u200937.5\u00a0\u00b0C. Amodiaquine is a 4-aminoquinoline derivative with anti-inflammatory properties in addition to antimalarial properties compared with Lumefantrine, which is a fluorine with mainly antimalarial properties , 39. It Failure rates following treatment of Ghanaian children with uncomplicated malaria using ASAQ and AL have remained less than 5% between 2005 and 2018 warranting their continuous use in the country. Children at higher risk of treatment failure have been those receiving AL; those with parasites on day 3; those residing within the coastal zone; those with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C on day 1 post-treatment (for ASAQ); and those with axillary temperature\u2009<\u200937.5\u00a0\u00b0C on day 1 (for AL). These predictors of treatment failure should guide management of uncomplicated malaria in Ghana.Additional file 1. Proportional hazard assumption test results.Additional file 2. Kaplan Meier survival curve for PCR-corrected ASAQ and AL treatment failure."}
+{"text": "There is paucity of data to support their continuous use in home-based treatment of uncomplicated Plasmodium falciparum malaria in Cameroon.\u00a0Hence, this study aimed to assess the effectiveness and safety of AS-AQ versus AL for home-based treatment of uncomplicated P. falciparum malaria among children 6\u2013120\u00a0months in Yaound\u00e9, Cameroon.Many studies have reported high efficacy and safety of artesunate-arm, open-label, randomized, controlled trial comparing the equivalence of AS-AQ  and AL (control group) was carried out from May 2019 to April 2020 at two secondary hospitals in Yaound\u00e9. Participants were randomized to receive either AS-AQ or AL. After the first dose, antimalarial drugs were given at home, rather than under direct observation by a study staff. The conventional on-treatment and post-treatment laboratory and clinical evaluations were not done until day 3 of the full antimalarial treatment course. The evaluation of effectiveness was mainly based on per protocol polymerase chain reaction adjusted adequate clinical and parasitological response (PP PCR adjusted ACPR) on day 28 post-treatment. Safety was based on assessment of adverse events (AEs) and severe adverse events (SAEs) from day 1 to day 28.A two-AQ (n\u2009=\u2009114) and AL (n\u2009=\u2009128). The PP PCR adjusted day 28 cure rates were . Expected mild to moderate adverse events were reported in both arms . The most common adverse events included: transient changes of hematologic indices and fever.A total of 242 children were randomized to receive AS-AQ and AL are effective and safe for home management of malaria in Yaound\u00e9. The evidence from this study supports the parallel use of the two drugs in routine practice. However, the findings from this study do not describe the likely duration of antimalarial effectiveness in holoendemic areas where multiple courses of treatment might be required.This study demonstrated that ASTrial registration: This study is a randomized controlled trial and it was retrospectively registered on 23/09/2020 at ClinicalTrials.gov with registration number NCT04565184.The online version contains supplementary material available at 10.1186/s12879-022-07101-2. The median age was 58.5\u00a0months in the AS-AQ arm while in the AL arm it was 64.5\u00a0months.Out of 114 participants who received ASFurthermore, there were no statistically significant differences observed in all the categorical and quantitative variables evaluated in the two comparable independent treatment arms (P\u2009>\u20090.05). This permitted the comparison between the two drug groups at baseline Table .Table 1B-to-treat (ITT) analyses were 82.5%  for AS-AQ and 83.6%  for AL. The ITT cure rates adjusted by PCR were 83.2%  for AS-AQ and 85.6%  for AL. Conversely, the PCR unadjusted in vivo effectiveness obtained from per protocol (PP) analyses were 95.9%  for AS-AQ and 93.0%  for AL. The PCR adjusted ACPR derived from PP analyses were 96.9%  for AS-AQ and 95.5%  for AL. The ITT and PP (PCR uncorrected and corrected) cure rates were not significantly different across arms (P\u2009>\u20090.05) (Table The in vivo effectiveness on day 28 using unadjusted PCR intention-AQ and 5 for AL) when adjusted by PCR. Recrudescence accounted for only one case of LPF in the AL arm after PCR correction. There were four cases of reinfections (1 for AS-AQ arm and 3 for AL arm) registered during the period of follow-up (Table Treatment failures for the two drugs were due to early treatment failure (ETF), late clinical failure (LCF) and late parasitological failure (LPF) unadjusted and adjusted by PCR. There were 8 cases of treatment failures (3 for ASup Table .-AQ and 96.5% for AL. The cure rates did not differ when compared on day 7 and/or day 14 for either arm (P\u2009=\u20091.000)  rates on day 7 and day 14 were: 96.9% for AS0) Table .Table 3A\u2013Meier survival cure rates without PCR correction were 96.5%  for AS-AQ and 93.8%  for AL. The two groups were not statistically different when compared (P\u2009=\u20090.363)  for AS-AQ and 96.1%  for AL. Similarly, the two groups were not statistically different when compared (P\u2009=\u20090.605) (Table The Kaplan5) Table .-AQ arm and 99 (86.1%) in the AL arm of study participants did not have fever on day 3 (P\u2009=\u20090.162). The proportion of children who cleared their parasites on day 3 were 70 (69.3%) for AS-AQ and 85 (73.9%) for AL (P\u2009=\u20090.453) (Table It was observed that 93 (92.1%) of study participants in the AS-AQ [100/101 ] and AL [115/116 ] drug arms. The adherence rate was not statistically different between the two treatment groups [P\u2009=\u20091.000] (Table Good adherence to treatment guidelines were recorded in the AS-AQ and 5 (4.0%) for AL (P\u2009=\u20091.000). In the AS-AQ arm, the highest proportion of gametocyte  was recorded on day 3 and day 7 of follow-up while the lowest proportion was documented on day 0 . In contrast, in the AL arm the highest prevalence of gametocyte  was registered on day 0 while the lowest rate of  was recorded on day 3, day 7, and day 14. There was complete gametocyte clearance on day 21 and 28 in both drug arms. None of the study participants in the two drug arms had more than one episode of gametocyte carriage post-treatment.The total number of the study participants carrying gametocytes on day 0, 3, 7, 14, 21 and/or 28\u00a0by microscopy were 5 (4.5%) for AS-AQ arm and 99 (86.1%) participants in the AL arm had mild to moderate adverse events from day 1 to day 28 based on per protocol analysis. Most of the study participants in both drug arms had at least two or more adverse events due to multiple responses. Out of 44 adverse events reported, the most common were: leucocytosis , lymphocytosis percentage , granulocytopenia percentage , lymphocytosis number , thrombocytosis  and fever .A total of 83 (84.7%) participants in the AS-%  and low diastolic blood pressure . All the adverse events resolved post-treatment  [median (IQR)]  and platelet count-PLT (\u00d7\u2009109/L) [median (IQR)]   did not significantly change over time. However, 9 biological parameters had a significant change between day 0 and day 7. A total of 53 (58.9%) and 66 (63.5%) of participants had abnormal values in the AS-amodiaquine (AS-AQ) versus artemether-lumefantrine (AL) for a period of 28\u00a0days among children infected with uncomplicated P. falciparum malaria in Yaound\u00e9, Cameroon. The effectiveness of the two drugs was assessed under less optimal conditions to mimic what happens in the communities 17\u00a0years after their adoption and implementation in Cameroon. The approach adopted for this study is a modification of the standard procedure recommended by WHO for monitoring the efficacy of ACTs [The present study had as primary objective to determine the effectiveness and safety of artesunate of ACTs . The sta-ACPR value above the WHO benchmark of 90% for AS-AQ. Similar adjusted PCR cure rates for AS-AQ have been recorded before in Cameroon [-AQ is higher than the range of 60.0% to 91.7% obtained from studies conducted at the slope of Mount Cameroon [-ACPR of the present study is lower than range of 98.1%-100.0% registered in supervised clinical trials conducted in Cameroon [-ACPR is lower than the rates of 96.0\u2013100.0% reported in malaria endemic countries in sub-Saharan Africa [-analysis on the therapeutic efficacy of AL (Coartem\u00ae) for the treatment of P. falciparum malaria in Africa gave a pooled PCR adjusted 97.0% [-AQ was slightly above this benchmark (91.2%). The differences observed in the effectiveness rates of AS-AQ and AL may be due to heterogeneity of study settings, participants, malaria transmission patterns, sample sizes, and development of resistance against artemisinin partner drugs. Even though, mutations conferring resistance to the artemisinins have not been reported in Cameroon, there is a need to regularly monitor the effectiveness and efficacy of antimalarial drugs as recommended by the WHO.The evaluation of in vivo effectiveness using per protocol (PP) analysis gave an adjusted PCRCameroon  and BurkCameroon . The adjCameroon  and otheCameroon , 44\u201348. Cameroon , BurkinaCameroon , 15, GhaCameroon  and SeneCameroon . The assCameroon . The adjn Africa , 49\u201351, n Africa  and Soutn Africa . A systeed 97.0% . The adjed 97.0% , 48 and -AQ and AL. This is in agreement with the findings of the comparative effectiveness and efficacy studies in which the two drugs were used [-AQ versus AL disagrees with the results reported in Burkina Faso [-AQ and AL for the treatment of uncomplicated P. falciparum malaria.The comparison of in vivo effectiveness on day 28 did not show any significant difference between ASere used , 44, 48.ina Faso , 46. The-treatment. This observation tallies with those of other studies that showed an increase in the effectiveness [-AQ and AL after correction with PCR. The AL drug arm documented more cases of reinfections after 14\u00a0day of follow-ups when compared with the AS-AQ drug arm [Furthermore, the present study registered very few cases of reinfection 14\u00a0days posttiveness , 48, 54 tiveness \u201315 of ASdrug arm , 48, 54.drug arm , 45, 54.drug arm . The difdrug arm , 56, 57,drug arm , 59, maldrug arm , immunitdrug arm , pharmacdrug arm , 63 and drug arm .-up days was found to be higher in the AS-AQ group when compared to the AL group. These observations are in harmony with those reported in Senegal/Ivory coast [-analysis from individual patient data conducted by the Worldwide Antimalarial Resistance Network (WWARN) Gametocyte Study Group confirmed that AL is more effective than the AS-AQ fixed dose combination in preventing gametocyte carriage shortly after treatment [-artemisinin partner drugs and the timing of artemisinin dosing are important determinants responsible for post-treatment gametocyte dynamics [In both drug arms, fever and parasite disappeared gradually by day 3. Rapid fever and parasite clearance rates have been reported before in Senegal/Ivory Coast , 66, Camry coast , 66 and ry coast . A meta-reatment . The autdynamics . Informa-up. There was no severe adverse event registered among the study participants. Previous pharmacovigilance studies on AS-AQ and AL have mostly reported the presence of common adverse events associated with the gastrointestinal tract and neuropsychiatric systems [-reporting by the patients, parents or guardians. A major setback of this method is recall bias. Some parents or guardians may actually have over-reported perceived adverse events because of anxiety about the possibility of adverse events.In addition, expected mild to moderated adverse events were documented in the two drug arms that resolved during follow systems , 18, 19.-AQ and AL arms. It was also shown that AS-AQ and AL drugs had varying effects on alanine aminotransferase (ALAT) and creatinine (CREA) levels on day 0 and day 7. These results corroborate with those realized in Senegal and Ivory Coast [-up is an indication that AS-AQ and AL are well tolerated and safe.The study drugs had a statistical significant effect on 9 biological parameters on day 0 and day 7 in the ASry Coast . These ory Coast . The res-AQ and AL in Cameroon, which should provide important support for policy-making. (2) Both regimens were studied concurrently in the town of Yaound\u00e9 in Cameroon seventeen years after policy implementation. The existence of data from previous efficacy trials of the same regimens in the same region was also important. (3) The fact that the study was performed in a holoendemic region in Cameroon is actually important, because the national malaria control program needs data from diverse transmission settings in order to devise rational policy. (4) The design was intended to \"mimic the routine standard of care\" by omitting clinical supervision of treatment, clinical evaluation, and laboratory testing after day zero and before day three. This is actually a strength given the study team's emphasis on effectiveness.The major strengths of this study are: (1) This work filled significant gaps in knowledge about the performance of AS-urban settings of Yaound\u00e9. These areas are characterized with holoendemic malaria transmission. This may not present the reality of what happens in the other malaria transmission settings in Cameroon and globally. (3) The estimated sample size may have been inadequate to establish the equivalence of the two drugs. (4) Families who were willing to be adherent to medication and clinical/laboratory evaluation schedules may not have been representative of the entire population from which the subjects were drawn. Hence, study results may not be generalizable.The major limitations of this study are: (1) There was no direct clinical observation of antimalarial drug dosing (after dose 1), and standard clinical and laboratory evaluations were not conducted after day zero and before day 3. (2) The study was conducted in urban and peri-AQ and AL are effective and well tolerated for home management of uncomplicated P. falciparum malaria among children in Yaound\u00e9, Cameroon. The PP cure rates adjusted by PCR and safety parameters for the two drugs were not statistically different and are said to be substantively the same when compared. The two regimens seem to have retained their effectiveness and safety profiles for over 17\u00a0years, in spite of frequent administration likely driven by the intensity of malaria transmission. The evidence from this study supports the government\u2019s policy on the parallel use of AS-AQ and AL in routine practice in the Southern regions of Cameroon. However, the findings from this study do not describe the likely duration of antimalarial effectiveness in holoendemic areas where multiple courses of treatment might be required. It was also reported that the lower limit of confidence interval was slightly below or above the WHO threshold of 90% for AL and AS-AQ respectively. Thus, there is a need to continuously monitor the effectiveness and safety profile of AS-AQ and AL in Cameroon.This study demonstrated that ASAdditional file 1: Study treatmentAdditional file 2: Reference ranges of heart rate, systolic blood pressure, diastolic blood pressure, respiratory rate, hematology parameters and biochemistry parametersAdditional file 3: Kaplan\u2013Meier survival curve for the effectiveness of AS-AQ versus AL (PP without and with PCR correction)Additional file 4: Evolution of biological parameters among study participants treated with AS-AQ and AL (Day 0 and Day 7) (PP)"}
+{"text": "Plasmodium vivax malaria elimination requires radical cure with chloroquine/primaquine. However, primaquine causes hemolysis in glucose-6-phosphate dehydrogenase-deficient (G6PDd) individuals. Between February 2016 and July 2017 in Odisha State, India, a prospective, observational, active pharmacovigilance study assessed the hematologic safety of directly observed 25 mg/kg chloroquine over 3 days plus primaquine 0.25 mg/kg/day for 14 days in 100 P. vivax patients (\u2265 1 year old) with hemoglobin (Hb) \u2265 7 g/dL. Pretreatment G6PDd screening was not done, but patients were advised on hemolysis signs and symptoms using a visual aid. For evaluable patients, the mean absolute change in Hb between day 0 and day 7 was \u22120.62 g/dL  for males (N = 53) versus \u22120.24 g/dL  for females . Hemoglobin declines \u2265 3 g/dL occurred in 5/99 (5.1%) patients ; none had concurrent clinical symptoms of hemolysis. Based on G6PD qualitative testing after study completion, three had a G6PD-normal phenotype, one female was confirmed by genotyping as G6PDd heterozygous, and one male had an unknown phenotype. A G6PDd prevalence survey was conducted between August 2017 and March 2018 in the same region using qualitative G6PD testing, confirmed by genotyping. G6PDd prevalence was 12.0% (14/117) in tribal versus 3.1% (16/509) in nontribal populations, with G6PD Orissa identified in 29/30 (96.7%) of G6PDd samples. Following chloroquine/primaquine, notable Hb declines were observed in this population that were not recognized by patients based on clinical signs and symptoms. Hypnozoites are carried asymptomatically, allowing the parasite to persist undetected and sheltered from malaria control efforts.P. vivax malaria relapse requires 8-aminoquinoline administration.\u2013\u2013,,G6PDd is an X-linked enzymopathy prevalent in populations currently or historically exposed to malaria.The G6PD enzyme is highly polymorphic, with deficient variants associated with different degrees of reduced enzyme stability.P. vivax cases are treated with chloroquine (25 mg/kg over 3 days) and supervised primaquine (0.25 mg/kg/day for 14 days), except for infants under 1 year old, pregnant and lactating women, and individuals known to be G6PDd.The India National Malaria Treatment Guidelines require that confirmed \u2013,P. falciparum is predominant in Odisha, around 10% of cases are P. vivax and the proportion of P. vivax tends to increase in areas approaching malaria elimination.\u2013P. vivax malaria radical cure is complicated by heterogeneity in G6PDd prevalence across the state, being on average around 3%,,,\u2013,The State of Odisha is the most malaria-endemic state in India, with 46 million people at risk of infection.,To improve access to malaria diagnosis and treatment in Odisha, an operational research initiative, the Comprehensive Case Management Project (CCMP), was launched in 2013.P. vivax malaria patients in Odisha by measuring hemoglobin (Hb) levels. Pretreatment G6PD testing is not generally available in the region, and point-of-care G6PD kits are not deployed. To reflect usual practice, patients in this study were not screened for G6PDd before primaquine administration. However, to analyze primaquine hematologic safety, qualitative G6PD testing was conducted following study completion. Additionally, G6PDd prevalence in the region was surveyed. The aim was to assess the safety of current practice, that is, using clinical signs and symptoms to identify hemolysis following chloroquine/primaquine in relation to the prevalence of G6PDd in the region. The results will inform policy on implementing well-tolerated and effective universal P. vivax radical cure with chloroquine/primaquine to support malaria elimination.This study evaluated the hematologic safety of directly observed 14-day primaquine in A prospective observational active pharmacovigilance study was conducted between February 2016 and July 2017 in Odisha State, India, at five primary health centers: Paiksahi and Thakurgarh (Angul district), Rasol and Hindol , and Saranggarh  within the CCMP . Following completion of the observational study, a G6PD prevalence survey (phenotype and genotype) was carried out in the same areas between August 2017 and March 2018. Both studies were conducted according to the principles of the Declaration of Helsinki and Good Clinical Practice and conformed to local and international laws on the study of human subjects. The studies were approved by the Institutional Ethics Committee of National Institute of Malaria Research. Before enrollment to either component, all participants or their parents/guardians completed informed written consent, and school-aged children also had to provide their assent.P. vivax malaria based on the results of bivalent rapid diagnostic tests (RDTs) used according to manufacturer\u2019s instructions, in line with national guidelines. Deployed RDTs were SD Bioline Malaria Ag Pf/Pv , EzDx\u2122 Malaria Pv/Pf Ag , and Onsite Malaria Pf/Pv Ag . Eligible patients were at least 1 year old with RDT-confirmed P. vivax malaria, Hb of at least 7 g/dL assessed using HemoCue\u00ae point-of-care testing , and willing and able to participate in the study. The ASHAs referred patients to study sub-centers for Hb assessments and therapy initiation.ASHAs routinely diagnose Patients were treated in accordance with national antimalarial drug policy, dosed by body weight with 25 mg/kg chloroquine over 3 days plus primaquine 0.25 mg/kg per day for 14 days. All treatment was directly observed by ASHAs or healthcare workers. All patients were advised of the signs and symptoms of hemolysis and the need to seek medical advice in alignment with CCMP protocols.Following enrollment, all patients underwent a physical examination and had a medical history taken. At the study sub-center, before treatment initiation, duplicate baseline Giemsa-stained blood slides were prepared for confirmation of malaria diagnosis by two trained microscopists using standard methods.P. vivax case who completed treatment. Patients were asked to report symptoms of acute hemolytic anemia, that is, excessive tiredness, dizziness, breathlessness, dark-colored urine, or pale skin. ASHAs used a visual instruction tool to educate and question patients regarding any notable symptoms . If any symptoms were identified, the patient was referred to the nearest primary health center for assessment and management.Patients were followed up daily during their 14-day primaquine course by ASHAs. ASHAs were compensated (Rs 150) by the NVBDCP/National Health Mission for each \u00ae. Anemia was defined using World Health Organization criteria.Hemoglobin levels were measured at baseline (day 0) and days 3, 7, 14, 28, and 42 using HemoCue,Following study completion, G6PD qualitative testing was conducted to inform the analysis of Hb levels. Approximately 3 mL blood was collected in an ethylenediaminetetraacetic acid (EDTA) Vacutainer from each subject and G6PD phenotyping was performed within 24 hours of sample collection using the dichlorophenol indophenol (DCIP) dye discoloration test as per published methods.The primary study endpoint was the absolute and proportionate fall in Hb from day 0 to day 7, and Hb recovery from day 28 to day 42. The number and proportion of patients who had adverse events, required hospitalization, underwent blood transfusion, or had treatment discontinued, were also evaluated. Data were presented using descriptive statistics. Statistical comparisons used the Mann\u2013Whitney test. Statistical analysis was performed in GraphPad Prism (version 9.0).P. vivax cases of 90%, with 5% absolute precision and 95% confidence intervals (CIs), 138 patients with P. vivax malaria were needed to estimate the prevalence of Hb declines.Assuming a prevalence of fall in Hb among A cluster sampling method was followed with villages in the catchment areas of the pharmacovigilance study chosen at random. Study enrollment was offered to all individuals in the sampled villages. All subjects who provided written informed consent were included in the study. Assuming an average prevalence of G6PDd of 10%, relative precision of 20%, and desired CI  of 90%, in a population of 300,000, a sample size of 604 subjects was required. Statistical comparisons used the Mann\u2013Whitney test .\u2013A 3 mL-blood sample was collected in an EDTA Vacutainer from each subject for G6PD phenotyping using the DCIP test as described above.N = 42, Paiksahi N = 18), followed by Dhenkanal  and Kandhamal (Saranggarh N = 16). The population from Kandhamal differed from the other regions, comprising all males over 14 years old. Two patients had missing day 7 values . One patient was lost to follow up with no post-baseline evaluations. Overall, 11 patients were lost to follow up , with 89% (89/100) of patients having baseline, day 28 and day 42 Hb values. Although efficacy was not formally evaluated in this study, there were no cases of recurrent malaria infection reported over the 42-day follow up.Of the 100 patients enrolled, 54% were over 14 years old and 61% had baseline mild/moderate anemia  of male patients were G6PD normal, G6PD enzyme activity status was unknown for the remaining 17 patients with no known G6PDd patients. In females, 87.0% (40/46) had normal G6PD enzyme activity based on qualitative testing, G6DP status was unknown for four patients, and there were two G6PDd female patients, both of which were confirmed by genotyping as heterozygous for G6PD Orissa.N = 53) versus females  (Table P = 0.093) (Table P = 0.0097), as was the mean percentage decline in Hb (P = 0.018) (Table P > 0.47)  was \u22121.4 g/dL  and in females (N = 45) was \u22120.97 g/dL  (P = 0.062)   of males and 54.8% (23/42) of females, and by day 42, 70.2% (33/47) of males and 59.5% (25/42) of females had Hb levels equivalent or higher than at baseline . At day 42, 31.5% (28/89) of patients had mild anemia and 16.9% (15/89) had moderate anemia, but none had severe anemia.N = 89). In these patients, the Hb nadir occurred most commonly at day 3 (N = 33) with a mean Hb decline of \u22121.3 g/dL . Hb declines were most pronounced on day 14 (N = 20) with a mean Hb decline of \u22121.8 g/dL  (P = 0.5) . The nadir for Hb declines \u2265 3 g/dL only occurred on days 7 (N = 1) and 14 (N = 4).A post hoc analysis considered only those patients with a post-baseline Hb decline (An Hb decrease of \u2265 10% was noted for 53.7% 29/54) of males and 44.4% (20/45) of females  of patients. Analysis of the relationship to G6PD phenotype determined at study completion showed that these patients included one G6PDd female, 13 patients with G6PD-normal phenotype , and five patients with unknown G6PD status  .There were six patients with an absolute Hb value < 7 g/dL and/or Hb decreases \u2265 3 g/dL. Two patients had Hb values < 7 g/L, both were from Angul, one 4-year-old male  had a baseline Hb of value of 7.6 g/Hb, declining to 6.7 g/dL on day 3 and 6.8 g/dL on day 14, with recovery to 11.4 g/dL on day 28. One 37-year-old male (G6PD phenotype unknown) had a baseline Hb value of 10.2 g/dL, declining to 7.9 g/dL on day 7 and of 6.8 g/dL on day 14, with recovery to 10.3 g/dL on day 42. For the five patients with Hb decreases \u2265 3 g/dL, three were tribal males  and two were nontribal females . This G6PDd heterozygous patient had a baseline Hb level of 12.0 g/dL, a nadir of 9.0 g/dL on day 14, and recovery to 12.3 g/dL by day 42 . An additional G6PDd heterozygous female had a maximum Hb decline of \u22120.3 g/dL from 10.6 g/dL at baseline to 10.3 g/dL on day 7, with recovery to 11.0 g/dL by day 14.Treatment was generally well tolerated Figure . The majFor the 94 patients followed up on day 7, eight adverse events occurred in four patients that could potentially indicate hemolytic anemia Figure . HoweverThe G6PDd prevalence study included 626 participants. Based on qualitative testing (DCIP), 5.1%  of participants were G6PDd or G6PD intermediate (Table N = 15) versus 1.97  IU/gHb in females (N = 15) (P = 0.0075) overall  and 3.0  IU/gHb in heterozygous females (N = 11)  versus nontribal persons (N = 16), either between males (P = 0.39) or females (P = 0.16)  G6PD enzyme level was 0.92 0.48, 1.29) IU/gHb in males  in Hb levels, which recover after day 3, are caused by hemodilution on rehydration. Consistent with previous studies,The disparity between clinical and laboratory findings was surprising as patients had been instructed to report all potentially relevant symptoms of hemolysis, even if they were not worsening, and were followed up daily by ASHAs. As 61.0% (61/100) of patients had mild-to-moderate anemia at study entry, it is possible they were adapted to living with low Hb levels and so did not manifest clinical signs of hemolysis. Also, in this area of high malaria endemicity, patients may expect to feel poorly, with common symptoms not appearing noteworthy, and patients may have been less diligent in their observations once their fever resolved at day 2 or 3. Dark-colored urine was not reported by any patient, and this might reflect the available toilet facilities, which make such observations difficult. These findings suggest that our current practice would not be expected to identify cases that might require medical intervention. Moreover, because declines \u2265 3 g/dL occurred at day 7 and day 14 following treatment initiation, patients may be distant from health services.,,Of the five patients with Hb decreases \u2265 3 g/dL, one was G6PDd, three had a G6PD-normal phenotype, and one had an unknown G6PD status. Thus, primaquine-induced hemolysis was suspected for at least one patient with a \u2265 3 g/dL Hb decline, and considered as a possible causality for the patient whose G6PD status was unknown. However, we did not expect to see notable Hb declines in three patients with a G6PD-normal phenotype. Because G6PD genotyping was only performed on samples identified as G6PDd following qualitative testing, we cannot exclude that these phenotypically \u201cG6PD-normal\u201d patients were false-negative results. For example, in cases of acute hemolysis associated with malaria, the erythrocyte population is enriched for young erythrocytes and reticulocytes, which have near-normal G6PD enzyme levels.,,The general health of tribal populations is poorer and socioeconomic status lower than nontribal populations.P. vivax relapses have been reported from areas where P. vivax has a short-latency relapse periodicity.,\u2013After 42 days of follow up, recovery to baseline Hb levels had not been achieved in 29.8% of males (14/47) and 40.5% of females (17/42). However, the hematologic consequences of not administering primaquine could not be evaluated in this study. Longer term hematologic benefits and wider health benefits from primaquine therapy versus repeated ,The G6PDd survey used a qualitative test, with G6PDd samples confirmed using G6PD quantitative testing and genotyping. The overall G6PDd prevalence was 4.8% (30/626). This is higher than identified in the Kalahandi and Rayagada districts of Odisha state (3.0% [59/1981]),,Consistent with previous studies, G6PD Orissa was the most prevalent G6PDd variant in Odisha, detected in 29/30 of the G6PDd samples confirmed by genotyping.,,As expected from previous reports using qualitative G6PD testing, the proportion of G6PDd males (6.2%) was higher than that of females (3.9%). Because heterozygous females have a mixture of G6PD-normal and G6PDd erythrocytes, only a proportion of G6PDd heterozygous females will be detected using phenotypic G6PD testing.P. vivax relapse is well established.,,,There were some key limitations across the two studies. Most notably, because this study was conducted in an operational context to evaluate current practice, there was no chloroquine-only control arm. The efficacy of primaquine versus chloroquine alone at preventing P. vivax patients in Odisha following treatment with chloroquine/primaquine. The visual instruction tool plus frequent and specific questioning by ASHAs did not identify patients with Hb declines that might require intervention. Qualitative G6PD testing (DCIP) conducted after study completion did not reliably predict patients\u2019 risk of Hb decline. The declines in Hb observed could be caused by primaquine/chloroquine administration, malaria, or other factors, and they may be multi-factorial. The survey confirmed that G6PDd prevalence was higher in the tribal versus nontribal population.In summary, notable declines in Hb were observed in some It is clear that radical cure is a key tool for malaria elimination in Odisha, but clinical observations could not be relied upon to identify patients with physiologically relevant Hb declines. Given the high prevalence of G6PDd alleles in Odisha, particularly in tribal populations, rather than attempting to detect primaquine-induced hemolysis, it would be preferable to avoid exposing those at risk by wider deployment of pretreatment G6PD testing. However, as the DCIP qualitative testing method was inadequate to predict individuals at risk of Hb declines, the utility of advanced point-of-care quantitative diagnostics should be evaluated.Supplemental materials"}
+{"text": "Malaria is a significant cause of morbidity and mortality throughout the world and particularly sub-Saharan Africa. The World Health Organization and many national bodies, including Burundi, recommend artemisinin-based therapy as first-line treatment for uncomplicated and severe malaria. Implementing this recommendation requires healthcare professionals' acceptance of this treatment as the optimal choice.A survey was conducted among Burundian healthcare professionals from June to September 2017 to assess prescribing preferences regarding artemisinins versus quinine for treating malaria. Healthcare professionals were surveyed from 32 health facilities in 10 provinces. Respondents included both physicians and nurses who provided responses about their antimalarial treatment preferences for a variety of clinical scenarios. Comparisons among healthcare professionals, their level of training, work setting, and length of work experience were examined using a series of stratified analyses, where the Pearson Chi-square statistic and odds ratios with 95% confidence intervals were calculated.Respondents included 101 doctors and 196 nurses. Seventy-nine percent of respondents worked in hospitals, while 58% had more than 5 years of work experience. Although 94% of respondents correctly identified artemisinin-based treatment as first-line therapy according to the national protocol, 24-40% of respondents preferred the use of quinine in various hypothetical clinical scenarios. Overall, nurses were at greater odds of preferring quinine versus artemisinins compared with physicians. Availability of artemisinins was associated positively with artemisinin preference. These results did not vary by duration of work experience.Though knowledge of artemisinin-based therapy was recognised by the majority of respondents as the recommended antimalarial treatment, a high proportion of Burundian healthcare professionals, especially nurses, prefer using oral and IV quinine in a number of clinical scenarios. These findings identify a significant barrier to the satisfactory implementation of a life-saving treatment in accordance with national and international recommendations. Most countries in Africa have been recommending artemisinin combination therapy (ACT) for first-line treatment of uncomplicated malaria since at least 2005, with Burundi adopting this recommendation in 2003.3 More recently, the South East Asian Quinine Artesunate Malaria Trial (SEAQUAMAT)for adults and the Artesunate versus quinine in the treatment of severe falciparum malaria in African children (AQUAMAT) trialfor children demonstrated decreased mortality for intravenous artesunate versus quinine in the treatment of severe falciparum malaria, a finding further supported by a 2012 Cochrane review.6 The World Health Organization (WHO) Guideline for the Treatment of Malaria has adopted artemisinin-based treatment for both categories since 2010.7 An African logistical study showed that artesunate is better, cheaper, and easier to administer than its longstanding forerunner quinine.8Malaria is a significant cause of morbidity and mortality throughout the world, and particularly in sub-Saharan Africa.lity see .2 One of1 Though progress continues to be made, obstacles to complete implementation merit analysis. Successful implementation depends on a multiplicity of factors, including availability and acceptance by healthcare professionals (HCP).9Despite the enormous benefit of this evidence-based recommendation, the WHO World Malaria Report 2019 mentions that only 48% of febrile children in the public sector of sub-Saharan Africa who sought care were given an antimalarial drug, and that only 80% of those given an antimalarial were given an ACT.3 In terms of availability, a Burundian study in 2011 showed that ASAQ was present in 88% of public sector venues.10 More recently, a 2019 national guideline changed the first-line ACT to artemether-lumefantrine (A-L), which has recently become widely available in Burundi. Second-line therapy is dihydroartemisinin-piperaquine, with oral quinine moving from second-line therapy to being used only in cases where there are contraindications to ACT  was officially replaced by artesunate-amodiaquine (AS-AQ) in november 2003, though quinine use is still widespread.s to ACT .1112 In two Cameroonian studies regarding implementation of AS-AQ, HCPs' hesitancy was due to adverse effects of amodiaquine, lack of availability, and doubt of efficacy.14 Two Burkina Faso studies also showed poor compliance to ACTs for uncomplicated malaria, despite adequate knowledge of national protocol recommendations, citing as reasons adverse effects and availability.16 A Kenyan study showed quinine prescription persisting particularly in children over 5 years and adult populations.17Acceptability and preference of these antimalarial drugs by health-care professionals and the population has been evaluated in a variety of heterogeneous studies across sub-Saharan Africa. A Rwandan study of non-artemisinin antimalarials found HCP non-compliance to be associated with ignorance of the protocol, doubt of efficacy, and fear of adverse effects.18 Preference for treating severe malaria with IV quinine was elsewhere seen in a 2016 Congolese study where less than 2% received IV artesunate.19 A 2015 Sudanese study also found overuse of IV quinine instead of oral ACTs for patients that had no criteria for severe malaria.20In regard to severe malaria, since 2012, Burundi's national protocol has recommended IV artesunate as first-line therapy with IV quinine as a second-line alternative when IV artesunate is unavailable. We were not able to find current availability data and IV quinine use remains widespread.In light of the paucity of studies about HCP preferences towards any artemisinin-based therapy in our region of Africa, and in particular of studies that evaluate such preferences for injectable artesunate for severe malaria, our study sought to address these topics in the context of the country of Burundi. If HCP preferences diverge from national and international recommendations, identification of this divergence could highlight an important gap in the pathway from correct knowledge to correct implementation.21 Seventeen hospitals were surveyed . Fifteen nursing-staffed health centers were surveyed .This cross-sectional study was performed between June and September 2017, roughly 5 years after the 2012 national guideline instituted injectable artesunate for severe malaria. In an effort to approach greater generalizability of our findings across Burundi, we elected to perform convenience sampling of facilities in ten of the eighteen provinces of Burundi, as well as convenience sampling of HCPs at those facilities. Five of the provinces  are in the hyper-endemic zone for malaria, and the other five provinces  are in the hypo-endemic zone, which was significantly affected by the malaria epidemic announced in March 2017.Assuming an alpha = 0.5 and a baseline 0.5 acceptance rate given a lack of previous study results in the region, we estimated a sufficient sample size of 98 physicians and 98 nurses to detect a change of 0.17 between the two groups. Given that physicians and nurses form the backbone of malaria diagnosis and treatment in the Burundian health system, this study uses the term healthcare professionals to refer to these two groups, though the general definition includes those trained to work in a broader range of health fields. All physicians and nurses present at a visit by the principal investigator to the above facilities were given the opportunity to complete the survey, including both consultant-level and generalist physicians, as well as nurses of all qualifying professional levels. A convenience sample of 297 respondents was obtained, including 101 physicians  and 196 nurses, 43 of which were registered nurses (level A0) and 153 of which were enrolled nurses (level A2 or A3).A written questionnaire designed by the principal investigator was administered in French directly by the principal investigator, with the possibility of asking clarifying questions in Kirundi if desired by the participant.Respondents first identified their place of work  followed by their profession . They also self-reported their professional experience as greater than or less than 5 years. Each respondent then reported the availability of intravenous (IV) artesunate, IV quinine, oral AS-AQ and oral quinine in their place of work. Next, respondents answered 6 questions about their preferred anti-malarial to prescribe in various scenarios. The 3 categorical response options were artesunate (or AS-AQ in the case of uncomplicated disease), quinine , and other choice . Respondents were then asked which antimalarial they would prefer to receive if they or a close member of their family were diagnosed with uncomplicated or severe malaria. These scenarios were followed by a question about which antimalarial causes more side effects, with the same categorical choices. Finally, their knowledge of the current Burundian recommendation was assessed by asking them to identify the first-line antimalarial according to the national protocol.p<0.05), odds ratios (OR) and 95% confidence intervals (CI).All survey responses were transferred into a data spreadsheet with respondents coded by number. The survey responses were then uploaded from the data spreadsheet into SPSS  for data analysis. All responses were categorical. Through a series of stratified analyses, specifically stratified by healthcare professionals (i.e. physicians and nurses), years of healthcare service, and availability of artesunate and ASAQ at the respondent's facility, we examined associations with the scenario outcomes. Measures of association for each of these stratified analyses used the Pearson chi-square test  Institutional Review Board (IRB #18-167). All respondents provided consent by completing the survey instrument. No personal identifiers were collected.Of the total 297 HCPs surveyed, 101 (34%) were physicians and 196 (66%) were nurses. Two hundred thirty five (79%) respondents worked in hospitals, and 62 (21%) in nurse-run health centers. One hundred twenty four (42%) respondents had less than 5 years' healthcare work experience .All healthcare professionals indicated that oral quinine was available at their place of work, with 281 (95%) respondents reporting the availability of IV quinine . ArtemisP<.01). Similar results were found for pediatric cases  as well as treatment choice if the respondent or their family member had malaria .Two hundred seventy nine (94%) HCPs surveyed correctly identified artemisinin-based therapy as the recommended first-line therapy according to the Burundian national protocol. However, when presented with a variety of clinical scenarios, many of these HCPs still preferred quinine to artemisinin-based therapy  respondents who preferred quinine for the first trimester of pregnancy  versus 103 (35%) for the second and third trimesters . 198 (67%) respondents believed that quinine causes more side effects than AS-AQ/Artesunate.Stratified subgroup analyses showed significant differences between doctors and nurses in all three scenarios involving severe malaria, with physicians at greater odds than nurses of preferring intravenous artesunate  than intravenous quinine for adults , for children , and for self or a family member . Stratified subgroup analyses of HCPs with more or less than five years' work experience showed no statistically significant differences in any antimalarial preferences .However, there were no significant differences between physicians and nurses for the three scenarios involving simple malaria, which compared oral AS-AQ and oral quinine (P=.003). A similar association was found when comparing the availability of IV artesunate and the health-care professionals' preference for using this medicine (P=.007).When treatment preferences were analyzed based on availability of AS-AQ in the respondent's place of work, availability of AS-AQ was associated with respondents' preference for using it  respondents' preferences were in agreement with the national protocol in the one scenario that recommends quinine therapy (first trimester of pregnancy). However, later in pregnancy, when artemisinin-based therapy is recommended, only 188 (63%) of respondents' preferences agreed with the protocol, a level of agreement very similar to the other non-pregnancy scenarios.22In our study, this gap between knowledge and practical preference was significantly more pronounced in the nursing workforce compared to physicians. This is especially consequential since nurses are prescribing these antimalarials throughout the country's health centers. Years of health care experience does not appear to modify this distinction. Both of these findings were contrary to a Tanzanian study which found longer work experience and non-doctor prescribers associated with increased ACT usage.What are the root causes of this problem of acceptance for a medicine whose efficacy has an excellent evidence base and whose role at the front of the national protocol is nearly universally known? To be sure, further studies are needed to explore this question more directly.One might posit that the amodiaquine portion of AS-AQ is a primary reason for hesitation in artemisinin-based therapy. This is supported by the HCPs who voluntarily wrote in A-L as \u201cother\u201d when asked their preferences. However, this study clearly shows that the significant differences between nurses and doctors resides in the treatment of severe malaria, where amodiaquine is not implicated. Thus, this problem could persist even with the replacement of AS-AQ by A-L in the 2019 national protocol.23 Whereas quinine is an ancient medicine with which the population is very familiar, AS-AQ and artesunate are less known at certain facilities. In facilities where these antimalarial drugs are available (and thus known), preference for them is increased. In a parallel situation, a 2014 Ghanian study showed IV quinine having the same problem as artesunate currently has when HCPs persisted in prescribing chloroquine for severe malaria though it had been replaced by quinine in the national protocol, suggesting a simple persistence in old habits.24 Perhaps HCPs are more reluctant to use an unfamiliar medicine especially with sicker patients.As shown above, the statistically significant correlation between both AS-AQ and artesunate availability and treatment preference for these recommended medicines suggests that unfamiliarity may play a role in HCPs' hesitation, which may decrease over time as seen elsewhere.We see here a strong suggestion that consistent knowledge of a national protocol may be accompanied by HCP hesitancy about implementing it. The issue of HCP's acceptance of artemisinin-based therapy needs to be further addressed in order to assure solid implementation of this life-saving treatment for the country's number one cause of morbidity and mortality.Though this study raises an important question, there are several limitations, including an inability to confirm the respondents' reported antimalarial availability, an inability to analyze the cluster effect of different sites, and a lack of a validated survey instrument. Lastly, the convenience sampling and lack of randomization of this study was not designed to be nationally representative, and the varying numbers of respondents from different health facilities may introduce selection bias. 25 Additionally, it would be helpful to examine how these HCPs' preferences influence their actual practice, since practice is influenced by more factors than just preference.26Regarding future avenues of study, a more expanded, randomized and nationally representative sample would be warranted, and this could also investigate to see if there are regional differences . It is also necessary to explore HCPs' reasons for preferring another antimalarial drug to artemisinin-based therapy. In regards to artemisinin implementation, HCPs in other studies have cited doubts about efficacy, concerns about cost, and confusion in protocol implementation as reasons for non-compliance, though this reasons seems especially unexplored for IV artesunate.On the level of practical action, several ways forward are suggested by this data. Simply continuing to increase availability of oral ACTs and IV artesunate may be useful in augmenting exposure and acceptance of these antimalarials. Additional educational interventions may be useful, especially for the nursing workforce. However, this study suggests that knowing the recommendation for artemisinin-based therapy is not sufficient. Thus, educational interventions should include the rationale for the preference of artemisins, both in terms of increased efficacy and less side effects, in order to maximize acceptance and thus implementation. Lastly, the inclusion of HCP's preferences for such important antimalarial drugs could be useful to monitor by national malaria programs in order to continually assess further intervention needs.5It is unclear whether this preference of many HCPs for IV quinine in severe malaria is equally present in other malaria-endemic countries, but this may also warrant further study. Given the magnitude of the mortality benefit of IV artesunate over IV quinine for severe malaria, this could represent an urgent, high-impact area of research and intervention, since rapid improvement of protocol implementation could save numerous lives, as opposed to gradually waiting on the HCP population to become habituated to a newer therapy.This study is the first in Burundi to evaluate HCP preferences towards artemisinin-based therapy versus quinine, which has revealed an important gap between knowledge of a therapeutic recommendation and practical preference, suggesting a problem of acceptance by health care professionals, which must be addressed for successful implementation of current malaria recommendations. In particular, the persistent preference for IV quinine over artesunate in severe malaria by many HCPs requires further investigation. This particular question is under-explored in the medical literature, and if present elsewhere, necessitates urgent research and intervention in order to efficiently apply this important life-saving advance in treatment. More comprehensive education about the advantages of artemisinin-based therapy and monitoring of HCP preferences by national malaria programs could also mitigate the effect of this implementation challenge."}
+{"text": "Background: Typhoid and paratyphoid fever (enteric fever) is a common cause of non-specific febrile infection in adults and children presenting to health care facilities in low resource settings such as the South Asia. \u00a0A 7-day course of a single oral antimicrobial such as ciprofloxacin, cefixime, or azithromycin is commonly used for its treatment. Increasing antimicrobial resistance threatens the effectiveness of these treatment choices. We hypothesize that combined treatment with azithromycin (active mainly intracellularly) and cefixime (active mainly extracellularly) will be a better option for the treatment of clinically suspected and culture-confirmed typhoid fever in South Asia.Methods: This is a phase IV, international multi-center, multi-country, comparative participant-and observer-blind, 1:1 randomised clinical trial. Patients with suspected uncomplicated typhoid fever will be randomized to one of the two interventions: Arm A: azithromycin 20mg/kg/day oral dose once daily (maximum 1gm/day) and cefixime 20mg/kg/day oral dose in two divided doses (maximum 400mg bd) for 7 days, Arm B: azithromycin 20mg/kg/day oral dose once daily (max 1gm/day) for 7 days AND cefixime-matched placebo for 7 days. We will recruit 1500 patients across sites in Bangladesh, India, Nepal, and Pakistan. We will assess whether treatment outcomes are better with the combination after one week of treatment and at one- and three-months follow-up.Discussion: Combined treatment may limit the emergence of resistance if one of the components is active against resistant sub-populations not covered by the other antimicrobial activity. If the combined treatment is better than the single antimicrobial treatment, this will be an important result for patients across South Asia and other typhoid endemic areas.Clinicaltrials.gov registration: NCT04349826 (16/04/2020) Salmonella enterica serovar Typhi) and paratyphoid fever , collectively referred to as typhoid fever in this protocol, are common causes of non-specific febrile infection in adults and children presenting to health care facilities in low resource settings such as the South Asia region. South Asia has been described as the largest hub for typhoid fever in the world and antimicrobial resistance has become a critical issue. In the last 20 years, treatment of typhoid fever with a 7-day course of a single oral antimicrobial, such as ciprofloxacin, cefixime or azithromycin, given in an out-patient setting has led to patient recovery in 4 to 6 days without the need for expensive hospitalization. Increasing antimicrobial resistance in Asia and sub-Saharan Africa threatens the effectiveness of these treatments and increases the risk of prolonged illness and severe disease.Typhoid  with cefixime and azithromycin in typhoid fever leave many clinicians unsure about the optimum antimicrobial treatment for typhoid fever in this region.The fluoroquinolone class of antimicrobials have been a common choice to treat typhoid fever across South Asia during the last two decades. Now low and high-level resistance is so widespread that the entire class is no longer a reliable treatment choice. The reported fever clearance times with azithromycin treatment (10\u201320 mg/kg/day for 5\u20137 days) is usually 4\u20136 days and clinical failure rates range from 0% to 18% with a microbiological failure rate from 0% to 3.2%. A number of the studies with azithromycin have demonstrated a delayed microbiological clearance, indicated by positive blood culture during the treatment course. Azithromycin activity is predominantly intracellular with studies from Vietnam indicating that up to one third ofS. Typhi may be found in the extracellular compartment. The low levels of azithromycin in plasma may be insufficient to ensure adequate killing activity. In contrast, the extended-spectrum cephalosporin cefixime is predominantly active in the extracellular compartment althoughin-vitro evidence also suggests some intracellular activity. The relative lack of intracellular activity by cefixime may be one reason for the variable treatment results and prolonged fever clearance times in patients with typhoid infection. For each of these antimicrobials, the exposure of the extracellular or intracellular bacteria to sub-therapeutic drug levels will increase its likelihood of resistance emergence.The outcome for patients treated with cefixime (10\u201320mg/kg/day for 7\u201314 days) in typhoid fever are particularly mixed with prolonged fever clearance times of 6\u20138 days, clinical failure rates of between 6% and 27% and microbiological failure rate between 0% and 4.5%. In an RCT of 105 adults with confirmed typhoid fever in Nepal, a combination of azithromycin and cefixime for out-patients and azithromycin and ceftriaxone for in-patients was superior to azithromycin alone with shorter fever clearance times. Unpublished data suggests that there is no evidence for significant antagonism againstS. Typhiin-vitro with the combination of azithromycin and ceftriaxone/cefixime  will be used instead of cefixime in the single drug arm so that neither the patient nor the study team know which patient is receiving which treatment. We will assess whether treatment outcomes are better with the combined treatment after one week and at one and three month follow-up. This is a phase IV, international multi-centre, multi-country, comparative participant and observer-blind, 1:1 randomised clinical treatment trial. Patients with suspected uncomplicated typhoid fever will be randomised to one of the two interventions: Arm A: azithromycin 20mg/kg/day oral dose once daily (maximum 1gm/day) AND cefixime 20mg/kg/day oral dose in two divided doses (maximum 400mg bd) for 7 days, Arm B: azithromycin 20mg/kg/day oral dose once daily (max 1gm/day) for 7 days AND cefixime-matched placebo for 7 days. The drug dosing table can be found as extended data1. Oxford University Clinical Research Unit (OUCRU)- Patan Academy of Health Sciences, Kathmandu, Nepal. Recruitment will take place in the out-patient department of Patan Hospital, the Civil Services Hospital, and Kathmandu, Nepal-Korea Friendship Hospital, Bhaktapur.2. Aga Khan University (AKU) Hospital. In addition to the main AKU campus, recruitment will take place at the National Institute of Child Health (NICH), two AKU secondary care Hospitals and the Civil Hospital, Karachi.3. The Christian Medical College, Vellore, South India. Recruitment will take place from the Paediatrics Department, CMC, Anatapura Health Centre, and Chinnalapuram Community Health Centre, CMC.4. The International Centre for Diarrheal Disease Research, Bangladesh . Recruitment will take place at the out-patient department in the hospital.th April 2020 (NCT04349826). This protocol has been written according to the SPIRIT guidelines.This trial was registered on clinicaltrials.gov on the 16Primary objectiveTo determine whether combined treatment with seven days of an azithromycin and cefixime is superior to seven days of azithromycin and placebo in preventing treatment failure in patients with clinically suspected or confirmed uncomplicated typhoid fever.Secondary objectivesTo compare the fever clearance time (FCT) in patients in each treatment armTo compare the time from onset of treatment to treatment failure in patients in each treatment armTo compare the time from onset of symptoms to treatment failure in patients in each treatment armTo compare the occurrence of adverse events in each treatment armS. Typhi orS. Paratyphi in the patients with blood culture confirmed typhoid fever at onset in each treatment armTo compare the clearance of faecal carriage ofTo compare the cost-effectiveness of the treatment in each treatment armExploratory objectivesTo determine the pharmacokinetics of each drug in the two treatment armsTo determine the diagnosis in participants who do not have blood culture confirmed typhoid feverTo compare the faecal micro biome of patients with confirmed and suspected typhoid feverEndpointsPrimary endpointS. Typhi orS. Paratyphi on day 7 of treatment regardless of the presence of fever  OR blood culture-confirmed typhoid fever relapse within 28 days of initiation of treatment.A composite outcome of treatment failure by the 28th day after the initiation of treatment will be defined by either of the following events: 1.Clinical failure: persistence of fever on day 7 (168 h) post treatment initiation OR The need for rescue treatment as judged by the Trial Clinician OR The development of any complication  OR Syndromic enteric fever relapse within 28 days of initiation of treatment. 2.Microbiological failure: a positive blood-culture forSecondary endpoints1. The fever clearance time (FCT) will be the time from the first dose of a study drug until a temperature of <\u200937.5\u00b0C (axillary); <\u200938.0\u00b0C  has been achieved for at least 48 hours2. The time to treatment failure will be the time from the first dose of a study drug until an event occurs defined as a treatment failure3. The time to treatment failure will be the time from the day of the first symptom until an event occurs defined as a treatment failure. As subjects are only under observation from the initiation of treatment onwards, they will be considered as left-truncated at that timepoint.4. Adverse events will be graded S. Typhi orS. Paratyphi with timepoints explicitly stated.5. Positive culture of faeces sample for6. The incremental cost-effectiveness ratio (ICER) will comprise of the total costs per case, real outpatient and in-patient costs, total direct and indirect costs for the family and healthcare system and health outcomes converted to disability adjusted life years (DALYs). The cost per DALY averted will be compared against multipliers of the gross domestic product (GDP)/capita in each of the four countries to establish the cost-effectiveness of the combination regimen.7. Additional diagnostic tests on the samples collected from participants who do not have blood culture confirmed typhoid fever8. Analysis of the faecal microbiome in collected faecal samplesAssessment of endpointsParticipants, or their family members, will be trained how to monitor the temperature and then contacted on a twice daily basis by telephone for monitoring and recording of temperature and enquiry using symptom checklist. Temperature and symptom logs will be maintained during the twice daily telephone calls to measure fever clearance time and when the patient is seen on day 7. Each enrolled subject will measure temperature either in the axillary area or orally and stick to one method and not interchange so the documentation will not be confusing. If there is persistence of fever and/or symptoms on day 7 the patient will be reassessed, rescue treatment administered if appropriate and the patient will be judged to be a treatment failure. In addition, adverse events or signs of any complication (using a check list) developing at any time will be closely monitored and patients will be referred to the hospital if necessary, under the supervision of one of the trial physicians.Patients still febrile and unwell on or before day 7 will be reassessed. In case of treatment failure a decision to admit or continue treatment as an outpatient will be made by the site Principal Investigator (PI). The choice of antibiotics used for cases of treatment failure, will depend on the initial report of blood culture if positive or the prevailing antibiotic sensitivity patterns if negative. The site PI will decide if rescue treatment is required. This may be ceftriaxone 60\u201380 mg/kg once daily (Max 4gm od) or meropenem 10\u201320 mg/kg three times daily or 0.5\u20131gm three times daily for body weight \u2265 50kg) according to local guidelines and follow-up will continue .Severe or complicated typhoid fever will be defined if any of the following develop:\u2022\u00a0\u00a0\u00a0Clinically significant gastrointestinal bleeding\u2022\u00a0\u00a0\u00a0Fall in the Glasgow Coma Scale score \u2022\u00a0\u00a0\u00a0Perforation of the gastrointestinal tract (confirmed at surgery)\u2022\u00a0\u00a0\u00a0Haemodynamic shock  or ultrasound evidence of a pericardial effusion)\u2022\u00a0\u00a0\u00a0Hepatitis   or alanine aminotransferase (ALT) )\u2022\u00a0\u00a0\u00a0Clinical diagnosis of cholecystitis \u2022\u00a0\u00a0\u00a0Pneumonia (respiratory symptoms and new chest X-ray infiltrates)S. Typhi orS. Paratyphi from blood collected on day 7, regardless of the presence of fever, will be judged to be treatment failure. Rescue treatment will be administered if necessary at the discretion of the site PI. Similarly, blood culture positive or syndromic relapse of typhoid fever during the follow-up through until day 28 will be judged a treatment failure and further treatment will be given under the guidance of the site PI. Participants will be followed-up approximately until resolution. If the illness is not resolved at that time of follow-up, an additional follow-up will be arrangedA positive culture ofS. Typhi orS. Paratyphi in the patients who were blood /stool culture positive for these organisms at entry to the study will be determined at 7 days, 14 days, 28 days and 3 months to check for clearance. If faecal carriage is still present at the 3 month follow up visit the Site PI will decide the need for further follow-up and treatment.Faecal carriage of. We will measure the incremental cost-effectiveness ratio (ICER) which will comprise of the total costs per case, real outpatient and in-patient costs, total direct and indirect costs for the family and healthcare system and health outcomes converted to Disability Adjusted Life Years (DALYs). The cost per DALY averted will then be compared against multipliers of the GDP/capita in each of the four countries to establish the cost-effectiveness of the combination regimen.We will collect data on health facility and household costs and loss of income during the illness and up to 28 days to enable a cost of illness analysis for typhoid fever in these settings, as well as a comprehensive economic evaluation of the treatment strategies. The difference in direct purchase costs of the treatment strategies in the four countries are modest . Therefore when considering these costs alone, any clinical benefits associated with the combined regimen may imply that the combined regimen is cost-effective. The economic benefits of the combined regimen, however, may extend further to include other health facility and household costs averted as most people in the subcontinent do not have health insurance; demonstrating these will be useful for policy guidance. We will therefore collect data on health facility and household costs and loss of income during the illness and up to 28 days to enable a cost of illness analysis for typhoid fever in these settings, as well as a comprehensive economic evaluation of the treatment strategies. The analysis will take a societal perspective and include the costs of AMR per antibiotic consumed using methods from a recent publication on this topicThe end of trial is the point at which the last participant has completed their follow-up period, all the data has been entered and queries resolved, the last sample has been processed and the database locked.Inclusion criteria\u2022\u00a0\u00a0\u00a0A history of fever at presentation for \u2265 72 hours and a documented fever (\u226537.5\u00b0C (axillary) or \u226538\u00b0C )\u2022\u00a0\u00a0\u00a0Age \u2265 2 years (and \u2265 10kg) to 65 years\u2022\u00a0\u00a0\u00a0No clear focus of infection on initial clinical evaluation\u2022\u00a0\u00a0\u00a0Malaria rapid diagnostic test (RDT) negative; dengue non-structural protein NS1 RDT negative; scrub typhus RDT negative; C-reactive protein (CRP) rapid test \u226510 mg/L\u2022\u00a0\u00a0\u00a0Able to take oral treatment\u2022\u00a0\u00a0\u00a0Able to attend for follow-up and can be contacted by telephone\u2022\u00a0\u00a0\u00a0Written fully informed consent to participate in the study including assent for children in addition to parental/legal guardian consent.Exclusion criteriaThe participant may not enter the trial if ANY of the following apply:\u2022\u00a0\u00a0\u00a0History of fever for >14 days\u2022\u00a0\u00a0\u00a0Pregnant or positive pregnancy test or breast-feeding\u2022\u00a0\u00a0\u00a0Presence of clinical symptoms or signs indicating a focal infection such as pneumonia; urinary infection, meningitis, eschar\u2022\u00a0\u00a0\u00a0Obtundation, haemodynamic shock, visible jaundice, gastrointestinal bleeding or any signs of severe disease that may require immediate hospitalisation\u2022\u00a0\u00a0\u00a0Being treated for tuberculosis (TB) or human immunodeficiency virus (HIV) or severe acute malnutrition\u2022\u00a0\u00a0\u00a0Patients with cardiac disease\u2022\u00a0\u00a0\u00a0Patient requiring intravenous antibiotics for any reason\u2022\u00a0\u00a0\u00a0Previous history of hypersensitivity to any of the treatment options\u2022\u00a0\u00a0\u00a0Either of the trial drugs are contraindicated for any reason (e.g. drug interactions)\u2022\u00a0\u00a0\u00a0Has received azithromycin or cefixime in the last five days\u2022\u00a0\u00a0\u00a0Receiving another antimicrobial and responding clinically to the treatment as judged by the attending clinician.\u2022\u00a0\u00a0\u00a0Being on another drug  that may also cause prolonged QT interval\u2022\u00a0\u00a0\u00a0Positive polymerase chain reaction (PCR) test for coronavirus disease 2019 (COVID-19) before or after randomisationScreening and eligibility. The non-specific clinical presentation of typhoid fever can make it difficult to distinguish typhoid from other causes of an acute undifferentiated febrile illness such as malaria and dengue based on clinical history, physical examination and initial laboratory investigations alone. There are no satisfactory rapid diagnostic tests for typhoid fever, and patients are treated for presumed typhoid fever (in high typhoid fever incidence areas) or fever without a focus that require antimicrobials (in low enteric fever incidence areas). We will use the RDTs available for other common causes of fever in our region to rule out malaria, dengue, and scrub typhus, and a CRP cut-off to select the sub-group of patients more likely to have a bacterial infection such as typhoid fever. In a large fever study from South East Asia, a CRP at a threshold of 10mg/L had sensitivity for detecting bacterial infections of 95% with a specificity of 49%. In 251 adults and children with blood culture positive typhoid fever in Vietnam, 242 (96.4%) had a CRP \u226510mg/L at presentation . In addition, with the appearance of COVID-19, this disease will have to be kept in the differential diagnosis as a cause of suspected or confirmed typhoid fever. We plan to do a PCR test for COVID-19 after randomization of our patients as there is usually a 24 hour test turnaround time. If the patient tests positive, the patient will be taken out of the study and followed up in a regular fashion as per the local hospital rules. The screening for COVID-19 will be site specific and may be based on reverse transcription polymerase chain reaction (RT-PCR) or antigen testing as per the local guidelines of the country.Informed consent. Prospective, written informed consent will be obtained from the participant, or the parent or person with legal responsibility  for a child (if participant\u2019s age is \u2264 18 years old), after explanation of the aims, methods, benefits and potential hazards of the trial and before trial specific procedures are performed. The consent forms can be found as extended data. Any patients potentially eligible for the study will be screened by the research nurse (RN) and community medical auxiliaries (CMA). Those meeting the criteria will be approached for informed consent by the study doctor.Written and verbal versions of the participant information and informed consent will be presented to the participant, or the parent or person with legal responsibility for a child, detailing: the exact nature of the trial; what it will involve for the participant; the implications and constraints of the protocol; the known side effects and any risks involved in taking part. It will be clearly stated that the participant, or the parent or person with legal responsibility for a child, is free to withdraw from the trial at any time for any reason without prejudice to future care, without affecting their legal rights and with no obligation to give the reason for withdrawal. The participant, or the parent or person with legal responsibility for a child, will also be informed that they can choose to have their remaining blood samples destroyed and not stored for future analysis at the end of the trial.The participant, or the parent or person with legal responsibility for a child, will be allowed time to consider the information, and the opportunity to question the Investigator or other independent parties to decide whether they will participate in the trial.The participant, or parent or person with legal responsibility a child, must personally sign and date the latest approved version of the informed consent form. For children, where understanding is considered adequate, they will be asked to sign the assent form as per local ethics committee guidance. For children below the considered age, assent will be verbal, along with the consent from guardian. The person who obtained the consent must be suitably qualified and experienced, and have been authorised to do so by the Chief/Principal Investigator. A copy of the signed informed consent will be given to the participant. The original signed form will be retained at the trial site.If the participant, or the parent or person with legal responsibility for a child, is illiterate then a third party independent of the study may act as a witness to attest that the information in the consent form and any other written information was accurately explained to, and apparently understood by, the participant, or the parent, or person with legal responsibility for a child, and that informed consent was freely given. In this event, the witness will also sign and date the consent form.Randomisation and treatment allocation. Participants will be randomly allocated to one of two treatment arms resulting in a 1:1 final disposition. The statistician in charge of randomization list preparation will set up statistical code to generate the randomization list and transfer it to the study pharmacist in accordance to the standard operation procedures (SOPs). The study pharmacist will then change the random seed, i.e. the initialization of the random numbers generator, in the statistical code in order to blind the biostatistician and then run the code to prepare the final randomization list. The generated randomization lists will be securely incorporated within the trial database. A reliable manual back-up system will also be available. The computer-generated randomisation list will use block randomization with stratification by site and age (children <16 years) and adults (\u226516 years). The recruitment in most sites will take place during working hours as these are uncomplicated typhoid patients.Randomisation will be performed using a web-based system accessed using an authorized username and password at each site and administered by the Oxford University Clinical Research Unit- Nepal Clinical Trial Unit independent from the study. Before treatment allocation the patient\u2019s eligibility and informed signed consent will be confirmed and entered into the database. The allocation code will be held by the CTU pharmacist in Nepal.Initial trial visit. Once consented, the following procedures will be completed and all details will be recorded in the case report form (CRF).1. A member of the study team will collect demographic information (including age and address) and participant contact details.2. A history and full physical examination will be performed. This will include current and past medical history and medications. Height/length and weight will be collected for all children <5 years of age.3. Any additional laboratory samples will be collected and all available results will be reviewed . Caregivers are instructed to be consistent and measure temperature always at the same location  for each patient and approximately at the same time. Face to face follow up by participant attendance at the clinic will be at 7 days, two weeks and one month. For patients who had positive blood or stool culture forS. Typhi orS. Paratyphi at the time of presentation there will be a final clinic follow-up at three months.The telephone calls will be made to ensure that clinical progress is satisfactory and for monitoring of symptoms. Any problems faced by participants can be discussed and possible adverse effects identified.Community medical assistants (CMA) will visit the patients at their home on day 2 and day 4 of treatment to ensure that clinical progress is satisfactory and for monitoring. Special attention will be paid to the assessment of adherence to study medications, drug-related adverse events and disease related events. Adherence will be assessed using standardised questionnaires and pill counts. At the first visit their home location will be recorded using global positioning system (GPS).All patients will return to the clinic for follow-up on day 7, day 14, day 28 (and day 90 if typhoid fever confirmed at onset) after the start of treatment for follow up clinical history, physical examination and sampling.To ensure an optimal retention rate, the participants will be contacted by phone to remind them of their next visit. In addition, patients who have missed a visit will be contacted by phone for a maximum of three times after which a maximum of three home visits can be conducted. All contact attempts will be recorded.Initial trial visitEthylenediaminetetraacetic acid (EDTA) tube blood (1-1.5mL) for haematology  Heparinised blood (2mL) for sodium, potassium, urea, creatinine, semi quantitative C reactive protein (CRP) test, bilirubin, aspartate transaminase (AST) and alanine transaminase (ALT)EDTA blood (2mL) will be used for malaria RDT, dengue RDT, scrub typhus RDT, and CRP RDT. Remaining blood centrifuged and plasma and blood pellet stored at -20\u00b0C /-80\u00b0C for future diagnostic, biomarker, pharmacokinetic studies and genetic analysis. Centrifugation and blood storage will be done ONLY if the patient fulfils the inclusion criteria.Blood culture (3-8mL depending on age). Urine sample will be collected from all participants and stored at -20\u00b0C/-80\u00b0C for later and tested by bioassay for antimicrobial activity.S. Typhi or Paratyphi). DNA (deoxyribonucleic acid) will be extracted from the stool and stored for later diagnostic microbiome studiesFaecal sample will be collected from all participants for standard bacterial culture . For CRP screening we will use a semi-quantitative lateral flow test  that uses a drop of blood and gives a result in 5 minutes. It has thresholds of 10, 40 and 80mg/L so will give the right cutoff for inclusion, and some information on how high CRP is when it is \u226510mg/L.Follow up trial visitsS. Typhi orS. Paratyphi on day 0, 7, 14, 28. Also on day 90 if the day 0 blood/stool culture positive for salmonella.A blood culture will be repeated at the day 7 follow up visit and at any time during follow-up if there is a clinical deterioration in the participant or a clinical suspicion of relapse after review and assessment by the trial physician. Stool cultures forBlood samples drawn in the study clinic for diagnosis and confirmation of suspected typhoid fever will be handled, stored, processed, in accordance with standard operating procedures of the study Microbiology Laboratory. The results of these tests will be recorded in the participant CRF for use in this study.Remaining samples will be stored at the local laboratory for further analysis, depending on funding availability, after the conclusion of the trial. This analysis will include diagnostic testing (serology or PCR) to determine additional causes of infection in the blood culture negative group.Additionally DNA extraction for investigation of the genetic control of susceptibility to infectious diseases like typhoid and pharmacogenetic determinants may be carried out. DNA will be genotyped/sequenced and analysis will be performed .ADAPT5.At selected sites, additional samples will be taken for pharmacokinetic analysis. This will be one or two additional EDTA samples taken during the course of treatment. The cells and plasma will be separated and stored at -20\u00b0C/-80\u00b0C. These samples will subsequently be assayed for azithromycin and/or cefixime using high-performance liquid chromatography (HPLC). The data will be analysed using a population pharmacokinetic modelling approach using Pharmacokinetic/Pharmacodynamic Systems Analysis SoftwareS. Typhi/S. Paratyphi across South Asia and characterise relevant resistance and/or virulence mechanisms.Bacterial isolates will be stored at -20\u00b0C/-80\u00b0C for later whole genome sequencing. The WGS will be performed at study sites if they have capacity, or at the Wellcome Sanger Institute, UK. This will be to determine if recurrent positive blood cultures are due to relapse or re-infection, to understand the population structure ofWe will randomize 1500 subjects (750 per arm) within 2 years, with between 125 and 250 recruited per year at each site. Assuming a treatment failure risk of 15% in the group given azithromycin alone, the sample size will guarantee 92% power to detect an absolute risk reduction of 6% (from 15% to 9%) and 80% power to detect an absolute risk reduction of 5% (from 15% to 10%) in the group given the combined antimicrobials at the two-sided significance level of 5% allowing for up to 10% loss to follow up. In case of a lower treatment failure risk of 10% in the control arm, the trial will have 94% power to detect a 5% absolute risk reduction (from 10% to 5%) and 77% to detect a 4% reduction (from 10% to 6%).The statistical aspects of the study are summarised here with details fully described in a statistical analysis plan (SAP) which will be finalised before any analysis takes place. The primary endpoint of this trial is the composite outcome of treatment failure by the 28th day after the initiation of treatment. Subjects who withdraw from the study prior to day 28 without treatment failure will be treated as right-censored. The primary comparison of the absolute risk of treatment failure until day 28 between the treatment arms will be based on Kaplan-Meier estimates and corresponding standard errors according to Greenwood\u2019s formula.S. Typhi orS. Paratyphi vs. negative), age (children (<16 years) and adults (\u226516 years)), and country. Time to treatment failure from symptom onset will be analysed in the same way but subjects will be considered as left-truncated at the time of onset of treatment to account for the fact that they were not under observation before that timepoint.The time to treatment failure from the onset of treatment will be described using Kaplan-Meier curves and compared between the treatment arms with a Cox proportional hazards model with treatment assignment as the primary covariate and adjustment for blood culture result  and comparisons between the arms will be based on a parametric Weibull accelerated failure time model. A two-sided significance level of 5% will be applied to all efficacy analyses. The primary analysis population for all analyses is the modified intention to treat (ITT) population consisting of all patients who have been randomised to the trial and received at least one dose of study treatment. Analysis will be according to the randomized treatment arm. Subjects, who were mistakenly randomised or withdrew before the first dose of study treatment was given, will be excluded. Subjects who were unblinded will be included in the ITT population.S. Typhi orS. Paratyphi vs. negative), by age (children (<16 years) and adults (\u226516 years)), by country, and by study site. The culture-confirmed population consists of all patients with blood-culture confirmed typhoid fever who received at least one dose of study treatment. Analysis will be according to the randomized treatment arm. The primary and secondary endpoints will be analysed both in the ITT and the culture-confirmed population. Planned sub-group analyses for the primary endpoint and fever clearance time include analyses by blood culture result  as a guidance.R language for statistical computing version 3.6.2 software.The efficacy interim analysis will also include a futility analysis which will assess the conditional power to detect a 6% difference in failure rates at the final analysis. If the conditional power is low, the DSMC may recommend closing the trial for futility or increasing its sample size. The final decision making in case of a sample size increase will require input from the Trial Steering Committee, the JGHT Committee and the joint funders. All analyses will be done withData collection is the responsibility of the clinical trial staff at the site under the supervision of the site PI. The investigator is responsible for ensuring the accuracy, completeness, legibility, and timeliness of the data reported. Clinical data (including adverse events (AEs), concomitant medications, and expected adverse reactions data) and clinical laboratory data will be entered into CliRes, a 21 CFR Part 11-compliant data capture system provided by the OUCRU IT department. The data system includes password protection and internal quality checks, such as automatic range checks, to identify data that appear inconsistent, incomplete, or inaccurate. Clinical data will be entered directly in tablets connected to the central database collection site at the Nepal CTU. The sites can access their data in the central data base which will be password protected.The primary data is collected onto the CRF while interviewing the patients at the outpatient department in the hospital. There will be 1500 patients, with 5 hospital visits each, 2 home visits by CMA and data obtained from the telephone calls. On average, each patient will have a CRF, which contains around 23\u201325 pages with all patient details. The participants will be identified by a unique trial specific number and/or code in any database. The name and any other identifying detail will NOT be included in any trial data electronic file.Only authorized users with specific delegation will be given access to the database. These users will be trained according to their delegation before working with the database. All the changes made in the database will be tracked and will be kept as a part of documentation. Data storage will be in the central database in the Nepal CTU and backed up in Vietnam. All the sites will be able to access their data secured with password from the central site. All the metadata will be specified during the database development which includes data type, coding, table and column names, validation etc. The data required for the analysis from the extracted database will be derived from the metadata dictionary.All the physical data from the CRFs will be collected in an electronic tablet in each site which will feed into the central electronic data base which will be stored securely at the OUCRU Nepal server with regular backup in Vietnam. The data security standards in place at each site are detailed in the unit\u2019s system level security policy which adheres to the standard for information security management ISO27001. All the trials participants\u2019 anonymity and confidentiality will be maintained throughout the study period. The same will be ensured with the data of the participants. They will be identified by the specific study numbers rather than their name or any other identifiable characters. All the users involved in the data management will be trained before allocation to the specific tasks related to patient details.Access to data for outside party will be given only after approval by the Trial steering group which will be through application to the group. OUCRU CTU SOP ensures participants\u2019 anonymity and Confidentiality during the data sharing procedure. The Nepal CTU will ensure the trial is `discoverable\u2019 through trial registration.The definitions of the EU Directive 2001/20/EC Article 2 based on the principles of the International Conference on Harmonization good clinical practice (ICH GCP) apply to this trial protocol. Adverse effects will be classified and graded according to the Common Terminology Criteria for Adverse Events (CTCAE) system. All serious and grade 3 or 4 AEs will be compared between arms and reported by frequency per arm. An independent DMC will oversee the safety of the trial participants Definitions. The definitions of the principles of ICH GCP apply to this trial protocol. All grade 3 or 4, or serious AEs and ARs, whether expected or not, should be recorded in the CRF. Non-serious grade 1 or 2 AEs need not be recorded unless they are thought to be related to the IMP or they result in a change or interruption in treatment.Procedure for immediate reporting of serious adverse events (SAEs)Site study team will complete an SAE report form for all reportable SAEs.Where the SAE requires immediate reporting, the SAE report form will be scanned and emailed to trial coordinating team immediately i.e., within 24 hours of site study team becoming aware of the event. The team will acknowledge receipt of this report.Site study team will provide additional, missing or follow up information in a timely fashion.The assessment of expectedness will be conducted within 48 hours by the Site study team and the Coordinating Centre team within three days of reporting.All deaths will be reported to the trial coordinating team immediately i.e., within 24 hours of site study team becoming aware of the event and to the chairman of the Data Safety Monitoring Board (DSMB)All SAEs other than those defined in the protocol as not requiring reporting must be reported on the SAE reporting form to CTU within 24 hours of the trial coordinating team becoming aware of the event. SAEs will be reported as soon as possible to the site ethics committee (EC). A written report will be sent as soon as possible but not later than 2 working days (for SAE resulting in death or life threatening). Additionally, the detailed report of the SAE should be submitted to the IRBs within 15 days. Additional medical information of the SAE\u2019s development must be reported in other report until the trial subjects recover or stabilize without further change expected. Other AEs should be recorded, summarized and reported in the annual report form and the final report form. All SAEs will be reported to OxTREC in the annual review form and to the DMC in accordance to the DMC charter.The trial will be conducted in accordance with the current approved protocol, GCP, relevant regulations and standard operating procedures including precautions regarding COVID-19. A risk assessment and monitoring plan will be prepared before the study starts and will be reviewed as necessary over the course of the trial to reflect significant changes to the protocol or outcomes of monitoring activities.Regular monitoring will be performed according to the trial specific monitoring plan. Data will be evaluated for compliance with the protocol and accuracy in relation to source documents as these are defined in the trial specific monitoring plan. Following written standard operating procedures, the monitors will verify that the clinical trial is conducted and data are generated, documented and reported in compliance with the protocol, GCP and the applicable regulatory requirements.The Investigator will ensure that this trial is conducted in accordance with the principles of the Declaration of Helsinki and with relevant regulations, approved protocol and good clinical practice. Following Sponsor approval, the protocol, informed consent form, participant information sheet and proposed advertising material has been approved by the Oxford Tropical Research Ethics Committee (OXTREC) (reference number 28-20) and the Nepal Health Research Council (reference number 2291). The decision from the remaining review boards is pending.The rights of the participant to refuse to participate in the trial without giving a reason must be respected. After the participant has entered into the trial, the clinician must remain free to give alternative treatment to that specified in the protocol, at any stage, if he/she feels it to be in the best interest of the participant. The reason for doing so, however, will be recorded; the participant will remain within the trial for the purpose of follow-up and for data analysis by the treatment option to which they have been allocated. Similarly, the participant must remain free to change their mind at any time about the protocol treatment and trial follow-up without giving a reason and without prejudicing his/her further treatment.All participants will receive the best available treatment of typhoid fever, following local and national guidelines. They will benefit from the frequent and careful follow-up of their condition throughout the treatment of their typhoid fever and for up to 28 days from randomization.The risks of participation are few. Azithromycin and cefixime are commonly prescribed drugs and there is widespread experience and expertise concerning their safe use. The choice dose, route of administration and duration of study treatment follows the international guidance for the treatment of typhoid fever. Previous trials have demonstrated the safety of this or similar regimens.Against these minimal risks, trial participants may benefit from receiving these antibiotics for treatment of their suspected or confirmed typhoid fever. In addition, all participants will benefit from the careful observation and follow-up over 28 days from enrolment, which will allow complications of typhoid fever to be rapidly identified and managed.The risks and benefits of participation will be communicated in two ways. First, all potential participants or their family members will be given a participant information sheet clearly listing the risks and benefits of the trial. Second, all potential participants (or their families) will be able to discuss participation with their consulting doctor who will be able to address questions not covered or arising from the participant information sheet.Participants\u2019 confidentiality will be maintained throughout the trial. Data submitted to Nepal CTU and samples sent to central testing facilities will be identified only by the trial number and participant initials.The Principal Investigator shall submit once a year throughout the clinical trial, or on request, an annual progress report to the REC, HRA (where required), host organisation, funder (where required) and Sponsor. In addition, an End of Trial notification and final report will be submitted to the local and national ethics committee, host organisation and Sponsor within 12 months of completion of the study.The study will comply with the General Data Protection Regulation (GDPR) and Data Protection Act 2018, which require data to be de-identified as soon as it is practical to do so. The processing of the personal data of participants will be minimised by making use of a unique participant study number only on all study documents and any electronic database(s), with the exception of the CRF, where participant initials may be added. All documents will be stored securely and only accessible by study staff and authorised personnel. The study staff will safeguard the privacy of participants\u2019 personal data.Reasonable travel expenses for any visits additional to normal care will be reimbursed adequately to cover their costs on production of receipts, or a mileage allowance will be provided as appropriate. In addition, the cost of trial-related AEs that lead to hospital admission or treatment will be covered by the studyInsurance. The University has a specialist insurance policy in place which would operate in the event of any participant suffering harm as a result of their involvement in the research .Contractual arrangements. Appropriate contractual arrangements will be put in place between the University of Oxford and each site involved in the study.The investigators will co-ordinate dissemination of data from this study. For dissemination of research, we will give presentations in typhoid meetings in Nepal, India, Bangladesh, Pakistan and in international forums like The American Society of Tropical Medicine and Hygiene (ASTMH), The Royal Society of Tropical Medicine and Hygiene (RSTMH) and also in infectious disease societies in each of the collaborator countries. We will also engage with local World Health Organization (WHO) and health ministries to see if the findings from the study can have impact on policy. Finally, we hope to publish the findings in a weekly international medical journal for a wide impact. All publications, including manuscripts, abstracts, oral/slide presentations, and book chapters, etc., based on data from this study will be reviewed by each sub-investigator prior to submission. Authors will acknowledge that the study was funded by MRC. In accordance with MRC, all publications related to this study will be open access. Authorship will be determined in accordance with the International Committee of Medical Journal Editors (ICMJE) guidelines and other contributors will be acknowledged.Study documents  will be maintained for time specified by the study protocol, sponsor and regulatory authority. Data will be stored in centralised Oxford University Clinical Research Unit (OUCRU) server which is backed up regularly. These documents will be archived in the study site for defined retention period securely. No personal identifiers or participants information will be included in the archive., a 6% reduction in treatment failures in this population using this combination of drugs will mean at least 420,000 patients avoiding the need for further antimicrobial treatment and/or hospital admission, and consequently result in a lower financial burden and a reduced potential for onward transmission of the infection.In this study, we will conduct a randomised comparison of azithromycin alone or azithromycin combined with cefixime for treating clinically suspected or confirmed uncomplicated typhoid fever at four sites across south Asia. We aim to test the hypothesis that the treatment failure proportion will be 15% with azithromycin alone but 9% if the azithromycin is combined with cefixime. When translated to the number of patients with typhoid fever across south Asia this improvement in outcome would make a substantial real-world contribution to improving the health of the population in our region. With an estimated 7 million people suffering from typhoid fever in South Asia each yearS. Typhi andS. Paratyphi A there are sporadic reports of resistance to both azithromycin and third generation cephalosporins. The recent outbreak of MDR, fluoroquinolone and ceftriaxone resistant (XDR) typhoid in Pakistan is a warning of the potential for untreatable typhoid. Azithromycin resistance in enteric fever pathogens would leave a major gap in treatment options. This combination would still be efficacious if the infecting pathogen was resistant to one of the drugs. If it could delay or prevent the emergence of this resistance that would have an important public health benefit. Finally, combination treatment may reduce the emergence of resistance in Enterobacteriaceae in gastrointestinal microbiota. As the use of drug combinations would lead to increased costs and more potential side-effects it is critically important to establish if there is a measurable clinical benefit of combining these antimicrobials. This potential effect is particularly important as fixed dose combinations of these antimicrobials are already available despite government regulations in India. Hence even if this study shows that the combined therapy has no advantage over a single drug treatment for suspected enteric fever, it will be important to know this against the background of the availability and anecdotal usage of the combination.Cefixime and azithromycin are widely used antimicrobials in suspected or confirmed typhoid fever with excellent safety profiles. If the combination treatment is better than the single antimicrobial treatment, this will be an important result for patients across South Asia and other suspected typhoid endemic areas. We will also additionally investigate the financial implications for families and health system. Combined treatment may also limit the emergence of resistance if one of the components is active against resistant sub-populations not covered by the other antimicrobial\u2019s activity. This is part of the rationale for using combination chemotherapy in other infections such as malaria, TB and HIV. In focusing on the fluoroquinolones concluded that this class of antimicrobial performed well in treating typhoid but that clinicians needed to take into account local resistance patterns. Recent data suggests that across most of South Asia the levels of non-susceptibity to fluoroquinolones is too high for this to be a reliable treatment option. The second systematic review focused on the role of azithromycin, in which seven RCTs involving 773 patients were identified. In comparison to the older fluoroquinolones (213 participants), there were fewer clinical failures (RR 0.46 (0.25-0.82)) while in comparison with ceftriaxone (132 participants) there was a significant reduction in the chance of relapse (RR 0.1 (0.01-0.76)). Both these reviews noted that most trials had been conducted on in-patients and may not therefore be representative of settings where most enteric fever is managed as an out-patient. There were also few studies in each comparison with studies that were too small to make firm conclusions on the presence or absence of important differences. The authors recommended a need for multi-centred, adequately powered trials with robust methods and analytical design. Many RCTs have only analysed the culture positive cases rather than \u2018intention-to-treat \u2018. The ACT-South Asia attempts to address these points by conducting a large multi-centre study of the out-patient treatment of clinically suspected and confirmed uncomplicated typhoid fever in the largest hub for typhoid fever in the world.There are two published systematic reviews that address the antimicrobial treatment of typhoid fever. One review is a pressing reason why this trial is needed now as there is clearly a risk of these XDR organisms spreading to other highly endemic sites in South Asia or for such organisms to emerge in other endemic locations. This study is of relevance to all of the proposed study sites as healthcare workers at each site contend with the management of patients with typhoid fever on a daily basis. Including sites from across the sub-continent will make the results generalisable to other locations in South Asia where similar problems with typhoid fever are found. Most typhoid fever in sub-Saharan Africa currently still responds to fluoroquinolone treatment but reports of fluoroquinolone resistance are emerging and alternative regimens will be needed if resistance become widespread. In addition, in our analysis, the effect of the intervention in the culture-negative patients may provide useful data to inform empiric treatment algorithms for both blood culture negative acute febrile illness in this region and also for medical facilities in vast swathes of South Asia where blood cultures are unavailable. Finally if the combination treatment arm results in less fecal carriage of the typhoid organisms, this strategy may indeed be helpful in the epidemiological control of the spread of the disease by carriers potentially helping with elimination of the disease in South Asia if the other interventions like vaccination and proper sanitation are also successful.The outcome of this first major regional collaborative RCT designed to deal with a rampant scourge will be of crucial importance for clinicians working in South Asia as it may help to guide the best empirical treatment for suspected typhoid fever and reduce the rates of treatment failure, the risk of complications and hospital admission, the overuse of more expensive second line antimicrobials and extra healthcare costs. The appearance of XDR typhoid in PakistanRecruitment was planned to start from September 2020. However, study initiation has been significantly affected by the pandemic all sites. Nepal has started patient enrolment and first patient was enrolled on 23 May 2021. Nine patients have been recruited in the trial already. Pakistan will probably be the next site to start enrolment soon followed by the other sites.No data are associated with this article.https://doi.org/10.5287/bodleian:OBopRno5q.Oxford University Research Archive (ORA): Dataset for ACT-South Asia protocol manuscript.This project contains the following extended data:- DSMC charter for ACT South Asia_V1.pdf (Data & Safety Monitoring Committee)- PIS_informed_consent_adults.pdf (Patient information sheet and consent form for adults)- Drug_dosing_table-ACT South Asia.pdf (Drug dosing table)- PIS_and _assent form.pdf (Patient information sheet and assent form for children)https://doi.org/10.5287/bodleian:OBopRno5q.Oxford University Research Archive (ORA): SPIRIT checklist for \u2018Azithromycin and cefixime combination versus azithromycin alone for the out-patient treatment of clinically suspected or confirmed uncomplicated typhoid fever in South Asia; a randomised controlled trialCreative Commons Attribution 4.0 International license (CC-BY 4.0).Data are available under the terms of the What about those who will be found to have another diagnosis, are they included in the ITT analysis?Leptospirosis which is \u00a0a neglected disease (probably due to lack of\u00a0 diagnostic methods). More efforts should be carried on to identify it. Since blood (serum ) is taken on day 0 and day 7, that sample can be kept for all kinds of serology including leptospirosis.As I mentioned previously one of the common febrile illnesses in these regions (mainly in\u00a0 agriculture population) isThe primary analysis of this study is the intention to treat, including those with culture negative. This group of patients on one hand represents the real group which is seen in the clinics in daily life. But, on the other hand , a major part of them might have\u00a0 another  infections. Therefore all efforts should be done to get an accurate diagnosis of those who were culture negative. In this regard:Is the study design appropriate for the research question?PartlyIs the rationale for, and objectives of, the study clearly described?YesAre sufficient details of the methods provided to allow replication by others?NoAre the datasets clearly presented in a useable and accessible format?YesReviewer Expertise:NAI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. Thank you very much, Dr Eli Schwartz.\u00a0 In answer to your first query, yes indeed those that are found to have another diagnosis will be included in the ITT analysis. And we will also have a sub-group analysis of just those who are culture positive for typhoid fever. So, this will be a wide-ranging study the results of which should impact the treatment of undifferentiated febrile illness in South Asia including typhoid fever. In response to your second query regarding serological diagnosis of leptospirosis and other common ( often neglected) disease-causing organisms, we are taking this matter up seriously and actively looking for means to carry out these tests. Thank you for the opportunity to review the authors responses to reviewer comments, which are clear and sufficiently detailed.Is the study design appropriate for the research question?PartlyIs the rationale for, and objectives of, the study clearly described?YesAre sufficient details of the methods provided to allow replication by others?YesAre the datasets clearly presented in a useable and accessible format?Not applicableReviewer Expertise:Infectious diseases, study designI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. This is an extremely important study which could have an important impact globally on the approach for treating people with typhoid. It could also have an economic impact as well as possibly contribute to changes in the epidemiology of the disease (see below). It is unfortunate that this trial took such a long time to be conducted. From an historical perspective, it is interesting and important to elaborate on the role of the field of travel medicine in the development of this therapeutic solution.rd generation cephalosporins (despite being\u00a0 sensitive in-vitro) and to azithromycin when each of them was given alone. The proposed approach in 20091 was to give these two drugs, azithromycin and 3rd generation intravenous (IV) cephalosporins, together. The idea was that these two drugs would cover the extracellular and intracellular compartments, the two compartments where the typhoid bacteria is shifting between. The high efficacy of fluoroquinolones before the development of NA-resistant strains can perhaps be attributed to their excellent distribution in both the intra- and extracellular compartments. Treating enteric fever (typhoid and paratyphoid fever) became a challenge during the two decades when resistance to quinolones, the magic bullet against typhoid fever, became evident. Interestingly, the call for a new approach was declared first from the Travel Medicine practitioners who reported\u00a0 the unsatisfactory response to both\u00a0 32 In this observational study, combined therapy shortened the febrile days from 6 to 3 days. This reference somehow was omitted in the current proposal by Abhishek Giri,\u00a0et al. This proposal was introduced as the novel treatment for typhoid fever and became the policy in Sheba Medical Ctr. In Israel. It was further approved during a major paratyphi outbreak in Israeli travelers in Nepal, reported in 2014. The Israeli concept was further tested in a comparative study done among local populations in Nepal [appears as Ref# 17 in the proposal]. In this trial, patients with typhoid and paratyphoid fever were included . The other advantage of the Nepalese study was the inclusion of an oral cephalosporin arm in non-hospitalized patients (and not only IV ceftriaxone) which is appropriate in the environment of most of the endemic countries where the majority of patients are treated as outpatients. The results showed that the combination of third-generation cephalosporins and azithromycin may confer a more effective therapy, reducing the time to defervescence and to the clearance of bacteremia. The current large scale, multi-country trial, as proposed now by the team, is most welcome. The design of the study is rigorous, including a placebo arm and a much larger number of participants. As the author mentioned, successful results will certainly lead to an adoption of this policy and bring with it great benefits to the patients as well as have a positive impact on the economy. An interesting outcome which might be monitored using the combined therapy is whether we will also see a \u00a0decrease in the fecal-carriage rates  and in this case the spread of the disease might be limited, having an important epidemiological impact. This aspect should be highlighted in the manuscript.Since the absorption of oral cephlosporins is not steady, results might be different from IV ceftriaxone treatment. I wonder whether in this huge trial a small fraction of hospitalized patients can be recruited to test the combination of IV ceftriaxone + azithro against one of the drugs + placebo. .It is not clear (or perhaps I missed it) whether those with negative cultures will be included in final analysis. If yes, what about all those who subsequently will have\u00a0 (with the more sophisticated tests) another microbial diagnosis?Leptospirosis which is not mentioned on the list. Should a rapid test or PCR be added to the list?There are lists of rapid tests that will be performed; one of the common febrile illnesses in these regions isBlood culture- As I know there are different blood-culture bottles in these countries with different amount of blood, which gives different sensitivity.\u00a0 Are these centers going to be supplied from a central supplier? There is no clear instruction about the amount of blood.The Placebo mentioned is sugar pills. I am concerned that this may prevent a real blinding of the study.There is redundancy in secondary end-point and the statement below appears 3 times. Other aspects:\u2022To compare the time from onset of treatment to treatment failure in patients in each treatment arm\u2022To compare the time from onset of symptoms to treatment failure in patients in each treatment arm\u2022To compare the time from onset of symptoms to treatment failure in patients in each treatment arm \u00a0-RefSchwartz E. Typhoid and paratyphoid fever. In: Schwartz E, editor. Tropical Diseases in Travelers. New York, NY: John Wiley & Sons; 2009: 144-153.Meltzer E, Stienlauf S, Leshem E, Sidi Y, Schwartz E. A large outbreak of Salmonella Paratyphi A infection among israeli travelers to Nepal. Clin Infect Dis. 2014 Feb;58(3):359-64. doi: 10.1093/cid/cit723. Epub 2013 Nov 5. PMID: 24198224.Is the study design appropriate for the research question?PartlyIs the rationale for, and objectives of, the study clearly described?YesAre sufficient details of the methods provided to allow replication by others?NoAre the datasets clearly presented in a useable and accessible format?YesReviewer Expertise:Travel and Tropical MedicineI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. We would like to thank Eli Schwartz for his comprehensive input that has definitely enhanced the presentation of the protocol in its clarity and focus. Below is our point-by-point response to both the Reviewers and suitable changes have also been made in the track-changed manuscript. Reviewer comments in italics.CommentsThis is an extremely important study which could have an important impact globally on the approach for treating people with typhoid. It could also have an economic impact as well as possibly contribute to changes in the epidemiology of the disease (see below). It is unfortunate that this trial took such a long time to be conducted. From an historical perspective, it is interesting and important to elaborate on the role of the field of travel medicine in the development of this therapeutic solution.rd\u00a0generation cephalosporins (despite being\u00a0 sensitive in-vitro) and to azithromycin when each of them was given alone. The proposed approach in 20091\u00a0was to give these two drugs, azithromycin and 3rd\u00a0generation intravenous (IV) cephalosporins, together. The idea was that these two drugs would cover the extracellular and intracellular compartments, the two compartments where the typhoid bacteria is shifting between. The high efficacy of fluoroquinolones before the development of NA-resistant strains can perhaps be attributed to their excellent distribution in both the intra- and extracellular compartments. Treating enteric fever (typhoid and paratyphoid fever) became a challenge during the two decades when resistance to quinolones, the magic bullet against typhoid fever, became evident. Interestingly, the call for a new approach was declared first from the Travel Medicine practitioners who reported\u00a0 the unsatisfactory response to both\u00a0 32\u00a0In this observational study, combined therapy shortened the febrile days from 6 to 3 days. This reference somehow was omitted in the current proposal by Abhishek Giri,\u00a0et al. This proposal was introduced as the novel treatment for typhoid fever and became the policy in Sheba Medical Ctr. In Israel. It was further approved during a major paratyphi outbreak in Israeli travelers in Nepal, reported in 2014. The Israeli concept was further tested in a comparative study done among local populations in Nepal [appears as Ref# 17 in the proposal]. In this trial, patients with typhoid and paratyphoid fever were included . The other advantage of the Nepalese study was the inclusion of an oral cephalosporin arm in non-hospitalized patients (and not only IV ceftriaxone) which is appropriate in the environment of most of the endemic countries where the majority of patients are treated as outpatients. The results showed that the combination of third-generation cephalosporins and azithromycin may confer a more effective therapy, reducing the time to defervescence and to the clearance of bacteremia. The current large scale, multi-country trial, as proposed now by the team, is most welcome. The design of the study is rigorous, including a placebo arm and a much larger number of participants. As the author mentioned, successful results will certainly lead to an adoption of this policy and bring with it great benefits to the patients as well as have a positive impact on the economy. An interesting outcome which might be monitored using the combined therapy is whether we will also see a \u00a0decrease in the fecal-carriage rates  and in this case the spread of the disease might be limited, having an important epidemiological impact. This aspect should be highlighted in the manuscript. Other aspects:Since the absorption of oral cephlosporins is not steady, results might be different from IV ceftriaxone treatment. I wonder whether in this huge trial a small fraction of hospitalized patients can be recruited to test the combination of IV ceftriaxone + azithro against one of the drugs + placebo. . It is not clear (or perhaps I missed it) whether those with negative cultures will be included in final analysis. If yes, what about all those who subsequently will have\u00a0 (with the more sophisticated tests) another microbial diagnosis?Response We thank the Reviewer for these very important comments emphasizing the points that have been raised in the protocol. Indeed studies  and returned travelers from Nepal published in the CID in 2014) carried out by Dr. Eli Schwartz and his team actually helped trigger the concept for this proposal. We mistakenly referenced another article instead of the correct CID article by Meltzer et al ( # 28). The mistake has now been rectified and we thank the reviewer for pointing this out. We have also now further emphasized at the end of the manuscript the fact that if the combination treatment arm results in the less fecal carriage, this will clearly be helpful in the control of the spread of the disease by carriers. Regarding the other Aspects portion,\u00a0 testing ceftriaxone will not be possible as this is a study of outpatients, not hospitalized patients, as mentioned by the Reviewer. Finally, we are indeed trying to obtain funding to carry out studies in the culture-negative group to ascertain etiology. We beg Reviewer two's pardon in that we missed responding to his last 4 important queries. Our responses are in bold and the reviewers comments are in italics.\u00a0which is not mentioned on the list. Should a rapid test or PCR be added to the list?There are lists of rapid tests that will be performed; one of the common febrile illnesses in these regions is\u00a0LeptospirosisThe reviewer raises an important point but there are no rapid reliable tests including PCR available for us at this stage to include this in the protocol. \u00a0Blood culture- As I know there are different blood-culture bottles in these countries with different amount of blood, which gives different sensitivity.\u00a0 Are these centers going to be supplied from a central supplier? There is no clear instruction about the amount of blood.\u00a0Under lab testing we have indeed mentioned in the protocol that 3 to 8 ml will be sampled from the patient depending on the age of the patient . There will be no central supplier.The Placebo mentioned is sugar pills. I am concerned that this may prevent a real blinding of the study.\u00a0We have tasted both the\u00a0placebo and the real cefixime and they taste no different.There is redundancy in secondary end-point and the statement below appears 3 times.\u00a0To compare the time from onset of treatment to treatment failure in patients in each treatment armTo compare the time from onset of symptoms to treatment failure in patients in each treatment armTo compare the time from onset of symptoms to treatment failure in patients in each treatment arm\u00a0 \u00a0We did not see this redundancy in the Secondary\u00a0Endpoints. Perhaps the Reviewer was referring to Secondary Objectives. But again there is no redundancy because we are trying\u00a0to compare onset of symptoms as well as onset of treatment separately to treatment failure.Thank you very much for your kind consideration.Dr Basnyat, PI Main comments The rationale for performing this study to assess the efficacy of monotherapy versus combined therapy for treatment of typhoid fever is well-justified, and the protocol is generally clear and sufficiently detailed.in patients with uncomplicated typhoid fever. In fact, the ITT analysis will answer whether combination therapy is superior to azithromycin minor in preventing treatment failurein patients with suspected typhoid fever, but not specifically in those with uncomplicated typhoid fever. Both questions have clinical importance, but the precise main research question being addressed should be accurately stated, and should be directly matched to the inclusion criteria and the primary endpoints . However, I have suggested that the study design is only partly appropriate for the research question because patients with suspected typhoid fever will be included, thereby mixing outcomes of patients with typhoid and those with other conditions. I note that the main analysis will be as ITT, but that primary and secondary endpoints will be analysed both as ITT and among the culture-confirmed population. I realise that the ITT design has practical and logistic advantages, but some non-typhoidal bacterial infections will also respond to these antibiotics. Consequently, since the main analysis is ITT, there is an apparent mismatch with the stated key primary outcome \u2013 to determine whether combination therapy is superior to azithromycin minor in preventing treatment failure My concern about the precise research question also relate to the power calculations, where there is a lack of clarity regarding whether it is suspected or proven infection that serves as the main endpoint. No data are provided on the proportion of suspected cases that \u2013 after exclusion of malaria, dengue, scrub typhus and COVID-19 (and a CRP <10) \u2013 are likely to actually have typhoid, so the impact on the study\u2019s power to assess efficacy endpoints in the subset with proven typhoid infection (culture positive) is unclear. Additionally, the sample size calculations assume treatment failure of 15%, but it is not clear whether this assumption refers to an estimated failure rate of treatment for typhoid infection specifically, or overall treatment failure in the context of an ITT analysis. Clarification here is needed. Finally, a 10% loss to follow-up out to 28 days is perhaps optimistic and should be justified.Minor comments On page 6, under Secondary Objectives, the following is stated twice \u2018To compare the time from onset of symptoms to treatment failure in patients in each treatment arm\u201d A secondary endpoint  is stated as \u201cPositive culture of faeces sample for S. Typhi or S. Paratyphi\u201d \u2013 suggest that the relevant timepoint(s) be explicitly stated Will enrolled patients be supplied with a thermometer so they can check their temperature at home? If a patient has a positive blood culture with antibiotic sensitivities that demonstrate resistance to the study medication(s), will a change to therapy before day 7 be instituted, or will this depend on clinical response? What is the reason for limiting the upper age to 65 years? An exclusion criterion is cardiac disease \u2013 does this extend to all types of cardiac disease, or is it only QT interval or arrhythmias? Page 8 states that children with adequate understanding may be able to sign the assent form, yet asper Page 10, this includes consent for DNA extraction for investigation of human genetic variables. I question the ethics of allowing children to consent to this aspect of the protocol. On page 9, under Follow-up trial visits, it is stated \u201cFor patients who had positive blood or stool culture for S. Typhi or S. Paratyphi at the time of admission\u2026..\u201d \u2013 since patients must have uncomplicated infection, I presume they won\u2019t  be admitted, so should this be \u201cpresentation\u201d rather than \u201cadmission\u201d? Bottom of 11/top P12 \u2018Clinical data will be entered directly in tablets will be utilized from the various sites to enter data directly to the central database collection site at the Nepal CTU.\u201d \u2013 something is missing from this sentence, which currently does not make sense. Further to my main comment above, the Discussion (page 14) also seems to mix potential impacts on suspected and proven typhoid infections. Will some children in the recruitment catchment potentially be vaccinated, and if so will this be documented?Is the study design appropriate for the research question?PartlyIs the rationale for, and objectives of, the study clearly described?YesAre sufficient details of the methods provided to allow replication by others?YesAre the datasets clearly presented in a useable and accessible format?Not applicableReviewer Expertise:Infectious diseases, study designI confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. We would like to very much thank Karin Leder for her comprehensive input that has definitely enhanced the presentation of the protocol in its clarity and focus. Below is our point-by-point response and suitable changes have also been made in the track-changed manuscript. Reviewer comments are in italics.Comments:The rationale for performing this study to assess the efficacy of monotherapy versus combined therapy for the treatment of typhoid fever is well-justified, and the protocol is generally clear and sufficiently detailed.in patients with uncomplicated typhoid fever. In fact, the ITT analysis will answer whether combination therapy is superior to azithromycin minor in preventing treatment failure\u00a0in patients with suspected typhoid fever, but not specifically in those with\u00a0uncomplicated typhoid fever. Both questions have clinical importance, but the precise main research question being addressed should be accurately stated and directly matched to the inclusion criteria and the primary endpoints .However, I have suggested that the study design is only partly appropriate for the research question because patients with suspected typhoid fever will be included, thereby mixing outcomes of patients with typhoid and those with other conditions. I note that the main analysis will be as ITT, but that primary and secondary endpoints will be analyzed both as ITT and among the culture-confirmed population. I realize that the ITT design has practical and logistic advantages, but some non-typhoidal bacterial infections will also respond to these antibiotics. Consequently, since the main analysis is ITT, there is an apparent mismatch with the stated key primary outcome \u2013 to determine whether combination therapy is superior to azithromycin minor in preventing treatment failure\u00a0 My concern about the precise research question also relate to the power calculations, where there is a lack of clarity regarding whether it is suspected or proven infection that serves as the main endpoint.No data are provided on the proportion of suspected cases that \u2013 after exclusion of malaria, dengue, scrub typhus and COVID-19 (and a CRP <10) \u2013 are likely to actually have typhoid, so the impact on the study\u2019s power to assess efficacy endpoints in the subset with proven typhoid infection (culture positive) is unclear. Additionally, the sample size calculations assume treatment failure of 15%, but it is not clear whether this assumption refers to an estimated failure rate of treatment for typhoid infection specifically, or overall treatment failure in the context of an ITT analysis. Clarification here is needed. Finally, a 10% loss to follow-up out to 28 days is perhaps optimistic and should be justified.Response: We thank the reviewer for raising this very important issue and pointing out where there is a lack of clarity. In this RCT we are comparing the response of azithromycin alone and azithromycin plus cefixime in clinically suspected and confirmed typhoid fever. This is outlined in the title, but we have not outlined this clearly in the text  and this now has been corrected in the text. We plan a sub-analysis of the culture-positive and culture-negative participants, but the primary analysis is the intention to treat. This represents the real group of patients that clinicians will be treating.1 Although more sensitive than blood culture, bone marrow culture is not feasible for out-patient managed typhoid. Other diagnostic approaches using serology or blood PCR also lack sensitivity and specificity.2,3 Many of the previous randomized controlled trials in typhoid fever have been criticized for restricting the analysis to blood culture-confirmed cases and not analyzing the outcome by intention to treat .4,5 Establishing the best endpoint in a typhoid treatment trial is not easy because of the challenges of case detection. Blood culture is the most widely accepted method of confirming the diagnosis but lacks sensitivity. In a systematic review, the sensitivity of blood culture in 529 true positive cases was 61% (95% CI 52-70%) compared with a bone marrow culture sensitivity of 96% (95% CI 92-99%) . Designing entry criteria to the study which therefore capture patients with typhoid fever is difficult. We know that there is an overlap in the clinical features of typhoid fever with other local infections. The clinical entry criteria in this RCT have been designed to try and optimize the proportion of patients who have typhoid fever. These criteria include clinical features consistent with an undifferentiated fever and the use of RDTs to exclude other common infections that may mimic typhoid. But it is evident that the patients recruited will include patients with blood culture-positive typhoid fever, patients with typhoid who are blood culture-negative, and patients with another infection whose clinical features are similar to typhoid.4 In this study the proportion of patients with culture-positive typhoid fever was 87 (27%) of 326 participants. We do not know how many of the other participants had blood culture-negative typhoid fever. If this proportion of 27% is replicated in the ACT-SA study there will be approximately 200 culture-positive participants in each arm of the study. In a recently published RCT, the NUFIT study, comparing azithromycin and trimethoprim-sulfamethoxazole in undifferentiated febrile illness in Nepal, the entry criteria are close to this study.5 The outcome in all of these studies was based on blood culture positive typhoid patients and the studies were not blinded.7-13 Failures in the azithromycin arm varied between 0% and 18% across the different trials . Each of these trials used slightly different azithromycin regimens and varied in the outcome measure used. The only RCT to use the 7-day 20mg/kg/day regimen that we propose to use in this trial (and the largest RCT)13 recorded a failure in 9.1% (13/142) of subjects. The next largest RCT which used an outcome definition close to the one which will be used in this study and an azithromycin dose of 10mg/kg/day for 7 days recorded a failure in 18% (11/62) of subjects. 12 The most recent of these trials was completed in Vietnam when there was no azithromycin resistance reported. 13 The estimated proportion of 15% of treatment failures with azithromycin was based on a review of the failure rates in the seven RCTs that were included in azithromycin systematic reviews.Figure 1: Random and fixed-effects meta-analysis of failure proportions with azithromycin monotherapy treatment of typhoid observed in earlier randomised controlled trials.7-13 Footnote: For individual studies, Pearson-Clopper confidence intervals are given. The combination was via the inverse variance method based on the logit-transform with a continuity correction of 0.5 in studies with zero cell frequencies. DerSimonian-Laird estimator for tau^2.\u00a0 Sporadic reports of azithromycin are beginning to appear and in the review of sensitivity data across the four sites involved in this trial resistance to azithromycin was present in 48 (2%) of 3617 isolates. Based on this evidence we considered it reasonable to estimate a failure rate of 15% in the azithromycin arm. If it is the case that we have 200 blood culture-positive patients in each arm of the trial, this will give an 80% power to detect a difference of 15% failure in the azithromycin arm and 6% in the azithromycin and cefixime arm at the two-sided significance level of 5%. Notably, in the NUFIT study which was published after our analysis of failure rates in RCTs was compiled and which was conducted double-blind, the treatment failure proportion with azithromycin at 28 days was 23 (14%) of 163 in the intention to treat analysis and 11 (30%) of 37 among the culture-positive participants. In our experience in past typhoid treatment RCTs, the dropout rate has varied between 10 and 20%. This is why we chose a 10% dropout proportion in the ACT-SA study although we will need to monitor this closely.Salmonella Typhi cases are detected by blood culture? A systematic literature review. Ann Clin Microbiol Antimicrob 2016; 15:32Mogasale V,\u00a0 Ramani E, Mogasale VV, Park JY. What proportion ofWijedoru L, Mallett S, Parry CM. Rapid diagnostic tests for typhoid and paratyphoid (enteric) fever (Review). Cochrane Database of Systematic Reviews 2017, Issue 5. Art CD008892Moore CE, Pan-Ngum W, Wijedoru LPM, et al. Evaluation of the diagnostic accuracy of a typhoid IgM flow assay for the diagnosis of typhoid fever in Cambodian children usiang a Bayesian latent class model assuming an imperfect gold standard. Am J Trop Med Hyg 2014; 90:114-120.Giri A, Karkey A, Dongol S, eta l. Trimethoprim-Suphamethoxazole versus azithromycin for the treatment of undifferentiated febrile illness in Nepal: A double-blind, randomized, placebo-controlled trial. Clin Infect Dis 2020; 73:e1478.Effa EE, Burkiwa H. Azithromycin for treating uncomplicated typhoid and paratyphoid fever (enteric fever) (Review) Cochrane Database of Systematic Reviews 2008, Issue 4. Art CD006083Effa EE, Lassi ZS, Critchley ZA, Garner P, Sinclair D, Olloaro P, Bhutta ZA. Fluoroquinolones for treating typhoid and paratyphoid fever (enteric fever) (Review) Cochrane Database of Systematic Reviews 2011, Issue 10. Art CD004530J Antimicrob Chemother 1999; 44(2): 243-50.Butler T, Sridhar CB, Daga MK, et al. Treatment of typhoid fever with azithromycin versus chloramphenicol in a randomized multicentre trial in India.Antimicrob Agents Chemother 1999; 43(6): 1441-4.Girgis NI, Butler T, Frenck RW, et al. Azithromycin versus ciprofloxacin for treatment of uncomplicated typhoid fever in a randomized trial in Egypt that included patients with multidrug resistance.Antimicrob Agents Chemother 2000; 44(7): 1855-9.Chinh NT, Parry CM, Ly NT, et al. A Randomized Controlled Comparison of Azithromycin and Ofloxacin for Treatment of Multidrug-Resistant or Nalidixic Acid-Resistant Enteric Fever.Clin Infect Dis 2000; 31(5): 1134-8.Frenck RW, Jr., Nakhla I, Sultan Y, et al. Azithromycin versus ceftriaxone for the treatment of uncomplicated typhoid fever in children.Clin Infect Dis 2004; 38(7): 951-7.Frenck RW, Jr., Mansour A, Nakhla I, et al. Short-course azithromycin for the treatment of uncomplicated typhoid fever in children and adolescents.Antimicrob Agents Chemother 2007; 51:819-25Parry CM, Ho VA, Phuong LT, et al. Randomized controlled comparison of ofloxacin, azithromycin, and an ofloxacin-azithromycin combination for treatment of multidrug-resistant and nalidixic acid-resistant typhoid fever.Dolecek C, Tran TP, Nguyen NR\u00a0 et al. A multicenter randomised controlled trial of gatifloxacin versus azithromycin for the treatment of uncomplicated typhoid fever in children and adults in Vietnam. PLoS ONE 2008; 3:e2188.Minor comments\u00a0 \u00a0On page 6, under Secondary Objectives, the following is stated twice \u2018To compare the time from onset of symptoms to treatment failure in patients in each treatment arm\u201dThank you very much. We have now deleted the redundant portion.A secondary endpoint  is stated as \u201cPositive culture of feces sample for S. Typhi or S. Paratyphi\u201d \u2013 suggest that the relevant timepoint(s) be explicitly statedWe have now added this point about explicitly stating the timepoint.Will enrolled patients be supplied with a thermometer so they can check their temperature at home?Yes.If a patient has a positive blood culture with antibiotic sensitivities that demonstrate resistance to the study medication(s), will a change to therapy before day 7 be instituted, or will this depend on clinical response?This will depend on the clinical response.What is the reason for limiting the upper age to 65 years?With azithromycin administration, one of our earlier reviewers suggested this to play it safe to avoid QT prolongation problems in the elderly population.An exclusion criterion is a cardiac disease \u2013 does this extend to all types of cardiac disease, or is it only QT interval or arrhythmias?All types including coronary artery disease.Page 8 states that children with adequate understanding may be able to sign the assent form, yet as per Page 10, this includes consent for DNA extraction for investigation of human genetic variables. I question the ethics of allowing children to consent to this aspect of the protocol.We agree. And as stated in the protocol we will rigorously follow the local ethical guidelines.On page 9, under Follow-up trial visits, it is stated \u201cFor patients who had positive blood or stool culture for S. Typhi or S. Paratyphi at the time of admission\u2026..\u201d \u2013 since patients must have an uncomplicated infection, I presume they won\u2019t  be admitted, so should this be \u201cpresentation\u201d rather than \u201cadmission\u201d?Thank you very much for picking this up. We have now changed it to presentation.Bottom of 11/top P12 \u2018Clinical data will be entered directly in tablets will be utilized from the various sites to enter data directly to the central database collection site at the Nepal CTU.\u201d \u2013 something is missing from this sentence, which currently does not make sense.Thank you very much. We have now tried to clarify this obvious confusion which the Reviewer kindly picked up.Further to my main comment above, the Discussion (page 14) also seems to mix potential impacts on suspected and proven typhoid infections.We have now tried to make this more clear in the Discussion including as addressed in the main comment section of this Reviewer.Will some children in the recruitment catchment potentially be vaccinated, and if so will this be documented?Yes."}
+{"text": "Triple antimalarial combination therapies combine potent and rapidly cleared artemisinins or related synthetic ozonides, such as arterolane, with two, more slowly eliminated partner drugs to reduce the risk of resistance. We aimed to assess the safety, tolerability, and efficacy of arterolane\u2013piperaquine\u2013mefloquine versus arterolane\u2013piperaquine and artemether\u2013lumefantrine for the treatment of uncomplicated falciparum malaria in Kenyan children.Plasmodium falciparum malaria were recruited. Eligible patients were aged 2\u201312 years and had an asexual parasitaemia of 5000\u2013250\u2009000 parasites per \u03bcL. The exclusion criteria included the presence of an acute illness other than malaria, the inability to tolerate oral medications, treatment with an artemisinin derivative in the previous 7 days, a known hypersensitivity or contraindication to any of the study drugs, and a QT interval corrected for heart rate  longer than 450 ms. Patients were randomly assigned (1:1:1), by use of blocks of six, nine, and 12, and opaque, sealed, and sequentially numbered envelopes, to receive either arterolane\u2013piperaquine, arterolane\u2013piperaquine\u2013mefloquine, or artemether\u2013lumefantrine. Laboratory staff, but not the patients, the patients' parents or caregivers, clinical or medical officers, nurses, or trial statistician, were masked to the intervention groups. For 3 days, oral artemether\u2013lumefantrine was administered twice daily (target dose 5\u201324 mg/kg of bodyweight of artemether and 29\u2013144 mg/kg of bodyweight of lumefantrine), and oral arterolane\u2013piperaquine  and oral arterolane\u2013piperaquine\u2013mefloquine (mefloquine dose 8 mg/kg of bodyweight) were administered once daily. All patients received 0\u00b725 mg/kg of bodyweight of oral primaquine at hour 24. All patients were admitted to Kilifi County Hospital for at least 3 consecutive days and followed up at day 7 and, thereafter, weekly for up to 42 days. The primary endpoint was 42-day PCR-corrected efficacy, defined as the absence of treatment failure in the first 42 days post-treatment, of arterolane\u2013piperaquine\u2013mefloquine versus artemether\u2013lumefantrine, and, along with safety, was analysed in the intention-to-treat population, which comprised all patients who received at least one dose of a study drug. The 42-day PCR-corrected efficacy of arterolane\u2013piperaquine\u2013mefloquine versus arterolane\u2013piperaquine was an important secondary endpoint and was also analysed in the intention-to-treat population. The non-inferiority margin for the risk difference between treatments was \u22127%. The study is registered in ClinicalTrials.gov, NCT03452475, and is completed.In this single-centre, open-label, randomised, non-inferiority trial done in Kilifi County Hospital, Kilifi, coastal Kenya, children with uncomplicated P falciparum were screened, of whom 217 were randomly assigned to receive either arterolane\u2013piperaquine (n=73), arterolane\u2013piperaquine\u2013mefloquine (n=72), or artemether\u2013lumefantrine (n=72) and comprised the intention-to-treat population. The 42-day PCR-corrected efficacy after treatment with arterolane\u2013piperaquine\u2013mefloquine  was non-inferior to that after treatment with artemether\u2013lumefantrine . The 42-day PCR-corrected efficacy of arterolane\u2013piperaquine\u2013mefloquine was non-inferior to that of arterolane\u2013piperaquine . Vomiting rates in the first hour post-drug administration were significantly higher in patients treated with arterolane\u2013piperaquine  or arterolane\u2013piperaquine\u2013mefloquine  than in patients treated with artemether\u2013lumefantrine . Upper respiratory tract complaints , headache , and abdominal pain  were the most frequently reported adverse events. There were no deaths.Between March 7, 2018, and May 2, 2019, 533 children with This study shows that arterolane\u2013piperaquine\u2013mefloquine is an efficacious and safe treatment for uncomplicated falciparum malaria in children and could potentially be used to prevent or delay the emergence of antimalarial resistance.UK Department for International Development, The Wellcome Trust, The Bill & Melinda Gates Foundation, Sun Pharmaceutical Industries Plasmodium falciparum can emerge independently in Africa, as it did in Rwanda.P falciparum malaria in both children and adults across India and AfricaPlasmodium vivax malaria.Artemisinin-based combination therapies (ACTs) are first-line drugs for the treatment of uncomplicated falciparum malaria in all malaria endemic countries. ACTs have substantially contributed to the decrease in malaria transmission in the current millennium.Evidence before this studyPlasmodium falciparum and Plasmodium vivax malaria in both children and adults across India. There were no studies on triple antimalarial combinations including arterolane\u2013piperaquine. A study in healthy Thai adult volunteers found that exposure to artemisinin dihydroartemisinin was decreased when dihydroartemisinin\u2013piperaquine was combined with mefloquine. The TRACII trial showed that dihydroartemisinin\u2013piperaquine\u2013mefloquine and artemether\u2013lumefantrine\u2013amodiaquine were highly efficacious, safe, and well tolerated in patients with P falciparum malaria.We searched PubMed for articles published in English between database inception and Aug 31, 2020, using the search terms \u201carterolane\u201d AND \u201cmalaria\u201d, which resulted in 34 articles. In addition, we searched PubMed with the same date and language restrictions using the search terms \u201cmalaria\u201d AND (\u201ctriple ACT\u201d OR \u201cTACT\u201d), which resulted in 35 articles. Arterolane maleate is a synthetic, rapidly acting, potent ozonide antimalarial. The fixed-dose combination of arterolane\u2013piperaquine has been shown to be efficacious and safe for the treatment of Added value of this studyIn this study, we show that both arterolane\u2013piperaquine and arterolane\u2013piperaquine\u2013mefloquine are highly efficacious, safe, and well tolerated treatments for uncomplicated falciparum malaria in Kenyan children. The pharmacokinetic profile of arterolane was not affected by the addition of mefloquine to arterolane\u2013piperaquine.Implications of all the available evidenceDeploying arterolane-based triple-combination therapies could delay the development of antimalarial drug resistance against arterolane and its partner drugs, because the chance of parasites developing resistance to all three drugs is the product of the chance of developing resistance to each individual drug.P falciparum malaria. For patients with a positive rapid diagnostic test but no signs of severe or complicated malaria or other disease, written informed consent was obtained from their parent or guardian, after which patients were admitted to Kilifi County Hospital. Patients directly presenting with fever at Kilifi County Hospital were screened for P falciparum malaria by use of a rapid diagnostic test and a blood film.We did a single-centre, open-label, randomised, non-inferiority trial with children in Kilifi County Hospital in Kilifi, coastal Kenya . ParticiP falciparum infection, defined as a positive blood smear with asexual forms of P falciparum , an asexual parasitaemia of 5000\u2013250\u2009000 parasites per \u03bcL, and a fever (a tympanic temperature >37\u00b75\u00b0C or a history of fever within the past 48 h before enrolment). They were also able to take oral medication, were willing and able to comply with the study protocol for the duration of the study, and had a parent or guardian provide written informed consent. An independent witness was sought in case of an illiterate parent. The exclusion criteria were: signs of severe or complicated malaria according to WHO guidelines;Eligible participants were aged 2\u201312 years and had uncomplicated The protocol was approved by the Oxford Tropical Research Ethics Committee in the UK and the Kenya Medical Research Institute Scientific and Ethics Review Unit in Kenya. The trial was monitored collaboratively by the Mahidol-Oxford Tropical Medicine Research Unit (MORU) and Kenya Medical Research Institute Wellcome Trust Research Programme clinical trials support groups.The admitting clinician enrolled all participants, assigned them to the trial groups, and was involved with subsequent patient care and reviews. By use of block sizes of six, nine, and 12, and opaque, sealed, sequentially numbered envelopes, patients were randomly assigned (1:1:1) to receive either arterolane\u2013piperaquine, arterolane\u2013piperaquine\u2013mefloquine, or artemether\u2013lumefantrine. Randomisation sequences were prepared by use of a computer code, with a computer seed included in the program to allow for reproducibility, before the start of the trial by the trial statistician (MM), who conducted the final analysis. Using the randomisation sequences, the envelopes containing treatment allocation information were prepared by the MORU clinical trial support group. All samples were deidentified and the laboratory staff were masked to group assignment. The patients, the patients' parents or caregivers, clinical or medical officers, nurses, and the trial statistician were not masked to the intervention groups. All treatments were directly observed.For 3 consecutive days, oral artemether\u2013lumefantrine was administered twice daily with a fatty snack or drink (containing at least 2 g of fat) to maximise absorption, whereas oral arterolane\u2013piperaquine and oral arterolane\u2013piperaquine\u2013mefloquine were administered once daily with a non-fatty snack or water.In case of vomiting within the first 30 min after study drug administration, a full dose of the study drug was readministered; in case of vomiting between 30 min and 60 min post-administration, a half dose of the study drug was readministered. All patients were admitted to Kilifi County Hospital for at least 3 consecutive days and followed up at day 7 and, thereafter, weekly for up to 42 days. During each day of admission and each day of follow-up, a standardised symptom questionnaire, physical examination, and a measurement of vital signs were done. A 12-lead electrocardiograph was done at screening, baseline, hour 4, hour 24, hour 28, hour 48, and hour 52. For this study, the QTc interval was calculated by use of both Bazett's correction method and Fridericia's correction method (QTcF). Biochemistry and full blood count measurements were done at baseline, day 3, day 7, and day 28. If baseline biochemistry values were abnormal (grade 3 or 4) or the QTc interval was prolonged by more than 60 ms at any timepoint compared with baseline, arterolane\u2013piperaquine\u2013mefloquine or arterolane\u2013piperaquine were discontinued and replaced by artemether\u2013lumefantrine.P falciparum parasite densities were assessed microscopically at screening, baseline, hours 4, 6, 8, and 12, and thereafter every 6 h until two consecutive blood films were negative. In every participant, parasite densities were also assessed at hours 24, 48, and 72 (even if two consecutive negative blood films were seen before these timepoints). A venous or capillary blood film was examined at each weekly follow-up until day 42 to detect recurrent infection. A recurrent infection was defined as blood smear positivity for asexual P falciparum. According to local guidelines, all recurrent infections were treated with artemether\u2013lumefantrine. Whole blood was collected on dried blood spots at baseline and the day of recurrent infections.Asexual P falciparum resistance to artemisinin (Pfkelch13 non-synonymous mutations) and piperaquine (Pfplasmepsin2/3 gene amplification) were identified by use of the SPOTMalaria V2 platform, which uses a multiplexed amplicon sequencing method, implemented on Illumina sequencers. For PCR correction, DNA extraction and purification were done using the standardised kit . Primer sequences, PCR amplification, and analyses were based on methods described previously.msp1, msp2, and glurp alleles matched those that were present at baseline and as a reinfection if there were one or more allelic differences.Genetic markers of The primary endpoint was the 42-day PCR-corrected efficacy, defined as the absence of treatment failure in the first 42 days after treatment, of arterolane\u2013piperaquine\u2013mefloquine versus artemether\u2013lumefantrine. The PCR-corrected efficacy denotes the absence of recrudescent infections during follow-up. The PCR-uncorrected efficacy denotes the absence of recrudescence and reinfections during follow-up. The 42-day PCR-corrected efficacy of arterolane\u2013piperaquine\u2013mefloquine versus arterolane\u2013piperaquine was an important secondary endpoint.Other prespecified secondary endpoints were parasite clearance half-lives, slide-positive parasitaemia at day 3, fever clearance times, 28-day PCR-corrected efficacy and 28-day and 42-day PCR-uncorrected efficacy, the proportion of patients completing a full treatment course, vomiting rates within 1 h of study drug administration, the prevalence of adverse events and serious adverse events, changes in heart rate at any timepoint, prolongation of the QTc interval (>60 ms or >500 ms) at hours 4, 24, 28, 48, and 52, and the pharmacokinetic profile of arterolane. Detailed results from the genomics and transcriptomics analyses will be reported separately.Serious adverse events were reported to the sponsor, the appropriate ethics committees, the regulator, and an independent data and safety monitoring board within 24 h of awareness by the study team. Serious adverse events were defined as per the International Council for Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use guidelines for good clinical practice. The data and safety monitoring board met before the start of the trial and evaluated unblinded safety data after recruitment of 30 patients and then 100 patients. All adverse events were graded according to the Division of Acquired Immune Deficiency Syndrome Table for Grading the Severity of Adult and Paediatric Adverse Events , in which grade 1 is mild, grade 2 is moderate, grade 3 is severe, and grade 4 is potentially life-threatening.P falciparum infection were included in the treatment failure group in the intention-to-treat analysis. Patients who presented with a malaria reinfection, withdrew consent, or were lost to follow-up were included in the treatment success group in the intention-to-treat analysis. Patients with any of these events or in whom the study drug was replaced by artemether\u2013lumefantrine  were excluded from the per-protocol analysis and were censored from the Kaplan-Meier survival analysis. We repeated our analyses of the primary outcome and secondary efficacy outcomes in the per-protocol population.We hypothesised that the 42-day PCR-corrected efficacy of arterolane\u2013piperaquine\u2013mefloquine would be non-inferior to artemether\u2013lumefantrine. Based on previous experience elsewhere in Africa, we assumed that the efficacy of artemether\u2013lumefantrine in Kenya was 98%.t test. The incidences of vomiting within the first hour after drug administration were compared between study groups by use of a \u03c72 test. p-values are given and statistical significance was declared at 5%. All aforementioned analyses were done in Stata, version 15.Parasite clearance half-lives were estimated by use of the Worldwide Antimalarial Resistance Network's parasite clearance estimator.max) and the time to reach the maximum concentration (Tmax) were derived from the observed data. The terminal elimination rate constant (\u03bb) was estimated by use of the software's best fit functionality . The elimination half-life was calculated as ln(2/\u03bb). Exposure (area under the concentration time curve [AUC]) was calculated with the trapezoidal method by use of the linear method for ascending concentrations and the log-linear method for descending concentrations. Exposure was calculated to the last timepoint (AUClast) and, by use of \u03bb to extrapolate from the last observed concentration, to infinity (AUC\u221e). Standard equations were used to calculate apparent elimination clearance and apparent volume of distribution. To evaluate the potential pharmacokinetic differences between arterolane, piperaquine, and mefloquine, concentrations at each sampling timepoint (including the 48 h and 72 h samples) were compared between groups by use of a Mann-Whitney U-test in GraphPad Prism, version 8.2.1. A data and safety monitoring board evaluated unblinded safety data after recruitment of 30 patients and then 100 patients. The study is registered in ClinicalTrials.gov, NCT03452475.Collected pharmacokinetic data were analysed by use of a non-compartmental approach in Pkanalix, version 2019R2, to assess differences in the pharmacokinetic profile of arterolane with or without mefloquine coadministration. We assumed that arterolane, which is not locally available, was undetectable at baseline (at 0 h) and concentrations less than the lower limit of quantification were replaced with a value equal to half the lower limit of quantification. Because of the sparse sampling design, a non-compartmental analysis was done on the median concentration at each sampling timepoint  comparing the children receiving arterolane\u2013piperaquine with the children receiving arterolane\u2013piperaquine\u2013mefloquine. The analysis was done only after the first dose. The maximum concentration , masked to the treatment group. Arterolane\u2013piperaquine (Synriam) was provided for the study by Sun Pharmaceutical Industries. Other study drugs were purchased against their commercial value. The funders of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report.P falciparum were screened  for patients treated with arterolane\u2013piperaquine, 100%  for patients treated with arterolane\u2013piperaquine\u2013mefloquine, and 96%  for patients treated with artemether\u2013lumefantrine . The 42-P falciparum. When reconsidering and imputing these two patients as having reinfections, the 42-day PCR-corrected efficacy of artemether\u2013lumefantrine increases to 99% .The two patients with recurrent infections for which PCR correction was not possible were treated with artemether\u2013lumefantrine. These infections were interpreted conservatively as recrudescent, although it is more probable that these were actually reinfections with In the intention-to-treat population, the 42-day PCR-uncorrected efficacy was 90% for arterolane\u2013piperaquine, 78% for arterolane\u2013piperaquine\u2013mefloquine, and 50% for artemether-lumefantrine , reflectParasite clearance half-lives could be calculated in 208 of 217 patients . Half-liPfkelch13 genotypes for 211 (97%) of 217 baseline samples. Of these, 203 (96%) did not carry any non-synonymous mutations. The Ala578Ser mutation, which is present throughout Africa and not associated with artemisinin resistance, was found in four samples. The other four samples were from mixed infections and contained rare, non-synonymous mutations , whose effect on artemisinin sensitivity is unknown. Amplification of the Pfplasmepsin2/3 gene was found in none of the 103 samples in which amplification status could be determined.We obtained The proportions of patients that completed a full treatment course were similar between treatments . ReasonsVomiting rates were significantly higher in patients treated with arterolane\u2013piperaquine (p=0\u00b70013) or arterolane\u2013piperaquine\u2013mefloquine (p=0\u00b70006) than in patients treated with artemether\u2013lumefantrine . All pat. QTcF and QTcB intervals longer than 500 ms were not observed in any patient  or arterolane\u2013piperaquine\u2013mefloquine  than after treatment with artemether\u2013lumefantrine  patient . The pre patient . Changes patient . Heart r patient . The pre patient .Headache, abdominal pain, and symptoms of upper respiratory tract infection were the most frequently reported adverse events . OverallThe most common biochemical adverse event was an abnormality in plasma creatinine concentrations . Mild-toA total of six serious adverse events fulfilling predefined criteria were reported, of which four were in patients treated with arterolane\u2013piperaquine, one was in a patient treated with arterolane\u2013piperaquine\u2013mefloquine, and one was in a patient treated with artemether\u2013lumefantrine . Grade 4last, Cmax, Tmax, the apparent elimination clearance, the apparent volume of distribution, and the elimination half-life) are shown in the The median concentration\u2013time profiles for the two groups given arterolane are shown in To our knowledge, this clinical trial is the first to compare the efficacy and safety of the triple antimalarial combination therapy, arterolane\u2013piperaquine\u2013mefloquine, with arterolane\u2013piperaquine and artemether\u2013lumefantrine for the treatment of uncomplicated falciparum malaria in Kenyan children. In addition, the study used a new weight-based dosing schedule for arterolane\u2013piperaquine. The efficacy of arterolane\u2013piperaquine\u2013mefloquine was non-inferior to that of artemether\u2013lumefantrine and arterolane\u2013piperaquine, and all three treatment combinations were well tolerated.P falciparum, was low for treatment with artemether\u2013lumefantrine, as result of a short post-treatment prophylactic effect, as has been described previously.Related to the much shorter plasma half-life of lumefantrine  compared with mefloquine  and piperaquine , the 42-day PCR-uncorrected efficacy, which includes reinfections with Prolongation of the QTc interval at hour 52 was significantly greater with arterolane\u2013piperaquine or arterolane\u2013piperaquine\u2013mefloquine than with artemether\u2013lumefantrine. Importantly, the addition of mefloquine to arterolane\u2013piperaquine did not further increase the QTc interval, which supports similar observations comparing QTc intervals after the addition of mefloquine to dihydroartemisinin\u2013piperaquine.Combining arterolane with two partner drugs in a triple therapy could preserve the efficacy of each individual drug, as the chance of parasites developing resistance to all three drugs is the product of the chance of developing resistance to each individual drug, assuming there are no interactions between resistance mechanisms. The cost of a triple antimalarial therapy will be slightly higher than the cost of a standard ACT. However, these increases in costs should be considered against the costs associated with the emergence of multidrug-resistant malaria, which would probably increase the morbidity and mortality of malaria and could set back successes in malaria control and elimination.Pfkelch13-mutated (artemisinin-resistant) strains, suggesting cross-resistance.P falciparum infections.Like the artemisinins, synthetic ozonides contain an endoperoxide bridge considered necessary for their parasiticidal potency, and in-vitro studies have shown reduced parasiticidal potency of arterolane (OZ277) in P falciparum.Our study had several limitations. The unblinded design could have affected the assessment of adverse events and the attribution of relatedness to the study drugs. However, objective measures, such as parasite clearance half-lives, treatment efficacy, electrocardiograph readings, and laboratory outcomes, are very unlikely to have been affected by the unblinded design of our study. Because our study relied on self-reporting of symptoms and all participants were young children, it is probable that the prevalence of complaints is an underestimation. The number of patients recruited to our study was small, and it is possible that less frequent side-effects were not identified. Furthermore, the study was done in a hospital setting, and each drug dose was administered and observed by a staff member, which might have resulted in a higher study drug adherence than that expected in a non-supervised setting. The most common reasons for study exclusion were having a parasitaemia out of our prespecified range or a negative blood smear. Future studies could evaluate the use of these drugs in clinical settings where diagnosis relies on rapid diagnostic tests because of the unavailability of microscopy. Further studies assessing treatment adherence in a non-clinical, non-supervised setting are needed. In addition, the relevance of the increased rates of vomiting we observed with triple ACTs compared with artemether\u2013lumefantrine, with respect to treatment efficacy, should be assessed. The addition of primaquine is unlikely to have affected the comparison of efficacy and safety between the three study groups because primaquine does not affect asexual-stage In conclusion, dosed according to weight, the efficacy of arterolane\u2013piperaquine\u2013mefloquine was non-inferior to that of artemether\u2013lumefantrine and arterolane\u2013piperaquine, and all combinations were safe and well tolerated, in the treatment of uncomplicated falciparum malaria in Kenyan children.dgc@kemri-wellcome.org. The data request form can be found online on the KWTRP Harvard Dataverse repository. Individual patient data will be available under managed access through the data access committee. There will be no restrictions on analysis; all requests will be evaluated by the data access committee for a decision in line with consent for data sharing and findability, accessibility, interoperability, and reusability (known as FAIR) principles. The mechanism will be open access and managed via email (dgc@kemri-wellcome.org) for the data requests to be considered by the data access committee. Related documents, such as the study protocol, statistical analysis plan, and informed consent form, will be made available (with publication) on request to the corresponding author . The data will be open access for the informed consent form, protocol, and statistical analysis plan. Once the data are in the Harvard Dataverse repository, a request for these documents can be made from the dataset page.With publication, deidentified, individual participant data that underlie this Article, along with a data dictionary describing variables in the dataset, will be made available to researchers whose proposed purpose of use is approved by the Kenya Medical Research Institute Wellcome Trust Research Programme (KWTRP)data access committee. Data will be shared directly or through access on the Harvard Dataverse repository. To request the dataset directly, please send a signed data request form to We declare no competing interests."}
+{"text": "In Phase II/III randomized controlled clinical trials for the treatment of acute uncomplicated malaria, pyronaridine\u2013artesunate demonstrated high efficacy and a safety profile consistent with that of comparators, except that asymptomatic, mainly mild-to-moderate transient increases in liver aminotransferases were reported for some patients. Hepatic safety, tolerability, and effectiveness have not been previously assessed under real-world conditions in Africa.Plasmodium falciparum reinfection, clinical effectiveness at day 28 was 98.6%  98.3 to 98.9) in the per-protocol population. There was no indication that comorbidities or malnutrition adversely affected outcomes. The key study limitation was that postbaseline clinical biochemistry was only evaluated when clinically indicated.This single-arm, open-label, cohort event monitoring study was conducted at 6 health centers in Cameroon, Democratic Republic of Congo, Gabon, Ivory Coast, and Republic of Congo between June 2017 and April 2019. The trial protocol as closely as possible resembled real-world clinical practice for the treatment of malaria at the centers. Eligible patients were adults or children of either sex, weighing at least 5 kg, with acute uncomplicated malaria who did not have contraindications for pyronaridine\u2013artesunate treatment as per the summary of product characteristics. Patients received fixed-dose pyronaridine\u2013artesunate once daily for 3 days, dosed by body weight, without regard to food intake. A tablet formulation was used in adults and adolescents and a pediatric granule formulation in children and infants under 20 kg body weight. The primary outcome was the hepatic event incidence, defined as the appearance of the clinical signs and symptoms of hepatotoxicity confirmed by a >2\u00d7 rise in alanine aminotransferase/aspartate aminotransferase (ALT/AST) versus baseline in patients with baseline ALT/AST >2\u00d7 the upper limit of normal (ULN). As a secondary outcome, this was assessed in patients with ALT/AST >2\u00d7 ULN prior to treatment versus a matched cohort of patients with normal baseline ALT/AST. The safety population comprised 7,154 patients, of mean age 13.9 years (standard deviation (SD) 14.6), around half of whom were male . Patients experienced 8,560 malaria episodes; 158 occurred in patients with baseline ALT/AST elevations >2\u00d7ULN. No protocol-defined hepatic events occurred following pyronaridine\u2013artesunate treatment of malaria patients with or without baseline hepatic dysfunction. Thus, no cohort comparison could be undertaken. Also, as postbaseline clinical chemistry was only performed where clinically indicated, postbaseline ALT/AST levels were not systematically assessed for all patients. Adverse events of any cause occurred in 20.8%  of patients, most frequently pyrexia  and vomiting . Adjusting for Pyronaridine\u2013artesunate had good tolerability and effectiveness in a representative African population under conditions similar to everyday clinical practice. These findings support pyronaridine\u2013artesunate as an operationally useful addition to the management of acute uncomplicated malaria.ClinicalTrials.govNCT03201770. Gaston Tona Lutete and co-workers report on safety and effectiveness of the antimalarial drug pyronaridine-artesunate in African countries. Plasmodium falciparum malaria.Pyronaridine\u2013artesunate has shown high efficacy and an acceptable safety profile in randomized controlled clinical trials for the treatment of acute uncomplicated Transient, asymptomatic, mostly mild-to-moderate increases in liver aminotransferases have been observed with pyronaridine\u2013artesunate in some patients.We assessed pyronaridine\u2013artesunate hepatic safety, tolerability, and effectiveness in adults and children for the treatment of acute uncomplicated malaria under real-world conditions in Africa, including patients with elevated baseline liver aminotransferases.In a single-arm, open-label, cohort event monitoring study conducted across 5 African countries, patients of any age, weighing >5 kg with acute uncomplicated malaria received fixed-dose pyronaridine\u2013artesunate once daily for 3 days.Patient exclusion and inclusion criteria were consistent with the approved prescribing information, and patients with elevated baseline liver enzymes could be treated under the protocol.The study included 8,560 malaria episodes, 158 of which occurred in patients with baseline alanine aminotransferase (ALT) or aspartate aminotransferase (AST) abnormalities.P. falciparum malaria.There were no hepatic events following pyronaridine\u2013artesunate treatment. Tolerability was as reported in previous studies, and treatment effectiveness at day 28 was 98.6% in patients with acute uncomplicated Pyronaridine\u2013artesunate was an operationally effective and well-tolerated antimalarial therapy in this unselected population of patients from 5 African countries. Africa carries the greatest malaria burden, with an estimated 215 million cases in 2019 and 94% of global deaths, mainly in children [Plasmodium falciparum are established in Southeast Asia, and where these strains are also resistant to partner drugs, high clinical failure rates are reported [Kelch 13 artemisinin resistance mutations have been verified in Africa [Artemisinin-based combination therapy (ACT) is recommended for first-line therapy of acute uncomplicated malaria, with several artemisinin and partner drug combinations deployed worldwide. Artemisinin-resistant reported . De novon Africa , and parn Africa .P. falciparum [Pyronaridine\u2013artesunate has demonstrated high efficacy in acute uncomplicated malaria in both Africa and Asia \u201318, incllciparum \u201310. In clciparum ,11,14. Alciparum ,11,14. Alciparum . Althouglciparum ,11,14. SAlthough the clinical relevance of the observed transient asymptomatic liver aminotransferase elevations is unclear, all pyronaridine\u2013artesunate clinical trial populations to date have excluded patients with underlying hepatic dysfunction. Consequently, it is currently unknown if underlying hepatic dysfunction will progress to clinical manifestations of liver impairment with pyronaridine\u2013artesunate. It is important, therefore, to clarify the risk\u2013benefit of pyronaridine\u2013artesunate in an unselected, representative population treated under normal clinical conditions in accordance with current drug prescribing recommendations.This large clinical trial was conducted as part of the postmarketing requirements specified by the European Medicines Agency (EMA) for pyronaridine\u2013artesunate. The pyronaridine\u2013artesunate-approved product prescribing information does not require prior measurement of liver enzymes before treatment . The priIn summary, the aim of this study was to characterize pyronaridine\u2013artesunate hepatic safety, tolerability, and effectiveness in a real-world setting in a clinically relevant population across a network of representative study sites in Central and West Africa.This single-arm, open-label cohort event monitoring study was conducted at 6 health centers in Cameroon, Democratic Republic of Congo, Gabon, Ivory Coast, and Republic of Congo between 22 June 2017 and 10 April 2019. The protocol adhered to Good Clinical Practice, the Declaration of Helsinki, and relevant regulations in the respective countries and was reviewed and accepted as a valid postmarketing safety assessment by the EMA. The authors vouch for the accuracy and completeness of the data and the fidelity of the trial conduct, analysis, and reporting to the protocol. A copy of the protocol and statistical analysis plan is provided in the Supporting information . This stAll patients, or their parents/guardians, provided written informed consent, and children able to understand the study gave signed assent. The protocol was conducted in compliance with the Declaration of Helsinki (amended Tokyo 2004) and the directives in the respective countries, in particular concerning the submission to Ethics Committees and the protection of personal data. Additionally, the study was conducted in accordance with the principles of the Good Clinical Practice according to the International Council for Harmonization (ICH) Harmonized Tripartite Guidance (CPMP/ICH/135/95). The ethics committee at each study site granted ethical approval as follows. Cameroon: Comit\u00e9 National d\u2019Ethique de la Recherche pour la Sant\u00e9 Humaine; Democratic Republic of Congo: Comit\u00e9 d\u2019Ethique de l\u2019Universit\u00e9 Protestante au Congo; Gabon: Comit\u00e9 Nationale d\u2019Ethique et de la Recherche Scientifique and Comit\u00e9 d\u2019Ethique Institutionnel Albert Schweitzer Hospital, Lambar\u00e9n\u00e9; Ivory Coast: Comit\u00e9 Nationale d\u2019Ethique et de la Recherche Scientifique and Comit\u00e9 National d\u2019Ethique des Sciences de la Vie et de Sant\u00e9; and Republic of Congo: Institutional Ethics Committee of the Fondation Congolaise pour la Recherche M\u00e9dicale.Eligible patients were male or female, of any age, with body weight of at least 5 kg. All patients had acute uncomplicated malaria and a fever or history of fever within the previous 24 hours and/or anemia. Exclusion criteria were severe malaria, clinical signs or symptoms of hepatic injury , known severe liver disease , known allergy to artemisinin and/or pyronaridine, known pregnancy, lactation, treatment with pyronaridine\u2013artesunate in the previous 28 days, or if the patient was considered by the investigator to be at particular risk.Fixed-dose pyronaridine\u2013artesunate  was administered once daily for 3 days with 2 formulations provided according to body weight and without regard to food intake. Daily doses for pyronaridine\u2013artesunate tablets (180:60 mg) were 1 (\u226520 to <24 kg), 2 (\u226524 to <45 kg), 3 (\u226545 to <65 kg), or 4 (\u226565 kg); and for granule sachets (60:20 mg) were 1 (\u22655 to <8 kg), 2 (\u22658 to <15 kg), or 3 (\u226515 to <20 kg). To reflect real-world practice, patients could be included more than once in the study, following a 28-day washout period between consecutive pyronaridine\u2013artesunate treatments. Patients received the first pyronaridine\u2013artesunate dose under supervision and were provided with the remaining doses to be taken unsupervised on the subsequent 2 consecutive days.At enrollment on day 0, eligible patients underwent physical examination, with demographic data and medical history noted.Blood samples (5 mL) were collected for hematology, clinical chemistry, and confirmation of malaria diagnosis. Blood samples were stored for retrospective analysis of baseline clinical chemistry and viral hepatitis assessment: hepatitis A, B, C, and E  and hepatitis delta antibody if positive for hepatitis B. All laboratory analysis was performed at the site where the patient was recruited by technically certified staff, and results were validated by external independent laboratories. Standard quality control procedures were in place, including daily internal controls and the quality control process assessed by the clinical monitors.P. falciparum recurrence, PCR genotyping was used to differentiate between recrudescence and reinfection by comparing blood spot samples taken at baseline versus recurrence, using published criteria [Malaria was diagnosed using a rapid diagnostic test (RDT) or Giemsa-stained thick blood smear. The RDT was consistent with local treatment guidelines and used according to the manufacturer\u2019s instructions by staff trained according to the local standard operating procedure . Microsccriteria .Adverse events were assessed without knowledge of baseline ALT/AST. Patients were to present at the clinic should any adverse event occur within 28 days of pyronaridine\u2013artesunate dosing. Home follow-up visits were conducted by CHWs on days 7 and 28 to assess treatment adherence, clinical condition, adverse events, and concomitant medications. CHWs were trained before the study and retrained during the study by clinical investigators in the assessment of safety and tolerability using questionnaire forms. In addition, investigators accompanied CHWs to field visits to assure quality and consistency in adverse event assessment. All clinical information provided by CHWs was assessed by clinical investigators to ensure accurate medical recording of all safety and tolerability signs. CHWs were provided with pictorial information specifically identifying the signs and symptoms of liver injury. Furthermore, any potential hepatotoxic signs or symptoms noted by CHWs were specifically discussed with the study doctor, with patients referred to the clinic if necessary.Hepatic events prompted blood sampling for clinical chemistry and hematology and a full hepatitis panel. Assessment at the health facility was triggered by serious or severe adverse events or adverse events of special interest, i.e., hepatotoxicity , serious liver reactions, and hypersensitivity . Female participants had a urinary pregnancy test at day 28, with provision in the case of pregnancy for extended follow-up until the birth or other outcome.The safety population included all enrolled patients who received at least 1 dose of study medication for any malaria episode. The primary outcome was the incidence of hepatic events in the safety population following pyronaridine\u2013artesunate treatment of malaria patients with baseline ALT or AST >2\u00d7ULN, defined as clinical signs and symptoms of possible hepatotoxicity, i.e., fatigue, nausea, abdominal pain, itching, or signs of jaundice , associated with a rise in ALT/AST >2\u00d7 the baseline value.2.Safety outcomes were the incidence of adverse events in the safety population overall and categorized by normal/abnormal/unknown baseline ALT/AST, any underlying hepatic disease identified on the hepatitis panel, HIV status (where known), age, sex, weight, and nutritional status. Malnutrition was defined for patients <5 years old as a mid-upper arm circumference <115 mm, in patients aged \u22655 years to \u226419 years using the World Health Organization age- and sex-specific body mass index (BMI)-for-age z-scores , and forTreatment adherence was self-assessed by patients and also estimated by CHWs based on empty packaging provided by patients versus the number of tablets/sachets dispensed.P. falciparum reinfection. Day 28 effectiveness subgroup analysis was planned for by country, dosing (granules versus tablets), malnutrition, and age. Additional outcomes were time between malaria episodes and the frequency of repeat episodes. The intention-to-treat population included all malaria episodes treated with at least 1 dose of study medication with confirmed positive parasitemia at baseline. The per-protocol population included all malaria episodes in the intention-to-treat population in patients who were treated with a full course of study medication (adherence per patient self-assessment), had a day 28 effectiveness endpoint, did not vomit after study drug administration (except where vomiting occurred after the first dose and dosing was successfully repeated), and who had no prior or concomitant medication, which had an antimalarial effect and so could interfere with the treatment outcome. All outcomes and trial populations were prespecified . To better mimic the clinical reality, patients could be included more than once in the study. For the outcome of clinical effectiveness, each malaria episode was treated as an independent event, with no adjustment for repeated measures. This was to better reflect the endpoint of operational effectiveness, i.e., to include patient-related as well as drug-related contributors and to be consistent with previous analyses of randomized controlled trials of repeated treatment with pyronaridine\u2013artesunate ,25. For Of the 8,609 malaria episodes that met study eligibility criteria, 8,560 were treated with at least 1 dose of study medication, comprising the safety population . Of the There were no malaria episodes with protocol-defined hepatic events in any patient in the safety population. No further planned secondary analyses of the primary endpoint could, therefore, be conducted.Adverse events of any cause occurred in 20.8%  of patients, most frequently pyrexia  and vomiting  . There wAdverse event frequency was not affected by sex, weight, nutritional status, HIV status, or first versus repeated malaria episode . The natHigher adverse event rates were noted for patients aged under 1 year  and those \u226518 years of age  versus those aged 5 to <18 years   Tables. There were 9.6%  of patients with adverse events considered related to pyronaridine\u2013artesunate treatment, most commonly vomiting  and headache  . TreatmeSerious adverse events were reported in 0.4%  of patients, with the most frequent being malaria  and anemia  . Four paThere were 3 deaths during the study , none of which were considered related to pyronaridine\u2013artesunate treatment. The case of severe malaria occurred in a 6-year-old girl from the Democratic Republic of Congo who was enrolled with acute uncomplicated malaria, presenting with asthenia and vomiting. She was diagnosed with severe malaria 1.5 hours following the first dose of pyronaridine\u2013artesunate and admitted to hospital after symptoms of asthenia worsened. Pyronaridine\u2013artesunate was discontinued, and the patient treated with intravenous quinine. She also received intravenous phenobarbital and diazepam for convulsions and cefotaxime and gentamycin for suspected meningitis, but her condition rapidly worsened, and she unfortunately died later the same day. As the patient died of severe malaria within 24 hours of her first dose of pyronaridine\u2013artesunate, it is usual to consider the cause as disease progression, and so the death was not attributed to drug treatment. The death from diarrhea occurred in a 66-year-old man who received 6 pyronaridine\u2013artesunate tablets over 3 days, who died 29 days after the first dose of study medication from severe diarrhea, with no further information available. The other death was that of a 9-year-old boy who drowned in a river 12 days after the first administration of pyronaridine\u2013artesunate.N = 7,154); in all cases, this was hypersensitivity occurring only in 1 episode: 6 occurred in the first episode , and the other 3 occurred for the first time in episodes 2, 3, and 4, respectively . All patients were aged \u22655 years with normal baseline ALT/AST, 4 had urticaria, and 1 each had maculo-papular rash, papular rash, hypersensitivity , lip swelling, and eye pruritis; all were of mild-to-moderate intensity.An adverse event of special interest occurred in 9 patients . Plasmodium ovale (n = 70) and Plasmodium malariae (n = 55) occurred as mixed Plasmodium infections; 1 P. ovale/P. malariae coinfection and 1 Plasmodium vivax mono-infection were identified  in the per-protocol population and 90.9%  in the intention-to-treat population. The unadjusted cure rate in the per-protocol population was 93.2%  and 85.9%  in the intention-to-treat population . There were no hepatic events in 8,560 episodes of uncomplicated Pyronaridine\u2013artesunate efficacy and safety have been already demonstrated in multicenter, randomized, controlled clinical trials conducted for regulatory purposes in a selected patient population. In those studies, raised ALT >5\u00d7ULN was more frequently observed with pyronaridine\u2013artesunate versus comparators, but clinical signs and symptoms of hepatotoxicity were not manifest and all patients recovered without sequelae or intervention ,14\u201318,25There are some key limitations to this study as designed. Firstly, this study aimed to assess whether there was any clinical evidence of hepatotoxicity following pyronaridine\u2013artesunate administration according to the approved prescribing information. There is no requirement to measure ALT/AST in patients without signs or symptoms of liver dysfunction prior to treatment in normal clinical practice. Thus, the current study was explicitly designed to identify clinical hepatic events only. Although this reflects normal practice and resource availability in the study setting, not all patients with hepatitis will manifest clinical symptoms, and our study does not exclude the possibility of occurrences of subclinical liver injury. The use of the retrospective analysis of baseline blood samples was employed to confirm that patients with AST/ALT >2\u00d7ULN had, in fact, been recruited and to allow comparison with a matched cohort of patients with normal baseline ALT/AST (secondary endpoint). Postbaseline clinical biochemistry was only conducted where clinically indicated, for example, for biochemical confirmation of any clinically suspected signs of hepatotoxicity. In fact, postbaseline biochemistry for any reason was indicated in only 7 patients. Consequently, no conclusions can be drawn on the impact of pyronaridine\u2013artesunate on postbaseline ALT/AST levels for patients with elevated baseline values. Secondly, in line with the prescribing information for pyronaridine\u2013artesunate, patients with clinical signs or symptoms of hepatic injury  or known severe liver disease  were excluded . Thus, nRandomized controlled clinical trials of pyronaridine\u2013artesunate provide an extensive safety database compared to a number of different antimalarial drugs, and the safety profile is well documented ,14\u201318,25The pharmacodynamic outcome of this trial was clinical effectiveness under real-world conditions, whereas previous randomized clinical trials reported clinical efficacy as defined using the standard WHO outcome of adequate clinical and parasitological response (ACPR) . Across In the per-protocol population, the difference between PCR-adjusted effectiveness  and unadjusted effectiveness  suggests that around 5% of patients were reinfected within 28 days of pyronaridine\u2013artesunate treatment. Pyronaridine has a half-life of about 14 to 18 days , and posP. falciparum malaria.In summary, in this large clinical trial, conducted in a real-world setting in a representative population of African malaria patients, pyronaridine\u2013artesunate had excellent tolerability, safety, and effectiveness, and no clinically evident hepatic events occurred in patients with or without ALT/AST elevations at baseline. The study supports pyronaridine\u2013artesunate as an operationally valuable therapy for the treatment of acute uncomplicated S1 STROBE Checklist(PDF)Click here for additional data file.S1 Protocol and SAP(PDF)Click here for additional data file.S1 Methods(PDF)Click here for additional data file.S1 Table(PDF)Click here for additional data file.S2 Table(PDF)Click here for additional data file.S3 Table(PDF)Click here for additional data file.S4 Table(PDF)Click here for additional data file.S5 Table(PDF)Click here for additional data file.S6 Table(PDF)Click here for additional data file.S7 Table(PDF)Click here for additional data file.S8 Table(PDF)Click here for additional data file.S9 Table(PDF)Click here for additional data file.S10 Table(PDF)Click here for additional data file.S11 Table(PDF)Click here for additional data file.S1 Fig(PDF)Click here for additional data file."}
+{"text": "Integrated drug efficacy surveillance (iDES) was formally introduced nationally across Thailand in fiscal year 2018 (FY2018), building on a history of drug efficacy monitoring and interventions. According to the National Malaria Elimination Strategy for Thailand 2017\u20132026, diagnosis is microscopically confirmed, treatment is prescribed, and patients are followed up four times to ensure cure.Plasmodium falciparum and 28\u00a0days for Plasmodium vivax. The primary outcome was the crude drug efficacy rate, estimated using Kaplan\u2013Meier methods, at day 42 for P. falciparum treated with dihydroartemisinin\u2013piperaquine plus primaquine, and day 28 for P. vivax treated with chloroquine plus primaquine; day 60 and day 90 efficacy were secondary outcomes for P. vivax.Routine patient data were extracted from the malaria information system for FY2018\u2013FY2020. Treatment failure of first-line therapy was defined as confirmed parasite reappearance within 42\u00a0days for P. falciparum malaria and at day 28 for P. vivax malaria has been increasing, with FY2020 follow-up rates of 61.5% and 57.2%, respectively. For P. falciparum malaria, day 42 efficacy in FY2018 was 92.4% (n\u2009=\u2009249), in FY2019 93.3% (n\u2009=\u2009379), and in FY2020 98.0% (n\u2009=\u2009167). Plasmodium falciparum recurrences occurred disproportionally in Sisaket Province, with day 42 efficacy rates of 75.9% in FY2018 (n\u2009=\u200959) and 49.4% in FY2019 (n\u2009=\u200949), leading to an update in first-line therapy to pyronaridine\u2013artesunate at the provincial level, rolled out in FY2020. For P. vivax malaria, day 28 efficacy (chloroquine efficacy) was 98.5% in FY2018 (n\u2009=\u20092048), 99.1% in FY2019 (n\u2009=\u20092206), and 99.9% in FY2020 (n\u2009=\u20092448), and day 90 efficacy (primaquine efficacy) was 94.8%, 96.3%, and 97.1%, respectively.The proportion of patients with outcomes recorded at day 42 for In Thailand, iDES provided operationally relevant data on drug efficacy, enabling the rapid amendment of treatment guidelines to improve patient outcomes and reduce the potential for the spread of drug-resistant parasites. A strong case-based surveillance system, integration with other health system processes, supporting biomarker collection and molecular analyses, and cross-border collaboration may maximize the potential of iDES in countries moving towards elimination. Plasmodium falciparum have been reported throughout the region, associated with resistance to both artemisinins and their partner drugs ) , P. falciparum malaria and for 28\u00a0days for P. vivax malaria has been increasing each year since the national launch of iDES  of P. vivax cases had a day 28 outcome recorded  and Thai residents (64.8% [127/196]), but lower for short-term migrants (8.0% [2/25])  of long-term migrants, 62.7% (1821/2905) of Thai residents, and 8.9% (52/587) of short-term migrants followed up at day 28  in FY2018, to 77.2% (4103/5315) in FY2019, and 84.2% (3549/4215) in FY2020.P. falciparum (dihydroartemisinin\u2013piperaquine\u2009+\u2009primaquine) or P. vivax (chloroquine\u2009+\u2009primaquine) malaria, and who had at least one follow-up visit. Recurrences occurred in different provinces in different years, though Sisaket, Tak, and Yala had P. vivax recurrences in all three iDES years  was 92.4%  in FY2018, 93.3%  in FY2019, and 98.0%  in FY2020 (Fig.\u00a0P. falciparum cases in Sisaket and Ubon Ratchathani. Two cases received pyronaridine\u2013artesunate, with no recurrence. One received dihydroartemisinin\u2013piperaquine; after parasite reappearance at day 60, the case was successfully treated with pyronaridine\u2013artesunate.For P. vivax malaria, chloroquine/primaquine day 28 efficacy was at least 98% across all iDES years, suggesting good clinical efficacy for chloroquine (Fig.\u00a0For ine Fig.\u00a0. Day 60 ine Fig.\u00a0. Sisaketine Fig.\u00a0. Howeverine Fig.\u00a0.Fig. 7EfThailand has made significant progress towards malaria elimination, surpassing its 2020 milestone reductions for both malaria incidence and mortality . HoweverIt was clear that as malaria incidence declined in Thailand, TES would no longer be feasible in many provinces. The reorganization of the health system to target malaria elimination offered an opportunity to integrate drug efficacy surveillance. However, retroactive adaptation of the MIS to incorporate the iDES module was complex and has required continued refinement of the platform to adapt to changing epidemiology and corresponding data needs. The DVBD can examine the data in almost real time, thereby enabling limited resources to be targeted to the provinces, villages, or health facilities most in need of support to improve iDES compliance. The DVBD and its partners are working to further improve data visualizations to facilitate the data analyses most needed by subnational officers, such as health care workers interested in patient follow-up and treatment outcomes.In order to move from TES to iDES, it was necessary for Thailand to have a strong case-based surveillance system and the capacity for universal and supervised anti-malarial treatment. For countries that have not yet reached the elimination phase, TES remains the most appropriate drug efficacy surveillance method . For iDERapid diagnostic tests for malaria have been an invaluable tool in areas of high-to-moderate transmission to identify cases and direct appropriate therapy. However, iDES requires microscopic malaria diagnosis to confirm parasite clearance. Maintaining microscopy skills in an elimination setting is challenging as some laboratories see very few cases. With sustained support from external funding partners, Thailand has invested in a strong cadre of trained microscopists stationed throughout the country, and 33 professionals hold current expert certification from the WHO. However, other countries in the GMS and elsewhere may need to consider how to build these skills as malaria burden reduces, and as an iDES system becomes the recommended programme for case-based surveillance and follow-up.P. falciparum has been sporadic, with difficulties in managing their storage and processing. Thus, only crude efficacy rates are reported here, but in a country aiming for elimination it is expected that all recurrences are recrudescences. The collection of samples for molecular resistance markers has also been sub-optimal while subnational officers gain new skills. The DVBD anticipates enhanced molecular surveillance and streamlined processes to triangulate clinical and laboratory data as a National Reference Laboratory database is developed and with additional training for health workers and laboratory staff.Despite being included in the iDES protocol, the collection of dried blood spots for PCR analysis of Follow-up rates for iDES have been increasing steadily since its introduction, supported by a network of village health volunteers and through community education. High burden provinces in western Thailand Fig.\u00a0 have lowAdherence to the national treatment guidelines is necessary to optimize patient outcomes and to support malaria elimination. Although adherence rates have been improving, further progress will require additional resources. For example, Thailand\u2019s village health volunteers are key in accessing remote locations and engaging with patients on a personal level, and additional training of these health workers is planned in FY2021, alongside iDES capacity building. In Sisaket, iDES identified issues related to re-treatment with first-line therapy following failure, leading to subsequent treatment failure. These repeated treatment failures are programmatic and may result from stock-outs of second-line anti-malarial therapies, patients presenting at different clinics, or patient or prescriber choice. Being able to find and interrogate these cases allows interventions to be targeted at the causes of non-adherence to treatment guidelines in specific locations.P. falciparum parasites are prevalent [P. falciparum cases.Although follow-up rates and treatment adherence are not perfect in terms of ensuring individual case outcome, data penetration has been sufficient to enable policy decisions on anti-malarial drug treatment at the provincial level, with dihydroartemisinin\u2013piperaquine switched to pyronaridine\u2013artesunate for two provinces. Although pyronaridine\u2013artesunate shows high efficacy in regions in the GMS where multidrug-resistant revalent \u201331, normP. vivax malaria elimination. Although generally high efficacy rates were observed, a disparity was evident in Sisaket versus other provinces. The high day 28 failure rate in Sisaket in FY2018 suggests sub-optimal chloroquine efficacy, though efficacy was 100% in FY2020. Mutations associated with chloroquine resistance have been detected in P. vivax isolates from Thailand [P. vivax re-infection, despite declining case numbers. Investigations and discussions on the appropriate response to P. vivax malaria in Sisaket are ongoing in FY2021, and the findings may also affect the management of P. vivax malaria elsewhere in Thailand.iDES data also underline the importance of Thailand . NotablyThailand , which cThailand , but thiiDES has provided operationally relevant data on drug efficacy; however, there are some limitations. Most importantly, it is difficult to obtain a complete dataset for data collected routinely, which may introduce a bias towards patients that are more easily reached. As follow-up improves, this bias should diminish. Similarly, although health workers attempt to verify drug adherence, not all treatment is directly observed. Thus, although malaria recurrence in this report is attributed to treatment failure, it could be a result of poor adherence. From an operational perspective, it is valuable to be able to differentiate these two sources of recurrence, so health workers are receiving continued training to observe the drug pack as a proxy for consumption. A final consideration is that MIS data can be adjusted to reflect new information received or to correct records, and so should be regarded as cross-sectional.P. falciparum. There is evidence that iDES can support appropriate management of imported malaria as part of a comprehensive prevention of reintroduction program [Given the low and declining malaria incidence in Thailand, without iDES it is unlikely that a pattern of treatment failure would be detected in time to avert an outbreak of drug-resistant  program . MaintaiThe investment in malaria elimination in Thailand is considerable, but the potential benefits are even greater . As coun"}
+{"text": "Dihydroartemisinin\u2013piperaquine (DHA\u2013PPQ) has been adopted as first-line therapy for uncomplicated falciparum malaria in Indonesia since 2010. The efficacy of DHA\u2013PPQ was evaluated in 2 sentinel sites in Keerom District, Papua and Merangin District, Jambi, Sumatra from April 2017 to April 2018.Clinical and parasitological parameters were monitored over a 42-day period following the World Health Organization standard in vivo protocol and subjects meeting the inclusion criteria were treated with DHA\u2013PPQ once daily for 3\u00a0days, administered orally.Plasmodium falciparum were detected in 7 cases and categorized as late treatment failures (LTF) at days 21, 35, and 42 and one of which was reinfected by Plasmodium vivax at day 42. Two cases were confirmed as recrudescent infection and 4 were re-infection. The PCR-corrected DHA\u2013PPQ efficacy for P. falciparum was 97.9% (95% CI 92.7\u201399.7). Of the 80 cases of P. vivax that were followed up, 71 cases were completely cured and classified as ACPR  and 9 cases showed recurrent infection at days 35 and 42, and classified as LTF. In Sumatra, of the 751 subjects screened, 35 vivax subjects enrolled, 34 completed the 42\u00a0day follow up. Thirty-three cases were completely cured and classified as ACPR  and 1 recurrent infection was observed and classified as LTF. No delay in parasite clearance nor severe adverse reaction was observed. Analysis of the Pfk13 gene in P. falciparum cases from Papua revealed no mutations associated with artemisinin resistance in the 20 SNPs previously reported. Analysis of the Pfpm2 gene at day 0 and day of recurrence in recrudescent cases revealed the same single copy number, whereas 3 of the 4 re-infection cases carried 2\u20133 Pfpm2 gene copy numbers.In Papua, 6339 subjects were screened through active and passive cases detection. Of the 114 falciparum and 81 vivax cases enrolled, 102 falciparum and 80 vivax cases completed the 42\u00a0day follow up, and 12 falciparum and 1 vivax cases were either lost to follow up or withdrawn. Kaplan\u2013Meier analysis of microscopy readings of 102 falciparum cases revealed 93.1% (95% CI 86.4\u201397.2) as Adequate Clinical and Parasitological Response (ACPR). No delay in parasite clearance nor severe adverse reaction was observed. Recurrent parasites of Treatment of falciparum and vivax malaria cases with DHA\u2013PPQ showed a high efficacy and safety. In Indonesia, reports to date revealed that artemisinin-based combination therapy (ACT), particularly dihydroartemisinin\u2013piperaquine (DHA\u2013PPQ), is highly effective to treat any human malaria cases. Although certain studies reported few cases of delayed parasite clearance , this evPlasmodium falciparum and Plasmodium vivax malaria in Indonesia. The additional objective is to observe the gametocyte carriage during the follow-up period.Malaria control programme in Indonesia has successfully brought down the malaria cases within the last few decades and in 2017, more than half of the district and municipality have been certified as malaria free areas. However, malaria cases remain high in eastern provinces, such as Papua, West Papua, Molucca and East Nusa Tenggara. In Western part of the country, malaria is either eliminated or significantly reduced and only several malaria foci left in Sumatra, Java, Bali and Kalimantan. In 2020, Indonesia reported 254,055 malaria cases with Annual Parasite Incidence (API) 0.94 cases per 1000 population and 74% of infections reported from Papua Province . UnfortuThe study was conducted in Keerom District, Papua and Merangin District, Jambi, Sumatra from April 2017\u2013April 2018  recruited. Participants were recruited from malaria-infected persons found during active case detection (ACD) or passive case detection (PCD) in Papua and Sumatra. Persons were aged between 1 and 65\u00a0years, weighed more than 5 kg, had fever or history of fever in the preceding 24\u00a0h, with slide-confirmed malaria with parasitaemia of\u2009\u2265\u2009500/ul asexual parasites for P. falciparum and\u2009\u2265\u2009250/ul asexual parasites for P. vivax. Persons were excluded with the following exclusion criteria: pregnant, had a history of allergy to the study drugs or study drug\u2019s derivative, had previously completed treatment with an anti-malarial drug in the preceding 2\u00a0weeks, or had a medical history of untreated hypertension or chronic heart, kidney, or liver disease [The study was designed as one arm prospective evaluation of clinical and parasitological responses to directly observed treatment for uncomplicated malaria. Until 2017, as the treatment failure rate to DHP\u2013PPQ in Papua and Sumatra were unknown, a rate of 30% had been chosen. At a confidence level of 95% and a precision around the estimate of 10%, and with a 20% increased to allowed lost to follow-up and withdrawn during the 42-day follow-up period, it was estimated a total of 120 patients  on filter paper using 3MM Whatman  and kept in individual plastic zip lock for parasite genotyping. A standard physical examination, blood smears and DBS were also collected from finger pricks on days 1, 2, 3, 7, 14, 21, 28, 35 and 42  with timAll study participants were given a treatment of dihydroartemisinin (DHA) and piperaquine (PPQ) from primary health centre , containing 40\u00a0mg DHA and 320\u00a0mg PPQ per tablet and was administered once a day for 3\u00a0days in front of study team, as a weight per dose regimen of 2.25 and 18\u00a0mg/kg of DHA\u2013PPQ  and follGenomic DNA (on day of enrolment and day of recrudescence) was extracted from DBS using Chelex-100 ion exchanger  according to a previously published procedure . ExtractPfk13 gene for artemisinin resistance was performed according to previously published method. The DNA was amplified by nested PCR and sanger sequencing method to detect the SNPs: G449A, N458Y, T474I; M476I; A481V; Y493H; T508N; P527T; G533S; N537I; R539T; I543T; P553L; R561H; V568G; P574L; C580Y; D584V; E612D; S623C of P. falciparum K13 [Amplification of arum K13 . BioEditP. falciparum plasmepsin 2 gene determination consisted of several stages. Initially, DNA was extracted from the blood spots on filter paper according to the Wooden method [Pfpm2 and Pftub genes using quantification of the real time polymerase chain reaction (RT-qPCR) and assay parameters according to the Witkowski method. The primers used for Pfpm2 gene were 5'-TGGTGATGCAGAAGTTGGAG-3' and 5'-TGGGACCCATAAATTAGCAGA-3', while for Pftubulin these were 5'-TGATGTGCGCAAGTGATCC-3' and 5'-TCCTTTGTGGACATTCTTCCTC-3' [Pfpm2 and Pftub. The 3D7 strain were quantified in 6 replicates for Pfpm2 and Pftub. Interpretation of results and run validation followed the Witkowski method. The 3D7 strain line was included in each run as standard control for one copy of Pfpm2 gene in 6 replicates. Pfpm2 copy number was calculated by the 2-\u0394\u0394Ct method [Copy number of n method  and puriTCCTC-3' . Each cot method  and the This study used the excel Kaplan\u2013Meier analysis template provided by the WHO. The template calculated automatically data in the entry 1 and 2 files. The results are expressed as success and failure cumulative incidence, with 95% confidence intervals.Plasmodium malariae; and 0.05% had Plasmodium ovale. The remainders were (2.1%) mixed infections.The subjects screening and recruitment in Papua and Jambi, Sumatra is shown in Fig.\u00a0P. falciparum cases and 81 (4.1%) P. vivax cases met inclusion criteria  of the 1984 ria Fig.\u00a0. The remThe demographic characteristics of the enrolled study subjects in Papua and Sumatra has been shown in Table In Sumatra, 751 subjects screened through passive and active case detection and 51 (6.7%) were found positive for vivax malaria Fig.\u00a0. Of the P. falciparum were detected in 7 cases at days 21, 35, day 42. Classification of the treatment outcomes by microscopy is presented in Table P. vivax, 2 cases were confirmed as recrudescent infection and the remaining 4 cases were re-infection. Therefore, the PCR-corrected DHA\u2013PPQ efficacy for falciparum was 97.9% (95% CI 92.7\u201399.7). No delay in parasite clearance nor severe adverse reaction was observed in any study participants.Of the 114 falciparum cases enrolled, 102 cases completed the 42\u00a0day follow up, and 12 cases were either lost to follow up or withdrawn. Recurrent parasites of Pfk13 gene to observe the 20 SNPs associated with artemisinin resistance revealed that all P. falciparum isolates carried the wildtype allele.PCR amplification and DNA sequencing of the msp1, msp2, and glurp genes  revealed the same single copy number, whereas 3 of the 4 re-infection cases carried 2\u20133 copy numbers (Table Late treatment failure (LTF) was observed in 7 study participants with microscopy reading Table  and genoes Table . The deled Table . Three Lrs Table .Table 4GOf the 81 vivax cases enrolled, 80 cases completed the 42\u00a0day follow up, and 1 case were lost to follow up. Classification of the treatment outcomes by microscopy was presented in Table Of the 35 vivax cases enrolled, 34 cases completed the 42\u00a0day follow up, and 1 case were lost to follow up. Classification of the treatment outcomes by microscopy for vivax cases in Sumatra was presented in Table P. falciparum and 15 persons with P. vivax malaria in Papua while in Sumatra 18 persons carried gametocyte at enrollment  poses a substantial threat to the currently endorsed malaria elimination programme as it may increase not only malaria morbidity but also re-introduction of malaria in areas where elimination have been achieved. Results of this study clearly indicate that DHA\u2013PPQ is still highly effective in both study sites, Papua and Sumatra. For the artemisinin particularly, this study showed no delay in parasite clearance and this is also supported by the absence of mutations associated with artemisinin resistance in mutation , 21, regPfpm2 gene [Plasmodium falciparum isolates that survived the PPQ treatment. Analysis of the Pfpm2 gene also supports for the existence of P. falciparum isolates that carried more than one copy of Pfpm2 gene among the samples in Papua. Previous therapeutic efficacy study to evaluate the efficacy of DHA\u2013PPQ in Southern part of Papua (Mimika Regency) did not find any P. falciparum isolates that carry more than one copy number in Pfpm2 gene [plasmepsin 2\u20133 gene cluster has been identified as an important molecular determinant of piperaquine resistance in P. falciparum [Evidence for the existence of parasite isolates that are slightly resistant to PPQ in Papua also alerts to the proper deployment of the drug in the area , 23. Pippm2 gene , and as pm2 gene . In Camblciparum \u201327 and rlciparum .The finding for DHA\u2013PPQ late treatment failure in this study alerts to the proper treatment of malaria in the area and also anticipate having second-line ACT to replace the piperaquine as partner drug. Currently, the Indonesia national policy to use quinine as second line drug is regarded to be impractical as it introduces longer treatment period and also more often side effects. In this regard, consideration of using another artemisinin-based combination, such as Artemether\u2009+\u2009lumefantrine or Artesunate\u2009+\u2009mefloquine at least for falciparum cases might be rational. Overall, the results of this study are reassuring and suggest that in the absence of artemisinin resistance, the artemisinin regimen in any artemisinin-based combination may delay de novo emergence of resistance to the partner drug, such as DHA\u2013PPQ, artemether\u2013lumefantrine and artesunate\u2013mefloquine. DHA\u2013PPQ has been used in Indonesia as the first line anti-malarial drug since 2008  and it tThe high gametocyte carriage at enrolment in vivax cases (18.5% and 51.4% in Papua and Sumatra respectively) Table  is likelIn Indonesia, DHA\u2013PPQ procurement is highly regulated by the Indonesian Ministry of Health, and the drug is only available at government health facilities and selected private-sector facilities, which are able to confirm that the prescription should be based on malaria positivity by microscopy or rapid diagnostic test. With this tight regulation it is anticipated that DHA\u2013PPQ will continue to play a role in the treatment of uncomplicated malaria in Indonesia until malaria is successfully eliminated in the country. On the other aspect, implementation of evidence-based vector control may also contribute to mitigate transmission and delay the emergence of anti-malarial drug resistance.The therapeutic efficacy study conducted in two sentinel sites in Papua and Sumatra, Indonesia during 2017\u20132018 revealed that DHA\u2013PPQ is still highly effective in both sites. The appearance of recurrent falciparum infection in small number of cases in Papua alert to the possible emergence of piperaquine resistance in the area and deserve further investigation to contain its spread and anticipate for the rational option of a second-line ACT. Further studies are required in different regencies in Papua, particularly those in border area with Papua New Guinea to determine the spread of resistance to DHA\u2013PPQ."}
+{"text": "Background and Purpose: To assess the safety and effectiveness of oral methylprednisolone (oMP) in comparison with intramuscular adrenocorticotropic hormone (imACTH) and oral prednisolone (oP) therapies in children with infantile spasms (IS).Methods: In this prospective, open-label, non-blinded, uncontrolled observational study, children (aged 2\u201324 months) with newly diagnosed IS presenting with hypsarrhythmia or its variants on electroencephalogram (EEG) were included. It was followed by imACTH, oP, or oMP (32\u201348 mg/day for 2 weeks followed by tapering) treatments. Electroclinical remission/spasm control, relapse, and adverse effects were evaluated in the short-term (days 14 and 42) and intermediary-term  intervals.Results: A total of 320 pediatric patients were enrolled: 108, 107, and 105 in the imACTH, oMP, and oP groups, respectively. The proportion of children achieving electroclinical remission on days 14 and 42 was similar among the three groups . The time to response was significantly faster in the oMP group . Spasm control at 3, 6, and 12 months was also similar in the three groups . The relapse rate in the imACTH group (24.10%) was lower than oMP (30.77%) and oP groups (33.33%), and the time taken for relapse in the imACTH group  was longer than oMP  and oP groups , but the differences were not statistically significant . The occurrence of adverse effects was similar among the three groups.Conclusions: The short and intermediary-term efficacy and recurrence rates of oMP are not inferior to those of imACTH and oP for the treatment of IS. Significantly, the time to achieve electroclinical remission with oMP was quicker than that with imACTH and oP. Considering its convenience, affordability, and the absence of irreversible side effects, oMP can serve as a form of first-line treatment for newly diagnosed IS. Infantile spasm (IS), a subtype of epileptic spasms, is an epileptic encephalopathy and is frequently associated with hypsarrhythmic interictal electroencephalograms (EEG) and psychomotor retardation . The incThe United Kingdom Infantile Spasms Study suggested that the ideal treatment agent for IS should achieve rapid control of spasms, normalize the EEG changes early, maintain a spasm-free state, and thus contribute to a better intermediary-term developmental outcome . CurrentAccording to the reports of the Sri Lanka Infantile Spasms Study (SLISS), the two most commonly used forms of hormonal therapies, the intramuscular adrenocorticotropic hormone  and the oral prednisolone , exhibit no difference in the short- (28 days) and intermediary-term  control of newly diagnosed IS and can be considered as equal options for first-line treatment , 10. HowThe reported efficacy of ivMP (30 mg/kg/day) in children with IS is ~60\u201383%, with freedom from spasms being achieved very early (2\u20133 days following treatment) among responders , 13. RecThis study was designed as a prospective, open-label, non-blinded, uncontrolled observational study to evaluate the efficacy and tolerability of oMP in children with newly diagnosed IS. From January 2016 to October 2020, children aged 2\u201324 months with newly diagnosed epileptic spasms in clusters with electroencephalographic evidence of hypsarrhythmia or its variants were enrolled and assessed for eligibility in the Department of Neurology of Wuhan Children's Hospital. Children with a diagnosis of IS based on the definition proposed by Lux et al.  and thosBefore treatment allocation, the baseline characteristics of all eligible participants were assessed using detailed clinical histories and examinations. Participants' ages at onset, perinatal details, family histories, and developmental statuses were recorded. The results of investigations such as routine blood tests, liver and kidney function, blood electrolytes, arterial blood gas, blood lactate, blood ammonia, urine ketones, blood tandem mass spectrometry, neuroimaging , EEG, metabolic testing, and Gesell development schedules test were documented. The etiology of IS was classified according to six etiologic categories defined by the International League Against Epilepsy Task Force, including structural, genetic, infectious, metabolic, immune, and unknown .The parents or guardians of all eligible participants were provided with detailed information about the standard treatment options available for IS, including the total cost, efficacy, and potential adverse effects of imACTH, oP, and oMP therapies. They were asked to make an informed choice based on the feasibility and expense of the three treatments. A chest X-ray and tuberculin test were performed to screen for tuberculosis before starting hormonal therapy. The baseline weight and blood pressure were measured, and pyridoxine was administered to exclude pyridoxine-dependent seizures.The imACTH group received ACTH  intramuscularly (25 IU/day) for 2 weeks. If spasms continued until day 7 or reappeared from days 8 to 14, the dose was increased to 40 IU/day. After 2 weeks of treatment, all children received oral prednisolone  at an initial 2 mg/kg dose, which was tapered off and stopped over 4 weeks. The oP group received oral prednisolone at a dose of 10 mg four times/day for 2 weeks. If spasms continued until day 7 or reappeared from days 8 to 14, the dose was increased to 20 mg three times/day. Prednisolone was tapered and stopped over 4 weeks: reductions were made once a week, with 30 mg daily, then 20 mg, then 10 mg, and then 5 mg; if on the higher dose of treatment, the doses were 40 mg daily, then 20 mg, then 10 mg, and then 5 mg.The oMP group received oral methylprednisolone  at a dose of 8 mg four times/day for 2 weeks. If spasms continued until day 7 or reappeared from day 8 to day 14, the dose was increased to 16 mg three times/day. After 2 weeks of treatment, methylprednisolone was tapered and stopped over 4 weeks: reductions were made once a week, with 24 mg daily, then 16 mg, then 8 mg, and then 4 mg; if on the higher dose of treatment, the doses were 32 mg daily, then 16 mg, then 8 mg, and then 4 mg.Currently, there are no guidelines or expert consensus for the subsequent treatment of IS following hormone therapy. In this work, all patients received oral topiramate  after the end of 6 weeks of hormone therapy.After the initial treatment, patients continued to be followed up at different time points. The initial follow-up included reviews at 14 and 42 days posttreatment, followed by subsequent reviews at 3, 6, and 12 months posttreatment. The first two time points were considered markers of a short-term response, and the three later time points were considered markers of an intermediary-term response. Parents or guardians were trained to document spasms and maintain seizure diaries. At all follow-up visits, the children were evaluated by the same pediatric neurologist. Those who relapsed were treated again with hormonal therapies or other oral anticonvulsants at their parents' discretion. Drug compliance was monitored by asking the parents or guardians directly. The numbers of patients who did not present for follow-up and those who died during follow-up were noted separately for the different time points.The primary outcomes measured were the short- and intermediary-term responses. The evaluated outcome indicators for short-term response were the number of children with electroclinical remission  at the time of review on days 14 and 42.The evaluated outcome indicators for intermediary-term response were the number of children with spasm control (defined as the absence of witnessed spasms for \u226528 consecutive days from the time of the last witnessed spasm) at 3, 6, and 12 months after initial therapy.The secondary outcomes measured were the time to response (defined as the first day after initiation of treatment on which spasms were not seen) and relapse , and the time taken for relapse .Adverse effects (defined as any untoward or unintended responses) were thought to be related to the treatments. The anticipated potential adverse reactions included cushingoid features, weight gain, increased appetite, frequent crying spells, irritability, infections, drowsiness, gastrointestinal upset , and electrolyte imbalances. Other parental concerns regarding side effects were also noted.Categorical data are summarized in proportions, and quantitative data are presented as medians and quartiles. The significance of the differences in electroclinical remission and relapses between the three treatment groups were assessed at 14 and 42 days and 3-, 6-, and 12-month intervals using the Chi-square test. The effect size was calculated using the risk ratio  and absolute risk reduction  with 95% confidence intervals (CIs). The number needed to treat (NNT) was calculated based on the number of patients who needed methylprednisolone to achieve spasm remission, compared with a patient who did not. The Kaplan\u2013Meier survival method was used to plot the relapse and response rates under different groups. The significance level was set at 0.05, and all tests were two-tailed. Statistical analyses were performed using SPSS version 22.0.This study was reviewed and approved by the Medical Ethics Committee of Wuhan Children's Hospital, Tongji Medical College, Huazhong University of Science & Technology (2015019). The parents of the patients signed written informed consent, agreed with their children's participation in this study, and allowed the use of the relevant data and information for scientific research.A total of 335 children with clinically confirmed epileptic spasms were evaluated. Eventually, only 320 children were included in the study to compose the three treatment groups. A total of 108, 107, and 105 children were allocated to the imACTH, oMP, and oP groups, respectively. The baseline characteristics of the three groups showed no significant differences .p = 0.869), 3 (p = 0.801), 6 (p = 0.685), and 12 months (p = 0.791). Four deaths occurred during the 12-month follow-up, all of which occurred after completing the 6-week initial treatment and were related to other systemic diseases.Details of enrollment, treatment allocation, and follow-up  are outlined in p = 0.362). Electroclinical remission by the end of day 42 was higher in the oMP group  than in the imACTH  and oP groups , although the difference was not statistically significant (p = 0.470).The time course of treatment and outcome evaluation is presented in P = 0.025). The proportion of infants with an electroclinical remission and the corresponding required days within 42 consecutive days post initial treatment is graphically presented in The total number of days required for electroclinical remission was significantly lower in children treated with oMP  compared with those treated with imACTH  and oP  (P = 0.775). In addition, the proportions of children in whom spasm control was achieved also showed no significant difference at 6 (p = 0.667) and 12 months (p = 0.779) in the three groups , oMP , and oP  groups showed no significant difference (e groups .p = 0.539). In addition, no significant difference was noted in the time taken for relapse within each treatment group (p = 0.530), although children within the oMP group  tended to relapse earlier than those in the imACTH  and oP groups  during the 12-month follow-up period . AlthougAdverse effects observed in the three groups included cushingoid features, weight gain, increased appetite, frequent crying spells, irritability, drowsiness, increased susceptibility to infection, gastrointestinal upset , and electrolyte imbalances . The mosThe most accepted standard first-line treatment for IS is hormonal therapy, which mainly refers to imACTH and oP. ACTH is preferred but is expensive and difficult to obtain in some countries. Several trials and meta-analyses have confirmed that high-dose oP (40\u201360 mg/day) is not inferior to ACTH and is a more cost-effective treatment for IS , 21, 22.2/day) was associated with a greater early response rate than oP by analyzing the multicenter database of the National Infantile Spasms Consortium, whereas Wanigasinghe et al. found that the time required for spasm cessation was quicker in children treated with oP compared with imACTH at a dose of 40\u201360 IU every other day (3.85 vs. 8.65 days) in the SLISS trial. In this study, the total number of days required for spasm control was ~8 days in children treated with oP and imACTH (25\u201340 IU/d). In addition, oP and imACTH showed no significant difference in the time until a response in 3/5 randomized controlled trials, and the mean time to spasm cessation ranged from 3 to 8 days in a metaanalysis conducted by Kuo et al.  of the SLISS trial conducted by Wanigasinghe et al. . Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.Z-SL, CY, J-SH, and G-FW: study conception and design. H-MZ, M-QL, SH, and X-LD: data acquisition. C-HY, CY, and H-MZ: analysis and data interpretation. H-MZ and C-HY: drafting of the manuscript. All authors contributed to the article and approved the submitted version.This work was supported by the Wuhan Municipal Health Commission of Clinical Research Projects (NO. WX17B11).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher."}
+{"text": "Current tuberculosis treatments leave patients with clinically significant lung injury and increased all-cause mortality post-cure. Adjunctive host-directed therapies could protect the lungs, improve long-term survival, and shorten treatment duration; however, few have been tested clinically. Therefore, we aimed to assess the safety and preliminary efficacy of four host-directed therapies for tuberculosis.Mycobacterium tuberculosis, a sputum Xpert cycle threshold of less than 20, and moderately advanced or far advanced disease on chest radiography. By use of numbers generated in blocks of ten and stratification by site, eligible patients were randomly assigned (1:1:1:1:1) to receive one of the four oral host-directed treatments plus standard tuberculosis treatment or standard treatment alone (the control group). Host-directed treatments were: CC-11050 ; everolimus ; auranofin ; and ergocalciferol . All study participants received oral rifabutin-substituted standard tuberculosis treatment for 180 days. Patients and clinicians were not masked to treatment assignment. Spirometry and sputum culture with solid and liquid media were done at baseline and up to 180 days at specified intervals throughout treatment. The primary endpoint was safety and tolerability up to day 210. Secondary preliminary efficacy endpoints were treatment effects on sputum microbiology (culture status at day 56 and the hazard ratio for stable culture conversion up to day 180) and lung function  measured by spirometry at day 56, day 180, and day 540. Safety was analysed in the intention-to-treat population and preliminary efficacy primarily in the per-protocol population. The trial is registered at ClinicalTrials.gov, NCT02968927. Post-treatment follow-up was completed in 2020.In this prospective, open-label, phase 2, randomised controlled trial, patients with pulmonary tuberculosis were recruited at three clinical sites in South Africa. Eligible patients were aged 18\u201365 years, HIV-1-negative, and had rifampicin-susceptible 1 in the control group was 61\u00b77% of predicted (95% CI 56\u00b73\u201367\u00b71) at baseline and 69\u00b71% (62\u00b73\u201375\u00b78) at day 180. Patients treated with CC-11050 and everolimus had increased recovery of FEV1 at day 180 relative to the control group , whereas auranofin and ergocalciferol recipients did not. None of the treatments had an effect on FVC during 180 days of follow-up or on measures of sputum culture status over the course of the study.Between Nov 18, 2016, and Sept 27, 2018, 200 patients were screened and randomly assigned to different treatment groups . 11 treatment-emergent serious adverse events occurred either during treatment or within 30 days after treatment discontinuation, of which three were attributable to a host-directed treatment. Life-threatening thrombocytopenia occurred in an auranofin recipient; apparent intra-abdominal sepsis leading to death occurred in another auranofin recipient and was classified as a suspected unexpected serious adverse reaction. Tuberculous spondylitis occurred as an apparent paradoxical reaction in a patient receiving ergocalciferol. Two patients in the control group had life-threatening, treatment-attributable liver injury. No treatment-emergent, treatment-attributable serious adverse events occurred in patients receiving CC-11050 or everolimus. Mean FEV1, a key measure of lung function and predictor of all-cause mortality. Further studies of these candidates are warranted.CC-11050 and everolimus were safe and reasonably well tolerated as adjunctive therapies for tuberculosis, and analysis of preliminary efficacy suggests they might also enhance the recovery of FEVThe Bill & Melinda Gates Foundation and the South African Medical Research Council. Evidence before this study1 is an independent, generalisable predictor of all-cause mortality. Tuberculosis leaves most patients with permanent impairment of lung function, including of FEV1. There is growing recognition that long-term all-cause mortality risks are increased after tuberculosis, to a degree consistent with impairment of FEV1. We searched PubMed for articles published in English between database inception and Dec 15, 2019, using the search terms \u201ctuberculosis\u201d, \u201cclinical trial\u201d, and \u201cspirometry\u201d. The search did not identify any studies of interventions to protect the lung done more recently than the 1960s. Several clinical trials of vitamin D have been done in patients with tuberculosis. Only one found that vitamin D enhanced sputum culture conversion at week 8 compared with standard treatment alone; none examined lung function. Studies of the type 4 phosphodiesterase inhibitor CC-11050 in experimental mouse and rabbit models of tuberculosis found that it improved the resolution of lung pathology and the clearance of Mycobacterium tuberculosis infection. One previous study of a non-specific phosphodiesterase inhibitor (pentoxifylline) found no benefit on short-term or long-term outcomes but did not examine effects on lung function.FEVAdded value of this study1. Interventions to enhance the recovery of FEV1 in patients with tuberculosis might help to reduce the increased long-term mortality risks observed after treatment.This phase 2 study found that adjunctive treatment with CC-11050 or everolimus in adults with pulmonary tuberculosis and baseline predictors of poor outcome was safe and well tolerated, and might have resulted in improved recovery of FEVImplications of all the available evidenceMore studies of CC-11050 and everolimus in patients with tuberculosis are warranted to further investigate their effects on lung function and assess their impact on long-term health outcomes.Tuberculosis is a leading cause of morbidity and mortality globally.3There is growing interest in the potential of adjunctive host-directed therapies to improve treatment options for patients with tuberculosis. As modulators of the host immune response, these treatments have the potential to protect the lungs, improve long-term survival, and shorten treatment duration by reducing lung inflammation, improving lesional drug penetration, and inducing antimicrobial activity in phagocytic cells.Mycobacterium tuberculosis infection.M tuberculosis.M tuberculosis.Therefore, we aimed to assess the safety and preliminary efficacy of four adjunctive host-directed therapies in patients with hard-to-treat tuberculosis: CC-11050, everolimus, auranofin, and vitamin D . CC-11050 is a type 4 phosphodiesterase inhibitor with anti-inflammatory properties and has shown efficacy in preclinical models of M tuberculosis/rifampin  test results were asked to consent for further screening to enter the study.In this prospective, open-label, phase 2, randomised controlled trial, patients with pulmonary tuberculosis were recruited at three clinical sites in Greater Johannesburg and Pretoria, South Africa: the Tembisa Clinical Research Centre, Tembisa; the Clinical HIV Research Unit, Johannesburg; and the Setshaba Research Centre, Soshanguve. Patients attending neighbouring public health clinics were asked to give permission for a review of laboratory records for possible study eligibility. Those with positive sputum Cepheid Xpert M tuberculosis and one or more probes with a cycle threshold less than 20. Cycle threshold indicates the number of cycles of DNA amplification required to result in detection of the target and is inversely related to the log of the target copy number. Patients also had moderately advanced or far advanced pulmonary tuberculosis on chest radiography , the London School of Hygiene & Tropical Medicine (reference number 10645), the South African Medicines Control Council (reference number 20160506), is registered as study 4297 in the South African Human Research Electronic Application System, and can be found online.By use of numbers computer-generated by the study statistician (SG) in blocks of ten and with stratification by site, patients were randomly assigned (1:1:1:1:1) to receive CC-11050, everolimus, auranofin, or ergocalciferol in addition to standard tuberculosis therapy, or standard tuberculosis therapy alone (control). At each study site, the individuals responsible for assigning patients to treatment groups were not involved in evaluating treatment outcomes. Envelopes containing allocation information were provided to each site by SG. Patients and clinicians were not masked to treatment assignment. However, microbiology and spirometry laboratory personnel and senior study leadership (RSW and GC) were masked to treatment assignment. Summaries of study progress prepared by SG that described outcomes by treatment group were available only to members of the study's data and safety monitoring committee.NCT01300208) and erythema nodosum leprosum (NCT03807362). The dose of everolimus was selected as the lowest showing human biological activity.Participants randomly allocated to adjunctive host-directed tuberculosis therapies received CC-11050 , everolimus , auranofin , or ergocalciferol  orally. Adjunctive CC-11050, everolimus, or auranofin were given from day 1 to day 112 as follows: 200 mg of CC-11050 twice a day with food; 0\u00b75 mg/day of everolimus; or 3 mg/day of auranofin for the first week, which was then increased to 6 mg/day. Ergocalciferol was given as a 5 mg dose on day 1, then 2\u00b75 mg on days 28 and 56. CC-11050 was given at the same dose used by Celgene in two phase 2 studies in patients with cutaneous lupus erythematosus  and liquid media  on days 0, 1, 7, 14, 21, 28, 35, 42, 56, 84, 112, 140, and 180, with particular attention to culture status on day 56. Patients were repeatedly encouraged to produce adequate sputum specimens, including through video instructional materials. Sputum induction was not done. M tuberculosis molecular testing, culture, and drug susceptibility testing, and a single commercial laboratory  for safety studies.Spirometry was done at the study sites by use of EasyOne Pro instruments  beginning on study day 1. The protocol originally specified that spirometry be done on days 1, 14, 28, 56, 84, 112, 140, and 180, with particular attention to lung function on days 56 and 180. A protocol amendment later specified that spirometry should additionally be done on day 540. All sites used a single microbiology laboratory  for The primary endpoint of the study was the safety and tolerability of the study treatments, in terms of treatment-emergent serious adverse events for CC-11050 and suspected unexpected serious adverse reactions for the other treatments up to day 210, and was centrally assessed. Secondary safety endpoints included treatment-emergent adverse events other than serious adverse events, categorised according to severity, drug-relatedness, and whether they lead to early withdrawal, and the proportion of patients with disease exacerbation (assessed at each scheduled visit) due to study treatments. Safety events were assessed by site personnel at each study visit and subsequently reviewed by the data and safety monitoring committee. Clinicians at each site provided initial assessments of severity (using adverse event grading tables from the Division of Allergy and Infectious Diseases) and treatment-relatedness. These assessments could be upgraded following central review.M tuberculosis (as indicated by sputum culture status) and on lung function. The proportion of patients with positive sputum culture status was assessed at day 56 using solid culture medium, because previous trial data suggest culture status at this timepoint is a predictor of tuberculosis recurrence.M tuberculosis was detected in more than one specimen after day 112. For the entire treatment period, stable culture conversion was defined as occurring on the day of the first of two consecutive sputum specimens obtained at least 2 weeks apart without growth of M tuberculosis. Specimens with liquid or solid cultures showing growth of M tuberculosis were considered positive. Instances in which patients were unable to provide a sputum specimen despite encouragement were considered negative. Cultures that were unevaluable because of contamination or other causes were considered missing. To assess lung function, FEV1 and forced vital capacity (FVC) were measured by spirometry, and change from control was assessed at numerous timepoints, with particular attention to days 56, 180, and 540. Spirometry findings were interpreted according to guidelines from the American Thoracic Society and the European Respiratory Society,1 was expressed as a percentage of the predicted value based on age, sex, height, and race (FEV1%).Secondary preliminary efficacy endpoints were key measures of the effects of host-directed therapies on the clearance of 18F-fluorodeoxyglucose-PET, CT imaging, and serum markers of inflammation, will be undertaken and reported separately.Analyses of other secondary microbiology and lung function outcomes and of early biomarkers of treatment response, including changes in NCT01300208) included only 48 patients with cutaneous lupus erythematosus. We also considered microbiological outcomes in choosing sample size. Near-complete (\u22641%) culture conversion at 2 months, measured by use of solid medium, has been proposed as a target for new 4-month regimens.With 40 patients per group, this study was predicted to have a 77% power, at a two-sided 0\u00b705 significance level, of detecting a significant difference in the proportion of serious adverse events between the control and experimental treatments, assuming proportions of 0\u00b705 (n=2) in the control group and 0\u00b73 (n=12) in the experimental groups. This sample size was considered appropriate to detect treatment-emergent serious adverse events attributable to CC-11050 in an exploratory trial in patients with tuberculosis because the sole previous phase 2 study . The secondary efficacy outcomes for microbiology and lung function were analysed in the per-protocol population, which included patients in the modified intention-to-treat population, but excluded those who did not complete treatment or were found to have inadequate adherence . The secondary efficacy analyses were repeated using the modified intention-to-treat population.1 and FVC at day 56, day 180, and day 540 were compared by ANCOVA. We did not adjust for multiple comparisons, but instead independently compared each candidate host-directed therapy with the control group, as each might be advanced independently on the basis of this analysis. Analyses were adjusted for baseline covariates. We considered adjusting for multiple baseline factors in each analysis, but found that the effect of additional factors was small. Missing data were excluded from analyses and not otherwise imputed or estimated.Safety data are presented descriptively, with particular attention to severity, treatment-relatedness, and action on study drug for serious adverse events and suspected unexpected serious adverse reactions. Logistic regression was used to assess associations of treatment group with culture-positive status measured after 56 days of treatment. We report unadjusted and adjusted odds ratios (ORs) and 95% CIs for culture status on solid medium, with the adjusted values controlling for baseline differences in time to positivity in automated liquid cultures. We individually compared experimental groups with the control group to calculate the hazard ratio (HR) for stable culture conversion up to 180 days using Cox proportional hazards regression. FEV1 were done to investigate adjustment for study site in the per-protocol population and to examine the use of more stringent spirometry criteria at day 180 in the per-protocol population (grades A\u2013D were considered acceptable). An additional post-hoc analysis fit a random effects model for the repeated measurements of FEV1% on a study day until day 180 using the modified intention-to-treat population and included an interaction between day and study group, and a random intercept for repeated measurement of FEV1%. Further post-hoc analyses separately examined treatment effects on culture conversion in liquid cultures in the per-protocol population using Cox proportional hazards methods.Post-hoc analyses of FEVNCT02968927 by ClinicalTrials.gov.All analyses were done by use of Stata (version 14). Study progress was monitored at regular intervals by an independent data and safety monitoring committee. This trial is registered as study 4297 in the South African Human Research Electronic Application System, as DOH-27\u20130616\u20135297 by the South African National Clinical Trial Registration, and as The Bill & Melinda Gates Foundation provided funding for this trial and contributed to the selection of host-directed therapy candidates and discussions of trial design. The South African Medical Research Council provided additional funding for administrative aspects of the study. Celgene provided CC-11050. The Aurum Institute served as a study sponsor. The funders of the study had no role in determining other aspects of study design, data collection, data analysis, data interpretation, or writing of the report.M tuberculosis burden  at baseline and 69\u00b71% (62\u00b73\u201375\u00b78) at 180 days, with much of the apparent increase occurring early during standard treatment. All groups except for the ergocalciferol group showed an apparent increase in mean FEV1 from baseline to day 14, although none of these increases reached statistical significance .1 can be impaired by the inability to fully inhale or rapidly exhale, which can be attributed to two predominant post-tuberculosis pathological mechanisms: fibrosis and bronchiectasis. The finding that enhanced recovery of FEV1 occurred relatively late (at day 180 but not day 56) during treatment is of particular interest. Inflammation due to recurring viral or bacterial lung infection is thought to drive progressive FEV1loss in COPD by promoting airway remodelling, a process that results in structural changes to the airways by depositing collagen in the extracellular matrix, altering the cellular composition of the epithelial barrier, and narrowing small airways.In analyses of preliminary efficacy, both CC-11050 and everolimus appeared to enhance the recovery of FEV1 is an independent, generalisable predictor of all-cause mortality, even in individuals with only mild to moderate impairment .1 declines to 70% of predicted (moderately impaired), and doubles again as FEV1 further declines to 50% of predicted (severely impaired).1. Multiple studies have confirmed that mortality risk is increased after tuberculosis.FEVM tuberculosis colony-forming unit counts to a greater extent than did isoniazid alone.M tuberculosis-infected rabbits, CC-11050 additionally reduced the expression of multiple matrix metalloproteinase genes (particularly MMP12) and reduced the extracellular deposition of collagen in the lung.30CC-11050, a type 4 phosphodiesterase inhibitor, was previously a backup compound for apremilast , sharing similar chemical structures and biological activities. Apremilast is now approved for multiple anti-inflammatory indications. CC-11050 decreases the production of pro-inflammatory cytokines, including tumour necrosis factor, by preventing the degradation of cyclic AMP. In preclinical models of active tuberculosis in the mouse and rabbit, treatment with CC-11050 plus isoniazid reduced the number and size of lung lesions, ameliorated lung pathology, and reduced lung 1 values at baseline in this study tended to be somewhat higher in everolimus recipients than in control recipients, the difference from controls at 180 days persisted after adjusting for this potential confounding factor. This clinical trial is, to our knowledge, the first of an mTOR inhibitor in tuberculosis.Everolimus, an inhibitor of mTOR, is used clinically at high doses for the treatment of some cancers, and at intermediate doses as an immunosuppressive agent for solid organ transplantation. Anti-inflammatory and anti-fibrotic effects of mTOR inhibitors have also been described.M tuberculosis, HIV, and severe acute respiratory syndrome coronavirus 2.Auranofin is an orally bioavailable gold salt introduced in the 1980s as a disease-modifying treatment for rheumatoid arthritis. In addition to their anti-inflammatory properties, gold salts show broad-spectrum antibacterial and antiviral activities in vitro, including against diverse intracellular pathogens such as Vitamin D is essential for host defences against tuberculosis.1 on day 14 is a true early treatment response or merely a learned improvement in test taking. Larger studies with longer follow-up will be necessary to directly assess effects on mortality.The findings of this trial are best considered in the context of its experimental medicine design. The study's main shortcomings are a small sample size and low statistical power. Experimental groups were compared with the control without adjustment for multiple comparisons. Patients and clinicians could not readily be masked to treatment assignment because of the multiplicity of experimental treatments, which potentially introduced bias. As an exploratory study, its key findings will require verification in future trials. Further studies will be necessary to assess effects in patients with common comorbidities such as HIV-1 infection and diabetes, in patients with rifampicin-resistant disease, and in greater numbers of female patients. Repeated spirometry on day 2 in future studies would help to elucidate whether the apparent improvement in FEV1, a key measure of lung function and predictor of long-term all-cause mortality. Further definitive studies of CC-11050 and everolimus in patients with pulmonary tuberculosis are warranted.In summary, two adjunctive host-directed therapies for\u2014CC-11050 and everolimus\u2014showed adequate safety and tolerability in patients beginning treatment for pulmonary tuberculosis. An analysis of preliminary efficacy suggests that these therapies might preserve FEVstudy protocol see https://bit.ly/2MA6bJLFor the rwallis@auruminstitute.org.Data collected for the study, including deidentified individual participant data, a data dictionary defining each field in the set, and the statistical analysis plan will be made available to other researchers on request from the time of publication of this manuscript. Research proposals can be submitted to the corresponding author (RSW) at"}
+{"text": "Dihydroartemisinin-piperaquine (DP) is a long-acting artemisinin combination treatment that provides effective chemoprevention and has been proposed as an alternative antimalarial drug for intermittent preventive therapy in pregnancy (IPTp). Several pharmacokinetic studies have shown that dose adjustment may not be needed for the treatment of malaria in pregnancy with DP. n\u2009=\u20095) followed by a three-compartment disposition model. Gestational age did not affect the pharmacokinetic parameters of piperaquine. After three rounds of monthly IPTp, 9.45%  of pregnant women had trough piperaquine concentrations below the suggested target concentration (10.3\u2009ng/ml). Translational simulations suggest that providing the full treatment course of DP at monthly intervals provides sufficient protection to prevent malaria infection. Monthly administration of DP has the potential to offer optimal prevention of malaria during pregnancy. Dihydroartemisinin-piperaquine (DP) is a long-acting artemisinin combination treatment that provides effective chemoprevention and has been proposed as an alternative antimalarial drug for intermittent preventive therapy in pregnancy (IPTp). Several pharmacokinetic studies have shown that dose adjustment may not be needed for the treatment of malaria in pregnancy with DP. However, there are limited data on the optimal dosing for IPTp. This study aimed to evaluate the population pharmacokinetics of piperaquine given as IPTp in pregnant women. Pregnant women were enrolled in clinical trials conducted in Kenya and Indonesia and treated with standard 3-day courses of DP, administered in 4- to 8-week intervals from the second trimester until delivery. Pharmacokinetic blood samples were collected for piperaquine drug measurements before each treatment round, at the time of breakthrough symptomatic malaria, and at delivery. Piperaquine population pharmacokinetic properties were investigated using nonlinear mixed-effects modeling with a prior approach. In total, data from 366 Kenyan and 101 Indonesian women were analyzed. The pharmacokinetic properties of piperaquine were adequately described using a flexible transit absorption ( Plasmodium falciparum can develop placental malaria, with sequestration of the parasite in the placental vasculature. Placental malaria adversely affects both the mother and the infant, with adverse outcomes including maternal anemia and death, abortion, stillbirth, preterm delivery, low birth weight, infant mortality, and poor long-term child development  (8\u2013Dihydroartemisinin-piperaquine (DP) is a highly efficacious and well-tolerated antimalarial treatment. The long half-life of piperaquine provides extended malaria chemoprevention for up to 6 weeks. This antimalarial combination has therefore been suggested for malaria chemoprevention in several populations, including pregnant women. In Thai adults, a monthly dose of DP demonstrated superior protective efficacy  compared to dosing every 2 months (86% efficacy with a 95% CI of 81 to 90%) . Similar5\u2013.0001) 8\u2013. One of Pharmacokinetic (PK) properties in pregnancy may be altered because of several physiological changes, including increased water and fat contents, increased renal function, and altered enzymatic expression and degree of plasma protein binding. For example, exposures to artemether, artesunate, chloroquine, dihydroartemisinin, lumefantrine, sulfadoxine, pyrimethamine, atovaquone, proguanil, and cycloguanil are altered during pregnancy \u201316. In c\u2013P. falciparum infections during pregnancy  of women with malaria have been investigated in several pregnant populations \u201325. Seve\u201319\u201319\u2013regnancy . The stuThe main results of these two clinical trials have been reported previously , 27. ParCmax) for the Kenyan and Indonesian pregnant women were predicted to be 212\u2009ng/ml  and 232\u2009ng/ml , respectively. Observed trough piperaquine concentrations accumulated substantially with repeated monthly IPTp, but predicted peak concentrations remained similar during the entire duration of IPTp  before the subsequent IPTp dose if taken monthly , blood smear, placental blood PCR, or placental tissue histology and was detected in 3.3% (3/92) of Indonesian women and 31.7% (112/353) of Kenyan women . Howeverregnancy , simulat monthly .Our analysis highlights that a standard 3-day treatment course of DP, provided monthly as IPTp, appears to provide sufficient protection from malaria infection in pregnant women. This finding was apparent in both Kenya and Indonesia, with only 7 of 350 pregnancies presenting with placental malaria infection at delivery. An estimated 90.6%  of women are likely to maintain piperaquine trough concentrations above 10.3\u2009ng/ml, and 66.8%  are likely to maintain concentrations above 13.9\u2009ng/ml, concentration thresholds previously found to be associated with 95% and 99% malaria protection after three rounds of DP dosing, respectively . HoweverF) relied heavily on the prior model, resulting in estimated 95% confidence intervals including the prior estimates. On the other hand, the clinical trial data were informative in determining the elimination clearance and predicting the trough concentrations in this study, resulting in a significantly different clearance in this study compared to that of the prior study .Even though our analysis utilized samples collected from two studies and almost 500 recruited pregnant women , most samples were collected close to trough concentrations, resulting in insufficient data to develop a robust absorption and distribution model of piperaquine. Piperaquine pharmacokinetics are normally described using a multiphasic disposition model and transit compartment absorption , 30\u201332. 30\u2013This study did not recruit nonpregnant patients, and therefore, it was not possible to determine if, overall, pregnancy had an effect on the pharmacokinetic properties of piperaquine. The potential effects of pregnancy on the pharmacokinetic properties of piperaquine are still unclear. Several previous studies have shown that pregnancy has no effect on the pharmacokinetic properties of piperaquine , 22. HowCAP-VEN). Since the different study sites provided different samples (venous plasma versus capillary dried blood spots), the population estimates of the conversion factor between capillary dried blood spot concentrations and venous plasma concentrations might be interpreted as a combination of sampling differences, ethnicity differences, and other unknown site-specific differences. However, previously reported results have not shown any evidence of clinically important ethnic differences associated with the pharmacokinetic properties of the nine antimalarial drugs used in the standard artemisinin-based combination therapy (ACT) regimens recommended by the WHO  in Indonesia and did not show any increase in absolute QTc or QTc prolongation with repeated cycles of monthly DP dosing . Peak piperaquine concentrations were estimated to be approximately 17-fold higher than trough concentrations. Therefore, the remaining piperaquine concentrations at the end of the monthly round, associated with the accumulation of piperaquine trough concentrations, had a very minor impact on the peak piperaquine concentrations due to the relatively small contribution to total peak concentrations  and D. PP dosing . This suP. falciparum infection . An altered administration schedule during the first round of IPTp might lead to poor drug adherence. The most important aspect of preventive treatment is adherence, and pregnant women are scheduled to visit the antenatal care (ANC) clinic on a monthly basis. The monthly dosing regimen is the most practical way to administer these preventive treatments and is likely to result in high efficacy when taken as instructed. Thus, adherence to the full 3-day course of DP is the main concern, and missing any of the home-administered doses (2nd and/or 3rd dose) will result in subtherapeutic piperaquine concentrations for a substantial duration of time before the next round of IPT dosing.Monthly dosing of DP provides better protection against malaria than less frequent dosing , 24, 37.This study has several limitations. IPTp is recommended for pregnant women during the 2nd and 3rd trimesters . PregnanThe population pharmacokinetic properties of piperaquine were successfully evaluated in pregnant women receiving IPTp. Five transit compartments followed by a three-compartment disposition model described the pharmacokinetic properties of piperaquine adequately. Gestational age and other baseline covariates had no significant effect on the pharmacokinetic properties of piperaquine. Modeling and simulation suggested that more than 90.3% of pregnant women who receive three monthly courses of IPTp achieved piperaquine exposures associated with protection against acquired malaria infections. Predicted peak concentrations did not accumulate with repeated dosing courses, suggesting that IPTp with DP is not likely to increase the risk for QT interval prolongation associated with piperaquine exposure, but further cardiac safety data are still needed. The PK/PD analysis presented here suggested that monthly IPTp with DP is likely to be highly protective against placental malaria.This population pharmacokinetic study included pregnant women from two distinct randomized three-arm clinical trials. Both trials included intermittent screening and treatment in pregnancy (ISTp) with DP (\u201cISTp-DP\u201d) and IPTp with DP (\u201cIPTp-DP\u201d). In Kenya, the third arm consisted of IPTp with SP, and in Indonesia, this was single screening and treatment with DP (SSTp-DP) , 27. BotAll pregnant women received a 3-day fixed oral combination of dihydroartemisinin and piperaquine  (40\u2009mg dihydroartemisinin and 320\u2009mg piperaquine tetraphosphate per tablet), dosed by weight at enrollment according to the manufacturer\u2019s recommendations, approximately once a month in the Indonesian study and at 4- to 6-week intervals in the Kenya study , 27. Theg for 10 min), and plasma samples were stored at \u221280\u00b0C until shipment on dry ice. In Indonesia, capillary samples were collected by finger prick according to the same schedule. A drop of blood was collected on filter paper , and the filter paper was dried horizontally (1\u2009h at 50% humidity) and packed into separate plastic bags with silica gel. The plasma samples from Kenya and dried blood spot samples from Indonesia were transported for drug analysis to the Department of Clinical Pharmacology, Mahidol-Oxford Tropical Medicine Research Unit (MORU), Bangkok, Thailand.In Kenya, a baseline venous blood sample was collected from all women prior to antimalarial administration. Trough venous samples (1.5\u2009ml) were collected before each monthly drug administration. An additional venous sample was collected at the time of delivery. All blood samples were centrifuged . Pirana version 2.9.0 , Perl-spij = \u0398 \u00d7 exp, where \u0398ij is the pharmacokinetic parameter estimate for the ith patient at the jth occasion, \u0398ij is the typical pharmacokinetic parameter estimate of the population, \u03b7i,\u0398 is the between-patient variability of parameter \u0398 in the ith patient, and \u03baij,\u0398 is the between-occasion variability of parameter \u0398 in the ith patient at the jth occasion. Both between-patient variability and between-occasion variability were assumed to be normally distributed with a zero mean and \u03c92 variance. Estimated between-patient variability below 10% or variability estimated with poor precision  was fixed to zero. Residual unexplained variability was modeled as an additive error on the log-transformed observed concentrations, which is essentially equivalent to an exponential error on an arithmetic scale.Pharmacokinetic parameters were assumed to be log-normally distributed and therefore implemented as exponential between-patient and between-occasion variabilities as \u0398i) was measured at enrollment only and introduced into the pharmacokinetic model as a fixed allometric function on all volume (exponent of n = 1.00) and clearance (exponent of n = 0.75) parameters, scaled to the median body weight (48.5\u2009kg) of the prior study population, as follows: \u0398i = \u0398 \u00d7 exp \u00d7 (BWi/48.5)n.Individual body weight (BWP\u2009<\u20090.05) and elimination (P\u2009<\u20090.001) approach. The corrected gestational age was implemented as a time-varying covariate. The effect of gestational age was also modeled separately using a full covariate approach in which the gestational age was implemented as a continuous covariate on all pharmacokinetic parameters in the final pharmacokinetic model. Secondary pharmacokinetic parameter estimates were derived from the post hoc pharmacokinetic parameter estimates of the final pharmacokinetic model.The population conversion factor (CF) between venous plasma concentrations and capillary blood concentrations was estimated without between-patient variability. All other covariates  were investigated by a stepwise addition . Potential model misspecification and systematic errors were evaluated by basic goodness-of-fit diagnostics. Eta and epsilon shrinkages were used to assess the ability to detect model misspecifications in goodness-of-fit diagnostics   . Model discrimination between two hierarchical models was determined by a likelihood ratio test, based on the chi-square distribution of the OFV  . The 5thP. falciparum infection during pregnancy in a previous IPTp study in Uganda , Eijkman Institute for Molecular Biology (project N:57), and Litbangkes, Ministry of Health, Jakarta (LB02.01/5.2/KE059/2013). This clinical trial was registered at the ISRCTN registry under number ISRCTN34010937. Written informed consent was obtained from all participants."}
+{"text": "Plasmodium falciparum in north eastern states of India since 2013 after confirming the resistance against sulfadoxine-pyrimethamine. In the present study, therapeutic efficacy of artemether-lumefantrine and k13 polymorphism was assessed in uncomplicated P. falciparum malaria.Malaria is a major public health problem in India and accounts for about 88% of malaria burden in South-East Asia. India alone accounted for 2% of total malaria cases globally. Anti-malarial drug resistance is one of the major problems for malaria control and elimination programme. Artemether-lumefantrine (AL) is the first-line treatment of uncomplicated P. falciparum malaria were administered with fixed dose combination (6 doses) of artemether-lumefantrine for 3days and clinical and parasitological response was recorded up to 28days as per World Health Organization protocol. Nucleotide sequencing of msp1 and msp2 gene was performed to differentiate between recrudescence and reinfection. Amplification and sequencing of k13 propeller gene region covering codon 450680 was also carried out to identify the polymorphism.This study was conducted at four community health centres located in Koraput district of Odisha, Bastar district of Chhattisgarh, Balaghat district of Madhya Pradesh and Gondia district of Maharashtra state. Patients with uncomplicated msp1 and msp2 confirmed 2 cases each of recrudescence and reinfection, respectively. Mutation analysis of k13 propeller gene showed one non-synonymous mutation Q613H in one isolate from Bastar.A total 376 malaria patients who fulfilled the enrolment criteria as well as consented for the study were enrolled. Total 356 patients were followed up successfully up to 28days. Overall, the adequate clinical and parasitological response was 98.9% and 99.4% with and without PCR correction respectively. No case of early treatment failure was observed. However, four cases (1.1%) of late parasitological failure were found from the Bastar district of Chhattisgarh. Genotyping of P. falciparum malaria among all age groups. No functional mutation in k13 was found in the study area. The data from this study will be helpful in implementation of artemether-lumefantrine in case of treatment failure by artesunate plus sulfadoxine-pyrimethamine.The study results showed that artemether-lumefantrine is highly effective in the treatment of uncomplicated  Plasmodium falciparum monoinfections [Malaria is a major public health problem in India and accounts for about 88% of malaria burden in South-East Asia. India alone accounted for 2% of total malaria cases globally . In Indifections .P. falciparum was reported from Assam in 1973 within India [P. falciparum cases [First information of chloroquine resistant in India , and resin India . Emergenin India . SP resiin India . In 2010Fixed dose combination of artemether (20mg) and lumefantrine (120mg) is commercially available. Artemether has a half-life of 13h, as it absorbs and metabolizes quickly leading to fast clearance of parasite from blood. Due to longer half-life of lumefantrine (36days), it further eliminates the residual parasite from the malaria patient and helps to prevent the recrudescence . Therefok13 propeller gene has found to be associated with artemisinin resistance in Southeast Asia [k13 in India.Mutation in ast Asia . Many muP. falciparum malaria was assessed in four states, Madhya Pradesh, Chhattisgarh, Maharashtra and Odisha and mutation in k13 propeller gene was analysed to confirm the artemisinin resistance.Periodic monitoring of therapeutic efficacy is recommended by World Health Organization (WHO) to control the malaria and avoid the spread of resistant parasite in the population. Therefore, therapeutic efficacy of AL for the treatment of uncomplicated P. falciparum and Plasmodium vivax were reported from these sites. The selected CHCs were having sufficient facility for management of uncomplicated malaria. All these sites were at distance of 40 to 70km from their district headquarter for the management of severe malaria cases at district hospitals.This study was a one-arm prospective study conducted at four sites during June 2017 to Dec 2017. These four community health centres (CHCs) located in Koraput district of Odisha, Baster district of Chhattisgarh, Balaghat district of Madhya Pradesh and Gondia district of Maharashtra state  were excluded from the study as the request for pregnancy test and initiation of contraceptive was not acceptable in local areas.Febrile patients aged between 6months and 60years of age attending the CHC hospitals were screened for malaria parasite by microscopy and patients with uncomplicated P. falciparum , fever or history of fever within 24 previous hours, ability to swallow oral medication and willing to comply with the study protocol for the duration of the study were included [Symptomatic patients aged 6months (>5kg body weight) and 60years of age with uncomplicated malaria due to monoinfection of included .P. falciparum species as detected by microscopy; febrile conditions caused by a disease other than malaria or another known underlying chronic or severe disease; female patients with positive pregnancy test or breastfeeding, who were unable to drink, had severe vomiting, presence of lethargy or decreased consciousness, inability to sit or stand, were all excluded [Patients with general danger signs or signs of severe falciparum malaria; severe malnutrition; mixed or monoinfections other than excluded .As the treatment failure rate to (artemether-lumefantrine) in the area is unknown, 5% was assumed. At a confidence level of 95% and a precision around the estimate of 5%, a minimum of 73 patients must be included. With a 20% increase to allow loss to follow-up and withdrawals during the 28day follow-up period, 88 patients were required in the study per site.P. falciparum malaria and fulfilling the enrolment criteria were enrolled in the study. The demographic information  were recorded. Two to three drops of finger prick blood was also blotted on to 3 MM filter paper  at the time of enrolment and during the follow-up for molecular study.The patient screening and enrolment started from June 2017 till December 2017. Patients presenting with fever or history of fever and symptoms of malaria were screened for malaria parasites by microscopy. Patients positive for Enrolled patients were clinically examined by on duty medical officer and artemether-lumefantrine tablets were administered orally according to body weight, twice a day over 3days  as per Indian National Malaria Drug Policy 2013 . Female Patient develops danger signs or severe malaria on Day 1, Day 2 or Day 3 and there are parasites in the blood, or the number of parasites in the blood on Day 2 is greater than that on the day of enrolment (Day 0), or axillary temperature on Day 3 is37.5C and there are parasites in the blood on Day 3, and Day 3 parasitaemia25% of those counted at the day of enrolment (Day 0).Danger signs or severe malaria in the presence of parasitaemia on any day between day 4 and day 28 in patients who did not previously meet any of the criteria of early treatment failure; andPresence of parasitaemia on any day between day 4 and day 28 with axillary temperature37.5C in patients who did not previously meet any of the criteria of early treatment failure.Presence of parasitaemia on any day between day 7 and day 28 with axillary temperature<37.5C in patients who did not previously meet any of the criteria of early treatment failure or late clinical failure.Absence of parasitaemia on day 28, irrespective of axillary temperature, in patients who did not previously meet any of the criteria of early treatment failure, late clinical failure or late parasitological failure.Counting of parasite was done on Giemsa-stained thick blood films until 200 white blood cells (WBCs) counted by light microscopy. Parasite density , was calculated as number of asexual parasites divided by the number of WBCs counted, and then multiplying it by an assumed WBC number (6000 per L). When the number of asexual parasites was<100 per 200 WBCs in follow-up smears, counting was done against at least 500 WBCs. A blood slide sample was considered negative when no asexual parasite seen after examination of 1000 WBCs or 100 fields of thick smear. The presence of gametocytes on the day the patient was enrolled or on the day of follow-up was also recorded.Blood smears of enrolled patients, including follow-up smears, were examined by two independent WHO level 1 qualified microscopists. If any discordance was found, a third reading was performed by another senior microscopist. Counting of parasite was also performed by two independent microscopists. If the difference between two readings varied by>25%, counting was again performed by a third microscopist. Each reader was unaware to the result of other reader.Data from both clinical and parasitological assessments for each participant were entered and therapeutic efficacy was analysed using WHO standardized Microsoft Excel data collection sheet. Furthermore, survival analysis among total subjects from all study sites was performed using STATA software version 14.Plasmodium species [msp1, msp2 and k13 gene was performed as per previously published protocol [msp1 & msp2) in the pre-treatment (Day 0) and post-treatment samples (day of parasite observed). Reinfection was diagnosed when all alleles for at least one of the markers differed between the two samples.If any patient was found positive for malaria parasite during follow up period, two to three drops of finger prick blood was also collected on 3 MM Whatman filter paper for identification of mixed infection using molecular tools. Genomic DNA was isolated and species-specific nested PCR was carried out using primers targeting the 18s rRNA gene for the identification of  species . PCR andprotocol , 18. RecSequencing result were analysed with analysis software v5.2 (Applied Biosystems) and sequence alignment was performed using GeneDoc software .P. falciparum monoinfection. A total 12% (224/1915) case of P. vivax and 4% (86/1915) mixed infection of P. falciparum and P. vivax was recorded. Out of 1602 mono P. falciparum cases, a total 376 malaria patients who fulfilled the enrolment criteria as well as consented for the study were enrolled. Therapeutic efficacy was determined in 356 (94.7%) patients who had completed their 28days follow-up while 20 patients were either withdrawn from the study or loss to follow up due to various reasons  with 84% (1602/1915) Background characteristics of the enrolled patients are summarized in Table Among the total 376 enrolled cases, 8 patients were loss to follow-up because of remote area and migration of population for their earning and livelihood. Therefore, these patients could not be traced. A total of 12 patients were withdrawn from the study as they were unable to complete the treatment due to persistent vomiting or failure to attend the scheduled visits during the first three days. However, 356 patients were followed up successfully up to 28days. The adequate clinical and parasitological response (ACPR) without PCR correction was 98.9%; 95% CI (97.199.7) with four cases  of late parasitological failure (LPF) . To understand whether treatment failures were recrudescence or reinfection, DNA samples were sequenced for P. falciparum merozoite surface protein 1 and 2. Out of four cases, two contain the same allele for both the genes msp1 and msp2 among both time (0day and day of failure), whereas other two patients had re-infection in which msp1 and msp2 allele were different at the time of treatment failure.Samples from 0day and day of late parasitological failure (LPF) were analysed for k13 gene. Out of these, successful sequencing was done from 308 samples. Alignment of these sequences from reference sequence  showed one Non-synonymous mutation Q613H in one sample from Madhya Pradesh only, while rest of the samples were wild type.Total 352 samples were taken for amplification of P. falciparum malaria [Plasmodium species mixed infection also have been reported from these study area [In the present study, efficacy of AL was tested in four states . These states are major contributor of malaria in India. Among these, Chhattisgarh 18%) and Odisha (11.7%) is the second and third highest contributor of malaria in the country . AL is r% and Odiudy area , 21. Preudy area 25, Braziudy area , 27.Four cases of LPF were observed from Bastar district of Chhattisgarh in the present study. After genotyping, two cases were reinfection and two cases of recrudescence were revealed. Previous study showed 1 case of LCF and 2 cases of LPF from Bastar region while another study from India also reported 1 case of ECF , 22. Park13 propeller gene is associated with artemisinin resistance. In this study, only one non-synonymous mutation (Q613H) was found in one isolate (0.28%) which is not linked to resistance. Previous study by our group from Madhya Pradesh, India showed M579T (1.6%) which is also not linked to artemisinin resistance [Polymorphism in sistance . Other ssolate 0.% which isistance  and noneP. falciparum malaria in all age groups and in the areas where more than one Plasmodium species are prevalent. Study revealed no functional mutation in k13 gene. This indicates no immediate threat of artemisinin resistance in study area. However regular monitoring of antimalarial drug efficacy and genotyping is important to tract the emerging resistance and alleles circulating in the malaria endemic areas.AL was highly effective for treatment of uncomplicated"}
+{"text": "Plasmodium falciparum malaria transmission. Safety monitoring concerns and the lack of a universal validated and approved primaquine pharmacovigilance tool is a challenge for a national rollout in many countries. This study aimed to explore the acceptance, reliability and perceived effectiveness of the primaquine roll out monitoring pharmacovigilance tool (PROMPT).Primaquine is a gametocytocidal drug recommended by the World Health Organization (WHO) in a single-low dose combined with artemisinin-based combination therapy (ACT) for the treatment and prevention of This study was conducted in three dispensaries in the Coastal region of Eastern Tanzania. The study held six in-depth interviews with healthcare providers and six participatory focus group discussions with malaria patients (3) and parents/guardians of sick children (3). Participants were purposively sampled. Thematic analysis was conducted with the aid of NVivo qualitative analysis software.The respondents\u2019 general acceptance and perceived effectiveness of the single-low dose primaquine and PROMPT was good. Screening procedure for treatment eligibility and explaining to patients about the possible adverse events was considered very useful for safety reasons. Crushing and dissolving of primaquine tablet to get the appropriate dose, particularly in children, was reported by all providers to be challenging. Transport costs and poor access to the health facility were the main reasons for a patient failing to return to the clinic for a scheduled follow-up visit. Treatment was perceived to be safe by both providers and patients and reported no case of a severe adverse event. Some providers were concerned with the haemoglobin drop observed on day 7.Single-low dose primaquine was perceived to be safe and acceptable among providers and patients. PROMPT demonstrated to be a reliable and user-friendly tool among providers. Further validation of the tool by involving the National Malaria Control Programme is pivotal to addressing key challenges and facilitating primaquine adoption in the national policy. About 3.2\u00a0billion people are at risk of malaria despite the progressive global decline of malaria prevalence. In 2019, there were over 200\u00a0million new cases of malaria and 409,000 deaths; approximately 94% occurred in sub-Saharan Africa More than two-thirds of the providers found the PROMPT data collection form useful as it helped them ensure that they had noted down all the necessary information required in making a treatment decision. They were not used to only one person completing a similar form in a specific fixed order. They mentioned the latter as one of the reasons for a prolonged time in attending to malaria patients. They suggested that the form should be shortened and that this should be incorporated with the existing medical information system.\u201cI only have experience of one patient presented with dark urine after using the treatment, we told him to come back on the following day and confirmed his urine was dark, but we did not check his Hb because day 0 Hb was around 12 points and something (12\u00a0mg/dl). We then gave him 4 sachets of ORS (for 4\u00a0l) and encouraged him to use them at home; after two days, he called us again and was happily informing us he is doing ok\u201d. IDI, Nurse aged 46All HCP reported no cases of severe adverse events such as death, severe anaemia or case referred for blood transfusion or any reason for using SLD primaquine in the management of uncomplicated malaria. The treatment safety concern mentioned by nearly all providers was the drop of haemoglobin level in many patients observed on day 7 of follow up but not to the level that may require a blood transfusion. Two providers each mentioned to have seen the patient coming back complaining of dark urine, and after 2\u00a0days, the urine was clear. HCPs mentioned managing this condition by prescribing oral rehydration therapy (ORS) and was used at home for 2\u00a0days. None of the providers received a patient with serious complaints linked to primaquine, such as stomachache, nausea or vomiting.\u201cThe treatment has reduced a big number of patients returning to our facility with malaria in a few days after completing malaria treatment. Now it can take several months for patients to return to the facility for malaria treatment\u2026 I can say this new medicine has also reduced the number of malaria attended cases\u201d. Nearly half of the providers mentioned the treatment has helped reduce the turn-up of patients to the facility due to recurrence of malaria. This was observed since they started including SLD primaquine in treating patients with ACT.\u2026 every month, my children were suffering from malaria, but since they started getting this treatment, at least a month has now passed without having malaria in my family\u201d. FGD, parents/guardian.A similar experience was also mentioned in one FGD with parents and guardians:The study has shown that a single-low dose primaquine regimen and pharmacovigilance monitoring tool among healthcare providers and patients in a real-world environment is feasible and acceptable. Although some providers considered it time-consuming, patient screening procedures and treatment information were perceived to be good and clear to both providers and patients. Primaquine administration had practical challenges in some patients who needed to crush and dissolve first the tablet to get an appropriate dose. According to the schedule, returning to the clinic for treatment follow-up on day 7 was challenging for patients due to varied reasons.The treatment was generally perceived as safe by both patients and providers with no reported severe adverse events. Providers were concerned about the haemoglobin level drop observed on the scheduled follow-up visit on day 7. Screening for the treatment eligibility criteria as per the PROMPT that includes assessment of the specific patient safety condition through medical history, clinical and laboratory investigations assisted the clinician in identifying life-threatening conditions, particularly severe anaemia. Routine laboratory screening for haemoglobin level is often ignored in patients diagnosed with malaria in many low-income countries, reportedly due to resources constraints . This maNo study facility could examine blood slides to determine parasitaemia level or type of malaria parasite because they had no laboratory technician. Therefore, malaria diagnosis in the present study was based on RDT, the test that is considered more reliable and saves time in resource-limited countries, countries mostly challenged by having unskilled personnel . The curGood providers patients interaction, including understanding patient\u2019s medical condition, provides the patient with relevant detailed information about the treatment, and importance for patients to return to the clinic for follow-up, as emphasized in the PROMPT, have been reported to improve the quality of care and adherence to treatment . PatientThe limited number of health providers in many resource-limited countries, including Tanzania, are overwhelmed with many patients in the clinic. This has been reported to affect the quality of care in these countries . Time coMost patients appreciated the PROMPT patient information sheet provided by healthcare providers on the first day of treatment, though some could not read and write. The latter should not ignore the value of supplement information material to patients because multiple information channels have reported improving understanding and knowledge of a particular subject matter . DisclosPROMPT defined therapeutic dose range is a novel idea from MEI research work that aims to simplify primaquine dosing regimen and minimize errors that may affect drug safety and efficacy to the patients . Most prAdministration of primaquine was reported to be challenging by all providers, particularly where there is a need to crush the tablet first in order to get the appropriate dose based on the patient\u2019s body weight. This is due to a small tablet size of around 6\u00a0mm in diameter, making it almost impossible to split it evenly. Unfortunately, the lack of the WHO prequalification primaquine tablet with lower strength has been reported to be a challenge by manufacturers due to limitations related to formulation processes . A possiSeveral studies have shown follow-up on patients after or through the treatment process to improve patient satisfaction, medical safety and ensure the health-related quality of life , 35. MosP. falciparum malaria [There was no severe adverse outcome observed or reported by either providers or patients. This agrees with the rich available information about the safety of SLD primaquine in treating  malaria \u20139. Provi malaria . WhetherThis study provided findings on acceptability and the safety monitoring of SLD primaquine in routine healthcare practice in rural and the lowest level of health facility delivery in low-income countries. Acceptance of the SLD primaquine and the use of PROMPT among healthcare providers in managing malaria is encouraging. The challenges observed in the present study calls upon the need to validate and customize the primaquine safety monitoring tool to a local context by working with the national malaria control programme before SLD roll-out."}
+{"text": "Determine the minimum dosage of alanyl-glutamine (Ala-Gln) required to improve gut integrity and growth in children at risk of environmental enteropathy (EE).z-scores less than \u22121. We randomized children to 10 days of nutritional supplementation: Ala-Gln at 3 g/day, Ala-Gln at 6 g/day, Ala-Gln at 12 g/day, or an isonitrogenous dose of glycine (Gly) placebo at 12.5 g/day. Our primary outcome was urinary lactulose-mannitol excretion testing. Secondary outcomes were anthropometry, fecal markers of inflammation, urine metabolic profiles, and malabsorption .This was a double-blinded randomized placebo-controlled dose-response trial. We enrolled 140 children residing in a low-income community in Fortaleza, Brazil. Participants were 2 to 60 months old and had weight-for-age (WAZ), height-for-age (HAZ), or weight-for-height (WHZ) P=0.05). We observed significant but transient improvements in WHZ at day 10 in 2 Ala-Gln groups, and in WHZ and WAZ in all Ala-Gln groups at day 30. We detected no effects on fecal inflammatory markers, diarrheal morbidity, or urine metabolic profiles; but did observe modest reductions in fecal energy and fecal lactoferrin in participants receiving Ala-Gln.Of 140 children, 103 completed 120 days of follow-up (24% dropout). In the group receiving the highest dose of Ala-Gln, we detected a modest improvement in urinary lactulose excretion from 0.19% on day 1 to 0.17% on day 10 (Intermediate dose Ala-Gln promotes short-term improvement in gut integrity and ponderal growth in children at risk of EE. Lower doses produced improvements in ponderal growth in the absence of enhanced gut integrity. Childhood undernutrition and diarrhea remain leading causes of death and disability in low- and middle-income countries (LMIC), unacceptably impeding progress towards 2030 global targets for childhood nutrition and survival . EnvironGut-trophic \u201crepair nutrients\u201d remain an area of intense interest in the search for adjunct therapies for EE . GlutamiWe designed the Intervention and Mechanisms of Alanyl-Glutamine for Inflammation, Nutrition, and Enteropathy (IMAGINE) study to address the following questions: what is the lowest dose of Ala-Gln that improves intestinal barrier function and nutritional status in children at risk of underweight, wasting, or stunting? 2) By what immune or metabolic mechanisms, does Ala-Gln exert these potential benefits?NCT01832636) was a prospective randomized double-blinded placebo-controlled dose-response trial conducted in the Parque Universit\u00e1rio urban community in Fortaleza, Brazil. A detailed description of this population is available in previous publications ; Ala-Gln at 6 g/day (n=35); Ala-Gln at 12 g/day (n=35); or glycine (Gly) at 12.5 g/day (n=35). Glycine, unlike glutamine, is not a preferential fuel for intestinal epithelial cells. Glycine was selected as a placebo to control for effects of dietary amino acid supplementation at a dose isonitrogenous to the highest dose of Ala-Gln. The Ala-Gln therapeutic supplement  or Gly placebo  was mixed with 50mL of milk, formula, or juice, and study personnel directly observed administration and ingestion. The study design is illustrated in Participants over 10 kg ingested a solution of 5 g of lactulose  and 1 g of mannitol  dissolved in 20mL of water after at least 3 hours of fasting. Children under 10 kg received 2 mL/kg of the solution. Urine was collected over the following 5 hours and 0.236 mg/mL of chlorhexidine  was added before storage at \u221220\u00b0C. Percent excretion of lactulose and mannitol was measured by high-performance liquid chromatography with pulsed amperometric detection .Enzyme-linked immunosorbent assays were carried out to detect the following fecal measures: alpha-1-antitrypsin  as a measure of fecal protein loss and intestinal permeability, myeloperoxidase  as a marker of intestinal mucosal neutrophil activity, neopterin  as a marker of T-helper cell 1 activity, and Reg-1\u03b2  as a measure of intestinal epithelial regeneration. Specimens were stored at \u221220\u00b0C before analysis. After thawing at room temperature, they were diluted in a buffer with protease inhibitors and analyzed via standard curves provided by the assay manufacturers. Fecal lactoferrin (LFF) was used as a marker of intestinal inflammation and was measured via agglutination assay using the LUEKO-TEST kit from TechLab  using previously described methods .g for 15 minutes at 4 \u00b0C then passed through a 0.45mm filter. Fecal cytokines  were measured using the Bio-Plex Pro Human Cytokine 8-plex Assay and the Bio-Plex Manager software was used for data acquisition and analysis .Fecal samples (2.5 g) were initially diluted with a 5.0mL protease inhibitor solution containing 100mL of 1\u00d7PBS, 100 mg of phenylmethylsulfonyl fluoride and 100 mg of soy trypsin inhibitor . This solution was centrifuged at 10,000 Approximately 25 g of feces from a single stool was collected and stored at \u221280 \u00b0C . Specimehttp://links.lww.com/MPG/B879) for methods for anthropometric measurements, urine metabolomics, stool microbiology, diarrheal morbidity, and adverse events.See P=0.05 and 90% power, we calculated a requirement of 22 participants in each experimental group. This calculation was based on data from preliminary studies in the same population (lactulosemannitol ratio=0.13\u00b10.04). A total of 140 participants, or 35 participants per group were enrolled to account for a possible dropout rate of 30%.As an objective measure of overall intestinal barrier function, the lactulose-mannitol test was selected as the primary outcome variable and was used to calculate the sample size. To detect a 30% reduction in lactulose-mannitol ratios with an alpha level of igrowup R package. Means (Ms) and standard deviations (SDs) are reported for normally distributed data and medians (Mdn) with interquartile ranges (IQR) for nonnormally distributed data. Lactulose-mannitol excretion tests and fecal markers were log-transformed before analysis. Unpaired t-tests and Mann-Whitney U- tests were used to analyze normally distributed or nonparametric data, respectively. Chi-squared tests were used to compare the prevalence of abnormal results per group over time. A P-value <0.05 was considered significant.Analyses were conducted on an intention-to-treat basis using GraphPad Prism  and the open-source WHO Of 140 children, 104 (74%) completed all 3 postintervention visits. A total of 36 participants did not complete the full protocol: 22 before day 1; 6 between 30 and 90 days; and 9 between 90 and 120 days. Reasons for noncompletion included change of address , voluntary dropout , and discontinuation for unknown reasons .z-scores, diarrheal history, or ULM ratios, which were within the range of values for healthy children in this community (http://links.lww.com/MPG/B879). At enrollment, 19.5% (n=128) of patients had abnormal testing for stool parasites including Giardia lamblia (10.1%), Ascaris lumbricoides (5.5%), Entamoeba coli (3.1%), Cryptosporidium (2.3%), Trichomonas (2.3%), and Enterobius vermicularis (1.5%). Only 2 patients had evidence of 2 concomitant stool parasites. Although fecal markers of inflammation and fecal cytokines were within the range previously reported in healthy controls, fecal calorimetry was slightly elevated compared with previously reported normal values with M (SD) of 5143.2 (505.5) . Fu. Fuz-sco (505.5) .In the Ala-Gln 12 g/day (intermediate dose) group, we detected a significant decrease in percent lactulose excretion between day 1 and 10 , suggesthttp://links.lww.com/MPG/B879).A significant improvement in WHZ was noted in the Ala-Gln 3 g/day (\u0394WHZ=0.095) and Ala-Gln 12 g/day (\u0394WHZ=0.09) groups at day 10 when compared with Gly (\u0394WHZ=\u22120.054). At day 30, a significant improvement in both WHZ  and WAZ  was seen in all treatment groups. This improvement, however, did not persist at day 90 or 120 (http://links.lww.com/MPG/B879).As an increase in ponderal growth at day 30 was observed in the absence of any improvement in barrier function, urine metabolic profiles were analyzed in the children randomized to Ala-Gln 6 g/day to explore the possibility that it was promoting weight gain through metabolic alterations. PCA and OPLS-DA models were constructed to compare the urinary metabolic profiles of children receiving Ala-Gln 6 g/day or the placebo Glycine at baseline and day 10 or 30. No clear between-group variation was, however, observed in either the PCA or OPLS-DA analyses (http://links.lww.com/MPG/B879).There was a trend towards improved absorption in the treatment groups that was not statistically significant. Over 10 days, there was an increase in fecal energy  in the participants receiving Gly. Conversely, fecal energy decreased  in the participants receiving Ala-Gln 3 g/day. Although fecal energy increased in participants receiving 6 and 12 g/day of Ala-Gln , these were relatively modest when compared with the Gly group (P=0.0247) decrease in lactoferrin was observed between day 1  and day 10  in the group receiving Ala-Gln 3 g/day. No changes were noted in fecal A1AT, MPO, NEO, Reg-1B, or cytokines between treatment groups or over time (http://links.lww.com/MPG/B879). The percentage of participants found to have enteric parasites significantly increased (P=0.028) from 19.5% (n=128) to 33.7% (n=74) on day 30, with Giardia accounting for the majority of this increase . Interestingly, Ala-Gln was associated with increasing Giardia prevalence  but Gly was not. Despite this, there was no significant difference in parasite prevalence between groups at any point and a stratified analysis did not show any effect of parasite presence on urinary lactulose-mannitol excretion testing or anthropometry. Fecal GM-CSF was elevated in participants with parasites (P<0.0001).Fecal markers of inflammation and cytokine profiles were obtained on days 1 (n=135) and 10 (n=98). A significant (http://links.lww.com/MPG/B879).A total of 34 (28.8%) adverse events (AEs) occurred in the 118 participants. There were no significant AEs (SAEs) resulting in death, threat to life, hospitalization, or significant disability or incapacity . The mosThe goal of our study was to determine the minimum dose at which Ala-Gln improves barrier function and nutritional status in children in Northeastern Brazil at risk for EE, and to clarify the mechanism through which it exerts these benefits. In this study, we found that 12 g/day of Ala-Gln improved intestinal barrier function as measured by urinary lactulose, which is in agreement with previous work in this patient population using a higher dose of 24 g/day . At this\u22121 \u00b7\u00b7day\u22121) and found no improvements in barrier function (\u22121 \u00b7 day\u22121)  \u201322. It i\u00b7 day\u22121)  and noneGiardia in establishing colonization through a number of potential pathways. Glutamine is a precursor for arginine, which is the main fuel source for Giardia, and also an effector of Giardia-induced intestinal epithelial cell quiescence (In considering trials of higher doses or longer periods of supplementation, safety considerations are important. Although early studies suggested that parenteral glutamine was of benefit to postoperative or trauma patients ,25, it wiescence . Alanineiescence . It is iThere is robust in-vitro evidence that glutamine modulates intercellular tight junctions and can reverse experimentally induced increases in intestinal cell permeability \u201334. In tOur results suggest glutamine has a positive, albeit transient, effect on ponderal growth through mechanisms independent of improvement in intestinal barrier function. Previous clinical trials of glutamine supplementation are mixed, with some showing an improvement in weight gain ,20,41 anAlthough we observed an intriguing relative decrease in fecal energy in patients receiving Ala-Gln versus placebo, we cannot confidently attribute superior ponderal growth in the treatment groups to enhanced intestinal absorption. Quantifying energy absorption in an LMIC community setting is challenging, and the multiday long collection periods standardly employed for metabolic balance studies would have been difficult for a study of this size. To our knowledge, utilization of spot fecal calorimetry in children with EE is unprecedented and limited in its inability to account for variation in meals, fecal energy, or stool volume. In a comparison of cystic fibrosis patients, a spot fecal calorimetry measurement was comparable with a 72-hour collection method .The search for repair nutrients to curtail stunting and its sequelae remains open to the imagination. EE is thought to result from interactions of marginal diets and heightened exposure to gut pathogens early in life leading to a maladaptive cycle of dysbiosis, inflammation, and impaired permeability and absorption. Such stresses can lead to metabolic derangements with implications for growth, development, and health. For example, children from a single community with either Kwashiorkor, marasmus, stunting, or normal growth have unique metabolic phenotypes , and metThe IMAGINE study was limited by several factors. First, we did not include a 24 g/day dose of Ala-Gln arm that would have allowed us to make direct comparisons with our earlier trial. Second, the participants we recruited were less stunted and had lower lactulose excretion than those recruited using similar criteria in previous trials from the same population . Hence, Giardia infection, the durability of Ala-Gln-mediated improvements in barrier function or ponderal growth, effects in populations with a higher burden of EE and stunting, and combination therapies addressing pathogen burden and inflammation.In conclusion, intermediate and lower doses of Ala-Gln promote transient improvements in ponderal growth in children at risk for EE in the absence of a consistent measurable improvement in intestinal barrier function. Despite having assessed a wide span of measures, we were unable to identify a single overriding mechanism through which Ala-Gln exerts this benefit, but would posit that it may modulate inflammation or absorption. Future studies should elucidate the relationship between Ala-Gln and Supplemental material"}
+{"text": "Plasmodium vivax commonly co-exists with Plasmodium falciparum, and which accounts for\u2009~\u200940% of the total number of malaria infections in the country. Regardless of the growing evidence over many decades of decreasing sensitivity of this parasite to different anti-malarial drugs, there has been no comprehensive attempt made to systematically review and meta-analyse the efficacy of different anti-malarial drugs against P. vivax in the country. However, outlining the efficacy of available anti-malarial drugs against this parasite is essential to guide recommendations for the optimal therapeutic strategy to use in clinical practice. The aim of this study was to synthesize evidence on the efficacy of anti-malarial drugs against clinical P. vivax malaria in Ethiopia.Ethiopia is one of the few countries in Africa where P. vivax were included in the review.\u00a0Data were analysed using Review Manager Software. Cochrane Q (\u03c72) and the I2 tests were used to assess heterogeneity. The funnel plot and Egger\u2019s test were used to examine risk of publication bias.All potentially relevant, peer-reviewed articles accessible in PubMed, Scopus, Web of Science, and Clinical Trial.gov electronic databases were retrieved using a search strategy combining keywords and related database-specific subject terms. Randomized controlled trials (RCTs) and non-randomized trials aiming to investigate the efficacy of anti-malarial drugs against I2\u2009=\u200986%, p\u2009<\u20090.0001) and publication bias . Different anti-malarial drugs showed varied efficacies against vivax malaria. The duration of follow-up significantly affected the calculated efficacy of any given anti-malarial drug, with longer duration of the follow-up (42\u00a0days) associated with significantly lower efficacy than efficacy reported on day 28. Also, pooled PCR-corrected efficacy and efficacy estimated from altitudinally lower transmission settings were significantly higher than PCR-uncorrected efficacy that estimated for moderate transmission settings, respectively.Out of 1294 identified citations, 14 articles that presented data on 29 treatment options were included in the analysis. These studies enrolled 2144 clinical vivax malaria patients. The pooled estimate of in vivo efficacy of anti-malarial drugs against vivax malaria in Ethiopia was 97.91% (95% CI: 97.29\u201398.52%), with significant heterogeneity (P. vivax mono-infections. The addition of primaquine (PQ) to CQ is recommended, as this is the only approved way to provide radical cure, and thus ensure sustained efficacy and longer protection against P. vivax. Continuous surveillance of the efficacy of anti-malarial drugs and clinical trials to allow robust conclusions remains necessary to proactively act against possible emergence and spread of drug-resistant P. vivax in Ethiopia.The overall efficacy of anti-malarial drugs evaluated for the treatment of vivax malaria in Ethiopia was generally high, although there was wide-ranging degree of efficacy, which was affected by the treatment options, duration of follow-up, transmission intensity, and the confirmation procedures for recurrent parasitaemia. Regardless of evidence of sporadic efficacy reduction reported in the country, chloroquine (CQ), the first-line regimen in Ethiopia, remained highly efficacious, supporting its continuous utilization for confirmed The online version contains supplementary material available at 10.1186/s12936-021-04016-2. Plasmodium vivax is the most widespread malaria parasite species, and it infects around 14 million people globally every year [P. vivax was considered a species that seldom circulated in sub-Saharan Africa. In Ethiopia, and some East African countries, it is a clear source of malaria infections and clinical disease [Plasmodium parasites in most malaria-endemic regions of the world, together with various other biological challenges, which threaten further progress.ery year , 3. Befoery year , P. viva disease . During  disease . The rem disease . HoweverP. vivax in Ethiopia during the last two decades [P. vivax-endemic countries however, this drug is no longer in use as CQ-resistant\u00a0P. vivax (CRPv) parasites have emerged and become widely disseminated [P. vivax\u00a0could increase vulnerability to other health problems and ultimately lead to severe outcomes [Regardless of the growing evidence for the decreasing efficacy of chloroquine (CQ) against  decades , CQ rema decades . In someeminated , or becaoutcomes . In addioutcomes , such asoutcomes , 15. Sinoutcomes , 17. Hypoutcomes  or spleeoutcomes , are seroutcomes , 21.P. vivax to CQ [Many studies from Ethiopia have reported a decreasing sensitivity of ax to CQ \u201324, althax to CQ \u201327. Althax to CQ , 28\u201330. The study was conducted in accordance with Preferred Reposting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. The protocol for this review was registered at PROSPERO International Prospective Register of Systematic Reviews, with ID: CRD42020201761 .African Journal Online (AJOL) (n\u2009=\u20092). Grey literature and non-published data were not included in the review. Results from different database searches were aggregated and any duplicated data/studies were removed.Related articles were gathered from the major electronic databases: PubMed n\u2009=\u20091057), Web of Science (n\u2009=\u200987), Scopus n\u2009=\u2009132), and Clinical Trial.gov (n\u2009=\u200918)  independently screened titles and abstracts of all records identified by the search strategy for potential inclusion in the review. Thereafter, full-text copies of articles deemed potentially relevant were retrieved and their eligibility was assessed. Disagreements between individual judgements were resolved through discussion. All excluded studies were listed and reasons given for their exclusion , and malaria transmission stratification (low (1751 and 2000\u00a0m), moderate (1001 and 1750\u00a0m), and high (<\u20091000\u00a0m)) as per 2021 mapping by the Ministry of Health of Ethiopia [Using a form, the two authors (TK and KB) independently extracted data on study characteristics such as author\u2019s names, study site/region and study period, methodological characteristics ), treatment options , AL alone OR combined with PQ (AL plus PQ)), and doses, follow-up days (28 or 42), gender, age, and outcome characteristics , those excluded/withdrawal, and re-infection with Ethiopia .The risk of bias for each included study was assessed independently using the Cochrane Handbook for Systematic Reviews of Interventions\u00a0. The criStDev = p\u00a0is a proportion of the population with the treatment success. Then, SE was calculated from the StDev using the formula, SE\u2009=\u2009StDevData were analysed using the Cochrane Review Manager (version 5.4) for qualitative and quantitative synthesis. Pooled, estimated treatment efficacy for each study was reported. Standard error of the mean (SE) for each study was calculated from the standard deviation obtained using the formula, 2) and the I2 tests. For the Cochrane\u2019s test, a p-value of the \u03c72 test less than 0.05 was considered as significant statistical heterogeneity. I2 values of 25%, 50% and 75% were considered to represent low, medium and high heterogeneity, respectively.\u00a0Due to considerable heterogeneity , a random effects model was used to obtain the pooled, estimated in vivo efficacy of anti-malarial drugs against clinical vivax malaria.Heterogeneity between studies was assessed using Cochrane\u2019s Q , OR AL alone or in combination of PQ), follow-up durations (28 or 42\u00a0days), and confirmatory tests for recurrent parasitaemia (PCR-corrected and PCR-uncorrected). Forest plots were used to display point estimates and confidence intervals. Publication bias for studies included in the meta-analysis was assessed quantitatively using the Egger\u2019s test and qualitatively by constructing a funnel plot and looking for asymmetry. ArcGIS software version 10.0 was used to sketch a map showing districts/regions from where anti-malarial drug efficacy estimates were reported.A total of 1296 citations/records were initially identified. After the duplicates were excluded, 1109 unique citations were screened and assessed for eligibility. From the remaining 1109 screened at title/abstract level, a total of 1077 records considered irrelevant for the purposes of the study were excluded. At the second phase of record assessment, a total of 32 eligible studies with available full text were carefully reviewed and 14 articles were included for qualitative and quantitative meta-analysis  in different years and seasons , 26\u201328. All 14 articles were written in English, and 10 of them reported results from single-arm, open-label, prospective cohort trials, each of which investigated the efficacy of CQ alone , 37\u201341. P. vivax-infected patients. About 91.89 and 96.08% of the patients achieved parasite clearance on day 2 and day 3, respectively. Likewise, fever clearance was achieved for 80, 89.46 and 96.15% of the patients on day 1, day 2 and day 3, respectively. In all studies except one, complete parasite and fever clearance were achieved on day 7 [All studies included in this meta-analysis reported the efficacy of anti-malarial drugs in clearing parasites and fever in on day 7 , for eacon day 7  study, oThe majority of studies, except for two , 29, fulI2\u2009=\u200986%, p\u2009<\u20090.0001). Indeed, the efficacy of anti-malarial drugs against P. vivax across individual studies varied considerably, ranging from 73.3% for AL on day 28 [p\u2009<\u20090.0001) , with a very significant high level of heterogeneity . The combination of CQ plus PQ showed greater and consistent therapeutic efficacy  than CQ alone. On the other hand, AL alone, irrespective of post-treatment follow-up periods, showed significantly the lowest  efficacy against P. vivax compared to other treatment options, but its supplementation with PQ resulted in enhanced efficacy . The efficacy of the different anti-malarial drugs against clinical vivax malaria considered in the current meta-analysis did appear to significantly affect the pooled estimate prevalence of P. vivax  , was 96.85% , with a high level of heterogeneity  significantly affected the overall pooled efficacy of anti-malarial drugs against P. vivax . Treatment efficacy of anti-malarial drugs reported on day 28 showed significantly higher efficacy  compared to the efficacy reported on day 42  . None of the included studies reported data from high malaria transmission settings in Ethiopia   than in altitudinally lower transmission areas . The transmission setting significantly affected the overall calculated efficacy of anti-malarial drugs  as compared to the overall pooled estimated efficacy reported for all treatment options (28 and 42\u00a0days)  . There was significantly reduced therapeutic efficacy as regards PCR-uncorrected efficacy reports  as compared to treatment failures that were PCR-corrected   revealed significant heterogeneity and differences between the pooled efficacy of anti-malarial drugs , which is well above the recommended WHO threshold for anti-malarial efficacy (\u2265\u200990%). This pooled, estimated efficacy was affected by the treatment options, duration of the follow-up, transmission intensity, and confirmatory tests for the recurrent parasitaemia. In all the analyses, there was substantial unexplained, high heterogeneity within the studies included. Hence, the validity of the effect estimated for each sub-group is uncertain as individual studies varied in terms of treatment type, follow-up duration, and confirmatory tests for the efficacy.This systematic review and meta-analysis was conducted with the aim of reviewing studies that reported results of in vivo anti-malarial drug therapy for clinical vivax malaria in Ethiopia. Those studies that focused on the therapeutic efficacy of different anti-malarial drugs against Pv in the country.The main drug investigated in all individual studies included in this review was CQ, the current first-line treatment for vivax malaria in Ethiopia. For this drug, the pooled, estimated efficacy was 96.85% (95% CI: 95.85\u201397.86). The slight efficacy improvement observed on day 28 post-treatment for CQ  in comparison with the greater overall, pooled, estimated efficacy for CQ (on day 28 and day 42) could be attributed to the drug\u2019s longer, indirectly monitored elimination time, which could reflect continued protection against re-infection and suppression of early relapses . AlthougP. vivax is the dominant malaria parasite and accounts for\u2009~\u200950\u201370% [P. vivax transmission areas. The rest of the studies were conducted in the southwest of the country , where P. falciparum is the dominant malaria parasite, and P. vivax accounts for only\u2009<\u200940% of total infections (\u2018low P. vivax transmission areas\u2019) [P. vivax has been less prevalent , anti-malarial drugs have shown excellent efficacy, estimated at 98.18%, (95% CI: 97.5\u201398.85) [Most of the articles included in this review reported data from studies conducted in north-central and central Rift Valley areas, and southwest of Ethiopia. Nine out of 16 studies reported data from north-central and central Rift Valley regions , where ~\u200950\u201370% , 45\u201347 o~\u200950\u201370% , these a areas\u2019) , 27, 37. areas\u2019) , 49. In  areas\u2019) , 28\u201330. 5\u201398.85) , 39, 41.P. vivax infection. Findings in this review further highlight the emergence and wider spread of CQ-resistant P. vivax strains in different parts of the country [P. vivax, mainly mutation of the genes responsible for the observed resistance, including Pvmdr-1 and Pvcrt-0, from the same study sites  where the development and expansion of CRPv in the country was confirmed [Pvcrt and\u00a0Pvmdr-1genes, two of the non-synonymous mutations at Y976F and F1076L were identified in the majority of the CQ-resistant P. vivax isolates.CQ has been in use for more than 60\u00a0years for the treatment of vivax malaria in Ethiopia . Althoug country \u201327. Manyonfirmed \u201355. In tPv has become a serious concern [Treatment with CQ and PQ, which offers a blood schizontocidal and hypnozoitocidal therapy (CQ 25\u00a0mg/kg for 3\u00a0days plus PQ 0.25\u00a0mg/kg for 14\u00a0days) significantly improved the therapeutic efficacy to 99.99%, even under conditions of longer follow-up, although the observation was based on few studies. As has been indicated above, the efficacy of the blood schizontocidal drug CQ can slowly diminish and fall to below MEC with loss of protection against re-infection with new parasites or relapse of the initial infection. Its supplementation with PQ could help to clear hypnozoites from the liver and protect against relapses. Besides its efficient hypnozoitocidal activity, reports showed that PQ could enhance the efficacy of CQ even in a setting where CR concern \u201359. The  concern , 17. HenP. vivax (85.43%) irrespective of the duration of follow-up. Because of its shortest elimination half-life (3\u20136\u00a0days), and its fastest-dropping concentration to below MEC, this drug combination (AL) could not protect patients from any relapse or re-infection that might appear as of the 21st day after initial infection in tropical regions [Pv in different regions of Ethiopia, CQ has still shown superior efficacy over AL for the treatment of vivax malaria in Ethiopia.AL, on the other hand, which is first-line treatment for falciparum malaria in Ethiopia , showed  regions . DespiteP. vivax\u00a0parasitaemia\u00a0following treatment is an indicator of treatment failure. However, classifying this treatment failure into recrudescent or new infections that appeared during follow-up in high malaria transmission areas is crucial, albeit currently challenging [Recurrent\u00a0llenging . PCR-corllenging . This collenging , 28\u201330 cP. vivax in Ethiopia and which were finally selected for inclusion were few. Secondly, the studies incorporated in the review lacked consistency in respect of follow-up: in some of the studies, the primary endpoint was 28\u00a0days, whereas it was 42\u00a0days for others. Such discrepancies had a significant effect on the pooled estimate of efficacy of anti-malarial drugs for vivax malaria. Variation in the experimental design among the studies also created significant challenges as regards using similar tools for efficacy analysis and drawing clear conclusions concerning the efficacy estimates for the drugs. In most of the studies considered for this review, recurrent parasitaemias were neither genotyped nor compared with the pre-treatment parasitaemia, and recurrent parasites were not checked to evaluate whether they were perhaps due to re-infection with different strains of the parasite or possibly due to the result of relapse involving different genotype. In addition, most of the studies were focused on CQ efficacy testing. For other anti-malarial drugs or combinations, the available studies were insufficient to make comparisons, and to assess their effects on the overall, estimated pooled efficacy. Furthermore, some published studies included only short methodological and results sections, and it was difficult to extract relevant information/data for further analysis. High heterogeneity of study design, which requires further explanation and determination of the causes was another challenge encountered during the course of the current review processes.Some of the limitations of this analysis were: firstly, the number of studies that focused on in vivo anti-malarial drug efficacy testing against The efficacy of different anti-malarial drugs evaluated for the treatment of vivax malaria in Ethiopia has shown a wide range of variability. Drug efficacy was mainly affected by the treatment options, duration of follow-up, malaria transmission settings, and the recurrent parasitaemia confirmation procedures. Those anti-malarial drugs supplemented with PQ showed excellent efficacy (up to 99.9%) when compared to any other options irrespective of the duration of follow-up and treatment options. By contrast, AL alone showed significantly lower efficacy against clinical vivax malaria. Regardless of strong evidence for the decreasing efficacy of CQ, the first-line regimen for the treatment of vivax malaria in Ethiopia, this review shows that CQ still has good efficacy in the country, and that urgent replacement with other anti-malarial drugs may not be needed nor justifiable, at least in the short term. On the other hand, supplementation of CQ with PQ could enhance efficacy, and might serve as an optional regimen for the treatment of vivax malaria in the country, provided a patient\u2019s safety in terms of haemolysis risk is minimized. Regular monitoring and continuous surveillance of the efficacy of CQ remains necessary to minimize the risk of the spread of CQ-resistance.Additional file 1: Fig. S1. Risk of bias assessment graph (a) and summary (b) of studies on in vivo efficacy of antimalarial drugs against P. vivax malaria in EthiopiaAdditional file 2: Fig. S2. Funnel plot for publication bias assessment of studies on in vivo efficacy of antimalarial drugs against clinical P. vivax malaria in Ethiopia.Additional file 3: Table S1. Summary of search keywords/terms. Table S2. Excluded studies and reasons for exclusion of studies on in vivo efficacy of anti-malarial drugs against clinical vivax malaria in Ethiopia. Table S3. ROB-2 tools for randomized and non-randomized studies on in vivo efficacy of anti-malarial drugs against clinical vivax malaria in Ethiopia"}
+{"text": "Plasmodium falciparum malaria.The duration of trial follow-up affects the ability to detect recrudescent infections following anti-malarial treatment. The aim of this study was to explore the proportions of recrudescent parasitaemia as ascribed by genotyping captured at various follow-up time-points in treatment efficacy trials for uncomplicated P. falciparum malaria recrudescence. Simulation studies were conducted based on the constructed distributions to quantify the absolute overestimation in efficacy due to sub-optimal follow-up.Individual patient data from 83 anti-malarial efficacy studies collated in the WorldWide Antimalarial Resistance Network (WWARN) repository with at least 28\u00a0days follow-up were available. The temporal and cumulative distributions of recrudescence were characterized using a Cox regression model with shared frailty on study-sites. Fractional polynomials were used to capture non-linear instantaneous hazard. The area under the density curve (AUC) of the constructed distribution was used to estimate the optimal follow-up period for capturing a Overall, 3703 recurrent infections were detected in 60 studies conducted in Africa  and 23 studies conducted in Asia and South America . Using molecular genotyping, 519 (14.0%) recurrences were ascribed as recrudescent infections. A 28\u00a0day artemether-lumefantrine (AL) efficacy trial would not have detected 58% [95% confidence interval (CI) 47\u201374%] of recrudescences in African children and 32% [95% CI 15\u201345%] in patients of all ages in Asia/South America. The corresponding estimate following a 42\u00a0day dihydroartemisinin-piperaquine (DP) efficacy trial in Africa was 47% [95% CI 19\u201390%] in children under 5\u00a0years old treated with\u2009>\u200948\u00a0mg/kg total piperaquine (PIP) dose and 9% [95% CI 0\u201322%] in those treated with\u2009\u2264\u200948\u00a0mg/kg PIP dose. In absolute terms, the simulation study found that trials limited to 28\u00a0days follow-up following AL underestimated the risk of recrudescence by a median of 2.8 percentage points compared to day 63 estimates and those limited to 42\u00a0days following DP underestimated the risk of recrudescence by a median of 2.0 percentage points compared to day 42 estimates. The analysis was limited by few clinical trials following patients for longer than 42\u00a0days  and the imprecision of PCR genotyping which overcalls recrudescence in areas of higher transmission biasing the later distribution.Restricting follow-up of clinical efficacy trials to day 28 for AL and day 42 for DP will miss a proportion of late recrudescent treatment failures but will have a modest impact in derived efficacy. The results highlight that as genotyping methods improve consideration should be given for trials with longer duration of follow-up to detect early indications of emerging drug resistance.The online version contains supplementary material available at 10.1186/s12936-021-03980-z. Before the introduction of molecular genotyping, the World Health Organization (WHO) recommended that anti-malarial clinical trials conducted in areas of high transmission should be restricted to a short follow-up duration 2\u00a0weeks). This was in part to reduce potential confounding from new infections which then could not be distinguished from recrudescences . One of \u00a0weeks. TPlasmodium falciparum genes, paving the way for classifying probabilistically individual recurrent infections as either homologous  or heterologous   in areas of high transmission. With 42\u00a0days of follow-up, the overestimation in efficacy fell to 0.9 percentage points  , the median hazard ratio (HR) for AL relative to DP (for recrudescence) was 2.00 [IQR: 1.29\u20132.78] at day 28, 1.30 [IQR: 1.07\u20131.68] on day 42 and fell to parity from day 49 onwards , any remaining parasites can grow and expand, eventually leading to patent infection and symptomatic illness; during this period, the instantaneous risk of recrudescence rises. In a population of patients the hazard of recrudescence rises as blood concentrations of the antimalarial drug fall below MIC values and then falls again as the majority of recrudescences have occurred. New infections are also constrained by residual concentrations of the slowly eliminated antimalarial drugs and \u201cbunch\u201d together after blood levels fall below prevailing MIC values [Using data from 83 anti-malarial clinical trials conducted in Asia, Africa and South America, this analysis characterized the temporal and cumulative distributions of recrudescence after treatment with AL or DP in patients with uncomplicated C values .Characterization of the temporal trend enabled the construction of the probability distribution of the infections estimated to be recrudescences. In African children less than 5\u00a0years old treated with DP, the peak distribution of recrudescence was around day 39 after initiation of therapy in children who were adequately dosed , but this fell to day 30 in children treated with a lower total dose of piperaquine (<\u200948\u00a0mg/kg). This is consistent with patients treated with a lower dose having blood concentrations which fall below the MIC sooner, allowing the parasite biomass to become patent earlier.In patients treated with AL there were significant regional differences in the probability distributions of recrudescent infections with a shift to the left in Asia and South America  compared to Africa (in children\u2009<\u20095\u00a0years). This may reflect suppression of the parasite growth by host immunity, which is acquired earlier in life in Africa, where children often have multiple infections per year, whilst immunity is acquired more slowly in low endemic settings in Asia and South America . HoweverTo estimate the degree to which absolute efficacy might be overestimated, simulation studies for AL and DP regimens were undertaken. Compared to the efficacy estimates at day 63, studies of AL restricted to 28\u00a0days follow-up overestimated drug efficacy by a median of 2.8% (range: 0.6%\u20135.5%), whereas studies of DP restricted to 42\u00a0day follow-up overestimated efficacy in adequately dosed DP by a median of 2.0% (range: 0%\u20134.6%). These results suggest that follow-up of 28\u00a0days for AL and 42\u00a0days for DP may result in failure to detect early signs of recrudescent parasitaemia occurring after the end of follow-up. Overall, the impact of follow-up on estimated treatment efficacy was modest, and any additional gain in accuracy in deriving these estimates must be weighed against the increased logistical challenge of studies with longer follow-up and a higher risk of misclassification when defining recrudescences and reinfections.Recrudescent parasitaemia is the primary determinant for the selection and onward transmission of de novo resistant parasites . Such seThe analysis presented has a number of important limitations. First, the estimation of the hazard function is vulnerable to the method used for estimation, especially on the distal part of the distribution (See Additional file The derived empirical distribution of recrudescence indicates that the currently recommended minimum follow-up for anti-malarial efficacy trials do not capture a significant proportion of PCR confirmed recrudescences occurring after AL and DP treatment. Whilst the overall impact of this limitation on the estimated efficacy of these anti-malarial regimens was modest in absolute terms, extension of the duration of follow-up to 42\u00a0days for AL and 63\u00a0days for DP, particularly with more precise methods of genotyping, would facilitate detection of early signs of emerging drug resistance, which can manifest through delayed parasite recrudescence.Additional file 1. Further methodological details and results.Additional file 2. List of studies used."}
+{"text": "In Uganda, artemether-lumefantrine (AL) is first-line therapy and dihydroartemisinin-piperaquine (DP) second-line therapy for the treatment of uncomplicated malaria. This study evaluated the efficacy and safety of AL and DP in the management of uncomplicated falciparum malaria and measured the prevalence of molecular markers of resistance in three sentinel sites in Uganda from 2018 to 2019.Pfk13 and Pfmdr1 genes were Sanger sequenced and plasmepsin-2 copy number was assessed by qPCR.This was a randomized, open-label, phase IV clinical trial. Children aged 6\u00a0months to 10\u00a0years with uncomplicated falciparum malaria were randomly assigned to treatment with AL or DP and followed for 28 and 42\u00a0days, respectively. Genotyping was used to distinguish recrudescence from new infection, and a Bayesian algorithm was used to assign each treatment failure a posterior probability of recrudescence. For monitoring resistance, Pfk13 gene were found in the successfully sequenced samples. In the AL arm, the NFD haplotype  was the predominant Pfmdr1 haplotype, present in 78 of 127 (61%) and 76 of 110 (69%) of the day 0 and day of failure samples, respectively. All the day 0 samples in the DP arm had one copy of the plasmepsin-2 gene.There were no early treatment failures. The uncorrected 28-day cumulative efficacy of AL ranged from 41.2 to 71.2% and the PCR-corrected cumulative 28-day efficacy of AL ranged from 87.2 to 94.4%. The uncorrected 28-day cumulative efficacy of DP ranged from 95.8 to 97.9% and the PCR-corrected cumulative 28-day efficacy of DP ranged from 98.9 to 100%. The uncorrected 42-day efficacy of DP ranged from 73.5 to 87.4% and the PCR-corrected 42-day efficacy of DP ranged from 92.1 to 97.5%. There were no reported serious adverse events associated with any of the regimens. No resistance-associated mutations in the DP remains highly effective and safe for the treatment of uncomplicated malaria in Uganda. Recurrent infections with AL were common. In Busia and Arua, the 95% confidence interval for PCR-corrected AL efficacy fell below 90%. Further efficacy monitoring for AL, including pharmacokinetic studies, is recommended.Trial registration The trail was also registered with the ISRCTN registry with study Trial No. PACTR201811640750761The online version contains supplementary material available at 10.1186/s12936-021-04021-5. Plasmodium falciparum, and one of the fundamental strategies for malaria case management is prompt diagnosis and treatment with artemisinin-based combination therapy (ACT), as recommended by the World Health Organization (WHO) [Malaria is a major public health concern in Uganda, with over 11 million cases registered in 2019 [on (WHO) . In Uganon (WHO) . Similaron (WHO) \u201311. Evidon (WHO) , 11, 12.P. falciparum parasites, and therefore the WHO recommends periodic monitoring [P. falciparum kelch 13 (Pfk13) gene are associated with artemisinin partial resistance, and 10 of these SNPs have a validated association: F446I, N458Y, M476I, Y493H, R539T, I543T, P553L, R561H, P574L, and C580Y [Although ACT remains efficacious in Africa, the success of anti-malarial treatment can be affected by the presence of drug-resistant nitoring . In ACT,nitoring . Howevernitoring , 15 and nitoring . In addind C580Y , 17. Mannd C580Y , 18 and nd C580Y . In bothPfmdr-1 gene, the most commonly studied SNPs include N86Y, Y184F, S1034C, N1042D, and D1246Y. The Pfmdr1 86Y mutation has been associated with chloroquine and amodiaquine tolerance, while the N86 wild type codon has been implicated in decreased susceptibility to lumefantrine [plasmepsin-2 multi-gene cluster on the parasite chromosome 14 and a non-synonymous SNP in a putative exonuclease gene (PF3D7_1362500) on chromosome 13, exo-E415G, has been associated with both in vitro piperaquine resistance and clinical treatment failures [In the fantrine . The 184fantrine . The S10fantrine . A gene failures .P. falciparum anti-malarial resistance in the Pfk13, Pfmdr1 and plasmepsin-2 genes.Monitoring drug efficacy and resistance for early detection is required and enhanced by implementing timely treatment policies in order to mitigate this threat . The focA randomized, open-label phase IV clinical trial was conducted from September 2018 to February 2019 in three health centres in Uganda: Aduku Health Centre IV in Kwania district, Northern Uganda; Arua Regional Referral Hospital in Arua district, northwestern Uganda; and, Masafu District Hospital in Busia district, Eastern Uganda , Basel, Switzerland; 20\u00a0mg artemether/120\u00a0mg lumefantrine) or DP (D-Artepp\u00ae (Forsun), Shanghai, China; 40\u00a0mg dihydroartemisinin/320\u00a0mg piperaquine) supplied by WHO. For both treatment arms, no food was given to the study participants, but the parents were advised to feed their children after administration of the study medication. All the doses of the study medication were administered as directly observed treatment by the study nurse at the study clinic and the study participants were monitored for 30\u00a0min after administration of study drugs. AL was administered twice a day for three days , and DP was administered once a day for three days . The dosage was determined using the weight-based dosage as indicated in the WHO malaria treatment guidelines [Study participants were administered one of two ACT: AL  or 42\u00a0days (DP). Study participants were also encouraged to visit the study clinic on any other day they felt unwell. At each follow-up visit, clinical response to treatment was monitored through a standardized history and physical examination, and parasitological response was assessed through examination of thick and thin blood films. Haemoglobin level was measured on day 0 and at the time of clinical treatment failure (early treatment failure (ETF) or late treatment failure (LCF). Haemoglobin was systematically measured in all patients from day 21 onwards using a portable spectrophotometer (Hemocue). A dried blood spot (DBS) was collected on the day of enrolment and on follow-up days for the parasite molecular studies.Treatment failures, as defined by WHO , were trMalaria parasitaemia was diagnosed using thick smears stained for 10\u00a0min with 10% Giemsa. Follow-up thick and thin smears were stained with 2% Giemsa for 30\u00a0min and used to determine the actual parasite density, species and presence of gametocytes. Parasitaemia was measured by counting the number of asexual parasites against 200 leucocytes in thick blood films, and thin films were used for detection of the different parasite species. Parasite density per \u00b5l of blood was calculated by multiplying the total number of parasites counted by 40, assuming that 1\u00a0\u00b5l of blood had a mean count of 8000 leucocytes , 26. WheP. falciparum.Molecular markers of anti-malarial drug resistance and microsatellite markers were analysed at the Centers for Disease Control and Prevention (CDC) Malaria Laboratory in Atlanta, USA , 28. Par\u00ae V2.6.3 . A Bayesian probabilistic algorithm was used to distinguish recrudescent from new infections, accounting for classification uncertainty with multi-parasite genetic diversity [Paired samples with recurrent infections (day 0 and day of recurrent infection) were analysed to distinguish recrudescence from re-infection using seven neutral microsatellite markers  over six chromosomes , 32. Fraiversity , 33.Pfk13 propeller domain (codon positions: 389-649), as previously described [Pfmdr1  mutations were investigated in paired day 0 and day of recurrent infection samples in the AL arm using a previously described method [Pfk13 and Pfmdr1 were used as reference sequences. Heterozygous SNPs (double peaks representing mixed infections) were identified using the heterozygous caller plug-in tool in Geneious with a minor allele threshold of at least 30%. For samples with mixed infections and SNP variations at multiple sites, each possible haplotype constructed from the observed SNPs was reported for Pfmdr1. Detection of P. falciparum plasmepsin 2 copy number was performed in paired day 0 and recurrent infection samples in the DP arm using an Agilent Mx3005 real-time PCR machine , according to previously described protocols [Sanger sequencing was used to investigate escribed , in all d method . SNPs werotocols Parasitaemia, assessed by microscopy within 28 (AL and DP) or 42 (DP) days of treatment, was either unadjusted or PCR-adjusted to distinguish recrudescence from new infection, and was used to calculate outcomes. Recrudescence was defined as the recurrence of asexual parasitaemia of the same genotype(s) that caused the original illness, due to incomplete clearance of asexual parasites after anti-malarial treatment, while new infection was an infection that followed a primary infection which is often  different from that which caused the initial infection , 14.Outcomes were classified according to WHO guidelines  and inclBoth AL and DP were studied at each site, with a target of 100 study participants in each treatment arm. The sample size calculation was based on estimated risks of treatment failure  of the treatment regimens to be studied . Based oPfmdr1 gene and plasmepsin-2 gene copy number, samples from subjects who failed therapy were evaluated, and for Pfk13, samples from all subjects were evaluated. Incident adverse events were reported by arm.Data were double-entered into Microsoft Access and analysed using Stata, version 14.2 (Stata). Per-protocol  and cumulative efficacy were calculated by arm and site at 28- or 42-days follow-up. Unadjusted outcomes were censored for loss to follow-up or exclusion. Outcomes adjusted by genotyping were censored for loss to follow-up or exclusion, new infections or failure of genotyping. The posterior probabilities of recrudescence generated using the Bayesian algorithm to distinguish recrudescence from re-infection were used to generate the per-protocol  efficacies, and posterior sampling was used to generate the Kaplan\u2013Meier estimates and 95% confidence intervals. For Markers Among 862 patients screened between September 2018 and February 2019, 263 were excluded during screening and 599 were randomized to receive therapy with either AL or DP; baseline characteristics are shown in Table The primary efficacy outcomes are shown in Table Secondary outcomes included the proportion of patients with parasites on days 1\u20133 and proportion of patients with fever after therapy  and at the time of recurrent infection were assessed. No artemisinin partial resistance-associated mutations in the Pfk13 gene was found in the 338/370 (91.4%) successfully sequenced paired day 0 and day of recurrent infections samples. The Pfmdr1 gene was investigated in paired day 0 and day of recurrent infection samples in the AL study arm. SNP results for each of the investigated codons are reported in Table Pfmdr1 haplotype construction was done using data for codons 86, 184 and 1246 in samples that had complete data at these codons  [Pfk13 polymorphisms and their possible association with reduced efficacy of artemisinin in Uganda. Analysis of the Pfmdr1 gene showed that NFD (N86/184F/D1246) and NYD (N86/Y184/D1246), both associated with reduced susceptibility to lumefantrine, were the most common haplotypes observed.In this study, no ctively) . There iPfmdr1 N86 polymorphisms associated with decreased sensitivity to lumefantrine could be contributing to the high levels of recurrent parasitaemia in the AL treatment arm.Prior studies in Uganda have consistently shown high efficacy for DP and AL , 40. TheAs reported in prior studies, treatment with DP provides a longer period of post-treatment prophylaxis than AL , leadingmsp1, msp2 and up to four microsatellites to differentiate recrudescent from new infections [This is the first time that seven neutral microsatellites were used for molecular correction of therapeutic efficacy monitoring results in Uganda, which may partially account for the variation in efficacy between this study and past studies. Previous studies in Uganda used fections , 40. Thefections . The Bayfections . With thOne limitation of this study was that co-administration of AL with a fatty food, as recommended by the product insert, was not consistently enforced. Future studies should ensure a fatty food is provided when administering AL and incorporate laboratory analysis of lumefantrine level to determine whether low drug levels are contributing to low efficacy findings. Another limitation was the expanded age range, 6\u00a0months to 10\u00a0years, due to low transmission in some study areas. The confounding factor of acquired immunity, which would be more probable in older children, may have resulted in overestimation of efficacy compared with studies recruiting children 6\u201359\u00a0months.Pfk13 mutations or increased plasmepsin-2 copy number associated with artemisinin or piperaquine resistance, respectively. Although both AL and DP are still considered appropriate for the treatment of uncomplicated malaria by the Uganda National Malaria Control Division, there is continued need for routine monitoring of the efficacy of anti-malarials, particularly for AL in Busia district.DP offered PCR-corrected efficacy greater than 90% in all three study sites. The point estimate for the PCR-corrected efficacy for AL was below 90% in some areas. There were no Additional file 1: Table S1. Raw genotyping data."}
+{"text": "BackgroundPlasmodium falciparum to antimalarial drugs remains a major impairment in the treatment and eradication of malaria globally. Following the introduction of artemisinin-based combination therapy (ACT), there have been reports of delayed parasite clearance. In Kenya, artemether\u2013lumefantrine (AL) is the recommended first-line treatment of uncomplicated malaria. This study sought to assess the efficacy of AL after a decade of use as the preferred method of managing malarial infections in Kenya. We assessed clinical and parasitological responses of children under 5 years between May and November 2015 in Chulaimbo sub-County, Kisumu, Kenya. Patients aged between 6 and 60 months with uncomplicated P. falciparum mono-infection, confirmed through microscopy, were enrolled in the study. The patients were admitted at the facility for 3 days, treated with a standard dose of AL, and then put under observation for the next 28 days for the assessment of clinical and parasitological responses. Of the 90 patients enrolled, 14 were lost to follow-up while 76 were followed through to the end of the study period. Seventy-five patients (98.7%) cleared the parasitaemia within a period of 48 h while one patient (1.3%) cleared on day 3. There was 100% adequate clinical and parasitological response. All the patients cleared the parasites on day 3 and there were no re-infections observed during the stated follow-up period. This study, therefore, concludes that AL is highly efficacious in clearing P. falciparum parasites in children aged \u22656 and \u226460 months. The study, however, underscores the need for continued monitoring of the drug to forestall both gradual ineffectiveness and possible resistance to the drug in all target users.The resistance of  P. falciparum, P. ovale, P. malaria, P. vivax and P. knowlesi, with P. falciparum being the most severe and the leading cause of morbidity and mortality worldwide . Pregnant women and children are at a higher risk, more so school-going children aged between 5 and 15 years old.In Kenya, CQ-resistant ched 70% . Like otched 70% . Thereafched 70% , 15. AL ched 70% . Based oched 70% , 17. In The Kenya national malaria control guidelines recommended dihydroartemisinin\u2013piperaquine (DHA/PPQ) as the second-line treatment for uncomplicated malaria in the country. Usually, parenteral artesunate is the drug of choice for severe malaria; however, when unavailable, parenteral quinine is given as an alternative. During the period of transition, both ACTs and SP drugs were the drugs of choice . The proP. falciparum-positive patients, especially in Southeast Asia [P. falciparum malaria with AL as stipulated in the WHO protocol.In spite of the successes in uptake, the curative effect of ACT, as with most other drugs, has tended to decline gradually, with studies showing a slowing action of parasite clearance in ast Asia , 20. Dueast Asia . TherefoP. falciparum transmission.We conducted the trial in Chulaimbo Sub-County Hospital, Lake Region of Kisumu County in Western Kenya . The areChildren aged between 6 and 60 months who visited the outpatient clinic at Chulaimbo Sub-County Hospital with signs of uncomplicated malaria from the month of May 2015 to November 2015 were sampled for the present study.P. falciparum and parasitaemia in the range of 2000\u2013200\u00a0000 asexual parasites per microliter of blood, without symptoms of severe and complicated malaria, such as prostration, breathing difficulties, severe anaemia, convulsions and inability to drink or vomiting.We recruited children aged 6 months or above 60 months whose parents gave written informed content. In addition, the study population had the following common characteristics: they all visited the health facility with symptoms of malaria; had the body weight of \u22655\u00a0kg; confirmed mono-infection with Among other things, patients below 6 months or above 60 months or whose body weight was <5\u00a0kg were not considered for participation. Similarly, those with a history of fever for more than 24\u00a0h with temperatures above 37.5\u00a0\u00b0C; multiple infections apart from malaria, such as evidence of liver infections; inability to take drugs orally, or was put under antimalarial medication within the previous 2 weeks were not considered potential participants. The same applied to potential participants whose parents/guardians remained uncooperative or failed to give a signed consent.in vivo study intended to assess the susceptibility of AL to drug resistance after previously being used to treat uncomplicated malaria. The assessment was conducted in accordance with the WHO guidelines [This was a single-arm, prospective, idelines .n\u00a0=\u00a0Z2p(1\u2013P)/e2 as described by Lwanga and Lameshow [The sample size determination was calculated using the formula Lameshow , where:Z\u00a0=\u00a0standard normal deviation of the required confidencen\u00a0=\u00a0the desired sample sizep\u00a0=\u00a0proportion of the target population estimated to have suffered from and received treatment for Malaria. According to the Kenya Malaria Indicator Survey, the prevalence of malaria stands at 38% or 0.38 [ or 0.38 .e\u00a0=\u00a0margin error or the desired precisionZ with 1.96, 0.38 for P and 0.1 for e, the minimum sample size was 1.962\u00a0\u00d7\u00a00.38 (1\u22120.38)/0.12\u00a0=\u00a090.Therefore, upon substituting Approximately, 0.05\u00a0ml of blood from a finger prick was collected. Thick and thin smears were prepared on two different slides. We examined slides stained with 10% Giemsa under the microscope to detect the presence of malaria parasites and estimated parasite densities. This was followed with a thin film blood slide stained with 3% Giemsa for parasite species determination and establishment of the presence of gametocytes. Parasitaemia was measured by counting the number of asexual parasites against 200 leucocytes in thick blood films. Parasite density per \u03bcl of blood was calculated by multiplying the total count by 40, assuming that 1\u00a0\u03bcl of blood had a mean count of 8000 leucocytes . The bloTreatment with AL was done for 3 days in line with WHO weight-based regime . A fixedUpon completion of the dose and recovery, the patients were allowed to go home. Follow-up visits were done on days 7, 14, 21 and 28 or at any time the patient felt unwell. Parents or guardians were informed and encouraged to bring their children to the clinic whenever they showed signs of being unwell and not necessarily wait for scheduled visits. For scheduled visits, parents who did not show up by mid-day of the appointed day were visited at home by a member of the study team and asked to report to the health facility. If a patient was not traced for scheduled follow-up, he/she was classified as lost to follow-up. During the scheduled visits, both clinical and parasitological assessments were performed.in vivo drug trial protocol of 2009 [The classification of treatment outcome was either ETF, LCF, LPF or ACPR based on the WHO  of 2009 .Of the 90 patients enrolled, 76 completed the follow-up at day 28 with ACPR. There was no treatment failure observed.At recruitment, 19.7% (15/76) of the study participants were severely parasitaemic (>10\u00a0000 parasites/\u03bcl) while the rest 80.3% (61/76) were found moderately parasitaemic (1000\u20139999 parasites/\u03bcl). After medication, parasitaemia in the patients under observation was observed to decline as follows: 55.3% of parasitaemia cleared on day 1, 94.3% on day 2 while all the parasitaemia cleared on day 3.Febrile individuals, those with \u226537.5 axillary temperature, accounted for 71.1% (54/76) of the total study population on the day of recruitment. The number decreased to 28.9% (22/76) on day 1 followed by a total return to normal temperatures of 36.5\u00a0\u00b0C on day 2. There were not any febrile cases recorded on subsequent days.et al. 1991 [g for 2\u00a0min. In the final step, the supernatant was transferred into another new tube, vortexed again before being transferred again to a new tube. The DNA product was then stored at \u221220\u00a0\u00b0C.DNA was extracted from dried blood spot as described by Warhurst al. 1991 . Brieflyet al. [2, 400\u00a0nM, 200\u00a0nM of primers, 1\u00a0U of Taq Polymerase and 1\u00a0\u03bcl of DNA template. Amplification was done using PCR that included the initial denaturation at 94\u00a0\u00b0C for 3\u00a0min, followed by 30 cycles of denaturation at 94\u00a0\u00b0C for 1\u00a0min, annealing at 55\u00a0\u00b0C for 2\u00a0min, extension at 72\u00a0\u00b0C for 2\u00a0min and the final extension of 72\u00a0\u00b0C for 10\u00a0min. Finally, the reaction was held at 4\u00a0\u00b0C. The amplicons were analysed by electrophoresis in 1.5% molecular grade agarose gel and visualised by UV trans-illuminator. The expected band size of 800\u00a0bp was scored against 100 base pair DNA ladder.The primers pairs designed by Demas et al.  of 18R-1\u03c72 test and two-sample t test. The time taken before parasite clearance after AL administration was determined using a Kaplan\u2013Meier estimator . Differences between the lost respondents and those that completed the study were analysed using the stimator .P. falciparum following a Pf18sRNA analysis. The response rate and the difference between those who got lost to follow-up and those stayed through were largely insignificant across analytical categories, as follows: gender ; age (mean and s.d. of 32.1 and 10.7 for those who fell off and a mean and s.d. of 32.1 and 11.2 for those who completed). The same trend was observed for weight with a mean and s.d. of 15.6 and 3.2 for those that never completed and a mean and s.d. of 14.1 and 2.9 (t(87)\u00a0=\u00a01.722, P\u00a0>\u00a00.05) for those who completed.A total of 151 children were screened for eligibility to participate in the study from May to November 2015. Ninety of them fulfilled the inclusion criteria and were enrolled in the study. Of the 90 that were recruited, 76 (84.4%) completed the 28-day period while 14 (15.4%) were lost to follow-up . Table 1The therapeutic efficacy outcome is shown in Pfmdr1 gene and K13 propeller gene.A sudden and unprecedented upsurge of resistance to anti-malarial drugs can extensively reverse gains made in the fight and control of malaria , 27. In P. falciparum kelch 13 resulting in a prolonged parasite clearance time, with C580Y being the most noted [in vitro was brought about by the pfkelch13 M476I mutation while sustained parasite survival ex vivo was caused by Y493H, I543T, R539T and C580Y mutations while in vivo parasite, the half-life was amplified by Y493H, R539T, with the most prevalence being C580Y mutation [Pfmdr1 has been associated with increased inhibitory concentration of lumefantrine in in vitro studies [pfmdr1 SNPs including N86 [Although the current study did not include the examination of molecular markers associated with AL resistance, a number of related studies have done so with profound outcomes . Severalst noted . The kelst noted . Anothermutation , 38. Themutation , 39. Res studies . In addiding N86  as well ding N86 .P. falciparum malaria in endemic regions globally [P. falciparum in the study area. Despite the study showing the absence of ETF with low recurrent malaria (1 LTF), the outcome of the Ethiopian study points to a highly therapeutic efficacy of both partners of AL [Among the artemisinin-based combination therapies, artesunate\u2013amodiaquine, DHA/PPQ and AL are currently recommended ACTs for the treatment of uncomplicated globally . Even whglobally , studiesglobally . In the rs of AL .Therefore, factors such as host immunity, nutritional factors, initial parasitaemia level, pharmacokinetics and pharmacodynamics may influence the therapeutic efficacy of a drug apart from inherent parasite susceptibility . Any of P. falciparum, especially at its sexual stages, alongside the capacities of the partner drug on the gametocytes has been adequately demonstrated [Yet another study in Ethiopia recorded a paltry five treatment failures: 1 (1.1%) LTF and 4 (4.5%) were LPF compared with a majority 84 (94.4%) ACPR. The study authenticated the findings of the present study which indicate that the treatment of uncomplicated malaria using AL has a high clearance rate of a 100% ACPR. Indeed, the above cited studies have not only shown high efficacy of AL in the treatment of uncomplicated malaria but also correspond with findings from most east African countries. In addition, the ability of AL to rapidly eliminate nstrated . ParasitP. falciparum [There were no gametocytes detected from day 0 to 28 in the current study. A study in Thailand showed that when artemisinin derivatives were introduced as a component of the first-line treatment, there was a tremendous reduction in the incidence of clinical lciparum . Howeverlciparum . Gametoclciparum . Moreovelciparum . The antlciparum . This shThis study observed that fever was linked to discomfort and was the key clinical manifestation in children with fever above 37.5\u00a0\u00b0C. The AL combination has been acknowledged as a rapid fever reducer and a long-acting drug to prevent recrudescence , 48, 49.P. falciparum in Kenya and beyond. It is observed that the fact that AL has been reliably in use for a decade explains why it is the best-choice medication to treat uncomplicated malaria caused by P. falciparum. Be that as it may, intensive and \u2018regular surveillance of ACT partner drugs needs be conducted\u2019 to not only ensure early detection of resistance to P. falciparum but also guarantee informed decisions by policy makers on matters of malaria treatment.This study concludes that AL continues to be effective against uncomplicated malaria caused by In this study, only those participants that remained under observation for the period of the study (28 days) were relied on for the conclusions presented in this study. It was therefore difficult to decisively rule out treatment failures that may not have been detected or accounted for, especially among participants that got lost to follow-up. We indeed attest that variables used  may not adequately reveal the whole extent of the impact of failure to account for the lost participants. Nevertheless, much as the sample size was small, including those lost to follow-up, the study outcome may reliably be generalised to related contexts."}
+{"text": "Plasmodium falciparum infections in Equatorial Guinea. This study was designed to evaluate the efficacy of these artemisinin-based combinations and detect mutations in P. falciparum kelch13-propeller domain gene (Pfkelch13).Artesunate-amodiaquine (ASAQ) and artemether-lumefantrine (AL) are the currently recommended first- and second-line therapies for uncomplicated P. falciparum infection and other inclusion criteria were sequentially enrolled first in ASAQ and then in AL at each site, and followed up for 28\u00a0days.\u00a0Clinical and parasitological parameters were assessed. The primary endpoint was PCR-adjusted adequate clinical and parasitological response (ACPR). Samples on day-0 were analysed for mutations in Pfkelch13 gene.A single-arm prospective study evaluating the efficacy of ASAQ and AL at three sites: Malabo, Bata and Ebebiyin was conducted between August 2017 and July 2018. Febrile children aged six months to 10\u00a0years with confirmed uncomplicated Pfkelch13 mutations (E433D and A578S), and none of them is the known markers associated with artemisinin resistance.A total 264 and 226 patients were enrolled in the ASAQ and AL treatment groups, respectively. Based on per-protocol analysis, PCR-adjusted cure rates of 98.6% to 100% and 92.4% to 100% were observed in patients treated with ASAQ and AL, respectively. All study children in both treatment groups were free of parasitaemia by day-3. Of the 476 samples with interpretable results, only three samples carried non-synonymous Pfkelch13 mutations associated with artemisinin resistance. Continued monitoring of the efficacy of these artemisinin-based combinations, at least every two years, along with molecular markers associated with artemisinin and partner drug resistance is imperative to inform national malaria treatment policy and detect resistant parasites early.The study confirmed high efficacy of ASAQ and AL for the treatment of uncomplicated falciparum infections as well as the absence of delayed parasite clearance and Trial registration ACTRN12617000456358, Registered 28 March 2017; http://www.anzctr.org.au/trial/MyTrial.aspx Plasmodium falciparum resistance to anti-malarial drugs poses a constant threat to the successful treatment of malaria infections. The emergence and spread of chloroquine and sulfadoxine/pyrimethamine resistances have led the WHO to recommend artemisinin-based combination therapy (ACT) for the treatment of uncomplicated falciparum infection [Countries in the World Health Organization (WHO) African Region bear most of the malaria burden and account for 94% of the estimated global malaria cases (213 million cases) and deaths  in 2019 [P. falciparum kelch 13 (Pfkelch13) propeller domain gene have been found to confer artemisinin partial resistance expressed by delayed parasite clearance [Pfkelch13 parasites (R561H) confering artemisinin partial resistance has been shown to emerge in 2014\u20132015 in one site in Rwanda followed by its expansion in another site located apart 100\u00a0km, several months later [Improved access to effective antimalarial treatments has contributed significantly to a marked reduction in the burden of malaria in recent years . AL folllearance , 5. Sinclearance  leading learance \u201313. MorePfK13 nonsynonimous mutation (M579I), associated with delayed parasite clearance and increased in vitro parasite survival rate as measured by the Ring-stage Survival Assay [Pfkelch13 gene to inform national malaria treatment policy.In Equatorial Guinea, malaria is a major health problem for the entire population 1,355,982) with an estimated 321,438  cases and 652 (440\u20131000) deaths in 2019 ,355,982 . EquatorEquatorial Guinea is located in Central Africa and is divided into two regions: the mainland area, which lies between Cameroon and Gabon, and the island region . About 75% of the country's population lives in the mainland. The study was conducted in selected health facilities in three study sites: Malabo in Bioko Island and Bata and Ebebiyin in Litoral and Ki\u00e9-Ntem provinces in the mainland, respectively Fig.\u00a0. The heaP. falciparum monoinfection, parasitaemia of 500 to 200,000 asexual parasites/\u00b5l, willingness to comply with the study visit schedule, and informed consent from parents or guardians  and glurp (glutamate-rich protein) loci, as described elsewhere [msp1 and msp2 and disparate results are resolved by glurp. Such an analysis demands concommittant results from at least two markers for classification of reinfection or recrudescence compared to three with the standard WHO methodology. If results for only 2 markers were available and results for a third marker were missing, the PCR correction was classified as undertermined. Samples were analysed at Institut Pasteur in Paris, France.Filter paper blood samples were collected from each patient on day-0 and in case of recurrence, on the day of parasite recurrence (from day-7). Samples were dried and stored in individual plastic bags containing desiccant. Each dried blood spot was punched out with a sterile puncher, and the spots were placed in numerical order in a 96-well plate. Parasite DNA was extracted using QIAamp DNA Blood Mini Kit (Qiagen). The DNA samples (day-0 and day of recurrence) were analysed for genotyping of the highly polymorphic regions lsewhere . The genlsewhere . As an elsewhere . In thisPfkelch13 gene (PF3D7_1343700) previously described to be associated with artemisinin resistance [. [P. falciparum (PF3D7_1343700) as a reference sequence. Electropherograms with mixed alleles were considered as mutant for the purpose of mutation frequency estimation. The quality control was assessed by including three blinded quality-control samples  in each 96-well sequencing plate.DNA extracted from day-0 dried blood spots were analysed to detect the presence of mutations in the propeller domain of  PF3D7_133700 prev PF3D7_133700 prevTreatment response was classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF) and adequate clinical and parasitological response (ACPR) before and after PCR correction using the WHO protocol . The priThe study was approved by the Ministry of Health and Social Welfare of Equatorial Guinea (No. 731\u2013150), the Ethics Committee of Spanish National Health Institute, Carlos III (CEI PI 57_2016-v3) and the WHO Research Ethics Review Committee (ERC.000286). Parents or guardians were informed about the study procedure, its benefits and potential risks, and their consent to enroll their children was obtained in writing before enrollment. They gave their written consent before enrolling their children in the study. If a patient, parent, or guardian was illiterate, he or she chose a witness to co-sign the consent form.http://www.who.int/malaria/publications/atoz/9789241597531/en/). Enrolled patients who were lost to follow-up or withdrawn from the study, had recurrent parasitaemia with new infection, or unknown PCR (indeterminate or missing) were excluded from per-protocol analysis. However, these cases were included in the Kaplan\u2013Meier analysis up to the day of loss or withdrawal from the study. Descriptive statistics including percentages, mean, standard deviation, and range were used. Student's t-test was used for analysis of continuous variables (parasite density and age) and Fisher's exact test was used for categorical data. A p-value of\u2009<\u20090.05 was considered significant.Data were double entered, cleaned, and analysed using the software programme WHO excel  and 226 with AL . The target sample size per drug and per site (n\u2009 = \u200988) was achieved except for the group treated with AL in Malabo site due to low malaria transmission. Baseline characteristics  of the study children in the different sites and treatment groups were comparable  showed ACPR of 100% (95% CI 94.6\u2013100%), 100% (95% CI 95.4\u2013100%) and 98.6% (95% CI 92.5\u2013100%) in patients treated with ASAQ in Malabo, Bata and Ebebiyin, respectively  recurrences, changed to recrudescence in 12/41 (29.2%) recurrences, and was undertermined for 7/41 (17.1%) recurrences  had non-synonymous mutations , carried Previous studies evaluating the efficacy of ASAQ in Equatorial Guinea reported a high cure rate with 97.3% ACPR in 2006 before the recommendation of ACT  and 96.6It\u00a0is\u00a0worth noting that malaria transmission varies in the study sites, with a low level in Malabo (Bioko Island) and a moderate to high level in the mainland where\u00a0Bata and Ebebiyin\u00a0sites are located .\u00a0A survePfkelch13 mutations associated with artemisinin resistance in South East Asia, may indicate the absence of partial artemisinin resistance in Equatorial Guinea. Ih addition, the data based on 476 clinical samples of P. falciparum, support that the indigenous M579I mutant, claimed to be associated with delayed parasite clearance and increased in vitro parasite survival rate [All study patients cleared their parasitaemia by day 3, indicating the absence of delayed parasite clearance, and together with lack of the known val rate , did notPfkelch13 mutant parasites associated with artemisinin resistance. Continued monitoring of the efficacy of these artemisinin-based combinations, at least every 2 years, is imperative to inform national malaria treatment policy. In addition, recent evidence of the de novo emergence of the Pfkelch13 mutation (R561H) associated with artemisinin resistance in Africa calls for monitoring molecular markers associated with artemisinin and partner drug resistance to detect resistant parasites early. In this study, the 2/3 algorithm increased the failure rate at high transmission site compared to the standard WHO methodology. Further comparison and validation in different transmission setting are needed before this new suggested algorithm can be systematically implemented for PCR correction.The study confirmed that ASAQ and AL remain highly effective in treating uncomplicated falciparum infections more than a decade after their use in Equatorial Guinea and that there are no known"}
+{"text": "Plasmodium falciparum transmission in areas aiming for elimination of malaria or areas facing artemisinin resistance. We aimed to determine the efficacy of pyronaridine\u2013artesunate and dihydroartemisinin\u2013piperaquine with and without single low-dose primaquine for reducing gametocyte density and transmission to mosquitoes.Pyronaridine\u2013artesunate is the most recently licensed artemisinin-based combination therapy. WHO has recommended that a single low dose of primaquine could be added to artemisinin-based combination therapies to reduce P falciparum malaria mono-infection and gametocyte carriage on microscopy, haemoglobin density of 9\u00b75 g/dL or higher, bodyweight less than 80 kg, and no use of antimalarial drugs over the past week. Participants were randomly assigned (1:1:1:1) to one of four treatment groups: pyronaridine\u2013artesunate, pyronaridine\u2013artesunate plus primaquine, dihydroartemisinin\u2013piperaquine, or dihydroartemisinin\u2013piperaquine plus primaquine. Treatment allocation was concealed to all study staff other than the trial pharmacist and treating physician. Dihydroartemisinin\u2013piperaquine and pyronaridine\u2013artesunate were administered as per manufacturer guidelines over 3 days; primaquine was administered as a single dose in oral solution according to bodyweight . The primary endpoint was percentage reduction in mosquito infection rate  at 48 h after treatment compared with baseline (before treatment) in all treatment groups. Data were analysed per protocol. This trial is now complete, and is registered with ClinicalTrials.gov, NCT04049916.We conducted a four-arm, single-blind, phase 2/3, randomised trial at the Ou\u00e9less\u00e9bougou Clinical Research Unit of the Malaria Research and Training Centre of the University of Bamako . Participants were aged 5\u201350 years, with asymptomatic Between Sept 10 and Nov 19, 2019, 1044 patients were assessed for eligibility and 100 were enrolled and randomly assigned to one of the four treatment groups (n=25 per group). Before treatment, 66 (66%) of 100 participants were infectious to mosquitoes, with a median of 15\u00b78% (IQR 5\u00b74\u201331\u00b79) of mosquitoes becoming infected. In individuals who were infectious before treatment, the median percentage reduction in mosquito infection rate 48 h after treatment was 100\u00b70% (IQR 100\u00b70 to 100\u00b70) for individuals treated with pyronaridine\u2013artesunate plus primaquine  and dihydroartemisinin\u2013piperaquine plus primaquine , compared with \u22128\u00b77% (\u221254\u00b78 to 93\u00b72) with pyronaridine\u2013artesunate  and 50\u00b74% (13\u00b78 to 70\u00b79) with dihydroartemisinin\u2013piperaquine . There were no serious adverse events, and there were no significant differences between treatment groups at any point in the frequency of any adverse events (Fisher's exact test p=0\u00b796) or adverse events related to study drugs (p=0\u00b764). The most common adverse events were headaches (40 events in 32 [32%] of 100 participants), rhinitis (31 events in 30 [30%]), and respiratory infection (20 events in 20 [20%]).P falciparum transmission. The new pyronaridine\u2013artesunate plus single low-dose primaquine combination is of immediate relevance to regions in which the containment of partial artemisinin and partner-drug resistance is a growing concern and in regions aiming to eliminate malaria.These data support the use of single low-dose primaquine as an effective supplement to dihydroartemisinin\u2013piperaquine and pyronaridine\u2013artesunate for blocking The Bill & Melinda Gates Foundation.For the French, Spanish and Swahilil translations of the abstract see Supplementary Materials section. Evidence before this studyPlasmodium vivax hypnozoiticidal therapy. All three studies assessed gametocyte density using microscopy, and none determined gametocyte transmissibility.We searched PubMed on Jan 21, 2021, with no date or language restrictions, for studies assessing treatment with pyronaridine\u2013artesunate (using the search terms \u201cPyronaridine-artesunate\u201d OR \u201cPyramax\u201d OR \u201cAS-PYR\u201d with down selection based on gametocyte detection), or pyronaridine\u2013artesunate plus primaquine ([\u201cPyronaridine-artesunate\u201d OR \u201cPyramax\u201d OR \u201cAS-PYR\u201d] AND \u201cPrimaquine\u201d). Among the 16 clinical or randomised controlled trials (2010\u201318) that met the search criteria, three studies presented data from a single trial done in Kenya (2018), in which gametocytes in patient blood were quantified by molecular methods up to 14 days after pyronaridine\u2013artesunate treatment . Further studies were identified in which gametocytes present before and after pyronaridine\u2013artesunate treatment were quantified by microscopy. None of the identified studies determined gametocyte infectivity to mosquitoes at any time. Narrowing this search to include studies assessing pyronaridine\u2013artesunate in combination with primaquine identified only five relevant clinical trials. Two of these trials were focused on pharmacokinetic interaction between pyronaridine\u2013artesunate and primaquine in healthy volunteers. The three other studies determined the safety and efficacy of pyronaridine\u2013artesunate plus primaquine for treatment of malaria infection: two using a single low dose of 15 mg, one using the 14-day dosing regimen of primaquine for An additional search of PubMed was done on the same date for studies assessing plasmodium infectivity to mosquitoes with mosquito feeding assays after any artemisinin-based combination therapy (ACT) treatment. For this search we used the terms \u201cMalaria\u201d AND (\u201cACT\u201d OR \u201cArtemisinin\u201d OR \u201cArtesunate\u201d OR \u201cDihydroartemisinin\u201d) AND (\u201cDMFA\u201d OR \u201cMFA\u201d OR \u201cMosquito feeding\u201d OR \u201cMosquito feeding assay\u201d). We identified 12 studies reporting mosquito infection data after ACT treatment. Excluding studies without specific timepoint identification, three distinct trials measured mosquito infection up to day 14 after treatment with dihydroartemisinin\u2013piperaquine or artemether\u2013lumefantrine: one done in Kenya (with data reported in two publications) and two in Cambodia. In one Cambodian study, only three (6%) of 48 individuals were infectious to mosquitoes at baseline. No studies assessed mosquito infection after day 14 and no studies were identified that used mosquito feeding assays following pyronaridine\u2013artesunate treatment.Added value of this studyThis is the first clinical trial assessing the malaria transmission-reducing efficacy of pyronaridine\u2013artesunate treatment alone or in combination with single low-dose primaquine, which has been recommended by WHO. The results suggest that pyronaridine\u2013artesunate treatment alone has little efficacy against gametocytes and fails to prevent malaria transmission to mosquitoes shortly after treatment, whereas pyronaridine\u2013artesunate plus single low-dose primaquine is highly effective as a schizonticidal and transmission-blocking drug combination. In addition to these insights, the study design allowed side-by-side comparison of pyronaridine\u2013artesunate and pyronaridine\u2013artesunate plus single low-dose primaquine with a standard second-line ACT (dihydroartemisinin\u2013piperaquine), alone and in combination with single low-dose primaquine. These data also provide the first direct evidence of continued transmission to mosquitoes more than 14 days after ACT treatment: some individuals treated with dihydroartemisinin\u2013piperaquine or pyronaridine\u2013artesunate without primaquine continued to infect mosquitoes for up to 28 days after treatment.Implications of all the available evidencePlasmodium falciparum malaria transmission. Before this study, the transmission-reducing efficacy of pyronaridine\u2013artesunate with or without single low-dose primaquine was unknown. Pyronaridine\u2013artesunate is a new and therefore valuable alternative antimalarial therapy in areas under threat from resistance or partial resistance to current first-line drug combinations. The data on the effects of pyronaridine\u2013artesunate alone provide a resource to policy makers considering treatment prioritisation. Our data on the efficacy of pyronaridine\u2013artesunate and dihydroartemisinin\u2013piperaquine with single low-dose primaquine support the WHO recommendation that ACTs be combined with single low-dose primaquine to rapidly clear gametocytes and prevent transmission in areas fighting the spread of antimalarial drug resistance or in areas aiming to eliminate malaria.The results of this study add to a growing pool of evidence supporting the use of single low-dose primaquine alongside ACT treatment for the immediate prevention of onward Plasmodium life stage that can be transmitted to mosquitoes. Drugs are a key component of malaria control strategies and yet, outside of mass drug administration,Gametocytes are the only Plasmodium falciparum malaria in all malaria endemic regions.P falciparum transmission in areas of artemisinin drug resistance or areas aiming for malaria elimination.Pyronaridine\u2013artesunate is a recommended treatment for uncomplicated P falciparum gametocytes, and to examine the long-term duration of gametocyte circulation and infectivity to mosquitoes after treatment.The transmission-reducing efficacy of pyronaridine\u2013artesunate and pyronaridine\u2013artesunate with single low-dose primaquine have not yet been tested, and there are indications that pyronaridine\u2013artesunate might inhibit primaquine metabolism.P falciparum gametocytes by microscopy ; haemoglobin density of 9\u00b75 g/dL or higher; age 5\u201350 years; bodyweight less than 80 kg; no clinical signs of malaria, defined by fever (\u226537\u00b75\u00b0C); no signs of chronic or severe disease; no allergies to any of the study drugs; and no reported use of antimalarial drugs over the past week. G6PD status was not tested. Exclusion criteria were pregnancy, known allergy to study treatments, clinical signs of severe malaria or hepatic injury or renal impairment, history of liver disease or renal impairment, family history of congenital prolongation of the corrected QT (QTc) interval, current or previous treatment with drugs metabolised by the enzyme cytochrome P450 2D6 (CYP2D6) or known to extend the QTc interval, and blood transfusion in the past 90 days. Before screening and study enrolment, participants provided written informed consent (if they were aged \u226518 years), assent with written parental consent (12\u201317 years), or written parental consent (<12 years).We conducted a four-arm, single-blind, phase 2/3, randomised controlled trial at the Ou\u00e9less\u00e9bougou Clinical Research Unit of the Malaria Research and Training Centre of the University of Bamako . Screening and recruitment were done in the town of Ou\u00e9less\u00e9bougou and surrounding villages. Ethics approval was granted by the Ethics Committee of the Faculty of Medicine, Pharmacy, and Dentistry of the University of Science, Techniques, and Technologies of Bamako , and the Research Ethics Committee of the London School of Hygiene & Tropical Medicine . Before the commencement of screening, our study team of clinicians and technicians met with community leaders, village health workers, and heads of households from each village to explain the study and obtain approval to conduct the study. Village health workers then used a door-to-door approach to inform households of the date and location where consenting and screening would take place. Participants were invited to enrol into the trial if they met the following criteria: positive for Participants were individually randomly assigned (1:1:1:1) in blocks of 12, to one of four treatment groups: pyronaridine\u2013artesunate , pyronaridine\u2013artesunate plus single low-dose primaquine , dihydroartemisinin\u2013piperaquine , or dihydroartemisinin\u2013piperaquine with single low-dose primaquine. An independent statistician at the Malaria Research and Training Centre randomly generated the treatment assignment using Stata version 16, which was linked to participant identification number. The statistician prepared sealed, opaque envelopes with the participant identification number on the outside and treatment assignment inside, which were sent to the study pharmacist. The study pharmacist provided treatment according to the contained assignment; consequently they and the study physician were not masked to treatment assignment, but all other investigators and staff involved in assessing all laboratory outcomes were masked. Participants could ask the study physician which treatment they received at any time.ACT treatments were administered over 3 days  under the direct supervision of the trial pharmacist, as per manufacturer instructions . All treBlood samples were taken for haemoglobin density measurement , thick film microscopy, and molecular analysis of gametocyte density before treatment (day 0) and on days 1 (haemoglobin only), 2, 7, 14, 21, 28, 35, 42, and 49 after treatment . AdditioAnopheles gambiae females were allowed to feed for 15\u201320 min on venous blood samples  through a warmed glass membrane feeder system .For infectivity assessments, 75 insectary-reared Thick film microscopy was performed as described previously,Mosquito infectivity was assessed at three levels: mean number of oocysts in a sample of mosquitoes , the proportion of mosquitoes infected with any number of oocysts , and infectivity of the study participant to any number of mosquitoes . The primary outcome measure was the median percentage change in mosquito infection rate at day 2 (48 h after treatment) within each group compared with baseline (pretreatment), with the same measure at day 7 as a secondary outcome. Percentage change was reported as percentage reduction  and presented with IQR. Other secondary transmission outcomes were mosquito infectivity  at days 2, 7, and at subsequent timepoints in the dihydroartemisinin\u2013piperaquine and pyronaridine\u2013artesunate treatment groups (to measure duration of infectivity without single low-dose primaquine). Secondary gametocyte outcome measures for all timepoints were gametocyte density, prevalence, sex ratio , circulation time, and area under the curve (AUC) of density over time. Safety outcomes at all timepoints were haemoglobin density and the total number of adverse events . Differences in all transmission, gametocyte, and safety outcomes were compared between treatment groups as secondary comparisons. Some secondary and exploratory measures were not included in this report because assays could not be completed due to COVID-19-related constraints; these were measures of histidine rich protein 2/3 (HRP2/3) density, parasite genotyping, and molecular measures of asexual parasite density.Sample size was informed by a previous trial at the same site and with the same outcomes.t tests (t score for difference compared with day 0), and two-way t tests . The proportion of gametocytes that were male was calculated for all values with total gametocyte densities of at least 0\u00b72 gametocytes per \u03bcL.t tests (t score for difference between ACT matched treatment groups). AUC of gametocyte density per participant over time was calculated using the linear trapezoid method using the first 28 days of observation only,10 adjusted AUC values, with adjustment for baseline gametocyte density . All other analyses of quantitative data were performed using Wilcoxon sign rank tests  and Wilcoxon rank sum tests (Z score for difference between ACT matched treatment groups at each timepoint). All comparisons were defined before study completion and analyses were not adjusted for multiple comparisons. For all analyses, the threshold for statistical significance was set at p<0\u00b705. Statistical analyses were done using STATA version 16.0 and SAS version 9.4.Mosquito infection data were analysed at timepoints after baseline only for those individuals who were infectious at baseline ; ClinicalTrials.gov, NCT04049916.This trial is registered with The funder of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report.Between Sept 10 and Nov 19, 2019, 1044 patients were assessed for eligibility and 100 were enrolled and randomly assigned to one of the four treatment groups . Before treatment, 66 (66%) of 100 participants were infectious to mosquitoes, with a median of 15\u00b78% (IQR 5\u00b74\u201331\u00b79) of mosquitoes becoming infected .At 48 h after treatment (day 2), 16 (64%) of 25 participants in the pyronaridine\u2013artesunate group, 19 (76%) of 25 in the dihydroartemisinin\u2013piperaquine group, none of 25 in the pyronaridine\u2013artesunate plus primaquine group, and one (4%) of 24 in the dihydroartemisinin\u2013piperaquine plus primaquine group infected any number of mosquitoes . In indivs dihydroartemisinin\u2013piperaquine plus primaquine, pyronaridine\u2013artesunate vs pyronaridine\u2013artesunate plus primaquine), the addition of primaquine to either treatment resulted in significant reductions in infectivity to mosquitoes, grouped average mosquito infection rate, and oocyst density by day 2 after treatment  of 25 participants in the pyronaridine\u2013artesunate group, 17 (68%) of 25 in the dihydroartemisinin\u2013piperaquine group, none of 24 in the pyronaridine\u2013artesunate plus primaquine group, and one (4%) of 24 in the dihydroartemisinin\u2013piperaquine plus primaquine group were infectious to mosquitoes , representing a small but significant reduction in haemoglobin density relative to baseline in the dihydroartemisinin\u2013piperaquine (mean change \u22123\u00b788% [95% CI \u22126\u00b796 to \u22120\u00b781]), dihydroartemisinin\u2013piperaquine plus primaquine (\u22126\u00b704% [\u20138\u00b752 to \u22123\u00b756]), and pyronaridine\u2013artesunate plus primaquine (\u22126\u00b785% [\u20139\u00b781 to \u22123\u00b788]) treatment groups . HaemoglOverall, 75 (75%) of 100 participants experienced a total of 173 adverse events during follow-up, 162 (94%) of which were categorised as mild severity and 11 (6%) as moderate . There wP falciparum transmission to mosquitoes by 48 h after treatment in asymptomatic gametocyte carriers in Mali. By contrast, P falciparum transmission to mosquitoes continued for up to 28 days after treatment with dihydroartemisinin\u2013piperaquine or pyronaridine\u2013artesunate alone. In line with these findings, gametocyte circulation time and AUC were significantly lower in the groups who received primaquine than in the non-primaquine groups; however, decreases in gametocyte density and changes in gametocyte sex ratio were preceded by the observed reduction in infectivity.This study showed that a single low dose of primaquine (0\u00b725 mg/kg) added to either dihydroartemisinin\u2013piperaquine or pyronaridine\u2013artesunate resulted in total or near-total reduction in P falciparum infection (NCT03814616). ACTs differ in their efficacy against gametocytes, with considerably longer gametocyte persistence following dihydroartemisinin\u2013piperaquine than after artemether\u2013lumefantrine.Pyronaridine\u2013artesunate is being deployed as the first-line antimalarial in areas of west Africa and has the potential to replace current first-line ACTs facing the threat of antimalarial drug resistance.Plasmodium vivax radical cure,P falciparum gametocyte clearance or transmission blockade; pyronaridine\u2013artesunate plus primaquine treatment resulted in total transmission reduction within 48 h, and dihydroartemisinin\u2013piperaquine plus primaquine resulted in near-total reduction. As in previous studies,In efforts to prolong the lifespan of first-line antimalarials and stem the spread of emergent resistance, supplementing ACTs with low doses of primaquine (as recommended by WHO)This study has several limitations. First, we recruited high-density gametocyte carriers, which allowed us to collect high-quality data on post-treatment transmissibility,P falciparum transmission.Given the concerns about the spread of partial artemisinin and partner-drug resistance in southeast Asia, alternative effective antimalarial therapies are needed to ease drug pressure and maintain treatment efficacy. The emergence of de novo kelch13-mutation-associated artemisinin resistance in Africa also highlights the importance of expanding the arsenal of available antimalarial treatments.Anonymised data reported in the manuscript will be made available to investigators who provide a methodologically sound proposal to the corresponding author. The protocol is available upon request.We declare no competing interests."}
+{"text": "The present study was conducted to partially fill this gap.European travellers to endemic countries are at risk of malaria and may be affected by a different range of co-morbidities than natives of endemic regions. The safety profile, especially cardiac issues, of artenimol (previously dihydroartemisinin)\u2013piperaquine (APQ) EurartesimParticipants were recruited through Health Care Provider\u2019s safety registry in 15 centres across 6 European countries in the period 2013\u20132016. Adverse events (AE) were collected, with a special focus on cardiovascular safety by including electrocardiogram QT intervals evaluated after correction with either Bazett\u2019s (QTcB) or Fridericia\u2019s (QTcF) methods, at baseline and after treatment. QTcB and/or QTcF prolongation were defined by a value >\u2009450\u00a0ms for males and children and >\u2009470\u00a0ms for females.2 (4.7). Among them, 75 reported a total of 129 AE (27 serious), 46 being suspected to be related to APQ (11 serious) and mostly labelled as due to haematological, gastrointestinal, or infection. Women and Non-African participants had significantly (p\u2009<\u20090.05) more AEs. Among AEs, 21 were due to cardiotoxicity (7.1%), mostly QT prolongation, while 6 were due to neurotoxicity (2.0%), mostly dizziness. Using QTcF correction, QT prolongation was observed in 17/143 participants (11.9%), only 2 of them reporting QTcF\u2009>\u2009500\u00a0ms (milliseconds) but no clinical symptoms. Using QTcB correction increases of >\u200960\u00a0ms were present in 9 participants (6.3%). A trend towards increased prolongation was observed in those over 65\u00a0years of age but only a few subjects were in this group. No new safety signal was reported. The overall efficacy rate was 255/257 (99.2%).Among 294 participants, 30.3% were women, 13.7% of Caucasian origin, 13.5% were current smoker, 13.6% current alcohol consumer and 42.2% declared at least one illness history. The mean (SD) age and body mass index were 39.8\u00a0years old (13.2) and 25.9\u00a0kg/mAPQ appears as an effective and well-tolerated drug for treatment of malaria in patients recruited in European countries. AEs and QT prolongation were in the range of those obtained in larger cohorts from endemic countries.Trial registration This study has been registered in EU Post-Authorization Studies Register as EUPAS6942The online version contains supplementary material available at 10.1186/s12936-021-03750-x. It is an effective artemisinin-based combination therapy which involves the simultaneous use of two blood schizontocidal compounds with independent modes of action, meeting all these needs [Imported malaria into non-endemic regions, particularly Europe and North America is an ongoing problem and most of the cases are in travellers visiting family and relatives . Treatmese needs , 3.The most common side-effects observed with APQ use in uncomplicated malaria patients (1\u201310 patients in 100) are anaemia, headache, corrected QT segment (QTc) prolongation and tachycardia .Preclinical studies with artemisinin-related products in rats and dogs showed electrocardiographic (ECG) effects, in particular prolongation of QTc \u20137. ConvePlasmodium falciparum malaria, including a warning not to take food within 3\u00a0h of ingesting APQ. The drug should not be prescribed to patients with risk factors for QTc prolongation . These include a history of symptomatic cardiac arrhythmias, clinically relevant bradycardia, any predisposing cardiac conditions for arrhythmia, electrolyte disturbances, and recent or on-going treatment known to prolong the QTc interval [Following these studies the European marketing authorization for APQ was obtained for the treatment of uncomplicated interval .While data from its use in malaria-endemic areas supports a very low risk of sudden death after APQ, which is not higher than the population baseline risk, there are few data available on APQ used for the treatment of uncomplicated malaria in returning travellers in Europe .In order to assess the actual safety profile and the potential for cardiotoxicity and QTc prolongation following APQ exposure in these patients, a European post-authorization safety registry was proposed with the goal to monitor all incoming patients with imported malaria. The present work is summarizing the results obtained in a study performed in patients from this registry.This study was an observational, registry-based, longitudinal, multicentric study assessing the safety profile of uncomplicated malaria patients receiving APQ treatment. Participants to the study were selected among patients included in a Health Care Provider (HCP) Safety Registry organized to monitor the outcome and safety of APQ treatment, consisting of 15 clinical centres across 6 European countries  authorized for the use of APQ. Standard APQ treatment was given at the manufacturer recommended dosage of 320\u00a0mg/40\u00a0mg coated-film tablets of either three tablets for participants weighting <\u200975\u00a0kg or four tablets in participants weighted \u2265\u200975\u00a0kg, administered three times at 24\u00a0h interval for 3 consecutive days.P. falciparum malaria that met the summary of product characteristics indications, any gender and age, and signed informed consent and evidence of receiving the APQ treatment on the day of enrolment [Participants were selected based on the following inclusion criteria: clinical and parasitological (microscopy or PCR) diagnosis of uncomplicated nrolment . Each HCSafety data were collected during the normal course of patient care by HCPs preferably during a 3-day hospitalization or, when hospitalization was refused or not possible, on an ambulatory basis. An ECG recording was recommended at baseline after patient enrolment (visit 1 defined as baseline day before or at the time of the first APQ administration) and after the last APQ dose . For the purposes of the analysis, the subpopulation having an ECG on the \u2018strict final treatment day\u2019, defined as the day of last APQ administration, was also individualized and named \u2018Strict QTc population\u2019. A follow-up visit (visit 3) was planned between 3 and 5 weeks after hospital discharge or after the out-patient treatment. If the visit was not feasible, patients were contacted by an independent study monitor for follow-up between day 15 and day 45. Any AE and concomitant medication were recorded.The primary objective was to study the safety profile associated to APQ treatments. The frequency and severity of all adverse events (AE) was recorded, with a particular attention to the main AE of Special Interest (AESI) for AQP treatment, i.e., cardiotoxicity , neurotoxicity , and phototoxicity (dermatitis or rash). Investigators were trained in AE/AESI reporting and had to specify if AEs were suspected to be related to APQ, and report the action taken and the outcome. Differences between pre-treatment baseline values and post-treatment values were measured for blood chemistry markers, such as serum alanine amino-transferase (ALAT), aspartate amino-transferase (ASAT) and creatinine, and for ECG recording parameters, specifically the QTc intervals. Measurements of blood chemistry markers such as serum ALAT, ASAT and creatinine were carried out in at each centre using standard methodologies.Measurements of ECG-based QTc interval (ms) were evaluated after correcting for the heart rate with Bazett\u2019s (QTcB) and Fridericia\u2019s formulae (QTcF) . QTc proThree populations were analysed: (1) the general \u2018safety population\u2019 that include all subjects receiving at least one dose of APQ; (2) the \u2018QTcF/QTcB population\u2019 defined as all patients having at least two ECG, one at baseline-visit 1 and one at the \u2018final treatment day\u2019, and (3) the \u2018Strict QTcF/QTcB population\u2019 with the second ECG at the \u2018strict final treatment day\u2019.Data were reported and tabled as mean\u2009+\u2009SD. Regarding the ECG measurements, differences in value of QTcF and QTcB were compared using ANOVA (p\u2009<\u20090.05).Change in the levels of ALAT, ASAT and creatinine were analysed with a non-parametric test of Wilcoxon\u2019s. Multivariate analysis was conducted with ANOVA including all covariates with a p\u2009<\u20090.20 and using a Backward selection process with a p\u2009<\u20090.05. The variables considered are the characteristics of the patients likely to or that could be associated with a variability of the QT space .A sample size of 300 patients was selected to ensure a large enough sample to capture the association between safety parameters (in particular QTc prolongation) and pre-specified factors, and to have a 90% probability of observing at least one cardiac AESI assuming an overall true incidence of these events of 0.008.The study followed a centralized regulatory submission at the European Medicines Agency . Ethical approval was received also by Hospital Ethics Committee in all the Countries participating in the study. Patients were enrolled after Informed consent signature.From May 2013 to August 2016, a total of 297 patients were included by 15 active centres (among 29 contacted and 23 agreed to participate): two centres in Belgium (n\u2009=\u200959 patients included), three in France (n\u2009=\u200993), two in Germany (n\u2009=\u200912), two in Italy (n\u2009=\u200977), five in Spain (n\u2009=\u200952) and one in the UK (n\u2009=\u20094). Most of active centres were located at university hospitals (80.0%) in infectious and tropical diseases departments (66.6%). Additionally, 43 patients meeting the selection criteria were not included in the study because patient\u2019s decision (34.3%), organizational reasons (20.0%) and medical reasons (17.1%) . A total of 49 patients prematurely discontinued the study , most often after the final treatment day Fig.\u00a0. They weWomen accounted for 30.3% of the participants Table . The majP. falciparum (n-273) by PCR (n\u2009=\u20096), thick smear (n\u2009=\u20094), P. falciparum antigen (n\u2009=\u20091), and positive but with unknown method for others (n\u2009=\u20094). Median P. falciparum counts were 25.0\u2009\u00d7\u2009103/\u00b5L (IQR: 3.5\u201380.0) (n\u2009=\u2009273). Co-infection with other species were identified in 6 patients . The most frequent symptoms of malaria were fever and headache. Haemoglobin was below the lower normal range for 38.2% of participants at baseline . Details of comorbidities can be viewed in  of patients who took concomitant medications at baseline, the most frequent were analgesics (77.2%), drugs for functional gastrointestinal disorders (21.3%), and antibiotics (21.3%). In total, between the month prior and the last day of APQ, 27.6% of participants had taken treatments known to prolong the QTc interval.APQ was initiated on median the day of malaria diagnosis (IQR: 0\u20131) and 97.3% of patients received APQ for 3\u00a0days. APQ dosages were generally consistent with patient weight , mainly 4 tablets per day for patient <\u200975\u00a0kg). The 24 patients but one weighing 100\u00a0kg or more received adequate dosage (12 pills). The APQ administration more than 3\u00a0h from food intake was respected in 182 of the 267 patients (68.2%) in whom information was available.P. falciparum overall efficacy rate was 99.2% with 255/257 having eliminated the parasitaemia at follow-up. Two patients were considered to have developed a recurrence of malaria patients as they were found to have a positive parasitaemia at visit 3.The No woman became pregnant after APQ administration. No substantial changes in hemoglobin and hematocrit were observed. Neutrophil granulocyte, platelet counts, liver parameters and C-reactive protein tended to improve during the course of the study. A total of 129 AEs were experienced by 75/294 patients (25.5%), of which 9 severe AE. In addition, 27 were serious AE were reported in the study   and normal renal function creatinine 12.7\u00a0mg/L. He had taken paracetamol 500\u00a0mg/ascorbic acid 200\u00a0mg/pheniramine maleate 25\u00a0mg prior to presentation. Following the diagnosis of malaria, he received the first dose of 3 tablets 320/40\u00a0mg of APQ adjusted to his weight of 65\u00a0kg (BMI 20.7\u00a0kg/m2). The time of his last meal to dosing was unknown. His fever and nausea were treated with metoclopramide 10\u00a0mg IV and paracetamol 1000\u00a0mg IV on the first day. His QTcF/QTcB were within the normal range at baseline and at his 2nd ECG on day 2, after APQ, the QTcF/QTcB changes were 5 and 9\u00a0ms, respectively. The ECG PR interval and QRS were in normal range at baseline and after APQ. Because of his vomiting, he was switched to atovaquone/proguanil at day 2 and for the subsequent 3\u00a0days (incomplete course of APQ). Five days after the last dose of APQ and 1\u00a0day after last dose of atovaquone/proguanil, he presented with an episode of ventricular tachycardia which reverted spontaneously. He was followed 48\u00a0days without a similar event. The ventricular tachycardia was judged to be related to his cardiomyopathy and not to the APQ treatment.Of the AE of special interest (AESIs), 28 occurred in 27 patients (9.2%). Among those reported, 21 cases (7.1%) were relation to cardiac abnormality . The clinical details on the subject with ventricular tachycardia; a 36-year-old black male with a previous diagnosis of athlete cardiomyopathy and known ECG abnormalities. The subject had never smoked and consumed alcohol less than once a week. This patient, at presentation, described abdominal pain, influenza like symptoms, fever, fatigue, arthralgia, myalgia, vomiting and headache. At baseline Others cardiac AESIs were of mild intensity and of which 19 were considered related to APQ and 3 led to drug discontinuation. In addition, 6 cases (2.0%) of neurotoxicity  were reported of which 4 cases were mild and 2 were of moderate intensity; 4 of them were considered APQ-related. Finally, 1 case (0.3%) of phototoxicity (rash of mild intensity) was raised, recovered after 3\u00a0weeks and was not considered drug related. Due to their rare occurrence, it was not possible to perform a factor analysis associated with AESIs.According to multivariate analysis, non-African patients and females were more likely to experience AEs (including AESIs) than African patients and males (p\u2009<\u20090.05), respectively  and 10 (4.3%) patients, respectively. These figures were 13 (5.5%) and 27 (11.4%) for QTcB. Only one patient  had a QTc value\u2009>\u2009500\u00a0ms at baseline (QTcB\u2009=\u2009557\u00a0ms and QTcF\u2009=\u2009478\u00a0ms), before APQ administration, but was lost to follow up without another ECG.The \u2018QTcF/QTcB population\u2019 was of 143 out of 234 participants, whereas the \u2018strict QTcF/QTcB population\u201d was of 60 out of 143.ECG changes are detailed in Table The mean change in QTcF and QTcB from baseline to the strict final treatment day (n\u2009=\u200960) was +\u200917.5 and +\u20092.6\u00a0ms, respectively. Multivariate assessment of the factors associated to changes in the eligible population is shown in Table Changes in ALAT, ASAT and creatinine from baseline to final treatment day were not significant. Factors associated with these changes are presented in Additional file P. falciparum malaria using a post-registration longitudinal registry in 15 HCP centres of 6 European countries. One in four participants reported a total of 129 AEs. Of the AEs, 46 (11 serious) were suspected by the clinician providing care to be related to APQ, the most common being gastrointestinal disorders. There were 28 AE of special interest , predominantly cardiological (n\u2009=\u200921) (7.1%), therefore it was not possible to analyse the factors associated with the development of neurotoxicity and phototoxicity events.Safety of APQ was studied in patients with diagnosis of uncomplicated acute Focusing on the cardiovascular safety outcome, on QTcF corrections, QT prolongation was observed in 5/143 participants serially assessed (11.9%), with no clinical symptoms. An increase of >\u200960\u00a0ms occurred in 9 participants (6.3%). Almost all of the QTc prolongations were less than 500\u00a0ms and judged as non-serious AE. In the two patients with QT prolongation over 500\u00a0ms following the first dose of APQ, the maximum value was 531\u00a0ms (QTcB). An association was observed between older (age\u2009>\u200965\u00a0years) individuals, and increase in QTcB, but with only four patients >\u200965\u00a0years it is difficult to draw conclusions without additional work. Furthermore, these differences were not observed with QTcF adjustments. One case of spontaneously reverting ventricular tachycardia, developed in a black male with a known cardiomyopathy, 5\u00a0days after his last incomplete course of APQ, and was felt by his HCP, to be unrelated to the APQ treatment.\u00ae development in the DM040011 study  and in the DM040010 study . In these studies, only 7 patients showed QTcF\u2009>\u2009500\u00a0ms (0.4%) [These findings are comparable to those observed during Eurartesims (0.4%) , 14.In a large prospective study  in four In another analysis of Ghanaian data of this effectiveness and safety African platform (INDEPTH-Network), safety analysis of APQ among 4563 patients (16.0% >\u200918\u00a0years old) did not identify any serious safety concerns . IncidenResults in patients from non-endemic European countries are, therefore, consistent with these studies from Africa or Asia, which describe infrequent episodes of reversible QTc prolongation with no clinical impact. Among other observations, changes in ALAT and ASAT were of limited interest given the small change in values between groups. No factors were significantly associated with the changes in creatinine values.Overall, few serious adverse event and no new safety signals were detected. At the multivariate analysis, AEs were significantly more often encountered among women and patients of non-African origin. This is the first time this association was recognized. This is challenging to interpret particularly where the outcome is AEs rather than adverse drug reactions. This could be related to disease manifestations in non-immune populations. Further research in these populations would be required to confirm and better understand these differences.The study showed a cure rate of 99.2%, in line with the expected APQ efficacy , 14.The strength of the study is to be a registry-based, series of patients with imported malaria exposed to APQ in six European countries with a systematic collection of all AEs in a context where concerns arise about the tolerance of anti-malarial treatments in Europe. It provides a better understanding of the use of APQ in people with co-morbidities. The study has also several limitations: it is not a comparative study; the Registry contained a small number of patients; hereby its statistical power is limited; not all patients had an ECG performed on the day of the last APQ intake. The number of lost to follow-up is significant and could have underestimated the frequency of AEs. However, the majority of loss to follow up occurred after the end of treatment and contributed to the analysis. Missing ECGs could produce a bias in the interpretation of ECG findings. Overall, the ultimate sample size achieved was less than the planned, and it is therefore possible that rare side effects may not have been detected due to the final small number of patients. The potassium value was not collected. Given these limitations, some differences are to be interpreted with caution.P. falciparum imported malaria.This is the first study addressing the safety profile of APQ treatment in a consistent number of patients with uncomplicated malaria imported to European countries in a context of routine medical prescribing of APQ. The description of the range of AE\u2019s and the analysis of the QTc interval and other safety data, led to the conclusion that there was no new safety signal or notable changes in their frequency as compared to previously identified signals in endemic populations. Although transient QTc prolongations up to a maximum of 531\u00a0ms was reported in one subject, no clinical consequence was observed. The efficacy rate at over 99%, was also as expected. APQ was found to be a well-tolerated artemisinin-based combination with efficacy and safety at least equivalent to the other artemisinin-based combinations available in the EU. It is also characterized by a simple administration modality, once a day for 3\u00a0days, which favors compliance to treatment and is a valuable option for use in the first-line treatment of uncomplicated Additional file 1. Illnesses and symptoms at Visit 1 by System Organ Class and Preferred Term\u2014safety registry of malaria patients treated with artenimol\u2013piperaquine.Additional file 2. Baseline biological results\u2014safety registry of malaria patients treated with artenimol\u2013piperaquine.Additional file 3. Overview of adverse events (AE) and serious adverse event (SAE) other than adverse event of special interest (AESI).Additional file 4. Factors associated with the occurrence of adverse event (AE) including adverse event of special interest (AESI).Additional file 5. Factors associated with the change in Alanine amino-transferase (ALAT), Aspartate amino-transferase (ASAT) and creatinin from baseline to the final treatment day."}
+{"text": "When people with human immunodeficiency virus (HIV) infection (PWH) develop malaria, they are at risk of poor anti-malarial treatment efficacy resulting from impairment in the immune response and/or drug-drug interactions that alter anti-malarial metabolism. The therapeutic efficacy of artemether-lumefantrine was evaluated in a cohort of PWH on antiretroviral therapy (ART) and included measurement of day 7 lumefantrine levels in a subset to evaluate for associations between lumefantrine exposure and treatment response.3 were randomized to daily trimethoprim-sulfamethoxazole (TS), weekly chloroquine (CQ) or no prophylaxis. After diagnosis of uncomplicated Plasmodium falciparum malaria, a therapeutic efficacy monitoring was conducted with PCR-correction according to WHO guidelines. The plasma lumefantrine levels on day 7 in 100 episodes of uncomplicated malaria was measured. A frailty proportional hazards model with random effects models to account for clustering examined the relationship between participant characteristics and malaria treatment failure within 28\u00a0days. Pearson\u2019s Chi\u2014squared test was used to compare lumefantrine concentrations among patients with treatment failure and adequate clinical and parasitological response (ACPR).Adults living with HIV (\u2265\u200918\u00a0years), on ART for\u2009\u2265\u20096\u00a0months with undetectable HIV RNA viral load and CD4 count\u2009\u2265\u2009250/mm411 malaria episodes were observed among 186 participants over 5\u00a0years. The unadjusted ACPR rate was 81% (95% CI 77\u201386). However, after PCR correction to exclude new infections, ACPR rate was 94% (95% CI 92\u201397). Increasing age and living in Ndirande were associated with decreased hazard of treatment failure. In this population of adults with HIV on ART, 54% (51/94) had levels below a previously defined optimal day 7 lumefantrine level of 200\u00a0ng/ml. This occurred more commonly among participants who were receiving an efavirenz-based ART compared to other ART regimens (OR 5.09 [95% CI 1.52\u20137.9]). Participants who experienced treatment failure had lower day 7 median lumefantrine levels (91\u00a0ng/ml [95% CI 48\u2013231]) than participants who experienced ACPR .Recurrent malaria infections are frequent in this population of PWH on ART. The PCR-adjusted efficacy of AL meets the WHO criteria for acceptable treatment efficacy. Nevertheless, lumefantrine levels tend to be low in this population, particularly in those on efavirenz-based regimens, with lower concentrations associated with more frequent malaria infections following treatment. These results highlight the importance of understanding drug-drug interactions when diseases commonly co-occur. HIV and malaria infections are endemic in sub-Saharan Africa, and previous research demonstrated high rates of clinical malaria illness among adults living with HIV infection after stopping trimethoprim-sulfamethoxazole prophylaxis . Thus, eNon-nucleoside reverse transcriptase inhibitors have been the most common antiretrovirals used in sub-Saharan Africa since the roll-out of antiretroviral therapy (ART)on the continent. These drugs are extensively metabolized by cytochrome p450 (CYP450) enzymes and thus interact with compounds that use similar pathways. In particular, efavirenz induces CYP 3A4 and CYP 2B6, which metabolize lumefantrine into desbutyl-lumefantrine, leading to reduced plasma levels of lumefantrine and increased risk of malaria treatment failure .While coHigh rates of malaria illness in PWH who discontinued trimethoprim-sulfamethoxazole (TS) or chloroquine (CQ) prophylaxis were reported from this randomized clinical trial, evaluating the role of anti-malarial prophylaxis among PWH on antiretroviral therapy in Malawi . ParticiThe study design and methodology for the parent clinical trial have been published previously . In summ3, and (3) CD4 count of at least 250/mm3. Additional inclusion/exclusion criteria are published with the protocol [Study participants were recruited from two sites in Malawi: Ndirande research clinic in Blantyre, an urban centre, and Tisungane ART clinic at Zomba Central Hospital, located in a more rural setting. Of note, the two sites have different malaria burdens: malaria parasite prevalence in children under 5\u00a0years old is 4% in Ndirande versus 28% in Zomba . Potentiprotocol .Plasmodium species using thin blood smears. Only infections with P. falciparum are included here.Participants who presented with malaria symptoms were evaluated at the study clinic and had blood collected for microscopy. Uncomplicated malaria was defined as objective fever (temperature\u2009\u2265\u200937.5\u00a0\u00b0C) or history of fever within the past 48\u00a0h and/or other symptoms such as nausea, vomiting and diarrhoea, headache, back pain, chills, or myalgia, plus detection of any malaria parasites in blood by microscopy. Participants with danger signs such as reduction or loss of consciousness and difficulty breathing were referred to the hospital and not enrolled in the therapeutic efficacy study. Two trained microscopists identified and quantified malaria parasitaemia from thick blood smears and determined P. falciparum malaria were treated with 80\u00a0mg artemether and 480\u00a0mg lumefantrine twice daily for 3\u00a0days. First doses were directly observed. All participants were followed up on days 1, 2, 3, 7, 14, 21 and 28 according to WHO standard for monitoring therapeutic efficacy, and blood smears and dried blood spots were collected at each visit.Participants diagnosed with uncomplicated In cases of recurrent malaria infection occurring on or after day 14, dried blood spots collected on filter paper from enrolment day and the day of recurrence of infection underwent extraction to distinguish new from recrudescent infection by genotyping merozoite surface protein-1 (MSP-1), MSP-2 and the glutamate-rich protein (GLURP) according to the publicly available protocol .Participants who consented to participate in a sub-study of lumefantrine drug concentration measurements were selected to submit blood specimens for day 7 drug levels. Participants were only enrolled 1\u00a0time. These participants were given 250\u00a0ml of milk to drink with each dose to ensure optimal and consistent lumefantrine absorption . Doses 1Plasma was shipped to the Parikh laboratory at the Yale School of Public Health  on dry ice, and immediately transferred to NorthEast BioLab  for drug level analysis of lumefantrine and desbutyl-lumefantrine using liquid chromatography-tandem mass spectrometry on an API 5000 triple-quadruple mass spectrometer . The calibration range was 10.9\u20133785\u00a0ng/mL for lumefantrine, and 1.9\u20131130\u00a0ng/mL for desbutyl-lumefantrine with the lower limit of quantification (LLOQ) at 10.0 and 1.9\u00a0ng/mL for lumefantrine and desbutyl-lumefantrine, respectively. The coefficient of variation was 1.4% and 5.6% for lumefantrine and desbutyl-lumefantrine, respectively.Treatment failure was diagnosed according to WHO criteria which classify outcomes in mutually exclusive groups [e groups .Early treatment failure was defined as any of the following: danger signs or severe malaria on day 1, 2 or 3 in the presence of parasitaemia; parasitaemia on day 2 higher than on day 0, irrespective of axillary temperature; parasitaemia on day 3 with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C; parasitaemia on day 3\u2009\u2265\u200925% of count on day 0.Late clinical failure was defined as any of the following: danger signs, severe malaria, or axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C in the presence of parasitaemia on any day between day 4 and day 28 in patients who did not previously meet early treatment failure criteria.Late parasitological failure was defined as any of the following: presence of parasitaemia on any day between day 7 and day 28 with axillary temperature\u2009<\u200937.5\u00a0\u00b0C in patients who did not previously meet early treatment failure or late clinical failure criteria.Adequate clinical and parasitological response (ACPR) was defined as: absence of parasitaemia on days 28, irrespective of axillary temperature, in patients who did not previously meet early treatment failure, late clinical failure or late parasitological failure criteria.After analysis of genotyping results, an infection was recrudescent if any genotype identified in the recurrent infection was also present at the initial infection. In PCR-adjusted results, only recrudescent infections were considered treatment failures.Kaplan Meier survival analysis was used to determine the cumulative success rate defined as not reaching a failure point during the time under observation. Individuals diagnosed with malaria who did not have a 28\u00a0day follow-up visit and did not meet treatment failure definitions were censored at their last visit. was estimated to assess for possible predictors of treatment failure, including prophylactic regimen, age, sex, ART regimen, presentation with fever, parasite density at presentation, and study site. A random effect for individual was included to account for multiple observations among the same individuals. For the analysis of prophylactic regimen, the CQ and TS treatment arms were combined due to few malaria episodes among individuals on prophylaxis.To examine the association between lumefantrine and desbutyl-lumefantrine concentrations and treatment failure, a Wilcoxon Rank Sum test was used to compare the distribution of lumefantrine concentration between patients with treatment failure and those with ACPR. Because the majority of desbutyl-lumefantrine levels were below the level of detection, these levels were classified as detectable or undetectable. Not all participants with concentration data had complete follow-up data. Due to the limited number of data points, participants who were followed up to at least 14\u00a0days without treatment failure were classified as treatment successes, participants who were not followed at either 14, 21, or 28\u00a0days were excluded. As a day 7 lumefantrine concentration of 200\u00a0ng/ml has been associated with higher likelihood of treatment success, the analysis examined whether this cut-off point was associated with treatment failure in this cohort using Pearson\u2019s Chi\u2014squared test. All statistical analysis was conducted in SAS version 9.4 .The study was approved by the Kamuzu University of Health Sciences Research Ethics Committee and the University of Maryland Baltimore Institutional Review Board. All participants provided written informed consent.From December 2012 to July 2018, clinical outcomes for 411 clinical malaria episodes among 186 participants . After PCR correction, the cumulative efficacy was 94% . The hazard of failure was higher among participants who received CQ or TS prophylaxis compared to those who did not , men compared to women, and participants who received efavirenz vs other ART regimens, though none of these differences achieved statistical significance . A statistically significant difference was observed between participants who experienced treatment failure vs. ACPR (91\u00a0ng/ml (IQR 48\u2013231) vs 190\u00a0ng/ml (IQR 101\u2013378), p-value\u2009<\u20090.008) (Table 59. A staDrug levels were low in this population with 54.3% of individuals having a day 7 lumefantrine concentrations less than 200\u00a0ng/ml. Among those with treatment failure, 65.0% had an undetectable desbutyl-lumefantrine level (Table In this evaluation of AL therapeutic efficacy among PWH who are well controlled on ART, unadjusted overall treatment efficacy was lower than expected at 81% but after PCR-correction to exclude new infections, the adjusted treatment efficacy rate was 94%. This meets the WHO requirement for first-line anti-malarial agents to have at least 90% efficacy in therapeutic efficacy studies. This also confirms that the observed high rate of recurrent infection is not likely due to drug resistance. Although these findings confirm that resistance to AL is not clinically detectable in Malawi, high rates of new infections may be due to low lumefantrine levels that fail to protect against subsequent infections.Although the 94% efficacy is an acceptable rate, the results are lower than 99% treatment efficacy documented in a population of children around the same time in Malawi . This maThe day seven 7 plasma lumefantrine levels were low in our population of adults living with HIV Recurrent parasitaemia was associated with lower lumefantrine levels. The low drug concentrations and recurrent parasitaemia were both more common in participants on efavirenz-based ART regimen than those taking other regimens, consistent with results of a previous meta-analysis [This study also supports the induction effect of efavirenz on lumefantrine metabolism through quantification of its primary metabolite, desbutyl-lumefantrine . EfavireAnopheles mosquitoes which increases the risk of treatment failure due to reinfection, though that was excluded by PCR correction. In addition, with higher density infections and more polyclonal infections, the PCR assay has more sensitivity to detect a low frequency recurrent clone. This explanation is supported by models that suggest false positive identification of recrudescent infections occur in higher transmission settings [A higher rate of treatment failure was observed among participants living in Zomba, a more rural area, compared to Blantyre, an urban centre. Malaria transmission intensity is higher in Zomba compared with Blantyre. Therefore, participants in Zomba were exposed to more infectious bites of settings .Although the study did not identify a concerning level of anti-malarial resistance as evidenced by the 94% efficacy rate, low drug levels have the potential to impact the emergence and spread of resistance . Sub-theThe PCR-corrected therapeutic efficacy of AL among PWH in this setting was within the required range, but patients on an efavirenz-based regimen experienced lower lumefantrine levels than patients on other ART highlighting the importance of drug-drug interactions. Drug-drug interactions for common infections can alter pharmacokinetics and pharmacodynamics and have significant impact on individual and public health outcomes. Although efavirenz is largely being replaced by dolutegravir in sub-Saharan Africa, it will remain in use in a smaller population. In addition, this report highlights the need to consider common infections and associated pharmacokinetic interactions in the wider drug development and drug policy decisions."}
+{"text": "The prevalence of superficial fungal infections in India is believed to have increased substantially in the past decade. We evaluated the treatment outcomes and risk factors associated with clinical response to a treatment course of itraconazole for the management of dermatomycosis in India.T. cruris or T. corporis, received itraconazole 200 mg/day (any formulation) orally for 7 days, and were followed for an additional 7 days.In this real-world, prospective pilot study (August 2019 to March 2020), adult participants (18\u201360 years), diagnosed with T. mentagrophytes species complex (91.7%) and T. rubrum (8.3%), was within the susceptibility range (0.015\u20130.25 mcg/mL). At day 14, 8/13 (61.5%) participants achieved a mycological response, 2/13 participants (15.4%) had a mycological failure and 90% showed a clinical response.The study was terminated early due to the COVID-19 pandemic. Of 40 enrolled participants  completed the study. The median (range) Clinical Evaluation Tool Signs and Symptoms total score at baseline was 5.5 (2\u201310). Clinical response of \u201chealed\u201d or \u201cmarkedly improved\u201d based on the Investigator Global Evaluation Tool at day 7 (primary objective) was 42.9% . Itraconazole minimum inhibitory concentration for identified microorganisms, COVID-19 pandemic affected patient recruitment and follow-up, so the findings call for a careful interpretation. Nevertheless, this real-world study reconfirmed the clinical efficacy and microbial susceptibility to itraconazole for the fungi causing dermatophytosis in India.NCT03923010.Trial registration number: Clinicaltrials.gov  Superficial cutaneous fungal infections or dermatomycosis/dermatophytosis are one of the most common dermatoses affecting around 20\u201325% of the world population . India hTrichophyton mentagrophytes pandemic in recent years [The clinical presentation of dermatomycosis varies from mild scaling and redness to severe inflammation \u201313. Althnt years , 22. Casnt years , 20, 23.Itraconazole, a triazole antifungal agent, has a broad spectrum of activity against infections caused by dermatophytes, yeasts, and many other fungi \u201327. ItraThe objective of this study was to evaluate real-world treatment outcomes and the risk factors associated with a poor clinical response with the use of a of itraconazole in patients with dermatomycosis.This real-world, prospective, non-randomized, multicenter, interventional, longitudinal study was conducted in India and consisted of an open-label treatment phase of 7 days followed by a follow-up phase of 7 days.Tinea cruris (T. cruris) or Tinea corporis (T. corporis) who demonstrated clinical response after 7 days of treatment. The secondary objectives were to estimate the proportion of participants with the mycological response after 14 days of follow-up and the association of clinical response with plasma drug concentrations (itraconazole and hydroxy-itraconazole) and baseline sensitivity pattern of causative fungi. Other exploratory objectives included the estimation of the proportion of participants with the clinical response after 14 days of follow-up and the extent to which clinical improvement at Day 7 predicts clinical improvement at Day 14.The primary objective of the study was to estimate the proportion of participants that received itraconazole for The study is registered with clinicaltrials.gov (NCT03923010). The study protocol, informed consent forms, and applicable study documents were approved by an independent ethics committee or institutional review board at each study site. They include Institutional Ethics Committee, NKP Salve Institute of Medical Science & Lata Mangeshkar Hospital, Nagpur, India; Institutional Ethics Committee, Postgraduate Institute of Medical Education and Research, Chandigarh, India; Institutional Ethics Committee, Sri Ramachandra Hospital, Chennai, India and Yenepoya Ethics Committee-1, University Road, Deralakatte, Mangalore, India. Signed informed consent was obtained from all participants at the start of the study. This study was conducted in accordance with the Declaration of Helsinki and is consistent with Good Clinical Practices and applicable regulatory requirements. The planned recruitment time for this study was 4 months. However, the period was extended to 10 months due to the Coronavirus Disease-2019 (COVID-19) pandemic. The study was eventually discontinued prematurely for the same reason.T. cruris or T. corporis infection, with or without a history of antifungal treatment were enrolled if they were prescribed oral itraconazole 200 mg/day for the management of their dermatophytosis. The participants were recruited from clinical practices of dermatologists from four sites  in India. Exclusion criteria included the history of ventricular dysfunction, liver or renal insufficiency; contraindications for the use of itraconazole; known achlorhydria or treatment for gastric acidity; the presence of other dermatoses e.g., psoriasis, seborrheic, or atopic dermatitis or infection with an organism with known or established resistance to itraconazole; co-existing fungal infection of other body areas, known allergies or receipt of CYP3A4 substrates or oral itraconazole within 14 days or systemic antifungal or corticosteroid therapy within 30 days or a topical antifungal or corticosteroid therapy within 14 days before screening.Adult participants aged \u226518 to \u226460 years with a clinical diagnosis of Participants were prescribed locally available itraconazole (either generic or reference). The dosage was a 200 mg capsule or 2 capsules of 100 mg each, once daily. Participants were instructed to take their prescribed dose of itraconazole orally each day after a full meal at home. The recommended duration of treatment was 7 days. The total duration was decided by the treating physician as part of the participant\u2019s clinical care, according to local standards of practice, and could be extended up to Day 14.Participants\u2019 demographic information was collected at baseline. The clinical assessment of the disease was evaluated by the treating physicians using two tools. The Clinical Evaluation Tool Signs and Symptoms  was used in the development program of itraconazole. It evaluates the presence and severity  of each of 6 signs and symptoms . The Investigator Global Evaluation (IGE) tool was used to classify the clinical response based on the percentage of clinical improvement from baseline. The CET SS total scores at Day 7 and Day 14 were compared with baseline scores to assess the percentage of clinical improvement, defined as a decrease in CET SS total score to 1 or 2, which was further used to classify the clinical efficacy using the Investigator Global Evaluation (IGE) tool, with scores 1 (absence of signs and symptoms) or 2 . Clinical response was thus defined as having scores 1 or 2  on the IGE tool of clinical improvement while the mycological response was defined as culture negative at Day 14 from participants positive at baseline.Pharmacokinetic (PK) assessments were conducted in participants who had at least 1 set of blood samples drawn at Day 7. Measurement of itraconazole and hydroxyitraconazole concentrations in plasma was performed using high performance liquid chromatography with tandem mass spectrometry. Blood samples for PK analysis were collected predose (at 24 hours (\u00b12 hours) after the previous dose on Day 6) and Day 13 and postdose (at 2 hours and 4.5 hours) after the Day 7 and Day 14 doses. Plasma concentrations of itraconazole and hydroxy-itraconazole were assessed against clinical outcomes.Skin scrapings were collected for culture and sensitivity analysis and the baseline resistance to itraconazole and its association with clinical outcomes was evaluated. Skin specimens were collected from the edge of lesions using sterile blunt scalpel. The KOH samples were analyzed locally at each study center and for mycological culture analysis samples were shipped to central laboratory . The fungal pathogens were identified by amplification and Sanger-based (dideoxy-termination) sequencing as per the Clinical and Laboratory Standards Institute (CLSI) MM18-A guidelines. A microdilution method, according to the Clinical and Laboratory Standards Institute (CLSI) M27 guidelines, was used to conduct antifungal sensitivity testing . The minSafety was assessed by monitoring adverse events (AEs) and laboratory parameters.As this was a pilot study, no formal hypothesis was tested. The sample size was not calculated based on power but rather selected to collect sufficient data to provide a first approximation of the distribution of efficacy and PK results across the large number of formulations that were planned to be studied. The target sample size was 50. All analyses were performed on participants receiving any generic itraconazole and not segregated out by the brand.Continuous variables were summarized using the number of observations, mean, standard deviation (SD), coefficient of variation (CV), median, and range as appropriate while the categorical values were described using the number of observations and percentages as appropriate. The point estimate and the corresponding 95% confidence intervals (CIs) for the efficacy endpoints were provided. One-Proportion Z-test was used to estimate the 95% CI for the percentage of participants who had a clinical response. The efficacy analyses were performed on the treated population that included participants who had the 7-day treatment regimen, had clinical evaluations at baseline and at Day 7, and completed the study. Missed data at Day 14 follow-up were imputed as \u201cnon-responders\u201d. For PK analysis, when an individual concentrate value was below the lower limit of quantification (LLOQ), the concentrate value was assigned a value of 0 for the analysis and presented in tables as below the quantification limit (BQL). In addition, when greater than 50% of the individual concentration values for a given time point were below the LLOQ, the mean, minimum, and median values were reported as BQL, and SD, %CV, and geometric mean were not reported for these analyses. For graphical analysis, plasma concentration values below the LLOQ were displayed as 0 for the linear plots. Logistic regression was used to determine the association of clinical response at Day 7 (dependent variable) with the baseline MIC sensitivity pattern (independent regressor) of causative fungi. SAS version 9.4 was used for calculations.The study was conducted from August 2019 to March 2020. Due to COVID-19 related lock-down, the study was prematurely terminated. Forty participants were enrolled of whom 37 received at least one dose of itraconazole, 12 (32%) received 7 days and 25 (67%) received 14 days of itraconazole. A total of 20 participants completed the study . The meaA total of 28 (75.6%) participants completed the 7-day treatment phase, and only 20 (50%) returned for the Day 14 visit and completed the study as per protocol. The most common reasons for discontinuation were COVID-19-related , personal (n = 6), and patient withdrawal (n = 4). Some participants did not return for either day 7 or day 14 visits. Thus, the number of participants available for clinical response evaluation is less than that on treatment.From 28 participants who completed 7 days of treatment and were evaluable for efficacy, 12  had a clinical response as \u201chealed\u201d or \u201cmarkedly improved\u201d based on the IGE Tool assessment.Twenty participants completed 14 days of follow-up and were evaluable for clinical response. Eighteen achieved a score of 1 (absence of signs and symptoms) or 2 , for clinical response of 90% . When all 8 participants that missed the Day 14 follow-up were imputed as \u201cnon-responders\u201d, the estimated proportion of the participants with clinical response was 64.3% .T. mentagrophytes species was the most common microorganism isolated (91.7%). The culture results were presented in A total of 28 fungal cultures were obtained at baseline, 24 (85%) were positive. T. mentagrophytes species complex isolated at baseline and the Day 14 visit . Two participants (15.4%) had mycological failure  itraconazole or reference itraconazole, concentrations of itraconazole and hydroxy-itraconazole were quantifiable in plasma prior to dosing on Day 7 in 23 of the 26 available samples with an LLOQ of 20 ng/mL for both the analytes. Of the 3 participants who had plasma concentrations BQL prior to dosing on Day 7, one participant also had no quantifiable plasma concentrations of itraconazole and hydroxy-itraconazole at 2 hours and 4.5 hours postdose. At Day 7, the mean plasma concentrations for itraconazole and hydroxy-itraconazole in non-responders  was comparable to that of responders , the lowest itraconazole drug concentration on Day 7 was greater than the MIC by 1.2 to 19.4 times the MIC at baseline.Clinical response at Day 7 was not associated with the highest or lowest plasma concentrations of itraconazole or hydroxy-itraconazole. Of the 3 participants with predose plasma concentrations BQL on Day 7, two participants were classified as responders  and 1 participant was classified as a nonresponder . All 3 participants had positive fungal cultures for T. cruris infection (moderate intensity) was recorded, leading to the withdrawal of the participant from the study, although, the investigator considered it to be unrelated to the study treatment. No serious AEs or deaths were reported during the study and there were no clinically significant changes in vital signs or laboratory parameters.The safety set included 37 participants who received at least 1 dose of itraconazole. There were no safety concerns with itraconazole. Only one treatment-emergent AE (TEAE) of worsening of This pilot study was planned to determine the clinical response after a treatment course of 7 days of itraconazole in patients suffering from superficial fungal infection in India. However, the study was prematurely terminated due to the COVID-19 pandemic, thus calling for a careful interpretation of the results. Nevertheless, the results provide interesting insights regarding the treatment outcomes and risk factors associated with clinical response in this patient population while they received itraconazole as part of their routine clinical care.Of the 40 participants enrolled in our study, approximately two-thirds were male, and the mean age was 35.5 years. This is consistent with prior reports showing male preponderance  and relatively high incidence in the age group of 20\u201340 years in India [Trichophyton species has emerged as a predominant organism with T. rubrum, T. mentagrophytes and T. verrucosum are the most commonly identified [T. mentagrophytes was the most common species (~92%) identified followed by T. rubrum (~8%). The MIC for itraconazole was within the susceptibility range (0.015 to 0.25 mcg/mL) in all isolates both at baseline and after treatment. At Day 7 more than 40% of the participants achieved clinical response . There was approximately a 2-fold improvement in the clinical response at Day 14 compared with Day 7 based on the completer analysis. This estimated ratio became 1.5 when participants who missed the Day 14 follow-up were categorized as \u201cnon-responders\u201d. The observed results are similar to those of the pivotal studies of itraconazole where >80% superficial dermatophytosis healed with the use of 100 mg oral itraconazole for 7 days [A dramatic change in the characteristics of dermatophytosis has been reported in India in recent years and entified . The culr 7 days .The PK properties of itraconazole should also be evaluated while considering the dose and duration of treatment . ItraconT. mentagrophytes species complex. This might be due to the fact that itraconazole accumulates at the infection site, making consistently high plasma concentrations unnecessary for clinical response [Interestingly, in our study, there was a lack of correlation between PK results and the clinical outcomes observed, along with an absence of itraconazole resistance in the causative organisms. The clinical response at Day 7 was neither associated with the highest or lowest observed plasma concentrations of itraconazole and hydroxy-itraconazole, nor with the MIC of the causative fungi. Thus, the treatment failure could not be attributed to either a lack of drug exposure or elevated MIC values for response . Owing tresponse \u201338.As per the recommendations of the Expert Consensus on the Management of Dermatophytosis in India (ECTODERM India), itraconazole is the preferred systemic antifungal therapy. However, in view of the perceived changes in both prevalence and clinical characteristics of dermatomycosis in India , the ECTODERM expert panel recommended itraconazole doses are 100\u2013200 mg once a day for 2\u20134 weeks in treating na\u00efve cases, and 200\u2013400 mg once a day prolonged periods (>4 weeks) in case of recalcitrant cases [Anecdotal reports of increased treatment failures with the use of itraconazole in India should be interpreted with caution. Although this is a small study, our findings suggest that resistance to itraconazole or PK factors may not be the cause of those reported findings. There are however other factors that cannot be excluded. One of them is the severity of the disease\u2014based on our clinical evaluation the cases included in this study were mild. It is possible that in more severe cases the response to itraconazole would be different. Another factor is adherence to treatment. It is well known that patients included in clinical trials have better adherence to treatment. Although this study tried to assess these patients in their regular practice, we included evaluations that were not part of their regular clinical care and that could have influenced their adherence to therapy and therefore the observed clinical outcomes. There have been other efforts to understand the epidemic of dermatophytosis in India with similar results , 39, 40.The conduct of the study was difficult from many perspectives; the COVID-19 pandemic affected patient recruitment and follow-up and consequently, the study understandably suffered from a significant drop-out rate. Though the small sample size and high discontinuation rate are important limitations, the study reconfirmed the clinical efficacy and microbial susceptibility to itraconazole for the fungi causing dermatophytosis in India.S1 Checklist(DOCX)Click here for additional data file.S1 File(PDF)Click here for additional data file.S2 File(PDF)Click here for additional data file.S3 File(PDF)Click here for additional data file.S4 File(DOCX)Click here for additional data file."}
+{"text": "Chemoprophylactics are a vital tool in the fight against malaria. They can be used to protect populations at risk, such as children younger than the age of 5 in areas of seasonal malaria transmission or pregnant women. Currently approved chemoprophylactics all present challenges. There are either concerns about unacceptable adverse effects such as neuropsychiatric sequalae (mefloquine), risks of hemolysis in patients with G6PD deficiency (8\u2010aminoquinolines such as tafenoquine), or cost and daily dosing (atovaquone\u2013proguanil). Therefore, there is a need to develop new chemoprophylactic agents to provide more affordable therapies with better compliance through improving properties such as pharmacokinetics to allow weekly, preferably monthly, dosing. Here we present a pharmacokinetic\u2013pharmacodynamic (PKPD) model constructed using DSM265 . The PKPD model mimics the parasite lifecycle by describing parasite dynamics and drug activity during the liver and blood stages. A major challenge is the estimation of model parameters, as only blood\u2010stage parasites can be observed once they have reached a threshold. By combining qualitative and quantitative knowledge about the parasite from various sources, it has been shown that it is possible to infer information about liver\u2010stage growth and its initial infection level. Furthermore, by integrating clinical data, the killing effect of the drug on liver\u2010 and blood\u2010stage parasites can be included in the PKPD model, and a clinical outcome can be predicted. Despite multiple challenges, the presented model has the potential to help translation from preclinical to late development for new chemoprophylactic candidates. WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC?Plasmodium falciparum growth.WHAT QUESTION DID THIS STUDY ADDRESS?The administration of chemoprophylactic medicines to healthy subjects to prevent them from developing malaria is a key strategy in protecting populations at high risk. However, current antimalarial drugs aimed at protecting people through chemoprophylaxis often come with concerning adverse effects, are expensive, or require courses of daily dosing. There is thus a need to discover and develop new chemoprophylactic drug candidates to provide more affordable therapies with better adherence. To effectively discover and develop chemoprophylactic medicines requires knowledge of how they kill or otherwise inhibit parasites in both the liver and blood stages of the lifecycle. Although data are available from induced blood\u2010stage malaria volunteer infection clinical studies and, separately, sporozoite\u2010infected human challenge studies, there is no known published work on the integration of the two to create a model for both the liver and blood stages of WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE?Is it possible to make inferences about parasite levels and activity in the liver stage of the malaria infection process, which cannot be measured directly, by mathematically modeling the pharmacokinetic and pharmacodynamic relationships between chemoprophylactic drugs, such as DSM265 and blood\u2010stage parasites, thereby increasing our ability to predict clinical outcomes and thus improving drug development?HOW MIGHT THIS CHANGE DRUG DISCOVERY, DEVELOPMENT, AND/OR THERAPEUTICS?The study shows that by combining qualitative and quantitative knowledge of parasite dynamics from two different study designs, induced blood\u2010stage malaria and sporozoite\u2010infected human challenge studies, to create a mathematically realistic pharmacokinetic\u2013pharmacodynamic model of malaria parasite and chemoprophylactic drug interaction, it was possible to infer information about liver\u2010stage parasite levels and activity. By integrating the two designs of study, it was also shown that it is possible for the model to predict clinical outcomes. By assuming similar characteristics between the liver\u2010 and blood\u2010stage activities, it was possible to predict the level of chemoprotection afforded by DSM265.One of the biggest challenges facing malaria treatment and prevention is a problem of knowledge concerning liver activity during the malaria parasite infection process. However, the ability to infer such information by way of mathematical modeling is a means of changing this. If blood\u2010stage activity is known, then pharmacokinetic\u2013pharmacodynamic models can be used to assess the level of chemoprotection delivered by compounds with similar liver\u2010 and blood\u2010stage activities in in vitro systems. This could be particularly useful in candidate selection for phase III clinical trials if such data were combined with knowledge from epidemiology and \u201cadherence behavior,\u201d augmenting the transition from preclinical to the latter stages of development for new and improved antimalarials.Despite an unprecedented period of success throughout the 21st century, malaria remains a significant global health challenge that in 2019 killed 409,000 people worldwide, of which 67% were children younger than the age of 5.Administration of chemoprophylactics to healthy subjects to prevent malaria development is a key tool in eradication. Some 25 million children aged younger than 5 are protected each season using a monthly treatment dose of sulfadoxine\u2013pyrimethamine plus amodiaquine costing around $1 per child\u2010year for the drug and $5 in program costs.Plasmodium falciparum) with rupturing of erythrocytes and reinfection. It is the periodic rupture at this stage that causes malaria clinical symptoms. To prevent malaria infection, molecules can either have causal prophylaxis, stopping the development of parasites in the liver , or suppressive prophylaxis, inhibiting the development of parasitemia .Development of chemoprophylactic molecules requires understanding of activity on all parasite life stages. After a mosquito bite, sporozoites travel to the liver and infect hepatocytes, where they multiply and mature into schizonts for 5\u20137\u2009daysP. falciparumTo prioritize new medicines, it is important to have a validated model of infection in humans that combines these two aspects, whereby predictions can be made regarding parasitemia supporting the development of suppressive prophylaxis, but also predictions on the prophylactic efficacy based on inferred liver activity. To help build such a model, data from DSM265, a new compound that selectively inhibits parasite dihydroorotate dehydrogenase (DHODH), was used. It shows in vitro activity against the liver and blood stages of P. falciparum\u2013infected erythrocytes were administered intravenously to healthy subjects. Once parasitemia reached a predefined threshold but was still below the clinical symptom threshold, DSM265 was administered. Parasitemia was monitored daily from infection to 1 month after dosing.In IBSM studies, P. falciparum, where parasites first invade the liver and later appear in blood. The drug to be tested was administered prior to infection, usually at one\u2010dose level with different time intervals between administration and infection, or different dose levels with a fixed time interval. Parasite levels in the liver cannot be directly measured, hence only parasite levels in blood were monitored. Parasitemia was initially below a limit of quantification (BLOQ) and became measurable if the administered drug dose was insufficient to kill either liver\u2010 or blood\u2010stage parasites. Using these studies, the presented PKPD model was developed to semimechanistically describe parasite dynamics in two compartments, the liver and blood, where transfer occurs only at maturation of merozoites at Day 6. Analogous to analyzing oral and i.v. PK by use of an absorption compartment with a lag time, the model deconvolutes drug activity for the unobserved liver stage to better capture the ability of DSM265 to provide protection.In spz HuCh, sporozoites were either injected intravenously or administered via mosquito bites.Data for this PKPD analysis were taken from existing publications. All participants gave written informed consent before being included in studies; all were approved by their institutional review boards.P. falciparum\u2013infected erythrocytes and treated with DSM265 7\u2009days after inoculation; PK and parasitemia were monitored in each subject. Parasitemia was measured prior to drug administration, providing information on the natural net growth of parasite in the blood stage. The phase IIa study was conducted in Peru with uncomplicated malaria patients (blood stage) who were treated with DSM265 at the time of diagnosis; both DSM265 concentrations and parasitemia were monitored.DSM265 dried blood spot (DBS) concentrations and parasitemia data were pooled from 15 different studies. These consisted of one first\u2010in\u2010human (FIH) study, two IBSM studies, one phase IIa study, two spz HuCh studies, and nine published studies to obtain control data from nontreated subjects. The FIH study was a PK single\u2010ascending\u2010dose study where DSM265 was administered to healthy volunteers; each subject provided a rich PK sampling profile.DSM265 has been described by McCarthy et al.A visualization of the chemoprophylaxis PKPD model is presented in Figure\u00a0PL, and parasites per milliliter in blood, PB, as the net balance of an exponential growth  and killing  due to an antimalarial drug. The release of merozoites from the liver to blood is described by the term kLB\u00b7TrLB\u00b7PL, where kLB is the rate of merozoites invading erythrocytes once they leave hepatocytes, and TrLB the cumulative fraction of hepatocytes that burst over time. PB being expressed in parasites per milliliter, the number of transferred merozoites is divided by VB, the total volume of blood assumed to be the standard human value of 5\u2009L for all individuals. PL was initialized at time 0 by Inoculum (the number of injected sporozoites) times Finc, the fraction of inoculated sporozoites that invade hepatocytes; many sporozoites are nonviable or do not reach the liver.The PKPD model describes the dynamic of parasite number in the liver, KillL and KillB were described by a turnover model as DSM265, a DHODH inhibitor, leads through a cascade to inhibit DNA and RNA syntheses.CDBS, Emax,X, EC50,X, hX, and kin,X are DSM265 DBS concentration, maximum killing rate, DBS concentration at which half of maximum effect is attained, the Hill coefficient, and the turnover rate, respectively.kLB was set to a fast value of six per hour  as merozoites invade erythrocytes rapidly.TrLB controlling time distribution of liver\u2010stage maturation was described by: T50 was the time by which half the hepatocytes burst after inoculation, and \u03c3LB was the distribution of release around time T50. Initially, T50 was set to 6\u2009days, as a mid\u2010value between 5 and 7\u2009days,\u03c3LB at 0.1\u2009h, thereby defining that more than 98% of merozoites were released between Day 6 \u221230\u2009min and Day 6 +30\u2009min.Regarding release of merozoites, P. falciparum in the blood stage. As infection started immediately in the blood, only the equation describing parasite dynamics in blood was used . Identifiability of hB requires parasitemia minima to be observed that, here, was BLOQ. Therefore, to achieve stable convergence, Emax,B, EC50,B , and kin,B were estimated with fixed values for hB between 1 and 10. Furthermore, as baseline parasitemia was different between the IBSM and phase IIa, study type was added as a covariate. The model with the smallest Akaike information criterion (AIC) was selected. As an indicator, minimum inhibitory concentration against blood stage, MICB, was estimated for each individual as: Parasitemia data from the IBSM and phase IIa studies were used, where individual PK parameters were added as regressors in the PKPD relationship of DSM265 against 6 cells/g of liver). The liver parasite growth rate was derived from biological knowledge. Each invaded hepatocyte releases 10,000 to 50,000 merozoites after the 5\u2010 to 7\u2010day liver stage.GRL,pop was calculated to be 0.072 per h. Only the initial number of invaded hepatocytes remained unknown, which is proportional to Finc. Using placebo data from all spz HuCh studies, Finc was evaluated in two steps. In Step 1, the blood\u2010stage equations were used to estimate the blood parasite growth rate GRB for each individual from the isolated growth phase data. In Step 2, Finc and its IIV were estimated using the PKPD model, with GRB as a regressor from Step 1, fixing the liver parasite growth rate for each individual, GRL,indiv, to be the population estimate GRL,pop times the ratio GRB,indiv/GRB,pop to maintain 100% correlation between the liver and blood growth rates relative to the population value. No distinction was made between volunteers who received five mosquito bites or an i.v. dose of 3200 sporozoites as the infection observed was similar. Therefore, each mosquito was assumed to deliver 640 sporozoites. As with the blood stage, minimum inhibitory concentration against liver stage, MICL, was estimated for each individual as: Liver parasite growth is not directly observable in clinical studies, as this would require biopsies to estimate infected hepatocytes. These are not performed because of a low likelihood of detection due to a low number of ejected sporozoites by mosquito bite  after i.v. inoculations to the number of volunteers in each treatment group. The 90% confidence interval (CI) for each treatment group was calculated using the Clopper\u2013Pearson intervalEmax,L was zero; the second assumed that liver and blood activities were equivalent, that is, Emax, EC50, h, and kin were the same. By comparing and contrasting the simulation predictions against the observed data, one can ascertain which scenario better described the cure success rates.To discriminate whether the final chemoprophylaxis model best described observed cure, the success rates were simulated as noted previously in two hypothetical scenarios for liver\u2010stage activity. The first scenario assumed that DSM265 was active only during the blood stage, that is, To evaluate the chemoprophylaxis PKPD model for predicting efficacy in a real\u2010world prophylactic paradigm, eight simplified scenarios were simulated. In all scenarios, infection occurred at Day 0 with the same level as in the spz HuCh studies (3200 sporozoites), with two doses of DSM265 at 400\u2009mg administered weekly. The first dose was administered between Day \u22127 and Day 0 prior to infection. For each scenario, success rates and 90% CI at Day 28 after infection were estimated by sampling PK and PD parameters, accounting for IIV and uncertainty, for 100 virtual trials each with 100 patients.EC50,L, where 70%\u201380% of samples were BLOQ, the analysis was more categorical than traditional minimizing of residuals between predictions and measured observations. BLOQ data and predictions are visualized in the goodness\u2010of\u2010fit diagnostics supplement  combined with the IQRtools package  and Monolix (Version 2018R2). Although there were few PK data points BLOQ, many parasitemia samples were BLOQ. Consequently, it was vital that the M3 method was implemented in Monolix to allow for categorical information from these samples to be included in the analyses. For sections of the analysis, for example, Step 5 estimation of ent Text\u00a0.Data from 190 participants were compiled: 55 from FIH, 15 from IBSM, 24 from a phase IIa study of DSM265 treating uncomplicated blood\u2010stage infection, 39 from spz HuCh studies (including 10 placebo), and 57 placebos from published studies. Data are summarized in Table\u00a0Q1, where IIV was set to zero due to high shrinkage. Regarding covariates, only dose level on relative bioavailability was statistically significant (reduction in AIC \u226510) and retained. The final PK model showed excellent overall diagnostic performance  and 2.5\u2009\u03bcg/ml  for volunteers and patients, respectively.In field studies with patients presenting with malaria infection, the parasite growth rate cannot be observed before drug administration as is possible in a VIS. Therefore, it was assumed that patients and volunteers have a similar growth rate, el Table\u00a0 showed ael Table\u00a0, Step 2.GRB following liver infection, the growth rate was estimated to be 0.062 per h with an IIV of 0.12 (Table\u00a0Where the blood\u2010stage equation (Equation 1.2) without hepatocyte transfer function was used to estimate the growth rate of parasites in blood\u2010stage 12 Table\u00a0, Step 3.12 Table\u00a0, Step 4,Emax for liver was equal to that in blood and that the turnover rate kin was the same for both pools allowed estimation of the liver\u2010stage EC50,L of 1.6\u2009\u03bcg/ml with an IIV of 0.45 . It is worth noting that EC50,L and MICL are expressed as blood concentrations that characterize activity in the liver and not as liver concentrations.Assuming that the 45 Table\u00a0, Step 5,For each treatment group, the observed and predicted success rates were compared to evaluate the predictability of the full PKPD model Figure\u00a0. The preAs sporozoites transit the liver prior to being observed in the blood, presystemic antimalarial drug activity will affect observed parasitemia; the unknown is the extent.MIC of 1.5\u2009\u03bcg/ml for the liver activity of DSM265, it may be considered surprising that individuals infected the furthest from the first drug administration were not fully protected. However, MIC is the concentration such that killing equals growth; it is not parasiticidal. For a patient to be malaria free, one needs far higher concentrations. Furthermore, there is IIV in the PK and PD parameters such that some patients will have too short a half\u2010life, too large a volume, and/or too high a MIC; these will be the failures.DSM265 protection was evaluated in eight simplified real\u2010life scenarios, where infection occurred on Day 0 with two 400\u2010mg doses of DSM265 administered weekly, with the first dose administered between Day \u22127 and Day 0 prior to infection. The model predicted that the closer to infection the first administration occurred, the better a subject would be protected . This may be useful in selecting candidate compounds for phase Ib (spz HuCh studies) and phase II clinical trials. The proposed scenarios described herein could be used to help select suitable doses and regimens to achieve the required success rates for phase II. However, to get closer to phase II study designs run under real\u2010world conditions, further factors need consideration. These include different infection rates in different geographic areas and different levels of infection, treatment durations, and compliance. This could be achieved by combining knowledge from epidemiology and \u201cadherence behavior\u201d. Future model development could also be adapted to include multiple mosquito bite scenarios, incorporating the unpredictability of real\u2010world conditions. Such a model could be used to estimate what drug dose would be required to produce higher rates of protection, for instance, in 75%\u201395% of cases, or to predict whether practical frequencies, such as monthly, could be viable.A chemoprophylaxis PKPD model with two compartments to mathematically represent liver\u2010 and blood\u2010stage parasitemia was created to support the development of chemoprophylactic drugs. However, because liver\u2010stage parasites cannot be directly observed, there remained the challenge of estimating parasite transfer and drug\u2010induced killing parameters. By assuming similar activity for DSM265 for both the liver and blood stages, as observed in vitro,It was also necessary to challenge the assumptions of the model. This was done by simulating spz HuCh assuming no liver activity or similar levels of activity for the liver and blood. This showed that liver activity cannot be ignored for DSM265, but imputing blood activity into liver activity already gave feedback on the level of chemoprotection afforded.T50, its growth rate GRL, distribution of merozoite release (controlled by \u03c3LB), differences between stages in killing rate , or in in vitro potency , their impact on estimation of liver\u2010stage PD parameters, and clinical outcomes. Due to the low numbers of parasites at the beginning of infection, parasite dynamics should be better described by a randomly varying stochastic model than a continuous model. However, although median parasite dynamic might still be well described, variability is more difficult to interpret as it is not straightforward to fix a threshold for cure (a parasite reduction threshold below that which can claim complete cure). This could explain why the model underpredicted the efficacy of DSM265 when administered 1 day prior to infection.One could also use sensitivity analysis to quantify the effect of various assumptions, such as duration of liver infection In conclusion, one of the biggest challenges facing malaria prevention is a problem of knowledge concerning liver activity during the malaria parasite infection process. However, the ability to infer such information by way of mathematical modeling is a means of changing this. If blood\u2010stage activity is known, then PKPD models can be used to assess the levels of chemoprotection delivered by a new compound. If liver\u2010stage activity were characterized in in vitro assays, one can refine those assessments, as explained previously. By combining with sensitivity analysis to derisk uncertainty on liver parameters, this could be useful in selecting suitable compounds for phase Ib and II clinical trials, especially if combined with knowledge from epidemiology and patient compliance. This would augment the transition from preclinical through translational medicine to later confirmatory stages of development for new and improved antimalarials.M.H.C.\u2010R. wrote the manuscript. M.H.C.\u2010R., N.A., N.G., and J.J.M. designed the research. M.H.C.\u2010R. performed the research. M.H.C.\u2010R., N.A., L.B., O.F.E., R.F., C.F., M.G., and J.W. analyzed the data. M.H.C.\u2010R., N.A., L.B., O.F.E., R.F., C.F., M.G., and J.W. contributed to the new analytical tools.www.mmv.org) and in its annual report. MMV receives support from Bill and Melinda Gates Foundation Grant INV\u2010007155.This work was funded in whole by Medicines for Malaria Venture (MMV). A full list of donors to MMV is available on its website (M.H.C.\u2010R., J.J.M., and N.G. are employees of Medicines for Malaria Venture, as was N.A. All other authors declared no competing interests for this work.Table S1Click here for additional data file.Table S2Click here for additional data file.Table S3Click here for additional data file.Text S1Click here for additional data file.Data S1Click here for additional data file.Data S2Click here for additional data file.Data S3Click here for additional data file.Data S4Click here for additional data file.Data S5Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria was investigated.The contribution of artefenomel to the clinical and parasiticidal activity of ferroquine and artefenomel in combination in uncomplicated This Phase 2a, randomized, open-label, parallel-group study was conducted from 11th\u00a0September 2018 to 6th November 2019 across seven\u00a0centres in Benin, Burkina Faso, Gabon, Kenya, and Uganda. Patients aged\u2009\u2265\u200914\u201369\u00a0years with microscopically confirmed infection  were randomized 1:1:1:1 to 400\u00a0mg ferroquine, or 400\u00a0mg ferroquine plus artefenomel 300, 600, or 1000\u00a0mg, administered as a single oral dose. The primary efficacy analysis was a logistic regression evaluating the contribution of artefenomel exposure to Day 28 PCR-adjusted adequate clinical and parasitological response (ACPR). Safety was also evaluated.0\u2013\u221e to Day 28 PCR-adjusted ACPR was not demonstrated when accounting for ferroquine AUC0\u2013d28, baseline parasitaemia, and other model covariates: odds ratio 1.1 . In the per-protocol population, Day 28 PCR-adjusted ACPR was 80.8%  with ferroquine alone and 90.3% , 90.9%  and 87.1%  with 300, 600, and 1000\u00a0mg artefenomel, respectively. Median time to parasite clearance (Kaplan\u2013Meier) was 56.1\u00a0h with ferroquine, more rapid with artefenomel, but similar for all doses (30.0\u00a0h). There were no deaths. Adverse events (AEs) of any cause occurred in 51.4% (18/35) of patients with ferroquine 400\u00a0mg alone, and 58.3% (21/36), 66.7% (24/36), and 72.7% (24/33) with 300, 600, and 1000\u00a0mg artefenomel, respectively. All AEs were of mild-to-moderate severity, and consistent with the known profiles of the compounds. Vomiting was the most reported AE. There were no cases of QTcF prolongation\u2009\u2265\u2009500\u00a0ms or\u2009>\u200960\u00a0ms from baseline.The randomized population included 140 patients. For the primary analysis in the pharmacokinetic/pharmacodynamic efficacy population (N\u2009=\u2009121), the contribution of artefenomel AUCThe contribution of artefenomel exposure to the clinical and parasitological activity of ferroquine/artefenomel could not be demonstrated in this study. Parasite clearance was faster with ferroquine/artefenomel versus ferroquine alone. All treatments were well tolerated.Trial registration: ClinicalTrials.gov, NCT03660839 .The online version contains supplementary material available at 10.1186/s12936-022-04420-2. Plasmodium falciparum malaria. However, the emergence and spread of parasite strains resistant to both the artemisinin and partner components in the Greater Mekong region has undermined clinical efficacy for several\u00a0approved ACT options [P. falciparum malaria [P. falciparum strains harbouring Pfkelch13 (k13) mutations indicative of artemisinin resistance have been found in travellers returning from Africa [P. falciparum associated with the A675V k13\u00a0mutation has been reported [Artemisinin-based combination therapy (ACT) is the current first-line treatment for uncomplicated  options , 2. In A malaria \u20135. Howevm Africa , 7, and m Africa . The emem Africa , 9, and m Africa , Angola m Africa , and Burm Africa . Importam Africa . Similarreported . HoweverConsidering the emergence of clinical resistance to ACT, new treatment options are clearly required. Ferroquine and artefenomel are novel anti-malarial drug candidates and a combination of these two drugs has been proposed as a treatment for uncomplicated malaria. New anti-malarial therapeutics should be developed as fixed-dose combinations as this is expected to improve patient adherence and reduce the risk of parasite resistance developing\u00a0to either drug, particularly if they have contrasting modes of action. When developing fixed-dose anti-malarial drug combinations, it is necessary to demonstrate the contribution of each component to overall efficacy and establish the safety and tolerability profile with co-administration \u201318.P. falciparum and was well tolerated [Artefenomel  is a synthetic peroxide, with a similar mechanism of action to artemisinin , 20. In olerated . In healolerated , and an olerated .P. falciparum [Ferroquine is a 4-aminoquinoline analog, and a strong inhibitor of hemozoin formation, with high efficacy against chloroquine-resistant and ACT-resistant lciparum \u201328. Singlciparum , and potlciparum . Pharmaclciparum . AlthougPlasmodium falciparum malaria\u2019 (FALCI) Phase 2 study, investigating artefenomel (800\u00a0mg) plus ferroquine  in African and Asian patients aged\u2009>\u20096\u00a0months to\u2009<\u200970\u00a0years with uncomplicated falciparum malaria [k13\u00a0C580Y mutation known to be associated with artemisinin resistance [The efficacy and safety of ferroquine/artefenomel was evaluated in the \u2018Ferroquine and Artefenomel in adults and children with  malaria . Efficacsistance .P.\u00a0falciparum malaria.FALCI was designed with only one dose level of artefenomel, and so there was a risk that the study would not be able to identify the contribution of artefenomel. Consequently, this parallel investigation was designed to specifically evaluate the artefenomel contribution to the efficacy of the combination in African adolescents and adults with uncomplicated This Phase 2a, randomized, open-label, parallel-group study was conducted from 11th\u00a0September 2018 to 6th November 2019 across seven\u00a0study centres in Benin (Cotonou), Burkina Faso (Banfora and Nanoro), Gabon (Libreville and Lambar\u00e9n\u00e9), Kenya (Kisumu), and Uganda . The primary objective was to show the contribution of artefenomel to the clinical and parasitological activity of artefenomel and ferroquine in combination by analysing the exposure\u2013response of artefenomel in patients with uncomplicated falciparum malaria. To achieve its objective, some patients would receive a sub-therapeutic dosing regimen, and this was considered in the design by using dedicated risk minimization activities, i.e., selection of a patient population at low risk of severe malaria, hospitalization for at least 48\u00a0h or longer based\u00a0upon the investigator\u2019s judgment, and administration of rescue therapy as soon as there was evidence of treatment failure or systematically on Day 29. The study protocol is provided as Additional file Investigational products were ferroquine 100\u00a0mg capsules  and artefenomel 300/400/600\u00a0mg granules formulation  presented in a sachet with alpha tocopherol polyethylene glycol 1000 succinate formulation and sucrose. Dose selection aimed to characterize the anti-malarial contribution of artefenomel to the combination. Artefenomel has been evaluated to doses up to 1200\u00a0mg with no safety concerns, and a wide range of doses (0\u20131000\u00a0mg) was selected to evaluate the exposure\u2013response. The 400\u00a0mg ferroquine dose was selected as a sub-therapeutic dose so as not to mask the contribution of artefenomel. Details of the dose selection methods are in Additional file Patients were randomized centrally using interactive response technology in a ratio of 1:1:1:1 to 400\u00a0mg ferroquine alone, or 400\u00a0mg ferroquine plus artefenomel 300, 600, or 1000\u00a0mg. The administration of treatments was open label as a single oral dose on Day 0 and was directly observed. Ferroquine was administered in a fasted condition. Artefenomel was prepared as a suspension in sterile water and given approximately 15\u00a0min after ferroquine. If vomiting occurred after ferroquine, the patient was not re-dosed, artefenomel was not administered, and the patient received rescue therapy. If the artefenomel dose was vomited within 5\u00a0min of administration, the patient was re-dosed. Vomiting within 5\u201335\u00a0min of artefenomel administration did not prompt redosing, but any remaining drug was given. Rescue anti-malarial therapy as per local recommendations was administered to patients before Day 28 if clinically indicated, if the ferroquine dose was vomited, or at Day 29 if not given previously.P. falciparum malaria  plus fever or a history of fever in the previous 24\u00a0h. All participants were required to use effective contraception and pregnant or lactating women were excluded. Key exclusion criteria were severe malaria [Plasmodium infection, clinically important medical conditions, severe vomiting or diarrhoea, severe malnutrition [To evaluate exposure\u2013response, a sub-therapeutic dosing regimen was to be administered. Thus, the study population was selected to be at low risk of severe malaria. Eligible participants were aged 14 to 69\u00a0years, body weight 35\u201395\u00a0kg, of either sex, presenting with microscopically confirmed uncomplicated  malaria , mixed Putrition , splenecScreening procedures included physical examination, medical history, 12-lead electrocardiogram (ECG), vital signs, clinical laboratory tests, viral hepatitis serology, and a pregnancy test. Patients were hospitalized for at least the first 48\u00a0h following treatment administration and longer if malaria symptoms or parasitaemia persisted. Patients were followed up on Days 3, 4, 5, 6, 7, 10, 14, 21 and 28. All patients received definitive approved anti-malarial treatment on Day 29 if they had not already received rescue therapy.P. falciparum marker genes msp1, msp2 and glurp, new infection was assumed when all the alleles in parasites from the post-treatment sample were different from those in the baseline sample, for one or more loci tested. Recrudescence was defined as at least one allele at each locus common to paired samples from baseline and at recurrence [Blood samples for parasite assessments were taken at screening, every 6\u00a0h until 36\u00a0h post-dose, at hours 48, 72, 96, 120, 144, and 168 post-dose, on Days 10, 14, 21, and 28, and at any time if clinically indicated. Giemsa-stained thick and thin blood films were prepared, and parasites identified and enumerated independently by two trained microscopists using standard procedures . ParasitAdverse events were assessed throughout the study. Additional post-treatment safety assessments were 12-lead ECGs, vital signs, haematology, and clinical laboratory tests  and ferroquine (11 sample time points)  at Day 28, defined as the absence of parasitaemia without previous treatment failure or rescue therapy, adjusted for re-infection using PCR genotyping .24, PRR48, and PRR72); parasite clearance rate; and time to parasite reduction by 50% (PC50) and 99% (PC99) of baseline parasitaemia, time for parasitaemia to reduce by 50% (TPC50) and 90% (TPC90) independent of baseline parasitaemia, and the estimated parasite reduction ratio at 24, 48 and 72\u00a0h post-dose  ; parasitaemia at baseline then every 6\u00a0h during the first 36\u00a0h post-dose, then at 48\u00a0h and every 24\u00a0h until Day 7; parasite clearance time; time to parasitaemia re-emergence, recrudescence, or reinfection; time elapsed below the LLQ of parasitaemia; fever clearance time; observed parasite reduction ratio at 24, 48 and 72\u00a0h post-dose , the frequency of serious adverse events, clinically important changes in clinical laboratory data, ECGs, vital signs, or physical examination. Adverse events of special interest were pregnancy, symptomatic overdose, increase in ALT\u2009\u2265\u20093xULN , QTcF\u2009\u2265\u2009500\u00a0ms, or QTcF prolongation\u2009>\u200960\u00a0ms from baseline.Cmax), concentration at Day 7 post-dose (Cd7), area under the concentration\u2013time curve from time 0 to infinity (AUC0\u2013\u221e) for artefenomel and ferroquine, and AUC from time 0 to Day 28 (AUC0\u2013d28) for ferroquine and desmethyl-ferroquine only.Pharmacokinetic assessments were secondary endpoints, but also supported the primary analysis evaluating the exposure\u2013response for artefenomel  population was a sub-set of the mITT population who were evaluable for Day 28 ACPR with no major protocol violations. The pharmacokinetic/pharmacodynamic (PK/PD) efficacy population was the primary efficacy analysis population and included patients in both the PK and mITT populations who had at least one evaluable PK sample for both artefenomel and ferroquine. Thus, patients who vomited or who did not receive a complete dose of study drug were not excluded from the PK/PD efficacy population.The safety population included all randomized patients who received one dose or a partial dose of the investigational drugs. The PK population was a sub-set of the safety population with at least one evaluable PK blood sample for either artefenomel or ferroquine. The microbiological intention-to-treat population (mITT) included all randomized patients who received the investigational drugs, had microscopically confirmed Sample size was based on the estimated efficacy for artefenomel  plus ferroquine (400\u00a0mg) derived from clinical trial simulations assuming a parasitaemia\u2009>\u20093000 parasites/\u00b5L  combined with MONOLIX (MLX2019R1) and the IQR package (v1.1.1) developed by IntiQuan  to support the entire workflow of a PK and logistic regression analysis from estimations to simulations. For simulations, IQR uses the library SUNDIALS (v2.9.0) from Computation (USA)  and ferroquine/desmethyl-ferroquine (AUC0\u2013d28) as covariates as well as baseline parasitaemia, age, body weight, sex, vomit status, and study centre  and P values were calculated for covariates. As a secondary efficacy analysis, the relationship between the estimated exposure of artefenomel and ferroquine and Day 28 crude ACPR was evaluated as described for Day 28 PCR-adjusted ACPR.The base model included ferroquine AUCFor other secondary efficacy outcomes and safety outcomes, statistical analysis was done using SAS (version 9.4). Day 28 PCR-adjusted and crude ACPR were summarized for the PP and mITT populations, with Clopper\u2013Pearson 95% CI. All other secondary efficacy outcomes were evaluated in the mITT population. Parasite clearance parameters were calculated using the WorldWide Antimalarial Resistance Network parasite clearance estimator (WWARN PCE) based on the linear portion of the individual natural logarithm parasitaemia\u2013time profiles . The timPharmacokinetic analysis was performed using non-linear mixed effect modelling as implemented in Monolix (version 2019R1), applying previously developed population PK models , 5.6% (2/36), and 3.0% (1/33) of patients in the 300\u00a0mg, 600\u00a0mg, and 1000\u00a0mg arms, respectively. After 35\u00a0min following dosing, vomiting was noted for 8.3% (3/36) of patients with 300\u00a0mg, 11.1% (4/36) with 600\u00a0mg, 24.2% (8/33) with 1000\u00a0mg artefenomel. No patients were given rescue medication because of vomiting.The PK/PD efficacy population included 132 patients, but five cases of reinfection and six with undetermined PCR results were considered missing for the Day 28 PCR-adjusted ACPR analysis, with 121 patients evaluable. Baseline demographic and clinical characteristics, drug exposure, and Day 28 PCR-adjusted ACPR outcome for this population are shown in Additional file 0\u2013\u221e and ferroquine AUC0\u2013d28 are shown in Fig.\u00a0Details of the PK analysis supporting the primary endpoint including concentration\u2013time profiles and summaries of estimated exposures for artefenomel, ferroquine and desmethyl-ferroquine relative to dosing are provided in Additional file\u00a0Day 28 PCR-adjusted ACPR was achieved by 94.4% 34/36) of patients who had an estimated AUC for both artefenomel and ferroquine above the median value, 92.0% (23/25) of patients when only exposure to artefenomel was above its median value, 88.0% (22/25) when only ferroquine exposure was above its median value, and 77.1% (27/35) when exposure to both drugs was below the median value , whereas artefenomel exposure was not. Univariate analysis indicated no effect on Day 28 PCR-adjusted ACPR of desmethyl-ferroquine exposure (P\u2009=\u20090.113), age (P\u2009=\u20090.192), sex (P\u2009=\u20090.0524), or body weight (P\u2009=\u20090.158). As exploring the contribution of artefenomel exposure to Day 28 PCR-adjusted ACPR was the primary objective, this covariate was included for the backward elimination analysis. The backward elimination suggested only the removal of artefenomel exposure.Full details of the exposure\u2013response analysis are provided in Additional file 0\u2013\u221e for Day 28 PCR-adjusted ACPR was 1.1    with ferroquine monotherapy. There was a trend for higher efficacy with artefenomel co-administration, though efficacy was similar across the artefenomel doses: 90.3%  with 300\u00a0mg, 90.9%  with 600\u00a0mg, and 87.1%  with 1000\u00a0mg  patients with reinfection in the ferroquine group, and 1/33 (3.0%), 2/36 (5.6%) and 0/32 (0%) with artefenomel 300, 600 and 1000\u00a0mg, respectively. Six of the recurrences had undetermined PCR results.Day 28 crude ACPR was 64.5%  for ferroquine alone, with a trend for higher efficacy in the artefenomel arms: 81.8%  with 300\u00a0mg, 77.8%  with 600\u00a0mg, and 78.1%  with 1000\u00a0mg . However, there was a trend over the 28-day study period for a lower probability of re-emergence and recrudescence with the combination arms versus ferroquine alone Fig.\u00a0.Fig. 6TiThe estimated median time to parasite clearance (Kaplan\u2013Meier) was 56.1\u00a0h  with ferroquine alone, but was more rapid in the combination arms, without an effect of artefenomel dose: 30.0\u00a0h  with 300\u00a0mg, 30.0\u00a0h  with 600\u00a0mg, and 30.0\u00a0h  with 1000\u00a0mg artefenomel  was 4.08 (SD 1.59) with ferroquine alone and ranged between 7.46 and 8.26 in the combination arms. Similarly, mean PC50 was 14.07 (SD5.6) compared with 7.74 to 9.47 for the combination arms  of patients in the ferroquine only group, and in the combination arms for 58.3% (21/36) with 300\u00a0mg, 66.7% (24/36) with 600\u00a0mg, and 72.7% (24/33) with 1000\u00a0mg artefenomel. The most common adverse events were malaria  with ferroquine alone, malaria and headache  with artefenomel 300\u00a0mg, malaria  with artefenomel 600\u00a0mg, and vomiting  with 1000\u00a0mg artefenomel  of patients with ferroquine, most commonly vomiting 3.6%; 5/140)  observed in a female patient with baseline elevations of ALT (1.1xULN), alkaline phosphatase (ALP) (3.2xULN), and total bilirubin (1.2xULN), and no history of hepato-biliary disorders. From Day 5, ALT increased to a maximum of 3.3xULN on Day 15, with concurrent ALP 5.4xULN, AST 1.3xULN, and total bilirubin 1.2xULN. Total bilirubin continued to rise, peaking at 3.4xULN on Day 25 with direct bilirubin at 4.7xULN and ALP at 5xULN. There were no symptoms, and ALT, AST, and total bilirubin values had returned to normal by Day 42 without intervention. Further elevations in ALT (1.7xULN), AST (1.5xULN), total bilirubin (1.3xULN), and ALP (2.6xULN) were noted on Day 96, but all measures had resolved to baseline levels by Day 141.There was no difference in the maximum post-baseline change in haematology parameters between treatment groups, which were consistent with recovery from malaria  to the clinical and parasiticidal activity of the artefenomel/ferroquine combination, defined as Day 28 PCR-adjusted ACPR, could not be identified in this study. The study also failed to identify the contribution of artefenomel exposure to the effect of the artefenomel/ferroquine combination for Day 28 crude ACPR. This occurred even though clinical trial simulations were performed to support the dose selection and sample size, and even though a wide range of individual artefenomel exposures (0\u201313.05\u00a0\u00b5g*h/ml) were observed.The contribution of artefenomel exposure , whereas the observed response with ferroquine alone (80.8%) was close to the upper limit of predicted efficacy. With a lower than predicted number of treatment failures across all treatment arms, it was statistically more difficult to detect a significant contribution of artefenomel.The reasons for a higher-than-expected response to ferroquine are unclear. However, this does not appear to result from higher than anticipated ferroquine or active metabolite exposures in this study population. Notably, limited exposure\u2013response data were available for ferroquine to inform the clinical trial simulations and no efficacy data for the artefenomel/ferroquine combination were available. Consequently, there was considerable uncertainty associated with the predictions for ferroquine exposure\u2013response. However, this uncertainty in the predicted ferroquine response was not considered in the clinical trial simulations.The upper limit for parasitaemia for enrolment in this study was 50,000 parasites/\u00b5L for ethical reasons, considerably lower than the maximum parasitaemia level used in the clinical trial simulations . Both in this study and in FALCI, higher baseline parasitaemia was shown to significantly decrease the odds of Day 28 PCR-adjusted ACPR . The lowIn FALCI, there was evidence that age influenced baseline parasitaemia . AfricanP. falciparum parasitaemia of\u2009\u2265\u20093,000 to\u2009\u2264\u200950,000 parasites/\u00b5L blood. Given that the ferroquine only arm was not expected to achieve clinical cure rates\u2009>\u200972%, this population was selected to minimize the risk of developing severe malaria. Note that data from Burkina Faso indicate that asymptomatic carriage of malaria is rare at parasite densities\u2009\u2265\u20092,500 parasites/\u00b5L blood, occurring at a rate of 5.3% (7/133) in patients aged\u2009>\u200915\u00a0years [Overall, eligible participants were aged 14 to 69\u00a0years, with 15\u00a0years . Similar15\u00a0years . Thus, aNo new safety signals were identified, and the safety findings were consistent with previous clinical studies , 40\u201343. Although there was one case of increased ALT, there was no suggestion of a relationship between artefenomel dosing and ALT increases. No patient had a QTcB or QTcF\u2009>\u2009500\u00a0ms or an increase from baseline\u2009>\u200960\u00a0ms. These findings contrast with FALCI, as increased ALT was reported in 2.1% (8/373) of patients and appeared to be associated with ferroquine . There wThis study illustrates the challenges in designing clinical trials to demonstrate the contribution of individual drugs to the overall efficacy of a combination therapy in a disease which if not rapidly treated can progress to a serious and life-threatening condition. To explore the contribution of the individual drugs, sub-therapeutic doses must be administered. However, the risk to the patients from disease progression must also be minimized. Consequently, in this study the patient population excluded children\u2009<\u200914\u00a0years of age who are most at risk of severe malaria, restricted baseline parasitaemia to the range\u2009\u2265\u20093000 to\u2009\u2264\u200950,000 parasites/\u00b5L blood and used drug doses that were not substantially sub-therapeutic.The contribution of artefenomel exposure to Day 28 PCR-adjusted ACPR could not be demonstrated. The main reason was likely the higher than anticipated efficacy with ferroquine alone which reduced the power of the study to identify the contribution of artefenomel to the drug combination, rather than a true lack of contribution. More rapid parasite clearance was demonstrated with the artefenomel/ferroquine combination compared to ferroquine alone, clearly showing the contribution of both drugs to parasite clearance. The combination was generally well tolerated, and the safety profile was consistent with the known profiles of the two compounds.Additional file 1: Study protocol.Additional file 2: Clinical trial simulation to determine the dose and sample sizes.Additional file 3: Pharmacokinetic analysis supporting the primary analysis.Additional file 4: Exposure\u2013response pharmacokinetic/pharmacodynamic analysis.Additional file 5. Supplementary tables and figures."}
+{"text": "The assessment in real\u2010life conditions of the safety and efficacy of new antimalarial drugs is of greatest interest. This study aimed to monitor and evaluate both clinical and biological safety of pyronaridine\u2010artesunate (PA) in real\u2010life conditions in Burkina Faso's health system. This was a single\u2010arm, open\u2010label study, where patients attending Nanoro health facilities with uncomplicated malaria were consented to be part of a cohort event monitoring (CEM). At inclusion (day\u20100), PA was administered orally once a day for 3\u2009days. Patients spontaneous reported any clinical adverse events (AEs) occurring within 28\u2009days following the treatment. Additionally, the study focused on AEs of special interest (AESI), namely clinical signs related to hepatotoxicity and increased alanine aminotransferase (ALT) and aspartate aminotransferase (AST). A nested subset of patients with blood sample collection at day\u20100 and day\u20107 were monitored to investigate the effect of PA on biochemistry parameters. From September 2017 to October 2018, 2786 patients were treated with PA. About 97.8% (2720/2786) of patients did not report any AE. The most commonly reported events were respiratory, thoracic, and mediastinal disorders (8.3 per 1000), infections and infestations (7.9 per 1000), and gastrointestinal disorders (7.2 per 1000). No clinical or biological hepatotoxicity event related to PA was reported during the follow\u2010up. Changes in biochemistry parameters remained within laboratory reference ranges. The study showed that PA is a well\u2010tolerated drug and should be considered as a good option by malaria control programs in countries where existing first\u2010line antimalarial drugs are continuously threatened by the emergence of drug resistance. ACTartemisinin\u2010based combination therapiesADRsadverse drug reactionsAEsadverse eventsAESIAEs of special interestALartemether\u2010lumefantrineALTalanine aminotransferaseASAQartesunate\u2010amodiaquineASTaspartate aminotransferaseCEMcohort event monitoringDHA\u2010PQPdihydroartemisinin\u2010piperaquineHCWshealth care workersHDSSHealth and Demographic Surveillance SystemMMVMedicines for Malaria VentureNMCPNational Malaria Control ProgramPApyronaridine\u2013artesunateSMCSeasonal Malaria ChemopreventionSSAsub\u2010Saharan Africa1With the contribution of international organizations such as Medicines for Malaria Venture (MMV) and other public\u2013private partnerships, new effective and well\u2010tolerated antimalarial drugs for large\u2010scale use are getting available on the market.Plasmodium falciparum and blood\u2010stage P. vivax according to multicenter clinical studies in Africa and Asia.P. falciparum demonstrated that PA is equivalent to the other marketed ACTs.In Burkina Faso, since 2008, fixed\u2010dose artemisinin\u2010based combination therapies (ACTs) were adopted as treatment for uncomplicated malaria.22.12 and lies between longitudes 1\u00b0892\u2009537 and 2\u00b083\u2009146 West and latitudes 12\u00b0857\u2009955 and 12\u00b0872\u2009863 North. Malaria is hyper\u2010endemic and peaks during the rainy season (July to November). The number of inhabitants covered by the HDSS was estimated at about 60\u2009000 in 2018, of whom 20% were children under 5\u2009years of age. At the same period, malaria represented the top one of all diseases, accounting for about 41.3% of outpatient medical consultations and 57.0% of hospitalizations. Children under 5\u2009years of age are the most vulnerable group affected by malaria, accounting for 58.5% of all malaria cases. In addition, the study area is characterized by a marked seasonality of malaria transmissionThe study was carried out from September 2017 to October 2018 in the health district of Nanoro through the Health and Demographic Surveillance System (HDSS).2.2This was a single\u2010arm, open\u2010label observational, non\u2010comparative phase IV study. All patients regardless of gender and age attending the health facilities in the Nanoro HDSS catchment area and for whom a diagnosis of uncomplicated malaria was posed (or suspected) by the public sector health care workers (HCWs) were invited to participate in the study. Patients were consented to be part of a cohort event monitoring (CEM). All consented participants (main cohort) were asked to return voluntarily to health facilities and to spontaneously report any occurrence of adverse events. Eligible patients were aged over 6\u2009months, weighed more than 5\u00a0kg and were able to take oral medications, and signed informed consent (a parent or guardian consented for children below 18\u2009years old). Patients were excluded if they had any of the following: the presence of clinical signs or symptoms of hepatic injury  or known to have severe liver disease ; known to be pregnant or lactating; severe malaria; known allergy to artemisinin and/or to pyronaridine. A nested subset of patients (nested cohort) with blood sample collection at day\u20100 and day\u20107 were monitored to investigate the effect of PA on blood biochemistry parameters.Shin Poong Pharmaceuticals Limited donated PA (Pyramax for brand name) but did not have any role in reviewing the protocol or the manuscript.2.3At the enrolment visit (day 0), in both groups, data on medical and drug history were obtained from each eligible patient and were recorded on an individual case report form (CRF). For each patient, a detailed clinical examination was performed by the HCWs and the findings were recorded on the appropriate CRF. Data on concomitant medications for each patient were also collected.Patients included in both passive and active monitoring cohorts were requested to return voluntarily to health facilities and to spontaneously report any AEs occurring within 28\u2009days after the administration of the first dose of PA, whereas patients included only in the active monitoring cohort had a scheduled follow up visit at day 7 (\u00b12\u2009days) at health facility and home visit at day 28. For the latter group, data on clinical conditions and concomitant treatments were collected at each scheduled visit. Additionally, blood smears for thick and thin film and blood spot was systematically obtained on days 0 before drug administration and day 7 after drug administration. Venous blood samples were collected on day 0 before drug administration and day 7 after drug administration. Then, plasma samples were processed to investigate specific liver function tests (LFTs), namely Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), total bilirubin, as well as renal function parameters namely creatinine and urea. Blood glucose was also measured.In case of unscheduled visit or whether the patient returned voluntarily to spontaneously report an adverse event within the 28\u2009days after PA administration, data on clinical condition and concomitant treatments were collected. Additionally, blood smears for thick and thin film were collected and a venous blood sample was drawn for biological investigation according to the clinical judgment of the study physician.Although in Burkina Faso ACTs are recommended for pregnant women in the second and third trimester of pregnancy,2.4For enrolled patients, PA was administered every 24\u2009h from the first administration for 3\u2009days. The first dose (day 0) was administrated under direct supervision of a HCW at the health facilities. After drug intake, patients were observed 60\u2009min. For those who vomited within 30\u2009min, a complete dose was re\u2010administered, whereas for those who experienced vomiting within 30\u201360\u2009min, a half dose was re\u2010administered. Re\u2010administration was attempted only once. Then, HCW explained to the patients how to take the second and third doses at home after 24\u2009h day 1) and after 48\u2009h (day 2) from the initiation of the treatment. PA was administered according to the patient body weight. Two types of presentation of PA were used to facilitate the dosing and administration: sachet (granules) for children under 20\u2009kg and tablets for children and adults weighting more than 20\u2009kg. The tablet presentation was dosed at 180/60\u2009mg of PA, whereas the sachet presentation was dosed at 60/20\u2009mg of PA. Daily dosing according to the weight is shown in Table\u00a0 and afte2.5Patients in the active monitoring cohort had a scheduled follow\u2010up visit on day 7 (\u00b12\u2009days) at the health facility. During this visit, the treatment adherence, that is, patients who complied with the recommended treatment according to age, was verified retrospectively through self\u2010reporting. Additionally, the occurrence of an AE between the drug intake and the day of the visit was reported by the HCWs. A check\u2010in was performed to detect whether symptoms of malaria persisted or whether an AE had occurred since the first administration of PA. Patients were contacted again on day 28 to ascertain the AE recovery or to notify any further new AE. Apart from these scheduled follow\u2010up visits in the active cohort, similarly to those included in the passive group, patients were encouraged to return to the health facility to report any occurrence of AE. AEs were documented by the clinical team as described by the participant or caregiver. Information was reviewed and coded using the Medical Dictionary for Regulatory Activities (MedDRA version 22.1 September 2019) system organ class (SOC).Adverse event was defined in accordance with the International Conference of Harmonization (ICH) guidelines for Good Clinical Practice (GCP) as: any untoward medical occurrence, irrespective of its suspected relationship to the study medications.At each contact, patients were assessed according to a standardized checklist, and information about current signs and symptoms was collected, including the start and end date. A severity grading scale , was used to grade severity of all symptoms in accordance with the toxicity grading scales developed by the WHO  and the National Institutes of Health (NIH), Division of Microbiology and Infectious Diseases.2.6The study's main outcome of interest was both clinical and biological safety within the 28\u2009days after starting PA treatment. The clinical outcome  was evaluated through the analysis of the AEs captured by the study clinical team; whereas biological safety was defined as any significant change in liver transaminases values (elevated AST/ALT). Safety was assessed in all patients who received at least one dose of PA. Special attention was given to the AEs classified as severe and/or adverse events of special interest (AESI) related to hepatotoxicity . Serious liver reactions and hypersensitivity were also considered as an outcome of interest (AESI).Safety outcomes also included any significant change in the total bilirubin, creatinine, and urea values.2.7For the CEM for the active or nested cohort, all visit data were captured using paper CRF and then double\u2010entered and verified using a database designed under an open\u2010source software, OpenClinica. For the CEM for passive or main cohort at enrolment visit, data were collected during the medical examination using an electronic CRF developed on open\u2010source software, Open Data Kit (ODK) Collect and, in case of unscheduled visit, data were captured with paper CRF and then double\u2010entered on OpenClinica.Statistical analyses were performed using R software . Descriptive analyses for the total participants and for each cohort were performed as required. In this present analysis, the estimates of the incidence of each AE were performed based on crude rates, with no attempt to carry out ca ausality assessment of individual cases. For the biological safety assessment i.e. the blood biochemistry sample analysis, Wilcoxon matched\u2010pairs signed\u2010rank test was used to compare pre\u2010treatment (day 0) and post\u2010treatment (day 7) values in each parameter. This comparison was performed in all participants who had laboratory results at both day 0 and day 7.33.1A total of 2832 patients with suspected uncomplicated malaria were screened to be enrolled in the study, and 2786 (98.4%) patients were included. Of these, 1761 agreed to be in the main cohort (passive group), whereas 1025 agreed to be in a nested cohort (active group). Table\u00a03.2No patient was excluded at day 0 for repeated vomiting after drug intake. However, 4.3% (121/2786) of all patients experienced a vomiting episode within 1\u2009h after drug administration at day 0. Among patients who experienced an episode of vomiting, about, 91.7% (111/121) vomited within 30\u2009min after drug taken, whereas, 8.3% (10/121) vomited between 30 and 60\u2009min after drug administration. Table\u00a03.3Adverse events suggesting clinical signs and symptoms of possible hepatotoxicity, namely abdominal pain, fatigue, nausea, itching, or signs of jaundice occurred mostly among participants with normal versus abnormal baseline ALT/AST values Table\u00a0.3.4During the follow\u2010up period, the majority of patients (97.8% [2720/2786]), who received at least one dose of PA did not report any adverse event. Overall, only 2.2% (60/2786) of patients reported at least one AE in both group. About 0.4% ((7/1761) in the passive group and 5.2% (53/1025) in the active group) patients reported a total of 94 AEs (11 in the passive group and 83 in the active group). Of the reported AEs, six were classified as serious (SAEs) by the study clinicians. The frequently (cumulative incidence per 1000) reported events classified according to the MedDRA System organ classification were respiratory, thoracic, and mediastinal disorders (8.3 per 1000), infections and infestations (7.9 per 1000), gastrointestinal disorders (7.2 per 1000), general disorders (5.4 per 1000) and skin and subcutaneous tissue disorders (1.8 per 1000)  occurred in participants under the age of 5\u2009years. Two cases of death were recorded during the course of the study. These SAEs were not related to the PA. The details of these cases of death are provided in the The administrated treatment was well tolerated in general. Thirteen adverse events were assessed by study clinicians to be related to the study drug Table\u00a0. No seri3.5In general, the median biochemical values of biochemistry parameters on day 7 after PA treatment were low compared with the baseline values on day 0 Figures\u00a0\u2013S3. All 4This current study was conducted in the mindset to collect safety data to support the scale\u2010up of PA access in Burkina Faso as well as other developing countries where routine pharmacovigilance is limited. To address this growing need for interest of safety data on PA, this study showed that PA was clinically and biologically well tolerated when used in real\u2010life conditions in population without clinical liver disorders. Our findings are consistent with previous studies and most frequently observed AEs were similar to these previous studies and malaria symptoms.9and because in such area of SMC, ASAQ one of the first\u2010line therapy should no longer be used.In June 2017, PA was added to the WHO Model List of Essential Medicines and Model List of Essential Medicines for Children.In this study, the findings did not show the occurrence of serious AEs related to PA treatment during the study period. These findings assert for the wide use of PA in the routine healthcare systems in malaria\u2010endemic areas. In 2012, PA tablets were granted a positive scientific opinion by the European Medicines Agency (EMA), but with a restricted label, mainly due to concerns over potential hepatic risk of the pyronaridine component, the number of children under 5\u2009years of age treated in the program, and safety, especially with repeated dosing. These restrictions were lifted in 2015 after being reviewed by the EMA of the interim results of the WANECAM repeated doses study. Currently, several scientific research data on hepatic events had provided enough evidence to overcome these concerns about the hepatic riskThe incidence of suspected AEs related to PA reported in our study was relatively low (4.3 per 1000) and most of them were classified as moderate. By investigating the timing of occurrence and type of AEs reported in our study, it is noticeable that most events reported seemed to be related more to malaria than to the intake of PA. In this study, about 5.2% of patients included in the active group reported an AE. This rate was lower than those reported in the previous randomized clinical trial (RCT) carried out among the African or Asian population.Several reasons could explain this low rate including the design of the study, which consisted of self\u2010reporting of adverse events. Indeed the latter could not have the same effectivity in detecting adverse events as that achieved in phase 2 and 3 clinical trials. In addition, in this study, it was found that more than 92% of reported AEs were graded as moderate. These findings would suggest that, the AEs with mild severity were not spontaneously reported by the population to the HCWs. The main difference could be due to the fact that safety assessment is more stringent (in particular more visits) in phases II/III studies (more than 50% of the patients' report AEs) than in phase IV studies (5% to 20% of patients reporting AEs). A study assessing the clinical safety of a newly registered antimalarial drug (using the same methodology as described in our study) suggested that the low rate of AE reported in Phase IV study than Phase II and III could be explained by the fact of anxiety of the patient taking a new drug. In addition, the blinded condition in which such drugs are administered may cause an increase of AE reporting.5In a context where existing first\u2010line antimalarial drugs are continuously threatened by the emergence of malaria parasite\u00a0resistance to antimalarial drugs, the evaluation of the newly registered antimalarial drug, PA, showed a good safety profile in patients of all the age groups in our study. Therefore, PA has deserve of being added to the list of existing first\u2010line treatments of uncomplicated malaria in Burkina Faso. These findings are useful for the malaria control program in countries for policy decision\u2010making.TH, TCM, SP, VI, RT, BR, and BF contributed to protocol development. TH, TCM, SP, VI, RT, CA, YWI, TMC, HSF, NAD BR, and BF contributed to data collection and the overall implementation of the study. RT, TH, PS, and YWI worked on data analysis and interpretation of the data. RT and TH drafted the manuscript and all authors read and approved the final manuscript.The study was sponsored by the INDEPTH Network as part of the funding for the INESS program.The authors report no conflicts of interest in this work.The protocol was approved by the institutional ethics committees of the \u201cInstitut de Recherche en Sciences de la Sant\u00e9, Bobo Dioulasso, Burkina Faso (deliberation n\u00b0 20\u20102017/CEIRES),\u201d and the Ethics Committee for Health Research of Burkina Faso (2017\u20106\u2010075/ CERS).Written informed consent was obtained from all patients or legal guardians before performing any study\u2010related activity.Appendix S1Click here for additional data file."}
+{"text": "Plasmodium vivax malaria. The study was not completed for operational reasons and emerging teratotoxicity data. For the eight adult male patients enrolled, adequate clinical and parasitological response at day 14 (primary endpoint) was 100% (8/8). Asexual parasites and gametocytes were cleared in all patients by 66 and 78 hours postdose, respectively. There were two recurrent P. vivax infections (days 20 and 28) and a new Plasmodium falciparum infection (day 22). MMV390048 exposure in P. vivax patients was lower than previously observed for healthy volunteers. Mild adverse events, mainly headache and gastrointestinal symptoms, were reported by eight patients. Single-dose MMV390048 (120 mg) rapidly cleared asexual parasites and gametocytes in patients with P. vivax malaria and was well tolerated.An open label, phase IIa study conducted in Ethiopia evaluated the efficacy, safety, tolerability, and pharmacokinetics of a single 120-mg dose of the phosphatidylinositol 4-kinase inhibitor MMV390048 in  Plasmodium strains requires the discovery and development of new antimalarial drugs.Malaria remains major threat to global health.Plasmodium life cycle stages, excepting late-stage hypnozoites in the liver.\u2013,\u2013MMV390048 is a phosphatidylinositol 4-kinase inhibitor with in\u00a0vitro activity against all This open label, adaptive, phase IIa study was designed to evaluate the efficacy, safety, tolerability, and pharmacokinetics of a single 120-mg dose of MMV390048 in adult patients with uncomplicated malaria. The study was conducted between October 6, 2017 and January 5, 2018 at two hospitals in Ethiopia (in Gondar and Jimma). Recruitment was suspended on December 4, 2017 to allow assessment of a teratogenicity signal in a concurrent investigation in rodents.The study protocol was approved by the independent Ethics Committees of the College of Public Health and Medical Sciences, Jimma University , the Institutional Review Board of the University of Gondar, Ethiopia, and the Ethiopian National Research Ethics Review Committee and Ethiopian Food and Drug Administration, Addis Ababa, and was registered with ClinicalTrials.gov (NCT02880241). Study conduct conformed to the national regulatory requirements of Ethiopia and the Declaration of Helsinki.P. vivax and three P. falciparum cohorts of 17 patients each . MMV390048 was supplied as 20-mg tablets . The first P. vivax and P. falciparum cohorts were to receive a single 120-mg oral dose of MMV390048 in the fasted state, with de-escalating dosing of subsequent cohorts determined by the results obtained. Four patients were to be enrolled into the P. vivax arm and followed until day 14 before enrolment into the P. falciparum arm was to begin, dependent on an acceptable review by the safety review team.Planned enrollment was for three P. vivax or P. falciparum monoinfection , fever or history of fever within the previous 48 hours for P. vivax and 24 hours for P. falciparum, and no signs or symptoms of severe or complicated malaria. Patients were admitted and received a single oral dose of 120-mg MMV390048 on day 0 and remained as inpatients until day 3 and two consecutive negative parasite assessments. Outpatient follow-up visits were made on days 7, 10/11, 14, 17/18, 21, 24/25, and 28. Rescue therapy for P. vivax was chloroquine. At the time of the study start, primaquine was not the standard of care in Ethiopia for patients living in malaria endemic regions. However, primaquine radical cure was administered to six of eight patients at the investigator\u2019s discretion after a negative glucose-6-phosphate dehydrogenase test.Eligible patients were adults aged 18\u201355 years, weighing 40\u201390 kg with microscopically confirmed Pf_ and Pv_18S rRNA quantitative polymerase chain reaction (qPCR) was consistent with published protocols.,Giemsa-stained thick and thin blood films for parasite identification and enumeration were prepared using standard methods.P. vivax malaria was unadjusted ACPR at day 14, defined as complete clearance of microscopically detected parasitemia without previous treatment failure. This enabled a rapid readout of clinical efficacy  and safety for dose adjustment of subsequent cohorts and progression to enrolment of the P. falciparum arm. Secondary efficacy outcomes for P. vivax were ACPR and recurrence rate at day 28. Safety outcomes included the frequency of adverse events up to day 35, and signs and symptoms of malaria up to day 28. Parasite clearance kinetics and MMV390048 pharmacokinetics were also planned analyses. Summary statistics were prepared for efficacy and safety data; no inferential statistical analysis was performed for this abbreviated dataset.The primary outcome for 2. All were infected with P. vivax malaria with a mean (SD) pre-dose asexual parasite count of 6,406 (6767) parasites/\u03bcL.The eight enrolled patients were males, self-defined as black, mean age 24.5 years (range 20\u201350 years), with a mean (SD) body mass index of 18.1 (1.2) kg/mP. vivax infection was reported in two patients (days 20 and day 28), both of whom received primaquine. A new P. falciparum infection was detected on day 22 in one patient (no primaquine). Thus, ACPR at day 28 was 62.8% (5/8) in the modified intention-to-treat analysis, and 71.4% (5/7) when the patient with the new P. falciparum infection was excluded.The primary endpoint of ACPR at day 14 was 100% (8/8). Asexual parasites were cleared by 24 hours postdose in four patients, by 48 hours in two patients, and by 66 hours in the remaining two patients . All patP. vivax recurrence on days 20 and 28.Gametocytes were detected in all patients at baseline and were cleared by 24 hours postdose in four patients, by 30 hours in two patients, and by 78 hours in two patients . GametocUsing qPCR, parasite clearance was achieved between 20 and 161 hours , with thCmax (peak plasma concentration) was comparable with studies in healthy volunteers, t1/2  and AUCinf (area under the concentration\u2013time curve from 0 hour to infinity) were lower than previously reported  and spontaneously resolved by day 6 and was concurrent with an upper respiratory tract infection that started on day 3. One patient had sinus tachycardia on day 1 that resolved the same day and was considered related to malaria infection. This patient also had decreased hemoglobin on day 2  that had resolved by day 6 and was considered related to drug treatment and malaria infection.There were three adverse events of special interest occurring in two patients. One patient had neutropenia, considered possibly drug related. This event started on day 2 . MMV390048 drug exposures were lower than expected based on previous findings in healthy volunteers. There were no safety or tolerability concerns with MMV390048 administration.In summary, a single oral dose of 120-mg MMV390048 rapidly cleared asexual parasites and gametocytes in eight male patients with"}
+{"text": "In 2020, Dihydroartemisinin-Piperaquine (DHAP) was adopted as a second-line antimalarial for treatment of uncomplicated malaria in Ghana following a review of the country\u2019s antimalarial medicines policy. Available data obtained in 2007 had shown PCR-uncorrected therapeutic efficacy of 93.3% using a 28-day follow-up schedule. In 2020, the standard 42-day follow-up schedule for DHAP was used to estimate efficacy levels among febrile children aged 6 months to 9 years in three malaria sentinel sites representing the three main ecological zones of the country- savannah, forest, and coastal. PCR genotyping distinguished between recrudescence and re-infection using merozoite surface protein 2 (MSP2)-specific primers for FC27 and 3D7 strains. Per protocol analyses showed day 28 efficacy of 100% in all three sentinel sites with day 42 PCR-corrected efficacy ranging between 90.3% (95% CI: 80.1 \u2013 96.4%) in the savannah zone and 100% in the forest and coastal zones, yielding a national average of 97.0% (95% CI: 93.4 \u2013 98.8). No day 3 parasitemia was observed in all three sites. Prevalence of measured fever (axillary temperature \u2265 37.5\u00b0C) declined from 50.0 - 98.8% on day 0 to 7.1-11.5% on day 1 whilst parasitemia declined from 100% on day 0 to 1.2 - 2.3% on day 1. Mean haemoglobin levels on days 28 and 42 were significantly higher than pre-treatment levels in all three sites. We conclude that DHAP is highly efficacious in the treatment of uncomplicated malaria in Ghana. This data will serve as baseline for subsequent DHAP efficacy studies in the country. In 2021Plasmodium falciparum malaria  included Dihydroartemisinin-Piperaquine (DHAP) as an artemisinin-based combination therapy (ACT) option for the treatment of uncomplicated  malaria . This de malaria . DHAP wa malaria . The lon malaria .In Ghana, DHAP and AL were considered alternate ACTs for patients unable to tolerate ASAQ three years after the introduction of ASAQ as first-line drug for the treatment of uncomplicated malaria . At that22.1The study was conducted in the War Memorial Hospital (WMH) in Navrongo (representing the savannah zone); Hohoe Municipal Hospital (HMH) in Hohoe (representing the forest zone); and Ewim polyclinic (EWP) in Cape-Coast  Figure\u00a012.2The study was a one-arm prospective evaluation of the clinical, parasitological and hematological responses of children treated with DHAP in three of Ghana\u2019s ten sentinel sites for monitoring therapeutic efficacy of first-line and second-line antimalarials.2.3Plasmodium falciparum infection and a parasite density ranging between 1,000 and 250,000 per \u00b5L. Children with severe malaria or whose parents refused to give consent were excluded from the study. As per the WHO protocol on methods for surveillance of antimalarial drug efficacy, a prior intake of an antimalarial was not an exclusion criterion  based on their weight in kilograms as per the manufacturer\u2019s instructions ; drug administration ; parasitological examination ; and hemoglobin level assessment  using an automated hematology analyser (Mindray BC-2800\u2122).All malaria blood slides for parasitological examination were examined by two microscopists. Slides with discordant readings were re-examined by a third senior microscopist. Discordant readings were related to presence/absence of asexual/sexual parasites, species diagnosis, and day 0 parasite density within range of inclusion criterion . Parasite recrudescence was distinguished from re-infection by PCR genotyping using merozoite surface protein 2 (MSP2)-specific primers for FC 27 and 3D7 strains. List of specific sequence of primers used have been published elsewhere . Samples2.4\u00ae template for therapeutic efficacy tests. The primary study outcomes were day 3 parasitemia and days 28 and 42 PCR-uncorrected and PCR-corrected efficacy outcomes. Day 3 parasitemia was evaluated as the proportion of patients seen on day 3 with parasitemia. Per protocol and Kaplan Meier survival analyses were used to describe the patterns of PCR-uncorrected and PCR-corrected treatment outcomes on days 28 and 42 post-treatment. Treatment outcomes analyzed were defined as early treatment failure (ETF), late parasitological failure (LPF), late clinical failure (LCF), and adequate clinical and parasitological response (ACPR) as per WHO protocol  of the Noguchi Memorial Institute for Medical Research, University of Ghana (FWA 00001824) approved this study (NMIMR-IRB CPN 032/05-06a amend. 2020). Written informed consent was obtained from each parent/guardian prior to commencement of the study. Each parent/guardian was presented with details of the study: objectives, methods, anticipated risks and benefits. They were also informed of their right to withdraw their children from the study at anytime during the study period without penalty.33.1A total of 226 of the 326 children screened met the inclusion criteria and were enrolled into the study in the three sites Figure\u00a023.2No day 3 parasitemia was detected in all three sites. Per protocol analyses on day 28 showed no ETF, LCF, and LPF yielding a day-28 DHAP cure rate of 100% in all three sites. Extending the follow-up period to the standard 42-day period showed PCR-uncorrected and PCR-corrected cure rates of 100% for HMH; PCR-uncorrected and PCR-corrected cure rates of 96.4% (95% CI: 86.4-99.4) and 100%, respectively, for EWP; PCR-uncorrected and PCR-corrected cure rates of 71.8% (95% CI: 60.3-81.1) and 90.3% (95% CI: 79.5-96.0), respectively, in WMH Table\u00a03.Kaplan-Meier survival analyses showed a PCR-uncorrected cumulative treatment success incidence of 1.000 for all three sites between Day 0 and Day 28 .3.3The prevalence of measured fever (axillary temperature \u2265 37.5\u00b0C) significantly decreased from Day 0 to Day 1 in all three sites: 87.4% (95% CI: 78.1-93.3) to 11.5% (95% CI: 6.0-20.6) in HMH; 50.0% (95% CI: 36.5-63.5) to 7.1% (95% CI: 2.3-18.1) in EWP; and 98.8% (95% CI: 93.1-99.9) to 7.3% (95% CI: 3.0-15.8) in WMH. On Day 2, no child in HMH and WMH had axillary temperature \u2265 37.5\u00b0C Figure\u00a04Parasite prevalence on Day 2 ranged between 1.2% (95% CI: 0.1-7.5) in WMH and 2.3% (95% CI: 0.4-8.8) in HMH. No child was parasitemic on Day 3 and Day 7  but significantly decreased to 0% on days 1 and 2 .Vomiting was the main adverse event reported. Prevalence of vomiting on day 0 was significantly highest in HMH compared with EWP and WMH  studied over the years. In 2005, ASAQ was studied in EWP and WMH with PCR uncorrected cure rates of 96.1% (95% CI: 88.0 \u2013 99.0) in EWP and 98.7% (95% CI: 92.1 \u2013 99.9) in WMH . In 2010PCR-corrected cure rates for all three sites on day 42 were over 90% (failure rates below WHO\u2019s threshold of 10% for partner drug resistance) , and theAs with other ACTs, DHAP achieved rapid parasite and fever clearance in all three sites. Parasite prevalence declined from 100% on day 0 to 1.2-2.3% on day 2 whilst prevalence of measured fever declined from 50.0-98.8% on day 0 to 7.1-11.5% on day 1. Also, mean hemoglobin levels significantly increased in all three sites after treatment with DHAP. These observations compare well with findings from previous Ghanaian studies  and stud5Findings of this study suggest that DHAP is highly efficacious in Ghana achieving over 90% cure rates in the treatment of uncomplicated malaria, and has the advantage of a longer prophylactic effect over new infections compared with ASAQ and AL. There is also no evidence of artemisinin partial resistance as per WHO definition . Subsequent DHAP efficacy studies, using current data as baseline, are critical in monitoring the usefulness of DHAP as second line ACT for uncomplicated malaria in Ghana.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Institutional Review Board, Noguchi Memorial Institute for Medical Research, University of Ghana. Written informed consent to participate in this study was provided by the participants\u2019 legal guardian/next of kin.BA, PB, NP, AA, ND-Q, NQ, EA, FO-A, KM and KK participated in the design and supervision of the study. ND-Q, SM and NQ performed PCR genotyping to distinguish re-infections from recrudescence. BA and EA performed the statistical analyses. BA drafted the manuscript. and shared with authors. All authors contributed to the article and approved the submitted version."}
+{"text": "We read with interest the comment by Kremsner and Krishna on our paper published on this Journal. The comment correctly points out that we documented an adequate clinical and parasitological response (ACPR) of 44% with quinine plus clindamycin compared with 97% on artemether-lumefantrine assessed on day 28 after starting treatment. We recognize the long-standing research interest by Kremsner and Krishna on the role of clindamycin-based combinations in the treatment of falciparum malaria, demonstrated by their publications. In this comment, Kremsner and Krishna are concerned with the level of ACPR on the quinine plus clindamycin arm in our study. They cite a few of their studies conducted in Gabon and Brazil that evaluated the efficacy of a 3-day course of 12-hourly quinine plus clindamycin treatment for participants with either uncomplicated or severe malaria. In the cited studies, administration of a relatively higher dose of quinine (15\u00a0mg/kg bd) combined with a relatively lower dose of clindamycin (7\u00a0mg/kg bd) was consistently associated with cure rates of between 88 and 100% (excluding reinfections). They correctly observe that 54% of children treated with quinine plus clindamycin in our study were still parasitaemic on day 3. However, they confirm that the mean parasite clearance with quinine plus clindamycin was slow (48 to 65\u00a0h) even in their previous studies and they commonly observed that up to 50% of participants were still parasitaemic on day 3 but \u201cwithout the need for rescue treatment\u201d. In conclusion, Kremsner and Krishna highlight the continuing need to evaluate non-artemisinin-based combinations for the treatment of uncomplicated malaria using appropriate study designs and correct dosages to avoid exposing participants to undertreatment and possible misinterpretation of the findings.We do not agree with the statement by Kremsner and Krishna that our study was probably not well-designed, which they have not elaborated. We followed the standard methods for the design of randomized controlled parallel-group clinical trials for assessing the treatment efficacy of anti-malarials \u20136. SpeciOverall, all the studies cited by Kremsner and Krishna had methodological flaws, had used a 3-day 12\u00a0hourly regimen of quinine plus clindamycin with comparable doses, long mean parasite clearance times but higher cure rates. It is important to note that there were probably spatio-temporal differences in parasite resistance patterns between Gabon and western Kenya. Only one of the cited studies was closely comparable to our study because they used a well-designed study to compare the efficacy of quinine plus clindamycin to an artemisinin-based combination therapy (artesunate plus clindamycin) in the treatment of African children with uncomplicated falciparum malaria . In conc"}
+{"text": "This exploratory study investigated four repurposed anti-infective drug regimens in outpatients with COVID-19.This phase 2, single centre, randomised, open-label, clinical trial was conducted in South Africa between 3rd September 2020 and 23rd August 2021. Symptomatic outpatients aged 18\u201365 years, with RT-PCR confirmed SARS-CoV-2 infection were computer randomised (1:1:1:1:1) to standard-of-care (SOC) with paracetamol, or SOC plus artesunate-amodiaquine (ASAQ), pyronaridine-artesunate (PA), favipiravir plus nitazoxanide (FPV\u00a0+\u00a0NTZ), or sofosbuvir-daclatasvir (SOF-DCV). The primary endpoint was the incidence of viral clearance, i.e., the proportion of patients with a negative SARS-CoV-2 RT-PCR on day 7, compared to SOC using a log-binomial model in the modified intention-to-treat (mITT) population.The mITT population included 186 patients: mean age (SD) 34.9 (10.3) years, body weight 78.2 (17.1) kg. Day 7 SARS-CoV-2 clearance rates  were: SOC 34.2% (13/38), ASAQ 38.5% , PA 30.3% , FPV\u00a0+\u00a0NTZ 27.0%  and SOF-DCV 23.5% . Three lower respiratory tract infections occurred ; two required hospitalisation . There were no deaths. Adverse events occurred in 55.3% (105/190) of patients, including one serious adverse event .There was no statistical difference in viral clearance for any regimen compared to SOC. All treatments were well tolerated.10.13039/501100004167Medicines for Malaria Venture, with funding from the UK Foreign, Commonwealth and Development Office, within the Covid-19 Therapeutics Accelerator in partnership with 10.13039/100004440Wellcome, the 10.13039/100000865Bill and Melinda Gates Foundation, and Mastercard. ClinicalTrials.gov, with 88 studies posted before 3rd September 2020, though none had reported results at that time. Notably, since this study was conducted, several drugs have shown efficacy in high-risk non-hospitalised patients with COVID-19 . However, there is limited evidence regarding the efficacy of these agents against more recent SARS-CoV-2 variants, and there are still no approved treatments for non-hospitalised COVID-19 patients who are not at high risk.Before this study was conducted there was little published evidence for therapeutic interventions in outpatients with COVID-19. A PubMed search (3rd September 2020) was conducted using the following search terms: COVID-19 OR severe acute respiratory syndrome coronavirus 2 OR SARS-CoV-2 AND treatment AND outpatient filtered for \u2018clinical trial\u2019. Of the six results returned, only two reported therapeutic interventions: a study conducted in the US and Canada found that hydroxychloroquine did not substantially reduce symptom severity in outpatients with early, mild COVID-19; in contrast a study conducted in Iran found that both hydroxychloroquine and febuxostat improved symptoms of fever, cough, and tachypnoea. The weight of evidence has since shown hydroxychloroquine to be ineffective in COVID-19. The same search terms were used for This exploratory study of outpatient treatment of COVID-19 assessed the antiviral efficacy and safety of the artemisinin-based antimalarial drugs artesunate-amodiaquine and pyronaridine-artesunate, as well the antiviral favipiravir and antiparasitic nitazoxanide in combination. The study also contributes further evidence regarding sofosbuvir-daclatasvir in outpatients with COVID-19. All investigational treatments were evaluated in comparison to standard-of-care (SOC) with paracetamol. Note that the study included the SARS-CoV-2 Alpha (B.1.1.7), Beta (B.1.351), and Delta (B.1.617.2) variants, identified by genome sequencing, but did not evaluate antiviral efficacy against the Omicron (B.1.1.529) variant.The antiviral efficacy of the four repurposed anti-infective drugs assessed was not improved over SOC in outpatients with COVID-19, but due to a lack of power the study could not rule out important clinical differences in either direction (benefit or harm). All four drug regimens were well tolerated. Future studies should consider using larger sample sizes, different doses, different populations , or alternative endpoints.Caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the COVID-19 pandemic brought significant mortality, morbidity, and economic losses globally.7,in\u00a0vitro screening effort had identified numerous candidates for repurposing against SARS-COV-2.Drug repurposing is an established route to accelerate the development of new treatments.,This exploratory study investigated the antiviral efficacy of four repurposed drug regimens versus standard-of-care (SOC) with paracetamol in outpatients with COVID-19. These were the antimalarial drugs artesunate-amodiaquine (ASAQ) and pyronaridine-artesunate (PA)in\u00a0vitro efficacy data and pharmacokinetic simulations to predict lung tissue drug concentrations.in\u00a0vitro efficacy,The choice of investigational regimens was finalised in May 2020. ASAQ, FPV, and NTZ were selected based on The investigational regimens were also considered for their known safety profiles in humans, immediate availability, and ease of delivery to low-resource settings. This study aimed to evaluate these promising COVID-19 outpatient treatments by demonstrating an increase in the proportion of patients with early viral clearance (day 7) versus SOC.ClinicalTrials.gov with the identifier NCT04532931.This exploratory phase 2, single centre, randomised, open-label, clinical trial was conducted in an outpatient setting in Johannesburg, South Africa between 3rd September 2020 and 23rd August 2021 see . This clAll patients provided signed informed consent. The study was conducted according to Good Clinical Practice, the Belmont Report, the Declaration of Helsinki, and South African law. The protocol was approved by the South African Health Products Regulatory Agency and the Human Research Ethics Committee , University of the Witwatersrand (ref: 200602B). Gauteng Provincial Department of Health provided the approval for the study to recruit from public clinics (ref: GP202008_202). An independent data monitoring committee was convened to monitor safety and efficacy.2) \u226595%, respiratory rate \u226424 breaths/minute, heart rate <120 beats/minute, and a normal mental state. Key exclusion criteria were pregnancy or lactation, QTc prolongation, or serum potassium <3.5\u00a0mmol/L. Patients >65 years of age were excluded to meet the eligibility criteria for mild COVID-19 disease in accordance with South African national guidelines.Eligible patients were male or female outpatients, aged \u226518 to \u226465 years, body weight \u226545\u00a0kg, with a positive SARS-CoV-2 RT-PCR test and symptoms starting \u226496\u00a0h prior to randomisation , with oxygen saturation  using a centralised automated randomisation system to one of five arms: SOC (paracetamol), or SOC plus one of ASAQ, PA, FPV\u00a0+\u00a0NTZ, or SOF-DCV. Given the complexity of maintaining blinding over the different dosing regimens and durations for the five arms, and the urgent need to identify COVID-19 treatments, all drugs were administered open label.Drug treatment was started on day 1: paracetamol 1000\u00a0mg 6-hourly as needed; ASAQ  200/540\u00a0mg once daily for 3 days; PA , once daily for 3 days dosed by body weight, 540/180\u00a0mg for 45 to <65\u00a0kg, and 720/240 for \u226565\u00a0kg; FPV , 1600\u00a0mg 12-hourly for 1 day, then 600\u00a0mg 12-hourly for 6 days; NTZ  1000\u00a0mg 12-hourly for 7 days with food; SOF-DCV , 400/60\u00a0mg once daily for 7 days. All treatments were given orally. The first dose was supervised, except for SOC which was taken as needed within the daily limit.Enrolment visits required an in-person consultation and physical examination. To limit the risk of SARS-CoV-2 transmission, follow-up and interim study visits were conducted via telemedicine, telephone, or text/direct messaging. Patients self-quarantined and were transported to the study site for sample collection or visited at home in line with national infection control guidelines. All patients received counselling on infection control.At screening (day 0), patient eligibility was assessed, demographic characteristics noted, and a blood sample taken for serum potassium. A mid-nasal swab and saliva sample were collected for RT-PCR detection and quantification of SARS-CoV-2, and viral culture. Post-screening patient assessments are shown in To estimate adherence, drug containers were evaluated at day 28. Blood samples for pharmacokinetic assessments were collected on day 7 for all experimental treatment arms, plus day 3 for the ASAQ and PA arms. Drug plasma or blood levels were determined at a central site  using validated protocols .Mid-nasal swab samples were collected on days 3, 7, 10, 14, and 28 for standardised qualitative and quantitative RT-PCR assays to confirm SARS-CoV-2 infection and investigate the change in viral RNA load . Serology was assessed at baseline , and repeated at day 28. Viral cultures (Vero E6 cell line) were performed for positive RT-PCR samples using published methods.2 and completed the FLU-PRO\u00ae Plus questionnaire and FLU-PRO\u00ae Plus Global Additional Diary Items.,Patients reported daily vital signs and SpO2 values <93%, or respiratory symptoms (chest/respiratory FLU-PRO domain score >0); protocol-defined lower respiratory tract infection (LRTI) incidence (resting SpO2 <93% at two readings 2\u00a0h apart plus subjective dyspnoea and/or cough); mortality incidence.The primary efficacy outcome was the incidence of SARS-CoV-2 clearance, defined as the proportion of patients with a mid-nasal swab negative for SARS-CoV-2 on qualitative RT-PCR on day 7. Secondary efficacy outcomes were: viral clearance at day 7 assessed by viral culture; viral clearance at days 3, 10, 14, 21, and 28 assessed by RT-PCR; time to SARS-CoV-2 clearance; estimated SARS-CoV-2 viral load; hospitalisation incidence to day 28, time to first hospitalisation, and the number of days hospitalised; disease severity as measured by the WHO Ordinal Scale for Clinical Improvement; FLU-PRO\u00ae Plus questionnaire responses; time to first zero WHO Ordinal Scale score; the proportion of days with fever, SpOSafety outcomes were the incidence and severity of adverse events, and changes in vital signs. Exploratory outcomes included SARS-CoV-2 seroconversion and the drug exposure\u2013response relationship for the primary efficacy outcome. See Based on a viral clearance rate of 20% by day 7 in the control arm, a sample size of 50 patients per arm  provided \u226580% power to detect an increase in viral clearance to 50% for the investigational arms, assuming a two-sided 5% type 1 error rate and 20% loss to follow-up. The efficacy assumptions were based on published data of 65% viral clearance at day 7 with FPV versus 20% for lopinavir/ritonavir.The safety population included patients who received at least one dose of randomised treatment. The modified intention-to-treat (mITT) population included patients with a confirmed SARS-CoV-2 infection who received at least one treatment dose. The as-treated population was defined as the patients who completed treatment and had viral load results for day 7 and were analysed for a sensitivity analysis of the primary outcome by what treatment they received. The pharmacokinetic population included patients who received at least one treatment dose with at least one pharmacokinetic result  and corresponding pulation . All staThe study was sponsored by Shin Poong Pharm. Co. Ltd. The study was funded by Medicines for Malaria Venture with grants obtained from UK Aid from the UK Foreign, Commonwealth and Development Office under the provision of the COVID-19 Therapeutics Accelerator in partnership with Wellcome, the Bill and Melinda Gates Foundation, and Mastercard. Additional funding was provided as follows: Shin Poong Pharm. Co. Ltd. funded the Data Safety Monitoring Board for this study; Medicines for Malaria Venture funded Naomi Richardson for writing, editorial, and graphic support services, and Julia Flynn for project management. Shin Poong Pharm. Co. Ltd. and Medicines for Malaria Venture were involved in development and approval of the protocol. Medicines for Malaria Venture provided trial, data management, and monitoring staff, conducted the statistical analysis, and developed the clinical trial report.2   . Sensiti=\u00a00.049) .Table\u00a02IP\u00a0=\u00a00.2879)  . Viral c\u00a00.2879) .By day 28, 69.7% (129/185) of evaluable patients had viral clearance assessed by RT-PCR. The proportion of patients with viral clearance at days 3, 10, 14, 21, and 28 was not improved versus SOC for any experimental treatment .P\u00a0=\u00a00.923) (P\u00a0=\u00a00.553)    for SOC, and was similar across the treatment arms (=\u00a00.923) a. Using =\u00a00.553) b. Subgronk test) a or highnk test) b.Fig.\u00a02T10 copies/mL)   and declpies/mL) , and in pies/mL) .Fig.\u00a04ChThere were three cases of protocol-defined LRTI , two of which required hospitalisation. One 40-year-old female (PA) was hospitalised and received supplemental oxygen for LRTI and respiratory distress (days 6\u201311); and one 43-year-old female (SOF-DCV) with COVID-19 pneumonia and respiratory distress (days 5\u201310) was categorised as hospitalised but received supplemental oxygen at home as a hospital bed was not available. All three LRTI cases resolved.Based on the WHO Ordinal Scale for Clinical Improvement, there were no statistically significant differences between SOC and the investigational arms in disease severity at days 7, 14, 21, or 28 , or time2 values <93%, or respiratory symptoms, except for the number of days with respiratory symptoms with ASAQ  versus SOC   . Mean to=\u00a00.026) .Mean adherence was estimated in the safety population as ASAQ 100% (n\u00a0=\u00a039), PA 100% (n\u00a0=\u00a038), FPV 97.3% (n\u00a0=\u00a038), NTZ 96.4% (n\u00a0=\u00a038), and SOF-DCV 94.8% (n\u00a0=\u00a036). Pharmacokinetic data were sparse, precluding the characterization of the full pharmacokinetic profile. Drug plasma or blood concentrations showed high inter-patient variability , with noThere were 238 adverse events of any cause reported in 55.3% (105/190) of patients, with an incidence by treatment arm of SOC 35.9% (14/39), ASAQ 46.2% (18/39), PA 55.3% (21/38), SOF-DCV 58.3% (21/36), and FPV\u00a0+\u00a0NTZ 81.6% (31/38). Overall, the most common adverse events were nausea (12.6% [24/190]), dizziness (11.6% [22/190]), and diarrhoea (11.6% [22/190]). Gastrointestinal adverse events  were particularly frequent with FPV\u00a0+\u00a0NTZ ; chromatThe majority (97.9% [233/238]) of adverse events were grade 1 or grade 2 in severity. There were four grade 3 adverse events: diarrhoea (SOC), suicidal ideation (SOC), and two cases of respiratory distress  . None ofAdverse events considered to be drug related were reported in 0% (0/39) of patients in the SOC arm, 27.8% (10/36) for SOF-DCV, 28.2% (11/39) for ASAQ, 31.6% (12/38) for PA, and 55.3% (21/38) for FPV\u00a0+\u00a0NTZ . The mosThere were no clinically important changes in vital signs throughout the study, though there was a trend for a lower pulse rate with ASAQ versus other arms .At baseline, 29.3% (53/181) of patients were positive for SARS CoV-2 serology, probably indicating prior exposure . By day This study explored the potential of four repurposed drug regimens for the outpatient treatment of COVID-19. The study population was relatively young and at low risk of disease progression.,,Notably, there were no cases of protocol defined LRTI in the SOC arm. There was no improvement in efficacy outcomes for any experimental arm versus SOC. Adverse events reported for the investigational regimens were broadly consistent with their known safety profiles,Due to the epidemiological uncertainty of the timing and duration of waves, recruitment was slow outside the short-lived peaks of transmission, with around 93% of patients failing screening because of a negative RT-PCR test. Hence, recruitment was stopped at 192 patients, below the planned target of 250, but as the drop-out rate was lower (10%) than expected (20%), 38 patients per arm retained 80% power based on original assumptions. Nevertheless, the day 7 clearance rate in the SOC arm (34.2%) was higher than the initial hypothesis of 20%, probably because of prior exposure\u201429.3% of patients had positive baseline serology. Based on the higher-than-expected rate of viral clearance in the SOC arm, 152 patients per arm would be needed to detect a 50% efficacy rate in the experimental arms with 80% power. Thus, the study was under powered for the primary outcome. Note that sub-group analyses  were not powered to detect differences between the groups.Notably, during the period of this study, few of the planned COVID-19 clinical studies in outpatients completed. Our five-arm study was logistically challenging given the extreme pressure on the South African health system. National restrictions to reduce SARS-CoV-2 transmission required a suite of innovative measures, including remote electronic data capture, the engagement, support, and training of patients in conducting and reporting assessments, and the use of mobile clinics. The high completion rate for patients recruited to the study indicates that study procedures were well accepted.In\u00a0vivo efficacy of pyronaridine in reducing viral titres and lung pathology was shown in SARS-CoV-2 infected transgenic mice expressing human host receptor hACE2 (K-18-hACE), possibly via inhibition of an essential viral protease, PL pro.There are limited data on ASAQ or PA, or FPV and NTZ in combination for the treatment of COVID-19. ASAQ and PA were used at doses approved for the treatment of uncomplicated malaria. For FPV, two studies in COVID-19 outpatients reported a median time to viral clearance of 10 days (n\u00a0=\u00a0112) versus 8 days with placebo,For SOF-DCV, a trial in outpatients reported no effect on hospitalisations versus hydroxychloroquine, but a reduction in fatigue and dyspnoea after 1 month.,P\u00a0=\u00a00.001) in death or hospitalisation.The main limitations of this study are the use of virological outcomes and its size. Viral clearance was chosen as the primary outcome in May 2020, when there was no consensus regarding the most appropriate study design. This allowed a logistically feasible sample size for the assumed efficacy rates. Also, the use of five very different dosing regimens necessitated an open-label study design, and viral clearance was an objective measure to compare efficacy that would not be influenced by this design. However, there are limitations to using virological endpoints. As neither quantitative nor qualitative RT-PCR differentiate between infectious and replication incompetent virus, viral clearance and viral load assessed through RT-PCR may be inappropriate endpoints to evaluate efficacy.In conclusion, a higher proportion of patients receiving SOC achieved viral clearance at day 7 than expected, and none of the treatment regimens showed a virological efficacy benefit. There was generally no improvement for secondary virological endpoints and symptomatic endpoints. Study participants were young, and 29.3% were seropositive for SARS-CoV-2 at baseline, indicating prior exposure. The combination of this, a limited sample size, and virological endpoints now considered less relevant by current treatment study standards, means that due to a lack of power definitive conclusions related to efficacy cannot be made in either direction (benefit or harm). Since the predictive relationship between the magnitude and timing of viral RNA reduction and viral infectivity or clinical benefit has not been fully established, results should also be interpreted with caution. All treatments were, however, well tolerated. Future studies may consider using larger sample sizes, different drug doses, including patient populations at risk of developing severe disease, and evaluating alternative end points, to demonstrate efficacy.NC, HJ, MFC, AO, AH, NA, SD, and WDFV contributed to the conception and design of the trial. NC, CK, HJ, MFC, YD, SA-B, RM, and ACM were involved in the acquisition of data. NC, CJ, BK, SA-B, RM, AO, AH, DW, NA, SD, and WDFV were involved in data analysis and interpretation. NC, RM, ACM, and DW verified the underlying data. All authors contributed to the development of the paper, provided critical review, and approved the final version for submission. All authors had access to the primary data and are able to take responsibility for the accuracy and completeness of the results. All authors had final responsibility for the decision to submit the paper.https://www.mmv.org/about-us/contact-us) referencing this publication. Data will be available for at least five years from publication of this study.De-identified participant data are available on reasonable request and with completion of a signed data access agreement from (NC declares grants and non-financial support from Shin Poong Pharm. Co. Ltd. during the conduct of the study and grants and non-financial support from ViiV Healthcare and Gilead Sciences, grants, personal fees and non-financial support from Johnson & Johnson, personal fees from Cipla and Frontiers Biotech outside the submitted work. NA, ACM, and SD are employees of Medicines for Malaria Venture. MFC is a former employee of Medicines for Malaria Venture. HJ is a consultant for Medicines for Malaria Venture and was a consultant medical monitor for this study. CJ and BK are employees of Shin Poong Pharm. Co. Ltd. CJ has a patent pharmaceutical composition for COVID-19 treatment pending. SA-B and RM are employees of Artemida Pharma which received funding from Shin Poong Pharm. Co. Ltd. RM consulted for Shin Poong Pharm. Co Ltd. and is the Shin Poong Pharm. Co. Ltd. qualified person for pharmacovigilance, responsible for providing interpretation of the safety data of this study and other studies involving pyronaridine-artesunate. SA-B consulted for Shin Poong Pharm. Co. Ltd. during the study for the operational aspects and in particular all activities related to the Data Monitoring Committee. AO is a director of Tandem Nano Ltd., co-inventor of patents relating to drug delivery, and has received grants and consultancy from AstraZeneca and Janssen, and grants and personal fees from Merck, ViiV healthcare, and Gilead outside of the submitted work. AH received a research grant from the funders for advice on statistical design and analysis during the conduct of the study. DW is an employee of DATAMAP. WDFV reports grants from Unitaid during the conduct of the study; grants from the Bill and Melinda Gates Foundation, the South African Medical Research Council, USAID, National Institutes of Health, FIND, Children's Investment Fund Foundation, personal fees from Virology Education, grants, personal fees and non-financial support from ViiV Healthcare, personal fees and non-financial support from Gilead Healthcare, grants, personal fees and non-financial support from MSD, grants, personal fees and non-financial support from Johnson & Johnson, and personal fees from Mylan/Viatris, Adcock-Ingram, Aspen, Abbott, Roche, and Sanofi outside the submitted work. CK and YD declare no competing interests."}
+{"text": "Plasmodium falciparum infection is often accompanied by disturbances of hematological and biochemical parameters. The objective of this study was to evaluate the changes in biochemical and hematological parameters during uncomplicated malaria in patients treated with ACTs.  Artemisinin-based Combination Therapies (ACTs) are widely used in the treatment of uncomplicated malaria. Plasmodium falciparum malaria were pooled from different open-randomized trial evaluating the efficacy of Artesunate-Mefloquine (ASMQ), Artesunate-Amodiaquine (ASAQ), Artemether-Lumefantrine (AL), and Dihydro-artemisinin-Piperaquine (DHAPQ) combinations. Biochemical  and hematological (hemoglobin and platelet levels) parameters were performed at baseline (D0) and at day 7 after treatment (D7). Data were analyzed as both continuous and categorical variables with 95% confidence interval. Risks and trends were calculated using multivariate logistic random effect models.  Data from patient with uncomplicated p = 0.57). At D7, hemoglobin level was 9.63\u2009g/dl and anemia was significantly more frequent (78.29% [p = 0.002]). The mean platelet count at day 0 was 154075.5 platelets/mm3 of blood and 339328.7 platelets/mm3 at day 7. Thrombocytopenia was about 53.61% and was associated with malaria (aOR = 3.4\u2009[2.18 \u2212 5.3]p < 10\u22123). 19.58% of patients had abnormal ALT and 40.28% had abnormal AST at D0. 27.22% of patients had normal bilirubin at D0. Renal function was normal in all patients in the study. Normalization of transaminases was noted between D0 and D7. The percentage of subjects with normal bilirubin increased between D0 and D7. Renal function did not vary significantly between D0 and D7.  A total of 720 patients with completed biological data were included in the analysis . The mean age of the patients was 9.43 \u00b1 9.1 years. Male subjects represented 58.47% . The mean hemoglobin level at inclusion (D0) was 9.79\u2009g/dl and anemia (Hb < 11\u2009g/dl) was 71.43% (aOR = 1.16\u2009[0.68 \u2212 1.98] Results from this analysis showed that subjects with high parasitaemia had a greater risk of anemia and thrombocytopenia. Artemisinin combinations were well-tolerated as no major biological disturbances were noted. The effects of ACTs on hematologic and biochemical parameters were not different. Malaria is a major public health problem in developing countries. According to World Health Organization (WHO), 241 million cases of malaria are recorded each year and there are approximately 627000 deaths. More than 95% of these cases and 96% of the deaths occur in Africa and mainly in children under 5 years old  .Following WHO recommendations, Senegal, through its National Malaria Control Program (NMCP), changed its policy for the management of uncomplicated malaria cases by adopting artemisinin-based combination therapy in 2006 .Several studies have shown that artemisinin-based combination therapies are effective and well-tolerated in the treatment of uncomplicated malaria.Plasmodium falciparum malaria on certain biological markers in the context of efficacy clinical trials on artemisinin combination therapy in Senegal.As a line extension of the ongoing surveillance system and to better document the modifications of biological parameters during malaria infection, this study was assessing the effect of This study was designed within the framework of open-randomized trials evaluating the efficacy and safety of artemisinin-based combination therapies in Senegal. The combinations Artesunate-Mefloquine (ASMQ), Artesunate-Amodiaquine (ASAQ), Artemether-Lumefantrine (AL), and Dihydro-artemisinin-Piperaquine (DHAPQ) were studied during randomized open trials conducted in the health districts of Dakar  and Kaolack . In Dakar, the study was conducted in Deggo health post which is located at 20\u2009km from the capital city. In Dakar, malaria incidence is between 5% to 15% habitants. Kaolack region is located in the central part of Senegal, 200\u2009km from Dakar. The study was carried out in Keur Soc\u00e9 and Medina Baye health posts where malaria incidence is more than to 15% habitants.P. falciparum is the predominant species and transmission is mainly due to Anopheles gambiae s.l.In these areas, malaria is highly seasonal during the rainy season (July to October) with a peak of transmission from September to December. P. falciparum malaria, parasitaemia between 1000 and 100000 trophozoites/\u03bcl, and patients able to take oral medication. Patients with severe and complicated malaria, according to the WHO definition p = 0.0002) compared to subjects younger than 15 years. Results from the analysis also showed an association between gender and thrombocytopenia. Male subjects had a 10% greater risk of developing thrombocytopenia (aOR = 1.1\u2009[0.81 \u2212 1.5]p = 0.52) compared to female subjects. An association between parasitaemia and thrombocytopenia was also found. Subjects with moderate parasitaemia had twice the risk of thrombocytopenia (aOR = 2.01\u2009[1.26 \u2212 3.2]p = 0.0035) compared to subjects with low parasitaemia. Subjects with high parasitaemia had a 3-fold increased risk of thrombocytopenia (aOR = 3.4\u2009[2.18 \u2212 5.3]p < 10\u22123) compared to subjects with moderate and low parasitaemia . Depending on the treatment group, hemoglobin level decreased from D0 to D7 in AL and ASMQ groups. In contrast, in DHAPQ and ASAQ groups, an increase was noted. In DHAPQ group, the mean hemoglobin level increased from 10.03\u2009g/dl to 10.7\u2009g/dl between D0 and D7 (p = 0.001). In ASAQ arm, mean hemoglobin level was lower at D0 (8.57\u2009g/dl) compared to D7 (8.63\u2009g/dl) without significant difference (p = 0.76). In AL group, mean hemoglobin level was 10.03\u2009g/dl at D0 versus 9.7\u2009g/dl at D7 (p = 0.049). A decrease in hemoglobin level was noted between D0 and D7 in the ASMQ group from 9.63\u2009g/dl to 9.1\u2009g/dl (p = 0.0001) (A decrease in hemoglobin level was observed between D0 and D7 from 9.76\u2009g/dl to 9.63\u2009g/dl. The difference in mean between D0 and D7 was 0.14\u2009g/dl ( 0.0001) .p = 0.07). In contrast, in AL and ASMQ groups, a decrease of 0.27 \u00b1 0.08\u2009g/dl [95% CI (0.1-0.44)] and 0.54 \u00b1 0.08\u2009g/dl [95% CI (0.27-0.81)] was noted in AL and ASMQ groups, respectively (p = 0.56). Between AL (0.27\u2009g/dl) and DHAPQ (-0.062\u2009g/dl) groups and DHAPQ and ASMQ groups (0.54\u2009g/dl), the differences were significative (p < 10\u22123) ] and \u22120.65 \u00b1 0.14\u2009g/dl [95% CI (-0.94--0.37)], respectively, in ASAQ and DHAPQ group ( < 10\u22123) .p value = 0.002). Anemia at D7 in ASAQ and ASMQ groups was 95.9% and 86.5%. In DHAPQ and AL groups, it was 58.6% and 77.7% . At baseline, more than 50% of patients in each group had thrombocytopenia. However, at D7, a significant decrease in thrombocytopenia was observed in different treatment groups. At D7, thrombocytopenia was more frequent in ASAQ and ASMQ groups with 10.2% and 9.4%, respectively  compared to D0 (80.4%). Patients with normal ALT were higher in AL group (88.1%) and DHAPQ group (81.7%). The number of patients with normal AST increased significantly between D0 and D7 in all treatment groups .p value = 1), between ASAQ (9.3\u2009IU/l) and AL (p = 1) and between ASAQ and ASMQ . ALT levels decreased by 3.15\u2009IU/l in the DHAPQ group. When comparing DHAPQ to the other groups, the difference was significant (p < 0.05). The mean AST decreased by 12.8\u2009IU/l in ASAQ group versus 14.4\u2009IU/l in AL group . The same trends were observed between ASAQ and ASMQ groups (p = 0.29), between ASAQ and DHAPQ (p = 0.07), and between AL and DHAPQ (p = 0.6). The differences were significative between ASMQ and AL and between ASMQ and DHAPQ (p < 10\u22123) (No significant difference was observed between ASMQ (11.7\u2009IU/l) and AL (7.6\u2009IU/l) ( < 10\u22123) .Mean bilirubin decreased significantly between D0 and D7 in all treatment groups except DHAPQ group where an increase was noted but not significative .The percentage of subjects with normal bilirubin increased between D0 and D7 in patients in the DHAPQ, AL, and ASMQ groups. In contrast, in the ASAQ group, any subject had normal bilirubin levels between D0 and D7 .p = 0.0001). In the AL, DHAPQ, and ASMQ groups, creatinine levels decreased by 0.31\u2009mg/l, 0.89\u2009mg/l, and 0.41\u2009mg/l, respectively, between D0 and D7 (p < 10\u22123)  .p < 10\u22123).Pairwise analysis between the different groups showed significant differences  for the management of uncomplicated malaria. These ACTs have proven to be effective and well tolerated. In Senegal, ACTs have been scaled up since 2006. The scale-up of ACTs must be accompanied by the establishment of a pharmacovigilance system to better document possible side effects of ACTs. This study was conducted to evaluate the changes in biochemical and hematological parameters during malaria infection in patients treated with ACTs.p = 0.57). The more the parasitaemia is high, the more the hemoglobin level is low.The results from this analysis have demonstrated an association between parasitaemia and anemia. Subjects with moderate parasitaemia had a 10% greater risk of anemia  .p < 0.001) p < 10 \u2212 3) compared to those with moderate and low parasitaemia. Same trends concerning the association between malaria and thrombocytopenia were previously described by in sub-Saharan Africa [An association between parasitaemia and thrombocytopenia was noted in our study. Subjects with moderate parasitaemia had a 2-fold increased risk of thrombocytopenia  [Zwang et al., when comparing the changes in hematologic parameters occurring in patients with uncomplicated llow up) .Concerning the liver enzymes, a normalization of transaminases was observed between D0 and D7 in all treatment groups The number of patients with normal transaminase was higher at day 7 (86.4% for ALT and 79.2% for AST) compared to the enrollment (28% for ALT and 59.7% for AST). This has been previously described by Sylla et al. in Senegal .Plasmodium falciparum malaria [An elevation of liver enzymes was noted in Nigeria in patients with uncomplicated  malaria . This ca malaria , 27.Regarding the mean of creatinine, no significant variation was observed between day 0 and day 7 in the different treatment arms. However, a significant decrease of bilirubin level was noted at day 7 in the three treatment arms. These results are in line with what were previously noted in Senegal \u201313.Results from this analysis confirm the complexity of explaining hematological and biochemical changes occurring in acute malaria. The results noted patients with high parasitaemia had a greater risk of anemia and thrombocytopenia. No significant difference was noted between the different treatment groups at day 7 in term of ALAT and ASAT level as well as bilirubin and creatinine concentration. However, this analysis cannot distinguish between the changes occurring while patients recover from uncomplicated malaria and those from drug adverse reactions. Analyzing these results is not simple and will require confirmation on extended databases. For this reason, it will be necessary to obtain and collect information from as many studies. Finally, establishing reference laboratory ranges that are locally relevant should be a priority."}
+{"text": "Seasonal malaria chemoprevention (SMC) consists of the intermittent administration of a 3\u00a0day course of anti-malarial medications during the months of highest malaria risk in the Sahel region, where malaria transmission is highly seasonal. SMC is an effective intervention to reduce episodes of uncomplicated and severe malaria in children. However, morbidity cannot be lowered without adherence to medications. The objective of this study was to examine SMC medication adherence and to identify the attitudes and practices of caregivers during the 2020 SMC campaign in the Dosso region..This study was conducted based on data from independent monitoring using random cluster sampling. Adherence levels and the attitudes and practices of caregivers were evaluated using data from caregivers\u2019 self-reports and analysed according to Bernard Vrijens\u2019 taxonomyAt the initiation of treatment phase, 99% of children (N\u2009=\u20092296) received their first administration of medication, with 90% of caregivers (N\u2009=\u20091436) knowing that the medications help prevent malaria. However, only 56% of caregivers (N\u2009=\u20091856) reported that treatment initiation took place under direct observation by the distributor. At the implementation of treatment phase, 90% of children (N\u2009=\u20092132) took their medication on the second day and 84% (N\u2009=\u20091068) took it the third day. \u201cForgetting,\u201d \u201cnot having time,\u201d and \u201cthe mother\u2019s absence\u201d were the main reasons caregivers gave to explain discontinuation of the 3\u00a0day course of medication.This simple, low-cost survey demonstrated that coverage of SMC and adherence by caregivers to completing the full 3\u00a0day medication course was high. The survey also showed that knowledge, attitudes, and practices of some caregivers regarding adherence to medications during the SMC campaign could be improved. Expanding distributors\u2019 training, developing and providing them with tools for interpersonal communication, and strengthening supervision could lead to even higher adherence. Malaria is a major public health challenge. With an estimated 241 million new cases and 627,000 deaths in 2020, this disease is a major cause of morbidity and mortality [In 2012, the WHO recommended seasonal malaria chemoprevention (SMC)  for the SMC is recognized by the WHO as an effective intervention to reduce episodes of uncomplicated and severe malaria in children by approximately 75%  and therAdherence to a medication regimen is the process by which patients take their medications as prescribed by a health care provider. Some investigators have refined the definition of adherence to include data on taking the prescribed number of pills each day and taking the pills within a prescribed time frame . Poor adDespite the large volume of literature on adherence to medications, a \u201cgold standard\u201d has not been clearly established to measure adherence. The available methods can be categorized as direct and indirect methods. Examples of direct methods of measuring adherence are administering the medications under direct observation, measuring concentrations of a drug or its metabolite in the blood or urine, and detecting or measuring a biological marker for the drug in the blood. Direct approaches are expensive, complicated, and likely to be biased by the patient. However, for some medications, measuring these levels is a good, commonly used method to evaluate adherence. The indirect methods for measuring adherence include asking the patient about the ease with which s/he takes the prescribed medications, evaluation of clinical response, tablet counting, collecting questionnaires from patients, using electronic drug monitors, measuring physiological markers, having the patient keep a medication diary, and evaluation of the children\u2019s adherence by asking for the help of a caregiver , 15. Thi\u201d and is the process of monitoring and supporting patients' adherence to medications by health care systems, providers, patients, and their social networks. The third element is \u201cadherence-related sciences\u201d. This element includes the disciplines that seek to understand the causes or consequences of differences between the prescribed and actual exposures to medications.In 2022, the National Malaria Control Programme and the U.S. President\u2019s Malaria Initiative (PMI) through the PMI Impact Malaria project in Niger decided to take a second look at the data from the independent monitoring of 2020 SMC campaign using Bernard Vrijens\u2019 taxonomy . AccordiThe hypothesis in this study was that adherence to SMC medication might be a problem in Niger. During campaign supervision missions in Niger, discrepancies were observed between the recommended/expected behaviours and those applied by the distributors  and caregivers . While adherence to SMC medication in Niger has been shown to be very high , more reThis study aimed to estimate adherence to anti-malarial medication in children aged 3 to 59\u00a0months during the 2020 SMC campaign in Dosso region  with an average disease incidence fluctuating between 250 and 450 cases per 1000 inhabitants .Direction r\u00e9gionale de la sant\u00e9 publique (DRSP) receives technical and financial support from the U.S. PMI Impact Malaria project.To implement the SMC campaign in the eight health districts in the region, the Dosso Since 2016, Dosso has been conducting annual SMC campaigns. The implementation has increased from one health district in 2016 to all eight health districts of the region in 2019. The number of children aged 3\u201359\u00a0months to be reached in the intervention in 2020 was 529,380. Since 2016 when the intervention started in the Dosso region, close to 90% of the children in the intervention districts were reached with SMC. However, the incidence of malaria recorded in health centres remained high. Therefore, questions were raised about the effectiveness of SMC in the region. The caregivers\u2019 poor adherence to the full 3\u00a0day course of medications was suggested as a possible explanation for the continued high incidence of malaria.Caregivers of children aged 3\u201359\u00a0months in the Dosso region were approached and asked to complete the survey.The independent monitoring targeted two health districts (HD) during each cycle of the 2020 SMC campaign in the Dosso region, i.e., eight HD for the four cycles. In each HD, three health areas were randomly selected using a list of the district\u2019s health areas. Then, a second random draw was performed from the list of localities (villages/neighbourhoods) to select three localities to be surveyed per health area. Within a locality, a random selection process was used to select twenty family compounds  by visiting every second family compound. In each selected family compound, the caregivers of all eligible children were interviewed systematically.The surveys started on the second day of each cycle and were conducted for 3\u00a0days. This meant that if a child received the first dose on day 3 of the distribution period and researchers visited on that day, only questions about day 1 could be asked. Independent monitors visited the family compounds, identified eligible children, interviewed caregivers about the current cycle, and verified that the SMC cards were filled out correctly. During the four cycles, 72 localities (villages/neighborhoods) and 1440 family compounds were monitored in the eight districts of the Dosso region.Each health district, health area, and locality were monitored only once during the campaign, i.e., the households visited after cycle 4 were not the same households as were visited after cycle 1. Nor were the households visited on day 3 the same as the households visited on day 2 of a single cycle.The caregivers rather than the children under five were interviewed. One caregiver could respond about more than one child during a single survey. As this was programme monitoring rather than a study, no information on the age or sex of the caregiver was collected. The broad age category (3\u201311\u00a0months and 12\u201359\u00a0months) and the sex of the child was collected. A structured questionnaire was used with pre-defined answer categories. The tool was closely inspired by the survey tool used in Mali for many years. It was digitized in KoboCollect and pre-tested before going to the field.The survey monitors were independent of the SMC campaign, in that they were recruited from outside of the health sector  so that the same people who were distributing or supervising the distributors would not be evaluating their own work. The household surveys were conducted for the NMCP, with funding from PMI Impact Malaria, and the reports were published as grey literature in Niger before this secondary analysis and manuscript were conceived.initiation, for example, medications administered on the first day may not be taken under the distributor\u2019s direct observation, or the child may refuse to drink the medication or spit out/vomit up the medication.At treatment During treatment implementation, caregivers may forget to administer the medications on subsequent days or may not administer the medications according to the dosage, or the caregiver may simply decide to stop administering the medication for some other reason .The project team analysed the data using the first element of Bernard Vrijens\u2019 taxonomy . This apThe indicators analysed were adherence to the medications on the first, second and third days of the 3\u00a0day course and the evaluation of the caregivers\u2019 attitudes and practices.Percentage of target children that received the SP and AQ: number of children that received medication on day 1 of the three-day course among the total number of eligible children visitedPercentage of the SP administration observed but not administered by the drug distributor: number of children that received medication on day 1 of the three-day course under direct observation among the total number of eligible children monitored.Percentage of target children that received medication on day 2 of the three-day course: number of children that received medication on the 2nd day of the three-day course among the total number of eligible children monitoredPercentage of target children that received medication on day 3 of the three-day course: number of children that received medication on the 3rd day of the three-day course among the total number of eligible children monitored.Note that the question on direct observation was asked in two parts: (1) If the child was treated, who administered the first dose? The surveyor was prompted to select caregiver or distributor from a drop-down box. If the answer to that first question was caregiver, a second question popped up: (2) If the dose was administered by the caregiver, was the administration supervised by the community distributor?A caregiver could be a mother, father, grandmother, aunt, older sibling, or other family member or even neighbor. Regardless of their relation to the child, ninety percent of caregivers surveyed knew that the SMC medications were targeting malaria in children see Fig.   during the first cycle and remained stable at 99.0% for the other cycles. The team observed the same trends when the monitors verified that the SMC card was marked Table .Table 1PAccording to 56% (n\u2009=\u20091035) of caregivers surveyed, treatment initiation on the first day took place under direct observation of the distributors. There is an upward trend of the observed administration of the first dose between the first cycle (39%) and subsequent cycles  of children took the AQ on the second day and 84.0% (n\u2009=\u2009897) took the AQ on the third day of the three-day course (Table Only minor adverse drug reactions (ADR) were reported by the caregivers; 6.8% (346/5111) of children among those monitored. The most reported ADR were vomiting (27.7%), drowsiness (24.6%), diarrhoea (14.2%), and loss of appetite (11.0%). Less frequently reported ADR included the occurrence of itching (5.5%), fever (5.5%), abdominal pain (4.6%), and rash (4.6%).Out of a total of 1436 caregivers, only 163 (11%) interrupted their child\u2019s treatment. The reasons were: 34% forgot, 21% did not have time and 12% were absent  interrupted their child's treatment. \u201cForgetting\u201d (34%), \u201cnot having time\u201d (21%), and \u201cthe mother\u2019s absence\u201d (12%) were the main reasons caregivers gave to explain discontinuation of the SMC treatment in the Dosso region. Most people in the area studied are farmers and many of them may forget to administer the medications or may not be at home at the scheduled time to administer the medications, due to the demands of field work at that time. This observation suggests the need to develop behavioural interventions or a combination of behavioral interventions , 38.As with any observational study, there is a risk of selection and observation bias. To minimize these biases, participants surveyed during the monitoring were randomly selected. The data was collected by individuals not involved in the implementation of the SMC campaign and who were unaware of the design and issue being studied.For the interpretation of the results, certain factors must be considered. First, the adherence evaluation method of self-reporting is subject to different biases, such as memory bias and social desirability bias. To reduce memory bias, data collection took place during the campaign\u2019s implementation, i.e., no more than one to three\u00a0days after the first dose of medication. To counter social desirability bias, SMC cards and empty blister packs were also checked. Secondly, this study was conducted in the Dosso region, which mostly consists of districts located in rural areas and where the malaria burden is mixed. The results can, therefore, be generalized in regions with the same profile. However, they may be less generalizable to urban areas where caregivers have different lifestyles.This study identified the attitudes and practices that influenced adherence to anti-malarial treatment in children 3\u201359\u00a0months of age during the 2020 SMC campaign in the Dosso region of Niger. The study found that adherence levels were very high at treatment initiation and dropped off a bit on the second and third days of each three-day course. The main attitudes and practices influencing high levels of adherence would be the caregivers\u2019 acceptance of the medications and good tolerability of the SMC medications, while \u201cforgetting,\u201d \u201cnot having time,\u201d and \u201cthe mother\u2019s absence\u201d were the factors that contributed most to non-adherence. Despite the caregivers\u2019 acceptance of SMC, distributors should continue to supervise the administration of the first dose and use this face-to-face time to strengthen the caregivers\u2019 knowledge about adherence to the medications on subsequent days, potential adverse drug reactions, and what to do if they occur.This study suggests that malaria programmes should carefully weigh up the potential benefit and cost based on evidence before introducing initiatives to increase full adherence such as three-day directly observed treatment. The study findings do not suggest that there is a major problem with adherence."}
+{"text": "Cipargamin (KAE609) is a potent antimalarial in a phase II trial. Here we report efficacy, pharmacokinetics, and resistance marker analysis across a range of cipargamin doses. These were secondary endpoints from a study primarily conducted to assess the hepatic safety of cipargamin (hepatic safety data are reported elsewhere).Plasmodium falciparum malaria. Cipargamin monotherapy was given as single doses up to 150 mg or up to 50 mg once daily for 3 days, with artemether-lumefantrine as control. Key efficacy endpoints were parasite clearance time (PCT), and polymerase chain reaction (PCR)\u2013corrected and uncorrected adequate clinical and parasitological response (ACPR) at 14 and 28 days. Pharmacokinetics and molecular markers of drug resistance were also assessed.This phase II, multicenter, randomized, open-label, dose-escalation trial was conducted in sub-Saharan Africa in adults with uncomplicated Pfatp4 gene, G358S, was detected in 65% of treatment failures. Pharmacokinetic parameters were consistent with previous data, and approximately dose proportional.All single or multiple cipargamin doses \u226550 mg were associated with rapid parasite clearance, with median PCT of 8 hours versus 24 hours for artemether-lumefantrine. PCR-corrected ACPR at 14 and 28 days was >75% and 65%, respectively, for each cipargamin dose. A treatment-emerging mutation in the P. falciparum malaria, and recrudescent parasites frequently harbored a treatment-emerging mutation. Cipargamin will be further developed with a suitable combination partner.Cipargamin, at single doses of 50 to 150 mg, was associated with very rapid parasite clearance, PCR-corrected ACPR at 28 days of >65% in adults with uncomplicated ClinicalTrials.gov (NCT03334747). Plasmodium falciparum malaria.Cipargamin, at single doses of 50\u2013150 mg, was associated with rapid parasite clearance (parasite clearance time ~8 hours) and PCR-corrected adequate clinical and parasitological response at day 29 of \u226565% in adult patients with uncomplicated  Plasmodium falciparum and Plasmodium vivax. This compares favorably with artemisinin-based therapies such as artemether-lumefantrine  [P. falciparum [Cipargamin (KAE609/NITD609) is a novel spiroindolone antimalarial . In a ph4 hours) . Parasitlciparum . The eliP. falciparum [Plasmodium by inhibiting the Na+ transporting plasma membrane ATPase Plasmodium falciparum ATPase 4 (PfATP4) [PfATP4 gene have been generated in vitro under prolonged drug pressure, leading to decreased cipargamin susceptibility [Cipargamin is active against all intraerythrocytic stages of lciparum  and gamelciparum . Spiroin(PfATP4) , 9. Mutatibility , 10, 11.Transient liver function test elevations were observed in some trial participants , 12, 13,This was a multicenter, randomized, open-label, dose-escalation phase II trial, conducted in Mali, Gabon, Ghana, Uganda, and Rwanda. The protocol and all amendments were reviewed by the Independent Ethics Committee or Institutional Review Board for each center. The trial was conducted according to International Conference on Harmonisation (ICH) E6 Guidelines for Good Clinical Practice. The trial is registered with ClinicalTrials.gov (NCT03334747).P. falciparum malaria . Exclusion criteria included mixed Plasmodium infections and severe malaria according to World Health Organization (WHO) criteria [Eligible patients were adults (\u226518 years old and\u2005\u226545 kg body weight) with microscopic confirmation of acute uncomplicated criteria . Full exPatients were treated in 5 cohorts, using ascending single or multiple doses of cipargamin . It was Recruitment to cohorts was sequential. After completion of each cohort, dose escalation of patients to the next cohort was determined by decision criteria . After being contacted by the investigator, the IRT assigned a randomization number  that linked the patient to a treatment arm and specified a unique medication number for the study drug to be dispensed to the patient. The randomization number was not communicated to the investigator. Randomization to a cohort could be suspended if patients experienced safety events.Patients received close monitoring in an inpatient setting for at least the first 3 days, followed by frequent outpatient monitoring for a total of 4 weeks. Patients were required to yield 2 consecutive negative blood smears for parasites and clearance of fever to be discharged. Cipargamin patients who met protocol-specified treatment failure criteria received artemether-lumefantrine as rescue medication.Pfatp4 analysis are provided in the Blood samples were taken for parasite counts (Giemsa-stained thick and thin films) at baseline; at 2, 4, 8, 12, 24, 36, 48, 60, and 72 hours; then at days 4, 7, 10, 14, 21, and 28 after starting treatment, and at unscheduled visits. At least 200-thick film visual fields were examined. Parasite counts were made per 200 leukocytes . The sampling schedule and method for pharmacokinetic analysis are provided in Planned efficacy outcomes were as follows: PCR-corrected and uncorrected adequate clinical and parasitological response (ACPR) at 14 and 28 postdose; PCT; fever clearance time (FCT); proportions of patients with parasitemia at 12, 24, and 48 hours; parasite reduction ratio at 24 hours (PRR24); incidence of reinfection and recrudescence at 28 days; incidence of early treatment failure ; time to maximum plasma concentration (Tmax); areas under the concentration-time curve from time zero to the time of the last quantifiable concentration (AUClast), zero to infinity (AUCinf), and zero to 24 hours (AUC0-24h); and elimination half-life (T1/2).Pharmacokinetic parameters calculated were maximum plasma concentration . Artemether-lumefantrine groups from all cohorts were pooled for analysis. The ACPR was calculated for each treatment by cohort, with 95% confidence intervals (CIs) provided using the Clopper-Pearson method, as were rates of ETF, LCF and LPF, and proportions of patients with parasitemia by time point. For PCT and FCT, descriptive statistics  were presented for each treatment by cohort using the Kaplan-Meier method. Incidence rates of recrudescence and reinfection at day 29 were estimated using the Kaplan-Meier method based on the subset of full analysis set patients with clearance of initial infection before day 15. For PRR24 hours, descriptive statistics and 95% CI for the geometric mean were provided by cohort and treatment. If the asexual parasite count at hour 24 was 0, the half value of detection limit was used to calculate the ratio. The detection limit was assumed to be 20 parasites/\u00b5L. Pharmacokinetic analyses were based on the pharmacokinetic analysis set . Noncompartmental analysis was used and descriptive statistics were provided. Two-sided 90% CIs were only calculated for AUCThe trial started on 16 November 2017 and was completed on 23 November 2019. A total of 188 patients  were randomized in 5 cohorts , 137 to P. falciparum count, which tended to be higher in cohorts 4 and 5  than cohorts 1 to 3 (850 to 6430). Cohorts 1 and 2 used potentially subtherapeutic doses: in view of this, the range of parasitemia permitted for inclusion in the study was low (500 to 50 000 parasites/\u00b5L).Patient demographics were comparable across treatment groups and consistent with the intended target population . CohortsAll cipargamin doses 50 mg or higher were associated with very rapid parasite clearance, with a median PCT of 8 hours compared with 24 hours for artemether-lumefantrine, with an apparent dose response plateauing at the 50-mg single dose . Figure Parasite reduction ratios at 24 hours increased with cipargamin dose. Peak median PRR24 around 1000 was achieved with the 50-mg single dose, whereas median PRR24 for the 25-mg single dose was approximately 600 and was below 100 for the 10-mg single dose. Due to the rapid parasite clearance with higher cipargamin doses, such that postbaseline parasite levels were undetectable in many patients, and the exclusion of patients with more than 50 000 parasites/\u00b5L from the study, the observed PRR24 for those doses may not reflect the actual values possible.The FCT could not be meaningfully assessed in cohorts 1 to 3 due to the small numbers of patients with pyrexia, which was probably related to antipyretic treatment with paracetamol (acetaminophen). In cohorts 4 and 5, FCT was shorter with all cipargamin dose regimens (means ranging from 5.7 to 9.9 hours) than the pooled artemether-lumefantrine group (13.2 hours).The PCR-corrected ACPRs at 14 and 28 days of over 75% and 65%, respectively , were acPfatp4 gene. One specific mutation in the Pfatp4 gene, G358S, was detected in samples from 22 of 34 patients treated with cipargamin at the time of treatment failure  and were approximately dose-proportional. Pharmacokinetic parameters after multiple doses were consistent with those after single doses. High variability in exposure (coefficient of variation: 25\u201353%) was observed across cohorts similarly to previous studies [Summary statistics for pharmacokinetic parameters are presented in The exposure\u2013response relationship for PRR24 was explored. PRR24 increased steeply with cipargamin exposure. Peak PRR24 appeared to be at an exposure of 15\u201320 \u03bcg.hr/mL and remained stable at higher exposures . For patIn recent years, mortality from malaria has declined, mainly due to the deployment of vector control measures and antimalarials based on artemisinin derivatives . ArtemisCipargamin is a novel spiroindolone antimalarial that has shown rapid parasite clearance in phase II studies , 2, 5. TThis study was not powered to assess efficacy, and the sequential cohorts and differences in baseline parasite counts complicate comparisons of efficacy between treatment arms. Patients were adults in sub-Saharan Africa , so efficacy results may not be easily generalizable to other age groups or regions , 5.In previous studies, cipargamin treatment was notable for the rapid clearance of parasites , 5. ResuThe ACPR, reported here across a wide cipargamin dose range, including 5 single-dose arms, was similar across treatment groups at day 15, but at day 29, recrudescence, LCF, and LPF were more frequent with cipargamin than with artemether-lumefantrine. This might be expected as cipargamin was used as monotherapy and the elimination half-life is approximately 24 hours. Other antimalarials with similar or shorter half-lives have also shown high rates of recrudescence when used as monotherapies , 24. In PfATP4 gene that lead to decreased susceptibility have been selected in vitro under prolonged drug pressure of cipargamin, which indicates the polymorphic nature of the gene [PfATP4 mutation (G358S) was common. The G358S mutant was first described from in vitro experiments and was generated under selection pressure for SJ733 [PfATP4 mutations were not detected in any of the baseline samples, this suggests that there is no pre-existing cipargamin resistance. Treatment with cipargamin monotherapy appears to have selected resistant parasites that arise spontaneously and are likely to be present in small numbers in patients with high parasitemia and undetectable at baseline [As with other PfATP4 inhibitors, mutations in the the gene , 11, 25.or SJ733 , a dihydor SJ733 . Since tP. falciparum malaria. This monotherapy study confirms the need for fixed-dose combination therapy to avoid recrudescence and selection for resistant parasites. Cipargamin, with its high potency, rapid parasite clearance, and potential for single-dose cure will be further developed for uncomplicated malaria with a suitable combination partner.In conclusion, cipargamin, at single doses of 50 mg to 150 mg, was associated with very rapid parasite clearance and PCR-corrected ACPR at day 29 of at least 65% in adult patients with uncomplicated Clinical Infectious Diseases online. Consisting of data provided by the authors to benefit the reader, the posted materials are not copyedited and are the sole responsibility of the authors, so questions or comments should be addressed to the corresponding author.Supplementary materials are available at ciab716_suppl_Supplementary_MaterialsClick here for additional data file."}
+{"text": "Plasmodium vivax malaria, Plasmodium falciparum malaria, and DHF admitted to the Hospital for Tropical Diseases, Bangkok, Thailand.Although platelet indices are routinely available using automated blood cell counters, the clinical applications of these parameters for malaria and dengue hemorrhagic fever (DHF) have not been substantially implemented. We conducted this study to investigate the potential role of platelet indices as a prognostic marker in adult patients with P. falciparum malaria, 71 with P. vivax malaria, and 52 with DHF. We evaluated the study groups\u2019 baseline clinical features and alterations of platelet indices during the first 4\u00a0days of admission.We enrolled 219 eligible patients, comprising 96 with P. vivax and P. falciparum; however, mean platelet volume (MPV) was significantly higher in patients with P. falciparum. Comparisons of the initial platelet indices in malaria and DHF showed that only PC and PCT were significantly lower in DHF. Although MPV in DHF tended to be lower than in malaria, a statistically significant difference was observed only with P. falciparum. Moreover, the results also showed no significant alterations in the platelet indices among the study groups during the first 4\u00a0days of admission.Upon admission, the initial laboratory findings showed no statistically significant difference in platelet count (PC), plateletcrit (PCT), or platelet distribution width (PDW) between patients with P. vivax and P. falciparum malaria mimicking DHF. Although a significant reduction in PC and PCT in DHF might be a clue for differential diagnosis of malaria, the use of MPV and PDW might be impractical. We suggest that appropriate laboratory diagnoses for malaria and dengue infections are still needed for the differential diagnosis of acute febrile patients who have a risk of malaria or dengue infections. To clarify the clinical utility of platelet indices in patients with dengue and malaria, further studies are required that particularly include patients with different severities, geographical areas, and levels of health care settings.Clinical presentations of DHF and malaria are nonspecific and may overlap with other common tropical diseases. Alterations of initial platelet indices may be investigated in  Anopheles and Aedes mosquitoes, respectively. Malaria and DHF are life-threatening diseases affecting subtropical and tropical countries, particularly Southeast Asia and Asia\u2013Pacific regions . In patients with P. vivax, the median PC tended to increase up to day 4 after admission, compared with the baseline level. Only the median PC of patients with P. falciparum reached a normal level (150\u2009\u00d7\u2009103/\u03bcL) within 4\u00a0days after admission , but a lower value was observed in patients with DHF (0.06%). The median PCT of P. falciparum decreased to 0.05% on day 2. However, on day 3, the median PCT increased to 0.06% and continually increased to 0.12% on day 4. With regard to patients with P. vivax, an increasing trend in the median PCT was observed during the first 4\u00a0days after admission. The median PCT on day 1 and day 2 was equal at 0.07% and increased to 0.11% on day 3 and 0.12% on day 4, respectively. We also found that the median PCT in DHF tended to be constant during the study period. The median levels on day 1 and day 2 were 0.06%. The level on day 3 declined slightly to 0.05%, and then the value returned to 0.06% on day 4.A similar pattern was observed in the dynamicity of PCT Fig.\u00a0. We noteP. vivax (8.3 fL) and DHF (8.6 fL) was slightly lower than in P. falciparum malaria patients (9.6 fL). The median values in P. falciparum malaria patients seemed to be constant on days 1, 2, and 3 and then dropped slightly to 8.75 fL on the fourth day. The median MPV in patients with P. vivax was constant on day 1 (8.3 fL) and 2 (8.35 fL) and then raised up to 10.15 fL on day 3 and 10.55 fL on day 4, respectively. In patients with DHF, an increasing trend in the median MPV was found on day 2 and day 3 (9.25 fL and 9.7 fL), respectively. However, the median level declined slightly on day 4 (9.1 fL).Interestingly, a negative trend was observed in the median MPV during the study period Fig.\u00a0. On day P. falciparum (57%), P. vivax (56.1%), and DHF (55.9%) were closely presented. In P. falciparum, an increasing level was observed on day 3 (59.25%) but decreased to 52.45% on day 4. The value of P. vivax, however, was decreased to 51.6% and then increased up to 61.4% on day 2 and day 4, respectively. Interestingly, the median PDW in patients with DHF increased to 60.5% on day 3 and then decreased to 60% on the fourth day  is a clinical laboratory condition that is usually observed in patients suffering from dengue infection and P. falciparum and P. vivax malaria. However, severe bleeding and platelet transfusion are rarely required [P. falciparum, P. vivax, and DHF, respectively. In our findings, a reduction in the patterns of PC did not differ statistically among the different disease groups, leading to the assumption that similar mechanisms govern this condition. With regard to thrombocytopenia and coagulation abnormality conditions in patients with dengue and malaria, most patients with DHF in our study had a PC level of\u2009<\u2009150\u2009\u00d7\u2009103/\u00b5L and an initial median PC that was statistically lower than in patients who were infected with malaria. In patients with DHF, up to 38% presented with a PC level of\u2009<\u200950,000/\u00b5L, and about 26.9% of the study patients were clinically associated with mild abnormal mucosal bleeding. This might be due to the intrinsic decrease in platelet values in adult patients superimposed on the bone marrow\u2019s direct effect of the dengue virus. Other reasons could be the alteration in cytokines, which interferes with the function of the bone marrow, or the immune-mediated destruction of platelets rather than platelet sequestration [Thrombocytopenia  as potential laboratory markers in patients suspected of having malaria or dengue , 23, 24.PDW is a laboratory marker of platelet anisocytosis, indicating variability in the volume and size distribution of platelets. The average size of platelets may be indicated by the MPV levels for clinical bone marrow activity. Total platelet biomass could be implied by the PCT value, showing a percentage of platelet volume occupied in blood. Therefore, PCT might be useful as a screening tool for detecting a quantitative platelet abnormality, because it is a combination between MPV and PC , 27. AccBecause platelet indices are machine specific, in cases of thrombocytopenia, these indices might not be reported in blood specimens in some machines. Moreover, this result of platelet indices might be interfered with by abnormal red blood cell parameters, such as in patients with thalassemia disease, who were not included in this study. Another limitation of this study was the days of admission at the early stage. The day of admission might not reflect patients\u2019 true thrombocytopenia-related condition, particularly in patients with dengue infection, as they might be in different infection phases (febrile versus toxic phases), which could affect PC and platelet indices. Moreover, changes in platelet parameters are dynamic; therefore, an analysis of the delta change in the platelet parameters might better explain the patient\u2019s condition, and a combination of platelet parameters might provide a more complete picture of the patient for further investigation.In conclusion, our findings show that alteration patterns in platelet indices, between DHF and malaria, might be impractical as a potential laboratory biomarker for clinical differential diagnosis. Although we found a significant reduction in the PC and PCT of patients with DHF, PDW and MPV did not demonstrate significance in comparison with malaria patients. Therefore, differential diagnosis using platelet indices as a clinical laboratory predictor in patients with dengue and malaria should not be used for clinical practice. We suggest that malaria should be microscopically detected and an appropriate laboratory diagnosis obtained for dengue infection for differential diagnosis, particularly in acute febrile patients who are at risk of malaria and dengue infections. Because both malaria and dengue infections share the same pathophysiology, the use of a PC and platelet indices might be more beneficial for determining the severity of the disease than the differential diagnosis of both diseases separately. Morbidity and mortality in patients with malaria and dengue could be reduced by appropriate prompt treatment after an early effective diagnosis. Therefore, further studies of platelet indices in patients with dengue and malaria, particularly among those with conditions of different severity, different geographical areas, and different levels of health care settings, may be required to construct practical measures for these patients."}
+{"text": "Hospital referral and admission in many- low and middle-income countries are not feasible for many young infants with sepsis/possible serious bacterial infection (PSBI). The effectiveness of simplified antibiotic regimens when referral to a hospital was not feasible has been shown before. We analysed the pooled data from the previous trials to compare the risk of poor clinical outcome for young infants with PSBI with the two regimens containing injectable procaine penicillin and gentamicin with the oral amoxicillin plus gentamicin regimen currently recommended by the World Health Organization (WHO) when referral is not feasible.Infant records from three individually randomised trials conducted in Africa and Asia were collated in a standard format. All trials enrolled young infants aged 0-59 days with any sign of PSBI . Eligible young infants whose caretakers refused hospital admission and consented were enrolled and randomised to a trial reference arm (arm A: procaine benzylpenicillin and gentamicin) or two experimental arms . We compared the rate of poor clinical outcomes by day 15  in reference arm A with experimental arms and present risk differences with 95% confidence interval (CI), adjusted for trial.P\u2009=\u20090.016) and intention-to-treat  analyses. Those who received arm C did not have an increased risk of poor clinical outcome compared to arm A for both per-protocol  and intention-to-treat  analyses. Overall, those who received arm B had a lower risk of poor clinical outcome compared to the combined arms A and C for both per-protocol  and intention-to-treat  analyses.A total of 7617 young infants, randomised to arm A, arm B, or arm C in the three trials, were included in this analysis. Most were 7-59 days old (71%) and predominately males (56%). Slightly over one-fifth of young infants had more than one sign of PSBI at the time of enrolment. Severe chest indrawing (45%), fever (43%), and feeding problems (25%) were the most common signs. Overall, those who received arm B had a lower risk of poor clinical outcome compared to arm A for both per-protocol  was superior to regimen A (injection procaine penicillin and injection gentamicin) and combined arms A and C  in terms of poor clinical outcome for the outpatient treatment of young infants with PSBI when inpatient treatment was not feasible.AFRINEST study [9] is registered with the Australian New Zealand Clinical Trials Registry: ACTRN12610000286044. SATT Bangladesh study [10] is registered with ClinicalTrials.gov: NCT00844337. SATT Pakistan study [11] is registered at ClinicalTrials.gov: NCT01027429. Of the estimated 2.5 million newborns who die each year, neonatal infections, including pneumonia, sepsis, and meningitis, are estimated to be the cause of 500\u2009000 deaths . Most ofConsidering this, a large, multi-stakeholder collaborative study was established to generate evidence to optimally treat young infants 0-59 days old who present with possible serious bacterial infection (PSBI) when treatment at a hospital is not feasible. These trials, termed the Simplified Antibiotics Treatment Trials (SATT), were conducted in Asia (Bangladesh and Pakistan) and in Africa ) . Three aBox 1The following treatment regimens were evaluated in the three trials:1. Arm A- intramuscular daily injections of procaine penicillin and gentamicin for 7 days.2. Arm B- oral amoxicillin and intramuscular gentamicin for 7 days.3. Arm C- intramuscular procaine penicillin and gentamicin for 2 days followed by oral amoxicillin for another 5 days.4. Arm D (Only in AFRINEST)- oral amoxicillin for 7 days plus intramuscular gentamicin only for the initial 2 days.Antibiotic dosages:1. Injection procaine benzylpenicillin in a dose of 50000 units/kg once per day intramuscularly.2. Injection gentamicin in the range 4.0 mg/kg once per day IM in the first week of life and 7.5 mg/kg once daily intramuscularly thereafter.Oral amoxicillin in suspension in a dose of 100 mg/kg per day (less than 2 kg are given 75 mg/kg per day), divided into two equal doses.The World Health Organization (WHO) recommended simplified antibiotic regimens for young infants with PSBI signs based on the three trials -11 if paWe conducted a pooled analysis across the three trials -11 to coInfant records from three individually randomised trials conducted in Africa and Asia were collated in a standard format. The details of the design of the methods used in each trial have been described in detail elsewhere -11,15-18All three trials enrolled young infants aged 0 to 59 days with at least one of the following five signs: fever (body temperature \u226538\u00b0C), hypothermia (body temperature <35.5\u00b0C), cessation of feeding well, movement only when stimulated, or severe chest indrawing. Young infants with signs of very severe disease, including convulsions, apnoea, inability to feed at all, unconsciousness, inability to cry, cyanosis, persistent vomiting or bulging fontanelle were excluded. Very low weight (<1500 g at the time of presentation) and hospital admission for illness in the past two weeks or previously enrolled in the study were also excluded. Caretakers of young infants clinically eligible for enrolment were advised a referral to a hospital for admission, and only young infants whose caretakers refused hospital admission and consented to inclusion in the trial were enrolled.Following recruitment, young infants were randomised to one of the following three treatment arms in Bangladesh and Pakistan: arm A, the trial reference arm, arm B, and arm C, while AFRINEST also included a third alternative treatment arm D  and on two other occasions during the second week after enrolment (day 11 and day 15). The primary outcome in all trials was treatment failure during the first week after enrolment, as defined in Box 21. Death from the day of enrolment till day 152. Treatment failure is the occurrence of any of the following:- Clinical deterioration at any time up to Day 8 based on the presence of at least one of the following danger signs: unconscious, convulsions, unable to feed, apnoea, cyanosis, bulging fontanelle, major bleeding, persistent vomiting.- Decision at any time up to day 8 by study personnel to change the antibiotic regimen or add another antibiotic for either of the following reasons:- New-onset infectious co-morbidity , or- Serious non-fatal antibiotic-associated adverse event .- Hospitalisation at any time up to Day 8 for any reason.- On or after Day 3: the occurrence of new signs of PSBI . (A \u201cnew\u201d sign was one that was not present at the time of enrolment.)- On Day 4, for infants with multiple signs at enrolment: Presence of at least two of the signs that were present on enrolment;- On Day 4, for infants with a single sign on enrolment: Presence of the same sign that was present on enrolment.- On or after Day 5: Recurrence of any of the following signs: fever, hypothermia, severe chest indrawing, movement only when stimulated, or poor feeding. (Recurrence implies the presence of the sign on enrolment AND documented resolution of the sign on at least one follow-up visit with the subsequent reappearance of the same sign on at least one follow-up visit on or after Day 5.)- Persistence on Day 8 of any of the five signs of PSBI that was present on enrolment.3. Non-fatal relapse Young infants were defined as adherent to treatment provided that they received 100% of scheduled antibiotic doses on days 1-3 or by the time of treatment failure, they received at least 50% of scheduled doses of each antibiotic on days 4 to 7 or by the time of treatment failure, and they did not receive any non-study injectable antibiotic prior to the day 8 assessment or treatment failure and did not receive any non-study oral antibiotic on days 1-3 prior to any treatment failure.In Bangladesh and Pakistan, young infants were classified as having adequate clinical follow-up if follow-up was completed during days 2-4, if follow-up was completed on at least one of days 5-8, and if the infant\u2019s vital status on day 8 was known.In AFRINEST, an infant was considered to have adequate clinical follow-up providing s/he was seen by an independent outcome assessor on day 4 and at least one of days 8, 11 and 15. In all trials, an infant was classified as eligible for inclusion in the per-protocol analysis providing that s/he was adherent to treatment and had an adequate clinical follow-up.The individual patient-level data, including the baseline characteristics, treatment regimen, and clinical outcome of each child, were pooled for the analyses. We excluded infants enrolled in arm D of the AFRINEST because arm D regimens were not considered in the Bangladesh and Pakistan trials. The primary aim was to obtain more precise estimates of the risk difference by comparing experimental treatment regimens (arm B or C) with standard regimens (arm A).www.stata.com).We performed both per-protocol and intention-to-treat analyses. We calculated the pooled risk difference with 95% confidence intervals (95% CI) and risk ratio with 95% CI for poor clinical outcomes between the experimental regimens (arm B or C) and arm A for outpatient treatment of young infants with PSBI, adjusting for trial. Additionally, we calculated the trial-specific risk difference with 95% CI and risk ratio with 95% CI for poor clinical outcomes between the experimental regimens (arm B or C) and arm A for outpatient treatment of young infants with PSBI. Moreover, we also calculated the pooled risk difference with 95% CI and risk ratio with 95% CI for poor clinical outcomes between arm B and combined arms A and C. All analyses were performed using Stata version 16  , 2490 inWe randomised 2544 infants to study arm A, 2531 to arm B and 2542 to arm C in the three trials -11. The P\u2009=\u20090.016), and the risk ratio was 0.81 . Similarly, for intention-to-treat analysis, 11.6% of 2544 infants in arm A, 9.8% of 2531 infants in Arb B, and 10.9% of 2542 infants in arm C had poor clinical outcomes. The risk difference between arm A and B was -1.8% , and the risk ratio was 0.84 . Those in arm C did not have an increased risk of poor clinical outcome compared to arm A for both per-protocol  and intention-to-treat  analyses.P\u2009=\u20090.035) and intention-to-treat  analyses  tool , which uEscherichia coli, other enteric gram-negative rods, streptococci, and pneumococci. It was impractical to give injectable benzylpenicillin or ampicillin four times a day in an outpatient setting. Injection procaine penicillin plus gentamicin was selected as the reference arm to treat PSBI based on the recommendations from the WHO consultation, the pharmacologic profile of antibiotics [The choice of these antibiotic regimens for outpatient use in the AFRINEST and SATT studies might be questioned by some. First, various antibiotic regimens had been used to manage neonatal sepsis at the community or an outpatient level in several research studies. Bang et al.  reportedibiotics ,24, and ibiotics . Both caibiotics -28. For ibiotics ,30. Fiftibiotics  did not ibiotics , and somibiotics . FinallyA potential explanation for procaine-penicillin being less effective than oral amoxicillin could be its slow dissolution after intramuscular injection, giving a maximum blood level at about 4 hours, which falls slowly over the next 15-20 hours . A 50\u200900Soon after its launch, the 2015 WHO PSBI management guideline when referral is not feasible was implemented in several countries to learn lessons on how to use it in diverse contexts. Summary results showed that the estimated coverage of PSBI treatment (including inpatient and outpatient treatment) in sick young infants was 76% when the guideline was used . If suchEquivalence trials comparing antibiotic regimen options for treatment of sick young infants with PSBI when inpatient treatment is not feasible are criticized by some experts because they excluded clinically severe cases and did not confirm neonatal sepsis microbiologically -38. If fThe main strength of our analyses is the large sample size from both African and Asian countries, which helps its generalizability. Second, a pooled analysis at the individual patient level was conducted with this large multi-country data, which supports the robustness of our results. The main limitations of this analysis are those inherent to the original trials. First, these trials were not blinded because it was unethical to give placebo injections to sick young infants to have an equal number of injections in all arms. Second, microbiological or other laboratory tests were not used to support the clinical diagnosis, as it was not feasible to set up such services at the first-level health facilities within the available resources.Analysis of pooled individual patient-level data from three large trials in Africa and Asia showed that the WHO-recommended simplified antibiotic regimen B  was superior to regimen A (injection procaine penicillin and injection gentamicin) and combined arms A and C  in terms of poor clinical outcome for the outpatient treatment of young infants with PSBI when inpatient treatment was not feasible. These findings can help to substantially expand our options to increase access to early and effective treatment for those who may otherwise go untreated."}
+{"text": "Plasmodium falciparum infection, hemoglobin, and grade point averages (GPA) based on standardized testing. Overall, 305 students contributed 4,564 observations. Compared with the control, SP plus artesunate and AQ plus artesunate reduced the odds of P. falciparum infection . We found strong evidence of increased mean hemoglobin concentrations (g/dL) in the SP plus artesunate group versus control . Collectively, schoolchildren given AQ plus artesunate had higher mean GPA  relative to control. Schoolgirls, compared with schoolboys, given SP plus artesunate had greater improvement in GPA ; interaction P = 0.048, respectively. The IPTsc decreases P. falciparum infections in schoolchildren. Treatment regimens that include longer-acting drugs may be more effective at decreasing malaria-related anemia and improving educational outcomes as observed among girls in this setting.Intermittent preventive treatment of malaria among schoolchildren (IPTsc) reduces clinical malaria, asymptomatic parasitemia, and anemia. The effects of IPTsc by gender have not been studied longitudinally. We investigated overall IPTsc efficacy and conducted a secondary analysis to explore gender-specific differences. We enrolled schoolchildren aged 6\u201313 years in an open-label, rolling-cohort randomized controlled trial between September 2007 and February 2013 in Kolle, Mali. Annually, schoolchildren received two full-treatment courses of sulfadoxine-pyrimethamine (SP) plus artesunate, or amodiaquine (AQ) plus artesunate, or no malaria treatment as control. We used mixed-effects generalized linear models to estimate differences in treatment outcomes across groups with interaction terms to explore gender-specific differences associated with  Plasmodium falciparum infection relative to other age groups is highest among school-age children\u2013P. falciparum infection fuelling malaria transmission and challenging malaria elimination efforts.,In 2020, an estimated 169 million cases of malaria resulting in 444,600 deaths were reported worldwide,P. falciparum infections account for 36% of medical consultations among school-aged children during the peak transmission season,About 200 million school-age children are at risk of malaria in Africa with the prevalence of infection often exceeding 50%.Intermittent preventive treatment (IPT) of malaria is an intervention designed to protect at-risk populations from clinical malaria and the consequences of asymptomatic parasitemiaP. falciparum infections, malaria-related anemia, and improves cognitive performance among older school-age children.P. falciparum prevalence is higher among boys than girls, whereas the overall incidence of clinical malaria is greater among girls.,There are currently no WHO recommendations, such as IPT for schoolchildren (IPTsc), despite mounting evidence that preventive therapy targeting this group decreases ,We previously reported results from the first 8 months of an IPTsc trial.P. falciparum infection ranges from 70% to 85%,Anopheles gambiae sensu lato comprises > 96% of all Anophelines.P. falciparum infection prevalence is 40\u201350%,Anophelines are An. funestus.The study was conducted in Kolle, a subsistence farming community of approximately 3,000 inhabitants situated 57 km southwest of Bamako, Mali. The rainy season, June to October, has an average of 176 mm rainfall,\u00a9, Microsoft Inc., Washington, WA) of study numbers and treatment group allocation. Children presenting for enrolment were assigned a study number in the sequence of presentation within their school grade. Participants were stratified by grade to ensure approximately equal numbers in each study group. Clinicians were aware of group allocation at the time of randomization. Once allocated, children remained in the same study group for the duration of the trial.The open-label, three-group, randomized controlled trial was conducted between September 2007 and February 2013 among schoolchildren who were randomized to receive IPTsc of SP plus artesunate, AQ plus artesunate, or no antimalarial therapy (control). The IPTsc was administered as two courses, 8 weeks apart from the first course early in the school term, overlapping with the high malaria transmission season (September or October). Monthly follow-up visits, including physical examinations and blood collection for measurement of hemoglobin and microscopic quantification of parasitemia, were conducted throughout the transmission season. Study outcomes were analyzed at the end of each year. Schoolchildren aged out of the trial follow-up when they completed sixth grade. Participants were randomized using a computer-generated shuffle . Monthly blood smears were read by two blinded, certified microscopists in the Malaria Research and Training Center. Discordant results were read by a third certified reader, which was considered the final read. Regardless of parasite status, students were given either: (1) SP as 25/1.25 mg/kg/day in a single-dose plus artesunate as 4 mg/kg once for 3 days; or (2) AQ as 10 mg/kg/day plus artesunate as 4 mg/kg/day for 3 days; or (3) no antimalarial treatment (control group). Vitamin C, rather than placebo, was given to first-year participants in the control group, though this was replaced by water in the following years. Children received their first course of annual study medication in September 2007, and in October in all the subsequent years. All doses were directly monitored. Participants were observed for signs of intolerance for a minimum of 30 minutes. If vomiting occurred during this observation period, treatment was readministered. Children returned the following 2 days to complete their respective regimens. Students were followed from 1 year to the next and contributed to the endpoints of the trial so long as they remained age eligible. Participants were instructed to come to the clinic for any illness between follow-up visits. Children with signs or symptoms of malaria had thick and thin smears read by microscopy and hemoglobin testing. Cases were managed with a re-dose of antimalarial treatment that matched the trial group allocation; students in the control group received SP in the first year of the trial to control SP efficacy at this time, and artemether-lumefantrine in the following years. Clinicians provided the supportive care and treatment of all other diagnoses according to national guidelines.Children received an initial history and physical examination while inclusion and exclusion criteria were reviewed. Trial staff collected finger-prick blood for the preparation of thick and thin blood smears and the measurement of hemoglobin concentration  based on standardized testing, during the monthly visits for the 5 months after the initial treatment of that year. Asymptomatic parasitemia was defined as participants having no signs of illness but at least one parasite per 100 fields on microscopy reading. The WHO definition was used for anemia defined as hemoglobin < 11g/dL.\u2122 database 2010 . Statistical analysis was performed using Stata SE version 16 . Baseline characteristics of participants were summarized by the allocation group using descriptive statistics. Pooled post-intervention outcomes aggregated by treatment group were also reported. Mixed-effects generalized linear models of the Gaussian family with an identity link were fitted to estimate differences in hemoglobin and GPA as calculated by standardized testing across allocation groups. A mixed-effects generalized linear model of the binomial family with a logit link was fitted to estimate the odds ratios for malaria prevalence across groups. In all mixed models, random intercepts were specified at the individual level with an unstructured variance\u2013covariance matrix to allow for different correlations between pairwise measurements on the same child at different times. A negative binomial generalized linear model with robust standard errors (SE) to adjust for clustering of repeated observations at the individual level was used to estimate differences in P. falciparum parasite density across the allocation groups. Interaction between allocation groups and gender were further fitted in each model to investigate differences in treatment effects by gender, with a likelihood ratio test to assess evidence for a subgroup effect. All models were further adjusted for grade, which was a stratification factor in the randomization; given the strong correlation between age and grade, this also served as an adjustment for age. Additionally, the interaction between allocation group and age was explored in all models to investigate whether any treatment effects varied as children got older and progressed through grades over time. For models of hemoglobin and parasitemia, adjustments for baseline values of the outcome were also included in the models. Marginal estimates of hemoglobin predicted by the mixed-effects model with interaction were plotted against grade to visually explore trends in hemoglobin across children of different ages in the three treatment groups.Our previous studies in Kolle found the prevalence of asymptomatic parasitemia in the control group to be 75%. To have 95% power to detect a 33% relative reduction in asymptomatic parasitemia at a 5% two-sided level of statistical significance, allowing for a 10% loss to follow-up, 97 children were required in each group for a total sample of 291. Children left the study after sixth grade; therefore, approximately 51 students were removed each year from the study by planned attrition. Data were double entered into a Microsoft AccessThe study protocol was approved by the Ethics Committee of the Faculty of Medicine and Odonto-Stomatology, Faculty of Pharmacy at the University of Sciences, Techniques and Technologies of Bamako, Bamako, Mali (N\u00b059/CE/FMPOS/2007 and amendment N\u00b059/CE/FMPOS/2008). This trial is registered with the Pan African Clinical Trials Registry (PACTR202109827515839).Among the 475 students 6\u201313 years of age in the village, 305 children presented for enrollment following the village crier\u2019s announcement of the study. Nine children declined to participate and 296 enrolled in 2007. Two students assigned to SP plus artesunate (0.7%) were lost to follow-up by January 2008: one was excluded for being younger than 6 years old, and the other ceased participation at the second treatment dose. We enrolled 11 new students to add to the study cohort in October 2008. Consequently, this analysis includes 305 students: 102 received SP plus artesunate, 103 were given AQ plus artesunate, and 100 were in the control group .P. falciparum infection was higher in the AQ plus artesunate group compared with the SP plus artesunate and control groups.Data on at least one characteristic were available from 305 children at baseline . There wP. falciparum infection was measured at the end of each malaria transmission season and pooled over the 6 years of the trial by allocation group: 24.1% of children in the control group had at least one infection, compared with 10.5% given SP plus artesunate, and 13.4% who received AQ plus artesunate (adjusted OR [aOR]) of infection: 0.34  for SP plus artesunate and 0.46  for AQ plus artesunate; P. falciparum prevalence . There was also no evidence of an adjusted difference in P. falciparum parasite densities in the SP plus artesunate or AQ plus artesunate groups compared with the control group  nor a gender-specific or age/grade-varying difference in effect .Prevalence of P = 0.002) in the SP plus artesunate, and 0.15  in the AQ plus artesunate relative to control. Nevertheless, there was evidence of a difference in treatment effect across the years of follow-up as children aged through the cohort . Trends by grade-level over the course of the study are shown in P value for interaction 0.601).Mean hemoglobin at the end of follow-up was 11.9 g/dL (SE: 0.1) among control versus 12.4 g/dL (SE: 0.1) for those given SP plus artesunate and 12.1 g/dL (SE: 0.1) who received AQ plus artesunate, with adjusted mean differences of 0.35 g/dL  among recipients of SP plus artesunate, and 0.36  among those given AQ plus artesunate, relative to control. There was some evidence of a gender-specific difference in the effect of treatment , with higher adjusted mean GPA in schoolgirls than schoolboys in the SP plus artesunate group , but there was a similar effect by gender in the AQ plus artesunate group . There was no evidence of an age/grade-varying difference in effect on GPA Mean GPA at the end of follow-up was 4.9 (SE: 0.1) in the control group, 5.0 (SE: 0.1) in the SP plus artesunate group, and 5.2 (SE: 0.1) in the AQ plus artesunate group, an adjusted mean difference of 0.04 units (95% CI: \u22120.30 to 0.38, P. falciparum infection among schoolchildren living in areas with high malaria endemicity. However, the impact of preventive treatment on anemia and education depends on the intervention drug. The antimalarial drugs used in this trial are formulations widely known to be safe and well tolerated. No cases of deaths related to the intervention or unusual adverse events were reported. These findings align with previous studies in Mali and other African countries.,\u2013Our study provides strong longitudinal evidence that preventive malaria treatment decreases ,P. falciparum infection, disease, and anemia indirectly affect educational achievement. Overall, AQ plus artesunate significantly increased GPA compared with control. This result aligns with a previous study that has shown a decrease in cognitive function linked to cerebral malaria and to asymptomatic parasitemia.P. falciparum diseaseWhile the study was not designed or powered to compare the drug regimens and head-to-head comparisons are not possible, differences in the efficacy of the regimens could be due to differences in drug half-life and protection from submicroscopic parasitemia. Sulfadoxine-pyrimethamine has a longer half-life than AQ.P. falciparum malaria and anemia compared with the control without evidence of a gender-specific effect of treatment on these outcomes. However, there was evidence of a gender-specific difference in GPA by standardized testing that trended toward an increase among schoolgirls, but not schoolboys, in the SP plus artesunate group. A potential explanation is that a larger proportion of the limitation in educational achievement for girls is attributable to P. falciparum infection. The etiologies of anemia and decreased cognitive function are multifactorial, including malaria, helminth infections, chronic inflammation, chronic undernutrition, and micronutrient deficiencies.\u2013P. falciparum operates to produce these effects. Interestingly, schoolgirls with asymptomatic malaria and anemia had significantly lower GPA compared with schoolboys in the SP plus artesunate group. Gender differences in P. falciparum infection have been previously reported in Africa.\u2013P. falciparum prevalence and hemoglobin levels are comparable and, therefore, both genders should be considered for IPTsc in a malaria control strategy.In our study, SP plus artesunate significantly decreased Our study adds to the growing body of evidence supporting IPTsc; however, there are some limitations. The study was an open label, which could have introduced bias in favor of the two treatment groups. Vitamin C was given to first-year participants in the control group, although this was replaced by water in the following years. Although vitamin C may minimally improve clinical outcomes among malaria cases,The high prevalence of malaria infection in schoolchildren underscores the need to reduce this burden and IPTsc is an efficacious intervention to achieve that aim. In addition, our longitudinal study supports further investigation of gender-specific effects of malaria interventions in children in sub-Saharan Africa."}
+{"text": "Plasmodium falciparum and Plasmodium vivax, respectively. The National Malaria Control and Elimination Programme in Ethiopia is considering dihydroartemisinin-piperaquine (DHA/PPQ) as an alternative regimen for P. falciparum and P. vivax.Routine monitoring of anti-malarial drugs is recommended for early detection of drug resistance and to inform national malaria treatment guidelines. In Ethiopia, the national treatment guidelines employ a species-specific approach. Artemether-lumefantrine (AL) and chloroquine (CQ) are the first-line schizonticidal treatments for P. falciparum and CQ against P. vivax. Patients 18\u00a0years or older with uncomplicated malaria mono-infection were recruited and randomized to AL or dihydroartemisinin-piperaquine (DHA/PPQ) against P. falciparum and CQ or DHA/PPQ for P. vivax. Patients were followed up for 28 (for CQ and AL) or 42\u00a0days (for DHA/PPQ) according to the WHO recommendations. Polymerase chain reaction (PCR)-corrected and uncorrected estimates were analysed by Kaplan Meier survival analysis and per protocol methods.The study assessed the clinical and parasitological efficacy of AL, CQ, and DHA/PPQ in four arms. Patients over 6\u00a0months and less than 18\u00a0years of age with uncomplicated malaria mono-infection were recruited and allocated to AL against P. falciparum; n\u2009=\u200975, DHA/PPQ- P. falciparum; n\u2009=\u2009142, CQ-P. vivax; n\u2009=\u200956, DHA/PPQ-P. vivax). High PCR-corrected adequate clinical and parasitological response (ACPR) rates were observed at the primary end points of 28\u00a0days for AL and CQ and 42\u00a0days for DHA/PPQ. ACPR rates were 100% in AL-Pf (95% CI: 96\u2013100), 98% in CQ-P. vivax (95% CI: 95\u2013100) at 28\u00a0days, and 100% in the DHA/PPQ arms for both P. falciparum and P. vivax at 42\u00a0days. For secondary endpoints, by day three 99% of AL-P. falciparum patients (n\u2009=\u2009101) cleared parasites and 100% were afebrile. For all other arms, 100% of patients cleared parasites and were afebrile by day three. No serious adverse events were reported.A total of 379 patients were enroled in four arms (n\u2009=\u2009106, AL-P. falciparum and P. vivax as an alternative option.This study demonstrated high therapeutic efficacy for the anti-malarial drugs currently used by the malaria control programme in Ethiopia and provides information on the efficacy of DHA/PPQ for the treatment of The online version contains supplementary material available at 10.1186/s12936-022-04350-z. Plasmodium falciparum accounting for 77% of the confirmed cases [P. falciparum, and chloroquine (CQ) remains the first-line treatment for P vivax in Ethiopia. The second-line treatment for P. falciparum uncomplicated malaria is oral quinine and for uncomplicated P. vivax malaria is AL [Malaria remains a disease of significant public health importance in Ethiopia despite the gains made through recent malaria control efforts. In 2020, the World Health Organization (WHO) estimated approximately 4.2 million malaria cases in Ethiopia with ed cases . Accordied cases . Prompt ia is AL . The EthP. falciparum throughout the country since the beginning of its use in 2004 [P. vivax has remained rare in Africa. In Ethiopia, CQ efficacy remains above 95% although sporadic reports of CQ failure suggest emerging resistance [The Ethiopian Public Health Institute (EPHI), collaborating with local and international partners ), has been monitoring the therapeutic efficacy of anti-malarial drugs that may be used for malaria management in the country. With the exception of data from a location in Arbaminch in 2008, where efficacy was reported to be 92.5% , AL efficacy remained greater than 95% against uncomplicated  in 2004 \u201310. Howe in 2004 , 12. Chlsistance \u201320. TherP. falciparum and P. vivax and well-tolerated in Africa and Asia [Although DHA/PPQ is a WHO-recommended treatment for malaria (regardless of species) and the first-line therapy in many countries in Asia and Africa, its efficacy has not been evaluated in Ethiopia to date. It has been demonstrated to be highly effective against both and Asia \u201325. Gastand Asia . QT proland Asia .P. falciparum and CQ or DHA/PPQ against uncomplicated P. vivax to provide on-going, evidence-based recommendations for the national malaria treatment guidelines.This study reports the therapeutic efficacy of AL or DHA/PPQ against uncomplicated P. falciparum and CQ or DHA/PPQ in patients with uncomplicated P. vivax.The study evaluated adequate clinical and parasitological responses (ACPR) to standard therapeutic doses of AL or DHA/PPQ in patients with uncomplicated The study was conducted in two sentinel sites in Ethiopia: 1) Felegeselam Health Centre, Pawe, Metekel Zone, Benishangul Gumuz Region and (2) Arbaminch Health Centre, Gamu-Gofa Zone, Southern Nations and Nationalities Peoples\u2019 (SNNP) Region . The study was based on the WHO recommendations for designing surveillance studies on anti-malarial drug efficacy [P. falciparum or P. vivax were enroled in the study. Plasmodium falciparum-infected patients 18\u00a0years of age and above were randomized to the AL or DHA/PPQ arms. All patients with P. falciparum older than six months and younger than 18\u00a0years were enroled in the AL arm. Similarly, patients with P. vivax 18\u00a0years of age and above were randomized to the CQ or DHA/PPQ arms. All patients older than six months and younger than 18\u00a0years were enroled in the CQ arm. Enrolment to DHA/PPQ arms were limited to adults 18\u00a0years of age and above as per the guidance of the Food, Medicine and Health Care Administration and Control Authority of Ethiopia (FMHACA).The study was designed as an open label, four arm trial conducted in two sites  was conducted to review the study protocol for the central study team. The site teams comprised of six people per site: two clinicians, two laboratory technologists, a porter/tracer, and a supervisor. The central team provided on-site training and supervision for the first two weeks of study enrolment. The site teams received additional regular supportive supervision throughout the study period.P. falciparum enroled in the study were treated with either AL or DHA/PPQ (if\u2009\u2265\u200918\u00a0years of age), and patients with P. vivax were treated with either CQ or DHA/PPQ (if\u2009\u2265\u200918\u00a0years of age). Artemether-lumefantrine  was administered twice daily for three days, and DHA/PPQ (40\u00a0mg DHA and 640\u00a0mg PPQ), Duo-CotecxinR, Holley-Cotec Pharmaceuticals, China) was administered once daily for 3\u00a0days according to the manufacturers\u2019 recommendations. CQ  was prescribed according to national treatment guidelines at 25\u00a0mg base/kg over 3\u00a0days . All study drugs were provided by the WHO Global Malaria Programme. All DHA/PPQ and CQ treatment doses were given under the direct supervision of study clinicians and study team members, whereas for AL only the morning dose was supervised, the evening dose was taken at home and patients were asked if they took the drug as instructed before administering the next dose. Patients were encouraged to eat fatty foods and expected to report completion of the second, evening AL dose.Patients with P. vivax mono- or mixed infection during enrolment or follow-up were offered treatment with 14\u00a0days of primaquine (0.25\u00a0mg/kg)  as per the national treatment guideline for radical cure at the end of the follow-up period or upon reaching a study endpoint.All patients were observed for adverse reactions or vomiting for 60\u00a0min following treatment administration. Patients vomiting their medication within the first 30\u00a0min received a repeat full dose; patients vomiting within 30\u201360\u00a0min received half the original dose. Patients with At enrolment, patients completed a medical examination and questionnaire, and a capillary blood sample was collected for blood film examination and haemoglobin measurement. In addition, three 50-\u03bcl capillary blood spots were stored on filter papers .Patients were asked to return for routine assessment, follow-up medical exam, and blood film examination on days 1, 2, 3, 7, 14, 21, 28, 35, and 42. For the primary efficacy outcome, AL and CQ were assessed to day 28 and DHA/PPQ to day 42 as recommended by WHO. Polymerase chain reaction (PCR) correction was only done to day 28 for AL and CQ arms. Patients were also asked to return to the clinic if they had signs or symptoms consistent with malaria or any adverse events on non-scheduled follow up days. Adverse events and concomitant medications were recorded at every visit, and repeat haemoglobin concentration was measured on days 0, 14, 28 and 42.P. falciparum and P. vivax [According to WHO recommendations, treatment responses were classified as early treatment failure (ETF), late clinical failure (LCF), late parasitological failure (LPF), or ACPR for P. vivax .Development of danger signs or severe malaria on day 1, 2 or 3, in the presence of parasitaemia; parasitaemia on day 2 higher than on day 0, irrespective of axillary temperature; parasitaemia on day 3 with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C; and parasitaemia on day 3\u2009\u2265\u200925% of count on day\u00a00.Development of danger signs or severe malaria in the presence of parasitaemia on any day between day 4 and day 28 in patients who did not previously meet any of the criteria of early treatment failure; and presence of parasitaemia on any day between day 4 and day 28 with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C in patients who did not previously meet any of the criteria of early treatment failure.Presence of parasitaemia on any day between day 7 and day 28 with axillary temperature\u2009<\u200937.5\u00a0\u00b0C in patients who did not previously meet any of the criteria of early treatment failure or late clinical failure.Absence of parasitaemia on day 28 or 42, irrespective of axillary temperature, in patients who did not previously meet any of the criteria of ETF, LCF or LPF.The primary endpoints were PCR-corrected ACPR on day 28 for AL and CQ and day 42 for DHA/PPQ. Other outcomes were loss to follow-up and withdrawals which included protocol violation, withdrawal of consent/assent, interference , and failure to complete study treatment .Blood samples collected by finger prick from febrile outpatients were stained by 10% Giemsa for 10\u201315\u00a0min for initial screening. Blood films were examined by two microscopists and all slides were read independently. When the patient was enroled, and at subsequent follow-up visits, thick and thin blood smears were prepared on a single slide for parasite detection and species identification. Slides were stained by 3% Giemsa for 45\u00a0min, and then parasites were counted on thick films as the number of asexual parasites per 200 white blood cells . A smear was declared negative if no parasites were seen after 1000 white blood cells were counted. The presence of gametocytes at enrolment or during follow-up was recorded. Asexual parasite density per microlitre (\u03bcL) was calculated on the assumption of 8000 leucocytes per \u03bcL blood. All collected slides were cross-checked by WHO-certified microscopists  after the study. If the two parasitaemia readings were in agreement (difference in parasite densities\u2009<\u200950%), the average results were recorded. If the two counts were discordant in terms of parasite species or density by\u2009>\u200950%, then a third, independent microscopist re-examined the blood slides. For parasite positivity and species, two concordant results were considered the final result, while for parasite density, the average of the two closest estimates of parasitaemia was considered final.Haemoglobin was measured from finger prick blood samples using a portable spectrophotometer  on days 0, 14, 28, and 42.P. falciparum positive samples, seven neutral microsatellite markers  were analyzed as previously published [P. vivax samples, seven microsatellite markers  were genotyped using published protocols [In order to differentiate recrudescence from re-infection, blood samples were collected on filter paper from a finger-prick on day 0 and on the day of parasite recurrence (day 7 onwards) for genotyping. Specimens were dried, stored in individual plastic bags with desiccants and protected from light, humidity, and extreme temperatures. The samples were genotyped at the U.S. Centers for Disease Control and Prevention (CDC) in Atlanta, GA, USA. Each dried blood spot was punched with a sterile puncher and DNA from the punched spots were extracted using QIAamp\u00ae DNA Mini Kit . For ublished . Microsarotocols , 28. BacP. falciparum recurrence, in addition to 20% of randomly selected day zero samples, were analysed for polymorphisms in the pfk13 propeller domain and pfmdr1 by performing Sanger sequencing [. The pfK13 domain from codon positions 389\u2013649 was assessed for presence of mutations known to be associated with artemisinin resistance as recommended by the WHO  using a previously published method [pfmdr1 gene that are associated with resistance to different anti-malarial drugs were analysed at codons N86Y, Y184F, S1034C, N1042D, D1246Y as reported previously [All samples of quencing . The pfKd method \u201332. FiveAssuming an ACPR rate of 95% and 95% confidence interval and 5% precision, a total of 73 patients per arm were calculated. Factoring in 20% loss to follow-up and withdrawals, 88 patients per each arm for a total of 352 patients across the four arms were targeted.SAS 9.3  and the WHO Excel-based data entry and analysis tool were used for analysis . Data weThe study protocol was approved by the Ethiopian Public Health Institute (EPHI), the National Ethical Committee in Ethiopia (3.10/171/2016) and FMHACA. In addition, the study was reviewed and approved by the Institutional Review Boards (IRBs) of Columbia University and the U.S. CDC (#6892). Written consent and/or assent was obtained from study participants or their guardians.P. falciparum or P. vivax fulfilled the inclusion criteria. As the number of cases per each site was not sufficient for per site analysis, data was pooled from the two study sites . One hundred and eighty-one P. falciparum-infected patients were enroled in AL (n\u2009=\u2009106) and DHA/PPQ (n\u2009=\u200975) arms. One hundred and ninety-eight P. vivax-infected patients were enroled in the CQ (n\u2009=\u2009142) and DHA/PPQ (n\u2009=\u200956) arms. Most P. falciparum infected patients were enroled in Arbaminch (n\u2009=\u2009156) and most P. vivax patients were enroled at Pawe (n\u2009=\u2009144) health centres. A total of 101 (95.3%) P. falciparum patients in the AL arm and 132 (93.0%) patients in the P. vivax-CQ arm reached the 28-day primary endpoint; 68 (90.7%) P. falciparum and 49 (87.5%) P. vivax patients in the DHA/PPQ arms completed the 42\u00a0days of follow up  in the P. falciparum-AL arm and 25\u00a0years (18\u201365) in the P. falciparum-DHA/PPQ arm. The median age was 14 (1\u201370) in the P. vivax-CQ arm and 23 (18\u201370) in the P. vivax-DHA/PPQ arm. Most of the study participants were male (56\u201371%) in all arms. Forty two percent of patients enroled had gametes: 77% (152/198) P. vivax-infected and 5% (9/181) P. falciparum-infected patients had gametes. The median day 0 gametocyte density for P. falciparum was zero per \u00b5l for both arms but with a range of 0\u20133,520 in the DHA/PPQ arm; for P. vivax, the median was 401 per \u00b5l  in the CQ arm and 320 per \u00b5l  in the DHA/PPQ arm. Median day zero haemoglobin levels were similar across the four arms  for AL and 100% for DHA/PPQ. The 42-day follow-up ACPR for DHA/PPQ was 100%. Two failures in the AL arm were observed, one on day 21 and one on day 28. Both failures were confirmed to be re-infection by PCR, one with a 33% and one with 11% probability of recrudescence. The 28-day follow-up PCR-corrected ACPR for CQ against P. vivax was 98% (95% CI: 94\u2013100), with the two PCR-corrected samples with probability of recrudescence above 99%. The 42-day follow-up ACPR for DHA/PPQ against P. vivax was 100% (95% CI 93\u2013100). The Kaplan\u2013Meier analysis is presented to show the censored estimates (Table The treatment outcomes for day 28 and 42 (limited to DHA/PPQ) are shown in Table P. falciparum arm and three (2%) asexual parasites on the CQ arm against P. vivax was observed on day two, that eventually cleared on day three. All patients cleared gametocytes. Two P. vivax-infected patients had gametocytes noted on day 28 in the CQ arm , and no resistance-related mutants were observed. One pfk13 non-synonymous mutation, E433D was observed in a day zero sample from Pawe. In the pfmdr1 gene, nine haplotypes were assessed, and the following mutations were observed: in one sample N86Y (2%), in two samples Y184Y/F (4%), in 35 samples Y184F (73%), and in one sample S1034N (2%). In total, 24/30 (80%) samples showed pfmdr1 mutant haplotypes in the P. falciparum-AL arm and 15/20 (75%) samples in the P. falciparum-DHA/PPQ arm. Prevalence and polymorphism of resistance markers are shown in Table A total of 50 Day 0 samples, 30 from the P. falciparum, and CQ and DHA/PPQ against uncomplicated P. vivax. All the study drugs showed high efficacy within their respective follow-up period, demonstrating that the first-line species-specific malaria treatments are efficacious in the study sites of Ethiopia. The PCR-corrected per-protocol analysis demonstrated 100% efficacy of AL against P. falciparum for the 28-day follow-up outcome and a 100% efficacy of DHA/PPQ against P. falciparum in the 42-day follow-up outcome. In a similar fashion, the PCR-corrected per-protocol analysis demonstrated 98% efficacy of CQ against P. vivax for the 28-day follow-up outcome, and 100% efficacy of DHA/PPQ for the 42-day follow-up outcome.The current study reported the therapeutic efficacy of AL and DHA/PPQ against uncomplicated P. falciparum, confirming the drug remains well-tolerated and effective for the intended use in Ethiopia [P. falciparum and P. vivax from elsewhere and the extended protection to day 42 attributed to piperaquine\u2019s longer half-life [The high efficacy reported for AL and CQ was consistent with previous studies in Ethiopia. Studies conducted since 2006 by EPHI and others have shown a high efficacy of AL against Ethiopia , 34, 35.alf-life \u201325.P. falciparum and P. vivax DHA/PPQ arms up to 42-day follow-up by microscopy, contrasting with other data indicating that piperaquine may encourage gametocyte production when provided as mono-therapy [Asexual and sexual parasites and fever were cleared by day three in all arms, reaffirming the sensitivity of the parasites circulating in the population to the respective drugs, especially the artesiminin component . Gametoc-therapy . As low -therapy \u201339. The -therapy , 40. AllP. falciparum samples sequenced for the PfK13 gene, none had a mutation associated with artemisinin resistance. These results were in contrast with previous studies that showed the presence of the k13 markers in Ethiopia and elsewhere in Africa [Pfmdr1 haplotypes have been associated with CQ, mefloquine, quinine, other quinolones, and/or artemisinin resistance [pfmdr1 markers. The markers were assumed to be related with piperaquine, suggesting the need for continuous molecular surveillance for anti-malarial resistance [Among the 50 n Africa , 41, 42.n Africa , 42. R56sistance , 31. A csistance .Chloroquine resistance against P. vivax has been reported in Ethiopia and in Southeast Asia , 34, 42.P. falciparum or P. vivax [P. falciparum in neighbouring Somalia and Eritrea [P. falciparum and P. vivax, as well as the long half-life, makes DHA/PPQ a good option for treatment and chemoprevention strategies in malaria elimination settings in Ethiopia and elsewhere [The high efficacy of DHA/PPQ has been reported in numerous African countries and Southeast Asia. Although only studied in adult subjects, this report is consistent with other studies showing high efficacy and no serious adverse events against either P. vivax . DHA/PPQP. vivax , 46. Non Eritrea , 24. Thelsewhere .P. falciparum-infected patients were reduced to 500 parasites/ul in order to increase enrolment rates; however, only nine patients had parasitaemia levels between 500 and 1000 parasites/ul and likely did not affect the efficacy outcomes greatly. The study only enroled adult patients (18 years and above) for the DHA/PPQ arm which resulted in lower baseline parasitaemia in the DHA/PPQ arms, increased median age for the study overall, lower enrolment and limited generalizability to younger age groups. The study was powered for efficacy outcomes per arm and cannot be used to compare results between arms. In addition, insufficient numbers of patients were enroled in a given health centre necessitating pooling of data from two study sites which resulted in not being able to provide site-specific efficacy estimates. Though sample size was not achieved, site-specific outcomes have been included which showed no major difference from the pooled analysis for the primary outcomes  over the 42\u00a0days of follow up for both P. falciparum and P. vivax supports the potential use of DHA/PPQ as an additional option for the treatment and chemoprevention of both P. falciparum and P. vivax in Ethiopia. Further comparisons investigating duration of protection and cost- effectiveness between DHA/PPQ, AL, and CQ in Ethiopia are warranted.This study demonstrated high therapeutic efficacy for the anti-malarial drugs currently used by the malaria control programme in Ethiopia and provides information on the efficacy of DHA/PPQ for the treatment of The findings and conclusions in this presentation are those of the authors and do not necessarily represent the official position of the U.S. Centers for Disease Control and Prevention or the U.S. Agency for International Development.Use of trade names is for identification only and does not imply endorsement by the U.S. Centers for Disease Control and Prevention/the Agency for Toxic Substances and Disease Registry, the U.S. Public Health Service, or the U.S. Department of Health and Human Services.Additional file 1. Microsatellite genotyping for determining recrudescence andreinfection.Additional file 2: Table S1. study profile and characteristics by site. Table S2. Treatment outcomes per site. Table S3. Proportion of slides negative for asexual parasites on day 2 and 3 per site. Table S4. Treatment outcome per site."}
+{"text": "Plasmodium vivax infection, but its Plasmodium falciparum transmission-blocking efficacy is unclear. We aimed to establish the efficacy and safety of three single low doses of tafenoquine in combination with dihydroartemisinin\u2013piperaquine for reducing gametocyte density and transmission to mosquitoes.Tafenoquine was recently approved as a prophylaxis and radical cure for P falciparum microscopy-detected gametocyte carriage, had a bodyweight of 80 kg or less, and had no clinical signs of malaria defined by fever. Participants were randomly assigned (1:1:1:1) to standard treatment with dihydroartemisinin\u2013piperaquine, or dihydroartemisinin\u2013piperaquine plus a single dose of tafenoquine (in solution) at a final dosage of 0\u00b742 mg/kg, 0\u00b783 mg/kg, or 1\u00b766 mg/kg. Randomisation was done with a computer-generated randomisation list and concealed with sealed, opaque envelopes. Dihydroartemisinin\u2013piperaquine was administered as oral tablets over 3 days , as per manufacturer instructions. A single dose of tafenoquine was administered as oral solution on day 0 in parallel with the first dose of dihydroartemisinin\u2013piperaquine. Tafenoquine dosing was based on bodyweight to standardise efficacy and risk variance. The primary endpoint, assessed in the per-protocol population, was median percentage change in mosquito infection rate 7 days after treatment compared with baseline. Safety endpoints included frequency and incidence of adverse events. The final follow-up visit was on Dec 23, 2021; the trial is registered with ClinicalTrials.gov, NCT04609098.In this four-arm, single-blind, phase 2, randomised controlled trial, participants were recruited at the Clinical Research Unit of the Malaria Research and Training Centre of the University of Bamako in Mali. Eligible participants were aged 12\u201350 years, with asymptomatic From Oct 29 to Nov 25, 2020, 1091 individuals were screened for eligibility, 80 of whom were enrolled and randomly assigned (20 per treatment group). Before treatment, 53 (66%) individuals were infectious to mosquitoes, infecting median 12\u00b750% of mosquitoes (IQR 3\u00b764\u201335\u00b700). Within-group reduction in mosquito infection rate on day 7 was 79\u00b795%  following dihydroartemisinin\u2013piperaquine only, 100%  following dihydroartemisinin\u2013piperaquine plus tafenoquine 0\u00b742 mg/kg, 100%  following dihydroartemisinin\u2013piperaquine plus tafenoquine 0\u00b783 mg/kg, and 100%  following dihydroartemisinin\u2013piperaquine plus tafenoquine 1\u00b766 mg/kg. 55 (69%) of 80 participants had a total of 94 adverse events over the course of the trial; 86 (92%) adverse events were categorised as mild, seven (7%) as moderate, and one (1%) as severe. The most common treatment-related adverse event was mild or moderate headache, which occurred in 15 (19%) participants . No serious adverse events occurred. No significant differences in the incidence of all adverse events (p=0\u00b773) or treatment-related adverse events (p=0\u00b762) were observed between treatment groups.P falciparum gametocyte clearance. All tafenoquine doses showed improved transmission reduction at day 7 compared with dihydroartemisinin\u2013piperaquine alone. These data support the case for further research on tafenoquine as a transmission-blocking supplement to standard antimalarials.Tafenoquine was well tolerated at all doses and accelerated Bill & Melinda Gates Foundation.For the French, Portuguese, Spanish and Swahili translations of the abstract see Supplementary Materials section. Evidence before this studyPlasmodium vivax liver stages and thereby form an essential component of P vivax radical cure. Primaquine is also a potent Plasmodium falciparum gametocytocide; a single low dose of primaquine combined with standard artemisinin-combination therapy (ACT) prevents transmission by rapidly sterilising and killing P falciparum gametocytes. The potency of tafenoquine as a P falciparum gametocytocide has so far remained unstudied. We searched PubMed on June 22, 2021, with no restrictions on publication date or language, for studies assessing the transmission-blocking and gametocytocidal abilities of tafenoquine treatment with search terms: ([Tafenoquine] OR [Krintafel] OR [Arakoda] OR [WR-238605]) AND . Among the 16 results (1992\u20132019) meeting the search criteria, none of the identified studies assessed the P falciparum gametocytocidal and transmission-blocking properties of tafenoquine in naturally infected gametocyte carriers. The results consisted of the following: four reviews; two studies assessing the sporontocidal activity of tafenoquine against P vivax; one study assessing glucose-6-phosphate dehydrogenase deficiency prevalence in Ethiopia; one study comparing in-vitro sensitivity of P falciparum isolates against three 8-aminoquinolines; one study investigating drug interactions between tafenoquine and ACTs; two articles regarding the development of tafenoquine; one study in which the prophylactic efficacy of tafenoquine was studied in a human P falciparum challenge model; and four studies wherein gametocytocidal and sporontocidal activity of tafenoquine was tested in avian and rodent models.Primaquine and its long-acting analogue, tafenoquine, belong to the class of 8-aminoquinolines that can clear Added value of this studyPlasmodium. The long-acting 8-aminoquinoline tafenoquine might have unique properties in blocking the transmission of malaria over a prolonged period and possibly multiple infections. To our knowledge, this is the first clinical trial assessing the P falciparum gametocytocidal and transmission-blocking properties of single low doses of tafenoquine. Three different single low doses of tafenoquine were combined with dihydroartemisinin\u2013piperaquine. The recommended dose of tafenoquine for radical cure of P vivax is 300 mg . The maximum dose used in the current study was one third of this recommended dose . The results show that tafenoquine accelerated P falciparum gametocyte clearance in a dose-dependent manner and greatly reduced transmission. Although findings from previous studies suggest that primaquine's efficacy is evident within the first 48 h after administration, tafenoquine's transmission-blocking occurred later; first detected on day 7 after initiation of treatment. These data support the case for further research on tafenoquine as a transmission-blocking supplement to standard antimalarials.Transmission-blocking antimalarials are important components of strategies to eliminate malaria and prevent the spread of drug-resistant Implications of all the available evidenceP falciparum gametocytocide.This study provides the first evidence that tafenoquine is gametocytocidal in humans and prevents transmission to mosquitoes by day 7 after treatment. Some individuals treated with dihydroartemisinin\u2013piperaquine alone were still infectious to mosquitoes up to the last day of mosquito feeding assays (day 14 after treatment), corroborating earlier reports on the persistence of transmissible gametocytes after artemisinin-combination therapy. By contrast, none of the individuals who received tafenoquine were infectious by day 14 and the majority of transmission events were prevented by day 7. The current findings highlight the importance of gametocytocidal and transmission-blocking supplements in areas trying to reduce the malaria infectious reservoir and identify tafenoquine as a new Plasmodium falciparum malaria, there is a clear need for interventions focused on reducing malaria parasite transmission.After years of success in malaria control, efforts to further reduce the global incidence of malaria are stalled. There were an estimated 229 million cases of malaria worldwide in 2019, up from 217 million in 2014.P falciparum asexual parasites but have little activity against mature gametocytes. Gametocytes are the only Plasmodium life stage that can be transmitted to mosquitoes, so their prevalence, density, and distribution within the human population define the malaria infectious reservoir. In areas consolidating malaria elimination or aiming to contain the spread of artemisinin resistance, WHO recommends that ACTs be combined with a single low dose of the 8-aminoquinoline compound, primaquine.Plasmodium vivaxP falciparum gametocytocide is suggested by in-vitroP vivax malaria with primaquine. Recent trials indicate that a single dose of 300 mg tafenoquine is considered safe among individuals with normal G6PD production and has equivalent transient haemolytic activity to standard 14-day primaquine regimens for P vivax treatment in women who are deficient in G6PD.Artemisinin combination therapies (ACTs) clear P falciparum gametocytes. Dosing for a gametocytocidal indication has not been assessed but is likely to be substantially lower than the required dose for radical cure of P vivax (as is the case with primaquine). In the current study, we assessed the gametocytocidal and transmission-blocking efficacy of a range of single low doses of tafenoquine  in combination with dihydroartemisinin\u2013piperaquine in Malian children and adults with normal G6PD production.Tafenoquine has not yet been tested in controlled trials for its ability to clear and prevent transmission of P falciparum gametocytes by microscopy ; absence of other non-P falciparum species on blood film; haemoglobin density of 10 g/dL or more; normal G6PD ; aged between 12\u201350 years; bodyweight 80 kg or less; no clinical signs of malaria defined by fever (\u226537\u00b75\u00b0C); no signs of acute, severe, or chronic disease; no allergies to any study drugs; reported no use of antimalarial drugs over the past week; consistent with the long half-life of tafenoquine, use of effective contraception for five half-lives (3 months) after the end of tafenoquine treatment. Exclusion criteria included pregnancy (tested at enrolment by urine and serum test) or lactation, use of other medication (except for paracetamol or aspirin), family history of congenital prolongation of the corrected QT interval, current or recent treatment with drugs known to extend the corrected QT interval, and blood transfusion in the past 90 days. Before screening and before study enrolment, participants provided written informed consent (aged \u226518 years) or assent with written parental consent (aged 12\u201317 years).This four-arm, single-blind, phase 2, randomised controlled trial was conducted at the Clinical Research Unit of the Malaria Research and Training Centre (MRTC) of the University of Bamako in Mali. Before the commencement of screening, our study team of clinicians and technicians met with community leaders, village health workers, and heads of households from each village to explain the study and obtain approval to conduct the study. Village health workers then used a door-to-door approach to inform households of the date and location where consenting and screening would take place. Participants were included in the trial if they met the following criteria: positive for Ethical approval was granted by the Ethics Committee of the Faculty of Medicine, Pharmacy, and Dentistry of the University of Science, Techniques, and Technologies of Bamako , and the Research Ethics Committee of the London School of Hygiene & Tropical Medicine .Participants were randomly assigned (1:1:1:1) to receive dihydroartemisinin\u2013piperaquine plus tafenoquine 1\u00b766 mg/kg, dihydroartemisinin\u2013piperaquine plus tafenoquine 0\u00b783 mg/kg, dihydroartemisinin\u2013piperaquine plus tafenoquine 0\u00b742 mg/kg, or dihydroartemisinin\u2013piperaquine alone. Enrolment continued until 80 participants were enrolled . An independent MRTC statistician randomly generated the treatment assignment using Stata (version 16), which was linked to participant identification number. The statistician prepared sealed, opaque envelopes with the participant identification number on the outside and treatment assignment inside, which were sent to the MRTC study pharmacist. The study pharmacist provided treatment according to the contained assignment and was consequently not masked to treatment assignment. All other investigators and staff involved in assessing all laboratory outcomes were masked. Participants could ask the study physician which treatment they received.P vivax is 300 mg .Dihydroartemisinin\u2013piperaquine was administered as oral tablets over 3 days , as per manufacturer instructions (see appendix 5 p 2 for dosing). A single dose of tafenoquine was administered as oral solution on day 0 in parallel with the first dose of dihydroartemisinin\u2013piperaquine. Tafenoquine dosing was based on bodyweight to standardise efficacy and risk variance . The maxAnopheles gambiae were allowed to feed for 15\u201320 min on venous blood samples  through a prewarmed glass membrane feeder system . All surviving mosquitoes were dissected on the seventh day after the feeding assay; midguts were stained in 1% mercurochrome and examined for the presence and density of oocysts by expert microscopists.Participants received a full clinical and parasitological examination on days 1, 2, 7, 14, 21, and 28 after receiving the first dose of the study drugs. Giemsa stained thick film microscopy was performed as described previously, with asexual stages counted against 200 white blood cells and gametocytes counted against 500 white blood cells.The primary outcome measure was median percentage change in mosquito infection rate between pretreatment and 7 days after treatment. Secondary outcomes were mosquito infection metrics  at other prespecified timepoints ; gametocyte prevalence, density, circulation time, area under the curve (AUC) of density over time, and sex ratio ; and safety assessments including incidence of clinical and laboratory adverse events. Differences in all transmission, gametocyte, and safety outcomes were compared between treatment groups  as secondary outcomes.Adverse events were graded by the study clinician for severity  and relatedness to study medication . A reduction in haemoglobin concentration of 40% or more from baseline was categorised as a haematological adverse event. An external data safety and monitoring committee was assembled before the trial, and safety data were discussed after enrolment of 40 participants, and after the final follow-up visit of the last participant.Sample size estimation was based on efficacy for single-dose primaquine  to provide a 95% or greater reduction in infectivity at 7 days after initiation of treatment compared with pretreatment using a membrane feeding assay.All outcomes were analysed in the per-protocol population. Mosquito infectivity was assessed at three levels: mean number of oocysts in a sample of mosquitoes , the proportion of mosquitoes infected with any number of oocysts , and infectivity of the study participant to any number of mosquitoes . Asexual parasite density was not measured by molecular methods due to the scarcity of nucleic acid extraction reagents.t tests (t score) for within-group analyses and linear regression adjusted for baseline levels of each measure for between-group analyses . Percentage change from baseline was analysed using two-way t tests. The proportion of gametocytes that were male was analysed for all values with total gametocyte densities of 0\u00b72 gametocytes per \u03bcL or more.t tests (t score), and the dose effect of tafenoquine on circulation time was determined using linear regression analysis. AUC of gametocyte density per participant over time was calculated using the linear trapezoid method,10 adjusted AUC values, with adjustment for baseline gametocyte density . All other analyses of quantitative data were done using Wilcoxon sign rank tests (z-score) and Wilcoxon rank-sum tests (z-score). All comparisons were defined before study completion and analyses were not adjusted for multiple comparisons. For all analyses, the threshold for statistical significance was set at p<0\u00b705. Statistical analysis was conducted using STATA (version 16.0) and SAS (version 9.4). The trial is registered with ClinicalTrials.gov, NCT04609098.Mosquito infection rate and oocyst density were analysed at timepoints after baseline only for those individuals who were infectious at baseline. The prevalence of gametocytes and infectious individuals were compared within and between treatment groups using one-sided Fishers exact tests. Haemoglobin and methaemoglobin levels were compared using paired The funder of the study had no role in study design, data collection, data analysis, data interpretation, or writing of the report.Between Oct 29 and Nov 25, 2020, 1091 individuals were screened for eligibility, 80 of whom were enrolled and randomly assigned  of 80 individuals completing this study visit (one in the dihydroartemisinin\u2013piperaquine plus tafenoquine 0\u00b742 mg/kg group did not complete the visit). 76 (95%) participants completed all visits to day 28 . The median number of mosquitoes dissected in an individual mosquito feeding experiment was 55 (IQR 45\u201362). Before treatment, 53 (66%) individuals were infectious to mosquitoes, with a median of 12\u00b750% (IQR 3\u00b764\u201335\u00b700)of mosquitoes becoming infected . The proGametocyte densities decreased after initiation of treatment in all study groups, although the decrease was more rapid in the tafenoquine groups than in the dihydroartemisinin\u2013piperaquine-only group . Total gThere was a small but statistically significant within-group reduction in haemoglobin density in the dihydroartemisinin\u2013piperaquine-only and dihydroartemisinin\u2013piperaquine plus tafenoquine 0\u00b783 mg/kg groups at days 2 and 7 compared with baseline, with no significant reductions in any other treatment group or at any other timepoint . The greOverall, 55 (69%) of 80 participants had a total of 94 adverse events during follow-up, of which 86 (92%) were categorised as mild, seven (7%) as moderate, and one (1%) as severe . The sinP falciparum gametocytocidal and transmission-blocking properties of single low doses of tafenoquine. By day 7, post-treatment transmission potential was greatly reduced in the 0\u00b742 and 0\u00b783 mg/kg tafenoquine dose groups, and completely annulled in individuals given the highest (1\u00b766 mg/kg) tafenoquine dose.To our knowledge, this was the first clinical trial specifically designed to determine the P falciparum gametocytocidal activity of tafenoquine.P vivax in which tafenoquine had a longer time to gametocyte and parasite clearance compared with chloroquine plus primaquine.Plasmodium gallinaceum model in which transmission to mosquitoes persisted for 4 days after initiation of tafenoquine monotherapy treatment and declined thereafter.P vivax relapsesACTs differ in their gametocytocidal properties.Despite not having a direct comparison to dihydroartemisinin\u2013piperaquine plus single low-dose primaquine, the current study suggests inferior transmission-blocking properties of the tested doses of tafenoquine compared with single low-dose primaquine tested in similar populations in multiple recent studies. Nonetheless, tafenoquine's long half-life could be a major advantage. Depending on the duration of its gametocytocidal activity, tafenoquine might play a role in preventing transmission of infections that are acquired after initiation of treatment. A long-acting gametocytocide could also be of relevance to prevent the transmission of drug-resistant parasites. Treatment failure has been associated with increased appearance of gametocytes after initiation of treatment,P vivax treatment trial indicate that that dihydroartemisinin\u2013piperaquine might inhibit tafenoquine activity against P vivax hypnozoites.P vivax are only recommended for use with chloroquine.P falciparum, it seems logical that single low-dose tafenoquine will need to be combined with ACT. If the safety of these tafenoquine doses in G6PD mixed populations can be demonstrated, our data indicate this combination will be effective at clearing asexual parasites and transmission blockade.Whether low-dose gametocytocides are included in standard treatment or mass administrations, the choice of partner schizonticide requires careful consideration. The current study shows that dihydroartemisinin\u2013piperaquine plus single low-dose tafenoquine has substantial gametocytocidal activity at doses of 1\u00b766 mg/kg, but it is noteworthy that preliminary observations from an unpublished P vivax malaria and G6PD enzyme activity of at least 70% in Columbia and Vietnam.To be used widely, tafenoquine's safety is a key consideration. We observed no increased number of adverse events in tafenoquine treatment groups compared with dihydroartemisinin\u2013piperaquine-only. Non-symptomatic transient increases in alanine aminotransferase were observed in two individuals who received the lowest tafenoquine dose; this might be associated with general infection-related liver-injury that is treatment independent.P falciparum gametocytocidal activity. A design with a follow-up over multiple malaria episodes or in which tafenoquine is administered without an efficacious schizonticidal drug would allow assessment of the duration of tafenoquine's activity. With increasing evidence for the transmission-blocking effects of primaquine and tafenoquine in individual gametocyte carriers, there is an urgent need for community trials to establish the added value of these gametocytocides in first-line antimalarial treatment and in mass treatment campaigns to reduce the transmission of (drug-resistant) malaria.In the current study, we established the short-term effect of tafenoquine on transmission from highly infectious individuals. This population allows a detailed assessment of tafenoquine's transmission-blocking properties with high discriminative power. Our study shows that a dose of 1\u00b766 mg/kg tafenoquine blocks transmission by day 7 but this effect might have been apparent at earlier moments when we did not do feeding experiments (days 3\u20137). In addition, our study provides no evidence for the duration of tafenoquine's transmission blocking activity. Future trials are required to examine the timing of tafenoquine activity with respect to both the initiation and duration of its Anonymised data reported in the manuscript will be made available to investigators who provide a methodologically sound proposal to the corresponding author. The protocol is available upon request.We declare no competing interests."}
+{"text": "Plasmodium falciparum malaria in Liberia. Intermittent preventive treatment with sulfadoxine/pyrimethamine is also recommended for pregnant women. The therapeutic efficacy of Artesunate\u2013amodiaquine and Artemether\u2013lumefantrine, and the frequency of molecular markers associated with anti-malarial drug resistance were investigated.Artesunate\u2013amodiaquine (ASAQ) and Artemether\u2013lumefantrine (AL) are the recommended treatment for uncomplicated https://www.who.int/malaria/publications/atoz/9789241597531/en/). Eligible children were recruited and monitored clinically and parasitologically for 28\u00a0days. Polymorphisms in the Pfkelch 13, chloroquine resistance transporter (Pfcrt), multidrug resistance 1 (Pfmdr-1), dihydrofolate reductase (Pfdhfr), and dihydropteroate synthase (Pfdhps) genes and copy number variations in the plasmepsin-2 (Pfpm2) gene were assessed in pretreatment samples.The therapeutic efficacy of ASAQ and AL was evaluated using the standard World Health Organization protocol , while the Pfmdr-1 86Y  and 184F  mutations were less frequent. The Pfdhfr triple mutant (51I/59R/108\u00a0N) was the predominant allele (49.2%). For the Pfdhps gene, it was the 540E mutant (16.0%), and the 436A mutant (14.3%). The quintuple allele (51I/59R/108\u00a0N-437G/540E) was detected in only one isolate (1/357).Of the 359 children enrolled, 180 were treated with ASAQ (89 in Saclepea and 91 in Bensonville) and 179 with AL (90 in Sinje and 89 in Kakata). Of the recruited children, 332 (92.5%) reached study endpoints. PCR-corrected per-protocol analysis showed ACPR of 90.2% (95% CI: 78.6\u201396.7%) in Bensonville and 92.7% (95% CI: 83.4.8\u201396.5%) in Saclepea for ASAQ, while ACPR of 100% was observed in Kakata and Sinje for AL. In both treatment groups, only two patients had parasites on day 3. No artemisinin resistance associated Pfcrt and Pfmdr-1 and the risk of parasite recrudescence in patients treated with ASAQ was observed. Parasites with signatures known to be associated with artemisinin and piperaquine resistance were not detected. The very low frequency of the quintuple Pfdhfr/Pfdhps mutant haplotype supports the continued use of SP for IPTp. Monitoring of efficacy and resistance markers of routinely used anti-malarials is necessary to inform malaria treatment policy.This study reports a decline in the efficacy of ASAQ treatment, while AL remained highly effective, supporting the recent decision by NMCP to replace ASAQ with AL as first-line treatment for uncomplicated falciparum malaria. No association between the presence of the mutations in Trial registration ACTRN12617001064392.The online version contains supplementary material available at 10.1186/s12936-022-04140-7. Malaria is a major health problem, with an estimated 241 million cases and 627,000 deaths worldwide, increasing from 227 million and 558,000 cases and deaths, respectively, in 2020 . Howeveri.e., genetic changes in Plasmodium falciparum genome), associated with anti-malarial drug resistance are complementary tools. Partial artemisinin resistance, defined as delayed parasite clearance following artesunate monotherapy or ACT [P. falciparum kelch13 (Pfkelch13) gene have been demonstrated to be a major determinant associated with artemisinin resistance [Pfkelch13 mutations have been detected worldwide, but only 21 have been validated  or are suspected to be associated  with partial artemisinin resistance [Pfkelch13 mutations known to be associated with artemisinin resistance in Africa has historically been rare and sporadic [Pfkelch13 mutants (R561H in Rwanda and A675V and C469Y in Uganda) associated with delayed parasite clearance and in vitro artemisinin resistance [In addition to TES, molecular markers  gene, a marker of piperaquine resistance [Pfpm2 gene:\u2009<\u20095% in Mozambique [Pfpm2 in polyclonal infections (MOI\u2009>\u20091) are challenging.\u00a0Single Nucleotide Polymorphisms (SNP) in the Plasmodium chloroquine resistance transporter (Pfcrt) and Plasmodium multi drug resistance 1 (Pfmdr-1) have been suspected to be associated with resistance to ACT partner drugs, lumefantrine and amodiaquine [High prevalence of multiple copies of the sistance , has beesistance \u201318. Studzambique , 30.5% azambique , and up diaquine , 22, butdiaquine .Pfdhfr)\u00a0and\u00a0dihydropteroate synthase (Pfdhps) genes increases the risk of SP treatment failure [Pfdhfr and 437G\u2009+\u2009540E in the Pfdhps genes) is a significant predictor of SP treatment failure [Pfdhps (A581G) on the genetic background of the quintuple mutant has been shown to confer a higher degree of resistance to SP [Sulfadoxine/pyrimethamine (SP) is the recommended drug for intermittent preventive treatment of pregnant women (IPTp) living in areas of moderate to high malaria transmission in Africa to prevent the deleterious effects of malaria on maternal and fetal outcomes . Accumul failure .\u00a0A quint failure , 27. Acqce to SP  and is ace to SP , 30 and ce to SP  when itsP. falciparum malaria [Pfkelch13, Pfcrt, Pfmdr-1, Pfdhfr, Pfdhps) and CNV (Pfpm2) associated or suspected to be associated with resistances to anti-malarial drugs.In Liberia, malaria transmission is perennial with an estimated 1,809,994 cases and 2,232 deaths in 2019 . ASAQ wa malaria . Current malaria . Since t malaria . To provStudy patients were recruited from four health facilities in four counties: (i) Bensonville Hospital, Bensonville, Montserrado County; (ii) Saclepea Comprehensive Health Center, Saclepea, Nimba County; (iii) Charles Henry Rennie Hospital, Kakata, Margibi County; and (iv) Sinje Health Center, Garwula, Grand Cape Mount County Fig.\u00a0. The effP. falciparum mono-infection with parasitaemia of 2000 to 200,000 asexual parasites/\u00b5l by microscopy. Other inclusion criteria included willingness to comply with the study visit schedule and informed consent from parents or guardians. Children with exclusion criteria, including the presence of general danger signs or signs of severe falciparum malaria, mixed or mono-infection with non-falciparum species, or severe malnutrition and febrile conditions due to diseases other than malaria, received appropriate care and treatment according to national guidelines.Potential study children who visited the study health facilities between December 2017 and May 2018 were screened for the following criteria: age 6\u201359\u00a0months, axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C and/or history of fever in the past 24\u00a0h, and Children recruited at the Bensonville and Saclepea sites received daily dose of ASAQ for 3\u00a0days according to the recommended weight bands: one tablet of 25 artesunate\u2009+\u200967.5 amodiaquine for 4.5 to\u2009<\u20099\u00a0kg body weight, one tablet of 50 artesunate\u2009+\u2009135 amodiaquine for 9\u2013\u2009<\u200918\u00a0kg body weight, and one tablet of 100 artesunate\u2009+\u2009270 amodiaquine for 18 to\u2009<\u200936\u00a0kg body weight. A dose of artesunate 4 (range 2\u201310) mg/kg\u2009+\u2009amodiaquine 10 (range 7.5\u201315) mg/kg body weight once daily for 3\u00a0days was the target. Children from Kakata and Sinje were given twice daily dose of AL for 3\u00a0days according to recommended weight bands: one tablet for those weighing 5\u201314\u00a0kg and two tablets for 15\u201324\u00a0kg. AL was given with milk or fatty meal. All treatments were administered under direct observation by the study team and patients were observed for 30\u00a0min. If the first dose was vomited, the treatment was administered again. If vomiting recurred, the patient was given artesunate injection according to national guidelines and the patient was withdrawn from the study. Patient with treatment failure were treated with quinine 10\u00a0mg/kg BW three times a day for seven days. Prequalified ASAQ  were obtained from WHO /HQ.Study children were followed for up to 28\u00a0days at scheduled visits on days 1, 2, 3, 7, 14, 21, and 28, and at unscheduled visits when symptoms worsened or recurred. The allowable time window for the weekly follow-up was\u2009\u00b1\u20091\u00a0day. A clinical assessment and parasitological examination were performed at each visit. Existing general Community Health Volunteers (gCHVs) in the study areas were engaged and trained to trace patients if they did not show up for the scheduled appointment.Thick and thin blood smears collected on the day of recruitment (day 0) and follow-up days were stained with Giemsa and asexual parasites were counted against White Blood Cells (WBC) in the thick blood smear using the WHO procedure . Assuminmsp1, msp2 and glurp [msp1/msp2 and\u2009\u00b1\u200920\u00a0bp for glurp. The genotypes of parasites on day 0 and on the day of parasite recurrence were compared to distinguish recrudescence (same genotype) from new infection (different genotype). Both the current WHO-recommended algorithm [Filter paper blood samples were collected from each patient on day 0 and on the day of parasite recurrence (from day 7 onward), stored in individual plastic bags with desiccant and protected from light, moisture and extreme temperature until analysis. Each dried blood spot was cut out sterilely and placed in an Eppendorf tube. DNA was extracted using the QIAamp DNA Blood Mini Kit (Qiagen) as previously described . Paired nd glurp . The fralgorithm  and the lgorithm  were usePfkelch13 gene associated with artemisinin resistance [Pfcrt and Pfmdr-1 genes associated or suspected to be associated with 4-aminoquinolines and aminoalcohol resistance [Pfdhfr and Pfdhps genes linked to pyrimethamine and sulfadoxine resistance. Amplicons from targeted sequences were generated using nested PCR assays as previously described [Pfkelch13, at codons 72\u201376, 93, 97, 145, 218, 343, 350 and 353 for Pfcrt, at codons 86, 184, 1034, 1042 and 1246 for Pfmdr-1, at codons 51, 59, 108, 164 for Pfdhfr and at codons 436, 437, 540, 581, 613 for Pfdhps were assessed with the CLC Main Workbench 20 software (Qiagen). Electropherograms with mixed alleles were considered as mutant for the purpose of mutation frequency estimation. Quality control was assessed by including blinded quality control samples  in each 96-well sequencing plate. DNA from D0 samples were also analysed to estimate copy number variations in the Pfpm2 gene, which is associated with piperaquine resistance, using the method described previously [Pfpm2 gene copy number was estimated samples without assessing the number of clones (MOI) and a Pfpm2 copy number\u2009>\u20091.5 was defined as an amplification of the gene. All D0 samples were screened for Pfkelch13 and Pfdhfr/Pfdhps mutations, while a subsample of 30% from each site was analysed for Pfcrt and Pfmdr-1 mutations and Pfpm2 copy number variations. The samples for genotyping and molecular markers were analysed at the Institut Pasteur, Paris, France.Day 0 DNA was also analysed for the presence of point mutations in the sistance , the Pfcsistance , and theescribed , 41 and eviously . Pfpm2 gPfkelch13, Pfdhfr, Pfdhps, Pfcrt, Pfmdr-1, and CNV in Pfmp2 genes.Treatment response was classified based on parasitological and clinical response as recommended by WHO : early tEthical approval for the study protocol was obtained from the University of Liberia-Pacific Institute for Research & Evaluation Institutional Review Board (UL-PIRE IRB) and the WHO Research Ethics Review Committee (ERC.0002892). Parents/guardians were informed of the study procedure, its benefits and potential risks and gave written informed consent for their children to participate in the study prior to enrollment.http://www.who.int/malaria/publications/atoz/9789241597531/en/). Per-protocol and Kaplan\u2013Meier analyses were used to analyse treatment outcomes according to the WHO protocol [Pfcrt K76T, Pfmdr1 N86Y andY184F mutations was compared between day 0 samples from cured and recrudescent patients using Fisher's exact test and an estimation of relative risk. A difference is considered significant if the p-value\u2009<\u2009is 0.05.A minimum sample of 73 children per site was estimated based on a 5% treatment failure rate for ASAQ and AL and with a 95% confidence level and 5% precision. Twenty percent (n\u2009=\u200915) was added to account for loss to follow-up and withdrawal during the 28-day follow-up period. The final target sample was 88 per site. Data were double-entered, validated against the case sheet in case of discrepancies, and analysed using the WHO excel software programme  and 179 (89 in Kakata and 90 in Sinje) were recruited for the ASAQ and AL clinical efficacy studies, respectively. Baseline characteristics of the recruited children were comparable between the sites except for mean parasite density  reached the study endpoints. Of the remaining 27 cases, 24  were lost to follow-up and three (2 in Bensonville and 1 in Saclepea) were withdrawn during follow-up due to missing treatment dose for day-1 (one case) or for day-2 (two cases). Before PCR correction, per-protocol analysis revealed an ACPR of 63.9% (51.7\u201374.9%) and 86.4% (77.4\u201392.8%) for ASAQ treatment in Bensonville and Saclepea, respectively, while ACPR of 94.0% (86.7\u201398.0%) and 100% (95.9\u2013100%) were observed for AL in Kakata and Sinje, respectively  and one non-synonymous mutant (V637I in one sample at Saclepea).Of the 359 pretreatment samples, 358 (99.7%) yielded interpretable results, most of which (353/358) carried the Pfpm2 copy number was performed on 112 randomly selected day-0 samples (25% of day-0 samples per site). Interpretable data were obtained for 100 samples (89.3%). Samples that were not interpretable were likely samples with insufficient DNA quantity  or poor DNA quality. Parasites in all samples carried a single copy of the Pfpm2 gene (CNV\u2009<\u20091.5).Estimation of Pfcrt and Pfmdr-1 genes. These samples included all the day 0 isolates from patients classified as recurrent  and a random selection of day-0 samples from patients classified as cured  or lost to follow-up .A total of 91\u00a0day 0 samples from patients treated with AL or ASAQ were selected and used to assess the frequency of mutations in the Pfcrt 76\u00a0T mutant was highly frequent , with little variation in frequencies between sites (80.0% to 95.2%) , followed by the 86Y mutation . Across the study sites, the proportions of the two mutants were not much different  was observed. Pfmdr-1\u00a0N86 and 86Y were equally distributed in the AL group and none of the genotype was associated with treatment failure.Molecular analysis revealed that the Pfdhfr and Pfdhps genes were successfully amplified in 99.7% (358/359) and 99.4% (357/359), respectively. For the Pfdhfr gene, the triple mutant (51I/59R/108\u00a0N) was the predominant allele and accounted for 49.2%, with little variation between sites , followed by the 540E allele (16.0%), the 436A allele (14.3%), and the 436A/437G double mutant allele (9.0%). No significant differences were observed between sites. The 581G mutation, associated with loss of protective efficacy of IPTi and IPTp, was rare and observed in seven samples .\u00a0The most frequent Pfdhfr/Pfdhps haplotypes were 51I/59R/108\u00a0N-wild type (25.2%), followed by 108\u00a0N-wild type (14.8%) and wild type-wild-type (7.8%), as described in Table The  99.7% 35/359 and Using the WHO recommended PCR analysis , the curIn the current study, parasite genotyping to distinguish between reinfection and recrudescence (true failures) was analysed using both the WHO  and the In a recent WHO analysis, overall, the proportion of recurrent parasitsemia classified as recrudescence was higher with the 2/3 algorithm than with the WHO method (p\u2009<\u20090.001). However, this did not always translate into a significant difference in Kaplan\u2013Meier estimates of treatment outcome. Differences in the Kaplan\u2013Meier estimates of treatment outcome were more evident in areas of moderate to high transmission than in areas of low to moderate transmission in particular for artemether\u2013lumefantrine. Though there is no gold standard, an expert committee recommended that WHO methodology  should bPfkelch13 mutation known to be associated with artemisinin resistance, indicates absence of artemisinin resistance in Liberia. Until recently, Pfkelch13 mutations known to be associated with artemisinin resistance were rare or absent in Africa [Pfkelch13 R561H mutant was detected in 7.3% of samples collected from the Masaka site in Rwanda between 2013 and 2015, but without delayed parasite clearance [Pfkelch13 A675V or C469Y, candidate mutations, and prolonged parasite clearance half-life following artemisinin monotherapy was reported from Uganda [Pfkelch13 mutations in Africa, as recommended by the WHO [In the current study, parasites were cleared by day 3 in all but two patients, which, together with the absence of the n Africa , 62\u201365. learance . A subsem Uganda . These fPfpm2 gene were detected in the current study. CNV data, however, from polyclonal infections are not completely reliable. Indeed, it remains a challenge to assess copy number variations in multi-genome infections, as minor variants with amplified plasmepsin 2\u20133 maybe be missed. A recent study found a high frequency of multiple copies of the Pfpm2 gene in African samples, varying from 11 to 34% [Pfpm2 gene amplification should be closely monitored in Africa.Dihydroartemisinin\u2013piperaquine has recently been adopted as a second-line treatment for uncomplicated malaria and as a drug for Mass Drug Administration (MDA) in Africa. No parasites with multiple copies of the 1 to 34% . Given tPfcrt and Pfmdr-1 genes was observed, which could be explained by the predominant use of ASAQ in the country. However, no association between the presence of the mutations in Pfcrt and Pfmdr-1 and the risk of parasite recrudescence in patients treated with ASAQ was observed. Overall, this clearly points out that robust molecular markers associated with amodiaquine and lumefantrine are still lacking.A medium to high frequency of parasites carrying polymorphisms in the Pfdhfr and Pfdhps genes is an important tool to determine the status of SP resistance and guide IPTp policy. Although quintuple Pfdhfr/Pfhps mutations (N51I/C59R/S108N-A437G/K540E) have been associated with clinical SP treatment failure [Pfdhfr triple mutation (N51I/C59R/S108N) was the most common Pfdhfr allele detected, accounting for 49.2%. The very low frequency  of the quintuple mutant haplotype and the absence of the sextuple mutation (quintuple-581G) support the continued use of SP for IPTp in Liberia. Due to the high proportion of polyclonal infections detected by msp1/msp2/glurp genotyping, we cannot infer Pfdhfr/Pfdhps haplotypes with absolute certainty because the combination of SNPs could be deduced from different clones. The prevalence of Pfdhfr and Pfdhps mutations varies across the continent from absent to a low prevalence of quintuple mutations in West Africa [Pfdhfr and Pfdhps genes will continue to evolve to saturation under SP drug pressure in moderate to high transmission settings in Africa, where IPT with SP is recommended [Intermittent preventive treatment of malaria in pregnancy with SP (IPTp- SP) is one of the recommended core interventions in areas of moderate to high malaria transmission in Africa, including Liberia , 34. As  failure , 27, evi failure . However failure , 67. In t Africa \u201371 and at Africa \u201376. As eommended . TherefoPfcrt and Pfmdr-1 and the risk of parasite recrudescence in patients treated with ASAQ was observed. The very low frequency of the Pfdhfr/Pfdhps five-mutant haplotype supports the continued use of SP as an IPTp. The therapeutic efficacy of recommended artemisinin-based combination, molecular markers of resistance to artemisinin, partner drugs and SP should be closely monitored for early detection of resistant parasites and development of evidence-based malaria treatment and chemoprevention strategies.The findings of this study report a decline in the efficacy of ASAQ, while AL remains highly effective, supporting the recent decision by NMCP to replace ASAQ with AL as first-line treatment for uncomplicated falciparum malaria. There are no parasites carrying signatures known to be associated with artemisinin and piperaquine resistance. No association between the presence of the mutations in Additional file 1. Liberia genotyping raw data."}
+{"text": "Medicinal plants have been successfully used as an alternative source of drugs for the treatment of microbial diseases. Finding a novel treatment for malaria is still challenging, and various extracts from different wild desert plants have been reported to have multiple medicinal uses for human public health, this study evaluated the antimalarial efficacy of several Egyptian plant extracts.Plasmodium falciparum (3D7), and to treat infection with non-lethal Plasmodium yoelii 17XNL in an in vivo malaria model in BALB/c mice.We assessed the cytotoxic potential of 13 plant extracts and their abilities to inhibit the in vitro growth of Trichodesma africanum, Artemisia judaica, Cleome droserifolia, and Vachellia tortilis, with weak-to-moderate activity against P. falciparum erythrocytic blood stages with mean half-maximal inhibitory concentration 50 (IC50) of 11.7\u2009\u03bcg/ml, 20.0\u2009\u03bcg/ml, 32.1\u2009\u03bcg/ml, and 40.0\u2009\u03bcg/ml, respectively. Their selectivity index values were 35.2, 15.8, 11.5, and 13.8, respectively. Among these four candidates, T. africanum crude extract exhibited the highest parasite suppression in a murine malaria model against P. yoelii.In vitro screening identified four promising candidates, Our study identified novel natural antimalarial agents of plant origin that have potential for development into therapeutics for treating malaria.The online version contains supplementary material available at 10.1186/s12906-022-03566-5. Plasmodium. In 2019, approximately 229 million cases of malaria and 409,000 associated deaths were reported across 87 malaria-endemic countries ); they had respective median effective doses 50 (ED50s) of 30\u2009mg/kg/day and 72\u2009mg/kg/day at 4\u2009days post-infection and of 66\u2009mg/kg/day and 79\u2009mg/kg/day at 6\u2009days post-infection [A. judaica, C. droserifolia, T. africanum, or V. tortilis from 0 to 6\u2009days post-infection each caused significant parasite suppression, with mean suppression percentages ranges of 13.5\u201360.6%, 17.1\u201361.9, 35.2\u201365.5%, and 36.3\u201372.5%, respectively, despite the extract dose being lower compared with previously reported studies (Table SThree rodent-specific  in mice . Here, wpression . Anothernfection . In the P. falciparum in vitro and against the murine malaria-causing parasite P. yoelii in a mouse model. Although the administration of these extracts at a dose of 100\u2009mg/kg/day for a 7-day course of treatment did not achieve 100% inhibition of P. yoelii growth in BALB/c mice, the parasite suppression data suggests that these extracts may have potent antimalarial activity. Their efficacies are likely correlated with their multiple medicinal uses and their chemical constituents. Among the four tested candidates, the T. africanum crude extract possessed the highest parasite suppression ability in a short-term treatment course in vivo and had the highest IC50 in vitro against the human malaria-causing parasite P. falciparum, whereas V. tortilis extract showed moderate-to-weak effect against P. falciparum in vitro and induced partial inhibition against P. yoelii in vivo. These data support the use of these extracts in the future development of an antimalarial therapeutic. Further study will be needed to understand the mechanism of action and identify the main biological components of these crude extracts.This study showed that crude extracts of four wild plants collected from Egypt had antimalarial efficacy against the human malaria-causing parasite Additional file 1: Table S1. The plants used in this study and their reported medicinal uses. Table S2. Chemotherapeutic test of four plant extracts against the growth of Plasmodium yoelii in mice. Figure S1. Sampling map of the plant samples collected in Egypt. Figure S2. Images of the collected plant materials. Figure S3. Effect of wild plant extracts on the growth of plasmodium yoelii in male BALB/c mice. Figure S4.\u00a0Effect of wild plant extracts on bodyweight change in Plasmodium-infected mice. Figure S5. Effect of wild plant extracts on survival rate of Plasmodium-infected mice."}
+{"text": "Continuous monitoring of AL therapeutic efficacy is crucial in Ethiopia, as per the World Health Organization (WHO) recommendation. This study aimed to assess the therapeutic efficacy of AL in the treatment of uncomplicated P. falciparum infection.In 2004, Ethiopia adopted artemether-lumefantrine . The PCRcorrected cure rate was 98.6% (95% CI 92.3\u2013100). AL demonstrated a rapid parasite and fever clearance with no parasitaemia on day 2 and febrile cases on day 3. Gametocyte clearance was complete by day three. No serious adverse events were reported during the 28 days follow-up.P. falciparum malaria in Ethiopia. Periodic therapeutic efficacy studies and monitoring of markers of resistance are recommended for early detection of resistant parasites.The study demonstrated high therapeutic efficacy and good safety profile of AL. This suggests the continuation of AL as the first-line drug for the treatment of uncomplicated The online version contains supplementary material available at 10.1186/s12936-022-04436-8. Plasmodium falciparum  manufactured by Ipca Laboratories Ltd (India) was obtained from the WHO Addis Ababa office. Participants were treated twice daily for three consecutive days with the standard six-dose regimen of AL ; enrolment violation ; voluntary protocol violation ; involuntary protocol violation , leading to administration of rescue treatment).Treatment outcome of patients were classified according to the WHO guideline . On the Development of danger signs or severe malaria on day 1, 2 or 3, in the presence of parasitaemia; parasitaemia on day 2 higher than on day 0, irrespective of axillary temperature; parasitaemia on day 3 with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C; and parasitaemia on day 3\u2009\u2265\u200925% of count on day 0.Development of danger signs or severe malaria in the presence of parasitaemia on any day between day 4 and day 28 in patients who did not previously meet any of the criteria of early treatment failure; and presence of parasitaemia on any day between day 4 and day 28 with axillary temperature\u2009\u2265\u200937.5\u00a0\u00b0C in patients who did not previously meet any of the criteria of early treatment failure.Presence of parasitaemia on any day between day 7 and day 28 with axillary temperature\u2009<\u200937.5\u00a0\u00b0C in patients who did not previously meet any of the criteria of early treatment failure or late clinical failure.Absence of parasitaemia on day 28, irrespective of axillary temperature, in patients who did not previously meet any of the criteria of ETF, LCF or LPF.Safety was assessed by recording the nature and incidence of adverse events. Adverse events were assessed by direct interview. All patients were consistently asked about prior symptoms as well as new symptoms that had appeared since their last appointment. The incidence of any adverse event, independent of its relationship to the study drug, was used as a safety end-point. An adverse event was defined as any unfavourable, unintended sign, symptom, syndrome or disease that develops or worsens with the use of the study drug, regardless of whether it is related to the medicinal product.msp1, msp2, and glurp) were to make primary end point analysis. The current study used two of the three markers, the most polymorphic msp2, and the less polymorphic msp1  from a newly acquired infection (different parasite strain). A nested PCR analysis was conducted on paired dried blood spots (day 0 and day of treatment failure) for treatment outcomes classified as late treatment failures (LCF or LPF). PCR correction was made based on fragment analysis on gel electrophoresis. The WHO recommends the use of three markers . Additional assent was obtained for children from age 12 to 17 years.Data entry and analysis were done using the WHO designed Excel spreadsheet. Efficacy data was analysed using the Kaplan-Meier (K-M) and per-protocol (PP) analysis methods. In PP analysis, the efficacy estimate (the proportion cured) of AL was derived by taking all participants followed until treatment failure and adequate response while excluding those who were withdrawn and lost to follow-up. In the K-M approach, patients without treatment failure during the study period and who did not complete follow-up due to withdrawal or lost to follow-up were included in the analysis until the last recorded visit when they were censored. In addition to the criteria for withdrawal, patients were withdrawn from the analysis when PCR results were unclassifiable.P. falciparum infections, 331 were Plasmodium vivax infections, while 16 were mixed infections. A total of 88 consented patients who fulfilled the inclusion criteria were included in the study. Seventeen participants did not complete the study due to withdrawal and lost-to follow-up  were males and 32 (36.4%) were females. Of these, 8 9.1%) were under 5 years of age, 35 (39.8%) were in the age group 5\u201315 years, whereas 45 (51.1%) were adults (\u2265\u200915 years). The mean age of the participants was 18.6 with a mean weight of 41.6\u00a0kg. The mean axillary body temperature, parasite count and haemoglobin level at baseline were 38.1 \u00b0C, 11,920/\u00b5l and 12.2\u00a0g/dl, respectively  . The PCR-corrected Kaplan-Meier survival analysis, the cumulative incidence of success rate of AL was 98.7 (95% CI 91.4\u201399.8)  the day 3 positivity rate was 0.0% (95 CI 0.04.3).o C axillary temperature) on the day of enrolment (day 0). The rest had a history of fever during the previous 24\u00a0h. Of the febrile patients on day 0, 39 (68.4%) were afebrile on day 1. On day 2, 52 (91.2%) of the participants were cleared of fever and 100% of the participants were cleared of fever on day 3. With this regard, the fever clearance rate of AL in this study was 68.4% on day 1, 91.2% on day 2 and 100% on day 3. No febrile cases were detected afterwards. The two treatment failures observed were afebrile on the day of presentation nor had a history of fever in the previous day.Out of the 88 participants, 57 (64.8%) were febrile  and was 12.3\u00a0g/dl (Range: 8.6\u201316.1\u00a0g/dl) on day 14. A slight increase in the mean haemoglobin was observed on day 28 to 12.7\u00a0g/dl (Range: 10-16.3\u00a0g/dl).Based on routine evaluation of worsening or new symptoms following treatment initiation, a total of 33 adverse events (AEs) were reported by 21 (23.9%) of the participants. Of the reported AEs, headache (8%), nausea (6.8%), anorexia (5.7%) and vomiting (4.5%) were the most common. Most of the AEs were reported in the first week of follow-up   three times daily for seven days).According to the WHO recommendations, a result is classified as recrudescence if both paired samples have at least one identical band at both loci amplified. The distance migrated by fragments were visually inspected to verify recrudescence versus re-infection between day zero and day of failure samples for each marker (pretreatment and post treatment samples). Fragments with matching migrated distance were considered recrudescence. Accordingly, one of the treatment failures in the present study was classified as recrudescence. However, no amplification product was obtained for the second treatment failure. As the two markers gave no amplification product, PCR for P. falciparum in the study area. The PCR-corrected cure-rate of AL in our study is consistent with the PCR-corrected findings of previous studies conducted in Ethiopia [The present study demonstrates high efficacy of AL against uncomplicated Ethiopia , 10\u201315 aEthiopia \u201318 afterEthiopia . These rNo ETF or parasitaemia was detected on day 3. However, one LPF was classified as recrudescent malaria after PCR correction. In addition, AL also demonstrated a swift fever and gametocyte clearance, with no febrile cases or gametocyte carriage on day 3 and afterwards. The present study also revealed the favourable safety and tolerability profile of AL.pfk13 gene, ACT still appear efficacious [In Africa, despite detection of mutations in the icacious . In thisicacious \u201324. It iicacious . Howevericacious .According to a review, which did not detect the effect of fat, factors associated with lower day 7 lumefantrine concentration, a key factor for AL efficacy, were unsupervised  treatment, younger children (\u2264\u20094 years), and fever on admission . Hence, P. falciparum infections [In a TES, persistence of parasitaemia on day 3 after treatment initiation is interpreted as suspected artemisinin partial resistance. A 3 to 5% day 3 parasitaemia is expected in areas where parasites are fully sensitive . In the fections \u201337. A refections . These fP. falciparum [Artemisinin derivatives also have a gametocidal effect on sexual stages of lciparum . This halciparum , 14. Sevlciparum , 28\u201330. lciparum \u201341. Nonelciparum , 43.The wealth of safety data on AL supports the favourable safety and tolerability profile of the drug. Most of the reported AEs are of mild or moderate severity and share a commonality with malaria or with concomitant infections \u201347. DespThe study has the following limitations. The partial supervision of the study was a potential limitation. For logistical reasons, the second dose of AL was administered at home and verbally confirmed the next day. Another limitation of this study was the absence of day seven blood lumefantrine level measurement and advanced molecular marker analysis.P. falciparum malaria with rapid fever, parasite and gametocyte clearance in the study area was documented. Accompanying any future TES with advanced molecular analysis and pharmacokinetic data would help to better explain treatment failures and early detection of emerging and spreading resistance.The results of this study provide supporting evidence that AL remains highly efficacious two decades after its introduction in Ethiopia. High efficacy of AL in the treatment of uncomplicated Additional file 1. Table S1. Drug dosing and regimens.Additional file 2. Table S2. Specific primers for msp1 and msp2.Additional file 3. Table S3. Summary of genotyping of parasites."}
+{"text": "We read with interest the publication on malaria treatment by Obonyo et al. . This commentary questions the methodology, especially the chosen time points of treatment outcome measures. A recent clinical trial conducted on Kenyan children by Obonyo et al. compared malaria treatment with quinine (10\u00a0mg/kg quinine sulphate: bd) given with clindamycin (10\u00a0mg/kg: bd) for six doses with artemether\u2013lumefantrine (administered in weight adjusted rounded dosages) [The design of comparative clinical trials for anti-malarials should be based on optimized dosage regimes, which unfortunately Obonyo\u2019s is not. For example, in an earlier study in Gabon, Ramharter et al. have confirmed adequate responses when 3-day regimens of quinine (15\u00a0mg/kg: bd) are combined with clindamycin (7\u00a0mg/kg: bd) for children with malaria . The higEven with higher quinine doses used together with clindamycin, there is a much longer parasite clearance time compared with an artesunate and clindamycin regimen (mean clearance times 46\u00a0h vs 29\u00a0h for the latter combination), although the final day 28 cure rate is very high and similar in both groups [Short term regimes of quinine\u2009+\u2009clindamycin have been used in many clinical trials. Altogether, randomized clinical trials and clinical observations in Brazil and in Gabon, have shown between 88 and 100% final cure rates without counting for reinfections on day 28. Thus, the true cure rate may be higher when PCR genotyping would be used to discount reinfections. Up to half of patients had positive thick blood smears on day 3 in these studies without any effect on final cure rate and need of rescue treatment .The mean parasite clearance times ranged between less than 2 days in African adults with mild malaria and low parasitaemia on admission , and 65\u00a0Identifying non-artemisinin containing combination treatments for uncomplicated childhood malaria is an important objective, but it must be met by implementation of appropriate trial designs with correct dosing in each of the treatment arms in comparative trials. Failure to do so exposes children to the risk of under treatment and of serious misinterpretation of the value of particular regimens."}
+{"text": "Plasmodium vivax malaria has been reported in different endemic settings in Ethiopia. This highlights the need to assess alternative options for P. vivax treatment with artemisinin-based combination therapy, such as pyronaridine-artesunate. This treatment regimen has shown high efficacy for uncomplicated malaria in both Africa and Asia. However, limited data are available from Ethiopia. This study was conducted to assess the efficacy and safety of pyronaridine-artesunate for the treatment of uncomplicated P. vivax malaria in Northwest Ethiopia.Declining efficacy of chloroquine for the treatment P. vivax malaria were enrolled between March and July 2021. Patients were treated with pyronaridine-artesunate once daily for three days. Clinical and parasitological parameters were monitored over a 42-day follow-up period using the standard World Health Organization protocol for therapeutic efficacy studies.A single arm prospective efficacy study was conducted in the Hamusite area, Northwest Ethiopia. Fifty-one febrile adult patients with uncomplicated A total of 4372 febrile patients were screened with 51 patients enrolled and 49 completing the 42-day follow-up period. The PCR-uncorrected adequate clinical and parasitological response (ACPR) was 95.9%  on day 42. Two patients had recurrences  on days 35 and 42. The parasite clearance rate was rapid with fast resolution of clinical symptoms; 100% of participants had cleared parasitaemia on day 1 and fever on day 2. All 16 (31.4%) patients with gametocyte carriage on day 0 had cleared by day 1. There were no serious adverse events., pyronaridine-artesunate was efficacious and well-tolerated for the treatment of uncomplicated P. vivax malaria. In adults in the study setting, it would be a suitable alternative option for case management.In this small study Plasmodium falciparum and Plasmodium vivax are co-endemic in Ethiopia, with proportions of 60% and 40%, respectively [P. vivax is 7.9% by systematic reviews with a wide distribution in the central west region extending to the Northwest and Southwest regions of Ethiopia [Despite malaria morbidity and mortality having reduced substantially in Ethiopia over the last decade, malaria remains a major public health problem . Approxiectively . These pP. vivax\u00a0malaria is chloroquine (CQ) complemented by primaquine (PQ) (0.25\u00a0mg/kg/day) for 14\u00a0days for radical cure [. falciparum\u00a0malaria, followed by single-dose PQ (0.75\u00a0mg/kg) for gametocyte clearance, and the recommended regimen for mixed infection of P. falciparum and P. vivax is AL co-administered with PQ for 14\u00a0days. Dihydroartemisinin-piperaquine is recommended as second-line treatment for both P. falciparum and P. vivax malaria.According to the current national malaria diagnosis and treatment guidelines for Ethiopia updated in 2020, the first-line drug for treatment for uncomplicated P. vivax was in 1989 from Papua New Guinea [P. vivax has been less prevalent in Africa with the first report in Ethiopia in 1996 [P. vivax from different endemic areas of Ethiopia, with the risk of recurrence at day 28 ranging from 7.5% to 22% [The first report of chloroquine resistance in w Guinea . Evidencw Guinea  and has w Guinea . However in 1996 . Recent % to 22% \u201314. The % to 22% .P. vivax in Africa and Asia [P. vivax malaria in Ethiopia.Several studies have investigated the safety and efficacy of pyronaridine-artesunate treatment for and Asia \u201320. HoweThis study was conducted between March and July 2021 at the Hamusit Health Centre, Dera Woreda, South Gonder in Northwest Ethiopia  therapeutic efficacy protocols , to evalP. vivax detected by microscopy, parasitaemia \u2265\u2009250 asexual parasites per microlitre of blood, axillary temperature \u2265\u200937.5\u00a0\u00b0C or history of fever in the past 24\u00a0h and permanently living within the health centre catchment area (5\u201310\u00a0km radius) during the study period.Inclusion criteria were: age \u2265\u200918\u00a0years, monoinfection with P. falciparum and P. vivax), evidence of severe malaria, clinical signs and symptoms of hepatic injury , and known severe liver disease (cirrhosis), known allergy to the study medication, evidence of a non-malaria febrile illness , took an investigation anti-malarial drug within 28\u00a0days, haemoglobin concentration <\u20098.0\u00a0g/dL, known pregnancy and breastfeeding.Exclusion criteria were: mixed infection  provided as a tablet (180\u00a0mg pyronaridine and 60\u00a0mg artesunate). Patients were dosed according to body weight: 20 to <\u200924\u00a0kg, one tablet; 24 to <\u200945\u00a0kg, two tablets; 45 to <\u200965\u00a0kg, three tablets; \u2265\u200965\u00a0kg, four tablets per day. Treatments were given under direct supervision and patients were observed for one hour after each dose to monitor for vomiting or other side effects. Vomiting within 30\u00a0min after administration, led to re-administration of the full dose and vomiting between 30\u00a0min and one hour led to re-administration of a half dose. Patients who vomited a second time were withdrawn from the study and received parenteral artesunate therapy administered according to national guidelines [Patients with Patients were followed-up daily for the first 3\u00a0days after the first dose (day 0) and then weekly on days 7, 14, 21, 28, 35 and 42. Patients were also assessed on any unscheduled visit if symptoms occurred. Adverse events and severe adverse events were defined according to the WHO protocol for monitoring the therapeutic efficacy of anti-malarial drugs . PQ trea\u00ae, Angelholm, Sweden) on days 0, 14, 28, and 42. Females aged 12\u00a0years and older were screened for pregnancy before enrolment.Finger prick thick and thin blood smears were taken from all participants and prepared on two slides for detection of parasites at all follow-up visits. The first slides were prepared by staining with 10% Giemsa for 10\u201315\u00a0min for initial screening. The second slides were stained using 3% Giemsa for 30\u00a0min and read by two independent laboratory technicians from the health centre. If results were discordant, a third reading was performed by a senior laboratory technician. Parasite densities were recorded for all positive slides. The number of asexual parasites was counted per 200 white blood cells (WBC) and parasitaemia was estimated assuming WBC counts of 8000/\u00b5L. Gametocytes were counted against 500 WBCs. Before any blood smear was interpreted as negative, two hundred oil-immersion high-power fields on the thick film were read . To ensuP. vivax. Safety outcomes were the proportion of any symptom occurring during the study period.Efficacy outcomes were based on an assessment of the parasitological and clinical outcomes of anti-malarial treatment according to the WHO guidelines . PatientData were double-entered into the WHO Excel spreadsheet designed for therapeutic efficacy data. IBM SPSS (version 24) software was used to generate descriptive statistics . Based on WHO guidelines the ACPR on day 42 was calculated using the Kaplan\u2013Meier method .The study was approved by the Institutional Review Board (IRB) of the Ethiopian Public Health Institute (EPHI). Written informed consent was obtained from all of the study participants; signed by adults after understanding in their local language. Participant identities were kept confidential throughout.P. falciparum and 82 (19%) P. vivax. Out of 82 P. vivax infected patients, 31 (37.8%) were excluded . Thus, 51 patients were enrolled and treated with pyronaridine-artesunate. There were 2 (3.9%) patients censored during the follow-up period due to loss to follow-up. The target sample of 88 patients was unable to be achieved due to the study being conducted after the peak transmission season.A total of 4,372 febrile patients were screened at Hamusit Health Centre between March and July 2021 . 76.5% (39/51) of patients reported a previous episode of malaria.The baseline characteristics of the study patients are presented in Table The overall day 42 PCR-uncorrected ACPR was 95.9% . Two patients were treatment failures  with late clinical failure on day 35 and day 42 (Table We assessed fever and parasite clearance time for the first seven days to detect any delayed treatment response. All patients had cleared their parasitaemia and gametocytaemia by day 1 (within 24\u00a0h). Fever had cleared in all patients by day 2 Fig.\u00a0.Fig. 3FeNo severe adverse events were recorded following the administration of pyronaridine-artesunate. Some patients had symptoms on day 0 consistent with clinical signs of malaria, including headache 31.4% (16/51), cough 5.9% (3/51), anorexia 3.9% (2/51), vomiting 3.9% (2/51), abdominal pain 3.7% (7/51), nausea 2.0% (1/51), diarrhoea 2.0% (1/51)  of the 49 patients who completed 42\u00a0days follow-up having a P. vivax recurrence.This study found pyronaridine-artesunate to be highly efficacious for the treatment of uncomplicated P. vivax therapeutic efficacy studies, this study was undertaken in a malaria-endemic setting, with patients at risk of new infections during the follow up period. This risk is related to the duration of follow up, host immunity, and level of transmission [Recurrent parasitaemia can be caused by recrudescences from the same isolate, reinfection from a new infection, or relapses from hypnozoites in the liver . Similarsmission .P. vivax malaria is more difficult to control and eliminate than P. falciparum because it can relapse from dormant liver stages after clearance of the initial infection. Effective cure of vivax malaria requires treatment of both its schizontocidal and hypnozoiticidal stages. Pyronaridine-artesunate has schizonticidal efficacy but lacks activity against the P. vivax liver stage hypnozoites. To effectively prevent relapses and minimize the risk of local transmission, pyronaridine-artesunate needs to be administered with a hypnozoiticidal agent, such as primaquine.In general, P. falciparum drug efficacy studies to distinguish new infections from true parasite recrudescences [P. vivax, molecular typing recurrences cannot reliably differentiate between recrudescences, relapses or re-infections. This is because patients can harbour different populations of hypnozoites derived from repeated inoculations [The present study had two late clinical failures recorded after pyronaridine-artesunate treatment at day 35 and day 42. Genotyping of polymorphic loci is undertaken routinely in escences . Howeverulations . The difThe speed of parasite clearance after anti-malarial treatment can be used to assess the therapeutic response to an anti-malarial drug . PresencPyronaridine\u2013artesunate have been associated with asymptomatic mild-to-moderate transient rises in liver transaminases in some malaria patients , 31. AltP. vivax malaria cases in the study area, which reduces the precision of the final results. Study participants were 18\u00a0years and older as the Ethiopia Food and Drug Authority didn\u2019t allow the study to be conducted children younger than 18\u00a0years of age. There is the potential for variable efficacy with pyronaridine-artesunate in children which limits the generalizability of the study\u2019s findings across all age groups.There were some limitations to the current study. The first limitation was that pyronaridine-artesunate plasma concentrations were not measured, so the possibility that the recurrent parasitaemias might have resulted from insufficient drug exposure or parasite resistance cannot be determined. In addition, despite the study being conducted soon after peak transmission season, the planned number of patients could not be enrolled. This was further complicated by a low overall number of P. vivax in Ethiopia. Despite a low number of cases, this study suggests that pyronaridine-artesunate is likely efficacious for the treatment of P. vivax-infected adults in Northwest Ethiopia, with a high parasite clearance rate and rapid clinical response. Following further studies, pyronaridine-artesunate may be considered as a potential anti-malarial as part of the national malaria control and elimination programme.In summary, this is the first study to report the efficacy of pyronaridine-artesunate for uncomplicated"}
+{"text": "Plasmodium falciparum infections are a relatively rare but potentially deadly disease found in returning travellers. We compare the national treatment guidelines of non\u2010endemic countries with the WHO guidelines for the treatment of Plasmodium falciparum infections.Review. We identified non\u2010endemic countries with an incidence rate of imported malaria of at least one per 100,000 population and at least 50 cases annually. Using PubMed and Google Search, we reviewed national guidelines published before 1 March 2021.Thirteen guidelines were identified. For uncomplicated falciparum malaria, 11 of 13 countries (85%) recommend an artemisinin\u2010based combination therapy as first\u2010line regimen in adults, of which artemether\u2013lumefantrine was the most common. For severe malaria, all guidelines recommend the use of intravenous artesunate. Only three countries adjust treatment recommendations based on expected artemisinin resistance.Treatment guidelines for uncomplicated falciparum malaria in non\u2010endemic countries generally adhere to WHO recommendations but often fail to mention the risk of drug resistance in returning travellers. Artemisinin\u2010based Combination Therapies (ACTs) should be the first choice for all uncomplicated malaria cases. Furthermore, the choice between ACTs should be based on regional resistance patterns. Plasmodium species. These ACTs are artemether\u2013lumefantrine, artesunate\u2013amodiaquine, artesunate\u2013mefloquine, artesunate\u2013sulphadoxine\u2013pyrimethamine, dihydroartemisinin\u2013piperaquine and artesunate\u2013pyronaridine [Malaria remains an important cause of morbidity and mortality in large parts of the world. WHO currently recommends the use of artemisinin\u2010based combination therapies (ACTs) for all naridine . These Anaridine . In addiPlasmodium falciparum infections are a relatively rare but potentially fatal cause of disease found in returning travellers. Even though Europe is a non\u2010endemic region, each year around 7300 travellers return with malaria [ malaria . In the  malaria , 5.Plasmodium falciparum is already widely established in most parts of the world [Antimalarial resistance should be taken into account when choosing an antimalarial treatment. For instance, chloroquine\u2010resistant he world . In addihe world , 8, 9. Che world . In the Plasmodium falciparum is associated with the highest morbidity and mortality. The aim of this review is therefore to report how closely national treatment guidelines of non\u2010endemic countries follow WHO recommendations for Plasmodium falciparum malaria treatment, taking into consideration disease severity and artemisinin resistance \u2019 and the name of the country of interest. We included the latest guidelines published before 1 March 2021. Where needed, Google Translate was used.Plasmodium falciparum malaria in adults, children and pregnant women. We also assessed whether the prevalence of antimalarial resistance in the country of transmission was considered when choosing an antimalarial. Definitions of (un)complicated malaria and artemisinin drug resistance can be found in the WHO guidelines on malaria treatment [Guidelines on prophylaxis were excluded, as were sub\u2010national guidelines. We reviewed the guidelines while focussing on first\u2010 and second\u2010line treatments for uncomplicated and severe or complicated reatment . AdditioBased on the incidence of imported malaria, as specified in the Methods section, we selected 17 countries, of which 13 were found to have a national guideline: Australia , BelgiumPlasmodium falciparum malaria [Table Plasmodium falciparum. An ACT is the first choice for chloroquine\u2010resistant Plasmodium falciparum. Not a single country advises more than two ACTs as potential first\u2010line treatments.For uncomplicated ia Table  in adultEleven countries specify treatment recommendations for children. In seven of those guidelines, artemether\u2013lumefantrine is the first choice, followed by dihydroartemisinin\u2013piperaquine (three countries) and atovaquone\u2013proguanil (two countries).Treatment protocols for pregnant women are specified in 12 guidelines. Only The Netherlands recommend the use of artemether\u2013lumefantrine as first\u2010line treatment in the first trimester. Quinine is advised six times as first\u2010line treatment in the first trimester, (hydro)chloroquine and atovaquone\u2013proguanil both once. In the second or third trimester, 10 countries recommend the use of an ACT as first\u2010line treatment, most often artemether\u2013lumefantrine.Nine of the 12 countries that recommend artemether\u2013lumefantrine as an option, recommended to take it with food or a fatty drink.Plasmodium falciparum malaria [Table Regarding severe ia Table , all couPlasmodium falciparum malaria in travellers from the GMS. The Swiss guideline suggests taking atovaquone\u2013proguanil over artemether\u2013lumefantrine or dihydroartemisinin\u2013piperaquine for uncomplicated, or as follow\u2010on medication for severe Plasmodium falciparum malaria due to resistance\u2010related treatment failures in the GMS. The other countries do not adjust their recommendations for travellers from a region with artemisinin resistance.Only three guidelines  incorporate artemisinin resistance patterns in the GMS into their recommendations on treating patients from this region. The French guideline advises the combination of intravenous quinine and artesunate instead of artesunate monotherapy for severe malaria in this group. The Australian guideline also recommends to take the origin of the infection into account when treating a case of severe malaria and recommended seeking expert advice. Additionally, the Australian guideline offers the options of prolonging treatment with artemether\u2013lumefantrine, switching to atovaquone\u2010proguanil or switching to quinine combined with doxycycline or clindamycin for uncomplicated Malaria remains a potentially deadly disease in returning travellers. Drug resistance patterns and treatment options are rapidly evolving, stressing the need for up\u2010to\u2010date guidelines on malaria treatment. This study provides an overview of the state of malaria treatment guidelines across a multitude of non\u2010endemic counties. Comparing national treatment guidelines with WHO recommendations showed many countries still recommending suboptimal treatment regimens. Improvements can be made by prescribing ACTs as first\u2010line treatment and by taking the origin of infection into account.Plasmodium falciparum malaria. They are more effective and have fewer adverse effects than other antimalarials such as atovaquone\u2013proguanil, quinine or mefloquine [ACTs are already generally accepted as the best treatment regimens for uncomplicated floquine , 31, 32.Plasmodium falciparum, as chloroquine\u2010sensitive Plasmodium falciparum is still endemic in parts of Latin America. Their guideline suggests different treatments for those groups, preferring (hydroxy)chloroquine as first\u2010line treatment when possible. However, WHO recommendations make no distinction between chloroquine\u2010resistant strains of Plasmodium falciparum and recommend using an ACT for all forms of Plasmodium falciparum malaria [Canada still recommends the use of quinine in combination with doxycycline or clindamycin as first\u2010line treatment, due to unavailability of ACTs. This combination of drugs is accountable for more adverse effects than ACTs . Atovaqu malaria .Guidelines specifying treatment protocols for children did not differ much from adult treatment protocols. WHO recommends the use of ACTs here as well but with different dosages depending on body weight . In 10 cThe use of ACTs in the first trimester of pregnancy is still debated. Current WHO recommendations still advise the use of quinine with clindamycin in the first trimester due to limited clinical safety data for ACTs . An ACT In general, artemether\u2013lumefantrine was the most used ACT. Higher levels of lumefantrine are associated with lower rates of treatment failure . For thiRegarding severe malaria, all countries included in this review recommend intravenous artesunate over intravenous quinine. Parenteral artesunate is superior to quinine in both adults and children , 5. GuidSevere malaria should be treated with oral follow\u2010on medication to eradicate the remaining parasitaemia. WHO recommends a full dose of effective ACTs orally after the initial parenteral treatment. Combining the data from uncomplicated malaria treatment protocols, we found 10 of 13 countries including an ACT as first\u2010line follow\u2010up treatment. Three countries explicitly advise a non\u2010ACT as follow\u2010on medication: atovaquone\u2013proguanil (Germany), atovaquone\u2013proguanil or quinine with doxycycline/clindamycin (Canada), and artesunate or quinine with doxycycline (Ireland). Still, as described above, an ACT is superior to non\u2010ACTs such as atovaquone\u2013proguanil or quinine as follow\u2010on medication.Plasmodium falciparum malaria strain. In a recent prospective study, conventional treatment with dihydroartemisinin\u2013piperaquine showed therapy failure rates up to 93% in the GMS [With artemisinin and partner drug resistance spreading in the GMS, travellers returning from this region could be infected by an artemisinin and partner drug\u2010resistant  the GMS . Only th the GMS . HoweverSo far, artemisinin resistance seems to have been contained to the GMS. And although some partner drugs have been used in monotherapy at large scale in the last decades, this has not led to widescale ACT failure outside of the GMS . HoweverPlasmodium falciparum infection returning from Southeast Asia should not be treated with dihydroartemisinin\u2013piperaquine. Instead, patients returning from this region could be treated with artesunate\u2013mefloquine or artemether\u2013lumefantrine.Again, we believe taking a travellers' origin into consideration is necessary as some ACTs show treatment failure in the GMS. For example, a patient with a Plasmodium falciparum malaria. A similar review on the treatment of other malaria species will be conducted in the near future.A limitation of this study is the small number of countries included. However, most of the countries included face at least 50 cases of imported malaria annually. Because there is, to our knowledge, no overview on malaria prevalence outside endemic regions, other non\u2010endemic countries with substantial numbers of imported malaria may have been missed in this review. Furthermore, we have no insight into which extent clinicians follow the national guidelines or the WHO guidelines. This review assessed guidelines solely focussing on the treatment of In conclusion, many national malaria treatment guidelines are not in line with WHO recommendations. Too often, non\u2010ACTs such as atovaquone\u2013proguanil and quinine are recommended as first\u2010line treatment. When an ACT is advised as the first choice, few guidelines follow\u2010up with an alternative ACT as the second choice. An ACT should not only be the first but also the second choice of treatment, in children, adults and second and third trimester pregnant women. Furthermore, most guidelines do not mention artemisinin and partner drug resistance, especially in the GMS. Failing to distinguish between travellers from the GMS and other parts of the world could lead to therapy failure in the first group. We recommend keeping a travellers' origin in mind, especially in the case of treatment failure and choosing a suitable ACT based on regional resistance patterns."}
+{"text": "Although primaquine (PQ) is indicated for G6PD-deficient patients, data on weekly PQ use in Brazil are limited. We aimed to investigate malaria recurrences among participants receiving daily and weekly PQ treatments in a real-life setting of two municipalities in the Amazon between 2019 and 2020. Patients receiving weekly PQ treatment had a lower risk of recurrence than those receiving daily PQ treatment , using a model adjusted for study site. Weekly PQ use did not increase the risk of malaria recurrence. Further studies with larger populations are warranted. Plasmodium vivax malaria has been imposed by glucose 6-phosphate dehydrogenase deficiency (G6PDd) in malaria-endemic areasP. vivax malaria, is associated with severe and often life-threatening hemolysis, even in patients with enzymatic variants that are considered to be mild to moderate,-A great challenge to the elimination of P. vivax malariaAlthough safer for G6PDd patients than the standard treatment (daily PQ), a recent systematic review could not show whether a weekly PQ treatment is as effective as the 14-day PQ treatment in preventing recurrences of ,TM qualitative test  was implemented in Rio Preto da Eva, Amazonas state (February 2019 to early January 2020), while the StandardTM G6PD quantitative test  was implemented in both M\u00e2ncio Lima, Acre state (January to December 2020) and Rio Preto da Eva (January to August 2020). At the MTUs, when the thick blood smear tested positive for P. vivax, the PQ regimen was chosen by the healthcare workers based on the results of the G6PD test: chloroquine plus daily (0.5 mg/kg/day for 7 days) or weekly (0.75 mg/kg/week for 8 weeks) PQ was administered to patients with normal and deficient G6PD status, respectively, in accordance with the Brazilian Ministry of Health guidelinesThis study was part of the Safeprim mixed-method study, which evaluated the implementation of G6PDd screening tools in malaria treatment units (MTUs) of two municipalities in the Brazilian AmazonTM, data were collected using REDCap forms in which all mandatory fields were automatically transferred to SIVEP-Malaria, which at that time did not include information on G6PD deficiency. For the StandardTM test, data were entered directly into the SIVEP-Malaria database, which was later adapted to receive G6PDd information as well. All patients\u2019 data were collected and transferred to the national malaria reporting system  by the healthcare teams in each municipality. During the implementation of CareStartFunda\u00e7\u00e3o de Medicina Tropical Dr. Heitor Vieira Dourado, Manaus, Brazil (CAAE: 92012818.1.0000.0005).Descriptive statistics were used to analyze the demographic data. To assess the recurrence rates between patients with normal G6PD levels and G6PD deficiency, patients were followed up for 180 days after the end of treatment using a model adjusted by study site as the grouping variable. Probabilistic data linkage was used for the deduplication of successive cases from the same patient, as described elsewhereThe results of 122 G6PD-deficient and 1,322 normal G6PD patients were reported between February 2019 and December 2020 in both municipalities. When comparing the recurrence rates between patients with normal G6PD and those with G6PD deficiency and considering them as a proxy for daily versus weekly PQ treatment, 20 of 122 patients (16.4%) who were taking PQ weekly had recurrence within 180 days compared with 265 of 1,322 patients (20.0%) who were taking PQ daily  versus daily (20%) PQ. The weekly use of PQ was recently added to the updated Brazilian Ministry of Health guidelines,P. vivax, adherence among patients with normal G6PD levels may be ensured; however, this depends on the availability of suitable and reliable G6PDd screening tools at the point of careNon-adherence to standard treatment in the Brazilian Amazon can contribute to the recurrence of malariaThis study has several limitations. This was an exploratory analysis; hence, the main study was not specifically designed to perform this comparison. Treatment supervision cannot be ensured for both daily and weekly treatments, as this mostly depends on the availability of personnel in both municipalities. The entry of data into the national malaria database was dependent solely on the teams in each municipality. However, as a real-life evidence study, we aimed to assess the implementation pragmatically, including all aspects. Even the use of a screening tool and patients already diagnosed with G6PD deficiency could influence the weekly treatment, thus causing it to be more supervised than the standard treatment. Although the study was conducted during the coronavirus disease 2019 (COVID-19) pandemic, the malaria diagnosis was not interrupted since it was provided at the basic unit level.In conclusion, patients receiving weekly PQ treatment had a lower risk of recurrence than those receiving daily PQ treatment in these two municipalities. However, further studies are necessary to assess its efficacy in larger populations with different healthcare levels."}
+{"text": "Plasmodium has been reported in Madagascar, but their effects on gametocytes are not well documented. The present study aims to determine the emergence of gametocyte and gametocyte clearance after artesunate-amodiaquine (ASAQ) or artemether-lumefantrine (AL) treatment in children with uncomplicated Plasmodium falciparum malaria in 5 regions of Madagascar.Gametocytes are the sexual stages ensuring continuity of the development cycle of the parasite, as well as its transmission to humans. The efficacy of artemisinin-based anti-malarials against asexual stages of P. falciparum malaria, aged between 1 and 15\u00a0years, were assigned randomly to AL or ASAQ treatment. They come from 5 regions of Madagascar with different epidemiological facies related to malaria: Ankilivalo, Benenitra, Ampanihy, Ankazomborona and Matanga. Gametocytes were identified by microscopy, from t blood smears at day 1, day 2, day 3, day 7, day 14, day 21 and day 28 after treatment.558 children with uncomplicated At baseline, 9.7% (54/558) children [95% CI: 7.4\u201312.5%] had detectable gametocyte by microscopy. Among the 54 enrolled children, gametocytes emergence rate was high during the first days of treatment in both treatment arms (AL and ASAQ), especially on day 1. Gametocytes were undetectable from day 14 for AL arm while for ASAQ arm, gametocyte carriage was gradually decreased but persisted until day 21.Plasmodium falciparum malaria.This study demonstrates that AL has a more rapid effect on gametocyte clearance compared to ASAQ in children with uncomplicated The online version contains supplementary material available at 10.1186/s12936-022-04369-2. Plasmodium falciparum gametocytes are relatively insensitive to several anti-malarials  had detectable gametocyte by microscopy  from the 54 patients with gametocytes were analysed. The patients are all children between 1 and 15\u00a0years of age.The figure below shows the distribution of gametocyte carriage according to parasitaemia on day 0 Fig.\u00a0. The curComparing the ASAQ and AL arm, the blood smears collected during the follow-up days  showed no significant emergence of gametocytes during and after the treatment . Regarding the overall treatment outcome, the recovery rate following the 2 arms of treatment was 100%.Plasmodium species to mosquitoes. Understanding the factors influencing gametocytaemia before treatment and the gametocytocidal properties of anti-malarials is of great importance for the implementation of interventions aimed at reducing the transmission of plasmodial infection. According to the literature, gametocytogenesis is sometimes described as a stress response of the parasite which allows it to escape from an increasingly unfavorable environment [The carriage of gametocytes is essential for the transmission of ironment .The results indicate that the carriage of gametocytes is negatively associated with the value of haemoglobin at day 0. Studies in Thailand and The Gambia found that haemoglobin concentrations were lower in gametocyte carriers , 12 and It was observed no significant emergence of gametocytes between treatment arms. Studies report that gametocytes found in peripheral blood emerge within the first week of starting treatment , 17. HowRegarding the clearance of gametocytes, the results show that the gametocytes were undetectable from day14 for the AL arm while the carriage gradually decreases but the gametocytes do not completely disappear until day 28 for the ASAQ arm. These results are consistent with previous studies . It has Plasmodium falciparum malaria.This study demonstrates that AL has a more rapid effect on gametocyte clearance compared to ASAQ in children with uncomplicated These results will be useful for adjusting policy treatment and orienting strategies to combat malaria in Madagascar.Additional file 1: ASAQ dosage.Additional file 2: AL dosage."}
+{"text": "In this Phase 2 trial, we assessed the efficacy and safety of l\u2010arginine at a dose of 20\u2009g orally for 1\u2009week and were followed\u2010up for 3\u2009weeks. The primary endpoint was change in walking speed in the 10\u2010m walk test (10MWT). The main secondary endpoints were change in Timed Up and Go Test (TUGT) time, improvement in inflammatory markers in cerebrospinal fluid (CSF), safety, and tolerability.This open\u2010label, single\u2010arm, Phase 2 study enrolled patients diagnosed with HAM/TSP. Patients received \u22123.5%, 95% CI \u221210.8% to 3.7%; P\u00a0=\u20090.32), a significant improvement was detected between baseline and Day 14 . Significant improvements were also observed in selected secondary endpoints, including in TUGT time , and in neopterin concentration in CSF . Adverse events were infrequent, mild, and resolved rapidly.The study enrolled 20 patients  with a mean age of 67.8\u2009years (95% CI 62.3 to 73.3). Although the primary endpoint, the changes in 10MWT time between baseline (Day 0) and Day 7, did not reach statistical significance (mean percent change in time l\u2010arginine therapy improved motor function and decreased CSF inflammatory markers. l\u2010arginine thus represents a promising therapeutic option for patients with HAM/TSP.UMIN000023854. The retrovirus human T\u2010lymphotropic virus type 1 (HTLV\u20101) is the etiologic agent for adult T cell leukemia/lymphomal\u2010Arginine is a basic amino acid and has significant metabolic roles including a precursor for the synthesis of urea, ornithine, citrulline, and nitric oxide. Recent studies have reported that l\u2010arginine and its metabolism would be a potential therapeutic target in a number of diseases.l\u2010arginine is currently in clinical use for the treatment of urea cycle defectsl\u2010arginine has been shown to have a protective effect against the accumulation of misfolded proteins in neuronsl\u2010arginine is known to cross the blood\u2013brain barrier and has a neuroprotective effect in neurological disorders,l\u2010arginine could be considered as a candidate therapy for HAM/TSP.l\u2010arginine in patients with HAM/TSP.In this open\u2010label, single\u2010arm, Phase 2 trial, we evaluated the efficacy and safety of short\u2010term treatment with orally administered l\u2010arginine at a dose of 20\u2009g orally three times (TID) daily for 1\u2009week and were followed\u2010up for 3\u2009weeks. Full inclusion and exclusion criteria are detailed in the protocol in This was an open\u2010label, single\u2010center, Phase 2 study. Patients aged \u226520\u2009years who were diagnosed with HAM/TSP according to the World Health Organization guidelinesThe study protocol was reviewed and approved by the Institutional Review Board of Kagoshima University. The clinical trial was registered with University Hospital Medical Information Network Clinical Trials Registry . All patients gave written informed consent prior to enrollment.l\u2010arginine was assessed throughout the study based on adverse events, vital signs, and laboratory tests.The primary endpoint was change in walking speed (i.e. time taken) in a 10\u2010m walk test (10MWT) between baseline (Day 0) to Day 7. The secondary efficacy endpoints were change in walking speed in a 10MWT from baseline to Days 3 and 14; change in Timed Up and Go Test (TUGT) time from baseline to Days 3, 7, and 14; and change in cell counts, total protein level, and neopterin concentration in the cerebrospinal fluid (CSF) from baseline to Day 7. The safety of l\u2010arginine concentration, complete blood cell count, HTLV\u20101 proviral load (PVL) in peripheral blood mononuclear cells (PBMCs), and levels of biochemical markers, cytokines, and amino acids. CSF tests included cell counts and concentrations of neopterin and C\u2010X\u2010C motif chemokine ligand 10 (CXCL10), which are specific central nervous system (CNS) inflammatory markers in patients with HAM/TSP.Lower limb spasticity was investigated using the Modified Ashworth Scalel\u2010arginine, adiponectin, ornithine, citrulline, and tryptophan were analyzed by one\u2010way repeated measures analysis of variance followed by Dunnett's multiple comparison test. HTLV\u20101 PVL and concentrations of neopterin and CXCL10 in CSF were analyzed by paired t\u2010tests. All statistical analyses were performed using Prism version 9.3.1 (GraphPad Software). P\u00a0<\u20090.05 was considered statistically significant. Values are presented as the mean and 95% confidence intervals (CI).Percent changes in 10MWT, TUGT, and PBMC PVL; mean change in OABSS score; and mean change in serum concentrations of 4 cells (95% CI 406.3 to 1222.0) . A significant improvement in the TUGT measure (secondary endpoint) was also observed between baseline and Day 14  and Day 28  and by 1.4 points between baseline and Day 28  were decreased from 12.7\u00a0pmol/mL (95% CI 7.6 to 17.8) at baseline to 10.6\u00a0pmol/mL (95% CI 6.8 to 14.3) on Day 7, representing a significant change  between baseline and Day 7 and by 20.6%  between baseline and Day 14. Other serum inflammatory markers measured, including tumor necrosis factor\u2010\u03b1 (TNF\u2010\u03b1) and natural killer cell counts, were not significantly changed from baseline at any time point analyzed  and Day 7, the end of the dosing period . Thereafter, plasma l\u2010arginine levels decreased and returned to baseline levels (31.8\u2013149.5\u00a0\u03bcmol/L by Day 28)  Fig.\u00a0. Thus, pl\u2010arginine on the plasma profiles of other amino acids and identified significant changes in the plasma concentrations of three amino acids; ornithine, citrulline, and tryptophan  and Scientific Research (C) (21\u2009K07418 to E. Matsuura) from the Japan Society for the Promotion of Science (JSPS), Japan.The authors have declared that no conflicts of interest exist.Appendix\u00a0S1Click here for additional data file.Figure\u00a0S1Click here for additional data file.Figure\u00a0S2Click here for additional data file."}
+{"text": "Plasmodium vivax malaria has been documented in Ethiopia. Thus, there is a need to assess the efficacy of alternative schizontocidal anti-malarials such as dihydroartemisinin\u2013piperaquine (DHA\u2013PPQ) in P. vivax malaria-infected patients. This study was conducted to evaluate the therapeutic efficacy of DHA\u2013PPQ drug in South West Ethiopia.Declining efficacy of chloroquine against P. vivax malaria. The study was conducted from May 2021 to August 2021, based on the standard World Health Organization study protocol for surveillance of anti-malarial therapeutic efficacy. The study endpoint was adequate clinical and parasitological response on day 42.This is a single-arm, prospective therapeutic efficacy study in patients with uncomplicated A total of 86 patients with uncomplicated vivax malaria were enrolled. Of these, 79 patients completed the scheduled follow up; all showing adequate clinical and parasitological responses to day 42, with a successful cure rate of 100% (95% CI 96\u2013100). Parasitaemias were cleared rapidly (86% by day 1 and 100% by day 3), as were clinical symptoms (100% by day 1). Gametocyte carriage decreased from 44% on Day 0 to 1% on day 1 and 0% on Day 2. Mean haemoglobin concentrations increased between day 0 (mean 12.2\u00a0g/dL) and day 42 (mean 13.3\u00a0g/dL). Treatment was well tolerated and no severe adverse events were observed.P. vivax, with no recurrences to day 42, and no safety concerns. This treatment, which is also effective against P. falciparum, appears to be an ideal alternative for P. vivax as part of the malaria elimination programme.In summary, treatment with DHA\u2013PPQ demonstrated excellent efficacy for uncomplicated  In Ethiopia, the scale-up of anti-malarial interventions has been associated with a reduction in malaria cases and deaths in the last decade. Ethiopia\u2019s National Malaria Indicator survey reported a very low prevalence of malaria by microscopy (0.5%) , and thePlasmodium vivax is a widely distributed parasite worldwide [Plasmodium\u00a0falciparum\u00a0and P. vivax,\u00a0with relative proportions of about 70% and 30%, respectively [P. vivax develops gametocytes substantially earlier than P. falciparum, leading to the early release of gametocytes into the bloodstream from the liver before the appearance of clinical symptoms and facilitating transmission of vivax malaria. Hypnozoite relapse of P. vivax, after resolution of the initial infection, further complicates the control and elimination of P. vivax compared with P. falciparum.orldwide , but in ectively . These pP. falciparum led to its replacement with sulfadoxine-pyrimethamine (SP) in 1998 [P. falciparum or P. vivax is quinine [Chloroquine was used for decades as the first-line treatment for all malaria species in Ethiopia. Widespread declines in the efficacy of chloroquine to treat  in 1998 , 6. Sinc quinine . HoweverP. falciparum, P. vivax has the ability to form dormant hypnozoites in the liver that can cause a relapse of infection weeks to months after the initial attack [P. vivax infections are derived from hypnozoites rather than new infections [P. vivax without prior glucose-6-phosphate dehydrogenase (G6PD) testing in regions with all transmission levels [P. falciparum and P. vivax. This drug is approved by the World Health Organization (WHO) and registered in Ethiopia [Unlike l attack . About 8fections . Currentn levels . In EthiEthiopia .P. vivax in Ethiopia [P. vivax, with prolonged post-treatment prophylaxis beyond that provided by artemether-lumefantrine, suppressing the risk of recurrence for 6\u20138\u00a0weeks after treatment [Several studies have reported a decline in the efficacy of chloroquine against Ethiopia \u201314 highlEthiopia . The sloreatment , 17.P. vivax in Ethiopia. Such data are important to understand the potential utility of DHA\u2013PPQ in Ethiopia as an alternative to chloroquine or artemether-lumefantrine for the treatment of P. vivax. This study was conducted to assess the therapeutic efficacy and safety of DHA\u2013PPQ for the treatment of vivax malaria in the Seacha area of Ethiopia.There are a lack of data on the effectiveness of DHA\u2013PPQ for the treatment of Plasmodium vivax is the predominant parasite species and Anopheles arabiensis is the major vector. Based on the current malaria stratification and mapping of the country [P. vivax transmission.The study was conducted in Seacha area, Arba Minch Zuria District, located in the Gamo Gofa zone of the Southern Nations, Nationalities, and Peoples\u2019 Region of Ethiopia Fig.\u00a0. The stu country , this stP. vivax mono-infection, with an asexual parasitaemia of 250 parasites/\u03bcL or above and living within the facility catchment area . Participants were excluded if they were pregnant, breast-feeding, had mixed species infection , had a day 0 haemoglobin\u2009<\u20095.0\u00a0g/dL, were unable to take oral medication or had repeated vomiting, had known hypersensitivity to the study drugs, had evidence of severe malaria, heart or liver diseases, had severe malnutrition, or had evidence of a non-malarial febrile illness . Primaquine therapy was not provided until the day of first recurrence or day 42.Febrile patients visiting the outpatient department of the Seacha Health Centre who fulfilled the inclusion criteria set by the WHO protocol for the assessment of the therapeutic efficacy of anti-malarial drugs were eligible for inclusion . Inclusi, with a 20% increase to allow for loss to follow-up and withdrawal during the 42-day follow-up period, at least 88 patients were to be recruited.The sample size was determined according to the WHO protocol ; using tEligible patients were treated with a full three-day course of DHA\u2013PPQ. DHA\u2013PPQ  was provided by the Ethiopian Federal Ministry of Health (FMoH) through WHO support. Daily drug dosage was based on body weight as follows: half a tablet, 5 to\u2009<\u20098\u00a0kg; three-quarters of a tablet, 8 to\u2009<\u200914\u00a0kg; one tablet, 14 to\u2009<\u200925\u00a0kg; two tablets, 25 to\u2009<\u200936\u00a0kg; three tablets, 36 to\u2009<\u200960\u00a0kg, four tablets 60 to\u2009<\u200980\u00a0kg and five tablets,\u2009>\u200980\u00a0kg.All medication doses were given under the direct supervision of the study team. Medication given to young children was crushed, mixed with water, and administered as a suspension. Patients were observed for thirty minutes to 1\u00a0h after each dose to monitor for vomiting or other side effects. If vomiting occurred within 30\u00a0min after administration, the full dose was re-administered. If vomiting occurred between 30\u00a0min and 1\u00a0h after administration, half of the dose was re-administered. Patients who vomited a second time were withdrawn from the study and received parenteral therapy according to national guidelines. Standard dose primaquine (0.25\u00a0mg/kg daily for 2\u00a0weeks) was given to patients at the end of the follow-up period or at recurrence.Patients were followed-up daily for the first 3\u00a0days after the first dose (day 0) and then weekly on day 7, 14, 21, 28, 35, and 42. Clinical and laboratory evaluations were undertaken during each follow-up visit. Patients were also assessed on an unscheduled visit if symptoms occurred. Adverse events and severe adverse events were defined according to the WHO protocol for monitoring the therapeutic efficacy of anti-malarial drugs and were monitored at each follow\u2010up visit . Fever c\u00ae, Angelholm, Sweden), on days 0, 14, 28, and 42. Female study participants aged 12\u00a0years and older were screened for pregnancy on enrolment.Thick and thin blood smears were prepared on a single slide, for parasite detection and species identification, respectively. Two smears were prepared from all participants at all follow-up visits. The first slide was prepared by staining with 10% Giemsa for 10\u201315\u00a0min for initial screening. The second slide was stained using 3% Giemsa for 30\u00a0min and read by two independent laboratory technicians from the health centre; in case of discrepancy between the first and second readings, a third reading was performed. Parasite densities were recorded for all positive slides. The number of asexual parasites was counted per 200 white blood cells (WBC) and parasitaemia was estimated assuming WBC counts of 8000/\u00b5l. Gametocytes were counted against 500 WBCs. Before any blood smear was interpreted as negative, two hundred oil-immersion high power fields on the thick film were read . To ensuStudy endpoints were categorized into primary and secondary endpoints. The primary outcome was the day 42 overall efficacy of DHA\u2013PPQ stated as polymerase chain reaction (PCR)-uncorrected proportion with adequate clinical and parasitological response (ACPR). Secondary endpoints were the incidence of adverse events, fever and parasite clearance, and haemoglobin levels across the follow-up period. Data were double-entered into the WHO Excel spreadsheet designed for therapeutic efficacy data. Data were also entered into IBM SPSS (version 24) software to calculate descriptive statistics . Based on WHO guidelines, the adequate clinical and parasitological response on day 42 was calculated using the Kaplan\u2013Meier cumulative risk method .The study was approved by the Institutional Review Board (IRB) of the Ethiopian Public Health Institute (EPHI). Written informed consent was obtained from all of the study participants\u2009\u2265\u200918\u00a0years of age or by parents or guardians of children less than 18\u00a0years of age, with children 12\u201317\u00a0years of age providing written informed assent.P. vivax (163/2662) of which 88 (54%) were initially enrolled and treated with DHA\u2013PPQ. The main reasons for exclusion from enrolment were living too far away for follow-up , refusal to consent , pregnancy or lactation , concomitant disease  and non-residence . After slide rechecking by the WHO accredited microscopist, 2 further patients were excluded . In total, 86 patients were finally enrolled and followed-up according to WHO protocols. There were 7 (8%) patients who were censored during follow-up (1 protocol violation and 6 lost to follow-up)  of them completed the 42-days follow up. The treatment outcomes by microscopy are presented in Table % of themThe proportion of patients with noticeable gametocytaemia at enrolment was 44% (38/86). The proportion of patients that had no evidence of gametocytes on day-1 was 99% (85/86). No gametocytes were present on day-2.The mean haemoglobin concentration at baseline was 12.2\u00a0g/dL (standard deviation 1.4). There was a small increase in mean haemoglobin concentration from baseline to day 28 and day 42. No study participant developed severe anaemia (<\u20098\u00a0g/dL) during follow up.The mean body temperature at day 0 was 37.9\u00a0\u00b0C (95% CI 37.7\u201338.2), with all patients having fever resolution (<\u200937.5\u00a0\u00b0C) by day 1 , followed by abdominal pain n\u2009=\u20093, 4%) and vomiting  , in addition to prevention of relapses prior to day 42. The absence of recurrent infections prior to day 42 is consistent with a prolonged period of post-treatment prophylaxis following DHA\u2013PPQ . This isP. falciparum and P. vivax malaria. In remote malaria endemic areas, where the ability to differentiate Plasmodium species may be limited, use of DHA\u2013PPQ may be used to simplify treatment and avoid complications caused by using the wrong anti-malarial.Despite a high gametocyte carriage rate detected pre-treatment in the current study (44%), complete gametocyte clearance was observed by day 2. Thus, DHA\u2013PPQ demonstrated good transmission-blocking efficacy, rapidly preventing infection of mosquitoes. In addition, DHA\u2013PPQ can be used to be used to treat both The day 0 mean haemoglobin level was lower than those on day 28 and day 42. This is consistent with the expected increase in haemoglobin concentration following successful parasite clearance . HoweverP. vivax is in the Seacha region and whether the need for an alternative treatment option is urgent.The generalizability of this study was limited by being undertaken at a single health centre. In addition, it is unclear what the efficacy of chloroquine against Effective anti-malarial treatment is a cornerstone of effective malaria control and progression towards achieving elimination in Ethiopia. Therefore, periodic monitoring of currently used anti-malarial drugs and potential alternative ACTs through therapeutic efficacy studies is important to identify the early emergence of anti-malarial drug resistance. The findings from this study demonstrate similar efficacy of DHA\u2013PPQ to a previous unpublished study conducted between 2017 and 2018 in Pawi and Arba Minch .P. vivax infection in all ages in Ethiopia. It demonstrates that DHA\u2013PPQ is highly efficacious and safe for the treatment of uncomplicated P. vivax malaria in Arba Minch, Ethiopia. With potential increases in chloroquine resistance in vivax malaria in Ethiopia, DHA\u2013PPQ provides an alternative treatment option to quinine, with rapid parasite clearance and resolution of fever and effective gametocyte killing. Further assessment of DHA\u2013PPQ in different transmission settings in Ethiopia will be important to confirm these results.This is the first published study to assess the efficacy of DHA\u2013PPQ against"}
+{"text": "Plasmodium antigens were significantly associated with ASMQ treatment outcome but not AL. We used machine learning to develop predictive models of treatment outcome based on the immunoprofile data. The models predict treatment outcome for ASMQ with high (72\u201385%) accuracy, but could not predict treatment outcome for AL. This divergent treatment outcome suggests that humoral immunity may synergize with the longer mefloquine half-life to provide a prophylactic effect at 28\u201342 days post-treatment, which was further supported by simulated pharmacokinetic profiling. Our computational approach and modeling revealed the synergistic effect of pre-existing immunity in patients with drug combination that has an extended efficacy on providing long term treatment efficacy of ASMQ.The impact of pre-existing immunity on the efficacy of artemisinin combination therapy is largely unknown. We performed in-depth profiling of serological responses in a therapeutic efficacy study [comparing artesunate-mefloquine (ASMQ) and artemether-lumefantrine (AL)] using a proteomic microarray. Responses to over 200  Plasmodium falciparum malaria. TESs conducted at regular intervals in the same location can be used for the detection of the decline of drug efficacy over time. Key indicators monitored during ACTs TESs include proportion of patients who are parasitemic on day 3, and treatment failure by days 28 or 42 (Therapeutic efficacy studies (TESs) are used to monitor efficacy of antimalarial drugs including assessment of clinical and parasitological outcome for artemisinin-based combination therapies (ACTs), the first-line treatment for uncomplicated 28 or 42 . NaturalArtemether-lumefantrine (AL) is the most widely used ACTs in sub-Saharan Africa (sSA), followed by artesunate-amodiaquine (ASAQ) . A studyThe power of computational tools and mathematical modeling in resolving complex biological questions such as identifying correlates of protection \u201312 or biclinicaltrials.gov (NCT01976780). Cohort I study was conducted between June 2013 and November 2014, and assessed ASMQ and AL, while cohort II study was conducted between December 2014 and July 2015, and assessed DP and AL. Potential study participants (age 6 months- 65 years) with uncomplicated malaria were identified using malaria rapid diagnostic test (mRDT). Informed consent/assent from the participants, parents or legally authorized representatives was obtained prior to screening procedures. Study details are described in This was a randomized, open-label, two-cohort trial, each with two arms conducted in western Kenya, a high transmission, holoendemic region. The study was approved by Institutional Review Boards (IRBs) and Human Subjects Protection Branch, and was registered at \u00ae\u2014Holly Cotec Pharmaceuticals, China), ASMQ or AL . Details of drug treatment scheduling, dosing, and administration are provided in Enrolled participants were randomized for malaria treatments and treated with DP  as well as on samples collected from participants who had reappearance of parasites on follow-up visits as previously described ] was used as previously described  while machine learning was carried out using the R caret package. The R codes are available in GitHub1 and datasets are available on request.Detailed bioinformatics, data analysis, and modeling methods can be found in us study . All stan = 118 in each cohort) were enrolled of which 200 (84.7%) participants completed 42-day follow-up, 100 from each cohort; 52 in ASMQ arm and 48 in AL arm in cohort I and 50 in each arm in cohort II (p = 0.042) but not on day 42 (p = 0.280), and in cohort II, a significant difference was present for DP vs. AL on day 28 (p = 0.001) and on day 42 (p = 0.008). Of note, none of the study participants developed side effects including those in ASMQ arm.A total of 236 study participants  was successfully performed in some of the parasite isolates  in the ASMQ arm. Normalizing the mean signal intensities to with age . Among aMachine learning methods (random forest models confirmed by logistic regression models) were used to assess the degree to which humoral immunity to malaria could predict treatment outcome. For the ASMQ arm, 100 random forest models were built for predicting treatment outcome using antibody signals that were significantly different between participants that showed nPC-ACPR compared to those who did not. Models built for predicting treatment outcomes on day 28 achieved 85% accuracy (Kappa: 0.40) with an average AUCROC of 0.85 , and on To explore how humoral immunity and treatment interact, we simulated PK profile of ASMQ and AL out to day 28 and 42. The PK profiles of both regimens show that the concentration of artemisinin derivative drugs clears within 6 days post-treatment. In the AL regimen, lumefantrine is cleared by day 11 post-treatment, therefore is unlikely to have a major impact on day 28 and 42 outcomes . By compP < 0.01) between the time to malaria re-infection and classes of participants identified by machine learning models  of the drug required to be effective is likely to vary between individuals. Given that all participants showed nPC-ACPR at day 7, for participants that showed nPC-ACPR failure by day 28, it is plausible the drug concentration fell below individual\u2019s MIC between day 7 and 28. Based on our simulated PK profiles, the average mefloquine concentration during that time span was 2.35 mg/l. Likewise, for participants that showed nPC-ACPR at day 28, but nPC-ACPR failure by day 42, the mefloquine concentration fell below their MIC within the time span of day 28 and 42, during which an average concentration was estimated to be 0.25 mg/l. Therefore, based on our estimates, \u201cvulnerable\u201d participants who had nPC-ACPR failure by day 28 have an average MIC > 2.35 mg/l, while immune participants who show nPC-ACPR even out to day 42 have an average MIC of < 0.25 mg/l. In addition, Cox regression analysis showed a strong association  relatively small sample size; our univariate analysis statistical test accounts for sample size and utilizes a multiple test correction to account for the large number of parameters being measured. The immune correlates of outcome were identified in ASMQ cohorts, but not in AL cohorts. This would not have been possible in an under-powered study. (2) The small number of correlates identified in the ASMQ Day 42 cohort could be the result of the waning impact of immunity over time, post-treatment, as the mefloquine concentration decreases. In the future, it will be important to repeat these analyses using other ACTs especially DP due to its high efficacy and the long prophylactic life-span of piperaquine.In conclusion, we have demonstrated that data integration, machine learning, and modeling provide a comprehensive approach capturing the underlying complexity of malaria control in sSA. Further, we have shown ASMQ is a highly effective drug, making it an appropriate choice of possible first-line treatment in western Kenya, a region which account for most malaria transmission in the country.The original contributions presented in this study are included in the article/The studies involving human participants were reviewed and approved by the Kenya Medical Research Institute Scientific and Ethical Review Unit (KEMRI-SERU)\u2014KEMRI SSC numbers 2518 and 2722, as well as the Walter Reed Army Institute of Research Institutional Review Board (WRAIR IRB)\u2014WRAIR numbers 1935 and 1935A. The patients/participants provided their written informed consent to participate in this study.BA: study principal investigator (PI), funding, data analysis, manuscript draft writing, and reviewing. PL: bioinformatics analysis and draft review. IO: study oversite and recruitment of study participants. EB-L: data analysis, draft writing, and reviewing. RW: study pharmacists. GO, LC-B, LI, DJ, BenO, AC, RY, CO, RO, and GC: laboratory analysis of specimens. JC: immunological data analysis, funding, and manuscript review. AW: bioinformatics analysis and team lead. HA: specimen and personnel management, data analysis, and draft review. DO: data analysis and manuscript review. BerO: consultation and manuscript review. SC: bioinformatics analysis, draft writing, and reviewing. EK: study and team management, concept refinement, draft writing, and reviewing. All authors contributed to the article and approved the submitted version."}
+{"text": "Plasmodium vivax forms dormant liver stages that can reactivate weeks or months following an acute infection. Recurrent infections are often associated with a febrile illness and can cause a cumulative risk of severe anaemia, direct and indirect mortality, and onward transmission of the parasite. There is an increased risk of P. vivax parasitaemia following falciparum malaria suggesting a rationale for universal use of radically curative treatment in patients with P. falciparum malaria even in the absence of detectable P. vivax parasitaemia in areas that are co-endemic for both species.P. vivax parasitemia on day 63. Secondary endpoints include incidence risk on day 63 of symptomatic P. vivax malaria and the risk of any P. falciparum parasitaemia. Secondary safety outcomes include the proportion of adverse events and serious adverse events, the incidence risk of severe anaemia (Hb<5g/dl and <7g/dl) and/or the risk for blood transfusion, the incidence risk of \u2265 25% fall in haemoglobin with and without haemoglobinuria, and the incidence risk of \u2265 25% fall in haemoglobin to under 7g/dl with and without haemoglobinuria.This is a multicentre, health care facility-based, randomized, controlled, open-label trial in Bangladesh, Indonesia and Ethiopia. Patients with uncomplicated falciparum malaria, G6PD activity of \u226570% of the adjusted male median (AMM) and haemoglobin levels \u22658g/dl are recruited into the study and randomized to either receive standard schizonticidal treatment plus 7-day high dose primaquine  or standard care in a 1:1 ratio. Patients are followed up weekly until day 63. The primary endpoint is the incidence risk of any P. vivax and P. falciparum in different endemic locations. If found safe and effective universal radical cure could represent a cost-effective approach to clear otherwise unrecognised P. vivax infections and hence accelerate P. vivax elimination.This study evaluates the potential benefit of a universal radical cure for both NCT03916003. Registered on 12 April 2019.The online version contains supplementary material available at 10.1186/s13063-022-06364-z. The order of the items has been modified to group similar items , an 8-aminoquinoline (8-AQ), which can cause severe drug-induced haemolysis in individuals with glucose-6-phosphate-dehydrogenase (G6PD) deficiency. The risk of haemolysis after PQ depends on the dose administered and the G6PD activity of the individual exposed. The risk of serious haemolysis associated with G6PD deficiency makes clinicians reluctant to prescribe PQ without prior testing, which is often unavailable . When PQP. vivax recurrence than the control  [The recently completed multicentre IMPROV study compared the efficacy of a 7-day PQ regimen (1.0 mg/kg/day for 7 days) with a 14-day regimen (0.5 mg/kg/day for 14 days). PQ was given with either chloroquine (CQ)  or Dihydroartemisinin-piperaquine (DHA-PIP) (in Indonesia). In total, 2388 patients were enrolled and screened for G6PD deficiency using the fluorescent spot test (FST). The 7-day PQ regimen\u00a0(PQ7) was non-inferior to the 14-day regimen and 5-fold more efficacious at reducing P. falciparum and P. vivax, and the benefits of this treatment are likely to extend to patients presenting with uncomplicated malaria due to either species. The potential to provide a radical cure safely with a 7-day regimen of PQ provides major practical advantages. Our multicentre randomized, open-label trial compares the safety and efficacy of a high dose PQ treatment in G6PD normal patients with P. falciparum to reduce the risk of subsequent P. vivax episodes to the current standard practice of providing only schizonticidal treatment.There is a need for safe and effective radical cure of malaria in areas co-endemic for both P. vivax parasitaemia by day 63 in G6PD normal patients following uncomplicated P. falciparum malaria.To assess the safety and efficacy of a 7-day course of high dose PQ treatment regimen in preventing recurrent P. falciparum parasitaemia.To assess the efficacy of a 7-day course of high dose PQ treatment regimen in preventing P. falciparum.To assess the efficacy of 7-day high dose PQ treatment in reducing gametocyte carriage of P. falciparum\u00a0malaria.To assess the safety and tolerability of high dose PQ in patients with P. vivax parasitaemia in patients presenting with uncomplicated P. falciparum mono-infection.This study is a multicentre, health care facility-based, randomized, controlled, open-label superiority trial comparing high dose PQ treatment to prevent The study is\u00a0a multicentre study carried out in study sites in Ethiopia, Bangladesh and Indonesia.Healthy male adultNo blood transfusion or major surgery in the last 3 monthsWritten informed consentParticipants must meet the following criteria to be eligible for the pre-study:P. falciparum mono-infectionFever (axillary temperature \u226537.5 \u00b0C) or history of fever in the preceding 48 hAge >1 year (\u2265 18 years at the Ethiopia site)G6PD normal as defined by the Biosensor  at \u226570% of the adjusted male median (AMM) for each siteWritten informed consentAble to comply with all study procedures and timelinesPatients must meet the following criteria to be eligible for the trial:General danger signs or symptoms of severe malariaAnaemia, defined as Hb <8g/dlPregnant women as determined by urine \u03b2-HCG pregnancy testBreastfeeding womenKnown hypersensitivity to any of the drugs givenRegular use of drugs with haemolytic potentialBlood transfusion within the last 4 monthsIf the patients meet any of the following criteria at the screening, they will not be eligible for the trial:Patients attending one of the study health centres will be screened for enrolment into the trial. If patients are eligible and willing to participate in the study, they will be asked to sign a written informed consent form for enrolment. Written informed consent will be obtained by trained and authorized staff. In the case of minors, the legal guardian will be asked to sign on behalf of the minor. In addition, minors above the age of 11 years will be asked to provide written assent.In Ethiopia, patients will be asked for additional consent to provide up to 3 blood samples on day 5 within 1 to 6 h after treatment to determine drug levels. At the study site in Bangladesh, participants will be asked to provide additional consent for the collections of costs incurred by the patient during their illness to inform cost analysis. All patients will be asked to provide additional consent for the long-term storage of blood samples to conduct tests not included in the protocol. Those additional procedures will require additional ethical approval.P. falciparum gametocytes. All treatment will be directly supervised.All participants in the control arm will be treated for acute uncomplicated falciparum malaria according to local guidelines. In Ethiopia and Bangladesh, this will be with artemether-lumefantrine (AL) administered twice daily over three days according to the manufacturer\u2019s recommendations. In Indonesia, patients will be treated with Dihydroartemisinin-piperaquine (DHA-PIP) administered once daily over three days according to the manufacturer\u2019s recommendations. If the participant vomits the treatment within 60 min, a repeat dose will be administered. If the patient vomits the second dose, the patient will receive second-line treatment. In the control arm, patients will be prescribed a single dose of PQ if this is included in the national guidelines for the eradication of All participants in the intervention arm will be treated for acute uncomplicated falciparum malaria according to local guidelines and as outlined for the control group. In addition, patients in the intervention arm will be administered a high-dose PQ regimen over 7 days (1.0 mg/kg/day for 7 days), which will be commenced on day 0 and administered along with a small snack. All treatment will be directly supervised.Each participant has the right to withdraw from the study at any time. If the participant withdraws consent or is lost to follow-up for 4 or more weekly visits, the patient will be excluded/withdrawn. It will be left to the Investigator\u2019s clinical judgement whether or not an adverse event (AE) is of sufficient severity to require stopping the participant\u2019s treatment. If the participant is withdrawn due to an AE, the investigator will arrange for follow-up visits or telephone calls until the AE has resolved and conditions stabilized.Patients who fail to respond adequately to the study treatment will be administered rescue treatment according to national guidelines. Patients who develop severe malaria will be admitted to hospital and treated with the recommended parenteral drug (intravenous or intramuscular artesunate or intramuscular artemether) according to national guidelines.Patients will be asked to return to the health centre for directly observed treatment for all drug doses. If patients do not return all efforts will be made to restart treatment.At enrolment, all anti-malarial medication received in the preceding 4 weeks will be documented on the clinical record form (CRF). Regular medication at trial entry for conditions other than malaria, e.g. asthma, hypertension, etc., will be documented. Patients will be asked to continue to take these regular medications in the normal way. Any additional drugs taken during the trial period for whatever reason will be documented . Patients who discontinue their trial medication prematurely, or who fail to respond to trial medication and receive other anti-malarial therapy, will be recorded with a start and end date. Drugs with antimalarial activity should be avoided, unless prescribed by the attending clinician.The sponsor has insurance, which is in accordance with the legal requirements. This insurance provides coverage for damage to research subjects through injury or death caused by any activities of the study.P. vivax parasitaemia on day 63Primary: The incidence risk of any P. vivax parasitaemia on day 63The incidence risk of symptomatic P. vivax parasitaemia on days 28 and 42The incidence risk of any P. falciparum malaria on days 28, 42 and 63The incidence risk of any P. falciparum gametocytaemia between days 7 and 63The incidence risk of Parasite clearance on days 1, 2 and 3Fever clearance on days 1, 2 and 3Secondary:The proportion of patients vomiting their medication on the day of enrolment within 1 hour of administration;The proportion of patients vomiting any of their PQ doses within 1 hour of administration;The proportion of adverse events and serious adverse events;The incidence risk of severe anaemia (Hb<5g/dl) and moderately severe anaemia (<7g/dl) and/or the risk for blood transfusion between days 3 and 7;The incidence risk of \u226525% fall in haemoglobin since baseline with and without haemoglobinuria on days 3 and 7;The incidence risk of \u226525% fall in haemoglobin to under 7g/dl with and without haemoglobinuria on days 3 and 7.The participant timeline can be found in Supplement P. vivax after P. falciparum is highest after treatment with AL, which is the schizonticidal treatment used in two of the three study sites (Bangladesh and Ethiopia). Assuming a risk of P. vivax after P. falciparum infection on day 63 after AL treatment of 41% and a reduction of this risk to 20% in the intervention arm [This study was powered assuming that the risk of tion arm , a totalAssuming a loss to follow up rate of 20%, the sample size will be increased to 403 across the sites in Ethiopia and Bangladesh where AL is used. We aim to recruit between 300 and 350 patients in Ethiopia, up to 50 in Bangladesh depending on the speed of recruitment.We further\u00a0aim to recruit 100 patients in Indonesia, which uses the schizonticidal treatment DHA-PIP. If we do not observe between site differences then estimates from the three sites will be combined giving >99% power for the primary outcome and additional power for the secondary outcomes.Patients will be recruited at established clinical trial sites in malaria-endemic areas. Recruitment was initially planned over a period of 1 year but was extended due to COVID-19-related interruptions and halting of clinical trial recruitment.Patients with malaria who are G6PD normal will be allocated randomly to one of two arms. The ratio of enrolment is 1:1 and randomization is done in blocks of 8.Sealed envelopes are prepared by an independent statistician before the study starts. Individual envelopes will only be opened after the screening is completed for the patients qualified to be enrolled into the study.Allocation sequence is generated by an independent statistician. Enrollment and assignment of participants is done by qualified study staff who have received delegation by the study PI to conduct these tasks.Neither participants nor study staff will be blinded to the allocation.Since the study is not blinded no procedures for unblinding are required.Only patients with a G6PD activity \u2265 70% of the adjusted male median (AMM) as determined by the Biosensor  will be eligible for enrolment into the main trial. In order to determine 100% G6PD activity at each site, a total of 30 adult males attending the health facility will be sampled. Only patients negative for malaria as confirmed by rapid diagnostic test (RDT) and without fever will be asked to participate. Separate written informed consent will be requested. A fingerpick sample  will be collected and G6PD activity measured using the Biosensor. No follow-up of those participants will be required.For screening purposes, a finger-prick blood sample will be collected and used for malaria slide, Hb measurement and G6PD testing using the Biosensor. Women between the ages of 13 to 49 years willing to participate will be asked to take a urinary pregnancy test. Participants who have a G6PD deficient result will be informed of their status and will be provided with information on G6PD deficiency and its consequences in everyday life and consecutive malaria infections.Eligible patients who provide written informed consent will be enrolled into the trial. Following enrolment, a brief questionnaire will be completed including demographic information, medical history, treatment history, history of recent blood transfusions, and a brief physical examination will be performed. 7.5 ml of venous blood will be collected in all consenting participants, and 450 \u03bcl of capillary blood will be collected in those who decline venous blood collection. Blood will be used for subsequent host and parasite analyses. All participants will receive the first dose of schizonticidal treatment under supervision. Patients in Bangladesh and Ethiopia who receive AL will be instructed to take the second dose at home. Patients in the intervention arm will receive their first dose of PQ.Baseline (pre-dose) urinary sample (not more than 50 ml) will be collected from the participants  before the administration of high-dose PQ. Post-dose urinary sample (not more than 50 ml) will also be collected from the participants in the intervention arm within 3\u20136 h following PQ administration.Baseline (pre-dose) metHb level will be measured using a non-invasive Masimo MetHb device before the administration of high-dose PQ. The metHb level will also be measured following the dosing of PQ with 30-min interval up to 4 h (Indonesia site only).All participants will be reviewed and the next dose of schizonticidal treatment will be provided. Patients in Bangladesh and Ethiopia who receive AL will be instructed to take the second daily dose at home. Patients in the intervention arm will receive their second dose of PQ. Capillary blood will be collected, a malaria smear prepared and Hb measured.All participants will be reviewed and treated with schizonticidal treatment under supervision. Patients in Bangladesh and Ethiopia who receive AL will be instructed to take the second daily dose at home. Patients randomized to the intervention arm will receive their third dose of PQ treatment under observation. Capillary blood will be collected, a malaria smear prepared and Hb measured. The remaining sample will be used to assess G6PD activity using the Biosensor.Participants in the intervention arm will be reviewed daily and treated with supervised PQ. Participants will be screened for any evidence of haemoglobinuria (dark urine) or adverse symptoms. If there are clinical concerns of haemolysis, a capillary blood sample will be taken for Hb measurement. An aliquot of the sample on day 6 will also be used to confirm drug levels. Additional more intense sampling for drug levels will be done on day 5 in Ethiopia only. For those patients, a quantitative G6DP test using the Biosensor will also be done. Patients in the control arm will not have scheduled visits before day 7.All participants will be reviewed again on day 7 and weekly thereafter, or on any day when they have further symptoms, until day 63. A symptom questionnaire and a brief physical exam will be performed. A capillary blood sample will be collected for immediate Hb measurement and blood film examination. The remaining capillary blood will be stored in a microtainer. On days 7, 28 and 63, the same sample will be used for repeat testing of G6PD activity using the Biosensor. No more than 450 \u03bcl of capillary blood will be collected at each visit. In Indonesia, not more than 3 ml of venous blood will be collected on day 63.All participants with fever or symptoms indicative of malaria will be asked to return to the same health centre as on enrolment. A symptom questionnaire will be completed, a brief physical examination undertaken, and a blood film will be examined for malaria diagnosis (which will be read immediately). In those patients diagnosed with recurrent parasitaemia, a 7.5-mL (1 tube) venous blood sample will be collected, Hb concentration will be measured, and the remaining blood will be stored in an EDTA tube for further analysis.All patients will be counselled before enrolment about the requirements during the trial, in particular about the frequency of follow up visits. The importance of completing the follow up will be highlighted, but patients will also be reassured that they can drop out of the study at any time without providing a reason.Menzies School of Health Research, as a legal trial sponsor, will be responsible for data collection, storage, protection, retention and destruction. Source data will be collected on paper case record forms (CRF) which will be stored securely on site. The data will only be accessible to the investigators, research sponsors, study monitor, the auditor(s), regulatory authorities, ethics committee and delegated members of the study team, who will enter all clinical data. Patient data will be recorded in pseudonymized form (i.e. without reference to the patient\u2019s name) using exclusively the patient\u2019s identification code.WWARN.org) servers. The WWARN is registered with the Registry of Research Data Repositories (re3data.org).Data will be entered into the online database REDCap by users assigned individual usernames and passwords. The database includes range checks for data values and internal validation checks. In addition, logic checks will be performed as well as 10% random checks at each monitoring visit to ensure data correctness and integrity. The database will be closed, data extracted and stored on WorldWide Antimalarial Resistance Network  pellets and plasma will be separated by centrifugation and stored at \u2212 80\u00b0C and \u2212 20\u00b0C, respectively. 7.5 ml of EDTA-anticoagulated venous blood samples will be collected on D0 and day of recurrence. In the case of infants and children, a total of 0.5 ml/kg up to a total of 7.5 ml will be collected. The sample will be white blood cell (WBC)-depleted and both RBC pellets and plasma stored at \u2212 80\u00b0C and \u2212 20\u00b0C, respectively, for further laboratory analyses. A total volume of 12.5\u201320 ml blood will be collected within 63 days; this volume is well within the WHO guidelines for clinical trials.Slides for microscopy will be collected on all scheduled and unscheduled visits. Slides will be read immediately. Quality control and assurance will be performed according to standard SOPs.Hb will be measured in all patients at enrolment, days 1 and 2. In the intervention group, Hb will also be measured on any day between days 3 and 6 when clinically warranted. Subsequently, Hb will be measured weekly from day 7 onwards. Hb will be measured using a Hemocue\u2122 Hb  machine or another rapid diagnostic test such as the Carestart Hb machine .One hundred percent G6PD activity at each site will be determined before the start of the study using Biosensor  readings from 30 male adults.In the trial, G6PD activity will be measured quantitatively using the Biosensor on day 0 for screening. Further Biosensor readings will also be done on days 2, 7, 28 and 63 and opportunistically if blood is collected in the intervention arm between days 3 and 6.A urine \u03b2-HCG pregnancy test will be conducted on all women aged 13\u201349 years who are screened for the study. Anyone testing positive will be excluded from the study.Cell-free Hb (CFHb) will be analysed on samples collected on day 0. For this purpose, whole blood samples will be double spun to remove platelets. Samples will then be stored frozen and further processing will be done at the Menzies School of Health Research, Darwin, Australia.LDH/HRP2 ELISA assays for the assessment of the total biomass will be done from samples collected on day 0 and the day of recurrence. Plasma samples will be stored frozen and further processing will be done at the Menzies School of Health Research, Darwin, Australia.Plasmodium spp. detected will be typed for genetic fingerprinting (to determine new or different parasites from previous or later infections), and known and putative molecular markers of drug resistance. Whole-genome sequencing and molecular typing analyses of parasites will be done by using new-generation high throughput platforms  at the Sanger Institute, UK, or similar techniques at other facilities.Sub-patent infections  are common and can result in anaemia, increased risk of recurrent symptomatic parasitaemia, and ongoing transmission. Parasite DNA will be extracted from RBC pellets for PCR analysis. PCR will be undertaken for speciation and quantitation of sub-patent infections and genotyping. All Human DNA will be extracted from collected samples. All samples will be assessed using molecular techniques for known and unknown variants of the G6PD gene (Xq28) and other candidate markers related to malaria susceptibility and treatment outcomes such as alpha and beta thalassaemia, Gerbich and Ovalocytosis.Red blood cell polymorphisms will be assessed by molecular analysis. CYP2D6 polymorphisms which are known to affect PQ metabolism will also be determined by host genotyping. Where possible, all tests will be undertaken in-country. However, where the laboratory does not have the capacity, samples will be transferred to a reference laboratory.P. vivax recurrence. In addition, in Ethiopia  up to 3 random samples 1 to 6 h post-treatment will be collected on day 5 to collect detailed pharmacokinetic (PK) data. Samples will be capillary samples collected into microtainers (max 100 \u03bcl per sample). Drug concentrations will be measured at the Mahidol Oxford Research Unit, Bangkok, Thailand, or an institution with a similar capacity.Blood collected on day 6 and day of recurrence will be used to measure the plasma concentrations of PQ and its metabolites (such as carboxyprimaquine), piperaquine, lumefantrine and desethyl-lumefantrine. Day 6 concentrations will be correlated with the risk of Blood collected from capillary or venous sampling on baseline, days 7, 28 and 63 will be used for serological analysis to quantify antibody levels and complement fixation and to assess how these are correlated with treatment efficacy. Antibody levels against other infectious diseases (except HIV) that may be associated with malaria immune responses will be measured. Samples will be processed on site and stored at \u2212 80\u00b0C and shipped on dry ice to the Menzies School of Health Research, Darwin, Australia.Methylated Hb levels will be measured in patients at the Indonesian study site only. A non-invasive monitoring platform (Masimo) will be used. Measurements will be taken by placing a sensor on the fingertip of the patient. The measurements will be taken just before the daily PQ dose and continued up to approximately 4 h in 30-min intervals making a total of 9 measurements in total for 3 days.To provide a pragmatic comparison of the different drug treatments, the principle of intention-to-treat (ITT) will be the main strategy of analysis adopted for the primary and secondary endpoints. These analyses will be conducted on all patients assigned to the treatment groups as randomized, regardless of the study treatment received. An analysis based on a per-protocol (PP)\u00a0approach will be conducted following the causal framework presented in Hernan & Robins . The folIneligibility (either arising during the study or retrospectively having been overlooked at screening)Significant protocol deviation Incomplete treatment courseA causal diagram will be drawn to identify any pre- or post-randomization confounders to guide the analysis approach.The safety data will be analysed using a safety data population comprising all patients who received at least one dose of treatment.P. vivax parasite recurrence within 63 days of follow-up (primary outcome) will be calculated using the Kaplan-Meier (KM) method for each trial arm as well as a comparison of the relative hazards between trial arms (hazard ratio (95% CI)) estimated from a Cox regression model for the time to the first recurrent episode with stratification for study site. The first episode of P. vivax (including mixed infections with P. vivax) will be treated as a failure endpoint. Patients presenting with parasitaemia other than P. vivax will be censored at the day of occurrence.The incidence risk (95% CI) of P. vivax parasitaemia on day 63; any P. vivax parasitaemia on days 28 and 42; any P. falciparum parasitaemia on days 28, 42 and 63; and any P. falciparum gametocytaemia on days 7 and 63.Comparisons will be made between the intervention and control arm. The analyses will be performed in the ITT and the PP population with stratification for the site. The same analysis approach will be used for the secondary outcome measures: symptomatic For safety data, the proportion and number of adverse and serious adverse events will be presented by intervention and control arms. Values and changes from baseline especially for Hb will be calculated for each visit by the treatment group.More details on the analysis of primary and secondary outcomes and safety analyses are provided in Supplement No formal interim analysis is planned, unless otherwise advised by the Data Safety Monitoring Board (DSMB).No a priori subgroup analyses are planned.Patients can have an incomplete course of treatment or data on drug administration may be missing. No imputation of treatment courses will be made for patients with missing data. For patients with missing data on AEs, the most conservative approach will be used. Rules for the adjudication of microscopy related endpoints as well as safety endpoints can be found in the Statistical Analysis Plan  the principal investigator, responsible for the overall trial conduct, (ii) the data manager, who is responsible for database development and maintenance, (iii) the study coordinator, responsible for trial registration, training of study staff on site, monitoring and logistical issues as well as annual reporting and (iv) the statistician, responsible for data analysis .No trial steering committee has been appointed.A DSMB has been appointed for this study. The DSMB is responsible for safeguarding the interests of study participants, assessing predominantly the safety of study procedures, and for monitoring the overall conduct of the study. The DSMB members are independent and it is their responsibility to prevent patients being exposed to any excess risks by recommending trial suspension or termination early if the safety or efficacy results are sufficiently convincing. The responsibilities of the DSMB are (i) to determine how frequently interim reviews of trial data should be undertaken, (ii) to conduct interim safety data reviews, and (iii) to report (following each DSMB meeting) to the sponsor and to recommend whether the trial should continue, the protocol be modified or the trial be stopped.Reports of specific serious adverse events (SAEs) are sent for review to the DSMB as they occur. This includes SAE reports of patients with acute haemolysis defined as Hb<7g/dl, anyone requiring blood transfusion or macroscopic haemoglobinuria (Hillmen \u22655) plus a fractional fall in Hb \u226525% from baseline, independent of the study arm. This will also include any SAEs which the investigator categorized as possibly or definitely related to the study drug.Quarterly reports are sent to the DSMB for review including (i) all SAE reports; (ii) line listings of all adverse events stratified by severity and study arm; (iii) a flow diagram with the number of screened and enrolled patients, patients who have been withdrawn or lost to follow-up; (iv) line listing of all protocol violations, v) individual patient Hb data between day 0 and day 14; (vi) the number of patients with fall in Hb \u2265 25% or reporting microscopic haemoglobinuria.Further details about the DSMB charter may be obtained upon reasonable request.All participants with SAEs will be reviewed with a standard questionnaire. Patients suspected of severe malaria or sepsis will be treated according to local hospital guidelines.All patients with an SAE will be assessed to ascertain malaria and haematological status. Relevant investigations will be ordered according to the attending physician. All results of clinical investigations will be collated on an SAE report form.All SAEs will be reported by the site investigator to the study coordinator within one working day of awareness, who will, in turn, notify the DSMB and PI within 48 h of their notification of the event. The site PI will be responsible for notifying the local Ethical Committees as appropriate. All information received for a case will be detailed on a full SAE report form.Adverse events will be recorded for up to 42 days after the last day of administration of PQ. All related AEs that result in a participant\u2019s withdrawal from the study, or are present at the end of the study, will be followed up until a satisfactory resolution occurs. It will be left to the Investigator\u2019s clinical judgement whether or not an AE is of sufficient severity to require stopping the participant\u2019s treatment. A participant may also voluntarily withdraw from treatment due to what he or she perceives as an intolerable AE. If either of these occurs, the participant must undergo an end of study assessment and be given appropriate care under medical supervision until symptoms cease or the condition becomes stable.An increase of >20% in metHb level of a subject will be recorded as an AE and the subject will be closely monitored. The subject will receive the treatment of acute methemoglobinemia according to the increase and their clinical features (Indonesia only).Malaria can cause a temporary fall in Hb that can be exacerbated by PQ. All patients will be monitored closely for acute recovery from their initial episode of malaria, any adverse drug reactions, and subsequent recurrence of malaria. In patients suspected of acute haemolysis, the clinical judgement of the attending clinician and patient safety should always come first, regardless of the trial protocol.A detailed monitoring plan has been prepared. In brief, it states that a minimum of 2 monitoring visits  will be conducted during the course of the study. If virtual monitoring, each monitoring visit will take place using online video conferencing and secure, encrypted file sharing. No identifying information will be stored on the server. Monitoring frequency may be increased based on the following criteria: (i) high subject enrolment at a particular site, (ii) compliance issues (e.g. significant protocol violations), (iii) site staff turnover, requiring additional training, and (iv) results and findings from previous monitoring visit. Detailed procedures are captured in the monitoring plan, which can be requested. Auditing can also take place by national health authorities at the participating study sites.Changes to protocol modification will be communicated promptly to all involved parties and updated in the clinical trials registry. Any protocol deviations will be fully documented.Results will be published in peer-reviewed journals. All Investigators will be involved in reviewing drafts of the manuscripts, abstracts, press releases, and any other publications arising from the study. Authorship will be determined in accordance with the ICMJE guidelines and other contributors will be acknowledged. Results will further be disseminated at international conferences and other stakeholder meetings including National Malaria Control programs in the region. All efforts will be made to disseminate results to study participants in line with local requirements.P. vivax put a strain on healthcare systems. A unified treatment policy for malaria has potential to confer significant individual, public health and operational benefits in regions co-endemic for P. falciparum and P. vivax. The optimal use of limited financial resources could necessitate strategies for treating P. vivax through universal radical cure.Recurrent episodes of Recruitment started on 18 August 2019 at the study site in Bangladesh and has been completed. The trial was paused in 2020 due to the COVID19 pandemic. Recruitment at the study in Ethiopia started on 27 January 2021 and on 1 June 2021 at the Indonesian site. The study is anticipated to be completed in 2022. The current protocol version number is V6.0.Additional file 1. Rationale for study.Additional file 2. Participant timeline.Additional file 3. Statistical Analysis Plan."}
+{"text": "Asymptomatic Plasmodium carriers form the majority of malaria-infected individuals in most endemic areas. A proportion of these asymptomatically infected individuals carry gametocytes, the transmissible stages of malaria parasites, that sustain human to mosquito transmission. Few studies examine gametocytaemia in asymptomatic school children who may form an important reservoir for transmission. We assessed the prevalence of gametocytaemia before antimalarial treatment and monitored clearance of gametocytes after treatment in asymptomatic malaria children.P. falciparum parasitaemia by microscopy. One hundred and fifty-five (155) parasite positive children were treated under direct observation with dihydroartemisinin-piperaquine (DP). Gametocyte carriage was determined by microscopy seven days prior to treatment, day 0 before treatment, and on days 7, 14 and 21 post initiation of treatment.A total of 274 primary school children were screened for p\u00a0=\u00a00.05) and asexual parasite density (p\u00a0=\u00a00.08). In a variate analysis, persistent gametocytaemia 7 or more days after treatment was significantly associated with post-treatment asexual parasitaemia at day 7 (P\u00a0=\u00a00.027) and presence of gametocytes on the day of treatment (P\u00a0<\u00a00.001).The prevalence of microscopically-detectable gametocytes at screening (day \u22127) and enrolment (day 0) were 9% (25/274) and 13.6% (21/155) respectively. Following DP treatment, gametocyte carriage dropped to 4% (6/135), 3% (5/135) and 6% (10/151) on days 7, 14 and 21 respectively. Asexual parasites persisted in a minority of treated children, resulting in microscopically detectable parasites on days 7 , 14  and 21 . Gametocyte carriage was inversely correlated with the age of the participants (Though DP provides both excellent cure rates for clinical malaria and a long prophylactic half-life, our findings suggest that after treatment of asymptomatic infections, both asexual parasites and gametocytes may persist in a minority of individuals during the first 3\u00a0weeks after treatment. This indicates DP may be unsuitable for use in mass drug administration strategies towards malaria elimination in Africa. The usesmission , consistse of DP . As a rese of DP .P. falciparum infections, treated with DP. These data are from a major study evaluating P. falciparum gametocyte immunity in Ghanaian school children  and the London School of Hygiene and Tropical Medicine Ethics Committee (Ref: 2010_5775). In addition, individual informed consent and assent as well as community consent were obtained before the study was conducted. Approval was also obtained from the Ahafo Ano South District Education Directorate and the authorities of the Pokukrom Methodist Primary School since the study involved school children. Given that primary school children represent an age group that require written informed consent to be obtained from their parents before participation, community consent was obtained as part of the community entry to introduce the study and the respective class teachers served as guardians.2.2The study participants were primary school children between the ages of 6 and 12\u00a0years attending the Methodist School in Pokukrom in the Ahafo Ano South District of the Ashanti Region of Ghana. Pokukrom is within the Ahafo Ano South district and it is an area of high malaria transmission, and with two rainy seasons; a major wet season from April to July and a minor one from September to November . This pa2.3via finger-prick blood for microscopy after clinical examination by qualified trained nurses. The inclusion criteria were parasite positivity of any Plasmodium species including mixed species infections, any parasitaemia levels without any obvious signs and symptoms of malaria. The main exclusion criteria were malaria symptoms, including recent history of fever, signs and symptoms of chronic and severe diseases and malaria treatment in the last two weeks . Asymptomatic malaria-infected children were defined as children who were blood slide positive but without any clinical symptoms and signs of malaria upon clinical examination. The study design, participants, inclusion and exclusion criteria, environment and sampling procedures have been previously described . Brieflywo weeks .2.4Thick and thin blood smears prepared in the field at each visit were stained with 10% Giemsa the same day upon returning to the laboratory at the Department of Biology, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana. Double parasite readings were independently carried out by counting the number of parasitized erythrocytes per 200 white blood cells for asexual parasites but against 500 white blood cells in the case of sexual parasites. The parasitaemia obtained were expressed as parasites per microliter of blood by assuming there are 8000 leucocytes in one microliter of blood . The thi2.5Plasmodium species composition during follow-up has been reported previously ) at visit 2. by direct observation and supervision by a clinically qualified nurse during school after meals, according to Ghana Government treatment guidelines and recommendations of the approving ethics review committee. Thus, children were enrolled one week after screening, once their test results were known. As it is not national policy to delay treatment of asymptomatic children for a week, in between the screening and enrollment days nurses were at school to monitor children for possible symptoms of malaria such as rising temperatures. Those who showed symptoms of malaria during this time were advised and referred to the health facility. On the day of enrolment  all parasite positive children who were present at school were given treatment and those who were absent were advised to visit the health facility upon their next attendance in school and these were not included in the study. Enrollees were followed up weekly on TD7 (visit 3), TD14 (visit 4) and TD21 (visit 5) with finger-prick blood collections for blood smears on slide, filter paper blood spots and plasma separation. Similarly, nurses were available on all days to monitor the school children during the follow-up to ensure complete compliance of treatment and possible symptoms of malaria. Microscopy and PCR detection of asexual parasite carriage and eviously .2.6Participant data were recorded manually on an enrollment/case report form capturing demographic data, asexual parasite and gametocyte counts which were then entered into a customized Microsoft Access database. The data were double-entered independently to allow correction of errors and inconsistencies and securely backed up. The outcome measures were gametocytaemia at enrollment, the development and loss of gametocytes during follow-up.t-test was used to determine differences in geometric mean densities of sexual and asexual parasites when normally distributed and in the geometric mean age of gametocyte and non-gametocyte carriers. The Wilcoxon rank-sum test was used to test for association between asexual parasite density and gametocyte prevalence. Age was considered as a continuous variable while asexual parasite and gametocyte prevalence were analysed as binary variables. Asexual parasite and gametocyte densities were considered as continuous variables. Sample size determination was based on previous known asexual parasite prevalence of 65% in a nearby study area and gametocyte prevalence of 25% in a similar endemic area within West Africa. Logistic regression models were used to determine factors associated with gametocyte carriage and persistence over different sampling times. All analyses were performed using Stata software .Geometric mean of the independent microscopy readings was used for analyses. Estimations of gametocyte and asexual parasite prevalence on the day of screening (day \u22127) was based on the full sampled population and subsequent longitudinal analyses considered only those who were parasite-positive on day 0, and thus received treatment. Analyses comparing different days were based on per protocol data. Student's 33.1p-value\u00a0=\u00a00.127 (P\u00a0=\u00a00.446). Similarly, there was no significant difference statistically in the geometric mean of asexual parasite densities between the day of screening (geometric mean\u00a0=\u00a074.69 (CI: 53.16\u2013104.95) and the day of enrolment (geometric mean\u00a0=\u00a092.48 (CI: 58.65\u2013145.82), p-value\u00a0=\u00a00.283 , 181 (66%) were parasite-positive by microscopy. Of these, 155 remained parasitaemic the following week at visit 2 and received the 3-day treatment with DP (TD0). These were followed up weekly for 3\u00a0weeks until visit 5 (TD21). The mean age for enrolled and treated children was 10\u00a0years  while th\u00a0=\u00a00.127 . No evid\u00a0=\u00a00.283 . One hun\u00a0=\u00a00.283 .  of all participants did not carry gametocytes at any point in time. Eleven percent (13/116)  of them a priori, could potentially be associated with post-treatment gametocyte carriage. Age in years, stated gender, asexual parasite density in peripheral blood films at visit 1 or visit 2 and gametocyte density in peripheral blood films at visit 1 were not associated with gametocyte carriage in follow-up. However, persistent asexual parasites at TD7 (visit 3), pre-treatment gametocyte density at visit 2 (TD0) and presence of gametocytes at either visits 1 or 2 were found to have associations with gametocyte carriage in follow-up, and were deployed together in a multiple logistic regression model to identify which, if any of these inter-related parameters might dominate as a risk factor (Exploratory univariate analyses were carried out for risk factors that, k factor . Of thes4P. falciparum and treated with dihydroartemisinin-piperaquine. This is ancillary to a major study of antibody responses to P. falciparum gametocytes (Here, we examine potential risk factors for gametocyte carriage in a longitudinal study where gametocytaemia was monitored in a cohort of asymptomatic school children infected with etocytes . Microscetocytes . Asexualin vitro and in vivo but has incomplete activity against stages IV and V gametocytes (Anopheles mosquitoes in membrane-feeding assays (In this study we found a microscopic gametocyte prevalence of 13.6% among the asymptomatic enrolled population. In addition, there were some newly identified gametocyte carriers after DP treatment. Most antimalarial drugs including piperaquine do kill the earliest developmental stages of gametocytes but the later stages are unaffected . Dihydroetocytes . Other sg assays . Our obsg assays , we assude novo gametocyte production. In this context our observation of a positive association between \u201cpersisting\u201d asexual parasitaemia and the likelihood of gametocytaemia 1 to 3\u00a0weeks after DP treatment is relevant and needs verification in future studies designed to measure drug efficacy.The persistence of gametocytes after treatment is likely to reflect both persistent and newly-emergent gametocytes following ACT treatment, the latter indicating the presence of some persisting asexual parasites leading to At least 5% of the children at any sampling time point harbored detectable gametocytes, but estimated densities fluctuated owing either to the low sensitivity of microscopy in detecting gametocytes or to emergence of new gametocytes. Gametocytaemia was found to be inversely correlated with the age of the school children as shown in previous studies . We alsoDespite observations made in this study, there are several studies showing evidence of efficacious treatment outcomes of DP in symptomatic individuals from different malaria endemicities. In an area of unstable malaria transmission in Sudan, It should be noted that our study was not designed as a clinical trial and, being conducted in asymptomatic children, there was no drug efficacy end point. Despite the above caveat, it is still unclear why our study demonstrates markedly different clearance kinetics of asexual parasites, and suggests that DP is less effective in the absence of a high-level anti-parasitic immune response as evidenced by the presence of malaria symptoms. Other possible explanations for our findings include poor drug absorption or sub-standard medicines. We have evidence to rule out the latter as tests of drug quality revealed that the batches of DP used in the study met the required standards .in vivo.The primary limitation of this study is our reliance on standard microscopy for gametocyte detection, which is known to underestimate gametocytaemia . Detecti5In this study we showed that microscopically detected gametocytes are abundant in school children and persistently fluctuated between sampling time points. Asexual parasites also persisted in a minority of treated individuals. These data imply that asymptomatic children in populations receiving MDA with DP could be an important reservoir of both persisting asexual parasitaemia and post-intervention transmission. Further studies deploying molecular characterization of gametocytes in asymptomatic individuals are required to understand why gametocytes and asexual parasites persist after DP treatment. These findings will have important implications for successful deployment of DP in community treatment campaigns to reduce malaria transmission.BD and CJS conceived and designed the study. DA, NKB and JAL contributed to fieldwork. BD performed the experiments and analysed the data with TB and CJS. BD wrote the first draft of the paper. All authors read and approved the final manuscript.The data used and or analysed during the study are available from the corresponding author on reasonable request.The study was approved by the Ghana Health Service Ethics Committee and the Ethics Committee of the London School of Hygiene and Tropical Medicine. Informed consent was obtained from all individuals included in this study.10.13039/501100003419Ghana Education Trust Fund. The research and field studies were funded by the 10.13039/100009660London School of Hygiene and Tropical Medicine, UK.Bismarck Dinko was funded by a PhD Fellowship from the The authors declare that there are no competing interests."}
+{"text": "Plasmodium and is still one of the most important infectious diseases in the world. Several biological characteristics of Plasmodium vivax contribute to the resilience of this species, including early gametocyte production, both of which lead to efficient malaria transmission to mosquitoes. This study evaluated the impact of currently used drugs on the transmission of P. vivax. Participants received one of the following treatments for malaria: i) chloroquine [10 mg/kg on day 1 and 7.5 mg/kg on day 2 and 3] co-administered with Primaquine [0.5 mg/kg/day for 7 days]; ii) Chloroquine [10 mg/kg on day 1 and 7.5 mg/kg on day 2 and 3] co-administered with one-dose of Tafenoquine [300 mg on day 1]; and iii) Artesunate and Mefloquine  co-administered with Primaquine [0.5 mg/kg/day for 14 days]. Patient blood was collected before treatment and 4 h, 24 h, 48 h and 72 h after treatment. The blood was used to perform a direct membrane feeding assay (DMFA) using Anopheles darlingi mosquitoes. The results showed 100% inhibition of the mosquito infection after 4 h using ASMQ+PQ, after 24 h for the combination of CQ+PQ and 48 h using CQ+TQ. The density of gametocytes declined over time in all treatment groups, although the decline was more rapid in the ASMQ+PQ group. In conclusion, it was possible to demonstrate the transmission-blocking efficacy of the malaria vivax treatment and that ASMQ+PQ acts faster than the two other treatments.Malaria is caused by parasite of the genus  P. vivax present in infected people is important to block malaria transmission from human to mosquito vector. Here, we assessed the potential of currently used antimalarial drugs for vivax malaria therapy for blocking malaria transmission in the mosquito vector An. darlingi. We observed that the antimalarials tested were able to block the infection of mosquitoes; however, treatment based on the artemisinin combination proved to be the fastest for blocking transmission in mosquitoes, using blood taken 4 h after treatment. As such, patients in endemic areas who receive the first line vivax malaria treatment, chloroquine and primaquine, could need protection from bites of the malaria vector for at least 24 h after the initial dose in order to interrupt malaria transmission. Our observations provide an overview of the transmission-blocking profile of antimalarials currently used in Brazil for the treatment of vivax malaria and they also provide a platform for the evaluation of new antimalarials, such as tafenoquine or other compounds for combating the sexual stages of P. vivax, which may impact the epidemiology of malaria.Vivax malaria is the most prevalent and widespread human malaria outside the African continent. The control of vivax malaria is based on vector control and effective radical cure of infected people. However, the elimination of the sexual stages of  Plasmodium, which remains of global public health significance  co-administered with Primaquine [0.5 mg/kg/day for 7 days]; ii) Chloroquine [10 mg/kg on day 1 and 7.5 mg/kg on day 2 and 3] co-administered with one-dose of Tafenoquine [300 mg on day 1]; and iii) Artesunate and Mefloquine  co-administered with Primaquine [0.5 mg/kg/day for 14 days], were identified as the following groups: CQ+PQ, CQ+TQ and ASMQ+PQ, respectively .The CQ+PQ group were enrolled before the inclusion of tafenoquine as a vivax malaria treatment in Porto Velho, and the CQ+TQ group were enrolled from October 2021 onwards, when tafenoquine was included as a vivax treatment in Porto Velho, Rondonia state, and Manaus, Amazonas state, after a previous glucose-6-phosphate dehydrogenase (G6PD) test ,28. PatiAll drugs were administered orally with food, and were provided without cost to all participants. The first blood collection (0 h), considered the control, was conducted at CEPEM before initiating the treatment, and the first dose of treatment was initiated immediately after blood collection (under supervision at the recruitment clinic). After initiation of treatment, all subsequent blood collections  were carried out by a visiting health worker at the volunteers\u2019 residence. The collections were carried out before administration of the drug and under full supervision .Blood samples were collected in heparinized (9 mL) Vacutainer tubes, maintained in a water flask at 37\u00b0C and immediately transported to PIVEM, which took less than 40 minutes. At PIVEM, the heparinized blood was used for the direct membrane feeding assay (DMFA) and a thick blood smear was prepared in duplicate using 10 \u03bcL of whole blood for quantification of parasitemia and gametocytemia via microscopy.Thick blood smears from each patient and from each timed blood collection were prepared then, Giemsa stained and examined under light microscopy (x100) with an oil immersion lens. The density of the gametocyte and asexual stages were estimated by counting parasites per 800 leukocytes according to WHO (2010) .An. darlingi colony used in this study was established in 2018 by Araujo et al. ; and the inhibition intensity: 100 x [1 \u2013(mean number of oocysts in the test group)/(mean number of oocyst in the control group)]. Positive values denote a reduction in the percentage of mosquitoes infected and the number of oocysts per mosquitoes, negative values denote increases.Twenty-four vivax malaria volunteers were enrolled in our study: CQ+PQ (n = 11), CQ+TQ (n = 7) and ASMQ+PQ (n = 6). However, 5 volunteers from the CQ+PQ group and 1 volunteer from the CQ+TQ group were excluded from study because they were not at home during a visit, or the assays could not be completed due to COVID-19-related constraints. Additionally, experiments which the mean oocyst account in the control group was less than 2.5 were excluded of analysis  due the Participant characteristics were similar between the treatment groups, and these are presented in The number of mosquitoes dissected per group varied between 39 to 40. Before treatment (0 h), all participants were infectious to mosquitoes .An. darlingi , none of the participants in the CQ+PQ and ASMQ+PQ groups infected any mosquitoes , while tRegarding parasitemia density, 91.7% of patients had gametocytemia on admission and a median of 5,445 asexual parasites/\u03bcL. Individuals who were gametocytemia negative at 0 h (before beginning of treatment) infected mosquitoes . In geneP. vivax parasites and blocking human-to-mosquito transmission when compared with chloroquine [P. falciparum treatment, artemisinin derivates prevent gametocyte development, while primaquine is the only drug known to act on mature gametocytes [P. vivax [P. vivax in therapeutic doses [P. vivax gametocytes to mosquitoes [An. darlingi mosquitoes.In this study, we assessed the efficacy of the transmission blocking by using a DMFA with the blood from patients who received first-line treatment with CQ+PQ, patients who participated in TQ implementation studies and received CQ+TQ, and patients who returned to the clinic with vivax malaria from days 14 to 60 after first-line treatment and received ASMQ+PQ. Our data showed that only one of four patients who were treated with ASMQ+PQ were able to infect mosquitoes for up to 4 h. While three of four patients treated with CQ+PQ or CQ+TQ were able to infect mosquitoes for up to 4 h. However, only patients treated with CQ+TQ remained infective to mosquitoes at 24 h after initiation of treatment. Artemisinin compounds have already been described as faster in clearing asexual oroquine ,34\u201337. AP. vivax  and alsosquitoes . Klein esquitoes  showed tP. vivax infection due to the rapid appearance of gametocytes in the peripheral blood. Unlike P. falciparum, P. vivax gametocytes tend to be already present when patient seek diagnosis and treatment [P. vivax gametocyte (a maximum of 3 days) [The transmission-blocking activity of antimalarial drugs is an important determinant of their impact on public health. Early diagnosis and treatment of malaria are essential for malaria control, mainly for reatment ,41,42. D 3 days) , the gamP. vivax, was approved and has been implemented in two cities of Brazil (Manaus and Porto Velho) [P. vivax. Tafenoquine exerts an extraordinarily broad spectrum of activity including against the exo-erythrocytic liver and erythrocytic asexual forms, sexual stages and sporogonic development in the mosquito host [An. darlingi. We found that three of four treated patients maintained the infection for up to 24 h, with the infection intensity ranging from 1 to 58 oocysts per midgut. Gametocytes were detected for up to 48 h and asexual parasites for up to 24 h. Tafenoquine is a slowly eliminated 8-aminoquinoline  that contrasts with the very rapid elimination of primaquine  [P. vivax as it may prevent relapses of P. vivax malaria [P. vivax transmission.Recently in Brazil, tafenoquine, a new drug for the radical cure of o Velho) ,21,28. Tito host \u201349. In o of 4 h) . The lonP. vivax in Brazil (CQ+PQ) showed transmission blocking of < 24 h, preventing the risk for transmission of parasites to the mosquito vector the night after taking the first dose. Therefore, protection of patients from bites of malaria vectors for at least 24 h after the initial dose should be used to interrupt the man-mosquito cycle malaria.It was demonstrated herein that ASMQ+PQ acts faster than the two other treatments, however, this treatment is available for patients who presented recurrence, which means that these patients had already received the first-line treatment up to 60 days previously. The first-line treatment for P. vivax treatment in Brazil. A few studies have systematically characterized the gametocytocidal or sporontocidal properties of new antimalarial drugs [Data from CQ+TQ treatment should be continue investigated, considering that it is a new drug and because it is included as an option for al drugs ,20,49. TS1 Table(XLSB)Click here for additional data file.S2 Table(XLSB)Click here for additional data file.S3 Table(XLSB)Click here for additional data file."}
+{"text": "Plasmodium falciparum malaria were randomized to receive ASAQ or AL for three days (1:1). 347/352 (98.5%) randomized patients reached the study endpoint on day 28. Crude adequate clinical and parasitological response (ACPR) rates were 100% (95% CI: 98.8\u2013100%) in the ASAQ group and 96% (95% CI: 93.1\u201398.9%) in the AL group (per protocol population). However, the PCR-corrected ACPR rate was 97.7% (95% CI: 95.4\u2013100%) in the AL group. Two cases of recrudescence and three of re-infection were observed. Mild and moderate adverse events, including gastrointestinal and/or nervous disorders, were reported in 11.9% (42/352) of patients. We found that ASAQ and AL were safe and efficacious for treating uncomplicated P. falciparum malaria. They may be used for treatment at health facilities and at the community level, and for mass drug administration campaigns.Malaria is a major public health problem in Madagascar, particularly in coastal areas. We conducted a randomized, controlled, parallel-group study of artemisinin-based combination therapy (ACT) in Mananjary and Farafangana, two localities on the rainy south-east coast of Madagascar, from March to September 2018. The efficacy and safety of artesunate + amodiaquine (ASAQ) and artemether + lumefantrine (AL) were assessed according to the WHO protocol with a 28-day follow-up. Children aged 6 months to 14 years with uncomplicated  Plasmodium falciparum is the predominant species [P. falciparum malaria [Malaria remains a major public health problem on the island of Madagascar, where perennial transmission occurs in coastal areas and  species , 11. Eff malaria , 19, 25. malaria , 14 and This study was conducted according to the Declaration of Helsinki, guidelines on Good Clinical Practice, and existing national legal and regulatory requirements. The study protocol was submitted and approved by the Madagascar Biomedical Research Ethics Committee (013/MSANP/CERBM) on February 26, 2018. All participants (parents/guardians and patients aged 7\u201314 years) received oral and written information concerning the study. Written informed consent was obtained from parents or guardians of all participants. If patients were unable to sign, a fingerprint was applied to the consent form. Children aged between 7 and 14 years old gave written assent to participate in the study. All signed informed consent forms will be kept for 15 years after the end of the study.P. falciparum malaria in children, based on the standard WHO protocol [This study was a multisite, randomized, controlled study to assess the efficacy and tolerability of artesunate + amodiaquine (ASAQ) and artemether + lumefantrine (AL) combinations for treating uncomplicated protocol .The study was conducted at two sites with perennial malaria transmission on the south-eastern coastal area of Madagascar . At these sites, the average annual temperature ranges from 15\u00a0\u00b0C to 32\u00a0\u00b0C, and the average rainfall ranges from 1200 to 2500 mm , 5.P. falciparum, residing in the study area for the duration of the trial, and according to the following criteria: aged between 6 months and 14 years, presenting an axillary temperature\u00a0\u2265\u00a037.5\u00a0\u00b0C or suffering from fever within the last 24\u00a0h, weighing\u00a0\u2265\u00a05\u00a0kg, having brachial circumference\u00a0>\u00a0110\u00a0mm, hemoglobinemia\u00a0\u2265\u00a08g/dL, P. falciparum mono-infection with parasite density from 1000 to 100,000 asexual forms per microliter of blood, and capable of receiving oral treatment. Both study sites are in districts with moderate malaria transmission, defined as 50\u2013100 cases per 1000 population per year [Patients were included between March and September 2018 at a basic health center at each site. The study included patients infected with per year . Patientper year , 30 or aPatients were randomized to one of two treatment groups, namely once daily ASAQ and twice daily AL. Randomization was stratified according to patient age using separate computer-generated randomization lists. Three age strata were used: 6\u201359 months, 5\u20139 years, and 10\u201314 years. The treatment information was placed in a sealed envelope. Each study participant was randomly assigned (1:1) to receive either ASAQ or AL.Treatment dosages were determined according to patient body weight , 31. Tre. Each visit consisted of a physical examination, combining evaluation of clinical safety and measurement of vital signs. Axillary temperature was measured using an electronic thermometer. Blood pressure and pulse were measured after a 10-min rest in sitting position. Blood samples were collected for parasitology (parasite count and PCR). On day 0 and day 28, hemoglobin concentration levels were measured with Hemocue\u00ae Hb201 + , from capillary fingertip blood.The WHO protocol with a 28-day follow-up to assess the therapeutic efficacy of anti-malarial drugs was used . Follow-Clinical efficacy was assessed by grading the pre-existing clinical signs and combining them with the temperature values. Parasitological efficacy was based on asexual parasitemia, although a gametocyte count was also systematically performed. The following clinical symptoms were assessed systematically at each visit and graded as absent, mild, moderate, severe, or very severe: perspiration, headache, chills, pain (specifying topography), jaundice, asthenia, dizziness, anorexia, skin fold, skin rash, and pruritus. The severity grading scales were adapted from guidelines provided by the WHO  , 27, 32.Patients, parents, or guardians were free to withdraw from the study or to stop taking treatment at any moment, irrespective of the reason. Patients were withdrawn following the investigator\u2019s decision in case of onset of any danger signs of severe malaria or any adverse event justifying treatment discontinuation. Early study withdrawal was documented by the investigator on the case report form (CRF). When possible, patients were evaluated according to the end-of-study visit procedures. Patients presenting severe disease or adverse events were referred for medical care.The study team made a phone call to remind the patient prior to each visit. At each visit, patients were given an allowance for travel expenses. In cases where a patient missed a visit, the study team visited the patient\u2019s home up to three times. If the patient was not available for the follow-up after three attempts, they were declared \u201clost to follow-up\u201d. For patients lost to follow-up, the CRF was completed up to the last visit attended.No patients prematurely withdrawn from the study were replaced.P. falciparum mono-infection and determine parasite density. Finger prick blood samples were collected on filter paper .At enrolment and at successive follow-up visits, thin and thick smears were obtained and stained with May Gr\u00fcnwald-Giemsa 10%. They were examined at the sites by dual reading to identify P. falciparum parasites after observing 500 leukocytes and then converting this figure into parasites (trophozoites and/or gametocytes) per microliter of blood, assuming an average leukocyte count of 8000/\u03bcL.Microscopy: parasite densities were calculated by counting the number of asexual and sexual For quality control purposes, all slides obtained at enrolment and at successive follow-up visits for Mananjary and Farafangana were analyzed by microscopy practitioners at IPM using a procedure that was blinded to the original result. Microscopy was considered negative if no parasites were seen after observing 1000 leukocytes.msp1, msp2, and glurp gene loci were compared. The PCR was performed using the following primer pairs MSP1: MSP1C1\u00a0=\u00a05\u2032\u2013GTACGTCTAATTCATTTGCACG\u20133\u2032 and MSP1D1\u00a0=\u00a05\u2032\u2013CACATGAAAGTTATCAAGAACTTGTC\u20133\u2032 for PCR1, MSP1C2\u00a0=\u00a05\u2032\u2013GATTGAAAGGTATTTGAC\u20133\u2032 and MSP1D2\u00a0=\u00a05\u2032\u2013GCAGTATTGACAGTTTATGG\u20133\u2032 for PCR2; MSP2\u00a0=\u00a0MSP2D1: 5\u2032\u2013GAAGGTAATTAAAACATTGTC\u20133\u2032 and MSP2C1\u00a0=\u00a05\u2032\u2013GAGGGATGTTGCTGCCAACAG\u20133\u2032 for PCR1, MSP2D2\u00a0=\u00a05\u2032\u2013GAGTATAAGGAGAAGTATG\u20133\u2032 and MSP2C2\u00a0=\u00a05\u2032\u2013CTAGAACCATGCATATGTCC\u20133\u2032 for PCR2; and GLURP: GlurpC1\u00a0=\u00a05\u2032\u2013ACATGCAAGTGTTGATCC\u20133\u2032 and GlurpC2\u00a0=\u00a05\u2032\u2013GATGGTTTGGGAGTAACG\u20133\u2032 for PCR1 and GlurpG1\u00a0=\u00a05\u2032\u2013TGAATTCGAAGATGTTCACACTGAAC\u20133\u2032 and GlurpG3\u00a0=\u00a05\u2032\u2013TGTAGGTACCACGGGTTCTTGTGG\u20133\u2032 for PCR2. Three microliters of DNA were amplified for 35 cycles during the primary PCR. Two microliters of the product of that amplification were used for another 35 cycles of PCR. Each PCR was performed with 1 \u03bcM of each primer, 200 \u03bcM of each dNTP, and 2.5 mM MgCl2. The sizes of the amplification products were determined by agarose gel electrophoresis using Quantity One\u00ae software v. 6.4.1 . Possible outcomes were i) recrudescence, if the alleles of the pre-treatment and post-treatment samples were the same for msp1, msp2, and glurp; ii) reinfection, if the alleles of the pre-treatment and post-treatment samples were distinct for any of these three loci; iii) mixed recrudescence and reinfection, if similar alleles were found in the pre-treatment and post-treatment samples for all the markers as mentioned above, but with additional distinct alleles identified; and iv) indeterminate, if either or both the pre-treatment and post-treatment samples could not be amplified. Mixed recrudescent and re-infection cases were computed as recrudescent [DNA extraction was performed from DBS using QIAGEN kits , according to the manufacturer\u2019s instructions. The analyses were performed at the Parasitology Unit of the Institut Pasteur de Madagascar. For participants with recurrent parasitemia after day 7, paired polymerase chain reaction (PCR) blots (from day 0 and the day of parasitemia recurrence) were analyzed for parasite merozoite surface proteins (MSP-1 and MSP-2) and glutamate rich protein (GLURP) to distinguish between reinfection and recrudescence, as described elsewhere , 24. DayThe number of subjects was calculated based on 28-day PCR corrected efficacy. We took into account the success rate hypothesis (95%), a confidence interval of 95%, a type I (\u03b1) error of 5% and a type II (\u03b2) error of 20%, and a loss to follow-up rate of 15%. A minimum of 86 patients per arm was needed for each study site.Data were recorded using the WHO-standard template using the ODK (Open Data Kit) system on an Android tablet and added to the database in real time. Data cleaning was done regularly to identify and fix any data that were incorrect, inaccurate, or incomplete. Statistical analysis was performed using RStudio version 4.2.2 .The primary endpoint was 28-day PCR-corrected treatment efficacy. Treatment outcomes were assessed according to the WHO guidelines  for earl\u03c72 test or Fisher\u2019s exact test, and quantitative variables with the Wilcoxon test. The uncorrected and PCR-corrected per protocol efficacy for each site and drug was calculated by dividing the number of participants classified as ACPR by all participants reaching a study outcome. The sum of posterior probabilities of recrudescence was used to calculate the total number of recrudescent infections for the PCR corrected analysis.Descriptive analysis was conducted using frequency/percentage for qualitative variables and median/interquartile interval (IIQ) for quantitative variables. Categorical variables were compared with the The PCR-corrected per-protocol population consisted of all patients without a major protocol deviation, who had a primary endpoint at day 28, who did not have any reinfection. For Kaplan\u2013Meier cumulative efficacy estimates, re-infections, lost to follow-up, and patients who withdrew were censored on the day of re-infection or the last day of follow-up. Posterior sampling was used to generate the PCR-corrected Kaplan\u2013Meier estimates and 95% confidence intervals using the posterior probabilities of recrudescence.During the study period, 352 of 1380 eligible children were randomized for inclusion in one of the two therapeutic arms, 176 in the ASAQ arm and 176 in the AL arm. Overall, 50.6% (89/176) of patients in the ASAQ group and 50% (88/176) in the AL group were female. Figure 1\u03c72 test, p\u00a0=\u00a00.1).PCR-corrected and uncorrected clinical and parasitological cure rates on day 28 using a Kaplan\u2013Meier estimate of survival curve are shown in .In the ASAQ arm, the uncorrected cumulate efficacy was 100% (95%CI: 95.9\u2013100) in Mananjary and Farafangana. Overall, the efficacy of ASAQ was estimated at 100% (95%CI: 97.8\u2013100). There were no significant differences in therapeutic response rates adjusted by region and age .At enrolment, in the AL arm the median (IIQ) parasite densities varied from 19,156 parasites/\u03bcL of blood  in Mananjary to 24,404 parasites/\u03bcL of blood  in Farafangana, and in the ASAQ arm the median (IIQ) parasite densities varied from 12,972 parasites/\u03bcL of blood  in Mananjary to 16,945 parasites/\u03bcL of blood  in Farafangana. Ten patients remained parasitemic on day 3: 0.6% (1/176) in the ASAQ arm and 5.1% (9/176) in the AL arm . The gam\u03c72 test, p\u00a0=\u00a00.3). Overall, 11.9% (42/352) patients who reached a study endpoint presented at least one emergent adverse event. Among these adverse events, 54.8% (23/42) were in the ASAQ arm and 45.2% (19/42) in the AL arm. In the ASAQ arm, the most frequently reported AEs included: cough 30.4% (7/23), abdominal pain 21.7% (5/23), vomiting 17.3% (4/23), headache 13.04% (3/23), edema 4.3% (1/23), sleep disorder 4.3% (1/23), eye trauma 4.3% (1/23) and anorexia 4.3% (1/23). In the AL arm, the most frequently reported AEs included: cough 31.5% (6/19), flu-like symptoms 21.05% (4/19), abdominal pain 10.5% (2/19), fever (10.5% (2/19), trauma 10.5% (2/19), vomiting 5.2% (1/19), headache 5.2% (1/19), adenopathy 5.2% (1/19), and anorexia 5.2% (1/19). Of all adverse events, two in the AL arm fulfilled the severity criteria. None of the adverse events were considered treatment-related by the investigators. No significant difference was observed between the two treatment groups in terms of presence of adverse events (p\u00a0=\u00a00.1) and day 28 (p\u00a0=\u00a00.5). An increase in hemoglobin values was observed for 217 individuals , a decrease for 85 individuals , and no change for 42 individuals . The means of hemoglobin level differences between day 0 and day 28 were statistically different between the 2 treatment arms .Data for hemoglobin values at day 0 and day 28 were available for 344 individuals; no statistical difference was observed between treatment arms at day 0  than in the ASAQ group  were well tolerated. The events observed during the assessment were expected and have already been described with this type of antimalarial. Abdominal pain, vomiting, headache and anorexia were the symptoms frequently reported by patients who took the artemisinin derivative in both groups. The tolerability of the treatment regimens was similar in our study. In contrast, in Cote d\u2019Ivoire for example, it was shown that AL is more tolerable than ASAQ , 34.At both sites, the uncorrected and corrected efficacy was lower in the AL group than in the ASAQ group. It is known that AL administration unaccompanied by fatty foods, as recommended by the manufacturer, may account for this difference that has been found in various studies, because both the effectiveness and tolerability of AL are better when the drug is taken with higher fat foods , 22.P. falciparum malaria in the south-eastern parts of Madagascar. We believe that ASAQ and AL may be used as first-line treatments to support the national policy plan to strengthen malaria control, with the aim of eliminating this disease in Madagascar. Also, ASAQ and AL could be used for mass drug administration to save lives during the frequent malaria outbreaks on the eastern coast of Madagascar.Our results demonstrate that the fixed-dose combinations ASAQ and AL are safe and efficacious for treating uncomplicated"}
+{"text": "Plasmodium falciparum in Northwest Ethiopia.Early case detection and prompt treatment are important malaria control and elimination strategies. However, the emergence and rapid spread of drug-resistant strains present a major challenge. This study reports the first therapeutic efficacy profile of pyronaridine-artesunate against uncomplicated This single-arm prospective study with 42-day follow-up period was conducted from March to May 2021 at Hamusit Health Centre using the World Health Organization (WHO) therapeutic efficacy study protocol. A total of 90 adults ages 18 and older with uncomplicated falciparum malaria consented and were enrolled in the study. A standard single-dose regimen of pyronaridine-artesunate was administered daily for 3 days, and clinical and parasitological outcomes were assessed over 42 days of follow-up. Thick and thin blood films were prepared from capillary blood and examined using light microscopy. Haemoglobin was measured and dried blood spots were collected on day 0 and on the day of failure.Out of 90 patients, 86/90 (95.6%) completed the 42-day follow-up study period. The overall PCR-corrected cure rate  was very high at 86/87 (98.9%) (95% CI: 92.2\u201399.8%) with no serious adverse events. The parasite clearance rate was high with fast resolution of clinical symptoms; 86/90 (95.6%) and 100% of the study participants cleared parasitaemia and fever on day 3, respectively.P. falciparum in this study population.Pyronaridine-artesunate was highly efficacious and safe against uncomplicated The online version contains supplementary material available at 10.1186/s12936-023-04618-y. Plasmodium falciparum and Plasmodium vivax malaria parasites co-exist. Malaria affects more than 60% of the population and is endemic across 75% of the country\u2019s land mass. Malaria in Ethiopia is seasonal, unstable, and cases generally occur in areas with altitudes below 2000\u00a0m above sea level , which is consistent with a study done in Myanmar . Other pThe median haemoglobin level at baseline (14.2\u2009\u00b1\u20092.03) was significantly higher than on Day 14 (12.7\u2009\u00b1\u20091.75), Day 28 (13.4\u2009\u00b1\u20091.52) and Day 42 (13.5\u2009\u00b1\u20091.79). This is not in line with previous studies on ACT where, following parasite clearance, haemoglobin levels had recovered. Several studies on other artemisinin-based combinations, such as artemisinin\u2013lumefantrine, showed variable but consistent increase in haemoglobin level after treatment \u201329. HoweThe slight increase in median haemoglobin from day 14 to day 42 in this study is also consistent with another study . It suggRehman et al.  reportedSeveral adverse events were reported in this study. These events were similar to the symptoms of malaria, and there were no major adverse events observed. The most common adverse events observed were consistent with previous study. These studies have also reported that the adverse event profile for pyronaridine artesunate was similar to that of artemether\u2013lumefantrine and mefloquine\u2013artesunate in falciparum malaria \u201340. AlthThe study has several limitations: blood drug level was not measured, advanced methods such as the molecular detection of standard resistance markers are yet to be done. The potential reduction of haemoglobin level after pyronaridine artesunate treatment and transaminase level were not investigated. The study enrolled only on adults (excluding vulnerable children), and careful consideration may be required in generalizing the study outcomes.P. falciparum malaria in adults, with rapid parasite clearance and fever resolution. Serious drug adverse events were not observed, implying that the drug is safe for the study group. (Additional file In the first therapeutic efficacy study in Ethiopia, pyronaridine\u2013artesunate was highly efficacious for the treatment of uncomplicated Additional file 1: Annex 1.\u00a0Kaplan-Meier Analysis without PCR correction."}
+{"text": "In Nigeria, declining responsiveness to artemether\u2013lumefantrine (AL), the artemisinin-based combination therapy (ACT) of choice since 2005, has been reported. Pyronaridine\u2013artesunate (PA) is a newer fixed-dose ACT recently prequalified by the WHO for the treatment of uncomplicated falciparum malaria. However, PA data from the Nigerian pediatric population is scarce. Therefore, the efficacy and safety of PA and AL using the WHO 28-day anti-malarial therapeutic efficacy study protocol in Ibadan, southwest Nigeria, were compared.Plasmodium falciparum malaria were enrolled in southwest Nigeria. Enrollees were randomly assigned to receive PA or AL at standard dosages according to body weight for 3\u00a0days. Venous blood was obtained for hematology, blood chemistry, and liver function tests on days 0, 3, 7, and 28 as part of the safety evaluation.In an open-labelled, randomized, controlled clinical trial, 172 children aged 3\u2013144\u00a0months with a history of fever and microscopically confirmed uncomplicated 165 (95.9%) of the enrolled individuals completed the study. About half  of enrollees were male. Eighty-seven (50.6%) received AL, while 85 (49.4%) received PA. Day 28, adequate clinical and parasitological response for PA was 92.7%  and 71.1%  for AL (0.001). Fever and parasite clearance were similar in both groups. Two of six and eight of 24 parasite recurrences were observed among PA- and AL-treated children, respectively. PCR-corrected Day-28 cure rates for PA were 97.4% (76/78) and 88.1% (59/67) for AL (=\u20090.04) in the per-protocol population after new infections were censored. Hematological recovery at day 28 was significantly better among PA-treated patients (34.9% 2.8) compared to those treated with AL (33.1% 3.0) (0.002). Adverse events in both treatment arms were mild and similar to the symptoms of malaria infection. Blood chemistry and liver function tests were mostly within normal limits, with an occasional marginal rise.PA and AL were well-tolerated. PA was significantly more efficacious than AL in both the PCR-uncorrected and PCR-corrected per-protocol populations during this study. The results of this study support the inclusion of PA in the anti-malarial treatment guidelines in Nigeria.Clinicaltrials.gov: NCT05192265.The online version contains supplementary material available at 10.1186/s12936-023-04574-7. Plasmodium falciparum strains with reduced in vivo susceptibility to artemisinin, which was first reported in Western Cambodia  first received a positive scientific opinion from the European Medicines Agency under Article 58 for the tablet formulation in 2012. Pyramax\u2122 granules  in Nigeria. This underscores the importance of evaluating another effective and user-friendly anti-malarial with good efficacy and tolerability in the country. This clinical trial resulted from efforts to improve efficacy, tolerability, cost, and treatment regimens. As the country is endemic for malaria, it is also critical to continue assessing the efficacy of AL, the current first-line ACT of choice in Nigeria, in order to detect early loss of efficacy. There is currently no data on the safety and efficacy of PA in the Nigerian population. The goal of this study is to compare the safety and efficacy of PA and AL in children aged 3\u00a0months to 12\u00a0years (144\u00a0months) with symptomatic acute uncomplicated malaria in south-west Nigeria.Furthermore, Sowunmi et al.  recentlyThis comparative, randomized, open-label, parallel-group clinical trial followed WHO guidelines . Study dP. falciparum malaria were enrolled if they had a minimum asexual parasite density of 1000/\u00b5L, an axillary temperature of\u2009\u2265\u200937.5\u00a0\u00b0C or a history of fever within 24\u00a0h of presentation as reported by a parent or caregiver, lived within fifteen kilometers of the study center, and could take drugs orally. Additional inclusion criteria included the absence of ACT intake in the 2\u00a0weeks preceding enrollment and signed informed consent from prospective enrollees\u2019 parents or guardians to participate in the study. Children with a history of allergy to any of the study drugs, including artemisinin, and its derivatives, lumefantrine, or pyronaridine, were excluded from the study. Children with concurrent illnesses that could impair response evaluation, such as bacterial or viral infections, were also excluded. Additionally, children with clinical evidence of severe malaria were excluded from the study  were treated with artesunate-amodiaquine (ASAQ). Response to ASAQ was prompt, leading to rapid parasite clearance and resolution of symptoms.All the 30 children with parasite recurrence among per-protocol (PP) population\u2014 of AL-treated enrollees had parasite recurrence by day 28, irrespective of whether it was a reinfection or recrudescence. This is far from ideal for the children who go through another episode of malaria infection at such short intervals, with all the health and socio-economic consequences for them and their families. The recorded high efficacy of PA is in keeping with reports of PA efficacy in the treatment of acute uncomplicated malaria among Gabonese children . Details are shown in Table It is particularly noteworthy that PA demonstrated a statistically significant higher PCR uncorrected efficacy among under 5-year-olds during this study [96.0% vs. 63.6% for PA and AL, respectively, as shown in Table P. falciparum more rapidly than AL [. [There was no difference in parasite clearance time between PA and AL treated groups, and the findings during this study are consistent with Roth and colleagues' findings among Kenyan children . In cont than AL , 20. On  than AL . The occan AL [.  previousan AL [. . This isIn this study, both AL and PA had good safety profiles. The commonly observed adverse events were fever, chills, rigors, anorexia, and headache. These symptoms are like those seen in malaria patients and resolve as the infection is cleared. These findings are consistent with those reported by earlier researchers , 38, 40.Another notable finding from the present study is that from days 0 to 21, gametocyte carriage was consistently higher among PA-treated children compared to AL-treated children, but the observed differences were not statistically significant (Additional file This study's findings should be viewed with a few caveats in mind. The current study included only eight children under the age of 12\u00a0months, of which only five were randomly assigned to the pyronaridine\u2013artesunate group. Even though the ACPR for the PA group was 100 percent on day 28, the number of enrollees is insufficient to draw any conclusions about efficacy and safety in that age group. As a result, more information on this patient population is needed for pyronaridine\u2013artesunate.The efficacy and safety of PA among Nigerian children is both impressive and important as Nigeria is one of the highest burden countries for malaria in SSA especially as this is being recorded at a time that AL has started to show declining efficacy. This study provides empirical evidence for the inclusion of PA in the National Treatment Guidelines, and it is hoped that findings from routine TES will provide relevant updates on the efficacy of ACT in the country for their sustained recommendation.Additional file 1. Table S1: Paired sample genotyping data. Tables S1a\u2013S1h and Tables S2a\u2013S2h and Figs. S1\u2013S8 of results showing liver enzymes, serum bilirubin, urea, creatinine, glucose and glucose of children enrolled in the study.Additional file 2.Table S3:Tables S3a and S3b showing gametocyte carriage among children treated with artemether\u2013lumefantrine and pyronaridine\u2013artesunate during the study."}
+{"text": "Overall, the compliance level of health workers (HWs) with MFT strategy was 72.7% (95% CI: 69.7\u201375.5). The odds of using PHF as the first source of care increased after the intervention , and the reported adherence to the 3-day treatment regimen was 82.1%; (95% CI: 79.6\u201384.3). Qualitative results showed a high acceptance of the MFT strategy with positive opinions from all stakeholders. (4) Conclusions: Implementing an MFT strategy is operationally feasible and acceptable by stakeholders in the health systems in Burkina Faso. This study provides evidence to support the simultaneous use of multiple first-line artemisinin combination therapies in malaria-endemic countries such as Burkina Faso.(1) Background: Effective malaria case management relies on World Health Organization (WHO) recommended artemisinin-based combination therapies (ACTs), but partial resistance to artemisinin has emerged and is spreading, threatening malaria control and elimination efforts. The strategy of deploying multiple first-line therapies (MFT) may help mitigate this threat and extend the therapeutic life of current ACTs. (2) Methods: A district-wide pilot quasi-experimental study was conducted, deploying three different ACTs at the public health facility (PHF) level for uncomplicated malaria treatment from December 2019 to December 2020 in the health district (HD) of Kaya, Burkina Faso. Mixed methods, including household and health facility-based quantitative and qualitative surveys, were used to evaluate the pilot programme. (3) Results: A total of 2008 suspected malaria patients were surveyed at PHFs, of which 79.1% were tested by rapid diagnostic test (RDT) with 65.5% positivity rate. In total, 86.1% of the confirmed cases received the appropriate ACT according to the MFT strategy. The adherence level did not differ by study segment ( Despite recent progress, sub-Saharan Africa still bears the major burden of malaria . In 2020Historically, the emergence and global spread of chloroquine resistance has had a dramatic public health impact . ACTs weGlobally, the deployment of ACTs contributed substantially to reducing malaria burden . HoweverShould ACTs lose efficacy, this would result in a dramatic rebound in malaria morbidity and mortality as no replacement drug is available. To prevent this; optimized use of the current ACTs may provide a higher long-term barrier to the emergence and/or the spread of resistance, while new drug classes are being developed. Modelling studies have concluded that deploying multiple first-line therapies (MFT) for uncomplicated malaria is a promising strategy to extend the useful therapeutic life of the current ACTs by reducing drug pressure and slowing down the spread of resistance without putting lives at risk ,24,25,26To assess the feasibility and the acceptability of the MFT strategy for uncomplicated malaria case management in a high burden malaria endemic country, we conducted a pilot study in the HD of Kaya in Burkina Faso. The pilot MFT strategy consisted of deploying three different ACTs targeting three different segments of the population. Our hypothesis was that the MFT strategy could be managed by the HWs and accepted by stakeholders in the health system in Burkina Faso. This study reports on the feasibility and acceptability of this MFT pilot implementation.2 with an estimated population of about 432,600 inhabitants in 2020.This was a district-wide pilot implementation study conducted in the health district (HD) of Kaya, in the north-central region of Burkina Faso, about 100 km from the capital city Ouagadougou. The map of the HD is presented in This was a quasi-experimental study design with pre- and post-intervention assessments conducted before the deployment of MFT and one year after the intervention, respectively. A mixed method, including desk reviews, quantitative and qualitative surveys were conducted to assess the study outcomes.The intervention took place from December 2019 to November 2020. It consisted of the deployment of three ACTs targeting three different segments of populations at the PHF level for the management of uncomplicated malaria cases. PYR\u2013AS (Pyramax) granule formulation (20/60 mg) procured from Shin Poong  was allocated to the Segment I (children under 5 years of age); DHA\u2013PQP (D-Artepp) tablets (40/340 mg) obtained from Fosun Pharma was allocated to Segment II (patients aged 5 years and above); and AL tablets (10/120 mg) procured from the Ministry of Health in Burkina Faso was allocated to the Segment III (pregnant women after the first trimester). All ACTs (except AL) were procured by Medicines for Malaria Venture (MMV) and prepacked in weight- and age-specific doses . They weThe procurement process was aligned with the existing national system. ACTs were purchased through the central essential and generic drugs store and made available at the district drug stores. HF drug store managers supplied the required ACTs from the district store. The HWs were not paid for the programme implementation. The dispensation of ACTs to patients in the study area followed the routine practices. Thus, PYR-AS and AL were given free of charge to malaria patients in segments I and III, respectively. For Segment II patients, DHA-PQP were sold at a subsidised price (100 XOF (0.169 USD for six tablets)) to match the cost of generic standard treatment. Before the intervention, HD and regional management teams were trained for 3 days to become trainers and supervisors. They organized thereafter 10 training sessions over 10 days for 20\u201330 HWs and drug store managers each. Overall, 271 HWs and all generic drug store managers (GDSM) (50) were trained on the MFT strategy, and all study-related procedures. HWs joining the HF after the initial training sessions were peer-trained onsite and during the supervision visits. Compliance to the strategy was optimized through regular supervisions based on established procedures. Identified issues were discussed on the spot and appropriate corrective and preventative actions taken. Study participants were selected through multi-stage sampling approach. First, 10 out of 33 accessible HFs were randomly selected; then, in each HF, 16 weeks of consultations out of 52 weeks were randomly selected; and lastly, all suspected malaria cases in the selected weeks were numbered and sampled using a systematic sampling method. The number of patients selected per segment and per facility was proportional to the facility\u2019s outpatient attendance in the district. Thus, 47%, 49% and 4% of suspect malaria cases were needed for children under 5, patients aged 5 years and above, and pregnant women, respectively . We inclThe sampling strategy for the household survey and qualitative surveys is described elsewhere . BrieflyQualitative survey included in-depth individual interviews (IDI) and focus group discussions (FGDs) conducted with diverse respondents including stakeholders of the heath system and community members.Assuming that the proportion of febrile patients tested and treated according to the MFT strategy was unknown in the framework of this new strategy and considering a design effect of 1.5; a minimum sample size of 578 patients was required to estimate the proportion of confirmed malaria patients appropriately managed with a 95% CI around the estimated proportion of 50%. This sample size was adjusted by the unknown compliance level of HWs with MFT strategy and the RDT testing and positivity rate in the district. Adjusting for 10% , the final sample size was 2001 suspected malaria patients to be sampled from the outpatient consultation registers. The sample size for the household survey and qualitative survey is described elsewhere . BrieflyThe qualitative survey sampling was exhaustive for all stakeholders in the health system , health system managers, healthcare providers as well as community members . Quantitative and qualitative data were collected using data extraction forms, household survey questionnaire and FGD, and IDI guides.During the HF-based survey, data were extracted from the outpatient consultation registers, which captured patient characteristics, symptoms, malaria diagnosis and treatment given by trained nurses using KoboCollect on electronic tablets in February 2021. Data on study drugs and RDT supplies availability were collected monthly from all covered PHFs including information related to their availability at the start of the month, quantity received during the month, quantity dispensed to patients and expired, stock remaining at the end of the month and number of days of stock out during the month. Pre- and post-intervention household surveys were conducted in November 2018 and November 2020, respectively, and focused on the health-seeking behaviour of patients with fever in the preceding 4 weeks. Variables of interest included timeliness of treatment, providers of care, compliance with treatment course, and perceptions of drug effects including adverse events. The FGD and IDI structured interview explored the stakeholders\u2019 perceptions and opinions about the intervention, implementation challenges, health-seeking behaviours for fever, and treatment practices. The interviews were conducted in the local language or in the language best spoken by the participant and by trained social scientists.The primary feasibility outcomes included adherence of HWs to the MFT guidelines , and stuThe communities\u2019 acceptability outcomes were measured during the household surveys through their adherence to the 3-day treatment regimen, the changes before and after the intervention in (i) prompt care-seeking behaviour and (ii) care-seeking at PHFs. The adherence to the treatment regimen was defined as correctly adhering if the prescribed ACT was taken for 3 days as per guidelines. Prompt care-seek behaviour was defined as seeking care within 24 h of fever onset while care-seeking at PHFs consisted of the use of PHF as the first source of providers for fever within 4 weeks before the survey. Exploratory variables included time before and after intervention variable, socio-demographic factors , and advice given to the patient regarding the dosing regimen. p\u2009 < \u20090.05. Logistic regression with odds ratio and 95% confidence interval were used to measure the effect of the intervention on the promptness of care-seeking, the use of PHFs as first source of care and the community adherence to the ACT dosing regimen. Exploratory variables were selected based on rational grounds prior to analysis and included in the final model irrespective of their level of significance. Observations with missing values were excluded from the regression analysis. Quantitative data were analysed using Stata SE V.16.1. Proportions were used for most analyses, and groups were compared using Pearson\u2019s chi square test at significance For qualitative data analysis, all interviews were transcribed in full and transcripts were checked, coded, and processed using QDA Miner Lite V2.0.8 software. Data-derived codes developed through deductive and inductive coding and retrieving were used during analysis. Thematic analysis was performed, including the acceptability of the strategy by different stakeholders, and perceptions of the limits or constraints of the strategy. In total, 2008 suspected malaria patients were surveyed at the HF level, of which 45.1%, 50.3%, and 4.6% were in segments I, II and III, respectively. Most patients (54%) were female and lived in rural areas (59.8%) summarizes in p = 0.19). Overall, 72.7% (95% CI: 69.7\u201375.5) of cases received the correct ACT at the correct dose. From the household survey, overall adherence to 3-day regimen was 82.1%. Patients treated with PYR-AS  or DHA-PQP  were more likely to adhere to the 3-day treatment regimen compared to AL. RDT stock outs were reported in 2.5% (1/40) of HFs in January 2020 for 3 days and seven HFs (7/40) in October and November 2020. A total of 1394 and 1448 febrile patients/caregivers were interviewed at pre- and post-intervention surveys, respectively. p = 0.58) conversely to the promptness which significantly decreased from 66.5% at baseline to 54.3% (p < 0.001). Throughout the intervention no serious adverse events were reported. All adverse events (AE) reported during the household survey were mild or moderate. Overall, 10.4% (99/949) patients interviewed reported having had an AE following the drug intake. The three most frequent AEs were vomiting, pruritus, and anorexia .In total, 28 IDIs and 33 FGDs were conducted post intervention. HWs interviews showed that the intervention was well received. They had some good knowledge of the MFT strategy including drugs assignment to segments, dosing instructions; and found the approach straightforward to apply after appropriate training.\u201cThe MFT approach, I think it is good because we have three choices and what is interesting for children under 5, Pyramax is in a sachet [powder], it is in a single dose per day, and easily children accept.\u201d\u201cFor us [HWs], it does not tire us. Because once we have specified, we know that this drug is for this age group and such a treatment for such an age group. Now, you give it according to weight, quantity by weight. And since we were given the posters and put them up in the consultation rooms, I do not think there is a problem for me anyway.\u201d\u201cEveryone actually adheres to this new malaria treatment protocol. If you have noticed even since this morning, it is Pyramax, D-artepp except in the case of pregnant women for whom AL is given.\u201d Some HWs reported that this strategy has contributed to the improvement in the care to malaria patients.\u201cThis strategy has contributed to better management of uncomplicated malaria. We cannot really say the opposite.\u201dThe policymakers also positively appreciated this new strategy. \u201cThis is a good strategy. I think it is new in our country. It is an experience. Thank God! Burkina Faso has given itself the power to absorb this new strategy. I followed the topic from the beginning to the end, from the training until we also report it. I think it is a good strategy\u201d\u201cIt is a good approach; people have bought in.\u201dCommunity members were asked to give their perceptions of the MFT intervention. The results showed a good acceptance by the population in the study area. Most of the participants reported that they were happy with the MFT intervention. Their perceptions were based on experiences with the study drugs, including the perceived efficacy of the two new ACTs (PYR-AS and DHA-PQP). \u201cThese are good medicines [study drugs]; we appreciate the medicines we are given. They are effective.\u201d \u201cIn any case, the product [new product] treats well. In the meantime, my child had malaria, and I went to the hospital, they gave me the prescription, and I took the products, and when I gave it to the child, he quickly recovered his health.\u201dDespite stakeholders\u2019 high acceptance of the intervention, some challenges and difficulties were identified, including supplies management (drug stock-outs or expiry), health staff availability, lack of refresher training for HWs, non-adherence to study guidelines. \u201cIn the meantime, there was stock out for D-artepp, so we had to switch to AL.\u201d\u201cUnfortunately, several shortcomings were noted because some [local drug store managers] did not know what target to give. They did not know what was the target that was concerned in this study, what are the different prescription modalities.\u201d\u201cI went to the hospital last time and the drug was stocked out. They gave me a prescription and told me to go to town to buy it. I said I could not because my husband is not here. It was the doctor himself who took my prescription to go to the city to get the medicine and come back.\u201dTo the best of our knowledge, this is the first study to provide evidence of the feasibility of field deployment of an MFT strategy at PHFs level. The study showed that the deployment was well received by both population and HWs, and resulted in a high compliance of HWs with the guidelines. The adherence of caregivers or patients to the dosing regimens was very high, as the drugs were perceived as highly efficacious.Eighty-six percent of malaria patients received the appropriate ACT as per MFT guidelines, and 72.7% received the correct ACT at the correct dose; showing the HWs\u2019 ability to comply with the MFT strategy. The good adherence of HWs to the MFT strategy guidelines may be explained by the simplicity and good documentation of the intervention, and the active involvement of key players in the health system  at all steps of the deployment. Indeed, the project was carefully embedded into the country health system using the same supply and distribution channel of ACTs. Furthermore, the subsidizing of new ACTs, their assured availability, considerable efforts for training, sensitization, supervision, and monitoring before and during the intervention, have all certainly enhanced the adherence of HWs and communities to the new strategy. We found that about 27% of confirmed cases were not adequately managed. This high proportion of non-adherence to the guidelines underlines the need for sustained monitoring of HWs practices, strengthening training and sensitization through periodic refresher training sessions, and improving the key actors\u2019 commitment during the large-scale implementation of MFT.Several MFT strategy options exist ,25,27, aOur findings revealed that a large fraction of febrile patients was tested for malaria (79.1%) and nearly all (98.4%) of those tested positive received WHO-recommended treatment. Still, this testing rate is below the NMCP 100% target goal . All PHFCommunities appreciated the strategy and adhered to it. The perceived efficacy of the two new ACTs (PYR-AS and DHA-PQP) associated with low frequencies of side effects, the availability of drugs, and the fact that the intervention was performed by the HWs whom they knew and trusted were identified as factors associated with this adherence. The acceptability of the MFT strategy was reflected by an increase in PHF use as the communities\u2019 first source of care for febrile illness. In contrast, the proportion of patients seeking care within 24 h of fever onset decreased after the intervention, reflecting the need to support the program with sensitization and social behaviour change activities. Community members explained that they quickly recovered after taking ACTs prescribed by HWs. The perceived efficacy is a strong predictor of communities\u2019 treatment adherence . AlthougThe overall reported adherence to the treatment regimen was 82.1%. Similar adherence levels were previously reported in patients treated with ACTs . The achThe drug stock management was satisfactory during the intervention. No drug stock- out was reported in the district store during the study period conversely to the HFs that experienced study drug stock-outs more frequently during the malaria peak period (September to November). The data suggest that the workload of GDSMs increases at malaria peak period, resulting in inadequacy of drug use and inventory. The stock-outs were confirmed by the communities expressing their experiences when attending PHFs. The low stock-out rates reported in this study refer to the context of Burkina Faso, which had a low stock-out rate before this pilot study implementation . Our finThe study findings should be considered in the context of the following limitations and strengths. The mixed-method design provides some confidence on the robustness of the evidence generated. This could support a policy recommendation of the MFT approach as a cornerstone antimalarial resistance mitigation strategy.The study has several limitations: Firstly, this pilot study was restricted to the PHFs only. Therefore, the findings may not be applicable if private health facilities and the community-based cases management are to be considered. Secondly, recall and social desirability bias may be associated with household surveys with the 4-week recall period and the self-reported data collected at the community level.The MFT strategies have been suggested to mitigate the emerging partial artemisinin resistance in Africa. This was the first large-scale field project on the implementation of an MFT strategy for uncomplicated malaria treatment, and it has shown to be operationally feasible and highly acceptable by stakeholders in the public sector. This study provides evidence to support the simultaneous use of multiple first-line artemisinin combination therapies in malaria-endemic countries such as Burkina Faso. For a successful scale up of this strategy, the involvement of key stakeholders throughout will play a pivotal role. Sustained education and training, continuous availability of drugs are keys for success."}
+{"text": "Plasmodium falciparum infection in the second or third trimester. The primary hypothesis is the pairwise non-inferiority of PA as compared with either AL or DP.Malaria infection during pregnancy increases the risk of low birth weight and infant mortality and should be prevented and treated. Artemisinin-based combination treatments are generally well tolerated, safe and effective; the most used being artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP). Pyronaridine-artesunate (PA) is a new artemisinin-based combination. The main objective of this study is to determine the efficacy and safety of PA versus AL or DP when administered to pregnant women with confirmed A phase 3, non-inferiority, randomised, open-label clinical trial to determine the safety and efficacy of AL, DP and PA in pregnant women with malaria in five sub-Saharan, malaria-endemic countries . A total of 1875 pregnant women will be randomised to one of the treatment arms. Women will be actively monitored until Day 63 post-treatment, at delivery and 4\u20136 weeks after delivery, and infants\u2019 health will be checked on their first birthday. The primary endpoint is the PCR-adjusted rate of adequate clinical and parasitological response at Day 42 in the per-protocol population.This protocol has been approved by the Ethics Committee for Health Research in Burkina Faso, the National Health Ethics Committee in the Democratic Republic of Congo, the Ethics Committee of the Faculty of Medicine and Odontostomatology/Faculty of Pharmacy in Mali, the Gambia Government/MRCG Joint Ethics Committee and the National Bioethics Committee for Health in Mozambique. Written informed consent will be obtained from each individual prior to her participation in the study. The results will be published in peer-reviewed open access journals and presented at (inter)national conferences and meetings.PACTR202011812241529. This is a randomised clinical trial that will provide evidence on the safety and efficacy of a newly registered artemisinin-based combination, pyronaridine-artesunate (PA), for the treatment of uncomplicated malaria in African pregnant women during the second and third trimesters of pregnancy.The trial will generate additional data on the safety and efficacy of dihydroartemisinin-piperaquine and artemether-lumefantrine and will increase substantially the treatment options for malaria in pregnancy.As PA is the last artemisinin-based combination to have been registered for the treatment of malaria, the proposed trial will complement the information already available for malaria treatment in children and non-pregnant adults.Weaknesses include the lack of regular monitoring of children within the first year of birth and the non-inclusion of infected women in the first trimester.Pregnancy is associated with hormonal changes and reduced immunity that make pregnant women more vulnerable to malaria.5 6Plasmodium falciparum malaria during the second and third trimesters of pregnancy, namely artemether-lumefantrine (AL), artesunate-amodiaquine, mefloquine-artesunate and dihydroartemisinin-piperaquine (DP), are generally safe and efficacious.P. falciparum or Plasmodium vivax malaria.Currently available ACTs for the treatment of P. falciparum infection in the second or third trimester.To address this need, we designed a randomised open-label clinical trial to determine the efficacy and safety of PA versus AL or DP, when administered to pregnant women with confirmed The specific trial objectives are:P. falciparum malaria.To compare the efficacy of PA versus AL or DP in terms of (1) treatment success (see definition below) 28, 42 and 63 days after the start of treatment, with and without genotyping; (2) parasite clearance time, including submicroscopic malaria infections; (3) gametocyte carriage and clearance; (4) haematological recovery by Day 7, 14, 28, 42 and 63 after treatment and at delivery; (5) birth weight measured within 72 hours of delivery and (vi) placental To describe the safety profile of PA, AL and DP in terms of (1) tolerability and (2) adverse events (AEs), including serious adverse events and adverse events of special interest, during the 63-day post-treatment follow-up (mother), at delivery (mother and baby), 1\u2009month (mother and baby) and 1\u2009year after the end of pregnancy (baby).To explore the pharmacokinetics of pyronaridine in HIV-infected and non-HIV-infected pregnant women. This will be done on a small number of study subjects  in the two countries with the highest HIV prevalence, namely, the Democratic Republic of the Congo (DRC) and Mozambique.The trial will be implemented in five African countries with multiple sites of recruitment, as indicated in The different sites were chosen to include African pregnant women who are residents of different malaria transmission facies, including the Gambia, which is in the pre-elimination phase.The study (recruitment) started on February 2021 and the planned end date is June 2024.Although patients were not involved in the design of the study, this project was designed to respond to their main concern, which is the reduction of malaria adverse effects in pregnant population. The governments of the five African countries were engaged through their National Malaria Control Programme (NMCPs), whose objective is to provide the evidence base to make an additional ACT available for malaria treatment in pregnancy. In the event of a positive result, it will be important to integrate this new treatment into (inter)national guidelines. This will be obtained by disseminating as rapidly and efficiently as possible the trial\u2019s results to relevant stakeholders, for example, the WHO, NMCPs and the scientific community working on malaria treatment. Through meetings in all the involved countries, permission from the community, religious leaders and women representatives has been sought before the trial starts. Therefore, similar channels will be used to share the study results. Participation in our trial can be burdensome as participants may endure psychological, cost and physical impacts. However, the project provided special attention such as closer supervision and treatment of any illness to enrolled patients. Also, transport costs have been provided to participants living far from the recruitment centre. Nonetheless, it has been suggested to all participants to withdraw their consent at any time if they feel burdened by the study procedures.P. falciparum malaria in women in the second and third trimesters of pregnancy. The non-inferiority design was suggested based on the hypothesis that PA, which is a newly registered ACT, is not worse than the other ACTs in terms of safety (for AL) and efficacy (for DP). The rationale for two control groups is that AL is currently the most used treatment for malaria in pregnancy, while DP seems to be the ACT with the best tolerability and efficacy profile.This is an open-label, multicentre, randomised, non-inferiority clinical trial comparing PA with AL and DP for the treatment of The primary hypothesis is the pairwise non-inferiority of PA as compared with either AL or DP, defined as a difference in the Day 42 PCR-adjusted adequate clinical and parasitological response (ACPR) of <5% (non-inferiority margin).To be included, patients should satisfy all the inclusion criteria, while none of the exclusion criteria should be present.Gestation \u226516 weeks and <37 weeks as assessed by ultrasound when possible. If not, the height of the uterus or delay of menstruation will be used.P. falciparum monoinfection (by microscopy) of any density, regardless of symptoms and HIV status.Haemoglobin \u22657\u2009g/L.Age \u226515 years.Residence within the health facility catchment area.Willingness to adhere to study requirements and to deliver at the health facility.Ability to provide written informed consent; if the woman is a minor of age or not emancipated, the consent must be given by a parent or legal guardian according to national law .For the PK study, HIV-infected women should be on first-line antiretroviral treatment for at least 6 months.Known allergy to the study drugs.History of known pregnancy complications or poor obstetric history, such as repeated stillbirths or eclampsia.History or presence of major illnesses likely to influence pregnancy outcomes.Any significant illness at the time of screening requiring hospitalisation, including:Severe malaria.Any sign or symptom suggesting hepatic lesions  or severe liver disease classified as B or C by the Child\u2013Pugh score.Known history or evidence of clinically significant cardiovascular disorders or family history of long QT syndrome.Intent to move out of the study catchment area before delivery or delivery out of the catchment area.Prior enrolment in the study.Clear evidence of recent (1\u2009week) treatment with antimicrobials with antimalarial activity . For HIV-infected pregnant women to be included in the PK substudy, cotrimoxazole use is not an exclusion criterion.Twin/multiple pregnancy.Known history of G6PD deficiency or sickle cell disease.Women recruited in the trial will be randomly allocated to one of the three treatment arms of the study according to a randomisation list generated using R software prior to any study activity.The block randomisation technique will be used to achieve balance in the allocation of participants to treatment arms. A varying size of the blocks will be set up to reduce selection bias by using random block sizes and keeping the investigator blind to the size of each block, particularly in this open randomised control arm study. The randomisation will be done per study site within each country and according to the specific sample size allocated to each site.The data management team in each country, in coordination with the study statistician and the study data management centre, will print the randomisation code containing the study arm and put it into a sealed envelope numbered sequentially and containing the treatment arm to which the patient should be allocated.Women in the second or third trimester of pregnancy will be screened for malaria with a rapid diagnostic test based on the detection of histidine-rich protein 2 and parasite lactate; positive results will be confirmed by microscopy. Although malaria slides will be read by two certified microscopists, there is a small chance that mixed infections might be overlooked, and this can be considered a potential limitation of this study.P. falciparum infection will be asked to provide written informed consent covering all trial procedures and be assigned a screening number. All women meeting the entry criteria will be given a randomisation number. Data from screened and randomised women will be kept in a logbook.Women with a confirmed PA is a film-coated tablet containing 180 mg of pyronaridine tetraphosphate and 60 mg of artesunate. The tablets should be taken orally once daily for 3\u2009days and according to body weight as follows: 24 to <45\u2009kg (two tablets), 45 to <65\u2009kg (three tablets) and \u226565\u2009kg (four tablets). This dosage regimen provides a daily dose of 7.2\u201313.8\u2009mg/kg pyronaridine and 2.4\u20134.6\u2009mg/kg artesunate. PA can be administered at any time, regardless of food consumption.DP is a white film-coated tablet composed of 40 mg of dihydroartemisinin and 320 mg of piperaquine. DP tablets should be taken orally once daily for 3\u2009days as follows: 24 to <36\u2009kg (two tablets), 36 to <75\u2009kg (three tablets) and \u226575\u2009kg (four tablets).9 10AL tablets contain 80 mg of artemether and 480 mg of lumefantrine. This is a fixed-dose combination of artemether (a semi-synthetic artemisinin derivative) and lumefantrine .First dose, at the time of initial diagnosis (Day 0): one tablet.Second dose, at 8 hours after the first dose: one tablet.Third dose, in the morning of Day 1: one tablet.Fourth dose, in the evening of Day 1: one tablet.Fifth dose, in the morning of Day 2: one tablet.Sixth dose, in the evening of Day 2: one tablet.Investigator\u2019s brochures and all documentation on drug quality will be provided by the manufacturers. The drugs will be shipped to the different sites with temperature monitors that record the temperature continuously. Once at the sites, the drugs will be stored and used according to the manufacturers\u2019 instructions. Independent study monitors (CliniPharm) will ensure compliance with good clinical practice, including good Investigational Product management practice.Pregnant women (second or third trimester) will be recruited for the study and allocated to one of the three treatment groups. The choice of two control arms (AL and DP) is justified by AL being the most commonly used treatment for malaria in African pregnant women, while DP has several advantages compared with AL, namely higher efficacy and longer post-treatment prophylaxis.The primary endpoint is treatment efficacy, determined as the proportion of women with PCR-adjusted ACPR at Day 42, that is, all women not having met the criteria of treatment failure . Recurre10.1136/bmjopen-2022-065295.supp1Supplementary dataSafety is the main secondary endpoint and includes adverse events detected during active follow-up (63 days post-treatment), including significant changes in relevant laboratory values, those detected at delivery, at 4\u20136 weeks and 1\u2009year after birth.Other secondary endpoints include:PCR-adjusted ACPR at Days 28 and 63.PCR unadjusted ACPR on Days 28, 42 and 63.Parasite and fever elimination time.Gametocyte carriage and clearance.Hematologic recovery, that is, haemoglobin changes between Day 0 and Days 7, 14, 28, 42, 63 and at delivery.Placenta malaria .Average birth weight and prevalence of LBW (<2500\u2009g).0-\u221e. The main pharmacokinetic parameters that will be evaluated are absorption rate (ka), drug clearance (CL/F) and volume of distribution (Vd/F).We will explore (1) the drug exposure and key pharmacokinetic parameters of pyronaridine in HIV-uninfected pregnant women and (2) the drug exposure and key pharmacokinetic parameters of HIV-infected pregnant women on antiretroviral therapy. Drug exposure is defined as the area under the whole blood concentration versus the time curve from zero to infinity, AUC The sample size was estimated assuming an efficacy for PA (PCR adjusted at Day 42) of at least 95% and a non-inferiority margin of 5%; non-inferiority will be tested using raw pooling of country data using Wilson\u2019s interval of proportion difference. The lower limit of the Wilson score interval of 97.5% of the AL (or DP) proportion of ACPR \u2013 the PA proportion of ACPR \u2013 must be greater than \u22125% to claim non-inferiority. In addition, an adjusted non-inferiority analysis will allow for the country effect.Participants\u2019 assignment to the study arm is under the responsibility of study-qualified physicians. All physicians participating in the PYRAPREG study have been trained in good clinical practices and all study requirements.Pregnant women fulfilling the inclusion/exclusion criteria will be recruited during antenatal clinics over a period of about 30 months. Scheduled visits will be on Days 3, 7 and then every week until Day 63 post-treatment. A window period is allowed if study subjects are unable to attend on the scheduled date,that is, \u00b11\u2009day for Days 7 and 14; \u00b12 for Days 21 and 28; and\u00b13 from Day 35 to Day 63. Women will be encouraged to attend the antenatal clinic between scheduled visits if they are sick.Pregnant women recruited during the third trimester (before 37 weeks) may deliver before the end of the 63-day active follow-up. In this case, the assessment at delivery will be done as planned but the active follow-up will continue after delivery until Day 63. A blood sample to measure liver function tests (LFT) and bilirubin will be taken within 48 hours of delivery from babies whose mothers delivered within 2 weeks of their inclusion in the trial.The outcome of pregnancy, birth weight and maternal haemoglobin will be collected as soon as possible after delivery. A placenta biopsy will be collected for later histopathological analysis. The newborn will be examined for congenital abnormalities. Both the mother and the newborn will be reassessed for any AEs between 4 and 6 weeks and then after 1\u2009year (only the baby). Patients will be assessed as summarised in the study visit schedule .At each visit, both scheduled and unscheduled, the medical history since the last visit (including any treatment taken) and current signs and symptoms (if any) will be collected. A blood sample for a thick smear will be collected and the body temperature will be checked. Dried blood spots for later genotyping to distinguish between recrudescence and new infection will be collected at Day 0, before treatment, and at every study visit. Information on any AE will also be collected. Haematology (full blood count) will be performed at Days 0, 7, 14, 28, 42 and 63; biochemistry , alanine transaminase (ALT), alkaline phosphatase (ALP) and creatinine) at Days 0, 1 and 7. In the event of increased LFTs, >3\u00d7\u2009upper limit normal (ULN), the result will be verified by taking an additional sample to be analysed (within 24 hours). Subsequent blood samples for LFTs will be taken at 48-hour intervals until the results return to <2\u00d7 ULN. ECG will be performed on Day 0, before drug intake, and on Day 2, after drug intake. If abnormal on Day 2, the ECG will be repeated on Day 7 and every week until returns to normal. At the end of the active follow-up, on Day 63, field assistants will visit the study subjects monthly to maintain contact, but without collecting any data or biological samples.Babies born from women who deliver within or 2 weeks after the active follow-up will have a blood sample taken to measure LFT and bilirubin within 48 hours of delivery.The outcome of pregnancy, birth weight and maternal haemoglobin will be collected as soon as possible after delivery. A placenta biopsy will be collected and put immediately in a 10% buffered formalin container stored at 4\u00b0C at the study site for later histopathological analysis. The newborn will be examined for congenital abnormalities. Both the mother and the newborn will be reassessed twice after delivery for any AE: between 4 and 6 weeks and then after 1\u2009year.msp1), msp2 and glutamate-rich protein (glurp) genes, single-copy genes of the P. falciparum genome. PCR amplification of DNA from a single parasite clone will yield a unique amplification product. For all three genes, each PCR amplification product of a different size is considered from a different P. falciparum clone and reflects a different genotype. For samples collected from the same patient on Day 0 and on the day of recurrent parasitaemia (after Day 3), the length polymorphism of msp1, msp2 and glurp will be determined, that is, the number of bands in each PCR reaction and their respective sizes. The results will be interpreted as follows:Genotyping of recurrent infections will be performed by characterising the merozoites surface protein 1  is found in the sample collected on Day 0 and on the day of the recurrent parasitaemia.New infection: for at least one marker, the length polymorphism is different between the sample collected on Day 0 and on the day of recurrent parasitaemia.Indeterminate: samples that did not give a result due to an inability to amplify DNA on Day 0 and/or on the day of recurrent parasitaemia.The placental biopsy samples will be processed and embedded in paraffin wax using standard techniques. Paraffin sections 4 \u00b5m thick will be stained with H&E. Placental biopsies after reading will be classified according to the following definitionsHaematology (including haemoglobin) and biochemistry  will be performed during active follow-up; haematology will be performed prior to the first dose of treatment, on Day 0, and then on Days 7, 14, 28, 42 and 63. An additional test will be performed at any unscheduled visit. In addition, only haemoglobin will be measured at delivery. Biochemistry will also be done on Day 0, prior to treatment, then on Day 1 and Day 7. If LFTs increase more than three times the ULN, the result will be verified by taking another sample for analysis as soon as possible . Subsequent blood samples for LFTs will be taken at 48-hour intervals until results return to <2\u00d7\u2009ULN.The baseline characteristics will be described by treatment group and site.The primary analysis will be the assessment of the non-inferiority of the PA compared with the DP and AL for the PCR-adjusted ACPR at Day 42. It will use the combined data from the five countries, with adjustment for any centre effect, using an additive model for response rates . This will allow the evaluation of two pairwise treatment comparisons, that is, PA versus AL and PA versus DP.P. falciparum infection prior to treatment.For the efficacy analysis, both a modified-intention to treat (m-ITT) approach and a per-protocol (PP) approach will be adopted, with PP analysis being the main approach, as recommended for equivalence studies. The m-ITT population will include all participants who have received any amount of the study drug and have confirmed The PP population will consist of all participants meeting the following predefined criteria:Fulfilling the entry criteria specified in the clinical study protocol.Completed treatment, including not having vomited the study drug or, if vomited, receiving a repeat dose that was not vomited.No previous or concomitant medication would interfere with the treatment outcome.The PP population will be identified after locking the database, just before the statistical analysis.For the primary endpoint, that is, treatment efficacy on Day 42, the proportion of participants with PCR-adjusted ACPR will be determined by the treatment arm. Similar procedures will be applied to the m-ITT population.The safety population will include all participants randomised and treated with at least one dose of the three antimalarial treatments. Standard safety report tables summarise and list safety data. All AEs will be coded using the most recent version of the Medical Dictionary for Regulatory Activities (MedDRA) dictionary. Treatment-emergent AEs will be defined as all AEs that started after the first administration of the study drug. These AEs will be summarised by primary system organ class and preferred term, separately for each treatment regimen and overall. Similar summaries will be provided for treatment-emergent AEs considered to be related to study treatments. In addition, treatment-emergent AEs will be summarised by primary system organ class, preferred term, and maximal severity.Vital signs and routine safety laboratory data will be summarised descriptively by treatment regimen and overall, by time point. Absolute values and changes from the baseline will be presented. Safety laboratory data will be classified according to the normal ranges , and summaries of changes from baseline in these categories will be provided by treatment regimen and overall. Furthermore, safety laboratory values will be classified according to Common Terminology Criteria for Adverse Events (CTCAE) and shift tables of the baseline CTCAE category versus post-baseline categories will be presented.0-inf), which will be characterised based on the final individual model parameter estimates and compared with a separate cohort of non-pregnant malaria patients treated with PA. The effect of pregnancy will be evaluated and quantified either as a binary or continuous  covariate on all primary estimated pharmacokinetic parameters . Furthermore, we will explore the effect of HIV background  on the pharmacokinetics of pyronaridine by evaluating HIV status as a covariate. Statistical significance and selection of covariates in nested population pharmacokinetic models will be based on a likelihood ratio test to evaluate the difference in model fit.Pyronaridine exposure in patients\u2019 blood will be assessed in pregnant women, both infected and non-infected by HIV, by determining pyronaridine concentrations in EDTA whole blood samples collected on Day 0 before and 1\u2009hour, 2\u2009hours, 6\u2009hours and 10\u2009hours after the first PA dose, on Day 1 (24 hours after the first PA dose while before the second PA dose), Day 2 (before the last PA dose), and subsequently on Days 7, 14, 21, 28 and 42 after initiation of treatment. A population pharmacokinetic approach using non-linear mixed effects modeling will be employed to analyze the pharmacokinetic data. A compartmental population pharmacokinetic model will be developed, including a stochastic model describing between-subject and residual variability. In short, the main parameter of interest is overall whole blood pyronaridine exposure, defined as the area under the concentration-time curve until infinity  (prestudy visit) for its preparedness to carry out, followed by regular monitoring and closeout visits.The coordination of the whole project is in the hands of the sponsor, who will be the primary contact. He will be assisted by appropriate administrative and financial staff. A Coordinating Committee (CC), including one member of each institution, will be the main decision body of the consortium. For the trial, there will be three entities involved in its implementation and management, namely, the Data Safety and Monitoring Board (DSMB), the Trial Steering Committee (TSC), and the Trial Management Group. Both DSMB and TSC are composed of independent experts to provide the overall supervision of the trial, monitor trial progress, and advise on scientific credibility.This protocol has been approved by the Ethics Committee for Health Research (CERS) in Burkina Faso (Reference: 2020-3-047), the National Health Ethics Committee (CNES) in the Democratic Republic of Congo (Reference: 169/CNES/BN/PMMF/2019), the Ethics Committee of the Faculty of Medicine and Odontostomatology (FMOS)/Faculty of Pharmacy (FPHA) in Mali (Reference: 2020/46/CE/FMOS/FAPH), the Gambia Government/MRCG Joint Ethics Committee (Reference: 21818) and the National Bioethics Committee for Health (CNBS) in Mozambique (Reference: 313/CNBS/20).Written informed consent will be obtained from each individual prior to her participation in the study by the study investigators.The outcomes of the project will be communicated to the NMCP of the respective countries and to the Global Malaria Programme of the WHO. The results will be published in peer-reviewed open access journals and presented at (inter)national conferences and meetings.A work package is focused on the development of a plan for internal and external project communication, the development of communication tools, and the dissemination and exploitation of the project\u2019s findings. All these activities will be in line with H2020 guidelines on dissemination and publication of results and will highlight the contribution of the EDCTP in tackling societal and health challenges.Active recruitment.The trial sponsor is the University of Sciences, Techniques, and Technologies of Bamako (USTTB). Hamdallaye ACI 2000, Rue 405 - Porte 359, BamakoT\u00e9l\u00e9phone : +223 20 29 04 07http://usttb.edu.ml/The trial sponsor in the study has the responsibility to ensure that data and source documents are granted for trial monitoring, audits, DSMB and Ethics Committee reviews, and regulatory inspections as appropriate."}
+{"text": "Plasmodium vivax malaria in its bid for malaria elimination by 2030. The emergence of anti-malarial drug resistance would challenge the elimination goal. There is limited evidence on the emergence of chloroquine drug resistance. The clinical and parasitological outcomes of treatment of P. vivax with chloroquine plus radical cure using low dose 14\u2009days primaquine were assessed in an endemic area of Ethiopia.Ethiopia rolled out primaquine nationwide in 2018 for radical cure along with chloroquine for the treatment of uncomplicated Plasmodium vivax mono-species infected patients (n\u2009=\u2009102) treated with a 14 days low dose (0.25\u00a0mg/kg body weight per day) primaquine plus chloroquine  were followed for 42 days to examine clinical and parasitological outcomes. Samples collected at recruitment and days of recurrence were examined by 18\u00a0S based nested polymerase chain reaction (nPCR) and Pvmsp3\u03b1 nPCR-restriction fragment length polymorphism. Asexual parasitaemia and the presence of gametocytes were assessed on the scheduled days using microscopy. Clinical symptoms, haemoglobin levels, and Hillmen urine test were also assessed.A semi-directly observed 42-days follow up in-vivo therapeutic efficacy study was conducted from October 2019 to February 2020. Pvmsp3\u03b1 genotyping. No adverse effect was detected related to the low dose 14 days primaquine administrations.Of the 102 patients followed in this study, no early clinical and parasitological failure was observed. All patients had adequate clinical and parasitological responses within the 28 days of follow up. Late clinical (n\u2009=\u20093) and parasitological (n\u2009=\u20096) failures were observed only after day 28. The cumulative incidence of failure was 10.9%  on day 42. Among the paired recurrent samples, identical clones were detected only in two samples on day 0 and day of recurrences (day 30 and 42) using P. vivax before 28 days of follow up. Interpretation of CQ plus PQ efficacy should be done with caution especially when the recurrent parasitaemia occurs after day 28. Therapeutic efficacy studies with appropriate design might be informative to rule out chloroquine or primaquine drug resistance and/or metabolism in the study area.Co-administration of CQ with PQ in the study area is well tolerated and there was no recurrence of  Ethiopia achieved the Global Technical Strategy for Malaria (GTS) target; case incidence decreased by more than 40% in 2020 compared with the 2015 baseline [Plasmodium falciparum and Plasmodium vivax are co-endemic in Ethiopia, with the later contributing to ~\u200940% of the total reported cases [Plasmodium vivax is a harder species to eliminate [P. vivax [P. vivax [P. vivax is primaquine (PQ) [P. vivax [ed cases . Intensiliminate  partly dliminate . In EthiP. vivax . ChloroqP. vivax . The online (PQ) . EthiopiP. vivax . PQ admiP. vivax , and PQ P. vivax to CQ in 1989 [P. vivax was reported from the highlands of Ethiopia in 1996 [Chloroquine has been in use for several decades in Ethiopia. A recent report highlighted high rates of recurrent vivax parasitaemia on day 42 in CQ arm compared to CQ plus PQ arm . This co in 1989  subseque in 1989 \u201317. The  in 1996  followed in 1996 . Treatme in 1996 . Globall in 1996 , 25. TheP. vivax and (ii) the haematologic response, PQ toxicity, gametocytaemia, fever and parasite clearance in 42 days follow up.The aim of this study was to assess (i) the efficacy of CQ plus 14 days low dose of PQ as radical cure treatment for uncomplicated Anopheles arabiensis is the major vector [P. falciparum and P. vivax are endemic with highly seasonal transmission , haemoglobin (Hb) level\u2009\u2265\u20095\u00a0g/dL, asexual parasitaemia\u2009\u2265\u2009250 parasites/\u00b5L, aged above 6 months, ability to swallow oral medication, and a negative pregnancy test or not breastfeeding [Ethical clearance was obtained from the Armauer Hansen Research Institute (AHRI/ALERT) (PO42/18) and the National Research Ethics Review Committee (MoSHE//RD 141/1097/19). Enrolment was made after explaining the purpose of the study and obtaining written informed consent. Consenting febrile patients (axillary temperature \u2265\u00a037.5 or history of fever within the last 48\u00a0h) attending the outpatient department were screened as per the inclusion and exclusion criteria. Volunteer patients who fulfilled the following criteria were recruited: confirmed tfeeding .\u00ae) filter paper and measure haemoglobin (Hb)  using portable spectrophotometer . Hillmen urine test was performed on days 1, 2, 3, 7, and 14 as per the national guideline [A general physical examination was done by study physicians during enrolment. Baseline data, including demography, axillary temperature and anthropometric measures were recorded. At each visit, a symptom questionnaire and adverse or serious adverse events were recorded. At recruitment and during the scheduled follow up days, finger prick blood samples (300\u2009\u00b5L) were collected using EDTA-microtainer tube (Becton Dickinson) for malaria diagnosis using RDT and thick and thin blood films, to prepare dried blood spots (DBS) on Whatman\u2122 3MM (VWRuideline  with sliPlasmodium vivax merozoite protein 3\u03b1 (Pvmsp3\u03b1) genotyping was done on paired samples from day 0 and day of recurrence using nPCR-restriction fragment length polymorphism (RFLP) method [Blood slides were stained with 10% Giemsa  for 10\u00a0m) method .P. vivax recurrence at day 42. Parasitological recurrence was classified as treatment failure on the day it occurred; whereas lost to follow up (LFU), withdrawals (WTH), and parasitaemia with a different species were censored on the last day of follow up. Treatment failures were categorized as (i) early treatment failure (ETF) ; (ii) late clinical failure (LCF)  and (iii) late parasitological failure (LPF) (presence of parasitaemia on any day between day 7 and 28 or day 42 with axillary temperature\u2009\u2264\u00a037\u2009\u00baC in patients who did not previously meet any of the criteria of early treatment failure or late treatment failure) and (iv) adequate clinical and parasitological response (ACPR)  and drug adverse events  fever and asexual parasite clearance  and gametocytaemia at follow up days 1, 2 and 3.P. falciparum infection was censored.Data were double entered using Microsoft Excel sheet and analyzed using the WHO Microsoft Excel and STATA v.14. Frequency counts with percentages , median with ranges (for age and haemoglobin level), arithmetic mean with standard deviations (for temperature and haemoglobin level) and geometric mean with ranges  were done. Change in mean haemoglobin level was compared using Wilcoxon signed-rank test. The cumulative incidence of failure on days 28 and 42 was assessed by survival probability analysis using Kaplan-Meier method. Anyone lost to follow up or withdrew or presenting with P. vivax parasitaemia was not detected until day 28 of the follow up. There was no early treatment failure. Seventy-two (88.9%) patients had adequate clinical and parasitological response (ACPR) on day 42. The cumulative incidence of failure at day 42 was 10.9% (95% CI 5.8\u201319.9%). Sixteen patients were lost to follow up (n\u2009=\u200912) and withdrew consent/assent (n\u2009=\u20094)  were confirmed to be P. vivax with the rest being P. falciparum , mixed species , or negative  using 18\u00a0S based nPCR. Fourteen patients became microscopy positive for Plasmodium parasites within the 42 days of follow up . Of the 5 P. falciparum microscopy positive patients, one was negative by nPCR whilst all P. vivax infections were confirmed. Paired samples from day 0 and day of recurrent parasitaemia of the 3 LCF and 6 LPF were genotyped targeting Pvmsp3\u03b1 gene. Based on the length variants of the PCR products [Hha I enzyme. Overall, eleven different restriction patterns were found. Identical restriction digestion band patterns on day 0 and day of recurrence were observed only for two samples  of the participants had microscopically detectable gametocytes. The vast majority of participants reported that they did not have/use malaria infection prevention tools  were febrile on day one. Fever subsided in all patients on day 2 but on day 3 one patient was found febrile. Asexual parasites were detected in half of the patients on day 1  and only in three patients (3.13%) on day 2. Parasites were cleared completely in all patients on day 3. Similarly, gametocyte clearance mirrored asexual parasite clearance as detected by microscopy. Gametocytes were detected only in 9.90% (10/101) patients on day 1 and only in one patient on day 2. After day 3, no gametocytes were detected (Table\u00a0P\u2009<\u20090.001) was observed in the Hb level between day 0 and 42; the median Hb concentration on day 42  was higher than baseline  , headache (8.91%), nausea 5.69%), abdominal pain (4.95%), chill (3.96%) and vomiting (2.97%). The incidence of adverse events declined from day 1 onwards. Dark-colored urine was not reported except on day 1 in two (1.98%) patients  following the WHO protocol in a P. falciparum and P. vivax co-endemic setting in Ethiopia [P. vivax infections occurred within 28 days of follow up [Following the rollout of PQ for radical cure of Ethiopia . None ofollow up  implyingollow up . In presollow up . Studiesollow up , 20. Simollow up . Similarollow up  indicateP. vivax, including Ethiopia, the risk of P. vivax relapse is generally high [True PQ resistance is really difficult to build, there are case reports of recurring vivax malaria despite an adequate dose of PQ administered with an effective blood schizontocidal agent . It has lly high  and it illy high . In the lly high , 42 and P. vivax relapse is a major global public health concern [P. vivax has been documented in Somalia and South East Asia [PQ failure and  concern , 43. Resast Asia . True trast Asia , 25. Potast Asia , 47, durast Asia , individast Asia , 50, adhast Asia , 25 and ast Asia . Though ast Asia . PQ is aast Asia . Though,ast Asia , and theP. falciparum and one negative. During follow up from among these discordant nPCR mixed samples, one case had P. falciparum infection on day 28 and two were LFU on day 2 and 7. For the rest of cases whether they were mixed, P. falciparum or negative for PCR, parasitaemia were not detected until the end of the study. This achievement might be attributable to the schizonticidal effect of PQ on both P. vivax and P. falciparum [P. vivax cases and five were P. falciparum infection: P. falciparum infections were detected after day 14 which were P. vivax during recruitment. In most malaria-endemic regions, the gold standard for diagnosing malaria is still microscopic examination of stained blood smears. However, in regions where P. falciparum and P. vivax are co-endemic, widespread misdiagnosis has been documented [Nine samples (8.8%) from day 0 were discordant with PCR: six of them were mixed; two were lciparum . Accordilciparum , these dcumented \u201357. MisdP. vivax is ambiguous to assign the causes of recurrence to reinfection, recrudescence or relapse [Pvmsp3\u03b1 genotyping result indicated that among the 9 paired samples (day 0 and day of recurrent), 7 of them had Type A alleles and 2 samples had mixed infections with Type A and Type B alleles for the first sample and Type A and D alleles for the second sample on day 0 but on day of recurrence only Type A allele was detected for the two samples. Pvmsp3\u03b1 genotyping results were used to calculate adjusted cumulative risk of recurrence by day 42 by censoring for heterologous infections [Pvmsp3\u03b1 genotyping for the two paired samples that were Type A size variant, the restriction digestion bands were identical on day 0 and day of recurrence. These identical clones were detected on days 30 and 42 and the rest of recurrent P. vivax parasites were heterologous. As reinfection with a similar genotype had a probability\u2009\u2264\u00a00.002 [P. vivax relapse after PQ therapy [P. vivax relapse after PQ therapy could be reduced. The limitation in present study is that there is no information on the genetic diversity of P. vivax circulating in the study area to increase the level of confidence in judging reinfection with the same or different clone.PCR genotyping of  relapse , 58. In fections . In thisfections . Pvmsp3\u03b1\u2009\u2264\u00a00.002 , recurre\u2009\u2264\u00a00.002 , 62. ThiThe present study showed that the combined CQ plus PQ treatment is well tolerated and resulted in early clearance of both the asexual and sexual stages.P. vivax before 28 days of follow up. Interpretation of CQ plus PQ efficacy should be done with caution especially when the recurrent parasitaemia occurs after day 28. Given the observed failures in later days with the combination regimen, it would be important to conduct therapeutic efficacy studies with appropriate design to rule out CQ or PQ drug resistance in the study area.Co-administration of CQ with PQ in the study area is well tolerated and there were no recurrence of"}
+{"text": "Plasmodium falciparum malaria, but the emergence of the new pfkelch13 R561H mutation in Rwanda, associated with delayed parasite clearance, suggests that interventions are needed to slow its spread. Using a Rwanda-specific spatial calibration of an individual-based malaria model, we evaluate 26 strategies aimed at minimizing treatment failures and delaying the spread of R561H after 3, 5 and 10\u2009years. Lengthening ACT courses and deploying multiple first-line therapies (MFTs) reduced treatment failures after 5\u2009years when compared to the current approach of a 3-d course of artemether\u2013lumefantrine. The best among these options (an MFT policy) resulted in median treatment failure counts that were 49% lower and a median R561H allele frequency that was 0.15 lower than under baseline. New approaches to resistance management, such as triple ACTs or sequential courses of two different ACTs, were projected to have a larger impact than longer ACT courses or MFT; these were associated with median treatment failure counts in 5\u2009years that were 81\u201392% lower than the current approach. A policy response to currently circulating artemisinin-resistant genotypes in Africa is urgently needed to prevent a population-wide rise in treatment failures.Artemisinin combination therapies (ACTs) are highly effective at treating uncomplicated  pfkelch R561H mutations in Rwanda over 5\u2009years.A model investigating 26 possible malaria treatment policy intervention scenarios showed that introducing multiple first-line antimalarial therapies is the most effective single policy change in slowing the spread of artemisinin-resistant  Plasmodium falciparum malaria, but the continued evolution of drug resistance by malaria parasites has the potential to undermine these advances. Since the first appearance of artemisinin resistance in Cambodia in the 2000s2, the spread of molecular markers associated with artemisinin resistance has largely been concentrated in Southeast Asia4. However, the de novo appearance of confirmed markers for artemisinin resistance in Rwanda10 and Uganda12 signals the need for interventions to be considered in the African context13 to minimize the expected health, mortality and economic costs should artemisinin resistance become widespread14.The introduction of artemisinin combination therapies (ACTs) has been instrumental in reducing the burden of 15. Since adoption of AL, the P. falciparum kelch protein 13  R561H mutation has emerged and been validated as a marker for partial artemisinin resistance in samples collected as part of clinical drug efficacy studies between 2012 and 2015 (ref. 3). In contrast to the wild-type clearance rate of 2.7\u2009h, the 561H mutant is associated with a delayed clearance half-life of 7.2\u2009h16. This is similar to the clearance half-life of the 580Y mutation that emerged in Cambodia16. Following the original identification of 561H in the Gasabo district of Rwanda5, additional studies have found 561H in more districts, with recent allele frequency measurements ranging from 0.045 to 0.219 (refs. 10). These findings indicate that drug-policy interventions are now needed to delay the spread of 561H within the local P. falciparum population and to reduce the impact of treatment failures due to artemisinin-resistant parasites.The ACT artemether\u2013lumefantrine (AL) was adopted by Rwanda as the first-line therapy for uncomplicated falciparum malaria in 2006 as part of a comprehensive national strategic plan for malaria controlP. falciparum malaria: AL, artesunate\u2013amodiaquine (ASAQ), artesunate\u2013mefloquine (ASMQ), dihydroartemisinin\u2013piperaquine (DHA\u2013PPQ), artesunate\u2013sulfadoxine-pyrimethamine (AS\u2009+\u2009SP) and artesunate\u2013pyronaridine (AS\u2013Pyr)17. Within sub-Saharan Africa, the predominant therapies deployed are AL and ASAQ18, while the non-ACT formulation of sulfadoxine-pyrimethamine\u2009+\u2009amodiaquine (SPAQ) is commonly used for seasonal malaria chemoprophylaxis19. As such, sub-Saharan Africa faces a constrained drug landscape that requires national drug-policy interventions to be balanced between delaying drug-resistance evolution by the parasite\u2014which leads to increased drug failures over the long term\u2014and ensuring that therapies currently given are highly efficacious.As of March 2023, the World Health Organization (WHO) recommends the following six ACTs for the treatment of uncomplicated 22. Additionally, a more logistically complicated strategy of drug rotation is evaluated along with sequential therapy approaches  and the deployment of triple ACTs. Table Studies detecting the mutant 561H allele indicate both increasing allele frequency and geographic spread from 2014 to 2019; however, there may still be a window of opportunity to delay or prevent the fixation of 561H in Rwanda and avert high numbers of treatment failures. Accordingly, using a mathematical model of malaria transmission, we examined 26 possible drug-policy interventions (the majority of which use existing therapies) and their ability to slow down 561H evolution and reduce long-term treatment failures. These include the replacement of the existing first-line therapy, introduction of multiple first-line therapies (MFTs) and lengthening the dosing schedule for AL from a 3-d course of treatment to up to 5\u2009d of AL treatment in accordance with previous clinical trialsn\u2009=\u2009100 replicates) in which no interventions were implemented. Our spatially calibrated model initially uses the 2017 Malaria Atlas Project (MAP) projections for Rwanda\u2019s malaria prevalence ) through the end of 2020, after which prevalence is scaled down in agreement with current incidence estimates for Rwanda23. Following this transmission reduction to the 2021 malaria incidence, the model presumes that incidence will remain stable over time. The allele frequency of 561H is calibrated  scenario was run  of treated P. falciparum malaria cases will fail treatment with the present first-line therapy of AL, or about 154,000 individuals for the calendar year ). This monthly average is forecast to increase to 39,400  per month by 2033  and 5-d AL results in a 561H allele frequency of 0.69  under 4-d AL and 13,100  under 5-d AL. Treatment failure numbers are improved under a longer course of AL because of the combined effect of lower 561H frequency and higher treatment efficacy of the longer course. AL efficacy generally remained high in the model\u2019s parameterization with the majority of AL efficacies, across all genotypes in the simulation, above 85%.Among alternate first-line therapies, extending the course of AL from 3 to 4 or 5\u2009d is the most immediately available option due to stocks of AL already being present and available. Continued use of a 3-d AL course for 5\u2009years is projected to lead to a median 561H allele frequency of 0.82 (IQR: 0.73\u20130.86), whereas 4-d AL results in a 561H allele frequency of 0.76  and a median of 14,100  treatment failures per month. However, a switch to DHA\u2013PPQ results in an acceleration of the fixation of 561H with the allele frequency reaching 0.90  within 5\u2009years and fixation within 10\u2009years  after 5\u2009years and 83,300  after 10\u2009years. When conducting sensitivity analysis for scenarios where DHA\u2013PPQ efficacy remains relatively high, switching to DHA\u2013PPQ is still projected to reach the 10% treatment failure threshold within 5\u2009years of deployment, although the long-term treatment failure rates are lower . Nine combinations of AL, ASAQ and DHA\u2013PPQ with distribution ratios of 25/75, 50/50 and 75/25 were considered (Table P\u2009<\u200910\u22124 compared to 3-d AL). Within 10\u2009years, all MFT approaches are projected to exceed 10% treatment failure, although combinations of 50% AL\u2009+\u200950% ASAQ and 25% AL\u2009+\u200975% ASAQ have comparably acceptable outcomes with 15.5%  and 14.5%  treatment failure rates, respectively. The high percentage of treatment failures in MFTs incorporating DHA\u2013PPQ at high levels  monthly treatment failures ; Supplementary Table P\u2009<\u200910\u22124). The procurement and distribution required to deploy a drug rotation coupled with MFT suggest that compliance and operations would have a considerable influence on the success of this approach. In general, MFT strategies promote the emergence of a large number of distinct genotypes, but selection pressure is weak on each genotype, limiting their ability to reach high allele frequency , minimization of safety risks by changing partner drugs and potential exploitation of partner drugs selecting for opposite alleles. We explored this approach in the following two ways: following the protocol discussed in ref. 29 with six consecutive days of treatment from day 0 to day 5 inclusive, and a modified protocol in which the second course is taken on days 7, 8 and 9 . These are lower than the median 561H frequencies projected for all therapy switches, MFT approaches and rotations considered thus far. ASAQ followed by AL is also projected to have the lowest treatment failure rate 5\u2009years after introduction at 1.8% . All sequential courses are projected to have median treatment failure rates below 4% after 5\u2009years and below 12% after 10\u2009years. As in other scenarios, 10\u2009years of DHA\u2013PPQ use as part of a strategy of sequential ACT courses still results in strong selection pressure for PPQ resistance with long-term treatment failures increasing correspondingly  and a 561H allele frequency of 0.43  after 5\u2009years, while ASMQ\u2013PPQ resulted in 1,900  monthly treatment failures and a 561H allele frequency of 0.47 . Median treatment failure rates are projected to be 1.9% and 1.8%, respectively, after 5\u2009years. The usage of ASMQ\u2013PPQ comes with an increased risk of PPQ failure leading to projected treatment failures of 18.8%  at 10\u2009years after deployment. In contrast, treatment failures are likely to still be low if ALAQ is deployed reaching only 2.1%  10\u2009years after deployment  and artesunate\u2013mefloquine\u2013piperaquine (ASMQ\u2013PPQ)34. To evaluate the possible impacts that compliance with treatment regimens would have, additional scenarios were evaluated in which ASAQ, DHA\u2013PPQ and 3-, 4- or 5-d courses of AL were administered using low (25\u201370%), moderate (50\u201380%) and high (70\u201390%) compliance rates for complete courses , resulting in real-world compliance rates between 65% and 90% for a 3-d course of treatment10, from its initial identification5, together with confirmation of 561H at frequencies similar to those projected in our simulation, suggests that the 561H allele is likely present throughout Rwanda. As recommended by the WHO, the Rwandan National Malaria Control Program (NMCP) should consider several strategies for mitigating the spread of pfkelch13 alleles associated with artemisinin resistance13.The projected national frequency of the 561H allele in Rwanda under a continuation of status quo treatment with a 3-d course of AL suggests that treatment failures will increase over the next 5\u2009years and that drug-policy interventions are required to mitigate this risk as much as possible. The current spread of the 561H allele to other districts36, the incidence of adverse cardiac events recorded during clinical trials has been low38. Switching to one of several MFT options where future treatment failure rates can be kept close to 10% will ensure there are no concerns with extended artemisinin dosing, and the success of these MFT deployments will depend on operational capability around the supply and distribution of ACTs. Based on the structure of the distribution network for antimalarials in Rwanda and the current availability of different ACTs, an MFT deployment is more likely to be feasible than a custom rotation approach, although it lacks the logistical ease of a single recommended first-line therapy.Our findings suggest that over a 5-year time frame extending the use of AL from 3 to 5\u2009d may hold treatment failure rates at or near the 10% threshold and switching to an MFT strategy is also worth considering with the optimal MFT approach\u2014a 75% ASAQ and 25% DHA\u2013PPQ deployment\u2014projected to hold treatment failure rates to 9.9% after 5\u2009years. Extending the course of treatment with AL has minimal logistical considerations beyond ensuring sufficient quality of doses being distributed and appears to be beneficial compared to the status quo scenario. Although there are concerns regarding the cardiotoxicity of antimalarial drugs39, and a number of the key mutations associated with these phenotypes are now known40. However, there is no guarantee that the same PPQ-resistance mutations will emerge in Africa. Any DHA\u2013PPQ-centered strategy in Africa should be paired with routine rapid-turnaround molecular surveillance for known markers of PPQ resistance. Similarly, for lumefantrine and amodiaquine, projected scenarios of resistance evolution come with uncertainty, but the effects of currently circulating parasite mutations on the efficacies of AL and ASAQ can be estimated.The major uncertainty in these strategy comparisons is that the future course of PPQ resistance in Africa cannot be predicted through any modeling approaches, in vitro studies or clinical trials. The DHA\u2013PPQ-resistant lineage that emerged in Southeast Asia led to 58% treatment failure under 3\u2009d of DHA\u2013PPQ treatmentCertain new therapeutic approaches to resistance management will need specific national approval or WHO prequalification. Triple ACTs and sequential ACT courses have the advantage of higher total parasite killing than a 3-d ACT and thus an associated lower treatment failure rate. All sequential and triple ACT approaches are projected to keep treatment failure rates below 4% over a 5-year period, with no noticeable advantage seen for sequential approaches despite their higher total dose of artemisinin. A major limitation of our modeling approach is that each therapy\u2019s pharmacodynamics and pharmacokinetics are simplified to daily killing and drug elimination rates; a more detailed analysis will be required to understand whether we should expect true differences in failure rates for these therapies. However, for both triple ACTs and sequential ACT courses, the combination of three drugs used over a short period lowers the probability of multidrug resistance emerging during a treatment course and slows down the spread of artemisinin resistance over the long term. Triple ACTs will likely enter the WHO approval process in the next year, while approval for sequential courses may have to be sought on a case-by-case basis.41 from 2006 to 2014 that has not been systematically captured in any known data collections. Furthermore, while the data and model calibration indicate that 561H initially appeared in the Gasabo district, the design of the simulation renders it incapable of fitting the highly stochastic process that would have driven the initial appearance and spread of 561H. Additionally, although it is possible that the dominant strain of P. falciparum circulating in Rwanda may be fitness neutral42, the model takes a more traditional assumption and includes a fitness penalty for the 561H mutation.To further contextualize this study within the limitations inherent in modeling studies, due to the introduction of AL in 2006, other genomic mutations, such as those associated with reduced lumefantrine susceptibility, are present in Rwanda. The introduction of mutations in the model year 2014 represents a \u2018model fitting compromise\u2019 made necessary due to the stochastic nature of the rare mutation emergence processPyronaridine\u2013artesunate deployment was not considered in our simulations as pyronaridine-resistant falciparum phenotypes have not yet been described. Assuming similar rates of emergence and similar drops in ACT efficacy due to future pyronaridine resistance, pyronaridine\u2013artesunate would likely make a positive contribution as an addition to any of the MFT strategies shown in Table 43). The most recent annual case estimates for the fiscal period of July 2021\u2013June 2022 place P. falciparum case numbers at around 1\u2009million annually, with 1.16\u2009million reported for calendar year 2021 to the WHO44. With these recent gains, resistance management strategies should seek to keep absolute treatment failure counts below 10,000 monthly for the medium term.Another major limitation for our model calibration\u2014and any malaria modeling exercise evaluating the last 5\u2009years of epidemiological changes\u2014is that the effects of the COVID-19 pandemic on malaria control are not easily quantified. A mixed-methods study of three high-endemic districts in Rwanda  suggests that while the distribution of malaria testing shifted, the overall decline in uncomplicated malaria cases continued through 2020 (ref. 45)) are likely to reduce biting rates and number of cases in the coming years. A recent pilot study in a rural part of the Gasabo district with high malaria risk showed that larviciding activity had a moderate effect on reducing incidence; the NMCP also expects to expand larviciding as part of a new set of malaria control activities. These interventions may prove beneficial in the fight against drug resistance as they may eliminate pockets of transmission, including drug-resistant alleles, and are likely to slow the overall geographic spread of all genotypes. These activities and their effects are not included in the present modeling analysis.Rwanda\u2019s NMCP\u2019s current activities to expand indoor residual spraying activities (from 2\u20138 districts last decade to 12\u201315 districts this decade) and to introduce synergistic insecticide-impregnated nets  were used as the basis to calibrate local transmission parameters on a 5-by-5\u2009km (25\u2009km2) scale in Rwanda  to 410.26 per 1,000 (Nyamasheke district) in 2022 following the switch to the incidence-based prevalence calibration. This results in an incidence of 139.13 per 1,000 for symptomatic cases and an incidence of 87.76 per 1,000 for treated cases at the national level in 2022, consistent with reporting that the incidence rate has been declining from a high of 403 per 1,000 in 2016  to 465.62 per 1,000 (Gisagara district) in 2020 under the MAP pfkelch 561H mutation were available at the district level\u2014four close to the capital Kigali9, two from the eastern district of Kayonza6 and one each from the districts of Huye7, Kirehe and Ngoma10. To calibrate to these values, the simulation was seeded with a single mutation of 561H genotypes in Gasabo district  to generate a slowly growing exponential curve with district frequencies that are consistent with measured values  will have an oversized effect on human movement dynamics. To account for the ability of ACT partner-drug resistance to accelerate the fixation of artemisinin resistance24, mutations affecting other alleles  are enabled in the simulation on 1 January 2014, following the completion of model burn-in using a previously calibrated mutation rate24. This produces a slight model delay in pfcrt and pfmdr1 mutations that are associated with the use of lumefantrine given the adoption of AL by Rwanda in 2006, likely resulting in a slight model bias toward low frequencies of alleles associated with lower lumefantrine susceptibility.Because the mutation rate to 561H alleles is not known, these artificially introduced 561H genotypes are the only means by which 561H can be introduced into the simulation. The spatial spread of 561H is driven by human movement and migration within the simulation, which is based on the gravity model described in ref. PfPR2\u201310 as projected by MAP50 versus the simulated PfPR2\u201310 before 2021; the district-level clinical cases projected by the simulation and the projected 561H frequency. To calibrate the PfPR2\u201310, the local transmission intensity, or \u03b2, was determined using a constrained parameter space search. This was performed by first binning the population in each 25\u2009km2 cell using Jenks natural breaks optimization followed by scanning the domain of possible \u03b2 values using a fixed population  along with the relevant population and climatic variables. Upon determining the possible \u03b2 values, they were assigned to cells in the model by matching the population in each cell to the appropriate bin and assigning the calculated transmission intensity. Once this process was complete, the model was run and assessed against a target deviation of MAP PfPR2\u201310 values to within \u00b110% . Starting with the total clinical cases per 1,000, it is clear that the spatial distribution of model-generated clinical cases is distributed in a manner that is consistent with 2017 true incidence50, although the counts of both all clinical cases , supporting the model as having a 561H introduction that is properly calibrated. As a note of caution, although this calibration suggests a mechanism through which 561H may have spread in Rwanda, the model was not designed, configured or calibrated to explore the nature of the introduction event or original mutation event.The final point of model calibration and validation is the 561H frequency. Presently, 561H frequency data are only available for Huye, Kayonza, Kirehe and Ngoma districts, along with the province of Kigali, consisting of Gasabo, Kicukiro and Nyarugenge districts . This baseline scenario also controls for any deviations in the model versus real-world data for Rwanda by allowing all policy interventions to be compared to the same projected outcome and presumes that the calibrated malaria incidence remains stable over time. A total of 26 drug-policy intervention scenarios were evaluated within the simulation  followed by replacement of DHA\u2013PPQ with one of two MFT strategies using AL and ASAQ. Four interventions replicated the therapeutic arms of the sequential ACT therapy regimen proposed in ref. 29 with AL followed by either ASAQ or DHA\u2013PPQ, or AL preceded by ASAQ or DHA\u2013PPQ. However, the model scenario deviates from the protocol proposed in ref. 29 by applying sequential therapy to all treatment-seeking individuals as opposed to just children aged 6\u2013120\u2009months. The second of the sequential therapies was given either on treatment days 3, 4 and 5 or on treatment days 7, 8 and 9; results for both timings of treatment courses are presented. Finally, two interventions considered the replacement of AL with one of two triple therapies (ALAQ and ASMQ\u2013PPQ), which are yet to be approved46.The policy scenarios run may be summarized as follows. Five evaluated interventions involved a change in first-line therapy to a 4-d course of AL, 5-d course of AL, ASAQ and DHA\u2013PPQ. Nine of the interventions involved MFT approaches with drug distribution proportions ranging from 25/75 to 50/50 to 75/25  and the allele frequency trajectories only weakly tracked the observed data points , moderate (50\u201380%) and high (70\u201390%) compliance rates for complete courses  and pessimistic scenarios in which African strains are more likely to develop PPQ resistance (1.25\u00d7 and 1.5\u00d7 scenarios). For all four scenarios, the annual percentage of treatment failures exceeds the 10% treatment failure threshold within 5\u2009years after switching from AL to DHA\u2013PPQ . The details of the pharmacokinetic/pharmacodynamic model are described in ref. 48, and the model uses calibrated values of 0.58 for PPQ-sensitive and 1.4 for PPQ-resistant parasites, where 1.0 is set as the standard therapeutic dose given in the simulation. The simulation was then run with values ranging from 1.0 to 1.6 for the EC50 of PPQ. As expected, an EC50 value less than the configured value of 1.4 had fewer treatment failures at 5\u2009years while 1.5 and 1.6 produced more treatment failures  are tested with nonparametric Wilcoxon rank-sum tests, and all 2) cell is the primary spatial unit in the simulation, the same size as the resolution of the MAP PfPR2\u201310 projections used as part of the model calibration50. This cellular resolution results in a modeled space of 979 cells covering 24,475\u2009km2 or about 93% of the total area of Rwanda , and the population skews younger with about 54.3% being under the age of 20\u2009years61 , the relevant therapy was given to the individual and the efficacy was assessed using the parasite density at 28\u2009d after the first treatment. Individuals with a parasite density less than 10\u2009ml\u22121 of blood were counted as cleared (failed otherwise). Drug efficacies of longer courses\u2014for example, 5-d AL or two sequential consecutive courses of ACT\u2014are calculated via a one-compartment pharmacokinetic model and traditional Hill-function pharmacodynamic model, using a 1-d time step48.As part of the model calibration and validation, the drug efficacies were used as previously calculatedP. falciparum is 95.5%, while the efficacy of 5-d AL is 97.5%. The complete drug efficacies are included in Supplementary Tables 25.The lowest efficacy in the simulation is that of DHA\u2013PPQ on the 561H genotype, which also carries PPQ resistance (43.3% efficacy); this is also the reason that PPQ resistance spreads so quickly in the model simulations. The efficacy of 3-d AL on wild-type This study included researchers from the Rwanda Biomedical Research Center (who were colocated and in close collaboration with Rwanda\u2019s NMCP). Under the leadership of A. Uwimana and C. Ngabonziza, they planned the major study-design aspects for this paper and assisted the mathematical modeling team in identifying drug-resistance-management strategies that could be feasibly implemented in Rwanda. Research roles and responsibilities were discussed throughout 2022, including during in-person meetings in August and October 2022. Capacity co-enhancement collaborations have already begun by planning (1) a local modeling instance to be run in RBC Kigali with M. Kabera as the primary analyst and (2) a series of tutorials for Penn State scientists to understand the operational aspects of drug delivery and distribution in Rwanda.Further information on research design is available in the Any methods, additional references, Nature Portfolio reporting summaries, source data, extended data, supplementary information, acknowledgements, peer review information; details of author contributions and competing interests; and statements of data and code availability are available at 10.1038/s41591-023-02551-w.Supplementary InformationSupplementary Tables 1\u20134  and Figs. 1\u20137 .Reporting SummarySupplementary TablesSupplementary Table 1: 561H frequency, published studies; Supplementary Table 2: primary study results; Supplementary Table 3: piperaquine sensitivity studies; Supplementary Table 4: compliance rates for sensitivity analysis; Supplementary Table 5: results for sensitivity to compliance rate; Supplementary Table 6: drug efficacy primary studies; Supplementary Table 7: drug efficacy compliance studies."}
+{"text": "After treatment, virus load was reduced in all groups (p\u2009<\u20090.0001) but was greater in the 0.1% group compared to placebo (p\u2009=\u20090.007). In a subset of patients  viral load was strongly reduced on day 4 in the 0.1% group compared to placebo (p\u2009=\u20090.005). Negative PCR results appeared earlier and more frequently in the azelastine treated groups: being 18.52% and 21.43% in the 0.1% and 0.02% groups, respectively, compared to 0% for placebo on day 8. Comparable numbers of adverse events occurred in all treatment groups with no safety concerns. The shown effects of azelastine nasal spray may thus be suggestive of azelastine\u2019s potential as an antiviral treatment.With the changing epidemiology of COVID-19 and its impact on our daily lives, there is still an unmet need of COVID-19 therapies treating early infections to prevent progression. The current study was a randomized, parallel, double-blind, placebo-controlled trial. Ninety SARS-CoV-2 positive patients were randomized into 3 groups receiving placebo, 0.02% or 0.1% azelastine nasal spray for 11\u00a0days, during which viral loads were assessed by quantitative PCR. Investigators assessed patients\u2019 status throughout the trial including safety follow-ups (days 16 and 60). Symptoms were documented in patient diaries. Initial viral loads were logTrial registration: The study was registered in the German Clinical Trial Register . EudraCT number: 2020-005544-34. The active substance (azelastine hydrochloride) is a histamine-1 receptor antagonist, which shows anti-inflammatory effects via mast cell stabilization and inhibition of leukotriene and pro-inflammatory cytokine production10. In an in vitro screening of 1,800 approved drugs by use of a SARS-CoV-2-S pseudovirus entry inhibitor model, 15 drugs were identified as active inhibitors, but only seven of these drugs were identified as active against SARS-CoV-2, three of which were anti-histamines: clemastine, trimeprazine and azelastine hydrochloride5. Reznikov et al. analyzed 219,000 medical records in a retrospective data base survey study and demonstrated that azelastine showed the highest association between prior usage among these antihistamines and SARS-CoV-2 negative test results in patients above the age of 60 . Antiviral activity was subsequently verified in cell culture. Moreover, this group showed that azelastine has the potential to inhibit SARS-CoV-2 cell entry by binding to the angiotensin-converting enzyme 2 (ACE2) receptor and to inhibit intracellular virus replication through binding to the sigma-1 receptor6. Furthermore, three independent groups predicted interaction of azelastine hydrochloride with the main protease of SARS-CoV-2: main protease (Mpro) or 3C-like cysteine protease (3CLpro)9. Ghahremanpour et al. also provided experimental evidence for the inhibition of the enzyme in a kinetic activity assay7.Since the start of the Coronavirus Disease 2019 (COVID-19) pandemic, several independent research groups revealed azelastine\u2019s potential as a promising candidate for drug repurposing to reduce SARS-CoV-2 viral load and infection rates10. Antiviral efficacy was observed at an EC50 of\u2009~\u20096\u00a0\u00b5M, which is an approximately 400-fold lower concentration compared to commercially available azelastine nasal sprays. In a highly relevant and translational in vitro model using reconstituted human nasal tissue, a fivefold diluted commercially available azelastine nasal spray solution inhibited viral replication almost completely within 72\u00a0h after SARS-CoV-2 infection10.By application of a novel computational approach based on Shannon entropy homology, Konrat et al. identified azelastine as an anti-viral candidate and demonstrated pronounced anti-SARS-CoV-2 activity in vitroCOVID-19: Azelastine nasal spray Reduces Virus-load In Nasal swabs).The aim of our study was to support the preclinical evidence for azelastine\u2019s antiviral activity in patients tested positive for SARS-CoV-2. The study was termed CARVIN  was given on 3rd February 2021.Ethics approval was granted by the Ethics Committee of the Faculty of Medicine of Cologne University on the 10Informed consent was obtained from all participants prior to involvement in the study.All methods were carried out in accordance with relevant guidelines, and the principles of Good Clinical Practice and the Declaration of Helsinki were adhered to.This trial was conducted at the Department of Otorhinolaryngology, Head and Neck Surgery of the Faculty of Medicine of the University of Cologne, Germany. Outpatients visiting Corona test centres were informed about the possibility of participating in the trial. Patients aged 18 to 60\u00a0years were eligible to participate if tested positive for SARS-CoV-2 in a Corona test centre by PCR test within 48\u00a0h prior to inclusion and had to quarantine at home due to instructions of the local health authority. A complete list of inclusion and exclusion criteria is presented in Table 11. Short intervals of swab collection time points, particularly during early days of infection, and high number of PCR tests aimed to monitor SARS-CoV-2 viral loads as closely as possible, considering that only limited knowledge regarding details of viral clearance was publicly available at the time of the study development. Additionally, safety follow-ups were performed at 2 time-points. On day 16, an on-site visit (V8) for female patients was conducted to perform a urine pregnancy test and to assess the safety of the therapy. For male patients, the assessment was done via phone call. A final safety follow-up and assessment of the patient status  by phone call was done on day 60 (V9) for all patients.This was a prospective, randomized, double-blind, placebo-controlled dose-finding proof-of-concept study, in which azelastine nasal spray was used in 2 doses: the commercially available concentration of 0.1% and a fivefold lower concentration of 0.02%. After having given informed consent, patients tested positively for SARS-CoV-2 were examined to assess eligibility according to inclusion/non-inclusion criteria and subsequently randomized to one of the three study groups. The first administration of the nasal spray was carried out in the presence of the investigator; products were subsequently self-administered for 11\u00a0days (treatment phase). During the treatment phase, 7 visits (V1\u2013V7) took place on days 1, 2, 3, 4, 5, 8 and 11. Samples of day 1 represent pre-treatment baseline samples. During visits, nasopharyngeal swabs were taken for quantitative PCR measurements, and investigators assessed the patient status in accordance with the WHO clinical progression scalePatient reported outcomes were documented by patient diaries and questionnaires. Therefore, during the treatment phase, patients were required to document the severity of their COVID-19 related symptoms in an electronic diary on a daily basis. On days 1, 5, 8 and 11, patients completed the standardized SF-36 questionnaire of quality of life. A summary of study activities is displayed in Table Assignment of the treatment with the investigational medicinal product in the different doses vs. placebo to each treatment number was performed in a centrally conducted, computer-generated 1:1:1 randomization procedure. Treatment kits were manufactured by URSAPHARM Arzneimittel GmbH, Saarbruecken, Germany, according to the randomization list . Patients were assigned a treatment number in an ascending mode according to their chronological order of inclusion. Investigators and trial participants were masked to the treatment as investigational medicinal products were identical in appearance.\u00ae) was manufactured at URSAPHARM Arzneimittel GmbH, Saarbruecken, Germany). All nasal sprays were composed of hypromellose, disodium edetate, citric acid, disodium phosphate dodecahydrate, sodium chloride and purified water. Additionally, 0.02% azelastine nasal spray and 0.1% azelastine nasal spray were formulated by the addition of 0.2\u00a0mg/mL or 1\u00a0mg/mL azelastine hydrochloride, respectively. One puff of the respective nasal spray was applied per nostril, 3 times a day .The trial medication . Following sampling, swabs were placed into 3\u00a0mL Virus Transport Medium  and delivered to the laboratory as quickly as possible. If delivery took place within 24\u00a0h after sampling, samples were to be stored at\u2009<\u200925\u00a0\u00b0C, if storage period was greater than 24\u00a0h , samples had to be stored and shipped at 2\u20138\u00a0\u00b0C. Samples were processed on the day of receipt at the central processing laboratory  by vortexing and aliquoting the viral transport medium and stored at\u2009\u2212\u200980\u00a0\u00b0C until analysis.\u00ae SARS-CoV-2 Test on the cobas\u00ae 6800 system . For quantification of SARS-CoV-2-RNA in copies/mL, a standard curve derived from a dilution series of a SARS-CoV-2 cell culture isolate in VTM and adjusted to Ct values obtained from two samples with defined SARS-CoV-2-RNA copy numbers  was used. For calibration purposes of quantitative assessments, reference samples were included with each PCR run. The dual-target RT-PCR independently targets the ORF1a/b and the sarbecovirus E genes, and assays were considered positive if at least one target returned a positive result . Of note, in vitro tests carried out prior to the current study did not indicate any interaction between the study products and the PCR reaction  at the time of the study. Symptoms were evaluated on a 5-point scale from 1\u2009=\u2009symptom absent or present very weakly to 5\u2009=\u2009symptom present very strongly: anosmia, ageusia, cough, sore throat, shortness of breath, coryza, general weakness, headache, aching limb, loss of appetite, pneumonia, nausea, abdominal pain, vomiting, diarrhea, conjunctivitis, rash, lymph node swelling, apathy, somnolence. In addition, presence or absence of fever (\u2265\u200938.0\u00a0\u00b0C) was documented daily . Symptoms were analyzed as single symptom scores, and as the total symptom score (TSS) reflecting the sum of all 20 single symptoms and presence/absence of fever .Patients had to daily document their COVID-19 specific symptoms in an electronic patient diary. Those parameters were based on the COVID-19 symptoms published by the Robert Koch Institute \u2019, \u2018good (2)\u2019, \u2018moderate (1)\u2019 or \u2018poor (0)\u2019.11. In addition, investigators measured body temperature during V1\u2013V7 and oxygen saturation of the blood (using a finger pulse oximeter) on V1, V3, and V5, V6 and V7.The patient status was assessed at V1\u2013V7 and at V9 by the investigators with a 11-category ordinal score proposed by the WHO11), body temperature and blood oxygen saturation, quality of life  and safety  over time.The primary endpoint of the CARVIN study was the assessment of virus load kinetics of SARS-CoV-2 by determining the presence and amount of viral carriage via PCR. Applied treatment regimens aimed to explore differences regarding viral carriage upon treatment with azelastine compared to placebo. Secondary endpoints included the assessment of symptoms, patient status  to result in 23 patients per treatment group completing the study and being eligible for analysis.The sample size calculation was based on the expected reduction of virus load during the treatment considering 3 treatment arms. It was assumed that all treatment groups present identical baseline virus load at enrolment with a mean value of 5.5 logData was analysed primarily exploratively; there was no formal testing of a given hypothesis. Analyses were done on the entire data set (ITT) as well as on a subset population with high viral load defined by baseline Ct values below 25 (Ct\u2009<\u200925). Both descriptive and exploratory statistics were performed. Subgroups were analysed exploratorily .Continuous data were described by statistical estimates . Categorical data were described by absolute frequencies and percentage of valid cases. Ct values reported as \u201cnegative\u201d were replaced with the value 45, and respective cp/mL values with the value 1, and cp/mL values\u2009<\u20092116 (ORF 1a/b gene) and cp/mL values\u2009<\u20091950 (E gene) were replaced with the value 1.U test was performed, and significance levels were adjusted to p\u2009<\u20090.0167 based on the Bonferroni correction. Kaplan\u2013Meier survival analyses with log-rank test were performed to display the occurrence of negative PCR test results upon treatment. To evaluate the total load during the study, AUC was calculated using a linear equation.Study endpoints were presented by descriptive statistics, aiming to compare the course of viral load between the three treatment groups. Whereas PCR data of individual days served for daily comparisons between treatment groups, the area under the curve (AUC) value was used for the evaluation of the overall development of viral kinetics. While comparison of categorial variables between groups were performed by Chi square testing, continuous variables were compared using ANCOVA with the factors baseline, visit, and treatment group. All tests were performed two-sided and the type 1 error (\u03b1) was set to 5%. Three-group comparisons were analysed with Kruskal\u2013Wallis test. For pairwise comparisons between treatment groups, Mann Whitney 15. It should be noted that the SARS-CoV-2 alpha variant (B.1.1.7) was the dominant variant in Germany during the enrolment phase of the current study16.Preliminary results of the current study have been published as preprint90 patients were recruited between 09/03/2021 and 28/04/2021, constituting the safety analysis set. Of those, 81 patients belonged to the Intention-To-Treat (ITT) population, comprising randomised patients meeting the key eligibility criteria and having evaluable viral load data on day 1 (baseline) and on day 11 (end of treatment). Of those, 27 patients belonged to the 0.1% azelastine group, 28 patients to the 0.02% azelastine group and 26 patients to the placebo group  of the safety analysis set was male, and the average age was 35.67\u2009\u00b1\u200912.94\u00a0years. The mean bmi of participants was 24.91\u2009\u00b1\u20095.27. Small differences were found with regard to age and bmi, which were both slightly higher in the azelastine 0.1% group . Therefore, the primary analysis for the viral loads was conducted non-parametrically. For clarity reason, only cp/mL values of the ORF 1a/b gene are shown in the main text of the manuscript. As a sensitivity analysis based on the SARS-CoV-2 E gene PCR tended to show overall the same effects, PCR results of the E gene are shown in the supplementary material . Data on virus variants was available for 59 patients and 54 (92%) of those carried the alpha (B.1.1.7) variant.The median/mean viral load value (ORF 1a/b gene) of the ITT analysis set at enrolment was log10 4.45\u2009\u00b1\u20092.26 in the 0.1% azelastine group, log10 4.12\u2009\u00b1\u20092.01 in the 0.02% azelastine and log10 3.82\u2009\u00b1\u20091.61 in the placebo group .As expected, a continuous decrease in the mean virus load was observed in all study groups during the 11 treatment days. The reduction of virus load (reflected by decreases of ORF 1a/b gene copy numbers) from baseline to the end of treatment (day 11) was logoup Fig.\u00a0. The red10 5.04\u2009\u00b1\u20092.05 in the 0.1% azelastine group, log10 4.39\u2009\u00b1\u20091.74 in the 0.02% azelastine and log10 4.15\u2009\u00b1\u20091.34 in the placebo group. Of note, the decrease of viral load on day 4 was significantly greater in the 0.1% azelastine group (decrease by log10 1.90\u2009\u00b1\u20091.03) compared to placebo (decrease by log10 1.05\u2009\u00b1\u20090.70).Within the subgroup of patients with baseline Ct values below 25, a similar progression of viral load data was observed Fig.\u00a0. The virp\u2009=\u20090.007, Fig.\u00a0p\u2009=\u20090.022, Fig.\u00a0Evaluation of AUC values  showed that the 0.1% azelastine group exhibited a greater AUC value of 24.14\u2009\u00b1\u200913.12 (referring to greater decrease) compared to the placebo group with an AUC value of 18.89\u2009\u00b1\u20094.70  and 6 of the 28 (21.4%) patients in the 0.1% azelastine and 0.02% azelastine groups, respectively were negative for the ORF1a/b gene, compared to the 0 of 26 patients in the placebo group. At the end of the treatment, 48.2% of the patients of the 0.1% azelastine group showed no detection of the ORF 1a/b gene, whereas only 23.1% of patients of the placebo group showed negative PCR results (supplementary Table p\u2009=\u20090.112).Kaplan\u2013Meier survival analyses underlined those findings, indicating that mean times of a PCR result to turn negative was 9.96\u00a0days (95% CI: 9.02\u201310.90) in the 0.1% azelastine group, 10.21\u00a0days (95% CI: 9.57\u201310.86) in the 0.02% azelastine group and 11.00 (95% CI: 10.00\u201310.77) in the placebo group  suffered from moderate symptoms  on day 1 of the study (supplementary Table p\u2009=\u20090.004) and 4 (p\u2009=\u20090.011) in the 0.1% azelastine group compared to placebo .The WHO clinical progression scale progressively decreased in all treatment groups during the study. At V1, a comparable distribution of patients with a score of 1  or 2  was observed. At the end of the study (day 60), all except one single patient (placebo group) showed a score of 0.Overall, none of the participating patients had clinically relevant increased values of body temperature (data not shown). Similarly, no clinically relevant differences regarding blood oxygen saturation values were detected between groups (data not shown).59.3% (0.1% azelastine treatment), 50.0% (0.02% azelastine treatment) and 80.8% (placebo treatment) of patients assessed the overall tolerability of the treatment as \u2018very good\u2019, which mirrored the tolerability judgement of the investigators, which was assessed as \u2018very good\u2019 for 59.3% (0.1% azelastine treatment), 50.0% (0.02% azelastine treatment) and 80.8% (placebo treatment) of patients. The efficacy of the treatment was judged as \u2018good\u2019 or \u2018very good\u2019 by 75.0% (0.1% azelastine treatment), 74.1% (0.02% azelastine treatment) and 50.0% (placebo treatment) of patients. The investigators judged the efficacy as \u2018good\u2019 or \u2018very good\u2019 in 74.1% (0.1% azelastine treatment), 82.1% (0.02% azelastine treatment) and 73.1% (placebo treatment) of treated patients. Overall, no statistical differences between groups were determined.n\u2009=\u20094], loss of smell [n\u2009=\u20094], loss of taste [n\u2009=\u20093], [muscle] weakness [n\u2009=\u20092], tiredness/exhaustion [n\u2009=\u20092], muscle ache, concentration impaired, headache, and cough).The number of possibly and probably related adverse events was comparable between treatment groups , the sigma-1 receptor (among other factors) plays a role in viral replication. It has been suggested that azelastine can inhibit the entry of the SARS-CoV-2 into the nasal mucosa by binding to the ACE2 receptor and also act via binding to the main protease of SARS-CoV-2 and to the host cell\u2019s sigma-1 receptor, therewith facilitating both viral entry and replication-inhibiting effectsThe current proof-of-concept study served to investigate if nasally applied azelastine may have the potential to reduce the viral load  in patients tested positively for SARS-CoV-2.17. Thus, a nitric oxide nasal spray was shown to reduce the viral load in adult patients with mild COVID-19 infection, and an accelerated SARS-CoV-2 clearance compared to placebo was demonstrated18. The preventive application of a hydroxypropyl methyl cellulose nasal spray showed promising results in an observational survey, indicating that it may reduce SARS-CoV-2 infection rates19. Treatment of COVID-19 with a hypertonic solution containing seawater, xylitol, panthenol and lactic acid was shown to reduce the viral shedding time in patients with asymptomatic or mild COVID-1920, whereas application of povidone iodine nasal spray showed only poor influence on SARS-CoV-2 viral titres22.Since the start of the COVID-19 pandemic, its treatment via the nasal route has been studied for a range of drugs23. In this context, it is interesting to note that publications indicate that individuals vaccinated against SARS-CoV-2 have lower viral loads and are less contagious25.The availability of a self-administrable nasal spray reducing subsequent viral transmission would have great impacts for the community as correlations between SARS-CoV-2 viral load and infectiousness have been shown10 6.85\u2009\u00b1\u20091.31\u00a0cp/mL, which was higher than more recently reported SARS-CoV-2 viral load values26. The higher viral load value may be explained with the dominance of the alpha (B.1.1.7) SARS-CoV-2 variant during the enrolment phase , which is known to infect the human nasal mucosa more efficiently than the wild-type and has been associated with higher viral load14. Indeed, the majority of the study subjects carried this variant. Whether the current data can be extrapolated to other SARS-CoV-2 variants needs to be investigated. Within this context it is important to point out that in vitro data indicate efficacy of azelastine against various SARS-CoV-2 variants tested10.Our study population was characterized by an initial mean viral load of log27.Upon treatment, a gradual decline of viral load from baseline (day 1) to day 11 of treatment was observed in all three study groups. This is similar to the natural SARS-CoV-2 clearance time of approximately 2\u00a0weeks. However, examples of prolonged nasal positivity have also been reported, and many factors are known to have an influence on the individual viral load and clearanceImportantly, the AUC analysis depicting the viral load decrease based on the detection of the ORF 1a/b gene over the 11-day treatment period showed a significantly greater reduction of virus load in the 0.1% azelastine group compared to placebo. Bearing in mind that viral load might be a surrogate measure of infectiousness, those results are encouraging as they indicate that azelastine may be a promising candidate for preventing the spread of this disease.30 underline the importance of analysis of this subset population. It would be desirable to extend the investigation of azelastine nasal spray as potential antiviral treatment with in vitro culture experiments.Interestingly, significantly greater decrease in viral load was shown on day 4 of treatment in patients with high viral burden (Ct\u2009<\u200925) treated with 0.1% azelastine compared to placebo, indicating that azelastine treatment may be advantageous for this patient population, particularly at an early timepoint of infection. Recent publications indicating that in vitro infectivity correlates with high virus concentrations (Ct\u2009\u2264\u200925) in nasal swabs31. Recently, Shmuel et al. reported that a low pH hypromellose nasal powder spray containing common components of nasal sprays could reduce SARS-CoV-2 infection rates19. However, a rinsing and diluting effect of the placebo formulation would have led to an underestimation of the effect of the use of the azelastine nasal spray.Of note, we cannot rule out the possibility that the placebo  contributed to viral clearance. In a study examining the effect of azelastine nasal spray on upper respiratory infections in children, it was found that the placebo group, receiving hypertonic saline solution (twice daily) also produced a favourable response compared to those receiving no treatment32.The current study demonstrated a gradual decrease of patients\u2019 symptoms and improvements of quality of life. Although no significant differences between groups regarding the total symptom score was shown, it may be speculated that the 0.1% azelastine spray may have positive influences on single symptoms such as \u201cshortness of breath\u201d, which was improved significantly greater in this treatment group compared to placebo at early time points of infection. It would be desirable to study azelastine treatment in a greater COVID-19 population to get further insights on azelastine\u2019s effects on individual symptoms and to determine its potential on long-term symptoms. Quality of life was assessed with the SF-36 questionnaire as no COVID-19 specific patient-reported outcome measures were available at the time of study. We are aware that this limited the capture of COVID-19 specific issues as questions were not specifically aimed for COVID-19 patients. It would be desirable to use a validated, COVID-19 specific questionnaire in future studies, and first attempts for its development are promising10\u2009\u2212\u20090.57 on day 11, which was significantly greater compared to placebo (p\u2009=\u20090.01)33. Comparably, differences in reduction of log10 viral load (cp/mL) in our study were\u2009\u2212\u20090.63 (ORF 1a/b gene) comparing treatment with 0.1% azelastine to placebo.Patients of the current trial were eligible upon positive PCR test results, and if enrolled no later than 48\u00a0h after swab sampling. Thus, it should be kept in mind that treatment started at a time point where the peak of viral load had probably passed. Although it may be expected that the azelastine might be most efficacious during very early time points after infection, its application in the current study setting could only be started during the symptomatic phase of the disease. Importantly, this scenario corresponds to current COVID-19 treatment regimens , which are usually started at\u2009\u2264\u20095\u20137\u00a0days upon start of symptoms but are still efficacious. Thus, antibody therapy (bamlanivimab and etesevimab) in positively tested, non-hospitalized patients demonstrated that treatment resulted in decreased SARS-CoV-2 viral load by log34.Importantly, newly emerging virus variants have the potential to evade the immune response, thereby affecting the efficacy of specific therapies and underlining the importance of new treatment strategies. This is exemplified by the emergence of the highly immune evasive omicron variant that is resistant to many monoclonal antibodies authorized for clinical useGenerally, treatment with azelastine appeared safe in SARS-CoV-2 positive patients: no serious adverse events were reported in the current study, and the number of adverse events was comparable between groups. Of note, the known bitter taste of azelastine was only negatively reported by a single patient, and compliance between treatment groups was comparable (mean\u2009\u00b1\u2009SD: 97 0.12\u2009\u00b1\u20099.7% compliance), thus indicating that the taste did not negatively influence treatment adherence.Our study showed both strengths and limitations. Thus, eligibility criteria were designed carefully to investigate a clearly defined, homogeneous study population of low-risk patients with a narrow age range. In addition, intervals between swab sampling were short and the overall number of performed PCR tests was high to allow a very close determination of the viral clearance. However, the overall small number of participants limits conclusions, and results should be interpreted with care. Of note, the mean viral load value showed small variability, thereby supporting the power of the current study.Overall, the current results are encouraging; however, further studies should be carried out to strengthen the findings, and treatment should be extended to other age and risk groups and cover individuals with different levels of symptom severity.35.Of note, pharmacometric analyses of our data indicate that more frequent applications of the nasal spray may be more appropriate for efficient treatmentBearing in mind the low number of participants in the current proof-of-concept study, the results still build a promising foundation for a currently running phase III study, during which effects of azelastine nasal spray on symptom severity and progression to severe COVID-19 disease are investigated in a greater patient population.Our study results provide the first human data showing that azelastine hydrochloride nasal spray used in a 0.1% concentration may be effective in accelerating the reduction of virus load in the nasal cavity and improving symptoms reported by COVID-19 patients. Future studies will help understanding the impact of azelastine hydrochloride in treating SARS-CoV-2 infected patients.Supplementary Information 1."}
+{"text": "Intravenous artesunate (AS) is the first-line treatment for patients with severe imported malaria (SIM) worldwide. However, after 10 years of use in France, AS hasn\u2019t yet received marketing authorization.The purpose of this study was to assess the real-life effectiveness and safety of AS in the treatment of SIM in two Hospitals in France.We performed a bicenter retrospective and observational study. All patients treated with AS for SIM between 2014 and 2018 and 2016\u20132020 were included. The effectiveness of AS was evaluated by parasite clearance, number of deaths, and the length of hospital stay. The real-life safety was assessed by related adverse events (AE) and monitoring of biological blood parameters during the hospital stay and follow-up period.110 patients were included during the six-year study period. 71.8% of patients were parasite-negative of their day 3 thick and thin blood smears after AS treatment. No patients discontinued AS due to an AE and no serious AE were declared. Two cases of delayed post-artesunate hemolysis occurred and required blood transfusions.This study highlights effectiveness and safety of AS in non-endemic areas. Administrative procedures must be accelerated in order to obtain full registration and facilitate access to AS in France. Plasmodium falciparum malaria cases continue to occur, so that in 2019 there were 5 540 imported malaria cases reported in France. Among these, severe forms represented 14.8% of imported cases . The demographic data are reported in Table\u00a0Plasmodium falciparum species malaria, two patients were treated for P. ovale malaria and one patient for P. vivax malaria, following a first reading identifying a P. falciparum. Each patient was then treated with the appropriate oral molecule following the identification of the species on second reading.The average time between the onset of symptoms and the return was 9 days . For 80% of the patients in this series, this was their first malaria episode, while for 21 patients (19.1%), a previous history of having had malaria could be documented. Despite AS having received institutional authorization exclusively for treatment of Almost three-quarters of the patients (71.8%) were parasite-negative of their day 3 blood smears after AS treatment. Data were missing for 13 patients. Of the 17 patients with a positive thick and thin blood smears at D3 (15.5%), 12 were parasite-negative on D7, two on D10, and3 remained without follow-up data. There were no documented early or late parasitological failures. One patient with four severity criteria at admission died within the first three days after hospital management. However, this patient suffered from severe hepatic impairment with a high risk of severe bleeding due to an active pancreatic carcinoma.The clinical parameters, severe malaria characteristics and treatment modalities are detailed in Table\u00a0Patient follow-up information was available for only one of the two centers. Of these 78 patients, 61 (78.2%) had follow-up consultations as part of their post-treatment monitoring with AS. These consultations were mostly scheduled between D7 and D28 and extended to D60 for two cases. For 44.3% of the patients there weretwo consultationson approximately D7 and D28, while for 23 patients (37.7%) there was only one consultation. Five patients did not attend the scheduled consultations and three were followed up in the context of their return abroad after hospitalization. Two hospitalized patients in wards with few malaria patients (nephrology and pediatric units) did not have a scheduled follow-up consultation.The adverse drug reactions are shown in Table\u00a0None of the patients discontinued AS due to an adverse event, and no serious adverse events, or cutaneous or injection site reaction were declared. Sixty-three patients (57.3%) presented a biological disorder during the 7-day follow-up of AS therapy, without a major clinical impact or requirement for readmission, except for two patients. The main biological adverse event was post-AS anemia. On day 7 after hospital admission, 44 patients had anemia according to the WHO guidelines, with mild anemia for 43.2% of them (WHO grade 1). Three patients suffered from severe anemia (WHO grade 3). Of these, half (24/44) already had anemia on hospital admission, with a mean decrease in the Hb level of \u2013 0.94\u00a0g/dL (SD: 2.8). On day 3 after admission and AS therapy, the Hb level decreased by an average of \u2013 2.17\u00a0g/dL (SD: 1.42). Two cases of delayed hemolysis were detected during outpatient follow-up. The first patient had a Hb level of 6.1\u00a0g/dL and lactate dehydrogenase (LDH) at five times normal on day 17 after treatment initiation, without a clinical impact except for mucocutaneous pallor . The parasitemia level was3%, and four AS doses were administered followed by an oral relay with artemether/lumefantrine (Novartis Pharma SAS). The patient\u2019s care for delayed hemolysis was readmission as an outpatientfor blood transfusion (two units of packed red blood cells). PADH was reported in the national pharmacovigilance database. The second patient presented with hyperparasitemia at 18% and exhibited four severity criteria: acidosis, hyperlactatemia, acute renal failure and jaundice. The baseline creatinine level at admission was 477 \u00b5mol/L. Six AS doses were administered with oral relay on day 5 with piperaquine/artenimol (Alfasigma SpA) (3 days). During the hospital stay, their Hb levels decreased from 8.9\u00a0g/dL at baseline to 6.7\u00a0g/dL on day 3 and reached a nadir of 5.3 on day 12. After blood transfusion, their Hb level stabilized at 7.4\u00a0g/dL and became 8.6\u00a0g/dL at discharge on day 23. At a follow-up visit at day 60, their Hb level was 12\u00a0g/dL, with slightly elevated LDH. The parasitemia was negative on day 3 and day 7. At the same time, the patient suffered from multi-organ failure within three days of admission, with an increase in creatinine to 611 \u00b5mol/L associated with encephalopathy (hyperuricemia). The patient started hemodialysis on day 5 for two days. At discharge (D18), their creatinine level was 147 \u00b5mol/L and decreased to 83 \u00b5mol/L at the 28-day follow-up consultation. This patient was the only one who exhibited elevated creatinine levels at discharge among the 18 patients with acute renal failure at admission. In total, two patients had elevated creatinine levels after AS therapy. One patient reported neuropathy of both feet that spontaneously resolved and could be attributed to oral antimalarial therapy. An abnormal potassium level was detected for 14 patients (12.8%) during AS treatment, mainly hyperkaliemia .p\u2009<\u20090.014, Fig.\u00a0p\u2009=\u20090.52) and no difference in terms of the parasitemia level (p\u2009=\u20090.71). An increase in the number of severity criteria corresponded with an increase in the number of AS doses administrered , but there was not a relationship with anemia secondary effects (p\u2009=\u20090.15) or hepatic transaminases levels (p\u2009=\u20090.28).Table\u00a0This retrospective, observational bicenter study presents additional data for six-years of real-life use of AS in two University Hospital centers in France. This work reports the solid effectiveness and reassuring safety data of intravenous AS in the treatment of severe malaria in non-endemic areas. No recrudescence was reported, and only two cases of PADH were recorded among the 110 patients of both participating centers.The efficacy of intravenous artesunate is well established in endemic areas and has shown superiority in antiparasitic efficacy and survival compared to quinine, thereby making AS a first-line treatment for severe malaria \u20135. DespiThe main arguments in favor of superior efficiency of AS are the benefit derived from reducing the parasite clearance time, particularly in hyperparasitemic patients >\u200910%), as well as the reduction in the length of stay in intensive care units (ICU) and hospital treatments in European patients %, as wel.Eder et al. with respectively 21\u00a0h and 20\u00a0h in favor of AS [Reduction of the parasitic clearance time has been described by Kurth et al. and or of AS , 23.Thisor of AS . The cosIn our study, no parasitic failure or recrudescence were documented. In its 5-year synthesis report of AS use, the French national reference center reported a mortality rate of 3% (28/919) and nine cases (1%) of parasitic failure including two recrudescence cases (at day 24 and day 28 of treatment). Failure was defined as persistent circulating trophozoites at day 7 .Only two patients had a documented PADH in this study, and no serious adverse reactions were reported. These results support the good tolerance of AS in real life and are in line with the data described in other studies. However, not all patients in our cohort were systematically followed up during the 4-week post-treatment period. It is, therefore, possible that these results suffer from a degree of bias due to under-reporting. Rolling et al. demonstrated a better tolerance of AS compared to quinine, with delayed hemolysis observed for 60% of patients as well as temporary deterioration of renal function (60%) while quinine caused at least one adverse reaction in 71% of patients. Quinine is known to havea number of adverse effects such as hypoglycemia, hearing disturbances, cardiotoxicity, and cinchonism . PADH isThe present study has several limitations. Adverse events might be underreported due to the retrospective review of the medical charts as well as due to data based on patient self-reporting of information. Moreover, some data may be incomplete due to the sometimes incomplete follow-up of discharged patients (follow-up at four weeks). As a result, not all patients were systematically screened and followed during the four weeks post-treatment with artesunate, implying a likely degree of underreporting of PADH. In our study, comorbidities and length of stay were not collected. Also, LDH levels were not collected for the PADH description. Finally, the retrospective nature of the study does not guarantee the completeness of the data collected despite the consultation of electronic medical records and some data were missing, mainly regarding follow-up after discharge.In high level of care areas, AS has gradually replaced quinine since 2011. Nevertheless differences in access to AS in Europe have led to disparate recommendations. Several studies have demonstrated the efficacy and safety of AS for severe malaria treatment with a faster ICU discharge rate and cost-effective therapy in high level of care areas . In the In conclusion, this work highlights the effectiveness and safety of intravenous AS in non-endemic areas. In non-endemic countries, AS suffers from slow development due to the lack of full WHO Good Manufacturing Practice (GMP) qualification as well as the lack of prospective randomized trials in travelers which are no longer possible for ethical reasons.It is, therefore, essential that the administrative procedures that have been underway for more than 10 years are accelerated in order to obtain full registration, which will facilitate access to AS in France.The current administrative burden for the initiation of treatment is not in line with the use of AS as an emergency and first-line treatment for severe imported malaria and as a cost-effective therapeutic strategy in high-care areas. The data regularly collected by the national reference center and the ANSM since 2011 should help accelerate this process. Finally, the recent overhaul in 2021 of the French system of treatments under compassionate access aimed at simplifying access to these treatments, particularly for manufacturers, is an opportunity to accelerate the procedures underway in order to obtain full approval and widespread access to intravenous artesunate in France."}
+{"text": "Causes of blackwater fever, a complication of malaria treatment, are not completely clear, and immune mechanisms might be involved. Clinical management is not standardized. We describe an episode of blackwater fever in a nonimmune 12-year-old girl in Italy who was treated with steroids, resulting in a rapid clinical resolution. Plasmodium falciparum parasitemia (26%).On May 9, 2022 (day 1), a 12-year-old girl was admitted to the pediatric emergency department of Modena Hospital, Modena, Italy, for persistent fever (>2 days) and lethargy. She had returned to Italy from a family excursion to Nigeria 11 days prior to admission. She did not take malaria prophylaxis. Her initial hospital evaluations revealed severe thrombocytopenia, increased total bilirubin, and lactate dehydrogenase . HemogloP. falciparum starting on day 4. Because the girl\u2019s hemoglobin levels had dropped steadily from the time of admission . Blood smears were negative for dmission , she recP. falciparum infection, and colleagues from a referral center for tropical diseases confirmed the diagnosis and recommended administration of steroids. We prescribed a 5-day treatment course of oral prednisone (1.3 mg/kg), starting on day 13. We tapered off the dose over the next 15 days and administered another blood transfusion on day 16. Symptoms cleared completely the day after steroid treatment began, and urine samples became normochromic 7 days later. The patient was discharged in good clinical condition on Day 23. One month later, blood test results were unremarkable (Our team suspected blackwater fever (BWF), a complication of markable . No abnoBWF is a condition characterized by massive hemolysis after treatment for acute malaria, with clinical symptoms that include hemoglobinuria, anemia, jaundice, and fever ; most important, hemoglobinuria is not reported. Another differentiating factor between the 2 conditions is that hemolysis due to artesunate is extravascular and, in BWF, hemolysis is intravascular (Plasmodium strains) , characteristics of the human host , and parasite type (e.g., different Treatment with steroids, as was determined for this patient, has been previously instituted in a malaria-endemic setting ("}
+{"text": "Plasmodium vivax malaria poses a potentially life-threatening risk of haemolysis in G6PD-deficient patients. Herein, we review five events of acute haemolytic anaemia following the administration of primaquine in four malaria trials from Indonesia, the Solomon Islands, and Vietnam. Five males aged 9 to 48 years were improperly classified as G6PD-normal by various screening procedures and included as subjects in trials of anti-relapse therapy with daily primaquine. Routine safety monitoring by physical examination, urine inspection, and blood haemoglobin (Hb) assessment were performed in all those trials. Early signs of acute haemolysis, i.e., dark urine and haemoglobin drop >20%, occurred only after day 3 and as late as day 8 of primaquine dosing. All patients were hospitalized and fully recovered, all but one following blood transfusion rescue. Hb nadir was 4.7 to 7.9 g/dL. Hospitalization was for 1 to 7 days. Hb levels returned to baseline values 3 to 10 days after transfusion. Failed G6PD screening procedures in these trials led G6PD-deficient patients to suffer harmful exposures to primaquine. The safe application of primaquine anti-relapse therapy requires G6PD screening and anticipation of its failure with a means of prompt detection and rescue from the typically abrupt haemolytic crisis.Primaquine for radical cure of  Plasmodium vivax is widespread in many regions of the world and is particularly prevalent in countries in South Asia, Southeast Asia, Oceania, and parts of South America [P. vivax blood-stage infections are due to relapses [ America . It is e America . Its abi America . Infecti America . It has relapses .Plasmodium vivax malaria may be cured only with the administration of 8-aminoquinoline chemotherapeutics, namely primaquine or tafenoquine [P. vivax patients of unknown G6PD status must choose between a therapy inviting the risk of acute haemolytic anaemia or withholding that therapy and inviting subsequent relapses and onward transmission. This clinical dilemma is mitigated by ascertaining the patient\u2019s G6PD status, thus protecting the G6PD-deficient minority from toxic exposure to 8-aminoquinoline therapy and protecting the G6PD-normal majority from ongoing latency and often multiple attacks of malaria.Latency in enoquine ,5. This enoquine ; this isenoquine . In Indoenoquine . In Vietenoquine . In the enoquine . HealthcP. vivax malaria occurs. Point-of-care G6PD testing technologies aimed at solving this serious problem of access have recently emerged [Plasmodium vivax [Screening for G6PD deficiency in most malaria-endemic settings has proven impractical until very recently . The gol emerged ,13. The  emerged . The WHOum vivax .P. vivax malaria were administered varied regimens of primaquine in combination with varied blood schizontocidal therapies as a radical cure of latent malaria in order to prevent relapses. All subjects were screened for G6PD deficiency prior to enrolment using varied screening kits and technologies. All subjects were misclassified as G6PD-normal and experienced onset of severe acute haemolytic anaemia 4 to 8 days after initiation of primaquine therapy. We describe here the clinical course of these haemolytic crises, treatment, and recovery.In this report, we highlight the risk of primaquine-induced acute haemolytic anaemia despite access to G6PD screening, even in the rigorous setting of controlled clinical trials. This retrospective case series reviews five severe adverse haemolytic events that occurred in four different randomized controlled trials (RCT) conducted in Indonesia , the Solomon Islands , and Vietnam  between 2012 and 2022. The authors of this report are the investigators of those trials. In all those trials, subjects with confirmed or suspected patent P. vivax malaria and primaquine therapy against latency in North Sumatra, Indonesia (reg no. NCT04223674). A total of 1133 participants aged 6\u201315 years old were enrolled from February 2022\u2013May 2023. Serological screening was performed to predict recent P. vivax exposure, which may indicate the risk of harbouring hypnozoites for which primaquine was given as a radical cure to prevent future relapse. These participants were recruited in a school-based mass blood survey. Physical examination by the study physician and capillary blood collection from finger prick was done onsite, whereas Hb and G6PD screening was performed at the nearby site laboratory (1\u20132 km distance). Blood samples were transported (at 4\u20138 \u00b0C) within 4 h of collection. Hb was read using HemoCue , and G6PD activity (in U/g Hb) was measured using a point-of-care quantitative G6PD testing platform  [The subject was a 9-year-old boy enrolled in a randomized controlled trial involving serological screening for exposure to The subject was a local resident screened for eligibility on the 22nd of February 2022. He was asymptomatic with a methaemoglobin (metHb) of 0.4% by non-invasive oximeter . His Hb was 12.0 g/dL, and G6PD was recorded as 6.3 U/g Hb. Treatment commenced a week following enrolment. The regimen consisted of three daily doses of dihydroartemisinin and piperaquine phosphate  and a daily dose of primaquine  scheduled for 7 days. The subject (body weight 21.1 kg) was given 1.5 tablets of DHP and 1.25 tablets of PQ  and a change in urine colour (Hillmen 4). The subject did not meet protocol criteria for treatment cessation , and a fourth dose of PQ was administered.On the following day (day 5), the subject\u2019s Hb had dropped by 30.8% from baseline (from 12 g/dL to 8.3 g/dL), and his urine showed visible signs of haemolysis  of packed red cells starting on the first day of hospitalization. He remained in the hospital for four days, receiving supportive therapy, including supplemental oxygen and fresh frozen plasma due to a brief episode of upper gastrointestinal bleeding. He was discharged in good condition with a Hb level of 13.1 g/dL.G6PD genotyping conducted the day after discharge  revealedA confirmed G6PD-deficient subject had been misclassified as normal. This likely occurred as a result of error in managing the sample in the laboratory during en masse G6PD screening. Investigation had shown the instrument used for screening to be in proper function and use by the operators. The clinical research team responded to the event and findings by implementing a second G6PD test at the point-of-care immediately prior to initiating PQ administration. No further similar events occurred.Plasmodium vivax malaria was enrolled in a trial assessing the safety and efficacy of two primaquine dosing regimens (Artemether-Lumefantrin plus Primaquine vs. Dihydroartemisinin-piperaquine plus Primaquine) in the Solomon Islands (reg. no. ANZCTR 12617000329369). The trial was conducted from September 2017 to August 2019. A total of 629 individuals with P. vivax were screened, and 384 were enrolled. Ten of the excluded participants were identified as G6PD-deficient. Participants were recruited mainly by way of referral to the study team by clinical staff working in the local health facilities  or through active case detection (ACD) in the field. After informed consent was obtained, both CareStart\u2122 Malaria and CareStart\u2122 G6PD RDTs (RDT-1) were performed to confirm participant eligibility according to presence of P. vivax malaria and absence of G6PD deficiency, respectively. Baseline demographics and clinical characteristics were then collected, ensuring inclusion criteria were met. Participants were then transferred to the central study site for repeat G6PD screening with both CareStart\u2122 and Binax Now\u2122 RDTs (RDT-2 and RDT-3). Both RDT-2 and RDT-3 were performed under temperature-controlled conditions in line with the manufacturer\u2019s recommendations. All three RDT results were checked and verified by a second operator at the time of testing. Only participants identified as G6PD-normal on all three RDT tests were enrolled.A 48-year-old man with The subject was enrolled on the 2nd of November 2017 after being cleared for inclusion. His body weight was 54 kg, his metHb by pulse oximetry  was 0.0%, and Hb estimation by HemoCue  was 10.1 g/dL. He was randomized to receive primaquine  at a dose of 15 mg per day  at a dose of 11.25 mg per day . By the time both cases occurred, 72 participants had been enrolled in this trial. The subject was a 16-year-old male with mixed Upon admission, his blood was taken for further laboratory testing. His blood slide was negative for malaria, and his Hb was 6.7 g/dL (moderate anaemia). He received one unit of red cells and ferrous sulphate supplementation and was discharged the next day (day 9).The subject was subsequently reviewed on day 10, when his Hb was measured at 7.9 g/dL according to HemoCue. His respiratory rate was 20 breaths/min, and his heart rate was 62 beats/min. His metHb was 1.6%. By day 12, his Hb has increased to 10.3 g/dL with normal vital signs , and metHb of 0.7%. He appeared well and had resumed daily regular activities.Further quantitative testing with OSMMR2000-D G6PD Kit  demonstrated G6PD activity of 1.45 U/g Hb (WHO Class B ). SequenP. vivax . A total of 2336 participants from four countries  were enrolled during the period of July 2014\u2013November 2017 [A 20-year-old man was enrolled in a multicentre clinical trial to assess the noninferiority of a short duration of high-dose primaquine (7 days of 1 mg/kg/d vs. 14 days of 0.5 mg/kg/d) as a radical cure for P. vivax by light microscopy and had no other health issues. His Hb was 15.3 g/dL with normal G6PD screening results recorded. He was enrolled on the 16th of November 2014 and randomized to receive 7 days of daily 1 mg/kg BW primaquine  was positive for On day 2, after receiving his first dose of primaquine (1 mg/kg), his urine Hillmen increased two levels from baseline (2 to 4), but this was not accompanied by any complaint or physical abnormalities. On day 3, however, after having received two doses of primaquine (2 mg/kg), his Hb had dropped 21% from baseline to 12.1 g/dL. No further change was observed in his urine colour, and a third PQ dose was administered. On day 4, his Hb further decreased to 11.5 g/dL, but because he was asymptomatic with a stable Hillmen score at 4, he was given another PQ dose. On day 5, his Hb fell to 9.7 g/dL (37% drop from baseline), but his urine colour at Hillmen 4\u20135 remained stable. His temperature was 37.5 \u00b0C with a heart rate of 84 beats/min and a blood pressure of 110/70 mm Hg. The patient was given a fifth PQ dose despite the presence of cessation criteria . Clinical signs of acute haemolysis started to be noticeable on the following day (day 6), when his Hb level decreased to 7.6 g/dL, a 50% drop from baseline value; he was pale and slightly jaundiced with dark urine (Hillmen 5). He complained of abdominal pain and was feverish (37.6 \u00b0C) and tachycardic (pulse = 96 beats/min). His blood pressure was 120/80 mm Hg, and his respiratory rate was 24 breaths/min. Details on his clinical progression from baseline to the onset of the haemolytic episode are described in On the following day (day 8), his Hb decreased further to 6.6 g/dL. He had no fever, and his urine colour had lightened (Hillmen 3\u20134). His Hb dropped slightly further to 6.4 g/dL the day after (day 9), as shown in \u00ae Hb 201+ . First, 3 mL of venous blood was collected in EDTA tubes, stored temporarily in a cooler box, and transported to a site laboratory located in a town 69 km away within 4 h of collection. The laboratory performed G6PD FST . The results were conveyed to the field team within 24 h via mobile phone text message. A total of 25 of the 231 screened participants were identified as G6PD-deficient and excluded.The subject was a local 13-year-old boy enrolled in a trial assessing the efficacy of spatial repellent to reduce malaria transmission in Southwest Sumba, Indonesia during September 2012\u2013April 2013 . In thisThe boy was screened on 18 October 2012. His weight was 29 kg, and his Hb was 11.3 g/dL. His G6PD status was recorded as normal, and he was enrolled in the study. Therapy with standard dihydroartemisinin-piperaquine (DHP) was initiated immediately, followed by primaquine the next day. The regimen for DHP was two tablets for three days, and the primaquine dose was 0.6 mg/kg  for 14 days. On day 3 after receiving two doses of primaquine (approximately 1.2 mg/kg), he complained of nausea with epigastric pain but did not appear anaemic or jaundiced. His Hb was not measured, and urine colour was not checked. A third dose of primaquine was given . On the following day, he complained again of nausea and fever, but his temperature on examination was 36.4 \u00b0C. The site physician then gave him ranitidine and metoclopramide. As the complaints subsided, a fourth dose of primaquine was administered . The next day, the patient visited the clinic with nausea and vomiting. He was also pale, icteric, and tachycardic. His spleen was enlarged on physical examination. His urine was dark, and his Hb level was 7.2 g/dL. His clinical progression and Hb drop are summarized in At the hospital (6 h after cessation of primaquine), his Hb dropped further to 5.6 g/dL  after seemingly testing G6PD-normal with qualitative or quantitative screening tests. All events occurred in the setting of rigorous clinical trial processes and procedures, including well-defined ascertainment strategies, careful screening protocols, close monitoring, and well-prepared mitigation plans in the event of inadvertent treatment of G6PD-deficient subjects due to screening failure. All of these patients fully recovered, four of them after blood transfusion therapy.The five cases described here document the clinical implications of what clearly represents failures of screening processes intended to protect G6PD-deficient patients from exposure to 8-aminoquinoline therapy. The precise reasons underlying each of these failures are difficult to discern. However, it is notable that they occurred in four separate clinical trials in three different countries. All employed high standards of clinical practice and oversight, carefully complying with protocols designed to minimize haemolytic risk. It is particularly important to note that the four trials utilized a variety of testing platforms, with a total of five different tests used including the SD biosensor (Case 1), CareStart\u2122 RDT (Cases 2\u20134), Binax now RDT (Cases 2 and 3), FST R&D Diagnostics (Case 4), and FST Trinity Biotech qualitative test (Case 5). This points to two important possible factors at play here. Firstly, no G6PD field test can be assumed to have 100% sensitivity  even under ideal laboratory operating conditions ,20,21,22The cardinal laboratory sign of acute haemolytic anaemia is a \u201csteep\u201d drop in measured Hb levels in blood that may be accompanied by symptoms of anaemia and/or signs like dark urine, pale conjunctivae, and yellow sclera. In three cases, these signs were not detected early enough to prompt a decision to stop PQ, whilst in the other two patients, the clinical course may have been different if PQ had been stopped earlier, as mandated by the protocol (Case 4), or had extra vigilance been exercised by re-assessing the Hb concentration and urine colour (Case 5). In the four cases with available daily Hb data, the course of haemolysis appeared only slightly after the first two doses; Hb dropped only by 10\u201320%, well within the normal range for a patient undergoing antimalarial therapy . ExceptiClinical features in the days leading up to the haemolytic crisis were not specific, as the common symptoms (nausea and fever) often occur in the days following treatment for acute malaria, and abdominal discomfort is a common complaint from patients consuming PQ anti-relapse therapy, more so with chloroquine compared to dihydroartemisinin piperaquine . The MetThe failure to detect haemolysis early will result in continuing haemolytic crisis with continued exposure to PQ dosing. This may be particularly dangerous when the G6PD deficiency variant involved is of a severely impaired enzyme activity phenotype. All of the four known variants involved in five cases in this series\u2014Coimbra, Union, Viangchan, and Vanua Lava\u2014were of that phenotype. Under the new WHO classification of G6PD variants, these are all Class B, with residual enzymatic activity of <45% of normal . CoimbraIn two of the cases reported here (Cases 1 and 4), the haemolytic crisis did not halt with the cessation of dosing but continued for the 2\u20133 days leading up to hospitalization and transfusion. This has been observed by others ,30,31. IThe haemolytic events reported here highlight the danger daily primaquine poses to patients having G6PD deficiency; this may be equally or more challenging with single-dose tafenoquine anti-relapse therapy, a drug having a mean half-life of ~14 days  and thusThe experiences reported here demonstrate the importance of escalated vigilance (follow-up and patient education), particularly during days 3\u20137 of PQ administration, even when G6PD screening is implemented. This principle is exemplified in Case 4, where the decision to continue with primaquine despite a notable decline in haemoglobin levels was underscored by the patient\u2019s apparently healthy appearance and stable Hillmen colour. In retrospect, primaquine should have been stopped earlier, even if the declining haemoglobin level was consistent with malaria. Close monitoring, interrupting primaquine administration upon any signs of haemolysis, and resuming treatment only when considered safe emerge as pivotal measures when administering primaquine. More research is needed to identify early markers that may predict with confidence the early onset of a severe haemolytic crisis with continued primaquine exposure, e.g., clinical symptoms with increasing Hillmen/urine urobilinogen. The onset of dark urine was clearly a late sign in our series but has also been seen with accidental primaquine overdose and in favism, in which the acute development of dark urine was associated with acute catastrophic falls in haemoglobin ,36,37.Primaquine poses the potential danger of severe haemolysis in individuals with G6PD deficiency. The drug should be given with extra vigilance, especially during days 3\u20137 of treatment. Screening for G6PD deficiency is imperative before initiating the treatment, and there should be a robust plan in place to promptly rescue patients from a haemolytic crisis, should it arise."}
+{"text": "P. vivax prevalence following a TPT campaign. Among a targeted population of 2322; 1973 (85.0%) participated in the baseline mass blood survey (MBS) and only 52.0% of the total targeted population  completed the TPT. G6PD deficiency was found among 13.5% of total MBS participants and those were excluded from TPT. Of 1315 eligible samples, farmers and gold miners, males, and those aged 15 to 45\u00a0years had higher percentages of non-participation in TPT. Qualitative findings showed that most of the non-participation groups were outside the villages during TPT because of time-sensitive agricultural and other occupational or education-related purposes. In addition to mitigating of some inclusion criteria , strengthening community awareness and increasing engagement should be pursued to increase community participation.Targeted mass primaquine treatment (TPT) might be an effective intervention to facilitate elimination of vivax malaria in Myanmar by 2030. In this study, we explored the factors hindering coverage of a TPT campaign conducted in a malarious township of northern Myanmar. From August 2019 to July 2020, a cross-sectional exploratory design including quantitative and qualitative data was conducted in five villages with high  There have been remarkable improvements in the malaria burden in Myanmar over the last decade since the country committed to the goal of being malaria-free by 20302. Still, in 2020 there were 58,836 confirmed malaria cases and ten reported malaria deaths in Myanmar1. The trend suggested an 88.3% morbidity reduction compared to 2010 data1. The species composition has likewise been shifting during this same time period. In 2011, P. vivax accounted for only 46.0% of all infections but by 2020 it composed 74.2% of all cases1. The reported P. vivax cases likely include many relapse cases, either because of poor compliance with a 14-day primaquine (PQ) treatment or lack of PQ administration in case of contraindication (e.g. pregnant women)3. Relapse cases need to be addressed as they can lead to transmission and while most elimination efforts have focused on falciparum malaria, it will be necessary to focus on P. vivax as well in order to achieve malaria elimination2. The existence of latent P. vivax reservoirs therefore poses a threat hindering the achievement of malaria elimination5.Malaria remains a priority public health concern in the Southeast Asia (SEA) Region. The region has the second highest malaria burden in the world, following Africa. Among 11 member countries from SEA, Indonesia, India and Myanmar are the top three most malaria endemic countries6 were endorsed in Myanmar in 2015 and have been regularly updated according to the latest global guidelines. The NMTG provide detailed information for using 14-day PQ (0.25\u00a0mg/kg bodyweight) as radical cure for P. vivax infections. Worldwide, PQ is widely used, and several studies have shown its efficacy9. Previous studies have suggested that many reported P. vivax cases were relapses rather than new infections12. The latent stage hypnozoites in the human liver can reactivate, leading to clinical relapse weeks or months after getting the infection13. Radical cure with PQ can clear hypnozoites6, however the long duration of the 14-day regimen has been difficult with regard to adherence14. Shorter courses of PQ, or new hypnozoitocidal drugs like tafenoquine that only require a single dose, might lead to more effective outcomes but must also be balanced with the dangers of hemolysis in G6PD deficient patients 15. G6PD deficiency is an inherited, X-linked trait. A recent study in western Myanmar16 reported an approximate 10.0% prevalence of G6PD deficiency among malaria infected individuals. While there is diversity in G6PD variants, the most common variant from Myanmar has been the Mahidol variant or 1311\u00a0T/93C haplotype17. Currently the WHO recommends the 14-day PQ regimen18 and this recommendation has been reflected in the NMTG. Meanwhile, a straightforward tool that can detect the presence of hypnozoites remains unavailable, G6PD deficiency testing remains difficult in normal field settings, G6PD deficiency is high in some malarious settings, and adherence to the full 14-day regimen of PQ is a continuing challenge.The National Malaria Treatment Guidelines (NMTG)P. vivax parasites20. Targeted mass primaquine treatment (TPT) has been successful in eliminating temperate-zone P. vivax in China21. Studies on TPT to eliminate P. vivax in China21 and the Democratic People\u2019s Republic of Korea22 pointed out the importance of achieving high coverage and being aware of a greater risk of hemolysis among patients with G6PD deficiency. Still, the feasibility of eliminating P. vivax from tropical and subtropical areas like the Greater Mekong Subregion (GMS), where G6PD deficiency is highly prevalent, has not been evaluated23. Moreover, the current NMTG6 also lack information about the implementation of TPT activities. Therefore, a TPT study was conducted in a township in Myanmar from 2019 to 2020 to assess the feasibility of using this elimination focused tool in this setting24. The TPT campaign included mass administration of PQ targeted specifically to people without G6PD deficiency and living in P. vivax high burden villages24.Large-scale mass primaquine treatment is one approach to address reservoirs of latent 4 described the use of active surveillance through mass blood surveys (MBS) in active malaria foci to determine the point prevalence at a time or to fill the gap of the routine passive case detection coverage. Furthermore, MBS should be paired with TPT activities to evaluate potential changes in malaria prevalence before and after TPT22. Blood tests including G6PD investigations and monitoring Hb% must also be done prior to and during the mass treatment activities.The National Malaria Strategic Plan19. Historically, mass drug administration campaigns with low coverage have shown minimal effectiveness. Many campaigns seek to achieve a coverage of at least 80.0%, especially for falciparum malaria but also for vivax25. Particularly with regard to TPT for P. vivax, contraindications to PQ and general compliance with the full 14-day antimalarial regime are major challenges to achieving high population coverage22. Next, it is also important to address the challenges as well as the needs of the community to participate in a mass treatment activity. Therefore, this study aimed to document the factors hindering people's participation in a recent targeted mass treatment trial with primaquine in northern Myanmar.The relative success of mass drug administration is dependent on several factors, including transmission level, the antimalarial used, and coverage of the targeted population (linked to community participation)P. vivax endemic township in northern Myanmar from August 2019 to July 2020.The study incorporated a cross-sectional, mixed methods design including both quantitative and qualitative data collection among people living in a 26. Malaria diagnosis and treatment services are provided free of charge through the government basic health staff and village malaria volunteers. Currently, patients with P. vivax are treated according to the NMTG6 without G6PD testing and using Directly Observed Treatment (DOT).Myanmar is composed of 14 states and regions and one union territory. Among them, Sagaing is one of the highest malaria burden regions in northern Myanmar. Of 34 total townships in Sagaing, Banmauk township  team. Then from these ten villages, five villages  were randomly selected and targeted for a study on mass treatment with PQ . Conversely, children who were\u2009<\u20097\u00a0years old, pregnant women or breastfeeding mothers, people who were G6PD deficient, and people with Hb levels less than 8\u00a0g/dl were all excluded from TPT. During MBS, malaria microscopy as well as blood spots on filter papers for PCR analysis were also collected to detect malaria parasites. Study participants were included in the study regardless of their infection status. Those who were found to be malaria positive were prescribed schizonticidal drugs followed by the TPT regimen.27, the G6PD kit showed almost 95.0% sensitivity and 90.0% specificity. However, to cover the possible false negative results by RDT, all the participants were crosschecked for the presence of dark colored urine during 14-day PQ course by the research team. No participants reported presence of dark colored urine during the 14-day PQ treatment.For G6PD testing, the CareStart G6PD RDT was used. It is a qualitative point-of-care visual screening test that identifies G6PD deficient patients using a whole blood sample. It uses the visual dye-colorization assay method and has a result time of 10\u00a0min. According to a test performance evaluation in MyanmarThose eligible for TPT were followed up with a checklist by health officers to monitor the completeness of DOT and to monitor possible side effects of the daily PQ regimen. To find out the reasons for not being present in the villages during TPT, a simple question was asked to other family members about why those missed samples were away from the village.After the TPT operation, qualitative data were collected through face-to-face in-depth interviews using interview guidelines. The guideline covered whether respondents knew about the implementation of TPT in the villages and the main reasons for not being included in the treatment campaign entirely or partially. The qualitative data also covered the reasons for not participating in the TPT. People from the five study villages were categorized into TPT and non-TPT groups. Individuals who completed the 14-day PQ regime were defined as the TPT group and those who were totally absent from any TPT activity including the initial MBS were defined as the non-MBS group. Those who were involved in MBS and were eligible for the TPT but later did not receive the TPT, regardless of the reason, were defined as the non-TPT group. In this study, there were only a few people who did not fully comply with the treatment or complete TPT. They were excluded from the qualitative study to facilitate the representativeness of study results. The qualitative assessment was conducted only among the non-MBS and non-TPT groups.Inclusion criteria for the qualitative part included being older than 18\u00a0years of age, and being able to communicate. We also attempted to ensure that participants were not under the influence of any narcotics, based on physical appearances and through informal questions. Nevertheless, no one was excluded due to that reason. Balance in gender, age distribution, occupation and proportionate samples from each village were also considered. A total of 30 individuals were included in the qualitative component of this research.The data collection team was comprised of five members with university degrees who were trained by the researchers on data collection as well as ethical precautions for both quantitative and qualitative data collection. The team visited each household and collected census data from the household head or a representative. During in-depth interviews, one data collector would be the interviewee and the other would act as a note taker. The transcripts were written in the Burmese language. Each in-depth interview session lasts around 20\u00a0min.The baseline population census was entered into a Microsoft Excel  spreadsheet. Information regarding participation status, blood results and reasons for not being involved in TPT activity were recorded for each person. The charts were produced directly from the excel sheet. The data were then analyzed using the Statistical Package for the Social Sciences .The quantitative component of this work began with a cross sectional survey that included a questionnaire and basic clinical assessments. The survey included data on age, gender, ethnicity, pregnancy and/or breastfeeding status, G6PD status, and mean hemoglobin levels. Exploratory statistical analyses were done to assess the characteristics of individuals who had G6PD deficiency and of those who did not participate in the study. Finally, a statistical analysis was done among all the eligible samples (n\u2009=\u20091315) to be involved in the TPT to explore sociodemographic factors relating to non-participation in the TPT. In this study, non-participation includes those who did not participate at all, or who only participated partially in the TPT, starting from initial blood screening to the completion of 14\u00a0days of PQ. Crude and adjusted odds ratios were reported together with 95% confidence intervals and p-values. All collected variables were entered into simple and multiple logistic regression models regardless of their statistical significance.For the qualitative data, Burmese dialogs were entered in a Microsoft Excel file representing each respondent with an identity code and some general characteristics such as age, gender, and occupation. Each quote was then translated into English independently by PLA and MTS. KMW and MPK have ensured the accuracy and correctness of the translation. KO and DMP improved the English language and grammar. Finalized quotations were grouped under two situations: either non-participation in MBS or non-participation in TPT. The lead authors chose the most appropriate quotes, and other authors re-checked and agreed with the final ones.The Institutional Review Boards of the University of Public Health, Yangon, Myanmar (UPH-IRB: 2020/ IR Research/ 2) and the University of South Florida, USA, reviewed and approved the study protocol. All methods were performed in accordance with the relevant guidelines and regulations. The respondents signed the informed consent forms before commencing the interviews and informed consent forms were obtained from all participants.When implementing an initial mass blood survey (MBS), 349 individuals were away from the villages and missed the survey. Another 658 villagers were excluded because of the exclusion criteria, including 294 children who were\u2009<\u20097\u00a0years old, 51 women who were either pregnant or breastfeeding, 266 people who were G6PD deficient, and another 68 with Hb levels lower than 8\u00a0g/dl. The cumulative number was 658 because some individuals possessed both criteria . This left a total of 1315 villagers who were eligible to be included in TPT. At the actual time of administration, a further 86 villagers were absent from the village and 21 individuals began but did not complete the radical cure regimen, leaving 1208 individuals who completed TPT. Overall, these final samples represented 52.0% of a total of 2322 villagers in the study villages, which was lower than the target set of overall 80.0% coverage. Still, it reached 91.9% of a total of 1,315 who were eligible to participate , 1973 (85.0%) were tested for G6PD status. Of 1973 samples, 266 (13.5%) were diagnosed as G6PD deficient. Their general characteristics were analyzed to find out possible associations with the presence of G6PD deficiency. Kadu, Kanan and Shan ethnic males living in ManKat, PintSinTe and NantHnyin villages were more likely to be G6PD deficient  were more prone to test positive for G6PD deficiency. Individuals from Kadu , Kanan  and Shan  ethnic groups showed higher odds of being diagnosed with G6PD deficiency as well. There were also differences in G6PD deficiency prevalence among study villages. People residing in ManKat , PintSinTe , and NantHnyin  villages had high prevalence of G6PD deficiency (Table ).A total of 435  people were away from the villages during the TPT campaign. Most of them were male (64.4%), aged 15 to 29\u00a0years (47.4%), farmers (46.9%), annual family income was\u2009<\u20091,000,000MMK (61.6%), and those who had only completed primary (35.2%) or middle school (42.3%) levels. When they were asked for reasons for not being included in the TPT, more than one-third (43.0%) said they had stayed in farms, and a few (20.9%) mentioned that they lived in gold\u00a0mine areas , headache , epigastric pain symptoms , palpitation  and the last six cases (0.5%) revealing nausea and vomiting. To monitor for hemolysis, in addition, all participants were strictly followed up for dark colored urination. However, no one revealed presence of dark colored urine during the 14-day PQ treatment Fig. .The logistic regression indicated that three variables were significantly associated with non-participation in the TPT. First, participants in the 15 to 44\u00a0year old age range showed higher odds of not participating in the TPT when compared to the younger age group  and  for age groups (15\u201329\u00a0years) and (30\u201344\u00a0years), respectively. Second, males  were more likely to not participate in the TPT. Third, farmers  and gold miners  possessed the highest odds of non-participation when compared to students. Annual family incomes and level of education were not associated with participation in the TPT in our analyses .\u201cMy family completed all treatment except two of my children who missed receiving the blood test and taking the drug because they lived in Banmauk township to attend grade-9 and -10 classes. I informed them about the TPT campaign during a monthly visit to their schools, but their return to the village was impossible.\u201d .\u201cWe have heard about giving anti-malarial treatment to all people in our villages. However, most of our family could not return to the village as the time coincided with our cultivation period and when the roads were terrible.\u201d .\u201cIn my family, 4 out of 6 members joined this mass treatment campaign. The other two (my husband and elder son) went to another township to seek new job opportunities to solve our family\u2019s current financial difficulties.\u201d .\u201cI just came back from the goldmine 2\u00a0days ago. I have been notified that there was a mass treatment activity in this village. However, I could not return to the village to participate in the activity because of my responsibility for ongoing mining activities.\u201d .Although many villagers were aware that the TPT campaigns had health benefits at both individual and community-levels, they did not participate in the campaign. Most people who missed the initial blood screening reported being absent because they traveled to worksites such as farms and goldmines to find a better income for their families (many were heads of their respective households). In addition, a few high school students could not participate in the TPT because they had to live and study at their high schools away from their villages.\u201cI already had my blood tested (for G6PD deficiency) and was told I am eligible to receive antimalarial treatment. However, I had to visit my native town during the proposed days when the campaign was occurring. Hence, I did not get any treatment. If there is another chance, I am sure I will participate.\u201d .\u201cI had a chance to participate in the first mass blood survey organized by the research team. I was also informed of the drug treatment schedule. But the proposed time frame coincides with our cultivation time, and I have missed a chance to get the treatment, unfortunately.\u201d .\u201cI was informed that the malaria team will provide me 14-days of primaquine treatment under their observation as I am eligible to participate; however, while waiting, the manager called me to come into a gold mine. Therefore, I did not receive the treatment as I am afraid of adverse consequences of incomplete treatment.\u201d .\u201cTo be honest, I am interested in this mass treatment activity as this is the first activity ever to reduce the malaria burden in our village without any charges. However, I have to work in a gold mine for my family's income, and I missed the opportunity to participate in it.\u201d .Some people were eligible to receive the treatment, but they had to work in forested areas far from the village. Some expressed that they will indeed participate once there is another round of mass treatment. Many people know the potential advantages of TPT, but they were afraid of the consequences if they could not adhere to the full treatment course. Most people also know the TPT program is free of charge.28, but require high levels of community participation in order to achieve the population level benefits . Achieving high levels of community participation can be challenging for many reasons. In this analysis, we identify innate un-modifiable community characteristics that lead to non-participation, as well as other characteristics that may be addressed either through community engagement or through adaptations by the TPT implementing teams.Interventions such as mass administration of antimalarials may provide both individual and population-level benefits29, and were therefore excluded from this study. The evidence base with regard to use of PQ in pregnancy and while breastfeeding is scant. Testing for G6PD deficiency status for in utero fetuses is difficult, invasive, and rarely done and PQ administration for pregnant women (regardless of their G6PD status) is often contraindicated30. Some recent evidence suggests that PQ levels in breastmilk are quite low, but PQ administration in lactating women is still rare31. Finally, in this study children\u2009<\u20097\u00a0years were excluded from the targeted treatment with PQ based on recommendations from collaborators at Mahidol University and following Thai National guidelines for trials. Thus, almost one-third of all people who participated in the blood screening in these study villages were excluded due to having one or more of these exclusion criteria. Mitigation of some inclusion and exclusion criteria (e.g. expanding the age range to less than 1\u00a0year to be included in the TPT) might lead to increased coverage of TPT.Individuals with G6PD deficiency may experience severe side effects if they take PQ33. However, currently, 14-day PQ has been prescribed by healthcare providers including basic health staff and volunteers without G6PD testing6. Patients with P. vivax are instructed to regularly check their urine color during the treatment and if there is dark-colored urine or any unusual anemic symptoms, they should come back to the health facility6. In Myanmar, people living in malaria-endemic areas with a history of taking PQ have rarely reported any hemolysis features.G6PD deficiency is not a modifiable attribute, and in this study we found that the proportions of G6PD deficient individuals varied by study village, by ethnicity, and by gender. This is an important non-modifiable attribute that should be considered when planning mass administration of PQ, as a very high proportion of G6PD deficiency could make it impossible to achieve targeted coverage of PQ in a given community. The reported G6PD deficiency in Myanmar ranges between 10.0% to 20.0%6. Mass treatment with a shorter regimen (e.g. 7\u00a0days of treatment) or the 8-week PQ regimen (in case of mild or moderate G6PD deficiency) may also avoid loss of compliance or increase coverage35. Given that Shan, Kadu, and Kanan ethnic males had a higher risk of G6PD deficiency, they should be closely monitored with access to regular hemoglobin checks and emergency medical care in upcoming TPT operations.The G6PD-normal TPT participants in this study also reported no severe side effects of PQ during the treatment. G6PD screening by qualitative or quantitative methods is recommended in future trials. Based on the quantitative results, including non-severe and severe forms of G6PD deficiency in the TPT could be considered with different treatment regimens, which is also in line with the current NMTG24 done in this setting. Therefore, healthcare support and social development programs should come together in later trials to facilitate the involvement of working-age individuals. For example, mobile healthcare workers might be able to administer the treatment at farms or other work sites. Delivering DOT by family members rather than healthcare providers might be an alternative for villagers who usually work in the daytime and come back home at nighttime36.There were several other factors that were associated with non-adherence in this study, many of which could plausibly be addressed through community engagement or through modifications among the TPT teams. For example, working-age people showed a higher chance of non-participation in TPT. In the study villages, family incomes have mainly relied on agricultural-related or gold mining activities usually employed by working age groups. This group of people is responsible for their respective family's economic conditions and several reported needing to work rather than participate in healthcare activities including TPT. This finding is in line with a previous study38. Therefore, targeted health education should be delivered to the male group to persuade their interest to participate in future TPT campaigns, along with potential modifications to the actual administration of PQ.In Myanmar culture, males are often accountable for family matters especially with regard to income. Work needs often conflict with health awareness-raising activities. Some studies have documented poor levels of malaria knowledge and practices among males when compared to females39. Typically, they cultivate their nearby farms around the villages once the rains begin, which often corresponds with the malaria season. Many of them work in rural and remote areas, and roads and other infrastructure need to be developed across the country. While there have been improvements over the last decade, the country is in the developing stage amid an economic crisis impacted by COVID-19 pandemic and political affairs. Therefore, during the rainy season many people have to stay in farming areas as road conditions have deteriorated and travel is difficult. Consequently, farmers are often outside the purview of healthcare coverage, which normally focuses on villages and communities, and they experience a high risk of malaria infection especially during the rainy season when malaria transmission is high40.Likewise, Myanmar is an agricultural country and many people living in the nation earn their living from farm-related activities41. Implementing TPT activities inside or nearby goldmines through malaria volunteers might lead to good TPT coverage. Otherwise, these groups of missing people might serve as an ongoing reservoir and source of malaria transmission. A study22 concluded that one of the reasons hampering the attainment of full coverage for mass treatment was migration of the targeted participants. There are a few published articles on TPT activities solely targeting forest-related workers and this area of work should be further pursued. Antimalarial chemoprophylaxis with ACT42 or PQ43 also has a potent prophylactic effect to prevent malaria transmission. Targeting high-risk groups such as gold miners in the TPT campaign might help increase overall population coverage. Moreover, malaria preventive practices and treatment-seeking behaviors among forest-related workers have historically been considerably low45. Therefore, malaria education should be carried out as well as surveillance systems to detect newly emerged cases.Similarly, gold miners often must stay near mines (often in forested settings) day and night. Therefore, they could not come back to their households. Findings from the qualitative component also corroborated this. Thus, in the next TPT campaigns, implementing mass treatment activities in the dry season or setting up malaria volunteers at the worksite might solve this challengeThere are both strengths and limitations to this study. To our knowledge, this study was the first research exploring the overall feasibility of primaquine mass treatment in Myanmar. As part of the study, the prevalence of G6PD deficiency was estimated for this township and it can also be applicable for considering routine G6PD testing in the national malaria surveillance system in the future. In addition to quantitative data, the inclusion of qualitative data added to our understanding of the processes under study, especially with regard to the lived challenges and barriers to participating in TPT. There are also a few limitations to this study. The study villages were selected because of the burden of vivax malaria. Our analysis is representative of these study villages, but may not be representative of others. Likewise, these data were collected in late 2019, prior to the onset of the COVID-19 pandemic and the recent political troubles in Myanmar. The healthcare and healthcare administration context has drastically changed. While there are still practical and useful lessons that have emerged from this study, there are likely new challenges that will need to be overcome if targeted mass treatment with PQ is to be implemented in Myanmar.There were several potentially useful lessons-learned from this study, perhaps especially with regard to future PQ trials in Myanmar or other similar regions. Only half of the targeted population completed the TPT after excluding ineligible participants and community members who were absent for part or all of the study (e.g. those who were away from the villages for work). An alternative approach like giving a shorter PQ regime during the dry season may increase adherence among some participants. In addition, implementing targeted treatment activity around the field sites or inside the workplaces by DOT may be a good option. Mitigating some inclusion criteria (e.g. including the 1- to 7-year-old children into the TPT) may be considered in upcoming trials to achieve higher overall coverage. Strengthening community awareness of targeted treatment activities especially among working-aged male farmers and gold miners might increase participation.Supplementary Table S1.Supplementary Figure S1."}
+{"text": "Plasmodium vivax malaria. In areas of chloroquine resistance, artemisinin-based combination therapies are used to treat malaria. This study aimed to evaluate tafenoquine plus the artemisinin-based combination therapy dihydroartemisinin\u2013piperaquine for the radical cure of P vivax malaria.Tafenoquine, co-administered with chloroquine, is approved for the radical cure (prevention of relapse) of P vivax malaria were randomly assigned by means of a computer-generated randomisation schedule (1:1:1) to dihydroartemisinin\u2013piperaquine alone, dihydroartemisinin\u2013piperaquine plus a masked single 300-mg dose of tafenoquine, or dihydroartemisinin\u2013piperaquine plus 14 days of primaquine (15 mg). The primary endpoint was 6-month relapse-free efficacy following tafenoquine plus dihydroartemisinin\u2013piperaquine versus dihydroartemisinin-piperaquine alone in all randomly assigned patients who received at least one dose of masked treatment and had microscopically confirmed P vivax at baseline . Safety was a secondary outcome and the safety population comprised all patients who received at least one dose of masked medication. This study is registered with ClinicalTrials.gov, NCT02802501 and is completed.In this double-blind, double-dummy, parallel group study, glucose-6-phosphate dehydrogenase-normal Indonesian soldiers with microscopically confirmed Between April 8, 2018, and Feb 4, 2019, of 164 patients screened for eligibility, 150 were randomly assigned (50 per treatment group). 6-month Kaplan-Meier relapse-free efficacy  was 11% (95% CI 4\u201322) in patients treated with dihydroartemisinin\u2013piperaquine alone versus 21% (11\u201334) in patients treated with tafenoquine plus dihydroartemisinin\u2013piperaquine  and 52% (37\u201365) in the primaquine plus dihydroartemisinin-piperaquine group. Adverse events over the first 28 days were reported in 27 (54%) of 50 patients treated with dihydroartemisinin\u2013piperaquine alone, 29 (58%) of 50 patients treated with tafenoquine plus dihydroartemisinin\u2013piperaquine, and 22 (44%) of 50 patients treated with primaquine plus dihydroartemisinin\u2013piperaquine. Serious adverse events were reported in one (2%) of 50, two (4%) of 50, and two (4%) of 50 of patients, respectively.P vivax malaria, the benefit was not clinically meaningful. This contrasts with previous studies in which tafenoquine plus chloroquine was clinically superior to chloroquine alone for radical cure of P vivax malaria.Although tafenoquine plus dihydroartemisinin\u2013piperaquine was statistically superior to dihydroartemisinin\u2013piperaquine alone for the radical cure of ExxonMobil, Bill & Melinda Gates Foundation, Newcrest Mining, UK Government all through Medicines for Malaria Venture; and GSK.For the Indonesian translation of the abstract see Supplementary Materials section. Plasmodium vivax malaria imposes a substantial global health burden, with more than 3 billion people at risk of infection.P vivax consists of combining a blood schizontocide, chloroquine, with a liver stage hypnozoitocide, primaquine, to prevent relapse from hepatic latency. WHO recommends a 7-day or 14-day course of primaquine;P vivax malaria by 70% compared with chloroquine alone.6In many endemic countries, \u201cradical cure\u201d of Evidence before this studyPlasmodium vivax malaria has been previously established in randomised controlled studies. In the phase 3 DETECTIVE study done in Ethiopia, Peru, Brazil, Cambodia, Thailand, and the Philippines, recurrence-free efficacy rates at 6 months (modified intention-to-treat population) were 62\u00b74% (95% CI 54\u00b79\u201369\u00b70) in the tafenoquine group, 27\u00b77% (19\u00b76\u201336\u00b76) in the placebo group, and 69\u00b76% (60\u00b72\u201377\u00b71) in the primaquine group. In a patient-level meta-analysis of the phase 3 DETECTIVE study and the phase 3 GATHER study , recurrence-free efficacy rates at 6 months were 67\u00b70% (61\u00b70\u201372\u00b73) in the tafenoquine group and 72\u00b78% (65\u00b76\u201378\u00b78) in the primaquine group. Dihydroartemisinin\u2013piperaquine is an artemisinin-based combination therapy used as an alternative to chloroquine for treatment of malaria. A PubMed search for articles published from database inception to July 1, 2022 containing the terms \u201cvivax malaria\u201d, \u201ctafenoquine\u201d, \u201cdihydroartemisinin\u201d or \u201cartenimol\u201d, and \u201cpiperaquine\u201d, with no language restrictions, revealed only a single article of relevance: a drug\u2013drug interaction study in healthy volunteers investigating the pharmacokinetic interaction between tafenoquine and the artemisinin-based combination therapies, dihydroartemisinin-piperaquine, and artemether-lumefantrine. There were no reported studies evaluating the efficacy and safety of tafenoquine co-administered with artemisinin-based combination therapies for the radical cure of P vivax malaria.The efficacy of a single 300-mg dose of tafenoquine co-administered with chloroquine for the radical cure of Added value of this studyP vivax malaria. No clinically meaningful benefit was observed. The benefit of primaquine (15 mg) plus dihydroartemisinin\u2013piperaquine was clinically meaningful, although nearly half of all patients relapsed within 6 months of treatment.This is the first clinical study to evaluate the efficacy and safety of a single 300-mg dose of tafenoquine co-administered with dihydroartemisinin\u2013piperaquine for the radical cure of Implications of all the available evidenceP vivax malaria. These results are important for national malaria control programmes in countries which might consider using artemisinin-based combination therapies for P vivax malaria.This study does not support co-administration of a single 300-mg dose tafenoquine with dihydroartemisinin\u2013piperaquine for the radical cure of P vivax malaria were reported in 2019,P vivax malaria.P vivax infection in regions in which chloroquine resistance is commonThe emergence of chloroquine resistanceP vivax malaria. The study was designed to show superiority of a single 300-mg dose of tafenoquine co-administered with standard doses of dihydroartemisinin\u2013piperaquine, compared with dihydroartemisinin\u2013piperaquine alone, for prevention of relapse of P vivax malaria over 6 months in Indonesian soldiers returning to malaria-free military bases in Malang and Madiun following deployment to an area with high P vivax endemicity who were phenotypically glucose-6-phosphate dehydrogenase-normal (The INSPECTOR study (Indonesian Study Proving Efficacy of Combination Therapy on Relapse) is the first study to evaluate co-administration of tafenoquine with dihydroartemisinin\u2013piperaquine for the radical cure of e-normal .14P vivax malaria by microscopy at their army bases were invited to participate in the study. A clinic, laboratory, and pharmacy were established at both bases for the duration of the study.This double-blind, double-dummy, randomised, parallel group study enrolled soldiers from two battalions based in East Java , and were glucose-6-phosphate dehydrogenase-normal as assessed by the qualitative fluorescent spot test , as primaquine and tafenoquine can cause acute haemolytic anaemia in patients with glucose-6-phosphate dehydrogenase deficiency. Glucose-6-phosphate dehydrogenase status was confirmed shortly after randomisation by quantitative spectrophotometric assay .Patients were eligible for enrolment if they were male, aged at least 18 years, had microscopically confirmed P vivax malaria (as defined by WHO), severe vomiting, corrected QT interval (QTc) of at least 450 msec, screening haemoglobin concentration less than 8 g/dL, alanine aminotransferase greater than twice the upper limit of normal, consumption of antimalarial drugs or drugs known to prolong the QTc interval in the past 30 days, or any other contraindication to administration of dihydroartemisinin\u2013piperaquine or primaquine. A complete list of exclusion criteria is included in the The main exclusion criteria were severe The trial was designed to support the use of tafenoquine 300 mg with dihydroartemisinin\u2013piperaquine. The underlying relapse rate following a course of dihydroartemisinin\u2013piperaquine cannot be assumed from historical data owing to the natural variation in both infection rates and relapse rates among infected subjects. The dihydroartemisinin\u2013piperaquine alone group was included as a relapse-prevention placebo control to provide an efficacy benchmark in the same setting and at the same time as the study treatment groups. The inclusion of the primaquine group provided a context against which to interpret the observed tafenoquine efficacy rate. A statistical comparison for non-inferiority between tafenoquine and primaquine was not done; such a comparison would require a considerably larger sample size and was not the main objective of the study, which was to estimate the efficacy of tafenoquine.The study complied with Good Clinical Practice, the Declaration of Helsinki, and relevant regulatory requirements. Ethics approval was obtained from the Oxford Tropical Research Ethics Committee (Project 9-16) and the Faculty of Medicine Universitas Indonesia Ethics Committee . Written informed consent was obtained individually from study patients and affirmed by an independent peer witness. Army rank superiors were not permitted to witness or participate in the consenting process to avoid any perception of coercion. The protocol was amended once on April 20, 2017 (before study start) to include an additional secondary objective requested by the Indonesian regulatory agency: comparison of dihydroartemisinin\u2013piperaquine plus primaquine  with dihydroartemisinin\u2013piperaquine alone. The full protocol is available online (NCT 02802501).Eligible patients were randomly assigned in a 1:1:1 ratio to dihydroartemisinin\u2013piperaquine alone, dihydroartemisinin\u2013piperaquine plus tafenoquine, or dihydroartemisinin\u2013piperaquine plus primaquine. All patients received open-label dihydroartemisinin\u2013piperaquine for 3 days plus masked treatment as follows: tafenoquine 300-mg single dose on day 1 and placebo for primaquine on days 1\u201314; primaquine 15 mg on days 1\u201314 and placebo for tafenoquine on day 1; or placebo for tafenoquine on day 1 and placebo for primaquine on days 1\u201314. The study sponsor provided a computer-generated randomisation schedule to the site. Visually matched tafenoquine and primaquine placebos were used to maintain masking of site staff, patients, and sponsor personnel.As methaemoglobin increases are associated with use of 8-aminoquinolines such as primaquine and tafenoquine, methaemoglobin assessments were done by an independent site assessor to avoid unmasking. Additionally, independent sponsor staff processed the methaemoglobin data. The independent methaemoglobin site and sponsor staff did not have access to other study data.Study treatments were supplied by the sponsor as eurartesim tablets containing dihydroartemisinin 40 mg and piperaquine 320 mg , tafenoquine 150-mg tablets , and primaquine formulated as over-encapsulated 15-mg tablets . All patients received open-label, oral dihydroartemisinin\u2013piperaquine daily over 3 consecutive days (days 1\u20133), according to weight (three tablets <75 kg or four tablets \u226575 kg). Tafenoquine (or matched placebo) was given as a single oral 300-mg dose (two \u00d7 150-mg tablets) on either day 1 or day 2. Primaquine (or matched primaquine placebo) was administered as a single oral 15-mg daily dose for 14 consecutive days starting on day 1 or day 2. All study medications were administered under direct supervision by study staff in the clinic.Following informed consent, screening assessments were done and all patients who were eligible to receive dihydroartemisinin\u2013piperaquine on the basis of labelling received their first dose of dihydroartemisinin\u2013piperaquine at least 3 h after their last meal. Once laboratory results confirmed continued eligibility, patients were randomly assigned to masked study medication, with the first dose given, with food, at least 3 h after dihydroartemisinin\u2013piperaquine, on day 1 or day 2. Blood smears for parasite assessment were done twice daily until two consecutive negative smears were obtained. After completion of the dosing period, patients attended a further seven follow-up visits .P vivax positive blood smears with or without symptoms were treated with rescue medication: dihydroartemisinin\u2013piperaquine plus open-label primaquine 0\u00b75 mg/kg daily for 14 days (see appendix 2 p 22 for study design).Patients were evaluated by clinical assessments and laboratory investigations including Giemsa-stained blood smears for parasitology, 12-lead electrocardiograms, haematology and clinical chemistry tests, and methaemoglobin measurement by means of a non-invasive pulse oximeter  at screening and selected visits throughout the study. After their initial in-patient treatments, patients were instructed to promptly return to the clinic (open 24 h) if they had malaria symptoms. Patients who returned with P vivax clones were identical to baseline in all markers. Masked external quality assurance was done on 20% of samples for CYP2D6 analysis, 20 paired samples for parasite DNA analysis, all positive blood films, and 10% of negative blood films  genotyping and parasite microsatellite DNA analysis (to establish the proportion of heterologous and homologous relapses). Relapse was defined as genetically homologous when All patients who were relapse free at 6 months received high-dose open-label primaquine  for 14 days at the end of the study to minimise the likelihood of relapse.The primary outcome was relapse-free efficacy over 6 months, defined as clearance of initial infection without subsequent microscopically confirmed recurrence or receipt of other antimalarial treatment. Secondary outcomes were relapse-free efficacy over 4 months, time to fever clearance, time to parasite clearance, and percentage of patients with recrudescence  to ensure the blood stage efficacy of dihydroartemisinin\u2013piperaquine. Safety assessments included frequency and severity of adverse events and review of 12-lead electrocardiograms, vital signs, and laboratory values. Tafenoquine pharmacokinetic assessments were also planned.The incidence of genetically homologous infections was established by five microsatellite markers. Additionally, the influence of human CYP2D6 polymorphisms on relapse was explored post-hoc by means of a CYP2D6 activity score system see .The primary comparison of interest was relapse-free efficacy over 6 months for tafenoquine plus dihydroartemisinin\u2013piperaquine versus dihydroartemisinin\u2013piperaquine alone. By means of the log-rank test for the primary comparison to detect a clinically meaningful difference of 35% in relapse-free survival rates over 6 months, and assuming a 50% rate on dihydroartemisinin\u2013piperaquine alone,P vivax at baseline were included in the primary and secondary efficacy analyses . The per-protocol population included all patients in the microbiological intention-to-treat population for whom there were no major protocol violations. The safety population comprised all patients who received at least one dose of masked medication. The primary treatment comparison in the study was tafenoquine plus dihydroartemisinin\u2013piperaquine versus dihydroartemisinin\u2013piperaquine alone. Although other treatment comparisons were made , these were considered secondary and hence no adjustments for multiplicity were made.All randomly assigned patients who received at least one dose of masked treatment and had microscopically confirmed P vivax parasitaemia, took an antimalarial drug post-baseline without confirmed P vivax, or did not have a 6-month assessment within the defined time window. Further sensitivity analyses were done , which analysed the proportion of participants who were relapse free at 4 and 6 months by means of logistic regression analyses . Treatment by covariate interaction (at 10% significance level) was assessed by means of Cox's proportional-hazards model to evaluate the effect of battalion , baseline P vivax parasite count and patient weight on the treatment effect. No adjustments for other covariates were made. Clearance times for P vivax and fever were estimated by means of Kaplan-Meier methods. The effect of CYP2D6 activity score or metaboliser class on relapse within each treatment group was investigated by logistic regression . Other efficacy and safety endpoints were summarised by means of descriptive statistics. Statistical analyses were done by means of the SAS software, version 9.4. Masked safety data were reviewed on a monthly basis by a GSK\u2013MMV safety review board which included a reviewer independent of the study sponsors. This study is registered with ClinicalTrials.gov, NCT02802501.The primary endpoint, time to relapse over 6 months, was summarised by means of Kaplan-Meier estimates and analysed by means of a Cox's proportional-hazards model, adjusting for battalion, for the microbiological intention-to-treat (primary) and per-protocol (sensitivity) populations. Of note, the protocol described a log-rank test, but this was amended in the study analysis plan  to Cox's proportional hazards. Patients were censored if they did not show initial clearance of The study sponsor (GSK) was responsible for study monitoring, data management, data analysis, and writing of the clinical report.Between April 8, 2018, and Feb 4, 2019, of 164 patients screened for eligibility, 150 were randomly assigned to masked study treatment; 50 per treatment group  for dihydroartemisinin\u2013piperaquine alone, 21\u00b70% (10\u00b77\u201333\u00b76) for tafenoquine plus dihydroartemisinin\u2013piperaquine, and 52\u00b70% (37\u00b74\u201364\u00b77) for primaquine plus dihydroartemisinin\u2013piperaquine. Tafenoquine plus dihydroartemisinin\u2013piperaquine reduced the risk of relapse at any time by 55\u00b76% versus dihydroartemisinin\u2013piperaquine alone  in the dihydroartemisinin\u2013piperaquine alone group, 39 patients (78%) in the tafenoquine plus dihydroartemisinin\u2013piperaquine group and 24 patients (48%) in the primaquine plus dihydroartemisinin\u2013piperaquine group had microscopically confirmed Primaquine plus dihydroartemisinin\u2013piperaquine reduced the risk of relapse over 6 months by 74\u00b72% versus dihydroartemisinin\u2013piperaquine alone . The reduction in the 6-month relapse risk for tafenoquine plus dihydroartemisinin\u2013piperaquine versus dihydroartemisinin\u2013piperaquine alone was greater for battalion 2  of 150 being categorised as poor or intermediate metabolisers . Logistic regression analyses showed no significant effect of CYP2D6 activity score  or metabOverall, 88 (83%) of 106 of all first relapses were genetically heterologous by microsatellite genotyping and the proportion of heterologous to homologous relapses was not influenced by treatment .The adverse event profile was similar across treatment groups, although the proportion of patients in the primaquine plus dihydroartemisinin\u2013piperaquine group reporting any adverse event was lower than for other groups . Most adFive serious adverse events were reported during the 6-month follow-up of which two were considered possibly related to treatment by the investigator . One patA higher percentage of patients in the tafenoquine plus dihydroartemisinin\u2013piperaquine group (12 [24%] of 50) had a clinically significant QTcF prolongation (defined as \u226560 msec change from baseline) on the 12-lead electrocardiogram . In all cases, QTcF prolongation was asymptomatic and returned to normal within 1 week without intervention . Only onHaemoglobin values were similar between treatment groups  and no pOwing to various logistical factors including the global response to the COVID-19 pandemic, it has not, to date, been possible to transport samples for pharmacokinetic analyses to a validated laboratory outside of Indonesia. Pharmacokinetic data from study patients cannot therefore be included in this publication.P vivax malaria. A dihydroartemisinin\u2013piperaquine alone group provided a placebo control for the primary assessment of tafenoquine plus dihydroartemisinin\u2013piperaquine efficacy for relapse prevention. Inclusion of a placebo control was considered justifiable as all patients received dihydroartemisinin\u2013piperaquine for the acute illness and were closely monitored for relapse on an army base, thereby ensuring that any episode of recurrent malaria could be treated promptly. Tafenoquine plus dihydroartemisinin\u2013piperaquine reduced the risk of relapse versus dihydroartemisinin\u2013piperaquine alone . However, a 10% difference in relapse-free efficacy was not considered clinically meaningful. The efficacy of tafenoquine plus dihydroartemisinin\u2013piperaquine was lower than observed in previous studies of tafenoquine co-administered with chloroquine.This is the first study to evaluate the efficacy and safety of a single 300-mg dose of tafenoquine co-administered with an artemisinin-based combination therapy for the radical cure of The reasons for the lack of clinically relevant efficacy with tafenoquine plus dihydroartemisinin\u2013piperaquine are unknown. Although no pharmacokinetic data are yet available, a pharmacokinetic interaction is considered unlikely given that there was no clinically significant pharmacokinetic interaction between tafenoquine and dihydroartemisinin\u2013piperaquine in a previous healthy volunteer study.Previous studies of primaquine plus chloroquine indicate that diminished primaquine efficacy might be associated with impaired CYP2D6 activity.The soldiers' battalion had a significant effect on estimates of efficacy, whereas other covariates (weight and baseline parasite counts) exerted no significant influence on the primary endpoint. Although these observations did not change the overall interpretation of the study , there was a notably higher relapse rate across all treatment groups in the second battalion compared with the first battalion, which could not be explained by differences in study conduct or drug supplies. Despite both battalions having similar baseline characteristics and being deployed to the same region of Indonesian Papua for similar durations, sharp differences in transmission intensity within that region are believed to have occurred. Battalion 1 had been working in a sparsely populated and forested sector, whereas battalion 2 had been operating in a relatively settled (cleared forest) and populated area\u2014conditions conducive to relatively light and heavy malaria transmission, respectively\u2014on the basis of the dominant anopheline vectors in New Guinea, which prefer open sunlight to shady forests.P vivax burden); relapse-free efficacy at 6 months was 91\u00b79% (tafenoquine 600 mg plus chloroquine) versus 89\u00b72% (tafenoquine 300 mg plus chloroquine).Consideration has been given to whether the 300-mg dose of tafenoquine used in this study might be suboptimal.P vivax in Indonesian Papua, a region where the Chesson strain of P vivax originated, a type which is characterised by frequently occurring relapses and tolerance to low-dose primaquine.P vivax radical cure in southeast Asia, uptake is low because of the haemolytic risk associated with 8-aminoquinoline treatment in the absence of G6PD testing. Therefore, the current study used low-dose primaquine (15 mg/day for 14 days) in accordance with Indonesian national treatment guidelines. The 6-month relapse-free efficacy with this primaquine regimen was 52% compared with 11% for dihydroartemisinin\u2013piperaquine alone which is lower than in previous studies with primaquine 15 mg/day plus chloroquine done in other countriesPatients in this study were infected with The adverse event profile for tafenoquine plus dihydroartemisinin\u2013piperaquine was consistent with the known adverse event profiles for the individual drugs and there were no new safety signals. There were no deaths, and no patients withdrew from the study. Two patients had a serious adverse event, which the investigator considered related to study treatment, but neither received tafenoquine. QTc prolongation is a known risk with dihydroartemisinin\u2013piperaquine.P vivax in Indonesian Papua, a region with one of the highest malaria transmission rates in the world, possibly associated with higher frequencies of hypnozoite carriage.The current study has several limitations. It was only done in patients who had been infected with P vivax recurrences in the study were very probably due to relapse. However, it was not possible to know whether the absence of recurrent parasitaemia was due to efficacious treatment or simply natural hypnozoite depletion. Genetic homology is not validated as proof of whether recrudescence, reinfection, or relapse has occurred (a relapse can be either homologous or heterologous). Thus, as there are no definitive methods to establish the presence of hypnozoites in the liver or confirm whether P vivax parasitaemia is due to relapse or de novo re-infection, the model used in this study provides an optimal method of evaluating radical cure in the clinical setting.The study design in which patients were infected in a highly malarious region and returned to an area free of malaria transmission meant that P vivax malaria. Although primaquine plus dihydroartemisinin-piperaquine was more efficacious than tafenoquine plus dihydroartemisinin\u2013piperaquine, nearly half of all patients receiving primaquine 15 mg daily for 14 days plus dihydroartemisinin\u2013piperaquine also relapsed. The findings of this study do not alter the established favourable benefit\u2013risk profile of single 300-mg dose tafenoquine co-administered with chloroquine for radical cure of P vivax malaria.In conclusion, the results of this study do not support co-administration of single 300-mg dose tafenoquine with dihydroartemisinin\u2013piperaquine for the radical cure of This online publication has been corrected. The corrected version first appeared at thelancet.com/infection on July 25, 2023https://www.clinicalstudydatarequest.com/. A data access agreement will be required.Within 6 months of this publication, anonymised individual patient data, the annotated case report form, protocol, reporting and analysis plan, dataset specifications, raw dataset, analysis-ready dataset, and clinical study report will be available for research proposals approved by an independent review committee. Proposals should be submitted to , and funding from Medicines for Malaria Venture (MMV) and GSK. AS, KC, RN, DS, YWS, II, WB, CBP, and SL report that they or their institutions received funding from MMV to do the study and editorial support from GSK for this publication. LLE and AWS report institutional funding from Menzies School of Health Research, MMV, and GSK. CC-D reports institutional funding from MMV, GSK, and the Chan Zuckerberg Initiative. IE reports institutional funding from MMV, GSK, WHO, SPARK (University of Melbourne), and honoraria from the Indonesian Ministry of Health. SD is an employee of MMV; his institution received funding from ExxonMobil, the Bill & Melinda Gates Foundation (grant number INV-007155/19-BMGF-006), Newcrest Mining, and the UK Government to fund the INSPECTOR study. EC is a former independent contractor at GSK and holds shares in the company. EC developed the first draft of the manuscript and provided editorial services as an independent medical writer funded by GSK. KR, DF, AB, SJ, KF, HS, AM, MT, and LKT are employees of GSK and hold shares in the company. J-PK, NG, and JAG are former employees of GSK and hold shares in GSK. JKB reports institutional research funding from MMV, GSK, the Wellcome Trust, the Gates Foundation, FIND, Sanaria, UKAid, University of Oxford, US Centers for Disease Control and Prevention, and the Chinese Center for Disease Control; travel and speaking fees from the Belgian Society of Tropical Medicine, the International Conference of Tropical Medicine and Malariology, and Singapore Malaria Group Conference; and participation on the US National Health Institute Data Safety Monitoring Board.IS reports grants or contracts from Menzies School of Health Research, Darwin and the National Health and Medical Research Council"}
+{"text": "Plasmodium falciparum infections may be accounted for this difference. Hence, the aims of this study were to assess the complexity of infection (COI) and genetic diversity of P. falciparum parasites during malaria treatment in Bougoula-Hameau and Faladje in Mali.Effective approaches to fight against malaria include disease prevention, an early diagnosis of malaria cases, and rapid management of confirmed cases by treatment with effective antimalarials. Artemisinin-based combination therapies are first-line treatments for uncomplicated malaria in endemic areas. However, cases of resistance to artemisinin have already been described in South-East Asia resulting in prolonged parasite clearance time after treatment. In Mali, though mutations in the K13 gene associated with delayed clearance in Asia are absent, a significant difference in parasite clearance time following treatment with artesunate was observed between two malaria endemic sites, Bougoula-Hameau and Faladje. Hypothetically, differences in complexity of Pfcsp and Pfama1 encoding gene were amplified by nested PCR and sequenced using the Illumina platform. The parasite clearance time (PCT) was determined using the parasite clearance estimator of Worldwide Antimarial Resistance Network (WWARN). Data were analyzed with R\u00ae.Thirty (30) patients per village were randomly selected from 221 patients enrolled in a prospective artesunate monotherapy study conducted in Faladje and Bougoula-Hameau in 2016. All parasitemic blood samples of patients from enrollment to last positive slide were retained to assess malaria parasite COI and polymorphisms. DNA were extracted with a Qiagen kit and The median number of genetically distinct parasite clones was similar at enrollment, 7 (IQR of 5\u20139) in Faladje and 6 (IQR of 4\u201310) in Bougoula-Hameau . On the first day after treatment initiation, the COI was higher in Faladje  than in Bougoula-Hameau  with a p-value =0. 02. Overall, COI was high with higher PCT. Finally, there was a low genetic diversity between Faladje and Bougoula-HameauThis study demonstrated that the difference in PCT observed between the two villages could be due to differences in the complexity of infection of these two villages. It is transmitted to humans by the bite of a female Anopheles mosquito, although, other means of transmission such as transfusion and fetal-maternal route exist [Plasmodium: P. falciparum, P. malariae, P.ovale wallikeri, P.ovale cutisi, P. vivax and P. Knowlesi infect humans, P. falciparum being the most lethal species.Malaria is a febrile and hemolyzing erythrocytopathy due to the presence and the development in the liver and then in the blood of a hematozoa of the genus te exist , 2. Six Despite the substantial progress made in malaria treatment and prevention, it remains a serious public health challenge in Africa. In 2021, there were an estimated 241 million cases of malaria worldwide, with 95% occurring in Africa . WHO repArtemisinin-based combination therapies (ACT) are the recommended first line treatment of uncomplicated malaria in endemic countries . In MaliP. falciparum kelch 13 (Pfkelch13) propeller domain gene [P. falciparum with high survival rates by ring survival assay associated with the Pfkelch13 A675V mutation has already been reported in Uganda in 2018 [Pfkelch13 R561H mutation associated with delayed parasite clearance after AL treatment in Rwanda [However, there have been concerns of reduced artemisinin efficacy in Southeast Asia  since 20ain gene . Emergen in 2018 . This isn Rwanda . These tPfkelch13 gene were described in samples collected during the study, they were not associated with artemisinin resistance [Pfkelch13 A578S mutation was observed in one sample (1/98) in Bougoula-Hameau while no such mutation was found in Faladje (N = 118) [Pfkelch13 gene. The discrepancy may probably be due to other factors such as immunity or complexity of infection (COI).Following the detection of artemisinin resistance cases in Southeast Asia, a prospective therapeutic efficacy study of artesunate conducted in Mali in 2010\u20132011 reported no evidence of delayed parasite clearance [sistance . In viewThe complexity of malaria infection is defined as the number of genetically distinct parasite variants co-infecting a single host, which is an important indicator of malaria epidemiology . MalariaP. falciparum infections to Artesunate monotherapy in Mali. To test our hypothesis, we assessed the dynamic of complexity of infection and the genetic diversity of P. falciparum in two malaria endemic villages of Mali during treatment with artesunate monotherapy.We hypothesized that the complexity of infection and genetic diversity contribute to differences in sensitivity of 2.2.1.P. falciparum malaria in Bougoula-Hameau and Faladje. We enrolled 121 participants in Faladje and 100 in Bougoula-Hameau. The protocol of this study was approved by the Ethics Committee of the Faculty of Medicine, Pharmacy, and Stomatology and written informed consent was obtained from the parent or guardian of each child before inclusion. Participants were hospitalized and treated for 7 days with artesunate monotherapy and were until all malaria symptoms and parasitemia were resolved. They were then followed-up for 28 days. Brielfly, four (4) mg / kg of artesunate was administered  on day 0 then 2 mg / kg from day 1 to day 7. Slides and dry blood spots were collected every 8 hours until three consecutive slides were negative. Additional blood specimens were collected on days 2, 3, 7, 14, 21 and 28. Blood smears were screened for parasitemia by two trained microscopists according to WHO standard procedures [A prospective therapeutic efficacy study of artesunate was conducted from October 2015 to March 2016 in patients who were at least 6 months of age or older and diagnosed with uncomplicated ocedures .2.2.We randomly selected 30 patients per site using the \u201csample\u201d function of the R package. A total of 259 time points from the 60 patients were sampled and used for sequencing. These time points represented all the positive dried blood spots (DBSs) from enrollment to the last positive slide.2.3.P. falciparum DNA was extracted from DBS using QIAGEN kit according to the manufacturer\u2019s instructions (Qiagen\u00ae). After DNA extraction a qPCR assay was used to amplify the varATS gene to verify the presence of P. falciparum parasite DNA in the extracts [Pfama1 and Pfcsp to investigate the complexity of infection and parasite genetic diversity in infections from Bougoula-Hameau and Faladje. These genes were amplified by nested PCR using two sets of primers, Pfama1-F1 (GAA GTT CAT GGT TCA GGT ATA AG); Pfama1-R1 (GTA TGG TTT TTC CAT CAG AAC TGG) and Pfcsp-F1(GTC GGA ATT CAT GAT GAG AAA ATT AGC TATT); Pfcsp-R1 (CTA ATT AAG GAA CAA GAA GG) to amplify the outer regions of Pfama1 (1503 bp) and Pfcsp (1194 bp) respectively. Two others set of primers Pfama1-NF (GAT GCT GAA GTA GCT GGA ACTC); Pfama1-NR (GTG ATG CTC TTT TTT CTT CCC CCC) and Pfcsp-NF (TCG TCA AAC ACA AGG GTT CT); Pfcsp-NR (ACG ACA TTA AAC ACA CTG GAA CA) were used for nested PCR to amplify 1432 bp of Pfama1 and 1058 bp of Pfcsp. For primary and nested PCR, Pfama1 and Pfcsp were amplified under similar cycling conditions: 98\u00b0C for 1 min, followed by 35 cycles with 45 s of denaturation at 98\u00b0C, annealing at 63\u00b0C for 45 s, and elongation at 72\u00b0C for 1 min. After 35 cycles, a final elongation step at 72\u00b0C for 5 min. After amplification, the PCR product was visualized on 1,2% agarose gel. PCR products were quantified by Qubit and normalized prior to pooling amplicons of the two genes for each sample. Pooled amplicons were used for library prepared using NEBNext\u00ae Ultra\u2122 II kit following the manufacturer\u2019s instructions. Libraries were quantified on Qubit and fragment sizes visualized on the Agilent 2200 TapeStation system. Library concentrations were used to pool indexed samples and sequenced (paired end) on Illumina Miseq platform.extracts . We used2.4.Pfama1 and Pfcsp, the genetic diversity (Fst), the within-infection fixation indices (FWS) and run a principal component analysis  in order to identify structure in the distribution of genetic variation across the two villages at different of follow up time point  sequences of each gene using BWA and SAM files generated from this step were converted into BAM files . During me point .FWS and parasite clearance time, respectively. The significance level was set at 5%.The nucleotide diversity analysis was performed using DnaSP v5. We used Student and Wilcoxon test to compare the complexity of infection and nucleotide diversity within and between populations. Linear regression implemented in ggplot R package were used to assess the correlation between the complexity of infection and parasite density, and 3.In both villages the number of samples per time point was around 30 except for timepoint H32 in Bougoula-Hameau and Faladje where only respectively 3 and 21 patients had positive slides.3.1.The median of parasite clearance time (PCT) was assessed and compared between the two study sites. An analysis of the difference of PCT between the two villages showed that the median PCT was significantly lower  in Bougoula-Hameau (2 hours) compared to Faladje (3 hours) .3.2.a decrease in the COI in Bougoula-Hameau during the follow-up time, with samples collected 32 hours post-treatment having the least COI, while those collected before treatment and up to 16 hours post-treatment had the highest COI . However, the number of clones was higher  in Faladje compared to Bougoula Hameau one day after treatment initiation  , with the least complex infections having the highest FWS  was lower one day post treatment than at enrollment in Bougoula and Faladje with p-value respectively equal to 0.02 and 0.002  and one day after treatment initiation , indicating that the within-host parasite diversity was high in Faladje compared to Bougoula-Hameau (To assess the effect of artesunate therapy on nd 0.002 . The meaa-Hameau .ST) between Bougoula-Hameau and Faladje was generally low, with an FST value of 0.001 at baseline and 0.002 one day after treatment.  and within-infection fixation indices (FWS) between the two study sites.Malaria treatment with artemisinin-based combination drugs have averted millions of deaths since their introduction almost two decades ago , 20. RedOur findings suggest a decline in the mean nucleotide diversity parameter (\u03c0) from enrollment to 24 hours after initiation of treatment in Bougoula-Hameau, while the pattern was not stable between time points in Faladje. Faladje was characterized with long parasite clearance times associated with more multiple strain (polygenetic) infections. These results suggest that genetic diversity at commencement of treatment may have contributed to the parasite clearance rate, with more complex infection responding slower to drugs. As the parasite populations from the two villages showed no population substructure from Fst and PCA analysis, there is adequate gene flow between the two sites, difference in transmission immunity may also have affected clearance time.et al [et al [P. falciparum in Kenya and Tanzania. This is contrary to our findings, although in this case we used artesunate monotherapy instead of an ACT as conducted by Topazian et al [P. falciparum parasites during ACT treatment in Democratic Republic of the Congo [Artemisinin resistance is measured by slow parasite clearance rates. The relevance of this definition and the importance of these phenotypes in Mali where parasitemia is mostly high needs further investigation. These assessments require consideration of the direct effect of artemisinin and its companion drugs, and other factors including sequestration and immunity on drug efficacy. While a study by Lee et al  supportel [et al  have fouan et al . On the an et al , althoughe Congo . These sThe village with more complex infections, Falaje, had a higher clearance time. Higher diversity and infection complexity reflect the intensity of transmission, with higher transmission driving the acquisition of immunity, thus helping drugs to clear malaria parasites , 26. ThiP. falciparum population as observed in high transmission settings, while low transmission settings are characterized by more monoclonal infections [The inbreeding coefficient is used to understand malaria parasite population biology as it undergoes gamete formation and fertilization during mosquito lifecycle . Highly fections , 29. Comfections , 31. At fections . These dfections . The isoThe novelty and strength of this work is the significant positive association between parasites clearance time  and the complexity of infection and the significant difference between parasite clearance time and complexity of infection between the two villages. In addition, this study used artesunate monotherapy, allows for an unbiased monitoring of the efficacy of an artemisinin derivative, contrary to ACTs where there is interference from partner drug efficacy.Pfama1 and Pfcsp) instead of whole genome, missing important contributions that may have come from other genes. However, we believe our approach has not significantly impacted on our conclusions. We also did not determine artesunate pharmacokinetics during the in vivo efficacy study, which would have allow us determine any differences in plasma drug concentration, that could influence drug efficacy [Our study has some limitations. Genetic diversity analyses have been carried out using amplicon deep sequencing of two polymorphic genes (efficacy .5.This study described a significant difference in complexity of infection between Bougoula-Hameau and Faladje 24 hours after artesunate monotherapy treatment. Complexity of infection positively correlates with the parasite clearance time and could be a significant determinant in treatment efficacy of artemisinin derivatives and ACTs."}
+{"text": "Plasmodium falciparum infections in many African countries, including Kenya. Recurrent infections have been reported in patients treated with AL or DP, suggesting the possibility of reinfection or parasite recrudescence associated with the development of resistance against the two therapies. The Plasmodium falciparum cysteine desulfurase IscS (Pfnfs1) K65 selection marker has previously been associated with decreased lumefantrine susceptibility. This study evaluated the frequency of the Pfnfs1 K65 resistance marker and associated K65Q resistant allele in recurrent infections collected from P. falciparum-infected individuals living in Matayos, Busia County, in western Kenya.Malaria remains a public health concern globally. Resistance to anti-malarial drugs has consistently threatened the gains in controlling the malaria parasites. Currently, artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) are the treatment regimens against Pfnfs1 K65 resistance marker and K65Q mutant allele in the recurrent infections. Plasmodium falciparum msp1 and P. falciparum msp2 genetic markers were used to distinguish recrudescent infections from new infections.Archived dried blood spots (DBS) of patients with recurrent malaria infection on clinical follow-up days after treatment with either AL or DP were used in the study. After extraction of genomic DNA, PCR amplification and sequencing analysis were employed to determine the frequencies of the The K65 wild-type allele was detected at a frequency of 41% while the K65Q mutant allele was detected at a frequency of 22% in the recurrent samples. 58% of the samples containing the K65 wild-type allele were AL treated samples and while 42% were DP treated samples. 79% of the samples with the K65Q mutation were AL treated samples and 21% were DP treated samples. The K65 wild-type allele was detected in three recrudescent infections (100%) identified from the AL treated samples. The K65 wild-type allele was detected in two recrudescent DP treated samples (67%) while the K65Q mutant allele was identified in one DP treated (33%) recrudescent sample.The data demonstrate a higher frequency of the K65 resistance marker in patients with recurrent infection during the study period. The study underscores the need for consistent monitoring of molecular markers of resistance in regions of high malaria transmission.The online version contains supplementary material available at 10.1186/s12936-023-04587-2. Plasmodium genus. The deadliest species, Plasmodium falciparum, is the most prevalent species on the African continent, where 95% of total malaria cases occur [Globally, malaria remains a disease of significant public health concern. In 2020, 627,000 deaths and 241 million cases of the disease were reported [P. falciparum malaria due to the efficacy of combining a short-acting artemisinin derivative with a longer half-life partner drug that eradicates residual parasite load [P. falciparum parasites was first reported in South-East Asia [Artemisinin-based combination therapy (ACT) is used to treat ite load . The emeast Asia . ACT remast Asia \u20136. Howevast Asia  and Uganast Asia . The losast Asia . An addiast Asia . Anti-maP. falciparum kelch13 (PfK13) gene [P. falciparum chloroquine resistance transporter (PfCRT) and the P. falciparum multidrug resistance 1 transporter (PfMDR1) [pfmdr1 gene [Artemisinin resistance is mainly associated with mutations in several loci of the 13) gene \u201313. Resi(PfMDR1) . The mec(PfMDR1) . Resistadr1 gene . Studiesdr1 gene . TherefoP. falciparum infections in Kenya [In Kenya, 70% of the entire population is at risk of contracting malaria . Approxiin Kenya . Non-synin Kenya , 23.Plasmodium falciparum cysteine desulfurase IscS (Pfnfs1) (PF3D7_0727200) has been associated with lumefantrine resistance. In the Gambia, higher IC50 values for the wild-type K65 alleles and a strong temporal differentiation of the same allele over seven years were associated with the lumefantrine resistance selection [Pfnfs1 need further studies to determine their potential roles in drug action and resistance.election . The PfNelection . These celection \u201328. The election , and theelection . Given telection , genes sPfnfs1 gene and associated outcomes in recurrent infections. To investigate mutations in the target region of the Pfnfs1 gene, DNA was extracted from the DBS, followed by PCR amplification and sequencing of the isolates.This study analysed archived DBS collected on days 21, 28, and 42 post-AL and DP treatment to determine the frequency of mutations in the target region of the 2. The region experiences two rainy seasons annually, from March to June and October to November. The area has a high malaria transmission and is part of the Lake Victoria basin, thereby providing suitable breeding conditions for the main malaria vectors, Anopheles gambiae, and Anopheles funestus, which facilitate continuous malaria transmission, while P. falciparum is the most prevalent parasite species in the area [The study was carried out in Matayos, Busia County, Western Kenya. Matayos is located at a latitude of 0.3618\u00b0N and longitude of 34.168\u00b0E, lies at an altitude is 1214\u00a0m above sea level, and has an area of 196.2kmthe area .P. falciparum mono-infection, and parasitaemia levels of between 2000 and 200,000 parasites/\u03bcL of blood. The exclusion criteria included patients treated for malaria in the preceding fourteen days and voluntary withdrawals from the study. Malaria infection was confirmed using microscopy. The study analysed 71 DBS from patients with recurrent malaria infections. These DBS were collected from days 21, 28, 42, and 5 samples from unscheduled days post-drug treatment with either AL or DP.Archived DBS collected in 2016 from a two-arm therapeutic efficacy study (TES) on artemether-lumefantrine (AL), and dihydroartemisinin-piperaquine (DP) were used in this study. A randomized controlled study design was employed where participants were monitored during treatment for adherence to the treatment guidelines. The treatment follow up days were day 7, 14, 21, 28 and 42 post-treatment. The study was conducted without bias towards either sex or gender. The archived DBS were stored at \u2212\u00a020\u00a0\u00b0C. The study's eligibility criteria included: obtaining informed consent, a history of fever with a body temperature of\u2009\u2265\u200937.5\u00a0\u00b0C, P. falciparum msp2 (Pfmsp2) and P. falciparum msp1 (Pfmsp1) were utilized to differentiate recrudescent infections from new infection following the standard methods [Pfmsp2 gene was amplified using the forward primer 5\u2032-GAAGGTAATTAAAACATTGTC-3\u2032 and reverse primer 5\u2032-GAGGGATGTTGCTGCTCCACA-3 for the primary PCR and forward primer 5\u2032-GAGTATAAG GAGAAGTATG-3\u2032 and reverse primer 5\u2032-CTAGAACCATGCATATGTCC-3\u2032 for the secondary PCR. The Pfmsp1 gene was amplified using the forward primer 5\u02b9-CTAGAAGCTTTAGAAGATGCAGTATT-3\u02b9 and reverse primer 5\u02b9-CTTAAATAGTATTCTAATTCAAGTGGATCA-3 for the primary PCR and forward primer 5-AAATGAAGAAGAAATTACTACAAAAGG-3\u02b9 and reverse primer 5\u02b9-GCTTGCATCAGCTGGAGGGCTTGCACC-3\u02b9 for the secondary PCR. Both the Pfmsp2 and Pfmsp1 PCR reactions were conducted under similar conditions. Briefly, the reaction mixture consisted of 5\u2009\u00d7\u2009Firepol mastermix with 12.5\u00a0mM MgCl2 , 10\u00a0\u00b5M of both the forward and reverse primers, nuclease-free water and 1\u00a0\u00b5l of individual genomic DNA extract to a final reaction volume of 20\u00a0\u00b5l. The PCR amplification was carried out in the ProFlex PCR system (Applied Biosystems) using the following optimized conditions: Primary PCR: initial denaturation at 94\u00a0\u00b0C for 3\u00a0min, followed by 30 cycles of denaturation at 94\u00a0\u00b0C for 25\u00a0s, annealing at 42\u00a0\u00b0C for 1\u00a0min and elongation at 65\u00a0\u00b0C for 2\u00a0min followed by a final extension of 72\u00a0\u00b0C for 3\u00a0min. Secondary PCR: initial denaturation at 94\u00a0\u00b0C for 3\u00a0min, followed by 30 cycles of denaturation at 94\u00a0\u00b0C for 25\u00a0s annealing at 50\u00a0\u00b0C for 1\u00a0min and elongation at 72\u00a0\u00b0C for 3\u00a0min followed by a final extension of 72\u00a0\u00b0C for 2\u00a0min.Two highly polymorphic markers,  methods . The PfmPfnfs1 was performed using archived DBS from patients with recurrent parasitaemia. Parasite genomic DNA was extracted from the DBS using the QIAamp DNA mini kit  following instructions from the manufacturer. The target region of the Pfnfs1 gene was amplified by conventional PCR using the forward primer 5\u2019- TTTGTGTTAAAAGACCTCATCCC-3\u2019 and reverse primer 5\u2019- TCTTGGGTCAATCATTGTGGTT-3\u2019 designed using Benchling. Briefly, the reaction mixture consisted of 5\u2009\u00d7\u2009Q5 reaction buffer , 10\u00a0\u00b5M of both the forward and reverse primers, 10\u00a0mM dNTPs mix, Q5 high-fidelity DNA polymerase , nuclease-free water and 1\u00a0\u00b5l of individual genomic DNA extract to a final reaction volume of 25\u00a0\u00b5l. The PCR amplification was carried out in the ProFlex PCR system (Applied Biosystems) using the following optimized conditions: initial denaturation at 98\u00a0\u00b0C for 30\u00a0s, followed by 35 cycles of denaturation at 98\u00a0\u00b0C for 15\u00a0s, annealing at 58\u00a0\u00b0C for 15\u00a0s and elongation at 72\u00a0\u00b0C for 2\u00a0min followed by a final extension of 72\u00a0\u00b0C for 7\u00a0min. The PCR products were analysed in a 1.5% agarose gel and purified using the QIAquick PCR purification kit  as per manufacturer instructions. The purified PCR samples were sequenced using a 3730xl DNA Analyzer sequencer BigDye v3.1 (Applied Biosystems).Analysis of P. falciparum strain obtained from PlasmoDB. Frequencies were calculated as the percentage of sequences that carried a mutation out of the total number of sequences with data for the locus of interest.Molecular Evolutionary Genetics Analysis (MEGA) software version 10  was usedThe study was conducted at the Kenya Medical Research Institute (KEMRI). All experiments were carried out as per relevant national and international standards and approved by the Scientific and Ethics Review Unit (SERU) of the Kenya Medical Research Institute under approval KEMRI/SERU/CTMDR/095/4172. The original study was a therapeutic efficacy study on the efficacy of ACT and was under SERU approval number SSC 2276.Pfnfs1 gene was successfully amplified and sequenced in 64 samples: 44 AL-treated patients and 20-DP-treated patients. 26 AL-treated samples and 14 DP-treated samples sequenced were confidently scored. 24 samples (18 AL-treated and 6 DP-treated samples) were not confidently scored and thus not included for further analysis. On day 21 post-treatment, 13 AL-treated patients had recurrent parasitaemia, while DP-treated patients had no recurrent infection. On day 28 post-treatment, 20 patients exhibited recurrent infection: 17 from the AL-treated and three from DP-treated patients. Further follow-up revealed that 12 AL-treated and 14 DP-treated patients had recurrent parasitaemia yielding a total of 26 patients on day 42 post-treatment. Two AL-treated and 3 DP-treated samples showed recurrent parasitaemia on unscheduled treatment follow-up days  pathway has been proposed as a promising target for anti-malarial drugs, and Additional file 1: Fig S1: Gel image of PCR products from amplification of the Plasmodium falciparum msp2and Plasmodium falciparum msp1gene from selected patients isolates. Lane 1- 100bp ladder; Lane 2- Patient sample 1, Day 0, MSP2 Lane 3- Patient sample 1, Day 28, MSP2, Lane 4- Patient sample 1, Day 0, MSP1, Lane 5- Patient sample 1, Day 28, MSP1; Lane 6- 100bp ladder, Lane 7- Patient sample 2, Day 0, MSP2, Lane 8- Patient sample 2, Day 42, MSP2, Lane 9- Patient sample 2, Day 0, MSP1, Lane 10- Patient sample 2, Day 42, MSP1; Lane 11- 100bp ladder, Lane 12- Patient sample 3, Day 0, MSP2, Lane 13- Patient sample 3, Day 21, MSP2, Lane 14- Patient sample 3, Day 0, MSP1, Lane 15- Patient sample 3, Day 21, MSP1, Lane 16- 100bp ladder. Lane 2-5: New infections, Lane 7-10: Recrudescent infections, Lane 12-15: New infection.Additional file 2: Fig S2: Gel image of PCR products from amplification of the Plasmodium falciparum cysteine desulfurase IscSgene from selected patients isolates. Lane 1- 100bp ladder, Lane 2- Patient sample 1, Lane 3- Patient sample 2, Lane 4- Patient sample 3, Lane 5- Patient sample 4, Lane 6- Patient sample 5, Lane 7- Patient sample 6, Lane 8- Patient sample 7, Lane 9- Patient sample 8, Lane 10- Patient sample 9, Lane 11- Patient sample 10, Lane 12- Patient sample 11, Lane 13- Patient sample 12, Lane 14- Patient sample 13, Lane 15- Patient sample 14, Lane 16- 100bp ladder."}
+{"text": "Limited information exists on any interactions between hydroxyurea (HU) and antimalarials in sickle cell disease (SCD). We evaluated changes in clinical and laboratory parameters among children with SCD on HU therapy treated with artemether-lumefantrine (AL) for acute uncomplicated malaria (UM).Plasmodium falciparum identification of rapid diagnostic test (RDT) positive samples was done using nested polymerase chain reaction (PCR).A prospective, non-randomized, pilot study of 127 children with SCD  were recruited from three hospitals in Accra. UM participants were treated with standard doses of AL and followed up, on days 1, 2, 3, 7, 14, and 28. Venous blood was collected at baseline and follow-up days in participants with UM for determination of malaria parasitaemia, full blood count, reticulocytes, and clinical chemistry. Further, p =\u20090.74) and adverse events , were comparable (p =\u20090.94). Day 28 reticulocyte count was higher in the HU+ (0.24) (0.17 to 0.37) vs. no-HU, . Significant differences in lymphocyte ; bilirubin ; and alanine aminotransferase,  were observed during follow up.Among SCD participants with UM, admission temperature, neutrophils, alanine-aminotransferase, gamma-glutamyl-transferase, and haemoglobin significantly differed between HU recipients (HU+) and steady state, while white blood cell, neutrophils, reticulocytes, bilirubin, urea, and temperature differed significantly between non-HU recipients (no-HU), and steady state. Mean parasitaemia (HU+, 2930.3 vs. no-HU, 1,060, Parasite clearance and adverse event occurrence were comparable between SCD children treated with AL irrespective of HU status. However, distinct patterns of changes in laboratory indices suggest the need for larger, more focused studies. The highest burden of sickle cell disease (SCD) is in sub-Saharan Africa where over 300,000 children are born each year with SCD, and related childhood mortality rates remain high , 2. MalaHydroxyurea (HU) was the first drug approved as a disease modifying agent in sickle cell anaemia and is recommended for use, irrespective of phenotypic severity. HU has multiple benefits in SCD including increases in absolute haemoglobin (Hb), mean corpuscular volume (MCV) and foetal haemoglobin (HbF) levels, reductions in white blood cell (WBC), neutrophil, platelet, and reticulocyte counts, and increased nitric oxide availability. At the cellular level, these effects inhibit sickle polymerization, reduce inflammation, and improve blood rheology , 9.9/L significantly decreased malaria incidence within the study population. The NOHARM study also demonstrated that HU was safe in very young children (ages 1.00\u20133.99\u2009years), with no difference in malaria incidence or severity between children on HU or placebo. NOHARM study participants received malaria chemoprophylaxis, and there were 12 episodes of clinical malaria over a 1\u2009year follow-up period  days 28 and 42 cure rates in SCD patients with acute malaria, using recommended antimalarials such as artemisinin-based combination therapies (ACT). These studies were conducted prior to the introduction of HU as a disease modifying agent in Ghana, where 1 in 50 newborns is affected with SCD , 13. It Studies that evaluate changes in clinical and laboratory parameters in SCD patients on HU during acute malaria would provide information to help guide optimal malaria treatment policy, especially since HU use in Africa has become more widespread and is considered standard of care in SCD. Therefore, our main study objectives were to: (i) evaluate differences, if any, in selected haematological and clinical chemistry parameters between children with SCD on HU therapy treated with artemether-lumefantrine (AL) for uncomplicated malaria, compared to those not on HU therapy, (ii) determine parasite clearance and treatment efficacy and (iii) frequency of occurrence of adverse events (AEs) in both groups during a 28\u2009days follow up period.This was a prospective, non-randomized, pilot study conducted at three sites\u2014Department of Child Health, Korle Bu Teaching Hospital, Greater Accra Regional Hospital, and Princess Marie Louise Children\u2019s Hospital, all in Accra, Ghana. Study recruitment was from June 2021 to August 2022. All three institutions are state-funded public health facilities with specialist paediatricians and paediatric sickle cell clinics. Folate supplementation is given routinely to all children with SCD at the study sites. For patients on HU, Hb F levels are not routinely checked due to the prohibitive cost of testing. Malaria chemoprophylaxis is not used, although families are routinely counselled on other preventive methods like insecticide-treated bed nets. National guidelines for uncomplicated malaria recommend treatment with ACT if there is either a positive malaria rapid diagnostic test (RDT) or blood film (microscopy) showing malaria parasites.Inclusion criteria were, (i) history of fever within the previous 72\u2009h or axillary temperature \u226537.5\u00b0C at the time of presentation to the health facility, (ii) positive malaria RDT, and (iii) decision by attending physician to treat as uncomplicated malaria with ACT. Children with signs and symptoms of severe malaria using WHO criteria  were excluded.One hundred and twenty-seven children, ages one to 15\u2009years, were prospectively recruited as follows: twenty-three (23) children with SCD and RDT-positive uncomplicated malaria who received AL (10 on HU and 13 not on HU) were enrolled and followed up. Additionally, 104 children with SCD in \u201csteady state\u201d reporting to the study sites for routine quarterly follow up clinic visits (58 on HU and 46 not on HU) were enrolled as comparators for the SCD RDT-positive participants. Steady state SCD was defined as clinically asymptomatic with no pain, fever or other intercurrent illness in the 4\u2009weeks prior to recruitment, no blood transfusions in the preceding 4\u2009months , and a nBaseline screening included clinical history, physical examination, malaria RDT , and completion of a structured case record form by study physicians. We collected 2\u2009mL of venous blood into ethylenediaminetetraacetic acid (EDTA) and heparinized tubes for determination of malaria parasitaemia, full blood count, reticulocytes (RET) and clinical chemistry, in all participants with a positive RDT test result. RDT-positive individuals were treated with a 3\u2009days course of AL per manufacturer\u2019s weight-based dosing guidelines. AL was supervised by caregivers of patients treated on outpatient department (OPD) basis. For those who were admitted, the nursing staff supervised their AL course.A complete physical examination, assessment for new or persistent symptoms, AEs, and malaria parasitaemia were done on follow up days 1, 2, 3, 7, 14, and 28. Full blood count, RET and clinical chemistry were repeated on days 3, 7, 14, and 28. Participants in steady state did not require follow up. AEs were assessed during follow up visits and those of grade two or higher [according to the Common Terminology Criteria for Adverse Events (CTCAE) version 5] were documented for all participants.Plasmodium falciparum identification of RDT positive samples was done by extracting genomic DNA from dried blood spots using a chelex extraction procedure and using nested polymerase chain reaction (PCR) approach to amplify the COX III gene, with primers specific for the parasite mitochondrial DNA.Thick and thin blood smears were prepared, stained with Giemsa, and parasite density was determined by counting the number of asexual stage parasites relative to 200 WBC, multiplied by the measured WBC count. Each slide was read independently by two certified microscopists and the average of two readings recorded. Full blood count with leucocyte differential, absolute neutrophil count (ANC) and RET were done using an automated haematology analyzer . Clinical chemistry  was done with the HumaLyte Plus 5 and HumaStar 100 chemistry analyzer systems (Germany). p-values less than 0.05 were considered statistically significant.Continuous data were analyzed as median and/or interquartile range (IQR) or as mean with standard deviation (SD), when normally distributed. Frequencies with percentages were used for categorical data. Overall group differences in the endpoints by patient sub-groups were evaluated using the Kruskal\u2013Wallis test. The Wilcoxon rank-sum rank test was utilized for pair-wise group comparisons. Correlations were expressed as Spearman rank coefficients. Approval for the study was obtained from the KBTH Institutional Review Board (KBTH\u2014IRB/00010/2020) and the Ghana Health Service Ethics Review Committee (GHS-ERC: 014/02/21). Informed consent was obtained from parents or guardians of all recruited participants, and assent obtained from children \u2265age 10\u2009years.The median (range) duration of HU treatment (for those on HU) was 28 (3\u201360) months and the median (range) HU dose was 25 (18\u201335) mg/kg/day.Selected demographic parameters (age and sex) were comparable between those with acute malaria and those in steady state, whether they were on HU or otherwise. Among those on HU (HU+), participants with acute malaria had significantly higher admission temperature, lower Hb, higher ANC, ALT, and gamma-glutamyl-transferase (GGT) than steady state participants. Among the no-HU group, participants with acute malaria had higher admission temperature, WBC, ANC, RET, total bilirubin (BIL) and urea than participants in steady state .n\u2009=\u200910) were all HbSS genotype, while the no-HU participants comprised HbSS (n\u2009=\u20099) and HbSC (n\u2009=\u20094) genotypes. Cardiac murmurs and hepatomegaly were observed more frequently in the HU+ group on day 0, while jaundice and splenomegaly were more prevalent in the no-HU group (The HU+ participants (HU group . Aside fp\u2009=\u20090.38), and by PCR among 60.9% (8/10 HU+ and 6/13 no-HU) , in the HU+ group  of the RDT-positive SCD participants  . GeometrU+ group . None ofU+ group .p\u2009=\u20090.024). Similarly, the change in RET count between HU+ [0.24 (0.17 to 0.37)] and no-HU [0.15 (0.09 to 0.27)] was significant on day 28 (p\u2009=\u20090.022). Graphical presentation of trends in haematological parameters from day 0 to day 28 are shown  but reverted to slightly higher levels by day 28. The change in lymphocyte count (day 0 and day 7) between HU+  [2.74 (\u22125.38 to 58.57)] and no-HU [\u22120.34 (\u22123.19 to 4.44)] groups was significant (re shown .p\u2009=\u20090.018 for both parameters). ALT changes during subsequent follow up days: days 0 and 7 (p\u2009=\u20090.010) and days 0 and 14 (p\u2009=\u20090.023), were also significant. The patterns of changes in other clinical chemistry parameters from day 0 to day 28 are shown (Day 0 (pre-treatment) higher ALT and AST levels in the HU+ group declined sharply, attaining a nadir on day 3. Comparatively, lower day 0 GGT and urea levels in the HU+ group trended upwards between days 3 and 14, reverting to comparable levels as the no-HU group, by day 28. Between days 0 and 3, mean (95% CI) changes in BIL [HU+ \u22124.44 (\u221216.36 to 20.74)]; no-HU [\u221218.37 (\u2212108.79 to \u22127.16)]; and ALT HU+ [\u22124.00 (\u221248.55 to 6.00)], no-HU [7.00 (\u221222.00 to 22.00)], were significant (re shown .p\u2009=\u20090.94). Elevated bilirubin  was the most common adverse event  adverse events occurred in the HU+ group, compared with 14.3% (10/70) in the no-HU group, by day 28 , it was significantly higher in the RDT-positive no-HU group compared to steady state no-HU. SCD causes alterations in blood cell indices characterized by a reduction in Hb and increase in RET, while malaria also causes accelerated breakdown of both parasitized and uninfected red cells, potentially worsening the effect on Hb and RET , 22. AltP. falciparum and P. vivax infections  similar haemoglobin levels at malaria diagnosis in the HU+ and no-HU groups. It may also explain the higher prevalence of anaemia-associated clinical signs at presentation in the HU+ group. The increased Hb by days 7 and 28, in both groups, are in keeping with complete recovery from the acute malaria infection.There were comparable ANC between the HU+ and no-HU malaria groups at baseline and significantly higher ANC in the HU+ group on days 3 and 14 but a reverse trend by day 28. However, the days 3 through 14 ANC among the HU+ group was skewed by the single post-splenectomy participant who had persistent parasitaemia through day 14. Neutrophils play a crucial role in the body\u2019s defense against pathogens and are implicated in the phagocytosis of malaria parasite-infested RBC and free merozoites. Neutrophils also produce reactive oxygen species that inhibit parasite growth . While tin vitro (in vivo (Lymphocyte counts on days 3 and 28 were comparable between the HU+ and no-HU groups. However, the corresponding counts were markedly different between days 7 and 14. While the implication of these dynamics are unknown, T cell lymphocytes play a role in priming phagocytic cells to capture or kill malaria parasites as well as helping B lymphocytes produce functional anti-parasitic antibodies . Lymphocin vitro  and spec(in vivo .Despite lack of any significant differences in biochemical parameters on Day 0 between both malaria groups, the temporal patterns showed marked variations in AST and ALT between days 0 and 28. The nadir of ALT and AST occurred in the HU+ group on day 3, with no change in BIL on day 3\u2014a finding that was not seen in the no-HU group. The latter group had an increase in the AST and ALT values on day 3, consistent with previous reports of self-limiting, disease and/or drug-related liver injury , 34. AltBeing the most prescribed ACT in sub-Saharan Africa , severalChildren with SCD treated with AL for acute uncomplicated malaria had rapid parasite clearance and high day 28 cure rates irrespective of whether they were on prior HU therapy. However, distinct pattern of changes in haematological and blood chemistry parameters between HU+ and no-HU groups in this pilot study, including rapid decline in ALT and AST among the HU+, in contrast with elevated ALT and BIL levels in the no-HU group during the acute illness phase, suggest that further focused studies are needed to evaluate the significance of these observations in the specific context of long term safety of HU during acute malaria and its treatment. Furthermore, our finding of recurrent parasitaemia in a single splenectomized patient, highlights the need for studies to refine the role of the spleen in SCD patients with acute malaria and to advocate for antimalarial preventive interventions for high-risk SCD patients in endemic areas.Although the 28\u2009days follow up schedule was consistent with WHO recommendations for antimalarial efficacy studies, a longer follow up would be a more optimal design in terms of determining the natural history and dynamics of changes in haematological or clinical chemistry parameters and long term safety implications. A heterogeneous SCD population with different phenotypes would be a more robust approach to unravelling the clinical and para-clinical effects of HU in this population.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving humans were approved by Institutional Review Board, Korle Bu Teaching Hospital; and the Ghana Health Service Ethics Review Committee. The studies were conducted in accordance with the local legislation and institutional requirements. Written informed consent for participation in this study was provided by the participants\u2019 legal guardians/next of kin.CS: Conceptualization, Data curation, Investigation, Methodology, Supervision, Validation, Writing \u2013 original draft, Writing \u2013 review & editing. SKA: Conceptualization, Data curation, Investigation, Methodology, Supervision, Validation, Writing \u2013 original draft, Writing \u2013 review & editing. AA: Data curation, Investigation, Methodology, Supervision, Validation, Writing \u2013 original draft, Writing \u2013 review & editing. MN: Conceptualization, Investigation, Methodology, Supervision, Validation, Writing \u2013 review & editing. YB: Conceptualization, Investigation, Methodology, Supervision, Validation, Writing \u2013 review & editing. NS: Conceptualization, Investigation, Methodology, Supervision, Validation, Writing \u2013 review & editing. SYA: Investigation, Methodology, Supervision, Validation, Writing \u2013 review & editing. AS: Data curation, Formal analysis, Investigation, Methodology, Validation, Writing \u2013 review & editing. LH: Conceptualization, Funding acquisition, Methodology, Project administration, Resources, Supervision, Validation, Writing \u2013 review & editing. MO: Conceptualization, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Validation, Writing \u2013 review & editing. GA: Conceptualization, Data curation, Formal analysis, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Validation, Visualization, Writing \u2013 original draft, Writing \u2013 review & editing."}
+{"text": "Plasmodium falciparum  for the selection of optimal drug combinations. First, we showed that the replication of P. falciparum was robust and highly reproducible in the PfalcHuMouse model by retrospective analysis of historical data. Second, we compared the relative value of parasite clearance from blood, parasite regrowth after suboptimal treatment (recrudescence), and cure as variables of therapeutic response to measure the contributions of partner drugs to combinations in vivo. To address the comparison, we first formalized and validated the day of recrudescence (DoR) as a new variable and found that there was a log-linear relationship with the number of viable parasites per mouse. Then, using historical data on monotherapy and two small cohorts of PfalcHuMice evaluated with ferroquine plus artefenomel or piperaquine plus artefenomel, we found that only measurements of parasite killing  as a function of drug exposure in blood allowed direct estimation of the individual drug contribution to efficacy by using multivariate statistical modeling and intuitive graphic displays. Overall, the analysis of parasite killing in the PfalcHuMouse model is a unique and robust experimental in vivo tool to inform the selection of optimal combinations by pharmacometric pharmacokinetic and pharmacodynamic (PK/PD) modeling.The development of new combinations of antimalarial drugs is urgently needed to prevent the spread of parasites resistant to drugs in clinical use and contribute to the control and eradication of malaria. In this work, we evaluated a standardized humanized mouse model of erythrocyte asexual stages of  Plasmodium that infect, feed on, and destroy human erythrocytes ; and third, the lack of standardized methods of data analysis to gauge and compare the contributions of partner drugs in combinations, particularly when the parasites used were not human pathogens .The selection of partner drugs in combinations has been classically addressed by using t ratios \u20139. Howev malaria , 11, prain vivo methodology for the prediction of optimal drug combinations employing a humanized mouse model of human malaria caused by Plasmodium falciparum. In this model, NODscidIL2R\u03b3cnull (NSG) mice are engrafted with human erythrocytes (hE), thereby rendering mice susceptible to the P. falciparum strain 3D70087/N9  .To address these problems, we aimed at developing an e model) . The Pfae model) , resultiothelium , 15. Becothelium \u201332 whichIn this work, we examined the robustness of the PfalcHuMouse model and the relative relevance of parasite clearance, recrudescence, and cure as variables of therapeutic response. These variables were used to investigate the individual contributions of antimalarial drugs in combination by using small experimental studies supported by historical data generated during early phases of discovery of the partner drugs.in vivo. First, the parasitemia at the onset of treatment was originally set up to match the parasite burden in humans as a function of body weight . This normalization approach was chosen because body weight correlates with blood volume and has been used for allometric scaling of many physiological constants among mammals. Thus, the number of parasites in adult patients weighing 70\u2009kg at hospital admission had been previously estimated to be 7\u2009\u00d7\u20091011 parasites (10 parasites/kgbody weight\u2009=\u200910). In the PfalcHuMouse model, an inoculum of 30\u2009\u00d7\u2009106 infected erythrocytes per mouse produced an average parasitemia of 1.42%  at treatment start . Thus, for an estimated volume of distribution of 1.5\u2009mL and 25 g of body weight, the estimated average density of parasites in PfalcHuMice was 9.74 for log10 parasites/kgbody weight, which was on the order of the human estimate. Interestingly, 96% of parasitemia determinations at day 1 of the study fell between 0.66 and 2.7%, making the log10 percent parasitemia on day 1 almost normally distributed, with a rate of 2% for unexpectedly low parasitemias  the distribution of parasitemia at the onset of drug treatment and (ii) the rate of parasite growth over two asexual parasite cycles arasites  . The avsitemias .P. falciparum 3D70087/N9in vivo was measured as the ratio between parasitemia at day 5 (pDay5) and pDay1 in untreated control individuals. The mean log10 pDay5/pDay1 was 1.006    and d. Tn/pDn+2, where pD stands for parasitemia and n indicates the day of the assay), which was a simplified alternative of the classical approach and is frequently used in clinical studies. Conversely, PR depends on the number of viable parasites remaining in the body that could proliferate after noneffective treatment and is not systematically used to measure parasite killing in vivo. To test PR validity, we hypothesized that the time point for recrudescent parasitemia to reach the value at the onset of drug treatment  correlated with the number of parasites that survived antimalarial treatment , parasite recrudescence (PR), and cure . PC depereatment . In our in vivo. To characterize this variable, we evaluated the linearity of DoR as an indicator of the number of viable parasites in the PfalcHuMouse model remaining after drug-mediated parasite clearance. To assess this contention without genetically modifying P. falciparum 3D70087/N9 , we injected different numbers of P. falciparum-infected erythrocytes by the intravenous route and measured the DoR in mice as the time required to achieve a parasitemia of 1.42% in peripheral blood in the absence of treatment (10 inoculum size over at least 6 logs (from 101 to 3\u2009\u00d7\u2009107). In this system, 1 log decrease in parasite numbers represented 2.85\u2009days increase of DoR . Extrapolation of the regression line indicated that the expected DoR for a single inoculated parasite was 23.7\u2009days, counted from the day of inoculation. Accordingly, the expected range of maximum DoR in standard efficacy studies encompassing 4\u2009days of administration should be between 28\u2009days for a rapidly parasiticidal drug and 35 for a slowly parasiticidal drug, counted from the day of last drug administration. However, the predictive power of the extrapolation of the DoR for a single parasite must be interpreted with caution because of the breakdown of linearity for inocula of \u226410 parasites, likely reflecting the stochastic nature of parasite growth from extremely low starting numbers.DoR is not routinely used as a quantitative variable to assess parasite killing reatment . We founn+2/pDn and DoR for a set of oral antimalarials with known mechanism of action at different dose levels and schedules of administration (n\u2009=\u200941 mice). The drugs for which curative exposures were available in the data set, namely, chloroquine, lumefantrine, and pyronaridine, did not result in recrudescence after day 35, roughly consistent with the theoretical calculation described above  plus ferroquine and artefenomel plus piperaquine, which have been recently evaluated in clinical trials , 38, as ferroquine], AUCpiperaquine, and AUCartefenomel) as a PK explanatory variable instead of the dose administered to PfalcHuMice, which is severely affected by the vehicles of administration  and those deemed cured  were excluded from the analysis because their actual DoR was indeterminate, as pointed out in the previous section. The data shown in In the case of the combination piperaquine plus artefenomel, a quantitative comparison with their respective monotherapies of their effect on DoR was not possible at all because all individuals treated with the combination were cured over the range of exposures in which piperaquine and artefenomel monotherapies were not curative  and d. Tn\u2009=\u200950; Tjur\u2019s R2 = 0.31), which was qualitatively consistent with findings in clinical trials  showed that the therapeutic effect of ferroquine and artefenomel was best explained by the sum of the effect size of the exposure of each drug according to the equationl trials .A significant improvement on the coefficient of determination could be obtained by considering an interaction term between drug exposures in the logistic model. However, we observed a high degree of correlation among the explanatory variables (variance inflation factor (VIF) higher than 10 for all explanatory variables). Therefore, the data set did not provide convincing evidence to include such interaction term in the logistic model. To further explore the possibility of the existence of a term of interaction, we examined the effect of adding a virtual individual cured in monotherapy with artefenomel at plausible exposure levels on the logistic function of probability. The results consistently showed that including a factor of mechanistic interaction between the drugs would be preferable over a simple model of summation of individual drug effects . Therefore, cure analysis predicted some level of mechanistic interaction between ferroquine and artefenomel that was not apparent in the analysis of DoR by multiple linear regression, likely due to discarding individuals not eligible for the last type of analysis.n\u2009=\u200940 individuals; Tjur\u2019s R2 = 0.91). Modeling the logistic function by including virtual individuals cured with artefenomel monotherapy at plausible exposures and balanced numbers predicted that the contributions of the two drugs would be similar. In this combination, the introduction of a term of interaction led to no convergence of the logistic model because there was no overlap between the sets of cured and noncured individuals . This result supported the possibility of a mechanistic interaction among the drugs as well, which was not detected in the corresponding clinical study , respectively, led to 95% probability of cure . No cures were available at TAD\u2019s database for monotherapy with artefenomel up to 4.7\u2009\u00d7\u200910\u22123\u2009nmol/106 iE with the schedules of treatment used in the experimental studies . The graphic analysis suggested that artefenomel was at least 10-fold less potent than ferroquine for killing P. falciparum parasites in vivo  , 44.P. falciparum 3D70087/N9. This fact markedly enhanced the predictive power to modeling the effect of drugs in vivo. Of note, in practice, our definition of DoR is independent of the limit of quantification of the technique used to measure parasitemia, conversely to the traditional time of parasite first appearance in blood, which critically depends on that limit. In our model implementation, the level of parasitemia at P0  is in the optimal range of accuracy, sensitivity, and specificity of the flow cytometry technique  as a variable of therapeutic response presented in this paper represents a novelty with respect to the usual statistical treatment of this variable. In general, recrudescence has been traditionally viewed as a \u201ctime-to-event\u201d variable  amenable for survival analysis techniques . In thisechnique . Howevered model . TherefoP. falciparum strains mentioned above. This technical characteristic of cure analysis opens the possibility of extending this analysis to PfalcHuMice infected with field isolates , and human monoclonal antibodies (MAbs).In summary, our results showed that the analysis of cure after treatment with drug combinations in the PfalcHuMouse model was the most informative technique for studying mechanistic antiparasiticidal interactions of candidate drugs https://www.nc3rs.org.uk/arrive-guidelines). The human biological samples were sourced ethically, and their research use was in accord with the terms of the informed consents.Studies with mice were performed at The Art of Discovery and approved by The Art of Discovery Institutional Animal Care and Use Committee (TAD-IACUC). This committee is certified by the Biscay County Government  to evaluate animal research projects from Spanish institutions according to point 43.3 from Royal Decree 53/2013, from 1 February (BOE-A-2013-1337). All experiments were carried out in accordance with European Directive 2010/63/EU. The results from the animal experiments are reported following ARRIVE guidelines except for disclosure of business trade confidential information  mice were purchased from Charles River Laboratories  and adapted to the TAD  animal facility for at least 1\u2009week before entering experimental procedures. The mice were used in experimental studies when their weight was in the range of 23 to 30 g. The mice were provided with ultrafiltered water  and gamma-irradiated standard pelleted diet  P. falciparum 3D70087/N9 has been described previously (0087/N9 was produced at TAD by expansion in human erythrocyte (hE)-engrafted NSG mice. P. falciparum 3D70087/N9 was maintained in vivo by serial passages to avoid the loss of competence to grow in humanized mice after in vitro culture.eviously . The strwww.biobancovasco.org; Galdakao, Basque Country, Spain), Centro de Transfusiones de la Comunidad de Castilla y Le\u00f3n , and Banc de Sang I Teixits . Blood samples were processed following standard operation procedures with appropriate approval of the Ethical and Scientific Committees. Blood was aliquoted and stored at 4\u00b0C until use. Prior to injection, hE were washed twice with RPMI 1640 and 25\u2009mM HEPES  containing 7\u2009\u00d7\u200910\u22123\u2009mM hypoxanthine  at room temperature. Finally, hE were resuspended at 50% or 75% hematocrit in RPMI 1640, 25% decomplemented human serum (Sigma-Aldrich), and 3.1\u2009mM hypoxanthine and warmed at 37\u00b0C for 20\u2009min prior to injection in mice.Concentrates of hE from malaria-negative donors were generously provided by the Basque Biobank , chloroquine , pyrimethamine (AK Scientific), atovaquone (Sigma-Aldrich), and ferroquine  were formulated in 1% methylcellulose  and 0.1% Tween 80  in double-distilled water. Piperaquine (AK Scientific) and artefenomel  were formulated in 0.1% hydroxypropyl-methylcellulose (Sigma-Aldrich), 0.5% benzyl alcohol (Sigma-Aldrich), 0.4% Tween 80, 0.9% sodium chloride  in double-distilled water. In some studies, piperaquine was formulated in double-distilled water. Pyronaridine  was formulated in 0.5% hydroxypropyl-methylcellulose and 0.1% Tween 80.0). The treatments were administered by oral gavage with 20-gauge straight reusable feeding needles . The volume of administration for the per os (p.o.) route was 10\u2009mL/kg of body weight unless otherwise stated.Before drug administration, each infected mouse was randomly assigned to its corresponding treatment. Drug treatment started when infected mice had ~1.4% patent parasitemia in peripheral blood  were taken at different time points after dosing, mixed with 25 \u03bcL of 10.9\u2009mM potassium oxalate and 59.5\u2009mM sodium fluoride in Milli-Q water, and immediately frozen on a thermal block at \u221280\u00b0C. The frozen samples were stored at \u221280\u00b0C until analysis. Blood from P. falciparum 3D70087/N9-infected HuMice harboring a parasitemia of ~1.4% was used for preparation of standard curves, calibration, and quality control samples. The drugs were extracted from 10 \u03bcL of lysates, obtained by protein precipitation of blood samples, by using standard liquid-liquid extraction methods. The samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) for quantification in Waters Micromass ultrahigh-performance triple quadrupole (UPLC-TQD)  mass spectrometers.The concentrations of drugs were measured in whole blood of 6 total erythrocytes counted for detection). Parasitemia was expressed as the percentage of parasitized erythrocytes with respect to the total erythrocytes in circulation and/or as the absolute concentration of circulating parasitized erythrocytes. For qualitative assessment of blood samples from infected mice, blood smears stained with 10% (vol/vol) Giemsa in saline buffer  were prepared and analyzed as described previously  after pharmacological treatment were maintained until day 60 of study. Blood samples were regularly taken every 2 to 3\u2009days and tested from the presence of circulating parasites.Multivariate linear regression, logistic regression, and descriptive statistics were carried out using GraphPad Prism 9.0 . The comparison of regression models was executed using the Akaike information criterion . Probabi0 to last of the drug in blood (h \u00d7 nmol/mL) divided by AUC0 to 96 h of the curve of the concentration of parasites in peripheral blood (h\u2009\u00d7\u2009106 infected hE/mL).The drug exposure as a function of parasitemia was calculated as the ratio between the AUCAll unique materials are available upon reasonable request."
\ No newline at end of file